Background: Heart failure with preserved ejection fraction (HFpEF) is a heterogeneous syndrome that may emerge from overlapping systemic processes associated with comorbidities. We assessed whether unique clusters of circulating proteins are associated with specific clinical characteristics and functional status at baseline and follow-up in a well-phenotyped cohort of patients with HFpEF., Methods: We evaluated 368 proteins associated with cardiovascular disease and inflammation in prerandomization blood samples from 763 VITALITY-HFpEF (Vericiguat to Improve Physical Functioning in Daily Living Activities of Patients With HFpEF) participants who had a left ventricular ejection fraction ≥45% and a heart failure decompensation event within 6 months. Proteins were clustered, and their associations with clinical characteristics, baseline, and 24-week functional outcomes (Kansas City Cardiomyopathy Questionnaire Physical Limitation Score, 6-minute walk distance [6MWD], and Fried frailty phenotype) were estimated with linear regression. Elastic net regression was used to derive a proteomic summary composite to predict changes in 24-week functional outcomes., Results: Four unique protein clusters were identified, containing 24, 66, 197, and 81 proteins. At baseline, 2 protein clusters with the hub proteins caspase-3 and Dickkopf-related protein 1 were associated with increased frailty, whereas the cluster with tumor necrosis factor receptor 1 as a hub protein was associated with lower Kansas City Cardiomyopathy Questionnaire Physical Limitation Score and shorter 6MWD. By contrast, the cluster with protein C as a hub protein was associated with less frailty and longer a 6MWD. The 24-week increase in 6MWD was negatively correlated with the protein cluster with caspase-3; the protein C cluster was correlated with less frailty at 24 weeks. The baseline proteomic summary composite predicted observed changes in Kansas City Cardiomyopathy Questionnaire Physical Limitation Score and 6MWD at 24 weeks (r=0.42 and 0.30; P <0.001 for both)., Conclusions: Proteomics differentiate specific baseline functional traits associated with HFpEF and may facilitate phenotyping in a heterogeneous disease. These proteins also provide insights into the diverse pathophysiology of HFpEF and which patients may improve functional status during follow-up., Registration: URL: https://www.clinicaltrials.gov; Unique identifier: NCT03547583., Competing Interests: Dr deFilippi has received research funding to Inova from Abbott Diagnostics, Roche Diagnostics, Siemens Healthineers, and Ortho Diagnostics and consults for FujiRebio, Roche Diagnostics, Siemens Healthineers, and Ortho Diagnostics. Dr P. Shah received National Institutes of Health (NIH) K23 Career Development Award 1K23HL143179; related grant support paid to institution from Merck, Bayer, and Roche; unrelated grant support from Abbott; related consulting for Merck; and unrelated consulting for Procyrion. Dr S.J. Shah has received research grants from Actelion, Corvia, and NIH; consulting fees/honoraria from Abbott, Amgen, Aria, AstraZeneca, Axon, Bayer, Boehringer Ingelheim, Boston Scientific, Bristol Myers Squibb, Cariora, Cardiovascular Systems Inc, Cyclerion, Cytokinetics, Eisai, ekoi.ai, GlaxoSmithKline, Imara, Ionis, Ironwood, Janssen, Keyto, Lilly Medical, MyoKardia, Novartis, Pfizer, Prothena, Regeneron, Sanofi, Shifamed, Tenax, and United Therapeutics. Dr Lam has received research grants from Bayer, National Medical Research Council of Singapore, Boston Scientific, Roche Diagnostic, Medtronic, Vifor Pharma, and AstraZeneca; consulting fees from Merck, Bayer, Boston Scientific, Roche Diagnostic, Vifor Pharma, AstraZeneca, Novartis, Amgen, Janssen Research & Development LLC, Menarini, Boehringer Ingelheim, Abbott Diagnostics, Corvia, Stealth BioTherapeutics, Novo Nordisk, JanaCare, Biofourmis, Darma, Applied Therapeutics, MyoKardia, Cytokinetics, WebMD Global LLC, Radcliffe Group Ltd, and corpus; and patent PCT/SG2016/050217 pending, a patent 16/216929 pending and cofounder and nonexecutive director of eKo.ai. Dr Butler is a consultant for Abbott, American Regent, Amgen, Applied Therapeutic, AskBio, Astellas, AstraZeneca, Bayer, Boehringer Ingelheim, Boston Scientific, Bristol Myers Squibb, Cardiac Dimension, Cardiocell, Cardior, CSL Bearing, CVRx, Cytokinetics, Daxor, Edwards, Element Science, Faraday, Foundry, G3P, Innolife, Impulse Dynamics, Imbria, Inventiva, Ionis, Lexicon, Lilly, LivaNova, Janssen, Medtronics, Merck, Occlutech, Owkin, Novartis, Novo Nordisk, Pfizer, Pharmacosmos, Pharmain, Pfizer, Prolaio, Regeneron, Renibus, Roche, Salamandra, Sanofi, scPharmaceuticals, Secretome, Sequana, SQ Innovation, Tenex, Tricog, Ultromics, Vifor, and Zoll. Dr Roessig is an employee of Bayer AG. Dr O’Connor received research funding from Merck and consulting fees from Bayer, Dey Pharma LP, and Bristol Myers Squibb Foundation. Dr Westerhout received consulting fees from Bayer. Dr Armstrong received consulting fees from Merck, Bayer, Boehringer Ingelheim, and Novo Nordisk and research grants from Merck, Bayer, Boehringer Ingelheim/Eli Lilly, and CSL Limited. The other author reports no conflicts.