Top

Published in:

01-04-2017 | Original Article

Development of T cells carrying two complementary chimeric antigen receptors against glypican-3 and asialoglycoprotein receptor 1 for the treatment of hepatocellular carcinoma

Authors: Cheng Chen, Kesang Li, Hua Jiang, Fei Song, Huiping Gao, Xiaorong Pan, Bizhi Shi, Yanyu Bi, Huamao Wang, Hongyang Wang, Zonghai Li

Published in: Cancer Immunology, Immunotherapy | Issue 4/2017

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.

Available only for authorised users

Li ZH, Chen C, Li KS, Jiang H, Song F, Gao HP, Pan XR, Shi BZ, Bi YY, Wang HM, Wang HY (2016) Development of T-cells carrying two complementary chimeric antigen receptors against GPC3 and ASGR1 for the treatment of hepatocellular carcinoma. Mol Ther 24(supplement 1):S79. doi:10.1038/mt.2016.78 CrossRef

Cancer IAfRo (2014) World cancer report 2014. WHO, Geneva

Llovet JM, Real MI, Montana X, Planas R, Coll S, Aponte J, Ayuso C, Sala M, Muchart J, Sola R, Rodes J, Bruix J, Barcelona Liver Cancer G (2002) Arterial embolisation or chemoembolisation versus symptomatic treatment in patients with unresectable hepatocellular carcinoma: a randomised controlled trial. Lancet 359(9319):1734–1739. doi:10.1016/S0140-6736(02)08649-X CrossRefPubMed

Llovet JM, Ricci S, Mazzaferro V, Hilgard P, Gane E, Blanc JF, de Oliveira AC, Santoro A, Raoul JL, Forner A, Schwartz M, Porta C, Zeuzem S, Bolondi L, Greten TF, Galle PR, Seitz JF, Borbath I, Haussinger D, Giannaris T, Shan M, Moscovici M, Voliotis D, Bruix J, Group SIS (2008) Sorafenib in advanced hepatocellular carcinoma. N Engl J Med 359(4):378–390. doi:10.1056/NEJMoa0708857 CrossRefPubMed

Kershaw MH, Westwood JA, Parker LL, Wang G, Eshhar Z, Mavroukakis SA, White DE, Wunderlich JR, Canevari S, Rogers-Freezer L, Chen CC, Yang JC, Rosenberg SA, Hwu P (2006) A phase I study on adoptive immunotherapy using gene-modified T cells for ovarian cancer. Clin Cancer Res 12(20 Pt 1):6106–6115. doi:10.1158/1078-0432.CCR-06-1183 CrossRefPubMedPubMedCentral

Park JR, Digiusto DL, Slovak M, Wright C, Naranjo A, Wagner J, Meechoovet HB, Bautista C, Chang WC, Ostberg JR, Jensen MC (2007) Adoptive transfer of chimeric antigen receptor re-directed cytolytic T lymphocyte clones in patients with neuroblastoma. Mol Ther 15(4):825–833. doi:10.1038/sj.mt.6300104 CrossRefPubMed

Carpenito C, Milone MC, Hassan R, Simonet JC, Lakhal M, Suhoski MM, Varela-Rohena A, Haines KM, Heitjan DF, Albelda SM, Carroll RG, Riley JL, Pastan I, June CH (2009) Control of large, established tumor xenografts with genetically retargeted human T cells containing CD28 and CD137 domains. Proc Natl Acad Sci USA 106(9):3360–3365. doi:10.1073/pnas.0813101106 CrossRefPubMedPubMedCentral

Kalos M, Levine BL, Porter DL, Katz S, Grupp SA, Bagg A, June CH (2011) T cells with chimeric antigen receptors have potent antitumor effects and can establish memory in patients with advanced leukemia. Sci Transl Med 3(95):95ra73. doi:10.1126/scitranslmed.3002842 CrossRefPubMedPubMedCentral

Porter DL, Levine BL, Kalos M, Bagg A, June CH (2011) Chimeric antigen receptor-modified T cells in chronic lymphoid leukemia. N Engl J Med 365(8):725–733. doi:10.1056/NEJMoa1103849 CrossRefPubMedPubMedCentral

10.

Gao H, Li K, Tu H, Pan X, Jiang H, Shi B, Kong J, Wang H, Yang S, Gu J, Li Z (2014) Development of T cells redirected to glypican-3 for the treatment of hepatocellular carcinoma. Clin Cancer Res 20(24):6418–6428. doi:10.1158/1078-0432.CCR-14-1170 CrossRefPubMed

11.

