Genotype-phenotype relationships in U.S. melanoma-prone families with CDKN2A and CDK4 mutations

BACKGROUND: Two genes have been implicated in the development of cutaneous malignant melanoma (CMM). CDK4 (the gene encoding cyclin-dependent kinase 4, an oncogene) has exhibited germline mutations found in only three melanoma-prone families to date. CDKN2A is a tumor suppressor gene that encodes p16 (which inhibits activity of the cyclin D1-CDK4 complex) with germline mutations detected in 10%-25% of melanoma-prone families, some of whom are also prone to pancreatic cancer. METHODS: We compared 104 CMM patients from 17 CDKN2A families and 12 CMM case subjects from two CDK4 families. We used nonparametric statistics to test for differences in median age at first CMM diagnosis, numbers of CMMs, and numbers of nevi. The three recurrent mutations were haplotyped. All P values were two-sided. RESULTS: The median age at CMM diagnosis (P =.70) and the median numbers of CMMs (P =.73) did not differ between CMM case subjects from CDKN2A versus CDK4 families. Assessment of CMM case subjects from CDKN2A families with and without pancreatic cancer revealed no statistically significant differences in median age at diagnosis (P =.80) or in tumor number (P =.24). There was, however, a statistically significant difference in age-adjusted median numbers of nevi (P =.004), and CMM case subjects from CDKN2A families without pancreatic cancer had greater numbers of nevi. Recurrent CDKN2A mutations were a change from valine to aspartic acid at codon 126 (n = 3) and from glycine to tryptophan at codon 101 (n = 3). Six CDKN2A families had pancreatic cancer. Both CDK4 families carried a mutation resulting in an arginine-to-cysteine substitution at codon 24. Analyses of recurrent CDKN2A and CDK4 mutations suggested common haplotypes. CONCLUSIONS: The recurrent CDKN2A mutations were observed in families with and without pancreatic cancer, which suggests that other factors may be involved in the development of pancreatic cancer. Despite hypothetical differences in the mechanisms of action between CDKN2A and CDK4, clinical factors were indistinguishable between CMM case subjects from CDKN2A versus CDK4 families.     

Contribution of germline mutations in the BRCA and PALB2 genes to pancreatic cancer in Italy

Pancreatic adenocarcinoma (PC) is the third most common cancer associated with BRCA mutations. Most notice has been given to BRCA2, while the association between BRCA1 and PC is less widely reported. Recently, PALB2 has been implicated in both PC and breast cancer (BC) susceptibility. We selected 29 Italian PC patients from a case-control study of PC according to their personal and family history of both PC and breast/ovarian cancer (BC/OC) and tested them for presence of germline mutations in BRCA1, BRCA2 and PALB2. We identified no germline mutations or deletions in PALB2, but detected 7 BRCA mutations (4 in BRCA1 and 3 in BRCA2). These findings suggest that PALB2 does not play a major role in PC susceptibility in our population. As we found an almost equal frequency of germline mutations in BRCA1 and BRCA2, germline alterations in either of these genes may explain a subset of Italian families presenting both PC and BC/OC. Moreover, as we began the observation of these families from probands who are affected by PC, we provide here a direct assessment of the role of PALB2 and BRCA mutations in PC susceptibility.     

Association of MC1R variants and risk of melanoma in melanoma-prone families with CDKN2A mutations.

