Prospective evaluation of finger two-point discrimination and carpal tunnel syndrome among women with breast cancer receiving adjuvant aromatase inhibitor therapy.

PURPOSE: Aromatase inhibitors (AIs) are associated with musculoskeletal symptoms and risk of developing carpal tunnel syndrome (CTS), which can impair quality of life and prompt treatment discontinuation. The incidence of CTS and clinical utility of diagnostic tests such as 2-point discrimination (2-PD) have not been prospectively examined among women receiving AIs. METHODS: Postmenopausal women with stage 0-III hormone receptor-positive breast cancer who were enrolled in a randomized clinical trial investigating adjuvant AIs (Exemestane and Letrozole Pharmacogenetics, ELPh) underwent prospective evaluation of 2-PD with the Disc-criminator™ (sliding aesthesiometer) and completed a CTS questionnaire at baseline, 3, 6, and 12 months, following initiation of AI. Changes in mean 2-PD were analyzed with multivariable mixed effects modelling. A p value < 0.05 was considered statistically significant. RESULTS: Of 100 women who underwent baseline 2-PD testing, CTS was identified by questionnaire in 11% at baseline prior to AI initiation. Prevalence of CTS at any time in the first year was 26%. A significant increase in worst 2-PD score was observed from baseline to 3 months (3.7 mm to 3.9 mm, respectively, p = 0.03) when adjusted for age, prior chemotherapy, randomized treatment assignment, and diabetes. There were no significant differences in treatment discontinuation due to CTS between the arms. CONCLUSION: For women receiving adjuvant AI, 2-PD scores were significantly worse at 3 months compared to baseline. Studies are required to assess whether change in 2-PD is an adequate objective assessment for CTS with AI therapy. Early diagnosis of CTS may expedite management, improve AI adherence, and enhance breast cancer outcomes.

Racial disparities in the rate of cardiotoxicity of HER2-targeted therapies among women with early breast cancer.

BACKGROUND: Human epidermal growth factor receptor 2 (HER2)-targeted therapies are highly effective at preventing breast cancer recurrence but are associated with cardiotoxicity in some patients, and minimal data are available regarding racial disparities in the incidence of this toxicity. The authors conducted a retrospective study to analyze the association of black or white race with treatment-induced cardiotoxicity and incomplete therapy among patients with HER2-positive early breast cancer. METHODS: Women with HER2-positive, stage I through III breast cancer who initiated (neo)adjuvant HER2-targeted therapy (trastuzumab with or without pertuzumab) from January 2005 to March 2015 at the authors' institution were eligible. We analyzed differences in the incidence of cardiotoxicity (a decline in the left ventricular ejection fraction to <50% AND an absolute drop in the left ventricular ejection fraction of ≥10% from baseline) and incomplete therapy (<52 weeks of HER2-targeted therapy) between black and white women in univariate and multivariable analyses. RESULTS: The authors identified 59 black patients and 157 white patients who had a median follow-up 5.2 years. The median patient age was 53 years and was similar for black and white patients. The 1-year cardiotoxicity incidence was 12% overall (95% confidence interval [CI], 7%-16%), 24% in black women (95% CI, 12%-34%), and 7% in white women (95% CI, 3%-11%). Black patients had a significantly greater probability of incomplete therapy compared with white patients (odds ratio, 4.61; 95% CI, 1.70-13.07; P = .002). High correlation was observed between a cardiotoxicity event and incomplete therapy (96% concordance). CONCLUSIONS: Black patients have a higher rate of cardiotoxicity and resultant incomplete adjuvant HER2-targeted therapy than white patients. This patient population may benefit from enhanced cardiac surveillance, cardioprotective strategies, and early referral to cardiology when appropriate. Cancer 2018;124:1904-11. © 2018 American Cancer Society.

A phase II study evaluating the efficacy of zoledronic acid in prevention of aromatase inhibitor-associated musculoskeletal symptoms: the ZAP trial.

