Citations

Abstract Purpose: Mammographic breast density (MBD) is decreased by tamoxifen, but the effect of aromatase inhibitors is less clear. Experimental Design: We enrolled early-stage postmenopausal patients with breast cancer initiating adjuvant aromatase inhibitor therapy and ascertained mammograms before and at an average 10 months of aromatase inhibitor therapy. We matched cases to healthy postmenopausal women (controls) from a large mammography screening cohort on age, baseline body mass index, baseline MBD, and interval between mammograms. We estimated change in MBD using a computer-assisted thresholding program (Cumulus) and compared differences between cases and matched controls. Results: In predominantly White women (96%), we found 14% of the 387 eligible cases had a MBD reduction of at least 5% after an average of 10 months of aromatase inhibitor therapy. MBD reductions were associated with higher baseline MBD, aromatase inhibitor use for more than 12 months, and prior postmenopausal hormone use. Comparing each case with her matched control, there was no evidence of an association of change in MBD with aromatase inhibitor therapy [median case–control difference among 369 pairs was −0.1% (10th and 90th percentile: −5.9%, 5.2%) P = 0.51]. Case–control differences were similar by type of aromatase inhibitor (P's 0.41 and 0.56); prior use of postmenopausal hormones (P = 0.85); baseline MBD (P = 0.55); and length of aromatase inhibitor therapy (P = 0.08). Conclusions: In postmenopausal women treated with aromatase inhibitors, 14% of cases had a MBD reduction of more than 5%, but these decreases did not differ from matched controls. These data suggest that MBD is not a clinically useful biomarker for predicting the value of aromatase inhibitor therapy in White postmenopausal women. Clin Cancer Res; 19(8); 2144–53. ©2013 AACR.

Purpose In patients with hormone-dependent postmenopausal breast cancer, standard adjuvant therapy involves 5 years of the nonsteroidal aromatase inhibitors anastrozole and letrozole. The steroidal inhibitor exemestane is partially non–cross-resistant with nonsteroidal aromatase inhibitors and is a mild androgen and could prove superior to anastrozole regarding efficacy and toxicity, specifically with less bone loss. Patients and Methods We designed an open-label, randomized, phase III trial of 5 years of exemestane versus anastrozole with a two-sided test of superiority to detect a 2.4% improvement with exemestane in 5-year event-free survival (EFS). Secondary objectives included assessment of overall survival, distant disease–free survival, incidence of contralateral new primary breast cancer, and safety. Results In the study, 7,576 women (median age, 64.1 years) were enrolled. At median follow-up of 4.1 years, 4-year EFS was 91% for exemestane and 91.2% for anastrozole (stratified hazard ratio, 1.02; 95% CI, 0.87 to 1.18; P = .85). Overall, distant disease–free survival and disease-specific survival were also similar. In all, 31.6% of patients discontinued treatment as a result of adverse effects, concomitant disease, or study refusal. Osteoporosis/osteopenia, hypertriglyceridemia, vaginal bleeding, and hypercholesterolemia were less frequent on exemestane, whereas mild liver function abnormalities and rare episodes of atrial fibrillation were less frequent on anastrozole. Vasomotor and musculoskeletal symptoms were similar between arms. Conclusion This first comparison of steroidal and nonsteroidal classes of aromatase inhibitors showed neither to be superior in terms of breast cancer outcomes as 5-year initial adjuvant therapy for postmenopausal breast cancer by two-way test. Less toxicity on bone is compatible with one hypothesis behind MA.27 but requires confirmation. Exemestane should be considered another option as up-front adjuvant therapy for postmenopausal hormone receptor–positive breast cancer.

Genome‐wide association studies (GWAS) of complex traits have generated many association signals for single nucleotide polymorphisms (SNPs). To understand the underlying causal genetic variant(s), focused DNA resequencing of targeted genomic regions is commonly used, yet the current cost of resequencing limits sample sizes for resequencing studies. Information from the large GWAS can be used to guide choice of samples for resequencing, such as the SNP genotypes in the targeted genomic region. Viewing the GWAS tag‐SNPs as imperfect surrogates for the underlying causal variants, yet expecting that the tag‐SNPs are correlated with the causal variants, a reasonable approach is a two‐phase case‐control design, with the GWAS serving as the first‐phase and the resequencing study serving as the second‐phase. Using stratified sampling based on both tag‐SNP genotypes and case‐control status, we explore the gains in power of a two‐phase design relative to randomly sampling cases and controls for resequencing (i.e., ignoring tag‐SNP genotypes). Simulation results show that stratified sampling based on both tag‐SNP genotypes and case‐control status is not likely to have lower power than stratified sampling based only on case‐control status, and can sometimes have substantially greater power. The gain in power depends on the amount of linkage disequilibrium between the tag‐SNP and causal variant alleles, as well as the effect size of the causal variant. Hence, the two‐phase design provides an efficient approach to follow‐up GWAS signals with DNA resequencing.

