The Chaperone Protein GRP78 Promotes Survival and Migration of Head and Neck Cancer After Direct Radiation Exposure and Extracellular Vesicle-Transfer.

Michael Schneider,Rosemarie Kell,Michael J. Atkinson,Klaudia Winkler,Simone Moertl,Michael W. Pfaffl

doi:10.3389/fonc.2022.842418

Abstract

Background and PurposeIncreased levels of the chaperone protein GRP78 have been implicated in poorer outcomes of cancer therapy. We have therefore explored the functional connection between the expression of GRP78 and the development of radioresistance and metastatic behavior in HNSCC.Material and MethodsThe association between gene expression of GRP78 and survival in HNSCC patients was examined using the TCGA database. The influence of ionizing radiation on the GRP78 levels in HNSCC cell lines, their secreted extracellular vesicles (EV) and non-irradiated EV-recipient cells was investigated by Western Blot and FACS. The consequences of chemical inhibition or experimental overexpression of GRP78 on radioresistance and migration of HNSCC cells were analyzed by clonogenic survival and gap closure assays.ResultsElevated levels of GRP78 RNA in HNSCC correlated with poorer overall survival. Radiation increased GRP78 protein expression on the surface of HNSCC cell lines. Experimental overexpression of GRP78 increased both radioresistance and migratory potential. Chemical inhibition of GRP78 impaired cell migration. EVs were identified as a potential source of increased GRP78 content as elevated levels of surface GRP78 were found in EVs released by irradiated cells. These vesicles transferred GRP78 to non-irradiated recipient cells during co-cultivation.ConclusionsWe have identified the chaperone protein GRP78 as a potential driver of increased radioresistance and motility in HNSCC. The uptake of GRP78-rich EVs originating from irradiated cells may contribute to a poorer prognosis through bystander effects mediated by the transfer of GRP78 to non-irradiated cells. Therefore, we consider the chaperone protein GRP78 to be an attractive target for improving radiotherapy strategies.

Highlights

Therapy options for head and neck squamous cell carcinomas (HNSCC) have improved over the last decades, tobacco consumption is declining and HPV-positive status is a good prognostic marker for tumor treatment
In HNSCC, we have previously demonstrated that Extracellular vesicle (EV) released from irradiated cells are able to increase the motility and radioresistance of recipient cells and proteomic analyses showed increased vesicular levels of Glucose-regulated protein 78kDa (GRP78) [17, 19]
We report that surface GRP78 expression is increased by exposure to ionizing radiation in two HNSCC cell lines

Summary

Introduction

Therapy options for head and neck squamous cell carcinomas (HNSCC) have improved over the last decades, tobacco consumption is declining and HPV-positive status is a good prognostic marker for tumor treatment. Especially HPVnegative and advanced-stage tumors still have a poor prognosis and an overall 5-year survival rate of approximately 60% [1, 2]. HNSCC are highly invasive and metastatic and frequently acquire treatment resistance. This resistance results in an increase in treatment doses of radiation and chemotherapy, which in turn leads to worse off-target toxicity. Standard therapy protocols combine surgery with radiotherapy and chemoradiotherapy despite the severe side effects and low effectiveness [2, 3], especially in non-resectable or incompletely resected tumors [4]. Increased levels of the chaperone protein GRP78 have been implicated in poorer outcomes of cancer therapy. We have explored the functional connection between the expression of GRP78 and the development of radioresistance and metastatic behavior in HNSCC

Methods

Results

Discussion

Conclusion