Abstract 1818: Enhanced antitumor and anti-angiogenic effects of Apatinib combined with chemotherapy in a zebrafish model of non-small cell lung cancer
Authors: Xiang Yi , Zhang Wenchao , Wu Zhuanbin , Qian Zijun , Zhou Jianping , Wang Xiaofei , Wang Weiqin , Zhou Ling , Liu Jing , Feng Yun , Zhou Min , Shi Guochao , Gao Beili 
Published: July 2017

Abstract
Background: Apatinib is a highly selective VEGFR2 inhibitor, but its effects on non-small cell lung cancer (NSCLC) has not been widely reported and whether there is synergistic effect of Apatinib and conventional chemotherapeutic drugs is also unclear.
Methods: In vitro, MTT assay was used to evaluate the combination effects of Apatinib and chemotherapy agents (Gemcitabine, Paclitaxel and Pemetrexed) against A549 (EGFR negative mutation cell line) lung cancer cells. In vivo, we firstly assessed the safety of Apatinib and chemotherapy agent in zebrafish model by observing organ developmental malformation and zebrafish embryos mortality induced by drug toxicity. The inhibition effect on the development of blood vessel in zebrafish was used to assess antiangiogenic property of Apatinib. By means of microinjection, A549 cells stained with red fluorescent cell tracer CM-Dil fluorescence were grafted into the yolk sac of zebrafish embryo. Then, zebrafish were incubated with Apatinib, chemotherapy agents or Apatinib combined chemotherapy agents, respectively. The tumor xenografts volume was measured by estimating the relative fluorescence intensity. Q-RTPCR was used to detect the expression of some genes associated with angiogenesis in order to identify molecular mechanism of the combination effects of Apatinib and chemotherapy agent.
Results: Apatinib had a direct inhibitory effect on A549 cells with the IC50 values ranging from 2.693 to 5.384μmol. Apatinib combined with Pemetrexed gave the most optimal anti-tumor effect compared with Gemcitabine or Paclitaxel in vitro. In zebrafish model, the LC50 value of Apatinib to zebrafish embryos was only half it of Pemetrexed to zebrafish embryos, suggesting that Apatinib is a low toxicity agent. In addition, Apatinib strongly inhibits the process of angiogenesis but not the developed and mature vasculature, suggesting that Apatinib can effectively inhibit tumor angiogenesis in lung cancer patient, which is aided in suppressing tumor growth. As for antitumor effects, Apatinib or Pemetrexed, whatever alone or combination all significantly inhibited tumor growth and the co-treatment of Apatinib and Pemetrexed gave the most optimal anti-tumor effect, suggesting that the combination of apatinib and Pemetrexed may be a promising alternative therapy for lung cancer patients. Finally, the qRT-PCR showed that in addition to synergistically inhibiting VEGFR2 gene expression, co-treatment of Apatinib and Pemetrexed also synergistically inhibit Efnb2a, Robo4 and FGFR4 gene expression.
Conclusions: Apatinib combined with chemotherapies, especially Pemetrexed, synergistically enhanced anti-tumor and anti-angiogenic effects in zebrafish model of NSCLC mainly through inhibiting VEGFR2, Efnb2a, Robo4 and FGFR4 pathways.
Note: This abstract was not presented at the meeting.
Citation Format: Yi Xiang, Wenchao Zhang, Zhuanbin Wu, Zijun Qian, Jianping Zhou, Xiaofei Wang, Weiqin Wang, Ling Zhou, Jing Liu, Yun Feng, Min Zhou, Guochao Shi, Beili Gao. Enhanced antitumor and anti-angiogenic effects of Apatinib combined with chemotherapy in a zebrafish model of non-small cell lung cancer [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 1818. doi:10.1158/1538-7445.AM2017-1818