Heritable germline epimutation of MSH2 in a family with hereditary nonpolyposis colorectal cancer.

Epimutations in the germline, such as methylation of the MLH1 gene, may contribute to hereditary cancer syndrome in human, but their transmission to offspring has never been documented. Here we report a family with inheritance, in three successive generations, of germline allele-specific and mosaic hypermethylation of the MSH2 gene, without evidence of DNA mismatch repair gene mutation. Three siblings carrying the germline methylation developed early-onset colorectal or endometrial cancers, all with microsatellite instability and MSH2 protein loss. Clonal bisulfite sequencing and pyrosequencing showed different methylation levels in different somatic tissues, with the highest level recorded in rectal mucosa and colon cancer tissue, and the lowest in blood leukocytes. This mosaic state of germline methylation with different tissue distribution could act as the first hit and provide a mechanism for genetic disease inheritance that may deviate from the mendelian pattern and be overlooked in conventional leukocyte-based genetic diagnosis strategy.

Molecular characterization of the 2011 Hong Kong scarlet fever outbreak.

A scarlet fever outbreak occurred in Hong Kong in 2011. The majority of cases resulted in the isolation of Streptococcus pyogenes emm12 with multiple antibiotic resistances. Phylogenetic analysis of 22 emm12 scarlet fever outbreak isolates, 7 temporally and geographically matched emm12 non-scarlet fever isolates, and 18 emm12 strains isolated during 2005-2010 indicated the outbreak was multiclonal. Genome sequencing of 2 nonclonal scarlet fever isolates (HKU16 and HKU30), coupled with diagnostic polymerase chain reaction assays, identified 2 mobile genetic elements distributed across the major lineages: a 64.9-kb integrative and conjugative element encoding tetracycline and macrolide resistance and a 46.4-kb prophage encoding superantigens SSA and SpeC and the DNase Spd1. Phenotypic comparison of HKU16 and HKU30 with the S. pyogenes M1T1 strain 5448 revealed that HKU16 displays increased adherence to HEp-2 human epithelial cells, whereas HKU16, HKU30, and 5448 exhibit equivalent resistance to neutrophils and virulence in a humanized plasminogen murine model. However, in contrast to M1T1, the virulence of HKU16 and HKU30 was not associated with covRS mutation. The multiclonal nature of the emm12 scarlet fever isolates suggests that factors such as mobile genetic elements, environmental factors, and host immune status may have contributed to the 2011 scarlet fever outbreak.

Epigenetic-genetic interactions in the APC/WNT, RAS/RAF, and P53 pathways in colorectal carcinoma.

PURPOSE: Early events in colorectal tumorigenesis include mutation of the adenomatous polyposis coli (APC) gene and epigenetic hypermethylation with transcriptional silencing of the O(6)-methylguanine DNA methyltransferase (MGMT), human mut L homologue 1 (hMLH1), and P16/CDKN2A genes. Epigenetic alterations affect genetic events: Loss of MGMT via hypermethylation reportedly predisposes to guanine-to-adenine or cytosine-to-thymine (G:C-->A:T) transition mutations in KRAS and P53, and silencing of hMLH1 leads to high levels of microsatellite instability (MSI-H)/mutator phenotype, suggesting that epigenetic-genetic subtypes exist. EXPERIMENTAL DESIGN: We evaluated the relationships of aberrant methylation of APC, MGMT, hMLH1, P16, N33, and five MINTs to mutations in APC, KRAS, BRAF, and P53 in 208 colorectal carcinomas. RESULTS: We found that APC hypermethylation was age related (P = 0.04), in contrast to the other genes, and did not cluster with CpG island methylator phenotype (CIMP) markers. Hypermethylation of APC concurrently with either MGMT or hMLH1 was strongly associated with occurrence of G-to-A transitions in APC [odds ratio (OR), 26.8; P < 0.0002 from multivariable logic regression model], but C-to-T transitions had no associations. There was no relationship of hypermethylation of any gene, including MGMT, with G-to-A or C-to-T transitions in KRAS or P53, although APC hypermethylation was associated with P53 mutation (P < 0.0002). CIMP with MSI-H due to hMLH1 hypermethylation, or CIMP with loss of MGMT expression in non-MSI-H tumors, was associated with BRAF mutation (OR, 4.5; P < 0.0002). CIMP was also associated with BRAF V600E T-to-A transversion (OR, 48.5; P < 0.0002). CONCLUSIONS: Our findings suggest that the heterogeneous epigenetic dysregulation of promoter methylation in various genes is interrelated with the occurrence of mutations, as manifested in epigenetic-genetic subgroups of tumors.

