Validation of a clinical breast cancer risk assessment tool combining a polygenic score for all ancestries with traditional risk factors.

PURPOSE: We previously described a combined risk score (CRS) that integrates a multiple-ancestry polygenic risk score (MA-PRS) with the Tyrer-Cuzick (TC) model to assess breast cancer (BC) risk. Here, we present a longitudinal validation of CRS in a real-world cohort. METHODS: This study included 130,058 patients referred for hereditary cancer genetic testing and negative for germline pathogenic variants in BC-associated genes. Data were obtained by linking genetic test results to medical claims (median follow-up 12.1 months). CRS calibration was evaluated by the ratio of observed to expected BCs. RESULTS: Three hundred forty BCs were observed over 148,349 patient-years. CRS was well-calibrated and demonstrated superior calibration compared with TC in high-risk deciles. MA-PRS alone had greater discriminatory accuracy than TC, and CRS had approximately 2-fold greater discriminatory accuracy than MA-PRS or TC. Among those classified as high risk by TC, 32.6% were low risk by CRS, and of those classified as low risk by TC, 4.3% were high risk by CRS. In cases where CRS and TC classifications disagreed, CRS was more accurate in predicting incident BC. CONCLUSION: CRS was well-calibrated and significantly improved BC risk stratification. Short-term follow-up suggests that clinical implementation of CRS should improve outcomes for patients of all ancestries through personalized risk-based screening and prevention.

The association between age at breast cancer diagnosis and prevalence of pathogenic variants.

PURPOSE: Young age at breast cancer (BC) diagnosis and family history of BC are strongly associated with high prevalence of pathogenic variants (PVs) in BRCA1 and BRCA2 genes. There is limited evidence for such associations with moderate/high penetrance BC-risk genes such as ATM, CHEK2, and PALB2. METHODS: We analyzed multi-gene panel testing results (09/2013-12/2019) for women unaffected by any cancer (N = 371,594) and those affected with BC (N = 130,151) ascertained for suspicion of hereditary breast and/or ovarian cancer. Multivariable logistic regression was used to test association between PV status and age at BC diagnosis (≤ 45 vs. > 45 years) or family history of BC after controlling for personal/family non-BC histories and self-reported ancestry. RESULTS: An association between young age (≤ 45 years) at diagnosis and presence of PVs was strong for BRCA1 (OR 3.95, 95% CI 3.64-4.29) and moderate for BRCA2 (OR 1.98, 95% CI 1.84-2.14). Modest associations were observed between PVs and young age at diagnosis for ATM (OR 1.22, 95% CI 1.08-1.37) and CHEK2 (OR 1.34, 95% CI 1.21-1.47) genes, but not for PALB2 (OR 1.12, 95% CI 0.98-1.27). For women with BC, earliest age of familial BC diagnosis followed a similar pattern. For unaffected women, earliest age of family cancer diagnosis was significantly associated with PV status only for BRCA1 (OR 2.34, 95% CI 2.13-2.56) and BRCA2 (OR 1.25, 95% CI 1.16-1.35). CONCLUSIONS: Young age at BC diagnosis is not a strong risk factor for carrying PVs in BC-associated genes ATM, CHEK2, or PALB2.

Cancer Risk Associated With PTEN Pathogenic Variants Identified Using Multigene Hereditary Cancer Panel Testing.

PURPOSE: PTEN-associated clinical syndromes such as Cowden syndrome (CS) increase cancer risk and have historically been diagnosed based upon phenotypic criteria. Because not all patients clinically diagnosed with CS have PTEN pathogenic variants (PVs), and not all patients with PTEN PVs have been clinically diagnosed with CS, the cancer risk conferred by PTEN PVs calculated from cohorts of patients with clinical diagnoses of CS/CS-like phenotypes may be inaccurate. METHODS: We assessed a consecutive cohort of 727,091 individuals tested clinically for hereditary cancer risk, with a multigene panel between September 2013 and February 2022. Multivariable logistic regression models accounting for personal and family cancer history, age, sex, and ancestry were used to quantify disease risks associated with PTEN PVs. RESULTS: PTEN PVs were detected in 0.027% (193/727,091) of the study population, and were associated with a high risk of female breast cancer (odds ratio [OR], 7.88; 95% CI, 5.57 to 11.16; P = 2.3 × 10-31), endometrial cancer (OR, 13.51; 95% CI, 8.77 to 20.83; P = 4.2 × 10-32), thyroid cancer (OR, 4.88; 95% CI, 2.64 to 9.01; P = 4.0 × 10-7), and colon polyposis (OR, 31.60; CI, 15.60 to 64.02; P = 9.0 × 10-22). We observed modest evidence suggesting that PTEN PVs may be associated with ovarian cancer risk (OR, 3.77; 95% CI, 1.71 to 8.32; P = 9.9 × 10-4). Among patients with similar personal/family history and ancestry, every 5-year increase in age of diagnosis decreased the likelihood of detecting a PTEN PV by roughly 60%. CONCLUSION: We demonstrate that PTEN PVs are associated with significantly increased risk for a range of cancers. Together with the observation that PTEN PV carriers had earlier disease onset relative to otherwise comparable noncarriers, our results may guide screening protocols, inform risk-management strategies, and warrant enhanced surveillance approaches that improve clinical outcomes for PTEN PV carriers, regardless of their clinical presentation.

