Xiaohong R. Yang

Differences in Risk Factors for Breast Cancer Molecular Subtypes in a Population-Based Study

Analysis of gene expression data suggests that breast cancers are divisible into molecular subtypes which have distinct clinical features. This study evaluates whether pathologic features and etiologic associations differ among molecular subtypes. We evaluated 804 women with invasive breast cancers and 2,502 controls participating in a Polish Breast Cancer Study. Immunohistochemical stains for estrogen receptor α, progesterone receptor, human epidermal growth factor receptors (HER2 and HER1), and cytokeratin 5 were used to classify cases into five molecular subtypes: luminal A, luminal B, HER2-expresing, basal-like, and unclassified. Relative risks were estimated using adjusted odds ratios and 95% confidence intervals. We observed that compared with the predominant luminal A tumors (69%), other subtypes were associated with unfavorable clinical features at diagnosis, especially HER2-expressing (8%) and basal-like (12%) tumors. Increasing body mass index significantly reduced the risk of luminal A tumors among premenopausal women (odds ratios, 0.71; 95% confidence intervals, 0.57-0.88 per five-unit increase), whereas it did not reduce risk for basal-like tumors (1.18; 0.86-1.64; Pheterogeneity = 0.003). On the other hand, reduced risk associated with increasing age at menarche was stronger for basal-like (0.78; 0.68-0.89 per 2-year increase) than luminal A tumors (0.90; 0.95-1.08; Pheterogeneity = 0.0009). Although family history increased risk for all subtypes (except for unclassified tumors), the magnitude of the relative risk was highest for basal-like tumors. Results from this study have shown that breast cancer risk factors may vary by molecular subtypes identified in expression studies, suggesting etiologic, in addition to clinical, heterogeneity of breast cancer. (Cancer Epidemiol Biomarkers Prev 2007;16(3):439–43)

T (brachyury) gene duplication confers major susceptibility to familial chordoma

Using high-resolution array-CGH, we identified unique duplications of a region on 6q27 in four multiplex families with at least three cases of chordoma, a cancer of presumed notochordal origin. The duplicated region contains only the T (brachyury) gene, which is important in notochord development and is expressed in most sporadic chordomas. Our findings highlight the value of screening for complex genomic rearrangements in searches for cancer-susceptibility genes.

Rare missense variants in POT1 predispose to familial cutaneous malignant melanoma

Although CDKN2A is the most frequent high-risk melanoma susceptibility gene, the underlying genetic factors for most melanoma-prone families remain unknown. Using whole-exome sequencing, we identified a rare variant that arose as a founder mutation in the telomere shelterin gene POT1 (chromosome 7, g.124493086C>T; p.Ser270Asn) in five unrelated melanoma-prone families from Romagna, Italy. Carriers of this variant had increased telomere lengths and numbers of fragile telomeres, suggesting that this variant perturbs telomere maintenance. Two additional rare POT1 variants were identified in all cases sequenced in two separate Italian families, one variant per family, yielding a frequency for POT1 variants comparable to that for CDKN2A mutations in this population. These variants were not found in public databases or in 2,038 genotyped Italian controls. We also identified two rare recurrent POT1 variants in US and French familial melanoma cases. Our findings suggest that POT1 is a major susceptibility gene for familial melanoma in several populations.

Association of MC1R Variants and Host Phenotypes With Melanoma Risk in CDKN2A Mutation Carriers: A GenoMEL Study

