Transgenic CD8αβ co-receptor rescues endogenous TCR function in TCR-transgenic virus-specific T cells

Abstract

BackgroundGenetically engineered virus-specific T cells (VSTs) are a platform for adoptive cell therapy after allogeneic hematopoietic stem cell transplantation. However, redirection to a tumor-associated antigen by the introduction of a transgenic T-cell receptor (TCR) reduces anti-viral activity, thereby impeding the possibility of preventing or treating two distinct complications—malignant relapse and viral infection—with a single cell therapy product. Availability of CD8αβ co-receptor molecules can significantly impact class I restricted T-cell activation, and thus, we interrogated whether transgenic CD8αβ improves anti-viral activity mediated by native VSTs with or without a co-expressed transgenic TCR (TCR8).MethodsOur existing clinical VST manufacturing platform was adapted and validated to engineer TCR+ or TCR8+ VSTs targeting cytomegalovirus and Epstein-Barr virus. Simultaneous anti-viral and anti-tumor function of engineered VSTs was assessed in vitro and in vivo. We used pentamer staining, interferon (IFN)-γ enzyme-linked immunospot (ELISpot), intracellular cytokine staining (ICS), cytotoxicity assays, co-cultures, and cytokine secretion assays for the in vitro characterization. The in vivo anti-tumor function was assessed in a leukemia xenograft mouse model.ResultsBoth transgenic CD8αβ alone and TCR8 had significant impact on the anti-viral function of engineered VSTs, and TCR8+ VSTs had comparable anti-viral activity as non-engineered VSTs as determined by IFN-γ ELISpot, ICS and cytotoxicity assays. TCR8-engineered VSTs had improved anti-tumor function and greater effector cytokine production in vitro, as well as enhanced anti-tumor function against leukemia xenografts in mice.ConclusionIncorporation of transgenic CD8αβ into vectors for TCR-targetable antigens preserves anti-viral activity of TCR transgenic VSTs while simultaneously supporting tumor-directed activity mediated by a transgenic TCR. Our approach may provide clinical benefit in preventing and treating viral infections and malignant relapse post-transplant.

Rapid Generation of TCR and CD8αβ Transgenic Virus Specific T Cells for Immunotherapy of Leukemia

“…For broader applicability of such multi-antigen targeted therapies, the complexity of the production processes needs to be reduced (recently reviewed in (10)). Here, we investigated at small scale if Interferon-g (IFN-g) cytokine capture (CC) selected virus-specific memory T cells from healthy donors are sufficiently enriched and activated to directly proceed to retroviral transduction introducing an HLA-A*02:01 restricted survivin targeted TCR (11) in combination with CD8ab (TCR8) to redirect VSTs to a broad tumor-associated antigen (12,13). We have previously demonstrated that the incorporation of CD8ab as a transgene restores anti-viral activity of TCR transgenic VSTs, and redirects CD4+ T cells to the class I restricted cognate antigen (12,13).…”

“…Here, we investigated at small scale if Interferon-g (IFN-g) cytokine capture (CC) selected virus-specific memory T cells from healthy donors are sufficiently enriched and activated to directly proceed to retroviral transduction introducing an HLA-A*02:01 restricted survivin targeted TCR (11) in combination with CD8ab (TCR8) to redirect VSTs to a broad tumor-associated antigen (12,13). We have previously demonstrated that the incorporation of CD8ab as a transgene restores anti-viral activity of TCR transgenic VSTs, and redirects CD4+ T cells to the class I restricted cognate antigen (12,13). CC is attractive because it is compatible with fully closed production of VSTs independently of donor HLA and can select T cells with diverse TCR repertoires recognizing various immunogenic epitopes (14)(15)(16).…”

Adding Help to an HLA-A*24:02 Tumor-Reactive γδTCR Increases Tumor Control

“…Vγ9Vδ2TCR ( 30 ) and CD8αβ-independent αβTCRs ( 32 ) have been also reported to reprogram CD4 + T cells, which not only have the ability to exert tumor cell killing but also induce maturation of professional antigen-presenting cells. Transfer of CD8αβ in combination with intermediate affinity tumor reactive αβTCR has been reported to support tumor control in vitro and in vivo ( 49 , 50 ), and for high affinity αβTCR with artificial signaling domains adding CD8α alone has been shown to reprogram CD4 + T cells ( 36 ). Within this context, our data show that CD8αα in combination with a natural γδTCR serves as costimulatory receptor, as opposed to the well-described inhibitory function of CD8αα on αβT cells within the context of a natural αβTCR.…”

T-Cell Dysfunction as a Limitation of Adoptive Immunotherapy: Current Concepts and Mitigation Strategies

“…The addition of CD8αβ to the transgenic TCR vector has been used to rescue endogenous MHC class I-restricted anti-viral TCR function [ 192 , 193 ]. These TCR-transgenic VSTs have a predominant central-memory phenotype and their anti-viral reactivity is preserved together with their anti-tumor function [ 194 ]. The insertion of the CD8αβ co-receptor also improved antigen recognition by the TCR/MHC complex, recruitment of the tyrosine kinase Lck to the immune synapse, and proper activation of signaling components for T-cell activation, in addition to allowing CD4 + T cells to recognize MHC class I-restricted antigens [ 195 , 196 ].…”