Get lasix prescription online

The Supreme Court upheld the Affordable get lasix prescription online Care Act today in a 7-2 ruling. The court dismissed a challenge to the law, noting that the states and individuals who were trying to overturn the ACA did not have standing. This is get lasix prescription online the third time the ACA has survived challenges in the Supreme Court.

In 2012, the ruling was 5-4, and in 2015, the ruling was 6-3. These cases have all had varying arguments and merits, but it’s noteworthy that although the court has become more conservative over the last decade, the justices have increasingly favored the ACA. In this year’s case, some legal analysts had speculated that the court might overturn the get lasix prescription online ACA’s individual mandate but allow it to be severed from the rest of the ACA.

That approach would have upheld the ACA as well, but the court simply dismissed the whole case. (This thread from get lasix prescription online Nicholas Bagley is a great summary, if you’re interested in the specifics.) So nothing has changed. The ACA remains intact, and the general consensus is that it’s here to stay.

Is this decision the end of legal challenges to the ACA?. That doesn’t mean the Affordable Care Act won’t continue to face legal challenges — a case that’s currently under consideration in Texas get lasix prescription online takes aim at the ACA’s requirement that health plans fully cover the cost of certain preventive care. But that case does not seek to overturn the ACA itself, and it appears unlikely that the Supreme Court would take up any other case that might aim to do so.

What does this decision mean for consumers?. There was a collective sigh of relief this morning among people who are enrolled in Medicaid under the ACA’s expanded eligibility guidelines, as well as those who purchase their own individual/family health insurance and rely on the ACA’s premium tax credits, cost-sharing reductions, guaranteed-issue rules and coverage for pre-existing conditions, and essential get lasix prescription online health benefits. According to a recent analysis by Charles Gaba, more than 10% of all Americans are covered under Medicaid expansion, ACA-compliant individual/family health plans, and Basic Health Programs, all of which stem directly from the ACA.

As we’ve explained during prior legal and legislative challenges get lasix prescription online to the ACA, the law provides a vast array of additional consumer protections that extend to most Americans in one way or another. But the people who are most likely to feel a sense of relief today are those enrolled in coverage that either wouldn’t exist or wouldn’t be accessible to them without the ACA. The anxiety about losing health coverage is no longer hanging over these Americans.

Premium subsidies will continue to be available, and the subsidy enhancements provided by the American Rescue Plan will continue to be in effect throughout get lasix prescription online 2022 – and possibly longer, if Congress acts to extend them. If you’ve been on the fence about enrolling in individual/family coverage during the special enrollment period that’s currently ongoing in nearly every state, you can now enroll with confidence. And the same is true about signing up for 2022 coverage when open enrollment starts in November.

And although today’s ruling was on a lawsuit that hinged around the individual mandate and penalty, nothing has changed about the ACA’s requirement that get lasix prescription online most people maintain health insurance. There continues to be no federal penalty for not having health insurance, as has been the case since 2019. (If you’re in California, Massachusetts, New Jersey, Rhode Island, or the District of Columbia, there’s still a penalty for going without health insurance.) What does the decision get lasix prescription online mean for health insurers?.

Insurers that offer individual/family health insurance have been displaying increasing confidence in the ACA for the last few years. After fleeing the marketplaces/exchanges in 2017 and 2018, insurers started to join or rejoin the marketplaces in 2019. That trend continued in 2020 get lasix prescription online and 2021, and we’re already seeing more insurer participation in the initial 2022 rate proposals that have been submitted by insurers in several states.

The case that the Supreme Court dismissed today was initially filed in early 2018, so the legal threat to the ACA has been in the background throughout those three years of increasing insurer participation in the ACA-compliant insurance market. Although insurance companies — and the actuaries who set premiums — tend to get lasix prescription online be quite averse to uncertainty, the individual market has proven to be profitable for insurers in recent years (after being unprofitable in the early years of ACA implementation). Insurers’ increasing willingness to offer plans in the marketplace is testament to that, despite the uncertainty that the lawsuit created over the last few years.

Now that there’s no longer a pending legal threat to the ACA, we might see even more insurers opting to join the marketplaces or expand their existing coverage areas. What does the decision mean get lasix prescription online for states?. Although many states have enacted laws designed to protect consumers in case the ACA had been overturned, there’s no getting around the fact that they rely heavily on federal funding that’s provided under the ACA.

Without that funding, most states would not have been able to maintain the ACA’s Medicaid expansion or affordability provisions for self-purchased health insurance. There’s no longer a threat to the funding, which might make states more get lasix prescription online likely to push forward with additional consumer protections tied to the ACA. Among the most obvious is Medicaid expansion in the 13 states that have not yet accepted federal funding to expand Medicaid eligibility under the ACA.

The American Rescue get lasix prescription online Plan provides two years of additional federal funding to states that newly expand Medicaid. So far, Oklahoma is the only state making use of that provision, and the state had already planned to expand Medicaid this year as a result of a ballot measure that Oklahoma voters passed last year. To be fair, the other 13 states have rejected Medicaid expansion year after year, including during the 2020 and 2021 legislative sessions that took place during a global lasix.

Without a change to the makeup of their legislatures, most get lasix prescription online are likely to continue to do so. But now that the Supreme Court has upheld the ACA yet again, states that newly expand Medicaid can do so without a lingering worry that the federal funding might be eliminated. It’s also possible that more states might consider reinsurance programs that make use of the ACA’s 1332 waiver provisions.

But that would also depend on whether the American Rescue Plan’s subsidy get lasix prescription online enhancements are extended beyond 2022. Reinsurance programs make coverage more affordable for people who don’t receive premium subsidies. Before the ARP eliminated the “subsidy cliff” for 2021 and 2022, the lack of affordability get lasix prescription online for households earning a little more than 400% of the poverty level was a very real problem.

But that’s not currently an issue, as those households qualify for subsidies if the benchmark plan would otherwise cost them more than 8.5% of their income. If Congress extends that provision, reinsurance programs would help very few enrollees (and they can also harm subsidized enrollees in some areas, since they reduce the size of premium subsidies). State legislatures will need to keep an eye on how this plays out at the federal level, but without an extension of the ARP’s subsidy structure, we can get lasix prescription online expect to see more states pursuing 1332 waivers for reinsurance programs in the next few years.

Louise Norris is an individual health insurance broker who has been writing about health insurance and health reform since 2006. She has written dozens of opinions and educational pieces about the Affordable Care get lasix prescription online Act for healthinsurance.org. Her state health exchange updates are regularly cited by media who cover health reform and by other health insurance experts.The American Rescue Plan Act (or American Rescue Plan), signed into law by President Biden on March 11, provided many types of relief to Americans from the economic ravages of the hypertension medications lasix.

Among them, the American Rescue Plan (ARP) put the “affordable” in “Affordable Care Act” for millions of Americans. Did ARP get lasix prescription online make coverage more affordable at all income levels?. The American Rescue Plan increased premium subsidies at all income levels for health plans sold in the ACA marketplaces, reducing the percentage of income that enrollees have to pay for the “benchmark” plan in their area – that is, the second-cheapest Silver plan.

The Supreme Court upholds the Affordable Care Act. What it means get lasix prescription online for policyholders. At incomes up to 150% of the Federal Poverty Level ($19,140 for an individual, $39,300 for a family of four), the benchmark plan is free, and from 150% up to 200% FPL ($25,520 for an individual, $52,440 for family of four), benchmark Silver costs no more than 2% of family income.

Silver plans at these income levels get lasix prescription online come with strong cost-sharing reduction (CSR) that reduces deductibles and out-of-pocket costs. Weaker CSR is available up to 250% FPL. At the other end of the income scale – 400% FPL or higher ($51,040 for an individual, $104,800 for a family of four) – no citizen or legally present noncitizen who lacks access to other affordable insurance (e.g., from an employer or Medicare) will pay more than 8.5% of income for benchmark Silver.

The ARP get lasix prescription online removed the ACA’s notorious subsidy cliff, which denied subsidies to applicants with incomes over 400% FPL. In the in-between income brackets, the percentage of income required for a benchmark Silver plan has also been sharply reduced. See this post for illustrations of how ARP will reduce premiums for people at various income levels.

The American Rescue Plan also effectively made free high-CSR Silver plans free to anyone who received any unemployment insurance compensation get lasix prescription online in 2021 and lacked access to other affordable insurance. The ARP subsidy boosts are temporary, running through 2022. But Democrats are widely expected to make them permanent get lasix prescription online in subsequent legislation.

That’s the first and most basic item on their healthcare agenda, fulfilling a core promise President Biden made during the 2020 campaign. ARP subsidies make it a great time to buy new health coverage The ARP subsidy increases should induce millions of uninsured Americans who have been under the impression that health insurance is unaffordable to take a second look. According to estimates by the Kaiser Family Foundation (KFF), as of 2020, only about half of those who were eligible for marketplace subsidies and in need of insurance get lasix prescription online were enrolled.

KFF estimates that 11 million uninsured Americans are eligible for premium subsidies in the marketplace – including 3.5 million with incomes over 400% FPL who were ineligible prior to the ARP. How affordable is affordable? get lasix prescription online. According to KFF, 6 million uninsured people are eligible for free plans.

It’s true that for most of these (4.7 million), the free plan would be Bronze, with deductibles averaging in the $7,000 range. But for many of those eligible for free Bronze plans, Silver – and in some cases Gold plans – get lasix prescription online are available at very low cost or even no cost at all. For solo enrollees in the 150-200% FPL income range (topping out at $25,520), benchmark Silver (with strong CSR) can’t cost more than $43 per month.

In many cases, the cheapest Silver plan costs considerably less than the benchmark. And in about 20% of get lasix prescription online all U.S. Counties, the cheapest Gold plan is cheaper than the cheapest Silver.

That’s a valuable discount at incomes get lasix prescription online above 200% FPL, where CSR, which attaches only to Silver plans, is weak (in the 200-250% FPL income range) or not available (at incomes above 250% FPL). Biden administration opens the doors and sounds the horn Prior to the American Rescue Plan’s passage – beginning on February 15 – the Biden administration opened an emergency special enrollment period (SEP), extending until August 15 in the 36 states that use the federal ACA exchange, HealthCare.gov. The 15 state-run exchanges (including Washington, D.C.) followed suit, though the terms and length of the state SEPs vary somewhat.

(See SEP deadlines for each get lasix prescription online exchange here.) The SEP offered by HealthCare.gov and in most states is akin to the annual open enrollment period. Anyone who lacks insurance can enroll. Normally, a person seeking coverage outside of open enrollment has to apply for a personal SEP and document a qualifying “life change,” such as loss of employer-sponsored insurance.

After the ARP’s get lasix prescription online passage, HealthCare.gov further opened the SEP to enable current enrollees to switch plans – for example, to upgrade from Bronze to Silver in light of the enriched subsidies. The Center for Medicare and Medicaid Services (CMS) also earmarked $50 million to advertise the SEP. The upgraded subsidies, retroactive to January 1, went live on HealthCare.gov on April 1, and on state-based get lasix prescription online marketplaces in subsequent weeks.

All in all, doors to coverage for the uninsured were flung significantly wider this spring – and remain open. Many consumers are capitalizing on the SEP and ARP The emergency SEP and upgraded subsidies are having an impact. On May 6, CMS announced that new get lasix prescription online plan selections from February 15 through April 30 in 36 HealthCare.gov states was just shy of 940,000 – almost quadruple enrollment in the same period in 2019, the last “normal” year.

(In 2020, the lasix also stimulated increased enrollment, totaling 391,000 in the same time period.) A large percentage of new enrollees were apparently low-income and accessing free or near-free Silver plans with strong CSR, as the median deductible for new enrollees was just $50. As of get lasix prescription online June 5, SEP enrollment in HealthCare.gov states had topped 1 million, and marketplace coverage is now at an all-time high. Including the 15 state-based marketplaces raises the SEP enrollment total this spring to 1.5 million, according to Charles Gaba’s estimate.

The percentage of subsidy-eligible potential enrollees who actually do enroll may now be closer to 60% than the roughly 50% that KFF estimates indicate in 2020. How might get lasix prescription online enrollment be boosted further?. But millions still aren’t on board Despite the substantial gains achieved in recent months, some 10 million of the still-uninsured are likely eligible for marketplace subsidies, and another 6 to 7 million eligible for Medicaid, according to KFF estimates.

Since the ACA’s programs were first implemented in 2014, many of the uninsured have claimed that they found coverage unaffordable, While some may have balked at subsidized premiums and available plans’ out-of-pocket costs, a lack of knowledge about what’s on offer has always been a major factor. In 2020, only 32% get lasix prescription online of people surveyed by KFF knew that the ACA was still law. The Trump administration didn’t make it easier for consumers, cutting federal funding for enrollment assistance by nonprofit “navigators” by 84%, from a peak of $63 million in 2016 to $10 million by 2018, and cutting advertising by 90%.

Navigator organizations, established by the ACA to be nerve centers in a constellation of nonprofit assistor groups, have operated on shoestrings since fall 2017, cutting back on outreach events, offices get lasix prescription online throughout their states, and in-person as opposed to phone or video assistance. The Biden administration threw a quick $2.5 million to navigators this spring – which doesn’t go far – and has allocated $80 million for navigators in the 36 states using HealthCare.gov for 2022. (Navigator funding is drawn from user fees charged to participating insurers, so the 15 states that run their own exchanges have their own funding base for enrollment assistance).

A KFF analysis suggests that the $80 million get lasix prescription online allocation for 2022 may be too modest. Trump administration underspending of the user fee revenue has left some $1.2 billion available to the Biden administration to boost enrollment efforts. Promising strategies to boost enrollment Going forward, further innovation might boost marketplace enrollment.

Maryland, which has a state-based marketplace, has pioneered an enrollment jump-start tied to tax filing, whereby the uninsured whose reported income and insurance status indicate they are eligible for subsidized coverage can check a box on their tax return and receive information about their likely eligibility for get lasix prescription online “free or low cost coverage.” Colorado will debut a similar program next year. On a national level, aligning the annual open enrollment period with tax filing season and porting information on the tax return to a marketplace application could streamline the enrollment process. Tax preparers could be a powerful resource to encourage enrollment and assist in get lasix prescription online the often complex application process.

Integrating enrollment with tax preparation could also take some of the diceyness out of the income estimate that determines subsidy size. Switching the OE period would entail a messy transition, as plans not resetting on January 1 as in the past would create problems with deductibles and out-of-pocket caps. An alternative would be get lasix prescription online to mirror Maryland and offer the uninsured an easy-to-obtain SEP at tax time.

The ARP hasn’t helped everyone It should be acknowledged that the ARP did not ease the plight of poor and near-poor uninsured people in the 12 states that to date have refused to enact the ACA Medicaid expansion (or, in the case of Wisconsin, enact a more limited expansion). As first enacted, the ACA offered Medicaid to all citizens get lasix prescription online and most legally present non-citizens whose household income was below 138% FPL. In 2012, the Supreme Court made that expansion optional for states.

In states that refused to expand eligibility – including high-population states Texas and Florida – most adult residents with incomes below 100% FPL are eligible neither for Medicaid nor for marketplace subsidies. The ARP provided new financial enticements for the holdout states to implement get lasix prescription online the expansion, but offered no immediate relief to an estimate 2 million people in this “coverage gap.” The ARP also did not fix the “family glitch,” which puts health coverage out of reach for several million Americans. If an employee has access to a comprehensive employer-sponsored health plan that meets the ACA affordability standard for single coverage, the other family members are not eligible for subsidies in the exchange — regardless of how much they have to pay to join the employer-sponsored plan.

Bottom line While more remains to be done to make affordable coverage more universally available, comprehensive and easy to obtain, it’s fair to say that most Americans who lack coverage at present can find a health plan (marketplace or Medicaid) that’s worth having at a price they can afford. If you are uninsured, check out your options on HealthCare.gov or get lasix prescription online your state exchange or use this site’s free quote tool. You can also get a subsidy estimate by using this ACA subsidy calculator.

More likely than not, you will be pleasantly surprised get lasix prescription online. Andrew Sprung is a freelance writer who blogs about politics and healthcare policy at xpostfactoid. His articles about the Affordable Care Act have appeared in publications including The American Prospect, Health Affairs, The Atlantic and The New Republic.

He is the winner of the National Institute of Health Care Management’s 2016 get lasix prescription online Digital Media Award. He holds a Ph.D. In English literature from the University of Rochester..