Abstract Purpose: Controversy exists about CYP2D6 genotype and tamoxifen efficacy. Experimental Design: A matched case–control study was conducted using the Austrian Breast and Colorectal Cancer Study Group Trial 8 (ABCSG8) that randomized postmenopausal women with estrogen receptor (ER)-positive breast cancer to tamoxifen for 5 years (arm A) or tamoxifen for 2 years followed by anastrozole for 3 years (arm B). Cases had disease recurrence, contralateral breast cancer, second non–breast cancer, or died. For each case, controls were identified from the same treatment arm of similar age, surgery/radiation, and tumor–node—metastasis (TNM) stage. Genotyping was conducted for alleles associated with no (PM; *3, *4, *6), reduced (IM; *10, and *41), and extensive (EM: absence of these alleles) CYP2D6 metabolism. Results: The common CYP2D6*4 allele was in Hardy–Weinberg equilibrium. In arm A during the first 5 years of therapy, women with two poor alleles [PM/PM: OR, 2.45; 95% confidence interval (CI), 1.05–5.73, P = 0.04] and women with one poor allele (PM/IM or PM/EM: OR, 1.67; 95% CI, 0.95–2.93; P = 0.07) had a higher likelihood of an event than women with two extensive alleles (EM/EM). In years 3 to 5 when patients remained on tamoxifen (arm A) or switched to anastrozole (arm B), PM/PM tended toward a higher likelihood of a disease event relative to EM/EM (OR, 2.40; 95% CI, 0.86–6.66; P = 0.09) among women on arm A but not among women on arm B (OR, 0.28; 95% CI, 0.03–2.30). Conclusion: In ABCSG8, the negative effects of reduced CYP2D6 metabolism were observed only during the period of tamoxifen administration and not after switching to anastrozole. Clin Cancer Res; 19(2); 500–7. ©2012 AACR.

Abstract Purpose: Aflibercept is a novel decoy receptor that efficiently neutralizes circulating VEGF. A pediatric phase I trial was conducted to define the dose-limiting toxicities (DLT), maximum tolerated dose (MTD), and pharmacokinetics (PK) of aflibercept. Experimental Design: Cohorts of three to six children with refractory solid tumors received aflibercept intravenously over 60 minutes every 14 days, at 2.0, 2.5, or 3.0 mg/kg/dose. PK sampling and analysis of peripheral blood biomarkers were conducted with the initial dose. Results: Twenty-one eligible patients were enrolled; 18 were fully evaluable for toxicity. One of six patients receiving 2.0 mg/kg/dose developed dose-limiting intratumoral hemorrhage and two of six receiving 3.0 mg/kg/dose developed either dose-limiting tumor pain or tissue necrosis. None of the six patients receiving 2.5 mg/kg/dose developed DLTs, defining this as the MTD. The most common non-DLTs were hypertension and fatigue. Three patients with hepatocellular carcinoma, hepatoblastoma and clear cell sarcoma had stable disease for >13 weeks. At the MTD, the ratio of free-to-bound aflibercept serum concentration was 2.10 on day 8 but only 0.44 by day 15. A rapid decrease in VEGF (P < 0.05) and increase in placental growth factor (PlGF; P < 0.05) from baseline was observed in response to aflibercept by day 2. Conclusions: The aflibercept MTD in children of 2.5 mg/kg/dose every 14 days is lower than the adult recommended dose of 4.0 mg/kg. This dose achieves, but does not sustain, free aflibercept concentrations in excess of bound. Tumor pain and hemorrhage may be evidence of antitumor activity but were dose-limiting. Clin Cancer Res; 18(18); 5081–9. ©2012 AACR.