PURPOSE: Aromatase inhibitor-associated musculoskeletal symptoms (AIMSS) are common adverse events of AIs often leading to drug discontinuation. We initiated a prospective clinical trial to evaluate whether bisphosphonates are associated with reduced incidence of AIMSS. METHODS: In the single-arm trial, the Zoledronic Acid Prophylaxis (ZAP) trial, we compared the incidence of AIMSS against historical controls from the Exemestane and Letrozole Pharmacogenomics (ELPh) trial. Eligible women were postmenopausal with stage 0-III breast cancer planning to receive adjuvant AIs. AIMSS was assessed using the Health Assessment Questionnaire and Visual Analog Scale over 12 months in both trials. Participants in the ZAP trial received zoledronic acid prior to initiating letrozole and after 6 months; ELPh participants included in the analysis were taking letrozole but not bisphosphonates. We analyzed patient-reported outcomes (PROs) and bone density in the ZAP trial using mixed-effects linear regression models and paired t tests, respectively. RESULTS: From 2011 to 2013, 59 postmenopausal women enrolled in ZAP trial. All 59 (100%) women received baseline and 52 (88%) received 6-month zoledronic acid, and had similar characteristics to historical controls from the ELPh trial (n = 206). Cumulatively during the first year of AI, 37 and 67% of ZAP and ELPh participants reported AIMSS (p < 0.001), respectively. Within the ZAP trial, we did not observe significant changes in other PROs; however, we report improvements in bone mineral density. CONCLUSIONS: Compared to historical controls, zoledronic acid administered concomitantly with adjuvant AIs was associated with a reduced incidence of AIMSS. A randomized controlled trial is required to confirm these findings.

Adjuvant radiation use in older women with early-stage breast cancer at Johns Hopkins.

PURPOSE: In 2004, The National Comprehensive Cancer Network (NCCN) Guidelines incorporated omission of radiation therapy after breast-conservation surgery in women ≥70 years old with stage I, estrogen receptor-positive breast cancer who plan to receive endocrine therapy. One study demonstrated wide variation in implementing this change across 13 NCCN institutions. We evaluated the practice pattern at our institution. METHODS: We identified women ≥70 years old treated at our institution from 2009 to 2014. We calculated radiation therapy omission rate in those meeting the guidelines. We explored associations between radiation therapy omission, year of diagnosis, and patient characteristics with Wilcoxon rank sum tests and Fisher's exact tests. RESULTS: A total of 667 women met the inclusion criteria, and 117 (18 %) were candidates for radiation therapy omission. Mean age among the 117 was 76.3 years (Range: 70-95). Overall radiation therapy omission rate was 36.8 %, but varied greatly by year of diagnosis (Range: 7.7-54.5 %). This variation persisted after excluding women who did not receive endocrine therapy (Mean: 39.0 %, Range: 0.0-75.0 %). Factors associated with higher radiation therapy omission rates included older age and not having pathological nodal evaluation. The radiation therapy omission rate did not vary by race, tumor type, grade, or size. CONCLUSIONS: The implementation of the NCCN guideline has not been consistent at our institution. Our data suggest that other tools should be considered to apply the guidelines more consistently. We have implemented a quality improvement protocol that incorporates life expectancy estimate and geriatric assessment in women meeting the NCCN guideline at our institution.

Diagnostic accuracy of sentinel node identification is maintained with the addition of local lidocaine and subareolar radioactive colloid injection.

Preoperative sentinel node localization (SNL) using a subareolar injection of radiotracer technetium-99m-sulfur colloid (Tc(99m)SC) is associated with significant pain. Lidocaine use during SNL is not widely adopted partly due to a concern that it can obscure sentinel node identification and reduce its diagnostic accuracy. We prospectively identified women with a biopsy-proven infiltrating breast cancer who were awaiting a SNL. The women completed the McGill pain questionnaire, Visual Analog Scale, and Wong-Baker FACES Pain Rating Scale prior to and following SNL. We identified a retrospective cohort of women with similar demographic and tumor characteristics who did not receive lidocaine before SNL. We compared sentinel lymph node identification rates in the two cohorts. We used Wilcoxon rank sum tests to compare continuous measures and Fisher's exact test for categorical measures. Between January 2011 to July 2012, 110 women consented, and 105 were eligible for and received lidocaine prior to Tc(99m)SC injection. The post-lidocaine identification rate of SNL was 95 % with Tc(99m)SC, and 100 % with the addition of intraoperative methylene blue dye/saline. Pain range prior to and following the SNL was unchanged (P = 0.703). We identified 187 women from 2005 to 2009 who did not receive lidocaine during preoperative SNL. There was no significant difference in the success rate of SNL, with or without lidocaine (P = 0.194). The administration of lidocaine during SNL prevents pain related to isotope injection while maintaining the success rate. We have changed our practice at our center to incorporate the use of lidocaine during all SNL.

