Purpose Studying the plasma proteome of control versus constitutionally thin (CT) individuals, exposed to overfeeding, may give insights into weight-gain management, providing relevant information to the clinical entity of weight-gain resistant CT, and discovering new markers for the condition. Experimental Design Untargeted protein relative quantification of 63 CT and normal-weight individuals was obtained in blood plasma at baseline, during and after an overfeeding challenge using mass spectrometry-based proteomics. Results The plasma proteome of CT subjects presented limited specificity with respect to controls at baseline. Yet, CT showed lower levels of inflammatory C-reactive protein and larger levels of protective insulin-like growth factor-binding protein 2. Differences were more marked during and after overfeeding. CT plasma proteome showed larger magnitude and significance in response, suggesting enhanced "resilience" and more rapid adaptation to changes. Four proteins behaved similarly between CT and controls, while five were regulated in opposite fashion. Ten proteins were differential during overfeeding in CT only (including increased fatty acid-binding protein and glyceraldehyde-3-phosphate dehydrogenase, and decreased apolipoprotein C-II and transferrin receptor protein 1). Conclusions and Clinical Relevance This first proteomic profiling of a CT cohort reveals different plasma proteomes between CT subjects and controls in a longitudinal clinical trial. Our molecular observations further support that the resistance to weight gain in CT subjects appears predominantly biological. ClinicalTrials.gov Identifier: NCT02004821