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U.S. National Science Foundation

The Sociology Program supports basic research on all forms of human social organization societies, institutions, groups and demography and processes of individual and institutional change. The program encourages theoretically focused empirical investigations aimed at improving the explanation of fundamental social processes. This includes research on organizations and organizational behavior, population dynamics, social movements, social groups, labor force participation, stratification and mobility, family, social networks, socialization, and the sociology of science and technology. The program supports both original data collection and secondary data analysis that use the full range of quantitative and qualitative methodological tools. Theoretically grounded projects that offer methodological innovations and improvements for data collection and analysis are also welcomed. Principal Investigators should selectPD 98-1331in the program announcement/solicitation block on the proposal cover sheet for submission of regular research projects to the sociology program. Projects are evaluated using the two foundation-wide criteria, intellectual merit and broader impacts. In assessing the intellectual merit of proposed research, four components are key to securing support from the Sociology Program: (1) the issues investigated must be theoretically grounded; (2) the research should be based on empirical observation or be subject to empirical validation or illustration; (3) the research design must be appropriate to the questions asked; and (4) the proposed research must advance our understanding of social processes, structures and methods. NSF also offers a number of specialized funding opportunities through its crosscutting and cross-directorate activities; some of the sociology-related opportunities are listed below. Crosscutting Research & Training Opportunities: ADVANCE: Increasing the Participation and Advancement of Women in Academic Science and Engineering Careers Faculty Early Career Development (CAREER) Program Graduate Research Fellowship Program (GRFP) Major Research Instrumentation (MRI) Program Mid-scale Research Infrastructure Programs SBE Postdoctoral Research Fellowships (SPRF) Research Experiences for Undergraduates (REU) Research at Undergraduate Institutions (RUI) Small Business Innovation Research (SBIR) Program To get information about these programs and others, please visit thecross-cutting and NSF-wide active funding opportunitiessearch page. NSF's mission calls for the broadening of opportunities for and expanding participation of groups, institutions and geographic regions that are underrepresented in STEM disciplines, which is essential to the health and vitality of science and engineering. NSF is committed to this principle of diversity and deems it central to the programs, projects and activities it considers and supports. NSF is also committed to public access to publications and data, unless there are countervailing interests that prohibit or limit public access to data, including matters of personally identifiable information of research participants, privacy or other issues of vulnerability such as economic, social or other security interests, etc.). SeePublic Access to Results of NSF-Funded ResearchandData Management for NSF SBE Directorate Proposals and Awards for more information.

NIGMS - National Institute of General Medical Sciences

Spatiotemporal Regulation of DNA Metabolism Pathways PROJECT SUMMARY/ABSTRACT Myriad DNA lesions occur continuously, and they necessitate distinct DNA damage repair mechanisms for removal. DNA double stranded breaks (DSBs) are potentially deleterious lesions that can trigger extensive loss of genetic information, chromosome fusions, and other gross chromosome rearrangements. DSBs are repaired mostly by homology-driven (Homologous Recombination: HR) or homology-independent (Non-Homologous End Joining, NHEJ) mechanisms. The choice of the repair pathway is dictated by the cell cycle phase, with HR being the more accurate (conservative) mechanism. Pathologies, such as meiotic defects and infertility, developmental syndromes, and cancer could stem from defects in HR. In addition to their involvement in DSB repair, many HR proteins fulfill key roles in the resolution of stalled replication forks or difficult-to-replicate DNA structures such as centromeres, telomeres, or DNA-RNA hybrids (R-loops). Importantly, HR proteins must be selectively activated to only perform repair functions at DNA lesions and differentially regulated to fulfill their crucial functions at other DNA structures such as stalled or collapsed replication forks. Timely activation/deactivation of HR proteins is thus a pre-requisite for maintaining genomic stability and avoidance of pathologies. There is a major gap of knowledge in understanding how the activity of HR proteins is dampened at DNA structures that resemble DNA lesions but activated at pathological structures, and how they fulfill unique roles at DNA breaks and replication forks. We postulate that phosphorylation and dephosphorylation of tyrosine residues, an under-studied subject as compared to serine/threonine modifications, contribute to the dynamic regulation of HR proteins. In our effort to fill this crucial knowledge gap, we have provided compelling evidence that EYA4, a dual activity protein phosphatase, acts on key HR and NHEJ factors to exert seminal impact on DNA repair efficiency and pathway choice. Over the past several years, we have devised biochemical procedures for the expression and purification of EYA4, and have identified RAD51 and 53BP1, central components of HR and NHEJ, respectively, as substrates of this poorly characterized protein phosphatase. We have made considerable progress in delineating the contributions of key phospho-residues in RAD51 to its role in HR, and in 53BP1 to NHEJ. We will now conduct mechanistic studies to understand the cellular regulation of EYA4, and whether its role in DNA repair pathway choice impacts the preservation of stressed and damaged replication forks and affects the maintenance of telomeres.

