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RET germline mutations identified by exome sequencing in a Chinese multiple endocrine neoplasia type 2A/familial medullary thyroid carcinoma family.

Qi XP, Ma JM, Du ZF, Ying RB, Fei J, Jin HY, Han JS, Wang JQ, Chen XL, Chen CY, Liu WT, Lu JJ, Zhang JG, Zhang XN - PLoS ONE (2011)

Bottom Line: Massively parallel sequencing revealed four missense mutations of RET.In the maternal origin, p.C634Y caused bilateral MTC in all 5 cases and bilateral pheochromocytoma in 2 of the 5; the earliest onset age was 28 years.The results confirmed the successful clinical utility of whole exome sequencing, and our data suggested that the p.C634Y/V292M/R67H/R982C mutation of RET exhibited a more aggressive clinical phenotype than p.C634Y or p.V292M/R67H/R982C, while p.V292M/R67H/R982C presented a relatively milder pathogenicity of MTC and likely predisposed to FMTC.

View Article: PubMed Central - PubMed

Affiliation: Department of Urologic Surgery, The 117th PLA Hospital, Hangzhou, Zhejiang, China. qxplmd@vip.sina.com

ABSTRACT

Background: Whole exome sequencing provides a labor-saving and direct means of genetic diagnosis of hereditary disorders in which the pathogenic gene harbors a large cohort of exons. We set out to demonstrate a suitable example of genetic diagnosis of MEN 2A/FMTC (multiple endocrine neoplasia type 2/familial medullary thyroid carcinoma) using this approach.

Methodology/principal findings: We sequenced the whole exome of six individuals from a large Chinese MEN2A/FMTC pedigree to identify the variants of the RET (REarranged during Transfection) protooncogene and followed this by validation. Then prophylactic or surgical thyroidectomy with modified or level VI lymph node dissection and adrenalectomy were performed for the carriers. The cases were closely followed up. Massively parallel sequencing revealed four missense mutations of RET. We unexpectedly discovered that the proband's daughter with MEN 2A-related MTC presented a novel p.C634Y/V292M/R67H/R982C compound mutation, due to the involvement of p.C634Y in the proband with MEN 2A and p.V292M/R67H/R982C in the proband's husband with FMTC. In the maternal origin, p.C634Y caused bilateral MTC in all 5 cases and bilateral pheochromocytoma in 2 of the 5; the earliest onset age was 28 years. In the paternal origin, one of the six p.V292M/R67H/R982C carriers presented bilateral MTC (70 years old), one only had bilateral C-cell hyperplasia (44 years), two had bilateral multi-nodules (46 and 48 years) and two showed no abnormality (22 and 19 years).

Conclusions/significance: The results confirmed the successful clinical utility of whole exome sequencing, and our data suggested that the p.C634Y/V292M/R67H/R982C mutation of RET exhibited a more aggressive clinical phenotype than p.C634Y or p.V292M/R67H/R982C, while p.V292M/R67H/R982C presented a relatively milder pathogenicity of MTC and likely predisposed to FMTC.

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Portion of genomic DNA and RT-PCR product sequencing from tumor tissues.A, Direct sequencing consequences of RT-PCR products from PHEO tissue of III-12 indicated a heterozygous G/A mutation at codon 634, which was consistent with the results of DNA from blood and tumor tissues and whole exome sequencing. B, C, D, Direct sequencing consequences of RT-PCR products from MTC tissue of III-11 indicated heterozygous G/A, C/T, and G/A mutations at codons 292, 982, and 67, respectively, which was consistent with the results of DNA from blood and tumor tissues and whole exome sequencing.
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pone-0020353-g003: Portion of genomic DNA and RT-PCR product sequencing from tumor tissues.A, Direct sequencing consequences of RT-PCR products from PHEO tissue of III-12 indicated a heterozygous G/A mutation at codon 634, which was consistent with the results of DNA from blood and tumor tissues and whole exome sequencing. B, C, D, Direct sequencing consequences of RT-PCR products from MTC tissue of III-11 indicated heterozygous G/A, C/T, and G/A mutations at codons 292, 982, and 67, respectively, which was consistent with the results of DNA from blood and tumor tissues and whole exome sequencing.

Mentions: The Sanger sequencing scan of the whole RET gene was completely consistent with the whole exome sequencing. The proband (III-12) carried the p.C634Y mutation within exon 11, and so did her family members III-15, III-20, III-23 and IV-21. Unfortunately and interestingly, the proband's husband (III-11) and his family members II-5, III-3, III-8, IV-3 and IV-7 carried the p.R67H mutation within exon 2, the p.V292M mutation within exon 5 and the p.R982C polymorphism within exon 18 (Table 1). Thus, their daughter (IV-10) inherited four substitutions of p.C634Y/V292M/R67H/R982C (GenBank accession number for p.C634Y: JF276429; p.V292M/R67H/R982C: JF273638), of which p.C634Y, p.V292M and p.R982C were respectively confirmed by Kpn I, Nco I and Rsa I digestion (Fig. 2). Genomic DNA and RT-PCR product sequencing from tumor tissues were both consistent with the results from blood and presented heterozygosity (Fig. 3). The four alterations were absent in 100 healthy controls. Haplotype analysis of four microsatellite markers from the 9 related members (II-5, II-6, III-10, III-11, III-12, III-13, III-15, IV-10 and IV-11) confirmed that p.C634Y was of maternal origin and p.V292M/R67H/R982C was paternal and located in a common allele (Fig. 4).


