Delineation of an immunosuppressive gradient in hepatocellular carcinoma using high-dimensional proteomic and transcriptomic analyses

• Published in: Proceedings of the National Academy of Sciences - PNAS Vol. 114; no. 29; pp. E5900 - E5909
• Main Authors: Chew, Valerie, Lai, Liyun, Pan, Lu, Lim, Chun Jye, Li, Juntao, Ong, Raymond, Chua, Camillus, Leong, Jing Yao, Lim, Kiat Hon, Toh, Han Chong, Lee, Ser Yee, Chan, Chung Yip, Goh, Brian K. P., Chung, Alexander, Chow, Pierce K. H., Albani, Salvatore
• Format: Journal Article
• Language: English
• Published: United States National Academy of Sciences 18.07.2017
• Series: PNAS Plus
• Subjects: Antigen (tumor-associated); Antigens; Biological Sciences; CCL20 protein; CCR6 protein; CD223 antigen; CD4 antigen; CD8 antigen; Cell activation; Compartments; Composition effects; CXCL10 protein; CXCR3 protein; Cytometry; Dimensional analysis; Exhaustion; Foxp3 protein; Hepatocellular carcinoma; Immune system; Immunofluorescence; Immunological memory; Immunology; Immunoregulation; Immunosuppression; Immunotherapy; Leukocytes; Liver cancer; Lymphocytes; Lymphocytes T; Macrophages; Memory cells; Natural killer cells; PD-1 protein; PNAS Plus; Proteomics; Recruitment; Tissues; Tumor cells; Tumors; regulatory T cells; tumor microenvironment; CyTOF; hepatocellular carcinoma; resident memory T cells
• ISSN: 0027-8424; 1091-6490
• DOI: 10.1073/pnas.1706559114

The recent development of immunotherapy as a cancer treatment has proved effective over recent years, but the precise dynamics between the tumor microenvironment (TME), nontumor microenvironment (NTME), and the systemic immune system remain elusive. Here, we interrogated these compartments in hepatocellular carcinoma (HCC) using high-dimensional proteomic and transcriptomic analyses. By time-of-flight mass cytometry, we found that the TME was enriched in regulatory T cells (Tregs), tissue resident memory CD8⁺ T cells (TRMs), resident natural killer cells (NKRs), and tumor-associated macrophages (TAMs). This finding was also validated with immunofluorescence staining on Foxp3⁺CD4⁺ and PD-1⁺ CD8⁺ T cells. Interestingly, Tregs and TRMs isolated from the TME expressed multiple markers for T-cell exhaustion, including PD-1, Lag-3, and Tim-3 compared with Tregs and TRMs isolated from the NTME. We found PD-1⁺ TRMs were the predominant T-cell subset responsive to anti–PD-1 treatment and significantly reduced in number with increasing HCC tumor progression. Furthermore, T-bet was identified as a key transcription factor, negatively correlated with PD-1 expression on memory CD8⁺ T cells, and the PD-1:T-bet ratio increased upon exposure to tumor antigens. Finally, transcriptomic analysis of tumor and adjacent nontumor tissues identified a chemotactic gradient for recruitment of TAMs and NKRs via CXCR3/CXCL10 and CCR6/CCL20 pathways, respectively. Taken together, these data confirm the existence of an immunosuppressive gradient across the TME, NTME, and peripheral blood in primary HCC that manipulates the activation status of tumor-infiltrating leukocytes and renders them immunocompromised against tumor cells. By understanding the immunologic composition of this gradient, more effective immunotherapeutics for HCC may be designed.