National Institutes of Health

Through this funding opportunity announcement (FOA), the National Cancer Institute (NCI) invites applications for P50 Research Center Grants for Specialized Programs of Research Excellence (SPORE). The program will fund P50 SPORE grants to support state-of-the-art investigator-initiated translational research that will contribute to improved prevention, early detection, diagnosis, and treatment of an organ-specific cancer or a highly related group of cancers. For the purpose of this FOA, a group of highly related cancers are those that are derived from the same organ system, such as gastrointestinal, neuroendocrine, head and neck, and other cancers. Other programmatically appropriate groups of cancers may include those centered around a common biological mechanism critical for promoting tumorigenesis and/or cancer progression in organ sites that belong to different organ systems. For example, a SPORE may focus on cancers caused by the same infectious agent or cancers promoted and sustained by dysregulation of a common signaling pathway. In addition, a SPORE may focus on cross-cutting themes such as pediatric cancers or cancer health disparities. The research supported through this program must be translational and must stem from research on human biology using cellular, molecular, structural, biochemical, and/or genetic experimental approaches. SPORE projects must have the goal of reaching a translational human endpoint within the project period of the grant.

National Institutes of Health

Through this funding opportunity announcement (FOA), the National Cancer Institute (NCI) invites applications for P50 Research Center Grants for Specialized Programs of Research Excellence (SPORE). The program will fund P50 SPORE grants to support state-of-the-art investigator-initiated translational research that will contribute to improved prevention, early detection, diagnosis, and treatment of an organ-specific cancer or a highly related group of cancers. For the purpose of this FOA, a group of highly related cancers are those that are derived from the same organ system, such as gastrointestinal, neuroendocrine, head and neck, and other cancers. Other programmatically appropriate groups of cancers may include those centered around a common biological mechanism critical for promoting tumorigenesis and/or cancer progression in organ sites that belong to different organ systems. For example, a SPORE may focus on cancers caused by the same infectious agent or cancers promoted and sustained by dysregulation of a common signaling pathway. In addition, a SPORE may focus on cross-cutting themes such as pediatric cancers or cancer health disparities. The research supported through this program must be translational and must stem from research on human biology using cellular, molecular, structural, biochemical, and/or genetic experimental approaches. SPORE projects must have the goal of reaching a translational human endpoint within the project period of the grant.

National Institutes of Health

Through this Notice of Funding Opportunity (NOFO), the National Cancer Institute (NCI) invites applications for P50 Research Center Grants for Specialized Programs of Research Excellence (SPORE). This is a re-issuance of PAR-23-284. The program will fund P50 SPORE grants to support state-of-the-art investigator-initiated translational research that will contribute to improved prevention, early detection, diagnosis, and treatment of an organ-specific cancer or a highly related group of cancers. For the purpose of this NOFO, a group of highly related cancers are those that are derived from the same organ system, such as gastrointestinal, neuroendocrine, head and neck, and other cancers. Other programmatically appropriate groups of cancers may include those centered around a common biological mechanism critical for promoting tumorigenesis and/or cancer progression in organ sites that belong to different organ systems. For example, a SPORE may focus on cancers caused by the same infectious agent or cancers promoted and sustained by dysregulation of a common signaling pathway. In addition, a SPORE may focus on cross-cutting themes such as pediatric cancers or epigenetics. The research supported through this program must be translational and must stem from research on human biology using cellular, molecular, structural, biochemical, and/or genetic experimental approaches. SPORE projects must have the goal of reaching a translational human endpoint within the project period of the grant.

National Institutes of Health

<p>Through this Notice of Funding Opportunity (NOFO), the National Cancer Institute (NCI) invites applications for P50 Research Center Grants for Specialized Programs of Research Excellence (SPORE). This is a re-issuance of <a href="https://grants.nih.gov/grants/guide/pa-files/PAR-23-284.html">PAR-23-284</a>. The program will fund P50 SPORE grants to support state-of-the-art investigator-initiated translational research that will contribute to improved prevention, early detection, diagnosis, and treatment of an organ-specific cancer or a highly related group of cancers. For the purpose of this NOFO, a group of highly related cancers are those that are derived from the same organ system, such as gastrointestinal, neuroendocrine, head and neck, and other cancers. Other programmatically appropriate groups of cancers may include those centered around a common biological mechanism critical for promoting tumorigenesis and/or cancer progression in organ sites that belong to different organ systems. For example, a SPORE may focus on cancers caused by the same infectious agent or cancers promoted and sustained by dysregulation of a common signaling pathway. In addition, a SPORE may focus on cross-cutting themes such as pediatric cancers or epigenetics. The research supported through this program must be translational and must stem from research on human biology using cellular, molecular, structural, biochemical, and/or genetic experimental approaches. SPORE projects must have the goal of reaching a translational human endpoint within the project period of the grant.</p>

