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As the wind howled and the rain slammed down, a team of nurses, respiratory therapists and a doctor worked through the night to care for 19 tiny babies as Hurricane Laura slammed southwestern Louisiana.The babies, some on ventilators or eating through viagra cost http://natalievartanian.com/buy-viagra-online-without-a-prescription/ a feeding tube, seemed to weather the storm just fine, said Dr. Juan Bossano, the medical director of the neonatal intensive care unit at Lake Charles Memorial Hospital for Women. "They did very viagra cost well. They tolerated it very well. We had a very good day," he said.Laura made landfall early Thursday morning as a Category 4 storm, packing top winds of 150 mph (241 kph), and pushing a storm surge as high as 15 feet in some areas.Hours before it made landfall, officials had to viagra cost move the babies from the women's hospital to the main hospital in the system after it became clear that storm surge could inundate the women's hospital, located on the southern end of Lake Charles.

The hospital has its own generator and hospital administrator Alesha Alford said it was built to withstand hurricane force winds. But in the viagra cost single story facility, there's no room to move up and storm surge in that area was expected to hit nine feet. In a roughly two-hour operation the babies in the intensive care unit were transferred by ambulance to Lake Charles Memorial Hospital, a ten-story facility on the northern side of the city. Trucks carried needed equipment such as incubators.Alford said the storm hadn't yet hit but "the skies looked very ominous." She said everyone pitched in to get supplies moved to the other hospital."It went as smooth as could be because we had everyone viagra cost helping," she said.Alford said three mothers who couldn't be discharged from the women's hospital were also transferred. Two of them had their newborns with them while the child of the third mom was in the intensive care unit.

Parents of the other children in the neonatal intensive viagra cost care unit couldn't stay with them during the storm because there wasn't enough room so Bossano said one nurse was tasked with calling parents to keep them informed of how their children were doing. Bossano occasionally posted updates on Facebook.Once they got situated at the larger hospital and the winds picked up, Alford said the patients were moved into the hallways. To "protect our babies," mattresses were pushed up against the windows to prevent flying glass although none of the windows ended viagra cost up breaking.She said as huge gusts of wind started coming in, they could feel the building vibrate. In addition to Bossano, the medical staff consisted of two neonatal nurse practitioners, 14 nurses and three respiratory therapists who worked on 12-hour shifts. Some of the staff slept on air mattresses in the hallway, Alford said viagra cost.

After making it through the hurricane, the plan was to have the babies stay in Lake Charles. While electricity was out in the city, the hospital has its own viagra cost generator. But Alford said the city's water system has been so heavily damaged that it ultimately forced them to transfer the babies as well as other patients to other hospitals around the state Friday.Both Alford and Bossano repeatedly praised the nursing staff for their work in caring for the babies that in some cases were born weighing only a pound or two. Some of the nursing staff lost their houses in the storm, and they were worried about their own families, but they put those concerns aside to care for their tiny patients."Really the nurses and the respiratory therapists are the heroes here," Bosanno viagra cost said. "They showed that very clearly the way they performed."There aren’t many hospital visitors amid the erectile dysfunction treatment viagra.

But, if you were to walk through intensive-care units at one New York City hospital, you’d see internet-connected speakers—about the size of a stack of Post-it Notes—affixed to the bedrails of some patient beds.It’s part of a project by two Weill Cornell Medicine doctors to help family members speak with ICU patients, often intubated viagra cost or otherwise not able to hold up a phone themselves, from afar.“The patients could be completely sedated, they could be in a coma,” but families still want to be there with them, said Dr. Marc Schiffman, an interventional radiologist and one of the doctors who spearheaded bringing the devices into ICUs.The speakers, now in 11 units at Weill Cornell, are part of a two-way communication system from company Relay, originally developed as a walkie-talkie system of sorts for children to stay in touch with their parents throughout the day. Users on one viagra cost end record snippets of conversation using a mobile app, which are automatically played out loud through the small speaker.Users on the other end push a button on the device to record a response.“Whenever (families) have a story they want to recount, they can just talk into their phone,” Schiffman said. €œIt gives the families a sense of autonomy (and) connection,” even when the patient can’t respond.The effort, dubbed the VoiceLove Project, began about four months ago, at the height of the erectile dysfunction treatment viagra in New York City.Families and other visitors were no longer allowed inside Weill Cornell, but still wanted a way to connect with patients who were sick with erectile dysfunction treatment. Initially, that involved a nurse standing in the ICU and holding up a phone or tablet viagra cost so families could see the patient—a task that took time out of their already busy day, potentially exposed them to erectile dysfunction treatment and often meant using scarce personal protective equipment.“It really wasn’t a practical solution,” said Dr.

Tamatha Fenster, a minimally invasive gynecologic surgeon.So Fenster and Schiffman began brainstorming hands-free technologies they could install directly at the bedside. Schiffman drove viagra cost to a local Target store and bought a few Relay walkie-talkie devices. After testing it with families and patients in the ICU, the two decided it was a “grand slam,” Schiffman said.Since March, hospitals have been trying new ways to keep patients connected to families at home, said Bill Flatley, senior service delivery manager at consulting firm OST. He said he’s mainly seen hospitals repurpose technology usually used for telemedicine, like tablets and cameras mounted on telemedicine carts.It’s likely hospitals will have to continue to restrict visitors, at least as long as viagra cost there’s uncertainty around erectile dysfunction treatment. So it’s integral for staff to figure out processes that make it easy for families to talk to patients—without putting an additional burden on clinicians or expecting them to serve as tech support.For Fenster and Schiffman, deploying walkie-talkies in the ICU for the first time took some leg work.To scale the walkie-talkie system, Schiffman reached out to Relay’s team via the company’s website, and the company agreed to donate roughly 130 devices and waived the per-user subscription fee.

The doctors viagra cost and Relay have continued to work together on best practices for using the devices in ICUs, a use case Relay is marketing and could sell to other hospitals, according to Jon Schniepp, Relay’s senior vice president of marketing.But Fenster and Schiffman couldn’t just bring walkie-talkies into the ICU. In the hospital setting, there are additional quality and privacy concerns. To address those, the doctors created a disposable case, which made it easier to keep the device sterile and blocked passersby from accidentally pressing the button that would transmit sounds to a family’s Relay app.The two spent thousands of dollars out of their own pockets to devise viagra cost the best case design, Fenster said, working with an industrial designer in New Jersey to 3D print different models. The final plastic case, customized with the phrase “VoiceLove” on the front, costs about $10 per case to print and ship. They’ve started reaching out to acute-care and post-acute facilities in California, Texas and other erectile dysfunction treatment hot viagra cost spots to explain how the VoiceLove Project works, hoping to connect other groups with Relay and share the case design.

But the doctors say they’re still working out the logistics of getting the equipment to interested organizationsWhen Dr. George Wanna saw how devastated viagra cost St. George Hospital University Medical Center was by an explosion that shook Beirut, he felt a need to help his hometown. The Aug viagra cost. 4 blast in the city’s harbor ravaged St.

George’s, so Wanna launched a GoFundMe page to viagra cost help the hospital, where a good friend of his, Dr. Alexander Nehme, is chief medical officer.At deadline, more than $86,600 had been raised, with a goal of $100,000. €œThis is the first time in their viagra cost 140-year history when St. George’s Hospital was damaged so severely that it is unable to function,” said Wanna, chair of the otolaryngology department at New York Eye and Ear Infirmary of Mount Sinai and Mount Sinai Beth Israel in New York. €¨St.

George Hospital even remained open during Lebanon’s 15-year civil war, a conflict that wracked Beirut and forced Wanna to spend much of his childhood in bomb shelters. Wanna is also working with Mount Sinai to send medical supplies. €œSt. George Hospital is in need of everything needed to run a hospital—beds, ventilators, protective equipment.” The tragedy also affected Wanna’s family. His parents weren’t home when the blast struck and were unharmed.

But “my parents’ home was severely damaged by the blast. Sadly, we lost the lives of several of my dad’s relatives,” he said via email. Wanna, who spent his residency at Mount Sinai, is grateful to the system. €œThey have given me a chance to have the kind of life I could never have hoped for—they helped me build a home and a life in this great country.”Healthcare leaders tell stories about incidents of racism or discrimination in their careers.Dr. Garth GrahamVP and Chief Community Health OfficerCVS HealthDr.

Patrice HarrisImmediate Past PresidentAmerican Medical AssociationDr. James HildrethPresident and CEOMeharry Medical CollegeDr. Carol MajorAssistant Dean of Diversity and InclusionUniversity of California, Irvine School of MedicineDr. Suzet McKinneyCEO and Executive DirectorIllinois Medical DistrictMarvin O’QuinnPresident and COOCommonSpirit Health.

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The transpopulation represents a vulnerable population segment both socially and medically, with a higher incidence of does medicare cover viagra mental health issues. During the erectile dysfunction treatment outbreak, transgender persons have faced additional social, psychological and physical difficulties.1 2 In Italy and in several other countries access to healthcare has been difficult or impossible thereby hindering the start does medicare cover viagra or continuation of hormonal and psychological treatments. Furthermore, several does medicare cover viagra planned gender-affirming surgeries have been postponed.

These obstacles may have caused an additional psychological burden given the positive effects of medical and surgical treatments on well-being, directly and indirectly, reducing stressors such as does medicare cover viagra workplace discrimination and social inequalities.3 Some organisational aspects should also be considered. Binary gender policies may worsen inequalities and marginalisation of transgender subjects potentially increasing the risk of morbidity and mortality.As with the general population, does medicare cover viagra during the lockdown, the Internet and social media were useful in reducing isolation and, in this particular population, were also relevant for keeping in touch with associations and healthcare facilities with the support of telemedicine services.4 Addressing the role of the telemedicine in the transpopulation, between May and June 2020 we conducted an anonymous web-based survey among transgenders living in Italy (ClinicalTrials.gov Identifier NCT04448418). Among the 108 respondents, with a mean age of 34.3±11.7 years, 73.1% were transmen and 26.9% transwomen and 88.9% were undergoing gender-affirming hormonal treatment (GAHT).

One in four subjects does medicare cover viagra (24.1%) presented a moderate-to-severe impact of the viagra event (Impact of Event Scale score ≥26). The availability of telematic endocrinological visit was associated with better Mental Health Scores in the 12-items Short Form Health Survey(SF-12) (p=0.030) and better IES (p=0.006).Our survey suggests a positive effect of telemedicine as the availability of telematic endocrinological consultations may have relieved the distress caused by the viagra by offering the opportunity to avoid does medicare cover viagra halting GAHT. In fact, deprivation of GAHT may result in several negative effects such as the increase in short-term self-medication and in depression and suicidal behaviour not only for those waiting for the start of treatment but also for those already using hormones.5 In conclusion, does medicare cover viagra particular attention should be paid to vulnerable groups like the transpopulation who may pay a higher price during the viagra.

The use of telemedicine for continuation and monitoring of GAHT may be an effective tool for mitigating the negative effects of the viagra.AcknowledgmentsThe authors thank Julie Norbury does medicare cover viagra for English copy editing.The British Medical Association recently published their report on the impact of erectile dysfunction treatment on mental health in England, highlighting the urgent need for investment in mental health services and further recruitment of mental health staff.1 Like many others, they have predicted a substantial increase in demand on mental health services in the coming months. Their recommendations include a call for does medicare cover viagra detailed workforce planning at local, national and system levels. This coincides with the publication of the ‘NHS People Plan’ which also emphasised the need to maximise staff potential.2 The message from does medicare cover viagra both is clear, it is time for Trusts to revise and improve how they use their multidisciplinary workforce, including non-medical prescribers (NMPs).Pharmacists have been able to register as independent prescribers since 20063 and as such, can work autonomously to prescribe any medicine for any medical condition within their areas of competency.4 There has been a slow uptake of pharmacists into this role5 and while a recent General Pharmaceutical Council survey found only a small increase between the number of active prescribers from 2013 (1.094) to 2019 (1.590), almost a quarter of prescribers included mental health within their prescribing practice.6 More recently, we have started to see increasing reports of the value of pharmacist independent prescribers in mental health services.7 8Pharmacists bring a unique perspective to patient consultation.

Their expertise in pharmacology and medicine use means they are ideally placed to help patients optimise their medicines treatment4 and to ensure that patients are involved in decisions about their medicines, taking into account individual views and preferences. This approach is consistent with the guidance does medicare cover viagra on medicines optimisation from the National Institute for Health and Care Excellence9 and the Royal Pharmaceutical Society,10 and the Department of Health’s drive to involve patients actively in clinical decisions.11 An increased focus on precision psychiatry in urging clinicians to tailor medicines to patients according to evidence about individualised risks and benefits.12 13 However, it takes time to discuss medicine choices and to explore individual beliefs about medicines. This is especially relevant in does medicare cover viagra Psychiatry, where a large group of medicines (eg, antipsychotics) may have a wide range of potential side effects.

Prescribing pharmacists could provide leadership does medicare cover viagra and support in tailoring medicines for patients, as part of the wider multidisciplinary team.10The recent news that Priadel, the most commonly used brand of lithium in the UK, is planned to be discontinued14 is another example where a new and unexpected burden on psychiatric services could be eased by sharing the workload with prescribing pharmacists. The Medicines and Healthcare Products Regulatory Agency recommends that patients should have an individualised medication review in order to does medicare cover viagra switch from one brand of lithium to another.14 This is work that can be done by prescribing pharmacists who have an in-depth knowledge of the pharmacokinetics of lithium formulations.Importantly, this is a role that can be delivered using telepsychiatry and enhanced by the use of digital tools. Patients can meet pharmacists does medicare cover viagra from the comfort of their own home using video conferencing.

Pharmacists can upload and share medicines information on the screen while discussing the benefits, risks and individual medication needs with each client. Increasingly organisations are using technology whereby prescriptions can be prepared electronically and sent securely to patients or their medicines providers.15We know from systematic reviews that NMPs in general are considered to provide a responsive, efficient and convenient service5 and to deliver similar prescribing outcomes as doctors.16 Medical professionals who have worked with NMPs have found that this support permits them to concentrate on clinical issues that require medical expertise.5 A patient survey carried out in 2013 indicated that independent non‐medical prescribing was valued highly by patients and that generally there were few perceived differences in the care received from respondents’ NMP and their usual doctor.17 The literature also suggests that an NMP’s role is more likely to flourish when linked to a strategic vision of NMPs within an National Health Service (NHS) Trust, along with a well-defined area of practice.18Mental health trusts are does medicare cover viagra being asked to prepare for a surge in referrals and as part of this planning, they will need to ensure that they get the most out of their highly skilled workforce. There are active pharmacist prescribers in many trusts, however, this role is not does medicare cover viagra yet commonplace.19 Health Education England has already identified that this is an important area of transformation for pharmacy and has called on mental health pharmacy teams to develop and share innovative ways of working.19 The ‘NHS People Plan’ outlines a commitment to train 50 community-based specialist mental health pharmacists within the next 2 years, along with a plan to extend the pharmacy foundation training to create a sustainable supply of prescribing pharmacists in future years.2We suggest that Mental Health Trusts should urgently develop prescribing roles for specialist mental health pharmacists, which are integrated within mental health teams.

In these roles, prescribing pharmacists can actively support their multidisciplinary colleagues in does medicare cover viagra case discussion meetings. Furthermore, they should host regular medication review clinics, where patients can be referred to discuss their medicine options and, as advancements in precision therapeutics continue, have their treatment does medicare cover viagra individually tailored to their needs. This is the way forward for a modern and patient-oriented NHS in the UK..

The transpopulation represents a vulnerable population segment both socially and medically, with viagra cost a best place to buy viagra higher incidence of mental health issues. During the erectile dysfunction treatment outbreak, transgender persons have faced additional social, psychological and physical difficulties.1 2 viagra cost In Italy and in several other countries access to healthcare has been difficult or impossible thereby hindering the start or continuation of hormonal and psychological treatments. Furthermore, several viagra cost planned gender-affirming surgeries have been postponed. These obstacles may have caused an additional psychological burden given the positive effects of medical and surgical treatments on well-being, directly viagra cost and indirectly, reducing stressors such as workplace discrimination and social inequalities.3 Some organisational aspects should also be considered.

Binary gender policies may worsen inequalities and marginalisation of transgender subjects potentially increasing the risk of morbidity and mortality.As with viagra cost the general population, during the lockdown, the Internet and social media were useful in reducing isolation and, in this particular population, were also relevant for keeping in touch with associations and healthcare facilities with the support of telemedicine services.4 Addressing the role of the telemedicine in the transpopulation, between May and June 2020 we conducted an anonymous web-based survey among transgenders living in Italy (ClinicalTrials.gov Identifier NCT04448418). Among the 108 respondents, with a mean age of 34.3±11.7 years, 73.1% were transmen and 26.9% transwomen and 88.9% were undergoing gender-affirming hormonal treatment (GAHT). One in four subjects (24.1%) presented a moderate-to-severe impact of the viagra viagra cost event (Impact of Event Scale score ≥26). The availability of telematic endocrinological visit was associated with better Mental Health Scores in the 12-items Short Form Health Survey(SF-12) (p=0.030) and better IES viagra cost (p=0.006).Our survey suggests a positive effect of telemedicine as the availability of telematic endocrinological consultations may have relieved the distress caused by the viagra by offering the opportunity to avoid halting GAHT.

In fact, deprivation of GAHT may result in viagra cost several negative effects such as the increase in short-term self-medication and in depression and suicidal behaviour not only for those waiting for the start of treatment but also for those already using hormones.5 In conclusion, particular attention should be paid to vulnerable groups like the transpopulation who may pay a higher price during the viagra. The use of telemedicine viagra cost for continuation and monitoring of GAHT may be an effective tool for mitigating the negative effects of the viagra.AcknowledgmentsThe authors thank Julie Norbury for English copy editing.The British Medical Association recently published their report on the impact of erectile dysfunction treatment on mental health in England, highlighting the urgent need for investment in mental health services and further recruitment of mental health staff.1 Like many others, they have predicted a substantial increase in demand on mental health services in the coming months. Their recommendations include a call for detailed workforce planning at local, national and viagra cost system levels. This coincides with the publication of the ‘NHS People Plan’ which also emphasised the need to maximise staff potential.2 The message from both is clear, it is time for Trusts to revise and improve how they use their multidisciplinary workforce, including non-medical prescribers (NMPs).Pharmacists have been able to register as independent prescribers since 20063 and as such, can work autonomously to viagra cost prescribe any medicine for any medical condition within their areas of competency.4 There has been a slow uptake of pharmacists into this role5 and while a recent General Pharmaceutical Council survey found only a small increase between the number of active prescribers from 2013 (1.094) to 2019 (1.590), almost a quarter of prescribers included mental health within their prescribing practice.6 More recently, we have started to see increasing reports of the value of pharmacist independent prescribers in mental health services.7 8Pharmacists bring a unique perspective to patient consultation.

Their expertise in pharmacology and medicine use means they are ideally placed to help patients optimise their medicines treatment4 and to ensure that patients are involved in decisions about their medicines, taking into account individual views and preferences. This approach is consistent with the guidance on medicines optimisation from the National Institute for Health and Care Excellence9 and the Royal Pharmaceutical Society,10 and the Department of Health’s drive to involve patients actively in clinical decisions.11 An increased focus on precision psychiatry in urging clinicians to tailor medicines to patients according to evidence about individualised risks and benefits.12 13 However, it takes time to discuss medicine choices and to explore viagra cost individual beliefs about medicines. This is especially relevant in Psychiatry, where a viagra cost large group of medicines (eg, antipsychotics) may have a wide range of potential side effects. Prescribing pharmacists could provide leadership and support in tailoring medicines for patients, as part of the wider multidisciplinary team.10The recent news that Priadel, the most commonly used brand of lithium in the UK, is planned to be discontinued14 is another example where a new and unexpected burden on psychiatric services could be eased by sharing viagra cost the workload with prescribing pharmacists.

The Medicines and Healthcare Products Regulatory Agency recommends that patients should have an individualised medication review in order to switch from one brand of lithium to another.14 viagra cost This is work that can be done by prescribing pharmacists who have an in-depth knowledge of the pharmacokinetics of lithium formulations.Importantly, this is a role that can be delivered using telepsychiatry and enhanced by the use of digital tools. Patients can meet pharmacists from the comfort viagra cost of their own home using video conferencing. Pharmacists can upload and share medicines information on the screen while discussing the benefits, risks and individual medication needs with each client. Increasingly organisations are using technology whereby prescriptions can be prepared electronically and sent securely to patients or their medicines providers.15We know from systematic reviews that NMPs in general are considered to provide a responsive, efficient and convenient service5 and to deliver similar prescribing outcomes as doctors.16 Medical professionals who have worked with NMPs have found that this support permits them to concentrate on clinical issues that require medical expertise.5 A patient survey carried out in 2013 indicated that independent non‐medical prescribing was valued highly by patients and that generally there were few perceived differences in the care received from viagra cost respondents’ NMP and their usual doctor.17 The literature also suggests that an NMP’s role is more likely to flourish when linked to a strategic vision of NMPs within an National Health Service (NHS) Trust, along with a well-defined area of practice.18Mental health trusts are being asked to prepare for a surge in referrals and as part of this planning, they will need to ensure that they get the most out of their highly skilled workforce.

There are active pharmacist prescribers in many trusts, however, this role is not yet commonplace.19 Health Education England has already identified that this is an important area of transformation for pharmacy and has called on mental health pharmacy teams to develop and share innovative ways of working.19 The ‘NHS People Plan’ outlines a commitment to train 50 community-based specialist mental health pharmacists within the next 2 years, along with a plan to extend the pharmacy foundation viagra cost training to create a sustainable supply of prescribing pharmacists in future years.2We suggest that Mental Health Trusts should urgently develop prescribing roles for specialist mental health pharmacists, which are integrated within mental health teams. In these roles, prescribing pharmacists can actively support their multidisciplinary colleagues in case discussion viagra cost meetings. Furthermore, they should host regular medication review clinics, where patients can be referred to discuss their medicine options and, as advancements in precision therapeutics continue, have their treatment individually tailored to their viagra cost needs. This is the way forward for a modern and patient-oriented NHS in the UK..

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After voters expanded Medicaid in conservative states like Missouri and Oklahoma, health care advocates are renewing a push for expansion in Mississippi and other Southern states where Republican leaders have long been opposed.They say the changing tide has followed rising income inequality, joblessness and pressure from hospitals in economic turmoil — issues exacerbated by the erectile dysfunction viagra."There have been, in the last two years, votes on Medicaid expansion in some of the most conservative, Republican-leaning states in the country, and Medicaid expansion has never lost," said Eliot Fishman, senior director of Health Policy at Families USA, a health care advocacy organization.Fishman spoke Thursday during an online viagra cost per pill forum about Medicaid expansion hosted by the Mississippi Health Advocacy Program and the Mississippi Center for Justice.Medicaid expansion is an option under the health care overhaul that Can you get levitra without a prescription then-President Barack Obama signed into law in 2010. Many Democratic-controlled states agreed to expansion, mainly for people whose jobs don't provide health insurance.However, since Republican Donald Trump became president in January 2017, voters in Idaho, Nebraska, Utah, Oklahoma, viagra cost per pill Maine and most recently Missouri have approved Medicaid expansion by ballot measures. In Virginia, legislators passed Medicaid expansion after Democrats gained power."This is clearly an issue which you can no longer shut down voter interest by just saying the word 'Obamacare,' " Fishman said. "That power has waned."There are now 12 states — including Mississippi, Georgia, Alabama, Texas, South Carolina, North Carolina, viagra cost per pill Tennessee and Florida — that have not expanded Medicaid. A newly formed collaborative, "Southerners for Medicaid Expansion," is aiming to put pressure on viagra cost per pill the holdouts.Medicaid is a government health insurance program for the needy, aged, blind and disabled, and it is paid by state and federal money.

Because Mississippi is poor, the federal government pays nearly 78% of the cost.Under expansion, the federal government pays 90% of the cost in any state.About 25% of Mississippi's nearly 3 million residents are already enrolled in Medicaid, and opponents have said they don't want more people taking part in a government program.Roy Mitchell, executive director of the Mississippi Health Advocacy Program, said hospitals are in desperate need of the dollars. Uncompensated care viagra cost per pill costs in Mississippi exceed $600 million annually, according to a 2019 statement from the Mississippi Hospital Association."Let's face it, providers are businessmen. Despite their marketing, they are inherently out to make a profit, and they are going to have to wake up in Mississippi," Mitchell said. "I'm sure erectile dysfunction treatment viagra cost per pill did a good job of doing that."Addressing ailing hospitals has been controversial. Republican Gov viagra cost per pill.

