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Search indexed bioRxiv preprints in genomics, neuroscience, cell biology and bioinformatics. Read source abstracts and check manuscript versions; preprints are not peer reviewed.

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At least 1,153 records · Page 64Linked to original sources

bioRxiv2020

Identification of antiviral drug candidates against SARS-CoV-2 from FDA-approved drugs

COVID-19 is an emerging infectious disease and was recently declared as a pandemic by WHO. Currently, there is no vaccine or therapeutic available for this disease. Drug repositioning represents the only feasible option to address this global challenge and a panel of 48 FDA-approved drugs that have been pre-selected by an assay of SARS-CoV was screened to identify potential antiviral drug candidates against SARS-CoV-2 infection. We found a total of 24 drugs which exhibited antiviral efficacy (0.1 M < IC50 < 10 M) against SARS-CoV-2. In particular, two FDA-approved drugs - niclosamide and ciclesonide - were notable in some respects. These drugs will be tested in an appropriate animal model for their antiviral activities. In near future, these already FDA-approved drugs could be further developed following clinical trials in order to provide additional therapeutic options for patients with COVID-19.

microbiology↗

bioRxiv2020

A SARS-CoV-2-Human Protein-Protein Interaction Map Reveals Drug Targets and Potential Drug-Repurposing

An outbreak of the novel coronavirus SARS-CoV-2, the causative agent of COVID-19 respiratory disease, has infected over 290,000 people since the end of 2019, killed over 12,000, and caused worldwide social and economic disruption1,2. There are currently no antiviral drugs with proven efficacy nor are there vaccines for its prevention. Unfortunately, the scientific community has little knowledge of the molecular details of SARS-CoV-2 infection. To illuminate this, we cloned, tagged and expressed 26 of the 29 viral proteins in human cells and identified the human proteins physically associated with each using affinity-purification mass spectrometry (AP-MS), which identified 332 high confidence SARS-CoV-2-human protein-protein interactions (PPIs). Among these, we identify 66 druggable human proteins or host factors targeted by 69 existing FDA-approved drugs, drugs in clinical trials and/or preclinical compounds, that we are currently evaluating for efficacy in live SARS-CoV-2 infection assays. The identification of host dependency factors mediating virus infection may provide key insights into effective molecular targets for developing broadly acting antiviral therapeutics against SARS-CoV-2 and other deadly coronavirus strains.

systems biology↗

bioRxiv2020

The functional landscape of patient derived RNF43 mutations predicts Wnt inhibitor sensitivity

A subset of Wnt-addicted cancers are sensitive to targeted therapies that block Wnt secretion or receptor engagement. RNF43 loss-of-function mutations that increase cell surface Wnt receptor abundance cause sensitivity to Wnt inhibitors. However, it is not clear which of the clinically identified RNF43 mutations affect its function in vivo. We assayed 90 missense and 45 truncating RNF43 mutations found in human cancers, using a combination of cell-based reporter assays, genome editing, flow cytometry and immunofluorescence microscopy. Patent-derived xenograft (PDX) models with C-terminal truncating RNF43 mutations were tested for Wnt inhibitor sensitivity. We find that five common germline variants of RNF43 have wild-type activity. The majority of cancer-associated missense mutations in the RING and PA domains are either loss of function or hyperactivating. Hyperactivating mutants appear to function through formation of inactive dimers with endogenous RNF43 and/or ZNRF3. C-terminal truncation mutants including the common G659fs mutant, have discordant behavior in in vitro versus in vivo assays. PDXs and cell lines with C-terminal truncations show increased cell surface FZD, Wnt/{beta}-catenin signaling and are responsive to PORCN inhibition in vivo, providing clear evidence of RNF43 loss of function. In conclusion, RNF43 nonsense and frameshift mutations, including those in the C-terminal domain, and specific missense mutations in RING and PA are loss of function and predict response to upstream Wnt inhibitors in microsatellite stable cancers. This study expands the landscape of actionable RNF43 mutations, potentially extending the benefit of these therapies to additional patients. Statement of SignificanceLoss of function RNF43 mutations, first described in pancreatic cancers, drive progression of multiple cancers by increasing cellular sensitivity to Wnt ligands. These cancers are therefore uniquely sensitive to agents such as PORCN inhibitors that block Wnt production. As the PORCN inhibitors and other upstream inhibitors advance into clinical trials it is important to identify the right patients to treat with these upstream Wnt inhibitors. Hence a detailed map of mutations that are actionable is required. Here we systematically examined a spectrum of 135 patient-derived RNF43 mutations from multiple cancers. Using cell-based reporter assays, genome editing and patient-derived xenografts, we identify rules to guide patient selection. MSS cancers with either truncating mutations anywhere in the gene, including C-terminal truncations around the G659 position, or point mutations in well-defined functional domains, are likely to have RNF43 loss of function and hence a response to therapy.

