Tandem BRAF Mutations in Primary Invasive Melanomas

The RAS/RAF/MAPK pathway likely mediates critical cell proliferation and survival signals in melanoma. BRAF mutations have been found in a high percentage of melanoma cell lines and metastases; however, only a few studies with a limited number of specimens have focused on primary melanomas. We examined BRAF exon 15 mutational status in 37 primary invasive melanomas of varying thicknesses, which had undergone a standardized pathology review. BRAF mutational status was determined using direct manual sequencing of PCR products, followed by resequencing separately amplified DNA aliquots to confirm each mutation. BRAF exon 15 mutations were found in 17 of 37 (46%) primary melanomas. Tumor-specific tandem mutations, encoding either V599K, V599R, or V599E, were found in 5 of 17 (29%) melanomas with BRAF exon 15 mutations. Cloning of BRAF double base-pair substitutions confirmed that both base changes were on the same allele and can result in a positive charge at codon 599. BRAF mutations, including tandem mutations, were frequently found in both thin and thick primary melanomas, implying that these mutations can occur early in the progression of melanoma. The finding of tandem mutations in thin melanomas makes it more likely that they arise as a simultaneous rather than sequential event

Risk factors for breast cancer characterized by the estrogen receptor alpha A908G (K303R) mutation

Abstract Introduction Estrogen is important in the development of breast cancer, and its biological effects are mediated primarily through the two estrogen receptors alpha and beta. A point mutation in the estrogen receptor alpha gene, ESR1, referred to as A908G or K303R, was originally identified in breast hyperplasias and was reported to be hypersensitive to estrogen. We recently detected this mutation at a low frequency of 6% in invasive breast tumors of the Carolina Breast Cancer Study (CBCS). Methods In this report, we evaluated risk factors for invasive breast cancer classified according to the presence or absence of the ESR1 A908G mutation in the CBCS, a population-based case-control study of breast cancer among younger and older white and African-American women in North Carolina. Of the 653 breast tumors evaluated, 37 were ESR1 A908G mutation-positive and 616 were mutation-negative. Results ESR1 A908G mutation-positive breast cancer was significantly associated with a first-degree family history of breast cancer (odds ratio [OR] = 2.69, 95% confidence interval [CI] = 1.15 to 6.28), whereas mutation-negative breast cancer was not. Comparison of the two case subgroups supported this finding (OR = 2.65, 95% CI = 1.15 to 6.09). There was also the suggestion that longer duration of oral contraceptive (OC) use (OR = 3.73, 95% CI = 1.16 to 12.03; P trend = 0.02 for use of more than 10 years) and recent use of OCs (OR = 3.63, 95% CI = 0.80 to 16.45; P trend = 0.10 for use within 10 years) were associated with ESR1 A908G mutation-positive breast cancer; however, ORs for comparison of the two case subgroups were not statistically significant. Hormone replacement therapy use was inversely correlated with mutation-negative breast cancer, but the effect on mutation-positive cancer was unclear due to the small number of postmenopausal cases whose tumors carried the mutation. Mutation-negative breast cancer was associated with several reproductive factors, including younger age at menarche (OR = 1.46, 95% CI = 1.09 to 1.94) and greater total estimated years of ovarian function (OR = 1.82, 95% CI = 1.21 to 2.74). Conclusion These preliminary results suggest that OCs may interact with the ESR1 A908G mutant receptor to drive the development of some breast tumors

Racial Variation in Breast Tumor Promoter Methylation in the Carolina Breast Cancer Study

African American (AA) women are diagnosed with more advanced breast cancers and have worse survival than white women, but a comprehensive understanding of the basis for this disparity remains unclear. Analysis of DNA methylation, an epigenetic mechanism that can regulate gene expression, could help to explain racial differences in breast tumor clinical biology and outcomes

Body Mass Index Is Associated with Gene Methylation in Estrogen Receptor-Positive Breast Tumors