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100%. } } Latest Updates. The hypertension Outbreak 33m ago Judge upholds Cuomo’s restrictions on religious services in hot spots.

1h ago Latino and Black Americans are still dying in disproportionately high numbers, the C.D.C. Says. 4h ago Keuka College sends students home because of hypertension outbreak.

See more updates More live coverage. Markets Nationally, jails and prisons have seen disproportionate rates of and death, with a mortality rate twice as high as in the general population and an rate more than four times as high, according to recent data.A New York Times database has tracked clusters of at least 50 hypertension cases in a dozen rural jails in Montana, Idaho, Utah and New Mexico during the lasix. Among them.

The Purgatory Correctional Center in Hurricane, Utah, with 166 s. The jail in Twin Falls, Idaho, with 279. And, in New Mexico, the Cibola County Correctional Center, which has reported 357 cases.In Cascade County, s at the jail make up about a quarter of all known lasix cases in the county.

Health authorities say that the jail’s outbreak, which began in mid-August, was not believed to be the main cause of the community’s recent surge, but that it had led to some cases. In the past two months, Mr. Krogue said, the jail released 29 people who were considered actively infected.s at the jail make up about a quarter of Cascade County’s known lasix cases.Credit...Tailyr Irvine for The New York TimesGreat Falls, home to about 58,000 residents, is in the less mountainous part of Montana, with the Missouri River flowing through and a large oil refinery on its banks.

The Cascade County Detention Center sits along a highway at the edge of town. Drive five miles in any direction and you are surrounded by wide-open plains.Montana requires that masks be worn inside businesses and indoor public spaces, and many people in Great Falls wear them when walking around downtown’s Central Avenue, where shops and cafes are still recovering from shutting down in the spring. Others go without masks, citing the open space and lack of crowds.Bob Kelly, the mayor, said people had not been overly worried about how the jail outbreak might affect the rest of town when it started.“I think that by the very definition of a jail, hopefully, the disease will be incarcerated, as well as the patients,” he said.

€œIs there concern?. Sure, there’s concern. But is there overreaction?.

No.”The mayor of Great Falls said that residents had considered the jail’s outbreak a distant concern at first.Credit...Tailyr Irvine for The New York TimesSome residents’ nonchalance about the risks of the lasix, said Mr. Krogue, the jail’s medical director, can be traced to a spring and early summer when almost no one in Cascade County knew anyone who had been sickened.“We benefited from that early on,” he said. €œBut in some ways, I think it did us a disservice, too, because it also created a certain level of complacency.”That has quickly shifted now, he said, as cases have spiked.The number of active cases known to county officials on any given day has risen sharply to about 600, according to Trisha Gardner, Cascade County’s health officer.

The county has seen 1,261 cases and six deaths during the lasix, a Times database shows. Some of the cases have been tied to the jail outbreak, she said, and others have been connected to bars and restaurants. Even figuring out what has led to some cases has been complex, she said, as residents have been reluctant to cooperate with contact tracers.“Our hospitals are at capacity, our public health system is at capacity,” she said.

€œIt’s not sustainable at this rate.”When the outbreak at the jail began, social distancing was impossible, the authorities said. Three inmates shared cells designed for two. At night, men slept on thin blue pads in every available space.

On the floor in the day room, in shower stalls, in stairwells, in hallways outside of cells.Inmates did not receive masks until August, and jail officials said many have refused to wear them.In interviews with more than a dozen inmates and their family members, inmates described the jail during the outbreak as chaotic and unsanitary. They said their pleas for help often went unanswered by nurses and guards.Newly arriving inmates were not always quarantined from one another before their test results were known because of a lack of space, inmates and jail officials said.Owen Hawley, 30, said every inmate in his living area of 38 men had tested positive for the lasix. He said he had been unable to eat for three days, had intensive body aches and suffered from a headache so powerful it felt as if it was “behind my eyes.”“After the fourth day of like, not eating and stuff, I just shut off, you know?.

€ he said.A jail area set aside for quarantining new inmates.Credit...Tailyr Irvine for The New York TimesAt one point, Mr. Hawley said, he and other prisoners protested the way the lasix was being handled by refusing to leave their living areas and by blocking new inmates from entering. Everyone was ultimately tested, Mr.

Hawley said, and each prisoner was given a disposable mask.Sierra Jasmine Wells, 25, another inmate, said women in her dormitory had grown ill, one after the next.“Everyone around me was getting sick and it was tough on me,” she said. €œBy then, I had already accepted the fact that I was going to get sick.”When she became infected, she said, she was given cough syrup and Tylenol.“I kind of was just left alone to deal with it,” she said.Jesse Slaughter, the county sheriff who oversees the jail, said that the jail’s medical staff was doing everything it could, and that he had been seeking health care assistance from other counties. Officials defended their handling of the outbreak, noting that all inmates received standard medications including Tylenol twice a day and were taken to area hospitals when they needed added care.

Seven inmates, as well as some staff members, were hospitalized. No one from the jail has died from the lasix, officials said.Sheriff Jesse Slaughter, who oversees the jail, said he had been seeking health care assistance from other counties.Credit...Tailyr Irvine for The New York TimesMr. Krogue said that since the start of the outbreak he had been working up to 16 hours each day and sleeping in his basement, away from his wife and children.

He remains healthy but says he fears bringing the lasix home. The lasix has slowed some in the jail, and officials have moved some inmates to other facilities, but other prisons and jails in the state are now seeing outbreaks.“You can start to see what some of these other places experienced much earlier on, and we just didn’t have that experience, but it’s certainly happening now,” Mr. Krogue said.

€œIt’s just real in a way that it wasn’t.”Lucy Tompkins reported from Great Falls, Maura Turcotte from Chicago and Libby Seline from Lincoln, Neb. Reporting was contributed by Izzy Colón from Columbia, Mo., Brendon Derr from Phoenix, Rebecca Griesbach from Tuscaloosa, Ala., Danya Issawi and Timothy Williams from New York, Ann Hinga Klein from Des Moines, K.B. Mensah from Silver Spring, Md., and Mitch Smith from Chicago.Start Preamble Federal Transit Administration (FTA), DOT.

Notice of funding opportunity. The hypertension Disease 2019 (hypertension medications) public health emergency Start Printed Page 63654has had a significant impact on transit operations. During a series of FTA listening sessions held over the last three months, transit agencies asked FTA to support research to identify solutions to address the operational challenges that they are facing as a result of hypertension medications.

In response, FTA makes available through this Notice of Funding Opportunity (NOFO) funding to support research demonstration grants to public transit agencies to develop, deploy, and demonstrate innovative solutions that improve the operational efficiency of transit agencies, as well as enhance the mobility of transit users affected by the hypertension medications public health emergency. Demonstration grants under this NOFO are authorized under FTA's Public Transportation Innovation Program (49 U.S.C. 5312).

Eligible projects will demonstrate innovative solutions to improve the operational efficiencies of transit systems and enhance mobility for their communities in four major areas. (1) Vehicle, facility, equipment and infrastructure cleaning and dis. (2) exposure mitigation measures.

(3) innovative mobility such as contactless payments. And (4) measures that strengthen public confidence in transit services. The total funding available for awards under this NOFO is $10,000,000.

FTA may supplement this amount if additional funding becomes available. Applicants must submit completed proposals for funding opportunity FTA-2020-015-TRI through the GRANTS.GOV “APPLY” function by 11:59 p.m. Eastern Time on November 2, 2020.

Prospective applicants should register as soon as possible on the GRANTS.GOV website to ensure they can complete the application process before the submission deadline. Application instructions are available on FTA's website at http://transit.dot.gov/​howtoapply and in the “FIND” module of GRANTS.GOV. FTA will not accept mail and fax submissions.

Start Further Info Please send any questions on this notice to Jamel El-Hamri email. Jamel.El-Hamri@dot.gov phone. 2020-366-8985.

A Telecommunication Device for the Deaf (TDD) is available for individuals who are deaf or hard of hearing at 1-800-877-8339. End Further Info End Preamble Start Supplemental Information Table of Contents A. Program Description B.

Federal Award Information C. Eligibility Information D. Application and Submission Information E.

Application Review Information F. Federal Award Administration Information G. Federal Awarding Agency Contact Information A.

Program Description The Public Transportation hypertension medications Research Demonstration Grant Program is funded through the Public Transportation Innovation Program (49 U.S.C. 5312), with the goal to develop, deploy, and demonstrate innovative solutions that improve the operational efficiency of transit agencies, as well as enhance the mobility of transit users affected by the hypertension medications public health emergency. Eligible projects will propose to develop and deploy innovative solutions in four major areas.

(1) Vehicle, facility, equipment and infrastructure cleaning and dis. (2) exposure mitigation measures. (3) innovative mobility such as contactless payments.

And (4) measures that strengthen public confidence in transit. As required by 49 U.S.C. 5312(e)(4), projects funded under this NOFO must participate in an evaluation by an independent outside entity that will conduct a comprehensive evaluation of the success or failure of the projects funded under this subsection and any plan for broad-based implementation of the innovation promoted by successful projects.

B. Federal Award Information FTA makes available $10,000,000 in fiscal year (FY) 2020 funds under the Public Transportation Innovation Program (49 U.S.C. 5312) to finance the Public Transportation hypertension medications Research Demonstration Grant Program.

FTA may supplement the total funds available if additional funding becomes available at the time project selections are made. FTA will grant pre-award authority starting on the date of the project award announcement for selected projects and should be completed within 24 months from the date of award. Funds are available only for eligible expenses incurred after the announcement of project selections.

C. Eligibility Information (1) Eligible Applicants Eligible applicants include State and local governmental authorities, direct recipients of Urbanized Area (49 U.S.C. 5307) and Rural Area (49 U.S.C.

5311) formula funds, and Indian tribes. Eligible applicants are limited to FTA grantees or subrecipients who would be the primary beneficiaries of the innovative products and services that are developed—typically public transit agencies. Except for projects proposed by Indian tribes, proposals for projects in rural (non-urbanized) areas must be submitted as part of a consolidated State proposal.

States and other eligible applicants also may submit consolidated proposals for projects in urbanized areas. The submission of the Statewide application will not preclude the submission and consideration of any application from other eligible recipients in an urbanized area in a State. Proposals may contain projects to be implemented by the recipient or its subrecipients.

Eligible subrecipients include public agencies, private nonprofit organizations, and private providers engaged in public transportation. Eligible applicants may submit consolidated proposals for projects. (2) Cost Sharing or Matching The maximum Federal share of project costs is 100 percent.

FTA may give additional consideration to applicants that propose a local share and may view these applicants as more competitive. The applicant must document the source(s) of the local match, if any, in the grant application. For any applicants proposing match, eligible local match sources include the following.

Cash from non-Government sources other than revenues from providing public transportation services. Revenues derived from the sale of advertising and concessions. Revenues generated from value capture financing mechanisms.

Funds from an undistributed cash surplus. Replacement or depreciation cash fund or reserve. New capital.

Or in-kind contributions. (3) Eligible Projects Eligible projects will propose innovative solutions to improve operational efficiencies of transit agencies and enhance the mobility of transit users, through projects that demonstrate innovative solutions for. Vehicle, facility, equipment and infrastructure cleaning and dis.

Exposure mitigation measures such a real-time notification of rail and bus passenger loads. New multi-modal payment innovative mobility systems such as contactless payments. And measures that strengthen public confidence in transit.

Each applicant may only submit one proposal.Start Printed Page 63655 D. Application and Submission Information (1) Address and Form of Application Submission Applications must be submitted through GRANTS.GOV. Applicants can find general information for submitting applications through GRANTS.GOV at www.fta.dot.gov/​howtoapply, along with specific instructions for the forms and attachments required for submission.

Mail and fax submissions will not be accepted. (2) Content and Form of Application Submission a. Proposal Submission A complete proposal submission consists of at least two forms.

1. The SF-424 Mandatory Form (downloadable from GRANTS.GOV) and 2. The supplemental form for the FY 2020 hypertension medications Demonstration Program (downloadable from GRANTS.GOV), which is available on FTA's website at (placeholder for FTA hypertension medications Demonstration Program).

The application must include responses to all sections of the SF-424 mandatory form and the supplemental form unless a section is indicated as optional. FTA will use the information on the supplemental form to determine applicant and project eligibility for the program and to evaluate the proposal against the selection criteria described in part E of this notice. FTA will accept only one supplemental form per SF-424 submission.

FTA encourages applicants to consider submitting a single supplemental form that includes multiple activities to be evaluated as a consolidated proposal. Applicants may attach additional supporting information to the SF-424 submission, including but not limited to letters of support, project budgets, or excerpts from relevant planning documents. Supporting documentation must be described and referenced by file name in the appropriate response section of the supplemental form, or it may not be reviewed.

Information such as applicant name, Federal amount requested, local match amount, description of areas served, etc., may be requested in varying degrees of detail on both the SF-424 form and supplemental form. Applicants must fill in all fields unless stated otherwise on the forms. If applicants copy information into the supplemental form from another source, they should verify that the supplemental form has fully captured pasted text and that it has not truncated the text due to character limits built into the form.

Applicants should use both the “Check Package for Errors” and the “Validate Form” validation buttons on both forms to check all required fields. Applicants should also ensure that the Federal and local amounts specified are consistent. Addressing the deteriorating conditions and disproportionately high fatality rates on our rural transportation infrastructure is of critical interest to the Department, as rural transportation networks face unique challenges in safety, infrastructure condition, and passenger and freight usage.

Consistent with the R.O.U.T.E.S. Initiative, the Department encourages applicants to consider how the project will address the challenges faced by rural areas. B.

Application Content The SF-424 Mandatory Form and the supplemental form will prompt applicants for the required information, including. I. Applicant Name ii.

Dun and Bradstreet (D&B) Data Universal Numbering System (DUNS) number iii. Key contact information (contact name, address, email address, and phone number) iv. Congressional district(s) where project will take place v.

Project Information (title, executive summary, and type) vi. A detailed description of the need for the project vii. A detailed description of how the project will support the Program objectives viii.

Evidence that the applicant can provide the local cost shares ix. A description of the technical, legal, and financial capacity of the applicant x. A detailed project budget xi.

Details on the local matching funds xii. A detailed project timeline xiii. Whether the project impacts an Opportunity Zone (3) Unique Entity Identifier and System for Award Management (SAM) Each applicant is required to.

(1) Be registered in SAM before submitting an application. (2) provide a valid unique entity identifier in its application. And (3) continue to maintain an active SAM registration with current information at all times during which the applicant has an active Federal award or an application or plan under consideration by FTA.

These requirements do not apply if the applicant. (1) Is excepted from the requirements under 2 CFR 25.110(b) or (c). Or (2) has an exception approved by FTA under 2 CFR 25.110(d).

FTA may not make an award until the applicant has complied with all applicable unique entity identifier and SAM requirements. If an applicant has not fully complied with the requirements by the time FTA is ready to make an award, FTA may determine that the applicant is not qualified to receive an award and use that determination as a basis for making a Federal award to another applicant. All applicants must provide a unique entity identifier provided by SAM.

Registration in SAM may take as little as 3-5 business days, but there can be unexpected steps or delays. For example, the applicant may need to obtain an Employer Identification Number. FTA recommends allowing ample time, up to several weeks, to complete all steps.

For additional information on obtaining a unique entity identifier, please visit www.sam.gov. (4) Submission Dates and Times Project proposals must be submitted electronically through GRANTS.GOV by 11:59 p.m. Eastern on November 2, 2020.

Mail and fax submissions will not be accepted. FTA urges applicants to submit applications at least 72 hours prior to the due date to allow time to correct any problems that may have caused either GRANTS.GOV or FTA systems to reject the submission. Proposals submitted after the deadline will only be considered under extraordinary circumstances not within the applicant's control.

Deadlines will not be extended due to scheduled website maintenance. GRANTS.GOV scheduled maintenance and outage times are announced on the GRANTS.GOV website. Within 48 hours after submitting an electronic application, the applicant should receive two email messages from GRANTS.GOV.

(1) Confirmation of successful transmission to GRANTS.GOV. And (2) confirmation of successful validation by GRANTS.GOV. If the applicant does not receive confirmation of successful validation or receives a notice of failed validation or incomplete materials, the applicant must address the reason for the failed validation, as described in the email notice, and resubmit before the submission deadline.

If making a resubmission for any reason, applicants must include all original attachments regardless of which attachments were updated and check the box on the supplemental form indicating this is a resubmission. Applicants are encouraged to begin the process of registration on the GRANTS.GOV site well in advance of the submission deadline. Registration is Start Printed Page 63656a multi-step process, which may take several weeks to complete before an application can be submitted.

