Akt phosphorylation at Ser473 predicts benefit of paclitaxel chemotherapy in node-positive breast cancer

Abstract

Purpose: We tested the hypothesis that Akt-Ser473 phosphorylation (pAkt) predicts benefit from the sequential addition of paclitaxel to adjuvant doxorubicin plus cyclophosphamide (AC) chemotherapy in patients with node-positive breast cancer participating in the National Surgical Adjuvant Breast and Bowel Project (NSABP) B-28 trial.

Patients and methods: Primary tumors from the NSABP B-28 trial tissue microarray were available from 1,581 of 3,060 patients who were randomly assigned to receive either four cycles of AC alone or followed by four cycles of paclitaxel. Immunohistochemistry and quantitative analysis of pAkt were performed at the National Cancer Institute blinded to clinical outcome. Association between pAkt and clinical outcome was assessed using multivariate Cox modeling adjusting for age, tumor size, number of positive nodes, tumor grade, estrogen receptor status, and human epidermal growth factor receptor 2 status.

Results: With a median follow-up of 9.1 years, there were no differences in disease-free survival (adjusted hazard ratio [HR], 1.02; P = .81) or overall survival (HR, 0.97; P = .80) with and without receiving paclitaxel among 975 patients with pAkt-negative tumors. In 606 patients with pAkt-positive tumors, the sequential addition of paclitaxel resulted in a 26% improvement in disease-free survival (HR, 0.74; P = .02) or a 20% improvement in overall survival (HR, 0.80; P = .17).

Conclusion: pAkt significantly predicts disease-free benefit from the sequential addition of paclitaxel to AC chemotherapy in patients with node-positive breast cancer. Patients with pAkt-negative breast tumors do not appear to benefit from the addition of paclitaxel.

Fig 1.

Disease-free benefits in patients treated with or without paclitaxel according to Akt-Ser473 phosphorylation (pAkt) status. Disease-free survival according to (A) negative pAkt or (B) positive pAkt was analyzed with the use of the Kaplan-Meier method. HR, hazard ratio.

Fig 2.

Adjusted disease-free survival (DFS) among patients treated with or without paclitaxel according to estrogen receptor (ER) and Akt-Ser473 phosphorylation (pAkt) status. DFS was analyzed with the use of the Kaplan-Meier method by (A, C) negative pAkt or (B, D) positive pAkt, or by (A, B) positive ER or (C, D) negative ER expression.

Fig 3.

Adjusted disease-free survival (DFS) among patients treated with or without paclitaxel according to human epidermal growth factor receptor 2 (HER2) and Akt-Ser473 phosphorylation (pAkt) status. DFS was analyzed with the use of the Kaplan-Meier method by (A, C) negative pAkt or (B, D) positive pAkt, or by (A, B) positive HER2 or (C, D) negative HER2 expression.

Fig A1.

Recommendations for Tumor Marker Prognostic Studies (REMARK) diagram on the distribution of patients with Akt-Ser473 phosphorylation (pAkt) measurement in the National Surgical Adjuvant Breast and Bowel Project (NSABP) B-28 trial. AC, doxorubicin plus cyclophosphamide; AC-P, AC followed by paclitaxel; TMA, tissue microarray.

Fig A2.

Akt-Ser473 phosphorylation (pAkt) immunohistochemistry in MDA-MB-468 breast cancer cells. Antigen retrieval was performed in high pH (pH 10) (A) antigen retrieval buffer versus (B) low pH (pH6) buffer. The optimized method showed good reproducibility with a 5.7% coefficient of variation (three independent staining in MDA-MB-468 cells); ×600.

Fig A3.

Representative Akt-Ser473 phosphorylation (pAkt) staining core by core comparison from the two sets of National Surgical Adjuvant Breast and Bowel Project (NSABP) B-28 tissue microarray. The NSABP B-28 tissue microarray set 1 was used for data analysis and data presentation in this study. ×200; inset magnifications ×40.

Fig A4.

Distribution of 1,581 patients by (A) Akt-Ser473 phosphorylation (pAkt) levels and (B) pAkt staining in representative primary breast tumors from the National Surgical Adjuvant Breast and Bowel Project (NSABP) B-28 trial. The staining index (SI) for pAkt ranged from 0 to 62.8. The arrow (SI of > 2) indicates the positive cutoff, by which 38% of breast tumors are pAkt positive and 62% are pAkt negative.

Fig A5.

Overall survival (OS) among patients treated with or without paclitaxel according to Akt-Ser473 phosphorylation (pAkt) status. OS according to (A) negative pAkt or (B) positive pAkt was analyzed with the use of the Kaplan-Meier method. HR, hazard ratio.

Fig A6.

Disease-free survival (DFS) among patients treated with or without paclitaxel according to Akt-Ser473 phosphorylation (pAkt), human epidermal growth factor receptor 2 (HER2), or estrogen receptor (ER) status. DFS according to (A) negative HER2 and positive ER, (B) positive pAkt, negative HER2, and positive ER, or (C) negative pAkt, negative HER2 and positive ER was analyzed with the use of the Kaplan-Meier method. HR, hazard ratio.