Abstract 1072: Synergistic effect of gemcitabine and a Dclk1 inhibitor on pancreatic cancer cell survival

Abstract

Abstract Pancreatic cancer has the highest mortality rate of all major cancers and is one of the most lethal malignancies. There is a constant upward trend in the number of patients diagnosed with pancreatic cancer and the number of deaths due to pancreatic cancer. Gemcitabine (GEM) is often used in the treatment of pancreatic cancer (PDAC), but has limited effects. Doublecortin-like kinase 1 (Dclk1) is important in the progression of early pancreatic neoplastic lesions and PDAC. However, the functional role of Dclk1 in PDAC is unknown. To identify the substrate protein phosphorylated by Dclk1, we performed a protein microarray analysis on Dclk1 knockdown cells. The results of this analysis directed our studies toward Chk1, which is known to be a potential regulator of the cell cycle and experiences upregulation of phosphorylation after GEM treatment. In general, GEM treatment results in DNA damage to pancreatic cancer cells, an increase in phosphorylated Chk1 (p-Chk1), and arrests the cell cycle progression to repair the damaged DNA. On the basis of the preliminary data, we hypothesized that the decrease in Chk1 phosphorylation by Dclk1 inhibition circumvents cell cycle arrest and impairs the subsequent DNA repair. The aim of this study was to evaluate the synergistic effect of Dclk1 inhibition and GEM treatment on pancreatic cancer cell survival. We used the human pancreatic cancer cell line MIA PaCa-2 and LRRK2-IN-1 (LRRK) as the Dclk1 inhibitor for this study. First, we examined the effects of GEM or the Dclk1 inhibitor or both on cancer cell proliferation and the expression of p-Chk1. Significantly decreased cell proliferation was observed on co-treatment of GEM and LRRK compared to GEM treatment alone. In addition, the expression of p-Chk1 significantly decreased on co-treatment compared to GEM treatment alone. Second, we used flow cytometry to analyze the cell cycle after treatment with GEM and/or LRRK. Almost all cancer cells treated with GEM alone were arrested in the S phase of the cell cycle. The addition of LRRK allows the cell cycle to proceed in the same manner as untreated control cancer cells do. We also evaluated DNA damage by measuring the intensity of gamma-H2AX. Cancer cells that were co-treated experienced more DNA damage than with GEM treatment alone. The co-treatment induced apoptosis without the repair of DNA damage in the cancer cells. In conclusion, the combined treatment with GEM and a Dclk1 inhibitor decreased the cell survival rate compared to treatment with GEM alone through the suppression of p-Chk1. Targeting Dclk1 in combination with GEM might offer an excellent opportunity for future pancreatic cancer treatments. Citation Format: Daichi Kawamura, Yoshihiro Takemoto, Arata Nishimoto, Toshiki Tanaka, Yukari Hironaka, Kumiko Yoshida, Junichi Murakami, Naruji Kugimiya, Eijiro Harada, Koji Ueno, Tohru Hosoyama, Kimikazu Hamano. Synergistic effect of gemcitabine and a Dclk1 inhibitor on pancreatic cancer cell survival [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 1072. doi:10.1158/1538-7445.AM2017-1072