Development and Validation of a Breast Cancer Polygenic Risk Score on the Basis of Genetic Ancestry Composition.

PURPOSE: Polygenic risk scores (PRSs) for breast cancer (BC) risk stratification have been developed primarily in women of European ancestry. Their application to women of non-European ancestry has lagged because of the lack of a formal approach to incorporate genetic ancestry and ancestry-dependent variant frequencies and effect sizes. Here, we propose a multiple-ancestry PRS (MA-PRS) that addresses these issues and may be useful in the development of equitable PRSs across other cancers and common diseases. MATERIALS AND METHODS: Women referred for hereditary cancer testing were divided into consecutive cohorts for development (n = 189,230) and for independent validation (n = 89,126). Individual genetic composition as fractions of three reference ancestries (African, East Asian, and European) was determined from ancestry-informative single-nucleotide polymorphisms. The MA-PRS is a combination of three ancestry-specific PRSs on the basis of genetic ancestral composition. Stratification of risk was evaluated by multivariable logistic regression models controlling for family cancer history. Goodness-of-fit analysis compared expected with observed relative risks by quantiles of the MA-PRS distribution. RESULTS: In independent validation, the MA-PRS was significantly associated with BC risk in the full cohort (odds ratio, 1.43; 95% CI, 1.40 to 1.46; P = 8.6 × 10-308) and within each major ancestry. The top decile of the MA-PRS consistently identified patients with two-fold increased risk of developing BC. Goodness-of-fit tests showed that the MA-PRS was well calibrated and predicted BC risk accurately in the tails of the distribution for both European and non-European women. CONCLUSION: The MA-PRS uses genetic ancestral composition to expand the utility of polygenic risk prediction to non-European women. Inclusion of genetic ancestry in polygenic risk prediction presents an opportunity for more personalized treatment decisions for women of varying and mixed ancestries.

Clinical Validation of EndoPredict in Pre-Menopausal Women with ER-Positive, HER2-Negative Primary Breast Cancer.

PURPOSE: The EndoPredict prognostic assay is validated to predict distant recurrence and response to chemotherapy primarily in post-menopausal women with estrogen receptor-positive (ER+), HER2- breast cancer. This study evaluated the performance of EndoPredict in pre-menopausal women. EXPERIMENTAL DESIGN: Tumor samples from 385 pre-menopausal women with ER+, HER2- primary breast cancer (pT1-3, pN0-1) who did not receive chemotherapy in addition to endocrine therapy were tested with EndoPredict to produce a 12-gene EP molecular score and an integrated EPclin score that includes pathologic tumor size and nodal status. Associations of molecular and EPclin scores with 10-year distant recurrence-free survival (DRFS) were evaluated by Cox proportional hazards models and Kaplan-Meier analysis. RESULTS: After a median follow-up of 9.7 years, both the EP molecular score and the molecular-clinicopathologic EPclin score were associated with increased risk of distant recurrence [HR, 1.33; 95% confidence interval (CI), 1.18-1.50; P = 7.2 × 10-6; HR, 3.58; 95% CI, 2.26-5.66; P = 9.8 × 10-8, respectively]. Both scores remained significant after adjusting for clinical factors in multivariate analysis. Patients with low-risk EPclin scores (64.7%) had significantly improved DRFS compared with high-risk patients (HR, 4.61; 95% CI, 1.40-15.17; P = 4.2 × 10-3). At 10 years, patients with low-risk and high-risk EPclin scores had a DRFS of 97% (95% CI, 93%-99%) and 76% (95% CI, 67%-82%), respectively. CONCLUSIONS: The EPclin score is strongly associated with DRFS in pre-menopausal women who received adjuvant endocrine therapy alone. On the basis of these data, pre-menopausal women with EPclin low-risk breast cancer may be treated with endocrine therapy only and safely forgo adjuvant chemotherapy.

Performance of the IBIS/Tyrer-Cuzick model of breast cancer risk by race and ethnicity in the Women's Health Initiative.