Background Carrying the cyclin-dependent kinase inhibitor 2A (CDKN2A) germline mutations is associated with a high risk for melanoma. Penetrance of CDKN2A mutations is modified by pigmentation characteristics, nevus phenotypes, and some variants of the melanocortin-1 receptor gene (MC1R), which is known to have a role in the pigmentation process. However, investigation of the associations of both MC1R variants and host phenotypes with melanoma risk has been limited. Methods We included 815 CDKN2A mutation carriers (473 affected, and 342 unaffected, with melanoma) from 186 families from 15 centers in Europe, North America, and Australia who participated in the Melanoma Genetics Consortium. In this family-based study, we assessed the associations of the four most frequent MC1R variants (V60L, V92M, R151C, and R160W) and the number of variants (1, ≥2 variants), alone or jointly with the host phenotypes (hair color, propensity to sunburn, and number of nevi), with melanoma risk in CDKN2A mutation carriers. These associations were estimated and tested using generalized estimating equations. All statistical tests were two-sided. Results Carrying any one of the four most frequent MC1R variants (V60L, V92M, R151C, R160W) in CDKN2A mutation carriers was associated with a statistically significantly increased risk for melanoma across all continents (1.24 × 10−6 ≤ P ≤ .0007). A consistent pattern of increase in melanoma risk was also associated with increase in number of MC1R variants. The risk of melanoma associated with at least two MC1R variants was 2.6-fold higher than the risk associated with only one variant (odds ratio = 5.83 [95% confidence interval = 3.60 to 9.46] vs 2.25 [95% confidence interval = 1.44 to 3.52]; Ptrend = 1.86 × 10−8). The joint analysis of MC1R variants and host phenotypes showed statistically significant associations of melanoma risk, together with MC1R variants (.0001 ≤ P ≤ .04), hair color (.006 ≤ P ≤ .06), and number of nevi (6.9 × 10−6 ≤ P ≤ .02). Conclusion Results show that MC1R variants, hair color, and number of nevi were jointly associated with melanoma risk in CDKN2A mutation carriers. This joint association may have important consequences for risk assessments in familial settings.

Melanoma prone families with CDK4 germline mutation: phenotypic profile and associations with MC1R variants

Background CDKN2A and CDK4 are high risk susceptibility genes for cutaneous malignant melanoma. Melanoma families with CDKN2A germline mutations have been extensively characterised, whereas CDK4 families are rare and lack a systematic investigation of their phenotype. Methods All known families with CDK4 germline mutations (n=17) were recruited for the study by contacting the authors of published papers or by requests via the Melanoma Genetics Consortium (GenoMEL). Phenotypic data related to primary melanoma and pigmentation characteristics were collected. The CDK4 exon 2 and the complete coding region of the MC1R gene were sequenced. Results Eleven families carried the CDK4 R24H mutation whereas six families had the R24C mutation. The total number of subjects with verified melanoma was 103, with a median age at first melanoma diagnosis of 39 years. Forty-three (41.7%) subjects had developed multiple primary melanomas (MPM). A CDK4 mutation was found in 89 (including 62 melanoma cases) of 209 tested subjects. CDK4 positive family members (both melanoma cases and unaffected subjects) were more likely to have clinically atypical nevi than CDK4 negative family members (p<0.001). MPM subjects had a higher frequency of MC1R red hair colour variants compared with subjects with one tumour (p=0.010). Conclusion Our study shows that families with CDK4 germline mutations cannot be distinguished phenotypically from CDKN2A melanoma families, which are characterised by early onset of disease, increased occurrence of clinically atypical nevi, and development of MPM. In a clinical setting, the CDK4 gene should therefore always be examined when a melanoma family tests negative for CDKN2A mutation.

DNA Hypermethylation of ESR1 and PGR in Breast Cancer: Pathologic and Epidemiologic Associations

