Relationships between computer-extracted mammographic texture pattern features and BRCA1/2 mutation status: a cross-sectional study.

INTRODUCTION: Mammographic density is similar among women at risk of either sporadic or BRCA1/2-related breast cancer. It has been suggested that digitized mammographic images contain computer-extractable information within the parenchymal pattern, which may contribute to distinguishing between BRCA1/2 mutation carriers and non-carriers. METHODS: We compared mammographic texture pattern features in digitized mammograms from women with deleterious BRCA1/2 mutations (n = 137) versus non-carriers (n = 100). Subjects were stratified into training (107 carriers, 70 non-carriers) and testing (30 carriers, 30 non-carriers) datasets. Masked to mutation status, texture features were extracted from a retro-areolar region-of-interest in each subject's digitized mammogram. Stepwise linear regression analysis of the training dataset identified variables to be included in a radiographic texture analysis (RTA) classifier model aimed at distinguishing BRCA1/2 carriers from non-carriers. The selected features were combined using a Bayesian Artificial Neural Network (BANN) algorithm, which produced a probability score rating the likelihood of each subject's belonging to the mutation-positive group. These probability scores were evaluated in the independent testing dataset to determine whether their distribution differed between BRCA1/2 mutation carriers and non-carriers. A receiver operating characteristic analysis was performed to estimate the model's discriminatory capacity. RESULTS: In the testing dataset, a one standard deviation (SD) increase in the probability score from the BANN-trained classifier was associated with a two-fold increase in the odds of predicting BRCA1/2 mutation status: unadjusted odds ratio (OR) = 2.00, 95% confidence interval (CI): 1.59, 2.51, P = 0.02; age-adjusted OR = 1.93, 95% CI: 1.53, 2.42, P = 0.03. Additional adjustment for percent mammographic density did little to change the OR. The area under the curve for the BANN-trained classifier to distinguish between BRCA1/2 mutation carriers and non-carriers was 0.68 for features alone and 0.72 for the features plus percent mammographic density. CONCLUSIONS: Our findings suggest that, unlike percent mammographic density, computer-extracted mammographic texture pattern features are associated with carrying BRCA1/2 mutations. Although still at an early stage, our novel RTA classifier has potential for improving mammographic image interpretation by permitting real-time risk stratification among women undergoing screening mammography.

Circulating estrogens and estrogens within the breast among postmenopausal BRCA1/2 mutation carriers.

Accurately quantifying parent estrogens (PE) estrone (E1) and estradiol (E2) and their metabolites (EM) within breast tissue and serum may permit detailed investigations of their contributions to breast carcinogenesis among BRCA1/2 mutation carriers. We conducted a study of PE/EM in serum, nipple aspirate fluid (NAF), and ductal lavage supernatant (DLS) among postmenopausal BRCA1/2 mutation carriers. PE/EM (conjugated and unconjugated) were measured in paired serum/NAF (n = 22 women) and paired serum/DLS samples (n = 24 women) using quantitative liquid chromatography-tandem mass spectrometry (LC/MS/MS). The relationships between serum and tissue-specific PE/EM were measured using Pearson's correlation coefficients. Conjugated forms of PE/EM constituted the majority of estrogen in serum (88 %), NAF (59 %) and DLS (69 %). PE/EM in NAF and serum were highly correlated [E1 (r = 0.97, p < 0.0001), E2 (r = 0.90, p < 0.0001) and estriol (E3) (r = 0.74, p < 0.0001)] as they were in DLS and serum [E1 (r = 0.92, p < 0.0001; E2 (r = 0.70, p = 0.0001; E3 (r = 0.67, p = 0.0004)]. Analyses of paired total estrogen values for NAF and serum, and DLS and serum yielded ratios of 0.22 (95 % CI 0.19-0.25) and 0.28 (95 % CI 0.24-0.32), respectively. This report is the first to employ LC/MS/MS to quantify PE/EM in novel breast tissue-derived biospecimens (i.e., NAF and DLS). We demonstrate that circulating PE and EM are strongly and positively correlated with tissue-specific PE and EM measured in NAF and DLS among postmenopausal BRCA1/2 mutation carriers. If confirmed, future etiologic studies could utilize the more readily obtainable serum hormone levels as a reliable surrogate measure of exposure at the tissue level.

