In Vitro and in Silico Study of New Biscoumarin Glycosides from Paramignya trimera against Angiotensin-Converting Enzyme 2 (ACE-2) for Preventing SARS-CoV-2 Infection.

In Vietnam, the stems and roots of the Rutaceous plant Paramignya trimera (Oliv.) Burkill (known locally as "Xáo tam phân") are widely used to treat liver diseases such as viral hepatitis and acute and chronic cirrhosis. In an effort to search for Vietnamese natural compounds capable of inhibiting coronavirus based on molecular docking screening, two new dimeric coumarin glycosides, namely cis-paratrimerin B (1) and cis-paratrimerin A (2), and two previously identified coumarins, the trans-isomers paratrimerin B (3) and paratrimerin A (4), were isolated from the roots of P. trimera and tested for their anti-angiotensin-converting enzyme 2 (ACE-2) inhibitory properties in vitro. It was discovered that ACE-2 enzyme was inhibited by cis-paratrimerin B (1), cis-paratrimerin A (2), and trans-paratrimerin B (3), with IC50 values of 28.9, 68, and 77 µM, respectively. Docking simulations revealed that four biscoumarin glycosides had good binding energies (∆G values ranging from -10.6 to -14.7 kcal/mol) and mostly bound to the S1' subsite of the ACE-2 protein. The key interactions of these natural ligands include metal chelation with zinc ions and multiple H-bonds with Ser128, Glu145, His345, Lys363, Thr371, Glu406, and Tyr803. Our findings demonstrated that biscoumarin glycosides from P. trimera roots occur naturally in both cis- and trans-diastereomeric forms. The biscoumarin glycosides Lys363, Thr371, Glu406, and Tyr803. Our findings demonstrated that biscoumarin glycosides from P. trimera roots hold potential for further studies as natural ACE-2 inhibitors for preventing severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection.

Discovery of Novel Spike Inhibitors against SARS-CoV-2 Infection.

Severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) is responsible for the current coronavirus disease pandemic. With the rapid evolution of variant strains, finding effective spike protein inhibitors is a logical and critical priority. Angiotensin-converting enzyme 2 (ACE2) has been identified as the functional receptor for SARS-CoV-2 viral entry, and thus related therapeutic approaches associated with the spike protein-ACE2 interaction show a high degree of feasibility for inhibiting viral infection. Our computer-aided drug design (CADD) method meticulously analyzed more than 260,000 compound records from the United States National Cancer Institute (NCI) database, to identify potential spike inhibitors. The spike protein receptor-binding domain (RBD) was chosen as the target protein for our virtual screening process. In cell-based validation, SARS-CoV-2 pseudovirus carrying a reporter gene was utilized to screen for effective compounds. Ultimately, compounds C2, C8, and C10 demonstrated significant antiviral activity against SARS-CoV-2, with estimated EC50 values of 8.8 µM, 6.7 µM, and 7.6 µM, respectively. Using the above compounds as templates, ten derivatives were generated and robust bioassay results revealed that C8.2 (EC50 = 5.9 µM) exhibited the strongest antiviral efficacy. Compounds C8.2 also displayed inhibitory activity against the Omicron variant, with an EC50 of 9.3 µM. Thus, the CADD method successfully discovered lead compounds binding to the spike protein RBD that are capable of inhibiting viral infection.

Microwave-assisted synthesis of highly sulfated mannuronate glycans as potential inhibitors against SARS-CoV-2.

Algae-based marine carbohydrate drugs are typically decorated with negative ion groups such as carboxylate and sulfate groups. However, the precise synthesis of highly sulfated alginates is challenging, thus impeding their structure-activity relationship studies. Herein we achieve a microwave-assisted synthesis of a range of highly sulfated mannuronate glycans with up to 17 sulfation sites by overcoming the incomplete sulfation due to the electrostatic repulsion of crowded polyanionic groups. Although the partially sulfated tetrasaccharide had the highest affinity for the receptor binding domain (RBD) of the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) Omicron variant, the fully sulfated octasaccharide showed the most potent interference with the binding of the RBD to angiotensin-converting enzyme 2 (ACE2) and Vero E6 cells, indicating that the sulfated oligosaccharides might inhibit the RBD binding to ACE2 in a length-dependent manner.