Patient-reported outcomes in women with breast cancer enrolled in a dual-center, double-blind, randomized controlled trial assessing the effect of acupuncture in reducing aromatase inhibitor-induced musculoskeletal symptoms.

BACKGROUND: Aromatase inhibitors (AIs) have been associated with decrements in patient-reported outcomes (PROs). The objective of this study was to assess whether real acupuncture (RA), compared with sham acupuncture (SA), improves PROs in patients with breast cancer who are receiving an adjuvant AI. METHODS: Postmenopausal women with a stage 0 through III breast cancer who received an AI and had treatment-associated musculoskeletal symptoms were randomized to receive 8 weekly RA versus SA in a dual-center, randomized controlled trial. The National Surgical Adjuvant Breast and Bowel Project (NSABP) menopausal symptoms questionnaire, the Center for Epidemiological Studies Depression (CESD) scale, the Hospital Anxiety and Depression Scale (HADS), the Pittsburgh Sleep Quality Index (PSQI), the hot flash daily diary, the Hot Flash-Related Daily Interference Scale (HFRDI), and the European quality-of-life survey (EuroQol) were used to assess PROs at baseline and at 4weeks, 8 weeks, and 12 weeks. RESULTS: The intention-to-treat analysis included 23 patients in the RA arm and 24 patients in the SA arm. There were no significant differences in baseline characteristics between the 2 groups. Compared with baseline, scores in the RA arm improved significantly at week 8 on the CESD (P = .022), hot flash severity (P = .006), hot flash frequency (P = .011), the HFRDI (P = .014), and NSABP menopausal symptoms (P = .022); scores in the SA arm improved significantly on the EuroQol (P = .022),the HFRDI (P = .043), and NSABP menopausal symptoms (P = .005). Post-hoc analysis indicated that African American patients (n = 9) benefited more from RA than SA compared with non-African American patients (n = 38) in reducing hot flash severity (P < .001) and frequency (P < .001) scores. CONCLUSIONS: Both RA and SA were associated with improvement in PROs among patients with breast cancer who were receiving AIs, and no significant difference was detected between arms. Racial differences in response to acupuncture warrant further study.

A dual-center randomized controlled double blind trial assessing the effect of acupuncture in reducing musculoskeletal symptoms in breast cancer patients taking aromatase inhibitors.

Up to 50 % of women receiving aromatase inhibitor (AI) complain of AI-associated musculoskeletal symptoms (AIMSS) and 15 % discontinue treatment. We conducted a randomized, sham-controlled trial to evaluate whether acupuncture improves AIMSS and to explore potential mechanisms. Postmenopausal women with early stage breast cancer, experiencing AIMSS were randomized to eight weekly real or sham acupuncture sessions. We evaluated changes in the Health Assessment Questionnaire Disability Index (HAQ-DI) and pain visual analog scale (VAS) following the intervention compared to baseline. Serum estradiol, ß-endorphin, and proinflammatory cytokine concentrations were measured pre and post-intervention. We enrolled 51 women of whom 47 were evaluable, including 23 randomized to real and 24 to sham acupuncture. Baseline characteristics were balanced between groups with the exception of a higher HAQ-DI score in the real acupuncture group (p = 0.047). We did not observe a statistically significant difference in reduction of HAQ-DI (p = 0.30) or VAS (p = 0.31) between the two groups. Following eight weekly treatments, we observed a statistically significant reduction of IL-17 (p ≤ 0.009) in both groups. No significant modulation was seen in estradiol, ß-endorphin, or other proinflammatory cytokine concentrations in either group. We did not observe a significant difference in AIMSS changes between real and sham acupuncture. As sham acupuncture used in this study may not be equivalent to placebo, further studies with a non-acupuncture arm may be required to establish whether acupuncture is beneficial for the treatment of AIMSS.

Fixed-dose capecitabine is feasible: results from a pharmacokinetic and pharmacogenetic study in metastatic breast cancer.

