Shepherd, L. E.

567 Background: We proposed adjunctive statistical standardization of quantitated ER and PgR to improve inter-laboratory comparability of biomarker results and therapeutic management of breast cancer. Methods: We utilized CCTG MA.27 (NCT00066573), an adjuvant phase III trial of exemestane versus anastrozole in postmenopausal women with ER+ and/or PgR+ tumours. IHC ER and PgR HSCORE and % positivity (%+) were centrally assessed by machine image quantitation, and each statistically standardized to mean of 0, standard deviation of 1 following Box-Cox variance stabilization transformations of square for ER; for PgR, 1.) natural logarithm (0.1 added to 0 HSCOREs and 0 %+), 2.) square root. The primary endpoint was STEEP distant disease-free survival (DDFS) at the longest trial follow-up of median 4.1 years; a secondary endpoint was event-free survival (EFS). Survival was described with Kaplan-Meier plots and tested with the univariate Wilcoxon (Peto-Prentice) test statistic. We examined cut-points at standard deviations about mean of 0 (<-1; (-1,0]; (0,1]; >1) and explored single cut-points. Cox multivariate regressions were adjusted for age, T and N stage, grade, lymphovascular invasion, treatment, and baseline patient demographics; 2-sided Wald tests had nominal significance if p<0.05. Results: Of the 7576 women accrued, 3048 women had machine-quantitated image analysis results: 2900 (95%) for ER; 2726 (89%) for PgR. Statistically standardized HSCORE and %+ units differentiated both univariate DDFS and EFS; DDFS was significantly different by ER levels (p<0.001) and PgR levels (p<0.001). In multivariable assessments, ER HSCORE and %+ were not significantly associated (p=0.52-0.88) with the DDFS primary endpoint in models with PgR, while higher PgR HSCORE and %+ had significantly better DDFS (p=.001, in all instances) in models with ER. Conclusions: DDFS was superior for patients with higher ER and PgR standardized units compared with those with HSCOREs and %+ <-1. The adjunctive statistical standardization of ER and PgR performed here is similar to that mandated for clinical practice by the World Health Organization for BMD. [Table: see text]

Abstract Background: Adjuvant breast cancer therapy is informed by whether a tumour is positive or negative for the biomarkers ER, PgR, and HER2, often without regard to level of positivity. Quantitation has been proposed to improve therapeutic management. Adjunctive statistical standardization has been proposed to improve inter-laboratory comparability of biomarkers results. Methods: This primary report utilized adjunctive statistical standardization of machine-quantitated image analysis biomarker assessments. CCTG MA.27 (NCT00066573) is an adjuvant phase III trial of exemestane versus anastrozole in postmenopausal women with ER+ and/or PgR+ tumours. IHC ER, PgR, and HER2 were centrally assessed, with FISH (HER2;HER2/CEP17) determinations for equivocal IHC HER2. HSCOREs were statistically standardized to a mean of 0, standard deviation of 1 following Box-Cox variance stabilization transformations of square for ER and natural logarithm for PgR (0.1 was added to 0 HSCOREs). The primary endpoint was STEEP distant disease-free survival (DDFS) at the longest trial follow-up of median 4.1 years. Survival was described with Kaplan-Meier plots. The univariate Wilcoxon (Peto-Prentice) test statistic was used with usual designation of negative/positive (0; &gt;0), and standardized cut-points at standard deviations about mean of 0(&lt;-1; (-1,0]; (0,1]; &gt;1). Cox multivariate regressions adjusted for age, T and N stage, grade, lymphovascular invasion, treatment, and baseline patient demographics, utilized likelihood ratio tests. Nominal significance was p=0.05. Results: Of the 7576 women accrued, 3048 had machine-quantitated image analysis results: 2900 (95%) for ER; 2726 (89%) for PgR. Only 8 women were ASCO/CAP ER- (HSCORE 0); PgR HSCORE was 0 for 533. Statistically standardized units differentiated DDFS ER levels (p&lt; 0.001) and PgR levels (p&lt; 0.001). In adjusted multivariate analyses, higher ER HSCORE