Multi-omics liquid biopsy identifies mitochondrial dysfunction in geographic atrophy and supports the longevity-associated metabolite alpha-ketoglutarate as a therapeutic strategy

Multi-omics liquid biopsy identifies mitochondrial dysfunction in geographic atrophy and supports the longevity-associated metabolite α-ketoglutarate as a therapeutic strategy | medRxiv Skip to main content Home About Submit ALERTS / RSS Search for this keyword Advanced Search Multi-omics liquid biopsy identifies mitochondrial dysfunction in geographic atrophy and supports the longevity-associated metabolite α-ketoglutarate as a therapeutic strategy Tsai-Chu Yeh , Gabriel Velez , Architesh Prasad , Soo Hyeon Lee , Ditte K. Rasmussen , Aarushi Kumar , Madhumeeta Chadha , Mohamed Ziad Dabaja , Aneal M Singh , Steven Sanislo , Stephen Smith , Prithvi Mryuthyunjaya , Artis Montague , Alexander G. Bassuk , David Almeida , Antoine Dufour , Vinit B Mahajan doi: https://doi.org/10.64898/2026.03.12.26347263 Tsai-Chu Yeh 1 Molecular Surgery Laboratory, Stanford University , Palo Alto, CA 94304, USA 2 Department of Ophthalmology, Byers Eye Institute, Stanford University , Palo Alto, CA 94304, USA 3 National Yang-Ming Chiao Tung University , Taipei, Taiwan MD, MTM Find this author on Google Scholar Find this author on PubMed Search for this author on this site Gabriel Velez 1 Molecular Surgery Laboratory, Stanford University , Palo Alto, CA 94304, USA 2 Department of Ophthalmology, Byers Eye Institute, Stanford University , Palo Alto, CA 94304, USA MD, PhD Find this author on Google Scholar Find this author on PubMed Search for this author on this site Architesh Prasad 1 Molecular Surgery Laboratory, Stanford University , Palo Alto, CA 94304, USA 2 Department of Ophthalmology, Byers Eye Institute, Stanford University , Palo Alto, CA 94304, USA Find this author on Google Scholar Find this author on PubMed Search for this author on this site Soo Hyeon Lee 1 Molecular Surgery Laboratory, Stanford University , Palo Alto, CA 94304, USA 2 Department of Ophthalmology, Byers Eye Institute, Stanford University , Palo Alto, CA 94304, USA PhD Find this author on Google Scholar Find this author on PubMed Search for this author on this site Ditte K. Rasmussen 1 Molecular Surgery Laboratory, Stanford University , Palo Alto, CA 94304, USA 2 Department of Ophthalmology, Byers Eye Institute, Stanford University , Palo Alto, CA 94304, USA 4 Department of Biomedicine, Aarhus University , Denmark MD Find this author on Google Scholar Find this author on PubMed Search for this author on this site Aarushi Kumar 1 Molecular Surgery Laboratory, Stanford University , Palo Alto, CA 94304, USA 2 Department of Ophthalmology, Byers Eye Institute, Stanford University , Palo Alto, CA 94304, USA BS Find this author on Google Scholar Find this author on PubMed Search for this author on this site Madhumeeta Chadha 1 Molecular Surgery Laboratory, Stanford University , Palo Alto, CA 94304, USA 2 Department of Ophthalmology, Byers Eye Institute, Stanford University , Palo Alto, CA 94304, USA PhD Find this author on Google Scholar Find this author on PubMed Search for this author on this site Mohamed Ziad Dabaja 5 Departments of Physiology and Pharmacology & Biochemistry and Molecular Biology, Cumming School of Medicine, University of Calgary , Calgary, AB, T2N 4N1, Canada Find this author on Google Scholar Find this author on PubMed Search for this author on this site Aneal M Singh 1 Molecular Surgery Laboratory, Stanford University , Palo Alto, CA 94304, USA 2 Department of Ophthalmology, Byers Eye Institute, Stanford University , Palo Alto, CA 94304, USA BA Find this author on Google Scholar Find this author on PubMed Search for this author on this site Steven Sanislo 2 Department of Ophthalmology, Byers Eye Institute, Stanford University , Palo Alto, CA 94304, USA MD Find this author on Google Scholar Find this author on PubMed Search for this author on this site Stephen Smith 2 Department of Ophthalmology, Byers Eye Institute, Stanford University , Palo Alto, CA 94304, USA MD Find this author on Google Scholar Find this author on PubMed Search for this author on this site Prithvi Mryuthyunjaya 1 Molecular Surgery Laboratory, Stanford University , Palo Alto, CA 94304, USA 2 Department of Ophthalmology, Byers Eye Institute, Stanford University , Palo Alto, CA 94304, USA MD, MHS Find this author on Google Scholar Find this author on PubMed Search for this author on this site Artis Montague 2 Department of Ophthalmology, Byers Eye Institute, Stanford University , Palo Alto, CA 94304, USA MD, PhD Find this author on Google Scholar Find this author on PubMed Search for this author on this site Alexander G. Bassuk 6 Stead Family Department of Pediatrics and Neurology, The Iowa Neuroscience Institute, The HAWKEYE-IDDRC, The University of Iowa Carver College of Medicine MD, PhD Find this author on Google Scholar Find this author on PubMed Search for this author on this site David Almeida 7 Erie Retina Research & Center for