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Tailoring the migration capacity of regulatory T cells to enhance their efficacy as a cellular therapy

University of British Columbia

Cell-based therapy with CD4⁺FOXP3⁺ Regulatory T cells (Tregs) has the potential to be an effective treatment to limit organ transplant rejection, graft-versus-host disease and autoimmunity. Yet, challenges for the successful implementation of Treg therapy include difficulties in isolating and expanding homogeneous cell populations while preserving their suppressive function. Further, to be effective, Tregs may need to express homing receptors for migration to inflammatory or tissue sites. The aims of this research were to investigate an alternate source of therapeutic Tregs, to test whether Treg homing capacity could be tailored, and to investigate the migratory capacity of engineered antigen-specific Tregs. To evaluate pediatric thymus – routinely removed during cardiac surgery – as a source of therapeutic Tregs, I isolated thymic Tregs as CD4⁺CD25⁺ cells using different protocols. Large numbers of thymic Tregs could be isolated and expanded in vitro to clinically relevant numbers while maintaining high FOXP3-expression. Thymic Tregs did not secrete pro-inflammatory cytokines and were suppressive in vitro and in a humanized mouse model. Next, I tested whether the addition of cytokines or metabolites to the expansion culture could tailor the homing capacity of thymic Tregs towards inflammatory and specific tissue sites. Addition of IFN-gamma and IL-12 yielded CXCR3⁺TBET⁺FOXP3⁺ Th1-Tregs with migratory capacity to Th1-chemokines in vitro while maintaining their Treg phenotype. Adding retinoic acid to cultures induced expression of the gut-homing receptors alpha4beta7-integrin and CCR9 on Tregs and elevated their gut-relevant suppressive mechanisms. Homing-receptor-tailored Tregs were epigenetically stable even after long-term exposure to inflammatory conditions and they were suppressive in vivo. Finally, I investigated how engineered antigen-specificity affected trafficking of Tregs in vivo, by transducing them with a chimeric antigen receptor (CAR) targeting HLA-A2. I tracked homing of luciferase-transduced HLA-A2-CAR Tregs longitudinally in vivo and found that HLA-A2-CAR Tregs migrated to and proliferated at HLA-A2-expressing skin grafts and could migrate to draining lymph nodes. Tregs recovered from lymph nodes maintained FOXP3- and CAR-expression. Collectively, these studies show that thymic Tregs could be used as an ‘off-the-shelf’ cellular therapy and tailoring Treg homing capacity offers a new and clinically-applicable approach to improving the potency and specificity of Treg therapy.

Text

2019-10-31

Attribution-NonCommercial-NoDerivatives 4.0 International

http://hdl.handle.net/2429/67601

Experimental Medicine

University of British Columbia

UBCV

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