NIAID - National Institute of Allergy and Infectious Diseases

ABSTRACT Type 1 diabetes is an autoimmune disease where the immune system mistakenly attacks and destroys insulin-producing beta cells in the pancreas. Our research has discovered unique hybrid molecules, called Hybrid Insulin Peptides (HIPs), that form in beta cells when fragments of insulin fuse with other protein fragments. Various HIPs contributing to disease in humans and mice are generated by an enzyme called Cathepsin D and serve as key targets for the immune system’s attack on beta cells. Interestingly, while HIPs are consistently detectable in laboratory mice used to study diabetes, they are harder to detect in human tissue samples even when analyzing larger amounts. We discovered this difference stems from how HIPs are made: the human version of Cathepsin D requires more acidic conditions to function compared to the mouse version. This could explain why human and mouse diabetes look different under the microscope - mice show widespread inflammation throughout the pancreas, while humans show more localized damage. To better understand how HIPs form in human disease, we propose to create a new mouse model where we replace the mouse version of Cathepsin D with the human version. We expect these “humanized” mice will form HIPs less readily, similar to humans. This model will help us understand how environmental factors influence HIP formation and disease development, potentially identifying new ways to prevent or treat type 1 diabetes. This research could reveal important insights into why type 1 diabetes develops and how environmental factors might influence disease progression through their effects on HIP formation, potentially leading to more effective prevention strategies.

OD - NIH Office of the Director

Vesicating (blister-forming) chemical-threat agents such as sulfur mustard (SM) or mustard gas, nitrogen mustard (NM), lewisite, and phosgene oxime can cause moderate to severe injuries and pain to the skin, eyes, and lungs. SM and NM are highly reactive bifunctional alkylating agents that can covalently modify all major cellular biomolecules, such as DNA, proteins, and lipids; thus, they are highly toxic. The eyes are particularly vulnerable to vesicant injuries, which cause a biphasic pathology of an acute response of photophobia, corneal erosions and inflammation, and chronic or late effects with significant deterioration of corneal structure and function from neovascularization, epithelial defects, fibrosis, and opacity. No therapeutic drugs are available as Medical Countermeasures (MCMs) for vesicant damage to the eye, eyelid or other organs. The major obstacle in developing potential MCMs is our limited understanding of the complex pathophysiological response of the eye after vesicant exposure. In this application, we propose to test the hypothesis that vesicating ocular injury pathology involves bioactive sphingolipid (SPL) pathways for acute and chronic inflammation and subsequent cornea, conjunctiva, and eyelid damage, causing significant vision impairment and dry-eye symptoms. In preliminary studies, we developed and characterized an NM-induced ocular surface injury (NMOSI) in mice, exposing the entire ocular surface to NM instead of only the cornea. We observed a severe acute inflammatory response that resolves in a month and cause damage to the cornea, atrophied eyelid glands, almost complete loss of vision, and apparent dry-eye symptoms. We found increased activity of acid sphingomyelinase, concurrent reduction in the sphingomyelin, and increased ceramides, suggesting sphingomyelinase activation in ocular surface tissue at three days post-exposure. Here, we propose to characterize NMOSI models in mice and rabbits, focusing on conjunctival goblet cells and epithelial stem cells and how NM affects the eyelids and their glands and causes dry-eye symptoms (SA #1). We will determine the temporal and spatial relationship of NM to SPL pathway for acute toxicity in ocular surface tissue of mice and rabbits separately from the cornea, conjunctiva-sclera, and eyelids at different time points (SA #2). It is unknown whether NM or SM-induced SPL pathways are overlapping. Hence, we propose to study if the NM- induced SPL pathway activation is similar to SM exposure (SA #3). Lastly, we plan to map out the pathway of SPL activation and lipid signaling using in vitro assays with meibomian gland epithelial and corneal cell lines (SA #4). We expect to identify novel associations of bioactive lipids in the inflammatory and wound-healing pathways of vesicating ocular injury, which will aid in improving our understanding of pathophysiological mechanisms of the injury and aid in developing potential MCMs in the future.