Microsatellite instability and novel mismatch repair gene mutations in northern Chinese population with hereditary non-polyposis colorectal cancer.

OBJECTIVE: Hereditary non-polyposis colorectal cancer (HNPCC) syndrome is the most common cause of hereditary colorectal cancer with an early age of onset. Microsatellite instability (MSI) and germline mutation in one of the DNA mismatch repair (MMR) genes are found in the majority of HNPCC families and provide an opportunity for genetic diagnosis and prophylactic screening. The MMR gene mutation spectrum may vary across different populations and be influenced by founder mutations that prevail in specific ethnic groups. China is a big and ancient nation with enormous genetic diversity, which is especially notable between the northern and southern Chinese populations. A MMR gene mutation database for the southern Chinese population based in Hong Kong has been previously established. This study compares the MMR gene mutation spectrum and the MSI of HNPCC between the northern and southern Chinese populations. METHODS: Twenty-five HNPCC families from northern China were systematically analyzed. The MSI analysis was performed using five loci in the USA National Cancer Institute (NCI) panel (D2S123, D5S346, BAT-25, BAT-26 and BAT-40) by PCR from the tumor and normal tissue. MSH2, MSH6 and MLH1 were performed using immunohistochemical staining. Two founder mutations of MSH2 and MLH1 were examined by PCR base analyses using primers flanking the two deletion sites (c.1452_1455delAATG in MSH2 and 1.8 kb deletion involving exon 11 of MLH1). RESULTS: Of the 25 families collected, 19 met Bethesda guideline (BG) 1 and six met BG3. Twenty-two (15.7%) were extra-colonic cancers with gastric cancer (in seven patients) being the most common cancer type. Of the 25 tumors analyzed, 21 (84%) were high level microsatellite instability (MSI-H) and four (16%) were microsatellite stable (MSS). Eighteen (86%) of the 21 MSI-H tumors showed loss of either the MLH1 or the MSH2 protein. Three MSI-H tumors and all four MSS tumors showed no loss of expression of the three MMR proteins. Out of the 21 patients with MSI-H tumors, 12 (57%) showed pathogenic germline mutations in either MLH1 (n = 8) or MSH2 (n = 4). Overall, three novel mutations (in patients H22, H17 and H29) have been identified. One of them, c.503_4insA, caused a frameshift mutation in the MLH1 gene. The other two were found in the MSH2 gene, including a frameshift (c.899_890insAT) and a splice junction (IVS7-1G-->A, SA of Exon 8) mutation. CONCLUSIONS: The results suggest a distinctly different mutation spectrum of MMR genes between northern and southern Chinese populations and call for a systematic, nationwide study to facilitate the design of a MMR gene mutation detection strategy tailored for individual populations in China.

MSH2 c.1452-1455delAATG is a founder mutation and an important cause of hereditary nonpolyposis colorectal cancer in the southern Chinese population.

Hereditary nonpolyposis colorectal cancer (HNPCC) accounts for approximately 2% of all colorectal cancer (CRC) cases and is the most common hereditary CRC syndrome. We have previously reported a high incidence of microsatellite instability (MSI) and germline mismatch repair (MMR) gene mutations in young Hong Kong Chinese with CRC. Ongoing studies at the Hereditary Gastrointestinal Cancer Registry in Hong Kong have revealed a unique germline MSH2 c.1452-1455delAATG mutation that has not been reported in other ethnic groups. Detailed analysis showed that this specific MSH2 mutation constituted 21% of all germline MMR gene mutations and 36% of all MSH2 germline mutations identified. We designed a specific PCR-based diagnostic test on paraffin-embedded tissues and identified this germline mutation in 2 (1.5%) of 138 consecutive patients with early-onset CRC (<46 years of age at diagnosis). Haplotype analysis was performed using 11 microsatellite markers located between D2S391 and D2S123. All 10 families had the same disease haplotype, suggesting a founder effect. These 10 families all originated from the Chinese province of Guangdong, which historically included Hong Kong. It is the most populous of the Chinese provinces, with a population of >93 million. Further analysis suggested that this founder mutation may date back to between 22 and 103 generations ago. The identification of this MSH2 founder mutation has important implications for the design of mutation-detection strategies for the southern Chinese population. Since there were major emigrations from Hong Kong and Guangdong province during the 19th and 20th centuries, this finding is also significant for Chinese communities worldwide.