Identification of noninvasive and disease-specific biomarkers in hereditary angioedema using urinary proteomics

Background Hereditary angioedema (HAE) is a rare and potentially life-threatening disease. Noninvasive and disease-specific biomarkers are needed for the early diagnosis and clinical management of HAE. Objective We sought to apply untargeted proteomics profiling and targeted proteomics validation to identify pathogenic mechanisms and candidate biomarkers of HAE. Methods Data-independent acquisition (DIA)-based proteomics profiling was performed in urine samples of HAE patients and healthy controls. Bioinformatics analysis was used for functional annotation and pathway enrichment of differentially expressed proteins. Furthermore, promising biomarker candidates were validated in another independent clinical cohort using parallel reaction monitoring (PRM) targeted proteomics quantification. Results Different urinary proteomics profiles were identified among type 1 HAE, type 2 HAE and healthy controls. A total of 401 differentially expressed proteins were identified between type 1 HAE and healthy controls. Bioinformatics analysis showed that several biological processes and pathways were significantly enriched in HAE, including complement and coagulation cascades, cell adhesion molecules, immune response, proteolysis, and bradykinin catabolic process. Moreover, a promising biomarker panel (C1-INH, KNG1 and EGF) were validated in another independent clinical cohort. The area under the curve (AUC) value of this biomarker panel reached 0.910 for HAE diagnosis (sensitivity: 91.7, specificity: 88.9, P <0.001). Conclusions This study describes the first application of a DIA-PRM workflow to identify noninvasive and disease-specific biomarkers in HAE patients. These findings will contribute to the pathogenesis research and biomarker discovery of HAE. Key Messages 1. Different urinary proteomics profiles were identified among type 1 HAE, type 2 HAE and healthy controls. 2. Several biological processes and pathways were significantly enriched in HAE, including complement and coagulation cascades, cell adhesion molecules, immune response, proteolysis, and bradykinin catabolic process. 3. A urinary biomarker panel (C1-INH, KNG1, and EGF) could be a promising noninvasive diagnostic tool for HAE. ### Competing Interest Statement The authors have declared no competing interest. ### Funding Statement This study was funded by grants from the National Key Research and Development Program of China (No. 2016YFC0901501), the National Natural Science Foundation of China (No. 81472870), the CAMS Innovation Fund for Medical Sciences (CIFMS 2021-1-I2M-003) and Youth Science Fund of Peking Union Medical College Hospital (pumch201912152). ### 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: Ethics Committee of Peking Union Medical College Hospital gave ethical approval for this work (No. HS-2402). 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 and uploaded the relevant EQUATOR Network research reporting checklist(s) and other pertinent material as supplementary files, if applicable. Yes All data produced in the present study are available upon reasonable request to the authors * HAE : hereditary angioedema HC : heathy control LC-MS/MS : liquid chromatography coupled with tandem mass spectrometry DIA : data-independent acquisition PRM : parallel reaction monitoring DEP : differentially expressed protein C1-INH or SERPING1 : Protease C1 : inhibitor KNG1 : Kininogen-1 EGF : Pro-epidermal growth factor CLU : Clusterin ROC : receiver operator characteristics AUC : area under the curve