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Lauener, Madeline

University of British Columbia

Chronic graft-versus-host disease (cGvHD) occurs in ~25% of pediatric and ~60% of adult Hematopoietic Stem Cell Transplant (HSCT) patients and is a major cause of mortality. In large study cohorts (~500 patients), our group has shown a CD56ᵇʳⁱᵍʰᵗPerforin⁻ Natural Killer population, consistent with described regulatory NK cells (NKreg), to strongly associate with protection from cGvHD development. Therefore, I hypothesized that NKreg cells suppress cGvHD through inflammatory cell inhibition, and these cells can be distinguished as a unique cell subset which can be expanded. In my project, I characterized NKreg cells according to transcriptome, proteome, metabolome, phenotype, and function; evaluated the in vitro suppressive effect of NKreg cells on cGvHD immune populations; and optimized the expansion of functional NKreg cells in vitro. Transcriptome analysis revealed that NKreg cells express high RNA levels of Granzyme K, IL-7R, GPR183, RANK, GM-CSFR, CD62L, and LEF1. These NKreg cells can be isolated by sorting for CD56⁺CD16⁻ NK cells, and this population can suppress allogeneic CD4⁺ T cells, but not Treg cells, CD8⁺ T cells, CD56ᵈⁱᵐCD16⁺ cytolytic NK cells, or B cells, through a contact-dependent mechanism, which is partially reliant on the PD-1, LAG-3, and/or TRAIL inhibitory pathways. Additionally, NKreg cells demonstrate a lack of cytotoxicity towards K562, MOLT-4, and Jurkat leukemic cells, and CD4⁺ T cells. Further, I determined two alternative approaches for obtaining clinically relevant numbers of NKreg cells, achieving up to 300-fold expansion, with use of IL-2, TGF-β1, and rapamycin for cells from peripheral blood, or approximately 350-fold expansion when expanding and differentiating NKreg cells from cord blood-hematopoietic stem and progenitor cells. In further characterization of the expanded NKreg cells, I describe their metabolism to be consistent with cytolytic NK cells, though they also secrete regulatory cytokines, including IL-10, TGF-β1, adenosine, and TRAIL. This work has established that a CD56ᵇʳⁱᵍʰᵗCD16⁻ NKreg population associates with protection from cGvHD development and has several unique characteristics. Further investigations may utilize these findings to evaluate functional efficacy in in vivo models of GvHD and translate the expansion protocol to one which is clinically applicable, with the goal of using these cells as a cGvHD cellular therapy.

Text

2024-07-02

Attribution-NonCommercial-NoDerivatives 4.0 International

http://hdl.handle.net/2429/88549

Pathology and Laboratory Medicine

University of British Columbia

UBCV

http://creativecommons.org/licenses/by-nc-nd/4.0/