title: Development of T cells carrying two complementary chimeric antigen receptors against glypican-3 and asialoglycoprotein receptor 1 for the treatment of hepatocellular carcinoma.
abstract: Adoptive immunotherapy leveraging chimeric antigen receptor-modified T (CAR-T) cells holds great promise for the treatment of cancer. However, tumor-associated antigens often have low expression levels in normal tissues, which can cause on-target, off-tumor toxicity. Recently, we reported that GPC3-targeted CAR-T cells could eradicate hepatocellular carcinoma (HCC) xenografts in mice. However, it remains unknown whether on-target, off-tumor toxicity can occur. Therefore, we proposed that dual-targeted CAR-T cells co-expressing glypican-3 (GPC3) and asialoglycoprotein receptor 1 (ASGR1) (a liver tissue-specific protein)-targeted CARs featuring CD3ζ and 28BB (containing both CD28 and 4-1BB signaling domains), respectively, may have reduced on-target, off-tumor toxicity. Our results demonstrated that dual-targeted CAR-T cells caused no cytotoxicity to ASGR1+GPC3-tumor cells, but they exhibited a similar cytotoxicity against GPC3+ASGR1-and GPC3+ASGR1+HCC cells in vitro. We found that dual-targeted CAR-T cells showed significantly higher cytokine secretion, proliferation and antiapoptosis ability against tumor cells bearing both antigens than single-targeted CAR-T cells in vitro. Furthermore, the dual-targeted CAR-T cells displayed potent growth suppression activity on GPC3+ASGR1+HCC tumor xenografts, while no obvious growth suppression was seen with single or double antigen-negative tumor xenografts. Additionally, the dual-targeted T cells exerted superior anticancer activity and persistence against single-targeted T cells in two GPC3+ASGR1+HCC xenograft models. Together, T cells carrying two complementary CARs against GPC3 and ASGR1 may reduce the risk of on-target, off-tumor toxicity while maintaining relatively potent antitumor activities on GPC3+ASGR1+HCC.
authors.affiliation: State Key Laboratory of Oncogenes and Related Genes, Shanghai Cancer Institute, Renji Hospital, Shanghai Jiaotong University School of Medicine, No. 25/Ln2200, XieTu Road, Shanghai, China.
authors.forename: Cheng
authors.initials: C
authors.lastname: Chen
authors.affiliation: State Key Laboratory of Oncogenes and Related Genes, Shanghai Cancer Institute, Renji Hospital, Shanghai Jiaotong University School of Medicine, No. 25/Ln2200, XieTu Road, Shanghai, China.
authors.forename: Kesang
authors.initials: K
authors.lastname: Li
authors.affiliation: State Key Laboratory of Oncogenes and Related Genes, Shanghai Cancer Institute, Renji Hospital, Shanghai Jiaotong University School of Medicine, No. 25/Ln2200, XieTu Road, Shanghai, China.
authors.forename: Hua
authors.initials: H
authors.lastname: Jiang
authors.affiliation: State Key Laboratory of Oncogenes and Related Genes, Shanghai Cancer Institute, Renji Hospital, Shanghai Jiaotong University School of Medicine, No. 25/Ln2200, XieTu Road, Shanghai, China.
authors.forename: Fei
authors.initials: F
authors.lastname: Song
authors.affiliation: State Key Laboratory of Oncogenes and Related Genes, Shanghai Cancer Institute, Renji Hospital, Shanghai Jiaotong University School of Medicine, No. 25/Ln2200, XieTu Road, Shanghai, China.
authors.forename: Huiping
authors.initials: H
authors.lastname: Gao
authors.affiliation: State Key Laboratory of Oncogenes and Related Genes, Shanghai Cancer Institute, Renji Hospital, Shanghai Jiaotong University School of Medicine, No. 25/Ln2200, XieTu Road, Shanghai, China.
authors.forename: Xiaorong
authors.initials: X
authors.lastname: Pan
authors.affiliation: State Key Laboratory of Oncogenes and Related Genes, Shanghai Cancer Institute, Renji Hospital, Shanghai Jiaotong University School of Medicine, No. 25/Ln2200, XieTu Road, Shanghai, China.
authors.forename: Bizhi
authors.initials: B
authors.lastname: Shi
authors.affiliation: State Key Laboratory of Oncogenes and Related Genes, Shanghai Cancer Institute, Renji Hospital, Shanghai Jiaotong University School of Medicine, No. 25/Ln2200, XieTu Road, Shanghai, China.
authors.forename: Yanyu
authors.initials: Y
authors.lastname: Bi
authors.affiliation: CARsgen Therapeutics, Shanghai, China.
authors.forename: Huamao
authors.initials: H
authors.lastname: Wang
authors.affiliation: State Key Laboratory of Oncogenes and Related Genes, Shanghai Cancer Institute, Renji Hospital, Shanghai Jiaotong University School of Medicine, No. 25/Ln2200, XieTu Road, Shanghai, China. hywangk@vip.sina.com.|International Cooperation Laboratory on Signal Transduction, Eastern Hepatobiliary Surgery Institute, Second Military Medical University, 225 Changhai Road, Shanghai, China. hywangk@vip.sina.com.|National Center for Liver Cancer, Shanghai, China. hywangk@vip.sina.com.
authors.forename: Hongyang
authors.initials: H
authors.lastname: Wang
authors.affiliation: State Key Laboratory of Oncogenes and Related Genes, Shanghai Cancer Institute, Renji Hospital, Shanghai Jiaotong University School of Medicine, No. 25/Ln2200, XieTu Road, Shanghai, China. zonghaili@shsmu.edu.cn.
authors.forename: Zonghai
authors.initials: Z
authors.lastname: Li
mesh_terms: D000818:Animals; D037263:Asialoglycoprotein Receptor / Q000276:immunology; D019496:Cancer Vaccines / Q000276:immunology*; D006528:Carcinoma, Hepatocellular / Q000276:immunology / Q000628:therapy*; D045744:Cell Line, Tumor; D049109:Cell Proliferation; D016207:Cytokines / Q000378:metabolism; D053673:Glypicans / Q000276:immunology; D006801:Humans; D016219:Immunotherapy, Adoptive / Q000379:methods*; D008113:Liver Neoplasms / Q000276:immunology / Q000628:therapy*; D008213:Lymphocyte Activation; D051379:Mice; D016513:Mice, SCID; D009928:Organ Specificity; D011948:Receptors, Antigen, T-Cell / Q000235:genetics / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism*; D037182:T-Cell Antigen Receptor Specificity; D013601:T-Lymphocytes / Q000502:physiology* / Q000637:transplantation; D023041:Xenograft Model Antitumor Assays
chemical_list: C546927:ASGR1 protein, human; D037263:Asialoglycoprotein Receptor; D019496:Cancer Vaccines; D016207:Cytokines; D053673:Glypicans; D011948:Receptors, Antigen, T-Cell; D011993:Recombinant Fusion Proteins
keywords: Asialoglycoprotein receptor 1; Chimeric antigen receptors; Dual-targeted T cells; Glypican-3; Hepatocellular carcinoma