Although obesity is associated with breast cancer incidence and prognosis, the underlying mechanisms are poorly understood. Identification of obesity-associated epigenetic changes in breast tissue may advance mechanistic understanding of breast cancer initiation and progression. The goal of this study, therefore, was to investigate associations between obesity and gene methylation in breast tumors

DNA methylation profiles in primary cutaneous melanomas are associated with clinically significant pathologic features

DNA methylation studies have elucidated a methylation signature distinguishing primary melanomas from benign nevi and provided new insights about genes that may be important in melanoma development. However, it is unclear whether methylation differences among primary melanomas are related to tumor pathologic features with known clinical significance. We utilized the Illumina Golden Gate Cancer Panel array to investigate the methylation profiles of 47 primary cutaneous melanomas. Array-wide methylation patterns revealed a positive association of methylation with Breslow thickness and mutated BRAF, a negative association with mitotic rate, and a weak association with ulceration. Hierarchical clustering on CpG sites exhibiting the most variable methylation (n=235) divided the melanoma samples into three clusters, including a highly-methylated cluster that was positively associated with Breslow thickness and an intermediately-methylated cluster associated with Breslow thickness and mitotic rate. Our findings provide support for the existence of methylation-defined subsets in melanomas, with increased methylation associated with Breslow thickness

The estrogen receptor-α A908G (K303R) mutation occurs at a low frequency in invasive breast tumors: results from a population-based study

INTRODUCTION: Evidence suggests that alterations in estrogen signaling pathways, including estrogen receptor-α (ER-α), occur during breast cancer development. A point mutation in ER-α (nucleotide A908G), producing an amino acid change from lysine to arginine at codon 303 (K303R) results in receptor hypersensitivity to estrogen. This mutation was initially reported in one-third of hyperplastic benign breast lesions, although several recent studies failed to detect it in benign or malignant breast tissues. METHODS: We screened 653 microdissected, newly diagnosed invasive breast tumors from patients in the Carolina Breast Cancer Study, a population-based case-control study of breast cancer in African American and white women in North Carolina, for the presence of the ER-α A908G mutation by using single-strand conformational polymorphism (SSCP) analysis and (33)P-cycle sequencing. RESULTS: We detected the ER-α A908G mutation in 37 of 653 (5.7%) breast tumors. The absence of this mutation in germline DNA confirmed it to be somatic. Three tumors exhibited only the mutant G base at nucleotide 908 on sequencing, indicating that the wild-type ER-α allele had been lost. The ER-α A908G mutation was found more frequently in higher-grade breast tumors (odds ratio (OR) 2.83; 95% confidence interval (CI) 1.09 to 7.34, grade II compared with grade I), and in mixed lobular/ductal tumors (OR 2.10; 95% CI 0.86 to 5.12) compared with ductal carcinomas, although the latter finding was not statistically significant. CONCLUSION: This population-based study, the largest so far to screen for the ER-α A908G mutation in breast cancer, confirms the presence of the mutant in invasive breast tumors. The mutation was associated with higher tumor grade and mixed lobular/ductal breast tumor histology

InterMEL: An international biorepository and clinical database to uncover predictors of survival in early-stage melanoma

We are conducting a multicenter study to identify classifiers predictive of disease-specific survival in patients with primary melanomas. Here we delineate the unique aspects, challenges, and best practices for optimizing a study of generally small-sized pigmented tumor samples including primary melanomas of at least 1.05mm from AJTCC TNM stage IIA-IIID patients. We also evaluated tissue-derived predictors of extracted nucleic acids’ quality and success in downstream testing. This ongoing study will target 1,000 melanomas within the international InterMEL consortium.Medicin

Association Between NRAS and BRAF Mutational Status and Melanoma-Specific Survival Among Patients With Higher-Risk Primary Melanoma

NRAS and BRAF mutations in melanoma inform current treatment paradigms but their role in survival from primary melanoma has not been established. Identification of patients at high risk of melanoma-related death based on their primary melanoma characteristics before evidence of recurrence could inform recommendations for patient follow-up and eligibility for adjuvant trials