Higher risk of recurrence in early-stage breast cancer patients with increased levels of ribosomal protein S6

Abstract

PI3K/AKT/mTOR pathway is implicated in breast cancer progression and recurrence. The identification of PIK3CA and AKT1 mutations and loss of PTEN serve as selection criterion for targeted therapies involving selective inhibitors. However, they do not consistently align with pathway activation, and high-cost determinations limit their routine application. PI3K-downstream epigenetic regulatory mechanisms broaden the alterations that amplify pathway activity and, consequently, sensitivity to selective inhibitors. In this retrospective observational study, conducted within a cohort of early-stage breast cancer patients, we determined phosphorylated ribosomal protein S6 (pS6) at Ser240/244 by immunohistochemistry as an indicator of PI3K pathway activation. Log-Rank test and Cox proportional hazards regression were used to analyze the clinical relevance of pS6, alone and together with clinicopathological variables, regarding recurrence-free survival. ROC curves and the area under the curves were used to evaluate the calibration and discrimination properties of uni- and multivariate models. Our results show that a high percentage of pS6 positive tumor cells was associated with an unfavorable prognosis in a cohort of 129 hormone receptor positive/HER2 negative breast cancer patients (Hazard Ratio = 5.92; Log-Rank p = 9.5e-08; median follow-up = 53 months). When assessed in combination with lymph node status, the predictive capacity was higher compared to both univariate models individually. In conclusion, pS6 could represent a novel independent marker for predicting recurrence risk in luminal breast cancer.

Keywords: Breast cancer; Immunohistochemistry analysis; PI3K/AKT/mTOR pathway; Predictive markers; Prognostic markers; Tumor relapse.

Fig. 1

Analysis of pS6 as a risk predictor of recurrence in luminal breast cancer patients. (a) Immunohistochemistry images of tissues from four different patients, two representatives of low and two representatives of high pS6 level, determined by a cutoff of 42.5% pS6 positive tumor cells. Cytoplasmic perinuclear staining can be observed in all the samples. (b) Kaplan-Meier curves for recurrence-free survival (RFS) with low and high pS6 level. Log-Rank p, HR: hazard ratio (95% confidence interval), and Cox p are specified. Statistical significance was set at p < 0.05.

Fig. 2

Multivariate analysis of pS6 and clinicopathological parameters as prognostic factors for RFS. Forest plots show the results obtained with pS6 and co-variables including (a) age (with less than 50 years as reference), (b) tumor size (with less than 2 cm as reference), (c) lymph node (LN) status (with negative LN as reference), and (d) stage (with stage I as reference). HR: hazard ratio; 95% CI: 95% confidence interval. Statistical significance was set at *p < 0.05.

Fig. 3

Analysis of pS6 and lymph node (LN) status as a combined risk predictor of recurrence in luminal breast cancer patients. (a) Kaplan-Meier curves for RFS with low and high pS6 jointly with negative and positive lymph nodes (LN- and LN+, respectively). Log-Rank p, HR: hazard ratio (95% confidence interval) and Cox p are specified. (b) ROC curves for LN and pS6, individually and combined. AUC: area under the curve. p-value using DeLong’s test. Statistical significance was set at p < 0.05.