Mammographic density does not differ between unaffected BRCA1/2 mutation carriers and women at low-to-average risk of breast cancer.

Elevated mammographic density (MD) is one of the strongest risk factors for sporadic breast cancer. Epidemiologic evidence suggests that MD is, in part, genetically determined; however, the relationship between MD and BRCA1/2 mutation status is equivocal. We compared MD in unaffected BRCA1/2 mutation carriers enrolled in the U.S. National Cancer Institute's Clinical Genetics Branch's Breast Imaging Study (n = 143) with women at low-to-average breast cancer risk enrolled in the same study (n = 29) or the NCI/National Naval Medical Center's Susceptibility to Breast Cancer Study (n = 90). The latter were BRCA mutation-negative members of mutation-positive families or women with no prior breast cancer, a Pedigree Assessment Tool score <8 (i.e., low risk of a hereditary breast cancer syndrome) and a Gail score <1.67. A single experienced mammographer measured MD using a computer-assisted thresholding method. We collected standard breast cancer risk factor information in both studies. Unadjusted mean percent MD was higher in women with BRCA1/2 mutations compared with women at low-to-average breast cancer risk (37.3% vs. 33.4%; P = 0.04), but these differences disappeared after adjusting for age and body mass index (34.9% vs. 36.3%; P = 0.43). We explored age at menarche, nulliparity, age at first birth, menopausal status, number of breast biopsies, and exposure to exogenous hormonal agents as potential confounders of the MD and BRCA1/2 association. Taking these factors into account did not significantly alter the results of the age/body mass index-adjusted analysis. Our results do not provide support for an independent effect of BRCA1/2 mutation status on mammographic density.

Cardiovascular and Metabolic Responses to Carbon Dioxide Euthanasia in Conscious and Anesthetized Rats.

Euthanasia is a necessary component in research and must be conducted humanely. Currently, regulated CO2 exposure in conscious rats is acceptable, but data are divided on whether CO2 alone is more distressing than anesthesia prior to CO2. To evaluate distress in rats, we compared physiologic responses to CO2 euthanasia with and without isoflurane preanesthesia. Male Sprague-Dawley rats were implanted with telemetry devices to measure mean arterial pressure (MAP), heart rate (HR), and blood glucose. Animals recovered for 2 wk and were then exposed to either 5% isoflurane (n = 6) or 100% CO2 (n = 7; calculated 30% chamber volume/min displacement) in their home cages to induce loss of consciousness. Euthanasia was then completed with CO2 in both groups. MAP and HR increased when the gas delivery lids were placed on the home cages of both groups. Both MAP and HR gradually decreased with isoflurane exposure. MAP increased and HR decreased with CO2 exposure. Glucose levels remained stable throughout the procedure, except for a small drop in conscious animals initially exposed to 100% CO2. These data suggest that both gases affect the measured parameters in a similar manner, and that environmental factors, such as gas delivery lid placement, also change these measurements.

Ductal lavage in women from BRCA1/2 families: is there a future for ductal lavage in women at increased genetic risk of breast cancer?