BACKGROUND: The IBIS/Tyrer-Cuzick model is used clinically to guide breast cancer screening and prevention, but was developed primarily in non-Hispanic White women. Little is known about its long-term performance in a racially/ethnically diverse population. METHODS: The Women's Health Initiative study enrolled postmenopausal women from 1993-1998. Women were included who were aged <80 years at enrollment with no prior breast cancer or mastectomy and with data required for IBIS/Tyrer-Cuzick calculation (weight; height; ages at menarche, first birth, and menopause; menopausal hormone therapy use; and family history of breast or ovarian cancer). Calibration was assessed by the ratio of observed breast cancer cases to the number expected by the IBIS/Tyrer-Cuzick model (O/E; calculated as the sum of cumulative hazards). Differential discrimination was tested for by self-reported race/ethnicity (non-Hispanic White, non-Hispanic Black, Hispanic, Asian or Pacific Islander, and American Indian or Alaskan Native) using Cox regression. Exploratory analyses, including simulation of a protective single-nucleotide polymorphism (SNP), rs140068132 at 6q25, were performed. RESULTS: During follow-up (median 18.9 years, maximum 23.4 years), 6783 breast cancer cases occurred among 90,967 women. IBIS/Tyrer-Cuzick was well calibrated overall (O/E ratio = 0.95; 95% CI, 0.93-0.97) and in most racial/ethnic groups, but overestimated risk for Hispanic women (O/E ratio = 0.75; 95% CI, 0.62-0.90). Discrimination did not differ by race/ethnicity. Exploratory simulation of the protective SNP suggested improved IBIS/Tyrer-Cuzick calibration for Hispanic women (O/E ratio = 0.80; 95% CI, 0.66-0.96). CONCLUSIONS: The IBIS/Tyrer-Cuzick model is well calibrated for several racial/ethnic groups over 2 decades of follow-up. Studies that incorporate genetic and other risk factors, particularly among Hispanic women, are essential to improve breast cancer-risk prediction.

Comprehensive Breast Cancer Risk Assessment for CHEK2 and ATM Pathogenic Variant Carriers Incorporating a Polygenic Risk Score and the Tyrer-Cuzick Model.

PURPOSE: Breast cancer risks for CHEK2 and ATM pathogenic variant (PV) carriers are modified by an 86-single nucleotide polymorphism polygenic risk score (PRS) and individual clinical factors. Here, we describe comprehensive risk prediction models for women of European ancestry combining PV status, PRS, and individual clinical variables. MATERIALS AND METHODS: This study included deidentified clinical records from 358,095 women of European ancestry who received testing with a multigene panel (September 2013 to November 2019). Model development included CHEK2 PV carriers (n = 4,286), ATM PV carriers (n = 2,666), and women negative for other breast cancer risk gene PVs (n = 351,143). Odds ratios (ORs) were calculated using multivariable logistic regression with adjustment for familial cancer history. Risk estimates incorporating PV status, PRS, and Tyrer-Cuzick v7.02 were calculated using a Fixed-Stratified method that accounts for correlations between risk factors. Stratification of PV carriers into risk categories on the basis of remaining lifetime risk (RLR) was assessed in independent cohorts of PV carriers. RESULTS: ORs for association of PV status with breast cancer were 2.01 (95% CI, 1.88 to 2.16) and 1.83 (95% CI, 1.68 to 2.00) for CHEK2 and ATM PV carriers, respectively. ORs for PRS per one standard deviation were 1.51 (95% CI, 1.37 to 1.66) and 1.45 (95% CI, 1.30 to 1.64) in CHEK2 and ATM PV carriers, respectively. Using the combined model (PRS plus Tyrer-Cuzick plus PV status), RLR was low (≤ 20%) for 24.2% of CHEK2 PV carriers, medium (20%-50%) for 63.8%, and high (> 50%) for 12.0%. Among ATM PV carriers, RLR was low for 31.5% of patients, medium for 58.5%, and high for 9.7%. CONCLUSION: In CHEK2 and ATM PV carriers, risk assessment including PRS, Tyrer-Cuzick, and PV status has the potential for more precise direction of screening and prevention strategies.

Integrating Clinical and Polygenic Factors to Predict Breast Cancer Risk in Women Undergoing Genetic Testing.