Abstract Breast cancer recurrence is believed to be caused by a potently tumorigenic sub-population of cancer cells possessing stem cell attributes of resistance to chemotherapy, radiation and other forms of treatment. Recently, we and others have reported the generation of breast cancer stem cell (BCSC) characteristics by inducers of epithelial to mesenchymal transition (EMT). Both EMT and acquisition of BCSC properties are therefore important steps in cancer progression, although the physiological process by which BCSCs arise in vivo remains unclear. We show here that exposure to TGFβ and TNFα induces a stable BCSC phenotype, characterized by self-renewal, greatly increased tumorigenicity, and increased resistance to oxaliplatin, etoposide and paclitaxel. We further found that the TGFβ/TNFα-derived cells showed downregulated expression of Claudins 3, 4 and 7 and the luminal marker KRT18, demonstrating a shift to the claudin low molecular subtype, a recently identified breast cancer subtype characterized by the expression of mesenchymal and stem cell-associated markers and correlation with poor prognosis. Furthermore, pathway analyses of the generated BCSCs showed deregulation of a number of breast cancer-associated genes and pathways including inflammatory cytokine and receptors, chemokines, homeobox, hedgehog, notch and metastases genes. These results confirm the role of EMT and BCSCs in resistance of breast cancer to conventional chemotherapy and identify novel target genes and pathways for validation as potential therapeutic targets. Our demonstration that stable BCSCs can be produced by specific microenvironmental signals also provides a valuable tool for studying cancer stem cells, as well as a means for screening potential cancer drugs for efficacy of targeting cancer stem cells to prevent recurrence. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 3341. doi:10.1158/1538-7445.AM2011-3341

Abstract Introduction: The NSABP P-1 and P-2 breast cancer prevention trials have demonstrated that the SERMs tamoxifen and raloxifene can reduce the risk of breast cancer in high risk women. Methods: We performed a nested matched case-control GWAS utilizing DNA from participants enrolled in P-1 and P-2. 592 participants developed invasive breast cancer or ductal carcinoma in situ, and were matched with 1171 controls who did not. Genome-wide genotyping with the Illumina 610-Quad chip and CYP2D6 genotyping for pertinent alleles were performed. Functional genomic studies involved the use of cultured cells, with siRNA knockdown, followed by qRT-PCR and Western blot analyses. Results: Eleven SNPs with p-values < 3E-05 were identified. Twenty one additional SNPs with p < 4E-05 were identified by imputation around the genotyped SNPs on chromosomes (Chrs) 4, 8, 9, 13, 16 and 22, which were then genotyped with the Invader platform. Initial functional genomic studies focused on 6 SNPs on Chr16 (p-values 1.81-9.55E-06), all of which were in ZNF423, a gene encoding a putative zinc-finger protein. The Chr16 minor variant SNP sequences had less risk than the common variant (OR=0.7). Our functional studies showed that incubation of Hs578T cells stably transfected with estrogen receptor (ER) α with 0.1 nM estradiol (E2) induced expression of ZNF423, BRCA1/2 for WT but not variant SNP sequences. There was also a striking difference between E2-induced ZNF423, BRCA1/2 expression for variant and WT SNPs in lymphoblastoid cell lines (LCLs) stably transfected with ERα, with only WT showing induction. ChIP assay showed that ZNF423 could bind to the 5’-flanking region of BRCA1, and reporter gene assays showed that ZNF423 could regulate BRCA1/2 transcription. In LCLs stably transfected with ERα and genotyped for ZNF423 SNPs, estrogen-dependent expression of all 3 genes occurred only in the presence of the WT, but not variant ZNF423 SNP sequences. Blockade of ERα with 4-OH tamoxifen prevented induction of expression in cells with WT sequences, but resulted in gene dose-dependent increases in BRCA1/2 expression in cells with variant SNP sequences-suggested that the minor variant has a “protective” effect during clinical SERM therapy and verified by ChIP assays. Conclusions: The SNPs in ZNF423 identified during GWAS were associated with differential E2-dependent induction of ZNF423, BRCA1 and BRCA2-implying that ZNF423 is “upstream” for the estrogen-dependent induction of BRCA1/2 expression. However, LCLs with variant SNP sequences showed greatly enhanced expression of ZNF423, BRCA1/2 during 4-OH tamoxifen exposure, while cells with WT sequences did not. These results may reveal a novel mechanism for SERM-dependent regulation of BRCA1/2 expression and may have implications for patient selection for SERM therapy. Funded by U10-CA37377, U10-CA69974 and U14-GM61388. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 4727. doi:10.1158/1538-7445.AM2011-4727

