Seattle Children's Healthcare System

IMMUNE MECHANISMS OF PROTECTION AGAINST MYCOBACTERIUM TUBERCULOSIS CENTER (IMPAC-TB)

UNDER THIS OPTION WE WILL CONDUCT STUDIES IN RM TO VALIDATE THE EFFICACY OF THE RHCMV/TB VECTOR ORTHOLOG OF THE HUMAN CMV VACCINE CANDIDATE (VIR-1778) CURRENTLY SCHEDULED FOR TESTING IN A PHASE 1 CLINICAL TRIAL. WE WILL ALSO DEVELOP AND TEST RHCMV VECTORS THAT ARE MODIFIED TO REDUCE OR ELIMINATE THE DELETERIOUS IMMUNOREGULATORY EFFECTS OF THE CONTROL RHCMV VECTORS THAT LED TO ACCELERATED TB DISEASE, IN PARTICULAR VECTOR BACKBONE MODIFICATIONS THAT WILL KEEP THE RHCMV/TB VECTOR OUT OF THE LUNG, AS WELL AS OBVIATE ITS ABILITY TO INFECT MONOCYTE/MACROPHAGES, ENDOTHELIAL AND EPITHELIAL CELLS SAND TO PRODUCE IMMUNOREGULATORY VIRAL (V) IL-10 (YEARS 6-7). SUB-ACTIVITY 1: GENERATE A RHCMV/TB (AND RHCMV/CONTROL) VECTOR BASED ON THE VIR-1778 HCMV/TB VECTOR (FL-RHCMV + ∀ RH157.5-∀ RH157.4 + RH107 R MTB-7AG) AND 4 DIFFERENT MODIFIED VERSIONS OF THIS ORTHOLOG VECTOR DESIGNED TO OBVIATE THE DELETERIOUS IMMUNOREGULATION MENTIONED ABOVE. ALL VECTORS WILL BE MADE AS BOTH TB-7AG INSERT EXPRESSION VECTORS (USING THE SAME TB-7AG CASSETTE BEING USED IN VIR 1778), AND AS A CONTROL VECTOR EXPRESSING AN INSERT OF ROUGHLY SIMILAR SIZE EXPRESSING UNRELATED SIV AGS. SUB-ACTIVITY 2: SELECT 2 OF THE 4 BACKBONE DESIGNS FROM SUB-ACTIVITY 1 AND THE VIR-1778 ORTHOLOG VECTOR SET FOR IMMUNOGENICITY AND EFFICACY TESTING IN MONKEYS, COMPARING THE BACKBONES WITH MTB VS. CONTROL AG INSERTS VS. UNVACCINATED CONTROLS. THE FOLLOWING GROUPS WILL BE TESTED USING UP TO A TOTAL OF 56 MONKEYS: €¢ VECTOR A (VIR-1778 ORTHOLOG) AND MATCHING CONTROL AG INSERT VECTOR; 8 MONKEYS EACH €¢ VECTOR B* (MODIFIED VIR-1388 ORTHOLOG VECTOR #1) AND MATCHING CONTROL AG INSERT VECTOR; 7-8 MONKEYS EACH €¢ VECTOR C* (MODIFIED VIR-1388 ORTHOLOG VECTOR #2) AND MATCHING CONTROL AG INSERT; 7-8 MONKEYS EACH, AND €¢ 8 UNVACCINATED CONTROLS SUB-ACTIVITY 3: PERFORM TIMED NECROPSIES AT WEEK 4 OR 5 POST-CHALLENGE AND PERFORM MICROBIOLOGIC AND HISTOPATHOLOGIC/IN SITU ANALYSES TO MEASURE EXTENT OF MTB INFECTION. SUB-ACTIVITY 4: CONDUCT LONGITUDINAL AND END-POINT (NECROPSY) ASSAYS TO MONITOR TRANSCRIPTOMIC PREDICTORS OF VACCINE EFFICACY AND THE DEVELOPMENT AND RESPONSE TO MTB CHALLENGE OF MTB-SPECIFIC CD4+ AND CD8+ T CELL RESPONSES AND INNATE €œTRAINED€Ͽ½ IMMUNITY. VECTOR B AND C TO BE SELECTED FROM VECTORS WITH THE FOLLOWING ADDITIONAL MODIFICATIONS (THAT IS ADDED TO TO THE VIR-1378 ORTHOLOG VECTOR BACKBONE) €¢ SINGLE CYCLE VERSION: + RH108-FKBP €¢ VIL-10 DELETED: + ∀ RH143 €¢ MYELOID, ENDOTHELIAL AND EPITHELIAL TROPISM-RESTRICTED AND VIL-10 DELETED: + ∀ RH143 + MIR126/142/205 (RH108/RH156) €¢ DOUBLE-DELETED AND MYELOID, ENDOTHELIAL AND EPITHELIAL TROPISM-RESTRICTED AND VIL-10 DELETED: + ∀ RH143 + MIR126/142/205 (RH108/RH156) + ∀ RH158-∀ RH161