Tate Reeves and leaders in the Republican-controlled Mississippi Legislature have opposed Medicaid expansion. Reeves has said money should go instead to federally funded community health centers that help people in need.Reeves has said repeatedly that the viagra has not changed his mind viagra cost per pill about expansion. And Republican House Speaker Philip Gunn told reporters this month that he also remains opposed.The Mississippi Hospital Association in 2019 proposed "Mississippi Cares," which it called Medicaid reform but not expansion. It was modeled after an Indiana viagra cost per pill program enacted under then-Gov. Mike Pence viagra cost per pill.

It would expand Medicaid eligibility while setting $20 monthly premium payments and copays. The proposal gained no traction during this year's Mississippi legislative session.While advocates for Medicaid expansion say they are hopeful, they acknowledge difficulties.Out of the 12 nonexpansion states, Mississippi and Florida are the only two with a ballot initiative process.Mississippi law says that for an initiative to be placed on the ballot, at least 106,190 certified signatures must be gathered, and those must be evenly divided among the five congressional districts that Mississippi used 20 years ago viagra cost per pill. Even if signature-gathering is successful, the earliest a proposal is likely to be on the ballot is November 2022."For all the stars to align in a ballot initiative ... There's got to be a commitment on the part of providers in Mississippi, I think, and we have to also look realistically at the viagra cost per pill amount of resources that it takes to do a ballot initiative," Mitchell said. "But it is certainly not out of reach."Consultations via tablets, laptops and phones linked patients and doctors when society shut down viagra cost per pill in early spring.

Telehealth visits dropped with the reopening, but they're still far more common than before and now there's a push to make them widely available in the future.Permanently expanding access will involve striking a balance between costs and quality, dealing with privacy concerns and potential fraud, and figuring out how telehealth can reach marginalized patients, including people with mental health problems."I don't think it is ever going to replace in-person visits, because sometimes a doctor needs to put hands on a patient," said CMS Administrator Seema Verma, the Trump administration's leading advocate for telehealth.Caveats aside, "it's almost a modern-day house call," she added."It's fair to say that telemedicine was in its infancy prior to the viagra, but it's come of age this year," said Murray Aitken of the data firm IQVIA, which tracks the impact.In the depths of the erectile dysfunction shutdown, telehealth accounted for more than 40% of primary care visits for patients with traditional Medicare, up from a tiny 0.1% sliver before the public health emergency. As the government's flagship health care program, Medicare covers more than 60 million people, including those age 65 and older, viagra cost per pill and younger disabled people.A recent poll of older adults by the University of Michigan Institute for Healthcare Policy &. Innovation found that more than 7 in 10 are interested in using telehealth for follow-ups with their doctor, and nearly 2 out of 3 feel comfortable with video conferences.But privacy was an issue, especially for those viagra cost per pill who hadn't tried telehealth. The poll found 27% of older adults who had not had a telemedicine visit were concerned about privacy, compared with 17% of those who tried it.Those who tried telehealth weren't completely sold. About 4 in 5 were concerned the doctor couldn't physically examine them, and 64% worried viagra cost per pill the quality wasn't as good."After the initial excitement, in the afterglow, patients realize 'I can't get my treatment,' or 'You can't see this thing in the back of my throat over the computer,' " said Dr.

Gary LeRoy of Dayton, Ohio, a primary care doctor and president of the American Academy of Family Physicians.For Medicare beneficiary Jean Grady of Westford, Vermont, telemedicine was a relief. She needed a checkup required by Medicare to continue receiving supplies for her wearable insulin pump viagra cost per pill. Being in a high risk group for erectile dysfunction treatment, Grady worried about potential exposure in a doctor's waiting room, and viagra cost per pill even more about losing her diabetes supplies if she missed Medicare's checkup deadline."I would have had to go back to taking insulin by syringe," she said.Grady prepared for the virtual visit by calling her clinician's tech department and downloading teleconference software. She says she would do some future visits by video, but not all. For example, people with diabetes need periodic blood tests, and their feet must be checked for signs of circulatory problems.Still, quite a few follow-ups "could be done very efficiently and be just as useful to the physician and myself as going in and seeing them in person," Grady said.Many private insurance plans, including those in Medicare Advantage, offer some level of telemedicine coverage.But traditional Medicare has restricted it to rural viagra cost per pill residents, who generally had to travel to specially designated sites to connect.Under the erectile dysfunction public health emergency, the administration temporarily waived Medicare's restrictions so enrollees anywhere could use telemedicine.

Patients could connect from home. Making such changes permanent would require legislation from Congress, but viagra cost per pill there's bipartisan interest.Sen. Lamar Alexander, chairman of the Senate Health, Education, Labor and Pensions Committee, says he'd like to see broader access, without breaking the bank."Our job should be to ensure that change is done with the goals of better viagra cost per pill outcomes and better patient experiences, at a lower cost," said Alexander, R-Tenn.That's a tall order.Payment will be a sticky obstacle. For now, Medicare is paying clinicians on par for virtual and in-person visits."Policymakers seems to be in a rush to pass legislation, but I think it is worth taking a little more time," said Juliette Cubanski, a Medicare expert with the nonpartisan Kaiser Family Foundation. "Fraud is one big area that policymakers need to be cognizant of."Fraud-busters agree.Telehealth is so new that "we don't have at this point a real sense of where the huge risks lie," said viagra cost per pill Andrew VanLandingham, a senior lawyer with the Health and Human Services inspector general's office.

"We are sort of in an experimental phase."Despite the risks, advocates see opportunities.Expanded Medicare telehealth could:help move the nation closer to a long-sought goal of treating mental health the same as physical conditions. Sen. Ron Wyden, D-Ore., wants to use telemedicine as a springboard to improve mental health care. IQVIA data shows 60% of psychiatric consults took place by telehealth during the shutdown.increase access for people living in remote communities, in low-income urban areas and even nursing homes. Medicare's research shows low-income beneficiaries have had similar patterns of using telehealth for primary care as program enrollees overall.improve coordination of care for people with chronic health conditions, a goal that requires patient and persistent monitoring.

Chronic care accounts for most program spending.University of Michigan health policy expert Mark Fendrick says Medicare should figure out what services add value for patients' health and taxpayers' wallets, and pay just for those.Telehealth "was an overnight sensation," said Fendrick. "Hopefully it's not a one-hit wonder."As the wind howled and the rain slammed down, a team of nurses, respiratory therapists and a doctor worked through the night to care for 19 tiny babies as Hurricane Laura slammed southwestern Louisiana.The babies, some on ventilators or eating through a feeding tube, seemed to weather the storm just fine, said Dr. Juan Bossano, the medical director of the neonatal intensive care unit at Lake Charles Memorial Hospital for Women. "They did very well. They tolerated it very well.

We had a very good day," he said.Laura made landfall early Thursday morning as a Category 4 storm, packing top winds of 150 mph (241 kph), and pushing a storm surge as high as 15 feet in some areas.Hours before it made landfall, officials had to move the babies from the women's hospital to the main hospital in the system after it became clear that storm surge could inundate the women's hospital, located on the southern end of Lake Charles. The hospital has its own generator and hospital administrator Alesha Alford said it was built to withstand hurricane force winds. But in the single story facility, there's no room to move up and storm surge in that area was expected to hit nine feet. In a roughly two-hour operation the babies in the intensive care unit were transferred by ambulance to Lake Charles Memorial Hospital, a ten-story facility on the northern side of the city. Trucks carried needed equipment such as incubators.Alford said the storm hadn't yet hit but "the skies looked very ominous." She said everyone pitched in to get supplies moved to the other hospital."It went as smooth as could be because we had everyone helping," she said.Alford said three mothers who couldn't be discharged from the women's hospital were also transferred.

Two of them had their newborns with them while the child of the third mom was in the intensive care unit. Parents of the other children in the neonatal intensive care unit couldn't stay with them during the storm because there wasn't enough room so Bossano said one nurse was tasked with calling parents to keep them informed of how their children were doing. Bossano occasionally posted updates on Facebook.Once they got situated at the larger hospital and the winds picked up, Alford said the patients were moved into the hallways. To "protect our babies," mattresses were pushed up against the windows to prevent flying glass although none of the windows ended up breaking.She said as huge gusts of wind started coming in, they could feel the building vibrate. In addition to Bossano, the medical staff consisted of two neonatal nurse practitioners, 14 nurses and three respiratory therapists who worked on 12-hour shifts.

Some of the staff slept on air mattresses in the hallway, Alford said. After making it through the hurricane, the plan was to have the babies stay in Lake Charles. While electricity was out in the city, the hospital has its own generator. But Alford said the city's water system has been so heavily damaged that it ultimately forced them to transfer the babies as well as other patients to other hospitals around the state Friday.Both Alford and Bossano repeatedly praised the nursing staff for their work in caring for the babies that in some cases were born weighing only a pound or two. Some of the nursing staff lost their houses in the storm, and they were worried about their own families, but they put those concerns aside to care for their tiny patients."Really the nurses and the respiratory therapists are the heroes here," Bosanno said.

"They showed that very clearly the way they performed."There aren’t many hospital visitors amid the erectile dysfunction treatment viagra. But, if you were to walk through intensive-care units at one New York City hospital, you’d see internet-connected speakers—about the size of a stack of Post-it Notes—affixed to the bedrails of some patient beds.It’s part of a project by two Weill Cornell Medicine doctors to help family members speak with ICU patients, often intubated or otherwise not able to hold up a phone themselves, from afar.“The patients could be completely sedated, they could be in a coma,” but families still want to be there with them, said Dr. Marc Schiffman, an interventional radiologist and one of the doctors who spearheaded bringing the devices into ICUs.The speakers, now in 11 units at Weill Cornell, are part of a two-way communication system from company Relay, originally developed as a walkie-talkie system of sorts for children to stay in touch with their parents throughout the day. Users on one end record snippets of conversation using a mobile app, which are automatically played out loud through the small speaker.Users on the other end push a button on the device to record a response.“Whenever (families) have a story they want to recount, they can just talk into their phone,” Schiffman said. €œIt gives the families a sense of autonomy (and) connection,” even when the patient can’t respond.The effort, dubbed the VoiceLove Project, began about four months ago, at the height of the erectile dysfunction treatment viagra in New York City.Families and other visitors were no longer allowed inside Weill Cornell, but still wanted a way to connect with patients who were sick with erectile dysfunction treatment.

Initially, that involved a nurse standing in the ICU and holding up a phone or tablet so families could see the patient—a task that took time out of their already busy day, potentially exposed them to erectile dysfunction treatment and often meant using scarce personal protective equipment.“It really wasn’t a practical solution,” said Dr. Tamatha Fenster, a minimally invasive gynecologic surgeon.So Fenster and Schiffman began brainstorming hands-free technologies they could install directly at the bedside. Schiffman drove to a local Target store and bought a few Relay walkie-talkie devices. After testing it with families and patients in the ICU, the two decided it was a “grand slam,” Schiffman said.Since March, hospitals have been trying new ways to keep patients connected to families at home, said Bill Flatley, senior service delivery manager at consulting firm OST. He said he’s mainly seen hospitals repurpose technology usually used for telemedicine, like tablets and cameras mounted on telemedicine carts.It’s likely hospitals will have to continue to restrict visitors, at least as long as there’s uncertainty around erectile dysfunction treatment.

So it’s integral for staff to figure out processes that make it easy for families to talk to patients—without putting an additional burden on clinicians or expecting them to serve as tech support.For Fenster and Schiffman, deploying walkie-talkies in the ICU for the first time took some leg work.To scale the walkie-talkie system, Schiffman reached out to Relay’s team via the company’s website, and the company agreed to donate roughly 130 devices and waived the per-user subscription fee. The doctors and Relay have continued to work together on best practices for using the devices in ICUs, a use case Relay is marketing and could sell to other hospitals, according to Jon Schniepp, Relay’s senior vice president of marketing.But Fenster and Schiffman couldn’t just bring walkie-talkies into the ICU. In the hospital setting, there are additional quality and privacy concerns. To address those, the doctors created a disposable case, which made it easier to keep the device sterile and blocked passersby from accidentally pressing the button that would transmit sounds to a family’s Relay app.The two spent thousands of dollars out of their own pockets to devise the best case design, Fenster said, working with an industrial designer in New Jersey to 3D print different models. The final plastic case, customized with the phrase “VoiceLove” on the front, costs about $10 per case to print and ship.

They’ve started reaching out to acute-care and post-acute facilities in California, Texas and other erectile dysfunction treatment hot spots to explain how the VoiceLove Project works, hoping to connect other groups with Relay and share the case design. But the doctors say they’re still working out the logistics of getting the equipment to interested organizationsWhen Dr. George Wanna saw how devastated St. George Hospital University Medical Center was by an explosion that shook Beirut, he felt a need to help his hometown. The Aug.

4 blast in the city’s harbor ravaged St. George’s, so Wanna launched a GoFundMe page to help the hospital, where a good friend of his, Dr. Alexander Nehme, is chief medical officer.At deadline, more than $86,600 had been raised, with a goal of $100,000. €œThis is the first time in their 140-year history when St. George’s Hospital was damaged so severely that it is unable to function,” said Wanna, chair of the otolaryngology department at New York Eye and Ear Infirmary of Mount Sinai and Mount Sinai Beth Israel in New York.

€¨St. George Hospital even remained open during Lebanon’s 15-year civil war, a conflict that wracked Beirut and forced Wanna to spend much of his childhood in bomb shelters. Wanna is also working with Mount Sinai to send medical supplies. €œSt. George Hospital is in need of everything needed to run a hospital—beds, ventilators, protective equipment.” The tragedy also affected Wanna’s family.

His parents weren’t home when the blast struck and were unharmed. But “my parents’ home was severely damaged by the blast. Sadly, we lost the lives of several of my dad’s relatives,” he said via email. Wanna, who spent his residency at Mount Sinai, is grateful to the system. €œThey have given me a chance to have the kind of life I could never have hoped for—they helped me build a home and a life in this great country.”.

After voters expanded Medicaid in conservative states like Missouri and Oklahoma, health care advocates are renewing a push for expansion in Mississippi and other Southern states where Republican leaders have long been opposed.They say the changing tide has followed rising income inequality, joblessness and pressure from hospitals in economic turmoil — issues exacerbated by the erectile dysfunction viagra."There have been, in the last two years, votes on Medicaid expansion in some of the most conservative, Republican-leaning states in the country, and Medicaid expansion has never lost," said Eliot Fishman, senior viagra cost director of Health Policy at Families USA, a health care advocacy organization.Fishman spoke Thursday during an online forum about Medicaid expansion hosted by the Mississippi Health Advocacy Program and the Mississippi Center for Justice.Medicaid expansion is an option under the health care overhaul that then-President Barack Obama signed into law in 2010. Many Democratic-controlled states agreed to expansion, mainly for people whose jobs don't provide health insurance.However, since Republican Donald Trump became president in January 2017, voters in Idaho, Nebraska, Utah, Oklahoma, Maine and most recently Missouri have approved viagra cost Medicaid expansion by ballot measures. In Virginia, legislators passed Medicaid expansion after Democrats gained power."This is clearly an issue which you can no longer shut down voter interest by just saying the word 'Obamacare,' " Fishman said.

"That power has waned."There are now 12 states — including Mississippi, Georgia, Alabama, Texas, South Carolina, North Carolina, Tennessee and Florida viagra cost — that have not expanded Medicaid. A newly formed collaborative, "Southerners for Medicaid Expansion," is aiming to put pressure on the holdouts.Medicaid is a government health insurance viagra cost program for the needy, aged, blind and disabled, and it is paid by state and federal money. Because Mississippi is poor, the federal government pays nearly 78% of the cost.Under expansion, the federal government pays 90% of the cost in any state.About 25% of Mississippi's nearly 3 million residents are already enrolled in Medicaid, and opponents have said they don't want more people taking part in a government program.Roy Mitchell, executive director of the Mississippi Health Advocacy Program, said hospitals are in desperate need of the dollars.

Uncompensated care costs in Mississippi exceed $600 million annually, according to a 2019 statement from the Mississippi Hospital Association."Let's face viagra cost it, providers are businessmen. Despite their marketing, they are inherently out to make a profit, and they are going to have to wake up in Mississippi," Mitchell said. "I'm sure erectile dysfunction treatment did a good job of doing that."Addressing viagra cost ailing hospitals has been controversial.

Republican Gov viagra cost. Tate Reeves and leaders in the Republican-controlled Mississippi Legislature have opposed Medicaid expansion. Reeves has said viagra cost money should go instead to federally funded community health centers that help people in need.Reeves has said repeatedly that the viagra has not changed his mind about expansion.

And Republican House Speaker Philip Gunn told reporters this month that he also remains opposed.The Mississippi Hospital Association in 2019 proposed "Mississippi Cares," which it called Medicaid reform but not expansion. It was modeled viagra cost after an Indiana program enacted under then-Gov. Mike Pence viagra cost.

It would expand Medicaid eligibility while setting $20 monthly premium payments and copays. The proposal gained no traction during viagra cost this year's Mississippi legislative session.While advocates for Medicaid expansion say they are hopeful, they acknowledge difficulties.Out of the 12 nonexpansion states, Mississippi and Florida are the only two with a ballot initiative process.Mississippi law says that for an initiative to be placed on the ballot, at least 106,190 certified signatures must be gathered, and those must be evenly divided among the five congressional districts that Mississippi used 20 years ago. Even if signature-gathering is successful, the earliest a proposal is likely to be on the ballot is November 2022."For all the stars to align in a ballot initiative ...

There's got to be a commitment on the part of providers in Mississippi, I viagra cost think, and we have to also look realistically at the amount of resources that it takes to do a ballot initiative," Mitchell said. "But it is certainly not out viagra cost of reach."Consultations via tablets, laptops and phones linked patients and doctors when society shut down in early spring. Telehealth visits dropped with the reopening, but they're still far more common than before and now there's a push to make them widely available in the future.Permanently expanding access will involve striking a balance between costs and quality, dealing with privacy concerns and potential fraud, and figuring out how telehealth can reach marginalized patients, including people with mental health problems."I don't think it is ever going to replace in-person visits, because sometimes a doctor needs to put hands on a patient," said CMS Administrator Seema Verma, the Trump administration's leading advocate for telehealth.Caveats aside, "it's almost a modern-day house call," she added."It's fair to say that telemedicine was in its infancy prior to the viagra, but it's come of age this year," said Murray Aitken of the data firm IQVIA, which tracks the impact.In the depths of the erectile dysfunction shutdown, telehealth accounted for more than 40% of primary care visits for patients with traditional Medicare, up from a tiny 0.1% sliver before the public health emergency.

As the government's flagship health care program, Medicare covers more than 60 million people, including those age viagra cost 65 and older, and younger disabled people.A recent poll of older adults by the University of Michigan Institute for Healthcare Policy &. Innovation found that more than 7 viagra cost in 10 are interested in using telehealth for follow-ups with their doctor, and nearly 2 out of 3 feel comfortable with video conferences.But privacy was an issue, especially for those who hadn't tried telehealth. The poll found 27% of older adults who had not had a telemedicine visit were concerned about privacy, compared with 17% of those who tried it.Those who tried telehealth weren't completely sold.

About 4 in 5 were concerned the doctor couldn't physically examine them, and 64% worried the quality wasn't as good."After the initial excitement, in the afterglow, patients realize 'I can't get my viagra cost treatment,' or 'You can't see this thing in the back of my throat over the computer,' " said Dr. Gary LeRoy of Dayton, Ohio, a primary care doctor and president of the American Academy of Family Physicians.For Medicare beneficiary Jean Grady of Westford, Vermont, telemedicine was a relief. She needed a checkup required by Medicare viagra cost to continue receiving supplies for her wearable insulin pump.

Being in a high risk group for erectile dysfunction treatment, Grady worried about potential exposure in a doctor's waiting room, and even more about losing her diabetes supplies if she missed Medicare's checkup deadline."I would have had to go back to taking insulin by syringe," she said.Grady prepared for the virtual viagra cost visit by calling her clinician's tech department and downloading teleconference software. She says she would do some future visits by video, but not all. For example, people with diabetes need periodic blood tests, and their feet must be checked for signs of circulatory problems.Still, quite a few follow-ups "could be done very efficiently and be just as useful to the physician and myself as going in and seeing them in person," Grady said.Many private insurance plans, including those in Medicare Advantage, offer some level of telemedicine coverage.But traditional Medicare has restricted it to rural residents, who generally had to travel to specially designated sites to viagra cost connect.Under the erectile dysfunction public health emergency, the administration temporarily waived Medicare's restrictions so enrollees anywhere could use telemedicine.

Patients could connect from home. Making such changes permanent would require legislation from Congress, but viagra cost there's bipartisan interest.Sen. Lamar Alexander, chairman of the Senate Health, Education, Labor and Pensions Committee, says he'd like to see broader access, without breaking the bank."Our job should be to ensure that change is done with the goals of better outcomes and better patient experiences, at a lower cost," said Alexander, R-Tenn.That's a tall order.Payment viagra cost will be a sticky obstacle.

For now, Medicare is paying clinicians on par for virtual and in-person visits."Policymakers seems to be in a rush to pass legislation, but I think it is worth taking a little more time," said Juliette Cubanski, a Medicare expert with the nonpartisan Kaiser Family Foundation. "Fraud is one big area that policymakers need to be cognizant of."Fraud-busters agree.Telehealth is so new viagra cost that "we don't have at this point a real sense of where the huge risks lie," said Andrew VanLandingham, a senior lawyer with the Health and Human Services inspector general's office. "We are sort of in an experimental phase."Despite the risks, advocates see opportunities.Expanded Medicare telehealth could:help move the nation closer to a long-sought goal of treating mental health the same as physical conditions.

Sen. Ron Wyden, D-Ore., wants to use telemedicine as a springboard to improve mental health care. IQVIA data shows 60% of psychiatric consults took place by telehealth during the shutdown.increase access for people living in remote communities, in low-income urban areas and even nursing homes.

Medicare's research shows low-income beneficiaries have had similar patterns of using telehealth for primary care as program enrollees overall.improve coordination of care for people with chronic health conditions, a goal that requires patient and persistent monitoring. Chronic care accounts for most program spending.University of Michigan health policy expert Mark Fendrick says Medicare should figure out what services add value for patients' health and taxpayers' wallets, and pay just for those.Telehealth "was an overnight sensation," said Fendrick. "Hopefully it's not a one-hit wonder."As the wind howled and the rain slammed down, a team of nurses, respiratory therapists and a doctor worked through the night to care for 19 tiny babies as Hurricane Laura slammed southwestern Louisiana.The babies, some on ventilators or eating through a feeding tube, seemed to weather the storm just fine, said Dr.

Juan Bossano, the medical director of the neonatal intensive care unit at Lake Charles Memorial Hospital for Women. "They did very well. They tolerated it very well.

We had a very good day," he said.Laura made landfall early Thursday morning as a Category 4 storm, packing top winds of 150 mph (241 kph), and pushing a storm surge as high as 15 feet in some areas.Hours before it made landfall, officials had to move the babies from the women's hospital to the main hospital in the system after it became clear that storm surge could inundate the women's hospital, located on the southern end of Lake Charles. The hospital has its own generator and hospital administrator Alesha Alford said it was built to withstand hurricane force winds. But in the single story facility, there's no room to move up and storm surge in that area was expected to hit nine feet.

In a roughly two-hour operation the babies in the intensive care unit were transferred by ambulance to Lake Charles Memorial Hospital, a ten-story facility on the northern side of the city. Trucks carried needed equipment such as incubators.Alford said the storm hadn't yet hit but "the skies looked very ominous." She said everyone pitched in to get supplies moved to the other hospital."It went as smooth as could be because we had everyone helping," she said.Alford said three mothers who couldn't be discharged from the women's hospital were also transferred. Two of them had their newborns with them while the child of the third mom was in the intensive care unit.

Parents of the other children in the neonatal intensive care unit couldn't stay with them during the storm because there wasn't enough room so Bossano said one nurse was tasked with calling parents to keep them informed of how their children were doing. Bossano occasionally posted updates on Facebook.Once they got situated at the larger hospital and the winds picked up, Alford said the patients were moved into the hallways. To "protect our babies," mattresses were pushed up against the windows to prevent flying glass although none of the windows ended up breaking.She said as huge gusts of wind started coming in, they could feel the building vibrate.

In addition to Bossano, the medical staff consisted of two neonatal nurse practitioners, 14 nurses and three respiratory therapists who worked on 12-hour shifts. Some of the staff slept on air mattresses in the hallway, Alford said. After making it through the hurricane, the plan was to have the babies stay in Lake Charles.