Abstract Background: Exemestane, a steroidal aromatase inhibitor, reduced the incidence of invasive breast cancers by 65% among 4560 post-menopausal randomized to exemestane or placebo for 5 years on MAP.3. Differences in quality of life (QOL) were judged to be minimal, but only summary information was reported. Purpose: To provide more detailed information about effects of exemestane on menopause-specific and health-related qualities of life. Method: Participation in quality of life assessment was an eligibility criterion. Menopause-specific and health-related qualities of life were assessed using the MENQOL (4 scales; physical, vasomotor, psychosocial, sexual) and SF-36 (8 scales; physical health, role function — physical, bodily pain, general health, vitality, social function, role function — emotional, mental health, and 2 summary scales) instruments, respectively at baseline, 6 months and then yearly after randomization. Compliance with QOL questionnaire completion at each follow-up visit ranged from 93–98%, and did not differ by group. Change scores for each MENQOL and SF-36 scale, calculated for each assessment time relative to baseline, were compared using the Wilcoxon Rank-Sum test. Summary scores were used to summarize the QOL scores observed at each time point for each SF-36 dimension and overall mental (MCS) and physical component summaries (PCS) and MENQOL domains. Clinically important worsening of MENQOL change scores was defined as an increase of ≥0.5/8 points. SF-36 change scores were considered worsened if scores decreased by ≥ 5 points from baseline. Results: Both groups were balanced on scores for MENQOL and SF-36 at baseline. Median follow-up was 35 months and the proportion of women who stopped study medication early for toxicity reasons was 15% in the exemestane arm and 11% in the placebo arm. There was a statistically significant difference in change scores for vasomotor symptoms among women on exemestane during the first 4 years (p-values <0.01), compared to placebo. However, no between-group differences in vasomotor change met the criterion for clinical importance. Women on exemestane had statistically poorer sexual functioning (mean change = −0.02, SD=1.37) compared to placebo (mean change = −0.12, SD=1.32) during the first 6 months on study (p-value = 0.03) but the differences were not statistically significant thereafter or clinically important at any time. Among the 8 SF-36 scales, only bodily pain was statistically different between exemestane and placebo for the first 24 months on study medication (p-value <0.01), but no between-group difference in change scores exceeded 5 points. Overall SF-36 PCS and MCS assessing changes in overall physical and mental health-related QOL did not differ significantly by group at any assessment. Conclusion: Our assessment that early differences in vasomotor symptoms and pain were probably not clinically important is supported by the observation of no between-group differences when overall physical and mental health-related QOL changes were compared. Exemestane does not appear to have a major negative impact on the quality of life among these women. Citation Information: Cancer Res 2011;71(24 Suppl):Abstract nr S6-1.

Abstract Background: We have previously reported the results of surgical excision without irradiation for selected patients with DCIS in ECOG E5194, where the 5-year rates of local recurrence varied with age, grade, and lesion size (Hughes et al. J Clin Oncol 27:5319, 2009). New methods are needed to provide more accurate and reproducible assessment of recurrence risk. Methods: ECOG E5194 included 670 eligible patients with DCIS treated with surgical excision (≥ 3 mm negative margins) without irradiation, 228 of whom received tamoxifen. Patients had low or intermediate grade DCIS ≤ 2.5 cm, or high grade DCIS ≤ 1 cm. The Oncotype DX® assay was performed by quantitative RT-PCR using formalin fixed paraffin embedded tumor specimens from 327 patients (49% of the parent study). Recurrence Score® (RS) was calculated using the published algorithm. A new, prespecified DCIS Score™ was designed to predict recurrence using an optimized gene expression algorithm. The primary objective was to determine whether there was a significant association between the risk of an ipsilateral breast event (IBE) and the continuous DCIS Score in Cox models. 46 patients had an IBE (defined as ipsilateral local recurrence of DCIS [n=20] or invasive cancer [n=26]). Median follow-up was 8.8 years. Results: The 10-year IBE rates were 15.4% for low/intermediate grade DCIS and 15.1% for high-grade DCIS (as determined by central pathology review), and for invasive IBE, 5.6% and 9.8%, respectively. Comparison between local and expert grading showed substantial disagreement. Continuous DCIS Score was significantly associated with IBE (HR 2.34 per 50 units; 95% CI 1.15, 4.59; p=0.02) when adjusted for tamoxifen use (prespecified primary analysis) and with invasive IBE (HR 3.73; CI 1.34, 9.82; p=0.01). DCIS Score was significantly associated with outcome when evaluated by the prespecified risk groups (see Table). Similar results were observed with and without adjustment for tamoxifen use or for negative margin width. Features associated with IBE in multivariate models included menopausal status (HR 0.49; 95% CI 0.27, 0.90; p=0.02), tumor size (HR 1.52 per 5 mm; 95% CI 1.11, 2.01; p=0.01), and continuous DCIS Score (HR 2.41; 95% CI 1.15, 4.89; p=0.02). The standard RS, which is calculated using thresholding of many genes unlike the DCIS Score, was not associated with IBE or invasive IBE (p > 0.6). Conclusions: We have prospectively validated a multigene assay that quantifies recurrence risk and complements traditional clinical and pathologic factors in selected patients with DCIS treated with surgical excision without irradiation. The DCIS Score provides a new clinical tool for individualized selection of treatment for patients with DCIS. Citation Information: Cancer Res 2011;71(24 Suppl):Abstract nr S4-6.