NCI - National Cancer Institute

Despite tremendous advances in cancer detection, diagnosis and treatment, many gaps remain, and cancer continues to be a chronic disease and the second leading cause of death in the US. To further advance the health and longevity of all Americans, it is of paramount importance to train the next generation of biomedical scientists by providing the high-quality research experiences and mentorship. Studies suggest that undergraduate research experiences can support trainee career development to pursue research careers.  The goal of this new R25 program, Stanford’s Cancer Research Education Program (SCREP) is to provide necessary research experiences to undergraduate trainees enabling them to persist in STEM fields by cultivating a scientific identity by building their confidence in their ability to succeed in cancer research setting.  SCREP will provide undergraduate trainees with the opportunity to actively engage in cancer relevant research within the stellar scientific and educational environment at Stanford University and the mentors from the Stanford Cancer Institute (SCI). The SCREP is a fully funded 10-week summer cancer research program, during which undergraduate trainees will work in cancer laboratories of the SCI and receive training in a wide range of cancer research concepts and techniques combined with curated programming that includes career and research seminars, skill building workshops and social events to complement the research components of the program. Trainees will be provided with a multi-layered supportive ecosystem where Mentors, Program Administrator and a Peer Mentor will provide individualized support. Another layer of support will be provided through leveraging existing Stanford University Resources and career enhancement partnership programs to ensure trainees continue their profession growth. This supportive ecosystem that SCREP provides is intended to help undergraduates trainees cultivate a scientific identity by building their confidence in their ability to succeed in cancer research setting. Together, SCREP will create research and career development opportunities for all undergraduates to explore, experience and pursue cancer research and careers in medicine. SCREP’s impact and effectiveness will be measured longitudinally by tracking trainees’ persistence in STEM, level of confidence in their abilities, and educational and career trajectories post program. The program ultimately supports the National Cancer Plan to develop future biomedical scientists in cancer research and clinical care workforce.

National Park Service

In cooperation with the University of Georgia, this project supports the development of statistically sound long-term natural resource monitoring, including sampling designs and data analysis approaches, to ensure that recommendations stemming from the monitoring are based on statistically sound analyses that support management needs.

Kids and Paper

Kids and Paper has served children ages 5-12 with afterschool and summer program through homework help, SEL development through creative arts, healthy meals, and recreational activities at Magnuson Community Center, located at 7110 62nd Ave NE Seattle, WA. We are now looking to expand our educational impact by adding a new element to our program, specific STEM focused activities and tutoring that expose children to topics and hands-on learning. This will help students connect what they are learning with real-world applications. We will offer the new STEM program through the summer of 2023 and anticipate closing this project on September 30, 2023.

Xbot Robotics

Xbot Robotics will organize a series of free STEM robotics workshops for youth. They will be held from fall 2017 to spring 2018 at the Beacon Hill Library located at 2821 Beacon Avenue S. Recruitment for the workshops will be targeted towards youth who may lack access to STEM activities and robotic kits at home or in their communities.

NIDDK - National Institute of Diabetes and Digestive and Kidney Diseases

PROJECT SUMMARY This investigator’s proposal describes a 5-year project of vertebral body derived mesenchymal stem cells within a Thread Reinforced Encapsulation Device (THRED) to treat the acute and long-term effects of mesenteric ischemia. The proposal evaluates intestinal organ recovery after treatment with cellular therapy and evaluates the metabolic and cellular host responses to massive surgical small bowel resection. The proposal provides basic and translational applications of emerging technologies relevant to the surgical treatment of intestinal ischemia. Investigators hypothesize that H2S is a critical component of VB-MSC mediated intestinal protection during mesenteric ischemia treatment, and that encapsulated MSCs can provide a stable source of H2S via an implantable, retrievable delivery system. They have developed a novel mouse with a mutation to test their hypothesis that VB-MSCs release hydrogen sulfide that then reacts at Cysteine440 on eNOS to bring about improved mesenteric blood flow. Through additional models, they investigate how VB-MSCs can be safely delivered within an implantable, retrievable, nanoporous device that allows cells to release their beneficial paracrine mediators while protecting them from host immune deletion. They propose three Specific Aims: 1) Develop a novel cellular delivery system to effectively deploy hydrogen sulfide from VB-MSCs in mesenteric ischemia, 2) Evaluate the interaction of THRED packaged VB-MSCs, H2S, and Nitric Oxide (NO) on chronic mesenteric vasodilation and long-term intestinal adaptation, and 3) Deploy THRED packaged VB-MSCs in a porcine model of mesenteric ischemia as final preclinical testing of scalability and effectiveness. The investigator is a pediatric surgeon scientist who completed his K08 funding through the NIDDK and was previously awarded an Early Stage Investigator R01 through the NIDDK. His career goals are to use this R01 to develop novel therapies and diagnostic tools for intestinal ischemia. He has collaborated with Dr. Minglin Ma at Cornell University who has extensive experience with implantable devices. Dr. Ma’s group has invented the THRED device, which is an electrospun nanoporous membrane that allows stem cells to interact with their local environment, while also containing them in a specific anatomical location and protecting them from immune destruction. Dr. Markel has also collaborated with Dr Tim Lescun, a large animal veterinarian at Purdue University which is approximately 45 minutes away from Dr. Markel’s institution. Additional collaborators include Dr. Erik Woods from Ossium Health, who will supply the VB-MSCs. In summary, this research aims to understand the mechanism that VB-MSCs use to provide acute and chronic protection in mesenteric ischemia. It also looks to identify appropriate delivery strategies so that cells can be delivered in an implantable, retrievable device for therapeutic use. The proposal is highly innovative and the investigator has the appropriate support, collaborations, and infrastructure in place to carry out the study.