The pro-drug capecitabine is approved for treatment of anthracycline- and paclitaxel-resistant metastatic breast cancer. However, toxicity and large interpatient pharmacokinetic variability occur despite body surface area (BSA)-dosing. We hypothesized that a fixed-dose schedule would simplify dosing and provide an effective and safe alternative to BSA-based dosing. We conducted an open label, single-arm, two-stage study of oral capecitabine with fixed starting dose (3,000 mg total daily dose in two divided doses × 14 days q21 days) in patients with metastatic breast cancer. We correlated pharmacodynamic endpoints [e.g., efficacy (response) per RECIST and toxicity], adherence and pharmacokinetics/pharmacogenetics. Sample size of 45 patients was required to detect a 25 % response rate from null response rate of 10 % using a Simon two-stage design. Twenty-six patients were enrolled in the first-stage and 21 were evaluable after a median of four cycles of capecitabine. Two thirds of patients received either the same dose or a dose 500 mg lower than what would have been administered with a commonly used 2,000 mg/m(2) BSA-dosing schedule. Eight patients had stable disease but progressed after a median of seven cycles. Despite a clinical benefit rate of 19 %, no RECIST responses were observed following the first stage and the study was closed. Dose-reductions were required for grade 2 hand-foot syndrome (28 %) and vomiting (5 %). Adherence was similar when using both patient-reported and Medication Event Monitoring System methods. High interpatient variability was observed for capecitabine and metabolite pharmacokinetics, but was not attributed to observed pharmacogenetic or BSA differences. Single agent activity of capecitabine was modest in our patients with estrogen receptor-positive or -negative metastatic breast cancer and comparable to recent studies. BSA was not the main source of pharmacokinetic variability. Fixed-dose capecitabine is feasible, and simplifies dosing.

Pilot study of breast sensation after breast reconstruction: evaluating the effects of radiation therapy and perforator flap neurotization on sensory recovery.

BACKGROUND: Some sensation to the breast returns after breast reconstruction, but recovery is variable and unpredictable. We primarily sought to assess the impact of different types of breast reconstruction [deep inferior epigastric artery perforator (DIEP) flaps versus implants] and radiation therapy on the return of sensation. METHODS: Thirty-seven patients who had unilateral or bilateral breast reconstruction via a DIEP flap or implant-based reconstruction, with or without radiation therapy (minimum follow-up, 18 months; range, 18-61 months) were studied. Of the 74 breasts, 27 had DIEP flaps, 29 had implants, and 18 were nonreconstructed. Eleven breasts with implants and 10 with DIEP flaps had had prereconstruction radiation therapy. The primary outcome was mean patient-perceived static and moving cutaneous pressure threshold in nine areas. We used univariate and multivariate analyses to assess what independent factors affected the return of sensation (significance, P < 0.05). RESULTS: Implants provided better static (P = 0.071) and moving sensation (P = 0.041) than did DIEP flaps. However, among irradiated breasts, skin over DIEP flaps had significantly better sensation than did that over implants (static, P = 0.019; moving, P = 0.028). Implant reconstructions with irradiated skin had significantly worse static (P = 0.002) and moving sensation (P = 0.014) than did nonirradiated implant reconstructions. CONCLUSIONS: Without irradiation, skin overlying implants is associated with better sensation recovery than DIEP flap skin. However, with irradiation, DIEP flap skin had better sensation recovery than did skin over implants. Neurotization trended toward improvement in sensation in DIEP flaps.

A short-term biomarker modulation study of simvastatin in women at increased risk of a new breast cancer.

Observational studies have demonstrated a decreased incidence of cancers among users of HMG CoA reductase inhibitors (statins) and a reduced risk of recurrence among statin users diagnosed with early stage breast cancer. We initiated a prospective study to identify potential biomarkers of simvastatin chemopreventive activity that can be validated in future trials. The contralateral breast of women with a previous history of breast cancer was used as a high-risk model. Eligible women who had completed all planned treatment of a prior stage 0-III breast cancer received simvastatin 40 mg orally daily for 24-28 weeks. At baseline and end-of-study, we measured circulating concentrations of high-sensitivity C-reactive protein (hsCRP), estrogens, and fasting lipids; breast density on contralateral breast mammogram; and quality of life by Rand Short Form 36-Item health survey. Fifty women were enrolled with a median age of 53 years. Total cholesterol, LDL cholesterol, triglyceride, and hsCRP fell significantly during the study (P values < 0.001, <0.001, 0.003, and 0.05, respectively). Estrone sulfate concentrations decreased with simvastatin treatment (P = 0.01 overall), particularly among post-menopausal participants (P = 0.006). We did not observe a significant change in circulating estradiol or estrone concentrations, contralateral mammographic breast density, or reported physical functioning or pain scores. This study demonstrates the feasibility of short-term biomarker modulation studies using the contralateral breast of high-risk women. Simvastatin appears to modulate estrone sulfate concentrations and its potential chemopreventive activity in breast cancer warrants further investigation.

