Urinary proteomics combined with home blood pressure telemonitoring for health care reform trial-First progress report.

High blood pressure (BP) and type-2 diabetes (T2DM) are forerunners of chronic kidney disease and left ventricular dysfunction. Home BP telemonitoring (HTM) and urinary peptidomic profiling (UPP) are technologies enabling risk stratification and personalized prevention. UPRIGHT-HTM (NCT04299529) is an investigator-initiated, multicenter, open-label, randomized trial with blinded endpoint evaluation designed to assess the efficacy of HTM plus UPP (experimental group) over HTM alone (control group) in guiding treatment in asymptomatic patients, aged 55-75 years, with ≥5 cardiovascular risk factors. From screening onwards, HTM data can be freely accessed by all patients and their caregivers; UPP results are communicated early during follow-up to patients and caregivers in the intervention group, but at trial closure in the control group. From May 2021 until January 2023, 235 patients were screened, of whom 53 were still progressing through the run-in period and 144 were randomized. Both groups had similar characteristics, including average age (62.0 years) and the proportions of African Blacks (81.9%), White Europeans (16.7%), women 56.2%, home (31.2%), and office (50.0%) hypertension, T2DM (36.4%), micro-albuminuria (29.4%), and ECG (9.7%) and echocardiographic (11.5%) left ventricular hypertrophy. Home and office BP were 128.8/79.2 mm Hg and 137.1/82.7 mm Hg, respectively, resulting in a prevalence of white-coat, masked and sustained hypertension of 40.3%, 11.1%, and 25.7%. HTM persisted after randomization (48 681 readings up to 15 January 2023). In conclusion, results predominantly from low-resource sub-Saharan centers proved the feasibility of this multi-ethnic trial. The COVID-19 pandemic caused delays and differential recruitment rates across centers.

Predictive performance and clinical application of COV50, a urinary proteomic biomarker in early COVID-19 infection: a prospective multicentre cohort study.

BACKGROUND: The SARS-CoV-2 pandemic is a worldwide challenge. The CRIT-CoV-U pilot study generated a urinary proteomic biomarker consisting of 50 peptides (COV50), which predicted death and disease progression from SARS-CoV-2. After the interim analysis presented for the German Government, here, we aimed to analyse the full dataset to consolidate the findings and propose potential clinical applications of this biomarker. METHODS: CRIT-CoV-U was a prospective multicentre cohort study. In eight European countries (Austria, France, Germany, Greece, North Macedonia, Poland, Spain, and Sweden), 1012 adults with PCR-confirmed COVID-19 were followed up for death and progression along the 8-point WHO scale. Capillary electrophoresis coupled with mass spectrometry was used for urinary proteomic profiling. Statistical methods included logistic regression and receiver operating characteristic curve analysis with a comparison of the area under curve (AUC) between nested models. Hospitalisation costs were derived from the care facility corresponding with the Markov chain probability of reaching WHO scores ranging from 3 to 8 and flat-rate hospitalisation costs adjusted for the gross per capita domestic product of each country. FINDINGS: From June 30 to Nov 19, 2020, 228 participants were recruited, and from April 30, 2020, to April 14, 2021, 784 participants were recruited, resulting in a total of 1012 participants. The entry WHO scores were 1-3 in 445 (44%) participants, 4-5 in 529 (52%) participants, and 6 in 38 (4%) participants; and of all participants, 119 died and 271 had disease progression. The odds ratio (OR) associated with COV50 in all 1012 participants for death was 2·44 (95% CI 2·05-2·92) unadjusted and 1·67 (1·34-2·07) when adjusted for sex, age, BMI, comorbidities, and baseline WHO score; and for disease progression, the OR was 1·79 (1·60-2·01) when unadjusted and 1·63 (1·41-1·91) when adjusted (p<0·0001 for all). The predictive accuracy of the optimised COV50 thresholds was 74·4% (71·6-77·1%) for mortality (threshold 0·47) and 67·4% (64·4-70·3%) for disease progression (threshold 0·04). When adjusted for covariables and the baseline WHO score, these thresholds improved AUCs from 0·835 to 0·853 (p=0·033) for death and from 0·697 to 0·730 (p=0·0008) for progression. Of 196 participants who received ambulatory care, 194 (99%) did not reach the 0·04 threshold. The cost reductions associated with 1 day less hospitalisation per 1000 participants were million Euro (M€) 0·887 (5-95% percentile interval 0·730-1·039) in participants at a low risk (COV50 <0·04) and M€2·098 (1·839-2·365) in participants at a high risk (COV50 ≥0·04). INTERPRETATION: The urinary proteomic COV50 marker might be predictive of adverse COVID-19 outcomes. Even in people with mild-to-moderate PCR-confirmed infections (WHO scores 1-4), the 0·04 COV50 threshold justifies earlier drug treatment, thereby potentially reducing the number of days in hospital and associated costs. FUNDING: German Federal Ministry of Health.

