Giovanni Luca Gravina1, Francesco Marampon, David Sherris, Francesca Vittorini, Ernesto Di Cesare, Vincenzo Tombolini, Andrea Lenzi, Emmanuele A Jannini, Claudio Festuccia. 1. Division of Radiation Oncology, Department of Clinical and Applied Sciences and Biotechnologies, University of L'Aquila, L'Aquila, Italy; Department of Clinical and Applied Sciences and Biotechnologies, Laboratory of Radiobiology, University of L'Aquila, L'Aquila, Italy; Department of Experimental Medicine, Section of Medical Pathophysiology, Food Science and Endocrinology, Sapienza University of Rome, Rome, Italy; Department of Clinical and Applied Sciences and Biotechnologies, School of Sexology, University of L'Aquila, L'Aquila, Italy.
Abstract
BACKGROUND: P529, a Torc1/Torc2 inhibitor, has demonstrated its potential as a radiosensitizer. However the molecular mechanisms underlying this phenomenon still need to be elucidated. Aim of this study is to dissect molecular mechanisms regulating the radiosensitizing properties of P529 in a wide panel of prostate cancer models. METHODS: Six tumor cell lines and xenograft models were used for in vitro and in vivo studies. Clonogenic survival, apoptotic, autophagic, and senescence assays were used to examine the effects of ionizing radiation (IR) alone and in combination with P529. CRM1, survivin, GSK-3β, and DNA-DSBs expression and modulation, upon P529 and RT, were monitored by western blot. In vivo treatment response upon P529, irradiation or combination of P529 with IR was monitored by tumor volume, time to progression (TTP), and immunohistochemical analysis. RESULTS: P529 treatment induced significantly more apoptosis and DNA double-strand break (DSB) when combined with radiotherapy resulting in cellular radiosensitization and growth delay of irradiated tumor xenografts. Upon P529 treatment Rad51, DNA-PKcs, and Ku70 protein expression was downregulated, indicating delayed DNA double-strand damage repair. The radiosensitizing properties of P529 were partially linked to GSK-3β, cyclin-D1, and c-myc modulation with associated inhibition of CRM1-mediated nuclear export of survivin. Importantly, autophagy and tumor senescence were involved in the enhanced P529 radioresponse. CONCLUSIONS: Impaired DNA double-strand damage repair, inhibition of CRM1-mediated nuclear export of survivin, modulation of cyclin-D1 and c-myc with associated pro-apoptotic and autophagic and senescent events explain the radiosensitizing properties of P529 in preclinical models of prostate cancer.
BACKGROUND: P529, a Torc1/Torc2 inhibitor, has demonstrated its potential as a radiosensitizer. However the molecular mechanisms underlying this phenomenon still need to be elucidated. Aim of this study is to dissect molecular mechanisms regulating the radiosensitizing properties of P529 in a wide panel of prostate cancer models. METHODS: Six tumor cell lines and xenograft models were used for in vitro and in vivo studies. Clonogenic survival, apoptotic, autophagic, and senescence assays were used to examine the effects of ionizing radiation (IR) alone and in combination with P529. CRM1, survivin, GSK-3β, and DNA-DSBs expression and modulation, upon P529 and RT, were monitored by western blot. In vivo treatment response upon P529, irradiation or combination of P529 with IR was monitored by tumor volume, time to progression (TTP), and immunohistochemical analysis. RESULTS: P529 treatment induced significantly more apoptosis and DNA double-strand break (DSB) when combined with radiotherapy resulting in cellular radiosensitization and growth delay of irradiated tumor xenografts. Upon P529 treatment Rad51, DNA-PKcs, and Ku70 protein expression was downregulated, indicating delayed DNA double-strand damage repair. The radiosensitizing properties of P529 were partially linked to GSK-3β, cyclin-D1, and c-myc modulation with associated inhibition of CRM1-mediated nuclear export of survivin. Importantly, autophagy and tumor senescence were involved in the enhanced P529 radioresponse. CONCLUSIONS: Impaired DNA double-strand damage repair, inhibition of CRM1-mediated nuclear export of survivin, modulation of cyclin-D1 and c-myc with associated pro-apoptotic and autophagic and senescent events explain the radiosensitizing properties of P529 in preclinical models of prostate cancer.
Authors: Alonso Heredia; Nhut Le; Ronald B Gartenhaus; Edward Sausville; Sandra Medina-Moreno; Juan C Zapata; Charles Davis; Robert C Gallo; Robert R Redfield Journal: Proc Natl Acad Sci U S A Date: 2015-07-13 Impact factor: 12.779