Samuel Chuah1, Joycelyn Lee2, Yuan Song3, Hyung-Don Kim4, Martin Wasser5, Neslihan A Kaya6, Kyunghye Bang4, Yong Joon Lee7, Seung Hyuck Jeon7, Sheena Suthen1, Shamirah A'Azman1, Gerald Gien1, Chun Jye Lim1, Camillus Chua1, Sharifah Nur Hazirah1, Hong Kai Lee8, Jia Qi Lim9, Tony K H Lim10, Joe Yeong11, Jinmiao Chen8, Eui-Cheol Shin7, Salvatore Albani5, Weiwei Zhai12, Changhoon Yoo4, Haiyan Liu3, Su Pin Choo13, David Tai14, Valerie Chew15. 1. Translational Immunology Institute (TII), SingHealth-DukeNUS Academic Medical Centre, Singapore 169856, Singapore. 2. Division of Medical Oncology, National Cancer Centre Singapore, Singapore 169610, Singapore. 3. Immunology Programme, Life Sciences Institute, Immunology Translational Research Program and Department of Microbiology and Immunology, Yong Loo Lin School of Medicine, National University of Singapore, Singapore 117456, Singapore. 4. Department of Oncology, Asan Medical Center (AMC), University of Ulsan College of Medicine, 88, Olympic-ro 43-gil, Songpa-gu, Seoul 05505, Republic of Korea. 5. Translational Immunology Institute (TII), SingHealth-DukeNUS Academic Medical Centre, Singapore 169856, Singapore; Duke-NUS Medical School, Singapore 169857, Singapore. 6. Genome Institute of Singapore (GIS), Agency for Science, Technology and Research (A∗STAR), Singapore 138672, Singapore; School of Biological Sciences, Nanyang Technological University, Singapore 637551, Singapore. 7. Graduate School of Medical Science and Engineering, Korea Advanced Institute of Science and Technology (KAIST), Daejeon 34141, Republic of Korea. 8. Singapore Immunology Network (SIgN), A∗STAR, Singapore 138648, Singapore. 9. Genome Institute of Singapore (GIS), Agency for Science, Technology and Research (A∗STAR), Singapore 138672, Singapore. 10. Duke-NUS Medical School, Singapore 169857, Singapore; Department of Anatomical Pathology, Singapore General Hospital (SGH), Singapore 169856, Singapore. 11. Duke-NUS Medical School, Singapore 169857, Singapore; Department of Anatomical Pathology, Singapore General Hospital (SGH), Singapore 169856, Singapore; Institute of Molecular and Cell Biology (IMCB), Agency for Science, Technology and Research (A∗STAR), Singapore 138673, Singapore. 12. Genome Institute of Singapore (GIS), Agency for Science, Technology and Research (A∗STAR), Singapore 138672, Singapore; Key Laboratory of Zoological Systematics and Evolution, Institute of Zoology, Chinese Academy of Sciences, Beijing 100107, China; Center for Excellence in Animal Evolution and Genetics, Chinese Academy of Sciences, Kunming 650223, China. 13. Division of Medical Oncology, National Cancer Centre Singapore, Singapore 169610, Singapore; Curie Oncology, Mount Elizabeth Novena Specialist Centre, Singapore 329563, Singapore. 14. Division of Medical Oncology, National Cancer Centre Singapore, Singapore 169610, Singapore. Electronic address: david.tai.w.m@singhealth.com.sg. 15. Translational Immunology Institute (TII), SingHealth-DukeNUS Academic Medical Centre, Singapore 169856, Singapore; Duke-NUS Medical School, Singapore 169857, Singapore. Electronic address: valerie.chew@duke-nus.edu.sg.