THE UNIVERSITY OF WASHINGTON CRYO-EM GROUP (UW-CRYOEM), UNDER THE DIRECTION OF DAVID VEESLER, PH.D., WILL PERFORM THE FOLLOWING WORK TO SUPPORT THE BASIC PROJECT REQUIREMENT. UW-CRYOEM WILL BE RESPONSIBLE FOR SOLVING CRYOEM STRUCTURES DURING THIS CONTRACT YEAR (SEPT 1, 2024 THROUGH AUG 31, 2025). UW-CRYOEM MAY ALSO MANAGE (SHEPHERD) AND WILL SUPPORT OTHER SPECIAL STUDIES AS NEEDED. A. STRUCTURE DETERMINATION OF PROTEINS OR PROTEIN COMPLEXES BY CRYOEM: UW-CRYOEM (DR. VEESLER) WILL DETERMINE THE STRUCTURE OF PROTEIN OR PROTEIN COMPLEXES USING CRYOEM AS A PART OF THIS SUBCONTRACT. THESE TARGET(S) WILL ORIGINATE FROM NIAID CATEGORY A-C INFECTIOUS DISEASE ORGANISMS, AS WELL AS EMERGING AND RE-EMERGING DISEASE AND ONCE APPROVED BY NIAID WILL ENTER THE SSGCID PIPELINE FOR TRACKING THE UW-CRYOEM WORK. THE TARGETS SELECTED WILL BE IMPACTFUL AND MAY BE MANAGED AS HIGH VALUE TARGET (HVT), COMMUNITY REQUEST PROJECT (CRP), HIGHEST IMPACT PROJECT (HIP) AND OTHER SPECIAL STUDIES WITHIN THE SSGCID PIPELINE. PROTEINS FROM VIRAL AND/OR EUKARYOTIC TARGETS WILL BE EXPRESSED IN BACULOVIRUS-INFECTED INSECT CELLS OR MAMMALIAN CELLS (HEK), WHILE BACTERIAL TARGETS (IF SELECTED FOR CRYOEM) WILL BE EXPRESSED IN E. COLI. THE MOLECULAR WEIGHT AND STOICHIOMETRY OF THE PURIFIED PROTEINS WILL BE CONFIRMED USING ANALYTICAL SEC COUPLED WITH MULTI-ANGLE LIGHT SCATTERING (SEC-MALS). THE HOMOGENEITY AND STRUCTURAL STABILITY OF THE PURIFIED PROTEINS WILL ALSO BE ASSESSED BY HIGH-THROUGHPUT NEGATIVE STAIN ELECTRON MICROSCOPY USING AN AUTOMATED DATA COLLECTION AND PROCESSING PIPELINE. PROMISING SAMPLES WILL BE FURTHER EVALUATED BY COMPUTING LOW-RESOLUTION THREE-DIMENSIONAL RECONSTRUCTIONS THAT DEMONSTRATE THE COMPOSITIONAL AND CONFORMATIONAL HETEROGENEITY OF EACH SAMPLE. THIS INFORMATION CAN ALSO BE USED TO GUIDE FURTHER CONSTRUCT SELECTION AND OPTIMIZATION OF THE EXPRESSION AND PURIFICATION CONDITIONS (IF NECESSARY). SAMPLES WILL BE VITRIFIED WITH AN FEI VITROBOT DEVICE TO PLUNGE-FREEZE GRIDS IN LIQUID ETHANE UNDER CAREFULLY CONTROLLED CONDITIONS. THE CRYOEM EXPERIMENTS WILL BE CARRIED OUT ON A TITAN KRIOS ELECTRON MICROSCOPE, EQUIPPED