Advanced Surgical Exploration , Erie, PA, 16505. USA MD PhD Find this author on Google Scholar Find this author on PubMed Search for this author on this site Antoine Dufour 5 Departments of Physiology and Pharmacology & Biochemistry and Molecular Biology, Cumming School of Medicine, University of Calgary , Calgary, AB, T2N 4N1, Canada PhD Find this author on Google Scholar Find this author on PubMed Search for this author on this site Vinit B Mahajan 1 Molecular Surgery Laboratory, Stanford University , Palo Alto, CA 94304, USA 2 Department of Ophthalmology, Byers Eye Institute, Stanford University , Palo Alto, CA 94304, USA MD, PhD Find this author on Google Scholar Find this author on PubMed Search for this author on this site For correspondence: mahajanlab{at}gmail.com Abstract Full Text Info/History Metrics Data/Code Preview PDF Abstract Background Mitochondrial dysfunction is an emerging metabolic hallmark of age-related diseases, yet tools to directly profile mitochondrial pathways and test metabolic interventions in the living human eye remain limited. Multi-omics ocular liquid biopsy enables real-time proteomic and metabolomic profiling of the intraocular microenvironment, complementing systemic biomarkers and imaging surrogates. Here, we used this approach to define mitochondrial and tricarboxylic acid (TCA) cycle dysregulation in geographic atrophy (GA) and to assess whether oral α-ketoglutarate (α-KG) supplementation can modulate mitochondrial metabolites within the eye. Methods Mitochondrial and TCA cycle-related proteins were profiled in aqueous humor (AH) samples from patients with GA using DNA-aptamer-based proteomics. In a phase 0 study, a second cohort undergoing sequential cataract surgery provided paired AH samples collected at first-eye surgery and at second-eye surgery after interim α-KG supplementation. These samples underwent targeted metabolomic profiling using hydrophilic interaction liquid chromatography coupled with mass spectrometry. Results In GA, 64 mitochondrial proteins were differentially expressed, including coordinated TCA-cycle deficiencies marked by reduced expression of enzymes regulating TCA entry and flux, including PDHB and DLST. In the phase 0 cohort, oral α-KG supplementation significantly increased intraocular α-KG levels and the α-KG-to-succinate ratio (P < 0.05), with coordinated shifts across TCA intermediates consistent with enhanced TCA cycle flux. Conclusions AH proteomics demonstrated mitochondrial pathway depletion in GA, consistent with reduced oxidative bioenergetic capacity. AH metabolomics provided first-in-human in vivo evidence that systemic α-KG supplementation can modify intraocular metabolites and may enhance intraocular energy metabolism. These findings support ocular liquid biopsy as a precision-health framework for per-patient biomarker-guided metabolic trials in GA. Plain Language Summary Geographic atrophy (GA) is an advanced form of age-related macular degeneration and a major cause of irreversible vision loss. To better understand the biology of GA, we studied proteins and small molecules in aqueous humor, the fluid inside the eye. We found that eyes with GA showed clear signs of mitochondrial dysfunction, including disruptions in the tricarboxylic acid (TCA) cycle, a key pathway for energy production. This suggests that impaired cellular metabolism is an important feature of the disease. We then tested whether oral α-ketoglutarate (α-KG), a metabolite involved in mitochondrial function and previously shown to have life-extending effects in preclinical studies, could alter these metabolic pathways in the human eye. We found that α-KG supplementation not only increased intraocular α-KG levels but changed metabolic markers linked to mitochondrial activity, providing the first direct evidence that oral supplementation can reach the eye and measurably modify metabolism inside the living human eye. Together, these findings show that liquid biopsy can provide a direct molecular snapshot of the living human eye and may help accelerate the development of biomarker-guided therapies for ocular diseases. Questions What specific mitochondrial and TCA-cycle dysfunctions occur in the aqueous humor (AH) of patients with geographic atrophy (GA), and can oral α-ketoglutarate (α-KG) supplementation measurably remodel these metabolic pathways in the living human eye? Findings AH proteomics in GA patients revealed significant mitochondrial disruption and a coordinated depletion of TCA-cycle enzymes. In a paired-eye interventional metabolomics study, oral α-KG significantly increased intraocular α-KG levels and the α-KG-to-succinate ratio, proving that systemic therapy can drive measurable metabolic modulation within the human eye. Meaning Multi-omics liquid biopsy provides a direct, eye-specific readout of mitochondrial metabolism in GA and offers early human proof-of-concept that a systemic metabolic therapy can successfully reach and modify intraocular pathways, paving the way for biomarker-guided