Radiation and hypoxia-induced non-targeted effects in normoxic and hypoxic conditions in human lung cancer cells

Abstract

Purpose: Many cell lines with anaerobic metabolism do not show cytotoxic abscopal effect (AE) following irradiation. Further, there is no existing data on the radiation- and hypoxia (H)-induced AE. The purpose of this study was to investigate and compare the status of radiation-induced abscopal effect (RIAE) in normoxic and hypoxic conditions.

Methods: Lung cancer cells (A549, H460) were exposed either to hypoxia or normoxia and then irradiated (2 or 10 Gy). After 24 h, unirradiated hypoxic (H-CM) or normoxic (N-CM) conditioned media (CM) and irradiated hypoxic (H-RCM) or normoxic (N-RCM) CM was collected. Hypoxia-resistant clones (HR: A549/HR, H460/HR) were generated by continuous exposure of the cells to hypoxia. Unirradiated parental cells or HR were exposed to H-CM, N-CM, H-RCM or N-RCM. In some groups, 24 h after exposure to CM, cells were directly irradiated with 2 Gy. Cell growth was monitored using real-time cell electronic sensing system. Further, levels of hypoxia and HIF1α regulated angiogenesis related growth factors, basic fibroblast growth factor (bFGF), placental growth factor (PlGF), soluble fms-like tyrosine kinase (sFlt-1) and vascular endothelial growth factor (VEGF) were assessed in CM.

Results: In the radio-resistant A549 cells, H-RCM was much more effective in inducing growth delay compared to N-RCM. In the radio-sensitive H460 cells, both N-RCM and H-RCM induced growth delay. Interestingly, effects of N-RCM were completely reversed in HR cells. Exposure of cells to direct irradiation (2 Gy) 24 h after incubation with CM resulted in 50–60% reduction in cell proliferation in A549/HR cells and a very significant induction of death (>95%) in H460/HR cells. Direct irradiation of parental or HR clones of A549 and H460 cells exposed to H-CM 24 h with 2 Gy induced significant reduction in cell proliferation (from 40% to >95%) in all the cells. Further, levels of sFlt-1 correlated with growth delay in all the cells.

Conclusions: These results for the first time demonstrate that irradiation of hypoxic cells and exposing the cells to acute hypoxia lead to significant AE.

Keywords:

• Abscopal
• bystander
• hypoxia
• lung cancer
• radiation

Disclosure statement

No potential conflict of interest was reported by the authors.

Additional information

Notes on contributors

Slavisa Tubin

Slavisa Tubin, MD, is a board-certified Radiation Oncologist and Scientific Investigator at Institut für Strahlentherapie/Radioonkologie of the KABEG Klinikum Klagenfurt am Wörthersee, AUT. His research interests include the use of stereotactic radiotherapy technique for induction of the non-targeted effects of radiotherapy. Currently, he is a Project Leader and Coordinating Investigator of an international, multi-centric, prospective study on intentional induction of the abscopal and bystander effects among oligometastatic patients.

Mansoor M. Ahmed

Mansoor M. Ahmed, PhD, is currently working as a Program Director, Radiotherapy Development Branch, Radiation Research Program (RRP) at NCI/NIH covering a portfolio of grants in immunology, signal transduction, biomarkers, cancer stem cells and in-vitro & in-vivo models in radiotherapy. Prior to joining RRP, he served as a faculty at the University of Kentucky, Weis Center for Research at Geisinger Clinic and University of Miami, establishing programs in Molecular Radiation Biology. During his career, he has more than 100 peer-reviewed publications with 15 invited reviews and chapters. He recently co-authored a book titled “Hypofractionation: Scientific Concepts and Clinical Experiences”. He is associate editor of Radiation Research. He guest-edited several special-topical issues in Radiation Research, Translational Cancer Research, and Seminars in Radiation Oncology.

Seema Gupta

Seema Gupta, PhD, is currently working as a Scientific Writer at Georgia Cancer Center, Augusta, GA. She previously served as a Research Assistant Professor at Albert Einstein College of Medicine, Bronx, NY and University of Miami, Miami, FL. She has been working in the fields of radiation-induced - signal transduction and apoptosis; high dose radiation therapy and immune modulation; radiation-chemotherapy interaction and tumor micro-environment; and cancer metabolism.