Background: Everolimus (EVE) plus exemestane (EXE) more than doubled progression-free survival (PFS) while maintaining quality of life vs EXE alone in postmenopausal women with hormone-receptor positive (HR⁺), HER2-negative (HER2^-) advanced breast cancer (BOLERO-2 phase III; NCT00863655). PFS benefit was seen in all clinically defined subgroups. We evaluated genetic variations of a broad panel of cancer-related genes and explored their correlations with EVE benefit. Methods: Exon sequence and gene copy number variations were analyzed in 182 cancer-related genes by next-generation sequencing (NGS). Correlations with PFS were evaluated using both univariate and multivariate Cox models. Results: NGS data (>250x coverage) were successfully generated from archival tumor specimens from 227 patients (NGS population, 157 and 70 in EVE+EXE and EXE arms, respectively) whose baseline characteristics and clinical outcome were comparable with the trial population (PFS HR = 0.40 and 0.45, respectively). The treatment benefit of EVE+EXE over EXE is maintained in the subgroups defined by each of the nine genes with a mutation rate >10% (eg, PIK3CA, FGFR1, and CCND1), or when less frequently mutated genes (eg, PTEN, AKT1) were included in their respective pathways. Patients with no or only 1 genetic alteration in PI3K or FGFR pathways, or CCND1, had a greater treatment effect from EVE (HR = 0.27, 95% CI 0.18-0.41, adjusted by covariates, in 76% of the NGS population), indicating the value of these pathways for predicting sensitivity/resistance to EVE in this setting. Conclusions: This is the first global registration trial in which efficacy-predictive biomarkers were explored by correlating broad genetic variations with clinical efficacy. It demonstrated the feasibility of applying large-scale NGS and subsequent correlative analysis to such trials. The observations suggest that a large subgroup of patients (76%), defined by minimal genetic variations in the PI3K or FGFR pathways, or CCND1, derives the most benefit from EVE therapy (HR = 0.27 vs 0.40 for the full NGS population). These exploratory results and their implication in understanding the interplay of multiple pathways in tumor cells and testing new hypotheses for targeted combination therapies in HR⁺/HER2^- BC will be further investigated. Clinical trial information: NCT00863655.