Comparison of breast cancer recurrence risk and cardiovascular disease incidence risk among postmenopausal women with breast cancer.

The majority of breast cancers are diagnosed in postmenopausal women. Competing comorbidities, particularly cardiovascular disease (CVD), should be considered when individualizing adjuvant therapies for these women. We compared the 10-year predicted breast cancer recurrence risk with CVD risk among postmenopausal women with hormone receptor-positive (HR+), non-metastatic breast cancer. CVD risk factor data were prospectively collected from postmenopausal women with stage I-III, HR+ breast cancer initiating adjuvant aromatase inhibitor therapy. We compared predicted 10-year CVD risk, including the composite index heart age, computed from modified Framingham risk score, with predicted 10-year risk of breast cancer recurrence using Adjuvant! Online. We created multivariable logistic regression models to estimate the odds ratios (OR) and 95% confidence intervals (CI) for greater CVD risk than breast cancer recurrence risk. Among 415 women, mean age and heart age were 60 and 67 years, respectively. Overall, 43% of women had a predicted 10-year CVD risk equivalent to breast cancer recurrence risk and 37% had CVD risk higher than breast cancer recurrence risk. Predicted CVD risk was higher than breast cancer recurrence risk for stage I disease (OR: 6.1, 95% CI: 3.4-11.2) or heart age >65 (OR: 12.4, 95% CI: 7.0-22.6). The majority of postmenopausal women with HR+ early breast cancer had a predicted 10-year CVD risk that was equivalent to or higher than breast cancer recurrence risk. Physicians should weigh competing risks and offer early screening and cardiac prevention strategies for women at a greater risk for CVD.

Monitoring of neoadjuvant chemotherapy using multiparametric, ²³Na sodium MR, and multimodality (PET/CT/MRI) imaging in locally advanced breast cancer.

We prospectively investigated using advanced magnetic resonance imaging (MRI) and positron emission tomography/computed tomography (PET/CT) to identify radiological biomarkers for treatment response in patients receiving preoperative systemic therapy (PST) for locally advanced breast cancer. Patients with a stage II or III breast cancer receiving PST were selected and underwent positron emission tomography (PET), magnetic resonance imaging (MRI), and breast biopsies at baseline and after the first cycle of PST (days 7-8) during the full course of treatment. PET/CT was acquired after injection of 2-deoxy-2-[18F]-fluoro-D-glucose (¹8FDG, 0.22 mCi/kg) and quantified with standardized uptake value assessment (SUV). Diagnostic breast MRI and sodium (²³Na) was acquired at 1.5 T. Total tissue sodium concentration (TSC), response criteria in solid tumors (RECIST), and volumes were quantified. Treatment response was determined by pathological assessment at surgery. Immunohistochemistry values of the proliferative index (Ki-67) were performed on biopsy specimens. Six of nineteen eligible women (43 ± 11 years) who received PST underwent radiological imaging of ¹8FDG-PET/CT and MRI for at least two cycles of treatment. Five patients had a pathological partial response (pPR) and one had pathological non-response (pNR). TSC decreased 21% in responders with increases in the non-responder (P = 0.03). Greater reduction in SUV was observed in responders (38%) compared to the non-responder (22%; P = 0.03). MRI volumes decreased after cycle 1 by 42% (responders) and 35% (non-responder; P = 0.11). Proliferation index Ki-67 declined in responders in the first cycle (median = 47%, range = 29-20%), but increased (4%) in the non-responder. Significant decreases in TSC, SUV, and Ki-67 were observed in responders with increases in TSC and Ki-67 in non-responders. Our results demonstrate the feasibility of using multi-modality proton, ²³Na MRI, and PET/CT metrics as radiological biomarkers for monitoring response to PST in patients with operable breast cancer.

Docetaxel metabolism is not altered by imatinib: findings from an early phase study in metastatic breast cancer.