WITH A GATAN GIF ENERGY FILTER AND A K2 SUMMIT DIRECT ELECTRON DETECTOR. AUTOMATED DATA ACQUISITION AND PROCESSING ALLOWS A STREAMLINING OF ALL THE STEPS INVOLVED IN STRUCTURE DETERMINATION BY CRYOEM AT NEAR-ATOMIC RESOLUTION. THE LEGINON DATA COLLECTION SOFTWARE RUNS CONTINUOUSLY; ENABLING AUTOMATED TARGETING AND IMAGE ACQUISITION. LEGINON IS INTEGRATED WITH APPION, A DATA PROCESSING PIPELINE THAT CONNECTS DATA ACQUISITION TO PROCESSING ON A LOCAL CLUSTER, PROVIDING A RELATIONAL DATABASE OF ALL MICROGRAPHS TO EXPEDITE SUBSEQUENT PROCESSING. APPION AUTOMATES MANY IMAGE PROCESSING TASKS AND CREATES A SMOOTH WORKFLOW THAT ALLOWS REAL-TIME ASSESSMENT OF DATA QUALITY DURING ACQUISITION AND PREVENTS ERRORS BY TRACKING ALL THE DETAILS RELATED TO IMAGE ACQUISITION AND PROCESSING. B. PANDEMIC PREPAREDNESS AND RAPID RESPONSE TO OUTBREAKS/EPIDEMIC/PANDEMICS: UW-CRYOEM WILL LEAD A STUDY ON THE COMPUTATIONAL DESIGN OF STABILIZED NIPAH VIRUS ATTACHMENT AND FUSION GLYCOPROTEINS. C. QUALITY CONTROL, DATA COLLECTION AND REPORTING: UW-CRYOEM WILL ESTABLISH AND CONDUCT QUALITY CONTROL PROCEDURES AS NECESSARY TO ENSURE THE PRODUCTION OF HIGH-QUALITY DATA. UW-CRYOEM WILL ENSURE THAT THE STATUS OF ALL WORK PERFORMED FOR SSGCID IS UPDATED IN A TIMELY FASHION, THROUGH THE CENTRAL TARGET TRACKING DATABASE (CTTDB), TRACKING SPREADSHEETS AND/OR ANY OTHER TRACKING SYSTEM REQUIRED BY THE PROJECT. DR. VEESLER AND/OR STAFF FROM UW-CRYOEM WILL PARTICIPATE IN SSGCID PROJECT MEETINGS AND NIAID-REQUIRED MEETINGS AS REQUIRED TO ENSURE THE SUCCESS OF THE PROJECT. DR. VEESLER AND/OR STAFF FOMR UW-CRYOEM WILL CONTRIBUTE TO QUARTERLY AND SEMI-ANNUAL PROGRESS REPORTS AND ANY OTHER NIAID REQUIRED REPORTS AS NEEDED AND IN A TIMELY FASHION AND WILL CONDUCT THE PROJECT WORK WITH THE AIM OF PRODUCING IMPACTFUL PUBLICATIONS. TAKEN TOGETHER IN PARTS A AND B ABOVE, TE

THIS STATEMENT OF WORK (SOW) OUTLINES THE ACTIVITIES THAT COLORADO STATE UNIVERSITY (CSU) WILL BE PERFORMING THROUGHOUT THE PROPOSED WORK PERIOD. NOTE THAT THE TASK NUMBERS BELOW REFLECT THE NUMBERING OF THE OVERALL ACTIVITIES IN THE PRIME CONTRACT. THROUGH THE EXECUTION OF THIS CONTRACT, WE WILL EXAMINE BASIC INDUCTION OF INNATE AND ADAPTIVE IMMUNITY FROM VACCINATION, INCLUDING STUDIES THAT ADDRESS NONTUBERCULOUS MYCOBACTERIA (NTM) INTERFERENCE, RNA VACCINE DELIVERY, PRIME-BOOST REGIMENS (INCLUDING BCG PRE-EXPOSURE), TRANSCRIPTOMIC HOST SIGNATURES, SYSTEMS BIOLOGY AND DURABLE IMMUNE MEMORY FOR THE FURTHER ADVANCEMENT