Scripps Research Institute

TECHNOLOGY TO EMPOWER CHANGES IN HEALTH (TECH) NETWORK PARTICIPANT TECHNOLOGIES CENTER

CONSORTIUM FOR HIV/AIDS VACCINE DEVELOPMENT

CENTER FOR HIV/AIDS VACCINE IMMUNOLOGY AND IMMUNOGEN DISCOVERY

THE COMPREHENSIVE CENTER FOR CHEMICAL PROBE DISCOVERY AND OPTIMIZATION AT SCRIPPS

CENTER FOR ANTIVIRAL MEDICINES & PANDEMIC PREPAREDNESS (CAMPP) - SUMMARY THE ONGOING COVID-19 PANDEMIC HAS BROUGHT TO LIGHT AN URGENT NEED TO ENHANCE THE THERAPEUTIC PREPAREDNESS FOR FUTURE VIRAL OUTBREAKS AND PANDEMICS. THE OVERARCHING GOAL OF THE “CENTER FOR ANTIVIRAL MEDICINES & PANDEMIC PREPAREDNESS” (CAMPP) IS THUS TO DEVELOP NOVEL STRATEGIES AND ENHANCE THE DRUG DISCOVERY PIPELINES FOR DIRECT-ACTING ANTIVIRALS AGAINST RNA VIRUSES OF PANDEMIC CONCERN. THE SPECIFIC FOCUS OF CAMPP WILL BE TO DEVELOP ANTIVIRALS AGAINST CORONAVIRUSES SARS-COV-2 (SCV2), SARS AND MERS; FLAVIVIRUSES INCLUDING ZIKA, WEST NILE AND DENGUE VIRUS; AND HEMORRHAGIC FEVER VIRUSES INCLUDING THE FILOVIRUS EBOLA, AND BUNYAVIRUSES SEVERE FEVER WITH THROMBOCYTOPENIA SYNDROME VIRUS AND LASSA VIRUS. INFECTION BY ANY OF THESE AGENTS HAS THE POTENTIAL TO CAUSE SEVERE HUMAN DISEASE WITH SIGNIFICANT MORTALITY RATES AND CURRENT OPTIONS FOR ANTIVIRAL TREATMENTS ARE LIMITED. THE ANTIVIRAL DRUG REMDESIVIR AND SEVERAL MONOCLONAL ANTIBODY TREATMENTS HAVE RECEIVED EMERGENCY USE AUTHORIZATION (EUA) FOR TREATMENT OF COVID-19, AND THE POLYMERASE INHIBITOR MOLNUPIRAVIR BY MERCK IS EXPECTED TO BE GRANTED EUA IN THE NEAR FUTURE. MONOCLONAL ANTIBODIES ARE ALSO AVAILABLE TO TREAT EBOLA VIRUS INFECTION, HOWEVER, NONE OF THESE DRUGS CAN BE ADMINISTERED ORALLY, POSING ADDITIONAL CHALLENGES IN TREATING EARLY INFECTION. THERE ARE NO APPROVED TREATMENTS FOR THE CAMPP FLAVIVIRUSES AND HEMORRHAGIC FEVER VIRUSES. TOWARD THIS END, WE HAVE ASSEMBLED A WORLD CLASS MULTIDISCIPLINARY TEAM OF INVESTIGATORS WITH EXPERTISE IN VIROLOGY OF RELEVANT VIRUSES, STRUCTURAL AND COMPUTATIONAL BIOLOGY, CHEMOPROTEOMICS, PHARMACOLOGY AND ORGANOID/ANIMAL MODELS, WHO WILL WORK CLOSELY WITH THE DRUG DEVELOPMENT EXPERTS AT THE DRUG DISCOVERY DIVISION OF THE SCRIPPS RESEARCH INSTITUTE, CALIBR, TO FURTHER THE DEVELOPMENT OF FOUR MAJOR CLASSES OF PROMISING ASSETS IN OUR DRUG DISCOVERY PIPELINE. FIRST, WE PROPOSE TO DEVELOP A POTENTIALLY BEST-IN-CLASS, ORALLY BIOAVAILABLE CORONAVIRUS PROTEASE (CLPRO) INHIBITOR FOR CORONAVIRUSES INCLUDING SCV2, FROM A LATE-STAGE DRUG ASSET UNDERGOING ADME OPTIMIZATION THAT IS EXPECTED TO ENTER IND-ENABLING STUDIES WITHIN THE NEXT THREE YEARS. SECOND, WE WILL IDENTIFY AND OPTIMIZE RNA POLYMERASE INHIBITORS FOR SCV2 AND OTHER CAMPP VIRUSES, WITH THE GOAL OF REACHING IND-ENABLING STUDIES WITHIN THE NEXT FOUR YEARS. THE THIRD FOCUS OF OUR PROPOSAL IS TO DEVELOP ANTIVIRALS AGAINST OTHER ‘DRUGGABLE’ PROTEINS ENCODED BY SCV2 AND ADDITIONAL VIRUSES POSING A PANDEMIC THREAT; THESE ASSETS INCLUDE INHIBITORS OF SCV2 HELICASE, E-PROTEIN ENCODED ION CHANNEL ACTIVITY, ENTRY AND FUSION ACTIVITIES, AND NUCLEOCAPSID, WITH THE GOAL OF OBTAINING IN VIVO PROOF-OF-CONCEPT FOR A SUBSET OF THESE MID-STAGE ASSETS. FINALLY, WE PROPOSE TO TARGET TRADITIONALLY CONSIDERED ‘UNDRUGGABLE’ NON-ENZYMATIC PROTEINS INCLUDING SCV2 AND FLAVIVIRAL STRUCTURAL PROTEINS AS WELL AS RNA STRUCTURE, TO DEVELOP NOVEL STRATEGIES FOR ANTIVIRAL DRUG DEVELOPMENT. CAMPP PROVIDES A HIGHLY INTEGRATED INFRASTRUCTURE OF INVESTIGATORS, EXPERTISE AND EXTERNAL PHARMACEUTICAL AND FOUNDING PARTNERS THAT WILL ENSURE THE CENTER’S SUCCESS IN ACHIEVING OUR GOALS AND NAVIGATING CHALLENGES OF THE DRUG DISCOVERY PROCESS.

SYSTEMS APPROACH TO IMMUNITY AND INFLAMMATION

JOINT CENTER FOR STRUCTURAL GENOMICS

CNS EFFECTS OF ALCOHOL: CELLULAR NEUROBIOLOGY

CONSORTIUM FOR IMMUNOTHERAPEUTICS AGAINST VIRAL HEMORRHAGIC FEVERS

CONSORTIUM FOR VIRAL SYSTEMS BIOLOGY (CVISB)

SCRIPPS TRANSLATIONAL SCIENCE INSTITUTE

REVERSING IMMUNE DYSFUNCTION FOR HIV-1 ERADICATION - PROJECT SUMMARY ALTHOUGH THE RATE OF NEW HIV INFECTIONS HAS DECREASED, CONTAINMENT AND EVENTUAL ERADICATION OF THE HIV PANDEMIC REMAINS A TOP PRIORITY IN CONTEMPORARY BIOMEDICAL RESEARCH. ONE OF THE MAJOR CHALLENGES TO HIV CURE IS THE NEED TO RESTORE NORMAL IMMUNE FUNCTION IN ORDER TO EFFECTIVELY ELIMINATE THE ESTABLISHED VIRAL RESERVOIR. WE HAVE ASSEMBLED IN RID-HIV: “REVERSING IMMUNE DYSFUNCTION FOR HIV-1 ERADICATION”, BASIC AND CLINICAL SCIENTISTS WITH EXPERTISE IN VIROLOGY, IMMUNOLOGY, MICROBIOME BIOLOGY, EPIGENETICS, AND SYSTEMS BIOLOGY. IN ADDITION, MERCK RESEARCH LABORATORIES WILL INVEST SIGNIFICANT INTELLECTUAL, HUMAN AND MATERIAL RESOURCES TO COMPLEMENT THE EFFORTS OF THE ACADEMIC SCIENTISTS. THE RID-HIV COLLABORATORY WILL COLLECTIVELY FUNCTION TO EXPLORE THE UNDERLYING BASIS OF THE IMMUNE DYSREGULATION IN HIV-INFECTED INDIVIDUALS AND THE IMPACT IT HAS ON RESERVOIR PERSISTENCE AND VIRAL REBOUND CONTROL. WE WILL TEST FOR THE FIRST TIME SEVERAL INNOVATIVE CONCEPTS, INCLUDING IDENTIFYING EPIGENETIC MECHANISMS IMPRINTED BY THE MICROBIOME AND HOST AND BACTERIAL METABOLOMES THAT PREVENTS THE DEVELOPMENT OF EFFECTIVE INNATE AND ADAPTIVE IMMUNE RESPONSES THAT CAN CONTROL THE SIZE, QUALITY AND ANATOMICAL LOCALIZATION OF THE HIV RESERVOIR. THE OVERARCHING GOAL OF THE RID-HIV COLLABORATORY IS TO PROVIDE PRECLINICAL IN VIVO PROOF-OF- CONCEPT FOR A THERAPEUTIC PARADIGM THAT ENCOMPASSES IMMUNE RESTORATIVE TREATMENTS, USED IN CONCERT WITH ENHANCED VIRAL REACTIVATION AND ELIMINATION STRATEGIES, IN ORDER TO DELIVER A HIV-1 CURE. WE PROPOSE THREE HIGHLY INTEGRATED AND COMPLEMENTARY SCIENTIFIC RESEARCH FOCI (RFS), TO BE SUPPORTED BY RIGOROUS AND ITERATIVE MODELING OF OUTCOMES AND SHAPED BY OUR OUTREACH TO THE HIV COMMUNITY. IN RF1 WE WILL INVESTIGATE THE MECHANISMS WHEREBY HOST- AND MICROBIOME-DERIVED METABOLITES IMPACT INNATE IMMUNE RESPONSES AND INFLUENCE THE MAINTENANCE OF THE LATENT VIRAL RESERVOIR. IN RF2 WE WILL PURSUE THE HYPOTHESIS THAT IN ART/ATI CLINICAL COHORTS, METABOLITES THAT GOVERN INNATE IMMUNITY SHAPE THE ADAPTIVE IMMUNE RESPONSES THAT COULD PREVENT VIRAL REBOUND UPON TREATMENT INTERRUPTION. IN ADDITION, WE WILL EVALUATE THE CAPACITY OF ENGINEERED ALLOGENIC STEM MEMORY T CELLS TO PROVIDE SUPERIOR COGNATE HELP TO PROMOTE THE EFFECTOR FUNCTIONS OF ANTIVIRAL CD8 T CELLS, AND WILL ASSESS THE ABILITY OF FDA-APPROVED AND NOVEL IMMUNE MODULATORS TO RESET THIS BASELINE IMMUNE DYSFUNCTION AND ENHANCE THE FUNCTION OF THIS NOVEL CELL THERAPY PRODUCT. IN RF 3 WE WILL OPTIMIZE A BEST-IN-CLASS LATENCY REVERSAL AGENT (LRA) AND IDENTIFY CLINICAL-STAGE MOLECULES WITH SYNERGISTIC LRA ACTIVITY. CLEARANCE OF REACTIVATED CELLS WILL BE ENHANCED USING A NOVEL STRATEGY FOR NK CELL RECRUITMENT AND BY GENETICALLY MODIFYING B CELLS TO PRODUCE BROADLY NEUTRALIZING HIV-1 ANTIBODIES THAT ENHANCE RESERVOIR CLEARANCE. FINALLY, GENE EDITING WILL BE DEPLOYED FOR IN VIVO TARGETING AND ELIMINATION OF LATENT PROVIRUS NOT AMENABLE TO LRAS. THE OUTCOMES OF STUDIES IN RF1, RF2 AND RF3 WILL ENABLE THE SYNTHESIS OF A PREDICTIVE MATHEMATICAL MODEL TO ESTABLISH THE MOST LIKELY COMBINATIONS OF THERAPIES TO ACHIEVE AN HIV-1 CURE, AND WHICH WILL BE TESTED IN A CAPSTONE AIM TO ESTABLISH PROOF-OF-CONCEPT FOR THESE STRATEGIES IN NHP MODELS AND TO ENABLE TRANSLATION TO THE CLINIC.

SCRIPPS CLINICAL AND TRANSLATIONAL SCIENCE HUB - PROJECT SUMMARY / ABSTRACT THE SCRIPPS RESEARCH TRANSLATIONAL INSTITUTE (SRTI) IS THE FOUNDATION FOR THE SCRIPPS HUB AND IS DEDICATED TO ACCELERATING SCIENCE THAT WILL IMPROVE HUMAN HEALTH. SRTI’S EMPHASIS ON GENOMICS, DIGITAL MEDICINE, AND INFORMATICS/ANALYTICS FOSTERS A MULTI-DIMENSIONAL UNDERSTANDING OF INDIVIDUALIZED HUMAN HEALTH. THE SCRIPPS HUB HAS PREVIOUSLY INCLUDED THE SCRIPPS RESEARCH INSTITUTE (SR) AND SCRIPPS HEALTH (SH) AS PRINCIPAL PARTNERS, INCLUDING RADY CHILDREN’S INSTITUTE OF GENOMIC MEDICINE (RCIGM) FOR GENOMIC MEDICINE INITIATIVES AND CALIFORNIA INSTITUTE FOR MEDICAL RESEARCH (CALIBR) FOR DRUG REPURPOSING / DISCOVERY. IN THE NEW CTSA CYCLE, OUR PARTNERS HAVE EXPANDED TO INCLUDE SAN DIEGO STATE UNIVERSITY (SDSU), ENABLING US TO COMBINE FORCES IN COMPUTER SCIENCE, ARTIFICIAL INTELLIGENCE, BIOSENSORS, AND DIVERSITY INCLUSION AND EQUITY INITATIVES. SRTI’S EXPERTISE IN GENOMICS WAS HIGHLIGHTED DURING THE PANDEMIC. WE BECAME ONE OF THE COUNTRY’S MOST PRODUCTIVE SEQUENCING CENTERS FOR SARS-COV-2 BY RAPIDLY FORMING THE “SEARCH” ALLIANCE TO PROCESS SAMPLES WITH THE SAN DIEGO COUNTY DEPARTMENT OF HEALTH, UCSD, SHARP HEALTH, CALIFORNIA DEPARTMENT OF PUBLIC HEALTH, HELIX, AND OUR PARTNERS RCIGM AND SH. DIGITAL MEDICINE HAS BEEN ONE OF SRTI’S CORE STRENGTHS, HAVING PIONEERED THE FIRST REMOTE, SITE-LESS DIGITAL CLINICAL TRIAL AND NOW EXPANDING THIS METHODOLOGY TO ADDRESS MANY OTHER MEDICAL CONDITIONS SUCH AS HEALTH DURING PREGNANCY AND SLEEP DISORDERS. OUR DIGITAL TRIALS CENTER LAUNCHED THE DIGITAL ENGAGEMENT & TRACKING FOR EARLY CONTROL AND TREATMENT SCRIPPS (DETECT) STUDY, AND IN A MATTER OF WEEKS OUR TEAM WAS ABLE TO ACCURATELY PREDICT THE LIKELIHOOD OF COVID WITH SARS-COV-2 USING PASSIVELY COLLECTED RESTING HEART RATE DATA FROM WRISTBAND SENSORS AND LATER TO IDENTIFY A PHYSIOLOGIC SIGNATURE THAT CORRELATES WITH LONG COVID (POST-ACUTE SEQUELAE OR SARS-COV-2 INFECTION). THE SCRIPPS HUB WILL INNOVATE CLINICAL AND TRANSLATIONAL SCIENCE TO IMPROVE HUMAN HEALTH ACROSS THE LIFESPAN AND DIVERSE RACIAL, ETHNIC, GEOGRAPHIC AND SOCIOECONOMIC COMMUNITIES. THE HUB WILL PROVIDE A NURTURING ENVIRONMENT FOR EDUCATION, TRAINING, AND CAREER DEVELOPMENT WITH A FOCUS ON INDIVIDUALIZED HEALTH DOMAINS OF GENOMICS, DIGITAL MEDICINE, AND BIOMEDICAL INFORMATICS, TO EMPOWER TOMORROW’S DIVERSE WORKFORCE.

