Abstract 776: Study on the induced resistance reversal by JAK2 gene RNAi and inhibitor AG490 in ovarian cancer paclitaxel-resistant cells

Abstract

Abstract Objective: To explore the influence on OC3/TAX300 cell line and the change of chemosensitivity to paclitaxel after inhibiting JAK2 gene expression by constructing a recombined lentivirus vector which expressing small interfering RNA(siRNA) of JAK2 gene, and using JAK2 inhibitor AG490 to discuss the possible mechanism of JAK2 on ovarian cancer chemotherapy resistance Methods: Design and build JAK2 siRNA lentivirus vector by using RNA interference technology, and to transfect the drug resistant cell line OC3/TAX300. The expression levels of JAK2 was detected by RT-PCR and Western blotting. The cell proliferation condition was determined by MTT method, the cell ultrastructure change was observed by transmission electron microscope and the cell cycle and apoptosis were detected by flow cytometry; Western blot was used to detect the expression of STAT3 protein which is JAK2 downstream signal transduction factor. Use different concentration of AG490 solution to culture paclitaxel-resistant cell line OC3/TAX300 of ovarian cancer; JAK2 and STAT3 mRNA and protein expression level were detected by Real-time PCR and Western blot. Detect cell proliferation condition, observe the cell ultrastructure change and detect the cell cycle and apoptosis after treating the variety concentration of AG490 cultivate cells by taxol. Results: 1.The JAK2 siRNA could significantly inhibits the expression of JAK2 mRNA and protein (P<0.05) which confirmed by real-time PCR and western blot results. 2. MTT results showed that the cell growth of JAK2 gene silencing group was significantly inhibited by taxol treatment after 48h and 72h (all P<0.01); AG490 could enhance the sensitivity of cells to taxol(P<0.05); 3. Typical apoptotic cells could be found under the transmission electron microscope in both JAK2 gene silencing group and AG490 culture group; 4. Flow cytometry detection results showed that both the cell apoptosis rate and the cell cycle in G2/M phase proportion increased significantly in JAK2 gene silencing group (P<0.01); The application AG490 can make obviously increase of the cell apoptosis and the cell cycle in G2/M phase proportion induced by paclitaxel(P<0.05).5. Western blot results showed the expression level of STAT3 protein decreased after JAK2 gene silencing(P < 0.05). The expression of JAK2 and STAT3 mRNA and protein were inhibited with varying degrees after using different concentration of AG490 to culture cells(P<0.05).Conclusion: RNAi technology and using tyrosine kinase inhibitor AG490 can effectively restrain the JAK2 and STAT3 expression in ovarian cancer resistant cells. By restoring the paclitaxel effect on cell cycle block, AG490 could inhibit the cell proliferation, induce the cell apoptosis and strengthen the resistant cells of paclitaxel chemotherapy sensitivity. JAK2-STAT3 signal transduction pathway may be involved in the process. Note: This abstract was not presented at the meeting. Citation Format: Hongxia Li. Study on the induced resistance reversal by JAK2 gene RNAi and inhibitor AG490 in ovarian cancer paclitaxel-resistant cells. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 776. doi:10.1158/1538-7445.AM2014-776