OF THE EFFORT TO DESIGN AND EVALUATE AN EFFICACIOUS VACCINE AGAINST MYCOBACTERIUM TUBERCULOSIS (M.TB). THE WORK UNDER THIS CONTRACT MAY INCLUDE THE POSSIBLE DISTRIBUTION OF DE-IDENTIFIED HUMAN SUBJECT PERIPHERAL BLOOD MONONUCLEAR CELLS (PBMC), SERUM, AND OTHER CLINICAL SAMPLES TO SCRI AND THE PHOENIX PARTNERS. NHP-DERIVED SAMPLES AND/OR DOWNSTREAM DATA MAY BE SHARED WITH THE PARTNERS ON THIS

THE UNIVERSITY OF KANSAS (KU), UNDER THE DIRECTION OF SCOTT LOVELL, PH.D., WILL PERFORM THE FOLLOWING WORK TO SUPPORT THE BASIC PROJECT REQUIREMENT (9/1/2024-8/31/2025). KU WILL ACCEPT PROTEIN AND OTHER SAMPLES PRIMARILY GENERATED BY OTHER SSGCID SITES AND PREPARE THEM FOR X-RAY CRYSTALLOGRAPHY STRUCTURE DETERMINATION STUDIES, CONDUCT CRYSTALLIZATION EXPERIMENTS, COLLECT DIFFRACTION DATA, AND PERFORM STRUCTURE SOLUTION. KU WILL ALSO SUPPORT VARIOUS SPECIAL PROJECT STUDIES AS NEEDED AND LEAD ONE FUNCTIONAL STUDY. A. STRUCTURE DETERMINATION OF PROTEINS OR PROTEIN COMPLEXES BY X-RAY CRYSTALLOGRAPHY: KU WILL ACCEPT PROTEIN SAMPLES PRIMARILY RECEIVED FROM THE OTHER TEAM SITES FOR X-RAY CRYSTALLOGRAPHY STRUCTURE DETERMINATION STUDIES AND CONDUCT HIGH THROUGHPUT CRYSTALLIZATION EXPERIMENTS, INCLUDING CRYSTALLIZATION TRIALS, OPTIMIZATION, DATA COLLECTION AND INTERPRETATION. WHEN NECESSARY, KU WILL CONDUCT CO-CRYSTALLIZATION SCREENS OF THE TARGET IN COMPLEX WITH SMALL MOLECULES AND LIGANDS. KU WILL IDENTIFY AND HARVEST CRYSTALS THAT ARE SUITABLE FOR DIFFRACTION SCREENING AND CONDUCT LARGE SCALE CRYSTAL DIFFRACTION SCREENS, COLLECT DIFFRACTION DATA AND MONITOR QUALITY METRICS OF THE SUITABLE CRYSTALS. WHERE APPROPRIATE, KU WILL ALSO LIAISE WITH KUMC€„¢S CRYOEM CENTER TO HELP SOLVE STRUCTURES BY CRYOEM. KU WILL PERFORM STRUCTURE SOLUTION, MODEL REFINEMENT AND VALIDATION FOR X-RAY CRYSTALLOGRAPHY. A STRUCTURE DETERMINATION PROJECT WILL BE CONSIDERED COMPLETE WHEN THE REFINEMENT AND VALIDATION OF THE FINAL STRUCTURE MODELS DO NOT PROVIDE ANY CLEAR SIGNAL OF WHERE THE MODEL CAN BE IMPROVED OR WHEN 3D STRUCTURE QUALITY PARAMETERS SATISFY THE MINIMUM QUALITY REQUIREMENTS ESTABLISHED AND AGREED UPON IN COORDINATION WITH THE CENTER FOR STRUCTURAL BIOLOGY OF INFECTIOUS DISEASES. KU WILL SUPPORT HIGH VALUE TARGETS (HVTS), COMMUNITY REQUEST PROJECTS (CRP), HIGHEST IMPACT PROJECTS (HIP) AND OTHER SPECIAL STUDIES WITHIN THE SSGCID PIPELINE, AS DECIDED BY THE SSGCID SCIENTIFIC LEADERSHIP TEAM (SLT). B. FUNCTIONAL STUDIES KU WILL LEAD ONE FUNCTION STUDY, ENTITLED €œSTRUCTURE AND