Docetaxel is primarily metabolized by CYP3A4 and susceptible to alterations in clearance by CYP3A4 inhibition and induction. Imatinib is a CYP3A4 inhibitor. A phase I study of docetaxel and imatinib in metastatic breast cancer (MBC) was conducted to test the hypothesis that imatinib decreased docetaxel clearance. Docetaxel was administered weekly × 3 with daily imatinib, repeated every 28 days; during cycle 1, imatinib was started on day 8. Docetaxel and imatinib pharmacokinetics, and hepatic CYP3A4 activity (erythromycin breath test) were evaluated during cycles 1 and 2. Toxicity and efficacy were assessed. Twelve patients were enrolled to three docetaxel/imatinib dose levels: 20 mg/m(2)/600 mg (DL1), 25 mg/m(2)/600 mg (DL2), and 25 mg/m(2)/400 mg (DL2a). Median number of prior chemotherapy regimens was 2 (range, 0-8). Toxicities were primarily observed at DL2; dose-limiting toxicities were Grade 3 transaminase elevations and diarrhea, and 5 patients had grade 2 nausea. Two patients had partial responses (7 months); two stable disease (2 and 4 months); five had progressive disease. Despite a 42% decrease in CYP3A4 activity after 3 weeks of imatinib co-administration, docetaxel clearance was unchanged. Mean ± standard deviation steady-state imatinib trough concentration (2.6 ± 1.2 µg/ml) was approximately 2.6-fold higher than previously observed in other cancer populations, and likely contributed to the poor tolerability of the combination in MBC. In conclusion, imatinib inhibited CYP3A4 but did not affect docetaxel clearance. Clinically, further investigation of this combination in MBC is not warranted due to excessive toxicities. However, these unexpected pharmacokinetic findings support further investigation of mechanisms underlying docetaxel elimination pathways.

In vitro and in vivo oxidative metabolism and glucuronidation of anastrozole.

AIMS: Little information is available regarding the metabolic routes of anastrozole and the specific enzymes involved. We characterized anastrozole oxidative and conjugation metabolism in vitro and in vivo. METHODS: A sensitive LC-MS/MS method was developed to measure anastrozole and its metabolites in vitro and in vivo. Anastrozole metabolism was characterized using human liver microsomes (HLMs), expressed cytochrome P450s (CYPs) and UDP-glucuronosyltransferases (UGTs). RESULTS: Hydroxyanastrozole and anastrozole glucuronide were identified as the main oxidative and conjugated metabolites of anastrozole in vitro, respectively. Formation of hydroxyanastrozole from anastrozole was markedly inhibited by CYP3A selective chemical inhibitors (by >90%) and significantly correlated with CYP3A activity in a panel of HLMs (r= 0.96, P= 0.0005) and mainly catalyzed by expressed CYP3A4 and CYP3A5. The K(m) values obtained from HLMs were also close to those from CYP3A4 and CYP3A5. Formation of anastrozole glucuronide in a bank of HLMs was correlated strongly with imipramine N-glucuronide, a marker of UGT1A4 (r= 0.72, P < 0.0001), while expressed UGT1A4 catalyzed its formation at the highest rate. Hydroxyanastrozole (mainly as a glucuronide) and anastrozole were quantified in plasma of breast cancer patients taking anastrozole (1 mg day⁻¹); anastrozole glucuronide was less apparent. CONCLUSION: Anastrozole is oxidized to hydroxyanastrozole mainly by CYP3A4 (and to some extent by CYP3A5 and CYP2C8). Once formed, this metabolite undergoes glucuronidation. Variable activity of CYP3A4 (and probably UGT1A4), possibly due to genetic polymorphisms and drug interactions, may alter anastrozole disposition and its effects in vivo.

Promoter hypermethylation in sentinel lymph nodes as a marker for breast cancer recurrence.

PURPOSE: Promoter methylation of tumor suppressor genes in histologically negative sentinel lymph nodes (HNSN) of early stage breast cancer patients has not been extensively studied. This study evaluates the methylation frequency and pattern in HNSN to determine if detection of hypermethylation of one or more genes is associated with an increased recurrence risk in node negative breast cancer. EXPERIMENTAL DESIGN: In 1998, a prospective study of patients with early stage breast cancer and HNSN was initiated in order to correlate sentinel node analysis with clinical outcome. Nodal tissue was selected from 120 HNSN patients for methylation analysis in at least one and up to six sentinel nodes using a panel of nine genes. Corresponding primary breast tumors from 79 patients were also evaluated for hypermethylation. Methylation analysis was performed using nested Methylation Sensitive PCR (n-MSP). Logistical regression was used to evaluate the relationship between clinical recurrence and methylation status. RESULTS: Over a median follow-up of 79 months, 13 of the 120 patients had clinical recurrence. Hypermethylation of genes was frequently observed in HNSN, but there was no correlation of methylation pattern and clinical recurrence. However, increased frequency of gene methylation of the primary tumor correlated with clinical recurrence. CONCLUSIONS: Although hypermethylation of multiple genes occurs frequently in HNSN of breast cancer patients, it is not associated with breast cancer recurrence in the first 7 years of clinical follow-up.

