Abstract Background: The absolute improvement of invasive disease-free survival with intensive adjuvant CDK4/6 inhibitor (CDK4/6i) therapy ranges from 4.9% to 7.6%. Given the associated adverse events (AEs) and economic implications, identifying patients who are most likely to benefit from CDK4/6i can help avoid under- or overtreatment.Short-term adaptive neoadjuvant endocrine therapy (NET) may facilitate the selection of patients who could benefit from intensified therapy. This study aims to utilize adaptive NET to guide patient selection for combination endocrine therapy with CDK4/6i and to further explore molecular mechanisms underlying resistance to CDK4/6i. Methods: Postmenopausal patients with stage II-III HR+/HER2- breast cancer and a baseline Ki-67 index 10% were prospectively enrolled in a multicenter clinical trial (Clinical trial.gov Identifier: NCT05809024). All patients received 2 weeks of NET with aromatase inhibitors (AIs), followed by a second tumor biopsy. Based on post-treatment Ki-67 levels (cut-off: 10%), patients were stratified into two groups: (A) AI-responsive (Ki-67 ≤10%), who proceeded directly to surgery; and (B) AI-nonresponsive (Ki-67 10%), who received an additional cycle of AI combined with the CDK4/6i dalpiciclib prior to surgery. Tumor samples were collected at three time points—prior to NET, after AI therapy, and after AI plus CDK4/6i—for single-cell transcriptome sequencing and next-generation sequencing (NGS). Results: Between February 2023 and 2025, 120 patients were enrolled. After 2 weeks of AI therapy, 68.3% (82/120) achieved Ki-67 ≤10% (AI-responsive), while 31.7% (38/120) remained with Ki-67 10% (AI-nonresponsive). Among the AI-nonresponsive patients, 50.0% (19/38) remained Ki-6710% following one cycle of AI plus CDK4/6i (CDK4/6i-nonresponsive), while the remaining 50.0% achieved Ki-67 ≤10% (CDK4/6i-responsive). No new drug-related AEs were observed. Single-cell sequencing was conducted in 23 patients: AI-responsive (n=7), AI-nonresponsive (n=12), CDK4/6i-responsive (n=2), and CDK4/6i-nonresponsive (n=2). Seven distinct cell clusters were identified: B cells, CD8+T cells, endothelial cells, epithelial cells, fibroblasts, macrophages, and mast cells. Mast cells was significantly more abundant in the AI-nonresponsive group, and the malignant epithelial C2 cell cluster was notably enriched in the CDK4/6i-nonresponsive group. Bulk transcriptomic sequencing of 52 tumor samples revealed upregulation of FUT6, APOF, and SERPINA6 in the CDK4/6i-nonresponsive group, with FUT6 showing the most significant differential expression. NGS was performed on 61 patients. PIK3CA mutations were detected in 54.5% of patients before AI therapy, rising to 60.6% after AI therapy (72.7% in the AI-nonresponsive group; 53.8% in the AI-responsive group), and declining to 42.8% following AI plus CDK4/6i treatment (62.5% in the CDK4/6i-nonresponsive group; 16.6% in the CDK4/6i-responsive group). TP53 mutations were observed in 36.3% of patients before AI therapy, 42.6% after AI therapy, and 57.1% following AI plus CDK4/6i treatment (75.0% in the CDK4/6i-nonresponsive group; 33.3% in the CDK4/6i-responsive group). Conclusions: In patients initially nonresponsive to AI therapy, the addition of CDK4/6i significantly reduced Ki-67 levels, indicating that these individuals may benefit from intensified CDK4/6i-based therapy. The malignant epithelial C2 cell cluster and FUT6 were identified as potential therapeutic targets associated with CDK4/6i resistance. Moreover, persistent or emerging PIK3CA and TP53 mutations during adaptive therapy may serve as molecular indicators of resistance to sequential AI and CDK4/6i treatments. Citation Format: Y. Wang, Z. Bi, P. Qiu, J. Zhang, C. Zheng, Y. Liu, B. Cong, X. Sun, P. Chen, C. Wang. Ki-67-guided stratification and resistance mechanism analysis of CDK4/6 inhibitor therapy following adaptiveneoadjuvant endocrine treatment in HR+/HER2− breast cancer abstract. In: Proceedings of the San Antonio Breast Cancer Symposium 2025; 2025 Dec 9-12; San Antonio, TX. Philadelphia (PA): AACR; Clin Cancer Res 2026;32(4 Suppl):Abstract nr PS2-07-20.
Wang et al. (Tue,) studied this question.