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2111 Intracellular Retention of Tcrαβ/CD3 to Generate Novel Allogeneic CAR-T Cells (ThisCART19A) with Enhanced Antitumor Potency for Treating B-ALL

Program: Oral and Poster Abstracts
Session: 704. Cellular Immunotherapies: Early Phase and Investigational Therapies: Poster I
Hematology Disease Topics & Pathways:
Research, clinical trials, Clinical Research
Saturday, December 9, 2023, 5:30 PM-7:30 PM

Yongxian HU1*, Guoqing Wei2*, Shan Fu3*, Pingnan Xiao3*, Jingjing Feng, MD4*, Mingming Zhang3*, Xingbing Wang, MD5*, Dongrui Wang3*, Jun Li6* and He Huang3*

1Bone Marrow Transplantation Center, the First Affiliated Hospital, Zhejiang University School of Medicine, HANGZHOU, AL, China
2Bone Marrow Transplantation Center, The First Affiliated Hospital, Zhejiang University School of Medicine, Hangzhou, China
3Bone Marrow Transplantation Center, the First Affiliated Hospital, Zhejiang University School of Medicine, Hangzhou, China
4The First affiliated Hospital, School of Medicine, Hangzhou, China
5The Flirst Affiliated Hospital Of University of Science and Technology of China, Hefei, Anhui, CHN
6Fundamenta Therapeutics Co., Ltd., Suzhou, China

Background: Autologous CAR-T therapy has been complicated by long production time, high-cost and risks of manufacturing failure. Allogeneic CAR-T cells can overcome these hurdles, but would subsequently require specific strategies to inhibit allogeneic TCR responses and GvHD. Gene-editing technologies can efficiently deplete endogenous TCR, but also leads to off-target edits and chromosomal abnormality. Furthermore, genetic depletion of TCR disrupts the intracellular T cell activation signal and may compromise CAR-T cytotoxicity. It is thus necessary to develop non-gene-editing allogeneic CAR-T platforms and enhance the potency of allogeneic CAR-T cells.

Methods: We developed a novel non-gene-editing platform named ThisCART (TCRαβ/CD3 and/or HLA-I intracellular sequestered) to manufacture allogeneic CAR-T cells. The platform was based on the intracellular retention of TCRαβ/CD3 complex, allowing for allogeneic CAR-T production with a single lentiviral vector without genetic depletion of TCR. Allogeneic CD19 CAR-T cells (ThisCART19A) was a prototypic product for the platform. The construct contains a CD19-targeted CAR and a KDEL-tagged anti-CD3 single chain antibody (scFv) which prevents TCRαβ/CD3 from being secreted from the endoplasmic reticulum (ER) (Figure A). The efficacy and safety of ThisCART19A were tested in xenograft models. Finally, a phase I study was conducted to assess the safety, efficacy and pharmacokinetics in patients with relapsed or refractory (R/R) B-ALL (NCT05350787). All patients received intravenous fludarabine (30mg/m2/d), cyclophosphamide (300mg/m2/d) and etoposide (100mg/d) for 5 days followed by a single infusion of thisCART19A.

Results: The manufacturing platform of ThisCART19A was able to achieve over 150-folds of ex vivo CAR-T expansion in all batches, with the purity of products (CAR-positive/TCRαβ-negative) above 99%. In preclinical models, ThisCART19A did not induce GvHD, and exhibited superior antitumor function compared to conventional CD19 CAR-T cells. In the Phase I study (Figure B), 10 patients were enrolled and 8 received thisCART19A at doses of 3 (n =5) and 5 (n = 3)×106/kg. All patients were diagnosed as relapse/refractory acute B cell leukemia (R/R B-ALL). Three patients previously received CD19-or CD22-targeted therapies (autologous CAR-T, BiTE or ADC). Grade 3-4 treatment-related adverse events were reported in 8/8 (100%) patients, the most frequent being neutropenia (100%) and thrombocytopenia (87.5%), which most likely related to lymphodepletion. Grade 3-4 CRS was reported in 2/8(25%) patients, and ICANS was reported in 3/8 (37.5%) patients which were all reversible with steroid treatment. 7 patients were evaluable for efficacy analyses (one died from CRS and infection at 5 days post infusion), and MRD-negative CR/CRi was achieved in 100% of these patients. With a median follow-up of 146 days (range, 56 to 407), 4/7 patients remained MRD-negative. Two patients were bridged to allo-HSCT. The mean peak of CAR-T number by FCM was 5908.8(0.51-17457.9) cells/µL , which occurred on day 9 (7-9).

Conclusions: We report for the first time that intracellular retention of TCRαβ/CD3 complex can successfully manufacture allogeneic CAR-T cells, with superior activation potential. In patients with R/R B-ALL, ThisCART19A demonstrated acceptable safety, robust expansion and encouraging clinical response profiles. With streamlined single-vector-based production and enhanced CAR signaling, ThisCART platform represents an attractive alternative to gene-editing-based allogeneic CAR-T platforms.

Disclosures: No relevant conflicts of interest to declare.

*signifies non-member of ASH