Abstract 2309: CDK9 as a potential therapeutic target in sarcomas

Abstract

Abstract Sarcomas are a heterogeneous group of over 80 different tumors which arise from mesenchymal or connective tissue, accounting for 1% of adult malignancies. The annual incidence of soft tissue sarcoma (STS) is approximately 11,000 cases in the United States and STS is among the top five causes of cancer deaths for those under 20 years old. Cyclin-dependent kinase 9 (CDK9) is a serine/threonine protein kinase involved in transcription elongation through phosphorylation and activation of RNA polymerase II (RPB1), which increases the levels of key oncogenic genes like myeloid cell leukemia-1 (MCL-1) and c-Myc. CDK9 inhibition reduces their expression, leading to cell cycle arrest and apoptosis. TP-1287 is an orally delivered phosphate pro-drug of alvocidib, a potent CDK9 inhibitor. We hypothesized that alvocidib, the pharmaceutically active form of TP-1287, can inhibit the CDK9 signaling pathway and suppress tumor growth of sarcoma cells. Alvocidib demonstrated the suppression of the phosphorylation of RPB1 (p-RPB1) and MCL-1 expression in an A-673 Ewing sarcoma cell line and U-2 OS and MG-63 osteosarcoma cell lines after 6-hr treatment. In U-2 OS cells, 24-hr treatment with alvocidib induced the cleavage of PARP-1, an indicator of apoptosis, whereas less than 6-hr treatment with alvocidib failed to induce PARP-1 cleavage, suggesting that long-term CDK9 suppression would be necessary for the apoptosis induction in sarcoma cells. In cell viability assays using CellTiter-Glo, alvocidib demonstrated an IC50 of around 100 nM: 32 - 90 nM in sarcoma cell lines after 4-day treatment and 24 - 174 nM in sarcoma cells, which were obtained from patient derived xenograft (PDX) models, after 6-day treatment. In the sarcoma cells from PDX models, MCL-1 mRNA expression was moderately correlated with alvocidib sensitivity. Further bioinformatic analysis to predict alvocidib sensitivity is currently underway. In immunofluorescence analysis, alvocidib induced the activation of caspase-3/7, another indicator of apoptosis, after 2-day treatment in the sarcoma cells from PDX models. In an in vivo pharmacodynamic (PD) study using BALB/c mice, p-RPB1 and MCL-1 suppression in peripheral blood mononuclear cells (PBMCs) was observed after single administration of TP-1287. In an in vivo efficacy study using the A-673 model, TP-1287 showed a trend of tumor growth inhibition (TGI) at 2.5 mg/kg (QD), but not at 7.5 mg/kg (Q3D). These in vivo studies suggest that CDK9 signaling pathway was inhibited after TP-1287 administration in vivo and that continuous CDK9 inhibition would be needed for TGI in sarcoma models. In vivo efficacy and PD studies using sarcoma PDX models are currently ongoing. Taken together, alvocidib has shown activity in inhibiting the growth of sarcoma cells and TP-1287 could be a viable therapeutic option targeting the CDK9 pathway in sarcomas. TP-1287 is being investigated for solid tumors including sarcomas (clinicaltrials.gov, NCT03604783). Citation Format: Yuta Matsumura, Hiroki Umehara, Jun Oishi, Adam Siddiqui, Jason M. Foulks, Setsuko Yamamoto, Steven L. Warner. CDK9 as a potential therapeutic target in sarcomas [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 2309.