Urinary peptidomic profiles to address age-related disabilities: a prospective population study.

BACKGROUND: The Global Burden of Diseases, Injuries, and Risk Factors Study 2019 called for innovation in addressing age-related disabilities. Our study aimed to identify and validate a urinary peptidomic profile (UPP) differentiating healthy from unhealthy ageing in the general population, to test the UPP predictor in independent patient cohorts, and to search for targetable molecular pathways underlying age-related chronic diseases. METHODS: In this prospective population study, we used data from participants in the Flemish Study on Environment, Genes and Health Outcomes (FLEMENGHO), done in northern Belgium from 1985 to 2019, and invited participants to a follow-up examination in 2005-10. Participants were eligible if their address was within 15 km of the examination centre and if they had not withdrawn consent in any of the previous examination cycles (1985-2004). All participants (2005-10) were also invited to an additional follow-up examination in 2009-13. Participants who took part in both the 2005-10 follow-up examination and in the additional 2009-13 follow-up visit constituted the derivation dataset, which included their 2005-10 data, and the time-shifted internal validation dataset, which included their 2009-13 data. The remaining participants who only had 2005-10 data constituted the synchronous internal validation dataset. Participants were excluded from analyses if they were incapacitated, had not undergone UPP, or had either missing or outlying (three SDs greater than the mean of all consenting participants) values of body-mass index, plasma glucose, or serum creatinine. The UPP was assessed by capillary electrophoresis coupled with mass spectrometry. The multidimensional UPP signature reflecting ageing was generated from the derivation dataset and validated in the time-shifted internal validation dataset and the synchronous validation dataset. It was further validated in patients with diabetes, COVID-19, or chronic kidney disease (CKD). In FLEMENGHO, the mortality endpoints were all-cause, cardiovascular, and non-cardiovascular mortality; other endpoints were fatal or non-fatal cancer and musculoskeletal disorders. Molecular pathway exploration was done using the Reactome and Kyoto Encyclopedia of Genes and Genomes databases. FINDINGS: 778 individuals (395 [51%] women and 383 [49%] men; aged 16·2-82·1 years; mean age 50·9 years [SD 15·8]) from the FLEMENGHO cohort had a follow-up examination between 2005 and 2010, of whom 559 participants had a further follow-up from Oct 28, 2009, to March 19, 2013, and made up the derivation (2005-10) and time-shifted internal validation (2009-13) datasets. 219 were examined once and constituted the synchronous internal validation dataset (2005-10). With correction for multiple testing and multivariable adjustment, chronological age was associated with 210 sequenced peptides mainly showing downregulation of collagen fragments. The trained model relating chronological age to UPP, derived by elastic net regression, included 54 peptides from 17 proteins. The UPP-age prediction model explained 76·3% (r=0·87) of chronological age in the derivation dataset, 54·4% (r=0·74) in the time-shifted validation dataset, and 65·3% (r=0·81) in the synchronous internal validation dataset. Compared with chronological age, the predicted UPP-age was greater in patients with diabetes (chronological age 50·8 years [SE 0·37] vs UPP-age 56·9 years [0·30]), COVID­19 (53·2 years [1·80] vs 58·5 years [1·67]), or CKD (54·6 years [0·97] vs 62·3 years [0·85]; all p<0·0001). In the FLEMENGHO cohort, independent of chronological age, UPP-age was significantly associated with various risk markers related to cardiovascular, metabolic, and renal disease, inflammation, and medication use. Over a median of 12·4 years (IQR 10·8-13·2), total mortality, cardiovascular mortality, and osteoporosis in the population was associated with UPP-age independent of chronological age, with hazard ratios per 10 year increase in UPP-age of 1·54 (95% CI 1·22-1·95) for total mortality, 1·72 (1·20-2·47) for cardiovascular mortality, and 1·40 (1·06-1·85) for osteoporosis and fractures. The most relevant molecular pathways informed by the proteins involved deregulation of collagen biology and extracellular matrix maintenance. INTERPRETATION: The UPP signature indicative of ageing reflects fibrosis and extracellular matrix remodelling and was associated with risk factors and adverse health outcomes in the population and with accelerated ageing in patients. Innovation in addressing disability should shift focus from the ontology of diseases to shared disease mechanisms, in particular ageing-related fibrotic degeneration. FUNDING: European Research Council, Ministry of the Flemish Community, OMRON Healthcare.

