Stanford University

NIH Research Projects · FY 2025 · 2002-05

PROJECT SUMMARY Numerous environmental and cellular factors can induce DNA lesions or create other types of challenges that can slow DNA replication, leading to replication stress. Failure to alleviate this stress and restart stalled replica- tion forks can cause genome instability, which can drive cancer initiation and progression and affect the cellular response to chemotherapy. Hence, deciphering the cellular response to replication stress––the long-term goal of this application––is imperative for understanding fundamental aspects of tumorigenesis. One crucial aspect of the replication stress response involves replication fork reversal, a process that leads to the formation of a four-way junction structure by the remodeling of both nascent and parental DNA strands. There are a family of ATP-dependent translocases that promote fork reversal in vitro and in cells, but why so many translocases are involved in this process is not understood. We showed that one of these translocases, HLTF, prevents a stress-resistant mode of replication by promoting fork reversal, and we elucidated the basic mechanism by which cells continue replication upon HLTF loss. The overall goal of the proposed project is to further elucidate the mechanisms by which HLTF mediates the replication stress response, characterizing its unique role in this process. HLTF is a multi-functional protein and recent studies using a panel of HLTF mutants that are each deficient for one of its activities have revealed unexpected roles for these activities in HLTF’s known functions, raising a number of new and exciting questions. This application will address questions about the molecular mechanisms by which HLTF promotes fork slowing and fork reversal in cells and determine the impact of its loss on replisome stability, genome stability, and cell proliferation. Aim 1 will investigate which domains of HLTF are needed for fork slowing, fork reversal, and resistance to replication stress, and HLTF’s association with the replication fork. HLTF’s dynamic interaction with the fork will also be probed. Aim 2 will investigate a newly discovered function for HLTF in stabilizing the replication fork. These experiments will employ a combi- nation of molecular, cellular, genetic and proteomic approaches as well as single-molecule imaging and track- ing methods in Xenopus extracts to solve fundamental questions about how cells respond to replication stress.

NIH Research Projects · FY 2025 · 2002-04

ABSTRACT: Immunoregulatory cytokines engage transmembrane signaling receptors in order to mediate a wide range of functions including leukocyte proliferation, differentiation, and expansion through JAK/STAT activation. Most immunoregulatory cytokines possess both redundant and distinct activities that are critical to normal immune homeostasis, but this functional pleiotropy presents a major problem for the effective use of these cytokines as immunotherapeutic cancer drugs. Cytokine pleiotropy is a consequence of different cytokine receptors being expressed on multiple different counterbalancing cell types that serves to neutralize anti-tumor actions and lead to systemic toxicity. During the prior term of this award, we gained an appreciation for the extracellular structural architectures of a spectrum of different immune cytokine complexes, including those of IL-2, IL-10, IL-12, IL-21, IL-22, IL-23, IL-27, IFN, and IFN. In this renewal application, with these structural templates in hand, in Aim 1 we focus our studies on the cytokines IL-2, IFN, and IL-12. These immune master regulators engage different but overlapping branches of the immune system, and share the issues of pleiotropy and toxicity that, if uncoupled, could lead to powerful cancer immunotherapeutic agents. We propose to “tune” signaling through structure-based cytokine engineering to attempt to create variants with decoupled pleiotropy, cell subset preferences, enhanced anti-tumor efficacies, and reduced toxicity – both alone and in combination. In Aim 2, we wish to understand the mechanistic basis for how tuned cytokines can differentially activate signaling inside the cells. Based on a recent breakthrough in our 20-year quest to solve the full-length JAK structure, we continue to pursue structural information on how cytokine binding to their receptors activates Janus Kinase (JAK) molecules, by reconstituting and imaging activated JAK homo- and heterodimers bound to both cytokine and intracellular JAKs and STATs. In this fashion, by combining structural biology, protein engineering, cell signaling, and in vivo tumor studies, we propose to obtain a complete molecular snapshot of cytokine receptor signaling from the initial engagement of ligand through the activation of intracellular signaling cascades and leverage this information for the engineering of cancer immunotherapeutics.

NIH Research Projects · FY 2026 · 2002-01

Project Summary This project will provide data critical to advance our understanding of enzyme sequence/structure/function relationships. We will develop innovative microfluidic tools and techniques and apply them to systematically study enzymes at a high-throughput and quantitative level. In the prior granting period, we developed HT- MEK (High-Throughput Microfluidic Enzyme Kinetics), which enabled recombinant expression, purification, and deep functional characterization of >1500 enzymes in parallel and returns kinetic and thermodynamic constants (e.g. kcat/KM, kcat, KM, Ki) at unprecedented scale. Here, we extend this platform to provide a suite of techniques (HT-MES, High-Throughput Microfluidic Enzyme Stability) capable of quantifying kinetic and thermodynamic stabilities (e.g. ΔGfold, kunfold) with similar throughput. We will also enable site-specific incorporation of noncanonical amino acids to provide the capability to isolate and systematically the impact of particular physicochemical residue properties and to investigate the impacts of post-translational modifications. During the prior granting period, our application of HT-MEK to profile multiple substitutions at each position and multiple functional parameters for the model alkaline phosphatase PafA revealed that residues with similar functional effects on catalysis formed large and spatially contiguous ‘regions’ that extended from the active site to distal surfaces, and that different regions affected different aspects of function. Here, we will systematically apply HT-MEK/S to a variety of new systems to build upon these observations and develop and test new models of how enzymes attain their functions and their observed kinetic and thermodynamic constants. Specifically, we will map functional couplings between residues in PafA via double and multi-mutant cycles and test the degree to which observations from PafA generalize by applying HT-MEK/S to investigate other alkaline phosphatase superfamily members and to acyl phosphatases, which provide ideal enzymes for developing and testing predictive computational models. We will also profile impacts of all nonsynonymous single nucleotide substitutions on folding and catalysis for protein tyrosine phosphatases (PTPs), providing clinically-relevant information about potential health consequences of mutations that can direct development of mechanistically relevant therapies and therapeutics. Finally, we will extend the reach of HT-MEK/S via collaborative projects with the Keedy and Zalatan labs. In all cases, we seek to use high-throughput data to test computational predictions, provide much-needed ground truth data for use by others, and reveal previously unattainable insights into the functional and energetic inner workings of enzymes.