Friends of STEM Technology

Youth participants will attend a Science Technology Engineering and Math summer camp where they will have two opportunities to enhance their video skills. They will learn the technical design process all programmers use to create video games. They will also assist in creating, writing and producing a basic video that will be viewed in a public venue upon completion.

DEPT OF DEFENSE.DEPT OF THE NAVY.NAVSUP.NAVSUP WEAPON SYSTEMS SUPPORT.NAVSUP WSS MECHANICSBURG.NAVSUP WEAPON SYSTEMS SUPPORT MECH

STEM,FLUID VALVE

NIAID - National Institute of Allergy and Infectious Diseases

Abstract Human immunodeficiency virus (HIV) targets the immune cells and weakens our defense against many infections and cancer. Without treatment, HIV infection advances to acquired immunodeficiency syndrome (AIDS). Although combination antiretroviral therapy has dramatically improved clinical outcomes, the rapid development of drug-resistant HIV strains limits the selection of effective therapies available for patients. Thus, there is an urgent need to identify alternative viral targets for inhibition. An essential yet poorly understood step in HIV replication is the Rev-response element (RRE)-mediated nuclear export of viral RNAs. During HIV infection, partially spliced and unspliced viral RNAs need to be exported from the nucleus to the cytoplasm for viral protein synthesis and virion assembly. This process depends on a specific interaction between the viral protein Rev and RRE present in the incompletely spliced viral RNAs. Multimeric Rev proteins bind to the RRE structure and recruit the nuclear export complex for cytoplasmic translocation of the viral RNAs. Despite the essential function in HIV replication, the RRE-Rev complex is currently an unexploited target in HIV chemotherapy, largely due to the lack of the structural information on the full-length RRE. Using a tRNA-scaffold approach, we previously determined the crystal structure of RRE stem-loop II, the initial Rev protein binding site. In this proposal, we will (1) determine the structure of the full-length RRE, (2) test the sequential and cooperative mechanism of Rev assembly, and (3) develop antisense oligonucleotide and de novo protein inhibitors that disrupt RRE-Rev interaction. Since RRE-Rev interaction is absolutely required for viral RNA export and virion assembly, targeting this complex has strong potential to suppress HIV replication and reduce viral burden in HIV- 1 infected individuals.

NIAID - National Institute of Allergy and Infectious Diseases

Abstract The development of effective vaccines against complex pathogens like HIV-1 requires immunogens that elicit broadly neutralizing antibodies (bnAbs). Traditional vaccine development strategies, including live- attenuated or subunit vaccines, and newer approaches leveraging genomic data, have not consistently induced bnAbs against HIV-1. Challenges stem from the unique structural and functional properties of bnAbs, such as improbable somatic mutations, autoreactivity, and long heavy-chain complementarity determining region-3 (HCDR3) loops. While priming immunogens have shown promise in selecting bnAb precursors, the production of mature bnAbs remains elusive, even in models engineered to ensure the presence of the relevant B-cell receptors. Our preliminary findings highlight the critical role of the transition state between unbound and bound antibody-antigen interactions in determining HIV-1 neutralizing antibody affinities. These transient structural states, which influence affinity maturation, neutralization breadth, and viral evolution, are poorly understood. Current methodologies primarily focus on static structural determinants of binding, neglecting the dynamic encounter complexes and intermediates that govern antibody-antigen association. This proposal aims to elucidate the association transition states and related intermediates for V2 apex- and CD4- binding site-directed bnAbs. Using structural and kinetic analyses, we will determine how variations in HIV-1 Env influence these transition states and how bnAbs overcome association barriers posed by Env sequence diversity. By dissecting the residue-level processes involved in antibody-antigen association, we will identify critical factors driving bnAb maturation and viral escape mechanisms. Using this information and large affinity datasets based on antibody clone HIV-1 Envelope interactions, we will develop cutting edge artificial intelligence/machine learning models to design immunogens with favorable affinity gradients for maturing bnAbs. Our work will address significant gaps in the understanding of antibody-antigen association, transitioning from phenomenological models to precise structural definitions of these poorly understood states and will directly use this information to inform immunogen development.