Monitoring of Serum DNA Methylation as an Early Independent Marker of Response and Survival in Metastatic Breast Cancer: TBCRC 005 Prospective Biomarker Study.

Purpose Epigenetic alterations measured in blood may help guide breast cancer treatment. The multisite prospective study TBCRC 005 was conducted to examine the ability of a novel panel of cell-free DNA methylation markers to predict survival outcomes in metastatic breast cancer (MBC) using a new quantitative multiplex assay (cMethDNA). Patients and Methods Ten genes were tested in duplicate serum samples from 141 women at baseline, at week 4, and at first restaging. A cumulative methylation index (CMI) was generated on the basis of six of the 10 genes tested. Methylation cut points were selected to maximize the log-rank statistic, and cross-validation was used to obtain unbiased point estimates. Logistic regression or Cox proportional hazard models were used to test associations between the CMI and progression-free survival (PFS), overall survival (OS), and disease status at first restaging. The added value of the CMI in predicting survival outcomes was evaluated and compared with circulating tumor cells (CellSearch). Results Median PFS and OS were significantly shorter in women with a high CMI (PFS, 2.1 months; OS, 12.3 months) versus a low CMI (PFS, 5.8 months; OS, 21.7 months). In multivariable models, among women with MBC, a high versus low CMI at week 4 was independently associated with worse PFS (hazard ratio, 1.79; 95% CI, 1.23 to 2.60; P = .002) and OS (hazard ratio, 1.75; 95% CI, 1.21 to 2.54; P = .003). An increase in the CMI from baseline to week 4 was associated with worse PFS ( P < .001) and progressive disease at first restaging ( P < .001). Week 4 CMI was a strong predictor of PFS, even in the presence of circulating tumor cells ( P = .004). Conclusion Methylation of this gene panel is a strong predictor of survival outcomes in MBC and may have clinical usefulness in risk stratification and disease monitoring.

Combination Epigenetic Therapy in Advanced Breast Cancer with 5-Azacitidine and Entinostat: A Phase II National Cancer Institute/Stand Up to Cancer Study.

Purpose: In breast cancer models, combination epigenetic therapy with a DNA methyltransferase inhibitor and a histone deacetylase inhibitor led to reexpression of genes encoding important therapeutic targets, including the estrogen receptor (ER). We conducted a multicenter phase II study of 5-azacitidine and entinostat in women with advanced hormone-resistant or triple-negative breast cancer (TNBC).Experimental Design: Patients received 5-azacitidine 40 mg/m2 (days 1-5, 8-10) and entinostat 7 mg (days 3, 10) on a 28-day cycle. Continuation of epigenetic therapy was offered with the addition of endocrine therapy at the time of progression [optional continuation (OC) phase]. Primary endpoint was objective response rate (ORR) in each cohort. We hypothesized that ORR would be ≥20% against null of 5% using Simon two-stage design. At least one response was required in 1 of 13 patients per cohort to continue accrual to 27 per cohort (type I error, 4%; power, 90%).Results: There was one partial response among 27 women with hormone-resistant disease (ORR = 4%; 95% CI, 0-19), and none in 13 women with TNBC. One additional partial response was observed in the OC phase in the hormone-resistant cohort (n = 12). Mandatory tumor samples were obtained pre- and posttreatment (58% paired) with either up- or downregulation of ER observed in approximately 50% of posttreatment biopsies in the hormone-resistant, but not TNBC cohort.Conclusions: Combination epigenetic therapy was well tolerated, but our primary endpoint was not met. OC phase results suggest that some women benefit from epigenetic therapy and/or reintroduction of endocrine therapy beyond progression, but further study is needed. Clin Cancer Res; 23(11); 2691-701. ©2016 AACR.

Individualized Molecular Analyses Guide Efforts (IMAGE): A Prospective Study of Molecular Profiling of Tissue and Blood in Metastatic Triple-Negative Breast Cancer.