NIH Research Projects · FY 2025 · 2001-09

This is a renewal application to RFA-HL-24-010 to propose that the Division of Blood and Marrow Transplantation and Cellular Therapy (BMT-CT) at Stanford University continue as a Core Clinical Center for the BMT Clinical Trials Network (BMT CTN). Stanford’s Program will perform ~700 adult transplants/cellular therapies in 2023 including autologous and allogeneic transplants using cells from matched and mismatched related and unrelated donors, cord blood units, ex vivo manipulated cell products and genetically modified cells, as well as novel chimeric antigen receptor T cell products. Stanford’s BMT-CT Program participates in basic and clinical research as a single institution, and within regional, national and international consortia. The Division is supported by a NIH Program Project Grant with over 32 years of funding of basic scientific research and clinical translational trials that advance the field. The Program is strengthened by highly experienced biostatics and data management groups, and a cGMP-compliant, FACT and CLIA certified Cellular Therapy Facility for routine cell processing and the development of investigational cellular therapies. The Program is a high accrual BMT CTN center and has enrolled 579 patients to 25 CTN studies. In the past granting period, Stanford investigators served in leadership roles of both committees and clinical trial protocols. Additionally, Stanford faculty participated on several protocol development committees and on other steering committees and task forces. Stanford will continue to leverage the capabilities of its Program to support the research goals of the CTN. It is our collective mission to advance the field of BMT-CT for patients with rare and difficult to treat blood diseases through high quality multi-center clinical trials. The goal of the proposed protocol in the current application is to determine whether a novel strategy can prevent and reduce the risk of developing graft-versus-host disease (GVHD) in patients lacking HLA matched donors, and thereby improve upon traditional allogeneic BMT. We propose a phase 1/2 clinical trial to evaluate the safety and efficacy of Orca-T, an engineered donor graft in which a highly purified Treg product is administered the same day as hematopoietic stem and progenitor cells followed 2 days later by conventional CD3+ T cells. The target population is patients with ALL, MDS, or AML receiving myeloablative conditioning followed by HCT using HLA mismatched unrelated donors (MMURDs). This investigator-initiated study builds upon our prior preclinical and clinical work with the Orca-T graft engineering platform in the HLA matched donor setting. The primary outcome of the Phase 2 portion is efficacy, as defined by GVHD-relapse-free survival (GRFS) at one year following HCT. Based on aggregated data, we hypothesize that the Orca-T allograft in combination with dual-agent GVHD prophylaxis (tacrolimus plus ruxolitinib) will synergistically and significantly reduce rates of GVHD in adults undergoing myeloablative allogeneic HCT with MMURDs. The Stanford BMT-CT program is committed to continued participation in BMT CTN trials, contributing concepts for consideration by the Network, and participating in Network committees and citizenship activities.

NIH Research Projects · FY 2026 · 2001-07

Summary Non-melanoma skin cancer, the most common US tumor, encompasses basal cell (BCC) and squamous cell (SCC) carcinomas, and leads to extensive morbidity and mortality. ARO46786 now in its 20th year, has focused on BCC-to-SCC transition (BST) a common but understudied resistance mechanism that represents a significant challenge to therapy and poorer outcomes. We propose five major BST Keratocarcinoma (KC) cell states and show that combinatorial AP-1 and SRF co-factor interactions provide the transcriptional switch from HH sensitivity to RAS-MAPK dependence and that c-FOS-driven BST appears reversible. By contrast, interrogation of stromal heterogeneity identified a skin cancer-associated macrophage (SCAM) required for BCC growth in allografts and organoids. SCAMs are self-propagating, transplantable, and long-lived in the BCC tumor environment. Tumor- associated macrophages have been shown to have proliferative and anti-immune checkpoint activity highlighting the gap in knowledge about the skin microenvironment. These preliminary studies support our overarching hypothesis that BST and SCAMs mediate BCC tumor evolution and resistance. To further test this hypothesis, ARO46786 will: Elucidate the drivers and sensitivities of BST by refining the KC chromatin and tumor dependency map and elucidating the mechanism of the reversible BST transcription factor switch; and elucidate the origin and function of SCAMs in BCC growth by dissecting SCAM function on tumor epithelial growth and identifying the determinants of SCAM polarization and function. Project completion will fill major gaps in our understanding how the tumor epithelium and microenvironment contribute to resistance, and lead to the nomination of novel cancer therapies.

NIH Research Projects · FY 2026 · 2000-09

Adeno-associated viral vectors (rAAV) have shown promise in some liver-based clinical trials including hemophilia B. However, one of the limitations is the loss of the vector genomes during tissue growth and cell division limiting the duration of expression when treatment is initiated early in life. For success in treating a number of genetic diseases treatment early in life is required. We previously developed a non-nuclease mediated AAV-mediated homologous recombination (AAV- HR) approach by using genomic homology arms to insert a protein coding sequence onto the end of the endogenous Albumin gene such that after homologous recombination, the modified locus would make a chimeric mRNA and both the endogenous albumin protein and a second protein, in our case human factor IX and treated the bleeding diathesis in a murine model of hemophilia B. Subsequently, this approach has been used to treat other murine models of human hepatodeficiency disorders and a Phase I/II clinical trial for methylmalonic acidemia was initiated by LogicBio therapeutics. The limitation is the efficiency of the process remains low and we have recently established that off-target integration likely can produce a proportion of the therapeutic protein. Our goal is to further study the genomic sources of the AAV-HR produced therapeutic product as well as parameters that influence the efficiency of AAV-HR in cells and in vivo. To do this, we will use high-throughput sequencing approaches to molecularly characterize the origin of off-target transcribed RNAs produced after AAV-HR mediated transduction. We will establish if they arise from vectors that integrate into regions of microhomologies or via non-homologous end joining as well as identify which sets of these RNAs are translated. Our preliminary data suggests that regions of the host genome that are more robust in R-loop formation will likely be more efficient at AAV-HR. We will directly test this hypothesis and use the information to design improved homology arms for AAV-HR. We will also do an unbiased in vivo genetic screen in mice to establish which genes when expression is reduced results in higher AAV-HR. We will confirm that parameters that influence non-nuclease mediated AAV-HR also effects nuclease mediated AAV-HR. The results from these studies will not only provide new mechanistic insights into AAV- HR but provide new approaches for enhanced genome editing, which would broaden the application for treating human genetic diseases.