cancer biology↗

bioRxiv2020

Azithromycin and ciprofloxacin have a chloroquine-like effect on respiratory epithelial cells

There is interest in the use of chloroquine/hydroxychloroquine (CQ/HCQ) and azithromycin (AZT) in COVID-19 therapy. Employing cystic fibrosis respiratory epithelial cells, here we show that drugs AZT and ciprofloxacin (CPX) act as acidotropic lipophilic weak bases and confer in vitro effects on intracellular organelles similar to the effects of CQ. These seemingly disparate FDA-approved antimicrobials display a common property of modulating pH of endosomes and trans-Golgi network. We believe this may in part help understand the potentially beneficial effects of CQ/HCQ and AZT in COVID-19, and that the present considerations of HCQ and AZT for clinical trials should be extended to CPX.

microbiology↗

bioRxiv2020

Brain-wide structural and functional disruption in mice with oligodendrocyte specific Nf1 deletion is rescued by inhibition of NOS

Neurofibromin gene (NF1) mutation causes Neurofibromatosis type 1 (NF1), a disorder in which brain white matter deficits identified by neuroimaging are common, yet of unknown cellular etiology. In mice, Nf1 loss in adult oligodendrocyte causes myelin decompaction, and increases oligodendrocyte nitric oxide (NO) levels. Nitric oxide synthase (NOS) inhibitors rescue this pathology. Whether oligodendrocyte pathology is sufficient to affect brainwide structure and account for NF1 imaging findings is unknown. Here, we show that Nf1 gene inactivation in adult oligodendrocytes (Plp-Nf1fl/+ mice) results in a motor coordination deficit. Magnetic resonance imaging in awake mice shows that fractional anisotropy is reduced in Plp-Nf1fl/+ corpus callosum and that interhemispheric functional connectivity in motor cortex is also reduced, consistent with disrupted myelin integrity. Further, NOS-specific inhibition rescued both measures. These results demonstrate that oligodendrocyte defects account for aspects of brain dysfunction in NF1, which can be identified by neuroimaging and ameliorated by NOS inhibition. Significance statementThis study assesses the effects of myelin decompaction on motor behavior and brain-wide structural and functional connectivity, and the effect of nitric oxide synthase (NOS) inhibition by N-nitro-L-arginine methyl ester (L-NAME) on these imaging measures. We report that inducible oligodendrocyte-specific inactivation of the Nf1 gene, which causes myelin decompaction, results in reduced motor coordination. Using diffusion-based MRI we show reduced myelin integrity and using functional MRI we show reduced functional connectivity in awake passive mice. L-NAME administration results in rescue of the pathology at the mesoscopic level using imaging procedures that can be directly applied to humans to study treatment efficacy in clinical trials.

neuroscience↗

bioRxiv2020

The Spike Protein S1 Subunit of SARS-CoV-2 Contains an LxxIxE-like Motif that is Known to Recruit the Host PP2A-B56 Phosphatase

SARS-CoV-2 is highly contagious and can cause acute respiratory distress syndrome (ARDS) and multiple organ failure that are largely attributed to the cytokine storm. The surface coronavirus spike (S) glycoprotein is considered as a key factor in host specificity because it mediates infection by receptor-recognition and membrane fusion. Here, the analysis of SARS-CoV-2 S protein revealed two B56-binding LxxIxE-like motifs in S1 and S2 subunits that could recruit the host protein phosphatase 2A (PP2A). The motif in S1 subunit is absent in SARS-CoV and MERS-CoV. Phosphatases and kinases are major players in the regulation of pro-inflammatory responses during pathogenic infections. Moreover, studies have shown that viruses target PP2A in order to manipulate hosts antiviral responses. Recent researches have indicated that SARS-CoV-2 is involved in sustained host inflammation. Therefore, by controlling acute inflammation, it is possible to eliminate its dangerous effects on the host. Among efforts to fight COVID-19, the interaction between LxxIxE-like motif and the PP2A-B56-binding pocket could be a target for the discovery and/or development of a bioactive ligand inhibitor for therapeutic purposes. Indeed, a small molecule called Artepillin C (ArtC), a main compound in Brazilian honeybee green propolis, mimics the side chains of LxxLxE motif. Importantly, ArtC is known, among other effects, to have anti-inflammatory activity that makes it an excellent candidate for future clinical trials in COVID-19 patients.