QSAR modeling approaches to identify a novel ACE2 inhibitor that selectively bind with the C and N terminals of the ectodomain.

Due to the high rates of drug development failure and the massive expenses associated with drug discovery, repurposing existing drugs has become more popular. As a result, we have used QSAR modelling on a large and varied dataset of 657 compounds in an effort to discover both explicit and subtle structural features requisite for ACE2 inhibitory activity, with the goal of identifying novel hit molecules. The QSAR modelling yielded a statistically robust QSAR model with high predictivity (R2tr=0.84, R2ex=0.79), previously undisclosed features, and novel mechanistic interpretations. The developed QSAR model predicted the ACE2 inhibitory activity (PIC50) of 1615 ZINC FDA compounds. This led to the detection of a PIC50 of 8.604 M for the hit molecule (ZINC000027990463). The hit molecule's docking score is -9.67 kcal/mol (RMSD 1.4). The hit molecule revealed 25 interactions with the residue ASP40, which defines the N and C termini of the ectodomain of ACE2. The HIT molecule conducted more than thirty contacts with water molecules and exhibited polar interaction with the ARG522 residue coupled with the second chloride ion, which is 10.4 nm away from the zinc ion. Both molecular docking and QSAR produced comparable findings. Moreover, MD simulation and MMGBSA studies verified docking analysis. The MD simulation showed that the hit molecule-ACE2 receptor complex is stable for 400 ns, suggesting that repurposed hit molecule 3 is a viable ACE2 inhibitor.

In silico screening of natural compounds to inhibit interaction of human ACE2 receptor and spike protein of SARS-CoV-2 for the prevention of COVID-19.

A computational investigation was carried out to find out potential phytochemicals that could inhibit the binding of human angiotensin-converting enzyme-2 (ACE2) receptors to spike protein of SARS-CoV-2 which is an essential step to gain entry inside human cells and onset of viral infection known as Coronavirus disease (COVID-19). A library of phytochemicals was screened by virtual screening against ACE2 receptors resulting in twenty phytochemicals out of 686 which had binding energy (-11.8 to -6.9 kcal/mol). Drug-likeness gave five hits, but ADMET analysis yielded 4 nontoxic hit phytochemicals. Molecular dynamics simulation of four-hit compounds resulted in acceptable stability and good dynamics behavior. These phytochemicals are Hinokinin, Gmelanone, Isocolumbin, and Tinocordioside, from Vitis vinifera, Gmelina arborea, and Tinospora cordifolia. The above-mentioned phytochemicals may be promising ACE2 inhibitors and can prevent infection of SARS-CoV-2 by inhibiting the entry of the virus into host cells.Communicated by Ramaswamy H. Sarma.

In silico Evaluation of ACE2 Inhibition by Prunus armeniaca L. and in vivo Toxicity Study.

BACKGROUND: SARS-CoV-2 is a virus that uses ACE2 to enter the host cell. AIMS AND OBJECTIVES: This study aimed to evaluate the in silico inhibitory activity of polyphenols from Prunus armeniaca (P. armeniaca) on angiotensin-converting enzyme 2 (ACE2). METHODS: The efficacy of phytocompounds from P. armeniaca in inhibiting ACE2 was tested through molecular docking and dynamic analyses. The toxicological analysis of P. armeniaca was also evaluated. RESULTS: A total of twenty polyphenols were docked against the ACE2 active site, and four compounds showed interesting profiles. In vivo acute toxicity study demonstrated that the aqueous extract of Prunus armeniaca was safe. CONCLUSION: Four compounds from Prunus armeniaca seem to exert an inhibitory potential of ACE2.

Is targeting angiotensin-converting enzyme 2 (ACE2) a prophylactic strategy against COVID-19?

