Lipid nanoparticle library towards development next generation genomic medicines

Ionizable amino lipids are a major constituent of the lipid nanoparticles for delivering nucleic acid therapeutics (e.g., DLin-MC3-DMA in ONPATTRO , ALC-0315 in Comirnaty , SM-102 in Spikevax ). Scarcity of lipids that are suitable for cell therapy, vaccination, and gene therapies continue to be a problem in advancing many potential diagnostic/therapeutic/vaccine candidates to the clinic. Herein, we describe the development of novel ionizable lipids to be used as functional excipients for designing vehicles for nucleic acid therapeutics/vaccines in vivo or ex vivo use in cell therapy applications. We first studied the transfection efficiency (TE) of LNP-based mRNA formulations of these ionizable lipid candidates in primary human T cells and established a workflow for engineering of primary immune T cells. We then adapted this workflow towards bioengineering of CAR constructs to T cells towards non-viral CAR T therapy. Lipids were also tested in rodents for vaccine applications using self-amplifying RNA (saRNA) encoding various antigens. We have then evaluated various ionizable lipid candidates and their biodistribution along with the mRNA/DNA translation exploration using various LNP compositions. Further, using ionizable lipids from the library, we have shown gene editing of various targets in rodents. We believe that these studies will pave the path to the advancement in nucleic acid based therapeutics and vaccines, or cell gene therapy agents for early diagnosis and detection of cancer, and for targeted genomic medicines towards cancer treatment and diagnosis.

Conducting clinical genomics research during the COVID-19 pandemic: Lessons learned from the CSER consortium experience.

Clinical research studies have navigated many changes throughout the COVID-19 pandemic. We sought to describe the pandemic's impact on research operations in the context of a clinical genomics research consortium that aimed to enroll a majority of participants from underrepresented populations. We interviewed (July to November 2020) and surveyed (May to August 2021) representatives of six projects in the Clinical Sequencing Evidence-Generating Research (CSER) consortium, which studies the implementation of genome sequencing in the clinical care of patients from populations that are underrepresented in genomics research or are medically underserved. Questions focused on COVID's impact on participant recruitment, enrollment, and engagement, and the transition to teleresearch. Responses were combined and thematically analyzed. Projects described factors at the project, institutional, and community levels that affected their experiences. Project factors included the project's progress at the pandemic's onset, the urgency of in-person clinical care for the disease being studied, and the degree to which teleresearch procedures were already incorporated. Institutional and community factors included institutional guidance for research and clinical care and the burden of COVID on the local community. Overall, being responsive to community experiences and values was essential to how CSER navigated evolving challenges during the COVID-19 pandemic.

Precision public health approaches to health equity

Since President Barack Obama announced the Precision Medicine Initiative during his state of the union address in January 2015, the sciences of precision medicine and health equity have largely grown in parallel, though there have been some efforts to bring the two together. As research on health equity has evolved to name and consider structural racism, the penultimate goal of research in this area also as moved from efforts to identify and describe gaps between racial and ethnic groups to characterizing the context creates and perpetuates racial inequities and how best to mitigate them. In this presentation, I will briefly describe how syndemics, intersectionality, and individual tailoring may complement downstream efforts to characterize epigenetic and genomic efforts to develop biomedical interventions to achieve healthcare equity and health equity. After noting how the principles of precision medicine may be applied more broadly than the most common way it is operationalized through genomic medicine, I argue that that creating racial justice in health will require defining health equity more clearly and precisely. Consequently, I utilize the example of Black men and the context of COVID-19 to highlight how more precisely defining the structural context of the population of interest by using tools such as intersectionality and syndemics is fundamental to achieving equity. I highlight how achieving health equity will require creating, resisting, undoing, and mitigating structural racism and note what that means for cancer research. Precision medicine may help to mitigate the health effects of structural racism, and it will remain an important tool to promote population health;however, efforts to achieve health equity and racial justice will require interventions that change the contexts and conditions that create, exacerbate, and perpetuate structural inequities and the racial inequities in health outcomes that they produce and maintain.