NIH Research Projects · FY 2025 · 2000-08

Synaptic transmission between pre- and postsynaptic neurons occurs when the presynaptic neuron terminal is temporarily depolarized upon an action potential, opening Ca2+ channels near the active zones of synapses. Because the extracellular Ca2+ concentration is much higher than the cytoplasmic concentration, Ca2+ flows into the cytoplasm, triggering fusion of neurotransmitter-filled synaptic vesicles with the presynaptic membrane in less than a millisecond. Upon fusion, neurotransmitter molecules are released into the synaptic cleft, and then bind to receptors located in the postsynaptic membrane. Finally, the fusion machinery is recycled for further rounds of fusion in the presynaptic cell. Major questions about the molecular mechanisms of membrane fusion and protein recycling remain. The architecture of the fusion machinery between the synaptic vesicles and plasma membranes is unknown, and the molecular steps after Ca2+-triggering are unknown. Furthermore, our understanding of the molecular mechanisms governing synaptic release probability and presynaptic plasticity is incomplete. Obtaining three-dimensional images of synaptic proteins within their natural membrane environment will be an essential step towards answering these questions. We propose a stepwise, bottom-up approach starting with simpler systems and moving to increasingly more complex systems. First, we will employ a hybrid (ex vivo / in vitro) approach where synaptic vesicles are isolated from mouse brain homogenates and combined with synthetic acceptor vesicles. Functional tests of this hybrid system will be performed using a new single vesicle fusion assay that discriminates between different stages of membrane fusion and includes many presynaptic proteins, including but not limited to SNAREs, synaptotagmin, and complexin. The contact sites between isolated synaptic vesicles and synthetic vesicles will be imaged by cryo-electron tomography (cryo-ET) followed by subtomogram averaging to reveal the architecture of these presynaptic complexes in their membrane environment. Next, we plan to image the equivalent membrane contact sites in both isolated synaptosomes and in synapses of neuronal cultures grown on EM grids. We anticipate that reconstructions of presynaptic complexes obtained with the hybrid approach are the starting point for locating such complexes in synaptosomes and in synapses in neuronal cultures. These in vivo reconstructions might reveal new molecular interactions or associations. After fusion, the AAA+ protein NSF and associated SNAP co-factors are essential for disassembly of SNARE complexes and for quality control of newly formed SNARE complexes (in conjunction with Munc18 and Munc13). Previously, we determined single-particle cryo-electron microscopy (cryo-EM) structures of the complex of NSF, αSNAP, and the neuronal SNARE complex under non-hydrolyzing conditions. We now aim to investigate the molecular details of disassembly, what conformational changes are involved, and how ATP hydrolysis is coupled to these conformational changes by cryo-EM.

NIH Research Projects · FY 2025 · 1999-09

PROJECT SUMMARY A hallmark of the nervous system is its ability to transform and combine multiple sources of information to guide behavior, and to tune this signal processing through learning, for more accurate control of behavior. How this adaptive signal processing is implemented by neural circuits remains largely unknown, due in part to the presence of feedback loops that mix together and temporally shape neural signals in complex ways. This project uses an interdisciplinary experimental-computational approach to dissect the signal processing operations supporting the control of eye movements by vestibular and visual sensory inputs. Rigorous experimental protocols, computational models, and statistical fitting approaches will be developed to characterize the signal transformations occurring at multiple sites in the relevant neural circuitry, to disentangle the roles of feedforward and feedback signals in the control of eye movement behavior. These approaches will be used to analyze how specific changes in the way vestibular and visual signals are processed by the circuit support adaptive changes in the amplitude and timing of the eye movement responses to these sensory inputs. By advancing our understanding of how learning alters the way a circuit computes, this work bridges the key conceptual gap between neural plasticity at the synaptic level and learning at the behavioral level, and provides a scientific foundation for the development of improved clinical approaches for promoting recovery or replacement of vestibular function.

NIH Research Projects · FY 2026 · 1999-04

Project Summary/Abstract The Pacific Symposium on Biocomputing (PSB) is multidisciplinary conference covering current research in the theory and practice of computational methods as applied to significant biological problems. PSB 2025 – 2030 will be the 30th to 35th meeting of this series. Each February, individuals from the scientific community propose sessions and workshops for the PSB meeting to be held the following January. In the face of stiff competition, 5 to 6 sessions and 4 to 6 workshops are chosen. These sessions and workshops are often organized by junior scientists who are developing new research areas. Thus, PSB provides early opportunities for serious examination of emerging research areas. PSB also provides junior scientists the opportunity to gain a significant career boost by this activity. Our goals are for sessions to cover emerging areas and for workshops to bring attention to very newest developments. This bottom-up, crowdsourcing approach has worked well to reveal the newest developments in the diverse topics in biocomputing. At the meeting, session organizers present a short introduction to the session usually followed by a high-profile speaker, typically a biologist, who provides an overview highlighting the importance of the research area. Next, 4 to 10 oral presentations (based on the number of accepted papers) are delivered. Submitted papers are rigorously peer-reviewed, with typically ≤ 30% being accepted. These peer-reviewed papers are published in the annual PSB proceedings (viewed as a journal by PubMed), are open access, and indexed by PubMed. Additionally, the meeting has 4 to 6 workshops, two keynote speakers (scientific and ELSI), four invited speakers, and a continuously running poster session. The poster session is one aspect of the conference that provides junior scientists with opportunities to network with prominent scientists in the field. PSB has historically been held in Hawaii and the conference venue creates many opportunities for organic networking/collaborative discussions to arise. PSB has continually improved from participant feedback; specific examples of this improvement will be provided. The PSB meeting is highly regarded. PSB continues to foster the development of computational biology and bioinformatics by providing important, critical exposure to emerging areas and thus deserves continued support.

NIH Research Projects · FY 2026 · 1999-02

REGULATORS OF EPIDERMAL GENE EXPRESSION PROJECT SUMMARY Epidermal homeostasis is critical for skin barrier function and is disrupted in diseases such as psoriasis, atopic dermatitis, and skin cancer. During the current funding cycle, AR045192 identified new essential roles for specific heterogeneous nuclear ribonucleoproteins (HNRNPs) and ubiquitin- like proteins (UBLs) in epidermal homeostasis. These regulators can exert post-transcriptional and post-translational impacts, respectively, however their actions in the skin are poorly understood. This competing renewal will define how these newly uncovered HNRNPs and UBLs mediate opposing effects on epidermal growth and differentiation. For HNRNP RNA-binding proteins (RBPs), AR045192 knocked out all 33 HNRNPs to uncover new roles for HNRNPC and HNRNPU (pro-progenitor) as well as HNRPNH (pro-differentiation) in epidermal gene expression. HNRNPs exert diverse impacts across RNA lifecycles, however, their actions in epidermis are understudied. AR045192 therefore used non-isotopic ligation-based UV crosslinking immunoprecipitation-mass spectrometry to uncover dynamic assemblies of proteins on mRNAs adjacent to HNRNPC that cooperate with HNRNPC in progenitor mRNA surveillance. Aim I will further define the identity, assembly dependencies, and functions of proteins co-assembled on progenitor and differentiation mRNAs with HNRNPU, and HNRNPH to address the emergent model that specific HNRNPs assemble with distinct combinations of other RBPs on discrete sets of target RNAs to control epidermal homeostasis Conjugation to UBLs, including ubiquitin, NEDD8, and SUMO2, can alter a protein’s localization, interactions, stability, and function, however, roles for UBLs in epidermal homeostasis are not fully characterized. AR045192 knocked out 124 genes encoding all human UBLs, and the enzymes that conjugate them to their target proteins to uncover opposing UBL actions in epidermis. The NEDD8 UBL, along with its corresponding NAE1 and UBA3 enzyme subunits, was essential to maintain undifferentiated progenitor gene expression in human and mouse epidermal cells and tissues. In contrast, the SUMO2 UBL, along with its SAE1 and UBA2 enzyme subunits, was required for differentiation. HNRNPU was identified as NEDD8-tagged in undifferentiated keratinocytes and SUMO2-tagged in differentiating cells; disrupting this reciprocal tagging altered HNRNPU-bound target RNAs. Aim II will apply new methods to characterize the opposing actions of NEDD8 and SUMO2 UBLs in epidermal gene regulation. This effort will expand insight into how specific HNRNP and UBL regulators exert their newly identified essential impacts on epidermal gene expression.