FUNCTIONAL STUDIES OF RSBU FROM CHLAMYDIA,€Ͽ½ WITH THE GOAL TO PERFORM STRUCTURAL AND FUNCTIONAL ANALYSIS OF THE RSBU PROTEIN FROM C. TRACHOMATIS. C. QUALITY CONTROL, DATA COLLECTION AND REPORTING: KU WILL ESTABLISH AND CONDUCT QUALITY CONTROL PROCEDURES AS NECESSARY TO ENSURE THE PRODUCTION OF HIGH-QUALITY DATA. KU WILL ENSURE THAT THE STATUS OF ALL WORK PERFORMED FOR SSGCID IS UPDATED IN A TIMELY FASHION, THROUGH THE CENTRAL TARGET TRACKING DATABASE (CTTDB), TRACKING SPREADSHEETS AND/OR ANY OTHER TRACKING SYSTEM REQUIRED BY THE PROJECT. KEY STAFF MEMBERS FROM KU WILL PARTICIPATE IN SSGCID PROJECT MEETINGS AND NIAID-REQUIRED MEETINGS TO ENSURE THE SUCCESS OF THE PROJECT. KU STAFF WILL CONTRIBUTE TO QUARTERLY AND SEMI-ANNUAL PROGRESS REPORTS AND ANY OTHER NIAID REQUIRED REPORTS AS NEEDED AND IN A TIMELY FASHION AND WILL CONDUCT THE PROJECT WORK WITH THE AIM OF PRODUCING IMPACTFUL PUBLICATIONS. IT IS EXPECTED THAT SIXTY STRUCTURES WILL BE SOLVED THROUGH THIS WORK.

THIS STATEMENT OF WORK (SOW) OUTLINES THE ACTIVITIES THAT OXFORD UNIVERSITY WILL BE PERFORMING THROUGHOUT THE PROPOSED TOTAL WORK PERIOD. NOTE THAT THE TASK NUMBERS BELOW REFLECT THE NUMBERING OF THE OVERALL ACTIVITIES IN THE PRIME CONTRACT. OPTION 4 ACTIVITIES: CONDUCT TWO PHASE 1 CLINICAL TRIALS IN THE CONTROLLED HUMAN MYCOBACTERIAL CHALLENGE MODEL USING AN ESTABLISHED MODEL OF AEROSOLIZED BCG DELIVERED TO THE LUNGS OF HUMANS, FOLLOWED BY BRONCHOSCOPIC ASSESSMENT OF MUCOSAL IMMUNITY, ASSESS VACCINE-MEDIATED PROTECTION FROM THIS CHALLENGE. TO DATE WE HAVE DELIVERED BCG VIA THE AEROSOL ROUTE TO OVER 65 VOLUNTEERS AND REPEATED THE DOSE ESCALATION FROM 1X104CFU - 1 X 107CFU BCG WITH NO SAFETY CONCERNS. PROTOCOL SUMMARY OF THE TWO PHASE I CLINICAL TRIALS: TB0441: A CLINICAL CHALLENGE STUDY TO EVALUATE CONTROLLED HUMAN INFECTION WITH BCG ADMINISTERED BY THE AEROSOL INHALED ROUTE IN HISTORICALLY BCG-VACCINATED HEALTHY ADULT VOLUNTEERS PHASE: 1 STUDY DURATION: 18 MONTHS POPULATION: HEALTHY ADULT VOLUNTEERS, AGED 18-50. A TOTAL OF 12 SUBJECTS WILL BE ENROLLED. ESTIMATED TIME TO COMPLETE ENROLLMENT: APPROXIMATELY 9 MONTHS SUBJECT PARTICIPATION DURATION: 6 MONTHS FROM ENROLLMENT NUMBER OF SITES: ONE (UNIVERSITY OF OXFORD/ OXFORD UNIVERSITY HOSPITALS NHS FOUNDATION TRUST) DESCRIPTION OF AGENTS OR INTERVENTION: BCG STUDY OBJECTIVES: PRIMARY AIM IS TO QUANTIFY THE BCG RECOVERABLE FROM BRONCHIAL SAMPLES IN HEALTHY HISTORICALLY BCG-VACCINATED ADULT VOLUNTEERS SECONDARY AIM IS TO EVALUATE THE