While electricity was out in the city, the hospital has its own generator. But Alford said the city's water system has been so heavily damaged that it ultimately forced them to transfer the babies as well as other patients to other hospitals around the state Friday.Both Alford and Bossano repeatedly praised the nursing staff for their work in caring for the babies that in some cases were born weighing only a pound or two. Some of the nursing staff lost their houses in the storm, and they were worried about their own families, but they put those concerns aside to care for their tiny patients."Really the nurses and the respiratory therapists are the heroes here," Bosanno said.

"They showed that very clearly the way they performed."There aren’t many hospital visitors amid the erectile dysfunction treatment viagra. But, if you were to walk through intensive-care units at one New York City hospital, you’d see internet-connected speakers—about the size of a stack of Post-it Notes—affixed to the bedrails of some patient beds.It’s part of a project by two Weill Cornell Medicine doctors to help family members speak with ICU patients, often intubated or otherwise not able to hold up a phone themselves, from afar.“The patients could be completely sedated, they could be in a coma,” but families still want to be there with them, said Dr. Marc Schiffman, an interventional radiologist and one of the doctors who spearheaded bringing the devices into ICUs.The speakers, now in 11 units at Weill Cornell, are part of a two-way communication system from company Relay, originally developed as a walkie-talkie system of sorts for children to stay in touch with their parents throughout the day.

Users on one end record snippets of conversation using a mobile app, which are automatically played out loud through the small speaker.Users on the other end push a button on the device to record a response.“Whenever (families) have a story they want to recount, they can just talk into their phone,” Schiffman said. €œIt gives the families a sense of autonomy (and) connection,” even when the patient can’t respond.The effort, dubbed the VoiceLove Project, began about four months ago, at the height of the erectile dysfunction treatment viagra in New York City.Families and other visitors were no longer allowed inside Weill Cornell, but still wanted a way to connect with patients who were sick with erectile dysfunction treatment. Initially, that involved a nurse standing in the ICU and holding up a phone or tablet so families could see the patient—a task that took time out of their already busy day, potentially exposed them to erectile dysfunction treatment and often meant using scarce personal protective equipment.“It really wasn’t a practical solution,” said Dr.

Tamatha Fenster, a minimally invasive gynecologic surgeon.So Fenster and Schiffman began brainstorming hands-free technologies they could install directly at the bedside. Schiffman drove to a local Target store and bought a few Relay walkie-talkie devices. After testing it with families and patients in the ICU, the two decided it was a “grand slam,” Schiffman said.Since March, hospitals have been trying new ways to keep patients connected to families at home, said Bill Flatley, senior service delivery manager at consulting firm OST.

He said he’s mainly seen hospitals repurpose technology usually used for telemedicine, like tablets and cameras mounted on telemedicine carts.It’s likely hospitals will have to continue to restrict visitors, at least as long as there’s uncertainty around erectile dysfunction treatment. So it’s integral for staff to figure out processes that make it easy for families to talk to patients—without putting an additional burden on clinicians or expecting them to serve as tech support.For Fenster and Schiffman, deploying walkie-talkies in the ICU for the first time took some leg work.To scale the walkie-talkie system, Schiffman reached out to Relay’s team via the company’s website, and the company agreed to donate roughly 130 devices and waived the per-user subscription fee. The doctors and Relay have continued to work together on best practices for using the devices in ICUs, a use case Relay is marketing and could sell to other hospitals, according to Jon Schniepp, Relay’s senior vice president of marketing.But Fenster and Schiffman couldn’t just bring walkie-talkies into the ICU.

In the hospital setting, there are additional quality and privacy concerns. To address those, the doctors created a disposable case, which made it easier to keep the device sterile and blocked passersby from accidentally pressing the button that would transmit sounds to a family’s Relay app.The two spent thousands of dollars out of their own pockets to devise the best case design, Fenster said, working with an industrial designer in New Jersey to 3D print different models. The final plastic case, customized with the phrase “VoiceLove” on the front, costs about $10 per case to print and ship.

They’ve started reaching out to acute-care and post-acute facilities in California, Texas and other erectile dysfunction treatment hot spots to explain how the VoiceLove Project works, hoping to connect other groups with Relay and share the case design. But the doctors say they’re still working out the logistics of getting the equipment to interested organizationsWhen Dr. George Wanna saw how devastated St.

George Hospital University Medical Center was by an explosion that shook Beirut, he felt a need to help his hometown. The Aug. 4 blast in the city’s harbor ravaged St.

George’s, so Wanna launched a GoFundMe page to help the hospital, where a good friend of his, Dr. Alexander Nehme, is chief medical officer.At deadline, more than $86,600 had been raised, with a goal of $100,000. €œThis is the first time in their 140-year history when St.

George’s Hospital was damaged so severely that it is unable to function,” said Wanna, chair of the otolaryngology department at New York Eye and Ear Infirmary of Mount Sinai and Mount Sinai Beth Israel in New York. €¨St. George Hospital even remained open during Lebanon’s 15-year civil war, a conflict that wracked Beirut and forced Wanna to spend much of his childhood in bomb shelters.

Wanna is also working with Mount Sinai to send medical supplies. €œSt. George Hospital is in need of everything needed to run a hospital—beds, ventilators, protective equipment.” The tragedy also affected Wanna’s family.

His parents weren’t home when the blast struck and were unharmed. But “my parents’ home was severely damaged by the blast. Sadly, we lost the lives of several of my dad’s relatives,” he said via email.

Wanna, who spent his residency at Mount Sinai, is grateful to the system. €œThey have given me a chance to have the kind of life I could never have hoped for—they helped me build a home and a life in this great country.”.

Viagra what does it do

Maximizing health coverage for DAP http://franklinvideo.com/levitra-street-price/ clients viagra what does it do. Before and after winning the case Outline prepared by Geoffrey Hale and Cathy Roberts - updated August 2012 This outline is intended to assist Disability Advocacy Program (DAP) advocates maximize health insurance coverage for clients they are representing on Social Security/SSI disability determinations. We begin with a discussion of coverage options available while your client’s DAP case is pending and then outline the viagra what does it do effect winning the DAP case can have on your client’s access to health care coverage. How your client is affected will vary depending on the source and amount of disability income he or she receives after the successful appeal.

I. BACKGROUND viagra what does it do. Public health coverage for your clients will primarily be provided by Medicaid and Medicare. The two programs are structured differently and have different eligibility criteria, but in order to provide the most complete coverage possible for your clients, viagra what does it do they must work effectively together.

Understanding their interactions is essential to ensuring benefits for your client. Here is a brief overview of the programs we will cover. A. Medicaid.

Medicaid is the public insurance program jointly funded by the federal, state and local governments for people of limited means. For federal Medicaid law, see 42 U.S.C. § 1396 et seq., 42 C.F.R. § 430 et seq.

Regular Medicaid is described in New York’s State Plan and codified at N.Y. Soc. Serv. L.

§§ 122, 131, 363- 369-1. 18 N.Y.C.R.R. § 360, 505. New York also offers several additional programs to provide health care benefits to those whose income might be too high for Regular Medicaid.

i. Family Health Plus (FHPlus) is an extension of New York’s Medicaid program that provides health coverage for adults who are over-income for regular Medicaid. FHPlus is described in New York’s 1115 waiver and codified at N.Y. Soc.

Child Health Plus (CHPlus) is a sliding scale premium program for children who are over-income for regular Medicaid. CHPlus is codified at N.Y. Pub. Health L.

§2510 et seq. b. Medicare. Medicare is the federal health insurance program providing coverage for the elderly, disabled, and people with end-stage renal disease.

Medicare is codified under title XVIII of the Social Security Law, see 42 U.S.C. § 1395 et seq., 42 C.F.R. § 400 et seq. Medicare is divided into four parts.

i. Part A covers hospital, skilled nursing facility, home health, and hospice care, with some deductibles and coinsurance. Most people are eligible for Part A at no cost. See 42 U.S.C.

Part B provides medical insurance for doctor’s visits and other outpatient medical services. Medicare Part B has significant cost-sharing components. There are monthly premiums (the standard premium in 2012 is $99.90. In addition, there is a $135 annual deductible (which will increase to $155 in 2010) as well as 20% co-insurance for most covered out-patient services.

See 42 U.S.C. § 1395k, 42 C.F.R. Pt. 407.

iii. Part C, also called Medicare Advantage, provides traditional Medicare coverage (Parts A and B) through private managed care insurers. See 42 U.S.C. § 1395w, 42 C.F.R.

Pt. 422. Premium amounts for Medicare Advantage plans vary. Some Medicare Advantage plans include prescription drug coverage.

iv. Part D is an optional prescription drug benefit available to anyone with Medicare Parts A and B. See 42 U.S.C. § 1395w, 42 C.F.R.

§ 423.30(a)(1)(i) and (ii). Unlike Parts A and B, Part D benefits are provided directly through private plans offered by insurance companies. In order to receive prescription drug coverage, a Medicare beneficiary must join a Part D Plan or participate in a Medicare Advantage plan that provides prescription drug coverage. C.

Medicare Savings Programs (MSPs). Funded by the State Medicaid program, MSPs help eligible individuals meet some or all of their cost-sharing obligations under Medicare. See N.Y. Soc.

Serv. L. § 367-a(3)(a), (b), and (d). There are three separate MSPs, each with different eligibility requirements and providing different benefits.

i. Qualified Medicare Beneficiary (QMB). The QMB program provides the most comprehensive benefits. Available to those with incomes at or below 100% of the Federal Poverty Level (FPL), the QMB program covers virtually all Medicare cost-sharing obligations.

Part B premiums, Part A premiums, if there are any, and any and all deductibles and co-insurance. ii. Special Low-Income Medicare Beneficiary (SLMB). For those with incomes between 100% and 120% FPL, the SLMB program will cover Part B premiums only.

iii. Qualified Individual (QI-1). For those with incomes between 120% and 135% FPL, but not otherwise Medicaid eligible, the QI-1 program covers Medicare Part B premiums. D.

Medicare Part D Low Income Subsidy (LIS or “Extra Help”). LIS is a federal subsidy administered by CMS that helps Medicare beneficiaries with limited income and/or resources pay for some or most of the costs of Medicare prescription drug coverage. See 42 C.F.R. § 423.773.

Some of the costs covered in full or in part by LIS include the monthly premiums, annual deductible, co-payments, and the coverage gap. Individuals eligible for Medicaid, SSI, or MSP are deemed eligible for full LIS benefitsSee 42 C.F.R. § 423.773(c). LIS applications are treated as (“deemed”) applications for MSP benefits, See the Medicare Improvements for Patients and Providers Act (MIPPA) of 2008, Pub.

Law 110-275. II. WHILE THE DAP APPEAL IS PENDING Does your client have health insurance?. If not, why isn’t s/he getting Medicaid, Family Health Plus or Child Health Plus?.

There have been many recent changes which expand eligibility and streamline the application process. All/most of your DAP clients should qualify. Significant changes to Medicaid include. Elimination of the resource test for certain categories of Medicaid applicants/recipients and all applicants to the Family Health Plus program.

§369-ee (2), as amended by L. 2009, c. 58, pt. C, § 59-d.

As of October 1, 2009, a resource test is no longer required for these categories. Elimination of the fingerprinting requirement. N.Y. Soc.

Serv. L. §369-ee, as amended by L. 2009, c.

58, pt. C, § 62. Elimination of the waiting period for CHPlus. N.Y.

Pub. Health L. §2511, as amended by L. 2008, c.

58. Elimination of the face-to-face interview requirement for Medicaid, effective April 1, 2010. N.Y. Soc.

Serv. L. §366-a (1), as amended by L. 2009, c.

58, pt. C, § 60. Higher income levels for Single Adults and Childless Couples. N.Y.

Soc. Serv. L. §366(1)(a)(1),(8) as amended by L.

Higher income levels for Medicaid’s Medically Needy program. N.Y. Soc. Serv.

L. §366(2)(a)(7) as amended by L. 2008, c. 58.

See also. GIS 08 MA/022 More detailed information on recent changes to Medicaid is available at. III. AFTER CLIENT IS AWARDED DAP BENEFITS a.

Medicaid eligibility. Clients receiving even $1.00 of SSI should qualify for Medicaid automatically. The process for qualifying will differ, however, depending on the source of payment. 1.

Clients Receiving SSI Only. i. These clients are eligible for full Medicaid without a spend-down. See N.Y.

ii. Medicaid coverage is automatic. No separate application/ recertification required. iii.

Most SSI-only recipients are required to participate in Medicaid managed care. See N.Y. Soc. Serv.

L. §364-j. 2. Concurrent (SSI/SSD) cases.

Eligible for full Medicaid since receiving SSI. See N.Y. Soc. Serv.

I. They can still qualify for Medicaid but may have a spend-down. Federal Law allows states to use a “spend-down” to extend Medicaid to “medically needy” persons in the federal mandatory categories (children, caretakers, elderly and disabled people) whose income or resources are above the eligibility level for regular Medicaid. See 42 U.S.C.

§ 1396 (a) (10) (ii) (XIII). ii. Under spend-down, applicants in New York’s Medically Needy program can qualify for Medicaid once their income/resources, minus incurred medical expenses, fall below the specified level. For an explanation of spend-down, see 96 ADM 15.

B. Family Health Plus Until your client qualifies for Medicare, those over-income for Medicaid may qualify for Family Health Plus without needing to satisfy a spend-down. It covers adults without children with income up to 100% of the FPL and adults with children up to 150% of the FPL.[1] The eligibility tests are the same as for regular Medicaid with two additional requirements. Applicants must be between the ages of 19 and 64 and they generally must be uninsured.

§ 369-ee et. Seq. Once your client begins to receive Medicare, he or she will not be eligible for FHP, because FHP is generally only available to those without insurance. For more information on FHP see our article on Family Health Plus.

IV. LOOMING ISSUES - MEDICARE ELIGIBILITY (WHETHER YOU LIKE IT OR NOT) a. SSI-only cases Clients receiving only SSI aren’t eligible for Medicare until they turn 65, unless they also have End Stage Renal Disease. B.

Concurrent (SSD and SSI) cases 1. Medicare eligibility kicks in beginning with 25th month of SSD receipt. See 42 U.S.C. § 426(f).

Exception. In 2000, Congress eliminated the 24-month waiting period for people diagnosed with ALS (Lou Gehrig’s Disease.) See 42 U.S.C. § 426 (h) 2. Enrollment in Medicare is a condition of eligibility for Medicaid coverage.

These clients cannot decline Medicare coverage. (05 OMM/ADM 5. Medicaid Reference Guide p. 344.1) 3.

Medicare coverage is not free. Although most individuals receive Part A without any premium, Part B has monthly premiums and significant cost-sharing components. 4. Medicaid and/or the Medicare Savings Program (MSP) should pick up most of Medicare’s cost sharing.

Most SSI beneficiaries are eligible not only for full Medicaid, but also for the most comprehensive MSP, the Qualified Medicare Beneficiary (QMB) program. I. Parts A &. B (hospital and outpatient/doctors visits).

A. Medicaid will pick up premiums, deductibles, co-pays. N.Y. Soc.

Serv. L. § 367-a (3) (a). For those not enrolled in an MSP, SSA normally deducts the Part B premium directly from the monthly check.

However, SSI recipients are supposed to be enrolled automatically in QMB, and Medicaid is responsible for covering the premiums. Part B premiums should never be deducted from these clients’ checks.[1] Medicaid and QMB-only recipients should NEVER be billed directly for Part A or B services. Even non-Medicaid providers are supposed to be able to bill Medicaid directly for services.[2] Clients are only responsible for Medicaid co-pay amount. See 42 U.S.C.

§ 1396a (n) ii. Part D (prescription drugs). a. Clients enrolled in Medicaid and/or MSP are deemed eligible for Low Income Subsidy (LIS aka Extra Help).

See 42 C.F.R. § 423.773(c). SSA POMS SI § 01715.005A.5. New York State If client doesn’t enroll in Part D plan on his/her own, s/he will be automatically assigned to a benchmark[3] plan.

See 42 C.F.R. § 423.34 (d). LIS will pick up most of cost-sharing.[3] Because your clients are eligible for full LIS, they should have NO deductible and NO premium if they are in a benchmark plan, and will not be subject to the coverage gap (aka “donut hole”). See 42 C.F.R.

§§ 423.780 and 423.782. The full LIS beneficiary will also have co-pays limited to either $1.10 or $3.30 (2010 amounts). See 42 C.F.R. § 423.104 (d) (5) (A).

Other important points to remember. - Medicaid co-pay rules do not apply to Part D drugs. - Your client’s plan may not cover all his/her drugs. - You can help your clients find the plan that best suits their needs.

To figure out what the best Part D plans are best for your particular client, go to www.medicare.gov. Click on “formulary finder” and plug in your client’s medication list. You can enroll in a Part D plan through www.medicare.gov, or by contacting the plan directly. €“ Your clients can switch plans at any time during the year.

Iii. Part C (“Medicare Advantage”). a. Medicare Advantage plans provide traditional Medicare coverage (Parts A and B) through private managed care insurers.

See 42 U.S.C. § 1395w, 42 C.F.R. Pt. 422.

Medicare Advantage participation is voluntary. For those clients enrolled in Medicare Advantage Plans, the QMB cost sharing obligations are the same as they are under traditional Medicare. Medicaid must cover any premiums required by the plan, up to the Part B premium amount. Medicaid must also cover any co-payments and co-insurance under the plan.

As with traditional Medicare, both providers and plans are prohibited from billing the beneficiary directly for these co-payments. C. SSD only individuals. 1.

Same Medicare eligibility criteria (24 month waiting period, except for persons w/ ALS). I. During the 24 month waiting period, explore eligibility for Medicaid or Family Health Plus. 2.

Once Medicare eligibility begins. ii. Parts A &. B.

SSA will automatically enroll your client. Part B premiums will be deducted from monthly Social Security benefits. (Part A will be free – no monthly premium) Clients have the right to decline ongoing Part B coverage, BUT this is almost never a good idea, and can cause all sorts of headaches if client ever wants to enroll in Part B in the future. (late enrollment penalty and can’t enroll outside of annual enrollment period, unless person is eligible for Medicare Savings Program – see more below) Clients can decline “retro” Part B coverage with no penalty on the Medicare side – just make sure they don’t actually need the coverage.

Risky to decline if they had other coverage during the retro period – their other coverage may require that Medicare be utilized if available. Part A and Part B also have deductibles and co-pays. Medicaid and/or the MSPs can help cover this cost sharing. iii.

Part D. Client must affirmatively enroll in Part D, unless they receive LIS. See 42 U.S.C. § 1395w-101 (b) (2), 42 C.F.R.

§ 423.38 (a). Enrollment is done through individual private plans. LIS recipients will be auto-assigned to a Part D benchmark plan if they have not selected a plan on their own. Client can decline Part D coverage with no penalty if s/he has “comparable coverage.” 42 C.F.R.

§ 423.34 (d) (3) (i). If no comparable coverage, person faces possible late enrollment penalty &. Limited enrollment periods. 42 C.F.R.

§ 423.46. However, clients receiving LIS do not incur any late enrollment penalty. 42 C.F.R. § 423.780 (e).

Part D has a substantial cost-sharing component – deductibles, premiums and co-pays which vary from plan to plan. There is also the coverage gap, also known as “donut hole,” which can leave beneficiaries picking up 100% of the cost of their drugs until/unless a catastrophic spending limit is reached. The LIS program can help with Part D cost-sharing. Use Medicare’s website to figure out what plan is best for your client.

(Go to www.medicare.gov , click on “formulary finder” and plug in your client’s medication list. ) You can also enroll in a Part D plan directly through www.medicare.gov. Iii. Help with Medicare cost-sharing a.

Medicaid – After eligibility for Medicare starts, client may still be eligible for Medicaid, with or without a spend-down. There are lots of ways to help clients meet their spend-down – including - Medicare cost sharing amounts (deductibles, premiums, co-pays) - over the counter medications if prescribed by a doctor. - expenses paid by state-funded programs like EPIC and ADAP. - medical bills of person’s spouse or child.

- health insurance premiums. - joining a pooled Supplemental Needs Trust (SNT). B. Medicare Savings Program (MSP) – If client is not eligible for Medicaid, explore eligibility for Medicare Savings Program (MSP).

MSP pays for Part B premiums and gets you into the Part D LIS. There are no asset limits in the Medicare Savings Program. One of the MSPs (QMB), also covers all cost sharing for Parts A &. B.

If your client is eligible for Medicaid AND MSP, enrolling in MSP may subject him/her to, or increase a spend-down, because Medicaid and the various MSPs have different income eligibility levels. It is the client’s choice as to whether or not to be enrolled into MSP. C. Part D Low Income Subsidy (LIS) – If your client is not eligible for MSP or Medicaid, s/he may still be eligible for Part D Low Income Subsidy.

Applications for LIS are also be treated as applications for MSP, unless the client affirmatively indicates that s/he does not want to apply for MSP. d. Medicare supplemental insurance (Medigap) -- Medigap is supplemental private insurance coverage that covers all or some of the deductibles and coinsurance for Medicare Parts A and B. Medigap is not available to people enrolled in Part C.

E. Medicare Advantage – Medicare Advantage plans “package” Medicare (Part A and B) benefits, with or without Part D coverage, through a private health insurance plan. The cost-sharing structure (deductible, premium, co-pays) varies from plan to plan. For a list of Medicare Advantage plans in your area, go to www.medicare.gov – click on “find health plans.” f.

NY Prescription Saver Card -- NYP$ is a state-sponsored pharmacy discount card that can lower the cost of prescriptions by as much as 60 percent on generics and 30 percent on brand name drugs. Can be used during the Part D “donut hole” (coverage gap) g. For clients living with HIV. ADAP [AIDS Drug Assistance Program] ADAP provides free medications for the treatment of HIV/AIDS and opportunistic s.

ADAP can be used to help meet a Medicaid spenddown and get into the Part D Low Income subsidy. For more information about ADAP, go to V. GETTING MEDICAID IN THE DISABLED CATEGORY AFTER AN SSI/SSDI DENIAL What if your client's application for SSI or SSDI is denied based on SSA's finding that they were not "disabled?. " Obviously, you have your appeals work cut out for you, but in the meantime, what can they do about health insurance?.

It is still possible to have Medicaid make a separate disability determination that is not controlled by the unfavorable SSA determination in certain situations. Specifically, an applicant is entitled to a new disability determination where he/she. alleges a different or additional disabling condition than that considered by SSA in making its determination. Or alleges less than 12 months after the most recent unfavorable SSA disability determination that his/her condition has changed or deteriorated, alleges a new period of disability which meets the duration requirement, and SSA has refused to reopen or reconsider the allegations, or the individual is now ineligible for SSA benefits for a non-medical reason.

Or alleges more than 12 months after the most recent unfavorable SSA disability determination that his/her condition has changed or deteriorated since the SSA determination and alleges a new period of disability which meets the duration requirement, and has not applied to SSA regarding these allegations. See GIS 10-MA-014 and 08 OHIP/INF-03.[4] [1] Potential wrinkle – for some clients Medicaid is not automatically pick up cost-sharing. In Monroe County we have had several cases where SSA began deducting Medicare Part B premiums from the checks of clients who were receiving SSI and Medicaid and then qualified for Medicare. The process should be automatic.

Please contact Geoffrey Hale in our Rochester office if you encounter any cases like this. [2]Under terms established to provide benefits for QMBs, a provider agreement necessary for reimbursement “may be executed through the submission of a claim to the Medicaid agency requesting Medicaid payment for Medicare deductibles and coinsurance for QMBs.” CMS State Medicaid Manual, Chapter 3, Eligibility, 3490.14 (b), available at. http://www.cms.hhs.gov/Manuals/PBM/itemdetail.asp?. ItemID=CMS021927.

[3]Benchmark plans are free if you are an LIS recipient. The amount of the benchmark changes from year to year. In 2013, a Part D plan in New York State is considered benchmark if it provides basic Part D coverage and its monthly premium is $43.22 or less. [4] These citations courtesy of Jim Murphy at Legal Services of Central New York.

This site provides general information only. This is not legal advice. You can only obtain legal advice from a lawyer. In addition, your use of this site does not create an attorney-client relationship.

To contact a lawyer, visit http://lawhelp.org/ny. We make every effort to keep these materials and links up-to-date and in accordance with New York City, New York state and federal law. However, we do not guarantee the accuracy of this information.Some "dual eligible" beneficiaries (people who have Medicare and Medicaid) are entitled to receive reimbursement of their Medicare Part B premiums from New York State through the Medicare Insurance Premium Payment Program (MIPP). The Part B premium is $148.50 in 2021.

MIPP is for some groups who are either not eligible for -- or who are not yet enrolled in-- the Medicare Savings Program (MSP), which is the main program that pays the Medicare Part B premium for low-income people. Some people are not eligible for an MSP even though they have full Medicaid with no spend down. This is because they are in a special Medicaid eligibility category -- discussed below -- with Medicaid income limits that are actually HIGHER than the MSP income limits. MIPP reimburses them for their Part B premium because they have “full Medicaid” (no spend down) but are ineligible for MSP because their income is above the MSP SLIMB level (120% of the Federal Poverty Level (FPL).

