RESUMEN
Radiotherapy is an important treatment option in the therapy of multiple tumor entities among them head and neck squamous cell carcinoma (HNSCC). However, the success of radiotherapy is limited by the development of radiation resistances. Heterogeneous nuclear ribonucleoprotein K (hnRNPK) is a cofactor of p53 and represents a potential target for radio sensitization of tumor cells. In this study, we analyzed the impact of hnRNPK on the DNA damage response after gamma irradiation. By yH2AX foci analysis, we found that hnRNPK knockdown increases DNA damage levels in irradiated cells. Tumor cells bearing a p53 mutation showed increased damage levels and delayed repair. Knockdown of hnRNPK applied simultaneously with irradiation reduced colony-forming ability and survival of tumor cells. Taken together, our data shows that hnRNPK is a relevant modifier of DNA damage repair and tumor cell survival. We therefore recommend further studies to evaluate the potential of hnRNPK as a drug target for improvement of radiotherapy success.
Asunto(s)
Carcinoma de Células Escamosas/radioterapia , Neoplasias de Cabeza y Cuello/radioterapia , Ribonucleoproteína Heterogénea-Nuclear Grupo K/genética , Tolerancia a Radiación/genética , Proteína p53 Supresora de Tumor/genética , Carcinoma de Células Escamosas/genética , Carcinoma de Células Escamosas/patología , Línea Celular Tumoral , Supervivencia Celular/efectos de la radiación , Daño del ADN/efectos de la radiación , Técnicas de Silenciamiento del Gen , Neoplasias de Cabeza y Cuello/genética , Neoplasias de Cabeza y Cuello/patología , Histonas/genética , Humanos , Tolerancia a Radiación/efectos de la radiación , Carcinoma de Células Escamosas de Cabeza y Cuello , Células Madre/efectos de la radiaciónRESUMEN
Irradiation resistance is a major obstacle of head and neck squamous cell carcinoma (HNSCC) therapy, limiting treatment success and patient survival. The aim of our experiments was to identify irradiation-regulated proteins as potential drug targets. Two established HNSCC cell lines (HNSCCUM-01T and HNSCCUM-02T) were treated with a single 8Gy (Gray) fraction of irradiation. Changes in cellular protein expression were studied after 24h by means of 2D-electrophoresis and MALDI-TOF-mass spectrometry. Ninety-four differentially expressed proteins were identified. The expression levels of four proteins were regulated similarly in both cell lines after irradiation treatment, i.e., GRP78, PRDX, ACTC, and the heterogeneous nuclear ribonucleoprotein K (hnRNPK), suggesting a relevant role during irradiation response. hnRNPK as a p53 interacting protein was verified by Western blotting and immunocytochemical staining as well as functionally analyzed. Knock-down by the use of siRNA resulted in only slightly reduced viability, however, migratory activity was strongly reduced. Combined application of siRNA against hnRNPK and irradiation reduced migration almost completely. We conclude that hnRNPK is potentially implicated in the radiogenic response of HNSCC. The inhibition of hnRNPK might reduce the metastasizing potential of HNSCC especially in combination with irradiation and suggest that this molecule should be further evaluated in this context. BIOLOGICAL SIGNIFICANCE: We showed completely impaired migration of irradiated hnRNPK-knock-out HNSCC cells, suggesting this molecule as a potential drug target in combined treatment schedules.