Baumhoer D, Tornillo L, Stadlmann S, Roncalli M, Diamantis EK, Terracciano LM (2008) Glypican 3 expression in human nonneoplastic, preneoplastic, and neoplastic tissues: a tissue microarray analysis of 4,387 tissue samples. Am J Clin Pathol 129(6):899–906. doi:10.1309/HCQWPWD50XHD2DW6 CrossRefPubMed

12.

Morgan RA, Yang JC, Kitano M, Dudley ME, Laurencot CM, Rosenberg SA (2010) Case report of a serious adverse event following the administration of T cells transduced with a chimeric antigen receptor recognizing ERBB2. Mol Ther 18(4):843–851. doi:10.1038/mt.2010.24 CrossRefPubMedPubMedCentral

13.

Wilkie S, van Schalkwyk MC, Hobbs S, Davies DM, van der Stegen SJ, Pereira AC, Burbridge SE, Box C, Eccles SA, Maher J (2012) Dual targeting of ErbB2 and MUC1 in breast cancer using chimeric antigen receptors engineered to provide complementary signaling. J Clin Immunol 32(5):1059–1070. doi:10.1007/s10875-012-9689-9 CrossRefPubMed

14.

Kloss CC, Condomines M, Cartellieri M, Bachmann M, Sadelain M (2013) Combinatorial antigen recognition with balanced signaling promotes selective tumor eradication by engineered T cells. Nat Biotechnol 31(1):71–75. doi:10.1038/nbt.2459 CrossRefPubMed

15.

Fiume L, Di Stefano G, Busi C, Mattioli A, Bonino F, Torrani-Cerenzia M, Verme G, Rapicetta M, Bertini M, Gervasi GB (1997) Liver targeting of antiviral nucleoside analogues through the asialoglycoprotein receptor. J Viral Hepat 4(6):363–370CrossRefPubMed

16.

Coulstock E, Sosabowski J, Ovecka M, Prince R, Goodall L, Mudd C, Sepp A, Davies M, Foster J, Burnet J, Dunlevy G, Walker A (2013) Liver-targeting of interferon-alpha with tissue-specific domain antibodies. PLoS ONE 8(2):e57263. doi:10.1371/journal.pone.0057263 CrossRefPubMedPubMedCentral

17.

Mu H, Lin KX, Zhao H, Xing S, Li C, Liu F, Lu HZ, Zhang Z, Sun YL, Yan XY, Cai JQ, Zhao XH (2014) Identification of biomarkers for hepatocellular carcinoma by semiquantitative immunocytochemistry. World J Gastroenterol 20(19):5826–5838. doi:10.3748/wjg.v20.i19.5826 CrossRefPubMedPubMedCentral

18.

Witzigmann D, Quagliata L, Schenk SH, Quintavalle C, Terracciano LM, Huwyler J (2016) Variable asialoglycoprotein receptor 1 expression in liver disease: implications for therapeutic intervention. Hepatol Res 46(7):686–696. doi:10.1111/hepr.12599 CrossRefPubMed

19.

Szymczak AL, Workman CJ, Wang Y, Vignali KM, Dilioglou S, Vanin EF, Vignali DA (2004) Correction of multi-gene deficiency in vivo using a single ‘self-cleaving’ 2A peptide-based retroviral vector. Nat Biotechnol 22(5):589–594. doi:10.1038/nbt957 CrossRefPubMed

20.

Wang H, Zhou M, Shi B, Zhang Q, Jiang H, Sun Y, Liu J, Zhou K, Yao M, Gu J, Yang S, Mao Y, Li Z (2011) Identification of an exon 4-deletion variant of epidermal growth factor receptor with increased metastasis-promoting capacity. Neoplasia 13(5):461–471CrossRefPubMedPubMedCentral

21.

Kuroda H, Kutner RH, Bazan NG, Reiser J (2009) Simplified lentivirus vector production in protein-free media using polyethylenimine-mediated transfection. J Virol Methods 157(2):113–121. doi:10.1016/j.jviromet.2008.11.021 CrossRefPubMed

22.

Li K, Pan X, Bi Y, Xu W, Chen C, Gao H, Shi B, Jiang H, Yang S, Jiang L, Li Z (2016) Adoptive immunotherapy using T lymphocytes redirected to glypican-3 for the treatment of lung squamous cell carcinoma. Oncotarget 7(3):2496–2507. doi:10.18632/oncotarget.6595 PubMed

23.