Major risk factors for melanoma include many nevi, especially dysplastic nevi, fair pigmentation, freckling, poor tanning ability, and germ line mutations in the CDKN2A, CDK4, or MC1R genes. We evaluated the relationship between MC1R and melanoma risk in CDKN2A melanoma-prone families with extensive clinical and epidemiologic data. We studied 395 subjects from 16 American CDKN2A families. Major melanoma risk factors were assessed by clinical examination or questionnaire; MC1R was sequenced. Odds ratios were estimated by unconditional and conditional logistic regression models. We examined the distribution of MC1R variants and median ages at melanoma diagnosis in multiple primary melanoma (MPM) and single primary melanoma (SPM) patients. Presence of multiple MC1R variants was significantly associated with melanoma, even after adjustment for major melanoma risk factors. All 40 MPM patients had at least one MC1R variant; 65% of MPM patients versus only 17% of SPM patients had at least two MC1R variants (P < 0.0001). For all 69 melanoma patients combined, as well as the 40 MPM patients, there was a statistically significant decrease in median age at diagnosis as numbers of MC1R variants increased (P = 0.010 and P = 0.008, respectively). In contrast, no significant reduction in age at melanoma diagnosis was observed for SPM patients (P = 0.91). The current study suggests that the presence of multiple MC1R variants is associated with the development of multiple melanoma tumors in patients with CDKN2A mutations. Additional studies are needed to confirm these findings and to explore the mechanisms that may contribute to this relationship.     

PALB2 mutations in familial breast and pancreatic cancer

PALB2 (Partner And Localizer of BRCA2) binds to and co-localizes with BRCA2 in DNA repair. Germline mutations in PALB2 have been identified in approximately 1–2% of familial breast cancer and 3–4% of familial pancreatic cancer cases. The goal of this study was to evaluate the prevalence of PALB2 mutations in women with breast cancer without BRCA1/2 mutations who also had a personal or family history of pancreatic cancer. PALB2 mutation analysis was performed in 94 non-BRCA1/2 breast cancer patients with a personal or family history of pancreatic cancer. Two truncating PALB2 mutations, c.3549C>CA and c.2962C>CT, were identified resulting in a mutation prevalence of 2.1%. The proband found to carry the c.3549C>CA PALB2 mutation had a mother diagnosed with both breast and pancreatic cancer; this relative was subsequently confirmed to carry the identical mutation. The proband with the c.2962C>CT mutation had a father and paternal aunt diagnosed with pancreatic cancer; neither relative was available for testing. Two novel PALB2 missense variants were also found, one of which was deemed potentially deleterious. The prevalence rate of PALB2 mutations in a non-BRCA1/2 breast cancer population specifically selected for a family history of pancreatic cancer does not appear to be significantly increased compared to that observed in other breast cancer populations studied thus far. Further evaluation is needed to determine the prevalence of PALB2 mutations and the clinical utility of such testing in those individuals affected with both breast and pancreatic cancers.     

Novel germline PALB2 truncating mutations in African American breast cancer patients

BACKGROUND:

It has been demonstrated that the partner and localizer of breast cancer 2 (PALB2) acts as a bridging molecule between the breast cancer 1 (BRCA1) and BRCA2 proteins and is responsible for facilitating BRCA2‐mediated DNA repair. Truncating mutations in the PALB2 gene reportedly are enriched in patients with Fanconi anemia and breast cancer in various populations.

METHODS:

The authors evaluated the contribution of PALB2 germline mutations in 279 African American women with breast cancer, including 29 patients with a strong family history, 29 patients with a moderate family history, 75 patients with a weak family history, and 146 patients with nonfamilial or sporadic breast cancer.

RESULTS:

After direct sequencing of all the coding exons, exon/intron boundaries, and 5′ and 3′ untranslated regions of PALB2, 3 novel, monoallelic, truncating mutations (1.08%; 3 in 279 patients) were identified (c.758dupT [exon 4], c.1479delC [exon 4], and c.3048delT [exon 10]) together with 50 sequence variants, 27 of which were novel. None of the truncating mutations were identified in a group of 262 controls from the same population.

CONCLUSIONS:

PALB2 mutations were present in both familial and nonfamilial breast cancers among African Americans. Rare PALB2 mutations accounted for a small but substantial proportion of patients with breast cancer. Cancer 2012. © 2011 American Cancer Society.     

