Jane Green

Mutations of a mutS homolog in hereditary nonpolyposis colorectal cancer

Recent studies have shown that a locus responsible for hereditary nonpolyposis colorectal cancer (HNPCC) is on chromosome 2p and that tumors developing in these patients contain alterations in microsatellite sequences (RER+ phenotype). We have used chromosome microdissection to obtain highly polymorphic markers from chromosome 2p16. These and other markers were ordered in a panel of somatic cell hybrids and used to define a 0.8 Mb interval containing the HNPCC locus. Candidate genes were then mapped, and one was found to lie within the 0.8 Mb interval. We identified this candidate by virtue of its homology to mutS mismatch repair genes. cDNA clones were obtained and the sequence used to detect germline mutations, including those producing termination codons, in HNPCC kindreds. Somatic as well as germline mutations of the gene were identified in RER+ tumor cells. This mutS homolog is therefore likely to be responsible for HNPCC.

Comparative genomics identifies a flagellar and basal body proteome that includes the BBS5 human disease gene.

Cilia and flagella are microtubule-based structures nucleated by modified centrioles termed basal bodies. These biochemically complex organelles have more than 250 and 150 polypeptides, respectively. To identify the proteins involved in ciliary and basal body biogenesis and function, we undertook a comparative genomics approach that subtracted the nonflagellated proteome of Arabidopsis from the shared proteome of the ciliated/flagellated organisms Chlamydomonas and human. We identified 688 genes that are present exclusively in organisms with flagella and basal bodies and validated these data through a series of in silico, in vitro, and in vivo studies. We then applied this resource to the study of human ciliation disorders and have identified BBS5, a novel gene for Bardet-Biedl syndrome. We show that this novel protein localizes to basal bodies in mouse and C. elegans, is under the regulatory control of daf-19, and is necessary for the generation of both cilia and flagella.

Genome-wide association scan identifies a colorectal cancer susceptibility locus on chromosome 8q24

Using a multistage genetic association approach comprising 7,480 affected individuals and 7,779 controls, we identified markers in chromosomal region 8q24 associated with colorectal cancer. In stage 1, we genotyped 99,632 SNPs in 1,257 affected individuals and 1,336 controls from Ontario. In stages 2–4, we performed serial replication studies using 4,024 affected individuals and 4,042 controls from Seattle, Newfoundland and Scotland. We identified one locus on chromosome 8q24 and another on 9p24 having combined odds ratios (OR) for stages 1–4 of 1.18 (trend; P = 1.41 × 10−8) and 1.14 (trend; P = 1.32 × 10−5), respectively. Additional analyses in 2,199 affected individuals and 2,401 controls from France and Europe supported the association at the 8q24 locus (OR = 1.16, trend; 95% confidence interval (c.i.): 1.07–1.26; P = 5.05 × 10−4). A summary across all seven studies at the 8q24 locus was highly significant (OR = 1.17, c.i.: 1.12–1.23; P = 3.16 × 10−11). This locus has also been implicated in prostate cancer.

The Cardinal Manifestations of Bardet–Biedl Syndrome, a Form of Laurence–Moon–Biedl Syndrome

To determine the interfamilial and intrafamilial variation in the expression of the Bardet-Biedl syndrome (a form of Laurence-Moon-Biedl syndrome), we looked for the five recognized features of the disorder (retinal dystrophy, obesity, polydactyly, mental retardation, and hypogonadism), plus possible renal manifestations, in some or all of 32 patients with this disorder. All 28 patients examined had severe retinal dystrophy, but only 2 had typical retinitis pigmentosa. Polydactyly was present in 18 of 31 patients, but syndactyly, brachydactyly, or both were present in all. Obesity was present in all but 1 of 25 patients. Only 13 of 32 patients were considered mentally retarded. Scores on verbal subtests of intelligence were usually lower than scores on performance tasks. Seven of eight men had small testes and genitalia, which was not due to hypogonadotropism. All 12 women studied had menstrual irregularities, and 3 had low serum estrogen levels (1 of these had hypogonadotropism, and 2 had primary gonadal failure). The remaining women who were of reproductive age had endocrinologic evidence of reproductive dysfunction. Diabetes mellitus was present in 9 of 20 patients. Renal structural or functional abnormalities were universal (n = 21), and three patients had end-stage renal failure. We conclude that the characteristic features of Bardet-Biedl syndrome are severe retinal dystrophy, dysmorphic extremities, obesity, renal abnormalities, and (in male patients only) hypogenitalism. Mental retardation, polydactyly, and hypogonadism in female patients are not necessarily present.

