Abstract
We recently reported of the distribution of REarranged during Transfection (RET) mutations in multiple endocrine neoplasia 2 (MEN2) in Denmark. In the study, we only included sequence variants classified as pathogenic in the ARUP database on July 1, 2016 (1). Since then, the database has changed the classification of the I852M variant from pathogenic to uncertain (
Our index patient, a 69-year-old female, was referred due to a slowly growing goiter. The medical history was unremarkable apart from a T1bN0M0 breast carcinoma eight years before referral. Basal serum calcitonin was 9500 ng/L (<7.3 ng/L). Fine-needle aspiration, ultrasonography, and fluorodeoxyglucose positron emission tomography were consistent with MTC and possible neck metastases. Preoperative urine catecholamines were normal. The patient underwent total thyroidectomy and bilateral neck dissection of levels III–VII. Pathology revealed a unifocal T3N0M0 MTC. Postoperatively basal calcitonin normalized. Mutational analysis of RET using DNA from peripheral blood leucocytes and formalin-fixed paraffin-embedded MTC tissue identified the I852M (c.2556C>G) germline variant and a novel E632_V637delinsAA (c.1895_1910delinsCAGC) somatic in-frame variant, respectively (see Supplementary Data; Supplementary Data are available online at
There was no family history of MEN2A. Both parents died before the diagnosis of MTC in the index patient, at 96 and 76 years of age. A brother did not carry the variant, but two sons aged 37 and 41 years were identified as carriers. Both sons had normal basal serum calcitonin, parathyroid hormone, and urine catecholamines. Ultrasonography of the neck identified a thyroid cyst <10 mm in the oldest brother, and fine-needle aspiration indicated a benign cyst. Based on the literature at the time and thorough multidisciplinary discussions, the two sons were offered prophylactic thyroidectomies. They underwent surgery at 39 and 42 years of age. The histology of each showed C-cell hyperplasia by calcitonin staining but no MTC. All four grandchildren were tested and found to be I852M negative. Follow-up of all three I852M carriers has been without clinical or biochemical signs of MTC, pheochromocytoma, or hyperparathyroidism for 5.5 years.
Correct classification of RET germline variants is crucial due the implications for carriers of pathogenic variants. This is best exemplified by the RET L791F variant, which was initially reported as pathogenic and several years afterwards relegated as benign. We aimed to assess whether this was also the case for the I852M variant.
Our systematic literature review identified two families and one single patient (comprising a total of eight I852M carriers), three in silico studies and one in vitro study (Supplementary Data).
A RET variant is considered causative of MEN2 if it segregates with MEN2 manifestations in at least two individuals within a family. Neither of the reported families fulfills these criteria (3,4). Also, none of the identified I852M carriers exhibits more than one MEN2 manifestation.
A similar scenario was seen in our family, where only one MEN2 manifestation was present (MTC in the index patient). In this MTC, we identified a RET E632_V637delinsAA (c.1895_1910delinsCAGC) somatic variant that introduces deletion of six and inclusion of two novel amino acids. The variant has not previously been reported in the COSMIC database. However, several other somatic deletions such as the c.1892_1914delinsCG, c.1894_1917delinsCACCGT, c.1894_1906delinsAGCT, and c.1895_1918delinsTGCGGC have been reported in this area. The c.1895_1910delinsCAGC variant, as well as the four other reported variants, all lead to deletion of the important Cys-634, which is a well-known RET hot-spot codon associated with MEN2. Consequently, it seems likely that the E632_V637delinsAA somatic variant is an activating mutation responsible for the MTC in this patient.
Other methods to assess the pathogenicity of a RET variant include in silico studies, in vitro studies, and determination of allelic frequency. Among the identified in silico studies, two of three considered the I852M variant benign, while one considered it uncertain. Only the in vitro study classified the variant as pathogenic (4). Meanwhile, the dbSNP database reports the allelic frequency as 0.00015, which is considerably higher than the estimated prevalence of MEN2 (1:35,000), supporting that the variant is benign.
We found no solid evidence supporting that the I852M germline variant causes MEN2A. In fact, most of the published literature indicates that the variant is benign. However, until further evidence arises, the variant seems best classified as uncertain.
Footnotes
Acknowledgment
This work was supported by the University of Southern Denmark, the Region of Southern Denmark, Odense University Hospital, Copenhagen University Hospital, the Danish Cancer Society, the Danish Cancer Research Foundation, and the A.P. Moeller Foundation.
Author Disclosure Statement
The authors declare that no competing financial interests exist.