eP520: Screening results in 186 any-health-status adults in primary care clinics receiving clinical NGS for 431 health risk and recessive genes

Introduction: The ACMG has recommended returning clinically relevant results for certain genes when identified in research or as secondary findings in diagnostic testing. Research studies have shown that genomic population screening detects patients with previously unrecognized and often actionable health risks or genetic conditions, with acceptably low levels of harm. Cascade testing of relatives at risk is enabled. Screening for recessive disorder carrier status with gene sequencing panels is common in clinical practice. Preventative screenings routinely occur in primary care settings. The cost of reliably sequencing of many genes in a clinically reliable fashion is approaching levels where offering genomic screening tests may be contemplated for entire populations, and the results used for preventative health purposes, including clinical correlation, early screening, and education. In anticipation of universal genome sequence-based screening, integrated with existing health risk screenings, we piloted a novel implementation of clinical genomic population screening in primary care, mostly family medicine clinics. Screening involved clinical sequencing and reporting of 431 genes where variants are associated with personal health risks or recessive disease carrier status. Methods: Interested primary care providers (PCPs) in two Family Medicine practice systems were invited to participate and given onboarding education. Adult patients with any health status were introduced to The Genomic DNA Test and provided test information by their PCPs in the context of preventative health assessment. Patient education materials included paper, online, and video information, a ‘hotline,’ and optional free genetic counseling. Patients completing a bespoke, health system-approved, written clinical consent provided blood or occasionally saliva samples that were NGS sequenced according to validated procedures in a commercial CLIA-certified genetic testing laboratory. Laboratory reports were returned to the PCP and patient after a local genetics professional added a 1-to-3-page messaging document, the Genomic Medicine Action Plan (GMAP). The PDF-format reports and GMAP were placed in the patient’s electronic health record. Only pathogenic (P) and likely pathogenic (LP) variants were reported. Variant classification was according to Sherloc, the performing laboratory’s system. Patients or providers could request free post-test genetic counseling locally, and the performing lab offered free family member testing and limited-cost partner testing for health risk panel genes and recessive disorder panel genes, respectively. Patients with health risk results were defined as being heterozygous for a P/LP variant for a dominant condition or for a recessive condition where some heterozygotes are symptomatic or co-dominant, hemizygous for a P/LP variant for an X-linked recessive condition, or bi-allelic and plausibly in trans for an autosomal (or X-linked in a female) recessive condition. Many such conditions that are common have reduced or low penetrance, and were characterized as increased risk compared to those not having those variants. When increased risk was identified, the GMAP recommended appropriate medical responses and/or patient education. As part of quality assessment of the pilot, the frequencies of reported results and certain events are monitored. Results: Between November 2019 and October 2021, 186 patients with a median age of 58 years were tested by 20 PCPs at no cost to patients or insurance. Testing volumes declined during the COVID-19 pandemic and when other health system events made high demands on PCPs and their staff. Only 13.3% of patients had no reportable variants in any of the 431 genes. Eighty point nine percent were carriers for at least one recessive disease. The most common recessive genes showing carrier status were HFE, SERPINA1, GALT, CFTR, BTD, F5, DHCR7, PC, GAA, GJB2, PMM2, PAH, and PKHD1. Twenty-six percent had at least one potential health risk result identified, 20% if the common thrombophilias are excluded. The most common category was hereditary cancer risk (7.5%), followed by F5, F2, and SERPINC1 thrombophilia variants (6.5%), hereditary hemochromatosis 1 (HFE) (4.3%), cardiovascular disorders, mostly cardiomyopathies (3.8%), alpha-1-antitrypsin deficiency or other pulmonary disorder (3.8%), familial Mediterranean fever heterozygotes (1.6%), G6PD deficiency (1.1%), and lipid disorder (0.5%). Two patients had health risks in two areas, and two in three areas. Interestingly, BRCA1 and BRCA2 variants were only identified in males. Thirteen patients, about 7%, had an amended report issued during the period. This happened when an unreported variant of uncertain significance was reclassified as LP or P, or when LP became P, and the performing laboratory issued an amended report. Surprisingly few patients took advantage of the free genetic counseling. No patient adverse events were reported by the participating PCPs despite ongoing outreach, nor by patients. Conclusion: Genomic population health screening can be successfully implemented in primary care settings with use of limited but essential genetic professional assistance, after careful planning and input from other medical specialties. A significant proportion of adults not selected for health status harbors germline genetic variants associated with increased health risk. In the absence of a culture where routine genomic screening is expected and where patient genomic competency is high, PCP capacity limits are a barrier to universality. Inclusion of genes for both health risk results with variable degrees of penetrance and for recessive carrier status, and multiple simultaneous results, dictates careful messaging of the implications, while doing so in a primary care setting begs a concise and efficient process. Rates of carrier detection were in-line with predictions based on general population frequencies. Rates of health risk detections were higher than earlier research programs because a larger number of genes with a much broader scope of health risk was included, including disorders with low penetrance yet meaningful clinical relevance and carefully-designed care pathways meant to optimize care while avoiding unnecessary additional testing. We conclude that genomic population health screening of primary care patients where large numbers of genes are clinically sequenced is feasible in a real-world health system, and that value exists for some tested patients now. Research to overcome certain technical limitations of current clinical genomic testing methods and to better stratify risk level in the context of incomplete penetrance should enhance the value of universally-offered genomic population health screening in the future.