NIH Research Projects · FY 2025 · 1999-01

PROJECT SUMMARY This is a renewal application of R01GM058867 which has supported our foundational efforts in chemical glycobiology tool development since 1999. Our current goal is to leverage glycochemistry and glycobiology in the development of proximity-based technologies for use in basic research and drug development. Molecules designed to induce neo-interactions among biomolecules are now well- established research tools and emerging clinical candidates. This paradigm is embodied by proteolysis-targeting chimeras (PROTAC), bifunctional molecules that form ternary complexes with target proteins and E3 ligase enzymes, leading to their degradation by the proteasome. Targeted protein degradation mediated by PROTACs is a powerful platform technology for basic research and for drug development. But this approach is limited to intracellular proteins; extracellular proteins that are secreted or plasma membrane-associated are mostly inaccessible to PROTAC-mediated degradation. In the previous granting period, we leveraged glycan- binding lysosomal trafficking receptors in a new modality for targeted degradation of extracellular proteins called lysosome-targeting chimeras (LYTACs). These comprised a target binder (e.g., antibody or nanobody) linked to a glycan-based ligand that engages either the ubiquitously expressed cation-independent mannose-6-phosphate receptor (M6PR) or the liver- specific asialoglycoprotein receptor (ASGPR). We developed LYTACs for both soluble and membrane-associated proteins of therapeutic relevance, and we used the LYTAC platform as a research tool alongside CRISPR screens to define cellular determinants of lysosomal trafficking and protein degradation. In the next funding period, we will build upon this work with three specific Aims. In Aim 1, we will develop tumor-immune cell targeted chimeras (TICTACs) as a new modality for cancer immune therapy. These molecules are intended to deplete immune checkpoint receptors from tumor-associated macrophages via internalization without affecting peripheral macrophages, thereby mitigating autoimmune toxicities. In Aim 2, we will develop LYTAC-antibody-drug conjugates (LYTAC-ADCs) that incorporate lysosomal trafficking receptor ligands into the ADC structure. The LYTAC component will drive internalization, thereby expanding the ADC target space to include cancer antigens that are inherently poor internalizers. Finally, in Aim 3 we will develop trogocytosis-targeting chimeras (TrogoTACs) as a new proximity-based modality for targeted protein transfer from one cell type to another.

NIH Research Projects · FY 2025 · 1997-09

This is an application to renew the Stanford Neurosciences graduate program, which leads to the only neuroscience-specific PhD degree at Stanford University. This interdisciplinary program consists of 88 students and 84 faculty from 23 departments (10 clinical, 13 basic science/engineering) across 3 schools. The range of departments illustrates the breadth of research areas, spanning from molecular/cellular to systems and behavior, human cognition, and translational work. The objective is to identify, recruit, and train predoctoral PhD students to become the next generation of leaders in neuroscience. Faculty receive training on admission standards, outreach to prospective students, and mentorship, and the program supports an active and engaged community that supports these priorities. The training grant and its implementation are the foundation of the program, providing a main source of student support for the first two predoctoral years. Stanford adds considerable value by subsidizing stipends to meet the high cost of living, paid administrative and faculty support, subsidies for 2 additional years of stipend and tuition, and many scientific and educational resources. The curriculum is designed to meet NIH standards for excellence in broad-based research training, experimental design, neural basis of disease, quantitative literacy, statistical methodology, rigor and reproducibility, and professional skills development. These curricular goals are achieved from day one, when students participate in an immersive neuroscience course to promote collaboration through active learning and mini-rotations. The first year provides broad-based learning through core modules that engage students in different ways within neuroscience subfields (anatomy, cellular, molecular, genetics, developmental, systems, cognitive, computational) to generate and test hypotheses using modern experimental, quantitative, and technological approaches. In the journal club, students broadly read the scientific literature, make oral presentations, critique rigor and statistical methodology, and engage in professional development activities. Students receive formal training in quantitative approaches, statistical analysis, rigor and reproducibility, and responsible conduct in research. Students select advanced topic and statistical courses to meet their scientific goals. Research training is provided by first-year rotations, followed by choice of a thesis lab, passing of an oral qualifying exam, thesis project, yearly committee meetings, and oral and written defense. Students receive mentoring from first-year advisors, thesis advisors, committee members, and peers. Program directors meet regularly with each student cohort and hold town hall meetings. Leadership is encouraged through student-run initiatives such as the annual retreat, teaching opportunities in courses and summer programs, outreach to local schools, and participation with faculty in committees including admissions, mental health, and curriculum.

NIH Research Projects · FY 2025 · 1997-09

The long term goal of this Research Program is to understand how newly translated proteins fold in eukaryotic cells. The proposed research will focus on folding events as they occur at the ribosome during synthesis of a polypeptide and will examine the role of molecular chaperones in their folding process. The conceptual framework for understanding de novo protein folding originates from our work in the previous funding cycle, which showed that a network of chaperones named CLIPS (Chaperones Linked to Protein Synthesis) is physically and functionally linked to the translation machinery. Our working hypothesis is that the CLIPS chaperones are tasked with guiding newly synthesized polypeptides to their folded conformation. Chaperone-mediated folding pathways appear to involve the cooperation of different classes of CLIPS, including chaperones that act early in the folding process, such as the Nascent Chain Associated Complex (NAC), the Hsp70 proteins and the GIM/prefoldin complex, and the mechanistically distinct chaperone TRiC/CCT, which appears to act later in the folding process. Our general strategy to elucidate how chaperones mediate the folding of newly synthesized proteins relies on the close integration of in vitro and in vivo approaches with computational biology analyses. Our proposed experiments are aimed at obtaining functional, mechanistic and structural insights into the role of chaperones in de novo folding.

