Regulatory effects of antitumor agent matrine on FOXO and PI3K-AKT pathway in castration-resistant prostate cancer cells
We previously demonstrated that matrine could inhibit the proliferating, migrating, as well as invading processes of both PC-3 and DU145 cells. However, the underlying molecular mechanisms have not yet been clearly defined. In this study, using various techniques such as high throughput sequencing technology, bioinformatics, quantitative real-time PCR, and immunoblot analysis, we aimed to understand whether matrine serves as a novel regulator of FOXO and PI3K-AKT signaling pathway. DU145 and PC-3 cell lines were cultured for 24 h in vitro. Cells were treated with either matrine or control serum for 48 h, followed by extraction of total RNA. The RNA was sequenced using HiSeq 2500 high-throughput sequencing platform (Illumina). A gene library was established and quality analysis of read data carried out. Integrated database from the website DAVID was used to analyze Gene Ontology (GO), and Kyoto encyclopedia of genes and genomes (KEGG) pathway of differential genes was used for pathway analysis, screening for fold differences of more than two times. The FOXO and PI3K-AKT signaling pathways were screened, and expression levels of mRNA and core protein detected by real-time PCR and immunoblotting, respectively. High throughput sequencing and GO analysis revealed that differentially expressed genes before and after treatment played an important role in cell metabolic process, growth process, anatomical structure formation, cellular component organization, and biological regulation. KEGG signal pathway analysis revealed that FOXO and PI3K-AKT signal pathways had a significant difference between before and after matrine-treated androgen-independent prostate cancer cells PC-3 and DU145. Real-time PCR showed that matrine treatment led to a significant increase in the expression levels of FOXO1A, FOXO3A, FOXO4, and FOXO6 in DU145 and PC-3 cells (P<0.01 or P<0.05), whereas the PI3K expression levels decreased (P<0.01). Similarly, immunoblotting revealed a significant increase (P<0.05) in the expression levels of FOXO1A FOXO3A, FOXO4, and FOXO6 in both PC-3 and DU145 cells, whereas PI3K expression levels decreased (P<0.05). Matrine had a broad regulating effect on the mRNA expression profiles of both PC-3 and DU145 cells. Matrine may inhibit cell proliferation, migration, as well as invasion, and induce apoptosis in both PC-3 and DU145 cells through FOXO and PI3K-AKT signaling pathways. Matrine could therefore be used as a complementary drug to present chemotherapeutic agents, for treating androgen-independent prostate cancer.
Keywordsmatrine androgen-independent prostate cancer mRNA FOXO signaling pathway PI3K-AKT signaling pathway
Unable to display preview. Download preview PDF.
This work was supported by the National Natural Science Foundation of China (81472382), the National Natural Science Foundation of China for Young Scientists (81101947), the Guangdong Province Natural Science Foundation (2014A030313079), the Fundamental Research Funds for the Central Universities (14ykpy19), Guangdong Province Science and Technology for Social Development Project (2013B021800107), Guangzhou City in 2015 scientific research projects (7415600066401 to Hai Huang).
- Beelen, K., Hoefnagel, L.D.C., Opdam, M., Wesseling, J., Sanders, J., Vincent, A.D., van Diest, P.J., and Linn, S.C. (2014). PI3K/AKT/mTOR pathway activation in primary and corresponding metastatic breast tumors after adjuvant endocrine therapy. Int J Cancer 135, 1257–1263.CrossRefPubMedPubMedCentralGoogle Scholar
- Guerin, M., Qian, C., Zhong, Q., Cui, Q., Guo, Y., Bei, J., Shao, J., Zhu, X., Huang, W., Wu, J., Liu, R., Liu, Q., Wang, J., Jia, W., Zheng, X., and Zeng, Y. (2016). Translational oncology toward benefiting cancer patients: the Sun Yat-sen University Cancer Center experience. Sci China Life Sci 59, 1057–1062.CrossRefPubMedGoogle Scholar
- Kumar, N., Crocker, T., Smith, T., Pow-Sang, J., Spiess, P.E., Connors, S., Chornukur, G., Dickinson, S.I., Bai, W., Williams, C.R., Salup, R., and Fu, W. (2012). Prostate cancer chemoprevention targeting high risk populations: model for trial design and outcome measures. J Cancer Sci Ther 2011, pii: 007.PubMedPubMedCentralGoogle Scholar
- Li, Q., Lai, Y., Wang, C., Xu, G., He, Z., Shang, X., Sun, Y., Zhang, F., Liu, L., and Huang, H. (2016). Matrine inhibits the proliferation, invasion and migration of castration-resistant prostate cancer cells through regulation of the NF-κB signaling pathway. Oncol Rep 35, 375–381.CrossRefPubMedGoogle Scholar
- Trapnell, C., Roberts, A., Goff, L., Pertea, G., Kim, D., Kelley, D.R., Pimentel, H., Salzberg, S.L., Rinn, J.L., and Pachter, L. (2012). Differential gene and transcript expression analysis of RNA-seq experiments with TopHat and Cufflinks. Nat Protoc 7, 562–578.CrossRefPubMedPubMedCentralGoogle Scholar
- Wang, Y., Zhou, Y., and Graves, D.T. (2014). FOXO transcription factors: their clinical significance and regulation. BioMed Res Int 2014, 1–13.Google Scholar
- Wilk, A., Urbanska, K., Grabacka, M., Mullinax, J., Marcinkiewicz, C., Impastato, D., Estrada, J.J., and Reiss, K. (2012). Fenofibrate-induced nuclear translocation of FoxO3A triggers Bim-mediated apoptosis in glioblastoma cells in vitro. Cell Cycle 11, 2660–2671.CrossRefPubMedPubMedCentralGoogle Scholar