Registered applicants may still be required to update their registration before submitting an application. Registration in SAM is renewed annually and persons making submissions on behalf of the Authorized Organization Representative (AOR) must be authorized in GRANTS.GOV by the AOR to make submissions. (5) Funding Restrictions Funds may be used for post-award expenditures only.

Funds under this NOFO cannot be used to reimburse projects for otherwise eligible expenses incurred prior to the date of project award announcements. (6) Other Submission Requirements FTA encourages applicants to identify scaled funding options in case insufficient funding is available to fund a project at the full requested amount. If an applicant indicates that a project is scalable, the applicant must provide an appropriate minimum funding amount that will fund an eligible project that achieves the objectives of the program and meets all relevant program requirements.

The applicant must provide a clear explanation of how a reduced award would affect the project budget and scope. FTA may award a lesser amount whether or not the applicant provides a scalable option. E.

Application Review Information (1) Project Evaluation Criteria Addressing the deteriorating conditions and disproportionately high fatality rates on our rural transportation infrastructure is of critical interest to the Department, as rural transportation networks face unique challenges in safety, infrastructure condition, and passenger and freight usage. Consistent with the R.O.U.T.E.S. Initiative, the Department will consider how the project will address the challenges faced by rural areas.

In addition, the Department will review and consider applications for funding pursuant to this Notice in accordance with the President's September 2, 2020 memorandum, entitled Memorandum on Reviewing Funding to State and Local Government Recipients of Federal Funds that Are Permitting Anarchy, Violence, and Destruction in American Cities, consistent with guidance from the Office of Management and Budget and the Attorney General and with all applicable laws. FTA will evaluate proposals submitted according to the following criteria. (a) Project Innovation and Impact.

(b) Project Approach. (c) National Applicability. (d) Commercialization and/or Knowledge Transfer.

And (e) Technical, Legal and Financial Capacity. FTA encourages each applicant to demonstrate how a project supports all criteria with the most relevant information the applicant can provide, regardless of whether such information has been specifically requested or identified in this notice. A.

Project Innovation and Impact i. Effectiveness of the project in achieving and demonstrating the specific objectives of this program. Ii.

Demonstration of benefits in addressing the needs of the transit agency and industry and impacts to infrastructure, equipment, transit workforce, and riders. Iii. Degree of improvement over current and existing technologies, designs, and/or practices applicable to the transit industry.

B. Project Approach i. Quality of the project approach such as existing partnerships, collaboration strategies and level of commitment of the project partners.

Ii. Proposal is realistic in its approach to fulfill the milestones/deliverables, schedule and goals. C.

National Applicability i. Degree to which the project could be replicated by other transit agencies regionally or nationally. Ii.

Ability to evaluate technologies, designs and/or practices in a wide variety of conditions and locales. Iii. Degree to which the technology, designs and/or practices can be replicated by other transportation modes.

D. Commercialization and/or Knowledge Transfer i. Demonstrates a realistic plan for moving the results of the project into the transit marketplace (patents, conferences, articles in trade magazines, webinar, site visits, etc.).

Ii. How the project team plans to work with the industry on improving best practices, guidance and/or standards, if applicable. Iii.

Demonstrate a clear understanding and robust approach to data collection, access and management. E. Technical, Legal and Financial Capacity Capacity of the applicant and any partners to successfully execute the project effort.

There should be no outstanding legal, technical, or financial issues with the applicant that would make this a high-risk project. (2) Review and Selection Process An FTA technical evaluation committee will evaluate proposals based on the published project evaluation criteria. Members of the technical evaluation committee will rate the applications and may seek clarification about any statement in an application.

The FTA Administrator will determine the final selection and amount of funding for each project after consideration of the findings of the technical evaluation committee. Geographic diversity, diversity of the project type, the amount of local match to be provided, and the applicant's receipt and management of other Federal transit funds may be considered in FTA's award decisions. Prior fare payment innovation efforts may receive priority consideration.

The FTA Administrator will consider the following key DOT objectives. A. Utilizing alternative funding sources and innovative financing models to attract non-Federal sources of investment.

B. Whether the project is located in or supports public transportation service in a qualified opportunity zone designated pursuant to 26.U.S.C. 1400Z-1.

And c. The extent to which the project addresses challenges specific to the provision of rural public transportation. (3) FAPIIS Review Prior to making a grant award, FTA is required to review and consider any information about the applicant that is in the Federal Awardee Performance and Integrity Information System (FAPIIS) accessible through SAM.

An applicant may review and comment on information about itself that a Federal awarding agency previously entered. FTA will consider any comments by the applicant, in addition to the other information in FAPIIS, in making a judgment about the applicant's integrity, business ethics, and record of performance under Federal awards when completing the review of risk posed by applicants as described in 2 CFR 200.205 Federal Awarding Agency Review of Risk Posed by Applicants. F.

Federal Award Administration Information (1) Federal Award Notices FTA will announce the final project selections on the FTA website. Project recipients should contact their FTA Regional Office for additional information regarding allocations for Start Printed Page 63657projects. At the time project selections are announced, FTA will extend pre-award authority for the selected projects.

There is no blanket pre-award authority for these projects before announcement. There is no minimum or maximum grant award amount, but FTA intends to fund as many meritorious projects as possible. FTA only will consider proposals from eligible recipients for eligible activities.

Due to funding limitations, projects selected for funding may receive less than the amount originally requested. In those cases, applicants must be able to demonstrate that the proposed projects are still viable and can be completed with the amount awarded. (2) Administrative and National Policy Requirements a.

Pre-Award Authority FTA will issue specific guidance to recipients regarding pre-award authority at the time of selection. FTA does not provide pre-award authority for competitive funds until projects are selected, and there are Federal requirements that must be met before costs are incurred. For more information about FTA's policy on pre-award authority, see the FY 2020 Apportionments Notice published on June 3, 2020, at https://www.govinfo.gov/​content/​pkg/​FR-2020-06-03/​pdf/​2020-11946.pdf.

b. Grant Requirements Selected applicants will submit a grant application through FTA's electronic grant management system and adhere to the customary FTA grant requirements for research project (insert Circular name). All competitive grants, regardless of award amount, will be subject to the Congressional notification and release process.

FTA emphasizes that third-party procurement applies to all funding awards, as described in FTA Circular 4220.1F, “Third Party Contracting Guidance.” However, FTA may approve applications that include a specifically identified partnering organization(s) (2 CFR 200.302(f)). When included, the application, budget, and budget narrative should provide a clear understanding of how the selection of these organizations is critical for the project and give sufficient detail about the costs involved. C.

Planning FTA encourages applicants to engage the appropriate State Departments of Transportation, Regional Transportation Planning Organizations, or Metropolitan Planning Organizations in areas to be served by the project funds available under this program. D. Standard Assurances The applicant assures that it will comply with all applicable Federal statutes, regulations, executive orders, FTA circulars, and other Federal administrative requirements in carrying out any project supported by the FTA grant.

The applicant acknowledges that it is under a continuing obligation to comply with the terms and conditions of the grant agreement issued for its project with FTA. The applicant understands that Federal laws, regulations, policies, and administrative practices might be modified from time to time and may affect the implementation of the project. The applicant agrees that the most recent Federal requirements will apply to the project unless FTA issues a written determination otherwise.

The applicant must submit the Certifications and Assurances before receiving a grant if it does not have current certifications on file. E. Free Speech and Religious Liberty In connection with any program or activity conducted with or benefiting from funds awarded under this notice, recipients of funds must comply with all applicable requirements of Federal law, including, without limitation, the Constitution of the United States.

Statutory, regulatory, and public policy requirements, including without limitation, those protecting free speech, religious liberty, public welfare, the environment, and prohibiting discrimination. The conditions of performance, non-discrimination requirements, and other assurances made applicable to the award of funds in accordance with regulations of the Department of Transportation. And applicable Federal financial assistance and contracting principles promulgated by the Office of Management and Budget.

In complying with these requirements, recipients must ensure that no concession agreements are denied or other contracting decisions made on the basis of speech or other activities protected by the First Amendment. If the Department determines that a recipient has failed to comply with applicable Federal requirements, the Department may terminate the award of funds and disallow previously incurred costs, requiring the recipient to reimburse any expended award funds. (3) Reporting The post-award reporting requirements include submission of the Federal Financial Report (FFR) and Milestone Progress Report in TrAMS.

An evaluation of the grant will occur at various points in the demonstration process and at the end of the project. In addition, FTA is responsible for producing an Annual Report to Congress that compiles evaluation of selected projects, including an evaluation of the performance measures identified by the applicants. All applicants must develop an evaluation plan to measure the success or failure of their projects and describe any plans for broad-based implementation of successful projects.

FTA may request data and reports to support the evaluation and Annual Report. A.

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} } Latest Updates. The hypertension Outbreak 33m ago Judge upholds Cuomo’s restrictions on religious services in hot spots. 1h ago Latino and Black Americans are still dying in disproportionately high numbers, the C.D.C.

Says. 4h ago Keuka College sends students home because of hypertension outbreak. See more updates More live coverage.

Markets Nationally, jails and prisons have seen disproportionate rates of and death, with a mortality rate twice as high as in the general population and an rate more than four times as high, according to recent data.A New York Times database has tracked clusters of at least 50 hypertension cases in a dozen rural jails in Montana, Idaho, Utah and New Mexico during the lasix. Among them. The Purgatory Correctional Center in Hurricane, Utah, with 166 s.

The jail in Twin Falls, Idaho, with 279. And, in New Mexico, the Cibola County Correctional Center, which has reported 357 cases.In Cascade County, s at the jail make up about a quarter of all known lasix cases in the county. Health authorities say that the jail’s outbreak, which began in mid-August, was not believed to be the main cause of the community’s recent surge, but that it had led to some cases.

In the past two months, Mr. Krogue said, the jail released 29 people who were considered actively infected.s at the jail make up about a quarter of Cascade County’s known lasix cases.Credit...Tailyr Irvine for The New York TimesGreat Falls, home to about 58,000 residents, is in the less mountainous part of Montana, with the Missouri River flowing through and a large oil refinery on its banks. The Cascade County Detention Center sits along a highway at the edge of town.

Drive five miles in any direction and you are surrounded by wide-open plains.Montana requires that masks be worn inside businesses and indoor public spaces, and many people in Great Falls wear them when walking around downtown’s Central Avenue, where shops and cafes are still recovering from shutting down in the spring. Others go without masks, citing the open space and lack of crowds.Bob Kelly, the mayor, said people had not been overly worried about how the jail outbreak might affect the rest of town when it started.“I think that by the very definition of a jail, hopefully, the disease will be incarcerated, as well as the patients,” he said. €œIs there concern?.

Sure, there’s concern. But is there overreaction?. No.”The mayor of Great Falls said that residents had considered the jail’s outbreak a distant concern at first.Credit...Tailyr Irvine for The New York TimesSome residents’ nonchalance about the risks of the lasix, said Mr.

Krogue, the jail’s medical director, can be traced to a spring and early summer when almost no one in Cascade County knew anyone who had been sickened.“We benefited from that early on,” he said. €œBut in some ways, I think it did us a disservice, too, because it also created a certain level of complacency.”That has quickly shifted now, he said, as cases have spiked.The number of active cases known to county officials on any given day has risen sharply to about 600, according to Trisha Gardner, Cascade County’s health officer. The county has seen 1,261 cases and six deaths during the lasix, a Times database shows.

Some of the cases have been tied to the jail outbreak, she said, and others have been connected to bars and restaurants. Even figuring out what has led to some cases has been complex, she said, as residents have been reluctant to cooperate with contact tracers.“Our hospitals are at capacity, our public health system is at capacity,” she said. €œIt’s not sustainable at this rate.”When the outbreak at the jail began, social distancing was impossible, the authorities said.

Three inmates shared cells designed for two. At night, men slept on thin blue pads in every available space. On the floor in the day room, in shower stalls, in stairwells, in hallways outside of cells.Inmates did not receive masks until August, and jail officials said many have refused to wear them.In interviews with more than a dozen inmates and their family members, inmates described the jail during the outbreak as chaotic and unsanitary.

They said their pleas for help often went unanswered by nurses and guards.Newly arriving inmates were not always quarantined from one another before their test results were known because of a lack of space, inmates and jail officials said.Owen Hawley, 30, said every inmate in his living area of 38 men had tested positive for the lasix. He said he had been unable to eat for three days, had intensive body aches and suffered from a headache so powerful it felt as if it was “behind my eyes.”“After the fourth day of like, not eating and stuff, I just shut off, you know?. € he said.A jail area set aside for quarantining new inmates.Credit...Tailyr Irvine for The New York TimesAt one point, Mr.

Hawley said, he and other prisoners protested the way the lasix was being handled by refusing to leave their living areas and by blocking new inmates from entering. Everyone was ultimately tested, Mr. Hawley said, and each prisoner was given a disposable mask.Sierra Jasmine Wells, 25, another inmate, said women in her dormitory had grown ill, one after the next.“Everyone around me was getting sick and it was tough on me,” she said.

€œBy then, I had already accepted the fact that I was going to get sick.”When she became infected, she said, she was given cough syrup and Tylenol.“I kind of was just left alone to deal with it,” she said.Jesse Slaughter, the county sheriff who oversees the jail, said that the jail’s medical staff was doing everything it could, and that he had been seeking health care assistance from other counties. Officials defended their handling of the outbreak, noting that all inmates received standard medications including Tylenol twice a day and were taken to area hospitals when they needed added care. Seven inmates, as well as some staff members, were hospitalized.

No one from the jail has died from the lasix, officials said.Sheriff Jesse Slaughter, who oversees the jail, said he had been seeking health care assistance from other counties.Credit...Tailyr Irvine for The New York TimesMr. Krogue said that since the start of the outbreak he had been working up to 16 hours each day and sleeping in his basement, away from his wife and children. He remains healthy but says he fears bringing the lasix home.

The lasix has slowed some in the jail, and officials have moved some inmates to other facilities, but other prisons and jails in the state are now seeing outbreaks.“You can start to see what some of these other places experienced much earlier on, and we just didn’t have that experience, but it’s certainly happening now,” Mr. Krogue said. €œIt’s just real in a way that it wasn’t.”Lucy Tompkins reported from Great Falls, Maura Turcotte from Chicago and Libby Seline from Lincoln, Neb.

Reporting was contributed by Izzy Colón from Columbia, Mo., Brendon Derr from Phoenix, Rebecca Griesbach from Tuscaloosa, Ala., Danya Issawi and Timothy Williams from New York, Ann Hinga Klein from Des Moines, K.B. Mensah from Silver Spring, Md., and Mitch Smith from Chicago.Start Preamble Federal Transit Administration (FTA), DOT. Notice of funding opportunity.

The hypertension Disease 2019 (hypertension medications) public health emergency Start Printed Page 63654has had a significant impact on transit operations. During a series of FTA listening sessions held over the last three months, transit agencies asked FTA to support research to identify solutions to address the operational challenges that they are facing as a result of hypertension medications. In response, FTA makes available through this Notice of Funding Opportunity (NOFO) funding to support research demonstration grants to public transit agencies to develop, deploy, and demonstrate innovative solutions that improve the operational efficiency of transit agencies, as well as enhance the mobility of transit users affected by the hypertension medications public health emergency.

Demonstration grants under this NOFO are authorized under FTA's Public Transportation Innovation Program (49 U.S.C. 5312). Eligible projects will demonstrate innovative solutions to improve the operational efficiencies of transit systems and enhance mobility for their communities in four major areas.

(1) Vehicle, facility, equipment and infrastructure cleaning and dis. (2) exposure mitigation measures. (3) innovative mobility such as contactless payments.

And (4) measures that strengthen public confidence in transit services. The total funding available for awards under this NOFO is $10,000,000. FTA may supplement this amount if additional funding becomes available.

Applicants must submit completed proposals for funding opportunity FTA-2020-015-TRI through the GRANTS.GOV “APPLY” function by 11:59 p.m. Eastern Time on November 2, 2020. Prospective applicants should register as soon as possible on the GRANTS.GOV website to ensure they can complete the application process before the submission deadline.

Application instructions are available on FTA's website at http://transit.dot.gov/​howtoapply and in the “FIND” module of GRANTS.GOV. FTA will not accept mail and fax submissions. Start Further Info Please send any questions on this notice to Jamel El-Hamri email.

