Project Grant F31AI202969

ELUCIDATING JNK REGULATION OF KILLER IMMUNOGLOBULIN-LIKE RECEPTOR ACQUISITION DURING HUMAN NK CELL DEVELOPMENT - PROJECT SUMMARY/ABSTRACT HUMAN NATURAL KILLER (NK) CELLS PLAY A CRITICAL ROLE IN PROTECTION AGAINST MALIGNANCY AND RE-ESTABLISHING HOST IMMUNITY AFTER ALLOGENEIC HEMATOPOIETIC STEM CELL TRANSPLANT (ALLO-HSCT). KILLER IMMUNOGLOBULIN-LIKE RECEPTORS (KIRS) ARE EXPRESSED ON THE SURFACE OF MATURE HUMAN NK CELLS AND PLAY AN INTEGRAL ROLE IN REGULATING NK CELL FUNCTION. HOWEVER, FOR UNKNOWN REASONS, KIR EXPRESSION ON NK CELLS RECONSTITUTED AFTER ALLO-HSCT CAN BE LOW, CORRELATING WITH WORSE PATIENT OUTCOMES. ADDITIONALLY, WHILE A LARGE NUMBER OF NK CELLS CAN BE DEVELOPED FROM CD34+ HEMATOPOIETIC PROGENITORS IN-VITRO FOR CLINICAL USE, KIR EXPRESSION CAN BE LOW AND VARIABLE ON THESE CELLS, DESPITE KIR EXPRESSION BEING ASSOCIATED WITH SUPERIOR NK CELL FUNCTION. THEREFORE, WE SEEK TO DETERMINE THE MECHANISMS THAT DRIVE KIR EXPRESSION DURING NK CELL DEVELOPMENT IN ORDER TO IMPROVE NK CELL-BASED THERAPIES AND ULTIMATELY IMPROVE PATIENT OUTCOMES. EVIDENCE HAS SHOWN THAT KIRS ARE REGULATED THROUGH A CAREFULLY CHOREOGRAPHED BALANCE OF METHYLATION, TRANSCRIPT EXPRESSION FROM PROXIMAL, INTERMEDIATE, AND DISTAL PROMOTERS, AND ANTISENSE TRANSCRIPT PRODUCTION. DESPITE THIS, THE MECHANISMS WHICH INITIATE AND PROMOTE KIR ACQUISITION DURING NK CELL DEVELOPMENT REMAIN UNKNOWN. FINDINGS FROM OUR GROUP SUGGEST THAT PARADOXICALLY WITHHOLDING IL- 15, A CRITICAL CYTOKINE FOR NK CELL DEVELOPMENT, DURING THE FIRST TWO WEEKS OF A NOVEL 28 DAY "SWITCH CULTURE" SYSTEM CAN SIGNIFICANTLY AUGMENT THE GENERATION OF KIR+ NK CELLS FROM CD34+ HEMATOPOIETIC PROGENITORS. FURTHERMORE, OUR FINDINGS SUGGEST THAT KIR ACQUISITION CAN BE FURTHER BOLSTERED BY PHARMACOLOGICALLY INHIBITING SIGNALING FROM C-JUN N-TERMINAL KINASE (JNK) THROUGHOUT THE DURATION OF THIS CULTURE SYSTEM. WE HYPOTHESIZE THAT DURING NK CELL DEVELOPMENT, JNK SIGNALING SUPPRESSES KIR ACQUISITION THROUGH MEDIATION OF TRANSCRIPTION FACTOR BINDING AT KIR GENE PROMOTERS. TO ADDRESS OUR HYPOTHESIS, WE WILL: DETERMINE HOW JNK IS ACTIVATED IN OUR CULTURE SYSTEM AND HOW IT MODULATES DISTAL PROMOTER EXPRESSION (AIM 1) AND DETERMINE HOW JNK SIGNALING IMPACTS KIR INTERMEDIATE AND PROXIMAL ELEMENT EXPRESSION (AIM 2). TO ADDRESS THESE AIMS, WE WILL UTILIZE OUR IN-VITRO CULTURE SYSTEM COMBINED WITH ROBUST GENETIC EDITING TECHNIQUES, PCR, AND METHYLATION SEQUENCING AT KIR GENE LOCI. THROUGH THE IDENTIFICATION OF THESE MECHANISMS, WE WILL BE ABLE TO BETTER UNDERSTAND THE MECHANISMS BEHIND KIR REGULATION. FURTHERMORE, THROUGH THE UNDERSTANDING OF THESE MECHANISMS WE HOPE TO UNCOVER NOVEL AVENUES FOR CLINICAL THERAPIES WHICH CAN BE USED TO IMPROVE PATIENT OUTCOMES.