Maintaining up-to-date clinical practice guidelines (CPGs) is essential for evidence-based medicine, but rapid evidence growth and complex expert consensus formation limit timely updates. We developed GUIDE, a multi-agent copilot system for human-AI collaborative guideline generation and updating. GUIDE delegates labor-intensive tasks to specialized AI agents while preserving expert judgment and final arbitration. In 12 NICE ophthalmology datasets, GUIDE achieved 88.85% literature recall, 86.06% alignment in certainty-of-evidence ratings, and 91.67% recommendation agreement, requiring 162.17 minutes per dataset on average (28.68% human intervention). Performance remained stable across 20 additional datasets from cardiology, endocrinology, gastroenterology, and respiratory medicine. Ablation analysis showed that removing expert intervention reduced performance in judgment-intensive steps such as Risk-of-Bias assessment. In six post-cutoff datasets, GUIDE maintained robust end-to-end performance on unseen evidence. Prospective real-world validation showed that GUIDE reduced expert workload and produced more clinically acceptable recommendations than the original decade-old guideline (mean score, 3.73 vs. 1.53; agreement, 66.7% vs. 13.3%), with outcomes comparable to human-only updating (P = 0.588) but substantially less time and effort. GUIDE offers a practical framework for timely, trustworthy, expert-centric guideline development and validates computationally augmented consensus.

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