Background: Vitamin D deficiency is prevalent among breast cancer patients and has been associated with poorer outcomes. Preclinical studies suggest that vitamin D may enhance the cytotoxic effects of chemotherapy through various mechanisms. This randomized controlled trial evaluated the effect of high-dose vitamin D supplementation on response to neoadjuvant chemotherapy in patients with locally advanced breast cancer. Methods: In this double-blind, placebo-controlled trial, 240 patients with stage II-III breast cancer were randomized to receive either high-dose vitamin D3 (50,000 IU weekly) or placebo during neoadjuvant chemotherapy. The primary endpoint was pathological complete response (pCR) rate. Secondary endpoints included clinical response, breast conservation rate, changes in vitamin D levels, toxicity, quality of life, and biomarker analyses. Results: Of 240 enrolled patients, 235 completed the study (118 in vitamin D group, 117 in placebo group). Baseline characteristics were well-balanced between groups, with 56% of patients vitamin D deficient (<20 ng/mL). The vitamin D group achieved significantly higher pCR rates compared to placebo (36.4% vs. 22.2%; relative risk 1.64, 95% CI: 1.08-2.48; p=0.016). Subgroup analyses revealed greater benefit in hormone receptor-negative tumors (61.0% vs. 38.1%; p=0.036), triple-negative subtype (50.0% vs. 24.1%; p=0.048), and baseline vitamin D deficiency (44.1% vs. 20.3%; p=0.004). Breast conservation rates were higher in the vitamin D group (54.2% vs. 40.2%; p=0.034). Mild hypercalcemia was more common with vitamin D (6.7% vs. 1.7%), but no grade ≥3 hypercalcemia occurred. Quality of life scores were significantly better in the vitamin D group before surgery (65.9 vs. 59.8; p=0.007). Multivariate analysis confirmed vitamin D supplementation as an independent predictor of pCR (adjusted odds ratio 2.31, 95% CI: 1.25-4.27; p=0.007). Conclusion: High-dose vitamin D supplementation during neoadjuvant chemotherapy significantly improved pathological complete response rates in breast cancer, particularly in hormone receptor-negative tumors and patients with baseline vitamin D deficiency. This simple, low-cost intervention may represent an important addition to current neoadjuvant treatment strategies.