Prophylaxis against COVID-19 is greatly needed for vulnerable populations who have a higher risk of developing severe disease. Vaccines and neutralizing antibodies against SARS-CoV-2 are currently the main approaches to preventing the virus infection. However, the constant mutation of SARS-CoV-2 poses a huge challenge to the effectiveness of these prophylactic strategies. A recent study suggested that downregulation of angiotensin-converting enzyme 2 (ACE2), the receptor of SARS-CoV-2 entry into human cells, can decrease susceptibility to viral infection in vitro, in vivo, and in human lungs and livers perfused ex situ. These findings indicate the potential to use agents to reduce ACE2 expression to prevent COVID-19, but the efficacy and safety should be verified in clinical trials. Considering ACE2 performs physiological functions, risks due to its downregulation and benefits from prophylaxis against SARS-CoV-2 infection should be carefully weighed. In the future, updating vaccines against variants of SARS-CoV-2 might still be an important strategy for prophylaxis against COVID-19. Soluble recombinant human ACE2 that acts as a decoy receptor might be an option to overcome the mutation of SARS-CoV-2.

Advances in Targeting ACE2 for Developing COVID-19 Therapeutics.

Since the onset of the coronavirus pandemic in December 2019, the SARS-CoV-2 virus has accounted for over 6.3 million lives resulting in the demand to develop novel therapeutic approaches to target and treat SARS-CoV-2. Improved understanding of viral entry and infection mechanisms has led to identifying different target receptors to mitigate infection in the host. Researchers have been working on identifying and targeting potential therapeutic target receptors utilizing different candidate drugs. Angiotensin-converting enzyme-2 (ACE2) has been known to perform critical functions in maintaining healthy cardiorespiratory function. However, ACE2 also functions as the binding site for the spike protein of SARS-CoV-2, allowing the virus to enter the cells and ensue infection. Therefore, drugs targeting ACE2 receptors can be considered as therapeutic candidates. Strategies targeting the level of ACE2 expression have been investigated and compared to other potential therapeutic targets, such as TMPRSS2, RdRp, and DPP4. This mini review discusses the key therapeutic approaches that target the ACE2 receptor, which is critical to the cellular entry and propagation of the novel SARS-CoV-2. In addition, we summarize the main advantages of ACE2 targeting against alternative approaches for the treatment of COVID-19.

Sensitivity to Vaccines, Therapeutic Antibodies, and Viral Entry Inhibitors and Advances To Counter the SARS-CoV-2 Omicron Variant.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) keeps evolving and mutating into newer variants over time, which gain higher transmissibility, disease severity, and spread in communities at a faster rate, resulting in multiple waves of surge in Coronavirus Disease 2019 (COVID-19) cases. A highly mutated and transmissible SARS-CoV-2 Omicron variant has recently emerged, driving the extremely high peak of infections in almost all continents at an unprecedented speed and scale. The Omicron variant evades the protection rendered by vaccine-induced antibodies and natural infection, as well as overpowers the antibody-based immunotherapies, raising the concerns of current effectiveness of available vaccines and monoclonal antibody-based therapies. This review outlines the most recent advancements in studying the virology and biology of the Omicron variant, highlighting its increased resistance to current antibody-based therapeutics and its immune escape against vaccines. However, the Omicron variant is highly sensitive to viral fusion inhibitors targeting the HR1 motif in the spike protein, enzyme inhibitors, involving the endosomal fusion pathway, and ACE2-based entry inhibitors. Omicron variant-associated infectivity and entry mechanisms of Omicron variant are essentially distinct from previous characterized variants. Innate sensing and immune evasion of SARS-CoV-2 and T cell immunity to the virus provide new perspectives of vaccine and drug development. These findings are important for understanding SARS-CoV-2 viral biology and advances in developing vaccines, antibody-based therapies, and more effective strategies to mitigate the transmission of the Omicron variant or the next SARS-CoV-2 variant of concern.

Neuromodulation by selective angiotensin-converting enzyme 2 inhibitors.