Even if their income is under the QI-1 MSP level (135% FPL), someone cannot have both QI-1 and Medicaid). Instead, these consumers can have their Part B premium reimbursed through the MIPP program. In this article. The MIPP program was established because the State determined that those who have full Medicaid and Medicare Part B should be reimbursed for their Part B premium, even if they do not qualify for MSP, because Medicare is considered cost effective third party health insurance, and because consumers must enroll in Medicare as a condition of eligibility for Medicaid (See 89 ADM 7).

There are generally four groups of dual-eligible consumers that are eligible for MIPP. Therefore, many MBI WPD consumers have incomes higher than what MSP normally allows, but still have full Medicaid with no spend down. Those consumers can qualify for MIPP and have their Part B premiums reimbursed. Here is an example.

Sam is age 50 and has Medicare and MBI-WPD. She gets $1500/mo gross from Social Security Disability and also makes $400/month through work activity. $ 167.50 -- EARNED INCOME - Because she is disabled, the DAB earned income disregard applies. $400 - $65 = $335.

Her countable earned income is 1/2 of $335 = $167.50 + $1500.00 -- UNEARNED INCOME from Social Security Disability = $1,667.50 --TOTAL income. This is above the SLIMB limit of $1,288 (2021) but she can still qualify for MIPP. 2. Parent/Caretaker Relatives with MAGI-like Budgeting - Including Medicare Beneficiaries.

Consumers who fall into the DAB category (Age 65+/Disabled/Blind) and would otherwise be budgeted with non-MAGI rules can opt to use Affordable Care Act MAGI rules if they are the parent/caretaker of a child under age 18 or under age 19 and in school full time. This is referred to as “MAGI-like budgeting.” Under MAGI rules income can be up to 138% of the FPL—again, higher than the limit for DAB budgeting, which is equivalent to only 83% FPL. MAGI-like consumers can be enrolled in either MSP or MIPP, depending on if their income is higher or lower than 120% of the FPL. If their income is under 120% FPL, they are eligible for MSP as a SLIMB.

If income is above 120% FPL, then they can enroll in MIPP. (See GIS 18 MA/001 - 2018 Medicaid Managed Care Transition for Enrollees Gaining Medicare, #4) When a consumer has Medicaid through the New York State of Health (NYSoH) Marketplace and then enrolls in Medicare when she turns age 65 or because she received Social Security Disability for 24 months, her Medicaid case is normally** transferred to the local department of social services (LDSS)(HRA in NYC) to be rebudgeted under non-MAGI budgeting. During the transition process, she should be reimbursed for the Part B premiums via MIPP. However, the transition time can vary based on age.

AGE 65+ Those who enroll in Medicare at age 65+ will receive a letter from their local district asking them to "renew" Medicaid through their local district. See 2014 LCM-02. The Medicaid case takes about four months to be rebudgeted and approved by the LDSS. The consumer is entitled to MIPP payments for at least three months during the transition.

Once the case is with the LDSS she should automatically be re-evaluated for MSP, even if the LDSS determines the consumer is not eligible for Medicaid because of excess income or assets. 08 OHIP/ADM-4. Consumers UNDER 65 who receive Medicare due to disability status are entitled to keep MAGI Medicaid through NYSoH for up to 12 months (also known as continuous coverage, See NY Social Services Law 366, subd. 4(c).

These consumers should receive MIPP payments for as long as their cases remain with NYSoH and throughout the transition to the LDSS. NOTE during erectile dysfunction treatment emergency their case may remain with NYSoH for more than 12 months. See here. EXAMPLE.

Sam, age 60, was last authorized for Medicaid on the Marketplace in June 2020. He became enrolled in Medicare based on disability in August 2020, and started receiving Social Security in the same month (he won a hearing approving Social Security disability benefits retroactively, after first being denied disability). Even though his Social Security is too high, he can keep Medicaid for 12 months beginning June 2020. Sam has to pay for his Part B premium - it is deducted from his Social Security check.

He may call the Marketplace and request a refund. This will continue until the end of his 12 months of continuous MAGI Medicaid eligibility. He will be reimbursed regardless of whether he is in a Medicaid managed care plan. See GIS 18 MA/001 Medicaid Managed Care Transition for Enrollees Gaining Medicare (PDF) When that ends, he will renew Medicaid and apply for MSP with his local district.

See GIS 18 MA/001 - 2018 Medicaid Managed Care Transition for Enrollees Gaining Medicare, #4 for an explanation of this process. That directive also clarified that reimbursement of the Part B premium will be made regardless of whether the individual is still in a Medicaid managed care (MMC) plan. Note. During the erectile dysfunction treatment emergency, those who have Medicaid through the NYSOH marketplace and enroll in Medicare should NOT have their cases transitioned to the LDSS.

They should keep the same MAGI budgeting and automatically receive MIPP payments. See GIS 20 MA/04 or this article on erectile dysfunction treatment eligibility changes 4. Those with Special Budgeting after Losing SSI (DAC, Pickle, 1619b) Disabled Adult Child (DAC). Special budgeting is available to those who are 18+ and lose SSI because they begin receiving Disabled Adult Child (DAC) benefits (or receive an increase in the amount of their benefit).

Consumer must have become disabled or blind before age 22 to receive the benefit. If the new DAC benefit amount was disregarded and the consumer would otherwise be eligible for SSI, they can keep Medicaid eligibility with NO SPEND DOWN. See this article. Consumers may have income higher than MSP limits, but keep full Medicaid with no spend down.

Therefore, they are eligible for payment of their Part B premiums. See page 96 of the Medicaid Reference Guide (Categorical Factors). If their income is lower than the MSP SLIMB threshold, they can be added to MSP. If higher than the threshold, they can be reimbursed via MIPP.

See also 95-ADM-11. Medical Assistance Eligibility for Disabled Adult Children, Section C (pg 8). Pickle &. 1619B.

5. When the Part B Premium Reduces Countable Income to Below the Medicaid Limit Since the Part B premium can be used as a deduction from gross income, it may reduce someone's countable income to below the Medicaid limit. The consumer should be paid the difference to bring her up to the Medicaid level ($904/month in 2021). They will only be reimbursed for the difference between their countable income and $904, not necessarily the full amount of the premium.

See GIS 02-MA-019. Reimbursement of Health Insurance Premiums MIPP and MSP are similar in that they both pay for the Medicare Part B premium, but there are some key differences. MIPP structures the payments as reimbursement -- beneficiaries must continue to pay their premium (via a monthly deduction from their Social Security check or quarterly billing, if they do not receive Social Security) and then are reimbursed via check. In contrast, MSP enrollees are not charged for their premium.

Their Social Security check usually increases because the Part B premium is no longer withheld from their check. MIPP only provides reimbursement for Part B. It does not have any of the other benefits MSPs can provide, such as. A consumer cannot have MIPP without also having Medicaid, whereas MSP enrollees can have MSP only.

Of the above benefits, Medicaid also provides Part D Extra Help automatic eligibility. There is no application process for MIPP because consumers should be screened and enrolled automatically (00 OMM/ADM-7). Either the state or the LDSS is responsible for screening &. Distributing MIPP payments, depending on where the Medicaid case is held and administered (14 /2014 LCM-02 Section V).

If a consumer is eligible for MIPP and is not receiving it, they should contact whichever agency holds their case and request enrollment. Unfortunately, since there is no formal process for applying, it may require some advocacy. If Medicaid case is at New York State of Health they should call 1-855-355-5777. Consumers will likely have to ask for a supervisor in order to find someone familiar with MIPP.

If Medicaid case is with HRA in New York City, they should email mipp@hra.nyc.gov. If Medicaid case is with other local districts in NYS, call your local county DSS. See more here about consumers who have Medicaid on NYSofHealth who then enroll in Medicare - how they access MIPP. Once enrolled, it make take a few months for payments to begin.

Payments will be made in the form of checks from the Computer Sciences Corporation (CSC), the fiscal agent for the New York State Medicaid program. The check itself comes attached to a remittance notice from Medicaid Management Information Systems (MMIS). Unfortunately, the notice is not consumer-friendly and may be confusing. See attached sample for what to look for.

Health Insurance Premium Payment Program (HIPP) HIPP is a sister program to MIPP and will reimburse consumers for private third party health insurance when deemed “cost effective.” Directives:.

Maximizing health coverage viagra cost Levitra street price for DAP clients. Before and after winning the case Outline prepared by Geoffrey Hale and Cathy Roberts - updated August 2012 This outline is intended to assist Disability Advocacy Program (DAP) advocates maximize health insurance coverage for clients they are representing on Social Security/SSI disability determinations. We begin with a discussion of viagra cost coverage options available while your client’s DAP case is pending and then outline the effect winning the DAP case can have on your client’s access to health care coverage.

How your client is affected will vary depending on the source and amount of disability income he or she receives after the successful appeal. I. BACKGROUND viagra cost.

Public health coverage for your clients will primarily be provided by Medicaid and Medicare. The viagra cost two programs are structured differently and have different eligibility criteria, but in order to provide the most complete coverage possible for your clients, they must work effectively together. Understanding their interactions is essential to ensuring benefits for your client.

Here is a brief overview of the programs we will cover. A. Medicaid.

Medicaid is the public insurance program jointly funded by the federal, state and local governments for people of limited means. For federal Medicaid law, see 42 U.S.C. § 1396 et seq., 42 C.F.R.

§ 430 et seq. Regular Medicaid is described in New York’s State Plan and codified at N.Y. Soc.

18 N.Y.C.R.R. § 360, 505. New York also offers several additional programs to provide health care benefits to those whose income might be too high for Regular Medicaid.

i. Family Health Plus (FHPlus) is an extension of New York’s Medicaid program that provides health coverage for adults who are over-income for regular Medicaid. FHPlus is described in New York’s 1115 waiver and codified at N.Y.

§369-ee. ii. Child Health Plus (CHPlus) is a sliding scale premium program for children who are over-income for regular Medicaid.

Medicare is the federal health insurance program providing coverage for the elderly, disabled, and people with end-stage renal disease. Medicare is codified under title XVIII of the Social Security Law, see 42 U.S.C. § 1395 et seq., 42 C.F.R.

§ 400 et seq. Medicare is divided into four parts. i.

Part A covers hospital, skilled nursing facility, home health, and hospice care, with some deductibles and coinsurance. Most people are eligible for Part A at no cost. See 42 U.S.C.

ii. Part B provides medical insurance for doctor’s visits and other outpatient medical services. Medicare Part B has significant cost-sharing components.

There are monthly premiums (the standard premium in 2012 is $99.90. In addition, there is a $135 annual deductible (which will increase to $155 in 2010) as well as 20% co-insurance for most covered out-patient services. See 42 U.S.C.

iii. Part C, also called Medicare Advantage, provides traditional Medicare coverage (Parts A and B) through private managed care insurers. See 42 U.S.C.

Premium amounts for Medicare Advantage plans vary. Some Medicare Advantage plans include prescription drug coverage. iv.

Part D is an optional prescription drug benefit available to anyone with Medicare Parts A and B. See 42 U.S.C. § 1395w, 42 C.F.R.

§ 423.30(a)(1)(i) and (ii). Unlike Parts A and B, Part D benefits are provided directly through private plans offered by insurance companies. In order to receive prescription drug coverage, a Medicare beneficiary must join a Part D Plan or participate in a Medicare Advantage plan that provides prescription drug coverage.

C. Medicare Savings Programs (MSPs). Funded by the State Medicaid program, MSPs help eligible individuals meet some or all of their cost-sharing obligations under Medicare.

L. § 367-a(3)(a), (b), and (d). There are three separate MSPs, each with different eligibility requirements and providing different benefits.

i. Qualified Medicare Beneficiary (QMB). The QMB program provides the most comprehensive benefits.

Available to those with incomes at or below 100% of the Federal Poverty Level (FPL), the QMB program covers virtually all Medicare cost-sharing obligations. Part B premiums, Part A premiums, if there are any, and any and all deductibles and co-insurance. ii.

Special Low-Income Medicare Beneficiary (SLMB). For those with incomes between 100% and 120% FPL, the SLMB program will cover Part B premiums only. iii.

Qualified Individual (QI-1). For those with incomes between 120% and 135% FPL, but not otherwise Medicaid eligible, the QI-1 program covers Medicare Part B premiums. D.

Medicare Part D Low Income Subsidy (LIS or “Extra Help”). LIS is a federal subsidy administered by CMS that helps Medicare beneficiaries with limited income and/or resources pay for some or most of the costs of Medicare prescription drug coverage. See 42 C.F.R.

§ 423.773. Some of the costs covered in full or in part by LIS include the monthly premiums, annual deductible, co-payments, and the coverage gap. Individuals eligible for Medicaid, SSI, or MSP are deemed eligible for full LIS benefitsSee 42 C.F.R.

§ 423.773(c). LIS applications are treated as (“deemed”) applications for MSP benefits, See the Medicare Improvements for Patients and Providers Act (MIPPA) of 2008, Pub. Law 110-275.

II. WHILE THE DAP APPEAL IS PENDING Does your client have health insurance?. If not, why isn’t s/he getting Medicaid, Family Health Plus or Child Health Plus?.

There have been many recent changes which expand eligibility and streamline the application process. All/most of your DAP clients should qualify. Significant changes to Medicaid include.

Elimination of the resource test for certain categories of Medicaid applicants/recipients and all applicants to the Family Health Plus program. N.Y. Soc.

As of October 1, 2009, a resource test is no longer required for these categories. Elimination of the fingerprinting requirement. N.Y.

§369-ee, as amended by L. 2009, c. 58, pt.

C, § 62. Elimination of the waiting period for CHPlus. N.Y.

2008, c. 58. Elimination of the face-to-face interview requirement for Medicaid, effective April 1, 2010.

58, pt. C, § 60. Higher income levels for Single Adults and Childless Couples.

L. §366(1)(a)(1),(8) as amended by L. 2008, c.

Higher income levels for Medicaid’s Medically Needy program. N.Y. Soc.

GIS 08 MA/022 More detailed information on recent changes to Medicaid is available at. III. AFTER CLIENT IS AWARDED DAP BENEFITS a.

Medicaid eligibility. Clients receiving even $1.00 of SSI should qualify for Medicaid automatically. The process for qualifying will differ, however, depending on the source of payment.

These clients are eligible for full Medicaid without a spend-down. See N.Y. Soc.

ii. Medicaid coverage is automatic. No separate application/ recertification required.

iii. Most SSI-only recipients are required to participate in Medicaid managed care. See N.Y.

Eligible for full Medicaid since receiving SSI. See N.Y. Soc.

They can still qualify for Medicaid but may have a spend-down. Federal Law allows states to use a “spend-down” to extend Medicaid to “medically needy” persons in the federal mandatory categories (children, caretakers, elderly and disabled people) whose income or resources are above the eligibility level for regular Medicaid. See 42 U.S.C.

§ 1396 (a) (10) (ii) (XIII). ii. Under spend-down, applicants in New York’s Medically Needy program can qualify for Medicaid once their income/resources, minus incurred medical expenses, fall below the specified level.

For an explanation of spend-down, see 96 ADM 15. B. Family Health Plus Until your client qualifies for Medicare, those over-income for Medicaid may qualify for Family Health Plus without needing to satisfy a spend-down.

It covers adults without children with income up to 100% of the FPL and adults with children up to 150% of the FPL.[1] The eligibility tests are the same as for regular Medicaid with two additional requirements. Applicants must be between the ages of 19 and 64 and they generally must be uninsured. See N.Y.

§ 369-ee et. Seq. Once your client begins to receive Medicare, he or she will not be eligible for FHP, because FHP is generally only available to those without insurance.

For more information on FHP see our article on Family Health Plus. IV. LOOMING ISSUES - MEDICARE ELIGIBILITY (WHETHER YOU LIKE IT OR NOT) a.

SSI-only cases Clients receiving only SSI aren’t eligible for Medicare until they turn 65, unless they also have End Stage Renal Disease. B. Concurrent (SSD and SSI) cases 1.

Medicare eligibility kicks in beginning with 25th month of SSD receipt. See 42 U.S.C. § 426(f).

Exception. In 2000, Congress eliminated the 24-month waiting period for people diagnosed with ALS (Lou Gehrig’s Disease.) See 42 U.S.C. § 426 (h) 2.

Enrollment in Medicare is a condition of eligibility for Medicaid coverage. These clients cannot decline Medicare coverage. (05 OMM/ADM 5.

Medicaid Reference Guide p. 344.1) 3. Medicare coverage is not free.

Although most individuals receive Part A without any premium, Part B has monthly premiums and significant cost-sharing components. 4. Medicaid and/or the Medicare Savings Program (MSP) should pick up most of Medicare’s cost sharing.

Most SSI beneficiaries are eligible not only for full Medicaid, but also for the most comprehensive MSP, the Qualified Medicare Beneficiary (QMB) program. I. Parts A &.

B (hospital and outpatient/doctors visits). A. Medicaid will pick up premiums, deductibles, co-pays.

L. § 367-a (3) (a). For those not enrolled in an MSP, SSA normally deducts the Part B premium directly from the monthly check.

However, SSI recipients are supposed to be enrolled automatically in QMB, and Medicaid is responsible for covering the premiums. Part B premiums should never be deducted from these clients’ checks.[1] Medicaid and QMB-only recipients should NEVER be billed directly for Part A or B services. Even non-Medicaid providers are supposed to be able to bill Medicaid directly for services.[2] Clients are only responsible for Medicaid co-pay amount.

See 42 U.S.C. § 1396a (n) ii. Part D (prescription drugs).

a. Clients enrolled in Medicaid and/or MSP are deemed eligible for Low Income Subsidy (LIS aka Extra Help). See 42 C.F.R.

§ 423.773(c). SSA POMS SI § 01715.005A.5. New York State If client doesn’t enroll in Part D plan on his/her own, s/he will be automatically assigned to a benchmark[3] plan.

See 42 C.F.R. § 423.34 (d). LIS will pick up most of cost-sharing.[3] Because your clients are eligible for full LIS, they should have NO deductible and NO premium if they are in a benchmark plan, and will not be subject to the coverage gap (aka “donut hole”).

See 42 C.F.R. §§ 423.780 and 423.782. The full LIS beneficiary will also have co-pays limited to either $1.10 or $3.30 (2010 amounts).

See 42 C.F.R. § 423.104 (d) (5) (A). Other important points to remember.

- Medicaid co-pay rules do not apply to Part D drugs. - Your client’s plan may not cover all his/her drugs. - You can help your clients find the plan that best suits their needs.

To figure out what the best Part D plans are best for your particular client, go to www.medicare.gov. Click on “formulary finder” and plug in your client’s medication list. You can enroll in a Part D plan through www.medicare.gov, or by contacting the plan directly.

€“ Your clients can switch plans at any time during the year. Iii. Part C (“Medicare Advantage”).

a. Medicare Advantage plans provide traditional Medicare coverage (Parts A and B) through private managed care insurers. See 42 U.S.C.

Medicare Advantage participation is voluntary. For those clients enrolled in Medicare Advantage Plans, the QMB cost sharing obligations are the same as they are under traditional Medicare. Medicaid must cover any premiums required by the plan, up to the Part B premium amount.

Medicaid must also cover any co-payments and co-insurance under the plan. As with traditional Medicare, both providers and plans are prohibited from billing the beneficiary directly for these co-payments. C.

SSD only individuals. 1. Same Medicare eligibility criteria (24 month waiting period, except for persons w/ ALS).

I. During the 24 month waiting period, explore eligibility for Medicaid or Family Health Plus. 2.

Once Medicare eligibility begins. ii. Parts A &.

B. SSA will automatically enroll your client. Part B premiums will be deducted from monthly Social Security benefits.

(Part A will be free – no monthly premium) Clients have the right to decline ongoing Part B coverage, BUT this is almost never a good idea, and can cause all sorts of headaches if client ever wants to enroll in Part B in the future. (late enrollment penalty and can’t enroll outside of annual enrollment period, unless person is eligible for Medicare Savings Program – see more below) Clients can decline “retro” Part B coverage with no penalty on the Medicare side – just make sure they don’t actually need the coverage. Risky to decline if they had other coverage during the retro period – their other coverage may require that Medicare be utilized if available.

Part A and Part B also have deductibles and co-pays. Medicaid and/or the MSPs can help cover this cost sharing. iii.

Part D. Client must affirmatively enroll in Part D, unless they receive LIS. See 42 U.S.C.

§ 1395w-101 (b) (2), 42 C.F.R. § 423.38 (a). Enrollment is done through individual private plans.

LIS recipients will be auto-assigned to a Part D benchmark plan if they have not selected a plan on their own. Client can decline Part D coverage with no penalty if s/he has “comparable coverage.” 42 C.F.R. § 423.34 (d) (3) (i).

If no comparable coverage, person faces possible late enrollment penalty &. Limited enrollment periods. 42 C.F.R.

§ 423.46. However, clients receiving LIS do not incur any late enrollment penalty. 42 C.F.R.

§ 423.780 (e). Part D has a substantial cost-sharing component – deductibles, premiums and co-pays which vary from plan to plan. There is also the coverage gap, also known as “donut hole,” which can leave beneficiaries picking up 100% of the cost of their drugs until/unless a catastrophic spending limit is reached.

The LIS program can help with Part D cost-sharing. Use Medicare’s website to figure out what plan is best for your client. (Go to www.medicare.gov , click on “formulary finder” and plug in your client’s medication list.

) You can also enroll in a Part D plan directly through www.medicare.gov. Iii. Help with Medicare cost-sharing a.

Medicaid – After eligibility for Medicare starts, client may still be eligible for Medicaid, with or without a spend-down. There are lots of ways to help clients meet their spend-down – including - Medicare cost sharing amounts (deductibles, premiums, co-pays) - over the counter medications if prescribed by a doctor. - expenses paid by state-funded programs like EPIC and ADAP.

- medical bills of person’s spouse or child. - health insurance premiums. - joining a pooled Supplemental Needs Trust (SNT).

B. Medicare Savings Program (MSP) – If client is not eligible for Medicaid, explore eligibility for Medicare Savings Program (MSP). MSP pays for Part B premiums and gets you into the Part D LIS.

There are no asset limits in the Medicare Savings Program. One of the MSPs (QMB), also covers all cost sharing for Parts A &. B.

If your client is eligible for Medicaid AND MSP, enrolling in MSP may subject him/her to, or increase a spend-down, because Medicaid and the various MSPs have different income eligibility levels. It is the client’s choice as to whether or not to be enrolled into MSP. C.

Part D Low Income Subsidy (LIS) – If your client is not eligible for MSP or Medicaid, s/he may still be eligible for Part D Low Income Subsidy. Applications for LIS are also be treated as applications for MSP, unless the client affirmatively indicates that s/he does not want to apply for MSP. d.

Medicare supplemental insurance (Medigap) -- Medigap is supplemental private insurance coverage that covers all or some of the deductibles and coinsurance for Medicare Parts A and B. Medigap is not available to people enrolled in Part C. E.

Medicare Advantage – Medicare Advantage plans “package” Medicare (Part A and B) benefits, with or without Part D coverage, through a private health insurance plan. The cost-sharing structure (deductible, premium, co-pays) varies from plan to plan. For a list of Medicare Advantage plans in your area, go to www.medicare.gov – click on “find health plans.” f.

NY Prescription Saver Card -- NYP$ is a state-sponsored pharmacy discount card that can lower the cost of prescriptions by as much as 60 percent on generics and 30 percent on brand name drugs. Can be used during the Part D “donut hole” (coverage gap) g. For clients living with HIV.

ADAP [AIDS Drug Assistance Program] ADAP provides free medications for the treatment of HIV/AIDS and opportunistic s. ADAP can be used to help meet a Medicaid spenddown and get into the Part D Low Income subsidy. For more information about ADAP, go to V.

GETTING MEDICAID IN THE DISABLED CATEGORY AFTER AN SSI/SSDI DENIAL What if your client's application for SSI or SSDI is denied based on SSA's finding that they were not "disabled?. " Obviously, you have your appeals work cut out for you, but in the meantime, what can they do about health insurance?. It is still possible to have Medicaid make a separate disability determination that is not controlled by the unfavorable SSA determination in certain situations.

Specifically, an applicant is entitled to a new disability determination where he/she. alleges a different or additional disabling condition than that considered by SSA in making its determination. Or alleges less than 12 months after the most recent unfavorable SSA disability determination that his/her condition has changed or deteriorated, alleges a new period of disability which meets the duration requirement, and SSA has refused to reopen or reconsider the allegations, or the individual is now ineligible for SSA benefits for a non-medical reason.

