DETECTION OF RET MUTATIONS IN MULTIPLE ENDOCRINE NEOPLASIA TYPE 2A AND FAMILIAL MEDULLARY-THYROID CARCINOMA BY DENATURING GRADIENT GEL-ELECTROPHORESIS

Germline missense mutations within the coding region of the RET proto- oncogene have recently been described in patients with the dominantly inherited cancer syndromes, multiple endocrine neoplasia type 2a (MEN 2a) and familial medullary thyroid carcinoma (FMTC). To date, the sequ ence variations occur in RET exons 10 and 11 and alter highly conserve d cysteine residues in the proposed extracellular domain at codons 609 , 611, 618, 620, and 634. To expedite rapid screening of populations a t risk of MEN 2a or FMTC, we developed a PCR-based denaturing gradient gel electrophoresis (DGGE) strategy that detects polymorphisms occurr ing at all five Cys codons in both RET exons using identical gel condi tions. In this report, the screening results from DGGE analysis of 15 distinct MEN 2a and FMTC mutations are shown. Each mutation generated a clearly distinguishable and unique homo- and heteroduplex band patte rn. Given the highly efficient, reproducible, and sensitive nature of this approach, DGGE is particularly appropriate for rapid, large-scale screening of patients. Since prior knowledge of the RET mutation is u nnecessary for analysis, DGGE is potentially valuable for distinguishi ng germline from seemingly sporadic medullary thyroid cancer as well a s identifying novel sequence changes. (C) 1996 Wiley Liss, Inc.