PROTEIN CARBOHYDRATE INTERACTIONS IN CELL COMMUNICATION

SCRIPPS TRANSLATIONAL SCIENCE INSTITUTE

JOINT CENTER FOR STRUCTURAL GENOMICS (JCSG-2)

HIV MACROMOLECULAR INTERACTIONS AND IMPACT ON VIRAL EVOLUTION OF DRUG RESISTANCE

ELICITING NEUTRALIZING ANTIBODIES AND B CELL RESPONSES USING NOVEL HIV ENV IMMUNOGENS IN NON-HUMAN PRIMATES - NEUTRALIZING ANTIBODIES ARE LIKELY TO BE REQUIRED FOR AN EFFECTIVE HIV-1 VACCINE. HOWEVER, FEW CANDIDATE VACCINES EFFICIENTLY ELICIT BROADLY NEUTRALIZING ANTIBODIES (BNABS) FOLLOWING VACCINATION. RECENTLY, THE VRC WORKING GROUP HAS ELICITED FUSION PEPTIDE-DIRECTED BNABS IN GUINEA PIGS AND NON-HUMAN PRIMATES (NHPS). SIMILARLY, WE RECENTLY ELICITED BNABS IN RABBITS FOLLOWING HETEROLOGOUS NFL (UNCLEAVED) TRIMER-LIPOSOME PRIME:BOOSTING AT SCRIPPS WHERE WE ACTIVATED B CELL RESPONSES WITH TARGETED N-GLYCAN DELETIONS IN THE PRIMING IMMUNIZATIONS (DUBROVSKAYA ET AL, IMMUNITY 2019). WE ISOLATED TWO RABBIT BNABS THAT RECAPITULATE THE SERUM ACTIVITY. THESE BNABS ARE DIRECTED AGAINST TWO DISTINCT SITES OF ENV VULNERABILITY. THE ELICITATION OF NEUTRALIZING RESPONSES WAS ENHANCED BY TARGETED N-GLYCAN DELETION, HIGH-DENSITY LIPOSOMAL ARRAY AND HETEROLOGOUS TRIMER RESTORATIVE BOOSTING ENV. IN INDEPENDENT EXPERIMENTS IN GUINEA PIGS, WE HAVE ELICITED BNABS IN MULTIPLE ANIMALS ALSO USING A N-GLYCAN DELETION, HETEROLOGOUS ENV NFL PRIME:BOOST APPROACH. THESE RECENT OUTCOMES ARE ENCOURAGING INROADS TOWARD THE SUCCESSFUL SOLUTION OF A 3-DECADE-LONG PROBLEM. THE NEW ERA OF NEAR-NATIVE TRIMERIC SPIKE MIMICS, COUPLED WITH PARTICULATE ARRAY, STRUCTURE-INFORMED DESIGN AND HIGH-RESOLUTION ANALYSIS OF ENV-SPECIFIC B CELL AND LYMPH NODE RESPONSES, AFFORD NEW OPPORTUNITIES TO MORE EFFICIENTLY ELICIT BNABS. WE PROPOSE AN INTEGRATED, MULTI-FACETED APPROACH BLENDING THE EXPERTISE OF WORLD LEADERS IN HIV ENV TRIMER DESIGN, ANALYSIS OF B CELL RESPONSES AND ABS FOLLOWING VACCINATION, NHP IMMUNE TISSUE ANALYSIS AND EM-BASED ANALYSIS OF ONGOING IMMUNE RESPONSES AND HIGH-RESOLUTION AB:TRIMER INTERACTIONS. WE WILL USE WELL-ORDERED TRIMER PRIME:BOOSTING THAT ELICITED BNABS IN RABBITS AND GUINEA PIGS TO ELICIT SUCH RESPONSES IN NHPS, TRANSLATING SUCCESS IN SMALL ANIMALS TO NHPS USING NOVEL IMMUNOGEN DESIGN AND PRESENTATION IN PROJECT 1 (WYATT) AND IMMUNIZATION OF NFL TRIMERS INTO NHPS VIA CORE B (SILVESTRI). WE WILL USE “REAL-TIME” SERUM FAB-TO-TRIMER BINDING EVALUATED BY EM POLYCLONAL IGG EPITOPE MAPPING (EMPEM) IN CORE C (WARD) IN COMPLEMENT WITH RAPID MAB NGS-BASED MAB CLONING, SEQUENCING AND FUNCTIONAL EXPRESSION IN PROJECT 2 (KARLSSON HEDESTAM). IN COLLABORATION WITH THE VRC (MASCOLA) WE WILL DEFINE EITHER NON-NEUTRALIZING MABS TO MASK UNWANTED NON-NEUTRALIZING EPITOPES OR TO BETTER DISPLAY THE EPITOPES OF CROSS-NEUTRALIZING MABS. WE WILL COMPARE NFL TRIMER-LIPOSOMES TO CELL SURFACE NFL TRIMER ARRAY EXPRESSED FROM THE EXCITING MRNA LIPID ENCAPSULATION TECHNOLOGY. WE WILL ASSESS IF IMMUNIZATION IN THE JUVENILE NHP B CELL REPERTOIRE COMPARED TO ADULT MACAQUES WILL BETTER GENERATE BNABS AS IS OBSERVED DURING HUMAN INFECTION. TO FOLLOW OUR DISCOVERY OF A TIER 2 CD4BS-DIRECTED BNAB FOLLOWING TRIMER-LIPOSOME VACCINATION, TERMED E70, WE WILL TARGET THE CD4BS BY DIRECTED NFL TRIMER DEGLYCOSYLATION IN THE NHPS. BASED ON OUR RECENT DISCOVERY OF THE VERY BROADLY NEUTRALIZING VACCINE-INDUCED MAB, 1C2, WE WILL ALSO FOCUS ON THE GP41:120 TRIMER INTERFACE. LEVERAGING THESE INITIAL LEADS, WE WILL UNDERTAKE A MULTIFACETED, CROSS-COMPONENT INTEGRATED APPROACH TO GUIDE THE ELICITATION OF BNABS IN NHPS FOLLOWING VACCINATION WITH NEAR-NATIVE, UNCLEAVED NFL ENV TRIMERS.

GENOMICS FOR KIDNEY TRANSPLANTATION

SCRIPPS TRANSLATIONAL SCIENCE INSTITUTE (UL1)

JCSG CENTER FOR INNOVATIVE MEMBRANE PROTEIN TECHNOLOGIES

HIGH RESOLUTION ANALYSIS OF ENV-DIRECTED B CELLS TO ACCELERATE VACCINE DESIGN

ADULT STEM CELLS FOR THERAPY OF VISUAL DISORDERS

STUDIES OF JOINT AGING AND OSTEOARTHRITIS

NEUTROPHIL LINEAGE IN INFLAMMATION - ABSTRACT NEUTROPHILS CONSTITUTE THE FIRST LINE OF CELLULAR DEFENSE AGAINST PATHOGENIC MICROORGANISMS. IN RESPONSE TO PRO- INFLAMMATORY CUES, UNRESTRICTED NEUTROPHIL ACTIVATION INDUCES TISSUE DAMAGE. TO AVOID DELETERIOUS EFFECTS TO THE HOST, NEUTROPHIL NUMBERS, ACTIVATION, AND LIFESPAN MUST BE TIGHTLY REGULATED, BUT THE MOLECULAR MECHANISMS THAT CONTROL NEUTROPHILS IN THE CONTEXT OF INFLAMMATORY DISEASE REMAIN ELUSIVE. CARDIOVASCULAR DISEASE IS THE LEADING GLOBAL CAUSE OF DEATH. RECENT EVIDENCE SUPPORTS AN IMPORTANT ROLE FOR NEUTROPHILS IN THE DEVELOPMENT OF CORONARY ARTERY DISEASE (CAD). NEUTROPHILS ARE PRESENT IN EARLY AORTIC LESIONS AND IN RUPTURE-PRONE ATHEROSCLEROTIC PLAQUES, AND A POSITIVE CORRELATION BETWEEN PLASMA LEVELS OF NEUTROPHIL SECRETORY PROTEINS AND CAD HAS BEEN ESTABLISHED, SUGGESTING THAT NEUTROPHIL EXOCYTOSIS MEDIATES DETRIMENTAL EFFECTS IN CAD. FURTHERMORE, NEUTROPHIL PRODUCTION IS INCREASED IN THE BONE MARROW IN ATHEROSCLEROTIC MODELS AND NEWLY IDENTIFIED NEUTROPHIL PRECURSORS ARE NOW KNOWN TO MEDIATE INFLAMMATION. HOW NEUTROPHIL SUBSETS CONTRIBUTE TO DISEASE PROGRESSION IN CAD HAS NOT BEEN STUDIED AND THE REGULATION OF NEUTROPHIL DIVERSITY IN DISEASE IS UNKNOWN. THE INFLAMMASOME IS AN EMERGING DRIVER IN ATHEROSCLEROSIS; HOWEVER, THE ROLE OF THE NLRP3 INFLAMMASOME ACTIVATION SELECTIVELY IN NEUTROPHILS ON ATHEROGENESIS HAS NOT BEEN STUDIED AND THE MECHANISMS REGULATING THE FUNCTIONS OF NEUTROPHIL LINEAGE CELLS IN THE CONTEXT OF INFLAMMASOME ACTIVATION AND ATHEROGENESIS REMAIN UNKNOWN. IN THIS SYNERGISTIC PROGRAM, PROJECT 1 NEUTROPHIL DEVELOPMENT DURING INFLAMMATION AND ATHEROSCLEROSIS WILL STUDY HOW NEUTROPHIL HETEROGENEITY IS MODULATED IN HUMAN SUBJECTS WITH CAD, AND HOW THE NLRP3 INFLAMMASOME IN NEUTROPHIL PROGENITORS INFLUENCES GRANULOPOIESIS AND NEUTROPHIL HETEROGENEITY IN ATHEROSCLEROSIS. PROJECT 2 NEUTROPHIL MECHANISMS DURING INFLAMMATION AND ATHEROSCLEROSIS WILL TEST THE HYPOTHESIS THAT HYPERLIPIDEMIA DIFFERENTIALLY REGULATES VESICULAR TRAFFICKING AND ASSOCIATED FUNCTIONS OF NEUTROPHIL PRECURSORS IN CAD, ESTABLISH MECHANISMS OF NLRP3-INDUCED NEUTROPHIL EXOCYTOSIS DYSREGULATION AND IMPLEMENT TRANSLATIONAL APPROACHES TO DECREASE NEUTROPHIL INFLAMMATION IN CAD. PROJECT 3 NEUTROPHIL SURVIVAL AND DEMISE DURING INFLAMMATORY STATES WILL CHARACTERIZE THE EXPRESSION AND FUNCTION OF COMPONENTS OF THE NLRP3 INFLAMMASOME IN CELLS OF THE NEUTROPHIL LINEAGE, AND WILL DEFINE THE EFFECTS OF HYPERLIPIDEMIA-INDUCED INFLAMMATION AND THE ROLES OF DEATH RECEPTOR SIGNALING IN IL-1SS PRODUCTION, MITOCHONDRIAL APOPTOSIS IN VIABILITY OF NEUTROPHIL LINEAGE CELLS, AND NECROPTOSIS SIGNALING IN ATHEROGENESIS. OUR SYNERGISTIC AND UNIQUE PROGRAM USES THE COMPLEMENTARY EXPERTISE OF THREE RENOWN RESEARCHERS, EXPERTS IN THE AREAS OF NEUTROPHIL DEVELOPMENT, NEUTROPHIL INTRACELLULAR FUNCTION REGULATION AND INFLAMMATION, TO STUDY THE CENTRAL HYPOTHESIS THAT UNRESTRICTED ACTIVATION OF NEUTROPHIL PROGENITORS AND MATURE NEUTROPHILS IS A FUNDAMENTAL PROCESS IN CARDIOVASCULAR DISEASE. THESE STUDIES WILL LEAD TO NOVEL APPROACHES TO TREAT NEUTROPHIL-MEDIATED INFLAMMATION IN CAD.

THROMBUS FORMATION AND ANTITHROMBOTIC INTERVENTION

WEST AFRICAN EMERGING INFECTIOUS DISEASE RESEARCH CENTER (WA-EIDRC)

A GENE THERAPEUTIC APPROACH TO STABLE SUPPRESSION OF HIV-1 REPLICATION

CYTOCHROME P-450 POLYMORPHISM

SCALABLE SYNTHESIS AND NEW BOND DISCONNECTIONS

HIV INTERACTIONS IN VIRAL EVOLUTION

SINGLE-MOLECULE DNA SEQUENCING WITH ENGINEERED NANOPORES

ELECTROPHYSIOLOGY OF ALCOHOL IN EXTENDED AMYGDELA

BROADLY EFFECTIVE HCV VACCINE - PROGRAM SUMMARY THE OVERARCHING GOAL OF THIS P01 PROPOSAL IS TO DEVELOP NOVEL HEPATITIS C VIRUS (HCV) VACCINE CANDIDATES THAT CAN ELICIT POTENT BROADLY NEUTRALIZING ANTIBODY RESPONSES IN IMMUNIZATION. DESPITE HIGHLY EFFECTIVE ANTIVIRAL DRUGS AGAINST HCV NOW BEING AVAILABLE, THE CONTINUOUS INCREASE IN NEW INFECTIONS UNDERSCORES THE REAL-WORLD CHALLENGES IN COMBATING THIS HUMAN INFECTION WITHOUT A VACCINE. THIS P01 PROPOSAL “BROADLY EFFECTIVE HEPATITIS C VACCINE” IS BUILT ON THE HYPOTHESIS THAT AN HCV VACCINE EFFECTIVE AGAINST DIVERSE CIRCULATING HCV STRAINS CAN BE DEVELOPED THROUGH RATIONAL ENGINEERING OF VACCINES TO ENHANCE ANTIGEN IMMUNOGENICITY AND PRESENTATION OF CONSERVED NEUTRALIZING EPITOPES, AND TO TARGET WELL-DEFINED MULTIDONOR CLASS BROADLY NEUTRALIZING ANTIBODY (BNAB) RESPONSES TO HCV. MULTIDONOR CLASS ANTIBODY RESPONSES ARE ANTIBODIES SHARING COMMON GENETIC AND FUNCTIONAL FEATURES PRODUCED IN INFECTION OR VACCINATION AT THE POPULATION LEVEL. THIS P01 PROGRAM CONSISTS OF 2 RESEARCH PROJECTS, SUPPORTED BY AN ADMIN CORE AND 2 SCIENTIFIC CORES. THE OVERALL AIMS OF THE PROGRAM ARE: (1) TO DETERMINE THE STRUCTURES OF HCV ENVELOPE GLYCOPROTEINS IMPORTANT FOR RATIONAL VACCINE DESIGN; (2) TO RATIONALLY DESIGN HCV VACCINE ANTIGENS FOR ELICITATION OF BROADLY NEUTRALIZING ANTIBODIES THAT WILL BE EFFECTIVE AGAINST DIVERSE CIRCULATING VIRAL STRAINS; (3) TO DETERMINE THE ANTIBODY RESPONSES ELICITED BY HCV VACCINE ANTIGENS IN PRECLINICAL ANIMAL MODELS. SUCCESS IN THIS RESEARCH PROGRAM WILL RESULT IN BOTH BASIC SCIENTIFIC KNOWLEDGE AND A STRONG HCV VACCINE CANDIDATE FOR FUTURE PILOT PRODUCTION AND CLINICAL TESTING.

