Distinct mutations in MLH1 and MSH2 genes in hereditary non-polyposis colorectal cancer (HNPCC) families from China

Hereditary non-polyposis Colorectal Cancer (HNPCC) is an autosomal dominant inheritance syndrome. HNPCC is the most common hereditary variant of colorectal cancer (CRC), which accounts for 2-5% CRCs, mainly due to hMLH1 and hMSH2 mutations that impair DNA repair functions. Our study aimed to identify the patterns of hMSH2 and hMLH1 mutations in Chinese HNPCC patients. Ninety-eight unrelated families from China meeting Amsterdam or Bethesda criteria were included in our study. Germline mutations in MLH1 and MSH2 genes, located in the exons and the splice-site junctions, were screened in the 98 probands by direct sequencing. Eleven mutations were found in ten patients (11%), with six in MLH1 (54.5%) and five in MSH2 (45.5%) genes. One patient had mutations in both MLH1 and MSH2 genes. Three novel mutations in MLH1 gene (c.157_160delGAGG, c.2157dupT and c.-64G>T) were found for the first time, and one suspected hotspot in MSH2 (c.1168C>T) was revealed.     

Molecular diagnostics of cancer predisposition: hereditary non-polyposis colorectal carcinoma and mismatch repair defects

Hereditary non-polyposis colorectal carcinoma accounts for 5–13% of all colorectal carcinomas and is inherited in a dominant fashion. Two different forms can be distinguished. Type I is restricted to colorectal cancers, whereas type II patients acquire acolorectal, endometrial, gastric, small intestinal and transitional carcinomas of the upper urinary tract. Germline mutations in the human mismatch repair genes (hMSH2, hMSH6, hMLH1, hPMS2) account for the majority of hereditary non-polyposis colorectal carcinoma. As a result of the mismatch repair deficiency, replication errors are not repaired, resulting in a mutator phenotype. Simple repetitive sequences (microsatellites) are especially prone to replication errors and analysis of their stability combined with immunohistochemical analysis of mismatch repair protein expression provides a rapid diagnostic strategy. For patients either (1) fulfilling the Amsterdam criteria for HNPCC, (2) with synchronous or metachronous hereditary non-polyposis colorectal carcinoma-related tumors, (3) with hereditary non-polyposis colorectal carcinoma-related tumors before the age of 45 and/or (4) with right sided CRC and mucinous, solid, or cribriform growth patterns, screening for mismatch repair deficiencies should be performed. The identification of colorectal cancers displaying a mutator phenotype has implications for both treatment and prognosis.     

DHPLC mutation analysis of the hereditary nonpolyposis colon cancer (HNPCC) genes hMLH1 and hMSH2

Denaturing high-performance liquid chromatography (DHPLC) is an efficient method for detection of mutations involving a single or few numbers of nucleotides, and it has been successfully used for mutation detection in disease-related genes. Colorectal cancer is one of the most common cancers, and mutations in the genes for hereditary nonpolyposis colon cancer (HNPCC), hMLH1 and hMSH2, also involve mainly point mutations. Sequence analysis is supposed to be a screening method with high sensitivity; however, it is time-consuming and expensive. We therefore decided to test sensitivity and reproducibility of DHPLC for 71 sequence variants in hMLH1 and hMSH2 initially found by sequence analysis in DNA samples of German HNPCC patients. DHPLC conditions of the PCR products were based on the melting pattern of the wild-type sequence of the corresponding PCR fragments. All but one of the 71 mutations was detected using DHPLC (sensitivity of 97%). Running time per sample averaged only 7 min, and the system is highly automated. Thus DHPLC is a rapid and sensitive method for the detection of hMLH1 and hMSH2 sequence variants.     

Presymptomatic diagnosis using a deletion of a single codon in families with hereditary non-polyposis colorectal cancer

The diagnosis of hereditary non-polyposis colorectal cancer (HNPCC) is often confirmed by a mutation in one of several mismatch-repair genes, in particular MLH1, MSH2 and MSH6. Presymptomatic diagnosis requires the identification of a mutation causing the disease. Three different deletions of a single amino acid codon have previously been published as assumed pathogenic. The objective of this study was to determine if an MSH2 3 base pair in-frame deletion (N596del) could be used in presymptomatic screening of at-risk individuals. We report on five HNPCC families with the N596del mutation, identified after mutation screening of MSH2 and MLH1. All patients in the families were haplotyped using markers flanking the MSH2 gene. The haplotypes revealed that the five families with high probability descended from only two founders. The N596del segregated with the HNPCC phenotype with lod scores of 3.2 and 2.0 at the recombination fraction of 0.0 in the two founder families. Sequencing of MSH2 and MLH1 did not reveal other pathogenic mutations, and N596del was not identified in 50 healthy controls. The mutation has previously been found expressed in mRNA, and is located in a conserved domain. The results support the hypothesis that N596del is the disease causing mutation and not a clinically silent variation. On this basis, the application of the MSH2 N596del mutation, in presymptomatic screening of HNPCC families, is recommended.     

