key: cord-0880320-fsbeqsuu
authors: Tantry, Udaya S; Bliden, Kevin P; Cho, Alastair; Walia, Naval; Dahlen, Jeffrey R; Ens, Gordon; Traianova, Malina; Jerjian, Christophe; Usman, Abira; Gurbel, Paul A
title: First Experience Addressing the Prognostic Utility of Novel Urinary Biomarkers in Patients with COVID-19
date: 2021-05-26
journal: Open Forum Infect Dis
DOI: 10.1093/ofid/ofab274
sha: dd714c9ccb6e17e71273209aa599e241fafaca2b
doc_id: 880320
cord_uid: fsbeqsuu

Urine 11-dehydro-thromboxane B(2) (u11-dh-TxB(2)), 8-hydroxy-2'-deoxyguanosine, and liver-type fatty acid binding protein levels (L-FABP) at the time of hospitalization were higher in COVID-19 patients with adverse events versus without events. Higher u11-dh-TxB(2) and L-FABP levels were associated with longer hospitalization, more thrombotic events, and greater mortality, providing evidence for potential utility as early prognostic biomarkers for COVID-19.

COVID-19, a global pandemic, can be associated with rapid and severe disease progression leading to death. Standard biomarkers of inflammation and hypercoagulability such as Creactive protein, interleukin-6, procalcitonin, fibrinogen, and d-dimer are currently measured in blood samples as diagnostic and prognostic markers (1) . However, limited data exist on the use of urinary biomarkers and their relation to COVID-19 severity and outcomes. Early assessment of COVID-19 using urinary markers may facilitate rapid and effective intervention to prevent the progression of the disease to a severe state. Measurement of biomarkers in urine specimens may be advantageous compared to blood specimens due to the comparative ease of sample collection. Urine in some cases is also associated with improved analyte stability and reduced matrix-related assay interference.

We report a sub-analysis of the evaluation of hemostasis in hospitalized COVID-19 patients (TARGET-COVID) study (URL: https://www.clinicaltrials.gov; Unique identifier: NCT04493307). The study was performed in accordance with standard ethical principles and the design of the study was approved by the local institutional review board of the LifeBridge Health. All patients provided written consent.

We enrolled hospitalized patients who were diagnosed with COVID-19 by reverse transcription-polymerase chain reaction assay (n=123) and patients without COVID-19 but with pneumonia and an elevated D-dimer (COVID-19 negative, n=18). Blood and urine samples were collected within 48 hours of hospital admission and serial samples were collected on a subset of patients (n=22) at day 3 and day 5-8 from the baseline. Standard blood biomarkers were determined in the central hospital laboratory (Sinai Hospital, Baltimore, MD, USA). Urinary 11-dehydro-thromboxane B 2 (u11-dh-TxB 2 ) levels were determined using an enzyme-linked immunoassay and microalbumin levels were determined using the Dimension clinical chemistry system at Inflammatory Markers Laboratory (Wichita, KS) (2). Urinary 8-hydroxy-2' -deoxyguanosine (8-OHdG) levels were measured by enzyme-linked immunoassay at CEDx Labs (Nashua, NH, USA) and liver-type fatty acid binding protein (L-FABP) levels were determined using a rapid, point-of-care lateral flow A c c e p t e d M a n u s c r i p t 4 immunoassay (Timewell Medical, Tokyo, Japan) whose results were quantified using a CHR-631 Rapid Test Reader (Kaiwood Technology Co., Ltd., Tainan City, Taiwan) (3, 4) . Inhospital all-cause death, thrombotic events (venous thrombosis, pulmonary thromboembolism, myocardial infarction, and ischemic stroke), non-convalescent plasma transfusion and bleeding occurrences were recorded.

Sixty-seven percent of the patients were African Americans. There were no significant differences in age, sex, co-morbidities, Sequential Organ Failure Assessment Score (SOFA), or levels of urine biomarkers between COVID-19 positive and negative patients (p=ns for all measurements In our study, in patients who were on aspirin at the time of hospitalization, u11-dh TxB 2 levels were lower (p=0.001) compared to patients not on aspirin, but only 17% of patients had u11-dh TxB2 levels lower than the cut-off value for aspirin therapeutic response (<1520 pg/mg creatinine) (2 (Table) and in non-aspirin treated patients with events versus without events (p=0.001) (Figure 1 ). In addition, u11-dh TxB 2 levels were higher in patients with prolonged (≥10 days) hospitalization (p=0.02), death (p=0.004), and mechanical ventilation requirement (p<0.001) (Table) . Urinary 11-dh-TxB 2 levels were increased at day 3 (p=0.01) and returned to similar levels as baseline levels at day 5-8. In patients not treated with aspirin, there was a good correlation between u11-dh TxB 2 and d-dimer (AUC 0.60, p<0.001), but not between CRP or any other conventional markers tested.

Similarly, L-FABP levels were higher in patients with events versus without events, ( 47 and p=0.45, respectively) , whereas urinary creatinine levels were higher in patients with prolonged hospitalization (p=0.007) (Table) .