Improved understanding of the etiology of estrogen receptor-α (ERα)–negative and progesterone receptor (PR)–negative breast cancers may permit improved risk prediction. In vitro studies implicate DNA hypermethylation of the ERα and PR promoters in the pathogenesis of ERα-negative and PR-negative tumors, but results are not definitive. We evaluated 200 invasive breast cancers selected from a population-based case-control study. DNA extracted from fixed tumor tissue cores was tested using MethyLight to assess DNA methylation at four CpG islands: ESR1 promoters A and B; PGR promoters A and B; and a CpG shore, ESR1 promoter C. DNA methylation results were compared with levels of ERα and PR, tumor characteristics, and breast cancer risk factors. We observed mild to moderate DNA methylation levels in most tumors for ESR1 promoters A and B and PGR promoter B, and a few tumors showed mild methylation in PGR promoter A. In contrast, ESR1 promoter C showed a wide range of methylation and was weakly correlated with lower expression levels of ERα (β = −0.26; P < 0.0001) and PR (β = −0.25; P < 0.0001). The percentage of tumors with methylated PGR promoters A and B was significantly higher for tumors with low ERα (A, Fishers test P = 0.0001; B, P = 0.033) and PR levels (A, P = 0.0006; B, P = 0.001). Our data suggest that the relationships between DNA methylation of ESR1 and PGR promoters and protein expression are weak and unlikely to represent a predominant mechanism of receptor silencing. In contrast to CpG islands, ESR1 promoter C showed a wider range of methylation levels and inverse associations with ERα and PR expression. (Cancer Epidemiol Biomarkers Prev 2009;18(11):3036–43)

Variation in breast cancer hormone receptor and HER2 levels by etiologic factors: A population-based analysis

Evidence suggests that breast cancer hormone receptor status varies by etiologic factors, but studies have been inconsistent. In a population‐based case–control study in Poland that included 2,386 cases and 2,502 controls, we assessed ER‐α and PR status of tumors based on clinical records according to etiologic exposure data collected via interview. For 842 cancers, we evaluated ER‐α, ER‐β, PR and HER2 levels by semiquantitative microscopic scoring of immunostained tissue microarrays and a quantitative immunofluorescence method, automated quantitative analysis (AQUA™). We related marker levels in tumors to etiologic factors, using standard regression models and novel statistical methods, permitting adjustment for both correlated tumor features and exposures. Results obtained with different assays were generally consistent. Receptor levels varied most significantly with body mass index (BMI), a factor that was inversely related to risk among premenopausal women and directly related to risk among postmenopausal women with larger tumors. After adjustment for correlated markers, exposures and pathologic characteristics, PR and HER2 AQUA levels were inversely related to BMI among premenopausal women (p‐trend = 0.01, both comparisons), whereas among postmenopausal women, PR levels were associated directly with BMI (p‐trend = 0.002). Among postmenopausal women, analyses demonstrated that BMI was related to an interaction of PR and HER2: odds ratio (OR) = 0.86 (95% CI = 0.69–1.07) for low PR and HER2 expression vs. OR = 1.78 (95% CI = 1.25–2.55) for high expression (p‐heterogeneity = 0.001). PR and HER2 levels in breast cancer vary by BMI, suggesting a heterogeneous etiology for tumors related to these markers.

Assessment of Automated Image Analysis of Breast Cancer Tissue Microarrays for Epidemiologic Studies

Background: A major challenge in studies of etiologic heterogeneity in breast cancer has been the limited throughput, accuracy, and reproducibility of measuring tissue markers. Computerized image analysis systems may help address these concerns, but published reports of their use are limited. We assessed agreement between automated and pathologist scores of a diverse set of immunohistochemical assays done on breast cancer tissue microarrays (TMA). Methods: TMAs of 440 breast cancers previously stained for estrogen receptor (ER)-α, progesterone receptor (PR), human epidermal growth factor receptor 2 (HER2), ER-β, and aromatase were independently scored by two pathologists and three automated systems (TMALab II, TMAx, and Ariol). Agreement between automated and pathologist scores of negative/positive was measured using the area under the receiver operating characteristics curve (AUC) and weighted κ statistics for categorical scores. We also investigated the correlation between immunohistochemical scores and mRNA expression levels. Results: Agreement between pathologist and automated negative/positive and categorical scores was excellent for ER-α and PR (AUC range = 0.98-0.99; κ range = 0.86-0.91). Lower levels of agreement were seen for ER-β categorical scores (AUC = 0.99-1.0; κ = 0.80-0.86) and both negative/positive and categorical scores for aromatase (AUC = 0.85-0.96; κ = 0.41-0.67) and HER2 (AUC = 0.94-0.97; κ = 0.53-0.72). For ER-α and PR, there was a strong correlation between mRNA levels and automated (ρ = 0.67-0.74) and pathologist immunohistochemical scores (ρ = 0.67-0.77). HER2 mRNA levels were more strongly correlated with pathologist (ρ = 0.63) than automated immunohistochemical scores (ρ = 0.41-0.49). Conclusions: Automated analysis of immunohistochemical markers is a promising approach for scoring large numbers of breast cancer tissues in epidemiologic investigations. This would facilitate studies of etiologic heterogeneity, which ultimately may allow improved risk prediction and better prevention approaches. Cancer Epidemiol Biomarkers Prev; 19(4); 992–9. ©2010 AACR.