PURPOSE: Screening and prevention decisions for women at increased risk of developing breast cancer depend on genetic and clinical factors to estimate risk and select appropriate interventions. Integration of polygenic risk into clinical breast cancer risk estimators can improve discrimination. However, correlated genetic effects must be incorporated carefully to avoid overestimation of risk. MATERIALS AND METHODS: A novel Fixed-Stratified method was developed that accounts for confounding when adding a new factor to an established risk model. A combined risk score (CRS) of an 86-single-nucleotide polymorphism polygenic risk score and the Tyrer-Cuzick v7.02 clinical risk estimator was generated with attenuation for confounding by family history. Calibration and discriminatory accuracy of the CRS were evaluated in two independent validation cohorts of women of European ancestry (N = 1,615 and N = 518). Discrimination for remaining lifetime risk was examined by age-adjusted logistic regression. Risk stratification with a 20% risk threshold was compared between CRS and Tyrer-Cuzick in an independent clinical cohort (N = 32,576). RESULTS: Simulation studies confirmed that the Fixed-Stratified method produced accurate risk estimation across patients with different family history. In both validation studies, CRS and Tyrer-Cuzick were significantly associated with breast cancer. In an analysis with both CRS and Tyrer-Cuzick as predictors of breast cancer, CRS added significant discrimination independent of that captured by Tyrer-Cuzick (P < 10-11 in validation 1; P < 10-7 in validation 2). In an independent cohort, 18% of women shifted breast cancer risk categories from their Tyrer-Cuzick-based risk compared with risk estimates by CRS. CONCLUSION: Integrating clinical and polygenic factors into a risk model offers more effective risk stratification and supports a personalized genomic approach to breast cancer screening and prevention.

Multigene assessment of genetic risk for women for two or more breast cancers.

PURPOSE: The prevalence, penetrance, and spectrum of pathogenic variants that predispose women to two or more breast cancers is largely unknown. METHODS: We queried clinical and genetic data from women with one or more breast cancer diagnosis who received multigene panel testing between 2013 and 2018. Clinical data were obtained from provider-completed test request forms. For each gene on the panel, a multivariable logistic regression model was constructed to test for association with risk of multiple breast cancer diagnoses. Models accounted for age of diagnosis, personal and family cancer history, and ancestry. Results are reported as odds ratios (ORs) with 95% confidence intervals (CIs). RESULTS: This study included 98,979 patients: 88,759 (89.7%) with a single breast cancer and 10,220 (10.3%) with ≥ 2 breast cancers. Of women with two or more breast cancers, 13.2% had a pathogenic variant in a cancer predisposition gene compared to 9.4% with a single breast cancer. BRCA1, BRCA2, CDH1, CHEK2, MSH6, PALB2, PTEN, and TP53 were significantly associated with two or more breast cancers, with ORs ranging from 1.35 for CHEK2 to 3.80 for PTEN. Overall, pathogenic variants in all breast cancer risk genes combined were associated with both metachronous (OR 1.65, 95% CI 1.53-1.79, p = 7.2 × 10-33) and synchronous (OR 1.33, 95% CI 1.19-1.50, p = 2.4 × 10-6) breast cancers. CONCLUSIONS: This study demonstrated that several high and moderate penetrance breast cancer susceptibility genes are associated with ≥ 2 breast cancers, affirming the association of two or more breast cancers with diverse genetic etiologies.

Germline Pathogenic Variants in the Ataxia Telangiectasia Mutated (ATM) Gene are Associated with High and Moderate Risks for Multiple Cancers.

Pathogenic variants (PVs) in ATM are relatively common, but the scope and magnitude of risk remains uncertain. This study aimed to estimate ATM PV cancer risks independent of family cancer history. This analysis included patients referred for hereditary cancer testing with a multi-gene panel (N = 627,742). Cancer risks for ATM PV carriers (N = 4,607) were adjusted for family history using multivariable logistic regression and reported as ORs with 95% confidence intervals (CIs). Subanalyses of the c.7271T>G missense PV were conducted. Moderate-to-high risks for pancreatic (OR, 4.21; 95% CI, 3.24-5.47), prostate (OR, 2.58; 95% CI, 1.93-3.44), gastric (OR, 2.97; 95% CI, 1.66-5.31), and invasive ductal breast (OR, 2.03; 95% CI, 1.89-2.19) cancers were estimated for ATM PV carriers. Notably, c.7271T>G was associated with higher invasive ductal breast cancer risk (OR, 3.76; 95% CI, 2.76-5.12) than other missense and truncating ATM PVs. Low-to-moderate risks were seen for ductal carcinoma in situ (OR, 1.80; 95% CI, 1.61-2.02), male breast cancer (OR, 1.72; 95% CI, 1.08-2.75), ovarian cancer (OR, 1.57; 95% CI, 1.35-1.83), colorectal cancer (OR, 1.49; 95% CI, 1.24-1.79), and melanoma (OR, 1.46; 95% CI, 1.18-1.81). ATM PVs are associated with multiple cancer risks and, while professional society guidelines support that carriers are eligible for increased breast and pancreatic cancer screening, increased screening for prostate and gastric cancer may also be warranted. c.7271T>G is associated with high risk for breast cancer, with a 3- to 4-fold risk increase that supports consideration of strategies for prevention and/or early detection. PREVENTION RELEVANCE: This study estimated risks for multiple cancers associated with ATM pathogenic variants independent of family history. These results indicate that some common variants may be associated with higher breast cancer risks than previously appreciated and increased screening for prostate and gastric cancer may be warranted for carriers of ATM pathogenic variants.

