On Dec. 23rd, 2021, the Zhang Lab from Biomedical Pioneering Innovation Center (BIOPIC), Beijing Advanced Innovation Center for Genomics (ICG), School of Life Sciences in Peking University, collaborating with Weidong Han’s team from Chinese PLA General Hospital, published a paper entitled “Temporal single-cell tracing reveals clonal revival and expansion of precursor exhausted T cells during anti-PD-1 therapy in lung cancer” in Nature Cancer, which proposes the concept of clonal revival and provides new insights into mechanisms underlying anti-PD-1 treatment.

　　Immune-checkpoint blockade (ICB) is able to potentiate antitumor immunity by disrupting coinhibitory T-cell signaling and represents a paradigm shift in cancer treatment. However, the remarkable responses to such therapies are limited to a minority of patients with lung cancer, highlighting the need for additional therapeutic strategies. To facilitate the rational design of next-generation immunotherapies, it is of critical importance to first understand the molecular and immunologic mechanisms underlying ICB-induced tumor rejection. The researchers performed temporal single-cell RNA and paired T-cell receptor sequencing on tumor biopsies from lung cancer patients following PD-1-based therapies, and systematically investigated the differences between responders and non-responders (Fig. 1).

　　Figure 1: Scheme of the overall study design and major findings

　　Tumor-infiltrating T cells can be reactive not only to tumor antigens but also to a wide range of epitopes unrelated to cancer (such as human influenza virus) [1]. Thus, one challenge in the analysis is how to accurately identify tumor-reactive T cells and further analyze their temporal behaviors following treatment. Emerging evidence demonstrates that exhausted T (Tex) cells in tumors are specifically derived from tumor-specific T cells, whereas T cells responsible for acute infections do not give rise to Tex cells [2-4]. Thus, a Tex subset could be used as a proxy for a tumor-reactive T-cell compartment [5].

　　Accordingly, the researchers identified the Tex cell subset first by unsupervised clustering and further categorized all CD8 T cells into two groups based on whether they shared TCR sequences with Tex cells. As aforementioned, cells sharing identical TCRs with Tex cells were considered as tumor-reactive, whereas the remaining cells fell into the bystander compartment. The researchers observed increased levels of precursor exhausted T (Texp) cells in responsive tumors after treatment, characterized by low expression of coinhibitory molecules and high expression of GZMK. By contrast, nonresponsive tumors failed to accumulate Texp cells.

　　Three possible modes could explain the formation of Texp cells: (1) reprogramming of the terminal Tex subset into Texp cells by treatment; (2) local expansion of pre-existing Texp cells; and (3) replenishment with peripheral T cell. The researches ruled out the first of these and demonstrated the latter two modes. Previous studies reported that PD-1 pathway blockade could reverse the dysfunctional state of terminally exhausted T cells during chronic viral infection, presenting a widely presumed mechanism of ICB treatment. However, with concentrated efforts in addressing this question, emerging evidence showed that the epigenetic stability of terminal Tex cells is difficult to alter and could limit this reinvigoration [6], and this study further supported this notion in human cancer.

　　Howard Chang’s group from the Stanford University previously proposed the concept of clonal replacement, demonstrating that clones with new clonotypes appeared after treatment, with clonotypes different from those detected in pre-treatment tumors. In this study, the researches found that both new and pre-existing tumor-reactive clones could be replenished with peripheral T cells and thus termed this phenomenon “clonal revival”. This study provides insights into mechanisms underlying PD-1-based therapies, implicating clonal revival and expansion of Texp cells as steps to improve NSCLC treatment.

　　Figure 2: Clonal revival

　　Ph.D. candidate Baolin Liu, Dr. Xueda Hu and Dr. Kaichao Feng are the co-first authors of the paper. Prof. Zemin Zhang and Prof. Weidong Han are the co-corresponding authors of the paper. The project was funded by the National Natural Science Foundation of China, the Beijing Natural Science Foundation, the Beijing Advanced Innovation Centre for Genomics at Peking University and the Boehringer Ingelheim GmbH.

　　Link: https://www.nature.com/articles/s43018-021-00292-8

　　References:

　　[1] Simoni et al., Bystander CD8+ T cells are abundant and phenotypically distinct in human tumour infiltrates, Nature (2018).

　　[2] Caushi et al., Transcriptional programs of neoantigen-specific TIL in anti-PD-1-treated lung cancers, Nature (2021).

　　[3] Oliveira et al., Phenotype, specificity and avidity of antitumour CD8+ T cells in melanoma, Nature (2021).

　　[4] Ahmadzadeh et al. Tumor antigen-specific CD8 T cells infiltrating the tumor express high levels of PD-1 and are functionally impaired. Blood (2009).

　　[5] van der Leun et al., CD8 + T cell states in human cancer: insights from single-cell analysis, Nat Rev Cancer (2020).

　　[6] Pauken et al., Epigenetic stability of exhausted T cells limits durability of reinvigoration by PD-1 blockade, Science (2016).

　　[7] Yost et al., Clonal replacement of tumor-specific T cells following PD-1 blockade, Nat. Med. (2019).