Immunostaining and RT-PCR: different approaches to search for RET rearrangements in patients with papillary thyroid carcinoma

Abstract

Different techniques of molecular biology have been used to screen for RET rearrangements. More recently, immunohistochemistry has been used, assuming that RET is not expressed in normal thyroid follicular cells. The present study was designed to define the prevalence of RET expression in patients with papillary thyroid carcinoma, by immunohistochemistry and by RT-PCR; to search specifically for RET/PTC-1; -2; -3 rearrangements using RT-PCR, and to compare results obtained by immunohistochemistry with those obtained by RT-PCR. Immunohistochemistry was performed using a polyclonal antibody against tyrosine kinase domain of Ret protein. Screening for RET/PTC1-3 was performed using RT-PCR and specific primers for each rearrangement; complementarily, a subset of cases were tested using RET exon 10/11 primers designed to detect the expression of the wild-type RET. Positive staining was observed in 30 of 39 (77%) tumours. RET/PTC1-3 rearrangements were detected in 8 of 32 (25%) cases. Ten of 15 (67%) cases expressed the wild-type RET. Two tumours characterised by positive immunostaining, absence of RET 5' expression and absence of RET/PTC1-3 expression were considered as expressing a RET rearrangement different from RET/PTC-1, -2, or -3. In 3 of 10 tumours, expression of the wild-type RET coexisted with the expression of a RET rearrangement. Positive staining does not necessarily mean the presence of a rearrangement; it may correspond to the expression of the wild-type RET, RET rearrangement or both. On the contrary, positive staining without evidence for the expression of the extracellular domain of RET is highly suggestive of a RET rearrangement independently of the type. Refinement of diagnosis depends on RT-PCR with specific primers.