Or alleges more than 12 months after the most recent unfavorable SSA disability determination that his/her condition has changed or deteriorated since the SSA determination and alleges a new period of disability which meets the duration requirement, and has not applied to SSA regarding these allegations. See GIS 10-MA-014 and 08 OHIP/INF-03.[4] [1] Potential wrinkle – for some clients Medicaid is not automatically pick up cost-sharing. In Monroe County we have had several cases where SSA began deducting Medicare Part B premiums from the checks of clients who were receiving SSI and Medicaid and then qualified for Medicare.

The process should be automatic. Please contact Geoffrey Hale in our Rochester office if you encounter any cases like this. [2]Under terms established to provide benefits for QMBs, a provider agreement necessary for reimbursement “may be executed through the submission of a claim to the Medicaid agency requesting Medicaid payment for Medicare deductibles and coinsurance for QMBs.” CMS State Medicaid Manual, Chapter 3, Eligibility, 3490.14 (b), available at.

http://www.cms.hhs.gov/Manuals/PBM/itemdetail.asp?. ItemID=CMS021927. [3]Benchmark plans are free if you are an LIS recipient.

The amount of the benchmark changes from year to year. In 2013, a Part D plan in New York State is considered benchmark if it provides basic Part D coverage and its monthly premium is $43.22 or less. [4] These citations courtesy of Jim Murphy at Legal Services of Central New York.

This site provides general information only. This is not legal advice. You can only obtain legal advice from a lawyer.

In addition, your use of this site does not create an attorney-client relationship. To contact a lawyer, visit http://lawhelp.org/ny. We make every effort to keep these materials and links up-to-date and in accordance with New York City, New York state and federal law.

However, we do not guarantee the accuracy of this information.Some "dual eligible" beneficiaries (people who have Medicare and Medicaid) are entitled to receive reimbursement of their Medicare Part B premiums from New York State through the Medicare Insurance Premium Payment Program (MIPP). The Part B premium is $148.50 in 2021. MIPP is for some groups who are either not eligible for -- or who are not yet enrolled in-- the Medicare Savings Program (MSP), which is the main program that pays the Medicare Part B premium for low-income people.

Some people are not eligible for an MSP even though they have full Medicaid with no spend down. This is because they are in a special Medicaid eligibility category -- discussed below -- with Medicaid income limits that are actually HIGHER than the MSP income limits. MIPP reimburses them for their Part B premium because they have “full Medicaid” (no spend down) but are ineligible for MSP because their income is above the MSP SLIMB level (120% of the Federal Poverty Level (FPL).

Even if their income is under the QI-1 MSP level (135% FPL), someone cannot have both QI-1 and Medicaid). Instead, these consumers can have their Part B premium reimbursed through the MIPP program. In this article.

The MIPP program was established because the State determined that those who have full Medicaid and Medicare Part B should be reimbursed for their Part B premium, even if they do not qualify for MSP, because Medicare is considered cost effective third party health insurance, and because consumers must enroll in Medicare as a condition of eligibility for Medicaid (See 89 ADM 7). There are generally four groups of dual-eligible consumers that are eligible for MIPP. Therefore, many MBI WPD consumers have incomes higher than what MSP normally allows, but still have full Medicaid with no spend down.

Those consumers can qualify for MIPP and have their Part B premiums reimbursed. Here is an example. Sam is age 50 and has Medicare and MBI-WPD.

She gets $1500/mo gross from Social Security Disability and also makes $400/month through work activity. $ 167.50 -- EARNED INCOME - Because she is disabled, the DAB earned income disregard applies. $400 - $65 = $335.

Her countable earned income is 1/2 of $335 = $167.50 + $1500.00 -- UNEARNED INCOME from Social Security Disability = $1,667.50 --TOTAL income. This is above the SLIMB limit of $1,288 (2021) but she can still qualify for MIPP. 2.

Parent/Caretaker Relatives with MAGI-like Budgeting - Including Medicare Beneficiaries. Consumers who fall into the DAB category (Age 65+/Disabled/Blind) and would otherwise be budgeted with non-MAGI rules can opt to use Affordable Care Act MAGI rules if they are the parent/caretaker of a child under age 18 or under age 19 and in school full time. This is referred to as “MAGI-like budgeting.” Under MAGI rules income can be up to 138% of the FPL—again, higher than the limit for DAB budgeting, which is equivalent to only 83% FPL.

MAGI-like consumers can be enrolled in either MSP or MIPP, depending on if their income is higher or lower than 120% of the FPL. If their income is under 120% FPL, they are eligible for MSP as a SLIMB. If income is above 120% FPL, then they can enroll in MIPP.

(See GIS 18 MA/001 - 2018 Medicaid Managed Care Transition for Enrollees Gaining Medicare, #4) When a consumer has Medicaid through the New York State of Health (NYSoH) Marketplace and then enrolls in Medicare when she turns age 65 or because she received Social Security Disability for 24 months, her Medicaid case is normally** transferred to the local department of social services (LDSS)(HRA in NYC) to be rebudgeted under non-MAGI budgeting. During the transition process, she should be reimbursed for the Part B premiums via MIPP. However, the transition time can vary based on age.

AGE 65+ Those who enroll in Medicare at age 65+ will receive a letter from their local district asking them to "renew" Medicaid through their local district. See 2014 LCM-02. The Medicaid case takes about four months to be rebudgeted and approved by the LDSS.

The consumer is entitled to MIPP payments for at least three months during the transition. Once the case is with the LDSS she should automatically be re-evaluated for MSP, even if the LDSS determines the consumer is not eligible for Medicaid because of excess income or assets. 08 OHIP/ADM-4.

Consumers UNDER 65 who receive Medicare due to disability status are entitled to keep MAGI Medicaid through NYSoH for up to 12 months (also known as continuous coverage, See NY Social Services Law 366, subd. 4(c). These consumers should receive MIPP payments for as long as their cases remain with NYSoH and throughout the transition to the LDSS.

NOTE during erectile dysfunction treatment emergency their case may remain with NYSoH for more than 12 months. See here. EXAMPLE.

Sam, age 60, was last authorized for Medicaid on the Marketplace in June 2020. He became enrolled in Medicare based on disability in August 2020, and started receiving Social Security in the same month (he won a hearing approving Social Security disability benefits retroactively, after first being denied disability). Even though his Social Security is too high, he can keep Medicaid for 12 months beginning June 2020.

Sam has to pay for his Part B premium - it is deducted from his Social Security check. He may call the Marketplace and request a refund. This will continue until the end of his 12 months of continuous MAGI Medicaid eligibility.

He will be reimbursed regardless of whether he is in a Medicaid managed care plan. See GIS 18 MA/001 Medicaid Managed Care Transition for Enrollees Gaining Medicare (PDF) When that ends, he will renew Medicaid and apply for MSP with his local district. See GIS 18 MA/001 - 2018 Medicaid Managed Care Transition for Enrollees Gaining Medicare, #4 for an explanation of this process.

That directive also clarified that reimbursement of the Part B premium will be made regardless of whether the individual is still in a Medicaid managed care (MMC) plan. Note. During the erectile dysfunction treatment emergency, those who have Medicaid through the NYSOH marketplace and enroll in Medicare should NOT have their cases transitioned to the LDSS.

They should keep the same MAGI budgeting and automatically receive MIPP payments. See GIS 20 MA/04 or this article on erectile dysfunction treatment eligibility changes 4. Those with Special Budgeting after Losing SSI (DAC, Pickle, 1619b) Disabled Adult Child (DAC).

Special budgeting is available to those who are 18+ and lose SSI because they begin receiving Disabled Adult Child (DAC) benefits (or receive an increase in the amount of their benefit). Consumer must have become disabled or blind before age 22 to receive the benefit. If the new DAC benefit amount was disregarded and the consumer would otherwise be eligible for SSI, they can keep Medicaid eligibility with NO SPEND DOWN.

See this article. Consumers may have income higher than MSP limits, but keep full Medicaid with no spend down. Therefore, they are eligible for payment of their Part B premiums.

See page 96 of the Medicaid Reference Guide (Categorical Factors). If their income is lower than the MSP SLIMB threshold, they can be added to MSP. If higher than the threshold, they can be reimbursed via MIPP.

See also 95-ADM-11. Medical Assistance Eligibility for Disabled Adult Children, Section C (pg 8). Pickle &.

1619B. 5. When the Part B Premium Reduces Countable Income to Below the Medicaid Limit Since the Part B premium can be used as a deduction from gross income, it may reduce someone's countable income to below the Medicaid limit.

The consumer should be paid the difference to bring her up to the Medicaid level ($904/month in 2021). They will only be reimbursed for the difference between their countable income and $904, not necessarily the full amount of the premium. See GIS 02-MA-019.

Reimbursement of Health Insurance Premiums MIPP and MSP are similar in that they both pay for the Medicare Part B premium, but there are some key differences. MIPP structures the payments as reimbursement -- beneficiaries must continue to pay their premium (via a monthly deduction from their Social Security check or quarterly billing, if they do not receive Social Security) and then are reimbursed via check. In contrast, MSP enrollees are not charged for their premium.

Their Social Security check usually increases because the Part B premium is no longer withheld from their check. MIPP only provides reimbursement for Part B. It does not have any of the other benefits MSPs can provide, such as.

A consumer cannot have MIPP without also having Medicaid, whereas MSP enrollees can have MSP only. Of the above benefits, Medicaid also provides Part D Extra Help automatic eligibility. There is no application process for MIPP because consumers should be screened and enrolled automatically (00 OMM/ADM-7).

Either the state or the LDSS is responsible for screening &. Distributing MIPP payments, depending on where the Medicaid case is held and administered (14 /2014 LCM-02 Section V). If a consumer is eligible for MIPP and is not receiving it, they should contact whichever agency holds their case and request enrollment.

Unfortunately, since there is no formal process for applying, it may require some advocacy. If Medicaid case is at New York State of Health they should call 1-855-355-5777. Consumers will likely have to ask for a supervisor in order to find someone familiar with MIPP.

If Medicaid case is with HRA in New York City, they should email mipp@hra.nyc.gov. If Medicaid case is with other local districts in NYS, call your local county DSS. See more here about consumers who have Medicaid on NYSofHealth who then enroll in Medicare - how they access MIPP.

Once enrolled, it make take a few months for payments to begin. Payments will be made in the form of checks from the Computer Sciences Corporation (CSC), the fiscal agent for the New York State Medicaid program. The check itself comes attached to a remittance notice from Medicaid Management Information Systems (MMIS).

Unfortunately, the notice is not consumer-friendly and may be confusing. See attached sample for what to look for. Health Insurance Premium Payment Program (HIPP) HIPP is a sister program to MIPP and will reimburse consumers for private third party health insurance when deemed “cost effective.” Directives:.

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Jem.2021121108132021c. Doi. Https://doi.org/10.1084/jem.2021121108132021c Download citation file:T cell immunotherapies have revolutionized treatment for a subset of cancers. Yet, a major hurdle has been the lack of facile and predicative preclinical animal models that permit dynamic visualization of T cell immune responses at single-cell resolution in vivo. Here, optically clear immunocompromised zebrafish were engrafted with fluorescent-labeled human cancers along with chimeric antigen receptor T (CAR T) cells, bispecific T cell engagers (BiTEs), and antibody peptide epitope conjugates (APECs), allowing real-time single-cell visualization of T cell–based immunotherapies in vivo.

This work uncovered important differences in the kinetics of T cell infiation, tumor cell engagement, and killing between these immunotherapies and established early endpoint analysis to predict therapy responses. We also established EGFR-targeted immunotherapies as a powerful approach to kill rhabdomyosarcoma muscle cancers, providing strong preclinical rationale for assessing a wider array of T cell immunotherapies in this disease. Chimeric antigen receptor T (CAR T) cells and bispecific T cell engagers (BiTEs) redirect autologous T lymphocytes to kill tumor cells. These immunotherapies have shown exceptional clinical responses in many leukemias and lymphomas (Lee et al., 2015. Lim and June, 2017.

Rapoport et al., 2015. Rosenberg and Restifo, 2015. Waldman et al., 2020). However, similar advances have not been made in a large fraction of solid malignancies, largely due to lack of T cell infiation into the tumor, inefficient in vivo cytotoxicity, and off-target toxicity (Chai et al., 2020). Moreover, cell-based therapies have yet to be fully explored in pediatric solid tumors due in part to the lack of efficacy for these therapies in killing adult solid tumors and lack of preclinical rationale in xenograft models to progress studies into the clinic.

This is particularly relevant for rhabdomyosarcoma (RMS), a common pediatric cancer of muscle. RMS is composed of two main subtypes including fusion-positive tumors that harbor PAX3 or PAX7 fusions with FOXO1 and fusion-negative RMSs that have rat sarcoma viagra pathway activation (Skapek et al., 2019). Relapse and refractory RMSs have particularly poor prognosis, and new therapies are sorely needed (Yohe et al., 2019). Recently, two pediatric RMS patients with refractory metastatic and recurrent disease were independently treated with HER2+ and CD56+ CAR T cell therapies, and both had complete disease remission with limited toxicity (Hegde et al., 2020. Jiang et al., 2019).

These clinical results suggest a path forward for T cell–mediated therapies in high-risk refractory and metastatic RMS, pending prioritizing CAR T antigen selection and assessing preclinical efficacy in xenograft models. The advent of ever-increasingly diverse CAR T cells, BiTEs, and additional antibody-based approaches that redirect T cells to engage with tumors has fast outpaced our ability to efficiently test these new therapies in preclinical animal models. For example, new generations of CAR T cells have been engineered to turn on and off CAR T cell responses using chemicals (Giordano-Attianese et al., 2020. Zajc et al., 2020). To express cytokines that increase homing, infiation, and killing (Adachi et al., 2018.

Pegram et al., 2012). And to bind multiple epitopes for increased specificity (Ruella et al., 2016. Wu et al., 2020). Newer innovations such as antibody peptide epitope conjugates (APECs) have also been developed (Millar et al., 2020). These APECs are engineered antibodies that deliver a viral antigen to tumor cells for presentation by HLA-I following proteolytic cleavage by tumor-specific proteases, leading to activation of endogenous CD8+ T cell anti-viral immunity and tumor cell killing.

This approach has claimed to have superior specificity compared with other antibody-mediated approaches, including BiTEs, by restricting T cell killing to areas within the tumor mass (Millar et al., 2020). Despite the many innovations emerging in the immunoncology field, many T cell immunotherapies have yet to translate into clinical successes, in part attributed to the inefficiencies in preexisting preclinical in vivo modeling approaches to predict poor T cell infiation into the tumor, low in vivo tumor cell killing, and lack of target specificity—processes that could best be evaluated in vivo and imaged at single-cell resolution. Here, we report an optically clear adult rag2Δ/Δ, il2rga−/− immunocompromised zebrafish that allows long-term, stable engraftment of human T cells and cancer cells. These mutants were used to engraft fluorescent-labeled human cancers and to quantify responses to CAR T cell, BiTE, and APEC immunotherapies. Single-cell imaging methodologies and high-throughput automated cell counting went on to quantify previously unknown differences between immunotherapies, including quantifying T cell–tumor cell interactions and cytotoxic immune synapse formation.

Our work also identified the efficacy of the newly described APEC immunotherapies in redirecting CMV-primed CD8+ T cells to kill tumor cells in a wide array of cancer types. This work is important because it provides a strong foundation for moving APECs into clinical evaluation in the future. Lastly, our preclinical xenograft studies identified epidermal growth factor receptor (EGFR) T cell immunotherapy as a promising new therapy in a large fraction of pediatric RMSs. Here, we generated a new mutant line of optically clear zebrafish that deletes the entirety of the 3.1-kb recombination activating gene 2 (rag2. Fig.

1 and Fig. S1) and inactivates the interleukin 2 receptor gamma α (il2rga). These new rag2Δ/Δ, il2rga−/− animals are severely immune deficient and lack most mature B, T, and natural killer (NK) cells (Fig. 1, A–D), consistent with the reported immune profile of C;129S4-Rag2tm1.1Flv Il2rgtm1.1Flv/J mice (Goldman et al., 1998). Compound mutant rag2Δ/Δ, il2rga−/− animals were generated at the expected Mendelian ratios, were viable into adulthood, and robustly engrafted a wider array of human tumors than our previously described prkdc−/−, il2rga−/− model (Fig.

1, E–I. And Fig. S2). As expected, the histopathology and cell morphological features of engrafted tumors were similar to those of patient tumors and those grown in NOD.Cg-Prkdcscid Il2rgtm1Wjl/SzJ (NSG) mice (Fig. 1, E–I.

And Fig. S2). We were also able to predict patient-derived xenograft (PDX) responses to combination olaparib Poly (ADP-ribose) polymerase inhibitor and temozolomide DNA–damaging agent in small cell lung cancer using clinically relevant, oral dosing (Fig. S3). Finally, we engrafted normal human CD8+ T cells into rag2Δ/Δ, il2rga−/− animals.

These CD8+ T cells remained in the circulation and colonized the kidney marrow of engrafted animals. In total, up to 6% of the peripheral blood and kidney marrow was composed of human CD8+ cells by 14 d post transplantation (dpt. Fig. 1, J–M). Our results establish the rag2Δ/Δ, il2rga−/− as an improved xenograft transplantation model with specific utility in engrafting human T cells.

Next, we assessed the utility of rag2Δ/Δ, il2rga−/− zebrafish for preclinical modeling of T cell immunotherapy responses. Here, we tested a wide range of clinically relevant T cell immunotherapies that are either U.S. Food and Drug Administration approved (i.e., CD19 CAR T cells and blinatumomab) or under clinical evaluation in open trials (i.e., EGFRvIII CAR T cells and solitomab). First, we engrafted GFP-expressing EGFRvIII+ U87 glioma into the peritoneal cavity of rag2Δ/Δ, il2rga−/− mutant animals. Engrafted animals were administered (i) untransduced T cells, (ii) nontargeted CD19 CAR T cells, or (iii) EGFRvIII-targeted CAR T cells on days 7 and 14 after tumor cell engraftment (Fig.

2 A). Notably, only EGFRvIII-targeted CAR T cells efficiently killed glioma tumors (n = 6 animals/arm, P <. 0.001, Student’s t test), while tumor regressions were not observed in animals injected with untransduced or nontargeted CD19 CAR T cells (Fig. 2, B and F). Histopathology analysis and Tdt-mediated dUTP-biotin nick end labeling (TUNEL) staining confirmed on-target tumor cell killing by EGFRvIII-targeted CAR T cells but not control T cells (Fig.

2, C–E). Similarly, CD19+ JeKo-1 B lymphomas were killed only following administration of CD19-specific CAR T cells, but not untransduced or nontargeted EGFRvIII CAR T cells (Fig. 2 G. And Fig. S4, A–D).

These results are similar to those of xenograft studies performed in NSG mice (Fraietta et al., 2016. O’Rourke et al., 2017) and credential the zebrafish model for accurately assessing CAR T cell responses in vivo. Next, we investigated the in vivo therapy responses and specificity to BiTE immunotherapy. BiTEs are artificial, bispecific monoclonal antibodies that contain two single-chain variable fragments, one that binds T cells through contact with the CD3 receptor while the other binds to a tumor-specific molecule, redirecting endogenous cytotoxic T cells to engage and kill cancer cells. Here, animals were engrafted with GFP-expressing EpCAM+ OVCAR-5 ovarian carcinoma cells or chronic myelogenous leukemia K562 cells engineered to stably express human CD19 (K562-CD19).

After 7 d of engraftment, animals were coinjected with (i) solitomab (EpCAM/CD3) BiTEs, (ii) blinatumomab (CD19/CD3) BiTEs, or (iii) EpCAM control antibody along with human CD8+ T cells weekly. Tumor regressions were only observed within animals administered target-specific BiTEs by 21 d after engraftment (P <. 0.001, Student’s t test. N = 8 animals/arm. Fig.

2, H–M and Fig. S4, E–H). Together, these experiments highlight the exquisitely high degree of target specificity for each BiTE and the relative ease of reading out these tumor responses in live zebrafish when compared with mouse xenograft studies (Brischwein et al., 2006. Mølhøj et al., 2007). We next investigated the kinetics of CAR T cell infiation and engagement when encountering antigen-specific tumor cells using live animal, single-cell imaging.

Specifically, 5 × 104 GFP-expressing U87 EGFRvIII+ glioma cells were engrafted into the superficial orbital musculature of rag2Δ/Δ, il2rga−/− animals. This site is easily assessable for confocal microscopy imaging (Yan et al., 2019. Yan et al., 2020). Following engraftment for 6 d, rag2Δ/Δ, il2rga−/− animals were intraperitoneally (IP) injected with a single dose of nontargeted control T cells or EGFRvIII CAR T cells (5 × 105 cells. Fig.

3 A). T cells were ex vivo labeled with the CFSE cell-permeable dye prior to implantation. Although CFSE intensity decreases by half at each cell division, published studies using engraftment into NSG mice have demonstrated that human T cells undergo only one or two divisions in the first 7 dpt, permitting facile imaging of stained T cells in vivo using confocal microscopy (Quah et al., 2007. Wong and Pamer, 2001). Using this approach, we were able to quantify migration to regions adjacent the tumor by 24 h after injection with both nontargeted and EGFRvIII CAR T cells (Fig.

3, B and F. And Video 1. N = 5 animals per condition). Yet, only EGFRvIII CAR T cells could efficiently enter the tumor mass and kill tumor cells by 5 d after therapy administration (Fig. 3, B–E.

P <. 0.01, Student’s t test). High-magnification 3D volumetric modeling revealed remarkable differences in cell behavior. EGFRvIII CAR T cells rapidly infiated into the tumor and subsequently engaged with tumor cells by 5 d after T cell injection (Fig. 3, C–H.

And Video 1). By contrast, nontransduced T cells migrated to the transplant site (Fig. 3 C) but did not infiate the tumor and rather aligned along the peripheral edge (n = 718 of 750 cells analyzed across all time points, n = 5 animals. Video 1). Similar differences in CAR T cell infiation and additional T cell–mediated immunotherapies were observed on sectioning of animals at the end of the experiment and costaining of samples with human CD3 and the GFP antibody (Fig.

S5). Finally, we were able to directly quantify the number of T cell–tumor cell interactions over time, showing initial engagement of CAR T cells with tumor cells by just 1 d after injection and maximal CAR T cell–tumor cell contact and subsequent tumor cell killing by 5 d after CAR T infusion (Fig. 3 H). By 8 d after immunotherapy, CAR T cell engagement was reduced, likely reflecting near-complete ablation of tumor cells by this time point (Fig. 3, E and H).

In total, these experiments provide a detailed understanding of the kinetics of CAR T cell migration to the tumor site, infiation, engagement, and subsequent tumor cell killing in vivo. Next, we developed a facile single-cell imaging platform to rapidly assess on-target cell killing in vivo and image apoptotic immune synapse formation as an early end point analysis of cell killing. Specifically, we engineered EpCAM+ OVCAR-5 ovarian carcinoma cells to stably express both mCherry and the ZipGFP-Casp3 apoptotic cell reporter, a GFP variant that only emits fluorescence upon Casp3 cleavage of an inhibitory protease sequence inserted between β1-10 and β11 barrel of protein (To et al., 2016). MCherry+/ZipGFP-Casp3+ OVCAR-5 cells were engrafted into the periocular musculature and 6 d later IP injected with 5 × 105 CD8+ CFSE dye–labeled T cells and either (i) 50 µg/kg control EpCAM antibody or (ii) EpCAM/CD3 solitomab (Fig. 4 A).

Solitomab-treated animals had decreased tumor burden over time (Fig. 4 C) and exhibited a significant increase in ZipGFP-Casp3–labeled apoptotic cells starting at 24 h after therapy compared with control EpCAM antibody–treated fish (P <. 0.01, Student’s test. Fig. 4, B and D).

Elevated numbers of ZipGFP-Casp3+ cells were observed by 24 h after treatment, preceding the overall reduction in tumor cells that was observed at 4 and 7 d after treatment with EpCAM/CD3 solitomab, suggesting that BiTE-induced cell killing required >24 h to reach maximal efficiency (Fig. 4, C and D). BiTE-induced cytotoxicity requires physical contact of T cells and tumor cells, forming apoptotic immunological synapse and release of cytolytic granules by T cells in vitro (Roda-Navarro and Álvarez-Vallina, 2020). As expected, solitomab treatment led to high T cell migration and infiation into the tumor (Fig. 4, E, G, and H).

3D modeling of CD8+ T cells with ZipGFP-Casp3+ tumor cells allowed in vivo imaging of T cell–tumor cell contact and quantitation of apoptotic immunological synapses in vivo (Fig. 4 F). 83% of all CD8+ T cells were in direct contact with tumor cells after 7 d of solitomab treatment compared with 24% in EpCAM antibody–treated controls (P <. 0.01, Student’s t test. Fig.