Development and Validation of a Clinical Polygenic Risk Score to Predict Breast Cancer Risk.

PURPOSE: Women with a family history of breast cancer are frequently referred for hereditary cancer genetic testing, yet < 10% are found to have pathogenic variants in known breast cancer susceptibility genes. Large-scale genotyping studies have identified common variants (primarily single-nucleotide polymorphisms [SNPs]) with individually modest breast cancer risk that, in aggregate, account for considerable breast cancer susceptibility. Here, we describe the development and empirical validation of an SNP-based polygenic breast cancer risk score. METHODS: A panel of 94 SNPs was examined for association with breast cancer in women of European ancestry undergoing hereditary cancer genetic testing and negative for pathogenic variants in breast cancer susceptibility genes. Candidate polygenic risk scores (PRSs) as predictors of personal breast cancer history were developed through multivariable logistic regression models adjusted for age, cancer history, and ancestry. An optimized PRS was validated in 2 independent cohorts (n = 13,174; n = 141,160). RESULTS: Within the training cohort (n = 24,259), 4,291 women (18%) had a personal history of breast cancer and 8,725 women (36%) reported breast cancer in a first-degree relative. The optimized PRS included 86 variants and was highly predictive of breast cancer status in both validation cohorts (P = 6.4 × 10-66; P < 10-325). The odds ratio (OR) per unit standard deviation was consistent between validations (OR, 1.45 [95% CI, 1.39 to 1.52]; OR 1.47 [95% CI, 1.45 to 1.49]). In a direct comparison, the 86-SNP PRS outperformed a previously described PRS of 77 SNPs. CONCLUSION: The validation and implementation of a PRS for women without pathogenic variants in known breast cancer susceptibility genes offers potential for risk stratification to guide surveillance recommendations.

Impact of a Cancer Gene Variant Reclassification Program Over a 20-Year Period.

PURPOSE: Hereditary cancer genetic testing can inform personalized medical management for individuals at increased cancer risk. However, many variants in cancer predisposition genes are individually rare, and traditional tools may be insufficient to evaluate pathogenicity. This analysis presents data on variant classification and reclassification over a 20-year period. PATIENTS AND METHODS: This is a retrospective analysis of > 1.9 million individuals who received hereditary cancer genetic testing from a single clinical laboratory (March 1997 to December 2017). Variant classification included review of evidence from traditional tools (eg, population frequency databases, literature) and laboratory-developed tools (eg, novel statistical methods, in-house RNA analysis) by a multidisciplinary expert committee. Variants may have been reclassified more than once and with more than one line of evidence. RESULTS: In this time period, 62,842 unique variants were observed across 25 cancer predisposition genes, and 2,976 variants were reclassified. Overall, 82.1% of reclassification events were downgrades (eg, variant of uncertain significance [VUS] to benign), and 17.9% were upgrades (eg, VUS to pathogenic). Among reclassified variants, 82.8% were initially classified as VUS, and 47.5% were identified in ≤ 20 individuals (allele frequency ≤ 0.001%). Laboratory-developed tools were used in 72.3% of variant reclassification events, which affected > 600,000 individuals. More than 1.3 million patients were identified as carrying a variant that was reclassified within this 20-year time period. CONCLUSION: The variant classification program used by the laboratory evaluated here enabled the reclassification of variants that were individually rare. Laboratory-developed tools were a key component of this program and were used in the majority of reclassifications. This demonstrates the importance of using robust and novel tools to reclassify rare variants to appropriately inform personalized medical management.

Association of a Polygenic Risk Score With Breast Cancer Among Women Carriers of High- and Moderate-Risk Breast Cancer Genes.