NEURPSYCHOPHARMACOLOGY-MULTIDISCIPLINARY TRAINING

FOUR-DIMENSIONAL HETEROGENEITY OF FLUID PHASE BIOPSIES IN CANCER (4DB-CENTER)

MOLECULAR MECHANISMS LINKING AGING, ABETA PROTEOTOXICITY AND NEURODEGENERATION

DEVELOPMENT AND ADVANCEMENT OF BROAD-SPECTRUM RESPIRATORY ANTIVIRALS

MICRORNAS FOR THERAPY OF VISUAL DISORDERS

RESISTANCE DRIVEN STRUCTURAL DESIGN FOR HIV THERAPIES

DEVELOPMENT OF CARBOXYL- AND AMIDE-DIRECTED C-H ACTIVATION/C-C COUPLING REACTIONS

ANTIBODY EFFECTOR FUNCTION IN PROTECTION AGAINST HIV-1

AUTODOCK SOFTWARE DEVELOPMENT AND MAINTENANCE

CHEMICAL PROTEOMIC PLATFORMS FOR RADICALLY EXPANDING CANCER DRUGGABILITY

INTEGRATIVE NEUROSCIENCE INITIATIVE ON ALCOHOLISM - WEST - SUPPLEMENT

MANAGING ATHEROSCLEROSIS BY MODULATING HDL FUNCTION

DEVELOPMENT OF OREXIN-1 RECEPTOR ANTAGONISTS FOR DRUG ADDICTION

ROLE OF RAB27 IN GRANULOCYTE FUNCTION

CHEMOPROTEOMIC IDENTIFICATION AND THERAPEUTIC VALIDATION OF PROTEINS OF METABOLIC SIGNIFICANCE

PROTEOMIC ANALYSIS OF PLASMINOGEN RECEPTORS

PROBING THE BIOCHEMICAL MECHANISM OF AMYLOID DISEASE

DISSECTING POLYCLONAL SERA TO REVEAL CORRELATES OF PRODUCTIVE IMMUNE RESPONSES TO HIV

DESIGN AND TESTING OF GERMLINE-TARGETING AND BOOSTING IMMUNOGENS TO ELICIT 10E8-LIKE BROADLY NEUTRALIZING ANTIBODIES AGAINST HIV

MULTIDIMENSIONAL DEVELOPMENT OF HIGH-AFFINITY ANTI-GLYCAN ANTIBODIES TO FIGHT DEADLY BACTERIAL INFECTIONS - ABSTRACT THE DEVELOPMENT OF IMMUNOTHERAPIES FOCUSED ON THE SURFACE GLYCANS OF BACTERIA IS HYPOTHESIZED TO BE A POTENTIAL PARADIGM SHIFT IN THE FIGHT AGAINST LIFE-THREATENING AND ANTIBIOTIC-RESISTANT BACTERIA, AN EMERGING AND INCREASING HEALTH CONCERN FOR WHICH THERAPEUTIC OPTIONS ARE LIMITED. OUR PO1 TEAM WILL USE CHEMISTRY TO DECONSTRUCT AND DISPLAY BACTERIAL GLYCAN STRUCTURES ON AN ARTIFICIAL PLATFORM TO MAKE THEM IMMUNOGENIC AND RECOGNIZED BY THE IMMUNE SYSTEM. IMMUNE RESPONSES WILL BE ANALYZED AND DISSECTED BY BACTERIOLOGISTS, CELLULAR AND STRUCTURAL IMMUNOLOGISTS TO DETERMINE THE CHARACTERISTICS OF WHAT MAKES A VACCINE OR AN ANTIBODY AGAINST GLYCANS EFFECTIVE AS AN ANTIBIOTIC AND DEPLOYABLE IN PRE-CLINICAL STUDIES. THIS PROGRAM THAT ASSEMBLES SOME OF THE WORLD EXPERTS IN THEIR RESPECTIVE FIELDS IS AMBITIOUS AND INTENDS TO PIONEER THE EFFORT OF PLACING IMMUNOTHERAPY NEXT TO CHEMOTHERAPY FOR THE TREATMENT OF BACTERIAL INFECTIONS. OUR UNIQUE COMBINATION OF CHEMISTRY-IMMUNOLOGY- BACTERIOLOGY-STRUCTURAL BIOLOGY WILL PROVIDE THE NECESSARY MECHANISTIC UNDERSTANDING OF WHAT QUALIFIES A VACCINE OR AN ANTIBODY TO BE EFFECTIVE IN IMMUNOTHERAPY. THE TEAM IS ALREADY PRODUCTIVE AND HAS PUBLISHED THE PROOFS OF PRINCIPLE OF THE APPROACH ON WHICH THE SCIENCE OF THIS APPLICATION IS BASED: VERY HIGH AFFINITY ANTIBODIES CAN BE PRODUCED AGAINST BACTERIAL GLYCANS EXPOSED AT THE SURFACE OF ANTIBIOTIC RESISTANT BACTERIA AND ARE EFFECTIVE AT COMBATING INFECTIOUS CHALLENGES. WE WILL EXPAND OUR STRATEGY TO THE SURFACE GLYCANS OF THREE BACTERIAL PATHOGENS LISTED BY WHO AS “CRITICAL” OR “HIGH” PRIORITY: STAPHYLOCOCCUS AUREUS, KLEBSIELLA PNEUMONIAE, AND NEISSERIA GONORRHEA. THE FUNDAMENTAL KNOWLEDGE THAT WE WILL GAIN FROM OUR STUDIES SHOULD ESTABLISH A VERY DETAILED BLUEPRINT OF THE IMMUNE RECOGNITION OF GLYCANS AND GLYCOPEPTIDES BY THE IMMUNE SYSTEM. THE INTEGRATION OF THE CHEMISTRY, IMMUNOLOGY, AND STRUCTURAL BIOLOGY FACETS OF THE PROJECT DIRECTLY INTO THE BACTERIOLOGY AND IN VIVO MODELS, WILL IDENTIFY GLYCANS TARGETS AND STRATEGIES TO INITIATE PRE-CLINICAL STUDIES.

PRECLINIAL DEVELOPMENT OF JNK3 INHIBITORS TO TREAT PARKINSON'S DISEASE

KNOCKIN MICE EXPRESSING BROADLY NEUTRALIZING HIV ANTIBODIES 4E10 AND B12

ELUCIDATING CARDIOVASCULAR PHENOTAYPES EMPLOYING GENOME EDITING OF IPS CELLS

STRUCTURE AND FUNCTION OF PROTEIN C

EPIGENOMIC MODULATION OF CYSTIC FIBROSIS

CANCER GENES: TRANSLATING BASICS TO APPLICATIONS

PROTEOGENETICS

REGULATION OF PROTEIN C PATHWAYS

ANTITUMOR ANTIBIOTICS

THE MULTI-OMICS VACCINE EVALUATION (MOVE) CONSORTIUM - PROJECT SUMMARY AN HIV VACCINE REPRESENTS OUR BEST OPPORTUNITY FOR ELICITING DURABLE, PROTECTIVE IMMUNITY AND ENDING THE DECADES-LONG AIDS PANDEMIC. TO BE SUCCESSFUL, AN ANTIBODY-BASED HIV VACCINE MUST RELIABLY INDUCE BROADLY NEUTRALIZING ANTIBODIES (BNABS), MOST LIKELY VIA A SERIES OF IMMUNOGENS OR MIXTURES OF IMMUNOGENS DELIVERED SEQUENTIALLY. TO AVOID VIRAL ESCAPE, BNABS OF AT LEAST 2-3 DISTINCT SPECIFICITIES MUST BE INDUCED CONCURRENTLY. THUS, A MORE HOLISTIC VIEW OF HIV VACCINE COMPOSITION WOULD RESEMBLE A MATRIX OF IMMUNOGENS RATHER THAN A SEQUENCE. CONSTRUCTING A SINGLE SET OF IMMUNOGENS THAT RELIABLY INDUCE BREADTH AGAINST JUST ONE EPITOPE IS A DAUNTING CHALLENGE. SUCCEEDING IN THE EXPONENTIALLY MORE DIFFICULT TASK OF ASSEMBLING A COHESIVE IMMUNOGEN MATRIX WILL REQUIRE SIGNIFICANT IMPROVEMENTS IN THE SPEED AND GRANULARITY WITH WHICH WE CAN DESIGN AND EVALUATE VACCINES. HERE, WE PROPOSE A SOLUTION: BY EMBEDDING MULTI-OMICS TECHNOLOGY AT EACH STAGE OF IMMUNOGEN DEVELOPMENT, WE CAN UNLOCK A REVOLUTIONARY INCREASE IN SCALE WHILE SIMULTANEOUSLY IMPROVING THE DEPTH AND RESOLUTION OF OUR ANALYSES. THE MULTI-OMICS VACCINE EVALUATION (MOVE) CONSORTIUM BRINGS TOGETHER A MULTI- DISCIPLINARY TEAM OF EXPERT INVESTIGATORS WITH A LONG HISTORY OF PRODUCTIVE COLLABORATION. OUR CENTRAL HYPOTHESIS IS THAT DEEPLY INTEGRATING ADVANCED MULTI-OMICS APPROACHES THROUGHOUT OUR ITERATIVE VACCINE DEVELOPMENT PIPELINE WILL SPEED THE DISCOVERY OF AN HIV VACCINE BY ALLOWING US TO OPERATE CONCURRENTLY ACROSS MULTIPLE IMMUNOGENS. OUR OVERALL MISSION IS TO ACCELERATE DEVELOPMENT OF AN HIV VACCINE BY PARALLELIZING THE DESIGN AND TESTING OF A MATRIX OF COMPLEMENTARY IMMUNOGENS THAT RELIABLY INDUCE BROAD, DURABLE IMMUNITY. TO ACCOMPLISH OUR OBJECTIVES, WE PROPOSE THE FOLLOWING SPECIFIC AIMS: SPECIFIC AIM 1: PROFILE THE HUMAN IMMUNOGENICITY OF A NOVEL V2 APEX-FOCUSING ENV TRIMER. SPECIFIC AIM 2: EVALUATE Q23-ELICITED IMMUNE RESPONSES, CANDIDATE BOOSTING IMMUNOGENS, AND DELIVERY STRATEGIES IN HUMANIZED ANIMAL MODELS. SPECIFIC AIM 3: DEVELOP THE Q23 BACKBONE INTO A MULTI-EPITOPE PRIMING IMMUNOGEN.

UNUSUAL PHARMACOPHORES AND NEW TOOLS FOR CROSS-COUPLING

GERMLINE TARGETING INFLUENZA IMMUNOGENS

ACTIVITY DEPENDENT CONTROL OF VISUAL SYSTEM DEVELOPMENT

FAAH: STRUCTURE, FUNCTION, AND IN VIVO INHIBITION

STUDIES ON MICROTUBULE BINDING PROTEINS

NOVEL PROTEOMICS APPROACH TO HIV-ASSOCIATED NEUROCOGNITIVE DISORDER & DRUG ABUSE

CELL SPECIFIC PERTURBATIONS OF THE PROTEOME IN ALZHEIMER'S DISEASE - ALZHEIMER’S DISEASE (AD) IS THE LEADING CAUSE OF AGING-RELATED COGNITIVE DECLINE, AFFECTING MORE THAN 5 MILLION AMERICANS OVER 65 YEARS OLD, AND THE NUMBER OF PATIENTS IS EXPECTED TO CLIMB TO 13 MILLION AS BABY BOOMERS AGE. OUR ABILITY TO LEARN AND REMEMBER DECLINES WITH AGE DUE TO PROGRESSIVE CHANGES IN SYNAPTIC CONNECTIVITY AND FUNCTION. SYNAPTIC ABNORMALITIES ALSO COMMONLY PRECEDE NEURONAL LOSS DURING EARLY STAGES OF ALZHEIMER’S DISEASE (AD) AND OTHER NEURODEGENERATIVE DISORDERS. HOWEVER, WE ARE ONLY BEGINNING TO UNDERSTAND THE FULL SPECTRUM OF THESE ABNORMALITIES, THEIR CONTRIBUTIONS TO COGNITIVE DECLINE, AND THE UNDERLYING MECHANISMS. THIS CHALLENGE IS LARGELY ATTRIBUTED TO THE COMPLEXITY OF THE BRAIN AND ETIOLOGIES OF AGING AND NEURODEGENERATION. HALLMARKS OF ADVANCED ALZHEIMER’S DISEASE (AD) INCLUDE ACCUMULATIONS OF EXTRACELLULAR AMYLOID PEPTIDES AND INTRACELLULAR HYPERPHOSPHORYLATED TAU PROTEIN AS WELL AS CHRONIC NEUROINFLAMMATION. WHILE GENES FOR FAMILIAL AD HAVE BEEN IDENTIFIED, WHICH SHED SUBSTANTIAL LIGHT ON THE ETIOLOGY OF THE DISEASE, THE MECHANISMS BEHIND SPORADIC ONSET AD STILL REMAIN A MYSTERY. WHILE STUDIES OF TRANSCRIPTIONAL DYNAMICS IN THE BRAIN HAVE BEEN TRANSFORMATIVE, TRANSCRIPTIONAL DYNAMICS DO NOT CORRELATE WELL WITH PROTEIN DYNAMICS BECAUSE PROTEIN SYNTHESIS, TURNOVER AND SUBCELLULAR LOCALIZATION ARE MORE TIGHTLY REGULATED SPATIALLY AND TEMPORALLY THAN TRANSCRIPTION. STUDIES HAVE SUGGESTED THAT PROTEOSTASIS DECLINES WITH AGE, IMPAIRING CELLS FROM MANAGING THE INEVITABLE MISFOLDING OF PROTEINS. OUR TEAM WILL STUDY PROTEIN DYNAMICS IN ANIMAL MODELS BASED ON BIO-ORTHOGONAL NON-CANONICAL AMINO ACID (BONCAT) PROTEIN LABELING. THESE METHODS ALLOW US TO MEASURE DYNAMICS IN PROTEIN SYNTHESIS AND DEGRADATION IN SPECIFIC BRAIN CELL TYPES RELEVANT TO AD. THESE MEASUREMENTS WILL PROVIDE NEW INFORMATION ABOUT THE DISRUPTION OF NORMAL CELLULAR PROCESSES. THE OVERREACHING GOAL OF OUR COLLABORATIVE PROPOSAL IS TO BRIDGE CRITICAL GAPS IN KNOWLEDGE BY LEVERAGING THE STATE-OF-THE-ART METHODS FOR BIO-ORTHOGONAL NON-CANONICAL AMINO ACID TAGGING (BONCAT) AND QUANTITATIVE MASS SPECTROMETRY (MS) TO IDENTIFY BRAIN CELL-TYPE CONTRIBUTIONS TO SYNAPTIC AND NEURONAL DECLINE ASSOCIATED WITH AD.