SGLT2-Inhibition reverts urinary peptide changes associated with severe COVID-19: An in-silico proof-of-principle of proteomics-based drug repurposing.

Severe COVID-19 is reflected by significant changes in urine peptides. Based on this observation, a clinical test predicting COVID-19 severity, CoV50, was developed and registered as in vitro diagnostic in Germany. We have hypothesized that molecular changes displayed by CoV50, likely reflective of endothelial damage, may be reversed by specific drugs. Such an impact by a drug could indicate potential benefits in the context of COVID-19. To test this hypothesis, urinary peptide data from patients without COVID-19 prior to and after drug treatment were collected from the human urinary proteome database. The drugs chosen were selected based on availability of sufficient number of participants in the dataset (n > 20) and potential value of drug therapies in the treatment of COVID-19 based on reports in the literature. In these participants without COVID-19, spironolactone did not demonstrate a significant impact on CoV50 scoring. Empagliflozin treatment resulted in a significant change in CoV50 scoring, indicative of a potential therapeutic benefit. The study serves as a proof-of-principle for a drug repurposing approach based on human urinary peptide signatures. The results support the initiation of a randomized control trial testing a potential positive effect of empagliflozin for severe COVID-19, possibly via endothelial protective mechanisms.

CD99 and polymeric immunoglobulin receptor peptides deregulation in critical COVID-19: A potential link to molecular pathophysiology?

Identification of significant changes in urinary peptides may enable improved understanding of molecular disease mechanisms. We aimed towards identifying urinary peptides associated with critical course of COVID-19 to yield hypotheses on molecular pathophysiological mechanisms in disease development. In this multicentre prospective study urine samples of PCR-confirmed COVID-19 patients were collected in different centres across Europe. The urinary peptidome of 53 patients at WHO stages 6-8 and 66 at WHO stages 1-3 COVID-19 disease was analysed using capillary electrophoresis coupled to mass spectrometry. 593 peptides were identified significantly affected by disease severity. These peptides were compared with changes associated with kidney disease or heart failure. Similarities with kidney disease were observed, indicating comparable molecular mechanisms. In contrast, convincing similarity to heart failure could not be detected. The data for the first time showed deregulation of CD99 and polymeric immunoglobulin receptor peptides and of known peptides associated with kidney disease, including collagen and alpha-1-antitrypsin. Peptidomic findings were in line with the pathophysiology of COVID-19. The clinical corollary is that COVID-19 induces specific inflammation of numerous tissues including endothelial lining. Restoring these changes, especially in CD99, PIGR and alpha-1-antitripsin, may represent a valid and effective therapeutic approach in COVID-19, targeting improvement of endothelial integrity.

