CD98hc as a potential marker of radioresistance in head and neck squamous cell carcinoma

Background: CD98hc protein is encoded by the SLC3A2 (solute carrier family 3 member 2) gene. CD98hc contributes to the amino acid transport and regulation of the integrin signaling pathway, and is a putative marker of cancer stem cells (CSCs). Recent retrospective analyses showed a correlation of high SLC3A2 expression with poor locoregional control in patients with locally advanced head and neck squamous cell carcinoma (HNSCC) treated with a curatively intended cisplatin-based postoperative radiochemotherapy or primary radiochemotherapy. The aim of this study is the investigation of potential molecular mechanisms of action of CD98hc as a regulator of HNSCC radioresistance. Methods: The expression levels of CD98 in nine HNSCC cell lines were measured by Western blot analysis and correlated with corresponding tumor control dose 50 (TCD50) values. The fractions of plasma membrane proteins from Cal33 and FaDu cells and their radioresistant sublines established by fractionated irradiation with a total dose of ≥50Gy of X-rays were enriched by differential centrifugation and quantitatively analyzed by tandem mass spectrometry. To determine the role of SLC3A2 in the regulation of cell radioresistance, HNSCC cell lines and their respective irradiated sublines were transfected with SLC3A2 siRNAs and analysed by 2D and/or 3D radiobiological clonogenic assays after X-ray irradiation. The CRISPR/Cas9 system was used for the stable monoallelic knock-out of SLC3A2 in HNSCC cells. The efficiency of DNA repair after irradiation was examined by γH2A.X foci analysis. The Cancer Genome Atlas (TCGA) gene dataset for HNSCC patients treated with and without radiotherapy was used to analyse a potential correlation of SLC3A2 expression with expression of other genes and with clinical outcome of the HNSCC patients. Results and Discussion: The data of proteomic profiling, the results of Western blot analysis correlated to TCD50 values, the functional assessment of the SLC3A2 knockdown and knockout cells along with analysis of the TCGA datasets confirmed a connection between the CD98hc expression level and the cancer cell radiosensitivity in vitro and in vivo. These results support the hypothesis that CD98hc is a regulator of HNSCC radioresistance. The ongoing experimental studies will reveal if CD98hc may be used as potential target for HNSCC radiosensitization.