Background: The existence of metabolic adaptation, following weight loss, remains a controversial issue. To our knowledge, no study has evaluated the role of energy balance (EB) in modulating metabolic adaptation. Objectives: The aim of this study was to determine if metabolic adaptation, at the level of resting metabolic rate (RMR), is modulated by participants' EB status. A secondary aim was to investigate if metabolic adaptation was associated with weight regain. Methods: Seventy-one individuals with obesity (BMI: 34.6 ± 3.4 kg/m2; age: 45.4 ± 8.2 y; 33 men) enrolled in a 1000-kcal/d diet for 8 wk, followed by 4 wk of weight stabilization and a 9-mo weight loss maintenance program. Body weight/composition and RMR were measured at baseline, week 9 (W9), week 13 (W13), and 1 y (1Y). Metabolic adaptation was defined as a significantly different (lower or higher) measured compared with predicted RMR. Results: Participants lost on average 14 kg by W9, followed by weight stabilization at W13, and regained 29% of their initial weight loss at 1Y. Metabolic adaptation was found at W9 (-92 ± 110 kcal/d, P < 0.001) and W13 (-38 ± 124 kcal/d, P = 0.011) but was not correlated with weight regain. A significant reduction in metabolic adaptation was seen between W9 and W13 (-53 ± 101 kcal/d, P < 0.001). In a subset of participants who gained weight between W9 and W13 (n = 33), no metabolic adaptation was seen at W13 (-26.8 ± 121.5 kcal/d, P = 0.214). In a subset of participants with data at all time points (n = 45), metabolic adaptation was present at W9 and W13 (-107 ± 102 kcal/d, P < 0.001 and -49 ± 128 kcal/d, P = 0.013) but not at 1Y (-7 ± 129, P = 0.701). Conclusion: After weight loss, metabolic adaptation at the level of RMR is dependent on the EB status of the participants, being reduced to half after a period of weight stabilization. Moreover, metabolic adaptation does not predict weight regain at 1Y follow-up. These trials were registered at clinicaltrials.gov as NCT02944253 and NCT03287726.