was associated with better DDFS (p=0.05) with weak evidence of an association (p=0.11) for standardized HSCORE, and no significant association (respectively, p=0.28, p=0.54) in models with PgR. Higher PgR was associated with better DDFS (p=0.001) in all multivariate assessments, including those with ER. Conclusions: DDFS was superior for patients with higher ER and PgR standardized units compared with those with HSCOREs &lt;-1. Adjunctive statistical standardization, similar to that mandated for clinical practice by the World Health Organization for BMD, should improve inter-laboratory comparability of biomarker results for similar patient populations. Biomarker N DDFS DDFS 5-year (%) 95% CI ER total 2900 ER &lt;-1 506 86 (82, 91) ER (-1, 0] 934 94 (92, 96) ER ( 0, 1] 919 94 (92, 96) ER &gt;1 541 96 (93, 98) PgR total 2726 PgR &lt;-1 734 89 (86, 92) PgR (-1, 0] 439 92 (89, 95) PgR ( 0, 1] 967 95 (93, 96) PgR &gt;1.0 586 98 (97,100) Citation Format: Judy-Anne Chapman, Jane Bayani, Sandip SenGupta, John MS Bartlett, Tammy Piper, Mary Anne Quintayo, Shakeel Virk, Paul Goss, James Ingle, Matthew Ellis, George Sledge Jr, George Budd, Manuela Rabaglio, Rafat Ansari, Richard Tozer, David D'Souza, Haji Chalchal, Silvana Spadafora, Vered Stearns, Edith A. Perez, Karen Gelmon, Timothy Whelan, Catherine Elliott, Lois Shepherd, Bingshu Chen, Karen Taylor. Adjunctive statistical standardization of quantitated machine image analysis of Estrogen and Progesterone Receptors: CCTG MA.27 trial [abstract]. In: Proceedings of the 2023 San Antonio Breast Cancer Symposium; 2023 Dec 5-9; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2024;84(9 Suppl):Abstract nr PO1-27-10.

6018 Background: Disease free survival was significantly improved in women receiving letrozole after standard adjuvant tamoxifen in the MA.17 trial. Based on the results of MA.17 and of other trials of aromatase inhibitors in early stage breast cancer, chronic aromatase inhibitor therapy, in postmenopausal women free of breast cancer recurrence, is now being widely employed. We analyzed the toxicity of letrozole according to ethnic status among women enrolled in MA.17. Methods: The chi-square test was used for comparison of rates of side effects between the two groups, Caucasian vs. ethnic minority (defined as all non-Caucasians). In a subset of women, quality of life (QOL) was assessed by the SF-36 Health Survey. Mean change scores in QOL from baseline were compared between groups for summary measures and domains using the Wilcoxon test. Results: 352 minority women and 4,708 Caucasians were enrolled in MA.17, of which 183 minority women and 2,339 Caucasians were randomized to receive letrozole. Caucasians were older than minority women and had a slightly longer duration of treatment with prior tamoxifen. Tumor size and nodal status were not significantly different between the two groups. In women who received letrozole, minority women had significantly lower incidence of hot flashes (49% vs. 58%; p = 0.02), fatigue (29% vs. 39%; p = 0.005), and arthritis (2% vs. 7%; p = 0.006) compared with Caucasians. Mean QOL change scores of SF-36 domains for women who received letrozole were not different but minority women had better mental health at 6 month assessment (p = 0.02) and worse bodily pain at 12 month assessment (p = 0.046). Conclusions: Minority women tolerated letrozole considerably better than Caucasians in the MA.17 trial. These preliminary findings suggest that minority women respond differently to letrozole in terms of toxicity. Recent demonstration of genotypic variations in the aromatase gene in different ethnic groups plus likely pharmacogenomic differences suggests that further research is needed to clarify the clinical outcomes of aromatase inhibition in women of diverse ethnicities. Future research strategies should focus on examining in vivo genotype-phenotype correlations to determine the effects of genetic variation on response to anticancer therapy and on toxicities and end-organ effects. [Table: see text]