CFTR PROCESSING IN THE ENDOPLASMIC RETICULUM

BIOLOGICALLY ACTIVE CYCLIC PEPTIDES

NOVEL CHEMICAL AND IMMUNOLOGICAL APPROACHES TO INFLUENZA THERAPY

RISK FACTORS FOR ALCOHOLISM IN NATIVE AMERICANS

IN VIVO INCORPORATION OF UNNATURAL AMINO ACIDS

INTEGRINS AND OCULAR ANGIOGENESIS

MECHANISMS OF FORCE SENSING IN THE NERVOUS SYSTEM

STRUCTURE-FUNCTION OF CYTOPROTECTIVE COAGULATION PROTEASES AND THEIR RECEPTORS

NATIONAL RESOURCE FOR AUTOMATED MOLECULAR MICROSCOPY

EEG AND ERP MEASURES OF ALCOHOL'S EFFECTS

DISCOVERING SMALL MOLECULES ACTIVATORS OF STRESS RESPONSIVE SIGNALING

SMALL MOLECULE THERAPEUTICS FOR BOTULLINUM NEUROTOXIN A

THE NOCICEPTIN ORL1 SYSTEM: TREATMENT TARGET FOR RELAPSE

THE ROLE OF CYCLIN E IN GROWTH CONTROL AND TUMOROGENESIS

MEDICATION DEVELOPMENT FOR PROTRACTED ABSTINENCE IN ALCOHOLISM

RIBONUCLEOPROTEIN COMPLEXES REGULATING T-CELL ACTIVATION

GENETIC CONTROL OF DEVELOPMENT

BUILDING A SOMATOSENSORY ATLAS FOR INTEROCEPTION - PROJECT SUMMARY/ABSTRACT SOMATOSENSORY NEURONS ARE WELL-KNOWN FOR INNERVATING SKIN AND MUSCLE AND TRANSMITTING SENSATIONS SUCH AS TOUCH, PAIN, ITCH, AND PROPRIOCEPTION TO THE BRAIN. HOWEVER, THESE SAME TYPE OF HETEROGENEOUS SET OF NEURONS ALSO INNERVATE INTERNAL ORGANS AND ARE RESPONSIBLE TO CONVEY INFORMATION OF THE INTERNAL STATE OF OUR BODY. THESE INTEROCEPTIVE SIGNALS INCLUDE BOTH CONSCIOUS (BLADDER FULLNESS) AND SUBCONSCIOUS (BLOOD PRESSURE) SENSES. DESPITE THEIR IMPORTANCE, WE DO NOT KNOW THEIR PROJECTION PATTERNS AND PHYSIOLOGICAL ROLES. HERE, WE PROPOSE TO USE A SET OF INNOVATIVE TOOLS THAT WILL ALLOW US TO SPECIFICALLY LABEL, VISUALIZE, AND ISOLATE SENSORY NEURONS INNERVATING VARIOUS INTERNAL ORGANS. USING A VARIETY OF STATE-OF-ART MOLECULAR, IMAGING, AND GENOMIC TECHNIQUES, WE WILL BUILD AN ANATOMICAL DATABASE OF THESE NEURONS AND USE SNRNASEQ TO ESTABLISH THE MOLECULAR IDENTITIES OF INDIVIDUAL SENSORY NEURONS BASED ON THEIR INNERVATING ORGANS. FINALLY, THE ANATOMICAL, CELLULAR, AND MOLECULAR MAPS WILL BE STANDARDIZED, INDEXED, AND INTEGRATED INTO AN ATLAS THAT WILL ALLOW THE COMMUNITY TO FREELY ACCESS THE DATA.

BIOGPS: EXTENSIBLE WEB 2.0 GENE PORTAL FOR STRUCTURED AND UNSTRUCTURED ANNOTATION

INTEGRATIVE OMICS ANALYSIS OF HUMAN CARTILAGE IN AGING AND OSTEOARTHRITIS

GENE NETWORK PERTURBATIONS IN ALCOHOL DEPENDENCE - A SYSTEMS BIOLOGY APPROACH

PROTEIN DYNAMICS IN DIHYDROFOLATE REDUCTASE CATALYSIS

TRANSCRIPTIONAL CONTROL OF SYNAPTIC PLASTICITY BY CLASS IIA HDACS

LEADERSHIP IN AD/ADRD DRUG DISCOVERY - SUMMARY THIS R35 LEADERSHIP APPLICATION IS RELEVANT TO MILESTONE 6D OF THE GOALS/MILESTONES OF THE NIH ALZHEIMER’S DISEASE AND ALZHEIMER’S DISEASE RELATED DEMENTIAS (AD/ADRD) SUMMITS—“INITIATE DRUG DISCOVERY EFFORTS TO DEVELOP NOVEL THERAPEUTIC AGENTS.” OUR OVERRIDING GOAL IN THIS APPLICATION IS TO SUPPORT THE INFRASTRUCTURE THE PI IS ASSEMBLING FOR AD/ADRD DRUG DISCOVERY AT SCRIPPS RESEARCH, WHICH WILL CONTINUE WELL AFTER THE AWARD IS OVER. OUR MAJOR PLANNED MILESTONE IS AIMED TO INSPIRE PROGRAM BUILDING AND CREATE TEAM BUILDING FOR AD/ADRD DRUG DISCOVERY USING THE VERY CONSIDERABLE INSTITUTIONAL RESOURCES AT SCRIPPS RESEARCH AND ITS CALIBR DRUG DISCOVERY CENTER. WE ARE ALSO BUILDING ONE OF THE NATION’S TOP-RANKED GRADUATE SCHOOL PROGRAMS IN DRUG DISCOVERY VIA OUR MENTORING USING THE FACULTY OF SCRIPPS’ #1/#2-RANKED CHEMISTRY- BIOCHEMISTRY DEPARTMENTS IN THE WORLD. AS PROOF OF FEASIBILITY, OF THE SIX CURRENTLY FDA-APPROVED MEDICINES FOR AD, THREE WERE DEVELOPED OUT OF THE PI, DR. LIPTON’S LABORATORY. VIA THIS R35 LEADERSHIP APPLICATION, DR. LIPTON WILL MENTOR OTHERS TO DEVELOP AD/ADRD-RELATED DRUGS ACTING AT NOVEL TARGETS TOWARD DISEASE-MODIFYING THERAPY. SPECIFICALLY, DR. LIPTON WILL SERVE AS RESEARCH MENTOR FOR NEW INVESTIGATORS AND EARLY STAGE INVESTIGATORS (NI/ESI) IN AD/ADRD DRUG DISCOVERY. TO DATE, VIRTUALLY ALL AD/ADRD THERAPEUTICS EVALUATED IN HUMAN CLINICAL TRIALS HAVE SO FAR FAILED TO SHOW DISEASE-MODIFYING EFFECTS FOR AD/ADRD. FOR EXAMPLE, THE LACK OF SIGNIFICANT EFFICACY IN CLINICAL TRIALS WITH ASS-CENTERED THERAPEUTICS TO DATE DEMANDS A NEW PARADIGM FOR DEVELOPMENT OF EFFECTIVE AD THERAPEUTICS. TO OVERCOME THESE SIGNIFICANT GAPS IN KNOWLEDGE AND TO ADVANCE THERAPIES, IMPROVEMENTS IN THE MODELS AND STRATEGIES BY WHICH AD/ADRD RESEARCHERS FUNCTION AND EXECUTE NEED TO OCCUR. ONE SOLUTION TO THIS CHALLENGE IS TO UNITE THE BEST BASIC SCIENTISTS WITH TRAINING IN NEUROSCIENCE, STRUCTURAL BIOLOGY, BIOINFORMATICS AND COMPUTATIONAL BIOLOGY, WITH EQUALLY EXPERT TEAMS IN THE PHARMACEUTICAL INDUSTRY TRAINED IN HIGH-THROUGHPUT SCREENING, ASSAY DEVELOPMENT, MEDICINAL CHEMISTRY, CHEMICAL BIOLOGY AND PHARMACOLOGY. WITH A COLLECTIVE AND HARMONIZED TEAM ALONG WITH SIGNIFICANT MENTORING EFFORTS FOR JUNIOR FACULTY (NI/ESI), THE PI, DR. LIPTON, HAS INITIATED THREE MAJOR EFFORTS TO ADDRESS THIS UNMET MEDICAL NEED, WHICH WILL BE CARRIED OUT UNDER THE AUSPICES OF THE CURRENT R35 LEADERSHIP AWARD APPLICATION: · INVESTIGATION, IDENTIFICATION AND CHARACTERIZATION OF POTENTIAL AD/ADRD THERAPEUTIC TARGETS IN THE CONTEXT OF THE CENTRAL PATHOPHYSIOLOGICAL PROCESSES IN AD/ADRD. · EXECUTION OF DRUG DISCOVERY CAMPAIGNS TO DEVELOP INVESTIGATIONAL NEW DRUG (IND) CLINICAL CANDIDATES ITERATIVELY WITH THE TARGET ELUCIDATION EFFORTS, WHILE BUILDING UPON SCRIPPS DRUG DISCOVERY INFRASTRUCTURE TO ACCOMPLISH THIS. · CONTINUE TO MENTOR JUNIOR FACULTY AND BUILD A WORLD-CLASS GRADUATE SCHOOL AROUND THE CHEMICAL BIOLOGY OF DRUG DESIGN FOR AD/ADRD AT SCRIPPS (CURRENTLY RANKED #2 BY US NEWS FOR BIOCHEMISTRY).

IMMUNOLOGY TRAINING

ORGANIZATION AND FUNCTIONS OF THE NUCLEAR LAMINA

SINGLE NUCLEUS GENE EXPRESSION IN MODERATE AND COMPULSIVE DRUG SELF-ADMINISTRATION IN A RODENT MODEL OF HIV - SUMMARY THE ABUSE OF STIMULANTS SUCH AS METHAMPHETAMINE (METH) EXACERBATES THE DELETERIOUS EFFECTS OF HIV INFECTION. HERE, WE WILL CARRY OUT SINGLE NUCLEUS RNA-SEQ WITH THE GOAL OF IDENTIFYING CELL TYPES AND CELL STATES THAT ARE PIVOTAL IN THE EFFECTS OF HIV AND CHRONIC METHAMPHETAMINE (METH) SELF-ADMINISTRATION ON KEY BRAIN REGIONS RELEVANT TO THE EFFECTS OF PERSISTENT HIV INFECTION AND METH USE DISORDER IN HIV TRANSGENIC (TG) RATS, WHICH HARBOR A NON-REPLICATING HIV-1 TRANSGENE AND EXPRESS CHRONIC LOW-LEVELS OF MULTIPLE HIV-1 PROTEINS. THE OCCASIONAL BUT LIMITED USE OF A DRUG IS CLINICALLY DISTINCT FROM ESCALATED DRUG USE, WHICH IS CHARACTERIZED BY THE EMERGENCE OF CHRONIC COMPULSIVE DRUG-SEEKING AND TAKING. THUS, WE WILL USE AN ESTABLISHED, STATE-OF-THE-ART PARADIGM OF VOLUNTARY INTRAVENOUS DRUG SELF-ADMINISTRATION UNDER LONG ACCESS (LGA) CONDITIONS THAT LEADS TO ESCALATED (COMPULSIVE) METH INTAKE IN COMPARISON TO SELF-ADMINISTRATION UNDER SHORT ACCESS (SHA) CONDITIONS, WHICH LEADS TO A MODERATE AND STABLE PATTERN OF METH INTAKE. THE PARADIGM OF ESCALATED DRUG INTAKE UNDER LGA CONDITIONS IS HIGHLY RELEVANT TO THE HUMAN SUBSTANCE USE DISORDER (SUD) AS IT MODELS ALL 7 OF THE CRITERIA FOR DRUG ADDICTION IN THE DIAGNOSTIC AND STATISTICAL MANUAL OF MENTAL DISORDERS (DSM)-IV AND 7 OF THE 11 CRITERIA IN THE DSM-V. WE SHOWED THAT HIV TG RATS SELF-ADMINISTERING METH IN THIS PARADIGM DISPLAY INCREASED COMPULSIVITY, NEUROINFLAMMATION, AND NEURAL INJURY. THE PROJECT WILL ADDRESS THE FOLLOWING VEXING QUESTION ABOUT PERSISTENT HIV INFECTION IN THE CNS: WHAT ARE THE CELL TYPES AND CELL STATES THAT DRIVE NEUROINFLAMMATION, NEURODEGENERATION, AND COMPULSIVE METH ABUSE IN THE SETTING OF HIV THAT CAN REVEAL THE PATHOGENIC MECHANISMS BEHIND NEUROHIV DISEASE PROGRESSION, VIRUS EXPRESSION AND PERSISTENCE? THE OVERARCHING HYPOTHESIS BEHIND THE PRESENT PROJECT IS THAT THE EXPLORATION OF THE GENE REGULATORY NETWORK AT THE SINGLE CELL LEVEL WILL ELUCIDATE KEY MECHANISMS THAT UNDERLIE THE EFFECTS OF HIV AND METH ABUSE AND THEIR DETRIMENTAL INTERACTIONS ON NEUROHIV DISEASE PROGRESSION, VIRUS EXPRESSION, AND VIRUS PERSISTENCE AND WILL INDICATE NOVEL THERAPEUTIC TARGETS FOR NEUROINFLAMMATION, NEURODEGENERATION, AND COMPULSIVITY TO TAKE METH. TO TEST THIS HYPOTHESIS, WE WILL USE A VALIDATED SYSTEMS BIOLOGY STRATEGY FOR THE RECONSTRUCTION AND INTERROGATION OF GENOME-WIDE GENE REGULATORY NETWORKS TO IDENTIFY THE GENE NETWORK DYSREGULATIONS ASSOCIATED WITH THE EFFECTS OF HIV, COMPULSIVE METH USE, AND THEIR INTERACTIONS AT SINGLE CELL RESOLUTION GENE PROFILING BY SINGLE NUCLEUS RNA-SEQ. OVERALL, THIS COLLABORATIVE INTERDISCIPLINARY PROPOSAL INTEGRATING SINGLE CELL LEVEL TRANSCRIPTOMICS, STATE-OF-THE-ART BEHAVIOR METHODS IN HIV TG AND WILD-TYPE RATS, AND COMPUTATIONAL STRATEGIES IS EXPECTED TO IDENTIFY NOVEL MECHANISTIC HYPOTHESES THAT MAY LEAD TO TRANSFORMATIVE NEW THERAPEUTIC CONCEPTS FOR SUBSTANCE USE DISORDER (SUD) IN THE HIV SETTING, AND WILL ESTABLISH KEY RESOURCES FOR THE NEUROHIV FIELD TO BE MADE PUBLICLY AVAILABLE THROUGH THE SCORCH DATA COORDINATION CENTER.