Goldstein NI, Prewett M, Zuklys K, Rockwell P, Mendelsohn J (1995) Biological efficacy of a chimeric antibody to the epidermal growth factor receptor in a human tumor xenograft model. Clin Cancer Res 1(11):1311–1318PubMed

24.

Shi B, Abrams M, Sepp-Lorenzino L (2013) Expression of asialoglycoprotein receptor 1 in human hepatocellular carcinoma. J Histochem Cytochem 61(12):901–909. doi:10.1369/0022155413503662 CrossRefPubMedPubMedCentral

25.

Di Tommaso L, Destro A, Seok JY, Balladore E, Terracciano L, Sangiovanni A, Iavarone M, Colombo M, Jang JJ, Yu E, Jin SY, Morenghi E, Park YN, Roncalli M (2009) The application of markers (HSP70 GPC3 and GS) in liver biopsies is useful for detection of hepatocellular carcinoma. J Hepatol 50(4):746–754. doi:10.1016/j.jhep.2008.11.014 CrossRefPubMed

26.

Li J, Gao JZ, Du JL, Wei LX (2014) Prognostic and clinicopathological significance of glypican-3 overexpression in hepatocellular carcinoma: a meta-analysis. World J Gastroenterol 20(20):6336–6344. doi:10.3748/wjg.v20.i20.6336 CrossRefPubMedPubMedCentral

27.

Hass HG, Jobst J, Scheurlen M, Vogel U, Nehls O (2015) Gene expression analysis for evaluation of potential biomarkers in hepatocellular carcinoma. Anticancer Res 35(4):2021–2028PubMed

28.

Trere D, Fiume L, De Giorgi LB, Di Stefano G, Migaldi M, Derenzini M (1999) The asialoglycoprotein receptor in human hepatocellular carcinomas: its expression on proliferating cells. Br J Cancer 81(3):404–408. doi:10.1038/sj.bjc.6690708 CrossRefPubMedPubMedCentral

29.

Shindo G, Endo T, Onda M, Goto S, Miyamoto Y, Kaneko T (2013) Is the CD4/CD8 ratio an effective indicator for clinical estimation of adoptive immunotherapy for cancer treatment? JCT 4(8):1382–1390. doi:10.4236/jct.2013.48164 CrossRef

30.

Yang S, Gattinoni L, Liu F, Ji Y, Yu Z, Restifo NP, Rosenberg SA, Morgan RA (2011) In vitro generated anti-tumor T lymphocytes exhibit distinct subsets mimicking in vivo antigen-experienced cells. Cancer Immunol Immunother 60(5):739–749. doi:10.1007/s00262-011-0977-7 CrossRefPubMedPubMedCentral

31.

Sallusto F, Geginat J, Lanzavecchia A (2004) Central memory and effector memory T cell subsets: function, generation, and maintenance. Annu Rev Immunol 22:745–763. doi:10.1146/annurev.immunol.22.012703.104702 CrossRefPubMed

32.

Croft M (2003) Costimulation of T cells by OX40, 4-1BB, and CD27. Cytokine Growth Factor Rev 14(3–4):265–273CrossRefPubMed

33.

Rudd CE, Schneider H (2003) Unifying concepts in CD28, ICOS and CTLA4 co-receptor signalling. Nat Rev Immunol 3(7):544–556. doi:10.1038/nri1131 CrossRefPubMed

34.

Watts TH (2005) TNF/TNFR family members in costimulation of T cell responses. Annu Rev Immunol 23:23–68. doi:10.1146/annurev.immunol.23.021704.115839 CrossRefPubMed

35.

Kohn DB, Dotti G, Brentjens R, Savoldo B, Jensen M, Cooper LJ, June CH, Rosenberg S, Sadelain M, Heslop HE (2011) CARs on track in the clinic. Mol Ther 19(3):432–438. doi:10.1038/mt.2011.1 CrossRefPubMedCentral

36.

Stauss HJ, Morris EC (2013) Immunotherapy with gene-modified T cells: limiting side effects provides new challenges. Gene Ther 20(11):1029–1032. doi:10.1038/gt.2013.34 CrossRefPubMed

37.

Lanitis E, Poussin M, Klattenhoff AW, Song D, Sandaltzopoulos R, June CH, Powell DJ Jr (2013) Chimeric antigen receptor T Cells with dissociated signaling domains exhibit focused antitumor activity with reduced potential for toxicity in vivo. Cancer Immunol Res 1(1):43–53. doi:10.1158/2326-6066.CIR-13-0008 CrossRefPubMedPubMedCentral

38.