eP236: TeleKidSeq: Incorporating telehealth into clinical care of children from diverse backgrounds undergoing clinical genome sequencing

Introduction: The global pandemic required healthcare institutions and clinical research programs to adapt quickly to non-traditional care models. TeleKidSeq is a pilot study that emerged from the NYCKidSeq program, an NIH-funded Clinical Sequencing Evidence-Generating (CSER) Consortium site focused on incorporating genomic medicine into the care of diverse New York City children with suspected genetic disorders. Embracing the opportunity to study the use of telehealth in delivering genomic results, TeleKidSeq will examine the impact of innovative remote genetic counseling modalities in medically underserved populations. Studies focusing on the use of telehealth performed before the COVID-19 pandemic have shown that patients prefer in-person visits to virtual visits;however, with the increased familiarity and widespread use of virtual platforms, we anticipate an increase in the preference for telehealth visits. TeleKidSeq aims to fill the gaps in current knowledge on the impact of visual aids in telehealth in diverse urban patient populations. Methods: TeleKidSeq will recruit 496 pediatric participants (aged 0-21 years) with neurologic, immunologic, or cardiac conditions suspected to have an underlying genetic cause who receive care predominantly within two large health systems in the New York metropolitan area. The Mount Sinai Genomics Stakeholder Board, consisting of diverse stakeholders and key community advisors, provided guidance about our study design and materials. Participants will be English- or Spanish-speaking, and based on prior enrollment data from NYCKidSeq study, we expect more than 65% will be from populations underrepresented in medical research. Prior to enrollment, participants will be randomized to receive their genomic results from a genetic counselor via telehealth either with screen sharing (ScrS) or without screen sharing (NScrS). All participants will receive genome sequencing (GS) from a clinically validated laboratory. Additionally, we will use GUÍA, a web-based application designed to enhance the delivery of genomic test results, in both the ScrC and NScrS arms to facilitate delivery of individualized genomic results and clinical information in a personalized, highly visual, and narrative manner. Surveys administered at baseline, after results disclosure, and 6-months post-results disclosure will be used to evaluate study outcomes. The primary outcome of the TeleKidSeq study will be participants’ perceived understanding of their GS results with a comparison between the results disclosed via videoconferencing with ScrS and NScrS arms. Secondary outcomes will include: objective understanding of GS results;understanding of medical follow-up recommendations and the actionability of genome sequencing results;adherence to medical follow-up recommendations made based on genomic results;and satisfaction with and ease of use of the telehealth experience, compared across the two arms. Diagnostic yield, clinical utility and cost of GS will also be assessed. Results: Not applicable. Conclusion: Overall, the TeleKidSeq pilot study will contribute to innovations in communicating genomic test results to diverse populations through telehealth technology. In conjunction with NYCKidSeq, this work will inform best practices for the implementation of genomic medicine in diverse, English- and Spanish-speaking populations.