Deletions account for 17% of pathogenic germline alterations in MLH1 and MSH2 in hereditary nonpolyposis colorectal cancer (HNPCC) families

Hereditary nonpolyposis colorectal cancer (HNPCC) is due to defects in DNA mismatch repair (MMR) genes MSH2, MLH1, MSH6, and to a lesser extent PMS2. Of 466 suspected HNPCC families, we defined 54 index patients with either tumors of high microsatellite instability (MSI-H) and/or loss of expression for either MLH1, MSH2, and/or MSH6, but without a detectable pathogenic point mutation in these genes. This study cohort was augmented to 64 patients by 10 mutation-negative index patients from Amsterdam families where no tumors were available. Deletion/duplication screening using the multiplex ligation-dependent probe amplification (MLPA) revealed 12 deletions in MSH2 and two deletions in MLH1. These deletions constitute 17% of pathogenic germline alterations but elucidate the susceptibility to HNPCC in only 22% of the mutation-negative study cohort, pointing towards other mutation mechanisms for an inherited inactivation of MLH1 or MSH2. We describe here four novel deletions. One novel and one known type of deletion were found for three and two unrelated families, respectively. MLPA analysis proved a reliable method for the detection of genomic deletions in MLH1 and MSH2; however, sequence variations in the ligation-probe binding site can mimic single exon deletions.     

Identity,community and care in online accounts of hereditary colorectal cancer syndrome

Sociological literature has explored how shifts in the point at which individuals may be designated as diseased impact upon experiences of ill health. Research has shown that experiences of being genetically “at risk” are shaped by and shape familial relations, coping strategies, and new forms of biosociality. Less is known about how living with genetic risk is negotiated in the everyday and over time, and the wider forms of identity, communities and care this involves. This article explores these arrangements drawing on online bloggers’ accounts of Familial Adenomatous Polyposis (FAP). We show how accounts of genetic risk co-exist with more palpable experiences of FAP in everyday life, notably the consequences of prophylactic surgeries. We consider how the act of blogging represents but also constitutes everyday experiences of hereditary cancer syndrome as simultaneously ordinary and exceptional, and reflect on the implications of our analysis for understanding experiences of genetic cancer risk.     

GSTM1 and GSTT1 polymorphisms as modifiers of age at diagnosis of hereditary nonpolyposis colorectal cancer (HNPCC) in a homogeneous cohort of individuals carrying a single predisposing mutation

The variability in phenotype that occurs for so-called ‘single-gene disorders’ may be because of germline alterations in numerous primary and “modifier” genes. Within HNPCC families harbouring the same primary predisposing mutation, differences exist in the site of cancer, age of onset of disease symptoms and, consequently, survival until diagnosis of disease. The current study investigated a cohort of 129 individuals, from 13 different families, who harbour the identical nonsense mutation (C1528T) in the hMLH1 gene, predisposing them primarily to Lynch I syndrome. This cohort was screened for previously described polymorphisms in the glutathione-S-transferase genes, viz. GSTT1 and GSTM1. Male null carriers for both GSTT1 and GSTM1 were approximately three times more at risk of developing cancer at an earlier age when compared to non-null males. This work, particularly because of the relatively large “homogeneous” primary mutation cohort, provides evidence that genotypic changes distinct from the primary ‘HNPCC-causing’ mutation, influence the survival period until diagnosis of disease. It provides an impetus for expanding the study to include a wider range of candidate modifier genes. Such work may potentially lead to the development of individualised interval screening regimens for individuals with varying modifier genotypes—an attractive option in a resource-poor country.     

Maffucci syndrome and soft tissue sarcoma: a case report

Background

Maffucci syndrome, a congenital mesodermal dysplasia characterized by multiple enchondromas and hemangiomas, was first described in 1881, and 200 cases have been reported in the literature since then. Its etiology is unknown, there is no predilection for race or sex, and the development of lesions usually occurs in puberty. The risk of sarcomatous transformation is about 25%.

Case presentation

The initial investigation of the case reported here focused on the diagnosis and treatment of malignancy, and the first diagnostic hypothesis was thrombosed hemangioma. After histopathologic confirmation of soft tissue sarcoma, examinations were performed to stage the tumor and enchondromas were found in ribs. The final diagnosis was Maffucci syndrome with malignant transformation.

Conclusion

Treatment should aim at symptom relief and early detection of malignancies; no therapy is indicated for asymptomatic patients. As in the case reported here, bone or soft tissue lesions that grow or become painful should be biopsied.