NIH Research Projects · FY 2025 · 1997-05

Project Summary/Abstract: Hematopoietic cell transplantation (HCT) and cellular therapies are effective treatments for a broad range of hematological malignances, representing the first successful and widely applied cellular therapy for cancer. In this revised Program Project Grant competitive renewal (years 34-38), we will probe the cellular basis of transplantation biology (Project 1 and 3) and cell therapies with novel cellular populations, including iNKT cells (Project 1), CAR T cells (Projects 2 and 3), Treg cells (Project 2) and novel antibody constructs (Project 4). The overall goals of our Program Project Grant are to develop a more robust fundamental understanding of transplantation biology and cellular therapies and to address the major challenges of HCT including reducing transplantation risks, preventing graft-vs-host-disease (GvHD) and disease relapse. We will utilize innovative animal modeling, comprehensive biological and molecular analysis, novel imaging and biologically focused translational clinical trials. Our Program involves four highly interactive Projects focusing on: addressing transplantation risk (Projects 3 and 4), the biology and prevention of GVHD (Projects 1, 2 and 3), prevention and treatment of disease relapse with iNKT cells (Project 1), CAR T cells (Project 2 and 3) and targeted immunotherapy strategies focused on the stem cell antigen CD117 (Project 4). These Projects are supported by 3 Cores (Administration, Biostatistics and Data Management and Correlative Sciences). Through our highly interactive Program Project, we will gain novel insights into the biology of HCT and cellular therapies and develop innovative strategies to improve outcomes for patients with hematological malignancies. The knowledge gained has profound implications for extending these therapies to other cancers and for the treatment of patients with a broad range of immunological conditions such as autoimmune disorders and organ transplantation.

NIH Research Projects · FY 2025 · 1996-09

Acetaldehyde, a toxic metabolite of ethanol, contributes to many human pathologies. It is mainly detoxified by the mitochondrial aldehyde dehydrogenase 2 (ALDH2). In addition to the common ALDH2*2 of >500 million people of East Asian ancestry, we have identified additional common ALDH2 variants of other genetic ancestries. Ethanol exposure in cells expressing these less active ALDH2 variants increases aldehydic load, mitochondrial dysfunction, and cell death. Considering the critical role of aldehydes on health, that ethanol (via fermentation) was present throughout mammalian evolution, and that ALDH2 insufficiencies are common, we hypothesize that enzymes other than ALDH2 may provide an additional detoxification capacity against increased aldehydic load and the resulting cytotoxicity. We also hypothesize that inactive variants of these other enzymes may be common in specific genetic ancestry, rendering individuals who have ALDH2 insufficiency even more sensitive to ethanol-induced toxicity than carriers of ALDH2 insufficiency alone. If these hypotheses are correct, the assessment of ALDH2 genetic polymorphisms alone is not sufficient to assess disease risk. In this proposal, we focus on mitochondrial enzymes because of the central role of mitochondrial dysfunction in many human pathologies. Our specific aims for the project are: Aim 1: Identify variants common to specific ancestry groups of aldehyde metabolizing mitochondrial enzymes and determine whether they have reduced activity or stability in vitro Aim 2: Determine if inactive variants of these mitochondrial enzymes alone or together with ALDH2*2 (as a model of ALDH2 insufficiency) affect ethanol-induced mitochondrial dysfunction and inflammation in culture. Aim 3: Determine the impact of acute and chronic ethanol treatment, in vivo, in mice in which an inactive human ALDH2 variant and an inactive human variant of a second enzyme identified in Aim 2 were knocked in. Innovation: our work represents the first systematic study to begin identifying additional mitochondrial enzymes that complement ALDH2 in reducing ethanol- and aldehydes-induced toxicity. Our study is also the first to examine the potential risk from ethanol exposure in different human genetic ancestry using model systems. Significance: As ethanol is consumed by most adults in the world, if our hypotheses are correct that additional mitochondrial enzymes are critical in protecting from an increased mitochondrial aldehydic load, this study will have important clinical implications and may further contribute to precision medicine based on the activity of these understudied mitochondrial enzymes.

NIH Research Projects · FY 2025 · 1996-09

The analysis of large datasets from computational biology and medicine represents an important chal- lenge for Statisticians. These biomedical data typically have a large number of correlated features with rel- atively weak signals for predicting phenotypes of interest. Examples include DNA sequences and GWAS, mass-spectra, RNAseq and protein arrays. The broad goal of this ongoing three-investigator grant is to de- velop and study statistical techniques that enhance the analysis and interpretation of these data. The team combines experience in statistical modeling, algorithmic development, and theoretical analysis. Through four Specific Aims, the new projects focus on development and validation of state-of-the art statistical methods to use structure to learn from high-dimensional data to advance human population health. 1. Cluster-aware supervised learning. In “omics” settings, there are a large number of features that often exhibit sizable correlations. This aim proposes the Cluster-Aware Lasso, a statistical method which fits a lasso regression model that adaptively selects clusters of features using a hierarchical clustering-based approach, enforcing a notion of a tree-respecting solution. It will be validated on gene expression and mass spec data, and extensions to other supervised learning settings studied. 2. SNP Selection from GWAS summary statistics with FDR control. Genome-wide association studies often report findings for phenotypes in terms of summary statistics for individual SNPs. This aim develops a statistical method to identify causal SNPs while controlling the False Discovery Rate. It uses an estimate of the SNP correlation matrix based on linkage-disequilibrium data, an approximate multivariate lasso fit and model-X knockoff techniques, and will be validated on UK Biobank data. 3. Inference for high-dimensional genetic covariance matrices. Statistical estimation of large genetic covariance matrices is needed to learn whether genetic variation at phenome-wide scale is concen- trated in relatively few trait combinations, with implications for evolution and pleiotropy. This aim will explore biases in Restricted Maximum Likelihood and study alternative parametric and nonparametric methods of estimation both by asymptotic approximation and simulation. 4. Mixture lasso for multiple instance learning. It is often known whether a person is sick, but not which of their immune cells are responding to a particular illness, nor which parts of biopsied tissue are diseased. There is a label only for each patient, but data instances on a more granular level. The aim is to predict the labels of each data instance. This project proposes a supervised learning method based on mixtures and the lasso, with validation on viral sequence and mass spectrometry data. Working together, the investigators and their students will implement the new statistical tools into publi- cally available software, following a pattern established in earlier cycles of this grant.