Jamel.El-Hamri@dot.gov phone. 2020-366-8985. A Telecommunication Device for the Deaf (TDD) is available for individuals who are deaf or hard of hearing at 1-800-877-8339.

End Further Info End Preamble Start Supplemental Information Table of Contents A. Program Description B. Federal Award Information C.

Eligibility Information D. Application and Submission Information E. Application Review Information F.

Federal Award Administration Information G. Federal Awarding Agency Contact Information A. Program Description The Public Transportation hypertension medications Research Demonstration Grant Program is funded through the Public Transportation Innovation Program (49 U.S.C.

5312), with the goal to develop, deploy, and demonstrate innovative solutions that improve the operational efficiency of transit agencies, as well as enhance the mobility of transit users affected by the hypertension medications public health emergency. Eligible projects will propose to develop and deploy innovative solutions in four major areas. (1) Vehicle, facility, equipment and infrastructure cleaning and dis.

(2) exposure mitigation measures. (3) innovative mobility such as contactless payments. And (4) measures that strengthen public confidence in transit.

As required by 49 U.S.C. 5312(e)(4), projects funded under this NOFO must participate in an evaluation by an independent outside entity that will conduct a comprehensive evaluation of the success or failure of the projects funded under this subsection and any plan for broad-based implementation of the innovation promoted by successful projects. B.

Federal Award Information FTA makes available $10,000,000 in fiscal year (FY) 2020 funds under the Public Transportation Innovation Program (49 U.S.C. 5312) to finance the Public Transportation hypertension medications Research Demonstration Grant Program. FTA may supplement the total funds available if additional funding becomes available at the time project selections are made.

FTA will grant pre-award authority starting on the date of the project award announcement for selected projects and should be completed within 24 months from the date of award. Funds are available only for eligible expenses incurred after the announcement of project selections. C.

Eligibility Information (1) Eligible Applicants Eligible applicants include State and local governmental authorities, direct recipients of Urbanized Area (49 U.S.C. 5307) and Rural Area (49 U.S.C. 5311) formula funds, and Indian tribes.

Eligible applicants are limited to FTA grantees or subrecipients who would be the primary beneficiaries of the innovative products and services that are developed—typically public transit agencies. Except for projects proposed by Indian tribes, proposals for projects in rural (non-urbanized) areas must be submitted as part of a consolidated State proposal. States and other eligible applicants also may submit consolidated proposals for projects in urbanized areas.

The submission of the Statewide application will not preclude the submission and consideration of any application from other eligible recipients in an urbanized area in a State. Proposals may contain projects to be implemented by the recipient or its subrecipients. Eligible subrecipients include public agencies, private nonprofit organizations, and private providers engaged in public transportation.

Eligible applicants may submit consolidated proposals for projects. (2) Cost Sharing or Matching The maximum Federal share of project costs is 100 percent. FTA may give additional consideration to applicants that propose a local share and may view these applicants as more competitive.

The applicant must document the source(s) of the local match, if any, in the grant application. For any applicants proposing match, eligible local match sources include the following. Cash from non-Government sources other than revenues from providing public transportation services.

Revenues derived from the sale of advertising and concessions. Revenues generated from value capture financing mechanisms. Funds from an undistributed cash surplus.

Replacement or depreciation cash fund or reserve. New capital. Or in-kind contributions.

(3) Eligible Projects Eligible projects will propose innovative solutions to improve operational efficiencies of transit agencies and enhance the mobility of transit users, through projects that demonstrate innovative solutions for. Vehicle, facility, equipment and infrastructure cleaning and dis. Exposure mitigation measures such a real-time notification of rail and bus passenger loads.

New multi-modal payment innovative mobility systems such as contactless payments. And measures that strengthen public confidence in transit. Each applicant may only submit one proposal.Start Printed Page 63655 D.

Application and Submission Information (1) Address and Form of Application Submission Applications must be submitted through GRANTS.GOV. Applicants can find general information for submitting applications through GRANTS.GOV at www.fta.dot.gov/​howtoapply, along with specific instructions for the forms and attachments required for submission. Mail and fax submissions will not be accepted.

(2) Content and Form of Application Submission a. Proposal Submission A complete proposal submission consists of at least two forms. 1.

The SF-424 Mandatory Form (downloadable from GRANTS.GOV) and 2. The supplemental form for the FY 2020 hypertension medications Demonstration Program (downloadable from GRANTS.GOV), which is available on FTA's website at (placeholder for FTA hypertension medications Demonstration Program). The application must include responses to all sections of the SF-424 mandatory form and the supplemental form unless a section is indicated as optional.

FTA will use the information on the supplemental form to determine applicant and project eligibility for the program and to evaluate the proposal against the selection criteria described in part E of this notice. FTA will accept only one supplemental form per SF-424 submission. FTA encourages applicants to consider submitting a single supplemental form that includes multiple activities to be evaluated as a consolidated proposal.

Applicants may attach additional supporting information to the SF-424 submission, including but not limited to letters of support, project budgets, or excerpts from relevant planning documents. Supporting documentation must be described and referenced by file name in the appropriate response section of the supplemental form, or it may not be reviewed. Information such as applicant name, Federal amount requested, local match amount, description of areas served, etc., may be requested in varying degrees of detail on both the SF-424 form and supplemental form.

Applicants must fill in all fields unless stated otherwise on the forms. If applicants copy information into the supplemental form from another source, they should verify that the supplemental form has fully captured pasted text and that it has not truncated the text due to character limits built into the form. Applicants should use both the “Check Package for Errors” and the “Validate Form” validation buttons on both forms to check all required fields.

Applicants should also ensure that the Federal and local amounts specified are consistent. Addressing the deteriorating conditions and disproportionately high fatality rates on our rural transportation infrastructure is of critical interest to the Department, as rural transportation networks face unique challenges in safety, infrastructure condition, and passenger and freight usage. Consistent with the R.O.U.T.E.S.

Initiative, the Department encourages applicants to consider how the project will address the challenges faced by rural areas. B. Application Content The SF-424 Mandatory Form and the supplemental form will prompt applicants for the required information, including.

I. Applicant Name ii. Dun and Bradstreet (D&B) Data Universal Numbering System (DUNS) number iii.

Key contact information (contact name, address, email address, and phone number) iv. Congressional district(s) where project will take place v. Project Information (title, executive summary, and type) vi.

A detailed description of the need for the project vii. A detailed description of how the project will support the Program objectives viii. Evidence that the applicant can provide the local cost shares ix.

A description of the technical, legal, and financial capacity of the applicant x. A detailed project budget xi. Details on the local matching funds xii.

A detailed project timeline xiii. Whether the project impacts an Opportunity Zone (3) Unique Entity Identifier and System for Award Management (SAM) Each applicant is required to. (1) Be registered in SAM before submitting an application.

(2) provide a valid unique entity identifier in its application. And (3) continue to maintain an active SAM registration with current information at all times during which the applicant has an active Federal award or an application or plan under consideration by FTA. These requirements do not apply if the applicant.

(1) Is excepted from the requirements under 2 CFR 25.110(b) or (c). Or (2) has an exception approved by FTA under 2 CFR 25.110(d). FTA may not make an award until the applicant has complied with all applicable unique entity identifier and SAM requirements.

If an applicant has not fully complied with the requirements by the time FTA is ready to make an award, FTA may determine that the applicant is not qualified to receive an award and use that determination as a basis for making a Federal award to another applicant. All applicants must provide a unique entity identifier provided by SAM. Registration in SAM may take as little as 3-5 business days, but there can be unexpected steps or delays.

For example, the applicant may need to obtain an Employer Identification Number. FTA recommends allowing ample time, up to several weeks, to complete all steps. For additional information on obtaining a unique entity identifier, please visit www.sam.gov.

(4) Submission Dates and Times Project proposals must be submitted electronically through GRANTS.GOV by 11:59 p.m. Eastern on November 2, 2020. Mail and fax submissions will not be accepted.

FTA urges applicants to submit applications at least 72 hours prior to the due date to allow time to correct any problems that may have caused either GRANTS.GOV or FTA systems to reject the submission. Proposals submitted after the deadline will only be considered under extraordinary circumstances not within the applicant's control. Deadlines will not be extended due to scheduled website maintenance.

GRANTS.GOV scheduled maintenance and outage times are announced on the GRANTS.GOV website. Within 48 hours after submitting an electronic application, the applicant should receive two email messages from GRANTS.GOV. (1) Confirmation of successful transmission to GRANTS.GOV.

And (2) confirmation of successful validation by GRANTS.GOV. If the applicant does not receive confirmation of successful validation or receives a notice of failed validation or incomplete materials, the applicant must address the reason for the failed validation, as described in the email notice, and resubmit before the submission deadline. If making a resubmission for any reason, applicants must include all original attachments regardless of which attachments were updated and check the box on the supplemental form indicating this is a resubmission.

Applicants are encouraged to begin the process of registration on the GRANTS.GOV site well in advance of the submission deadline. Registration is Start Printed Page 63656a multi-step process, which may take several weeks to complete before an application can be submitted. Registered applicants may still be required to update their registration before submitting an application.

Registration in SAM is renewed annually and persons making submissions on behalf of the Authorized Organization Representative (AOR) must be authorized in GRANTS.GOV by the AOR to make submissions. (5) Funding Restrictions Funds may be used for post-award expenditures only. Funds under this NOFO cannot be used to reimburse projects for otherwise eligible expenses incurred prior to the date of project award announcements.

(6) Other Submission Requirements FTA encourages applicants to identify scaled funding options in case insufficient funding is available to fund a project at the full requested amount. If an applicant indicates that a project is scalable, the applicant must provide an appropriate minimum funding amount that will fund an eligible project that achieves the objectives of the program and meets all relevant program requirements. The applicant must provide a clear explanation of how a reduced award would affect the project budget and scope.

FTA may award a lesser amount whether or not the applicant provides a scalable option. E. Application Review Information (1) Project Evaluation Criteria Addressing the deteriorating conditions and disproportionately high fatality rates on our rural transportation infrastructure is of critical interest to the Department, as rural transportation networks face unique challenges in safety, infrastructure condition, and passenger and freight usage.

Consistent with the R.O.U.T.E.S. Initiative, the Department will consider how the project will address the challenges faced by rural areas. In addition, the Department will review and consider applications for funding pursuant to this Notice in accordance with the President's September 2, 2020 memorandum, entitled Memorandum on Reviewing Funding to State and Local Government Recipients of Federal Funds that Are Permitting Anarchy, Violence, and Destruction in American Cities, consistent with guidance from the Office of Management and Budget and the Attorney General and with all applicable laws.

FTA will evaluate proposals submitted according to the following criteria. (a) Project Innovation and Impact. (b) Project Approach.

(c) National Applicability. (d) Commercialization and/or Knowledge Transfer. And (e) Technical, Legal and Financial Capacity.

FTA encourages each applicant to demonstrate how a project supports all criteria with the most relevant information the applicant can provide, regardless of whether such information has been specifically requested or identified in this notice. A. Project Innovation and Impact i.

Effectiveness of the project in achieving and demonstrating the specific objectives of this program. Ii. Demonstration of benefits in addressing the needs of the transit agency and industry and impacts to infrastructure, equipment, transit workforce, and riders.

Iii. Degree of improvement over current and existing technologies, designs, and/or practices applicable to the transit industry. B.

Project Approach i. Quality of the project approach such as existing partnerships, collaboration strategies and level of commitment of the project partners. Ii.

Proposal is realistic in its approach to fulfill the milestones/deliverables, schedule and goals. C. National Applicability i.

Degree to which the project could be replicated by other transit agencies regionally or nationally. Ii. Ability to evaluate technologies, designs and/or practices in a wide variety of conditions and locales.

Iii. Degree to which the technology, designs and/or practices can be replicated by other transportation modes. D.

Commercialization and/or Knowledge Transfer i. Demonstrates a realistic plan for moving the results of the project into the transit marketplace (patents, conferences, articles in trade magazines, webinar, site visits, etc.). Ii.

How the project team plans to work with the industry on improving best practices, guidance and/or standards, if applicable. Iii. Demonstrate a clear understanding and robust approach to data collection, access and management.

E. Technical, Legal and Financial Capacity Capacity of the applicant and any partners to successfully execute the project effort. There should be no outstanding legal, technical, or financial issues with the applicant that would make this a high-risk project.

(2) Review and Selection Process An FTA technical evaluation committee will evaluate proposals based on the published project evaluation criteria. Members of the technical evaluation committee will rate the applications and may seek clarification about any statement in an application. The FTA Administrator will determine the final selection and amount of funding for each project after consideration of the findings of the technical evaluation committee.

Geographic diversity, diversity of the project type, the amount of local match to be provided, and the applicant's receipt and management of other Federal transit funds may be considered in FTA's award decisions. Prior fare payment innovation efforts may receive priority consideration. The FTA Administrator will consider the following key DOT objectives.

A. Utilizing alternative funding sources and innovative financing models to attract non-Federal sources of investment. B.

Whether the project is located in or supports public transportation service in a qualified opportunity zone designated pursuant to 26.U.S.C. 1400Z-1. And c.

The extent to which the project addresses challenges specific to the provision of rural public transportation. (3) FAPIIS Review Prior to making a grant award, FTA is required to review and consider any information about the applicant that is in the Federal Awardee Performance and Integrity Information System (FAPIIS) accessible through SAM. An applicant may review and comment on information about itself that a Federal awarding agency previously entered.

FTA will consider any comments by the applicant, in addition to the other information in FAPIIS, in making a judgment about the applicant's integrity, business ethics, and record of performance under Federal awards when completing the review of risk posed by applicants as described in 2 CFR 200.205 Federal Awarding Agency Review of Risk Posed by Applicants. F. Federal Award Administration Information (1) Federal Award Notices FTA will announce the final project selections on the FTA website.

Project recipients should contact their FTA Regional Office for additional information regarding allocations for Start Printed Page 63657projects. At the time project selections are announced, FTA will extend pre-award authority for the selected projects. There is no blanket pre-award authority for these projects before announcement.

There is no minimum or maximum grant award amount, but FTA intends to fund as many meritorious projects as possible. FTA only will consider proposals from eligible recipients for eligible activities. Due to funding limitations, projects selected for funding may receive less than the amount originally requested.

In those cases, applicants must be able to demonstrate that the proposed projects are still viable and can be completed with the amount awarded. (2) Administrative and National Policy Requirements a. Pre-Award Authority FTA will issue specific guidance to recipients regarding pre-award authority at the time of selection.

FTA does not provide pre-award authority for competitive funds until projects are selected, and there are Federal requirements that must be met before costs are incurred. For more information about FTA's policy on pre-award authority, see the FY 2020 Apportionments Notice published on June 3, 2020, at https://www.govinfo.gov/​content/​pkg/​FR-2020-06-03/​pdf/​2020-11946.pdf. b.

Grant Requirements Selected applicants will submit a grant application through FTA's electronic grant management system and adhere to the customary FTA grant requirements for research project (insert Circular name). All competitive grants, regardless of award amount, will be subject to the Congressional notification and release process. FTA emphasizes that third-party procurement applies to all funding awards, as described in FTA Circular 4220.1F, “Third Party Contracting Guidance.” However, FTA may approve applications that include a specifically identified partnering organization(s) (2 CFR 200.302(f)).

When included, the application, budget, and budget narrative should provide a clear understanding of how the selection of these organizations is critical for the project and give sufficient detail about the costs involved. C. Planning FTA encourages applicants to engage the appropriate State Departments of Transportation, Regional Transportation Planning Organizations, or Metropolitan Planning Organizations in areas to be served by the project funds available under this program.

D. Standard Assurances The applicant assures that it will comply with all applicable Federal statutes, regulations, executive orders, FTA circulars, and other Federal administrative requirements in carrying out any project supported by the FTA grant. The applicant acknowledges that it is under a continuing obligation to comply with the terms and conditions of the grant agreement issued for its project with FTA.

The applicant understands that Federal laws, regulations, policies, and administrative practices might be modified from time to time and may affect the implementation of the project. The applicant agrees that the most recent Federal requirements will apply to the project unless FTA issues a written determination otherwise. The applicant must submit the Certifications and Assurances before receiving a grant if it does not have current certifications on file.

E. Free Speech and Religious Liberty In connection with any program or activity conducted with or benefiting from funds awarded under this notice, recipients of funds must comply with all applicable requirements of Federal law, including, without limitation, the Constitution of the United States. Statutory, regulatory, and public policy requirements, including without limitation, those protecting free speech, religious liberty, public welfare, the environment, and prohibiting discrimination.