NIAID - National Institute of Allergy and Infectious Diseases

Hepadnaviruses are partially double-stranded DNA viruses that replicate by protein-primed reverse transcription. This family includes duck hepatitis B virus (DHBV) with which hepadnaviral reverse transcription was discovered and human hepatitis B virus (HBV) that kills 1,100,000 people annually. HBV therapy primarily employs nucleos(t)ide analog drugs that target the viral reverse transcriptase (RT) activity. Reverse transcription is catalyzed by the 4-domain viral polymerase (P) which has protein priming, RT, and ribonuclease H (RNase H) activities. The TP and spacer domains are unique to the hepadnaviruses. Reverse transcription starts with chaperone-mediated binding of P to the ε stem loop on the viral pregenomic RNA (pgRNA). Reverse transcription is primed by a tyrosine in P’s terminal protein domain (TP), templated by ε. The RT synthesizes the first strand of the viral DNA, and the RNase H destroys the pgRNA to permit synthesis of the second DNA strand. P is a monomer, and the covalent linkage between P and the DNA persists throughout reverse transcription. Hepadnaviral protein-primed reverse transcription differs greatly from retroviral reverse transcription, but its enzymology is poorly understood even though HBV P is a major drug target. This knowledge gap is in part due to the inability to determine the structure of P. We recently predicted the structure of P and validated the model. This revealed a novel fold in which the TP domain that primes reverse transcription is cupped over P’s catalytic core of P, with the priming tyrosine on a loop over the RT active site. This model makes mechanistic predictions regarding reverse transcription and provides guidance for how to test the hypotheses. Premise: The molecular model of P enables in-depth mechanistic analyses of P structure, nucleic acid binding, and DNA priming by the enzyme for the first time. Aim 1. What P sequences are needed for ε binding and priming? We will define the minimal active form(s) of P for RNA binding and DNA priming, and identify residues of P that contact ε and are essential for priming. Aim 2. What are the structural alterations to P associated with the shift from the priming-incompetent to priming-competent state? We will define how the TP domain binds to the catalytic core of P, explore P’s conformational shifts during priming, and determine how key RNA binding motifs are exposed during ε binding. Aim 3. How do conformational dynamics of P contribute to ε binding and priming? We will probe how P’s flexibility affects ε binding and DNA priming using molecular dynamics plus pharmacological and mutational analyses. We will test effects of mutations affecting RNA binding and DNA priming on viral replication in cells. This study will fill major gaps in our understanding of hepadnaviral reverse transcriptase enzymology by defining the interactions holding P in its novel conformation, how P binds to ε, and how enzyme flexibility contributes to the early phases of reverse transcription. It will also provide key information needed to develop non-active site inhibitors of HBV P to improve therapy for HBV patients.

National Institutes of Health

The NIH Research Education Program (R25) supports research education activities in the mission areas of the NIH. The over-arching goal of this R25 program is to support educational activities that foster a better understanding of biomedical, behavioral and clinical research and its implications. To accomplish the stated over-arching goal, this FOA will support creative educational activities with a primary focus on research experiences for high school or undergraduate students or science teachers during the summer academic break. The proposed program needs to fit within the mission of the participating IC that the application is being submitted to and should not have a general STEM focus (see below and Table of IC-Specific Information and Points of Contact).

National Institutes of Health

The NIH Research Education Program (R25) supports research education activities in the mission areas of the NIH. The over-arching goal of this R25 program is to support educational activities that foster a better understanding of biomedical, behavioral and clinical research and its implications. To accomplish the stated over-arching goal, this FOA will support creative educational activities with a primary focus on research experiences for high school or undergraduate students or science teachers during the summer academic break. The proposed program needs to fit within the mission of the participating IC that the application is being submitted to and should not have a general STEM focus (see below and Table of IC-Specific Information and Points of Contact).

NHLBI - National Heart Lung and Blood Institute

ABSTRACT Cardiovascular disease remains a leading cause of morbidity and mortality, despite significant advances in surgical techniques and medical therapies. The development of novel diagnostic, treatment, and prevention strategies requires an in-depth understanding of cardiovascular pathophysiology, paired with advanced engineering and computational expertise. However, a critical gap exists in formal training opportunities for medical students to integrate surgical and engineering skills, contributing to a growing deficit of cardiovascular surgeon-scientists. To address this need, we propose the Surgery, Technology and Engineering Mentorship for Medical Students (STEMS) short-term training program at Washington University in St. Louis. This program is founded on the premise that early engagement of post-baccalaureate medical students will inspire their pursuit of academic careers in cardiovascular surgery while accelerating the development of innovative technologies for diagnosing, treating, and preventing cardiovascular diseases. Research topics will include critical areas such as cardiac arrhythmia, coronary artery disease, peripheral vascular disease (arterial/venous), aortic aneurysmal disease, and neurovascular disease. With a diverse and accomplished faculty mentorship network, robust institutional support, and established multi-disciplinary research programs, the STEMS training program will equip trainees with synergistic skills in translational research and engineering to address complex challenges in this field. Graduates of this program will emerge with foundational knowledge to pursue impactful future academic and research pursuits in the diagnosis, management, and prevention of cardiovascular diseases.

