IMPE Abstracts

In pediatrics, thyroid tumor stratification is difficult to assess. Epithelial-mesenchymal transition (EMT), plays a role in tumor development. In human carcinomas Brachyury (Brachy) has been identified as a regulator of EMT associated to malignancy. The Insulin Like Growth Factors (IGFs) are mitogens that play important roles in both normal and neoplastic growth. To date, no information about Brachy and IGF1R expression in pediatric thyroid nodular disease is available.

Aim: To evaluate Brachy and IGF1R expression in thyroid nodular samples from pediatric patients and to study the effect of Brachy and IGF1R overexpression in a thyroid papillary carcinoma cell line (TPC) in vitro.

Methods: Paraffin-embedded samples from pediatric patients with Thyroid Papillary Carcinomas (TPCa), Follicular Adenomas (FA) or Benign Thyroid Nodular disease (BTN) were processed for Brachy and IGF1R immunostaining. TPC cells were used to obtain clones overexpressing Brachy (TPC.BR4 and TPC.BR6) and IGF1R (TPC.IGF1R5 and TPC.IGF1R7). Gene expression was quantified by rqPCR. Phaloidin IF staining, Viability and apoptosis assays (3 days) and wounding assays (24h) were carried out. Protein extracts were obtained from whole lysates and processed by western blotting (WB).

Results: 50 samples were analysed, 17 from BTN. Only TPCa and FA showed positive staining for Brachy (15/24TPCa;5/9FA) and IGF1R (11/24TPCa;4/9FA). In carcinomas, positivity for IGF1R was only detected when Brachy was present. In vitro, Brachy overexpression in TPC cells increased proliferation (** P<0,01 TPC.BR4, *** P<0,005 TPC.BR6) while IGF1R overexpression resulted in a lower proliferation rate (**** P<0,0001) compared to parental cells. Cell migration was also higher when Brachy was overexpressed (* P< 0.05 TPC.BR4 and TPC.BR6 vs TPC), e-cadherin expression was diminished, and mesenchymal markers (vimentin-fibronectin) were increased. The opposite profile was found in IGF1R overexpressing clones. Initial risk stratification showed an association between Brachy and high risk, while IGF1R positive immunostaining was associated with low risk initial assessment.

Conclusion: Pediatric TPCa showed positive labeling for both Brachy and IGF1R. While Brachy expression was associated to high risk, IGF1R expression was associated to low risk initial stratification. In vitro, Brachy overexpression led to a mesenchymal like phenotype in TPC.BR4 and TPC.BR6 clones. Conversely, IGF1R expression seems to favor epithelial features on TPC.IGF1R5 and TPC.IGF1R7 clones. These results suggest potential opposite roles for Brachy and IGF1R in the biology of thyroid tumors.