PIK3CA mutations associated with gene signature of low mTORC1 signaling and better outcomes in estrogen receptor-positive breast cancer

Abstract

PIK3CA mutations are reported to be present in approximately 25% of breast cancer (BC), particularly the estrogen receptor-positive (ER+) and HER2-overexpressing (HER2+) subtypes, making them one of the most common genetic aberrations in BC. In experimental models, these mutations have been shown to activate AKT and induce oncogenic transformation, and hence these lesions have been hypothesized to render tumors highly sensitive to therapeutic PI3K/mTOR inhibition. By analyzing gene expression and protein data from nearly 1,800 human BCs, we report that a PIK3CA mutation-associated gene signature (PIK3CA-GS) derived from exon 20 (kinase domain) mutations was able to predict PIK3CA mutation status in two independent datasets, strongly suggesting a characteristic set of gene expression-induced changes. However, in ER+/HER2- BC despite pathway activation, PIK3CA mutations were associated with a phenotype of relatively low mTORC1 signaling and a good prognosis with tamoxifen monotherapy. The relationship between clinical outcome and the PIK3CA-GS was also assessed. Although the PIK3CA-GS was not associated with prognosis in ER- and HER2+ BC, it could identify better clinical outcomes in ER+/HER2- disease. In ER+ BC cell lines, PIK3CA mutations were also associated with sensitivity to tamoxifen. These findings could have important implications for the treatment of PIK3CA-mutant BCs and the development of PI3K/mTOR inhibitors.

Fig. 1.

The PIK3CA-GS has better prognostic significance than mutation status alone. Kaplan-Meier curves according to PIK3CA mutation status by sequencing or by the PIK3CA-GS in (A) all patients in whom mutation status was assessed, (B) PIK3CA mt compared with prognostic subgroups of ER+ BC, (C) patients classified as PIK3CA WT by sequencing, (D) patients classified as PIK3CA mt by sequencing, and (E) ER+ BC classified as good prognosis (black: ER+/HER2−/L) and poor prognosis (green/blue: ER+/HER2−/H) molecular subtypes and prognosis according to PIK3CA-GS expression.

Fig. 2.

Box-plots comparing expression of (A) p-AKT and (B) p-S6 levels in PIK3CA mt and WT samples from the TAM dataset. Examples of PIK3CA mt and WT samples (C and E, respectively): pAKT was present in both the nuclei and cytoplasm, and pS6 staining was observed mainly in the cytoplasm from positive tissue specimens (D and F, magnification 400×, high-power field).

Fig. 3.

PIK3CA mt samples and mt-like samples have show decreased expression of downstream PI3K/AKT/mTOR proteins. Heat map represents unsupervised hierarchical clustering of BCs and corresponding protein data from the MDACC cohort. Patients are represented horizontally: PIK3CA mt status by sequencing (seq), according to gene signature (GS), ER and HER2 (FISH) status are indicated by filled (positive) and white (negative) boxes and Exon 9 and 20 mutations are gray and black boxes, respectively; Clustering was performed by using Pearson correlation metric and centroid linkage. Proteins and phosphorylated proteins are indicated vertically. Red represents over-expression, green represents underexpression.

Fig. 4.

Box plots showing relative expression of the PIK3CA mt (A) and PTEN-loss (B) gene signatures according to molecular subtype and PI3K pathway aberration from the Saal et al. (19) dataset. PIK3CA-sequenced mutants (HD+KD) have less expression of the PTEN-loss gene signature compared with BCs that are PTEN-deficient by IHC (P = 0.001) or HER2-amplified (P = 0.01), but still higher expression compared with the ER+/HER2−/L (Luminal-A) tumors (P = 0.002).

Fig. 5.

In a panel of ER+ BC cell lines, IC₅₀ was assessed to rapamycin and tamoxifen according to PIK3CA status. PIK3CA mutations (MCF7, T47D, KPL1) predict sensitivity to tamoxifen (A), but not rapamycin (B). Cell lines are ranked according to increasing PIK3CA-GS (C) The PIK3CA-GS could identify ER+ PIK3CA mt lines, but in the reverse direction to human BC (AUC, 0.9; P = 0.008; Fig. S1C), i.e., cell lines are associated with relatively high mTORC1 output. The rapamcyin-resistant ZR751 line was associated with the highest PIK3CA-GS value; black bars/circles represent PIK3CA mt cell lines in A, B, and C; error bars indicate 95% CI; each bar represents the mean IC₅₀ from triplicate plates.