RET germline mutations identified by exome sequencing in a Chinese multiple endocrine neoplasia type 2A/familial medullary thyroid carcinoma family.

Qi XP, Ma JM, Du ZF, Ying RB, Fei J, Jin HY, Han JS, Wang JQ, Chen XL, Chen CY, Liu WT, Lu JJ, Zhang JG, Zhang XN - PLoS ONE (2011)

Portion of genomic DNA and RT-PCR product sequencing from tumor tissues.A, Direct sequencing consequences of RT-PCR products from PHEO tissue of III-12 indicated a heterozygous G/A mutation at codon 634, which was consistent with the results of DNA from blood and tumor tissues and whole exome sequencing. B, C, D, Direct sequencing consequences of RT-PCR products from MTC tissue of III-11 indicated heterozygous G/A, C/T, and G/A mutations at codons 292, 982, and 67, respectively, which was consistent with the results of DNA from blood and tumor tissues and whole exome sequencing.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3105051&req=5

pone-0020353-g003: Portion of genomic DNA and RT-PCR product sequencing from tumor tissues.A, Direct sequencing consequences of RT-PCR products from PHEO tissue of III-12 indicated a heterozygous G/A mutation at codon 634, which was consistent with the results of DNA from blood and tumor tissues and whole exome sequencing. B, C, D, Direct sequencing consequences of RT-PCR products from MTC tissue of III-11 indicated heterozygous G/A, C/T, and G/A mutations at codons 292, 982, and 67, respectively, which was consistent with the results of DNA from blood and tumor tissues and whole exome sequencing.
Mentions: The Sanger sequencing scan of the whole RET gene was completely consistent with the whole exome sequencing. The proband (III-12) carried the p.C634Y mutation within exon 11, and so did her family members III-15, III-20, III-23 and IV-21. Unfortunately and interestingly, the proband's husband (III-11) and his family members II-5, III-3, III-8, IV-3 and IV-7 carried the p.R67H mutation within exon 2, the p.V292M mutation within exon 5 and the p.R982C polymorphism within exon 18 (Table 1). Thus, their daughter (IV-10) inherited four substitutions of p.C634Y/V292M/R67H/R982C (GenBank accession number for p.C634Y: JF276429; p.V292M/R67H/R982C: JF273638), of which p.C634Y, p.V292M and p.R982C were respectively confirmed by Kpn I, Nco I and Rsa I digestion (Fig. 2). Genomic DNA and RT-PCR product sequencing from tumor tissues were both consistent with the results from blood and presented heterozygosity (Fig. 3). The four alterations were absent in 100 healthy controls. Haplotype analysis of four microsatellite markers from the 9 related members (II-5, II-6, III-10, III-11, III-12, III-13, III-15, IV-10 and IV-11) confirmed that p.C634Y was of maternal origin and p.V292M/R67H/R982C was paternal and located in a common allele (Fig. 4).

Bottom Line: Massively parallel sequencing revealed four missense mutations of RET.In the maternal origin, p.C634Y caused bilateral MTC in all 5 cases and bilateral pheochromocytoma in 2 of the 5; the earliest onset age was 28 years.The results confirmed the successful clinical utility of whole exome sequencing, and our data suggested that the p.C634Y/V292M/R67H/R982C mutation of RET exhibited a more aggressive clinical phenotype than p.C634Y or p.V292M/R67H/R982C, while p.V292M/R67H/R982C presented a relatively milder pathogenicity of MTC and likely predisposed to FMTC.

View Article: PubMed Central - PubMed

Affiliation: Department of Urologic Surgery, The 117th PLA Hospital, Hangzhou, Zhejiang, China. qxplmd@vip.sina.com

ABSTRACT

Background: Whole exome sequencing provides a labor-saving and direct means of genetic diagnosis of hereditary disorders in which the pathogenic gene harbors a large cohort of exons. We set out to demonstrate a suitable example of genetic diagnosis of MEN 2A/FMTC (multiple endocrine neoplasia type 2/familial medullary thyroid carcinoma) using this approach.

Methodology/principal findings: We sequenced the whole exome of six individuals from a large Chinese MEN2A/FMTC pedigree to identify the variants of the RET (REarranged during Transfection) protooncogene and followed this by validation. Then prophylactic or surgical thyroidectomy with modified or level VI lymph node dissection and adrenalectomy were performed for the carriers. The cases were closely followed up. Massively parallel sequencing revealed four missense mutations of RET. We unexpectedly discovered that the proband's daughter with MEN 2A-related MTC presented a novel p.C634Y/V292M/R67H/R982C compound mutation, due to the involvement of p.C634Y in the proband with MEN 2A and p.V292M/R67H/R982C in the proband's husband with FMTC. In the maternal origin, p.C634Y caused bilateral MTC in all 5 cases and bilateral pheochromocytoma in 2 of the 5; the earliest onset age was 28 years. In the paternal origin, one of the six p.V292M/R67H/R982C carriers presented bilateral MTC (70 years old), one only had bilateral C-cell hyperplasia (44 years), two had bilateral multi-nodules (46 and 48 years) and two showed no abnormality (22 and 19 years).

Conclusions/significance: The results confirmed the successful clinical utility of whole exome sequencing, and our data suggested that the p.C634Y/V292M/R67H/R982C mutation of RET exhibited a more aggressive clinical phenotype than p.C634Y or p.V292M/R67H/R982C, while p.V292M/R67H/R982C presented a relatively milder pathogenicity of MTC and likely predisposed to FMTC.

Show MeSH
Related in: MedlinePlus