Biology subjects

Rezelj, V. V.

Publications and source records attributed to Rezelj, V. V..

5 recordsLinked to original sources

bioRxiv2021

Phospholipidosis is a shared mechanism underlying the in vitro antiviral activity of many repurposed drugs against SARS-CoV-2

Repurposing drugs as treatments for COVID-19 has drawn much attention. A common strategy has been to screen for established drugs, typically developed for other indications, that are antiviral in cells or organisms. Intriguingly, most of the drugs that have emerged from these campaigns, though diverse in structure, share a common physical property: cationic amphiphilicity. Provoked by the similarity of these repurposed drugs to those inducing phospholipidosis, a well-known drug side effect, we investigated phospholipidosis as a mechanism for antiviral activity. We tested 23 cationic amphiphilic drugs--including those from phenotypic screens and others that we ourselves had found--for induction of phospholipidosis in cell culture. We found that most of the repurposed drugs, which included hydroxychloroquine, azithromycin, amiodarone, and four others that have already progressed to clinical trials, induced phospholipidosis in the same concentration range as their antiviral activity; indeed, there was a strong monotonic correlation between antiviral efficacy and the magnitude of the phospholipidosis. Conversely, drugs active against the same targets that did not induce phospholipidosis were not antiviral. Phospholipidosis depends on the gross physical properties of drugs, and does not reflect specific target-based activities, rather it may be considered a confound in early drug discovery. Understanding its role in infection, and detecting its effects rapidly, will allow the community to better distinguish between drugs and lead compounds that more directly impact COVID-19 from the large proportion of molecules that manifest this confounding effect, saving much time, effort and cost. One Sentence SummaryDrug-induced phospholipidosis is a single mechanism that may explain the in vitro efficacy of a wide-variety of therapeutics repurposed for COVID-19.

pharmacology and toxicology↗

bioRxiv2021

BRD2 inhibition blocks SARS-CoV-2 infection in vitro by reducing transcription of the host cell receptor ACE2

SARS-CoV-2 infection of human cells is initiated by the binding of the viral Spike protein to its cell-surface receptor ACE2. We conducted a targeted CRISPRi screen to uncover druggable pathways controlling Spike protein binding to human cells. We found that the protein BRD2 is required for ACE2 transcription in human lung epithelial cells and cardiomyocytes, and BRD2 inhibitors currently evaluated in clinical trials potently block endogenous ACE2 expression and SARS-CoV-2 infection of human cells, including those of human nasal epithelia. Moreover, pharmacological BRD2 inhibition with the drug ABBV-744 inhibited SARS-CoV-2 replication in Syrian hamsters. We also found that BRD2 controls transcription of several other genes induced upon SARS-CoV-2 infection, including the interferon response, which in turn regulates the antiviral response. Together, our results pinpoint BRD2 as a potent and essential regulator of the host response to SARS-CoV-2 infection and highlight the potential of BRD2 as a novel therapeutic target for COVID-19.

cell biology↗

bioRxiv2020

An ultra-high affinity synthetic nanobody blocks SARS-CoV-2 infection by locking Spike into an inactive conformation

Without an effective prophylactic solution, infections from SARS-CoV-2 continue to rise worldwide with devastating health and economic costs. SARS-CoV-2 gains entry into host cells via an interaction between its Spike protein and the host cell receptor angiotensin converting enzyme 2 (ACE2). Disruption of this interaction confers potent neutralization of viral entry, providing an avenue for vaccine design and for therapeutic antibodies. Here, we develop single-domain antibodies (nanobodies) that potently disrupt the interaction between the SARS-CoV-2 Spike and ACE2. By screening a yeast surface-displayed library of synthetic nanobody sequences, we identified a panel of nanobodies that bind to multiple epitopes on Spike and block ACE2 interaction via two distinct mechanisms. Cryogenic electron microscopy (cryo-EM) revealed that one exceptionally stable nanobody, Nb6, binds Spike in a fully inactive conformation with its receptor binding domains (RBDs) locked into their inaccessible down-state, incapable of binding ACE2. Affinity maturation and structure-guided design of multivalency yielded a trivalent nanobody, mNb6-tri, with femtomolar affinity for SARS-CoV-2 Spike and picomolar neutralization of SARS-CoV-2 infection. mNb6-tri retains stability and function after aerosolization, lyophilization, and heat treatment. These properties may enable aerosol-mediated delivery of this potent neutralizer directly to the airway epithelia, promising to yield a widely deployable, patient-friendly prophylactic and/or early infection therapeutic agent to stem the worst pandemic in a century.