The Clinical Phenotype of Lynch Syndrome Due to Germ-Line PMS2 Mutations

BACKGROUND & AIMS Although the clinical phenotype of Lynch syndrome (also known as hereditary nonpolyposis colorectal cancer) has been well described, little is known about disease in PMS2 mutation carriers. Now that mutation detection methods can discern mutations in PMS2 from mutations in its pseudogenes, more mutation carriers have been identified. Information about the clinical significance of PMS2 mutations is crucial for appropriate counseling. Here, we report the clinical characteristics of a large series of PMS2 mutation carriers. METHODS We performed PMS2 mutation analysis using long-range polymerase chain reaction and multiplex ligation-dependent probe amplification for 99 probands diagnosed with Lynch syndrome-associated tumors showing isolated loss of PMS2 by immunohistochemistry. Penetrance was calculated using a modified segregation analysis adjusting for ascertainment. RESULTS Germ-line PMS2 mutations were detected in 62% of probands (n = 55 monoallelic; 6 biallelic). Among families with monoallelic PMS2 mutations, 65.5% met revised Bethesda guidelines. Compared with the general population, in mutation carriers, the incidence of colorectal cancer was 5.2-fold higher, and the incidence of endometrial cancer was 7.5-fold higher. In North America, this translates to a cumulative cancer risk to age 70 years of 15%-20% for colorectal cancer, 15% for endometrial cancer, and 25%-32% for any Lynch syndrome-associated cancer. No elevated risk for non-Lynch syndrome-associated cancers was observed. CONCLUSIONS PMS2 mutations contribute significantly to Lynch syndrome, but the penetrance for monoallelic mutation carriers appears to be lower than that for the other mismatch repair genes. Modified counseling and cancer surveillance guidelines for PMS2 mutation carriers are proposed.

Mutations in a member of the Ras superfamily of small GTP-binding proteins causes Bardet-Biedl syndrome

RAB, ADP-ribosylation factors (ARFs) and ARF-like (ARL) proteins belong to the Ras superfamily of small GTP-binding proteins and are essential for various membrane-associated intracellular trafficking processes. None of the ∼50 known members of this family are linked to human disease. Using a bioinformatic screen for ciliary genes in combination with mutational analyses, we identified ARL6 as the gene underlying Bardet-Biedl syndrome type 3, a multisystemic disorder characterized by obesity, blindness, polydactyly, renal abnormalities and cognitive impairment. We uncovered four different homozygous substitutions in ARL6 in four unrelated families affected with Bardet-Biedl syndrome, two of which disrupt a threonine residue important for GTP binding and function of several related small GTP-binding proteins. Analysis of the Caenorhabditis elegans ARL6 homolog indicates that it is specifically expressed in ciliated cells, and that, in addition to the postulated cytoplasmic functions of ARL proteins, it undergoes intraflagellar transport. These findings implicate a small GTP-binding protein in ciliary transport and the pathogenesis of a pleiotropic disorder.

Mutations in MKKS cause obesity, retinal dystrophy and renal malformations associated with Bardet-Biedl syndrome

Bardet-Biedl syndrome (BBS) is an autosomal recessive disorder predominantly characterized by obesity, retinal dystrophy, polydactyly, learning difficulties, hypogenitalism and renal malformations, with secondary features that include diabetes mellitus, endocrinological dysfunction and behavioural abnormalities. Despite an initial expectation of genetic homogeneity due to relative clinical uniformity, five BBS loci have been reported, with evidence for additional loci in the human genome; however, no genes for BBS have yet been identified. We performed a genome screen with BBS families from Newfoundland that were excluded from BBS1–5 and identified linkage with D20S189. Fine-mapping reduced the critical interval to 1.9 cM between D20S851 and D20S189, encompassing a chaperonin-like gene. Mutations in this gene were recently reported to be associated with McKusick-Kaufman syndrome (MKKS; ref. 8). Given both the mapping position and clinical similarities of these two syndromes, we screened MKKS and identified mutations in five Newfoundland and two European-American BBS pedigrees. Most are frameshift alleles that are likely to result in a non-functional protein. Our data suggest that a complete loss of function of the MKKS product, and thus an inability to fold a range of target proteins, is responsible for the clinical manifestations of BBS.

Carcinoma of the cervix and tobacco smoking: Collaborative reanalysis of individual data on 13,541 women with carcinoma of the cervix and 23,017 women without carcinoma of the cervix from 23 epidemiological studies - International collaboration of epidemiological studies of cervical cancer

