Electronic Thesis/Dissertation

 

On Natural Killer Cell Homing: Approaches to Improve Autologous NK Cell Immunotherapy Targeting Bone Marrow Malignancies Público Deposited

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Significant advances in the field of cancer immunotherapy have allowed for disease remission and improved quality of life for subsets of patients. The therapeutic potential of autologous ex vivo expanded natural killer (NK) cells to treat patients with hematological malignancies is currently being evaluated in the preclinical and clinical trial settings. NK cells are powerful cytotoxic lymphocytes and require close proximity to malignant cells for proper recognition and cytolytic reaction. Thus, the ability of infused NK cells to effectively treat patients with bone marrow (BM)-resident tumor may rely on efficient infiltration of NK cells into BM compartments following intravenous infusion. The research presented here provides the first evidence that ex vivo cellular cultivation leads to differential regulation of chemotactic receptor expression. This observed shift may guide NK cells from the circulation into liver and other inflammatory environments, ultimately deterring the BM trafficking of adoptively transferred NK cells. A comprehensive hypothesis is offered to addresses several biological and technical factors that may contribute to these phenotypic shifts. Importantly, these data reveal gene transcription and surface expression of CXCR4, a critical chemokine receptor for directing cellular migration into BM, is massively downregulated due to the ex vivo cultivation of NK cells. WHIM syndrome, a rare immunodeficiency that is known to be caused by a single gain-of-function mutation in CXCR4, manifests due to the sequestration of mature lymphocytes in the BM. RNA-sequencing analysis of ex vivo expanded NK cells that were isolated from WHIM syndrome patients revealed a vastly different gene transcription profile compared to expanded NK cells from healthy donors. In depth analysis of the data presented here provide novel insight into the unforeseen complex biology of this rare disease. Importantly, following infusion into mice, NK cells from these patients have an advantage for trafficking into BM over NK cells from healthy donors. In support of the objective to develop clinically feasible methods to genetically and molecularly modify NK cells for improved trafficking into BM niches, mRNA transfection of ex vivo expanded NK cells with gain-of-function CXCR4 variant mRNA was utilized to improve the BM homing of NK cells following infusion into mice. Furthermore, hematopoietic stem cells and multiple myeloma cells express E-selectin ligand carbohydrates (i.e., sialyl Lewis X moieties) that bind to BM E-selectin to initiate trans-migration into BM compartments. Installation of sialyl Lewis X onto the surface of ex vivo expanded NK cell by in vitro fucosylation was used to facilitate BM homing of infused autologous NK cells in the Rhesus macaque, however this did not result in improved BM trafficking. The collective findings presented here synthesize efforts to understand variables that may contribute to suboptimal BM infiltration of NK cells following adoptive transfer into patients and to employ novel techniques to improve autologous NK cell-based therapies.

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