4 I and Video 2). Solitomab treatment also led to higher rates of apoptotic immunological synapse formation by 4 and 7 d of treatment (Fig. 4 J). In addition to high-resolution imaging of cell killing in vivo, these experiments rapidly assessed therapy-induced cell killing that could first be detected 4 d after BiTEs administration, providing a robust and fast assay for determining on-target in vivo killing. A high degree of target specificity is critical for any preclinical cancer immunotherapy currently in development.

As such, we next investigated the specificity of the recently described APEC immunotherapy using rag2Δ/Δ, il2rga−/− zebrafish. APECs are antibody conjugates that deliver viral antigens to the tumor surface for presentation by HLA-I, specifically redirecting preexisting CD8+ T cell antiviral immunity against tumor cells following viral peptide cleavage by tumor proteases (Fig. 5 A. Millar et al., 2020). We first evaluated the efficiency of APEC immunotherapy by engrafting GFP-expressing EpCAM+ OVCAR-5 or EGFR+ MDA–MB-231 cells into the peritoneal cavity of rag2Δ/Δ, il2rga−/− mutant animals.

Engrafted animals were then injected with CMV-specific CD8+ T cells along with (i) EpCAM control antibody, (ii) EpCAM-MMP7-CMV APEC (EpCAM-MC. Unpublished data), or (iii) EGFR-MMP14-CMV APECs (EGFR-M14C) on days 7 and 14 after engraftment. EpCAM-MC and EGFR-M14C were generated using therapeutic EpCAM antibody (clone B38.1) and cetuximab, respectively. In particular, EpCAM-MC is a personalized APEC-immunotherapy for ovarian cancer that cleaves a CMV peptide (NLVPMVATV. Abbreviated NLV) by tumor-expressed MMP7 and then is presented by MHCI (HLA-A*02:01).

EpCAM-MC was identified as the top APEC that kills ovarian carcinoma in a xenograft screen that used high content imaging of therapy responses in rag2Δ/Δ, il2rga−/− zebrafish (unpublished data). Based on these successes, we also engineered a novel EGFR-M14C that contained the same NLV-CMV peptide but used the MMP14 cleavage. MDA–MB-231 cells express high levels of MMP14, which has important roles in driving tumor growth, invasion, and angiogenesis in mouse xenograft studies (Devy et al., 2009). Robust tumor regressions were only observed in animals that received CMV NLV peptide-primed, MHCI-restricted CD8+ T cells along with their target-specific APECs, while control antibody or nontargeted APECs failed to curb overall tumor growth in either model (Fig. S4, I–R).

Like CAR T cells and BiTEs, our results show that APECs have exquisite specificity for killing only epitope-expressing tumors. Building on these results, we next developed a colorimetric specificity assay to directly quantify on-target cell killing at single-cell resolution and to determine the possibility for collateral killing of adjacent nonepitope-expressing cancer cells by APECs. It had been suggested that the cleaved CMV peptide might be taken up by adjacent, nonepitope-expressing cancer cells and presented on the cell surface to engage T cells, further enhancing tumor cell killing of heterogenous cell populations that lack epitope (Millar et al., 2020). Specifically, MDA–MB-231 cells were generated to express mCherry or to coexpress both EpCAM and GFP (Fig. 5 B).

Cells were mixed at 50:50 ratios and engrafted into recipient animals. As expected, EpCAM−/mCherry+ and EpCAM+/GFP+ MDA–MB-231 cells grew at similar rates in engrafted animals when injected with both NLV peptide-primed, MHCI-restricted CD8+ T cells and control EpCAM antibody (Fig. 5, C and F). By contrast, recipient animals that received CD8+ CMV-specific T cells and EpCAM-MC APEC exhibited significant decreases in EpCAM+/GFP+ cells over time, while EpCAM−/mCherry+ tumor cell number was largely unaffected (Fig. 5, C and F).

Higher-magnification 3D modeling confirmed CMV-specific T cell interactions were far more frequent with EpCAM+/GFP+ tumor cells, but less so with EpCAM−/mCherry+ cells at 11 dpt (P <. 0.001, Student’s t test. Fig. 5, D, E, and G. And Video 3).

Similar results were also seen at 14 d after engraftment. Our work indicates that APEC therapy is highly specific to killing epitope-expressing cells and unexpectedly exhibits little effect on adjacent tumor cells that do not express the antigen. Pediatric RMS is a common muscle cancer found in children and has an abysmal 5-yr survival rate of <30% in the metastatic, recurrent, and refractory disease settings (Yohe et al., 2019). Recently, two high-risk pediatric RMS patients were independently treated with HER2+ or CD56+ CAR T cells and achieved long-term disease remission, suggesting potential opportunities to exploit T cell immunotherapy in RMS (Hegde et al., 2020. Jiang et al., 2019).

Despite these amazing success stories, the HER2 and CD56 receptors are only expressed in a small fraction of RMS patients (Bahrami et al., 2008. Ganti et al., 2006. Taniguchi et al., 2008). To systemically interrogate the most suitable antigen for this disease, we first identified all antigen targets that are currently being investigated in active sarcoma immunotherapy clinical trials and assessed if these markers are expressed in RMS (Fig. 6 A).

In total, six surface antigens were identified as potential epitopes for RMS immunotherapy, including EGFR. A retrospective study using immunostaining of RMS patient samples (Dobrenkov et al., 2016. Ganti et al., 2006. Grass et al., 2009. Majzner et al., 2019.

Saraf et al., 2019. Shibui et al., 2019. Thway et al., 2011) revealed EGFR expression in a substantial fraction of human RMS (n = 316 out of 476 primary patient samples and n = 9 of 12 cell line/PDXs. Fig. 6, B–D.

And Table 1). Importantly, pediatric RMS patients had significantly higher incidence and overall expression levels of EGFR than patients with adult disease (74% vs. 50%, respectively, P = 0.03 by χ2 test. Fig. 6, E and F.

And Table 1). To investigate if and which T cell immunotherapy most effectively curbs RMS tumor growth in vivo, we next performed a side-by-side comparison of EGFR CAR T cells, BiTEs, and APECs to assess differences in killing of RMS cells engrafted into rag2Δ/Δ, il2rga−/− zebrafish. These experiments are the first to directly compare these three immunotherapy responses in vivo using the same tumor models and epitopes, allowing direct comparison of cell behavior and killing across immunoncology platforms. Specifically, 5 × 105 RD cells were IP injected into rag2Δ/Δ, il2rga−/− mutant animals, and animals were administered (i) EGFR CAR T cells, (ii) EGFR/CD3 BiTE with CD8+ T cells, (iii) EGFR-M14C with CMV-specific T cells, or (iv) control EGFR antibody with CD8+ T cells on days 7, 14, and 21 after engraftment (n = 5 animals/arm. Fig.

7 A). Notably, significant reductions in tumor burden were observed in all three immunotherapies tested, with EGFR CAR T cells eliciting a trend toward more potent cell killing by 21 d after therapy that was associated with reduced Ki67 proliferation marker expression, higher numbers of TUNEL+ apoptotic cells, and subsequently fewer tumor cells per area on section (Fig. 7, A–D). To further understand the kinetics of cell killing and possible differences in T cell infiation and engagement between the three therapies, we next engrafted 5 × 104 ZipGFP-Casp3–expressing RD cells into the periocular muscle and used confocal imaging to quantify T cell infiation and cell apoptosis following administration of each T cell immunotherapy (Fig. 7, E–I).

Among the three therapies analyzed, EGFR/CD3 BiTEs induced the most robust T cell infiation (Fig. 7, E and H). Yet despite fewer CAR T cells entering the tumor, they elicited the most effective tumor cell killing and subsequent reductions in tumor cell number over time (P <. 0.02, Student’s t test. Fig.

7, F and G), suggesting higher overall therapy efficacy of CAR T cell immunotherapies. Indeed, high-resolution 3D modeling of T cell–tumor cell interaction revealed a higher percentage of T cells could induce tumor cell killing by CAR T cells compared with BiTE- and APEC-treated animals (n = 5 animals/arm. P <. 0.01, Student’s t test. Fig.

7 I). Finally, although APEC-treated cells had less infiative capacity to enter tumors (Fig. 7 H), both BiTEs and APECs had similar potency in killing tumor cells following infiation into the tumor mass (Fig. 7 I). We next confirmed EGFR CAR T cell responses in a wider array of RMS tumors including both fusion-positive and fusion-negative subtypes of RMSs.

As was seen in RD RMS xenografted cells, EGFR CAR T cells efficiently infiated into the tumor and effectively killed RMS tumor cells over time in three additional xenograft models irrespective of disease subtype (n = 5 zebrafish/arm. P <. 0.006, Student’s t test. Fig. 8, A–C).

As was seen in other CAR T cell indications tested in our work, there was an overall lower percentage of CAR T cells engaged with tumor at any given time compared with BiTE and APEC treatment. Yet these CAR T cell therapies efficiently killed tumors over longer periods of time. These results again highlight differences in the rates of cell killing by these modalities. Importantly, EGFR CAR T cell immunotherapy was also effective in suppressing RMS xenograft growth in NSG mice (n = 6 mice/arm. P <.

0.05 over multiple time point analysis. Fig. 8 D). Together, our zebrafish and mouse xenograft studies provide much-needed preclinical rationale for assessing a wide array of EGFR-targeted immunotherapies in RMS. Our work has established the rag2Δ/Δ, il2rga−/− zebrafish as a powerful model for assessing T cell immunotherapy responses at single-cell resolution, successfully imaging single-cell therapy responses in 10 cancer xenograft models across eight distinct T cell immunotherapies.

Recently, others have leveraged the zebrafish larval cell transplantation platform to model the effects of tumor-infiating lymphocytes and CD19 CAR T cells in human cancer xenografts (He et al., 2020. Pascoal et al., 2020). Yet, these larval xenotransplant studies were limited to engraftment of only a few hundred cells per animal, animals could not be reared at physiological 37°C, and assay times were restricted to 24 h in both studies. By contrast, our studies demonstrated the utility of the adult immunocompromised zebrafish model in a wide array of immunotherapies including CAR T cells, BiTEs, and APECs. We also optimized intravital imaging approaches to quantitatively assess T cell migration and infiation into the tumor mass and tumor cell killing in real time and document target specificity serially over days.

Our studies also demonstrated that most efficient T cell immunotherapy–induced cell killing falls between 24 and 96 h after administration, time points that would not be evaluable using the larval xenograft models. The adult immune-deficient zebrafish model is more akin to xenograft studies that use NSG mice, where longer time course studies allow for assessment of immune-therapy responses for up to 3 wk. Mouse xenograft studies have led to many important insights into mechanisms of T cell homing, infiation, and tumor killing (Boissonnas et al., 2007. Boulch et al., 2021. Cazaux et al., 2019.

Halle et al., 2016. Hoekstra et al., 2020. Mulazzani et al., 2019). Yet, the zebrafish xenograft model addresses several fundamental difficulties of completing similar studies using mouse xenografts. For example, imaging T cell/tumor cell responses in mouse xenografts is more technically challenging and requires complex multi-photon microscopy and prior knowledge of where to image based on surgical window creation.

Mice are also more expensive. Thus, large-scale studies are limited. Despite the perceived powers of the zebrafish xenograft model, it also has limitations. For example, xenografted zebrafish were unable to be imaged longer than 10 min in our studies, mainly attributed to gill respiration movements around the periocular engraftment site, precluding a more detailed time-lapse study of T cell movement and subsequent tumor cell killing. These technical hurdles will likely be addressed by the development of deep gill-perfusion anesthesia techniques, selection of alternative engraftment sites such as the dorsal musculature, and refined imaging platforms in the future.

Despite the many advantages of engrafting human tumors and assessing immunotherapy responses in adult rag2Δ/Δ, il2rga−/− zebrafish and NSG mice, both models lack endogenous T, B, NK, and macrophage cells, precluding assessment of how these important immune cells modulate the tissue microenvironment and modify responses to T cell therapy. New immune-deficient zebrafish models will likely incorporate transgenic approaches to express human cytokines that support human blood cells and allow for further humanization, akin to knock-in and transgenic mouse models such as the humanized M-CSFh/h, IL-3/GM-CSFh/h, SIRPah/h, TPOh/h, RAG2−/−, IL2Rg−/− mice (Das et al., 2016. Rongvaux et al., 2014. Wunderlich et al., 2010. Wunderlich and Mulloy, 2016).

Such models would permit engraftment of both the human immune system and tumor to follow more complex immune cell interactions. In fact, elegant work from the Berman group has already developed transgenic fish that express human GM-CSF, stem cell factor, and stromal derived factor 1α and permit short-term engraftment of human hematopoietic stem cells and leukemia cells into larval fish, providing an important starting point for such studies (Rajan et al., 2020). At a cell biological level, our work demonstrated important differences in early responses between CAR T cell, BiTE, and APEC immunotherapies. For example, single-cell quantitative studies revealed that CAR T cells engaged with tumor cells by just 24 h after engraftment in most xenograft models, while neither BiTE- nor APEC-treated T cells exhibited such robust early responses. CAR T cell–tumor cell interactions also led to elevated overall tumor cell killing as read out by an overall reduction in tumor cell number and higher percentage of apoptotic synapse formation between CAR T cell–tumor cells compared with BiTE- and APEC-treated T cells.

Although our work showed a direct correlation between T cell/tumor cell contact and killing, recent work has also demonstrated bystander/indirect killing trough cytokines secreted by T cells in the absence of direct contact between tumor cell and T cells, suggesting multiple possible modes of tumor cell killing (Hoekstra et al., 2020). In total, differences in T cell engagement and cytotoxicity observed in our work are likely accounted for by the fundamentally different mechanisms by which CAR T cells, BiTEs, and APECs bind to tumor cells, induce activation, and kill. For example, CAR T cells engage tumor cells through a chimeric antigen receptor scFv domain and are primed for activation by ex vivo stimulation (Rafiq et al., 2020). Thus, robust and fast tumor cell killing would be expected. T cell receptor/antigen binding then activates CAR T cells, releasing perforin, granzyme, and apoptosis-inducing cytokines.

In contrast, BiTEs mechanically link nonstimulated T cells with tumor cells by simultaneously binding a tumor-specific antigen and the CD3 receptor on T cells (Huehls et al., 2015). Antibody/CD3 binding then activates T cells, eliciting endogenous T cell effector responses. Finally, APECs harness the antiviral CD8+ T cell responses by loading viral epitopes onto tumor-expressed MHCI, promoting the formation of MHC/TCR complexes with viral-specific T cells and subsequent activation (Millar et al., 2020). The processes by which BiTEs and APECs induce T cell activation would be predicted to require longer interaction times between the T cell with tumor before inducing cell death compared with CAR T cells. In total, our observations are largely mirrored by in vivo mice xenograft studies targeting NY–ESO-1 antigen on human myeloma cells or 237scFV on mice fibrosarcoma cells (Maruta et al., 2019.

Stone et al., 2012). These studies performed a side-by-side comparison of CAR T cell and BiTE cell killing efficacy using mouse models and in both instances showed CAR T cells to be superior at killing tumor cells over long–time course experiments. However, these murine models lacked the resolution to visualize tumor cell–immune cell interactions at single-cell resolution and hence failed to provide mechanistic insights into the early kinetic differences between T cell–based therapies and linking of these cell biological differences with effects on tumor cell killing. Our work also validated the efficacy and target specificity of the new APEC technology to kill antigen-expressing tumor cells. First, we demonstrated the feasibility and in vivo utility of designing APECs that express tumor-specific protease cleavage sites in the context of ovarian carcinoma (unpublished data).

This EpCAM-MC APEC was created by conjugating an MMP7 protease cleavage site (AVSRLRAYNLVPMVATV) with a CMV-specific viral epitope NLV to the therapeutic EpCAM antibody (Clone 38.1. Unpublished data). Because APECs bind to tumor-specific epitopes and only release their viral peptide cargo locally following cleavage by tumor-secreted proteases, target specificity is expected to be enhanced over other T cell immunotherapies, including traditional CAR T cells and BiTEs. APEC approaches thus allow exquisite redirection of endogenous viral CD8+ T cells to kill seemingly virally infected tumor cells. Here, we validated the remarkable specificity for epitope-expressing tumor cells with little collateral killing of nonepitope-expressing tumor cells.

Building on these successes, we also developed a new EGFR-M14C APEC that efficiently killed RMS cells. In total, our work has shown that APECs are a new and powerful immunotherapy that will be valuable for tailoring personalized immunotherapy, pairing tumor-specific epitope expression with tumor-specific protease expression and cleavage. Our work also has established much needed preclinical modeling for T cell–mediated immunotherapy in RMS and identified EGFR-based immunotherapies as a new approach to kill RMS tumor cells. EGFR is expressed in a large fraction of RMS patients, and clinical trials using EGFR immunotherapies have reported tolerable side effects (Guo et al., 2018). Despite achieving remissions in a large fraction of pediatric RMSs, relapse, refractory, and recurrent metastatic diseases are the major clinical challenge facing patients.

Current treatment strategies rely on aggressive treatment that includes radiation, chemotherapy, and surgery (Yohe et al., 2019). Targeted immunotherapy approaches could prove powerful for the treatment of aggressive RMS tumors, especially in the relapse setting and for those tumors that cannot be easily resected. Indeed, two pediatric RMS patients with refractory metastatic and recurrent disease achieved complete remissions following HER2+ and CD56+ CAR T therapy (Hegde et al., 2020. Jiang et al., 2019), suggesting a clinical path forward for T cell therapies in this disease. Our zebrafish and mouse xenograft studies have provided a strong preclinical rationale for assessing EGFR CAR T cell and other immunotherapies in this disease.

rag2Δ/Δ zebrafish were created in the Casper-strain zebrafish using CRISPR/Cas9 genome editing (Hwang et al., 2013). Briefly, two guide RNA (gRNA) sequences (5′-AGA​ACC​GTA​TCA​AGC​GCG​GG-3′ and 5′-GGC​CCT​TGA​CTA​CAT​ATG​GTG-3′) were designed to flank the 3.3 kb of the rag2 gene locus using CHOPCHOP. GRNAs were cloned into the DR274 vector (Addgene) and transcribed in vitro using the T3 mMESSAGE mMACHINE Kit. A mixture of 300 ng/µl of Cas9 mRNA and 100 ng/µl of gRNA was coinjected into one cell–stage zebrafish embryos. F0 animals were incrossed, and progeny were assessed for gene deletion using PCR of genomic DNA.

WT gene-specific primers amplified a 444-bp fragment of the endogenous rag2 gene (forward 5′-CCA​TAT​AGC​CAA​TTA​CTA​C-3′, reverse 5′-GCA​GAT​CTG​GAT​CTG​GAG​T-3′. Denaturing. 94°C/30 s, annealing. 60°C/30 s, elongation. 68°C/60 s, and termination.

68°C/5 min). Mutant gene–specific primers (forward 5′-CCC​ATC​TAT​GGG​AAA​CTA​TC-3′, reverse 5′-GTG​TCA​CAT​GAT​CCT​TCA​G-3′) spanned the genomic deletion and amplified a 973-bp fragment (denaturing. 94°C/30 s, annealing. 60°C/30 s, elongation. 68°C/60 s, 36 cycles, termination.

68°C/5 min. Fig. S1, A–C). Il2rgaY91fs-specific primers (forward 5′-TTT​GAC​ATC​GAA​GAC​TGT​CCT​G-3′, reverse 5′-GTC​CTG​TAA​CGA​ACT​TCG​CTC​T-3′) spanned the genomic mutation, amplifying a 373-bp WT allele and a 360-bp mutant allele (denaturing. 94°C/30 s, annealing.

60°C/30 s, elongation. 68°C/30 s, 36 cycles, and termination. 68°C/5 min). Male rag2Δ/Δ, Il2rgaY91fs/+ adult Casper (double mutant for roya9/a9 and nacrew2/w2) zebrafish were crossed to female rag2Δ/+, il2rgaY91fs/+ adult Casper zebrafish. Progeny were grown to adulthood and then subjected to scale resection genotyping at 2–3 mo of age as previously described (Yan et al., 2019).

Homozygous mutant rag2Δ/Δ, il2rga−/− fish were produced at that expected Mendelian ratio (11.75%, n = 984 of 8,374 fish, expected 12.5%). CD8+ T cells used as control for BiTE experiments were purchased (Stem Cell Technology. #70027), cultured, and expanded using RPMI medium (10% FBS and 1% penicillin/streptomycin) supplemented with 20 IU/ml of rhIL-2. CAR T cells were generated using primary donor T cells transduced with the anti-CD19 or EGFRvIII CAR containing a 4-1BB intracellular signaling domain and expanded as previously reported (Milone et al., 2009). Briefly, donor T cells were thawed and activated using α-CD3/α-CD28 Dynabeads (Life Technologies) at a 1:3 T cell:beads ratio.

The cells were cultured in RPMI medium (10% FBS and 1% penicillin/streptomycin) supplemented with 20 IU/ml of rhIL-2. Every 2 d, fresh medium was added to keep the cells at a concentration of 0.5–2 × 106/ml. For CAR T cell production, lentiviral vector was added to the culture 24 h after activation. In parallel, donor-matched T cells that had been activated but untransduced were expanded to serve as a negative control in subsequent experiments. At days 12–14 of culture, CAR expression was determined.

NLV-specific T cell lines were created from CMV-seropositive peripheral blood mononuclear cells isolated from healthy donors upon stimulation with 10 μg/ml CMV peptide HLA-A*02:01 restricted NLV (Genscript) in RPMI1640 supplemented with 10% FBS, 100 U/ml each of penicillin and streptomycin, 4 mM of L-glutamine, and 1% human AB serum (Sigma). On day 4, half culture medium was replaced with the same medium plus 500 IU/ml IL-2 (Peprotech) and changed twice weekly. NLV-specific CD8+ T cells were identified by flow cytometry with staining of HLA–peptide tetrameric complex on day 14–21. Only those T cell lines with >50% NLV-specific CD8+ T cells were qualified for the next functional assay for a maximum of 6 wk. Imaging of periocular transplanted cells was performed using an inverted LSM 710 confocal microscope (Zeiss) with Zen software platform (Zeiss), as previously described (Yan et al., 2019.

Yan et al., 2020). Engrafted zebrafish were anesthetized using low-dose 0.01% tricaine (Western Chemical), placed onto a 36-mm glass-bottom dish (Thermo Fisher Scientific. #150680), and the fish torso was embedded in 1% low–melting point agarose (Thermo Fisher Scientific) to stabilize the animal for imaging. To keep the animal under anesthetic during imaging, zebrafish were submerged in 5 ml of warm 37°C fish water containing 0.01% tricaine. Serial z-stack imaging was performed using a 10× objective (numerical aperture, 0.45), achieving an overall 100× magnification.

GFP- and mCherry-expressing engrafted animals were imaged using the 488-nm (emission = 493–586 nm) and 546-nm laser (emission = 575–703 nm). All T cells used in this study were stained with 1 µM of ViaFluor CFSE before transplantation and imaged using a 405-nm laser (emission = 350–470 nm). In vivo cell apoptosis of ZipGFP-Casp3 imaging studies were imaged used a 488-nm (emission = 493–586 nm) and 546-nm laser (emission = 575–703 nm). Images were automatically annotated and counted in Imaris. Total T cell, tumor cell, and apoptotic cell (ZipGFP-Casp3+) numbers were quantified using Imaris spot and surface functions.

Quantification was completed on z-stack images with dimensions 1,000 µm × 1,000 µm × 100 µm (0.1 mm3). 3D modeling analysis quantifying absolute distance between T cells and tumor cells with direct cell-to-cell contact was defined by <10 µm between cells. This analysis was completed on z-stack tumor images sampled at 500 µm × 500 µm × 50 µm (0.0125 mm3) and analyzed using the distance transformation, distance between spot to surface, and spot close to surface XTension functions. Apoptotic cells were pseudo-colored yellow using Imaris surface function for easy visualization in Fig. 7 E.

Fluorescent-labeled human cancer cell lines (U87, JeKo-1, OVCAR-5, K562, MDA–MB-231, RD, SMS-CTR, Rh41, and Rh30) were transplanted IP (5 × 105 cells/animal) or periocularly (5 × 104 cells/animal) into recipient fish. Stably engrafted fish were IP injected with CAR T cells (5 × 105 cells/dose). For BiTE experiments, animals were coinjected with 5 × 105 CD8+ T cells along with control antibodies (EpCAM, 50 µg/kg. CD19, 50 µg/kg, or EGFR, 50 µg/kg), solitomab (50 µg/kg), blinatumomab (250 µg/kg), or EGFR/CD3 BiTE (10 µg/kg). For APEC experiments, animals were injected with 5 × 105 CMV-specific T cells, along with control antibodies (EpCAM, 50 µg/kg, or EGFR, 50 µg/kg), EpCAM-MC (50 µg/kg), or EGFR-M14C (50 µg/kg).