IMPACTING MITOCHONDRIA FUNCTION THROUGH ALTERED PROTEASE ACTIVITY

ROLE OF MECHANICALLY ACTIVATED ION CHANNELS IN SOMATOSENSATION

LACRIMAL GLAND REPAIR USING PROGENITOR CELLS

THE GENE WIKI: COMMUNITY INTELLIGENCE APPLIED TO GENE ANNOTATION

NOVEL HISTONE DEACETYLASE INHIBITORS AS THERAPEUTICS FOR HUNTINGTON'S DISEASE

CHEMOPROTEOMIC METHODS FOR SERINE HYDROLASE INHIBITOR DEVELOPMENT

AUTODOCK-FR: A MODULAR APPROACH TO FLEXIBLE RECEPTOR DOCKING

INFLUENZA VIRUS RECEPTORS ON HUMAN AIRWAY EPITHELIAL CELLS

DISCOVERY AND TREATMENT OF THE EARLY AUTOIMMUNE REACTION IN TYPE 1 DIABETES

C57BI/6 MOUSE LINES EXPRESSING CRE-RECOMBINASE IN THE NERVOUS SYSTEM

QUANTITATIVE PROTEOME ANALYSIS OF BRAIN DEVELOPMENT

USING A DISEASE-AFFECTED CELL TO SYNTHESIZE ITS OWN DRUG

FOXO TRANSCRIPTION FACTORS IN JOINT AGING AND OSTEOARTHRITIS PATHOGENESIS

INSTITUTIONAL CAREER DEVELOPMENT CORE

ASSEMBLY OF E. COLI RIBOSOMES IN VITRO AND IN CELLS

DYNAMICS OF THE BACTERIAL RIBOSOME AND PROTEOME

MODELING DRUGS OF ABUSE-HIV INTERACTIONS USING IPSC-DERIVED HUMAN CEREBRAL ORGANOIDS - SUMMARY HIV-INFECTED MICROGLIA RELEASE CYTOKINES, CHEMOKINES AND VIRAL GENE PRODUCTS, SUCH AS GP120 AND TAT, THAT ARE TOXIC TO NEURONS. IN THIS PROPOSAL, WE WILL USE HUMAN INDUCED PLURIPOTENT STEM CELLS (IPSCS) TO GENERATE CORTICAL AND DOPAMINERGIC NEURONS AS WELL AS MICROGLIA, WHICH WILL BE CULTURED AS ORGANOIDS TO MODEL HIV-DRUG ABUSE INTERACTIONS IN CONJUNCTION WITH IN-DEPTH GENOMIC APPROACHES AND ELECTROPHYSIOLOGY. WE SHOW ELECTROPHYSIOLOGICAL RESULTS THAT DEMONSTRATE THAT THE LEVELS OF DIFFERENTIATION OF THE NEURONS IN OUR ORGANOIDS ARE COMPARABLE TO IN VIVO AND EX VIVO PREPARATIONS AND ARE AMONG THE BEST THAT CAN BE FOUND IN THE SCIENTIFIC LITERATURE. TO IDENTIFY CANDIDATE HOST REGULATORS OF HIV EXPRESSION AND MEDIATORS OF HIV-INDUCED TISSUE DAMAGE AND DISEASE PROGRESSION, WE WILL EXPOSE MICROGLIA-CONTAINING CEREBRAL ORGANOIDS TO METHAMPHETAMINE (METH), A STIMULANT, AND MORPHINE, AN OPIATE, WHICH ARE MEMBERS OF TWO OF THE CLASSES OF DRUGS OF ABUSE THAT ARE MORE PREVALENT AMONG PEOPLE LIVING WITH HIV/AIDS (PLWHA). CEREBRAL ORGANOIDS WILL BE EXPOSED TO TOXIC HIV PRODUCTS SUCH AS TAT, GP120, DRUGS OF ABUSE AND COMBINATION ANTIRETROVIRAL THERAPY (CART). IN SOME EXPERIMENTS WE WILL ALSO INCORPORATE IPSC-DERIVED ASTROCYTES INTO MICROGLIA-CONTAINING CEREBRAL ORGANOIDS TO TEST THE ROLE OF ASTROCYTES IN NEUROPROTECTION AND NEURODEGENERATION. WE WILL CARRY OUT SINGLE-CELL GENE EXPRESSION PROFILING OF IPSC-DERIVED ORGANOIDS EXPOSED TO DRUGS OF ABUSE, HIV PRODUCTS, AND CART AND WE WILL EMPLOY AN ADVANCED SYSTEMS BIOLOGY STRATEGY TO GENERATE TESTABLE MECHANISTIC HYPOTHESES ON THE PATHOGENESIS OF NEURODEGENERATION AND TISSUE DAMAGE IN NEUROHIV (AIM 1) AND IDENTIFY CANDIDATE REGULATORS OF THE LTR PROMOTER THAT MAY SHED LIGHT ON THE REGULATION OF LATENCY AND REACTIVATION OF THE HIV PROVIRUS (AIM 2). IN PRELIMINARY STUDIES, THIS COMPUTATIONAL EXPERIMENTAL APPROACH ALLOWED US TO IDENTIFY CANDIDATE DRIVERS OF GENE EXPRESSION CHANGES ASSOCIATED WITH NEUROHIV AND NEURODEGENERATIVE DISEASES, INCLUDING ALZHEIMER’S DISEASE AND DRUG AND ALCOHOL ABUSE. THESE FINDINGS SUPPORT THE OVERARCHING HYPOTHESIS THAT DISSECTION OF THE GENE REGULATORY NETWORK OF THE CENTRAL NERVOUS SYSTEM WILL PAVE THE WAY FOR THE IDENTIFICATION OF NOVEL MECHANISTIC HYPOTHESES AND DRUGGABLE TARGETS TO IMPROVE NEUROPSYCHOLOGICAL FUNCTIONING OF PLWHA AND SUBSTANCE ABUSE COMORBIDITY.

LOWERING MITOCHONDRIAL ATP SYNTHASE ACTIVITY SLOWS AGING AND ALZHEIMER'S DISEASE

UNCLEAVED PREFUSION-OPTIMIZED TRIMERS ON NANOPARTICLES AS HIV VACCINES

STRUCTURAL AND MECHANISTIC BASIS FOR RNA SILENCING

QUALITY CONTROL IN RIBOSOME ASSEMBLY - THE FUNCTION OF REGULATORY PROTEINS

DESIGNING AND OPTIMIZING CANDIDATE HIV VACCINES AND BOOSTING PROTOCOLS

A SAFETY SWITCH FOR AN EFFECTIVE HIV-1 VACCINE

GLYCAN DEPENDENT EPITOPES OF HIV BROADLY NEUTRALIZING ANTIBODIES

MOLECULAR AND CELLULAR MECHANISMS OF THE LYSOSOMAL STORAGE DISEASE CYSTINOSIS

ENGINEER IMMUNE CELLS VIA CHEMOENZYMATIC GLYCAN EDITING

IDENTIFICATION AND THERAPEUTIC VALIATION OF ADIPOCYTE SERINE HYDROLASES OF METAB

REGULATION OF SIALOSIDE EXPRESSION IN LEUKOCYTES

AUTOMATED, OPTIMIZED, INTELLIGENT DATA COLLECTION FOR CRYO-EM - PROJECT SUMMARY CRYO-ELECTRON MICROSCOPY (CRYO-EM) IS NOW A WIDELY ESTABLISHED AND INDISPENSABLE METHOD FOR DETERMINING THE HIGH-RESOLUTION STRUCTURES OF BIOMEDICALLY IMPORTANT MOLECULES. GIVEN THAT THOUSANDS OF IMAGES, OFTEN ACQUIRED OVER THE COURSE OF SEVERAL DAYS, ARE REQUIRED TO OBTAIN SUCH STRUCTURES, AUTOMATION SOFTWARE HAS PLAYED A CRITICAL ROLE IN THE LARGE-SCALE ADOPTION OF THIS METHOD BY THE SCIENTIFIC COMMUNITY. IN JUST THE PAST FIVE YEARS, CRYO-EM HAS REVOLUTIONIZED OUR UNDERSTANDING OF ENTIRE BIOLOGICAL SYSTEMS, AND IN 2020 PROVIDED THE FIRST MOLECULAR DESCRIPTIONS OF SARS-COV-2 INTERACTION WITH NEUTRALIZING ANTIBODIES. THE WIDESPREAD ADOPTION OF CRYO-EM RECENTLY PROMPTED THE NIH TO INVEST IN THREE NATIONAL CENTERS THROUGH THE TRANSFORMATIVE HIGH RESOLUTION CRYO- ELECTRON MICROSCOPY PROGRAM, PROVIDING FREE, HIGH-END ELECTRON MICROSCOPE ACCESS TO BIOLOGISTS ACROSS THE COUNTRY. THE EXPONENTIAL INCREASE IN THE POPULARITY OF CRYO-EM HAS LED TO AN ASTONISHING NUMBER OF DEVELOPMENTS IN SAMPLE PREPARATION METHODOLOGIES AND IMAGE PROCESSING ALGORITHMS, WHICH HAVE IMPROVED ATTAINABLE RESOLUTION OF SINGLE PARTICLE RECONSTRUCTIONS. HOWEVER, COMPARATIVELY LITTLE PROGRESS HAS BEEN MADE IN OPTIMIZING THE QUALITY OF THE CRYO-EM DATA BEING COLLECTED. THE PIONEERING SOFTWARE PACKAGES LEGINON AND APPION DEMONSTRATED THE POWER OF AUTOMATED DATA ACQUISITION AND REAL-TIME PROCESSING (RESPECTIVELY), AND THERE ARE NOW NUMEROUS PROGRAMS FOR AUTOMATED DATA ACQUISITION AND REAL-TIME PROCESSING. DESPITE ADVANCES IN AUTOMATION, OPTIMALLY EXTRACTING THE HIGHEST QUALITY DATA FROM AN EM SAMPLE STILL REQUIRES MANUAL INVOLVEMENT OF AN EXPERT ELECTRON MICROSCOPIST. USER INTERVENTION AND EXPERTISE IS NECESSARY TO RUN THE APPROPRIATE IMAGE ANALYSES, INTERPRET THE RESULTS, AND MAKE INFORMED DECISIONS ON HOW THE PROCESSED RESULTS RELATE TO THE ONGOING DATA COLLECTION. HOWEVER, EVEN EXPERTS MUST CONTENT WITH THE FACT THAT THE “BEST GRID REGIONS” DIFFER DRASTICALLY FROM SAMPLE TO SAMPLE, AND THERE ARE NO ESTABLISHED TOOLS FOR AUTOMATICALLY AND QUICKLY ASSESSING THE QUALITY OF THE SPECIMEN ACROSS THE VARIOUS MICROENVIRONMENTS OF AN EM GRID. GIVEN THE EVER-INCREASING INCORPORATION OF CRYO-EM INTO LABS’ RESEARCH PROGRAMS, IT IS IMPERATIVE THAT DATA COLLECTION AND PROCESSING BE STREAMLINED TO MATCH THE GROWING NEEDS OF THE STRUCTURAL COMMUNITY. WE PROPOSE TO DEVELOP A SECOND GENERATION LEGINON/APPION SOFTWARE PACKAGE, “MAGELLON”, TO OVERCOME EXISTING BOTTLENECKS AND PROVIDE AN AVENUE TOWARD FULLY AUTOMATED DATA ACQUISITION THAT BYPASSES NEED FOR USER INPUT DURING DATA COLLECTION. IMPORTANTLY, THIS SOFTWARE WILL SUPPORT THE COMPUTATIONAL INFRASTRUCTURE TO ENABLE REAL-TIME IMAGE PROCESSING RESULTS TO INFORM ON AND MODIFY THE ONGOING DATA COLLECTION REGIME BY LEARNING WHERE TO ACQUIRE IMAGES IN REGIONS THAT WILL YIELD THE HIGHEST RESOLUTION STRUCTURES. WE WILL DEVELOP AND INCORPORATE NEW, FAST IMAGE ASSESSMENT ROUTINES, WHILE ALSO PROVIDING AN APPLICATION PROGRAMMING INTERFACE TO ENABLE THE INCORPORATION OF EXTENSIONS AND PLUGINS FROM DEVELOPERS IN THE COMMUNITY. FURTHER, MAGELLON WILL ENABLE STRAIGHTFORWARD, SEAMLESS IMPORT AND EXPORT OF DATA FROM ITS DATABASE TO ACCOMMODATE REMOTE DATA ACQUISITION AT ANY OF THE REGIONAL OR NATIONAL CRYO-EM CENTERS.

IMPACTING CELL GROWTH THROUGH ALTERED CIRCADIAN PROTEOLYSIS

DEVELOPING RELIABLE IN SILICO METHODS TO DESIGN SMALL MOLECULES TARGETING RNA

RESOURCE FOR STRUCTURE-BASED COMPUTATIONAL DRUG DISCOVERY AND DESIGN (RSD3) - PROJECT SUMMARY AUTOMATED DOCKING IS A COMPUTATIONAL METHOD THAT IS NOW ROUTINELY USED FOR IDENTIFYING SMALL MOLECULES THAT HAVE THE POTENTIAL TO BE NEW DRUGS. OVER THE PAST THREE DECADES WE HAVE DEVELOPED DOCKING METHODS AND OUR DOCKING SOFTWARE AUTODOCK IS NOT THE MOST CITED DOCKING SOFTWARE WITH OVER 40,000 CITATIONS IN THE PEER REVIEWED LITERATURE. AS SUCH, IT HAS BECOME AN IMPORTANT RESOURCE FOR THE COMMUNITY. WE SEEK TO CREATE A NATIONAL RESOURCE THAT WILL ALLOW US TO MAINTAIN AND MODERNIZE THIS SOFTWARE TO ADAPT TO EVOLVING HARDWARE PLATFORMS AND OPERATING SYSTEMS, TO KEEP THE SOFTWARE UP TO DATE AND RELEVANT BY INCORPORATING THE LATEST ALGORITHMIC DEVELOPMENTS, AND TO SUPPORT ITS LARGE USER COMMUNITY. IN PARALLEL WITH THESE EFFORTS, AN IMPORTANT GOAL OF THE PROPOSED RESOURCE IS TO CHART A PATH TOWARD A SELF-SUSTAINED SOFTWARE ECOSYSTEM WHERE CONTRIBUTORS FROM AROUND THE WORLD WILL CONTRIBUTE TO MAINTAIN AND FURTHER DEVELOP THIS SOFTWARE AFTER THE LIFETIME OF THIS AWARD. THIS GOAL HAS BEEN ACHIEVED BY OTHER SUCCESSFUL OPEN SOURCE PROJECTS SUCH AS DEBIAN AND PYTHON, AND THE WRITTEN INTEREST OF MANY COLLEAGUES BOLSTER OUR CONFIDENCE THAT THE AUTODOCK SOFTWARE TOO CAN REACH THIS GOAL. TO THIS END WE PROPOSE THREE SPECIFIC AIMS. OUR FIRST AIM IS TO MAINTAIN AND MODERNIZE THE SOFTWARE CODE. THIS CRITICAL WORK IS NEEDED FOR THE SOFTWARE TO REMAIN FUNCTIONAL AND ABLE TO ADDRESS THE EVOLVING NEEDS OF THE COMMUNITY. AS WE OVERHAUL THE SOFTWARE, WE WILL LEVERAGE NEWER TOOLKITS FOR GENERATING MODERN AND INTUITIVE GRAPHICAL USER INTERFACES. THESE INTERFACES ENABLE RESEARCHERS, SUCH AS CLINICAL PHYSICIANS OR CHEMISTS WITH LIMITED COMPUTATIONAL SKILLS FOR INSTANCE, TO USE DOCKING TO BETTER UNDERSTAND THE MECHANISM OF ACTION OF A DRUG AND OPTIMIZE IT. OUR SECOND AIM IS CONCERNED WITH MAKING OUR SOFTWARE TOOLS INTEROPERABLE WITH OTHER IMPORTANT MODELING SOFTWARE TOOLS SUCH AS MOLECULAR DYNAMICS FOR INSTANCE. NOT ONLY DOES THIS AUGMENT THE POTENTIAL OF DOCKING TO LEAD TO NOVEL THERAPEUTIC MOLECULES, BUT IT ALSO SERVES OUR GOAL TO CREATE A COMMUNITY OF DEVELOPERS WHO WILL ULTIMATELY MAINTAIN THIS SOFTWARE ECOSYSTEM. FINALLY OUR THIRD AND FINAL AIM IS ABOUT SUPPORTING THE LARGE USER COMMUNITY, GROWING IT AND ENSURING THAT THE SOFTWARE IS EASILY DISCOVERED, OBTAINED AND INSTALLED. WE HAVE A LONG TRACK RECORD OF DEVELOPING OPEN SOURCE SOFTWARE PROMOTING BEST PRACTICES IN SOFTWARE ENGINEERING, AND MAKING THESE TOOLS USABLE, USEFUL, AND AVAILABLE TO THE COMMUNITY. LIKEWISE WE HAVE SUPPORTED AND GROWN OUR USER COMMUNITY FOR MANY YEARS. THIS PUTS US IN A UNIQUE POSITION TO CREATE THE PROPOSED RESEARCH WHICH, IF FUNDED, WILL ALLOW US TO CONVERT THIS VALUABLE SOFTWARE CODE INTO A COMMUNITY SUPPORTED SOFTWARE ECOSYSTEM ENSURING THAT THIS SOFTWARE WILL CONTINUE TO SUPPORT THE DESIGN OF NOVEL THERAPEUTIC MOLECULES BEYOND THE LIFETIME OF THIS AWARD.