NIH Research Projects · FY 2026 · 1996-08

MECHANISMS OF EPIDERMAL HOMEOSTASIS PROJECT SUMMARY Biomolecular cues that mediate homeostasis are not fully elucidated. During the most recent funding cycle, AR043799 found that intracellular glucose increases during differentiation of diverse cell types where it controls multimerization of nucleic acid binding proteins, independent of its role in energetics. This new role for glucose is analogous to action as a second messenger. In epidermis, glucose directly bound specific RNA binding proteins (RBPs), including DDX21, as well as specific DNA binding protein (DBP) transcription factors (TFs), including IRF6, to alter their dimerization in ways essential for differentiation. To understand principles of glucose action in homeostasis, this competing renewal will define the function of glucose binding to additional RBPs and TFs. For RBPs, we found that DEAD-box DDX RBPs were among the most enriched glucose binding proteins essential for epidermal differentiation. Glucose binding dissociated DDX21 dimers, propelling DDX21 monomers out of the nucleolar rRNA production machinery into nuclear complexes controlling splicing of essential pro-differentiation mRNAs. In contrast, glucose binding altered DDX50 association with an entirely different set of interactors and had no effect on RNA splicing, indicating that glucose binding engages a diversity of pro-differentiation mechanisms for specific DDX RBPs. Aim I will characterize glucose-enabled DDX50 pro-differentiation functions as well as the impacts of glucose binding on the subcellular localization, protein interactions, RNA interactions, mRNA splicing, and RNA-dependent protein assemblies of 3 other glucose-binding RNA helicases essential for epidermal homeostasis, namely DDX1, DDX17, and DDX18. For TFs, we found that glucose binding to IRF6 – in contrast to its dissociating effects on both DDX21 and DDX50 RBP dimers - induced IRF6 homodimerization, along with IRF6 DNA binding, genomic targeting, and differentiation gene transcription. This raised the question as to how glucose impacts other glucose-binding TFs essential for epidermal homeostasis. Among the latter is the pro- differentiation TF, TFAP2A, which binds DNA as a dimer with other AP-2 subunits. Aim II will characterize the effects of glucose binding on TFAP2A dimerization and target gene induction. Epidermal differentiation is enabled by other known factors, including calcium, specific adhesion proteins, and dominant differentiation-driving TFs. Aim II will therefore also determine the interplay between physiologic elevation of intracellular glucose and differentiation enabled by representative important contextual factors in epidermal cells. This effort will define the function of glucose-binding regulators in epidermal differentiation to expand insight into the mechanistic actions of newly identified biomolecular cues in homeostasis.

NIH Research Projects · FY 2025 · 1996-04

ABSTRACT The overarching aim of this proposal, which is in response to PA-20-185, asks whether alcohol use exacerbates the motor and cognitive deficits of Mild Cognitive Impairment (MCI) (cross-sectionally), and whether drinking accelerates progression of MCI toward dementia (longitudinally). The answers have implications for harm reduction, especially if "non-hazardous levels" of alcohol consumption prove to be deleterious to individuals with MCI, in terms of postural stability and risk of falling that can affect activities of daily living and quality of life. We propose three specific aims: Specific Aim 1: Test cross-sectional and longitudinal relations between drinking rates and static balance and dynamic gait in MCI and control men and women. Hypothesis: Higher rates of alcohol use will be associated with greater instability and slower gait in MCI compared with lower drinking MCI and controls matched in age, sex, and alcohol consumption variables. Hypothesis: At follow-up, accelerated cognitive decline in those with MCI will be predicted by slower natural gait and greater alcohol consumption compared with MCI with faster gait and lower alcohol consumption. Specific Aim 2: Use causal modeling to test multi-factorial determinants of static and dynamic stability metrics: alcohol consumption, sex, sensory physiology, cognitive performance, hematological measures of nutrition, physical and cognitive activities history, and activities of daily living (ADL). Hypothesis: Cross-sectionally, factors related to greater instability in MCI will be greater alcohol consumption, poorer sensory testing, and markers of poorer nutrition. In turn, these biomarkers of compromised function will predict poorer ADLs and quality of life. Hypothesis: Longitudinal measures will confirm cross-sectional relations and identify metrics that track standing instability, gait slowing, poorer ADL, and decline toward dementia with greater alcohol consumption. Specific Aim 3: Identify brain mechanisms of instability factors identified with causal modeling. Hypothesis: Factors related to greater imbalance and gait slowing identified with causal models will be related to smaller and compromised cerebello-pontine-motor cortical nodes and circuitry, whereas factors defining cognitive and mnemonic status will be related to smaller limbic and parietal volumes and diminished integrity of related circuitry. Both the balance and cognitive relations will be mediated by alcohol consumption rates. Hypothesis: Longitudinal analysis will reveal that these relations are enduring. Further, greater rates of alcohol consumption in the testing interim will result in accelerated functional decline in stability, gait speed, and cognitive status and cerebello-pontine-cortical circuitry. Exploratory analysis will use hypothesis-free, data-driven, in-house machine/deep learning methods to seek reliable constellations of factors that predict postural instability, gait impairment, and biomarkers of falling.

NIH Research Projects · FY 2026 · 1995-09

PROJECT SUMMARY/ABSTRACT The Saccharomyces Genome Database (SGD) is the comprehensive resource providing the highest quality reference information about the genome, and its elements, of the budding yeast, Saccharomyces cerevisiae. S. cerevisiae research has yielded fundamental knowledge about eukaryotic genetics, genome maintenance and regulation, and a variety of cellular processes. SGD provides a comprehensive resource that captures the known biology of S. cerevisiae and facilitates experimentation in biological systems. S. cerevisiae knowledge informs genetic medicine via annotation of human disease-related phenotypes and gene function through functional complementation between yeast and human homologs. S. cerevisiae is the most well-studied and well-understood eukaryote and the experimental literature for this yeast contains the collected results of decades of research. SGD collects, organizes, and connects the results from a large variety of genetic, molecular, and biochemical assays, extending this information by assimilating the results of large-scale genomic assays. SGD also provides links to other fungi and model eukaryotes via orthology and incorporates formalized and controlled vocabularies to represent biological concepts. The detailed annotation of yeast genes that have human orthologs provides substantial functional insight into the human orthologs. SGD maintains and broadens relationships with the greater scientific community and makes technical improvements through the development of tools and the use of third-party software that enhances the work of scientists and educators. SGD is a founding member of both the Gene Ontology Consortium and the Alliance for Genome Resources, and also collaborates with many components of the NCBI and the EBI. SGD provides a service to the yeast scientific community, as well as reaching out to scientists in the greater biomedical research community to serve those who have a need for genetic information that can be provided via the collected data on yeast genes, their products, and their functions.