Here, neuromodulatory effects of selective angiotensin-converting enzyme 2 (ACE2) inhibitors were investigated. Two different types of small molecule ligands for ACE2 inhibition were selected using chemical genetic approach, they were synthesized using developed chemical method and tested using presynaptic rat brain nerve terminals (synaptosomes). EBC-36032 (1 µM) increased in a dose-dependent manner spontaneous and stimulated ROS generation in nerve terminals that was of non-mitochondrial origin. Another inhibitor EBC-36033 (MLN-4760) was inert regarding modulation of ROS generation. EBC-36032 and EBC-36033 (100 µM) did not modulate the exocytotic release of L-[14C]glutamate, whereas both inhibitors decreased the initial rate of uptake, but not accumulation (10 min) of L-[14C]glutamate by nerve terminals. EBC-36032 (100 µM) decreased the exocytotic release as well as the initial rate and accumulation of [3H]GABA by nerve terminals. EBC-36032 and EBC-36033 did not change the extracellular levels and transporter-mediated release of [3H]GABA and L-[14C]glutamate, and tonic leakage of [3H]GABA from nerve terminals. Therefore, synthesized selective ACE2 inhibitors decreased uptake of glutamate and GABA as well as exocytosis of GABA at the presynaptic level. The initial rate of glutamate uptake was the only parameter that was mitigated by both ACE2 inhibitors despite stereochemistry issues. In terms of ACE2-targeted antiviral/anti-SARS-CoV-2 and other therapies, novel ACE2 inhibitors should be checked on the subject of possible renin-angiotensin system (RAS)-independent neurological side effects.

The Race for ACE: Targeting Angiotensin-Converting Enzymes (ACE) in SARS-CoV-2 Infection.

The SARS-CoV-2 virus is spreading around the world, and its clinical manifestation COVID-19 is challenging medical, economic, and social systems. With more and more scientific and social media reports on the COVID-19 pandemic appearing, differences in geographical presentations and clinical management occur. Since ACE2 (angiotensin-converting enzyme 2) is the gatekeeper receptor for the SARS-CoV-2 virus in the upper bronchial system, we here focus on the central role of the renin-angiotensin aldosterone system (RAAS) in the SARS-CoV-2 virus infection, the role of pharmacological RAAS inhibitors, and specific genetic aspects, i.e., single nucleotide polymorphisms (SNP) for the clinical outcome of COVID-19. We aimed to bring together clinical, epidemiological, molecular, and pathophysiological and pharmacological data/observations on cardiovascular aspects in the actual SARS-CoV-2 virus pandemic. In detail, we will report controversies about the Yin-Yan between ACE2 and ACE1 and potential implications for the treatment of hypertension, coronary artery disease, and heart failure. Here, we summarize the encouraging and dynamic global effort of multiple biomedical disciplines resulted in astonishing fight against COVID-19 targeting the renin-angiotensin-aldosterone system, yet the race for ACE just begun.

Lactoferrin Inhibition of the Complex Formation between ACE2 Receptor and SARS CoV-2 Recognition Binding Domain.

The present investigation focuses on the analysis of the interactions among human lactoferrin (LF), SARS-CoV-2 receptor-binding domain (RBD) and human angiotensin-converting enzyme 2 (ACE2) receptor in order to assess possible mutual interactions that could provide a molecular basis of the reported preventative effect of lactoferrin against CoV-2 infection. In particular, kinetic and thermodynamic parameters for the pairwise interactions among the three proteins were measured via two independent techniques, biolayer interferometry and latex nanoparticle-enhanced turbidimetry. The results obtained clearly indicate that LF is able to bind the ACE2 receptor ectodomain with significantly high affinity, whereas no binding to the RBD was observed up to the maximum "physiological" lactoferrin concentration range. Lactoferrin, above 1 µM concentration, thus appears to directly interfere with RBD-ACE2 binding, bringing about a measurable, up to 300-fold increase of the KD value relative to RBD-ACE2 complex formation.

Identification of FDA-approved bifonazole as a SARS-CoV-2 blocking agent following a bioreporter drug screen.

We established a split nanoluciferase complementation assay to rapidly screen for inhibitors that interfere with binding of the receptor binding domain (RBD) of the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) spike glycoprotein with its target receptor, angiotensin-converting enzyme 2 (ACE2). After a screen of 1,200 US Food and Drug Administration (FDA)-approved compounds, we identified bifonazole, an imidazole-based antifungal agent, as a competitive inhibitor of RBD-ACE2 binding. Mechanistically, bifonazole binds ACE2 around residue K353, which prevents association with the RBD, affecting entry and replication of spike-pseudotyped viruses as well as native SARS-CoV-2 and its variants of concern (VOCs). Intranasal administration of bifonazole reduces lethality in K18-hACE2 mice challenged with vesicular stomatitis virus (VSV)-spike by 40%, with a similar benefit after live SARS-CoV-2 challenge. Our screen identified an antiviral agent that is effective against SARS-CoV-2 and VOCs such as Omicron that employ the same receptor to infect cells and therefore has high potential to be repurposed to control, treat, or prevent coronavirus disease 2019 (COVID-19).