NEI - National Eye Institute

PROJECT SUMMARY Millions of elderly individuals suffer from visual impairment due to dry age-related macular degeneration (AMD), for which effective treatments are limited. In advanced stages of dry AMD, the gradual loss of retinal pigment epithelial (RPE) cells leads to the death of overlying photoreceptors, causing progressive vision loss and eventual blindness. Transplantation of healthy RPE to replace lost/diseased RPE cells have shown in rodent studies to rescue rod and cone photoreceptors, improve retinal electrophysiological responses, and enhance visual thresholds over extended periods. However, these studies have used xenogeneic (across species) cell transplants, which require immune suppression (IS) to prevent the host’s immune system from rejecting the transplanted cells. Similarly, allogeneic (same species) RPE cell transplants in non-immune suppressed models, including pigs and nonhuman primates, are typically rejected within three weeks. Although IS appears currently necessary and sufficient to protect transplanted cells, it raises significant safety concerns, especially for elderly AMD patients who may experience toxic side effects from long-term or indefinite IS use. Additionally, IS introduces challenges such as patient compliance and the risk of rejection with suboptimal dosages. Current Phase I/II clinical trials using allogeneic RPE cells combine multiple IS medications to prevent rejection, but the majority of adverse effects stem from the IS regimen rather than the cell therapy itself. To address the complications of IS, the NIH has initiated an autologous RPE cell trial, despite the high logistical and cost barriers to broad implementation. In contrast, our approach focuses on developing a scalable allogeneic cell-based therapy that can avoid immune rejection, providing greater access, efficiency, and lower costs. We have demonstrated feasibility of this approach in multiple settings including short-term RPE cell transplants in the eye in non-immune suppressed NHPs. In the proposed studies, we will generate multiple lines of allogeneic induced pluripotent stem cells (iPSCs) from nonhuman primates (NHPs) and engineer them to lack expression of class I and II major histocompatibility complexes and to overexpress the “don’t eat me” signal, CD47. We will then optimize the differentiation of these modified iPSCs into RPE cells for transplantation studies in both normal and diseased NHP retinas. Transplantation studies will include short and long-term survival and in diseased retinal conditions to replicate acute version chronic rejection in normal and diseased retinal environments. Finally, we will optimize the use of a safety switch to enable selective removal of cells in the subretinal space should that ever be necessary. These studies will demonstrate the potential of gene-modified RPE cells to evade immune rejection while maintaining a high safety profile and will help identify factors in retinal disease environments that may affect the survival of transplanted RPE cells. Successful completion of these aims will lay the groundwork for translating these studies toward clinical application.

NCI - National Cancer Institute

Abstract H-FIRE is an emerging, minimally invasive focal ablation technique that utilizes low energy, microsecond-long electric pulses delivered locally within targeted tissues for several minutes. Tumor cell death is rapidly induced through a loss of cellular homeostasis and is not dependent on thermal effects. Due to its non-thermal mechanism, H-FIRE has been shown to preserve important tissue components such as the extracellular matrix, blood vessels, and nerves, as well as the immunogenicity of tumor antigens compared to other ablation modalities. H-FIRE can ablate predictable volumes of tissue without the need for neuroparalytics as is required for its FDA-approved predecessor, and safe and effective H-FIRE treatment has been demonstrated in rat glioma models and canine patients with spontaneous brain tumors. Key advantages of H-FIRE for GBM treatment include i) glioma and glioma stem-like cell specific ablation resulting from tuned H-FIRE pulses; ii) enhancement of ablation through combination with a targeted molecular adjuvant; iii) reversible BBB breakdown and enhanced delivery of molecular therapeutics from the dissipating H-FIRE field well beyond the tumor margin; iv) electrical feedback for monitoring and modeling treatment; and v) activation of innate and adaptive anti-tumor immune responses from intracranial H-FIRE. These advantages motivate the central hypothesis for this effort, specifically that H-FIRE combined with the cancer adjuvant immunomodulatory drug, indoximod, will overcome key drivers of therapy resistance by combining effective ablation of a core tumor mass with enhancing delivery and efficacy of a small molecule drug that is normally blocked from entry by the BBB. The project has two aims. In Aim 1, pulse parameters will be evaluated on their ability to completely and selectively ablate malignant tissue in comparison to normal brain tissue, to corroborate preliminary in vitro data using a more relevant in vivo glioma rat model. In Aim 2, the in vivo synergy resulting from the combination of H-FIRE with indoximod will be quantified using an orthotopic rat GBM model. Computational models and in vitro experimentation are highly adaptable, allowing real-time data collection and analysis. However, fully replicating the intricate brain environment and the complete surgical procedure involving the skull can only be accurately assessed using an in vivo animal model. In selecting the species and strain, female and male Fischer rats were chosen because no lower species model is available that uniquely support the electrode configuration for in vivo brain electroporation studies. A mammal is needed for adequate analysis of tissue ablation. Additionally, species matching will allow us to make comparisons to previously completed studies. These aims will lead to independently optimized H-FIRE protocols for i) ablation of a tumor bulk resulting in effective anti-tumor immune stimulation and ii) reduction of barriers to the efficacy of indoximod adjuvant therapy. Intraoperative MRI-guided electrode insertion will allow for highprecision electrosurgery and fine-tuned BBB disruption and brain cancer treatment efficacy in preparation for treating canine patients, with the expectation to translate those results to human patients

