Project Grant R41AI165350

CCR5 IMMUNOTOXINS AS COMPONENTS OF HIV CURE REGIMENS - THIS PROJECT WILL ESTABLISH PROOF-OF-CONCEPT FOR CCR5 IMMUNOTOXINS AS HIV RESERVOIR-DEPLETION AGENTS FOR USE IN CURE STRATEGIES. HIV CURE AND REMISSION STRATEGIES REQUIRE ELIMINATION OR REDUCTION OF THE HIV RESERVOIR. APPROACHES TO RESERVOIR REDUCTION PURSUED BY OTHER GROUPS INCLUDE GENE EDITING, I.E., DIRECT REMOVAL OF INTEGRATED PROVIRUS; LATENCY REVERSAL AND AN ACCOMPANYING "KILL" STRATEGY; AND "BLOCK-AND-LOCK", WHICH IS THE EFFECTIVE ELIMINATION OF RESERVOIR CELLS BY PERMANENTLY SUPPRESSING GENE EXPRESSION. WE INSTEAD PURSUED A STRATEGY OF DIRECTLY ELIMINATING POTENTIAL RESERVOIR CELLS IN EARLY INFECTION, HYPOTHESIZING THAT MOST EARLY RESERVOIR CELLS EXPRESS CCR5. USING A CD3/CCR5 BISPECIFIC ANTIBODY FOR RESERVOIR DEPLETION, WE DEMONSTRATED DELAYED REBOUND IN 4/7 AND APPARENT CURE OF 2/7 SIV-INFECTED INFANT MACAQUES. ONE CURED ANIMAL WAS NECROPSIED 204 DAYS AFTER ART WITHDRAWAL AND THE SECOND REMAINS AVIREMIC AFTER 1.6 YEARS. BOTH WERE DEPLETED OF CD8+ T CELLS TO ENCOURAGE VIRAL REBOUND, BUT NO REBOUND WAS SEEN. NO PROVIRAL DNA WAS DETECTED IN CIRCULATING CELLS AT ANY TIME POINT FOLLOWING ART WITHDRAWAL. SENSITIVE VIRAL OUTGROWTH ASSAYS FAILED TO RECOVER REPLICATION-COMPETENT VIRUS. NO PROVIRAL GENOMES WERE DETECTED IN THE TISSUES OF ONE SACRIFICED AND APPARENTLY CURED ANIMAL. THUS, RESULTS OBTAINED SO FAR SHOW THAT THESE ANIMALS HAVE ACHIEVED AT LEAST "FUNCTIONAL" AND PERHAPS STERILIZING CURE. THE BSAB WE EMPLOYED ACHIEVES VERY EFFICIENT, TRANSIENT DEPLETION OF CCR5+ CELLS BUT ALSO CAUSES AN INFLAMMATORY REACTION WITH CYTOKINE PRODUCTION AND TEMPORARY CD3+ LYMPHOPENIA. TENDEL IS THEREFORE DEVELOPING CCR5 IMMUNOTOXINS FOR USE IN CURATIVE ANTI-HIV REGIMENS. WE HYPOTHESIZE THAT CCR5 IMMUNOTOXINS CAN ACHIEVE SPECIFIC AND MINIMALLY TOXIC DEPLETION OF CCR5+ T CELLS FROM BLOOD, GUT, AND LYMPHOID TISSUES. SA1. PRODUCE OPTIMIZED IMMUNOTOXINS BASED ON CCR5 LIGAND-TOXIN FUSIONS. PREVIOUSLY PUBLISHED IMMUNOTOXINS WERE EFFECTIVE AT HIGH CONCENTRATION IN VITRO BUT FAILED IN-VIVO TESTS. IN THIS AIM WE DEVELOP NEW IMMUNOTOXINS BASED ON CCR5 LIGANDS COMBINED WITH LINKERS OF DIFFERENT LENGTHS AND WITH VARIOUS TOXIN CANDIDATES. WE THEN TEST THE CANDIDATES IN VITRO FOR SPECIFIC LETHALITY TO CCR5-EXPRESSING CELLS. SA2. TEST PHARMACODYNAMICS OF DEVELOPMENT CANDIDATES IN RHESUS MACAQUES, WITH A FOCUS ON CCR5 DEPLETION FROM GUT TISSUES WITHOUT IMMUNE ACTIVATION. HERE THE DEVELOPMENT CANDIDATES FROM AIM 1 ARE ASSESSED IN MACAQUES AND THE RESULTS COMPARED TO DEPLETION ACHIEVED BY THE BENCHMARK BSAB, WHICH IS ALREADY SHOWN TO BE POTENTIALLY CURATIVE. MOST IMPORTANTLY, THESE EXPERIMENTS TEST IF CANDIDATE IMMUNOTOXINS CAN ACHIEVE NEARLY 100% CCR5 DEPLETION FOR AT LEAST SEVERAL WEEKS IN RECIPIENT ANIMALS. OUR PRELIMINARY DATA SHOW THAT THE CCR5+ RESERVOIR IS AN "ACHILLES' HEEL" IN EARLY SIV AND PERHAPS HIV INFECTION. THE PROPOSED RESEARCH WILL PURSUE PROOF-OF-CONCEPT FOR NOVEL TENDEL IMMUNOTOXINS.