Objective: The main objective was to assess repeatability and learning effect of the 6-min walk test (6MWT) in a cohort of preoperative cancer patients referred to a prehabilitation program. As a secondary objective, we aimed to identify determinants of improvement in the second test. Materials and methods: Secondary analysis from a large prospective study on the implementation of a multimodal prehabilitation program in a real-life scenario. Eligible patients were assessed at baseline before starting the prehabilitation program. The 6MWT was conducted according to the American Thoracic Society (ATS)/European Respiratory Society (ERS) guidelines with two tests being performed under identical conditions separated by 30 min. The distance covered (in meters) and the physiological responses (heart rate, oxygen saturation, fatigue, and dyspnea) to each test were recorded and compared. Results: A total of 170 patients (60.9%) were analyzed. Repeatability of the distance covered with the 6MWT was excellent (ICC = 0.98; 95% CI: 0.92–0.99), but a mean increase of + 19.5 m (95% CI: 15.6–23.5 m; p = <.001) in the second test was found, showing a learning effect with limits of agreement between − 31.3 and 70.4 m. Coefficient of variation was 4%. No clinical factor was found to be associated with an improvement in the second test. Conclusions: The 6MWT showed excellent repeatability in preoperative cancer patients, but a significant learning effect is present. No associated factors with a clinically meaningful improvement in the second test were identified. In light of these findings, two attempts of the 6MWT should be encouraged in this population.