Supplementary Material for: Resistance to Fas-Mediated Apoptosis Does Not Correlate to Structural Alterations or Expression Changes of the Death Receptor in Papillary Thyroid Carcinomas

Background: Malignant cells exhibit significant resistance to FAS-mediated cell death, through different processes, including FAS mutations, soluble FAS expression, or FAS transcriptional dysregulation by P53, eventually escaping from immune surveillance. Since thyroid carcinomas were shown to be resistant to FAS-mediated apoptosis, we investigated the above mechanisms in thyroid carcinoma samples. Methods: Thirty-seven thyroid carcinoma samples were analyzed for mutations in FAS exon 9 and TP53 exons 5–8 and protein expression by means of immunohistochemistry. Moreover, thyroid carcinoma mRNA samples were subjected to reverse transcription – PCR, to evaluate the relative expression of transmembrane FAS versus its soluble form. Results: Analysis revealed indications for TP53 mutations in the anaplastic carcinomas, but not in the other thyroid specimens examined for TP53 or FAS exon 9 mutations. FAS receptor expression was observed in almost all thyroid specimens (97%) with significant up-regulation in papillary carcinomas. P53 nuclear staining was observed only in anaplastic carcinomas. Full-length FAS mRNA was detected in all specimens examined, with soluble FAS mRNA being either absent or present in very low amounts. Conclusions: Our results denote that FAS death domain or TP53 DNA-binding domain mutations, down-regulation of FAS receptor expression, or expression of FAS soluble isoform are not responsible for the seeming inhibition of FAS-mediated apoptosis in papillary thyroid carcinoma cells.