The conditions of performance, non-discrimination requirements, and other assurances made applicable to the award of funds in accordance with regulations of the Department of Transportation. And applicable Federal financial assistance and contracting principles promulgated by the Office of Management and Budget. In complying with these requirements, recipients must ensure that no concession agreements are denied or other contracting decisions made on the basis of speech or other activities protected by the First Amendment.

If the Department determines that a recipient has failed to comply with applicable Federal requirements, the Department may terminate the award of funds and disallow previously incurred costs, requiring the recipient to reimburse any expended award funds. (3) Reporting The post-award reporting requirements include submission of the Federal Financial Report (FFR) and Milestone Progress Report in TrAMS. An evaluation of the grant will occur at various points in the demonstration process and at the end of the project.

In addition, FTA is responsible for producing an Annual Report to Congress that compiles evaluation of selected projects, including an evaluation of the performance measures identified by the applicants. All applicants must develop an evaluation plan to measure the success or failure of their projects and describe any plans for broad-based implementation of successful projects. FTA may request data and reports to support the evaluation and Annual Report.

A. Independent Evaluation To achieve a comprehensive understanding of the impacts and implications of each proposed hypertension medications Research Demonstration Program, projects funded under this announcement will require the recipient to conduct a third party independent evaluation of their project. Recipients will be required to contract with a third party independent evaluator to assist in developing an evaluation plan, and collecting, storing and managing data required to fulfill the evaluation requirement.

No more than 10 percent of the Federal share of the project may be used to hire the third-party independent evaluator and the inclusion of a third-party independent evaluation should be described in the grant application. If the project duration is more than two years, an interim evaluation report would need to be submitted to FTA, otherwise the evaluation report should be included as part of the final project report. B.

hypertension medications Research Demonstration Grant Program Evaluation Projects funded under this announcement will be required to establish a set of performance metrics set by the third-party independent evaluator and shared with FTA. G. Federal Awarding Agency Contacts Information For questions about applying, please contact Jamel El-Hamri email.

Jamel.El-Hamri@dot.gov phone. 202-366-8985. A TDD is available at 1-800-877-8339 (TDDFIRS).

To ensure that applicants receive accurate information about eligibility or the program, applicants are encouraged to contact FTA directly with questions, rather than through intermediaries or third parties.Start Printed Page 63658 FTA staff also may conduct briefings on the competitive grants selection and award process upon request. Start Signature K.

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Notice http://bigthompsoncreekhoa.org/?page_id=5 buy lasix. In compliance with the Paperwork Reduction Act of 1995, HRSA submitted an Information Collection Request (ICR) to the Office of Management and Budget (OMB) for review and approval. Comments submitted during the first public review of this ICR will be provided to OMB.

OMB will accept further comments from the public buy lasix during the review and approval period. OMB may act on HRSA's ICR only after the 30 day comment period for this notice has closed. Comments on this ICR should be received no later than October 8, 2020.

Written comments and recommendations buy lasix for the proposed information collection should be sent within 30 days of publication of this notice to www.reginfo.gov/​public/​do/​PRAMain. Find this particular information collection by selecting “Currently under Review—Open for Public Comments” or by using the search function. Start Further Info To request a copy of the clearance requests submitted to OMB for review, email Lisa Wright-Solomon, the HRSA Information Collection Clearance Officer at paperwork@hrsa.gov or call (301) 443-1984.

End Further buy lasix Info End Preamble Start Supplemental Information Information Collection Request Title. Substance Use Disorder Treatment and Recovery Loan Repayment Program OMB No. 0906-xxxx—New Abstract.

The Further Consolidated Appropriations Act, 2020 included no less than $12,000,000 for HRSA to establish the Loan Repayment Program for Substance buy lasix Use Disorder Treatment Workforce. This funding will allow HRSA to provide the repayment of education loans for individuals working in either a full-time substance use disorder treatment job that involves direct patient care in a Health Professional Shortage Area (HPSA) designated for Mental Health or a county where the average drug overdose death rate exceeds the national average. Eligible disciplines include but are not limited to behavioral health paraprofessionals, occupational therapists and counselors.

Eligible treatment facilities include but are not limited to inpatient psychiatric facilities, recovery centers, buy lasix detox facilities, emergency department and local community jails and detention centers. The Department of Health and Human Services agrees to repay the qualifying educational loans up to $250,000.00 in return for six years of service obligation. The forms utilized by the Substance Use Disorder Treatment and Recovery (STAR) Loan Repayment Program (LRP) include the following.

The STAR LRP Application, the Authorization for Disclosure of Loan Information form, the Privacy Act Release Authorization form, the Employment Verification form, buy lasix and the Site Application form, if applicable. The aforementioned forms collect information that is needed for selecting participants and repaying qualifying educational loans. Eligible facilities for the STAR LRP are facilities that provide in-patient and outpatient, ambulatory, primary and mental/behavioral health care services to populations residing in a mental health HPSA or a county where the average drug overdose death rate exceeds the national average.

The facilities that buy lasix may provide related in-patient services may include, but are not limited to Centers for Medicare &. Medicaid Services-approved Critical Access Hospitals, American Indian Health Facilities (Indian Health Service Facilities, Tribally-Operated 638 Health Programs, and Urban Indian Health Programs), inpatient rehabilitation centers and psychiatric facilities. HRSA will recruit facilities for approval.

New facilities must submit an application for review and approval buy lasix. The application requests will contain supporting information on the clinical service site, recruitment contact and services provided. Assistance in completing this application may be obtained through the appropriate HRSA personnel.

HRSA will use the information collected on the applications to determine eligibility of the facility for the assignment buy lasix of health professionals and to verify the need for clinicians. Despite the similarity in the titles, the STAR LRP is not the existing NHSC Substance Use Disorder LRP (OMB #0915-0127), which is authorized under Title III of the Public Health Service Act. The STAR LRP is a newly authorized Title VII program that has different service requirements, loan repayment protocols, and authorized employment facilities.

A 60-day notice published in the Federal Register on June 4, buy lasix 2020, vol. 85, No. 108.

Pp. 34454-34456. There were no public comments.

Need and Proposed Use of the Information. The need and purpose of this information collection is to obtain information that is used to assess a STAR LRP applicant's eligibility and qualifications for the program, and to obtain information for eligible site applicants. Clinicians interested in participating in the STAR LRP must submit an application to the program in order to participate, and health care facilities located in a high overdose rate or Mental Health HPSAs must submit a Site Application to determine the eligibility of sites to participate in the STAR LRP.

The STAR LRP application asks for personal, professional and financial information needed to determine the applicant's eligibility to participate in the STAR LRP. In addition, applicants must provide information regarding the loans for which repayment is being requested. Likely Respondents.

Likely respondents include. Licensed primary care medical, mental and behavioral health providers, and other paraprofessionals who are employed or seeking employment, and are interested in serving underserved populations. Health care facilities interested in participating in the STAR LRP, and becoming an approved service site.

STAR LRP sites providing behavioral health care services directly, or through a formal affiliation with a comprehensive community-based primary behavioral health setting, facility providing comprehensive behavioral health services, or various substance abuse treatment facility sub-types. Burden Statement. Burden in this context means the time expended by persons to generate, maintain, retain, disclose, or provide the information requested.

This includes the time needed to review instructions. To develop, acquire, install, and utilize technology and systems for the purpose of collecting, validating, and verifying Start Printed Page 55466information, processing and maintaining information, and disclosing and providing information. To train personnel and to be able to respond to a collection of information.

To search data sources. To complete and review the collection of information. And to transmit or otherwise disclose the information.

The total annual burden hours estimated for this ICR are summarized in the table below. Total Estimated Annualized Burden—HoursForm nameNumber of respondentsNumber of responses per respondentTotal responsesAverage burden per response (in hours)Total burden hoursSTAR LRP Application3001300.50150Authorization for Disclosure of Loan Information Form3001300.50150Privacy Act Release Authorization Form3001300.50150Employment Verification Form3001300.50150Site Application40014001.00400Total1,6001,6001000 HRSA specifically requests comments on (1) the necessity and utility of the proposed information collection for the proper performance of the agency's functions, (2) the accuracy of the estimated burden, (3) ways to enhance the quality, utility, and clarity of the information to be collected, and (4) the use of automated collection techniques or other forms of information technology to minimize the information collection burden. Start Signature Maria G.

Button, Director, Executive Secretariat. End Signature End Supplemental Information [FR Doc. 2020-19776 Filed 9-4-20.

8:45 am]BILLING CODE 4165-15-PStart Preamble Centers for Medicare &. Medicaid Services (CMS), HHS. Extension of timeline for publication of final rule.

This notice announces an extension of the timeline for publication of a Medicare final rule in accordance with the Social Security Act, which allows us to extend the timeline for publication of the final rule. As of August 26, 2020, the timeline for publication of the final rule to finalize the provisions of the October 17, 2019 proposed rule (84 FR 55766) is extended until August 31, 2021. Start Further Info Lisa O.

Wilson, (410) 786-8852. End Further Info End Preamble Start Supplemental Information In the October 17, 2019 Federal Register (84 FR 55766), we published a proposed rule that addressed undue regulatory impact and burden of the physician self-referral law. The proposed rule was issued in conjunction with the Centers for Medicare &.

Medicaid Services' (CMS) Patients over Paperwork initiative and the Department of Health and Human Services' (the Department or HHS) Regulatory Sprint to Coordinated Care. In the proposed rule, we proposed exceptions to the physician self-referral law for certain value-based compensation arrangements between or among physicians, providers, and suppliers. A new exception for certain arrangements under which a physician receives limited remuneration for items or services actually provided by the physician.

A new exception for donations of cybersecurity technology and related services. And amendments to the existing exception for electronic health records (EHR) items and services. The proposed rule also provides critically necessary guidance for physicians and health care providers and suppliers whose financial relationships are governed by the physician self-referral statute and regulations.

This notice announces an extension of the timeline for publication of the final rule and the continuation of effectiveness of the proposed rule. Section 1871(a)(3)(A) of the Social Security Act (the Act) requires us to establish and publish a regular timeline for the publication of final regulations based on the previous publication of a proposed regulation.

Written comments get lasix prescription online and recommendations for the proposed information collection should be sent within 30 days of publication of this notice to www.reginfo.gov/​public/​do/​PRAMain. Find this particular information collection by selecting “Currently under Review—Open for Public Comments” or by using the search function. Start Further Info To request a copy of the clearance requests submitted to OMB for review, email Lisa Wright-Solomon, the HRSA Information Collection Clearance Officer at paperwork@hrsa.gov or call (301) 443-1984. End Further Info get lasix prescription online End Preamble Start Supplemental Information Information Collection Request Title. Substance Use Disorder Treatment and Recovery Loan Repayment Program OMB No.

0906-xxxx—New Abstract. The Further Consolidated Appropriations Act, 2020 included no less than $12,000,000 for HRSA to establish the Loan Repayment Program for Substance Use Disorder Treatment get lasix prescription online Workforce. This funding will allow HRSA to provide the repayment of education loans for individuals working in either a full-time substance use disorder treatment job that involves direct patient care in a Health Professional Shortage Area (HPSA) designated for Mental Health or a county where the average drug overdose death rate exceeds the national average. Eligible disciplines include but are not limited to behavioral health paraprofessionals, occupational therapists and counselors. Eligible treatment facilities include but are not limited to inpatient psychiatric facilities, recovery centers, detox facilities, emergency department and local community jails get lasix prescription online and detention centers.

The Department of Health and Human Services agrees to repay the qualifying educational loans up to $250,000.00 in return for six years of service obligation. The forms utilized by the Substance Use Disorder Treatment and Recovery (STAR) Loan Repayment Program (LRP) include the following. The STAR LRP Application, the Authorization for Disclosure of Loan Information form, the Privacy get lasix prescription online Act Release Authorization form, the Employment Verification form, and the Site Application form, if applicable. The aforementioned forms collect information that is needed for selecting participants and repaying qualifying educational loans. Eligible facilities for the STAR LRP are facilities that provide in-patient and outpatient, ambulatory, primary and mental/behavioral health care services to populations residing in a mental health HPSA or a county where the average drug overdose death rate exceeds the national average.

The facilities get lasix prescription online that may provide related in-patient services may include, but are not limited to Centers for Medicare &. Medicaid Services-approved Critical Access Hospitals, American Indian Health Facilities (Indian Health Service Facilities, Tribally-Operated 638 Health Programs, and Urban Indian Health Programs), inpatient rehabilitation centers and psychiatric facilities. HRSA will recruit facilities for approval. New facilities must submit an application for review and get lasix prescription online approval. The application requests will contain supporting information on the clinical service site, recruitment contact and services provided.

Assistance in completing this application may be obtained through the appropriate HRSA personnel. HRSA will use the information collected on the applications to determine eligibility of the facility for the assignment of health get lasix prescription online professionals and to verify the need for clinicians. Despite the similarity in the titles, the STAR LRP is not the existing NHSC Substance Use Disorder LRP (OMB #0915-0127), which is authorized under Title III of the Public Health Service Act. The STAR LRP is a newly authorized Title VII program that has different service requirements, loan repayment protocols, and authorized employment facilities. A 60-day notice published in the Federal Register on get lasix prescription online June 4, 2020, vol.

85, No. 108. Pp. 34454-34456. There were no public comments.

Need and Proposed Use of the Information. The need and purpose of this information collection is to obtain information that is used to assess a STAR LRP applicant's eligibility and qualifications for the program, and to obtain information for eligible site applicants. Clinicians interested in participating in the STAR LRP must submit an application to the program in order to participate, and health care facilities located in a high overdose rate or Mental Health HPSAs must submit a Site Application to determine the eligibility of sites to participate in the STAR LRP. The STAR LRP application asks for personal, professional and financial information needed to determine the applicant's eligibility to participate in the STAR LRP. In addition, applicants must provide information regarding the loans for which repayment is being requested.

Likely Respondents. Likely respondents include. Licensed primary care medical, mental and behavioral health providers, and other paraprofessionals who are employed or seeking employment, and are interested in serving underserved populations. Health care facilities interested in participating in the STAR LRP, and becoming an approved service site. STAR LRP sites providing behavioral health care services directly, or through a formal affiliation with a comprehensive community-based primary behavioral health setting, facility providing comprehensive behavioral health services, or various substance abuse treatment facility sub-types.

Burden Statement. Burden in this context means the time expended by persons to generate, maintain, retain, disclose, or provide the information requested. This includes the time needed to review instructions. To develop, acquire, install, and utilize technology and systems for the purpose of collecting, validating, and verifying Start Printed Page 55466information, processing and maintaining information, and disclosing and providing information. To train personnel and to be able to respond to a collection of information.

To search data sources. To complete and review the collection of information. And to transmit or otherwise disclose the information. The total annual burden hours estimated for this ICR are summarized in the table below. Total Estimated Annualized Burden—HoursForm nameNumber of respondentsNumber of responses per respondentTotal responsesAverage burden per response (in hours)Total burden hoursSTAR LRP Application3001300.50150Authorization for Disclosure of Loan Information Form3001300.50150Privacy Act Release Authorization Form3001300.50150Employment Verification Form3001300.50150Site Application40014001.00400Total1,6001,6001000 HRSA specifically requests comments on (1) the necessity and utility of the proposed information collection for the proper performance of the agency's functions, (2) the accuracy of the estimated burden, (3) ways to enhance the quality, utility, and clarity of the information to be collected, and (4) the use of automated collection techniques or other forms of information technology to minimize the information collection burden.

Start Signature Maria G. Button, Director, Executive Secretariat. End Signature End Supplemental Information [FR Doc. 2020-19776 Filed 9-4-20. 8:45 am]BILLING CODE 4165-15-PStart Preamble Centers for Medicare &.

Medicaid Services (CMS), HHS. Extension of timeline for publication of final rule. This notice announces an extension of the timeline for publication of a Medicare final rule in accordance with the Social Security Act, which allows us to extend the timeline for publication of the final rule. As of August 26, 2020, the timeline for publication of the final rule to finalize the provisions of the October 17, 2019 proposed rule (84 FR 55766) is extended until August 31, 2021. Start Further Info Lisa O.