PURPOSE: The clinical utility of next-generation sequencing (NGS) in breast cancer has not been demonstrated. We hypothesized that we could perform NGS of a new biopsy from patients with metastatic triple-negative breast cancer (TNBC) in a clinically actionable timeframe. EXPERIMENTAL DESIGN: We planned to enroll 40 patients onto a prospective study, Individualized Molecular Analyses Guide Efforts (IMAGE), to evaluate the feasibility of obtaining a new biopsy of a metastatic site, perform NGS (FoundationOne), and convene a molecular tumor board to formulate treatment recommendations within 28 days. We collected blood at baseline and at time of restaging to assess cell-free circulating plasma tumor DNA (ptDNA). RESULTS: We enrolled 26 women with metastatic TNBC who had received ≥1 line of prior chemotherapy, and 20 (77%) underwent NGS of a metastatic site biopsy. Twelve (60%) evaluable patients received treatment recommendations within 28 days of consent. The study closed after 20 patients underwent NGS, based on protocol-specified interim futility analysis. Three patients went on to receive genomically directed therapies. Twenty-four of 26 patients had genetic alterations successfully detected in ptDNA. Among 5 patients, 4 mutations found in tumor tissues were not identified in blood, and 4 mutations found in blood were not found in corresponding tumors. In 9 patients, NGS of follow-up blood samples showed 100% concordance with baseline blood samples. CONCLUSIONS: This study demonstrates challenges of performing NGS on prospective tissue biopsies in patients with metastatic TNBC within 28 days, while also highlighting the potential use of blood as a more time-efficient and less invasive method of mutational assessment. Clin Cancer Res; 23(2); 379-86. ©2016 AACR.

A Clinicopathologic Analysis of 45 Patients With Metaplastic Breast Carcinoma.

OBJECTIVES: Metaplastic breast carcinomas (MBCs) are rare, aggressive cancers lacking targeted therapy. Here, we review the clinicopathologic features, treatment, and outcomes of patients with MBC treated at our institution. METHODS: We searched clinical and pathology databases for patients with histologically confirmed MBC from 1999 to 2012. We estimated survival probabilities using the Kaplan-Meier method and evaluated prognostic factors using Cox regression. RESULTS: Forty-five cases were identified, including chondroid (24%), spindled (20%), sarcomatoid (16%), squamous (11%), and mixed (29%) histologic subtypes. Median tumor size was 3 cm, with 86% grade III and 69% triple-negative for estrogen receptor, progesterone receptor, and human epidermal growth factor receptor 2. Most had negative lymph nodes, and two patients had metastases at diagnosis. Six patients received neoadjuvant therapy, with one pathologic complete response. All patients underwent surgery, 60% received adjuvant radiation, and 58% had adjuvant chemotherapy. Five-year recurrence-free survival was 64%; 5-year overall survival was 69%. Tumor size, history of breast cancer, and mixed histology were associated with inferior outcomes. CONCLUSIONS: We report one of the largest single-institution series of patients with MBC. MBC is associated with a poor prognosis, despite low nodal involvement. Most patients in this series had high-grade, triple-negative tumors and were treated with optimal therapy.

Gene Methylation and Cytological Atypia in Random Fine-Needle Aspirates for Assessment of Breast Cancer Risk.

Methods to determine individualized breast cancer risk lack sufficient sensitivity to select women most likely to benefit from preventive strategies. Alterations in DNA methylation occur early in breast cancer. We hypothesized that cancer-specific methylation markers could enhance breast cancer risk assessment. We evaluated 380 women without a history of breast cancer. We determined their menopausal status or menstrual cycle phase, risk of developing breast cancer (Gail model), and breast density and obtained random fine-needle aspiration (rFNA) samples for assessment of cytopathology and cumulative methylation index (CMI). Eight methylated gene markers were identified through whole-genome methylation analysis and included novel and previously established breast cancer detection genes. We performed correlative and multivariate linear regression analyses to evaluate DNA methylation of a gene panel as a function of clinical factors associated with breast cancer risk. CMI and individual gene methylation were independent of age, menopausal status or menstrual phase, lifetime Gail risk score, and breast density. CMI and individual gene methylation for the eight genes increased significantly (P < 0.001) with increasing cytological atypia. The findings were verified with multivariate analyses correcting for age, log (Gail), log (percent density), rFNA cell number, and body mass index. Our results demonstrate a significant association between cytological atypia and high CMI, which does not vary with menstrual phase or menopause and is independent of Gail risk and mammographic density. Thus, CMI is an excellent candidate breast cancer risk biomarker, warranting larger prospective studies to establish its utility for cancer risk assessment. Cancer Prev Res; 9(8); 673-82. ©2016 AACR.