Background: Alpelisib (A), a PI3K inhibitor, is FDA-approved with fulvestrant for patients with PIK3CA-mutated HR+/HER2- MBC. Everolimus (E), an mTOR inhibitor, is FDA-approved with endocrine therapy (ET) regardless of PI3K/AKT/mTOR pathway mutation status. The efficacy and safety of sequential treatment with these pathway inhibitors is not well described. Methods: This single-center retrospective cohort study identified patients with HR+/HER2- MBC treated with both A and E sequentially, each in combination with ET, in either order from 2012-2023. We collected information on patient demographics, clinical and pathologic characteristics, treatment history, treatment adverse effects, and dates of disease progression (defined as radiographic or clinical progression leading to treatment change) from medical records. Patients treated first with E then A were categorized as Group 1 and those first with A then E as Group 2. Rates of prior CDK4/6i use were compared between groups with the chi-square test; differences in prior lines of therapy were assessed with the Wilcoxon rank sum test. Median PFS1 (PFS on 1^st pathway inhibitor) and PFS2 (PFS on 2^nd pathway inhibitor) were computed using the Kaplan-Meier method and the impact of prior treatment on PFS2 between groups was compared using Cox proportional hazards analysis. Results: 115 patients with HR+/HER2- MBC were included in this study. There were 62 (54%) patients in Group 1 and 53 (46%) patients in Group 2. Median prior lines of therapy in the metastatic setting for Group 1 was 4 (range 1-13) and for Group 2 was 3 (range 1-8; p=0.08). Median number of intervening therapies between E and A in Group 1 was 2 (range 0-8) and between A and E in Group 2 was 0 (range 0-4; p<0.001). 34/62 (55%) patients in Group 1 and 45/53 (85%) in Group 2 received prior CDK4/6i (p<0.001). In Group 1, 49/62 (79%) of patients discontinued E for progression and 13 (21%) discontinued for toxicity. Median PFS1 was 9.2 months (95%CI 5.5-13.0) and PFS2 was 5.0 months (95%CI 4.6-7.9). In Group 2, 34/53 (64%) of patients discontinued A for progression and 19 (36%) discontinued for toxicity. Median PFS1 was 9.3 months (95%CI 7.0-14.0) and PFS2 was 3.5 months (95%CI 3.0-4.1). After adjusting for prior CDK4/6i use and prior lines of therapy (total and intervening), PFS2 was not significantly different between the groups. Genomic information will be included at the time of presentation. Conclusions: This is the largest retrospective study to-date of patients treated sequentially with PI3K/AKT/mTOR pathway inhibitors. PFS was longer with the 1st pathway inhibitor than with the 2nd regardless of sequence. This has implications for optimal integration of capivasertib, an AKT inhibitor, into HR+ MBC treatment. Prospective studies testing optimal sequencing are needed.