BACKGROUND: Anlotinib, a small-molecule tyrosine kinase inhibitor (TKI), suppresses angiogenesis and tumor progression. As the mechanisms underlying the resistance to epidermal growth factor receptor (EGFR)-TKIs are complex and diverse, further exploration of new treatment strategies is necessary. Combination therapy with EGFR-TKIs and anlotinib targets multiple signaling pathways, enhancing efficacy in patients with EGFR-positive non-small cell lung cancer (NSCLC). This study evaluated the efficacy and safety of anlotinib with EGFR-TKIs in patients with NSCLC who developed resistance to postoperative adjuvant therapy. METHODS: From January 2020 to December 2023, 48 patients at the Department of Thoracic Surgery, the First Affiliated Hospital of Zhejiang University, who developed resistance to adjuvant therapy were included in this retrospective study. All patients received anlotinib (10-12 mg, po, d1-14, q3w) alongside their original EGFR-TKI regimen. The primary endpoint was progression-free survival (PFS), while secondary endpoints included 6- and 12-month PFS rates, overall survival (OS), and safety. PFS was defined as the time from the initiation of anlotinib plus EGFR-TKI to disease progression or death, and OS was defined as the time from the start of anlotinib plus EGFR-TKI to death from any cause. RESULTS: Among the 48 patients, 23 previously received first- or second-generation EGFR-TKIs, and 25 received third-generation EGFR-TKIs. As of March 25, 2024, the median follow-up duration was 33.3 months [95% confidence interval (CI): 23.2-43.3]. The median PFS was 9.5 months (95% CI: 4.8-14.3), and 6- and 12-month PFS rates were 70.8% and 47.9%, respectively. For patients previously treated with first-/second- and third-generation EGFR-TKIs, the median PFS was 10.3 months (95% CI: 6.1-14.4) and 7.7 months (95% CI: 4.8-10.6), with a 6-month PFS rate of 69.6% and 72.0%, respectively, and a 12-month PFS rate of 47.8% and 48.0%, respectively. The median OS was 31.0 months [95% CI: not reached (NR)-NR], with 6-month and 12-month rates of 91.7% and 85.4%, respectively. For patients previously treated with first-/second- and third-generation EGFR-TKIs, the median OS was NR and 20.3 months (95% CI: 10.7-30.0), respectively; meanwhile, the OS rates were 95.7% and 88.0% at 6 months, and 91.3% and 80.0% at 12 months, respectively. The incidence rates of any grade and grade ≥3 treatment-related adverse events (TRAEs) were 75.0% (36/48) and 10.4% (5/48), respectively. The most common TRAEs included hypertension (17/48, 35.4%), proteinuria (15/48, 31.3%), rash (11/48, 22.9%), fatigue (5/48, 10.4%), and diarrhea (4/48, 8.3%), and no new safety events were observed. Dose reduction and discontinuation of anlotinib were reported in four (8.3%) and five (10.4%) patients previously treated with first-/second- and third-generation EGFR-TKIs, respectively. CONCLUSIONS: Patients with NSCLC who developed resistance to postoperative EGFR-TKIs demonstrated promising efficacy and manageable safety, extending the treatment window and survival opportunities.