549 Background: MA.17 evaluated letrozole (LET) or placebo (PLAC) after 5 years of tamoxifen (Tam) and showed [median follow-up 30 months (mos)] significant improvement in disease-free survival (DFS) for LET [hazard ratio (HR) 0.57, p = 0.00008]. The trial was unblinded and PLAC patients (pts) were offered LET. An ITT analysis of all outcomes, before and after unblinding, based on the original randomization was performed. Methods: A stratified log-rank test was used to compare DFS, distant (D)DFS, overall survival (OS) and incidence of contra-lateral breast cancer (CBC). The Cox regression model used baseline stratification variables and two prespecified factors, menopausal status at the start of Tam and time on Tam. Subgroup analyses for DFS and OS were performed for the two prespecified subsets. All p-values were two-sided Results: 5187 pts were randomized at baseline and, at unblinding, 1655 of 2268 PLAC pts accepted LET. At median follow-up of 54 mos (range,16–86) 363 recurrences or CBC’s (144 LET and 219 PLAC) occurred; 118 LET and 176 PLAC pts had recurrent disease and 26 LET and 43 PLAC pts had CBC. 4 year DFS was 94.3% (LET) and 91.4% (PLAC) (HR 0.64; 95% CI, 0.52 - 0.79; p = 0.00002). Corresponding 4 year DDFS was 96.2% and 94.9% (HR 0.76; 0.58–0.99; p = 0.041). 4 year OS was 95.0% (LET) and 95.1% (PLAC) (HR 1.00; 0.78–1.28; p = 0.99). LET was equally effective in node +ve and -ve pts (i.e., similar HRs) in DFS. OS was not significantly different for LET and PLAC in any subgroup. The annual rate of CBC was 0.29% LET (0.18–0.40) and 0.47% PLAC (0.34–0.61); HR 0.61 (0.38–0.98) p = 0.037. 255 pts had died as of the data cut-off (128 LET and 127 PLAC). Conclusions: In this ITT analysis, pts originally randomized to LET within 3 months of stopping Tam did better than PLAC pts in DFS, DDFS and CBC, despite 73% of PLAC pts crossing to LET after unblinding. This highlights the strong beneficial effect of extended adjuvant therapy with LET. [Table: see text]

540 Background: Risk of death from other malignancies (OM) and other causes (OC) than breast cancer (BC) increases with age. Effects of baseline factors on type of death were assessed with competing risks analyses. Methods: In NCIC CTG MA.17, 5,187 women free of recurrent breast cancer after 5 years of tamoxifen were randomized to letrozole (L, 2,593 women) or placebo (P, 2,594 women). The primary endpoint was disease free survival (DFS), and secondary, overall survival (OS). Follow-up was to October 9, 2005: median 3.9 years, range <0.1 to 7.0 years. Effects of competing risks were examined for endpoints of BC, OM, and OC for 11 baseline trial factors: treatment, age, menopausal status, duration of prior tamoxifen, adjuvant radiotherapy, bone fracture, osteoporosis, cardiovascular disease, hormone receptor status, nodal status, adjuvant chemotherapy. Lagakos’ hierarchical method (Lagakos, Appl. Statist. 1978; 27:235–241) was used to test for differential effects of baseline factors on type of death (BC, OM, OC). Results: Rate of censoring was 97.8%, with 256 deaths (BC, 102; OM, 50; OC, 100; unknown, 4). Non-breast cancer deaths accounted for 60% of known deaths; 72%, for those ≥70 years; and 48%, for those <70 years. Two baseline factors differentially affected type of death. Women with cardiovascular disease were more likely to die from OC (p=0.02), while those with osteoporosis were more likely to die of OM (p=0.03). Age and nodal status had directionally similar effects. Older women had shorter survival from all 3 causes of death (p=0.01). Lymph node positivity was associated with worse survival (p=0.003). Conclusions: Extended L provides similar proportional benefit in improving DFS for all ages of women (Muss ref abstract SABCS 2006). However, the magnitude of competing non-breast cancer, and non-treatment related, causes of death needs to be considered more frequently, since with early detection and improved therapies, breast cancer patients may increasingly be expected to survive their disease to die from another cause. The novel association between baseline osteoporosis and other malignancies is being explored quantitatively. No significant financial relationships to disclose.