NIH Research Projects · FY 2025 · 1995-09

We propose to continue the highly successful Stanford Genome Training Program (SGTP) for another five years, comprising the 28th to the 32nd year of the program. Since its inception, the SGTP has trained 224 Graduate Students and 79 Postdoctoral Fellows with outcomes of outstanding research contributions and publications, and many trainees later assuming careers in leadership positions in academia and industry. In this next funding period, we will continue to select and train exceptional students via extensive programs that include rigorous coursework, skill-building in computational and quantitative biology, training in the responsible conduct of research as well as in reproducibility and rigor, ethics, and other activities essential to scientific growth. After initial laboratory rotations, students will perform their thesis research in the laboratory of one of our 56 outstanding participating faculty, who hold primary appointments in 16 departments of four Stanford schools; the majority of these investigators have trained students or postdocs supported by the SGTP. Collaborations and interactions among faculty and students from different groups are commonplace and facilitate student success and interdisciplinary research. Our well-resourced laboratories and facilities enable the very best science in a wide range of genomics-related research areas. The Stanford School of Medicine is highly supportive with programs that foster general skills, well-being, and career advancement, and provides a highly interactive environment conducive to productive collaboration and communication. We will continue our efforts to provide a successful program with all individuals. We are enthusiastic to continue to train the next generation of science leaders and highly skilled researchers, exhibiting creativity, integrity, and productivity, both for academia and the private sector.

NIH Research Projects · FY 2026 · 1995-08

Project Summary/Abstract  The intersection of genomics and infectious diseases has defined one of the most important leading edges in contemporary science and one of the most critical areas of advancement in medicine, and as a result, has provided a rich intellectual foundation for the training of postdoctoral scholars in infectious diseases. The objectives of the grant are to train M.D. and Ph.D. post-doctoral fellows in the application of genomics to infectious diseases, and to prepare them for successful, productive, independent careers that have a significant impact on the health-related research needs of the nation. We propose to prepare all trainees irrespective of whether they intend to conduct their research at the bench or from the bedside, in fundamentals of both genomics and applied clinical investigation, with the goal of enhancing the translation of scientific discoveries into clinical practice. We request support for five postdoctoral fellows each year - Ph.D. candidates who have completed their thesis work and M.D. candidates who have completed their clinical training will be evaluated by the Program's Steering Committee and offered admission on the basis of exceptional academic record, faculty interviews, and interest in, and aptitude for research. The training program will be interdisciplinary, involving faculty from Biology, Biochemistry, Bioengineering, Chemical Engineering, Chemistry, Emergency Medicine, Genetics, Medicine, Microbiology & Immunology, Pathology, and Pediatrics. All fellows will train for at least two years. Each trainee will be encouraged to take courses in clinical research and in basic sciences related to genomics. Additionally, each trainee will complete a two-year interdisciplinary core curriculum in applied genomics of infectious diseases. Beginning in the first year, each fellow will embark on an in-depth research project supervised by one or more of the Program's faculty; in many cases, joint-mentoring will involve faculty from different disciplines. In this way, many research projects will be cross-disciplinary. Seminars by trainees and an annual research retreat will promote interactions between program participants. While support from the Training grant will be for two years, all fellows will be encouraged to begin to seek independent support after the first year. We will also consider funding for a third year if needed for a trainee’s trajectory. The program intends to meet a recognized need for clinician-scientists in Infectious Diseases who are trained in functional and applied genomics – a recognized strength at Stanford University School of Medicine. This training grant has been, and continues to be the primary source of postdoctoral research training support for clinician-scholars. Trainees are expected to transition to independent positions in academia or industry, or positions in public health leadership and apply their education in genomics to address pressing issues in infectious diseases.

NIH Research Projects · FY 2025 · 1994-09

Project Abstract: This is an application for competitive renewal of an institutional training grant that to date has trained 89 postdoctoral fellows for careers in preclinical, clinical and translational research in psychiatric disorders. Physician trainees are eligible if they have completed the PGY 3 year of psychiatry training or PGY 4 year of neurology; psychology trainees are eligible if they have completed an APA approved clinical internship. Fellows work with a mentor who is a member of the Stanford University faculty for a period of two years on projects related to the clinical/biological (including brain imaging) phenomenology, basic neuroscience (including optogenetics and organoids), and translational treatment outcomes of psychiatric disorders. The Program includes seminars in research methodology, clinical research design and statistical analysis. Additionally, all fellows are required to take a course focused on ethics in medical research and are encouraged to take the course in reproducibility of evidence. They have access to a wide array of elective courses and workshops offered at Stanford University. Mentorship emphasizes the development of an independent career. Fellows are expected to design and conduct their own research projects and are mentored in applying for funding to support their research post fellowship. An Executive Committee provides program oversight including continual review trainees' progress towards their individual goals and the of the training program. The Program is now in its 29th year. Fifty postgraduate trainees (MDs, MD/PhDs and PhDs) have been in the program in the past 15 years. Graduating fellows have been successful in obtaining faculty positions around the US and Canada (e.g., Stanford University, Weil-Cornell, Medical College, Yale University, UCSF, U. Pennsylvania, U Texas at Austin, Baylor, etc.) A strong majority continue to be involved in mental health research and have obtained funding to support their research, including NIH Career Development Awards: R01's; NIMH Director Awards (e.g., BRAINS and Pioneer Awards), Simons Foundation Awards, Welcome Trust Awards, NARSAD Young Investigator Awards, Klingenstein Foundation Awards, and other foundation grants. Building on the success of our program in attaining its goals, we aim to continue to train promising clinically trained fellows to become independent mental health researchers working to advance mental health care.

