The Clinical And Genetic Analysis Of Multiple Endocrine Neoplasia Type 2

MEN2(Multiple Endocrine Neoplasia Type 2) is an autosomal dominant syndrome with an incidence about 1/30000. It was classified as three subtypes according to clincal features, MEN2 A, MEN2 B and FMTC(Familiar Medullary Thyroid Carcinoma). MEN2 A is characterized as MTC, about 50% carriers affect PCC(Pheochromocytoma), 20-30% affect PHPT(Primary Hyperparathyroid Adenoma); MEN2 B carriers affect MTC, PCC and ganglioneuromatos, marfanoid habitus, but PHPT is not included; FMTC is defined as solitary MTC component. RET proto-oncogene mutation was believed the molecular pathogenesis of MEN2. Correlation between RET genotype and MEN2 has been established. However, there is few data from Chinese population and no genetic mechanism other than RET has been reported.This study analysed the clinical features and geno-phenotype correlations in 135 MEN2 patients from 53 unrelated MEN2 pedigrees across China diagnosed in Shanghai Clinical Center for Endocrine and Metabolic Diseases. Hot spot of RET proto-oncogene(exon 8,10,11,13,14,15 and 16) were analyzed by PCR-Sanger sequencing. A total of 125 MEN2 A patients and 10 MEN2 B patients were diagnosed,11 types of RET mutation were identified as following: p.C634R(n=19,42.22%),p.C634Y(n=12,26.67%),p.C634W(n=4,8.89%),p.C634S(n=3,6.67%),p.C634F(n=2,4.44%),p.C620R(n=1,2.22%),p.C620S(n=1,2.22%),p.C611Y(n=1,2.22%),p.C618R(n=1,2.22%),p.E768D(n=1,2.22%)in MEN2 A and p.M918 T in MEN2 B. Four mutations as p.C620 R,p.C620 S,p.C611 Y and p.E768 D were firstly reported in Chinese population. When comparing mutations located in exon 10 and exon 11, carriers with exon 10 mutations showed no parathyroid tumor and later MTC onset age than exon 11 mutations carrier(p<0.05). For different mutant types at RET codon 634, there were significant difference regarding as age of initial PCC surgery. Hereditary MTC showed earlier age onset than sporadic MTC(36.27±14.34 vs 45.34±13.57,p<0.01)and habored higher rate of LN metastasis(62.50% vs 39.29%, p<0.05).This study further applied Affemtrix SNP 6.0 array and exome sequencing on matched MEN2 A tumors and germline DNA. Fewer mutations per tumor were observed in MEN2 A associated tumors compared to none-endocrine malignance(7.58 vs 65.60, P <0.01). A total of 32 somatic mutations were identified in the nine MEN2 associated tumors, of which 28(87.5%) were point mutations and four(12.5%) were small insertions, duplications or deletions. Recurrent alterations were found in 13 genes with either mutations or copy number alteration. We further validated somatic alterations of the above 13 genes in an independent set of patients via Sanger sequencing(MTC,N=20; PCC, N=20), but no recurrent mutations was found. We evaluated function of EIF4G1 mutation(p.E1147V) by overexpression in MTC cell line, TT cell. Mutant EIF4G1 led to increased cell proliferation and RET/MAPK phosphorylation in TT cell. This study suguests RET was the primary driver in MEN2 associated tumors while low-frequent alterations such as EIF4G1 might participate in MEN2 A associated tumorigenesis, possibly by regulating the activity of RET pathway.