Abstract 2228: Altering rictor with diosmetin reduces tumor progression in orthotopic prostate cancer model

Abstract

Abstract Rapamycin insensitive companion of mTOR (Rictor) is an essential subunit of mTOR complex 2, maintains the integrity of the complex and functions as full activator of Akt. Rictor has been implicated to be involved in growth and progression of malignancies. Reports suggested that overexpression of Rictor in prostate cancer tissues, which may have potential role in prostate cancer progression. We demonstrated silencing Rictor in PC-3 cells decreased p- Akt (Ser-473) expression; conversely p-Akt (Thr-308) and p-PKCα (Ser-657) expression was increasing. Additionally to observe the molecular mechanism of Rictor regulation, we took HAP1 cells (derived from male chronic myelogenous leukemia (CML) cell line KBM-7) were edited and silenced by CRISPR/Cas9 with 2 and 10 base pair Rictor in a coding exon 5. HAP1 haploid human cell line contains a single copy of all chromosomes except for a heterozygous 30-megabase fragment of Chromosome 15; these cells are instrumental for genetic screening, as gene inactivation is greatly facilitated by the absence of a second gene copy. In these cells we observed p-Akt (Ser-473) expression was significantly less with more pronounced in 10 base pair than 2 base pair. Further to determine potential role of diosmetin (5, 7-Trihydroxy-4′-methoxyflavone; a natural flavonoid present in citrus plant, has anti-mutagenic properties) on Rictor silenced 2 base pair and 10 base pair HAP1 cells, we treated these cells in concentration response fashion, and observed decrease in p-Akt (Ser-473) expression than control cells. Next, we determine invivo potential of Rictor silencing, we used luciferase tag PC-3 cells silenced with Rictor gene orthotopically implanted in ventral prostate of mice. After 8 weeks of Rictor silenced cells implanted mice, we observed less luciferase activity in prostate and occasional metastasis than control luciferase PC-3 cells. Moreover, diosmetin fed mice in dose dependent fashion (20 and 50µg/animal/day) represented decrease in tumor luciferase activity with reduced tumor volume and no metastasis than control mice. We observed metastasis only in control and rapamycin (8 mg/kg body weight intraperitoneally every alternate day) treated mice in spine, lung and kidney. Taken together, our studies demonstrate that diosmetin fed animals resulted in growth inhibition and induction of apoptosis by altering Rictor signaling cascade. We are demonstrating that diosmetin modulates molecular targets viz., Rictor and Akt to alter cellular events and elicit anticancer effects in prostate cancer cells. Citation Format: Rebecca Pakradooni, Ahmad Khalifa, Riddhi Patel, Sanjeev Shukla. Altering rictor with diosmetin reduces tumor progression in orthotopic prostate cancer model [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 2228. doi:10.1158/1538-7445.AM2017-2228