NIH Research Projects · FY 2025 · 1993-09

This application represents a competitive renewal of a highly successful institutional postdoctoral training grant based in Stanford’s Center for Interdisciplinary Brain Sciences Research (CIBSR), under the direction of Dr. Allan Reiss. The program is embedded in a research-intensive environment that is designed to facilitate the development of a new generation of investigators skilled in interdisciplinary research at the forefront of child psychiatry and neurodevelopment. Eligible candidates include MDs, MD/PhDs, and PhDs with potential for productive preclinical, clinical, or translational research careers. Physician candidates must have completed at least three years of training in psychiatry, pediatrics, or neurology, or plan to combine the T32 with their residency/fellowship. PhD candidates must have completed doctorates in relevant fields such as psychology, neuroscience, or computational sciences. Applicants with strong motivation to address precision medicine themes in pediatric neuropsychiatry are prioritized. Now in its 31st year, the program has trained 26 fellows (5 MDs, 3 MD/PhDs, 17 PhDs, 1 EdD) over the past 15 years. During the current grant period, the program met all goals: attracting 75 applicants, filling all slots (four per year), and retaining all but one trainee for 2–3 years (except one fellow who joined the faculty after one year), and prepared trainees for successful careers as independent investigators. The program fosters successful independent investigators and uses flexible mentor assignments with co-mentorship as the norm. The scientific scope is broad, spanning basic, clinical, and translational research. Several new junior mentors, including three Stanford postdoc alumni, have been added to partner with senior mentors, including three faculty who are graduates of Stanford postdoctoral fellowships. In the next 5-year period, 9–11 additional fellows will be trained. Plans are in place to increase MD and MD/PhD recruitment beyond the current 2 of 10 fellows. Each trainee will receive an individually tailored plan for didactics, mentorship, and research. We will focus the trainee’s curriculum and research training on bridging traditional, between-discipline gaps in methodology, and on focusing on the application of interdisciplinary solutions to complex precision medicine issues in child psychiatry and neurodevelopment.

NIH Research Projects · FY 2025 · 1993-02

This proposal is a competing renewal for our longstanding T32, the Stanford Cancer Imaging Training (SCIT) Program. Drs. Jeremy Dahl, PhD, and Bruce Daniel, MD, will lead this program, which features 25 mentors with independent funding and 10 (7 internal/3 external) distinguished program advisors. This is a 2-year program that trains 5 fellows (a mix of PhD and radiology-trained MDs) per year over a 5-year funding cycle. Our required coursework includes 2 courses in the clinical/cancer sciences, 2 in imaging science, 1 in biostatistics, 1 in medical ethics (“Responsible Conduct of Research”), 2 workshops in grant writing, an attendance at a minimum of 4 multidisciplinary tumor boards, and regular attendance during a continuing education workshop that covers topics in responsible conduct of research and rigor and reproducibility. In addition, trainees can select from a multitude of electives offered by various Stanford University faculty across numerous clinical, science, and engineering departments. Each trainee’s primary focus is a mentored cancer-imaging research project aimed at publications in peer-reviewed journals and presentations at National meetings. We pair each trainee with both a basic science and physician mentor, to provide guidance in course and research-topic selection and to develop a translational mindset. Through the SCIT program, we will continue our longstanding mission of training the next generation of researchers in the development and clinical application of advanced techniques for cancer imaging. We will recruit trainees from a nationwide pool to identify a strong set of trainees with a broad cancer imaging interests to develop the next generation U.S. research workforce. The need for the SCIT Program is even greater now than when it began in 1993. Radiology plays a key role in the diagnosis and treatment of cancer patients. Our Department is one of the very few that has been able to grow in response to this role and embrace what is now a multidisciplinary vision towards image-based cancer research. The SCIT Program leverages the Stanford Cancer Institute (an NIH-designated Comprehensive Cancer Center) and the Stanford Canary Center for Cancer Early Detection as well as many other Stanford resources and programs. All of our SCIT trainees were productive while in the program with nearly 90% who continue research activity in cancer imaging today. Current trainees are pursuing research in radiology-pathology fusion to predict treatment response of breast cancer, improving accuracy of prostate cancer detection on MRI with deep learning methods, optical coherence tomography histology to decrease the positive margin rate in lumpectomy for breast cancer, developing a partial ring time-of-flight positron emission tomography scanner with 3D event positioning to visualize and quantify cancer lesions, AI-based detection models to distinguish ductal carcinoma in situ from invasive breast cancer on pre-operative breast MRI, and optimizing MR lymphangiography to maximize clinically relevant data and drive therapeutic innovation.

NIH Research Projects · FY 2026 · 1991-04

PROJECT SUMMARY The Division of Neonatal and Developmental Medicine at Stanford University submits a competing renewal application to participate in the NICHD Neonatal Research Network (NRN). The Division and its faculty members have a long history of innovative research accomplishments in neonatal medicine. This expertise dovetails with the NRN goal of conducting definitive and rigorous multicenter trials and observational studies in newborns to improve survival without neurodevelopmental impairment. As a participant in the NRN since 1991, this center has proven to be highly productive, contributing to leadership, study design, protocol development, execution, analysis, and results dissemination. Led by PI, Krisa Van Meurs, Alternate PI, Valerie Chock, and Follow-up PI, Susan Hintz, the site neonatologists and subspecialty collaborators have wide-ranging expertise and significant clinical research experience with 613 manuscripts published in neonates since 2016. Within the NRN, Stanford investigators have 55 subcommittee assignments and appear as co-authors 117 times on NRN manuscripts. Dr. Van Meurs is Chair of the Publications Committee and coordinated 131 manuscript reviews, spearheaded a review of NRN policies leading to a reduction in time to publication, and was Editor for the Seminars in Perinatology volume describing NRN contributions over the last grant cycle. As Lead Follow-up PI and Chair of Follow-up Protocol Development, Dr. Hintz has been responsible for advancing protocols and proposals, and assuring the quality of NRN follow up visits, and enhancing rigor of training and certification procedures. During the COVID pandemic she led re-envisioning of certification processes, launched new tools, and developed individualized improvement plans for challenged sites. Cody Arnold is Co-PI for the ongoing Cycled Phototherapy trial. Valerie Chock led a NHLBI-funded secondary to the Transfusion in Prematures trial on near-infrared spectroscopy (NIRS), Dr. Van Meurs led a secondary to the Optimizing Cooling trial on amplitude-integrated EEG, and Courtney Wusthoff led a secondary study to the Preemie Hypothermia trial on EEG. Stanford neuroradiologists have been MRI central readers for all the NRN cooling trials. The Stanford site with its 2 satellite sites, El Camino Hospital and Santa Clara Valley Medical Center, provide an annual delivery base of >12,000 births with 59% born to high risk mothers, 140 NICU beds, and >1900 NICU admissions with 83% inborn. Two sites anticipate growth in deliveries over the next 3 years. Our aggregate population is diverse in comparison to the rest of the US with 25% Asian and 52% Hispanic. Over the last 6 grant cycles Stanford has demonstrated wide-ranging expertise, exceptional leadership, and strong collaborative abilities that have served the NRN well. One of the greatest strengths Stanford has to offer is our extensive and talented pool of young clinical investigators that will become the clinical research leaders of the future. In summary, the Stanford site has the neonatal and subspecialty faculty, physical space, research resources, professional staff, institutional support, and patient population necessary to continue as one of the finest centers participating in the NRN.