ELICITATION OF HIV BROADLY NEUTRALIZING ANTIBODIES FROM ENGINEERED B CELLS - PROJECT SUMMARY/ABSTRACT HIV BROADLY NEUTRALIZING ANTIBODIES (BNABS) ARE EFFECTIVE AGAINST THE VIRUS, BUT CANNOT BE ELICITED THROUGH VACCINATION DUE TO GENETIC LIMITATIONS IMPOSED BY THE HUMAN REPERTOIRE OF B CELL ANTIGEN-RECEPTORS (BCRS). WE HAVE RECENTLY SHOWN THAT MATURE PRIMARY B CELLS FROM WILD-TYPE MICE CAN BE ENGINEERED EX VIVO TO EXPRESS BNAB GENES AS FUNCTIONAL ANTIGEN RECEPTORS, AND THAT THESE CELLS CAN BE RETURNED TO THE HOST AND VACCINATED TO GENERATE DURABLE BNAB RESPONSES. BECAUSE THIS ANIMAL MODEL CANNOT SUPPORT HIV INFECTION, THERAPEUTIC OR PROTECTIVE EFFICACY OF THESE RESPONSES HAVE NOT YET BEEN EVALUATED. THE PROPOSAL DESCRIBED HERE: 1) EXPLORES NOVEL B CELL TARGETING APPROACHES THAT AIM TO IMPROVE HOW B CELLS ARE ENGINEERED AND FUNCTION IN VIVO; 2) DEFINES VACCINE PARAMETERS AND THE PROPERTIES OF ENGINEERED CELLS REQUIRED FOR REPRODUCIBLE ELICITATION OF DURABLE BNAB RESPONSES IN THE MOUSE MODEL AND; 3) TRANSLATES OUR SUCCESSFUL ‘ENGINEERED B CELL VACCINE’ (EBCV) PROTOTYPE TO THE RHESUS MACAQUE ANIMAL MODEL FOR EFFICACY TESTING USING CHIMERIC HIV/SIV VIRUSES (SHIVS). BNABS ELICITED IN NON-HUMAN PRIMATES (NHP) WILL BE TESTED FOR THEIR ABILITY TO PREVENT HETEROLOGOUS TIER-2 VIRUS INFECTION, SUPPRESS, OR MAINTAIN SUPPRESSION OF VIREMIA, IN ORDER TO EVALUATE THE POTENTIAL FOR EBCVS TO FUNCTION AS PROPHYLACTIC VACCINES OR AS AN HIV FUNCTIONAL CURE. SUCH AN APPROACH MAY HAVE SIGNIFICANT ADVANTAGES OVER OTHER GENE THERAPY STRATEGIES THAT AIM TO DURABLY SECRETE BNABS FROM MUSCLE OR LIVER CELLS. WHEN ELICITED FROM MODIFIED B CELLS, BNAB RESPONSES SHOULD BE: 1) ABLE TO MATURE IN AFFINITY IN RESPONSE TO A RAPIDLY EVOLVING PATHOGEN; 2) INCREASE TITERS IN THE PRESENCE OF ANTIGEN; 3) BE EXPRESSED AS ALL EFFECTOR ISOTYPES; 4) TOLERATED BY THE IMMUNE SYSTEM BECAUSE THEY ORIGINATE FROM B CELLS AND; 5) SUBJECT TO TOLERANCE MECHANISMS THAT SHOULD INACTIVATE THE CELLS EXPRESSING THEM IF THEY ARE HARMFUL. ALL STUDIES DESIGNED IN THIS PROPOSAL SUPPORT OUR LONG-TERM GOAL, THE DEVELOPMENT OF SAFE, EFFECTIVE AND ECONOMICALLY FEASIBLE ‘ENGINEERED B CELL VACCINES’ THAT WOULD GENERATE DURABLE HIV BNAB RESPONSES FROM GENOME MODIFIED B CELLS AS A STRATEGY FOR PREVENTION OR HIV FUNCTIONAL CURE.

DISCOVERY OF POTENT HUMAN ANTIBODIES FOR PREVENTION OF (RECURRENT) HCV INFECTION

FUNCTIONAL COOPERATION OF TISSUE FACTOR AND INTEGRINS

CATALYST-CONTROLLED, SITE-SELECTIVE C-H FUNCTIONALIZATION OF HETEROCYCLES

COOPERATIVE CENTER FOR THE STUDY OF HCV ANTIBODY RESPONSES AND VACCINE ANTIGENS

CHEMICAL PROBES FOR METABOLIC PATHWAY DISCOVERY IN HUMAN DISEASE

MRE11/RAD50 STRUCTURAL BIOLOGY FOR DNA DAMAGE RESPONSES

ROLES OF THE SMC5-SMC6 HOLOCOMPLEX IN GENOME STABILITY

TRANSMEMBRANE PROTEINS INVOLVED IN HUMAN TUMOR EXPANSION

TRANSCRIPTIONAL REGULATION OF CHONDROGENESIS

SYSTEMS BIOLOGY OF HIV, METHAMPHETAMINE AND ANTIRETROVIRALS INTERACTIONS

THE GENERATION AND PROTECTIVE FUNCTION OF LUNG TISSUE RESIDENT MEMORY T CELLS FOLLOWING SARS-COV-2 INFECTION OR VACCINATION - PROJECT SUMMARY/ABSTRACT CORONAVIRUS DISEASE 2019 (COVID-19), CAUSED BY SEVERE ACUTE RESPIRATORY SYNDROME CORONAVIRUS 2 (SARS- COV-2), HAS EXPLODED INTO A GLOBAL PANDEMIC CAUSING SIGNIFICANT LOSS OF LIFE, PRONOUNCED ECONOMIC IMPACT AND SIGNIFICANT LONG-TERM MEDICAL IMPACTS WHICH ARE STILL BEING CHARACTERIZED. DESPITE IMPRESSIVE PROTECTION AFFORDED BY THE CURRENT SARS-COV-2 VACCINES WHICH ARE PRIMARILY MEDIATED BY ANTIBODY NEUTRALIZATION OF VIRUS, THESE ANTIBODIES WANE OVER TIME AND NEW VIRAL VARIANTS HAVE EMERGED THAT ARE LESS SUSCEPTIBLE TOBY THESE ANTIBODIES. A MAJOR QUESTION IS WHETHER MEMORY T CELLS AFFORD MORE LONG-LASTING PROTECTION DURING SARS-COV-2 INFECTION. PRIOR STUDIES SHOWED THAT PEOPLE WHO RECOVERED FROM SARS-COV INFECTION FROM 2003 EXHIBITED SARS-COV- SPECIFIC MEMORY CD8+ T CELL RESPONSES IN PERIPHERAL BLOOD FOR UP TO 11 YEARS, VIRUS-SPECIFIC ANTIBODIES WERE NOT DETECTABLE AT 6 YEARS, AND THERE WAS NO CROSS-REACTIVITY WITH MERS-COV PEPTIDE1,2. FOR THE CURRENT PANDEMIC, WE ARE STARTING TO LEARN WHAT TYPES OF MEMORY T AND B CELLS FORM AFTER SARS-COV2 INFECTION IN THE BLOOD, HOWEVER WE HAVE NO IDEA THE LONGEVITY OF THESE RESPONSES. MORE IMPORTANTLY, WE KNOW VERY LITTLE ABOUT THE PULMONARY MEMORY T AND B CELLS THAT FORM IN THE LUNG AFTER SARS-COV-2 INFECTION AND, BASED ON WHAT HAS BEEN LEARNED FROM OTHER VIRUSES, THESE MAY BE THE MOST PROTECTIVE MEMORY CELLS NEEDED FOR SUPERIOR LONG- TERM IMMUNITY TO REINFECTION. IN THIS COLLABORATIVE PROPOSAL BETWEEN THE TEIJARO, KAECH AND FARBER LABS, WE BRING TOGETHER WORLD- CLASS EXPERTISE IN ANTI-VIRAL IMMUNITY AND LUNG PATHOGENESIS, IMMUNOLOGICAL MEMORY AND HUMAN IMMUNOLOGY TO STUDY THE DEVELOPMENT AND PROTECTIVE ROLE OF LUNG-RESIDENT TRM CELLS IN SARS-COV-2. IN AIM 1, WE WILL STUDY THE FUNDAMENTALS OF SARS-COV-2 TRM DEVELOPMENT IN THE LUNGS OF HACE2-TRANSGENIC MICE AND DETERMINE WHETHER TRM ARE REQUIRED FOR LONG- TERM IMMUNITY, THROUGH GENETIC PERTURBATIONS OF TGFBR2 AND SMAD4 IN T CELLS, WHICH ARE DIFFERENTIALLY REQUIRED FOR TRM DIFFERENTIATION3, AND THE DEPLETION OF CIRCULATING MEMORY CELLS. IN AIM 2, WE WILL EXAMINE IF LUNG TRM CELLS CONTRIBUTE TO THE PROTECTION AFFORDED BY MRNA VACCINES TO BETTER UNDERSTAND THE ROLE, IF ANY, OF MEMORY T CELLS IN MEDIATING PROTECTION TO SARS-COV-2 AND NOTABLE VARIANTS OF CONCERN (VOC). LASTLY, IN AIM 3, WE EXTEND AND COMPLEMENT OUR STUDIES IN MICE TO DR. FARBER’S HUMAN DONOR REPOSITORY TO ASSESS HUMAN MEMORY FORMATION FOLLOWING SARS-COV-2 INFECTION AND VACCINATION. WE HOPE TO ANSWER THE BASIC QUESTION OF WHICH TYPES OF MEMORY T CELLS FORM AFTER SARS- COV-2 INFECTION AND CONFER LONG-TERM PROTECTIVE IMMUNITY TO THIS VIRUS AND VOCS. THESE STUDIES WILL PROVIDE CRITICAL INFORMATION RELATED TO THE QUALITY OF IMMUNOLOGICAL MEMORY THAT FORMS AFTER SARS-COV- 2 INFECTION IN MICE AND HUMANS AND WILL SERVE TO GUIDE CURRENT AND FUTURE VACCINE DEVELOPMENT.

INNATE IMMUNE SURVEILLANCE OF HIV-1 DURING TRANSMISSION AND SYSTEMIC INFECTION

COMPOUND REPOSITIONING FOR ALZHEIMER'S DISEASE USING KNOWLEDGE GRAPHS, INSURANCE CLAIMS DATA, AND GENE EXPRESSION COMPLEMENTARITY

STUDIES ON PROTEIN LIPIDATION

ESTABLISHING STRATEGIES TO AMELIORATE AMYLOID PATHOLOGY IN LIGHT CHAIN AMYLOIDOSIS

IMMUNOPHARMACOTHERAPY FOR MITIGATING OPIOID ADDICTION

FUSION PEPTIDE DIRECTED IMMUNOGENS THAT ELICIT NEUTRALIZING AND PROTECTIVE ANTIBODIES IN NON-HUMAN PRIMATES

THERAPEUTIC USE OF AN ENHANCED FORM OF CD4-IG

OMICS ANALYSES OF HIV AND SUBSTANCE USE DISORDER

HUMAN ANTIBODIES TO HIV-1 BY REPERTOIRE CLONING

MODELING AND INTERACTING WITH THE VISIBLE MOLECULAR CELL

EFFECTS OF ADOLESCENT ALCOHOL EXPOSURE ON SLEEP AND AROUSAL IN ADULTHOOD

IDENTIFICATION OF SMALL MOLECULES FOR NEUROLOGICAL COMPLICATIONS OF HIV AND SUBSTANCE ABUSE COMORBIDITY

STUDY THE MECHANISMS UNDERLYING COMMON FRAGILE SITE PROTECTION

MECHANISMS OF MUTANT PRION PROTEIN AGGREGATION WITHIN ENDOLYSOSOMAL PATHWAYS - ULTRASTRUCTURAL STUDIES OF HUMAN PATIENT BRAINS WITH PRION DISEASE HAVE REVEALED THE ACCUMULATION OF MISFOLDED PRP WITHIN DYSTROPHIC NEURITES INSIDE ENDOLYSOSOMES. OTHER COMPLEX STRUCTURES CO- EXIST WITHIN THESE DYSTROPHIES AND ARE NOW CONSIDERED COMMON FEATURES OF PRION PATHOLOGIES, INCLUDING AUTOPHAGIC VACUOLE-LIKE MEMBRANE-BOUND ORGANELLES, LYSOSOMAL ELECTRON-DENSE BODIES, AND ENLARGED ENDOLYSOSOMES. THE MECHANISMS LEADING TO THE FORMATION OF THESE STRUCTURES REMAIN UNKNOWN. OUR STUDIES HAVE IDENTIFIED AN ENDOLYSOSOMAL PATHWAY IN MAMMALIAN NEURONS THAT WE CALL AXONAL RAPID ENDOSOMAL SORTING AND TRANSPORT-DEPENDENT AGGREGATION (ARESTA), THAT DRIVES THE FORMATION OF NEUROTOXIC AXONAL AGGREGATES OF A MISFOLDED MUTANT PRION PROTEIN (PRP) INSIDE ENDO-MEMBRANE STRUCTURES THAT WE CALL “ENDOGGRESOMES”. THE LONG-TERM GOAL OF THIS PROPOSAL IS TO CHARACTERIZE THE ENDOCYTIC PATHWAYS THAT PLAY A ROLE IN THE PATHOPHYSIOLOGY OF PRION DISEASES, ALZHEIMER’S DISEASE, AND OF ALZHEIMER’S DISEASE RELATED DEMENTIAS, THAT WILL PROVIDE ACTIONABLE TARGETS FOR THEIR PHARMACOLOGICAL TREATMENT. THE OBJECTIVES OF THIS PROPOSAL ARE (I) TO DETERMINE THE GENERALITY OF THE ARESTA PATHWAY IN FORMATION OF ENDOGGRESOMES IN AXONS OF NEURONS EXPRESSING VARIOUS FAMILIAL PRP MUTATIONS, AND TO CHARACTERIZE THE MOLECULAR AND ULTRASTRUCTURAL ARCHITECTURE OF NEUROTOXIC ENDOGGRESOMES; (II) TO DETERMINE THE MECHANISMS OF MUTANT PRP ENDOGGRESOME-MEDIATED AXONAL IMPAIRMENTS; AND (III) TO DETERMINE HOW THE ENDOLYSOSOMAL ARESTA PATHWAY MODULATES THE FORMATION OF AXONAL MUTANT PRP AGGREGATES IN VIVO. THE CENTRAL HYPOTHESES ARE (I) ARESTA DRIVES THE FORMATION OF ENDOGGRESOMES IN VARIOUS FAMILIAL PRION DISEASES BY INTERACTIONS WITH CO-FACTORS WITHIN ENDOCYTIC ROUTES; (II) MUTANT PRP ENDOGGRESOME-INDUCED PATHOLOGIES ACT AS AXONOTOXICITY HUBS THAT INHIBIT NEURONAL FUNCTION BY IMPAIRING LOCAL AXONAL CYTOSKELETAL-ORGANELLE INTERACTIONS, AND (III) ENDOLYSOSOMAL PATHWAYS MODULATE THE FORMATION AND PATHOLOGY OF MUTANT PRP AGGREGATES IN VIVO. THE PROPOSED RESEARCH IS INNOVATIVE BECAUSE IT PROVIDES A CONCEPTUAL FRAMEWORK FOR DEVELOPING MODELS THAT INCLUDE NOVEL ENDOLYSOSOMAL PATHWAY-MEDIATED MECHANISMS TO EXPLAIN HOW INTRA-AXONAL AGGREGATES FORM AND IMPAIR NEURONAL FUNCTION IN THE PRIONOPATHIES. THE PROPOSED RESEARCH IS SIGNIFICANT BECAUSE IT IDENTIFIES THE ENDOCYTIC PATHWAY AND SPECIFICALLY ARESTA AND ENDOGGRESOMES, AS ANTI- AGGREGATION TARGETS FOR THERAPIES TO INHIBIT AGGREGATE FORMATION AND REVERSE RELATED PATHOLOGIES. AS AMYLOID-B PEPTIDES, TAU, AND MOST PROTEINS THAT MISFOLD IN NEURODEGENERATION TRANSIT WITHIN ENDOCYTIC ROUTES AT SOME POINT IN THEIR PROCESSING ROUTES, OUR FINDINGS ARE EXPECTED TO BE RELEVANT TO ALZHEIMER’S DISEASE AND ALZHEIMER’S DISEASE RELATED DEMENTIAS.