Tobacco smoking has been classified as a cause of cervical cancer, but the effect of different patterns of smoking on risk is unclear. The International Collaboration of Epidemiological Studies of Cervical Cancer has brought together and combined individual data on 13,541 women with and 23,017 women without cervical carcinoma, from 23 epidemiological studies. Relative risks (RRs) and 95% confidence intervals (CIs) of carcinoma of the cervix in relation to tobacco smoking were calculated with stratification by study, age, sexual partners, age at first intercourse, oral contraceptive use and parity. Current smokers had a significantly increased risk of squamous cell carcinoma of the cervix compared to never smokers (RR = 1.60 (95% CI: 1.48–1.73), p<0.001). There was increased risk for past smokers also, though to a lesser extent (RR = 1.12 (1.01–1.25)), and there was no clear trend with time since stopping smoking (p‐trend = 0.6). There was no association between smoking and adenocarcinoma of the cervix (RR = 0.89 (0.74–1.06) and 0.89 (0.72–1.10) for current and past smokers respectively), and the differences between the RRs for smoking and squamous cell and adenocarcinoma were statistically significant (current smoking p<0.001 and past smoking p = 0.01). In current smokers, the RR of squamous cell carcinoma increased with increasing number of cigarettes smoked per day and also with younger age at starting smoking (p<0.001 for each trend), but not with duration of smoking (p‐trend = 0.3). Eight of the studies had tested women for cervical HPV‐DNA, and in analyses restricted to women who tested positive, there was a significantly increased risk in current compared to never smokers for squamous cell carcinoma (RR = 1.95 (1.43–2.65)), but not for adenocarcinoma (RR = 1.06 (0.14–7.96)). In summary, smokers are at an increased risk of squamous cell but not of adenocarcinoma of the cervix. The risk of squamous cell carcinoma increases in current smokers with the number of cigarettes smoked per day and with younger age at starting smoking.

Mutations in MKKS cause Bardet-Biedl syndrome

Bardet-Biedl syndrome (BBS) is an autosomal recessive disorder with locus heterogeneity. None of the ‘responsible’ genes have previously been identified. Some BBS cases (approximately 10%) remain unassigned to the five previously mapped loci. McKusick-Kaufma syndrome (MKS) includes hydrometrocolpos, postaxial polydactyly and congenital heart disease, and is also inherited in an autosomal recessive manner. We ascertained 34 unrelated probands with classic features of BBS including retinitis pigmentosa (RP), obesity and polydactyly. The probands were from families unsuitable for linkage because of family size. We found MKKS mutations in four typical BBS probands (Table 1). The first is a 13-year-old Hispanic girl with severe RP, PAP, mental retardation and obesity (BMI >40). She was a compound heterozygote for a missense (1042G→A, G52D) and a nonsense (1679T→A, Y264stop) mutation in exon 3. Cloning and sequencing of the separate alleles confirmed that the mutations were present in trans. A second BBS proband (from Newfoundland), born to consanguineous parents, was homozygous for two deletions (1316delC and 1324-1326delGTA) in exon 3, predicting a frameshift. An affected brother was also homozygous for the deletions, whereas an unaffected sibling had two normal copies of MKKS. Both the proband and her affected brother had RP, PAP, mild mental retardation, morbid obesity (BMI >50 and 37, respectively), lobulated kidneys with prominent calyces and diabetes mellitus (diagnosed at ages 33 and 30, respectively). A deceased sister (DNA unavailable) had similar phenotypic features (RP with blindness by age 13, BMI >45, abnormal glucose tolerance test and IQ=64, vaginal atresia and syndactyly of both feet). Both parents and the maternal grandfather were heterozygous for the deletions. Genotyping with markers from the MKKS region confirmed homozygosity at 20p12 in both affected individuals.

Gynecologic cancer as a sentinel cancer for women with hereditary nonpolyposis colorectal cancer syndrome

OBJECTIVE: Women with hereditary nonpolyposis colorectal cancer syndrome have a 40–60% lifetime risk for colon cancer, a 40–60% lifetime risk for endometrial cancer, and a 12% lifetime risk for ovarian cancer. A number of women with hereditary nonpolyposis colorectal cancer syndrome will have more than one cancer in their lifetime. The purpose of this study was to estimate whether women with hereditary nonpolyposis colorectal cancer syndrome who develop 2 primary cancers present with gynecologic or colon cancer as their “sentinel cancer.” METHODS: Women whose families fulfilled Amsterdam criteria for hereditary nonpolyposis colorectal cancer syndrome and who developed 2 primary colorectal/gynecologic cancers in their lifetime were identified from 5 large hereditary nonpolyposis colorectal cancer syndrome registries. Information on age at cancer diagnoses and which cancer (colon cancer or endometrial cancer/ovarian cancer) developed first was obtained. RESULTS: A total of 117 women with dual primary cancers from 223 Amsterdam families were identified. In 16 women, colon cancer and endometrial cancer/ovarian cancer were diagnosed simultaneously. Of the remaining 101 women, 52 (51%) women had an endometrial or ovarian cancer diagnosed first. Forty-nine (49%) women had a colon cancer diagnosed first. For women who developed endometrial cancer/ovarian cancer first, mean age at diagnosis of endometrial cancer/ovarian cancer was 44. For women who developed colon cancer first, the mean age at diagnosis of colon cancer was 40. CONCLUSION: In this large series of women with hereditary nonpolyposis colorectal cancer syndrome who developed 2 primary colorectal/gynecologic cancers, endometrial cancer/ovarian cancer was the “sentinel cancer,” preceding the development of colon cancer, in half of the cases. Therefore, gynecologists and gynecologic oncologists play a pivotal role in the identification of women with hereditary nonpolyposis colorectal cancer syndrome. LEVEL OF EVIDENCE: II-3