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Single-cell transcriptome analysis of the heterogeneous effects of differential expression of tumor PD-L1 on responding TCR-T cells.

ABSTRACT: Rationale: TCR-T cell therapy plays a critical role in the treatment of malignant cancers. However, it is unclear how TCR-T cells are affected by PD-L1 molecule in the tumor environment. We performed an in-depth evaluation on how differential expressions of tumor PD-L1 can affect the functionality of T cells. Methods: We used MART-1-specific TCR-T cells (TCR-T_MART-1), stimulated with MART-1_27-35 peptide-loaded MEL-526 tumor cells, expressing different proportions of PD-L1, to perform cellular assays and high-throughput single-cell RNA sequencing. Results: Different clusters of activated or cytotoxic TCR-T_MART-1 responded divergently when stimulated with tumor cells expressing different percentages of PD-L1 expression. Compared to control T cells, TCR-T_MART-1 were more sensitive to exhaustion, and secreted not only pro-inflammatory cytokines but also anti-inflammatory cytokines with increasing proportions of PD-L1⁺ tumor cells. The gene profiles of chemokines were modified by increased expression of tumor PD-L1, which concurrently downregulated pro-inflammatory and anti-inflammatory transcription factors. Furthermore, increased expression of tumor PD-L1 showed distinct effects on different inhibitory checkpoint molecules (ICMs). In addition, there was a limited correlation between the enrichment of cell death signaling in tumor cells and T cells and increased tumor PD-L1 expression. Conclusion: Overall, though the effector functionality of TCR-T cells was suppressed by increased expression percentages of tumor PD-L1 in vitro, scRNA-seq profiles revealed that both the anti-inflammatory and pro-inflammatory responses were triggered by a higher expression of tumor PD-L1. This suggests that the sole blockade of tumor PD-L1 might inhibit not only the anti-inflammatory response but also the pro-inflammatory response in the complicated tumor microenvironment. Thus, the outcome of PD-L1 intervention may depend on the final balance among the highly dynamic and heterogeneous immune regulatory circuits.

SUBMITTER: Ding R

PROVIDER: S-EPMC7978322 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:Although programmed cell death-1/programmed death-ligand 1 (PD-L1) inhibitors show remarkable antitumor activity, a large portion of patients with cancer, even those with high PD-L1-expressing tumors, do not respond to their effects. Most PD-L1 inhibitors contain modified fragment crystallizable region (Fc) receptor binding sites to prevent antibody-dependent cellular cytotoxicity (ADCC) against PD-L1-expressing non-tumor cells. However, natural killer (NK) cells have specific antitumor activity in the presence of tumor-targeting antibody through ADCC, which could enhance NK cell-induced cytotoxicity. We evaluated the antitumor efficacy of ADCC via anti-PD-L1 monoclonal antibodies (mAbs) and NK cells against several PD-L1-positive cancer cell lines. Various cancer cell lines were used as target cell lines. Surface PD-L1 expression was analyzed by flow cytometry. IMC-001 and anti-hPD-L1-hIgG1 were tested as anti-PD-L1 mAbs with ADCC and atezolizumab as an anti-PD-L1 mAb without ADCC. NK cell cytotoxicity was measured by 51Cr-release assay and CD107a degranulation assay. Also, live cell imaging was performed to evaluate cytotoxicity in a single-cell level. NK-92-CD16 (CD16-transduced NK-92 cell line) and peripheral blood mononuclear cells from healthy donors, respectively, were used as an effector cell. FcγRIIIa (CD16a)-V158F genotyping was performed for healthy donors. We demonstrated that the cytotoxicity of NK-92-CD16 cells toward PD-L1-positive cancer cell lines was significantly enhanced in the presence of anti-PD-L1 mAb with ADCC. We also noted a significant increase in primary human NK cell cytotoxicity against PD-L1-positive human cancer cells when cocultured with anti-PD-L1 mAb with ADCC. Moreover, NK cells expressing a FCGR3A high-affinity genotype displayed higher anti-PD-L1 mAb-mediated ADCC lysis of tumor cells than donors with a low-affinity genotype. These results suggest that NK cells induce an ADCC response in combination with anti-PD-L1 mAbs, which helps promote ADCC antitumor activity against PD-L1-positive tumors. This study provides support for NK cell immunotherapy against high PD-L1-expressing tumors in combination with ADCC through anti-PD-L1 mAbs.

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Single-cell transcriptome analysis of the heterogeneous effects of differential expression of tumor PD-L1 on responding TCR-T cells.

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