ALPHA-V INTEGINS AND ADENOVIRUS CELL ENTRY

PATHOGENESIS OF PERSISTENT VIRAL INFECTION

MECHANISMS OF GENE SILENCING IN FRIEDREICH'S ATAXIA

DYNAMIC INTERACTIONS OF THE S-NITROSOPROTEOME IN TYPE 2 DIABETES/METABOLIC SYNDROME AND ALZHEIMER?S DISEASE

IMMUNOPHARMACOTHERAPY FOR THE TREATMENT OF COCAINE ABUSE

HIV-1 VACCINE DESIGN EMPHASIZING BNAB TARGETS ON MEMBRANE ENV LIPOSOMES

EXPLORING SIGLEC-GLYCAN LIGAND INTERACTIONS USING CHEMOENZYMATIC APPROACHES - PROJECT SUMMARY/ABSTRACT REGULATION OF THE IMMUNE SYSTEM IS SUBSTANTIALLY INFLUENCED BY GLYCOSYLATION. CELL-SURFACE GLYCANS TUNE LIGAND-RECEPTOR BINDING AND SET A THRESHOLD FOR INITIATING THE DOWNSTREAM SIGNALING FOR IMMUNE CELL ACTIVATION. SIGLECS (SIALIC ACID-BINDING IMMUNOGLOBULIN-TYPE LECTINS) ARE A FAMILY OF REGULATORY RECEPTORS INVOLVED IN THESE PROCESSES. A SIGLEC CAN BIND TO BOTH CIS AND TRANS SIALYLATED GLYCAN LIGANDS THAT ARE EXPRESSED ON THE SAME OR INTERACTING CELLS, RESPECTIVELY. THE BINDING OF SIGLECS WITH THEIR LIGANDS CAN EITHER SEGREGATE SIGLECS FROM ACTIVATION RECEPTORS OR MOVE THEM CLOSER. THE ABILITY OF INHIBITORY SIGLECS TO MODULATE ACTIVATION RECEPTORS IS REGULATED BY SPATIAL PROXIMITY: RECRUITING SIGLECS TO THE IMMUNE SYNAPSE IN THE PROXIMITY OF ACTIVATION RECEPTORS WOULD TRIGGER INHIBITORY SIGNALING TO SUPPRESS IMMUNE-SYSTEM ACTIVATION, WHEREAS MOVING SIGLECS AWAY FROM ACTIVATION RECEPTORS WOULD ENABLE OPTIMAL SIGNALING THROUGH ACTIVATION RECEPTORS. FOR THE ABOVE REASONS, RECENTLY, SIGLECS HAVE BEEN DESCRIBED AS GLYCO-IMMUNE CHECKPOINTS. THROUGH THEIR INTERACTION WITH SIALYLATED GLYCANS ABERRANTLY EXPRESSED ON TUMOR CELLS, INNATE IMMUNE CELL-ASSOCIATED SIGLECS TRIGGER SIGNALING CASCADES TO INHIBIT IMMUNE-SYSTEM ACTIVATION. LIKEWISE, SIGLECS UPREGULATED ON TUMOR CELLS INTERACT WITH YET-TO-BE IDENTIFIED T-CELL MEMBRANE GLYCOPROTEINS TO SUPPRESS T CELL ANTI-TUMOR FUNCTIONS. ON THE POSITIVE SIDE, HOWEVER, INHIBITORY SIGNALING THROUGH SIGLECS CURBS INFLAMMATION DURING CELL DEATH INDUCED BY VIRAL INFECTION. DESPITE THESE INTRIGUING OBSERVATIONS, THE MECHANISMS UNDERLYING THE ABOVE PROCESSES ARE JUST STARTING TO BE ELUCIDATED. THE OVERARCHING GOAL OF THIS PROJECT IS TO USE A COMBINATION OF CHEMOENZYMATIC, BIOCHEMICAL AND GENETIC TOOLS TO EXPLORE SIGLEC-GLYCAN LIGAND INTERACTIONS AND THEIR THERAPEUTIC IMPLICATION. IN AIM 1, WE WILL DESIGN SIGLEC-BASED CHIMERIC SWITCH RECEPTORS AND CONVERT INHIBITORY SIGLECS INTO ACTIVATION RECEPTORS. IN AIM 2, WE WILL USE A CELL-BASED GLYCAN ARRAY PLATFORM TO SCREEN FOR HIGH-AFFINITY AND SPECIFIC LIGANDS OF SIGLECS. ONCE IDENTIFIED, WE WILL EXPLORE THEIR UTILITIES TO SUPPRESS OR HARNESS THE INHIBITORY SIGLEC SIGNALING FOR THERAPEUTIC APPLICATIONS. FINALLY, WE WILL USE OUR CHEMOENZYMATIC TOOLS TO INVESTIGATE HOW THE SIGLEC-CIS LIGAND INTERACTION IS INVOLVED IN MEDIATING THE SIGLEC–TRANS LIGAND INTERACTION AND ACCORDINGLY IMMUNE CELL ACTIVATION (AIM 3).

MECHANISMS OF PROTECTION OF UNIVERSAL THERAPEUTIC ANTIBODIES TO INFLUENZA A

SINGLE NUCLEUS GENE EXPRESSION IN MODERATE AND COMPULSIVE OPIOID SELF-ADMINISTRATION IN A RODENT MODEL OF HIV - SUMMARY THE ABUSE OF OPIOID DRUGS IS ASSOCIATED WITH TREATMENT NON-COMPLIANCE, GREATER RISK OF VIRAL TRANSMISSION, AND MORE RAPID CLINICAL PROGRESSION OF HIV DISEASE. THE OVERARCHING HYPOTHESIS BEHIND THE PRESENT PROJECT IS THAT THE ANALYSIS OF MOLECULAR PROFILES OF NEURONAL AND GLIA CELLS AT THE SINGLE CELL LEVEL IN DRUG ABUSE-RELEVANT BRAIN REGIONS BY SINGLE NUCLEUS RNA-SEQ (SNRNA-SEQ) WILL REVEAL KEY GENES THAT ARE DYSREGULATED BY THE INTERACTION OF HIV WITH OPIOID ABUSE, RESULTING IN NEURODEGENERATION AND COGNITIVE IMPAIRMENT. TO TEST THE PRESENT HYPOTHESIS, WE PROPOSE TO USE VALIDATED SYSTEMS BIOLOGY STRATEGIES FOR THE RECONSTRUCTION AND INTERROGATION OF A GENOME-SCALE INTEGRATED GENE REGULATORY NETWORK IN CONJUNCTION WITH SNRNA-SEQ FROM HIV TRANSGENIC (TG) RATS, WHICH HARBOR A NON-REPLICATING HIV-1 TRANSGENE EXPRESSING CHRONIC LOW-LEVELS OF MULTIPLE HIV-1 PROTEINS IN DISEASE-RELEVANT CELL TYPES, AND WILD-TYPE RATS. THE OCCASIONAL BUT LIMITED USE OF A DRUG IS CLINICALLY DISTINCT FROM DEPENDENT DRUG USE, WHICH IS CHARACTERIZED BY THE EMERGENCE OF DEPENDENCE AND A NEGATIVE EMOTIONAL STATE WHEN ACCESS TO THE DRUG IS PREVENTED THAT DRIVES NEGATIVE REINFORCEMENT, A POWERFUL SOURCE OF MOTIVATION FOR DRUG SEEKING. THEREFORE, WE WILL USE A STATE-OF-THE-ART PARADIGM OF VOLUNTARY INTRAVENOUS OPIOID SELF-ADMINISTRATION UNDER SHORT ACCESS (SHA) CONDITIONS, WHICH IS CHARACTERIZED BY A NON-DEPENDENT, “RECREATIONAL” PATTERN OF DRUG USE, AND LONG ACCESS (LGA) CONDITIONS, WHICH LEADS TO DEPENDENT DRUG INTAKE. ESCALATED DRUG INTAKE UNDER LGA CONDITIONS IS HIGHLY RELEVANT TO HUMAN SUBSTANCE USE DISORDER (SUD) AS IT HAS BEEN SUGGESTED THAT IT MODELS ALL 7 OF THE CRITERIA FOR DRUG ADDICTION IN THE DIAGNOSTIC AND STATISTICAL MANUAL OF MENTAL DISORDERS (DSM)-IV AND 7 OF THE 11 CRITERIA IN THE DSM-V. WE SHOWED THAT HIV TG RATS SELF-ADMINISTERING OXYCODONE IN THIS LGA PARADIGM OF ESCALATED SELF-ADMINISTRATION DISPLAY INCREASED NEURAL INJURY AND COGNITIVE IMPAIRMENT. THE PROJECT WILL ADDRESS THE FOLLOWING VEXING QUESTION ABOUT OPIOID ABUSE IN THE SETTING OF HIV INFECTION: WHAT ARE THE CELL TYPES AND CELL STATES THAT DRIVE NEUROINFLAMMATION, NEURODEGENERATION, VIRUS EXPRESSION, AND ESCALATED (DEPENDENT) OPIOID SELF-ADMINISTRATION AND COGNITIVE IMPAIRMENT IN THE SETTING OF HIV? OVERALL, THIS COLLABORATIVE INTERDISCIPLINARY PROPOSAL INTEGRATING SINGLE CELL LEVEL TRANSCRIPTOMICS, STATE-OF-THE- ART BEHAVIOR METHODS IN HIV TG AND WILD-TYPE RATS, AND COMPUTATIONAL STRATEGIES FOR THE DECONVOLUTION OF THE GENE REGULATORY NETWORK AT THE SINGLE CELL LEVEL WILL ELUCIDATE KEY MECHANISMS THAT UNDERLIE THE EFFECTS OF HIV AND OPIOID ABUSE AND THEIR DETRIMENTAL INTERACTIONS ON NEUROHIV PROGRESSION, VIRUS EXPRESSION AND PERSISTENCE. THE RESULTS WILL INDICATE TRANSFORMATIVE NEW MECHANISTIC HYPOTHESES THAT MAY LEAD TO NOVEL THERAPEUTIC CONCEPTS FOR OPIOID USE DISORDER (OUD) IN THE SETTING OF HIV AND WILL ESTABLISH KEY RESOURCES FOR THE NEUROHIV FIELD TO BE MADE PUBLICLY AVAILABLE THROUGH THE SCORCH DATA COORDINATION CENTER AND OTHER PUBLIC REPOSITORIES.

SINGLE MOLECULE STUDIES OF PROTEIN FOLDING MECHANISMS

CHEMOENZYMATIC GLYCAN EDITING FOR DECIPHERING BIOLOGICAL FUNCTIONS OF GLYCANS - PROJECT SUMMARY/ABSTRACT CELL-SURFACE GLYCANS PARTICIPATE IN NUMEROUS BIOLOGICAL PROCESSES, INCLUDING SIGNAL TRANSDUCTION, CELL-CELL COMMUNICATION AND DEVELOPMENT. ABERRANT GLYCOSYLATION IS A HALLMARK OF HUMAN DISEASE. AT A MOLECULAR LEVEL, GLYCANS REPRESENT THE FIRST POINTS OF CONTACT BETWEEN CELLS. HOWEVER, NOT DIRECTLY ENCODED IN THE GENOME, THESE BIOMOLECULES ARE CHALLENGING TO STUDY USING MOLECULAR BIOLOGY TECHNIQUES ALONE. METABOLIC OLIGOSACCHARIDE ENGINEERING (MOE) DEVELOPED IN LATE 1990’S HAS REVOLUTIONIZED THE WAY FOR THE LABELING AND VISUALIZATION OF GLYCANS IN LIVING ORGANISMS. IN THIS METHOD, CELLS’ OWN GLYCAN BIOSYNTHETIC MACHINERY IS HIJACKED TO INCORPORATE UNNATURAL MONOSACCHARIDES WITH LINKAGE PROMISCUITY. COMPLEMENTARY TO MOE, CHEMOENZYMATIC GLYCAN EDITING HAS EMERGED AS A VALUABLE TOOL TO PROBE AND MODIFY GLYCAN STRUCTURES WITHIN A CELLULAR ENVIRONMENT. UNLIKE MOE, CHEMOENZYMATIC GLYCAN MODIFICATION UTILIZES RECOMBINANT GLYCOSYLTRANSFERASES TO TRANSFER NATURAL OR UNNATURAL MONOSACCHARIDES WITH NOVEL FUNCTIONS FROM ACTIVATED NUCLEOTIDE SUGARS TO GLYCOCONJUGATES ON THE CELL SURFACE WITH LINKAGE SPECIFICITY. FOR THESE REASONS, CHEMOENZYMATIC GLYCAN MODIFICATION PROVIDES A FACILE AND MORE PRECISE WAY FOR PROBING THE FUNCTION OF GLYCANS IN THEIR NATIVE ENVIRONMENTS. BUILDING UPON OUR SUCCESSFUL APPLICATION OF CHEMOENZYMATIC GLYCAN EDITING, IN THE NEXT FIVE YEARS WE WILL EXPAND OUR CHEMOENZYMATIC TOOL KITS TO STUDY GLYCANS’ CELLULAR FUNCTIONS WITH A FOCUS ON THE SPECIAL ROLES OF N- ACETYLLACTOSAMINE (LACNAC), FUCOSE AND SIALIC ACID IN IMMUNE REGULATION. CELL-SURFACE LACNAC MEDIATES LIGAND-RECEPTOR BINDING AND SETS A THRESHOLD FOR INITIATING THE DOWNSTREAM SIGNALING FOR IMMUNE CELL ACTIVATION. LACNAC RESIDUES ARE DYNAMICALLY MODIFIED BY SIALIC ACID AND/OR FUCOSE. HOWEVER, THE SPECIFIC ROLES OF THESE MODIFICATIONS IN IMMUNE REGULATION AND DISEASE PROGRESSION REMAIN OBSCURE. WE ARE PARTICULARLY INTERESTED IN FINDING OUT: (1) IF CHANGES IN LACNAC AND FUCOSYLATION STATUS CAN SERVE AS GLYCAN SIGNATURES OF T CELL EXHAUSTION DURING WHICH T CELLS GRADUALLY LOSE THEIR CYTOKINE PRODUCTION, PROLIFERATION AND CYTOTOXIC CAPACITY; (2) CAN CELL-SURFACE IN SITU LACNAC FUCOSYLATION BE USED TO BOOST THE EFFICACY OF ANTITUMOR IMMUNITY OF T CELLS AND NK CELLS? IN PARALLEL, WE WILL DEVELOP CHEMOENZYMATIC TOOLS FOR PROFILING SIALYLATED GLYCOPROTEIN LIGANDS OF SIGLECS (SIALIC ACID-BINDING IMMUNOGLOBULIN-TYPE LECTINS) AND FOR THE IDENTIFICATION OF UNNATURAL, HIGH-AFFINITY AND SPECIFIC LIGANDS TO INTERROGATE SIGLEC FUNCTIONS. THROUGH THESE STUDIES, WE WILL GAIN A DEEPER UNDERSTANDING OF HOW LACNAC, FUCOSE AND SIALIC ACID ARE INVOLVED IN THE REGULATION OF THE IMMUNE CELL ACTIVATION, EFFECTOR FUNCTION AND EXHAUSTION. TOOLS DEVELOPED IN THIS PROJECT CAN ALSO BE USED TO STUDY OTHER TYPES OF GLYCANS AND THEIR INTERACTIONS WITH GLYCAN BINDING PROTEINS.