Female Breast Cancer Incidence Among Asian and Western Populations: More Similar Than Expected

BACKGROUND Previous reports suggested that female breast cancer is associated with earlier ages at onset among Asian than Western populations. However, most studies utilized cross-sectional analyses that may be confounded by calendar-period and/or birth cohort effects. We, therefore, considered a longitudinal (forward-looking) approach adjusted for calendar-period changes and conditioned upon birth cohort. METHODS Invasive female breast cancer data (1988-2009) were obtained from cancer registries in China, Hong Kong, South Korea, Taiwan, Singapore, and the United States. Age-period-cohort models were used to extrapolate longitudinal age-specific incidence rates for the 1920, 1944, and 1970 birth cohorts. RESULTS Cross-sectional age-specific incidence rates rose continuously until age 80 years among US white women, but plateaued or decreased after age 50 years among Asian women. In contrast, longitudinal age-specific rates were proportional (similar) among all Asian countries and the United States with incidence rates rising continuously until age 80 years. The extrapolated estimates for the most recent cohorts in some Asian countries actually showed later ages at onset than in the United States. Additionally, over successive birth cohorts, the incidence rate ratios (IRRs) for the longitudinal curves converged (narrowed) between Asian and US white women. CONCLUSIONS Similar longitudinal age-specific incidence rates along with converging IRRs indicate that the age effects for invasive breast cancer are more similar among Asian and Western populations than might be expected from a solely cross-sectional analysis. Indeed, the Asian breast cancer rates in recent generations are even surpassing the historically high rates in the United States, highlighting an urgent need for efficient prevention and treatment strategies among Asian populations.

Telomere Length and the Risk of Cutaneous Malignant Melanoma in Melanoma-Prone Families with and without CDKN2A Mutations

Introduction Recent evidence suggests a link between constitutional telomere length (TL) and cancer risk. Previous studies have suggested that longer telomeres were associated with an increased risk of melanoma and larger size and number of nevi. The goal of this study was to examine whether TL modified the risk of melanoma in melanoma-prone families with and without CDKN2A germline mutations. Materials and Methods We measured TL in blood DNA in 119 cutaneous malignant melanoma (CMM) cases and 208 unaffected individuals. We also genotyped 13 tagging SNPs in TERT. Results We found that longer telomeres were associated with an increased risk of CMM (adjusted OR = 2.81, 95% CI = 1.02–7.72, P = 0.04). The association of longer TL with CMM risk was seen in CDKN2A- cases but not in CDKN2A+ cases. Among CMM cases, the presence of solar injury was associated with shorter telomeres (P = 0.002). One SNP in TERT, rs2735940, was significantly associated with TL (P = 0.002) after Bonferroni correction. Discussion Our findings suggest that TL regulation could be variable by CDKN2A mutation status, sun exposure, and pigmentation phenotype. Therefore, TL measurement alone may not be a good marker for predicting CMM risk.