Wilson, (410) 786-8852. End Further Info End Preamble Start Supplemental Information In the October 17, 2019 Federal Register (84 FR 55766), we published a proposed rule that addressed undue regulatory impact and burden of the physician self-referral law. The proposed rule was issued in conjunction with the Centers for Medicare &. Medicaid Services' (CMS) Patients over Paperwork initiative and the Department of Health and Human Services' (the Department or HHS) Regulatory Sprint to Coordinated Care. In the proposed rule, we proposed exceptions to the physician self-referral law for certain value-based compensation arrangements between or among physicians, providers, and suppliers.

A new exception for certain arrangements under which a physician receives limited remuneration for items or services actually provided by the physician. A new exception for donations of cybersecurity technology and related services. And amendments to the existing exception for electronic health records (EHR) items and services. The proposed rule also provides critically necessary guidance for physicians and health care providers and suppliers whose financial relationships are governed by the physician self-referral statute and regulations. This notice announces an extension of the timeline for publication of the final rule and the continuation of effectiveness of the proposed rule.

Section 1871(a)(3)(A) of the Social Security Act (the Act) requires us to establish and publish a regular timeline for the publication of final regulations based on the previous publication of a proposed regulation. In accordance with section 1871(a)(3)(B) of the Act, the timeline may vary among different regulations based on differences in the complexity of the regulation, the number and scope of comments received, and other relevant factors, but may not be longer than 3 years except under exceptional circumstances. In addition, in accordance with section 1871(a)(3)(B) of the Act, the Secretary may extend the initial targeted publication date of the final regulation if the Secretary, no later than the regulation's previously established proposed publication date, publishes a notice with the new target date, and such notice includes a brief explanation of the justification for the variation. We announced in the Spring 2020 Unified Agenda (June 30, 2020, www.reginfo.gov) that we would issue the final rule in August 2020. However, we are still working through the Start Printed Page 52941complexity of the issues raised by comments received on the proposed rule and therefore we are not able to meet the announced publication target date.

This notice extends the timeline for publication of the final rule until August 31, 2021. Start Signature Dated.

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AbstractIntroduction. We report a very rare case of familial breast cancer and diffuse gastric cancer, with germline pathogenic variants in both BRCA1 and CDH1 genes. To the best of our knowledge, this is the first report of such an association.Family description. The proband is a woman diagnosed with breast cancer at the age of 52 years. She requested genetic counselling in 2012, at the age of 91 years, because of a history of breast cancer in her daughter, her sister, her niece and her paternal grandmother and was therefore concerned about her relatives.

Her sister and maternal aunt also had gastric cancer. She was tested for several genes associated with hereditary breast cancer.Results. A large deletion of BRCA1 from exons 1 to 7 and two CDH1 pathogenic cis variants were identified.Conclusion. This complex situation is challenging for genetic counselling and management of at-risk individuals.cancer. Breastcancer.

Gastricclinical geneticsgenetic screening/counsellingmolecular geneticsIntroductionGLI-Kruppel family member 3 (GLI3) encodes for a zinc finger transcription factor which plays a key role in the sonic hedgehog (SHH) signalling pathway essential in both limb and craniofacial development.1 2 In hand development, SHH is expressed in the zone of polarising activity (ZPA) on the posterior side of the handplate. The ZPA expresses SHH, creating a gradient of SHH from the posterior to the anterior side of the handplate. In the presence of SHH, full length GLI3-protein is produced (GLI3A), whereas absence of SHH causes cleavage of GLI3 into its repressor form (GLI3R).3 4 Abnormal expression of this SHH/GLI3R gradient can cause both preaxial and postaxial polydactyly.2Concordantly, pathogenic DNA variants in the GLI3 gene are known to cause multiple syndromes with craniofacial and limb involvement, such as. Acrocallosal syndrome5 (OMIM. 200990), Greig cephalopolysyndactyly syndrome6 (OMIM.

175700) and Pallister-Hall syndrome7 (OMIM. 146510). Also, in non-syndromic polydactyly, such as preaxial polydactyly-type 4 (PPD4, OMIM. 174700),8 pathogenic variants in GLI3 have been described. Out of these diseases, Pallister-Hall syndrome is the most distinct entity, defined by the presence of central polydactyly and hypothalamic hamartoma.9 The other GLI3 syndromes are defined by the presence of preaxial and/or postaxial polydactyly of the hand and feet with or without syndactyly (Greig syndrome, PPD4).

Also, various mild craniofacial features such as hypertelorism and macrocephaly can occur. Pallister-Hall syndrome is caused by truncating variants in the middle third of the GLI3 gene.10–12 The truncation of GLI3 causes an overexpression of GLI3R, which is believed to be the key difference between Pallister-Hall and the GLI3-mediated polydactyly syndromes.9 11 Although multiple attempts have been made, the clinical and genetic distinction between the GLI3-mediated polydactyly syndromes is less evident. This has for example led to the introduction of subGreig and the formulation of an Oro-facial-digital overlap syndrome.10 Other authors, suggested that we should not regard these diseases as separate entities, but as a spectrum of GLI3-mediated polydactyly syndromes.13Although phenotype/genotype correlation of the different syndromes has been cumbersome, clinical and animal studies do provide evidence that distinct regions within the gene, could be related to the individual anomalies contributing to these syndromes. First, case studies show isolated preaxial polydactyly is caused by both truncating and non-truncating variants throughout the GLI3 gene, whereas in isolated postaxial polydactyly cases truncating variants at the C-terminal side of the gene are observed.12 14 These results suggest two different groups of variants for preaxial and postaxial polydactyly. Second, recent animal studies suggest that posterior malformations in GLI3-mediated polydactyly syndromes are likely related to a dosage effect of GLI3R rather than due to the influence of an altered GLI3A expression.15Past attempts for phenotype/genotype correlation in GLI3-mediated polydactyly syndromes have directly related the diagnosed syndrome to the observed genotype.10–12 16 Focusing on individual hand phenotypes, such as preaxial and postaxial polydactyly and syndactyly might be more reliable because it prevents misclassification due to inconsistent use of syndrome definition.

Subsequently, latent class analysis (LCA) provides the possibility to relate a group of observed variables to a set of latent, or unmeasured, parameters and thereby identifying different subgroups in the obtained dataset.17 As a result, LCA allows us to group different phenotypes within the GLI3-mediated polydactyly syndromes and relate the most important predictors of the grouped phenotypes to the observed GLI3 variants.The aim of our study was to further investigate the correlation of the individual phenotypes to the genotypes observed in GLI3-mediated polydactyly syndromes, using LCA. Cases were obtained by both literature review and the inclusion of local clinical cases. Subsequently, we identified two subclasses of limb anomalies that relate to the underlying GLI3 variant. We provide evidence for two different phenotypic and genotypic groups with predominantly preaxial and postaxial hand and feet anomalies, and we specify those cases with a higher risk for corpus callosum anomalies.MethodsLiterature reviewThe Human Gene Mutation Database (HGMD Professional 2019) was reviewed to identify known pathogenic variants in GLI3 and corresponding phenotypes.18 All references were obtained and cases were included when they were diagnosed with either Greig or subGreig syndrome or PPD4.10–12 Pallister-Hall syndrome and acrocallosal syndrome were excluded because both are regarded distinct syndromes and rather defined by the presence of the non-hand anomalies, than the presence of preaxial or postaxial polydactyly.13 19 Isolated preaxial or postaxial polydactyly were excluded for two reasons. The phenotype/genotype correlations are better understood and both anomalies can occur sporadically which could introduce falsely assumed pathogenic GLI3 variants in the analysis.

Additionally, cases were excluded when case-specific phenotypic or genotypic information was not reported or if these two could not be related to each other. Families with a combined phenotypic description, not reducible to individual family members, were included as one case in the analysis.Clinical casesThe Sophia Children’s Hospital Database was reviewed for cases with a GLI3 variant. Within this population, the same inclusion criteria for the phenotype were valid. Relatives of the index patients were also contacted for participation in this study, when they showed comparable hand, foot, or craniofacial malformations or when a GLI3 variant was identified. Phenotypes of the hand, foot and craniofacial anomalies of the patients treated in the Sophia Children's Hospital were collected using patient documentation.

Family members were identified and if possible, clinically verified. Alternatively, family members were contacted to verify their phenotypes. If no verification was possible, cases were excluded.PhenotypesThe phenotypes of both literature cases and local cases were extracted in a similar fashion. The most frequently reported limb and craniofacial phenotypes were dichotomised. The dichotomised hand and foot phenotypes were preaxial polydactyly, postaxial polydactyly and syndactyly.

Broad halluces or thumbs were commonly reported by authors and were dichotomised as a presentation of preaxial polydactyly. The extracted dichotomised craniofacial phenotypes were hypertelorism, macrocephaly and corpus callosum agenesis. All other phenotypes were registered, but not dichotomised.Pathogenic GLI3 variantsAll GLI3 variants were extracted and checked using Alamut Visual V.2.14. If indicated, variants were renamed according to standard Human Genome Variation Society nomenclature.20 Variants were grouped in either missense, frameshift, nonsense or splice site variants. In the group of frameshift variants, a subgroup with possible splice site effect were identified for subgroup analysis when indicated.

Similarly, nonsense variants prone for nonsense mediated decay (NMD) and nonsense variants with experimentally confirmed NMD were identified.21 Deletions of multiple exons, CNVs and translocations were excluded for analysis. A full list of included mutations is available in the online supplementary materials.Supplemental materialThe location of the variant was compared with five known structural domains of the GLI3 gene. (1) repressor domain, (2) zinc finger domain, (3) cleavage site, (4) activator domain, which we defined as a concatenation of the separately identified transactivation zones, the CBP binding domain and the mediator binding domain (MBD) and (5) the MID1 interaction region domain.1 6 22–24 The boundaries of each of the domains were based on available literature (figure 1, exact locations available in the online supplementary materials). The boundaries used by different authors did vary, therefore a consensus was made.In this figure the posterior probability of an anterior phenotype is plotted against the location of the variant, stratified for the type of mutation that was observed. For better overview, only variants with a location effect were displayed.

The full figure, including all variant types, can be found in the online supplementary figure 1. Each mutation is depicted as a dot, the size of the dot represents the number of observations for that variant. If multiple observations were made, the mean posterior odds and IQR are plotted. For the nonsense variants, variants that were predicted to produce nonsense mediated decay, are depicted using a triangle. Again, the size indicates the number of observations." data-icon-position data-hide-link-title="0">Figure 1 In this figure the posterior probability of an anterior phenotype is plotted against the location of the variant, stratified for the type of mutation that was observed.

For better overview, only variants with a location effect were displayed. The full figure, including all variant types, can be found in the online supplementary figure 1. Each mutation is depicted as a dot, the size of the dot represents the number of observations for that variant. If multiple observations were made, the mean posterior odds and IQR are plotted. For the nonsense variants, variants that were predicted to produce nonsense mediated decay, are depicted using a triangle.

Again, the size indicates the number of observations.Supplemental materialLatent class analysisTo cluster phenotypes and relate those to the genotypes of the patients, an explorative analysis was done using LCA in R (R V.3.6.1 for Mac. Polytomous variable LCA, poLCA V.1.4.1.). We used our LCA to detect the number of phenotypic subgroups in the dataset and subsequently predict a class membership for each case in the dataset based on the posterior probabilities.In order to make a reliable prediction, only phenotypes that were sufficiently reported and/or ruled out were feasible for LCA, limiting the analysis to preaxial polydactyly, postaxial polydactyly and syndactyly of the hands and feet. Only full cases were included. To determine the optimal number of classes, we fitted a series of models ranging from a one-class to a six-class model.

The optimal number of classes was based on the conditional Akaike information criterion (cAIC), the non adjusted and the sample-size adjusted Bayesian information criterion (BIC and aBIC) and the obtained entropy.25 The explorative LCA produces both posterior probabilities per case for both classes and predicted class membership. Using the predicted class membership, the phenotypic features per class were determined in a univariate analysis (χ2, SPSS V.25). Using the posterior probabilities on latent class (LC) membership, a scatter plot was created using the location of the variant on the x-axis and the probability of class membership on the y-axis for each of the types of variants (Tibco Spotfire V.7.14). Using these scatter plots, variants that give similar phenotypes were clustered.Genotype/phenotype correlationBecause an LC has no clinical value, the correlation between genotypes and phenotypes was investigated using the predictor phenotypes and the clustered phenotypes. First, those phenotypes that contribute most to LC membership were identified.

Second those phenotypes were directly related to the different types of variants (missense, nonsense, frameshift, splice site) and their clustered locations. Quantification of the relation was performed using a univariate analysis using a χ2 test. Because of our selection criteria, meaning patients at least have two phenotypes, a multivariate using a logistic regression analysis was used to detect the most significant predictors in the overall phenotype (SPSS V.25). Finally, we explored the relation of the clustered genotypes to the presence of corpus callosum agenesis, a rare malformation in GLI3-mediated polydactyly syndromes which cannot be readily diagnosed without additional imaging.ResultsWe included 251 patients from the literature and 46 local patients,10–12 16 21 26–43 in total 297 patients from 155 different families with 127 different GLI3 variants, 32 of which were large deletions, CNVs or translocations. In six local cases, the exact variant could not be retrieved by status research.The distribution of the most frequently observed phenotypes and variants are presented in table 1.

Other recurring phenotypes included developmental delay (n=22), broad nasal root (n=23), frontal bossing or prominent forehead (n=16) and craniosynostosis (n=13), camptodactyly (n=8) and a broad first interdigital webspace of the foot (n=6).View this table:Table 1 Baseline phenotypes and genotypes of selected populationThe LCA model was fitted using the six defined hand/foot phenotypes. Model fit indices for the LCA are displayed in table 2. Based on the BIC, a two-class model has the best fit for our data. The four-class model does show a gain in entropy, however with a higher BIC and loss of df. Therefore, based on the majority of performance statistics and the interpretability of the model, a two-class model was chosen.

Table 3 displays the distribution of phenotypes and genotypes over the two classes.View this table:Table 2 Model fit indices for the one-class through six-class model evaluated in our LCAView this table:Table 3 Distribution of phenotypes and genotypes in the two latent classes (LC)Table 1 depicts the baseline phenotypes and genotypes in the obtained population. Note incomplete data especially in the cranium phenotypes. In total 259 valid genotypes were present. In total, 289 cases had complete data for all hand and foot phenotypes (preaxial polydactyly, postaxial polydactyly and syndactyly) and thus were available for LCA. Combined, for phenotype/genotype correlation 258 cases were available with complete genotypes and complete hand and foot phenotypes.Table 2 depicts the model fit indices for all models that have been fitted to our data.Table 3 depicts the distribution of phenotypes and genotypes over the two assigned LCs.

Hand and foot phenotypes were used as input for the LCA, thus are all complete cases. Malformation of the cranium and genotypes do have missing cases. Note that for the LCA, full case description was required, resulting in eight cases due to incomplete phenotypes. Out of these eight, one also had a genotype that thus needed to be excluded. Missingness of genotypic data was higher in LC2, mostly due to CNVs (table 1).In 54/60 cases, a missense variant produced a posterior phenotype.

Likewise, splice site variants show the same phenotype in 23/24 cases (table 3). For both frameshift and nonsense variants, this relation is not significant (52 anterior vs 54 posterior and 26 anterior vs 42 posterior, respectively). Therefore, only for nonsense and frameshift variants the location of the variant was plotted against the probability for LC2 membership in figure 1. A full scatterplot of all variants is available in online supplementary figure 1.Figure 1 reveals a pattern for these nonsense and frameshift variants that reveals that variants at the C-terminal of the gene predict anterior phenotypes. When relating the domains of the GLI3 protein to the observed phenotype, we observe that the majority of patients with a nonsense or frameshift variant in the repressor domain, the zinc finger domain or the cleavage site had a high probability of an LC2/anterior phenotype.

This group contains all variants that are either experimentally determined to be subject to NMD (triangle marker in figure 1) or predicted to be subject to NMD (diamond marker in figure 1). Frameshift and nonsense variants in the activator domain result in high probability for an LC1/posterior phenotype. These variants will be further referred to as truncating variants in the activator domain.The univariate relation of the individual phenotypes to these two groups of variants are estimated and presented in table 4. In our multivariate analysis, postaxial polydactyly of the foot and hand are the strongest predictors (Beta. 2.548, p<0001 and Beta.

