Abstract 6388: MERTK and ROCK inhibitors mediate synergistic AML cell death and enhance therapeutic activity in combination with cytotoxic chemotherapy

Abstract

Abstract Survival rates are suboptimal for adult and pediatric patients with acute myeloid leukemia (AML). Cytotoxic chemotherapies have short and long-term side effects and are often contraindicated for elderly patients. Targeted agents can reduce toxicity compared to chemotherapy; however, resistance to single-agents often develops, thus combination therapies may provide more durable responses. To increase efficacy and reduce exposure to cytotoxic agents, a targeted combination therapy (MERTK/ROCKi) was utilized alone and with doxorubicin or etoposide, frontline cytotoxic chemotherapeutics routinely used for AML treatment. MERTK is aberrantly expressed in >80% of AML patient samples and MERTK inhibitors are in clinical development. Both MERTK and rho-associated, coiled-coil-containing protein kinases 1 and 2 (ROCK1/2) regulate actin/microtubule dynamics and cell cycle progression and shMERTK or siROCK1 knockdown induces apoptosis in AML cells. Here, analysis of The Cancer Genome Atlas database revealed poorer overall survival in patients with higher levels of ROCK1 mRNA (p<0.01, n=172). MERTK inhibitors (MRX-2843 or UNC3997) and ROCK1/2 inhibitors (RKI-1447 or GSK269962A) synergized to reduce cell density in 4 of 6 AML cell lines tested and had an additive effect in the remaining cell lines. Flow cytometric analysis of cells stained with popro-1-iodide and propidium iodide (PI) dyes revealed synergistic induction of cell death in KG-1 (p<0.05), OCI-AML5 (p<0.05), and NB4 (p<0.1) cell cultures and an additive effect in Kasumi-1 cultures. Cells treated with the combination therapy failed to expand, even when cultured in the absence of inhibitor(s), and cell density was reduced in response to treatment with the combination therapy compared to single agents (p<0.01). Flow cytometric analysis of DNA content revealed an increased fraction of cells with G2/M DNA content in cultures treated with MRX-2843 (p<0.01). This effect was more pronounced in cultures treated with the combination therapy (p<0.05-p<0.0001) and was accompanied by an accumulation of cells with slightly reduced PI staining relative to untreated G2 cells. Thus, the combination therapy mediates anti-leukemia activity by multiple mechanisms, including abrogation of cell cycle progression and induction of cell death. Addition of MERTK/ROCKi therapy to doxorubicin or etoposide additively decreased cell density relative to chemotherapy alone in OCI-AML5 and KG-1 cultures (p<0.0001). Treatment with doxorubicin and MERTK/ROCKi therapy also decreased OCI-AML5 cell density compared to MERTK/ROCKi therapy alone (p<0.01). These data indicate that combination therapies targeting MERTK and ROCK1/2, alone or in conjunction with chemotherapy, may be particularly effective for treatment of AML and support further studies in animal models. Citation Format: Dawn E. Barnes, Xiaodong Wang, Stephen V. Frye, H. Shelton Earp, Deborah DeRyckere, Douglas K. Barnes. MERTK and ROCK inhibitors mediate synergistic AML cell death and enhance therapeutic activity in combination with cytotoxic chemotherapy [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 6388.