A population-based study of tumor gene expression and risk of breast cancer death among lymph node-negative patients

Abstract

Introduction: The Oncotype DX assay was recently reported to predict risk for distant recurrence among a clinical trial population of tamoxifen-treated patients with lymph node-negative, estrogen receptor (ER)-positive breast cancer. To confirm and extend these findings, we evaluated the performance of this 21-gene assay among node-negative patients from a community hospital setting.

Methods: A case-control study was conducted among 4,964 Kaiser Permanente patients diagnosed with node-negative invasive breast cancer from 1985 to 1994 and not treated with adjuvant chemotherapy. Cases (n = 220) were patients who died from breast cancer. Controls (n = 570) were breast cancer patients who were individually matched to cases with respect to age, race, adjuvant tamoxifen, medical facility and diagnosis year, and were alive at the date of death of their matched case. Using an RT-PCR assay, archived tumor tissues were analyzed for expression levels of 16 cancer-related and five reference genes, and a summary risk score (the Recurrence Score) was calculated for each patient. Conditional logistic regression methods were used to estimate the association between risk of breast cancer death and Recurrence Score.

Results: After adjusting for tumor size and grade, the Recurrence Score was associated with risk of breast cancer death in ER-positive, tamoxifen-treated and -untreated patients (P = 0.003 and P = 0.03, respectively). At 10 years, the risks for breast cancer death in ER-positive, tamoxifen-treated patients were 2.8% (95% confidence interval [CI] 1.7-3.9%), 10.7% (95% CI 6.3-14.9%), and 15.5% (95% CI 7.6-22.8%) for those in the low, intermediate and high risk Recurrence Score groups, respectively. They were 6.2% (95% CI 4.5-7.9%), 17.8% (95% CI 11.8-23.3%), and 19.9% (95% CI 14.2-25.2%) for ER-positive patients not treated with tamoxifen. In both the tamoxifen-treated and -untreated groups, approximately 50% of patients had low risk Recurrence Score values.

Conclusion: In this large, population-based study of lymph node-negative patients not treated with chemotherapy, the Recurrence Score was strongly associated with risk of breast cancer death among ER-positive, tamoxifen-treated and -untreated patients.

Figure 1

Twenty-one gene panel and calculation of recurrence score. (a) The final gene list (16 cancer-related and five reference genes) and summary score (Recurrence Score) algorithm for this assay were developed by analyzing the results of three independent preliminary breast cancer studies (for instance, training sets) with a total of 447 patients [11]. The Recurrence Score, on a scale from 0 to 100, is derived from the reference-normalized expression measurements in four steps. In the first step the expression for each gene is normalized relative to the expression of the five reference genes (b-actin, GAPDH, GUS, RPLPO, and TFRC). Reference-normalized expression measurements range from 0 to 15, where a 1-unit increase reflects approximately a twofold increase in RNA. (b) In the second step the HER2 Group Score, the ER Group Score, the Proliferation Group Score, and the Invasion Group Score are calculated from individual gene expression measurements. (c) In the third step the Recurrence Score unscaled (RS_U) is calculated using coefficients that were pre-defined based on regression analysis of gene expression and recurrence in the three training studies (Providence, Rush, and NSABP B-20 [12]). A plus sign indicates increased expression is associated with increased recurrence risk. A minus sign indicates that increased expression is associated with decreased recurrence risk.

Figure 2

Relative risks (RRs) for death associated with expression of single genes. Findings are stratified by tamoxifen treatment and ER status. The position of each symbol indicates the RR. The length of the horizontal line through the symbol indicates the 95% CI. The blue boxes indicate the RRs and 95% CIs for ER-positive patients treated with tamoxifen, the green pyramids indicate the RRs and 95% CIs for ER-positive patients not treated with tamoxifen, and the red downward pointing triangles indicate the RRs and 95% CIs for ER-negative patients not treated with tamoxifen. CI, confidence interval; ER, estrogen receptor; RR, relative risk.

Figure 3

Relative risks (RRs) of death associated with gene group scores used in calculation of the Recurrence Score. Findings are stratified by tamoxifen treatment and ER status. The position of each symbol indicates the RR. The length of the horizontal line through the symbol indicates the 95% CI. The blue boxes indicate the RRs and 95% CIs for ER-positive patients treated with tamoxifen, the green pyramids indicate the RRs and 95% CIs for ER-positive patients not treated with tamoxifen, and the red downward pointing triangles indicate the RRs and 95% CIs for ER-negative patients not treated with tamoxifen. CI, confidence interval; ER, estrogen receptor; RR, relative risk.