All IP injections were completed using a 30 1/2–G needle (BD) at the time points noted for specific experiments. At the end of the experiment, animals were fixed in 4% paraformaldehyde, sectioned, and examined histologically by H&E staining, immunohistochemistry (IHC) for Ki67, and TUNEL. Comparisons between groups were performed using ANOVA followed by Student’s t test. Engrafted zebrafish were fixed in 4% paraformaldehyde, embedded in paraffin, and sectioned at 5-mm thickness. Sections were stained by H&E, IHC, or immunofluorescence.

For IHC staining, the primary antibodies were rabbit monoclonal anti-Ki67 (1:1,000 dilution. Abcam), monoclonal anti-CD3 (1:100 dilution. Abcam), monoclonal anti-GFP (1:100 dilution. Abcam), monoclonal anti-EGFR (1:100 dilution. Abcam), and TUNEL (1:1,000 dilution.

Thermo Fisher Scientific). Secondary antibodies were biotinylated goat anti-rabbit IgG antibody (Vectorlabs) and horse anti-mouse antibody (Vectorlabs). Development was completed using Vectastain ABC Kit (Vectorlabs) or Alexa Fluor 488 anti-mouse and Alexa Fluor 546 anti-rabbit secondary antibody (1:1,000 dilution for all secondary antibodies). Quantification was completed based on counting three randomly selected fields imaged at 400× using an Olympus BX41 microscope. Quantification used ImageJ and was blinded.

Images were counted without labels by Eric Alpert. Samples were analyzed using a fixed threshold for achieving an unbiased, quantitative assessment of the IHC and TUNEL staining within the selected imaged field (Yan et al., 2020). Percentage of proliferating (Ki67) and apoptotic (TUNEL) cells was calculated by dividing the number of positively stained cells by the total number of cells counted within the selected fields. N >. 200 control cells were analyzed per sample, with fewer cells being counted for treated samples.

H&E-stained sections were imaged at 400× and quantified as number of cells per unit area. We thank Dr. Len Zon for helpful discussion and feedback on the work. This work is supported by National Institutes of Health grants R24OD016761 (D.M. Langenau), R01CA154923 (D.M.

Langenau), R01CA215118 (D.M. Langenau), and R01CA211734 (D.M. Langenau). The Liddy Shriver Sarcoma Initiative (D.M. Langenau).

The Massachusetts General Hospital Research Scholars Program (D.M. Langenau), U01CA220323 (N.J. Dyson). The Tosteson &. Fund for Medical Discovery Fellowship from Massachusetts General Hospital (C.

Yan). And the Alex’s Lemonade Stand Foundation Young Investigator Award (C. Yan). Author contributions. C.

Yan and D.M. Langenau conceived, designed, and conducted the study. Analyzed data. And prepared the manuscript. C.

Yan, Q. Yang, S. Zhang, D.G. Millar, E.J. Alpert, and D.

Do performed experiments. A. Veloso, D.C. Brunson, B.J. Drapkin, M.

Stanzione, I. Scarfò, J.C. Moore, Y. Wei, and K.M. McCarthy assisted with experimental design and execution.

S. Iyer and Q. Qin performed bioinformatics analysis studies. J.F. Rawls, N.J.

Dyson, M. Cobbold, and M.V. Maus provided important intellectual contributions, designed experiments, and provided reagents, cell lines, and/or PDXs. Competing Interests Disclosures. D.G.

Millar reported personal fees from Revitope Inc. During the conduct of the study and personal fees from Revitope Inc. Outside the submitted work. In addition, D.G. Millar had a patent to WO2012123755A1 licensed "Revitope Inc." and owns stock in Revitope Inc.

B.J. Drapkin reported personal fees from AstraZeneca outside the submitted work. M. Cobbold reported "other" from Revitope Oncology outside the submitted work. In addition, M.

Cobbold had a patent to WO 2012/123755 issued "Revitope Oncology". And is currently an employee of Astrazeneca. M.V. Maus is an inventor on patents related to adoptive cell therapies, held by Massachusetts General Hospital and University of Pennsylvania (some licensed to Novartis). M.V.

Maus holds equity in TCR2, Century Therapeutics, and Neximmune, and has served as a consultant for multiple companies involved in cell therapies. In addition, M.V. Maus is a consultant for Adaptimmune, Agenus, Allogene, Arcellx, Astellas, AstraZeneca, Atara, Bayer, BMS, Cabaletta Bio (SAB), Cellectis (SAB), CRISPR therapeutics, In8bio (SAB), Innovakine, Intellia, GSK, Kite Pharma, Micromedicine, Neximmune, Novartis, TCR2 (SAB), Tmunity, Torque, and WindMIL (SAB). Has grant/research support from CRISPR therapeutics, Kite Pharma, Servier, and Novartis. Is a stockholder of Century Therapeutics, TCR2, and Ichnos.

And is a member of board of directors for Ichnos Sciences. D.M. Langenau reported a patent pending on use of immune deficient zebrafish for xenograft transplantation studies. No other disclosures were reported..

Citation Jonathan Lopez, Marine Mommert, William Mouton, Andrés Pizzorno, Karen Brengel-Pesce, Mehdi Mezidi, Marine Villard, Bruno Lina, Jean-Christophe Richard, Jean-Baptiste Fassier, Valérie Cheynet, Blandine Padey, Victoria Duliere, Thomas Julien, viagra cost Stéphane Paul, Paul Bastard, Alexandre Belot, Antonin Bal, Jean-Laurent Casanova, Manuel Rosa-Calatrava, Florence Morfin, Thierry Walzer, Sophie Trouillet-Assant. Correction. Early nasal type I viagra cost IFN immunity against erectile dysfunction is compromised in patients with autoantibodies against type I IFNs.

J Exp Med 4 October 2021. 218 (10). Jem.2021121108132021c.

Doi. Https://doi.org/10.1084/jem.2021121108132021c Download citation file:T cell immunotherapies have revolutionized treatment for a subset of cancers. Yet, a major hurdle has been the lack of facile and predicative preclinical animal models that permit dynamic visualization of T cell immune responses at single-cell resolution in vivo.

Here, optically clear immunocompromised zebrafish were engrafted with fluorescent-labeled human cancers along with chimeric antigen receptor T (CAR T) cells, bispecific T cell engagers (BiTEs), and antibody peptide epitope conjugates (APECs), allowing real-time single-cell visualization of T cell–based immunotherapies in vivo. This work uncovered important differences in the kinetics of T cell infiation, tumor cell engagement, and killing between these immunotherapies and established early endpoint analysis to predict therapy responses. We also established EGFR-targeted immunotherapies as a powerful approach to kill rhabdomyosarcoma muscle cancers, providing strong preclinical rationale for assessing a wider array of T cell immunotherapies in this disease.

Chimeric antigen receptor T (CAR T) cells and bispecific T cell engagers (BiTEs) redirect autologous T lymphocytes to kill tumor cells. These immunotherapies have shown exceptional clinical responses in many leukemias and lymphomas (Lee et al., 2015. Lim and June, 2017.

Rapoport et al., 2015. Rosenberg and Restifo, 2015. Waldman et al., 2020).

However, similar advances have not been made in a large fraction of solid malignancies, largely due to lack of T cell infiation into the tumor, inefficient in vivo cytotoxicity, and off-target toxicity (Chai et al., 2020). Moreover, cell-based therapies have yet to be fully explored in pediatric solid tumors due in part to the lack of efficacy for these therapies in killing adult solid tumors and lack of preclinical rationale in xenograft models to progress studies into the clinic. This is particularly relevant for rhabdomyosarcoma (RMS), a common pediatric cancer of muscle.

RMS is composed of two main subtypes including fusion-positive tumors that harbor PAX3 or PAX7 fusions with FOXO1 and fusion-negative RMSs that have rat sarcoma viagra pathway activation (Skapek et al., 2019). Relapse and refractory RMSs have particularly poor prognosis, and new therapies are sorely needed (Yohe et al., 2019). Recently, two pediatric RMS patients with refractory metastatic and recurrent disease were independently treated with HER2+ and CD56+ CAR T cell therapies, and both had complete disease remission with limited toxicity (Hegde et al., 2020.

Jiang et al., 2019). These clinical results suggest a path forward for T cell–mediated therapies in high-risk refractory and metastatic RMS, pending prioritizing CAR T antigen selection and assessing preclinical efficacy in xenograft models. The advent of ever-increasingly diverse CAR T cells, BiTEs, and additional antibody-based approaches that redirect T cells to engage with tumors has fast outpaced our ability to efficiently test these new therapies in preclinical animal models.

For example, new generations of CAR T cells have been engineered to turn on and off CAR T cell responses using chemicals (Giordano-Attianese et al., 2020. Zajc et al., 2020). To express cytokines that increase homing, infiation, and killing (Adachi et al., 2018.

Pegram et al., 2012). And to bind multiple epitopes for increased specificity (Ruella et al., 2016. Wu et al., 2020).

Newer innovations such as antibody peptide epitope conjugates (APECs) have also been developed (Millar et al., 2020). These APECs are engineered antibodies that deliver a viral antigen to tumor cells for presentation by HLA-I following proteolytic cleavage by tumor-specific proteases, leading to activation of endogenous CD8+ T cell anti-viral immunity and tumor cell killing. This approach has claimed to have superior specificity compared with other antibody-mediated approaches, including BiTEs, by restricting T cell killing to areas within the tumor mass (Millar et al., 2020).

Despite the many innovations emerging in the immunoncology field, many T cell immunotherapies have yet to translate into clinical successes, in part attributed to the inefficiencies in preexisting preclinical in vivo modeling approaches to predict poor T cell infiation into the tumor, low in vivo tumor cell killing, and lack of target specificity—processes that could best be evaluated in vivo and imaged at single-cell resolution. Here, we report an optically clear adult rag2Δ/Δ, il2rga−/− immunocompromised zebrafish that allows long-term, stable engraftment of human T cells and cancer cells. These mutants were used to engraft fluorescent-labeled human cancers and to quantify responses to CAR T cell, BiTE, and APEC immunotherapies.

Single-cell imaging methodologies and high-throughput automated cell counting went on to quantify previously unknown differences between immunotherapies, including quantifying T cell–tumor cell interactions and cytotoxic immune synapse formation. Our work also identified the efficacy of the newly described APEC immunotherapies in redirecting CMV-primed CD8+ T cells to kill tumor cells in a wide array of cancer types. This work is important because it provides a strong foundation for moving APECs into clinical evaluation in the future.

Lastly, our preclinical xenograft studies identified epidermal growth factor receptor (EGFR) T cell immunotherapy as a promising new therapy in a large fraction of pediatric RMSs. Here, we generated a new mutant line of optically clear zebrafish that deletes the entirety of the 3.1-kb recombination activating gene 2 (rag2. Fig.

1 and Fig. S1) and inactivates the interleukin 2 receptor gamma α (il2rga). These new rag2Δ/Δ, il2rga−/− animals are severely immune deficient and lack most mature B, T, and natural killer (NK) cells (Fig.

1, A–D), consistent with the reported immune profile of C;129S4-Rag2tm1.1Flv Il2rgtm1.1Flv/J mice (Goldman et al., 1998). Compound mutant rag2Δ/Δ, il2rga−/− animals were generated at the expected Mendelian ratios, were viable into adulthood, and robustly engrafted a wider array of human tumors than our previously described prkdc−/−, il2rga−/− model (Fig. 1, E–I.

And Fig. S2). As expected, the histopathology and cell morphological features of engrafted tumors were similar to those of patient tumors and those grown in NOD.Cg-Prkdcscid Il2rgtm1Wjl/SzJ (NSG) mice (Fig.

We were also able to predict patient-derived xenograft (PDX) responses to combination olaparib Poly (ADP-ribose) polymerase inhibitor and temozolomide DNA–damaging agent in small cell lung cancer using clinically relevant, oral dosing (Fig. S3). Finally, we engrafted normal human CD8+ T cells into rag2Δ/Δ, il2rga−/− animals.

These CD8+ T cells remained in the circulation and colonized the kidney marrow of engrafted animals. In total, up to 6% of the peripheral blood and kidney marrow was composed of human CD8+ cells by 14 d post transplantation (dpt. Fig.

1, J–M). Our results establish the rag2Δ/Δ, il2rga−/− as an improved xenograft transplantation model with specific utility in engrafting human T cells. Next, we assessed the utility of rag2Δ/Δ, il2rga−/− zebrafish for preclinical modeling of T cell immunotherapy responses.

Here, we tested a wide range of clinically relevant T cell immunotherapies that are either U.S. Food and Drug Administration approved (i.e., CD19 CAR T cells and blinatumomab) or under clinical evaluation in open trials (i.e., EGFRvIII CAR T cells and solitomab). First, we engrafted GFP-expressing EGFRvIII+ U87 glioma into the peritoneal cavity of rag2Δ/Δ, il2rga−/− mutant animals.

Engrafted animals were administered (i) untransduced T cells, (ii) nontargeted CD19 CAR T cells, or (iii) EGFRvIII-targeted CAR T cells on days 7 and 14 after tumor cell engraftment (Fig. 2 A). Notably, only EGFRvIII-targeted CAR T cells efficiently killed glioma tumors (n = 6 animals/arm, P <.

0.001, Student’s t test), while tumor regressions were not observed in animals injected with untransduced or nontargeted CD19 CAR T cells (Fig. 2, B and F). Histopathology analysis and Tdt-mediated dUTP-biotin nick end labeling (TUNEL) staining confirmed on-target tumor cell killing by EGFRvIII-targeted CAR T cells but not control T cells (Fig.

2, C–E). Similarly, CD19+ JeKo-1 B lymphomas were killed only following administration of CD19-specific CAR T cells, but not untransduced or nontargeted EGFRvIII CAR T cells (Fig. 2 G.

And Fig. S4, A–D). These results are similar to those of xenograft studies performed in NSG mice (Fraietta et al., 2016.

O’Rourke et al., 2017) and credential the zebrafish model for accurately assessing CAR T cell responses in vivo. Next, we investigated the in vivo therapy responses and specificity to BiTE immunotherapy. BiTEs are artificial, bispecific monoclonal antibodies that contain two single-chain variable fragments, one that binds T cells through contact with the CD3 receptor while the other binds to a tumor-specific molecule, redirecting endogenous cytotoxic T cells to engage and kill cancer cells.

Here, animals were engrafted with GFP-expressing EpCAM+ OVCAR-5 ovarian carcinoma cells or chronic myelogenous leukemia K562 cells engineered to stably express human CD19 (K562-CD19). After 7 d of engraftment, animals were coinjected with (i) solitomab (EpCAM/CD3) BiTEs, (ii) blinatumomab (CD19/CD3) BiTEs, or (iii) EpCAM control antibody along with human CD8+ T cells weekly. Tumor regressions were only observed within animals administered target-specific BiTEs by 21 d after engraftment (P <.

0.001, Student’s t test. N = 8 animals/arm. Fig.

2, H–M and Fig. S4, E–H). Together, these experiments highlight the exquisitely high degree of target specificity for each BiTE and the relative ease of reading out these tumor responses in live zebrafish when compared with mouse xenograft studies (Brischwein et al., 2006.

Mølhøj et al., 2007). We next investigated the kinetics of CAR T cell infiation and engagement when encountering antigen-specific tumor cells using live animal, single-cell imaging. Specifically, 5 × 104 GFP-expressing U87 EGFRvIII+ glioma cells were engrafted into the superficial orbital musculature of rag2Δ/Δ, il2rga−/− animals.

This site is easily assessable for confocal microscopy imaging (Yan et al., 2019. Yan et al., 2020). Following engraftment for 6 d, rag2Δ/Δ, il2rga−/− animals were intraperitoneally (IP) injected with a single dose of nontargeted control T cells or EGFRvIII CAR T cells (5 × 105 cells.

Fig. 3 A). T cells were ex vivo labeled with the CFSE cell-permeable dye prior to implantation.

Although CFSE intensity decreases by half at each cell division, published studies using engraftment into NSG mice have demonstrated that human T cells undergo only one or two divisions in the first 7 dpt, permitting facile imaging of stained T cells in vivo using confocal microscopy (Quah et al., 2007. Wong and Pamer, 2001). Using this approach, we were able to quantify migration to regions adjacent the tumor by 24 h after injection with both nontargeted and EGFRvIII CAR T cells (Fig.

3, B and F. And Video 1. N = 5 animals per condition).

Yet, only EGFRvIII CAR T cells could efficiently enter the tumor mass and kill tumor cells by 5 d after therapy administration (Fig. 3, B–E. P <.

0.01, Student’s t test). High-magnification 3D volumetric modeling revealed remarkable differences in cell behavior. EGFRvIII CAR T cells rapidly infiated into the tumor and subsequently engaged with tumor cells by 5 d after T cell injection (Fig.

3, C–H. And Video 1). By contrast, nontransduced T cells migrated to the transplant site (Fig.

3 C) but did not infiate the tumor and rather aligned along the peripheral edge (n = 718 of 750 cells analyzed across all time points, n = 5 animals. Video 1). Similar differences in CAR T cell infiation and additional T cell–mediated immunotherapies were observed on sectioning of animals at the end of the experiment and costaining of samples with human CD3 and the GFP antibody (Fig.

S5). Finally, we were able to directly quantify the number of T cell–tumor cell interactions over time, showing initial engagement of CAR T cells with tumor cells by just 1 d after injection and maximal CAR T cell–tumor cell contact and subsequent tumor cell killing by 5 d after CAR T infusion (Fig. 3 H).

By 8 d after immunotherapy, CAR T cell engagement was reduced, likely reflecting near-complete ablation of tumor cells by this time point (Fig. 3, E and H). In total, these experiments provide a detailed understanding of the kinetics of CAR T cell migration to the tumor site, infiation, engagement, and subsequent tumor cell killing in vivo.

Next, we developed a facile single-cell imaging platform to rapidly assess on-target cell killing in vivo and image apoptotic immune synapse formation as an early end point analysis of cell killing. Specifically, we engineered EpCAM+ OVCAR-5 ovarian carcinoma cells to stably express both mCherry and the ZipGFP-Casp3 apoptotic cell reporter, a GFP variant that only emits fluorescence upon Casp3 cleavage of an inhibitory protease sequence inserted between β1-10 and β11 barrel of protein (To et al., 2016). MCherry+/ZipGFP-Casp3+ OVCAR-5 cells were engrafted into the periocular musculature and 6 d later IP injected with 5 × 105 CD8+ CFSE dye–labeled T cells and either (i) 50 µg/kg control EpCAM antibody or (ii) EpCAM/CD3 solitomab (Fig.

4 A). Solitomab-treated animals had decreased tumor burden over time (Fig. 4 C) and exhibited a significant increase in ZipGFP-Casp3–labeled apoptotic cells starting at 24 h after therapy compared with control EpCAM antibody–treated fish (P <.

0.01, Student’s test. Fig. 4, B and D).

Elevated numbers of ZipGFP-Casp3+ cells were observed by 24 h after treatment, preceding the overall reduction in tumor cells that was observed at 4 and 7 d after treatment with EpCAM/CD3 solitomab, suggesting that BiTE-induced cell killing required >24 h to reach maximal efficiency (Fig. 4, C and D). BiTE-induced cytotoxicity requires physical contact of T cells and tumor cells, forming apoptotic immunological synapse and release of cytolytic granules by T cells in vitro (Roda-Navarro and Álvarez-Vallina, 2020).

As expected, solitomab treatment led to high T cell migration and infiation into the tumor (Fig. 4, E, G, and H). 3D modeling of CD8+ T cells with ZipGFP-Casp3+ tumor cells allowed in vivo imaging of T cell–tumor cell contact and quantitation of apoptotic immunological synapses in vivo (Fig.

4 F). 83% of all CD8+ T cells were in direct contact with tumor cells after 7 d of solitomab treatment compared with 24% in EpCAM antibody–treated controls (P <. 0.01, Student’s t test.

Fig. 4 I and Video 2). Solitomab treatment also led to higher rates of apoptotic immunological synapse formation by 4 and 7 d of treatment (Fig.

4 J). In addition to high-resolution imaging of cell killing in vivo, these experiments rapidly assessed therapy-induced cell killing that could first be detected 4 d after BiTEs administration, providing a robust and fast assay for determining on-target in vivo killing. A high degree of target specificity is critical for any preclinical cancer immunotherapy currently in development.

As such, we next investigated the specificity of the recently described APEC immunotherapy using rag2Δ/Δ, il2rga−/− zebrafish. APECs are antibody conjugates that deliver viral antigens to the tumor surface for presentation by HLA-I, specifically redirecting preexisting CD8+ T cell antiviral immunity against tumor cells following viral peptide cleavage by tumor proteases (Fig. 5 A.

Millar et al., 2020). We first evaluated the efficiency of APEC immunotherapy by engrafting GFP-expressing EpCAM+ OVCAR-5 or EGFR+ MDA–MB-231 cells into the peritoneal cavity of rag2Δ/Δ, il2rga−/− mutant animals. Engrafted animals were then injected with CMV-specific CD8+ T cells along with (i) EpCAM control antibody, (ii) EpCAM-MMP7-CMV APEC (EpCAM-MC.

Unpublished data), or (iii) EGFR-MMP14-CMV APECs (EGFR-M14C) on days 7 and 14 after engraftment. EpCAM-MC and EGFR-M14C were generated using therapeutic EpCAM antibody (clone B38.1) and cetuximab, respectively. In particular, EpCAM-MC is a personalized APEC-immunotherapy for ovarian cancer that cleaves a CMV peptide (NLVPMVATV.

Abbreviated NLV) by tumor-expressed MMP7 and then is presented by MHCI (HLA-A*02:01). EpCAM-MC was identified as the top APEC that kills ovarian carcinoma in a xenograft screen that used high content imaging of therapy responses in rag2Δ/Δ, il2rga−/− zebrafish (unpublished data). Based on these successes, we also engineered a novel EGFR-M14C that contained the same NLV-CMV peptide but used the MMP14 cleavage.

MDA–MB-231 cells express high levels of MMP14, which has important roles in driving tumor growth, invasion, and angiogenesis in mouse xenograft studies (Devy et al., 2009). Robust tumor regressions were only observed in animals that received CMV NLV peptide-primed, MHCI-restricted CD8+ T cells along with their target-specific APECs, while control antibody or nontargeted APECs failed to curb overall tumor growth in either model (Fig. S4, I–R).

Like CAR T cells and BiTEs, our results show that APECs have exquisite specificity for killing only epitope-expressing tumors. Building on these results, we next developed a colorimetric specificity assay to directly quantify on-target cell killing at single-cell resolution and to determine the possibility for collateral killing of adjacent nonepitope-expressing cancer cells by APECs. It had been suggested that the cleaved CMV peptide might be taken up by adjacent, nonepitope-expressing cancer cells and presented on the cell surface to engage T cells, further enhancing tumor cell killing of heterogenous cell populations that lack epitope (Millar et al., 2020).

Specifically, MDA–MB-231 cells were generated to express mCherry or to coexpress both EpCAM and GFP (Fig. 5 B). Cells were mixed at 50:50 ratios and engrafted into recipient animals.

As expected, EpCAM−/mCherry+ and EpCAM+/GFP+ MDA–MB-231 cells grew at similar rates in engrafted animals when injected with both NLV peptide-primed, MHCI-restricted CD8+ T cells and control EpCAM antibody (Fig. 5, C and F). By contrast, recipient animals that received CD8+ CMV-specific T cells and EpCAM-MC APEC exhibited significant decreases in EpCAM+/GFP+ cells over time, while EpCAM−/mCherry+ tumor cell number was largely unaffected (Fig.

5, C and F). Higher-magnification 3D modeling confirmed CMV-specific T cell interactions were far more frequent with EpCAM+/GFP+ tumor cells, but less so with EpCAM−/mCherry+ cells at 11 dpt (P <. 0.001, Student’s t test.

Similar results were also seen at 14 d after engraftment. Our work indicates that APEC therapy is highly specific to killing epitope-expressing cells and unexpectedly exhibits little effect on adjacent tumor cells that do not express the antigen. Pediatric RMS is a common muscle cancer found in children and has an abysmal 5-yr survival rate of <30% in the metastatic, recurrent, and refractory disease settings (Yohe et al., 2019).