PURPOSE: Ductal lavage has been used for risk stratification and biomarker development and to identify intermediate endpoints for risk-reducing intervention trials. Little is known about patient characteristics associated with obtaining nipple aspirate fluid (NAF) and adequate cell counts (> or =10 cells) in ductal lavage specimens from BRCA mutation carriers. METHODS: We evaluated patient characteristics associated with obtaining NAF and adequate cell counts in ductal lavage specimens from the largest cohort of women from BRCA families yet studied (BRCA1/2 = 146, mutation-negative = 23, untested = 2). Fisher's exact test was used to evaluate categorical variables; Wilcoxon nonparametric test was used to evaluate continuous variables associated with NAF or ductal lavage cell count adequacy. Logistic regression was used to identify independent correlates of NAF and ductal lavage cell count adequacy. RESULTS: From 171 women, 45 (26%) women had NAF and 70 (41%) women had ductal lavage samples with > or =10 cells. Postmenopausal women with intact ovaries compared with premenopausal women [odds ratio (OR), 4.8; P = 0.03] and women without a prior breast cancer history (OR, 5.2; P = 0.04) had an increased likelihood of yielding NAF. Having breast-fed (OR, 3.4; P = 0.001), the presence of NAF before ductal lavage (OR, 3.2; P = 0.003), and being premenopausal (OR, 3.0; P = 0.003) increased the likelihood of ductal lavage cell count adequacy. In known BRCA1/2 mutation carriers, only breast-feeding (OR, 2.5; P = 0.01) and the presence of NAF (OR, 3.0; P = 0.01) were independent correlates of ductal lavage cell count adequacy. CONCLUSIONS: Ductal lavage is unlikely to be useful in breast cancer screening among BRCA1/2 mutation carriers because the procedure fails to yield adequate specimens sufficient for reliable cytologic diagnosis or to support translational research activities.

Tolerability of breast ductal lavage in women from families at high genetic risk of breast cancer.

BACKGROUND: Ductal lavage (DL) has been proposed as a minimally-invasive, well-tolerated tool for obtaining breast epithelial cells for cytological evaluation of breast cancer risk. We report DL tolerability in BRCA1/2 mutation-positive and -negative women from an IRB-approved research study. METHODS: 165 BRCA1/2 mutation-positive, 26 mutation-negative and 3 mutation unknown women underwent mammography, breast MRI and DL. Psychological well-being and perceptions of pain were obtained before and after DL, and compared with pain experienced during other screening procedures. RESULTS: The average anticipated and experienced discomfort rating for DL, 47 and 48 (0-100), were significantly higher (p < 0.01) than the anticipated and experienced discomfort of mammogram (38 and 34), MRI (36 and 25) or nipple aspiration (42 and 27). Women with greater pre-existing emotional distress experienced more DL-related discomfort than they anticipated. Women reporting DL-related pain as worse than expected were nearly three times more likely to refuse subsequent DL than those reporting it as the same or better than expected. Twenty-five percent of participants refused repeat DL at first annual follow-up. CONCLUSION: DL was anticipated to be and experienced as more uncomfortable than other procedures used in breast cancer screening. Higher underlying psychological distress was associated with decreased DL tolerability.

Prospectively Identified Incident Testicular Cancer Risk in a Familial Testicular Cancer Cohort.

BACKGROUND: Human testicular germ cell tumors (TGCT) have a strong genetic component and a high familial relative risk. However, linkage analyses have not identified a rare, highly penetrant familial TGCT (FTGCT) susceptibility locus. Currently, multiple low-penetrance genes are hypothesized to underlie the familial multiple-case phenotype. The observation that two is the most common number of affected individuals per family presents an impediment to FTGCT gene discovery. Clinically, the prospective TGCT risk in the multiple-case family context is unknown. METHODS: We performed a prospective analysis of TGCT incidence in a cohort of multiple-affected-person families and sporadic-bilateral-case families; 1,260 men from 140 families (10,207 person-years of follow-up) met our inclusion criteria. Age-, gender-, and calendar time-specific standardized incidence ratios (SIR) for TGCT relative to the general population were calculated using SEER*Stat. RESULTS: Eight incident TGCTs occurred during prospective FTGCT cohort follow-up (versus 0.67 expected; SIR = 11.9; 95% CI, 5.1-23.4; excess absolute risk = 7.2/10,000). We demonstrate that the incidence rate of TGCT is greater among bloodline male relatives from multiple-case testicular cancer families than that expected in the general population, a pattern characteristic of adult-onset Mendelian cancer susceptibility disorders. Two of these incident TGCTs occurred in relatives of sporadic-bilateral cases (0.15 expected; SIR = 13.4; 95% CI, 1.6-48.6). CONCLUSIONS: Our data are the first to indicate that despite relatively low numbers of affected individuals per family, members of both multiple-affected-person FTGCT families and sporadic-bilateral TGCT families comprise high-risk groups for incident testicular cancer. IMPACT: Men at high TGCT risk might benefit from tailored risk stratification and surveillance strategies.