Preclinical Studies and Phase II Trial of Venetoclax in Combination with Chidamide and Azacitidine in Relapsed/Refractory Acute Myeloid Leukemia

Background: The treatment for relapsed and refractory acute myeloid leukemia (R/R AML) remains challenging and leukemia stem cells (LSCs) play a causal role in R/R AML. Recent progress in genomics and molecular biology has uncovered a spectrum of mutations in AML suggesting that translational research combined with targeted therapy for LSCs has greater guiding significance to improve outcome in R/R AML. Therefore, we used a high-throughput screening platform to identify a novel and effective therapeutic combination, which is venetoclax, chidamide, and azacitidine (VCA). Next, to assess the efficacy of VCA, we performed preclinical studies on LSCs-like cell lines and patient-derived xenograft (PDX) mouse model. In addition, a subsequent phase II study was conducted to test the safety and preliminary efficacy of this combination.

Methods:Preclinical: We performed an automated drug screen to evaluate the drug sensitivity in CD34⁺CD38^- KG-1α and Kasumi-1 cells simultaneously. Ultimately select the optimal combination therapy of venetoclax, chidamide, and azacitidine (VCA) by using a high-throughput platform. To validate the efficacy of the combination in vivo, we used primary CD34⁺ AML PDX model. Mice were treated with single, dual, or triple of venetoclax (VEN, 30 mg/kg, oral gavage), chidamide (CS055, 7.5 mg/kg, oral gavage), and azacitidine (5-AZA, 2 mg/kg, intraperitoneal injection) every 3 days for 12 days consecutively.

Phase II: In a multi-center, single-arm, open-labeled phase II study, patients aged ≥18 years with R/R AML were treated with VEN at 200 mg orally daily for 14 days, CS055 at 30 mg orally twice per week for 14 days, and 5-AZA at 100 mg by continuous hypodermic injection (IH) daily for 7 days for up to two 28-day cycles. The primary objective was to assess the safety and efficacy of VCA. The primary efficacy measurement was overall response rate (ORR), which was the combined complete remission (CR), CR with incomplete hematologic recovery (CRi) and partial remission (PR).

ResultsPreclinical: VCA exhibited synergistic effects on attenuating cell viability and inducing apoptosis in CD34⁺CD38^- KG-1α and Kasumi-1 cells. Importantly, triple therapy intensively reduced leukemia burden and improved prognosis in a primary CD34⁺ AML derived PDX model without significant adverse effects versus single or double combination. Mechanistically, co-exposure to three agents led to the inhibition of anti-apoptotic BCL-2 family proteins (including BCL2, BCL-XL and MCL-1), associating with enhanced ubiquitination. Moreover, CS055-mediated HDAC inhibition increased F-box and WD repeat domain-containing protein 7 (FBW7) acetylation, thus resulting in FBW7 stabilization, which further enhanced the MCL-1 protein degradation.

Phase II: As of July 30^th, 2022, 29 patients were enrolled in the study. Patients had a median age of 51 years (range: 35-64). The median time on study was 31 (range: 4-not reached [NR]) days. 25 patients (86.2%) had confirmed responses. The ORR was 75% (12/16: 2 CR, 7 CRi and 3PR) with one complete and 88.9% (8/9: 5 CR, 2 CRi and 1 PR) with two completes. The most common non-hematologic adverse events (AE) were nausea (75%) and vomiting (25%), and no grade III or IV non-hematologic AE was observed. There was no tumor lysis syndrome (TLS) or treatment-discontinuation due to therapy.

Conclusions: This novel combination strategy of VEN, CS055 and 5-AZA has shown synergy in AML LSCs-like cells and PDX model. The phase II data suggests that VCA as a chemo-free therapy exhibits promising efficacy and is safe and well-tolerated in patients with R/R AML.

No relevant conflicts of interest to declare.