Recently, two high-risk pediatric RMS patients were independently treated with HER2+ or CD56+ CAR T cells and achieved long-term disease remission, suggesting potential opportunities to exploit T cell immunotherapy in RMS (Hegde et al., 2020. Jiang et al., 2019). Despite these amazing success stories, the HER2 and CD56 receptors are only expressed in a small fraction of RMS patients (Bahrami et al., 2008.

Ganti et al., 2006. Taniguchi et al., 2008). To systemically interrogate the most suitable antigen for this disease, we first identified all antigen targets that are currently being investigated in active sarcoma immunotherapy clinical trials and assessed if these markers are expressed in RMS (Fig.

6 A). In total, six surface antigens were identified as potential epitopes for RMS immunotherapy, including EGFR. A retrospective study using immunostaining of RMS patient samples (Dobrenkov et al., 2016.

Ganti et al., 2006. Grass et al., 2009. Majzner et al., 2019.

Saraf et al., 2019. Shibui et al., 2019. Thway et al., 2011) revealed EGFR expression in a substantial fraction of human RMS (n = 316 out of 476 primary patient samples and n = 9 of 12 cell line/PDXs.

Importantly, pediatric RMS patients had significantly higher incidence and overall expression levels of EGFR than patients with adult disease (74% vs. 50%, respectively, P = 0.03 by χ2 test. Fig.

6, E and F. And Table 1). To investigate if and which T cell immunotherapy most effectively curbs RMS tumor growth in vivo, we next performed a side-by-side comparison of EGFR CAR T cells, BiTEs, and APECs to assess differences in killing of RMS cells engrafted into rag2Δ/Δ, il2rga−/− zebrafish.

These experiments are the first to directly compare these three immunotherapy responses in vivo using the same tumor models and epitopes, allowing direct comparison of cell behavior and killing across immunoncology platforms. Specifically, 5 × 105 RD cells were IP injected into rag2Δ/Δ, il2rga−/− mutant animals, and animals were administered (i) EGFR CAR T cells, (ii) EGFR/CD3 BiTE with CD8+ T cells, (iii) EGFR-M14C with CMV-specific T cells, or (iv) control EGFR antibody with CD8+ T cells on days 7, 14, and 21 after engraftment (n = 5 animals/arm. Fig.

7 A). Notably, significant reductions in tumor burden were observed in all three immunotherapies tested, with EGFR CAR T cells eliciting a trend toward more potent cell killing by 21 d after therapy that was associated with reduced Ki67 proliferation marker expression, higher numbers of TUNEL+ apoptotic cells, and subsequently fewer tumor cells per area on section (Fig. 7, A–D).

To further understand the kinetics of cell killing and possible differences in T cell infiation and engagement between the three therapies, we next engrafted 5 × 104 ZipGFP-Casp3–expressing RD cells into the periocular muscle and used confocal imaging to quantify T cell infiation and cell apoptosis following administration of each T cell immunotherapy (Fig. 7, E–I). Among the three therapies analyzed, EGFR/CD3 BiTEs induced the most robust T cell infiation (Fig.

7, E and H). Yet despite fewer CAR T cells entering the tumor, they elicited the most effective tumor cell killing and subsequent reductions in tumor cell number over time (P <. 0.02, Student’s t test.

Fig. 7, F and G), suggesting higher overall therapy efficacy of CAR T cell immunotherapies. Indeed, high-resolution 3D modeling of T cell–tumor cell interaction revealed a higher percentage of T cells could induce tumor cell killing by CAR T cells compared with BiTE- and APEC-treated animals (n = 5 animals/arm.

7 I). Finally, although APEC-treated cells had less infiative capacity to enter tumors (Fig. 7 H), both BiTEs and APECs had similar potency in killing tumor cells following infiation into the tumor mass (Fig.

7 I). We next confirmed EGFR CAR T cell responses in a wider array of RMS tumors including both fusion-positive and fusion-negative subtypes of RMSs. As was seen in RD RMS xenografted cells, EGFR CAR T cells efficiently infiated into the tumor and effectively killed RMS tumor cells over time in three additional xenograft models irrespective of disease subtype (n = 5 zebrafish/arm.

8, A–C). As was seen in other CAR T cell indications tested in our work, there was an overall lower percentage of CAR T cells engaged with tumor at any given time compared with BiTE and APEC treatment. Yet these CAR T cell therapies efficiently killed tumors over longer periods of time.

These results again highlight differences in the rates of cell killing by these modalities. Importantly, EGFR CAR T cell immunotherapy was also effective in suppressing RMS xenograft growth in NSG mice (n = 6 mice/arm. P <.

0.05 over multiple time point analysis. Fig. 8 D).

Together, our zebrafish and mouse xenograft studies provide much-needed preclinical rationale for assessing a wide array of EGFR-targeted immunotherapies in RMS. Our work has established the rag2Δ/Δ, il2rga−/− zebrafish as a powerful model for assessing T cell immunotherapy responses at single-cell resolution, successfully imaging single-cell therapy responses in 10 cancer xenograft models across eight distinct T cell immunotherapies. Recently, others have leveraged the zebrafish larval cell transplantation platform to model the effects of tumor-infiating lymphocytes and CD19 CAR T cells in human cancer xenografts (He et al., 2020.

Pascoal et al., 2020). Yet, these larval xenotransplant studies were limited to engraftment of only a few hundred cells per animal, animals could not be reared at physiological 37°C, and assay times were restricted to 24 h in both studies. By contrast, our studies demonstrated the utility of the adult immunocompromised zebrafish model in a wide array of immunotherapies including CAR T cells, BiTEs, and APECs.

We also optimized intravital imaging approaches to quantitatively assess T cell migration and infiation into the tumor mass and tumor cell killing in real time and document target specificity serially over days. Our studies also demonstrated that most efficient T cell immunotherapy–induced cell killing falls between 24 and 96 h after administration, time points that would not be evaluable using the larval xenograft models. The adult immune-deficient zebrafish model is more akin to xenograft studies that use NSG mice, where longer time course studies allow for assessment of immune-therapy responses for up to 3 wk.

Mouse xenograft studies have led to many important insights into mechanisms of T cell homing, infiation, and tumor killing (Boissonnas et al., 2007. Boulch et al., 2021. Cazaux et al., 2019.

Halle et al., 2016. Hoekstra et al., 2020. Mulazzani et al., 2019).

Yet, the zebrafish xenograft model addresses several fundamental difficulties of completing similar studies using mouse xenografts. For example, imaging T cell/tumor cell responses in mouse xenografts is more technically challenging and requires complex multi-photon microscopy and prior knowledge of where to image based on surgical window creation. Mice are also more expensive.

Thus, large-scale studies are limited. Despite the perceived powers of the zebrafish xenograft model, it also has limitations. For example, xenografted zebrafish were unable to be imaged longer than 10 min in our studies, mainly attributed to gill respiration movements around the periocular engraftment site, precluding a more detailed time-lapse study of T cell movement and subsequent tumor cell killing.

These technical hurdles will likely be addressed by the development of deep gill-perfusion anesthesia techniques, selection of alternative engraftment sites such as the dorsal musculature, and refined imaging platforms in the future. Despite the many advantages of engrafting human tumors and assessing immunotherapy responses in adult rag2Δ/Δ, il2rga−/− zebrafish and NSG mice, both models lack endogenous T, B, NK, and macrophage cells, precluding assessment of how these important immune cells modulate the tissue microenvironment and modify responses to T cell therapy. New immune-deficient zebrafish models will likely incorporate transgenic approaches to express human cytokines that support human blood cells and allow for further humanization, akin to knock-in and transgenic mouse models such as the humanized M-CSFh/h, IL-3/GM-CSFh/h, SIRPah/h, TPOh/h, RAG2−/−, IL2Rg−/− mice (Das et al., 2016.

Rongvaux et al., 2014. Wunderlich et al., 2010. Wunderlich and Mulloy, 2016).

Such models would permit engraftment of both the human immune system and tumor to follow more complex immune cell interactions. In fact, elegant work from the Berman group has already developed transgenic fish that express human GM-CSF, stem cell factor, and stromal derived factor 1α and permit short-term engraftment of human hematopoietic stem cells and leukemia cells into larval fish, providing an important starting point for such studies (Rajan et al., 2020). At a cell biological level, our work demonstrated important differences in early responses between CAR T cell, BiTE, and APEC immunotherapies.

For example, single-cell quantitative studies revealed that CAR T cells engaged with tumor cells by just 24 h after engraftment in most xenograft models, while neither BiTE- nor APEC-treated T cells exhibited such robust early responses. CAR T cell–tumor cell interactions also led to elevated overall tumor cell killing as read out by an overall reduction in tumor cell number and higher percentage of apoptotic synapse formation between CAR T cell–tumor cells compared with BiTE- and APEC-treated T cells. Although our work showed a direct correlation between T cell/tumor cell contact and killing, recent work has also demonstrated bystander/indirect killing trough cytokines secreted by T cells in the absence of direct contact between tumor cell and T cells, suggesting multiple possible modes of tumor cell killing (Hoekstra et al., 2020).

In total, differences in T cell engagement and cytotoxicity observed in our work are likely accounted for by the fundamentally different mechanisms by which CAR T cells, BiTEs, and APECs bind to tumor cells, induce activation, and kill. For example, CAR T cells engage tumor cells through a chimeric antigen receptor scFv domain and are primed for activation by ex vivo stimulation (Rafiq et al., 2020). Thus, robust and fast tumor cell killing would be expected.

T cell receptor/antigen binding then activates CAR T cells, releasing perforin, granzyme, and apoptosis-inducing cytokines. In contrast, BiTEs mechanically link nonstimulated T cells with tumor cells by simultaneously binding a tumor-specific antigen and the CD3 receptor on T cells (Huehls et al., 2015). Antibody/CD3 binding then activates T cells, eliciting endogenous T cell effector responses.

Finally, APECs harness the antiviral CD8+ T cell responses by loading viral epitopes onto tumor-expressed MHCI, promoting the formation of MHC/TCR complexes with viral-specific T cells and subsequent activation (Millar et al., 2020). The processes by which BiTEs and APECs induce T cell activation would be predicted to require longer interaction times between the T cell with tumor before inducing cell death compared with CAR T cells. In total, our observations are largely mirrored by in vivo mice xenograft studies targeting NY–ESO-1 antigen on human myeloma cells or 237scFV on mice fibrosarcoma cells (Maruta et al., 2019.

Stone et al., 2012). These studies performed a side-by-side comparison of CAR T cell and BiTE cell killing efficacy using mouse models and in both instances showed CAR T cells to be superior at killing tumor cells over long–time course experiments. However, these murine models lacked the resolution to visualize tumor cell–immune cell interactions at single-cell resolution and hence failed to provide mechanistic insights into the early kinetic differences between T cell–based therapies and linking of these cell biological differences with effects on tumor cell killing.

Our work also validated the efficacy and target specificity of the new APEC technology to kill antigen-expressing tumor cells. First, we demonstrated the feasibility and in vivo utility of designing APECs that express tumor-specific protease cleavage sites in the context of ovarian carcinoma (unpublished data). This EpCAM-MC APEC was created by conjugating an MMP7 protease cleavage site (AVSRLRAYNLVPMVATV) with a CMV-specific viral epitope NLV to the therapeutic EpCAM antibody (Clone 38.1.

Unpublished data). Because APECs bind to tumor-specific epitopes and only release their viral peptide cargo locally following cleavage by tumor-secreted proteases, target specificity is expected to be enhanced over other T cell immunotherapies, including traditional CAR T cells and BiTEs. APEC approaches thus allow exquisite redirection of endogenous viral CD8+ T cells to kill seemingly virally infected tumor cells.

Here, we validated the remarkable specificity for epitope-expressing tumor cells with little collateral killing of nonepitope-expressing tumor cells. Building on these successes, we also developed a new EGFR-M14C APEC that efficiently killed RMS cells. In total, our work has shown that APECs are a new and powerful immunotherapy that will be valuable for tailoring personalized immunotherapy, pairing tumor-specific epitope expression with tumor-specific protease expression and cleavage.

Our work also has established much needed preclinical modeling for T cell–mediated immunotherapy in RMS and identified EGFR-based immunotherapies as a new approach to kill RMS tumor cells. EGFR is expressed in a large fraction of RMS patients, and clinical trials using EGFR immunotherapies have reported tolerable side effects (Guo et al., 2018). Despite achieving remissions in a large fraction of pediatric RMSs, relapse, refractory, and recurrent metastatic diseases are the major clinical challenge facing patients.

Current treatment strategies rely on aggressive treatment that includes radiation, chemotherapy, and surgery (Yohe et al., 2019). Targeted immunotherapy approaches could prove powerful for the treatment of aggressive RMS tumors, especially in the relapse setting and for those tumors that cannot be easily resected. Indeed, two pediatric RMS patients with refractory metastatic and recurrent disease achieved complete remissions following HER2+ and CD56+ CAR T therapy (Hegde et al., 2020.

Jiang et al., 2019), suggesting a clinical path forward for T cell therapies in this disease. Our zebrafish and mouse xenograft studies have provided a strong preclinical rationale for assessing EGFR CAR T cell and other immunotherapies in this disease. rag2Δ/Δ zebrafish were created in the Casper-strain zebrafish using CRISPR/Cas9 genome editing (Hwang et al., 2013).

Briefly, two guide RNA (gRNA) sequences (5′-AGA​ACC​GTA​TCA​AGC​GCG​GG-3′ and 5′-GGC​CCT​TGA​CTA​CAT​ATG​GTG-3′) were designed to flank the 3.3 kb of the rag2 gene locus using CHOPCHOP. GRNAs were cloned into the DR274 vector (Addgene) and transcribed in vitro using the T3 mMESSAGE mMACHINE Kit. A mixture of 300 ng/µl of Cas9 mRNA and 100 ng/µl of gRNA was coinjected into one cell–stage zebrafish embryos.

F0 animals were incrossed, and progeny were assessed for gene deletion using PCR of genomic DNA. WT gene-specific primers amplified a 444-bp fragment of the endogenous rag2 gene (forward 5′-CCA​TAT​AGC​CAA​TTA​CTA​C-3′, reverse 5′-GCA​GAT​CTG​GAT​CTG​GAG​T-3′. Denaturing.

94°C/30 s, annealing. 60°C/30 s, elongation. 68°C/60 s, and termination.

68°C/5 min). Mutant gene–specific primers (forward 5′-CCC​ATC​TAT​GGG​AAA​CTA​TC-3′, reverse 5′-GTG​TCA​CAT​GAT​CCT​TCA​G-3′) spanned the genomic deletion and amplified a 973-bp fragment (denaturing. 94°C/30 s, annealing.

60°C/30 s, elongation. 68°C/60 s, 36 cycles, termination. 68°C/5 min.

Fig. S1, A–C). Il2rgaY91fs-specific primers (forward 5′-TTT​GAC​ATC​GAA​GAC​TGT​CCT​G-3′, reverse 5′-GTC​CTG​TAA​CGA​ACT​TCG​CTC​T-3′) spanned the genomic mutation, amplifying a 373-bp WT allele and a 360-bp mutant allele (denaturing.

94°C/30 s, annealing. 60°C/30 s, elongation. 68°C/30 s, 36 cycles, and termination.

68°C/5 min). Male rag2Δ/Δ, Il2rgaY91fs/+ adult Casper (double mutant for roya9/a9 and nacrew2/w2) zebrafish were crossed to female rag2Δ/+, il2rgaY91fs/+ adult Casper zebrafish. Progeny were grown to adulthood and then subjected to scale resection genotyping at 2–3 mo of age as previously described (Yan et al., 2019).

Homozygous mutant rag2Δ/Δ, il2rga−/− fish were produced at that expected Mendelian ratio (11.75%, n = 984 of 8,374 fish, expected 12.5%). CD8+ T cells used as control for BiTE experiments were purchased (Stem Cell Technology. #70027), cultured, and expanded using RPMI medium (10% FBS and 1% penicillin/streptomycin) supplemented with 20 IU/ml of rhIL-2.

CAR T cells were generated using primary donor T cells transduced with the anti-CD19 or EGFRvIII CAR containing a 4-1BB intracellular signaling domain and expanded as previously reported (Milone et al., 2009). Briefly, donor T cells were thawed and activated using α-CD3/α-CD28 Dynabeads (Life Technologies) at a 1:3 T cell:beads ratio. The cells were cultured in RPMI medium (10% FBS and 1% penicillin/streptomycin) supplemented with 20 IU/ml of rhIL-2.

Every 2 d, fresh medium was added to keep the cells at a concentration of 0.5–2 × 106/ml. For CAR T cell production, lentiviral vector was added to the culture 24 h after activation. In parallel, donor-matched T cells that had been activated but untransduced were expanded to serve as a negative control in subsequent experiments.

At days 12–14 of culture, CAR expression was determined. NLV-specific T cell lines were created from CMV-seropositive peripheral blood mononuclear cells isolated from healthy donors upon stimulation with 10 μg/ml CMV peptide HLA-A*02:01 restricted NLV (Genscript) in RPMI1640 supplemented with 10% FBS, 100 U/ml each of penicillin and streptomycin, 4 mM of L-glutamine, and 1% human AB serum (Sigma). On day 4, half culture medium was replaced with the same medium plus 500 IU/ml IL-2 (Peprotech) and changed twice weekly.

NLV-specific CD8+ T cells were identified by flow cytometry with staining of HLA–peptide tetrameric complex on day 14–21. Only those T cell lines with >50% NLV-specific CD8+ T cells were qualified for the next functional assay for a maximum of 6 wk. Imaging of periocular transplanted cells was performed using an inverted LSM 710 confocal microscope (Zeiss) with Zen software platform (Zeiss), as previously described (Yan et al., 2019.

Yan et al., 2020). Engrafted zebrafish were anesthetized using low-dose 0.01% tricaine (Western Chemical), placed onto a 36-mm glass-bottom dish (Thermo Fisher Scientific. #150680), and the fish torso was embedded in 1% low–melting point agarose (Thermo Fisher Scientific) to stabilize the animal for imaging.

To keep the animal under anesthetic during imaging, zebrafish were submerged in 5 ml of warm 37°C fish water containing 0.01% tricaine. Serial z-stack imaging was performed using a 10× objective (numerical aperture, 0.45), achieving an overall 100× magnification. GFP- and mCherry-expressing engrafted animals were imaged using the 488-nm (emission = 493–586 nm) and 546-nm laser (emission = 575–703 nm).

All T cells used in this study were stained with 1 µM of ViaFluor CFSE before transplantation and imaged using a 405-nm laser (emission = 350–470 nm). In vivo cell apoptosis of ZipGFP-Casp3 imaging studies were imaged used a 488-nm (emission = 493–586 nm) and 546-nm laser (emission = 575–703 nm). Images were automatically annotated and counted in Imaris.

Total T cell, tumor cell, and apoptotic cell (ZipGFP-Casp3+) numbers were quantified using Imaris spot and surface functions. Quantification was completed on z-stack images with dimensions 1,000 µm × 1,000 µm × 100 µm (0.1 mm3). 3D modeling analysis quantifying absolute distance between T cells and tumor cells with direct cell-to-cell contact was defined by <10 µm between cells.

This analysis was completed on z-stack tumor images sampled at 500 µm × 500 µm × 50 µm (0.0125 mm3) and analyzed using the distance transformation, distance between spot to surface, and spot close to surface XTension functions. Apoptotic cells were pseudo-colored yellow using Imaris surface function for easy visualization in Fig. 7 E.

Fluorescent-labeled human cancer cell lines (U87, JeKo-1, OVCAR-5, K562, MDA–MB-231, RD, SMS-CTR, Rh41, and Rh30) were transplanted IP (5 × 105 cells/animal) or periocularly (5 × 104 cells/animal) into recipient fish. Stably engrafted fish were IP injected with CAR T cells (5 × 105 cells/dose). For BiTE experiments, animals were coinjected with 5 × 105 CD8+ T cells along with control antibodies (EpCAM, 50 µg/kg.

CD19, 50 µg/kg, or EGFR, 50 µg/kg), solitomab (50 µg/kg), blinatumomab (250 µg/kg), or EGFR/CD3 BiTE (10 µg/kg). For APEC experiments, animals were injected with 5 × 105 CMV-specific T cells, along with control antibodies (EpCAM, 50 µg/kg, or EGFR, 50 µg/kg), EpCAM-MC (50 µg/kg), or EGFR-M14C (50 µg/kg). All IP injections were completed using a 30 1/2–G needle (BD) at the time points noted for specific experiments.

At the end of the experiment, animals were fixed in 4% paraformaldehyde, sectioned, and examined histologically by H&E staining, immunohistochemistry (IHC) for Ki67, and TUNEL. Comparisons between groups were performed using ANOVA followed by Student’s t test. Engrafted zebrafish were fixed in 4% paraformaldehyde, embedded in paraffin, and sectioned at 5-mm thickness.

Sections were stained by H&E, IHC, or immunofluorescence. For IHC staining, the primary antibodies were rabbit monoclonal anti-Ki67 (1:1,000 dilution. Abcam), monoclonal anti-CD3 (1:100 dilution.

Abcam), monoclonal anti-GFP (1:100 dilution. Abcam), monoclonal anti-EGFR (1:100 dilution. Abcam), and TUNEL (1:1,000 dilution.

Thermo Fisher Scientific). Secondary antibodies were biotinylated goat anti-rabbit IgG antibody (Vectorlabs) and horse anti-mouse antibody (Vectorlabs). Development was completed using Vectastain ABC Kit (Vectorlabs) or Alexa Fluor 488 anti-mouse and Alexa Fluor 546 anti-rabbit secondary antibody (1:1,000 dilution for all secondary antibodies).

Quantification was completed based on counting three randomly selected fields imaged at 400× using an Olympus BX41 microscope. Quantification used ImageJ and was blinded. Images were counted without labels by Eric Alpert.

Samples were analyzed using a fixed threshold for achieving an unbiased, quantitative assessment of the IHC and TUNEL staining within the selected imaged field (Yan et al., 2020). Percentage of proliferating (Ki67) and apoptotic (TUNEL) cells was calculated by dividing the number of positively stained cells by the total number of cells counted within the selected fields. N >.

200 control cells were analyzed per sample, with fewer cells being counted for treated samples. H&E-stained sections were imaged at 400× and quantified as number of cells per unit area. We thank Dr.

Len Zon for helpful discussion and feedback on the work. This work is supported by National Institutes of Health grants R24OD016761 (D.M. Langenau), R01CA154923 (D.M.

Langenau), R01CA215118 (D.M. Langenau), and R01CA211734 (D.M. Langenau).

The Liddy Shriver Sarcoma Initiative (D.M. Langenau). The Massachusetts General Hospital Research Scholars Program (D.M.

Langenau), U01CA220323 (N.J. Dyson). The Tosteson &.

Fund for Medical Discovery Fellowship from Massachusetts General Hospital (C. Yan). And the Alex’s Lemonade Stand Foundation Young Investigator Award (C.

Yan and D.M. Langenau conceived, designed, and conducted the study. Analyzed data.

Wei, and K.M. McCarthy assisted with experimental design and execution. S.

Iyer and Q. Qin performed bioinformatics analysis studies. J.F.

Maus provided important intellectual contributions, designed experiments, and provided reagents, cell lines, and/or PDXs. Competing Interests Disclosures. D.G.

Millar reported personal fees from Revitope Inc. During the conduct of the study and personal fees from Revitope Inc. Outside the submitted work.

In addition, D.G. Millar had a patent to WO2012123755A1 licensed "Revitope Inc." and owns stock in Revitope Inc. B.J.

Drapkin reported personal fees from AstraZeneca outside the submitted work. M. Cobbold reported "other" from Revitope Oncology outside the submitted work.

In addition, M. Cobbold had a patent to WO 2012/123755 issued "Revitope Oncology". And is currently an employee of Astrazeneca.

M.V. Maus is an inventor on patents related to adoptive cell therapies, held by Massachusetts General Hospital and University of Pennsylvania (some licensed to Novartis). M.V.

Maus holds equity in TCR2, Century Therapeutics, and Neximmune, and has served as a consultant for multiple companies involved in cell therapies. In addition, M.V. Maus is a consultant for Adaptimmune, Agenus, Allogene, Arcellx, Astellas, AstraZeneca, Atara, Bayer, BMS, Cabaletta Bio (SAB), Cellectis (SAB), CRISPR therapeutics, In8bio (SAB), Innovakine, Intellia, GSK, Kite Pharma, Micromedicine, Neximmune, Novartis, TCR2 (SAB), Tmunity, Torque, and WindMIL (SAB).

Has grant/research support from CRISPR therapeutics, Kite Pharma, Servier, and Novartis. Is a stockholder of Century Therapeutics, TCR2, and Ichnos. And is a member of board of directors for Ichnos Sciences.

D.M. Langenau reported a patent pending on use of immune deficient zebrafish for xenograft transplantation studies. No other disclosures were reported..

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