Genetic variants in DNA repair genes and the risk of cutaneous malignant melanoma in melanoma-prone families with/without CDKN2A mutations

Cutaneous malignant melanoma (CMM) is an etiologically heterogeneous disease with genetic, environmental (sun exposure) and host (pigmentation/nevi) factors and their interactions contributing to risk. Genetic variants in DNA repair genes may be particularly important since their altered function in response to sun exposure-related DNA damage maybe related to risk for CMM. However, systematic evaluations of genetic variants in DNA repair genes are limited, particularly in high-risk families. We comprehensively analyzed DNA repair gene polymorphisms and CMM risk in melanoma-prone families with/without CDKN2A mutations. A total of 586 individuals (183 CMM) from 53 families (23 CDKN2A (+), 30 CDKN2A (-)) were genotyped for 2964 tagSNPs in 131 DNA repair genes. Conditional logistic regression, conditioning on families, was used to estimate trend p-values, odds ratios and 95% confidence intervals for the association between CMM and each SNP separately, adjusted for age and sex. p-Values for SNPs in the same gene were combined to yield gene specific p-values. Two genes, POLN and PRKDC, were significantly associated with melanoma after Bonferroni correction for multiple testing (p = 0.0003 and 0.00035, respectively). DCLRE1B showed suggestive association (p = 0.0006). 28 ～ 56% of genotyped SNPs in these genes had single SNP p < 0.05. The most significant SNPs in POLN and PRKDC had similar effects in CDKN2A (+) and CDKN2A (-) families. Our finding suggests that polymorphisms in DNA repair genes, POLN and PRKDC, were associated with increased melanoma risk in melanoma families with and without CDKN2A mutations.     

BRCA2 germline mutations in Japanese breast cancer families

Germline mutations of BRCA2 were examined in 20 Japanese breast cancer families without BRCA1 mutations, including one demonstrating cancer development in a male. Three different mutations, resulting in truncation of the BRCA2 protein, were detected in 3 different families. They were 9474insA (exon 24, termination at codon 3110), C8729A (exon 20, S2834 ter) and 982del4 (exon 9, termination at codon 275). The 982del4 mutation was detected in the family with a case of male breast cancer. Age at onset was young, with a range of 28-43 years, in the 2 female breast cancer families with truncation mutations. One probable missense mutation, A10462G (I3412V), was further detected in 2 families, although cosegregation of this allele with the breast cancer phenotype was not complete. The rate of BRCA2 mutations in Japanese families was suggested to be almost the same as in Western countries, and larger than it is the case for BRCA1. Int. J. Cancer 74:199-204, 1997. © 1997 Wiley-Liss, Inc.     

Gene-covariate interaction between dysplastic nevi and the CDKN2A gene in American melanoma-prone families.

The CDKN2A gene has been implicated in cutaneous malignant melanoma pathogenesis. Although CDKN2A mutations confer substantial risk for melanoma, clinicoepidemiological covariates including dysplastic nevi (DN), total nevi, and solar injury also enhance melanoma risk. To examine the relationship between CDKN2A and these three risk factors, we conducted combined segregation/linkage analysis using the class D regressive logistic model, as implemented in the computer program REGRESS. Genetic and covariate data were collected on 20 American melanoma-prone families, 13 of which had cosegregating CDKN2A mutations. Two types of analyses were conducted. The missing-indicator method used a missing-value indicator, set to 1 for unknown and 0 for known covariate status, and a second variable set to 1 for exposed and 0 for unexposed or unknown. The second method, complete-cases method, coded subjects with missing covariates as unknown for the affection status. The results for both analyses were very similar. Overall, there was a significant improvement in the likelihood when DN, total nevi or both covariates were added to the base model, which included dominant transmission of the CDKN2A gene and a linear increase of risk with the logarithm of age on the logit scale. In contrast, inclusion of solar injury did not significantly improve the likelihood for the base model. Significant evidence for a gene-covariate interaction was detected between DN and CDKN2A when DN was the only covariate in the model (missing-indicator method or complete-cases method) or when both DN and total nevi were in the model (complete-cases method only). Interestingly, in both methods, the odds ratio (OR) for DN was greater in subjects without mutations (OR, 20.1; 95% confidence interval, 4.8-92.8) versus those with CDKN2A mutations (OR, 3.3; 95% confidence interval, 1.1-10.0; complete-cases method). The CDKN2A-DN interaction illustrates the complex etiology of melanoma and needs to be confirmed in a larger sample of families.     