     

The colorectal cancer stem-like cell hypothesis: a pathologist's point of view

Colorectal cancer is the fourth most common cancer in men and the third most common cancer in women worldwide. The partial failure of classic therapeutic options makes scientists to doubt the efficacy of systemic treatments in targeting the essential cell populations and achieving cure as a final goal. Overgrowing data suggest that cancer is a disease closely linked to stem cells (SCs). It is well known that the first identification of cancer stem-like cells in acute myeloid leukaemia was soon followed by similar results in solid malignancies, including colorectal cancer, and the classic model for colon carcinogenesis supports the development of sudden mutations that will lead to the activation or inactivation of certain oncogenes or tumor suppressors. Thus, this process may go on for years before the first symptoms and the only cells able to withstand for many years, avoid apoptosis and have a high regenerative capacity are the progenitor cells found at the lower part of colon crypts. A more profound study of the mechanisms and molecular signalling pathways that control the basic characteristics of SCs, such as asymmetrical division or self-renewal, may help comprehend the basic mechanisms of cancer genesis and progression. This will result in the development of new therapeutic agents that may target chemoresistant cell populations and improve the therapeutic results. In the current review we point out the importance of cancer stem-like cells in colorectal oncology from a pathologist's point of view, stating the obvious correlation between histology, embryology and surgical pathology.     

Discrepancies between estimated and perceived risk of cancer among individuals with hereditary nonpolyposis colorectal cancer

Communicating cancer risk and recommending adequate control programs is central for genetic counseling. Individuals affected by hereditary nonpolyposis colorectal cancer (HNPCC) are at about 80% life-time risk of colorectal cancer and for female carriers 40-60% risk of endometrial cancer and 10-15% risk of ovarian cancer. The perceived risk among mutation carriers may, however, deviate from the risk communicated and has been demonstrated to influence adherence to control programs. We investigated the perceived cancer risk among HNPCC mutation carriers (n = 47) and correlated the findings to individual characteristics. A perceived risk of colorectal cancer above 60% was reported by 22/45 individuals, and only one out of five mutation carriers reported a perceived risk > 80%. Female mutation carriers, individuals below age 50, and individuals who received their oncogenetic counseling within 1 year prior to the study reported higher, albeit not significantly, perceived risks of colorectal cancer. Higher perceived risks were also reported by individuals who had lost a parent to HNPCC-related cancer at early age, whereas individuals with a personal history of cancer did not report a higher perceived risk. Regarding gynecological cancer, 6/18 females reported a perceived risk of 40-60% for endometrial cancer, whereas the remaining women both underestimated and overestimated their risk, and none of the women referred to the risk of ovarian cancer. We conclude that despite educational efforts and an increasing amount of data on the cancer risk in HNPCC, a minority of the mutation carriers report a perceived risk at the same level as that communicated during oncogenetic counseling.     

The value of MUTYH testing in patients with early onset microsatellite stable colorectal cancer referred for hereditary nonpolyposis colon cancer syndrome testing

MUTYH adenomatous polyposis (MAP) can mimic both the familial adenomatous polyposis (FAP) and hereditary nonpolyposis colon cancer (HNPCC) phenotypes. As a result of MAP's phenotypic overlap with FAP, some DNA diagnostic laboratories perform MUTYH testing in conjunction with APC testing in patients with suspected FAP or attenuated FAP (AFAP). In addition to testing FAP/AFAP samples for MUTYH mutations, we were interested whether there would also be value in testing samples referred for HNPCC testing. To determine this, we tested a consecutive series of 229 samples referred for HNPCC testing for the two most common MUTYH mutations in the Caucasian population. To enrich our study population with MAP cases, we only included samples from patients with early onset colorectal cancer (CRC diagnosed <50 years old) in whom HNPCC had been excluded by microsatellite instability testing (microsatellite stable or low microsatellite instability). Four biallelic (2%) and six monoallelic (3%) MUTYH mutation carriers were identified. No clinical factors predicted MUTYH mutation status. Specifically, a family history of vertical transmission of CRC or having few polyps (<15) did not rule out the possibility of biallelic MUTYH mutations. Thus, MUTYH mutation testing may be a reasonable cascade test in early onset CRC found to have proficient DNA mismatch repair, regardless of pattern of family history or number of polyps.     

Predictive testing for hereditary nonpolyposis colorectal cancer: subjective perception regarding colorectal and endometrial cancer, distress, and health-related behavior at one year post-test

This study evaluated illness representations, distress, and health-related behavior one year after disclosure of a predictive genetic test result for hereditary nonpolyposis colorectal cancer (HNPCC) in 36 carriers and 36 noncarriers. Post-test, no significant differences between carriers and noncarriers were found in perceived risk and perceived seriousness of colorectal and endometrial cancer or in perceived control over endometrial cancer. Confidence in the controllability of colorectal cancer by means of medical examinations was higher for carriers than noncarriers post-test. Mean levels of distress (cancer-specific distress, state anxiety, psychoneuroticism) were within normal ranges and none of the participants had an overall pattern (on all scales) of clinically elevated levels of distress. Carriers had significantly higher cancer-related distress one year posttest than noncarriers. In both groups, colorectal cancer-related distress decreased. Noncarriers additionally showed decreased endometrial cancer-related distress and state anxiety. Within the year after testing, none of the noncarriers had a colonoscopy and all carriers where adherent to the recommendations regarding colorectal cancer screening. Although interview data delineated individually different problems specifically related to predictive testing (e.g., worry, difficulties in relation to other relatives, burden of regular follow-up), generally, predictive testing for HNPCC does not seem to induce major psychological problems. Moreover, the presented data are promising regarding the impact on health-related behavior.