Importance: To date, few studies have examined the extent to which polygenic single-nucleotide variation (SNV) (formerly single-nucleotide polymorphism) scores modify risk for carriers of pathogenic variants (PVs) in breast cancer susceptibility genes. In previous reports, polygenic risk modification was reduced for BRCA1 and BRCA2 PV carriers compared with noncarriers, but limited information is available for carriers of CHEK2, ATM, or PALB2 PVs. Objective: To examine an 86-SNV polygenic risk score (PRS) for BRCA1, BRCA2, CHEK2, ATM, and PALB2 PV carriers. Design, Setting, and Participants: A retrospective case-control study using data on 150 962 women tested with a multigene hereditary cancer panel between July 19, 2016, and January 11, 2019, was conducted in a commercial testing laboratory. Participants included women of European ancestry between the ages of 18 and 84 years. Main Outcomes and Measures: Multivariable logistic regression was used to examine the association of the 86-SNV score with invasive breast cancer after adjusting for age, ancestry, and personal and/or family cancer history. Effect sizes, expressed as standardized odds ratios (ORs) with 95% CIs, were assessed for carriers of PVs in each gene as well as for noncarriers. Results: The median age at hereditary cancer testing of the population was 48 years (range, 18-84 years); there were 141 160 noncarriers in addition to carriers of BRCA1 (n = 2249), BRCA2 (n = 2638), CHEK2 (n = 2564), ATM (n = 1445), and PALB2 (n = 906) PVs included in the analysis. The 86-SNV score was associated with breast cancer risk in each of the carrier populations (P < 1 × 10-4). Stratification was more pronounced for noncarriers (OR, 1.47; 95% CI, 1.45-1.49) and CHEK2 PV carriers (OR, 1.49; 95% CI, 1.36-1.64) than for carriers of BRCA1 (OR, 1.20; 95% CI, 1.10-1.32) or BRCA2 (OR, 1.23; 95% CI, 1.12-1.34) PVs. Odds ratios for ATM (OR, 1.37; 95% CI, 1.21-1.55) and PALB2 (OR, 1.34; 95% CI, 1.16-1.55) PV carrier populations were intermediate between those for BRCA1/2 and CHEK2 noncarriers. Conclusions and Relevance: In this study, the 86-SNV score was associated with modified risk for carriers of BRCA1, BRCA2, CHEK2, ATM, and PALB2 PVs. This finding supports previous reports of reduced PRS stratification for BRCA1 and BRCA2 PV carriers compared with noncarriers. Modification of risk in CHEK2 carriers associated with the 86-SNV score appeared to be similar to that observed in women without a PV. Larger studies are needed to provide more refined estimates of polygenic modification of risk for women with PVs in other moderate-penetrance genes.

Biomarker Assessment of HR Deficiency, Tumor BRCA1/2 Mutations, and CCNE1 Copy Number in Ovarian Cancer: Associations with Clinical Outcome Following Platinum Monotherapy.

The current study evaluated three biomarkers [homologous recombination deficiency (HRD), tumor BRCA1/2 (tBRCA) mutations, and CCNE1 copy-number variation (CNV)] in ovarian tumors from patients enrolled on the SCOTROC4 clinical trial for associations with outcome following carboplatin monotherapy. Ovarian tumors (n = 250), with high-grade serous (HGSOC) subgroup analysis (n = 179) were classified as HRD positive (HRD score ≥42 or tBRCA mutation) and as CCNE1 amplification positive (CCNE1 CNV score >2.4). Seventy-four (30%) tumors were HRD positive, including 34 (14%) with tBRCA mutations. Forty-seven (19%) were CCNE1 amplification positive, all of which were tBRCA wild-type. HRD and tBRCA, but not CCNE1 amplification, were significantly associated with CA125 complete response in the entire cohort (HRD, P = 0.00015; tBRCA P = 0.0096), and the HGSOC subgroup (HRD, P = 0.0016; tBRCA P = 0.032). HRD and lack of CCNE1 amplification were associated with improved progression-free survival (PFS) and overall survival (OS) in the full cohort and HGSOC subgroup (HRD, P = 0.00021; CCNE1 status P = 0.038). HRD remained significant for OS and PFS after adjusting for clinical factors, while CCNE1 status only remained significant for PFS. Patients with HRD-positive tumors had greater PFS and OS benefit from platinum dose intensification than HRD-negative tumors (P = 0.049 and P = 0.035, respectively). An alternative exploratory HRD score threshold (≥33 or tBRCA mutation) was also significantly associated with both PFS and OS in the HGSOC subset.Implications: HRD, tumor BRCA1/2 mutations, and absence of CCNE1 amplification are associated with improved survival of ovarian cancer patients treated with platinum monotherapy and HRD-positive patients may benefit from platinum dose intensification. Mol Cancer Res; 16(7); 1103-11. ©2018 AACR.

Homologous recombination deficiency (HRD) status predicts response to standard neoadjuvant chemotherapy in patients with triple-negative or BRCA1/2 mutation-associated breast cancer.

