IMMUNE SYSTEM IMPAIRMENT HAS BEEN DOCUMENTED DURING AND AFTER LONG-DURATION SPACEFLIGHT WHICH COULD INCREASE THE RISK OF CANCER AND LATENT VIRAL REACTIVATION IN EXPLORATION CLASS MISSION CREW. NATURAL KILLER (NK)-CELLS ARE A CRITICAL COMPONENT OF THE IMMUNE SYSTEM AND PLAY A VITAL ROLE IN ANTI-TUMOR SURVEILLANCE AND VIRAL CONTROL. WE HAVE SHOWN THAT THE ANTI-TUMOR ACTIVITY OF NK-CELLS IS IMPAIRED IN ISS CREWMEMBERS DURING THE FLIGHT PHASE OF THE MISSION AND IS ALSO COMPROMISED WHEN NORMAL HUMAN NK-CELLS ARE EXPOSED TO SIMULATED MICROGRAVITY IN VITRO. AS SUCH IT IS CRITICAL TO DETERMINE THE EFFECTS OF MICROGRAVITY ON THE ANTI-TUMOR ACTIVITIES OF NKCELLS IN VIVO TO DETERMINE IF THIS POSES A SERIOUS RISK TO THE HEALTH AND SAFETY OF EXPLORATION CLASS MISSION CREW. MOREOVER IT IS EQUALLY IMPORTANT TO START VALIDATING POTENTIAL IMMUNE SYSTEM COUNTERMEASURES IN PREPARATION FOR LONG DURATION MISSIONS. THE MAIN GOAL OF THIS PROJECT IS TO DETERMINE THE EFFECTS OF SIMULATED MICROGRAVITY ON THE ANTI-TUMOR ACTIVITY OF HUMAN NK-CELLS AND GD-T CELLS IN VIVO USING A HUMANIZED EXPERIMENTAL MOUSE MODEL. WE WILL ALSO TEST THE EFFICACY OF A POTENTIAL COUNTERMEASURE THAT IS CURRENTLY USED IN ALLOGENEIC AND AUTOLOGOUS STEM CELL TRANSPLANT SETTINGS TO BOOST NK-CELL AND GD-T CELL FUNCTION IN VIVO AFTER TRANSPLANT. THIS GOAL WILL BE ACHIEVED THROUGH THE FOLLOWING SPECIFIC AIMS: SA1: DETERMINE THE EFFECT OF SIMULATED MICROGRAVITY ON THE ANTI-TUMOR CAPACITY OF HUMAN NK-CELLS AND GD-T CELLS IN VIVO. EXPERIMENTAL APPROACH: WE WILL USE OUR ESTABLISHED ROTATING CELL CULTURE MODEL TO EXPOSE HUMAN PBMCS EXPANDED NK-CELLS OR EXPANDED GD-T CELLS TO SIMULATED MICROGRAVITY OR CONTROLS (STATIC-1G AND ROTATIONAL CONTROL) PRIOR TO ADOPTIVE TRANSFER INTO HUMAN TUMOR BEARING NOD-SCID-IL2RGNULL (NSG) MICE. THE NSG MICE WILL BE ENGRAFTED WITH A HUMAN LUCIFERASE-LABELLED HUMAN LEUKEMIA CELL LINE (K562) AS BLOOD CANCERS ARE CONSIDERED A RISK FOR LONG DURATION EXPLORATION CREW. THE NSG MICE WILL ALSO BE ENGRAFTED WITH AN NK-CELL SENSITIVE SOLID TUMOR (HUMAN NEUROBLASTOMA; SK-N-SH CELL LINE) WHICH WILL ENABLE US TO STUDY TUMOR INFILTRATES BY ISOLATING TISSUES AND QUANTIFYING THE HUMAN IMMUNE CELLS IN THE TUMOR BY MULTI-PARAMETER FLOW CYTOMETRY. SPECTRAL IMAGING (LAGOX) WILL BE UTILIZED TO MEASURE TUMOR PROGRESSION AND DISSEMINATION OF THE LUCIFERASE-TAGGED HUMAN CANCER CELL LINES. HYPOTHESIS: EXPOSURE TO SIMULATED MICROGRAVITY WILL HAVE AN ADVERSE EFFECT ON THE IN VIVO DYNAMICS OF HUMAN NK AND GD-T CELLS IMPAIRING THEIR ABILITY TO INFILTRATE AND CONTROL HUMAN TUMOR GROWTH IN A HUMANIZED RODENT MODEL. SA2: DETERMINE THE EFFECT OF SYSTEMIC ADMINISTRATION OF IL-2 AND ZOL ON THE FUNCTION OF NK AND GD-T CELLS RESPECTIVELY. EXPERIMENTAL APPROACH: NSG MICE ADOPTIVELY TRANSFERRED WITH HUMAN PBMCS NK-CELLS AND GD-T CELLS WILL BE INFUSED WITH ZOLEDRONIC ACID (ZOL) AND IL-2 AS A POTENTIAL COUNTERMEASURE TO MICROGRAVITY-INDUCED IMMUNE SYSTEM IMPAIRMENT. ZOL AND IL-2 ARE CURRENTLY USED CLINICALLY TO INCREASE THE PROLIFERATION AND FUNCTION OF NK-CELLS AND GD-T CELLS IN BONE MARROW TRANSPLANT PATIENTS TO PREVENT DISEASE RELAPSE. HYPOTHESIS: SYSTEMIC ADMINISTRATION OF IL-2 AND ZOL WILL RESCUE THE IN VIVO ANTI-TUMOR ACTIVITY OF HUMAN NK-CELLS AND GD-T CELLS FOLLOWING THEIR EXPOSURE TO SIMULATED MICROGRAVITY. BY THE END OF THIS PROJECT WE WILL HAVE A BETTER UNDERSTANDING ON THE EFFECTS OF SIMULATED MICROGRAVITY ON ANTI-TUMOR IMMUNITY IN VIVO. WE WILL ALSO MAKE THE FIRST STEPS TO DETERMINE THE EFFICACY OF AN IMMUNE SYSTEM COUNTERMEASURE CURRENTLY USED IN A CLINICAL SETTING TO BOOST THE ANTI-TUMOR AND ANTI-VIRAL ACTIVITY OF HUMAN NK-CELLS AND GD-T CELL IN VIVO WHICH COULD HAVE APPLICATION FOR LONG-DURATION EXPLORATION CLASS MISSION CREW.