Our results suggest the prognostic utility of 11-dh TxB 2 , L-FABP, and 8-OHdG measured in urine in patients with COVID-19 at the time of hospitalization. Urinary 11-dh TxB 2 is a marker of whole-body inflammation and TxA 2 biosynthesis contributed by platelets, leukocytes, and endothelial cells (5) . The independent relation of urine u11-dh TxB 2 to adverse outcomes in patients with cardiovascular disease and diabetes treated with aspirin has been demonstrated in major clinical trials (6-8). Among COVID-19 patients in our study, we found significantly elevated levels of u11-dh TxB 2 in patients with events compared to patients without events and higher u11-dh TxB 2 levels were associated with prolonged hospitalization, death, and mechanical ventilation requirement. These observations highlight the potential prognostic utility of u11-dh TxB 2 in patients with COVID-19. This is in line with the observations of systemic cytokine storm, endothelial dysfunction, and elevated thrombotic risk in COVID-19. In addition, it is also interesting to note that aspirin therapy at the time of hospitalization in most patients was pharmacodynamically inadequate to provide a therapeutic response based on measurements of u11-dh TxB 2 (2). L-FABP, expressed in proximal renal tubules, is excreted in urine following hypoxia and is a more accurate indicator of acute kidney injury than serum creatinine (9) . L-FABP may assist in predicting the severity of COVID-19 at an early stage (10) . Due to the high metabolic demand of the kidneys, combined with the sensitivity and specificity of urinary L-FABP for renal ischemia, it has been hypothesized that elevations of urinary L-FABP may be indicative A c c e p t e d M a n u s c r i p t 6 of renal ischemia associated with COVID-19. In our study, urine L-FABP levels were higher in COVID-19 patients with events. These observations indicate that patients with more severe COVID-19 have evidence of renal ischemia and that acute renal injury may be, in part, responsible for the adverse outcomes observed in patients with COVID-19. 8-OHdG is a proposed biomarker of oxidative damage of deoxyribonucleic acid (Figure 2) . It has been shown to be associated with cardiovascular disease and endothelial dysfunction in patients with diabetes (11) (12) (13) . Cytokine storm, and a higher prevalence of diabetes among patients with events appear linked to the higher levels of 8-OHdG observed in these patients.

These pilot data are hypothesis-generating, although this is the largest study to comprehensively study urine biomarkers in COVID-19. Nevertheless, the biomarkers we studied indicate that endothelial dysfunction and inflammation (11-dh TxB 2 ), renal hypoxia and apoptosis (L-FABP), and oxidative stress (8-OHdG) are associated with the severity of the COVID-19. Most importantly, this study provides the first evidence for early prognostic information on COVID-19 severity, including death, using a simple, noninvasive urine sample. Evidence from larger studies are required to confirm the clinical validity and utility of these noninvasive markers.

A c c e p t e d M a n u s c r i p t 7

Diagnostic and prognostic value of hematological and immunological markers in COVID-19 infection: A meta-analysis of 6320 patients

Defining platelet response to acetylsalicylic acid: the relation between inhibition of serum thromboxane B2 and agonist-induced platelet aggregation

Probucol and atorvastatin decrease urinary 8-hydroxy-2'-deoxyguanosine in patients with diabetes and hypercholesterolemia

Combination of two urinary biomarkers predicts acute kidney injury after adult cardiac surgery

Aspirin resistance

Aspirin-resistant thromboxane biosynthesis and the risk of myocardial infarction, stroke, or cardiovascular death in patients at high risk for cardiovascular events

Clopidogrel for High Atherothrombotic Risk and Ischemic Stabilization, Management and Avoidance (CHARISMA) Investigators. Incomplete inhibition of thromboxane biosynthesis by acetylsalicylic acid: determinants and effect on cardiovascular risk

The ASCEND Study Collaborative Group, Thromboxane metabolite excretion is associated with serious vascular events in diabetes mellitus: a sub-study of the ASCEND trial

Evaluation of new acute kidney injury biomarkers in a mixed intensive care unit

Evaluation of Coronavirus Disease 2019 Severity Using Urine Biomarkers

8-Hydroxy-2-deoxyguanosine levels and cardiovascular disease: a systematic review and meta-analysis of the literature

8-Hydroxy2'-deoxyguanosine (8-OHdG): a critical biomarker of oxidative stress and carcinogenesis

Circulating Oxidative Stress Biomarkers in Clinical Studies on Type 2 diabetes and Its Complications

A c c e p t e d M a n u s c r i p t 8 

Platelet and Thrombosis Research, LLC, Baltimore, MD, USA.

Dr. Gurbel reports grants and personal fees from Bayer HealthCare LLC, Otitopic Inc, Amgen, Janssen, and US WorldMeds LLC; grants from Instrumentation Laboratory, Haemonetics, Medicure Inc, Idorsia Pharmaceuticals, and Hikari Dx; personal fees from UpToDate; Dr Gurbel is a relator and expert witness in litigation involving clopidogrel; in addition, Dr. Gurbel has two patents, Detection of restenosis risk in patients issued and Assessment of cardiac health and thrombotic risk in a patient.Dr. Tantry reports receiving honoraria from UptoDate and Aggredyne.Other author reports no disclosures. A c c e p t e d M a n u s c r i p t 12 Figure 2