First-in-Human Clinical Study of a Novel CD7-Targeted Chimeric Antigen Receptor (CAR)-T Cell Therapy for Refractory/Relapsed Mixed Phenotype Acute Leukemia (MPAL)
Authors: Zhang Xian , Yang Junfang , Li Jingjing , Shi Yanze , Su Yunchao , Liu Ying , Ba Min , Wu Yan , Li Wenqian , Li Jianqiang , Lu Peihua 
Published: November 2021

Abstract
Introduction
Mixed phenotype acute leukemia (MPAL) is a rare subtype of acute leukemia with features of both acute lymphoblastic leukemia (ALL) and acute myeloid leukemia (AML). The blast cells of MPAL express multilineage immunophenotypic markers and may have a shared B/T/myeloid phenotype. CD7 is expressed by the leukemic blasts and malignant progenitor cells of approximately 30% of AML patients. MPAL is usually associated with a relatively poor prognosis which is even worse compared to either AML or ALL, and even among patients that undergo allogeneic hematopoietic stem cell transplantation (allo-HSCT), underscoring the need for new therapies for these patients. CAR-T cell therapy has shown to be efficacious with a tolerable safety profile in a subset of hematological malignancies. Here, we explored the safety and efficacy of CAR-T therapy in treating refractory/relapsed (r/r) MPAL patients with CD7-positive in a phase I dose-escalation clinical study (https://clinicaltrials.gov NCT04938115).
Methods
Peripheral blood (PB) mononuclear cells were obtained from patient themselves or patient transplant donors for those who relapsed post-transplant by leukapheresis. T-cells were purified using CD3+ magnetic beads or CD4+ and CD8+ magnetic beads. The second-generation CD7 CAR with a 4-1BB costimulatory domain was manufactured per the manufacturer's protocol. This phase I dose escalation study was initiated to explore the safety and efficacy of CD7 CAR-T in treating patients with MPAL. Prior to the CAR-T cells infusion, patients received systemic bridging chemotherapy due to rapid disease progression and then all patients received intravenous fludarabine (30mg/m 2/d) and cyclophosphamide (300mg/m 2/d) (FC) lymphodepleting chemotherapy for 3 consecutive days (Day -5 to Day -3). The median time from leukapheresis to CAR-T cell infusion was 14 days.
Results
From Nov. 2020 to June 2021, 5 adult acute leukemia patients with CD7-positive (4 MPAL, 1 FLT3-mutated AML) were enrolled and infused with CD7 CAR T-cells, one with a low-dose (1.5×10 5/kg), 2 with a medium dose (5.0×10 5/kg), and 3 with a high dose infusion (1.0×10 6/kg). The median transduction efficiency of products was 70.4% (range: 32.0%-96.9%). As of July 30, 2021 cut-off date, the median observation time was 37 days (range: 35-232 days). Characteristics of enrolled patients are shown in Table 1. The median age was 33 years (range: 21-37 years), and the median bone marrow (BM) blasts percentage by morphology was 28.0% (range: 2.5-51.5%) at enrollment. Four patients had prior allo-HSCT history including one who had extramedullary disease (EMD) who relapsed following a 2 nd transplant and had also received CD19 CAR-T cell therapy prior to enrollment in the current trial. The median interval period from prior transplant to CD7 CAR-T cell infusion was 16 months (range: 8-23 months). Following infusion, the median peak of circulating CD7 CAR-T cell was 1.17×10 5copies/μg (0.651~7.04×10 5copies/μg) genomic DNA which occurred around Day 21 (Day14 - Day21) and 59.2% (4.28%~74.58%) occurring on Day18 (Day11-Day21) by q-PCR and flow cytometry respectively (Fig.1).
Four weeks post CD7 CAR-T cell infusion, 4/5 patients achieved complete remission (CR) or CR with incomplete blood recovery (CRi) in BM and all achieved minimal residual disease (MRD)-negative CR. One patient who relapsed post 2 nd transplant, also with EMD, achieved CR in BM, however, remission was not achieved in EMD through PET-CT evaluation on Day 35.
One MRD-negative CR patient subsequently underwent consolidative allo-HSCT 120 days post CD7 CAR T-cell infusion and has remained in MRD-negative CR. Other 3 patients without consolidative allo-HSCT remained in MRD-negative CR till the last follow-up. All 5 patients developed cytokine release syndrome (CRS)-three of 5 patients had grade I CRS, one patient had grade II and one patient had grade III CRS. None of the patients had neurotoxicity. Among the 4 patients who had prior allo-HSCT, 1 developed grade I graft-versus-host disease post CAR-T therapy.
Conclusion
This study demonstrated that CD7-targeted CAR-T therapy offers an opportunity to achieve CR for CD7-positive MPAL patients even for those who relapsed post-transplant. Safety was manageable, however, more data on additional patients and longer observation times are needed to further evaluate the efficacy of CD7 CAR-T products.
Figure 1 Figure 1.

Disclosures
Ba: SenlangBio: Current Employment.