Membrane translocation and activation of GnRH receptor sensitize prostate cancer cells to radiation

Abstract

Background

GnRH analogs are widely used as neoadjuvant agents for radiotherapy in prostate cancer (PCa) patients, with well-documented effects in reducing tumor bulk and increasing progression-free survival. GnRH analogs act locally in the prostate by triggering apoptosis of PCa cells via activation of the GnRH receptor (GnRHR). During PCa progression, the distribution of GnRHR within the cell is altered, with reduced expression in the cell membrane and remaining sequestered in the endoplasmic reticulum. Pharmacoperone IN3 is able to relocalize GnRHR to the cell membrane. The aim of this study was to evaluate the effect of radiation on PCa cells pretreated with leuprolide, alone or in combination with IN3, as radiosensitizers.

Material and methods

PC3 and human PCa primary cell cultures were treated with IN3 for 24 h, followed by different doses of leuprolide for 48 h and, finally, single doses of radiation (3, 6, and 9 Gy). After radiation, cell survival, apoptosis, cell cycle distribution, and colony growth were evaluated.

Results

Radiation reduced cell survival and increased apoptosis in a dose-dependent manner. This effect was also directly related to leuprolide concentration. Pretreatment with IN3 enhanced apoptosis and decreased cell survival, also observing a higher proportion of cells arrested in G2.

Conclusion

Neoadjuvant leuprolide increases radiation-mediated apoptosis of PCa cells. This effect was enhanced by pretreatment with pharmacoperone IN3. Clinical use of IN3 as a radiosensitizer combined with androgen deprivation therapy to improve survival of patients with PCa remains to be evaluated.

Keywords:

• Prostate cancer
• primary cell cultures
• leuprolide and GnRH analogs
• GnRH receptor (GnRHR)
• radiotherapy
• pharmacoperone IN3

Acknowledgments

Thanks to miss Graciela Caroca for her excellent technical assistance and Martín Mondaca, Pamela Poblete, Nelson Rocha and Jorge Corvalán from the Radiation Oncology Service of the National Cancer Institute for their support and willingness.

Disclosure statement

No potential conflict of interest was reported by the author(s).

Additional information

Funding

This study was supported by Grant N° 3090016 and 11121525 (C. Sánchez) and N° 1140417 (E. Castellón) from Fondo Nacional de Desarrollo Científico y Tecnológico (Fondecyt, ANID), Chile. Pharmacoperone IN3, was provided under agreement by Merck Sharp & Dohme Corp (New Jersey, USA).

Notes on contributors

Catherine Sánchez

Catherine Sánchez DVM, PhD, Clinical researcher on the role of extracellular vesicles in the mechanisms of carcinogenesis and tumor progression in prostate cancer. In addition, the use of exosomes as diagnostic biomarkers.

Alejandro Mercado

Alejandro Mercado MD, PhD, Urologist. Assistant professor at the University of Chile, as a clinical urologist he participates in the development and implementation of studies focused on improving the diagnosis/treatment of various urological pathologies.

Héctor R. Contreras

Héctor R. Contreras Professor at the University of Chile. His research focuses primarily on the mechanisms of prostate cancer progression and the regulation of the epithelium-mesenchymal transition.

Felipe Carvajal V

Felipe Carvajal V. MD, Radiologist. Assistant professor at the University of Chile, Radiotherapist Oncologist at the National Cancer Institute (Chile). Collaborator of various studies on response to radiological therapy in cancer.

Apolo Salgado

Apolo Salgado MD, Radiotherapist Oncologist. Head of the radiotherapy department at the National Cancer Institute (Chile). Extensive experience in radiotherapy and evaluation of new therapeutic strategies in cancer.

Christian Huidobro

Christian Huidobro MD, Urologist. As a clinician, he has participated in the implementation and development of several prostate cancer research studies.

Enrique A. Castellón

Enrique A. Castellón Professor at the University of Chile. Director of the Department of Basic-Clinical Oncology. His line of research has focused on the cellular and molecular biology of cancer, in particular, on the participation of Cancer Stem Cells in the progression and metastasis of human prostate cancer.