clinical trials in AMD. Competing Interest Statement The authors have declared no competing interest. Clinical Trial NCT07269704 Funding Statement TCY is supported by NIH grant 2T32EY027816. VBM is supported by NIH grants (R01EY031952, R01EY031360, R01EY030151, and P30EY026877), and Research to Prevent Blindness, New York, New York. GV is supported by NIH grant R38EY037090. Author Declarations I confirm all relevant ethical guidelines have been followed, and any necessary IRB and/or ethics committee approvals have been obtained. Yes The details of the IRB/oversight body that provided approval or exemption for the research described are given below: All study procedures were approved by the Stanford University Institutional Review Board, complied with HIPAA regulations, adhered to the Declaration of Helsinki, and written informed consent was obtained from all participants. I confirm that all necessary patient/participant consent has been obtained and the appropriate institutional forms have been archived, and that any patient/participant/sample identifiers included were not known to anyone (e.g., hospital staff, patients or participants themselves) outside the research group so cannot be used to identify individuals. Yes I understand that all clinical trials and any other prospective interventional studies must be registered with an ICMJE-approved registry, such as ClinicalTrials.gov. I confirm that any such study reported in the manuscript has been registered and the trial registration ID is provided (note: if posting a prospective study registered retrospectively, please provide a statement in the trial ID field explaining why the study was not registered in advance). Yes I have followed all appropriate research reporting guidelines, such as any relevant EQUATOR Network research reporting checklist(s) and other pertinent material, if applicable. Yes Data Availability All data produced in the present study are available upon reasonable request to the authors Copyright The copyright holder for this preprint is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity. It is made available under a CC-BY-NC 4.0 International license . 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Share Multi-omics liquid biopsy identifies mitochondrial dysfunction in geographic atrophy and supports the longevity-associated metabolite α-ketoglutarate as a therapeutic strategy Tsai-Chu Yeh , Gabriel Velez , Architesh Prasad , Soo Hyeon Lee , Ditte K. Rasmussen , Aarushi Kumar , Madhumeeta Chadha , Mohamed Ziad Dabaja , Aneal M Singh , Steven Sanislo , Stephen Smith , Prithvi Mryuthyunjaya , Artis Montague , Alexander G. Bassuk , David Almeida , Antoine Dufour , Vinit B Mahajan medRxiv 2026.03.12.26347263; doi: https://doi.org/10.64898/2026.03.12.26347263 Share This Article: Copy Citation Tools Multi-omics liquid biopsy identifies mitochondrial dysfunction in geographic atrophy and supports the longevity-associated metabolite α-ketoglutarate as a therapeutic strategy Tsai-Chu Yeh , Gabriel Velez , Architesh Prasad , Soo Hyeon Lee , Ditte K. Rasmussen , Aarushi Kumar , Madhumeeta Chadha , Mohamed Ziad Dabaja , Aneal M Singh , Steven Sanislo , Stephen Smith , Prithvi Mryuthyunjaya , Artis Montague , Alexander G. Bassuk , David Almeida , Antoine Dufour , Vinit B Mahajan medRxiv 2026.03.12.26347263; doi: https://doi.org/10.64898/2026.03.12.26347263 Citation Manager Formats BibTeX Bookends EasyBib EndNote (tagged) EndNote 8 (xml) Medlars Mendeley Papers RefWorks Tagged Ref Manager RIS Zotero Tweet Widget Facebook Like Google Plus One Subject Area Ophthalmology Subject Areas All Articles Addiction Medicine (638) Allergy and Immunology (917) Anesthesia (350) Cardiovascular Medicine (5005) Dentistry and Oral Medicine (496) Dermatology (428) Emergency Medicine (676) Endocrinology (including Diabetes Mellitus and Metabolic Disease) (1691) Epidemiology (16263) Forensic Medicine (36) Gastroenterology (1241) Genetic and Genomic Medicine (7278) Geriatric Medicine (751) Health Economics (1093) Health Informatics (5289) Health Policy (1467) Health Systems and Quality Improvement (1832) Hematology (599) HIV/AIDS (1381) Infectious Diseases (except HIV/AIDS) (16490) Intensive Care and Critical Care Medicine (1203) Medical Education (700) Medical Ethics (156) Nephrology (752) Neurology (7596) Nursing (377) Nutrition (1130) Obstetrics and Gynecology (1296) Occupational and Environmental Health (1040) Oncology (3746) Ophthalmology (1093) Orthopedics (409) Otolaryngology (465) Pain Medicine (501) Palliative Medicine (143) Pathology (721) Pediatrics (1858) Pharmacology and Therapeutics (755) Primary Care Research (786) Psychiatry and Clinical Psychology (6130) Public and Global Health (9992) Radiology and Imaging (2531) Rehabilitation Medicine and Physical Therapy (1519) Respiratory Medicine (1269) Rheumatology (661) Sexual and Reproductive Health (804) Sports Medicine (597) Surgery (803) Toxicology (109) Transplantation (319) Urology (308)

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