Associations of 9p21 variants with cutaneous malignant melanoma, nevi, and pigmentation phenotypes in melanoma-prone families with and without CDKN2A mutations

Chromosome 9p21 has been implicated in the pathogenesis of cutaneous malignant melanoma (CMM). In addition to CDKN2A, the major known high-risk susceptibility gene for CMM, recent studies suggest that other 9p21 genes may be involved in melanoma/nevi development. To identify 9p21 variants that influence susceptibility to CMM and number of nevi in CMM-prone families with and without CDKN2A mutations, we analyzed 562 individuals (183 CMM) from 53 families (23 CDKN2A+, 30 CDKN2A?) for 233 tagging SNPs in 21 genes at 9p21. Single SNP- and gene-based regression analyses were used to assess the risk of CMM, nevi count, skin complexion, and tanning ability associated with these SNPs and genes. We found that SNP rs7023329 in the MTAP gene was associated with number of nevi (P _trend = 0.003) confirming a recent finding by a genome-wide association study. In addition, three SNPs in the ACO1 gene, rs7855483 (P _trend = 0.002), rs17288067 (P _trend = 0.0009), and rs10813813 (P _trend = 0.005), showed the strongest associations with CMM risk. None of the examined 9p21 SNPs was associated with skin complexion, whereas two SNPs, rs10964862 in IFNW1 (P _trend = 0.003), and rs13290968 in TUSC1 (P _trend = 0.0006), were associated with tanning ability. Gene-based analyses suggested that the ACO1 gene was significantly associated with CMM (P = 0.0004); genes IFNW1 (P = 0.002) and ACO1 (P = 0.0002) were significantly associated with tanning ability. Our findings are consistent with recent proposals that additional 9p21 genes may contribute to CMM susceptibility in CMM-prone families. These genetic variants may, at least partially, exert their effects through nevi and tanning ability.     

Low prevalence of germline PALB2 mutations in Australian triple‐negative breast cancer

Triple‐negative breast cancer (TNBC) is a tumour classification that is defined by oestrogen receptor, progesterone receptor and human epidermal growth factor receptor 2 receptor negativity. TNBCs share a similar gene expression profile to BRCA‐mutated tumours, have been shown to carry a high proportion of BRCA mutations and have a more adverse prognosis compared to other types of breast tumours. PALB2 has been shown to be a moderate‐penetrance breast cancer susceptibility gene and is involved in the same DNA damage repair pathway as BRCA1 and BRCA2; this raises the possibility that germline PALB2 mutations may be involved in the pathogenesis of TNBCs. In our study, we sequenced the coding regions of PALB2 (including intron/exon boundaries) in genomic DNA from 347 patients diagnosed with TNBC to determine the prevalence of deleterious mutations in this population. Two novel truncating mutations (c.758dup and c.2390del) and one previously detected truncating mutation (c.3113+5G>C) were found. In addition, five variants predicted to be protein‐affecting were also identified. Our study shows that the prevalence of PALB2 germline mutations in individuals with TNBC is ～1%, similar to the prevalence of PALB2 germline mutation of 1% in familial non‐BRCA1/2 breast cancer cohorts.     

CDKN2A and CDK4 mutation analysis in Italian melanoma-prone families: functional characterization of a novel CDKN2A germ line mutation