NIBIB - National Institute of Biomedical Imaging and Bioengineering

Synthetic Genetic Controller Circuits for Transcription Factor-Directed Differentiation PI: Domitilla Del Vecchio1,2,3 co-I: James J. Collins2,3,4,5 co-I: Thorsten Schlaeger6 1Department of Mechanical Engineering, MIT; 2Department of Biological Engineering, MIT 3Synthetic Biology Center, MIT; 4Broad Institute of MIT & Harvard; 5The Wyss Institute 6 Stem Cell Transplantation Program, Boston Children’s Hospital PROJECT SUMMARY The ultimate goal of this project is to create synthetic genetic circuits that accurately control the level of cell fate- specific transcription factors (TFs) autonomously in response to cell state changes. The underlying hypothesis is that the level and timing of expression of critical TFs dictates the efficiency of cell conversion protocols and the quality of produced cells. Here, we focus on the differentiation of human induced pluripotent stem cells (hiPSCs) into hemogenic endothelial cells (HECs) from which all hematopoietic stem and progenitor cells (HSC/HPCs) arise. Current methods to derive definite HECs (dHECs), which have the potential to produce adult-type lymphoid cells and HSCs, remain largely inefficient and are also difficult to execute and scale, and, as a consequence, exhibit high degrees of variability in out- comes between different labs, hiPSC lines, and even between replicate experiments.These problems hamper analysis of the underlying developmental processes and pose formidable obstacles to clinical translation of hiPSC-derived blood cell products since ensuring the safety and cost-effectiveness of the product necessitates high differentiation efficiency and consistency. Prior work has demonstrated that SCL (S), LMO2 (L), GATA2 (G), and ETV2 (E) TFs, when expressed in mesodermal cells, activate dHEC gene regulatory networks (GRNs) across species but also that efficient forward programming to dHECs requires discovery and subsequent implementation of both optimal expression levels and tim- ing for the TFs. Yet, conventional methods for TF-mediated cell fate programming generally rely on indiscriminate overexpression with little control on cellular TF levels and without cell state sensing. This is largely due to our inability to precisely control TF profiles during cell fate programming, and this limitation has prevented discovering optimal tra- jectories and subsequently enforcing them. Here, we propose synthetic genetic controller circuits that overcome this hurdle. In Aim 1, we create genetic circuit designs that set TF levels and use them in an efficient in vitro differentiation protocol to discover the optimal combination of S, L, G, E levels and timing. In Aim 2, we develop a circuit architecture, based on a novel TET1-enabled positive feedback system, to prevent epigenetic silencing of genetic circuits once de- livered to hiPSCs. In Aim 3, we make our genetic controller circuits enforce autonomously the optimal SLGE TF levels found in Aim 1 in response to the hiPSC-to-mesoderm transition. We achieve this by a new autocatalytic ADAR-based RNA sense-and-respond system, which senses the mesoderm marker Brachyury (TBXT) and enforces user-defined TF levels in response to it. We anticipate that this process, by being autonomous as opposed to manual and by enforcing optimal TF trajectories, will result in a more efficient, repeatable, and robust hiPSCs to dHECs conversion protocol, thereby helping fill the gap to clinical translation. Although in this project we tailor the genetic circuit designs to controlling SLGE TFs after sensing mesoderm-specific transcripts, the designs can be readily modified to express different TFs in response to any other cell type- or state-specific transcript. Therefore, we believe that the synthetic biology technology that we will establish will have broad impact on any other cell fate programming as well as on cell-or gene-therapy projects where expression levels and timing, as well as resistance to silencing, are important.