Antitumor effects of cyclin dependent kinase 9 inhibition in esophageal adenocarcinoma

Abstract

Role of cyclin dependent kinase 9(CDK9) as a potential target in esophageal adenocarcinoma (EAC) is unknown. We investigated CDK9 protein expression in EAC and Barrett's esophagus and role of CDK9 in oncogenic processes of EAC in vitro and in murine xenografts. The CDK9 expression was significantly higher in EAC as compared to Barrett's esophagus in patient samples. Stable shCDK9 in SKGT4 reduced proliferation by 37% at day 4, increased apoptosis at 48 hours and induced G1 cell cycle arrest at 48 hours (58.4% vs. 45.8%) compared to controls SKGT4 cells. SKGT4-shCDK9 cell-derived tumors were significantly smaller than control SKGT4-derived tumors in xenografts (72.89mm3 vs. 270mm3). Pharmaceutical inhibition of CDK9 by Flavopiridol (0.1μm for 48 hours) and CAN508 (20 and 40μm for 72 hours) induced significant reduction in proliferation and 2-fold increase in apoptosis in SKGT4, FLO1 and OE33 cells. In xenograft models, CAN508 (60 mg/kg/dayx10 days) and Flavopiridol (4mg/kg/dayx10 days) caused 50.8% and 63.1% reduction in xenograft tumors as compared to control on post-treatment day 21. Reduction of MCL-1 and phosphorylated RNA polymerase II was observed with transient shCDK9 in SKGT4 cells but not with stable shCDK9. CAN508 (20 and 40 μm) and Flavopiridol (0.1, 0.2 and 0.3 μm) for 4 hours showed reduction in MCL-1 mRNA (84% and 96%) and protein. Mcl-1 overexpression conferred resistance to Flavopiridol (0.2 μm or 0.4 μm for 48 hours) and CAN 508 (20 or 40μm for 72 hours). Chromatin immunoprecipitation demonstrated significant reduction of binding of transcriptional factor HIF-1α to MCL-1 promoter in FLO-1 cells by CDK9 inhibitors.

Targeting cyclin-dependent kinase 9 by a novel inhibitor enhances radiosensitization and identifies Axl as a novel downstream target in esophageal adenocarcinoma

“…Previously, we showed that CDK9 inhibitors exert dose-dependent anti-proliferative effects against 6 esophageal adenocarcinoma cell lines [16]. Radioresistant OE33R cells were established by exposing radiosensitive OE33 cells to weekly doses of 2 Gy radiation and radiation resistance was achieved after 45 fractions of 2 Gy radiation.…”

“…BAY1143572 induces its anti-tumorigenic effects in adult T-cell leukemia/lymphoma by inhibiting pSer2 and pSer7 RNA Pol II, MYC, and MCL-1. Our studies [16] (unpublished data) indicate that CDK9/p-TEFb inhibition is the dominant mechanism of action for three CDK inhibitors: Flavopiridol [21, 22], CAN 508 and BAY1143572, in esophageal adenocarcinoma. In the present study, we show for the first time that inhibition of CDK9 potently enhances radiation sensitivity in various preclinical models of esophageal adenocarcinoma.…”

“…Cyclin dependent kinase (CDK) 9 is a promising target to enhance radiosensitization [13–16]. Previously, we showed overexpression of CDK9 in esophageal adenocarcinoma cells compared to matched normal esophageal epithelial cells and Barrett’s esophagus [16].…”

“…Cyclin dependent kinase (CDK) 9 is a promising target to enhance radiosensitization [13–16]. Previously, we showed overexpression of CDK9 in esophageal adenocarcinoma cells compared to matched normal esophageal epithelial cells and Barrett’s esophagus [16]. BAY1143572 (Atuveciclib), a novel, first-in-class CDK9 specific inhibitor more potently inhibits CDK9 (PTEFb) activity because it’s IC50 is 50 fold lower than the IC50 of other CDKs [17–19].…”

Comparative Modeling of CDK9 Inhibitors to Explore Selectivity and Structure-Activity Relationships

“…CDK9 can be activated by four distinct cyclins (T1, T2a, T2b, and K) to form one of four positive transcription elongation factor (P-TEFb) complexes that regulate RNA polymerase II [10]. CDK9 is found to be overexpressed in a variety of cancers including pancreatic [11], ovarian [12], cervical [13], esophageal [14] and osteosarcoma [15], with increased CDK9 correlated to poorer patient prognosis [11,15,12]. From in vitro studies and animal models, CDK9 has been implicated as a potential target for therapeutic intervention due to its role in MYC stabilization [16], chemo-and radio-resistance [17][18][19], and maintenance of cancer cell stemness [20,21].…”