PURPOSE: Defects in the homologous recombination (HR) DNA repair pathway sensitize tumors to therapeutics that target this pathway. A significant proportion of triple-negative breast cancers (TNBC) carry HR defects. The HRD assay is highly associated with sensitivity to neoadjuvant platinum-based chemotherapy in TNBC. Standard chemotherapy consists of some combination of an anthracycline, cyclophosphamide, and taxane. This study assesses the association of HR deficiency status with response to standard neoadjuvant chemotherapy in TNBC or BRCA1/2 mutation-associated breast cancer. METHODS: Tumor samples were retrospectively obtained from 45 TNBC patients and 2 BRCA1/2 mutant, hormone receptor-positive/HER2-negative breast cancer patients who received anthracycline- and/or taxane-based neoadjuvant chemotherapy at Stanford University or Cedars-Sinai Medical Centers. The HRD score and tumor BRCA1/2 mutation status were determined from baseline tumor biopsies. HR deficient tumors were those with a HRD score of ≥ 42 or a tumor BRCA1/2 mutation. Response was categorized by the residual cancer burden (RCB) index. RESULTS: HR deficient patients were more likely to achieve a pathologic complete response (pCR) compared with non-deficient patients (OR 13.06, CI 1.52-11.241, p = 0.0028). Among BRCA1/2 mutation wild-type patients, HR deficient patients were more likely to achieve a pCR (OR 16, 95% CI 1.65-160.41, p = 0.0041) compared with HR non-deficient patients. Further, HRD scores were highly concordant pre- and post-therapy (Spearman correlation > 99%). CONCLUSIONS: HR deficiency status is significantly associated with response to standard neoadjuvant chemotherapy in TNBC. This observation is consistent with the mechanisms of action of doxorubicin and cyclophosphamide as DNA damaging agents.

Breast and Ovarian Cancer Penetrance Estimates Derived From Germline Multiple-Gene Sequencing Results in Women.

PURPOSE: Multiple-gene, next-generation sequencing panels are increasingly used to assess hereditary cancer risks of patients with diverse personal and family cancer histories. The magnitude of breast and ovarian cancer risk associated with many clinically tested genes, and independent of family cancer history, remains to be quantified. METHODS: We queried a commercial laboratory database of 95,561 women tested clinically for hereditary cancer risk with a 25-gene (APC, ATM, BARD1, BMPR1A, BRCA1, BRCA2, BRIP1, CDH1, CDK4, CHEK2, MLH2, MSH2, MSH6, MUTYH, NBN, P14ARF, P16, PALB2, PMS2, PTEN, RAD51C, RAD51D, SMAD4, STK11, and TP53) next-generation sequencing panel. Multivariable logistic regression models accounting for family history were used to examine the association between pathogenic mutations and breast or ovarian cancer. As a confirmatory approach, a matched case-control analysis was conducted, defining cases as patients with breast or ovarian cancer and controls as women without cancer. RESULTS: One or more pathogenic mutations were detected in 6,775 (7%) of 95,561 women. Eight genes (ATM, BARD1, BRCA1, BRCA2, CHEK2, PALB2, PTEN, and TP53) were associated with breast cancer, with odds ratios (ORs) ranging from two-fold (ATM: OR, 1.74; 95% CI, 1.46 to 2.07) to six-fold (BRCA1: OR, 5.91; 95% CI, 5.25 to 6.67). Eleven genes (ATM, BRCA1, BRCA2, BRIP1, MLH1, MSH2, MSH6, NBN, STK11, RAD51C, and RAD51D) were associated with ovarian cancer, with OR ranging from two-fold (ATM: OR, 1.69; 95% CI, 1.19 to 2.40) to 40-fold (STK11: OR, 41.9; 95% CI, 5.55 to 315). Multivariable models and matched case-control analyses yielded similar results. CONCLUSION: Among nearly 100,000 clinically tested women, 7% carried a pathogenic mutation in one or more cancer-associated genes. Associated breast and ovarian cancer risks ranged from two- to 40-fold after controlling for family history. These results may inform cancer risk counseling.

Prediction of distant recurrence in resected stage I and II lung adenocarcinoma.

OBJECTIVES: Optimal procedures for adjuvant treatment and post-surgical surveillance of resected non-small-cell lung cancer remain under discussion. Pathological features are the main determinant of follow-up therapy but have limited ability to identify patients at risk of recurrence. Increasingly, molecular markers are incorporated into clinical decision-making, including measures of tumor growth. The CCP score is a quantitative, molecular measure of proliferation derived from the RNA expression of 31 cell cycle genes and a component of the molecular prognostic score (mPS). The mPS score is a linear combination of CCP score and pathological stage. CCP score and mPS are independent predictors of survival in resected lung adenocarcinoma. MATERIALS AND METHODS: CCP scores were determined by RT-qPCR for 318 patients diagnosed with stage I-II lung adenocarcinoma. Association of mPS and CCP score with distant recurrence and lung-cancer specific survival was assessed in Cox proportional hazards regression models adjusted for age, gender, tumor size, pathological stage and pleural invasion. Distant recurrence-free survival and lung-cancer specific survival by mPS risk group were calculated by Kaplan-Meier survival analysis. RESULTS: CCP scores were obtained for 205 stage I and 84 stage II patients. CCP score and mPS were independent markers of distant recurrence (CCP: HR 1.62, 95%CI 1.15-2.29, p=0.0055; mPS: HR 2.22, 95%CI 1.11-4.44, p=0.023). Patients with low mPS tumors were at significantly reduced risk of distant recurrence (log-rank p=4.2×10-5). Among stage I patients, stratification by mPS identified a patient group with increased risk of distant recurrence (36%, 95%CI 28-46%, log-rank p=0.0011) CONCLUSIONS: The molecular prognostic score stratifies early-stage, resected lung cancer patients for risk of distant recurrence and could be useful to inform treatment and surveillance decisions.

