The 6-minute walk test (6MWT) is a reliable and valid tool for determining an individual’s functional capacity, and has been used to predict summit success. The primary aim of the study was to evaluate whether a 6MWT in normobaric hypoxia could predict physiological responses and exercise performance at altitude. The secondary aim was to determine construct validity of the 6MWT for monitoring acclimatization to 3400 m (Cuzco, Peru).Twenty-nine participants performed six 6MWTs in four conditions: normoxic outdoor (NO), normoxic treadmill (NT), and hypoxic treadmill (HT) were each performed once; and hypoxic outdoor (HO) was performed three times, at 42 hours (HO1), 138 hours (HO2), and 210 hours (HO3) after arrival at Cuzco.One-way analysis of variance revealed no difference (P > .05) between NO and HO1 for 6MWT distance. HT and HO protocols were comparable for the measurement of delta heart rate (HR) and post-test peripheral oxygen saturation (%Spo2; P > .05). Acclimatization was evidenced by reductions (P < .05) in resting HR and respiratory rate (RR) between HO1, HO2, and HO3, and preservation of Spo2 between HO1 and HO2. Postexercise HR and RR were not different (P > .05) with acclimatization. The duration to ascend to 4215 m on a trek was moderately correlated (P < .05) to HR during the trek and the 6MWT distance during HT; no other physiological markers predicted performance.The 6MWT is a simple, time-efficient tool for predicting physiological responses to simulated and actual altitude, which are comparable. The 6MWT is effective at monitoring elements of acclimatization to moderate altitude.