Physical interaction between CDKN2A/p16 and CDK4 proteins regulates the cell cycle progression through the G1 phase and dysfunction of these proteins by gene mutation is implicated in genetic predisposition to melanoma. We analysed 15 Italian melanoma families for germ line mutations in the coding region of the CDKN2A gene and exon 2 of the CDK4 gene. One novel disease-associated mutation (P48T), 3 known pathological mutations (R24P, G101W and N71S) and 2 common polymorphisms (A148T and Nt500 G>C) were identified in the CDKN2A gene. In a family harbouring the R24P mutation, an intronic variant (IVS1, +37 G>C) of uncertain significance was detected in a non-carrier melanoma case. The overall incidence of CDKN2A mutations was 33.3%, but this percentage was higher in families with 3 or more melanoma cases (50%) than in those with only 2 affected relatives (25%). Noteworthy, functional analysis established that the novel mutated protein, while being impaired in cell growth and inhibition assays, retains some in vitro binding to CDK4/6. No variant in the p16-binding region of CDK4 was identified in our families. Our results, obtained in a heterogeneous group of families, support the view that inactivating mutations of CDKN2A contribute to melanoma susceptibility more than activating mutations of CDK4 and that other genetic factors must be responsible for melanoma clustering in a high proportion of families. In addition, they indicate the need for a combination of functional assays to determine the pathogenetic nature of new CDKN2A mutations.     

Germline mutations in BRIP1 and PALB2 in Jewish high cancer risk families

Germline mutations in BRCA1 and BRCA2 account for ~30 % of inherited breast cancer. BRIP1 and PALB2 are likely genes for breast cancer susceptibility, based on their roles in maintaining cellular integrity. Indeed, few pathogenic germline mutations in both genes are reported in ethnically diverse breast cancer families. There is a paucity of data on the putative contribution of both genes to inherited breast cancer in Jewish high risk families. High risk Jewish women, none of whom was a carrier of the predominant Jewish mutations in BRCA1/BRCA2, were screened for BRIP1 germline mutations by combined denaturing gradient gel electrophoresis, high resolution melting and sequencing. Direct sequencing of exons and flanking intronic sequences was used for PALB2 mutational analysis. Overall, 149 women, all of high risk, cancer prone families of Ashkenazi origin, were genotyped for BRIP1 mutations: 127 with breast cancer, 22 with ovarian cancer. No truncating mutations were noted and one novel (p.Ala745Thr) and two previously described missense mutations were detected. For PALB2, 93 women were genotyped (87 with breast cancer) of Ashkenazi (n = 32) and non Ashkenazi Jewish origin. Fifteen sequence variants were detected, of these, none was truncating, four were not previously reported, and two (p.Asp871Gly and p.Leu1119Pro) were seemingly pathogenic based on the PolyPhen2 protein prediction algorithm. These missense mutations were not detected in any of 113 healthy Ashkenazi and 109 Moroccan, cancer free controls. In conclusion, germline mutations in BRIP1 and PALB2 contribute marginally to breast cancer susceptibility in ethnically diverse, Jewish high risk families.     

BRCA1 andBRCA2 germline mutations in Japanese with hereditary breast cancer families

We examined germline mutations inBRCA1 andBRCA2 in 23 Japanese breast cancer families, using PCR-SSCP analysis. The same nonsense mutation (exon 5, Leu63ter) ofBRCA1 was detected in two different families. Three different mutations resulting in a truncatedBRCA2 protein (exon 9, 20, 24) were detected in three different families, including one male case of breast cancer. One base substitution mutation inBRCA2, A10462G, was detected in the other two families. Although the mean age of onset for breast cancer in families with theBRCA1- mutation was 50 years, the age of onset in families with theBRCA2-mutation was from 28 to 43 years. Among the 23 families examined, two families had members with ovarian cancers, three had members with prostate cancers, and one had a pancreatic cancer. However, none of these families was positive for theBRCA1 orBRCA2 mutation. Histopathologically, we observed a prevalence of histological grade 3 inBRCA2-associated familial breast cancers, because of nuclear atypia, structural atypia and mitotic activity. It is suggested thatBRCA2 may play a more important role thanBRCA1 in Japanese familial breast cancers, and these mutations are related to the aggressive nature and highly proliferative activity of the tumors.     

A common founder for the V126D CDKN2A mutation in seven North American melanoma-prone families

One of the most common melanoma-related CDKN2A mutations reported in North America is the V126D mutation. We examined nine markers surrounding CDKN2A in three American and four Canadian families carrying the V126D mutation. All seven families had a haplotype consistent with a common ancestor/founder for this mutation. In addition, the mutation appears to have originated 34-52 generations ago (1-LOD-unit support interval 13-98 generations).