Clinical genomic population health testing for 431 genes implemented through family medicine practices: the Vermont experience

Integration of genomics into health practice depends on successful implementation in non-research settings. We describe a medical home-centered implementation at the intersection of genomic medicine and population health in the UVM Health Network. In this clinical implementation, the hospital laboratory orchestrates a collaboration involving primary care providers (PCPs), patient and family advisors, health system administrators, clinical genetics services, oncologists and cardiologists, Vermont’s accountable care organization, and a commercial CLIA genomic testing laboratory. Phenotypically unselected adult primary care patients are offered “The Genomic DNATest” at no cost as part of their regular care. Testing is introduced by primary care providers and their staff using a brief animated video and printed decision aids with graded detail. Question resolution and pre- and post-test genetic counseling is offered at no cost using telephone, video, or in-person visits, and is coordinated bya single phone and email contact point, the Genomic Medicine Resource Center. 431 genes are sequenced for germline health risk and recessive carrier variants;only pathogenic and likely-pathogenic variants are reported. New reports are issued when reported and unreported variants are later reclassified. Test reports are reviewed by a clinical geneticist and genetic counselor. Two brief "action plans" are developed with PCP and patient focus in a single messaging document. This is prepended to the lab reports before release to the PCP, who reviews and then conveys them to the patient. PCPs and their staff receive initial training on the test and process and are invited to participate in an online community with monthly video case discussions. Among the first 72 patients tested, 17% had a health risk identified. This included dominantly inherited disorders and bi-allelic or hemizygous variants for common recessive disorders. Care pathways created in advance using multi-disciplinary expertise were activated for those. Free testing for blood relatives was made available. 76% of tested patients had at least one heterozygous recessive disease variant identified, and low-cost partner testingwas made available. Frequency of positive test results was in line with population frequency predictions. Pre- and post-test genetic counseling uptakewas lower than expected. This raised the question of unmet informational needs. A 2-page anonymous process quality survey mailed twice to the first 61 tested patients had a 31% return rate. Key findings included (1) pre-test engagement methods and decision aids were helpful;(2) the testing decision was influenced equally by value for the individual’s health, for their family’s health, and for researchers;(3) emotions during the ∼4-week time to results were neutral or excited, with none experiencing anxious feelings, and none reported the wait time as too long;(4) 21% reported contacting the Genomic Medicine Resource Center;(5) 16% reported referral to a specialist due to their result;(6) about half reported sharing the results with family members, but none reported any family members getting tested;(7) none indicated they were dissatisfied with the testing and result process, and only one responded they would not recommend others get the test;and (8) all agreed or somewhat agreed that the PCPs officewas the right place to do this testing.While this implementation was designed with scalability and a low management profile in mind, several systems-level barriers were encountered that contributed to lower engagement efforts and slower expansion than planned. This included lack of institutional information technology resources to surmount paper-based systems for requisitions, sample-routing, and consent forms;dependency of the patient engagement process during PCP visits on rooming and nursing staff during times of staffing shortages;susceptibility to practice model disruptions and priorities caused by the Covid-19 pandemic;and PCP time distraction resulting from user interface and polic changes in our EHR during the pilot. These barriers are targets for study and continuous process improvement activities. In summary, an example of clinical genomic population health testing using a medical-home focus has been successfully implemented in a non-research setting, supported by multi-disciplinary collaboration. This implementation depends on minimal staff, avoids financial barriers to access and genetic counseling, and offers a short, defined, test turnaround time as compared to similar biobank-based research programs. Tested patients find the program satisfactory, and meaningful test results are at least as common as in existing population health risk screening archetypes.