2 Background: The MA.17 trial demonstrated that extended adjuvant endocrine therapy with letrozole after 5-y of tamoxifen markedly reduced the risk of recurrence in women with ER+ early stage breast cancer. This trial provides an opportunity to assess the ability of biomarkers to predict late recurrences in ER+ breast cancer. The Breast Cancer Index (BCI), a continuous risk index based on the combination of HOXB13:IL17BR (H:I) and the molecular grade index (MGI), estimates the individual risk of recurrence in ER+ breast cancer patients. In this study, the prognostic utility of BCI to predict late recurrences was examined. Methods: FFPE tumor blocks were collected from patients who experienced a breast cancer recurrence up to unblinding of MA.17. Controls were matched 2:1 for age, tumor size, nodal status and prior chemotherapy, and were disease free for longer than cases. All cases were reviewed for standard histopathology and evaluated using the real-time RT-PCR BCI assay. Results: Patient characteristics for the case-control study were similar to that from the overall study. Characteristics for cases (N=83) and controls (N=166) were not significantly different except for treatment. A higher percentage of controls compared to cases tended to be categorized as low risk by BCI (58% vs 43%), while a lower percentage of controls than cases tended to be categorized as high risk by BCI (34% vs 24%). In univariate analysis, treatment, BCI, H:I and HOXB13, but not tumor grade or MGI, were significant predictors of late recurrence. After adjusting for standard variables (age, tumor grade and treatment), BCI (OR 2.37; P=0.03), H:I (OR 2.55; P=0.04) and HOXB13 (OR 1.35; P=0.02) remained significant predictors of recurrence. HOXB13 expression at diagnosis predicted patient benefit from extended endocrine therapy with letrozole. Conclusions: In this case-controlled study, the data demonstrate that BCI is a significant predictor of late recurrences in ER+ patients following 5-y of tamoxifen. The prognostic performance of BCI to predict late recurrences was largely dependent on HOXB13 expression. The integration of H:I and MGI within BCI provides prognostic utility for both early and late recurrences.

Abstract Background: In 2000, cyclophosphamide(C), epirubicin(E) and fluorouracil (F) (CEF) and doxorubicin (A) and C followed by paclitaxel (T) (AC/T) given every three weeks were commonly used regimens in Canada and the USA to treat women with node positive or high risk node negative operable breast cancer. In a previous trial in women with locally advanced breast cancer, 3 months of dose-dense EC was equivalent to six months of CEF. We hypothesized that the addition of 3 months of a taxane following dose-dense EC would be superior to CEF alone or AC/T. In a randomized trial in women with early breast cancer we compared CEF, dose dense EC/T and AC/T. An interim analysis (JCO, 2010) conducted when 50% of the events for the relapse-free-survival (RFS) analysis were observed, demonstrated that AC/T administered every 3 weeks was significantly inferior to CEF or EC/T. The results of the final RFS including CEF versus EC/T will be presented. Methods: Women, 60 years or younger, with operable axillary node positive or high risk node negative breast cancer were randomized to adjuvant CEF, EC/T or AC/T. CEF was given for 6 cycles and consisted of: C 75 mg/m2 orally on Days 1–14, and E 60 mg/m2 and F 500 mg/m2, IV on Days 1 and 8. CEF patients received concurrent antibiotic prophylaxis. EC/T consisted of 6 cycles of EC every 14 days; E 120 mg/m2 and C 830 mg/m2, both IV on Day 1 with filgrastim 5 [ug]/kg/day subcutaneous from Days 2–13 and epoetin alpha 40,000 units subcutaneous once weekly. After completion of EC, 4 cycles of T 175 mg/m2 every 21 days with filgrastim and epoetin alpha were given. The AC/T regimen consisted of A 60 mg/m2 and C 600 mg/m2 IV every 21 days for 4 cycles. This was followed by T 175 mg/m2 every 21 days for 4 cycles. The final analysis for RFS was planned when 453 events were observed permitting the detection of a hazard ratio (HR) of 1.43 between CEF and AC/T and a HR of 1.43 between EC/T and the best of the other two arms. Quality of Life data using the EORTC C-30 and the Breast Cancer Chemotherapy questionnaires were collected throughout the study. The results will be available at the time of the presentation. Results: A total of 2104 women were enrolled between December 2000 and April 2005. The required 453 events for the RFS analysis were observed by the clinical cut-off date of March 30, 2012. At this point 369 deaths had been observed. The median follow-up is 87.5 months. Outstanding data and queries are currently being collected and reviewed with an expected database lock and final analysis mid August, 2012. Febrile neutropenia remains higher on the CEF and EC/T arms while more neuropathy was seen in the taxane containing regimens. Conclusions: Although the two regimens considered a standard of care at the time the study was initiated are no longer widely used, the trial results will enhance our understanding of the magnitude of benefit of taxane following an anthracycline, the significance of cumulative anthracycline dose, and the role of dose density/ intensity as backbone of conventional chemo regimens. Citation Information: Cancer Res 2012;72(24 Suppl):Abstract nr P1-13-01.