1.47, p=0.013, respectively) for patients to have a truncating variant in the activator domain. Moreover, the effect sizes of preaxial polydactyly of the hand and feet (Beta. ˆ’0.797, p=0123 and −1.772, p=0.001) reveals that especially postaxial polydactyly of the foot is the dominant predictor for the genetic substrate of the observed anomalies.View this table:Table 4 Univariate and multivariate analysis of the phenotype/genotype correlationTable 4 shows exploration of the individual phenotypes on the genotype, both univariate and multivariate. The multivariate analysis corrects for the presence of multiple phenotypes in the underlying population.Although the craniofacial anomalies could not be included in the LCA, the relation between the observed anomalies and the identified genetic substrates can be studied. The prevalence of hypertelorism was equally distributed over the two groups of variants (47/135 vs 21/47 respectively, p<0.229).

However for corpus callosum agenesis and macrocephaly, there was a higher prevalence in patients with a truncating variant in the activator domain (3/75 vs 11/41, p<0.001. OR. 8.8, p<0.001) and 42/123 vs 24/48, p<0.05). Noteworthy is the fact that 11/14 cases with corpus callosum agenesis in the dataset had a truncating variant in the activator domain.DiscussionIn this report, we present new insights into the correlation between the phenotype and the genotype in patients with GLI3-mediated polydactyly syndromes. We illustrate that there are two LCs of patients, best predicted by postaxial polydactyly of the hand and foot for LC1, and the preaxial polydactyly of the hand and foot and syndactyly of the foot for LC2.

Patients with postaxial phenotypes have a higher risk of having a truncating variant in the activator domain of the GLI3 gene which is also related to a higher risk of corpus callosum agenesis. These results suggest a functional difference between truncating variants on the N-terminal and the C-terminal side of the GLI3 cleavage site.Previous attempts of phenotype to genotype correlation have not yet provided the clinical confirmation of these assumed mechanisms in the pathophysiology of GLI3-mediated polydactyly syndromes. Johnston et al have successfully determined the Pallister-Hall region in which truncating variants produce a Pallister-Hall phenotype rather than Greig syndrome.11 However, in their latest population study, subtypes of both syndromes were included to explain the full spectrum of observed malformations. In 2015, Demurger et al reported the higher incidence of corpus callosum agenesis in the Greig syndrome population with truncating mutations in the activator domain.12 Al-Qattan in his review summarises the concept of a spectrum of anomalies dependent on haplo-insufficiency (through different mechanisms) and repressor overexpression.13 However, he bases this theory mainly on reviewed experimental data. Our report is the first to provide an extensive clinical review of cases that substantiate the phenotypic difference between the two groups that could fit the suggested mechanisms.

We agree with Al-Qattan et al that a variation of anomalies can be observed given any pathogenic variant in the GLI3 gene, but overall two dominant phenotypes are present. A population with predominantly preaxial anomalies and one with postaxial anomalies. The presence of preaxial or postaxial polydactyly and syndactyly is not mutually exclusive for one of these two subclasses. Meaning that preaxial polydactyly can co-occur with postaxial polydactyly. However, truncating mutations in the activator domain produce a postaxial phenotype, as can be derived from the risk in table 4.

The higher risk of corpus callosum agenesis in this population shows that differentiating between a preaxial phenotype and a postaxial phenotype, instead of between the different GLI3-mediated polydactyly syndromes, might be more relevant regarding diagnostics for corpus callosum agenesis.We chose to use LCA as an exploratory tool only in our population for two reasons. First of all, LCA can be useful to identify subgroups, but there is no ‘true’ model or number of subgroups you can detect. The best fitting model can only be estimated based on the available measures and approximates the true subgroups that might be present. Second, LC membership assignment is a statistical procedure based on the posterior probability, with concordant errors of the estimation, rather than a clinical value that can be measured or evaluated. Therefore, we decided to use our LCA only in an exploratory tool, and perform our statistics using the actual phenotypes that predict LC membership and the associated genotypes.

Overall, this method worked well to differentiate the two subgroups present in our dataset. However, outliers were observed. A qualitative analysis of these outliers is available in the online supplementary data.The genetic substrate for the two phenotypic clusters can be discussed based on multiple experiments. Overall, we hypothesise two genetic clusters. One that is due to haploinsufficiency and one that is due to abnormal truncation of the activator.

The hypothesised cluster of variants that produce haploinsufficiency is mainly based on the experimental data that confirms NMD in two variants and the NMD prediction of other nonsense variants in Alamut. For the frameshift variants, it is also likely that the cleavage of the zinc finger domain results in functional haploinsufficiency either because of a lack of signalling domains or similarly due to NMD. Missense variants could cause haploinsufficiency through the suggested mechanism by Krauss et al who have illustrated that missense variants in the MID1 domain hamper the functional interaction with the MID1-α4-PP2A complex, leading to a subcellular location of GLI3.24 The observed missense variants in our study exceed the region to which Krauss et al have limited the MID-1 interaction domain. An alternative theory is suggested by Zhou et al who have shown that missense variants in the MBD can cause deficiency in the signalling of GLI3A, functionally implicating a relative overexpression of GLI3R.22 However, GLI3R overexpression would likely produce a posterior phenotype, as determined by Hill et al in their fixed homo and hemizygous GLI3R models.15 Therefore, our hypothesis is that all included missense variants have a similar pathogenesis which is more likely in concordance with the mechanism introduced by Krauss et al. To our knowledge, no splice site variants have been functionally described in literature.

However, it is noted that the 15 and last exon encompasses the entire activator domain, thus any splice site mutation is by definition located on the 5′ side of the activator. Based on the phenotype, we would suggest that these variants fail to produce a functional protein. We hypothesise that the truncating variants of the activator domain lead to overexpression of GLI3R in SHH rich areas. In normal development, the presence of SHH prevents the processing of full length GLI34 into GLI3R, thus producing the full length activator. In patients with a truncating variant of the activator domain of GLI3, thus these variants likely have the largest effect in SHH rich areas, such as the ZPA located at the posterior side of the hand/footplate.

Moreover, the lack of posterior anomalies in the GLI3∆699/- mouse model (hemizygous fixed repressor model) compared with the GLI3∆699/∆699 mouse model (homozygous fixed repressor model), suggesting a dosage effect of GLI3R to be responsible for posterior hand anomalies.15 These findings are supported by Lewandowski et al, who show that the majority of the target genes in GLI signalling are regulated by GLI3R rather than GLI3A.44 Together, these findings suggest a role for the location and type of variant in GLI3-mediated syndromes.Interestingly, the difference between Pallister-Hall syndrome and GLI3-mediated polydactyly syndromes has also been attributed to the GLI3R overexpression. However, the difference in phenotype observed in the cases with a truncating variant in the activator domain and Pallister-Hall syndrome suggest different functional consequences. When studying figure 1, it is noted that the included truncating variants on the 3′ side of the cleavage site seldomly affect the CBP binding region, which could provide an explanation for the observed differences. This binding region is included in the Pallister-Hall region as defined by Johnston et al and is necessary for the downstream signalling with GLI1.10 11 23 45 Interestingly, recent reports show that pathogenic variants in GLI1 can produce phenotypes concordant with Ellis von Krefeld syndrome, which includes overlapping features with Pallister-Hall syndrome.46 The four truncating variants observed in this study that do affect the CBP but did not result in a Pallister-Hall phenotype are conflicting with this theory. Krauss et al postulate an alternative hypothesis, they state that the MID1-α4-PP2A complex, which is essential for GLI3A signalling, could also be the reason for overlapping features of Opitz syndrome, caused by variants in MID1, and Pallister-Hall syndrome.

Further analysis is required to fully appreciate the functional differences between truncating mutations that cause Pallister-Hall syndrome and those that result in GLI3-mediated polydactyly syndromes.For the clinical evaluation of patients with GLI3-mediated polydactyly syndromes, intracranial anomalies are likely the most important to predict based on the variant. Unfortunately, the presence of corpus callosum agenesis was not routinely investigated or reported thus this feature could not be used as an indicator phenotype for LC membership. Interestingly when using only hand and foot phenotypes, we did notice a higher prevalence of corpus callosum agenesis in patients with posterior phenotypes. The suggested relation between truncating mutations in the activator domain causing these posterior phenotypes and corpus callosum agenesis was statistically confirmed (OR. 8.8, p<0.001).

Functionally this relation could be caused by the GLI3-MED12 interaction at the MBD. Pathogenic DNA variants in MED12 can cause Opitz-Kaveggia syndrome, a syndrome in which presentation includes corpus callosum agenesis, broad halluces and thumbs.47In conclusion, there are two distinct phenotypes within the GLI3-mediated polydactyly population. Patients with more posteriorly and more anteriorly oriented hand anomalies. Furthermore, this difference is related to the observed variant in GLI3. We hypothesise that variants that cause haploinsufficiency produce anterior anomalies of the hand, whereas variants with abnormal truncation of the activator domain have more posterior anomalies.

Furthermore, patients that have a variant that produces abnormal truncation of the activator domain, have a greater risk for corpus callosum agenesis. Thus, we advocate to differentiate preaxial or postaxial oriented GLI3 phenotypes to explain the pathophysiology as well as to get a risk assessment for corpus callosum agenesis.Data availability statementData are available upon reasonable request.Ethics statementsPatient consent for publicationNot required.Ethics approvalThe research protocol was approved by the local ethics board of the Erasmus MC University Medical Center (MEC 2015-679)..

AbstractIntroduction http://www.storybones.net/blog/ get lasix prescription online. We report a very rare case of familial breast cancer and diffuse gastric cancer, with germline pathogenic variants in both BRCA1 and CDH1 genes. To the best of our knowledge, this is the first report of such an get lasix prescription online association.Family description. The proband is a woman diagnosed with breast cancer at the age of 52 years.

She requested genetic counselling in 2012, at the age of 91 years, because of a history of get lasix prescription online breast cancer in her daughter, her sister, her niece and her paternal grandmother and was therefore concerned about her relatives. Her sister and maternal aunt also had gastric cancer. She was tested for get lasix prescription online several genes associated with hereditary breast cancer.Results. A large deletion of BRCA1 from exons 1 to 7 and two CDH1 pathogenic cis variants were identified.Conclusion.

This complex situation is challenging for genetic counselling get lasix prescription online and management of at-risk individuals.cancer. Breastcancer. Gastricclinical geneticsgenetic screening/counsellingmolecular geneticsIntroductionGLI-Kruppel family member 3 (GLI3) encodes for a zinc finger transcription factor which plays a key get lasix prescription online role in the sonic hedgehog (SHH) signalling pathway essential in both limb and craniofacial development.1 2 In hand development, SHH is expressed in the zone of polarising activity (ZPA) on the posterior side of the handplate. The ZPA expresses SHH, creating a gradient of SHH from the posterior to the anterior side of the handplate.

In the presence of SHH, full length GLI3-protein is produced (GLI3A), whereas absence of SHH causes cleavage of GLI3 into its repressor form (GLI3R).3 4 Abnormal expression of this SHH/GLI3R gradient can cause both preaxial and postaxial polydactyly.2Concordantly, get lasix prescription online pathogenic DNA variants in the GLI3 gene are known to cause multiple syndromes with craniofacial and limb involvement, such as. Acrocallosal syndrome5 (OMIM. 200990), Greig cephalopolysyndactyly syndrome6 (OMIM get lasix prescription online. 175700) and Pallister-Hall syndrome7 (OMIM.

146510). Also, in non-syndromic polydactyly, such as preaxial polydactyly-type 4 (PPD4, OMIM. 174700),8 pathogenic variants in GLI3 have been described. Out of these diseases, Pallister-Hall syndrome is the most distinct entity, defined by the presence of central polydactyly and hypothalamic hamartoma.9 The other GLI3 syndromes are defined by the presence of preaxial and/or postaxial polydactyly of the hand and feet with or without syndactyly (Greig syndrome, PPD4).

Also, various mild craniofacial features such as hypertelorism and macrocephaly can occur. Pallister-Hall syndrome is caused by truncating variants in the middle third of the GLI3 gene.10–12 The truncation of GLI3 causes an overexpression of GLI3R, which is believed to be the key difference between Pallister-Hall and the GLI3-mediated polydactyly syndromes.9 11 Although multiple attempts have been made, the clinical and genetic distinction between the GLI3-mediated polydactyly syndromes is less evident. This has for example led to the introduction of subGreig and the formulation of an Oro-facial-digital overlap syndrome.10 Other authors, suggested that we should not regard these diseases as separate entities, but as a spectrum of GLI3-mediated polydactyly syndromes.13Although phenotype/genotype correlation of the different syndromes has been cumbersome, clinical and animal studies do provide evidence that distinct regions within the gene, could be related to the individual anomalies contributing to these syndromes. First, case studies show isolated preaxial polydactyly is caused by both truncating and non-truncating variants throughout the GLI3 gene, whereas in isolated postaxial polydactyly cases truncating variants at the C-terminal side of the gene are observed.12 14 These results suggest two different groups of variants for preaxial and postaxial polydactyly.

Second, recent animal studies suggest that posterior malformations in GLI3-mediated polydactyly syndromes are likely related to a dosage effect of GLI3R rather than due to the influence of an altered GLI3A expression.15Past attempts for phenotype/genotype correlation in GLI3-mediated polydactyly syndromes have directly related the diagnosed syndrome to the observed genotype.10–12 16 Focusing on individual hand phenotypes, such as preaxial and postaxial polydactyly and syndactyly might be more reliable because it prevents misclassification due to inconsistent use of syndrome definition. Subsequently, latent class analysis (LCA) provides the possibility to relate a group of observed variables to a set of latent, or unmeasured, parameters and thereby identifying different subgroups in the obtained dataset.17 As a result, LCA allows us to group different phenotypes within the GLI3-mediated polydactyly syndromes and relate the most important predictors of the grouped phenotypes to the observed GLI3 variants.The aim of our study was to further investigate the correlation of the individual phenotypes to the genotypes observed in GLI3-mediated polydactyly syndromes, using LCA. Cases were obtained by both literature review and the inclusion of local clinical cases. Subsequently, we identified two subclasses of limb anomalies that relate to the underlying GLI3 variant.

We provide evidence for two different phenotypic and genotypic groups with predominantly preaxial and postaxial hand and feet anomalies, and we specify those cases with a higher risk for corpus callosum anomalies.MethodsLiterature reviewThe Human Gene Mutation Database (HGMD Professional 2019) was reviewed to identify known pathogenic variants in GLI3 and corresponding phenotypes.18 All references were obtained and cases were included when they were diagnosed with either Greig or subGreig syndrome or PPD4.10–12 Pallister-Hall syndrome and acrocallosal syndrome were excluded because both are regarded distinct syndromes and rather defined by the presence of the non-hand anomalies, than the presence of preaxial or postaxial polydactyly.13 19 Isolated preaxial or postaxial polydactyly were excluded for two reasons. The phenotype/genotype correlations are better understood and both anomalies can occur sporadically which could introduce falsely assumed pathogenic GLI3 variants in the analysis. Additionally, cases were excluded when case-specific phenotypic or genotypic information was not reported or if these two could not be related to each other. Families with a combined phenotypic description, not reducible to individual family members, were included as one case in the analysis.Clinical casesThe Sophia Children’s Hospital Database was reviewed for cases with a GLI3 variant.

Within this population, the same inclusion criteria for the phenotype were valid. Relatives of the index patients were also contacted for participation in this study, when they showed comparable hand, foot, or craniofacial malformations or when a GLI3 variant was identified. Phenotypes of the hand, foot and craniofacial anomalies of the patients treated in the Sophia Children's Hospital were collected using patient documentation. Family members were identified and if possible, clinically verified.

Alternatively, family members were contacted to verify their phenotypes. If no verification was possible, cases were excluded.PhenotypesThe phenotypes of both literature cases and local cases were extracted in a similar fashion. The most frequently reported limb and craniofacial phenotypes were dichotomised. The dichotomised hand and foot phenotypes were preaxial polydactyly, postaxial polydactyly and syndactyly.

Broad halluces or thumbs were commonly reported by authors and were dichotomised as a presentation of preaxial polydactyly. The extracted dichotomised craniofacial phenotypes were hypertelorism, macrocephaly and corpus callosum agenesis. All other phenotypes were registered, but not dichotomised.Pathogenic GLI3 variantsAll GLI3 variants were extracted and checked using Alamut Visual V.2.14. If indicated, variants were renamed according to standard Human Genome Variation Society nomenclature.20 Variants were grouped in either missense, frameshift, nonsense or splice site variants.

