Role of NFκB and HIF-1α in Follicular Thyroid Neoplasm Progression

Background: Several gene abnormalities have been described in follicular thyroid carcinomas (FTC), but no study evaluates topographically vascular (hypoxia inducible factor-1α, HIF-1α) and inflammatory signaling pathways (nuclear factor κB, NFκB) by tumor phenotype.

Design: We selected 9 hyperplastic nodules, 22 adenomas, 14 minimally-invasive FTC, 24 widely-invasive FTC, and 13 anaplastic carcinomas (WHO criteria). Total RNA was extracted from normal and neoplastic tissues by hot acidic phenol, DNase I-treated, phenol extracted and cleaned (RNeasy columns). T7-(dT24) oligomer was used for priming the first-strand cDNA synthesis and the resultant cDNA was phenol/ chloroform extracted, and used as template for cRNA synthesis (T7 MegaScript In Vitro Transcription Kit). The cRNA was fragmented, Cy3- and Cy5-labeled, and hybridized to the human GeneChip X3P Array noncompetitively. Cross-validated gene expression analyses (CGEA) were performed (expression factor>2, significance<0.01), and variables studied regarding the histological diagnosis. Significant variables were evaluated immunohistochemically using standard protocols for the pathway last effector.

Results: Tumor infiltrating lymphoxytes were absent in 68/73 (93%) neoplasms, regardless of histologic subtype. CGEA showed down-regulation of ubiquitin C and sequestosome 1 in FTC, which directly correlated with CDC34, SKP1/2 (HIF-1α degradation pathway), and inversely with IκBα (inhibition of NFκB pathway). Across all neoplasms the positive cell percentage was higher in the internal compartments. NFκB-p50 and p65 (p<001) as well as HIF-1α (p=0.049) immunoexpressions increase with increasing neoplastic grade accordingly, in particular in internal compartments. The expression of NFκB was low across all the lesions, the greatest degree of immunoexpression seen in internal compartments of anaplastic carcinomas (4%). The hyperplastic nodules showed an exceptional level of expression of HIF-1α in the internal compartment (over 16%, ∼25x the highest percentage observed in any other lesion)

Conclusions: Ubiquitylation down-regulation of follicular tumor cells results in peripheral down-regulated NFκB pathways (p65 and p50) and HIF-1α, but increase expression in internal compartments with more aggressive tumor behavior suggesting an involvement in follicular thyroid tumor progression.