Baseline Mutations and Up-Regulation of PI3K-AKT Pathway Serve as Potential Indicators of Lack of Response to Neoadjuvant Chemotherapy in Stage II/III Breast Cancer

Abstract

Background: Neoadjuvant chemotherapy (NAC) has been expanded to hormone receptor (HR) positive breast cancer (BC) patients with operable disease, to increase the likelihood of breast-conserving surgery. Genomic profiling at baseline would reveal NAC response relevant genomic features and signaling pathways, guiding clinical NAC utilization based on patients' genomic characteristics.

Methods: We prospectively studied stage II/III BC patients who were eligible for breast-conserving surgery. Patients received epirubicin and cyclophosphamide for 4 cycles, followed by another 4-cycle docetaxel, and human epidermal growth factor receptor (HER2) positive patients were additionally treated with herceptin when using docetaxel (EC-T(H)). NAC responses were evaluated as pathologic complete response (pCR) or non-pathologic complete response (non-pCR). Genomic features related to NAC responses were identified by profiling baseline tumor tissues sampled one day before NAC, using whole-exome sequencing. Differentially expressed genes and up-/down-regulated pathways were investigated by performing RNA-sequencing.

Results: A total of 25 stage II/III BC patients were enrolled, including 5 patients ultimately evaluated as pCR and 20 patients evaluated as non-pCR. PIK3CA (48%) and TP53 (40%) mutations were enriched in patients not achieving pCR. Mutated phosphatidylinositol-3-kinase-AKT (PI3K-AKT) pathway and homologous recombinational repair pathway were also more frequently observed in patients evaluated as non-pCR. Significant arm-level amplifications (8q24.23 and 17q12) and deletions (1p32.2, 4p14, 7q11.23, 10q21.3, 11q23.3, etc.) were identified among patients not achieving pCR, while patients achieving pCR displayed no significant copy number alterations. Significantly up-regulated expression of PI3K-AKT pathway genes was also detected among patients failed to achieve pCR, compared to patients achieving pCR.

Conclusion: Compared to BC patients achieving pCR to NAC, aberrant activation of PI3K-AKT pathway genes were more frequently observed in patients not achieving pCR, consistent with the significant up-regulation of PI3K-AKT pathway gene expression in the non-pCR subgroup. Together, these findings indicate that upregulated PI3K-AKT pathway serves as a potential indicator of lack of response to NAC in stage II/III BC patients, and other effective therapeutic options are urgently needed for those resistant patients.

Keywords: PI3K-AKT pathway; PIK3CA mutations; neoadjuvant chemotherapy (NAC); pathologic complete response (pCR); stage II/III breast cancer (BC).

Figure 1

The flowchart of enrollment and analyzable patients. A total of 60 stage II/III breast cancer (BC) patients planned to receive breast-conserving surgery and neoadjuvant chemotherapy (NAC) of epirubicin, cyclophosphamide plus docetaxel with or without herceptin (EC-T(H)) were eligible for participation. We excluded 32 patients without available baseline tumor tissues and 3 patients without NAC response records, and the remaining 25 patients were enrolled in this study. All these 25 patients were analyzable for whole-exome sequencing (WES) data analyses, but 3 patients without enough tumor tissue for RNA sequencing (RNA-Seq) were excluded from RNA-Seq data analyses.

Figure 2

Genetic profile of baseline tumor tissue samples. The baseline tumor tissue samples of 25 patients were performed by WES. Altered genes of the phosphatidylinositol-3-kinase-AKT (PI3K-AKT) pathway and homologous recombinational repair (HRR) pathway and altered genes over 10% frequency were displayed. The most frequently altered genes were PIK3CA, TP53, DDX11, MUC4, QRICH2, and TTN. PIK3CA and TP53 mutations were enriched in patients not achieving pCR, and mutated PI3K-AKT pathway and HRR pathway were also more frequently observed in patients not achieving pCR.

Figure 3

Baseline arm-level copy number alterations (CNAs) and responses to neoadjuvant chemotherapy (NAC). (A) Significant amplifications were detected in the chromosomal region 8q24.23 and 17q12, in patients not achieving pCR, but no significant amplifications were observed in patients achieving pCR. Of note, ERBB2, a gene closely related to breast cancer (BC) was located in 17q12. (B) Multiple significant deletions were detected in patients not achieving pCR, and five regions with the highest q-values were 1p32.3, 4p14, 7q11.23, 10q21.3,11q23.3.

Figure 4

Differentially expressed genes (DEGs) and up-/down-regulated signaling pathways between pathologic complete response (pCR) patients or non-pathologic complete response (non-pCR) patients. A total of 61 DEGs were identified between the pCR subgroup and the non-pCR subgroup, including 34 genes expressed at higher levels and 27 genes expressed at lower levels, in the pCR subgroup, with absolute fold-change greater than 2. The phosphatidylinositol-3-kinase-AKT (PI3K-AKT) pathway was significantly down regulated among patients achieving pCR, with 4 lower expressed genes. Other down-regulated pathways included protein digestion and absorption, focal adhesion, AGE-RAGE, ECM-receptor, etc. On the other hand, higher expressed genes belonged to the pathways of metabolic pathways, neuroactive ligand-receptor interaction, glutamatergic synapse, glycerophospholipid metabolism, glycerolipid metabolism, nicotine addiction pathways, etc.