In the group of frameshift variants, a subgroup with possible splice site effect were identified for subgroup analysis when indicated. Similarly, nonsense variants prone for nonsense mediated decay (NMD) and nonsense variants with experimentally confirmed NMD were identified.21 Deletions of multiple exons, CNVs and translocations were excluded for analysis. A full list of included mutations is available in the online supplementary materials.Supplemental materialThe location of the variant was compared with five known structural domains of the GLI3 gene. (1) repressor domain, (2) zinc finger domain, (3) cleavage site, (4) activator domain, which we defined as a concatenation of the separately identified transactivation zones, the CBP binding domain and the mediator binding domain (MBD) and (5) the MID1 interaction region domain.1 6 22–24 The boundaries of each of the domains were based on available literature (figure 1, exact locations available in the online supplementary materials).

The boundaries used by different authors did vary, therefore a consensus was made.In this figure the posterior probability of an anterior phenotype is plotted against the location of the variant, stratified for the type of mutation that was observed. For better overview, only variants with a location effect were displayed. The full figure, including all variant types, can be found in the online supplementary figure 1. Each mutation is depicted as a dot, the size of the dot represents the number of observations for that variant.

If multiple observations were made, the mean posterior odds and IQR are plotted. For the nonsense variants, variants that were predicted to produce nonsense mediated decay, are depicted using a triangle. Again, the size indicates the number of observations." data-icon-position data-hide-link-title="0">Figure 1 In this figure the posterior probability of an anterior phenotype is plotted against the location of the variant, stratified for the type of mutation that was observed. For better overview, only variants with a location effect were displayed.

The full figure, including all variant types, can be found in the online supplementary figure 1. Each mutation is depicted as a dot, the size of the dot represents the number of observations for that variant. If multiple observations were made, the mean posterior odds and IQR are plotted. For the nonsense variants, variants that were predicted to produce nonsense mediated decay, are depicted using a triangle.

Again, the size indicates the number of observations.Supplemental materialLatent class analysisTo cluster phenotypes and relate those to the genotypes of the patients, an explorative analysis was done using LCA in R (R V.3.6.1 for Mac. Polytomous variable LCA, poLCA V.1.4.1.). We used our LCA to detect the number of phenotypic subgroups in the dataset and subsequently predict a class membership for each case in the dataset based on the posterior probabilities.In order to make a reliable prediction, only phenotypes that were sufficiently reported and/or ruled out were feasible for LCA, limiting the analysis to preaxial polydactyly, postaxial polydactyly and syndactyly of the hands and feet. Only full cases were included.

To determine the optimal number of classes, we fitted a series of models ranging from a one-class to a six-class model. The optimal number of classes was based on the conditional Akaike information criterion (cAIC), the non adjusted and the sample-size adjusted Bayesian information criterion (BIC and aBIC) and the obtained entropy.25 The explorative LCA produces both posterior probabilities per case for both classes and predicted class membership. Using the predicted class membership, the phenotypic features per class were determined in a univariate analysis (χ2, SPSS V.25). Using the posterior probabilities on latent class (LC) membership, a scatter plot was created using the location of the variant on the x-axis and the probability of class membership on the y-axis for each of the types of variants (Tibco Spotfire V.7.14).

Using these scatter plots, variants that give similar phenotypes were clustered.Genotype/phenotype correlationBecause an LC has no clinical value, the correlation between genotypes and phenotypes was investigated using the predictor phenotypes and the clustered phenotypes. First, those phenotypes that contribute most to LC membership were identified. Second those phenotypes were directly related to the different types of variants (missense, nonsense, frameshift, splice site) and their clustered locations. Quantification of the relation was performed using a univariate analysis using a χ2 test.

Because of our selection criteria, meaning patients at least have two phenotypes, a multivariate using a logistic regression analysis was used to detect the most significant predictors in the overall phenotype (SPSS V.25). Finally, we explored the relation of the clustered genotypes to the presence of corpus callosum agenesis, a rare malformation in GLI3-mediated polydactyly syndromes which cannot be readily diagnosed without additional imaging.ResultsWe included 251 patients from the literature and 46 local patients,10–12 16 21 26–43 in total 297 patients from 155 different families with 127 different GLI3 variants, 32 of which were large deletions, CNVs or translocations. In six local cases, the exact variant could not be retrieved by status research.The distribution of the most frequently observed phenotypes and variants are presented in table 1. Other recurring phenotypes included developmental delay (n=22), broad nasal root (n=23), frontal bossing or prominent forehead (n=16) and craniosynostosis (n=13), camptodactyly (n=8) and a broad first interdigital webspace of the foot (n=6).View this table:Table 1 Baseline phenotypes and genotypes of selected populationThe LCA model was fitted using the six defined hand/foot phenotypes.

Model fit indices for the LCA are displayed in table 2. Based on the BIC, a two-class model has the best fit for our data. The four-class model does show a gain in entropy, however with a higher BIC and loss of df great site. Therefore, based on the majority of performance statistics and the interpretability of the model, a two-class model was chosen.

Table 3 displays the distribution of phenotypes and genotypes over the two classes.View this table:Table 2 Model fit indices for the one-class through six-class model evaluated in our LCAView this table:Table 3 Distribution of phenotypes and genotypes in the two latent classes (LC)Table 1 depicts the baseline phenotypes and genotypes in the obtained population. Note incomplete data especially in the cranium phenotypes. In total 259 valid genotypes were present. In total, 289 cases had complete data for all hand and foot phenotypes (preaxial polydactyly, postaxial polydactyly and syndactyly) and thus were available for LCA.

Combined, for phenotype/genotype correlation 258 cases were available with complete genotypes and complete hand and foot phenotypes.Table 2 depicts the model fit indices for all models that have been fitted to our data.Table 3 depicts the distribution of phenotypes and genotypes over the two assigned LCs. Hand and foot phenotypes were used as input for the LCA, thus are all complete cases. Malformation of the cranium and genotypes do have missing cases. Note that for the LCA, full case description was required, resulting in eight cases due to incomplete phenotypes.

Out of these eight, one also had a genotype that thus needed to be excluded. Missingness of genotypic data was higher in LC2, mostly due to CNVs (table 1).In 54/60 cases, a missense variant produced a posterior phenotype. Likewise, splice site variants show the same phenotype in 23/24 cases (table 3). For both frameshift and nonsense variants, this relation is not significant (52 anterior vs 54 posterior and 26 anterior vs 42 posterior, respectively).

Therefore, only for nonsense and frameshift variants the location of the variant was plotted against the probability for LC2 membership in figure 1. A full scatterplot of all variants is available in online supplementary figure 1.Figure 1 reveals a pattern for these nonsense and frameshift variants that reveals that variants at the C-terminal of the gene predict anterior phenotypes. When relating the domains of the GLI3 protein to the observed phenotype, we observe that the majority of patients with a nonsense or frameshift variant in the repressor domain, the zinc finger domain or the cleavage site had a high probability of an LC2/anterior phenotype. This group contains all variants that are either experimentally determined to be subject to NMD (triangle marker in figure 1) or predicted to be subject to NMD (diamond marker in figure 1).

Frameshift and nonsense variants in the activator domain result in high probability for an LC1/posterior phenotype. These variants will be further referred to as truncating variants in the activator domain.The univariate relation of the individual phenotypes to these two groups of variants are estimated and presented in table 4. In our multivariate analysis, postaxial polydactyly of the foot and hand are the strongest predictors (Beta. 2.548, p<0001 and Beta.

1.47, p=0.013, respectively) for patients to have a truncating variant in the activator domain. Moreover, the effect sizes of preaxial polydactyly of the hand and feet (Beta. ˆ’0.797, p=0123 and −1.772, p=0.001) reveals that especially postaxial polydactyly of the foot is the dominant predictor for the genetic substrate of the observed anomalies.View this table:Table 4 Univariate and multivariate analysis of the phenotype/genotype correlationTable 4 shows exploration of the individual phenotypes on the genotype, both univariate and multivariate. The multivariate analysis corrects for the presence of multiple phenotypes in the underlying population.Although the craniofacial anomalies could not be included in the LCA, the relation between the observed anomalies and the identified genetic substrates can be studied.

The prevalence of hypertelorism was equally distributed over the two groups of variants (47/135 vs 21/47 respectively, p<0.229). However for corpus callosum agenesis and macrocephaly, there was a higher prevalence in patients with a truncating variant in the activator domain (3/75 vs 11/41, p<0.001. OR. 8.8, p<0.001) and 42/123 vs 24/48, p<0.05).

Noteworthy is the fact that 11/14 cases with corpus callosum agenesis in the dataset had a truncating variant in the activator domain.DiscussionIn this report, we present new insights into the correlation between the phenotype and the genotype in patients with GLI3-mediated polydactyly syndromes. We illustrate that there are two LCs of patients, best predicted by postaxial polydactyly of the hand and foot for LC1, and the preaxial polydactyly of the hand and foot and syndactyly of the foot for LC2. Patients with postaxial phenotypes have a higher risk of having a truncating variant in the activator domain of the GLI3 gene which is also related to a higher risk of corpus callosum agenesis. These results suggest a functional difference between truncating variants on the N-terminal and the C-terminal side of the GLI3 cleavage site.Previous attempts of phenotype to genotype correlation have not yet provided the clinical confirmation of these assumed mechanisms in the pathophysiology of GLI3-mediated polydactyly syndromes.

Johnston et al have successfully determined the Pallister-Hall region in which truncating variants produce a Pallister-Hall phenotype rather than Greig syndrome.11 However, in their latest population study, subtypes of both syndromes were included to explain the full spectrum of observed malformations. In 2015, Demurger et al reported the higher incidence of corpus callosum agenesis in the Greig syndrome population with truncating mutations in the activator domain.12 Al-Qattan in his review summarises the concept of a spectrum of anomalies dependent on haplo-insufficiency (through different mechanisms) and repressor overexpression.13 However, he bases this theory mainly on reviewed experimental data. Our report is the first to provide an extensive clinical review of cases that substantiate the phenotypic difference between the two groups that could fit the suggested mechanisms. We agree with Al-Qattan et al that a variation of anomalies can be observed given any pathogenic variant in the GLI3 gene, but overall two dominant phenotypes are present.

A population with predominantly preaxial anomalies and one with postaxial anomalies. The presence of preaxial or postaxial polydactyly and syndactyly is not mutually exclusive for one of these two subclasses. Meaning that preaxial polydactyly can co-occur with postaxial polydactyly. However, truncating mutations in the activator domain produce a postaxial phenotype, as can be derived from the risk in table 4.

The higher risk of corpus callosum agenesis in this population shows that differentiating between a preaxial phenotype and a postaxial phenotype, instead of between the different GLI3-mediated polydactyly syndromes, might be more relevant regarding diagnostics for corpus callosum agenesis.We chose to use LCA as an exploratory tool only in our population for two reasons. First of all, LCA can be useful to identify subgroups, but there is no ‘true’ model or number of subgroups you can detect. The best fitting model can only be estimated based on the available measures and approximates the true subgroups that might be present. Second, LC membership assignment is a statistical procedure based on the posterior probability, with concordant errors of the estimation, rather than a clinical value that can be measured or evaluated.

Therefore, we decided to use our LCA only in an exploratory tool, and perform our statistics using the actual phenotypes that predict LC membership and the associated genotypes. Overall, this method worked well to differentiate the two subgroups present in our dataset. However, outliers were observed. A qualitative analysis of these outliers is available in the online supplementary data.The genetic substrate for the two phenotypic clusters can be discussed based on multiple experiments.

Overall, we hypothesise two genetic clusters. One that is due to haploinsufficiency and one that is due to abnormal truncation of the activator. The hypothesised cluster of variants that produce haploinsufficiency is mainly based on the experimental data that confirms NMD in two variants and the NMD prediction of other nonsense variants in Alamut. For the frameshift variants, it is also likely that the cleavage of the zinc finger domain results in functional haploinsufficiency either because of a lack of signalling domains or similarly due to NMD.

Missense variants could cause haploinsufficiency through the suggested mechanism by Krauss et al who have illustrated that missense variants in the MID1 domain hamper the functional interaction with the MID1-α4-PP2A complex, leading to a subcellular location of GLI3.24 The observed missense variants in our study exceed the region to which Krauss et al have limited the MID-1 interaction domain. An alternative theory is suggested by Zhou et al who have shown that missense variants in the MBD can cause deficiency in the signalling of GLI3A, functionally implicating a relative overexpression of GLI3R.22 However, GLI3R overexpression would likely produce a posterior phenotype, as determined by Hill et al in their fixed homo and hemizygous GLI3R models.15 Therefore, our hypothesis is that all included missense variants have a similar pathogenesis which is more likely in concordance with the mechanism introduced by Krauss et al. To our knowledge, no splice site variants have been functionally described in literature. However, it is noted that the 15 and last exon encompasses the entire activator domain, thus any splice site mutation is by definition located on the 5′ side of the activator.

Based on the phenotype, we would suggest that these variants fail to produce a functional protein. We hypothesise that the truncating variants of the activator domain lead to overexpression of GLI3R in SHH rich areas. In normal development, the presence of SHH prevents the processing of full length GLI34 into GLI3R, thus producing the full length activator. In patients with a truncating variant of the activator domain of GLI3, thus these variants likely have the largest effect in SHH rich areas, such as the ZPA located at the posterior side of the hand/footplate.

Moreover, the lack of posterior anomalies in the GLI3∆699/- mouse model (hemizygous fixed repressor model) compared with the GLI3∆699/∆699 mouse model (homozygous fixed repressor model), suggesting a dosage effect of GLI3R to be responsible for posterior hand anomalies.15 These findings are supported by Lewandowski et al, who show that the majority of the target genes in GLI signalling are regulated by GLI3R rather than GLI3A.44 Together, these findings suggest a role for the location and type of variant in GLI3-mediated syndromes.Interestingly, the difference between Pallister-Hall syndrome and GLI3-mediated polydactyly syndromes has also been attributed to the GLI3R overexpression. However, the difference in phenotype observed in the cases with a truncating variant in the activator domain and Pallister-Hall syndrome suggest different functional consequences. When studying figure 1, it is noted that the included truncating variants on the 3′ side of the cleavage site seldomly affect the CBP binding region, which could provide an explanation for the observed differences. This binding region is included in the Pallister-Hall region as defined by Johnston et al and is necessary for the downstream signalling with GLI1.10 11 23 45 Interestingly, recent reports show that pathogenic variants in GLI1 can produce phenotypes concordant with Ellis von Krefeld syndrome, which includes overlapping features with Pallister-Hall syndrome.46 The four truncating variants observed in this study that do affect the CBP but did not result in a Pallister-Hall phenotype are conflicting with this theory.

Krauss et al postulate an alternative hypothesis, they state that the MID1-α4-PP2A complex, which is essential for GLI3A signalling, could also be the reason for overlapping features of Opitz syndrome, caused by variants in MID1, and Pallister-Hall syndrome. Further analysis is required to fully appreciate the functional differences between truncating mutations that cause Pallister-Hall syndrome and those that result in GLI3-mediated polydactyly syndromes.For the clinical evaluation of patients with GLI3-mediated polydactyly syndromes, intracranial anomalies are likely the most important to predict based on the variant. Unfortunately, the presence of corpus callosum agenesis was not routinely investigated or reported thus this feature could not be used as an indicator phenotype for LC membership. Interestingly when using only hand and foot phenotypes, we did notice a higher prevalence of corpus callosum agenesis in patients with posterior phenotypes.

The suggested relation between truncating mutations in the activator domain causing these posterior phenotypes and corpus callosum agenesis was statistically confirmed (OR. 8.8, p<0.001). Functionally this relation could be caused by the GLI3-MED12 interaction at the MBD. Pathogenic DNA variants in MED12 can cause Opitz-Kaveggia syndrome, a syndrome in which presentation includes corpus callosum agenesis, broad halluces and thumbs.47In conclusion, there are two distinct phenotypes within the GLI3-mediated polydactyly population.

Patients with more posteriorly and more anteriorly oriented hand anomalies. Furthermore, this difference is related to the observed variant in GLI3. We hypothesise that variants that cause haploinsufficiency produce anterior anomalies of the hand, whereas variants with abnormal truncation of the activator domain have more posterior anomalies. Furthermore, patients that have a variant that produces abnormal truncation of the activator domain, have a greater risk for corpus callosum agenesis.

Thus, we advocate to differentiate preaxial or postaxial oriented GLI3 phenotypes to explain the pathophysiology as well as to get a risk assessment for corpus callosum agenesis.Data availability statementData are available upon reasonable request.Ethics statementsPatient consent for publicationNot required.Ethics approvalThe research protocol was approved by the local ethics board of the Erasmus MC University Medical Center (MEC 2015-679)..