An ACE2-blocking antibody confers broad neutralization and protection against Omicron and other SARS-CoV-2 variants of concern.

The ongoing evolution of SARS-CoV-2 has resulted in the emergence of Omicron, which displays notable immune escape potential through mutations at key antigenic sites on the spike protein. Many of these mutations localize to the spike protein ACE2 receptor binding domain, annulling the neutralizing activity of therapeutic antibodies that were effective against other variants of concern (VOCs) earlier in the pandemic. Here, we identified a receptor-blocking human monoclonal antibody, 87G7, that retained potent in vitro neutralizing activity against SARS-CoV-2 variants including the Alpha, Beta, Gamma, Delta, and Omicron (BA.1/BA.2) VOCs. Using cryo-electron microscopy and site-directed mutagenesis experiments, we showed that 87G7 targets a patch of hydrophobic residues in the ACE2-binding site that are highly conserved in SARS-CoV-2 variants, explaining its broad neutralization capacity. 87G7 protected mice and hamsters prophylactically against challenge with all current SARS-CoV-2 VOCs and showed therapeutic activity against SARS-CoV-2 challenge in both animal models. Our findings demonstrate that 87G7 holds promise as a prophylactic or therapeutic agent for COVID-19 that is more resilient to SARS-CoV-2 antigenic diversity.

A Cell-Free Assay for Rapid Screening of Inhibitors of hACE2-Receptor-SARS-CoV-2-Spike Binding.

We present a cell-free assay for rapid screening of candidate inhibitors of protein binding, focusing on inhibition of the interaction between the SARS-CoV-2 Spike receptor binding domain (RBD) and human angiotensin-converting enzyme 2 (hACE2). The assay has two components: fluorescent polystyrene particles covalently coated with RBD, termed virion-particles (v-particles), and fluorescently labeled hACE2 (hACE2F) that binds the v-particles. When incubated with an inhibitor, v-particle-hACE2F binding is diminished, resulting in a reduction in the fluorescent signal of bound hACE2F relative to the noninhibitor control, which can be measured via flow cytometry or fluorescence microscopy. We determine the amount of RBD needed for v-particle preparation, v-particle incubation time with hACE2F, hACE2F detection limit, and specificity of v-particle binding to hACE2F. We measure the dose response of the v-particles to known inhibitors. Finally, utilizing an RNA-binding protein tdPP7 incorporated into hACE2F, we demonstrate that RNA-hACE2F granules trap v-particles effectively, providing a basis for potential RNA-hACE2F therapeutics.

Different compounds against Angiotensin-Converting Enzyme 2 (ACE2) receptor potentially containing the infectivity of SARS-CoV-2: an in silico study.

Novel SARS coronavirus or SARS-CoV-2 is a novel coronavirus that was identified and spread from Wuhan in 2019. On January 30th, the World Health Organization declared the coronavirus outbreak as a Global Public Health Emergency. Although Remdesivir and Molnupiravir are FDA-approved drugs for COVID-19, finding new efficient and low-cost antiviral drugs against COVID-19 for applying in more countries can still be helpful. One of the potential sources for finding new and low-cost drugs is the herbal compounds in addition to repurposing FDA-approved drugs. So, in this study, we focused on finding effective drug candidates against COVID-19 based on the computational approaches. As ACE2 serves as a critical receptor for cell entry of this virus. Inhibiting the binding site of SARS-CoV-2 on human ACE2 provides a promising therapeutic approach for developing drugs against SARS-CoV-2. Herein, we applied a bioinformatics approach to identify possible potential inhibitors of SARS-CoV-2. A library of FDA-approved compounds and five natural compounds was screened using Smina docking. Top-docking compounds are then applied in Molecular Dynamics (MD) simulation to assess the stability of ACE2-inhibitor complexes. Results indicate that Luteolin and Chrysin represent high conformation stability with ACE2 during 120 ns of Molecular Dynamics simulation. The binding free energies of Luteolin and Chrysin were calculated by the Molecular Mechanics/Poisson-Boltzmann Surface Area method (MM/PBSA) which confirmed the relative binding free energy of these drugs to ACE2 in favor of the effective binding. So, Luteolin and Chrysin could sufficiently interact with ACE2 and block the Spike binding pocket of ACE2 and can be a potential inhibitor against the binding of SARS-CoV-2 to ACE2 receptor which is an early stage of infection. Luteolin and Chrysin could be suggestive as beneficial compounds for preventing or reducing SARS-CoV-2 transmission and infection which need experimental work to prove.

