Photon irradiation prompts autophagy in anaplastic thyroid cancer

Vollständiger Abstract

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Abstract Although promising results have been obtained for anaplastic thyroid cancer (ATC) therapy, this therapy still needs to be improved. In addition to the currently approved therapies, radiotherapy alone or combined with adjuvant immunotherapy could be beneficial for patients affected by ATC. The patients affected by this aggressive solid malignancy could benefit from the modulation of autophagy in cancer cells. This study focused on detecting autophagy players in ATCs and bursting autophagy process via photon irradiation to induce decay in irradiated ATCs. The transcript expression of autophagy genes was detected in tumor tissue resected from 19 patients and in C643 cells, four primary ATC cell lines and primary follicular thyroid cell line (Nthy-ori-3–1) photons irradiated with 4 or 6 Gy. The protein level of Beclin1 was detected by immunofluorescence in 10/19 patients. The levels of autophagy markers were detected by RT‒qPCR and western blotting in irradiated cells. Autophagy and maturation of autophagosome vesicles were monitored in C643 cells stably transfected with the GFP-RFP-LC3B plasmid. All patients included in the study exhibited significant overexpression of autophagy transcripts. Additionally, the Beclin1 protein was expressed in resected tumor tissue. Furthermore, the analysis of autophagy-related gene transcripts revealed significant increases in the expression of these genes in C643, Patient 2 and Patient 3 cells irradiated with 4 or 6 Gy. Additionally, irradiation with 4 or 6 Gy downregulated the expression of all the proteins involved in the autophagy process. Thus, the ongoing catabolic process was confirmed. Interestingly, the levels of AMPKα and its active phosphorylated form were strongly downregulated, which excluded its involvement in autophagy activation. 6 Gy photon irradiation caused an increase in both green and red fluorescence in C643-derived spheroids. An increase in fluorescence was detectable for up to 28 days. The spheroids gradually exhibited an increase in fluorescence, which probably caused the dismantling of their ultrastructure. This finding provides evidence of the bursting of autophagy and its ability to affect spheroid integrity. Photon irradiation exacerbates autophagy in anaplastic thyroid cancer cells and could represent a valid target for focused therapy against this aggressive malignancy.