Abstract Background: The third-generation aromatase inhibitors (AIs) are established adjuvant therapy for postmenopausal women with early stage breast cancer. Musculoskeletal adverse events (MS AE) have become increasingly recognized as an issue for many women receiving AIs resulting in decreased quality of life and, in some instances, discontinuation of the AI. MA.27 is an ongoing randomized clinical trial comparing anastrozole with exemestane that accrued 7576 patients (pts), including 6827 from North America, about 79% of whom provided DNA and consent for genetic testing. A review of withdrawals related to adverse events revealed that the most common cause was MS AE. The hypothesis for our study was that a genome-wide association (GWA) case control study would identify single nucleotide polymorphisms (SNPs) associated with MS AE.Methods: Cases were defined as pts with grade 3 or 4 MS AE (NCI CTCAE v3.0) or who went off treatment for any grade MS AE within the first 2 years and were matched to 2 controls based on treatment arm, length of follow-up, prior chemotherapy, age and prior celecoxib (at least 3 months prior to MS AE). The study was restricted to the 94% of pts self-identified as Caucasian. Genotyping was performed with the Illumina Human610-Quad. Genotype quality control prior to analysis required call rates ≥98%. Eigenstrat analyses were performed to control for population stratification.Results: The GWA study included 293 cases and 586 controls with 6 pts (0.7%) (4 cases, 2 controls) excluded for call rates &lt;98%. 551,358 SNPs were used in the analyses after removing 29,439 for a minor allele frequency &lt;1% and 82 with Hardy Weinberg Equilibrium p-values &lt;10-6. Eigenvectors did not impact the results when used as covariates. Four SNPs with the lowest p-values (3.3-8.2 x 10-6, Armitage test), 3 in high LD (r2&gt;.8) on chromosome (ch) 14 (rs7158782, rs7159713, rs2369049) and 1 on the X ch (rs6637820) were identified. The gene closest (4000-7000 bp) to the 3 SNPs on ch 14 was T-Cell Leukemia 1A (TCL1A), encoding a protein known to augment AKT kinase activity. Expression array data from lymphoblastoid cell lines from 300 subjects of 3 ethnicities revealed that all 3 SNPs were associated with decreased TCL1A expression after adjusting for ethnicity using 2 probe sets (p=0.006 to 0.03). Electromobility shift assay (EMSA) showed that 2 of these SNPs (rs7158782, rs7159713) displayed a shift and the former SNP showed substantially less binding for variant than wild type. The gene closest to the X ch SNP was Immunoglobulin Superfamily 1 (IGSF1), but EMSA did not show a shift for this SNP.Conclusions: MS AEs represent a major impediment to optimal use of AIs in women with breast cancer and this GWA study identified SNPs on ch 14 that provide a focus for further research to identify pts at risk for, and means to ameliorate, this adverse event.(Supported in part by NIH grants U01GM61388, U01GM63173, P50CA116201 and U10CA77202) Citation Information: Cancer Res 2009;69(24 Suppl):Abstract nr 15.