Abstract 3098: The effects of a pan-cyclin dependent kinase (CDK) inhibitor and its combination with cisplatin in nasopharyngeal carcinoma (NPC)
Authors: Lim Pei Li , Low John SW , Siemeister Gerhard , Goh Boon Cher , Hsieh Wen-son 
Published: August 2015

Undifferentiated NPC is a unique epithelial malignancy endemic to South-east Asia and North Africa. Cell cycle dysregulation, mediated by mechanisms such as over-expression of cyclin D1 and down-regulation of p16INK4A is frequently found in NPC. Our previous work suggest that NPC cells are sensitive to the effects of cell cycle inhibition. BAY1000394 (BAY) is a potent pan-CDK inhibitor exhibiting broad anti-proliferation activity across a number of different cancer types. In this study, the anti-tumor effects of BAY alone and in combination with cisplatin on NPC cell lines were examined in vitro (MTS assay) and in a xenograft model. Cell cycle effects and apoptosis were examined via flowcytometry. Protein levels of relevant cell cycle regulators and mediators of apoptosis were assessed via Western Blot.

Time-dependent inhibition of NPC cell lines HONE-1 and HK-1 by BAY was evaluated and demonstrated that growth inhibition of 50% of the cell population occurred by 48hrs of treatment. Mean IC50 of 30nM was seen. BAY inhibits phosphorylation of cellular CDK target proteins such as retinoblastoma (Ser807/811) (Rb), nucleophosmin (Thr199) (NPM) and RNA polymerase II (Ser2) (p-RNA POL II). Dephosphorylation of RNA POL II inhibits transcription of gene products resulting in rapid decline of MCL-1. Direct caspase-3 activation was observed with the concomitant appearance of active caspase-3; inducing PARP cleavage and inhibition of survivin. BAY was able to induce cell cycle arrest at multiple checkpoints thus enhancing apoptosis. Annexin V and PI staining after 96hrs of BAY exposure showed apoptosis of &gt;80% of BAY treated cell. In vivo dosing of BAY1000394 at 0.5mg/kg and 1mg/kg showed significant growth inhibition as compared to control (p-value &lt;0.05).

Combination studies of BAY with cisplatin displayed synergistic cell kill. Cell cycle analysis of HONE-1 cells treated at 10nM and 20nM BAY with doses of cisplatin ranging from 0.5μM to 2μM showed marked increase in cell death as compared to control and NP69 epithelial cells. Protein expression of activated caspase-3, PARP cleavage and MCL-1 inhibition were observed in dual drug treatment groups. No change in expressions these proteins were seen in treated NP69 cells. In vivo assays of both drugs also demonstrated growth inhibition of tumor bearing mice at BAY 0.5mg/kg and 1.0mg/kg with cisplatin 6mg/kg with good tolerance. Tumor protein expression of p-RNA POL II, NPM and MCL-1 expression were suppressed, suggesting inhibition of CDK9/cyclin T1.

Our data suggest that BAY inhibits NPC tumor cell proliferation in a time- and dose- dependent manner through inhibition of cell-cycle progression and induction of apoptosis. Combination studies with cisplatin suggest synergistic anti-tumor effect. This preliminary study strongly suggests that BAY and its combination with cisplatin has therapeutic potential in the treatment of NPC.

Citation Format: Pei Li Lim, John SW Low, Gerhard Siemeister, Boon Cher Goh, Wen-son Hsieh. The effects of a pan-cyclin dependent kinase (CDK) inhibitor and its combination with cisplatin in nasopharyngeal carcinoma (NPC). [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 3098. doi:10.1158/1538-7445.AM2015-3098