Adoptive transfer of T cells transduced with an HLA-independent T-cell receptor against tyrosinase-related protein 2
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Abstract
Adoptive T-cell receptor (TCR) therapy has shown promise in melanoma and other tumor types, but efficacy is often compromised when tumors evade detection by downregulating HLA class I molecules, impairing antigen presentation. Strategies that enable T-cell responses independent of classical HLA-mediated presentation are therefore needed, particularly in solid tumors, in which antigen loss and heterogeneity are major barriers. A promising target for TCR-based immunotherapy of melanoma is Tyrosinase-related protein 2 (TRP2). As a melanocyte differentiation antigen, TRP2 contributes to melanin biosynthesis and may have anti-apoptotic functions, while its restricted expression in normal tissues reduces the risk of off-tumor toxicity. Exploiting naturally occurring, HLA-independent TCRs against TRP2 offers a strategy to circumvent immune evasion through HLA downregulation while maintaining antigen-specific tumor recognition. In this dissertation, the efficacy and safety of a naturally occurring, HLA-independent αβ TCR against human and murine TRP2 were evaluated in vivo using xenograft and syngeneic melanoma models. The TCR, originally identified in a melanoma patient, was previously cloned, codon-optimized, and engineered with murine constant regions. In the xenograft model, NSG mice bearing HLA-deficient Ma-Mel-86b melanoma cells, either with endogenous TRP2 expression or virally transduced to overexpress TRP2, were treated with human anti-TRP2 TCR-T cells. Control mice received T cells transduced with either an HLA-A*02:01-restricted anti-NY-ESO-1 TCR or firefly luciferase alone. In the syngeneic model, C57BL/6 mice carrying murine melanoma cells (B78.H1) stably expressing human TRP2 received murine anti-TRP2 TCR-T cells with endogenous TCR α- and β-chains disrupted by CRISPR/Cas9 editing. In both models, tumor cells expressed NanoLuc luciferase, T cells expressed firefly luciferase, and IL-2 was given to support T-cell activity. Treatment efficacy was assessed by in vivo bioluminescence imaging, histology, and physical examination. Anti-TRP2 TCR-T cells consistently localized to tumors and persisted in vivo, eliminating TRP2-overexpressing tumors in both models while only delaying growth in natural TRP2-expressing tumors. Safety assessments revealed no systemic toxicity or relevant on-target, off-tumor effects. In the syngeneic model, localized vitiligo-like depigmentation occurred without evidence of internal organ toxicity. These results demonstrated the apparent safety and anti-tumor activity of the HLA-independent TRP2-specific TCR but also pointed to limited efficacy against naturally expressing tumors, likely due to inflammation-induced dedifferentiation and antigen downregulation. In vitro, the receptor tyrosine kinase inhibitor tivozanib preserved TRP2 expression under TNF-α- or IFN-γ-induced stress. Future studies will be needed to evaluate whether combining TCR therapy with tivozanib or other drugs that are able to reverse inflammation-induced dedifferentiation can improve control of tumors with natural TRP2 expression in vivo. In summary, this dissertation demonstrates in vivo anti-tumor activity and an apparent favorable safety profile of an HLA-independent TRP2-specific TCR. While tumors overexpressing TRP2 were effectively eliminated, efficacy against tumors with natural TRP2 expression was limited. More broadly, the results illustrated how HLA-independent TCRs bypass classical antigen presentation and may guide the development of safe and more versatile TCR therapies for solid tumors.