The AKR1C3-Activated Prodrug, Achm-025, Eradicates Disease in Preclinical Models of Aggressive T-Cell Acute Lymphoblastic Leukemia

Introduction: T-cell acute lymphoblastic leukemia (T-ALL) is an aggressive malignancy that is exceptionally difficult to cure after relapse. T-ALL expresses significantly higher levels of the enzyme aldo-keto reductase family 1 member C3 (AKR1C3) compared with B-cell ALL. To exploit this finding, we developed a novel prodrug, ACHM-025, which is selectively activated by AKR1C3 to form a potent cell-entrapped DNA alkylating agent. ACHM-025 was designed to improve drug specificity and minimize toxicity observed with currently used DNA alkylating agents, such as cyclophosphamide (CPM), a prodrug which is activated systemically via liver enzymes. We evaluated the in vivo efficacy and AKR1C3 selectivity of ACHM-025 against a panel of 25 pediatric T-ALL patient-derived xenografts (PDXs) alongside standard-of-care therapy.

Methods: AKR1C3 expression in T-ALL PDXs was determined by RNA-seq, immunoblotting and intracellular flow cytometry. For in vivo efficacy studies, PDXs were established as orthotopic models in immune-deficient NSG mice. Engraftment was assessed by enumerating the proportion of human versus mouse CD45⁺ cells (%huCD45⁺) in the peripheral blood. Treatment commenced when the %huCD45⁺ reached ≥1% (Day 0), and events were defined as %huCD45⁺ ≥25% or leukemia-related morbidity. Drug efficacy was assessed by mouse event-free survival (EFS) and stringent objective response measures.

For the single agent study, ACHM-025 (IP weekly, Days 0, 7, 14) was assessed against 25 T-ALL PDXs using a single mouse trial (SMT) format (one vehicle treated mouse, one drug treated mouse). For the consolidation therapy comparison, ACHM-025 (IP Days 0, 7) or CPM (IP Days 0, 7) combined with cytarabine (Ara-C; IP Days 0-4, 7-11) and 6-mercaptopurine (6MP; IP Days 0-4, 7-11) were assessed against a T-ALL PDX derived from a patient at relapse. For the relapsed/refractory (R/R) therapy study, ACHM-025 (IP Days 0, 7, 14) and nelarabine (IP Days 0-4, 14-18) were assessed against a T-ALL PDX derived from a patient at relapse.

Results: ACHM-025 dose-limiting toxicity in cynomolgus monkeys was neutropenia, where the pharmacokinetic equivalent dose in NSG mice was well tolerated. We first evaluated the in vivo efficacy of ACHM-025 as a single agent using the SMT format across an extended panel of 25 T-ALL PDXs, to address the impact of genetic heterogeneity in pediatric ALL on drug response. Remarkably, 7/25 PDXs treated with ACHM-025 did not relapse over 250 days after the last treatment and a total of 22/25 T-ALL PDXs scored an objective response. In comparison, vehicle treated EFS ranged from 3-26 days for the 25 T-ALL PDXs. Importantly, AKR1C3 expression (mRNA, protein or intracellular) provides a predictive biomarker of efficacy, as ACHM-025 was significantly more effective against T-ALL PDXs with high AKR1C3 expression versus those with low AKR1C3 expression (p<0.0001).

CPM is included in standard-of-care consolidation therapy for ALL in combination with Ara-C and 6MP. Comparing single agents, ACHM-025 was significantly more effective than CPM (T-C 25 vs. 57 days, p=0.0005, Table 1). The combination of ACHM-025, Ara-C and 6MP was significantly more effective than standard-of-care consolidation therapy (CPM, Ara-C and 6MP), more than doubling survival (T-C 36 vs. 75 days, p=0.0005, Table 1). Finally, we compared ACHM-025 with nelarabine, which is the only FDA approved agent for R/R T-ALL. Comparing single agents, ACHM-025 was substantially more effective than nelarabine (T-C 19 vs. 198 days, p=0.0005, Table 1). Importantly, relapsed disease remained sensitive to ACHM-025 re-treatment in vivo, with no evidence of acquired resistance. Remarkably, no mice treated with the ACHM-025/nelarabine combination relapsed over 250 days after the last treatment (Table 1).

Conclusions: ACHM-025 exerted profound in vivo efficacy against T-ALL PDXs and eradicated the disease in 7 aggressive T-ALL PDXs. ACHM-025 was significantly more effective than CPM both as a single agent and when used in combination with Ara-C/6MP. Notably, ACHM-025 in combination with nelarabine was curative when used to treat a chemoresistant T-ALL PDX in vivo. The in vivo efficacy of ACHM-025 directly correlated with AKR1C3 expression levels, providing a predictive biomarker for response. These data provide strong preclinical evidence highlighting the potential of ACHM-025 as a targeted and effective therapy for aggressive forms of T-ALL.