-Author name in bold denotes the presenting author
-Asterisk * with author name denotes a Non-ASH member
Clinically Relevant Abstract denotes an abstract that is clinically relevant.

PhD Trainee denotes that this is a recommended PHD Trainee Session.

Ticketed Session denotes that this is a ticketed session.

1967 Development of Anti-CD3 Chimeric Antigen Receptor (CAR)-T Cells for Allogeneic Cell Therapy of Peripheral T-Cell Lymphoma (PTCL)

Program: Oral and Poster Abstracts
Session: 703. Cellular Immunotherapies: Basic and Translational: Poster I
Hematology Disease Topics & Pathways:
Research, Biological therapies, Translational Research, Lymphomas, Chimeric Antigen Receptor (CAR)-T Cell Therapies, T Cell lymphoma, Diseases, Therapies, Lymphoid Malignancies
Saturday, December 10, 2022, 5:30 PM-7:30 PM

Hongliang Qian1*, Florence Pik Hoon Gay1*, Jane Wan Lu Pang1*, Yunqin Lee1*, Joshur Ang1*, Hwee Ching Tan1*, Erica SH Lek1*, Dario Campana2 and Ying Xim Tan3

1MediSix Therapeutics, Singapore, Singapore
2Department of Pediatrics and Cancer Science Institute, Yong Loo Lin School of Medicine, National University of Singapore, Singapore, Singapore
3Medisix Therapeutics, Singapore, Singapore

Infusion of CAR-T cells has produced dramatic clinical responses in patients with B-cell leukemias and lymphomas as well as multiple myeloma, but cell-based and potentially curative therapies for PTCL are not available. We sought to develop an effective CAR T-cell therapy for PTCL by targeting CD3.

PTCL develops from mature T-cells and tumors from most subtypes retain high and uniform CD3 expression. CD3 is not expressed in non-hematopoietic tissues but is highly expressed in T lymphocytes. Therefore, expression of an anti-CD3 CAR in T lymphocytes is likely to result in T-cell self-killing, precluding consistent manufacturing of sufficient CAR-T cell numbers. Indeed, when we transduced peripheral blood T-cells with a lentiviral vector delivering a second-generation CAR targeting CD3 (anti-CD3–41BB-CD3zeta), massive fratricide rapidly ensued. To solve this problem, we downregulated surface CD3 expression using a CD3 protein expression blocker (PEBL). CD3 PEBL is composed of an anti-CD3 single-chain variable fragment and an intracellular retention domain that anchors the cognate antigen in the endoplasmic reticulum and Golgi apparatus before degradation. Expression of CD3 PEBL in T-cells completely abolished cell surface CD3 expression, without affecting cell proliferation and cytotoxicity (Kamiya et al. Blood Adv 2018). When T-cells were transduced with lentiviral vectors containing anti-CD3 CAR and CD3 PEBL, fratricide was limited, resulting in high numbers of viable anti-CD3 PEBL-CAR T cells. With peripheral blood cells from 6 healthy donors, the median percentage of CAR+ cells obtained was 45.5% (range 30%-62%) and the median expansion achieved 9-10 days post T-cell activation and transduction was 46-fold (range 11-95). Anti-CD3 PEBL-CAR T cells had potent and specific cytotoxicity against CD3+ target cells in short- and long-term assays and proliferated vigorously in the presence of CD3+ target cells. We further assessed the anti-tumor efficacy of anti-CD3 PEBL-CAR T cells in immunodeficient mice engrafted with CD3+ Jurkat cells expressing firefly luciferase and eGFP. Anti-CD3 PEBL-CAR T cells produced a distinct anti-leukemic effect, with treated mice showing a marked reduction in leukemic cell burden (Fig. 1A).

Importantly, CD3 PEBL also prevented T-cell receptor (TCR)/CD3 display on the cell surface and abolished TCR-mediated signalling. Minimal signs of graft-versus-host disease (GvHD) were detected in immunodeficient mice treated with anti-CD3 PEBL-CAR T cells, whereas mice treated with non-transduced T-cells developed GvHD which caused substantial weight loss (Fig. 1B).

In conclusion, PEBL-mediated blockade of surface TCR/CD3 complex expression prevents fratricide mediated by anti-CD3 CAR and abrogates TCR-based antigen recognition by T-cells. The latter property is critical for the manufacturing of allogenic CAR-T cells without GvHD potential. The technology developed in this study fits well in current cGMP cell manufacturing protocols and it does not pose the risk of off-target gene editing. Taken together, these features warrant testing of this novel anti-CD3 CAR-T therapy in the treatment of relapsed/refractory PTCL.

Disclosures: Qian: MediSix Therapeutics: Current Employment, Current holder of stock options in a privately-held company. Gay: MediSix Therapeutics: Current Employment, Current holder of stock options in a privately-held company. Pang: MediSix Therapeutics: Current Employment, Current holder of stock options in a privately-held company. Lee: MediSix Therapeutics: Current Employment, Current holder of stock options in a privately-held company. Ang: MediSix Therapeutics: Current Employment, Current holder of stock options in a privately-held company. Tan: MediSix Therapeutics: Current Employment, Current holder of stock options in a privately-held company; Tessa Therapeutics: Ended employment in the past 24 months. Lek: MediSix Therapeutics: Current Employment, Current holder of stock options in a privately-held company. Campana: MediSix Therapeutics: Consultancy, Current equity holder in private company, Patents & Royalties; Nkarta Therapeutics: Current equity holder in publicly-traded company, Membership on an entity's Board of Directors or advisory committees, Patents & Royalties; Juno Therapeutics (Bristol-Myers-Squibb): Patents & Royalties; Smart Immune: Membership on an entity's Board of Directors or advisory committees. Tan: MediSix Therapeutics: Current Employment, Current holder of stock options in a privately-held company.

*signifies non-member of ASH