Abstract A047: Tumor specific γδ T cells expand and respond to PD-1 blockade
Authors: Lien Scott C. , Ly Dalam , Yang S.Y. Cindy , Wang Ben X. , Paul Michael St. , Gadalla Ramy , Noamani Babak , Boross-Harmer Sarah , Pugh Trevor J. , Spreafico Anna , Hirano Naoto , Razak Albiruni R.A. , Ohashi Pamela S. 
Published: December 2023

Abstract
With the breakthrough of checkpoint blockade, immunotherapies targeting PD-1/PD-L1 have had remarkable success in the clinic and are now widely used to treat a variety of malignancies. While the majority of research on T cell exhaustion and PD-1 blockade has been focused on conventional αβ T cells, the contribution of innate-like T cells such as γδ T cells to PD-1 blockade is not clear. γδ T cells are generally not restricted to conventional MHC-peptide molecules and can recognize a diverse range of ligands including phosphoantigens presented on butyrophilins and MHC class I-like family members MR1, and CD1 isoforms. Using flow cytometry, we evaluated the γδ T cell responses in tumor biopsies and peripheral blood from six MCC patients treated with pembrolizumab. Furthermore, bulk and single cell γδ TCR sequencing was used to measure diversity and clonal expansion in the blood upon PD-1 blockade. TCRs were cloned and expressed in Jurkat cells to screen for reactivity against tumor cells lines. Finally, we used CRISPR-Cas9 to knockout known γδ T cell ligands to identify potential tumor antigen recognized by γδ T cells. We identified a MCC patient who experienced a complete response to pembrolizumab treatment and had a ten-fold expansion of γδ T cells in their tumor biopsies. Tumor-infiltrating γδ T cells expressed high levels of inhibitory receptors PD-1 and TIGIT. Furthermore, γδ T cells characterized in peripheral blood were predominantly Vδ1 cells and had increased Ki-67 expression upon pembrolizumab treatment. From TCR sequencing of peripheral blood, we observed the emergence of a dominant γδ T cell clonotype that was also found in the tumor. Upon TCR gene transfer into Jurkat 76 cells, we identified a γδ TCR that was able to recognize multiple Merkel cancer cell lines. CRISPR knockout of B2M in tumor cells still retained γδ TCR reactivity, suggesting that γδ TCR recognition is independent of B2M expression. Together, these results show innate-like T cells such as γδ T cells also have the capacity to respond to checkpoint blockade. As mutations in antigen presentation can be a resistance mechanism to PD-1 blockade, our study provides the rationale to utilize γδ T cells for treating cancer variants that have escaped the immune system and warrants further investigation of the interplay between γδ T cells and checkpoint blockade.
Citation Format: Scott C. Lien, Dalam Ly, S.Y. Cindy Yang, Ben X. Wang, Michael St. Paul, Ramy Gadalla, Babak Noamani, Sarah Boross-Harmer, Trevor J. Pugh, Anna Spreafico, Naoto Hirano, Albiruni R.A. Razak, Pamela S. Ohashi. Tumor specific γδ T cells expand and respond to PD-1 blockade [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: Tumor Immunology and Immunotherapy; 2023 Oct 1-4; Toronto, Ontario, Canada. Philadelphia (PA): AACR; Cancer Immunol Res 2023;11(12 Suppl):Abstract nr A047.