Retrospective analysis of NGS data in pediatric genetic cohort: detection of rare variants that may impact influenza and COVID-19 infection

It is difficult to assign a precise frequency of infections that defines an increased susceptibility to infections reflecting an impaired immune response given the majority of patients with intact immune systems still contract multiple upper respiratory infections each year, usually of viral origin. In fact, the average child may experience up to six to eight upper respiratory infections each year. The frequency of these infections may be related to environmental exposures but also may be triggered by genetic susceptibility. As an example, respiratory disease complicates the management of several inherited metabolic diseases, either at presentation or as late-onset features. More recently, children of all ages have been shown to contract COVID-19;however, children with underlying medical conditions are at increased risk. COVID-19 has been known for almost a year now, with several studies identifying genetic risk factors are associated with severe COVID-19. In the context of a health system wide genomic medicine program “Genomic Answers for Kids” at Children’s Mercy, Kansas City, we performed a retrospective analysis of rare variants predicted to be deleterious at 12 known loci known to govern TLR3- and IRF7- dependant type I Interferon immunity of all patients/families (>2000) tested for suspected genetic disorders. We bioinformatically extracted all rare variants in those 12 genes linked to type I interferon pathway from our internal warehouse. From those, ~340 variants were further analyzed based on inheritance, minor allele frequency in population datasets, and in silico prediction. The vast majority of this subgroup of GA4 K patients were referred for a suspected neurological disorder with or without multiple congenital anomalies (~75%). Only 15% were referred for metabolic disorders. Of those, 50% have a known genetic diagnosis unrelated to Immune deficiency. Of the selected index cases, the medical records, and if available the outpatient records, were reviewed to document the occurrence of recurrent infection and/or COVID-19. Preliminary data showed 46 “extremely” rare variants were detected in 37 GA4 K patients;6/37 (16%) have ≥2 in 1–12 genes, one GA4 K patient has 4 “extremely” rare variants in IRF7, and 3/37 GA4 K patients are deceased (~1%). Moreover, a novel disease gene was uncovered in a previously undiagnosed family, of which we identified an additional two affected individuals from an international collaboration. Finally, in a family with apparently dominant transmission of tumid lupus we observe putative causal variant in gene UNC93B1 – linking chronic inflammatory disorder (with known type I interferon association) to mutations predisposing to COVID-19. Recurrent or persistent infection is usually a manifestation of primary immunodeficiency. While most children with recurrent infections have a normal immunity, it is important to remember a subset of patients have an unrecognized genetic susceptibility to infection. Further analysis and monitoring are on-going. As we are continuing to struggle with the COVID-19 pandemic, combined with flu season, understanding precisely who may be at higher risk of infection and complications is critical and could play an important role in ongoing efforts to in disease prevention and the development of better treatment protocols

ACE2 and FURIN variants are potential predictors of SARS-CoV-2 outcome: A time to implement precision medicine against COVID-19.

The severity of the new COVID-19 pandemic caused by the SARS-CoV-2 virus is strikingly variable in different global populations. SARS-CoV-2 uses ACE2 as a cell receptor, TMPRSS2 protease, and FURIN peptidase to invade human cells. Here, we investigated 1,378 whole-exome sequences of individuals from the Middle Eastern populations (Kuwait, Qatar, and Iran) to explore natural variations in the ACE2, TMPRSS2, and FURIN genes. We identified two activating variants (K26R and N720D) in the ACE2 gene that are more common in Europeans than in the Middle Eastern, East Asian, and African populations. We postulate that K26R can activate ACE2 and facilitate binding to S-protein RBD while N720D enhances TMPRSS2 cutting and, ultimately, viral entry. We also detected deleterious variants in FURIN that are frequent in the Middle Eastern but not in the European populations. This study highlights specific genetic variations in the ACE2 and FURIN genes that may explain SARS-CoV-2 clinical disparity. We showed structural evidence of the functionality of these activating variants that increase the SARS-CoV-2 aggressiveness. Finally, our data illustrate a significant correlation between ACE2 variants identified in people from Middle Eastern origins that can be further explored to explain the variation in COVID-19 infection and mortality rates globally.