Takezawa R, Shinzawa K, Watanabe Y, Akaike T (1993) Determination of mouse major asialoglycoprotein receptor cDNA sequence. Biochim Biophys Acta 1172(1):220–222CrossRefPubMed

39.

Soukharev S, Berlin WK, Hanover JA, Bethke B, Sauer B (2000) Organization of the mouse ASGR1 gene encoding the major subunit of the hepatic asialoglycoprotein receptor. Gene 241(2):233–240CrossRefPubMed

40.

Nakano K, Yoshino T, Nezu J, Tsunoda H, Igawa T, Konishi H, Tanaka M, Sugo I, Kawai S, Ishiguro T, Kinoshita Y (2007) Anti-glypican 3 antibody. US 2007/0190599 A l [United States Patent Application Publication]

41.

Weijtens ME, Hart EH, Bolhuis RL (2000) Functional balance between T cell chimeric receptor density and tumor associated antigen density: CTL mediated cytolysis and lymphokine production. Gene Ther 7(1):35–42. doi:10.1038/sj.gt.3301051 CrossRefPubMed

42.

Caruso HG, Hurton LV, Najjar A, Rushworth D, Ang S, Olivares S, Mi T, Switzer K, Singh H, Huls H, Lee DA, Heimberger AB, Champlin RE, Cooper LJ (2015) Tuning sensitivity of CAR to EGFR density limits recognition of normal tissue while maintaining potent antitumor activity. Cancer Res 75(17):3505–3518. doi:10.1158/0008-5472.CAN-15-0139 CrossRefPubMedPubMedCentral

43.

Long AH, Haso WM, Shern JF, Wanhainen KM, Murgai M, Ingaramo M, Smith JP, Walker AJ, Kohler ME, Venkateshwara VR, Kaplan RN, Patterson GH, Fry TJ, Orentas RJ, Mackall CL (2015) 4-1BB costimulation ameliorates T cell exhaustion induced by tonic signaling of chimeric antigen receptors. Nat Med 21(6):581–590. doi:10.1038/nm.3838 CrossRefPubMedPubMedCentral

44.

Holliger P, Hudson PJ (2005) Engineered antibody fragments and the rise of single domains. Nat Biotechnol 23(9):1126–1136. doi:10.1038/nbt1142 CrossRefPubMed

Title: Development of T cells carrying two complementary chimeric antigen receptors against glypican-3 and asialoglycoprotein receptor 1 for the treatment of hepatocellular carcinoma
Authors: Cheng Chen
Kesang Li
Hua Jiang
Fei Song
Huiping Gao
Xiaorong Pan
Bizhi Shi
Yanyu Bi
Huamao Wang
Hongyang Wang
Zonghai Li
Publication date: 01-04-2017
Publisher: Springer Berlin Heidelberg
Published in: Cancer Immunology, Immunotherapy / Issue 4/2017
Print ISSN: 0340-7004
Electronic ISSN: 1432-0851
DOI: https://doi.org/10.1007/s00262-016-1949-8

Webinar | 19-02-2024 | 17:30 (CET)

Keynote webinar | Spotlight on antibody–drug conjugates in cancer

Watch now

Antibody–drug conjugates (ADCs) are novel agents that have shown promise across multiple tumor types. Explore the current landscape of ADCs in breast and lung cancer with our experts, and gain insights into the mechanism of action, key clinical trials data, existing challenges, and future directions.

Dr. Véronique Diéras

Prof. Fabrice Barlesi

Developed by: Springer Medicine

At a glance: The ONWARDS insulin icodec trials

Springer Medicine

Abstract

Please log in to get access to this content

Other articles of this Issue 4/2017

Inclusion of BLIMP-1+ effector regulatory T cells improves the Immunoscore in a cohort of New Zealand colorectal cancer patients: a pilot study

Regulation of PD-L1 expression on murine tumor-associated monocytes and macrophages by locally produced TNF-α

In memory of Professor Enrico Mihich: Editor-in-Chief of Cancer Immunology, Immunotherapy 1982–2012

Combined treatment with ipilimumab and intratumoral interleukin-2 in pretreated patients with stage IV melanoma—safety and efficacy in a phase II study

Intratumoral Th2 predisposition combines with an increased Th1 functional phenotype in clinical response to intravesical BCG in bladder cancer

The kinase inhibitors R406 and GS-9973 impair T cell functions and macrophage-mediated anti-tumor activity of rituximab in chronic lymphocytic leukemia patients

Keynote webinar | Spotlight on antibody–drug conjugates in cancer