SCRIPPS TRANSLATIONAL SCIENCE INSTITUTE (UL1)

DEVELOPMENT OF ANTIGEN-SPECIFIC T CELL MEMORY

REGULATION OF ETHANOL EFFECTS ON SYNAPTIC TRANSMISSION

METABOLOMICS TECHNOLOGIES TO ADVANCE BIOMEDICAL RESEARCH

BIOPOLYMERS CONTAINING UNNATURAL BUILDING BLOCKS

SULFUR(VI) FLUORIDE EXCHANGE (SUFEX): NEW DEVELOPMENTS AND BIOLOGICAL APPLICATIONS

THE ROLE OF RAC1 IN CANCER

UNDERSTANDING BETA-SHEET STRUCTURE IN AQUEOUS SOLUTION.

AMICA: A NEW COSTIMULATORY MOLECULE GAMMADELTA T CELLS

EFFECTS OF INSULIN-DEPENDENT DIABETES RESISTANCE ALLELES ON CD8 TOLERANCE IN NOD

REGULATION OF MEMORY FORMATION BY THE GTPASE-ACTIVATING PROTEIN SYNGAP

NEUROPLASTICITY OF THE EXTENDED AMYGDALA CRF CIRCUITRY IN ALCOHOL DEPENDENCE

PROTEINS OF COAGULATION PATHWAYS

REGULATION OF G1-SPECIFIC GENE EXPRESSION IN YEAST

STUDIES OF THE DNA DAMAGE CHECKPOINTS

IMPROVING THE SCRIPPS RESEARCH INSTITUTE BSL3 CAPABILITIES TO COMBAT VIRUSES OF PANDEMIC CONCERN - PROJECT SUMMARY/ABSTRACT THE GOAL OF THE APPLICATION “IMPROVING THE SCRIPPS RESEARCH INSTITUTE BSL3 CAPABILITIES TO COMBAT VIRUSES OF PANDEMIC CONCERN” IS TO RENOVATE AND UPGRADE THE SCRIPPS RESEARCH INSTITUTE (TSRI) CURRENT ABSL3 AND BSL3 FACILITIES. THIS WILL ENHANCE TSRI RESEARCH CAPABILITIES TO COMBAT RNA VIRUSES OF PANDEMIC POTENTIAL. TSRI HAS ~ 2,000 SF DEDICATED TO ABSL3 AND ~ 1,500 SF DEDICATED TO BSL3. CURRENTLY, THESE FACILITIES SUPPORT RESEARCH PROJECTS FROM EIGHT TSRI FACULTY WHOSE COMBINED AREAS OF EXPERTISE COVER RESEARCH ON RNA VIRUSES OF PANDEMIC POTENTIAL, INCLUDING BUNYAVIRALES, CORONAVIRIDAE, FLAVIVIRIDAE, FILOVIRIDAE AND TOGAVIRIDAE. IN ADDITION, THE FACILITIES ARE SUPPORTING SEVERAL PROJECTS SPONSORED BY PHARMACEUTICAL COMPANIES. THE SPECIFIC STUDIES HAVE STRONG TRANSLATIONAL COMPONENTS AIMED AT DEVELOPING ANTIVIRAL DRUGS AND IMMUNOTHERAPEUTICS AGAINST THESE RNA VIRUSES. TSRI ABSL3 AND BSL3 SUITES, LOCATED IN THE MOLECULAR BIOLOGY BUILDING (MBB), HAVE BEEN OPERATING 24/7 SINCE THE LAST RENOVATIONS IN 2004. THE EXISTING MECHANICAL SYSTEMS HAVE BEEN MAINTAINED BEYOND ASHREA EQUIPMENT LIFE EXPECTANCY FOR THE LA JOLLA, CA. COASTAL CLIMATE. IN ADDITION, THE ORIGINAL DESIGN FROM 1985-87 HAS RESULTED IN UNDERUTILIZATION OF ~160 SF IN THE ABSL3 AND ~400 SF IN THE BSL3. TO CORRECT THESE DEFICIENCIES, TSRI HAS COMPLETED A CONCEPTUAL DESIGN WITH FPBA ARCHITECTS & TK1SC MECHANICAL ENGINEERS. THIS PLAN INCLUDES REPLACING THE AGING HVAC SYSTEMS OF THE ABSL3 AND BSL3 SPACE TO INCREASE AIRFLOW CAPACITY WITH N+1 REDUNDANCY AND UPDATE THE HEPA FILTRATION SYSTEM OF THE ABSL3. IN ADDITION, THE PROJECT WILL MODERNIZE THE BIOCONTAINMENT SUITES BY INCREASING HOLDING CAPACITY (ABSL3), CREATING A SELECT AGENT ISOLATION ROOM (BSL3), AND INSTALLING A NEW STERILIZER (ABSL3). THE CURRENT ABSL3/BSL3 FACILITIES LACK IMAGING, CELL SORTING, AND SINGLE CELL ANALYSIS CAPABILITIES. TO REMEDY THESE DEFICIENCIES, WE PROPOSE ADDING TO THE FACILITY THE EVOSTM IMAGING SYSTEM, THE SH800S CELL SORTER AND THE CHROMIUM IX SYSTEM. THESE IMPROVEMENTS WILL HELP TO ADDRESS THE INCREASING DEMANDS BY TSRI FACULTY FOR ABSL3/BSL3 SPACE, TIME AND IMPROVED FUNCTIONAL CAPABILITIES, WHILE ENSURING THE HIGHEST LEVEL OF SAFETY DURING DAILY OPERATIONS OF THE FACILITY. MOREOVER, RESEARCH ACTIVITIES PROPOSED UNDER THE ANTIVIRAL PROGRAM FOR PANDEMICS (APP) U19AI171443 AVIDD GRANT APPLICATION, WILL GREATLY BENEFIT FROM THE PROPOSED IMPROVEMENTS TO THE CURRENT TSRI ABSL3 AND BSL3 FACILITIES. THE PROJECT WILL BE MANAGED BY A WELL-INTEGRATED AND HIGHLY QUALIFIED TEAM, INCLUDING LEADERS FROM FACILITIES CONSTRUCTION AND ENGINEERING, THE INSTITUTIONAL ANIMAL PROGRAM AND RESOURCES, EHS AND BIOSAFETY, THE EXECUTIVE VP OFFICE, AND INVESTIGATORS DIRECTLY INVOLVED IN ABSL3/BSL3 RESEARCH ACTIVITIES. IN ADDITION, THE APPLICATION HAS THE UNWAVERING SUPPORT OF THE TSRI LEADERSHIP.

ANTISENSE MEDIATED TREATMENT OF HEPATITIS B VIRUS INFECTION

PTPN22 R619W IN NOD MICE

DISCOVERY AND ANALYSIS OF SYNTHETIC TLR AGONISTS AND ANTAGONISTS

ROLE OF NLRP3 SIGNALS IN ISCHEMIA/REPERFUSION-INDUCED ORGAN INJURY - PROJECT SUMMARY: THIS PROPOSAL EVALUATES HOW INTRACELLULAR SIGNALING PATHWAYS GENERATED BY THE CYTOPLASMIC INNATE IMMUNE RECEPTOR NLRP3 ARE REGULATED IN RENAL TUBULAR EPITHELIAL CELLS. RENAL TUBULE EPITHELIAL CELLS PLAY A CENTRAL ROLE IN ISCHEMIC KIDNEY INJURY, AND OUR LAB AND OTHERS HAVE SHOWN THAT GLOBAL NLRP3 BLOCKADE CAN PREVENT EXPERIMENTAL RENAL ISCHEMIA/REPERFUSION INJURY. NLRP3 SIGNALING CLASSICALLY RESULTS IN CELL DEATH (PYROPTOSIS), HOWEVER ALTERNATIVE NLRP3 SIGNALING PATHWAYS HAVE RECENTLY BEEN IDENTIFIED THAT LEAD TO SECRETION OF IL1B/IL18 WITHOUT CELL DEATH. THUS, SELECTIVELY TARGETING NLRP3-MEDIATED CELL DEATH SIGNALING MIGHT PREVENT RENAL TUBULE INJURY WHILE STILL PRESERVING THE HOST’S ABILITY TO SECRETE IL1B/IL18 AND MOUNT NEEDED HOST DEFENSE RESPONSES (E.G., ANTIMICROBIAL RESPONSES). THE PROJECT IS HIGHLY SIGNIFICANT FOR THE DEVELOPMENT OF TARGETED THERAPEUTICS FOR ISCHEMIC KIDNEY INJURY, WHICH OCCURS FREQUENTLY IN HOSPITALIZED PATIENTS AND IN ALL ORGANS PROCURED FOR TRANSPLANTATION. BROAD/LONG-TERM OBJECTIVES: THE LONG-TERM GOALS OF THE PROPOSED RESEARCH ARE TO DEFINE HOW NLRP3 CONTRIBUTES TO INJURIOUS TISSUE RESPONSES IN THE KIDNEY AND HOW ITS SIGNALING CAN BE EFFECTIVELY AND SELECTIVELY TARGETED. SPECIFIC AIMS: THE SPECIFIC OBJECTIVE OF THIS PROPOSAL IS TO TEST THE HYPOTHESIS THAT THE CYTOPLASMIC PATTERN RECOGNITION RECEPTOR NLRP3 CAN BE SPECIFICALLY TARGETED IN RENAL TUBULAR EPITHELIAL CELLS TO SELECTIVELY BLOCK NLRP3-MEDIATED RENAL TUBULE CELL DEATH, WHILE PRESERVING IL1B/IL18 RESPONSES. AIM 1 DEFINES THE MECHANISMS THAT DRIVE NLRP3 SIGNALING PATHWAYS IN PROXIMAL RENAL TUBULE CELLS. AIM 2 DETERMINES HOW NLRP3 SIGNALING CAN BE SELECTIVELY TARGETED IN THE PROXIMAL RENAL TUBULE CELLS. AIM 3 DETERMINES HOW NLRP3 SIGNALING, AND ITS SELECTIVE BLOCKADE, INFLUENCES RENAL TUBULAR INJURY/IL1B/IL18 SECRETION IN VIVO, WHEN NLRP3 EXPRESSION IS ISOLATED TO THE RENAL TUBULE EPITHELIUM. RESEARCH DESIGN AND METHODS FOR ACHIEVING THE STATED GOALS: AIM 1 COMPARES AND CONTRASTS ACTIVATION OF NLRP3 IN HUMAN AND MURINE PROXIMAL RENAL TUBULAR CELLS AND WILL DETERMINE HOW NLRP3 ACTIVATION CONTRIBUTES TO PYROPTOSIS AND/OR PRODUCTION OF IL1B/IL18. AIM 2 WILL DETERMINE HOW SPECIFIC NLRP3 ACTIVATION PATHWAYS CAN BE TARGETED IN THE TWO SPECIES TO PREVENT CELL DEATH AND PRESERVE CYTOKINE SECRETION. AIM 3 WILL FOCUS ON UNDERSTANDING HOW NLRP3 RESTRICTED TO THE RENAL TUBULE EPITHELIUM IN VIVO CAN BE TARGETED TO PREVENT RENAL TUBULAR INJURY WITHOUT IMPAIRING IL1B/IL18. IN THE THIRD AIM, TWO DIFFERENT MODELS OF NLRP3 EXPRESSION WILL BE USED; ONE WHERE THE RENAL TUBULES EXPRESS A CONDITIONAL LOSS-OF-FUNCTION OF NLRP3 AND ANOTHER WHERE THERE IS A HYPER-ACTIVATABLE FORM OF NLRP3. THE STUDIES HERE WILL DETERMINE WHETHER CANONICAL NLRP3-MEDIATED CELL DEATH SIGNALING CAN BE UNCOUPLED FROM NONCANONICAL SIGNALING TO PREVENT CELL DEATH WHILE PRESERVING SECRETION OF IL1B/IL18. HEALTH RELATEDNESS OF PROJECT: IF THE AIMS OF THIS PROPOSAL ARE MET, WE WILL LEARN HOW ACTIVATION OF NLRP3 (DEEMED A CENTRAL CELLULAR STRESS DETECTOR) CAN BE REGULATED TO TARGET A COMMON DISEASE, RENAL ISCHEMIA/REPERFUSION INJURY. THIS KNOWLEDGE IS CRUCIAL FOR THE DEVELOPMENT OF RATIONAL TARGETED THERAPIES FOR PREVENTION OR AMELIORATION OF RENAL ISCHEMIA/REPERFUSION INJURY IN CLINICAL SITUATIONS WHERE HYPOXIA IS ANTICIPATED.

MODULATING SIGNALING ENDOCANNABINOIDS AND FATTY ACID AMIDES

SYSTEMS BIOLOGY OF RNA MODIFICATIONS IN HIV/AIDS AND SUBSTANCE USE DISORDERS

MOUSE MODELS TO STUDY INHERITED FORMS OF DEAFNESS

MASSIVELY PARALLEL IDENTIFICATION OF PROTEIN LIGANDS

GENOMIC MODIFICATION WITH PURIFIED NUCLEASE PROTEINS FOR HIV-1 THERAPY

STRUCTURAL BASIS FOR CBP/P300 TRANSCRIPTIONAL REGULATION

GRADUATE RESEARCH FELLOWSHIP PROGRAM (GRFP)

INNATE REGULATION OF ADAPTIVE IMMUNITY

GENE-ENVIRONMENT INTERACTION: THE BRAIN CRF SYSTEM IN ALCOHOL PREFERRING MSP RATS

PROBING THE GERMLINES OF BROADLY NEUTRALIZING ANTI-HIV ANTIBODIES IN KNOCKIN MICE

EXTRA-TRANSLATIONAL ROLES OF AMINOACYL TRNA SYNTHETASES IN CONNECTION TO DISEASE

BIOLOGY OF ?-LINKED GLYCOSYLCERAMIDES IN THE IMMUNE SYSTEM

DEFINING GENOME STABILITY MECHANISMS AND THEIR REGULATION BY SUMO AND UBIQUITIN