Classification of genetic variants in genes associated with Lynch syndrome using a clinical history weighting algorithm.

BACKGROUND: Lynch syndrome is a hereditary cancer syndrome associated with high risks of colorectal and endometrial cancer that is caused by pathogenic variants in the mismatch repair genes (MLH1, MSH2, MSH6, PMS2, EPCAM). Accurate classification of variants identified in these genes as pathogenic or benign enables informed medical management decisions. Previously, we developed a clinical History Weighting Algorithm (HWA) for the classification of variants of uncertain significance (VUSs) in BRCA1 and BRCA2. The BRCA1/2 HWA is based on the premise that pathogenic variants in these genes will be identified more often in individuals with strong personal and/or family histories of breast and/or ovarian cancer, while the identification of benign variants should be independent of cancer history. Here we report the development of a similar HWA to allow for classification of VUSs in genes associated with Lynch syndrome using data collected through both syndrome-specific and pan-cancer panel testing. METHODS: Upon completion of algorithm development, the HWA was tested using simulated variants constructed from 79,214 probands, as well as 379 true variants. Positive (PPV) and negative predictive values (NPV) were calculated on a per gene basis. RESULTS: 25,500 pathogenic and 50,500 benign simulated variants were analyzed using the HWA and the PPVs and NPVs for each gene were greater than 0.997 and 0.999, respectively. The HWA was also evaluated using 100 trials for each of the 379 true variants. PPVs of >0.998 and NPVs of >0.999 were obtained for all genes. CONCLUSIONS: We have developed and implemented a HWA to aid in the classification of VUSs in genes associated with Lynch syndrome. The work presented here demonstrates that this HWA is able to classify MLH1, MSH2, and MSH6 VUSs as either benign or pathogenic with high accuracy.

Stratification of resectable lung adenocarcinoma by molecular and pathological risk estimators.

BACKGROUND: Mortality in early stage, resectable lung cancer is sufficiently high to warrant consideration of post-surgical treatment. Novel markers to stratify resectable lung cancer patients may help with the selection of treatment to improve outcome. METHODS: Primary tumour tissue from 485 patients, surgically treated for stage I-II lung adenocarcinoma, was analysed for the RNA expression of 31 cell cycle progression (CCP) genes by quantitative polymerase chain reaction (PCR). The expression average, the CCP score, was combined with pathological stage into a prognostic score (PS). Cox proportional hazards regression assessed prediction of 5-year lung cancer mortality above clinical variables. The PS threshold was tested for risk discrimination by the Mantel-Cox log-rank test. RESULTS: The CCP score added significant information above clinical markers (all patients, P=0.0029; stage I patients, P=0.013). The prognostic score was a superior predictor of outcome compared to pathological stage alone (PS, P=0.00084; stage, P=0.24). Five-year lung cancer mortality was significantly different between the low-risk (90%, 95% confidence interval (CI) 81-95%), and high-risk groups (65%, 95% CI 57-72%), P=4.2×10(-6)). CONCLUSIONS: The CCP score is an independent prognostic marker in early stage lung adenocarcinoma. The prognostic score provides superior risk estimates than stage alone. The threefold higher risk in the high-risk group defines a subset of patients that should consider therapeutic choices to improve outcome.

Analytical validation of a proliferation-based molecular signature used as a prognostic marker in early stage lung adenocarcinoma.

AIMS: The aim of these studies was to validate the analytical performance of a cell cycle progression (CCP) gene signature that provides prognostic information for early stage lung adenocarcinomas. MATERIALS & METHODS: Formalin-fixed paraffin-embedded (FFPE) lung resections were evaluated by quantitative RT-PCR for the expression of 31 target and 15 housekeeper genes comprising the CCP score. RESULTS: The signature had a standard deviation (SD) of 0.06 score units and a dynamic range spanning CCP scores between -13 and 14. The average amplicon efficiencies for target and housekeeper genes were 107% and 105%, respectively. All but one amplicon had a SD <0.5 CT. CONCLUSION: These studies demonstrate that the gene signature is robust and reproducible, making it suitable for use in a clinical setting.