Tvt CAR7: Phase 1 Clinical Trial of Base-Edited "Universal” CAR7 T Cells for Paediatric Relapsed/Refractory T-ALL

Background Genome editing can overcome HLA barriers to generate 'off-the-shelf’ CAR T cell therapies. Despite the success of CAR-T cell therapies in B-cell malignancies, the expression of shared T cell antigens has constrained the development of CAR T cells targeting T-cell malignancies, due to T cell fratricide. Targeted base editing using CRISPR guided cytidine deamination mediates highly precise C→U→T conversion which can directly disrupt gene expression without DNA breaks. This Phase I study will investigate the feasibility and safety of base edited (BE) allogeneic CAR T cells against CD7, disrupted for TCR, CD7 and CD52. The cells are used to secure remission ahead of allogeneic stem cell transplantation (allo-SCT).

Investigational medicinal product (IMP) Two allogeneic healthy donor derived BE-CAR7 T cell banks were manufactured from steady state apheresis harvests using a semi-automated process under compliant conditions. Cells were activated with an anti-CD3/CD28 reagent (Transact) and electroporated with codon optimised base editor (coBE) mRNA and three single guide RNAs targeting TRBC, CD52 and CD7. Subsequent lentiviral transduction delivered CAR7 (59-62% efficiency) and magnetic bead processing depleted residual TCRαβ+ T cells (residual 0.1-0.3%). Highly efficient disruption of CD7 and CD52 (98-99%) was confirmed by sequencing. Cells were cryopreserved and QC tested by flow cytometry, ddPCR for copy number (3.1-3.6) and replication competent lentivirus was excluded. Anti-leukemic potency was confirmed in vitro and in human:murine chimera experiments.

Trial sponsorship & approvals The TvT CAR7 study (ISRCTN15323014) is an open label, single centre, single arm and non-randomised clinical trial. The trial is sponsored by Great Ormond Street Hospital NHS trust and is supported by MRC, Wellcome Trust and NIHR. Clinical trial authorisation was provided by MHRA and ethical approval by a gene therapy advisory committee (GTAC) ahead of site initiation.

Study aims and Objectives The study aims to establish the safety and feasibility of BE-CAR7 to induce molecular remission in children with relapsed/refractory (r/r) CD7-positive T-ALL, ahead of a planned allo-SCT. Assessments include incidence of adverse events, duration of remission, disease-free survival and overall survival. Expansion, persistence and elimination of BE-CAR7 cells and immune recovery after allo-SCT are monitored.

Eligibility, treatment and recruitment The study opened on 14^th April 2022 at Great Ormond Street Hospital recruiting children across the UK aged between 6 months and 16 years with r/r CD7+ T-cell malignancies quantifiable in bone marrow (>10^-4 by flow or PCR). Exclusions include active uncontrolled infections, pre-existing GvHD or the detection of anti-HLA antibodies against the IMP. Eligible patients receive lymphodepletion with fludarabine (150 mg/sqm), cyclophosphamide (120 mg/kg) and alemtuzumab (1 mg/kg), followed by infusion of 0.2-2.0x10⁶ BE-CAR7 T cells (maximum 5x10⁴/kg TCRαβ T cells). Patients in molecular remission at day +28 proceed to allo-SCT, to remove BE-CAR 7 cells and promote immune reconstitution. The first subject was infused on 4^th May 2022. Grade 2 CRS and grade 1 ICANS were observed, but no GvHD. Bone marrow assessment at d+28 revealed morphological remission without count recovery and with PCR-MRD <10^-4 and the patient proceeded to reduced intensity allo-SCT as planned. This Phase 1 study aims to treat 10 children in the UK.

Conclusion The study has demonstrated the feasibility of manufacturing "off-the-shelf” GMP-compliant base edited CAR7 T cells and is investigating their safety and efficacy against paediatric r/r T-ALL.

Qasim:Kite & Virocell: Consultancy; Wugen: Consultancy; Novartis: Consultancy; Tessa Therapeutics: Consultancy.