Virtual Screening of Natural Chemical Databases to Search for Potential ACE2 Inhibitors.

The angiotensin-converting enzyme II (ACE2) is a multifunctional protein in both health and disease conditions, which serves as a counterregulatory component of RAS function in a cardioprotective role. ACE2 modulation may also have relevance to ovarian cancer, diabetes, acute lung injury, fibrotic diseases, etc. Furthermore, since the outbreak of the coronavirus disease in 2019 (COVID-19), ACE2 has been recognized as the host receptor of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). The receptor binding domain of the SARS-CoV-2 S-protein has a strong interaction with ACE2, so ACE2 may be a potent drug target to prevent the virus from invading host cells for anti-COVID-19 drug discovery. In this study, structure- and property-based virtual screening methods were combined to filter natural product databases from ChemDiv, TargetMol, and InterBioScreen to find potential ACE2 inhibitors. The binding affinity between protein and ligands was predicted using both Glide SP and XP scoring functions and the MM-GBSA method. ADME properties were also calculated to evaluate chemical drug-likeness. Then, molecular dynamics (MD) simulations were performed to further explore the binding modes between the highest-potential compounds and ACE2. Results showed that the compounds 154-23-4 and STOCK1N-07141 possess potential ACE2 inhibition activities and deserve further study.

Jumping from Fragment to Drug via Smart Scaffolds.

A focused drug repurposing approach is described where an FDA-approved drug is rationally selected for biological testing based on structural similarities to a fragment compound found to bind a target protein by an NMR screen. The approach is demonstrated by first screening a curated fragment library using 19 F NMR to discover a quality binder to ACE2, the human receptor required for entry and infection by the SARS-CoV-2 virus. Based on this binder, a highly related scaffold was derived and used as a "smart scaffold" or template in a computer-aided finger-print search of a library of FDA-approved or marketed drugs. The most interesting structural match involved the drug vortioxetine which was then experimentally shown by NMR spectroscopy to bind directly to human ACE2. Also, an ELISA assay showed that the drug inhibits the interaction of human ACE2 to the SARS-CoV-2 receptor-binding-domain (RBD). Moreover, our cell-culture infectivity assay confirmed that vortioxetine is active against SARS-CoV-2 and inhibits viral replication. Thus, the use of "smart scaffolds" based on binders from fragment screens may have general utility for identifying candidates of FDA-approved or marketed drugs as a rapid repurposing strategy. Similar approaches can be envisioned for other fields involving small-molecule chemical applications.

Foe and friend in the COVID-19-associated acute kidney injury: an insight on intrarenal renin-angiotensin system.

Since the first reported case in December of 2019, the coronavirus disease 2019 (COVID-19) has became an international public health emergency. So far, there are more than 228,206,384 confirmed cases including 4,687,066 deaths. Kidney with high expression of angiotensin-converting enzyme 2 (ACE2) is one of the extrapulmonary target organs affected in patients with COVID-19. Acute kidney injury (AKI) is one of the independent risk factors for the death of COVID-19 patients. The imbalance between ACE2-Ang(1-7)-MasR and ACE-Ang II-AT1R axis in the kidney may contribute to COVID-19-associated AKI. Although series of research have shown the inconsistent effects of multiple common RAS inhibitors on ACE2 expression and enzyme activity, most of the retrospective cohort studies indicated the safety and protective effects of ACEI/ARB in COVID-19 patients. This review article highlights the current knowledge on the possible involvement of intrarenal RAS in COVID-19-associated AKI with a primary focus on the opposing effects of ACE2-Ang(1-7)-MasR and ACE-Ang II-AT1R signaling in the kidney. Human recombinant soluble ACE2 or ACE2 variants with preserved ACE2-enzymatic activity may be the best options to improve COVID-19-associated AKI.