biochemistry↗

bioRxiv2019

Upstream translation initiation expands the coding capacity of segmented negative-strand RNA viruses

Segmented negative-strand RNA viruses (sNSVs) include the influenza viruses, the bunyaviruses, and other major pathogens of humans, other animals and plants. The genomes of these viruses are extremely short. In response to this severe genetic constraint, sNSVs use a variety of strategies to maximise their coding potential. Because the eukaryotic hosts parasitized by sNSVs can regulate gene expression through low levels of translation initiation upstream of their canonical open reading frames (ORFs), we asked whether sNSVs could use upstream translation initiation to expand their own genetic repertoires. Consistent with this hypothesis, we showed that influenza A viruses (IAVs) and bunyaviruses were capable of upstream translation initiation. Using a combination of reporter assays and viral infections, we found that upstream translation in IAVs can initiate in two unusual ways: through non-AUG initiation in virally encoded untranslated regions, and through the appropriation of an AUG-containing leader sequence from host mRNAs through viral cap-snatching, a process we termed start-snatching. Finally, while upstream translation of cellular genes is mainly regulatory, for sNSVs it also has the potential to create novel viral gene products. If in frame with a viral ORF, this creates N-extensions of canonical viral proteins. If not, it allows the expression of cryptic overlapping ORFs, which we found were highly conserved in IAV and widely distributed in peribunyaviruses. Thus, by exploiting their hosts capacity for upstream translation initiation, sNSVs can expand still further the coding potential of their extremely compact RNA genomes.

molecular biology↗

bioRxiv2019

Modeling cell infection via virus-producing cells rather than free infectious virus significantly improves fits of an in vitro viral kinetic data

Chikungunya and Zika viruses are arthropod-borne viruses that pose significant threat to public health. Experimental data show that during in vitro infection both viruses exhibit qualitatively distinct replication cycle kinetics. Chikungunya viral load rapidly accumulates within the first several hours post infection whereas Zika virus begins to increase at much later times. We sought to characterize these qualitatively distinct in vitro kinetics of chikungunya and Zika viruses by fitting a family of mathematical models to time course viral load datasets. We demonstrate that the standard viral kinetic model, which considers that new infections result only from free virus penetrating susceptible cells, does not fit experimental data as well as a model in which the number of virus-infected cells is the primary determinant of infection rate. We provide biologically meaningful quantifications of the main viral kinetic parameters and show that our results support cell-to-cell or localized transmission as a significant contributor to viral infection with chikungunya and Zika viruses.\n\nImportanceMathematical modeling has become a useful tool to tease out information about virus-host interactions and thus complements experimental work in characterizing and quantifying processes within viral replication cycle. Importantly, mathematical models can fill in incomplete data sets and identify key parameters of infection, provided the appropriate model is used. The in vitro time course dynamics of mosquito transmitted viruses, such as chikungunya and Zika, have not been studied by mathematical modeling and thus limits our knowledge about quantitative description of the individual determinants of viral replication cycle. This study employs dynamical modeling framework to show that the rate at which cells become virus-infected is proportional to the number or virus-infected cells rather than free extracellular virus in the milieu, a widely accepted assumption in models of viral infections. Using the refined mathematical model in combination with viral load data, we provide quantification of the main drivers of chikungunya and Zika in vitro kinetics. Together, our results bring quantitative understanding of the basic components of chikungunya and Zika virus dynamics.

microbiology↗