A urinary peptidomic profile predicts outcome in SARS-CoV-2-infected patients.

BACKGROUND: COVID-19 prediction models based on clinical characteristics, routine biochemistry and imaging, have been developed, but little is known on proteomic markers reflecting the molecular pathophysiology of disease progression. METHODS: The multicentre (six European study sites) Prospective Validation of a Proteomic Urine Test for Early and Accurate Prognosis of Critical Course Complications in Patients with SARS-CoV-2 Infection Study (Crit-COV-U) is recruiting consecutive patients (≥ 18 years) with PCR-confirmed SARS-CoV-2 infection. A urinary proteomic biomarker (COV50) developed by capillary-electrophoresis-mass spectrometry (CE-MS) technology, comprising 50 sequenced peptides and identifying the parental proteins, was evaluated in 228 patients (derivation cohort) with replication in 99 patients (validation cohort). Death and progression along the World Health Organization (WHO) Clinical Progression Scale were assessed up to 21 days after the initial PCR test. Statistical methods included logistic regression, receiver operating curve (ROC) analysis and comparison of the area under the curve (AUC). FINDINGS: In the derivation cohort, 23 patients died, and 48 developed worse WHO scores. The odds ratios (OR) for death per 1 standard deviation (SD) increment in COV50 were 3·52 (95% CI, 2·02-6·13, p <0·0001) unadjusted and 2·73 (1·25-5·95, p = 0·012) adjusted for sex, age, baseline WHO score, body mass index (BMI) and comorbidities. For WHO scale progression, the corresponding OR were 2·63 (1·80-3·85, p<0·0001) and 3·38 (1·85-6·17, p<0·0001), respectively. The area under the curve (AUC) for COV50 as a continuously distributed variable was 0·80 (0·72-0·88) for mortality and 0·74 (0·66-0·81) for worsening WHO score. The optimised COV50 thresholds for mortality and worsening WHO score were 0·47 and 0·04 with sensitivity/specificity of 87·0 (74·6%) and 77·1 (63·9%), respectively. On top of covariates, COV50 improved the AUC, albeit borderline for death, from 0·78 to 0·82 (p = 0·11) and 0·84 (p = 0·052) for mortality and from 0·68 to 0·78 (p = 0·0097) and 0·75 (p = 0·021) for worsening WHO score. The validation cohort findings were confirmatory. INTERPRETATION: This first CRIT-COV-U report proves the concept that urinary proteomic profiling generates biomarkers indicating adverse COVID-19 outcomes, even at an early disease stage, including WHO stages 1-3. These findings need to be consolidated in an upcoming final dataset. FUNDING: The German Federal Ministry of Health funded the study.

Urinary Proteomics Associates with COVID-19 Severity: Pilot Proof-of-Principle Data and Design of a Multicentric Diagnostic Study.

SARS-CoV-2 infection results in a mild-to-moderate disease course in most patients, allowing outpatient self-care and quarantine. However, in approx. 10% of cases a two- or three-phasic critical disease course with starting from day 7 to 10 is observed. To facilitate and plan outpatient care, biomarkers prognosing such worsening at an early stage appear of outmost importance. In this accelerated article, we report on the identification of urinary peptides significantly associated with SARS-CoV-2 infection, and the development of a multi-marker urinary peptide based test, COVID20, that may enable prognosis of critical and fatal outcomes in COVID-19 patients. COVID20 is composed of 20 endogenous peptides mainly derived from various collagen chains that enable differentiating moderate or severe disease from critical state or death with 83% sensitivity at 100% specificity. Based on the performance in this pilot study, testing in a prospective study on 1000 patients has been initiated. This article is protected by copyright. All rights reserved.