Abstract
BACKGROUND & AIMS: While immune checkpoint blockade (ICB) has shown promise in patients with hepatocellular carcinoma (HCC), it is associated with modest response rates and immune-related adverse events (irAEs) are common. In this study, we aimed to decipher immune trajectories and mechanisms of response and/or irAEs in patients with HCC receiving anti-programmed cell death 1 (anti-PD-1) therapy. METHODS: Pre- and on-treatment peripheral blood samples (n = 60) obtained from 32 patients with HCC (Singapore cohort) were analysed by cytometry by time-of-flight and single-cell RNA sequencing, with flow cytometric validation in an independent Korean cohort (n = 29). Mechanistic validation was conducted by bulk RNA sequencing of 20 pre- and on-treatment tumour biopsies and using a murine HCC model treated with different immunotherapeutic combinations. RESULTS: Single-cell analyses identified CXCR3+CD8+ effector memory T (TEM) cells and CD11c+ antigen-presenting cells (APC) as associated with response (p = 0.0004 and 0.0255, respectively), progression-free survival (p = 0.00079 and 0.0015, respectively), and irAEs (p = 0.0034 and 0.0125, respectively) in anti-PD-1-treated patients with HCC. Type-1 conventional dendritic cells were identified as the specific APC associated with response, while 2 immunosuppressive CD14+ myeloid clusters were linked to reduced irAEs. Further analyses of CXCR3+CD8+ TEM cells showed cell-cell interactions specific to response vs. irAEs, from which the anti-PD-1 and anti-TNFR2 combination was harnessed to uncouple these effects, resulting in enhanced response without increased irAEs in a murine HCC model. CONCLUSIONS: This study identifies early predictors of clinical response to anti-PD-1 ICB in patients with HCC and offers mechanistic insights into the immune trajectories of these immune subsets at the interface between response and toxicity. We also propose a new combination immunotherapy for HCC to enhance response without exacerbating irAEs. CLINICAL TRIAL NUMBER: NCT03695952. LAY SUMMARY: Response rates to immune checkpoint blockade (ICB) treatment in hepatocellular carcinoma (HCC) remain modest and adverse events are common. Herein, we identified early predictors of response and gained an in-depth understanding of the immunological mechanisms behind response and adverse events in patients with HCC treated with ICB. We also proposed a new combination immunotherapy for HCC that enhances response without exacerbating adverse events.
BACKGROUND & AIMS: While immune checkpoint blockade (ICB) has shown promise in patients with hepatocellular carcinoma (HCC), it is associated with modest response rates and immune-related adverse events (irAEs) are common. In this study, we aimed to decipher immune trajectories and mechanisms of response and/or irAEs in patients with HCC receiving anti-programmed cell death 1 (anti-PD-1) therapy. METHODS: Pre- and on-treatment peripheral blood samples (n = 60) obtained from 32 patients with HCC (Singapore cohort) were analysed by cytometry by time-of-flight and single-cell RNA sequencing, with flow cytometric validation in an independent Korean cohort (n = 29). Mechanistic validation was conducted by bulk RNA sequencing of 20 pre- and on-treatment tumour biopsies and using a murine HCC model treated with different immunotherapeutic combinations. RESULTS: Single-cell analyses identified CXCR3+CD8+ effector memory T (TEM) cells and CD11c+ antigen-presenting cells (APC) as associated with response (p = 0.0004 and 0.0255, respectively), progression-free survival (p = 0.00079 and 0.0015, respectively), and irAEs (p = 0.0034 and 0.0125, respectively) in anti-PD-1-treated patients with HCC. Type-1 conventional dendritic cells were identified as the specific APC associated with response, while 2 immunosuppressive CD14+ myeloid clusters were linked to reduced irAEs. Further analyses of CXCR3+CD8+ TEM cells showed cell-cell interactions specific to response vs. irAEs, from which the anti-PD-1 and anti-TNFR2 combination was harnessed to uncouple these effects, resulting in enhanced response without increased irAEs in a murine HCC model. CONCLUSIONS: This study identifies early predictors of clinical response to anti-PD-1 ICB in patients with HCC and offers mechanistic insights into the immune trajectories of these immune subsets at the interface between response and toxicity. We also propose a new combination immunotherapy for HCC to enhance response without exacerbating irAEs. CLINICAL TRIAL NUMBER: NCT03695952. LAY SUMMARY: Response rates to immune checkpoint blockade (ICB) treatment in hepatocellular carcinoma (HCC) remain modest and adverse events are common. Herein, we identified early predictors of response and gained an in-depth understanding of the immunological mechanisms behind response and adverse events in patients with HCC treated with ICB. We also proposed a new combination immunotherapy for HCC that enhances response without exacerbating adverse events.