CD123-directed chimeric antigen receptor T-cell (CART123) therapy for relapsed/refractory pediatric acute myeloid leukemia

Abstract Introduction Clinical outcomes of children, adolescents and young adults (CAYA) with second or greater relapsed or refractory (r/r) acute myeloid leukemia (AML) are dismal. Current salvage approaches are ineffective, highlighting an urgent need for novel therapies in this population. Investigation of an autologous 4-1BB CD123-directed CAR (CART123) in adults with r/r AML yielded responses in 25% (PMID 39333315). We now report initial safety and preliminary efficacy from a pediatric phase I clinical trial (NCT04678336) evaluating CART123 in a CAYA population with r/r AML. Methods Patients received a single infusion of CART123 following administration of lymphodepleting chemotherapy (LD). We first evaluated a dose level (DL) of 2 x106 CART123 cells/kg (DL1), and after safety was demonstrated in 6 patients, amended to a 3+3 dose escalation design, exploring a higher dose of 5 x106 CART123 cells/kg (DL2). Cytokine release syndrome (CRS) and immune effector cell-associated neurotoxicity syndrome (ICANS) were graded using consensus guidelines. Protocol-defined dose-limiting toxicities (DLTs) included prolonged grade 3 CRS, ICANS, and non-hematologic toxicities. Bone marrow (BM) evaluation was performed on day 28 to assess morphologic response, minimal residual disease (MRD) by flow cytometry and potential on-target/off-tumor myeloid aplasia. CART123 expansion was assessed in peripheral blood (PB) using quantitative polymerase chain reaction and serum cytokines were assessed by a Luminex multiplex immunoassay. Results Twelve patients were screened, 10 enrolled, and 9 had successful generation of CART123 products meeting target dose. One subject withdrew consent prior to CART123 manufacture, and 2 died of progressive disease prior to infusion. Seven patients were infused with CART123 between June 2021 and September 2024 (6 at DL1, 1 at DL2). Median age at infusion was 5 years (range 3-27), 2 (29%) were female, 1 (14%) was of Hispanic ethnicity, and 4 (56%) non-Hispanic patients were of non-white race. LD regimens included fludarabine and cyclophosphamide with (n=3) or without (n=4) azacitidine. All infused patients had highly refractory disease, with a median of 5 prior lines of treatment (range 3-10), and a median pre-infusion BM blast burden of 50% (range 0-95%). CD123 antigen expression on AML blasts was positive in 6 and negative in 1. DL1: All 6 (100%) patients treated at DL1 experienced CRS (grade 1 or 2 in 5, grade 4 in 1). No ICANS was observed and no DLTs were identified. Although CART123 proliferation occurred with a median peak detection in PB of 7,314 copies/µg genomic DNA (gDNA) (range 28-120,818), CART123 was undetectable by day 28 in 3/5 assessed patients. There were no clinical responses at this dose level. CD123 expression was retained on AML blasts in 5/5 patients with pre-infusion antigen positivity. DL2: The patient treated at DL2 experienced grade 4 CRS and grade 3 ICANS, both deemed DLTs due to duration, and both fully resolved by day 10 following infusion. Treatment for toxicity included tocilizumab, steroids and anakinra. Cytokine profiling demonstrated early elevation of IL-6, IFN-gamma, CXCL9 and GM-CSF. Upon resolution of CRS, the patient developed leukocytosis driven by emergence of PB blasts, with absolute blast count rising to 9,570/µL by day 10 following CART123 infusion. The patient then developed progressive monocytosis, with absolute monocyte count rising to a maximum of 25,280/µL on day 18, coinciding with disappearance of PB blasts. CART123 expansion peaked at 10,816 copies/µg gDNA on day 20, with ongoing detection in PB on day 28. This patient achieved an MRD-negative complete response with partial recovery of platelet count (CRp) on day 28 and proceeded to hematopoietic stem cell transplant (HSCT). Following HSCT, 100% donor chimerism was achieved, however, the patient relapsed on day +145 and died due to disease progression on day +174 post-HSCT. Conclusions CART123 manufacture is feasible in heavily pre-treated CAYA patients with r/r AML. For the patient treated at the higher dose, CART123 appeared to induce monocytic differentiation of AML blasts, and day 28 response of CRp facilitated consolidative HSCT. Detailed molecular characterization is underway to better understand response mechanisms. Future investigation of CART123 will continue dose exploration and evaluate co-therapy with cytokine blockade.