HUMAN CLINICAL RESPONSE TO BCG CHALLENGE BY THE AEROSOL INHALED ROUTE IN HEALTHY, HISTORICALLY BCG-VACCINATED ADULT VOLUNTEERS EXPLORATORY AIM IS TO IDENTIFY LABORATORY MARKERS OF THE IMMUNE RESPONSE THAT CORRELATE WITH THE LEVELS OF BCG RECOVERED FROM BRONCHIAL SAMPLES STUDY OUTCOME MEASURES: PRIMARY OUTCOME MEASURE: 1. BCG RECOVERABLE FROM BRONCHIAL SAMPLE BCG CULTURE BY MYCOBACTERIAL GROWTH INDICATOR TUBE; PCR QUANTIFICATION AND DNA STRIP ASSAY SPECIATION AS REQUIRED [TIME FRAME: UP TO DAY 168] SECONDARY OUTCOME MEASURE: 2. EVALUATION OF THE HUMAN CLINICAL RESPONSE TO BCG CHALLENGE ACTIVELY AND PASSIVELY COLLECTED DATA ON ADVERSE EVENTS; DETAILED PARTICIPANT SYMPTOM PROFILES, LUNG FUNCTION TEST RESULTS [TIME FRAME: UP TO DAY 168] OTHER PRE-SPECIFIED OUTCOME MEASURES: 3. LABORATORY MARKERS OF THE IMMUNE RESPONSE THAT CORRELATE WITH THE LEVELS OF BCG RECOVERED FROM BRONCHIAL SAMPLES. LABORATORY MARKERS OF INNATE AND ADAPTIVE IMMUNITY, INCLUDING EX-VIVO IFN-ïϿ½Â§ ELISPOT AND ELISA; RNA SEQUENCE ANALYSIS AND INTRACELLULAR CYTOKINE STAINING OF BLOOD AND BRONCHIAL SAMPLES. BCG CFU COUNTS, MGIT AND/ OR MOLECULAR DIAGNOSTICS ON INDUCED SPUTUM (IF PERFORMED) AND ON MATRIX FROM DUCKBILL MASK COLLECTION [TIME FRAME: UP TO DAY 168] DESCRIPTION OF STUDY GROUPS: GROUP 1: DOSE ESCALATION IN HISTORICALLY BCG VACCINATED VOLUNTEERS (N=12) 1A: 3 HISTORICALLY BCG VACCINATED VOLUNTEERS WILL RECEIVE 1 X 104 CFU AEROSOL INHALED BCG 1B: 3 HISTORICALLY BCG VACCINATED VOLUNTEERS WILL RECEIVE 1 X 105 CFU AEROSOL INHALED BCG 1C: 3 HISTORICALLY BCG VACCINATED VOLUNTEERS WILL RECEIVE 1 X 106 CFU AEROSOL INHALED BCG 1D: 3 HISTORICALLY BCG VACCINATED VOLUNTEERS WILL RECEIVE 1 X 107 CFU AEROSOL INHALED BCG TB0451: A PHASE I CLINICAL TRIAL TO EVALUATE (I) THE ADAPTIVE IMMUNE RESPONSE TO A CONTROLLED HUMAN INFECTION WITH AEROSOL BCG; AND (II) THE PROTECTIVE EFFICACY OF A PRIOR BCG VACCINATION AND/OR ID93+GLA-SE VACCINATION USING THIS MODEL PHASE: 1 STUDY DURATION: 30 MONTHS POPULATION: HEALTHY ADULT VOLUNTEERS, AGED 18-50. A TOTAL OF 48 SUBJECTS WILL BE ENROLLED. ESTIMATED TIME TO COMPLETE ENROLLMENT: APPROXIMATELY 21 MONTHS SUBJECT PARTICIPATION DURATION: BCG ONLY: 2 MONTHS FROM ENROLLMENT ID93 + GLA-SE + BCG: 6 MONTHS FROM ENROLLMENT NUMBER OF SITES: ONE (UNIVERSITY OF OXFORD/ OXFORD UNIVERSITY HOSPITALS NHS FOUNDATION TRUST) WITH A POSSIBILITY OF A SECOND SITE TO AID RECRUITMENT (I.E SOUTHAMPTON, BIRMINGHAM) DESCRIPTION OF AGENTS OR INTERVENTION: BCG ID93 + GLA-SE (2 ÂÂΜG