bioinformatics↗

bioRxiv2020

Identifying therapeutic drug targets for rare and common forms of short stature

While GWAS of common diseases has delivered thousands of novel genetic findings, prioritizing genes for translation to therapeutics has been challenging. Here, we propose an approach to resolve that issue by identifying genes that have both gain of function (GoF) and loss of function (LoF) mutations associated with opposing effects on phenotype (Bidirectional Effect Selected Targets, BEST). Bidirectionality is a desirable feature of the best targets because it implies both a causal role on the phenotype in one direction and that modulating the target activity might be safe and therapeutically beneficial in the other. We used height, a highly heritable trait and a model of complex diseases, to test our approach. Using 34,231 individuals with exome sequence data and height, we identified five genes (IGF1R, NPPC, NPR2, FGFR3, and SHOX) with evidence for bidirectional effects on stature. Rare protein-altering variants significantly increased risk for idiopathic short stature (ISS) (OR=2.75, p= 3.99x10-8). These genes are key members of the only two pathways successfully targeted for short stature: Growth Hormone/Insulin-like growth factor 1 axis and C-type Natriuretic peptide (CNP) for Achondroplasia, a monogenic form of dwarfism. We assayed a subset of NPR2 mutations and identified those with elevated (GoF) and diminished (LoF) activity and found that a polygenic score for height modulates the penetrance of pathogenic variants. We also demonstrated that adding exogenous CNP (encoded by NPPC) rescues the NPR2 haploinsufficiency molecular phenotype in a CRISPR-engineered cell line, thus validating its potential therapeutic treatment for inherited forms of short stature. Finally, we found that these BEST targets increase the probability of success in clinical trials above and beyond targets with other genetic evidence. Our results show the value of looking for bidirectional effects to identify and validate drug targets.

genetics↗

bioRxiv2020

In vitro screening of a FDA approved chemical library reveals potential inhibitors of SARS-CoV-2 replication

A novel coronavirus, named SARS-CoV-2, emerged in 2019 from Hubei region in China and rapidly spread worldwide. As no approved therapeutics exists to treat Covid-19, the disease associated to SARS-Cov-2, there is an urgent need to propose molecules that could quickly enter into clinics. Repurposing of approved drugs is a strategy that can bypass the time consuming stages of drug development. In this study, we screened the Prestwick Chemical Library(R) composed of 1,520 approved drugs in an infected cell-based assay. 90 compounds were identified. The robustness of the screen was assessed by the identification of drugs, such as Chloroquine derivatives and protease inhibitors, already in clinical trials. The hits were sorted according to their chemical composition and their known therapeutic effect, then EC50 and CC50 were determined for a subset of compounds. Several drugs, such as Azithromycine, Opipramol, Quinidine or Omeprazol present antiviral potency with 2<EC50<20{micro}M. By providing new information on molecules inhibiting SARS-CoV-2 replication in vitro, this study could contribute to the short-term repurposing of drugs against Covid-19.

microbiology↗

bioRxiv2020

Atazanavir inhibits SARS-CoV-2 replication and pro-inflammatory cytokine production

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is already responsible for far more deaths than previous pathogenic coronaviruses (CoVs) from 2002 and 2012. The identification of clinically approved drugs to be repurposed to combat 2019 CoV disease (COVID-19) would allow the rapid implementation of potentially life-saving procedures. The major protease (Mpro) of SARS-CoV-2 is considered a promising target, based on previous results from related CoVs with lopinavir (LPV), an HIV protease inhibitor. However, limited evidence exists for other clinically approved antiretroviral protease inhibitors, such as atazanavir (ATV). ATV is of high interest because of its bioavailability within the respiratory tract. Our results show that ATV could dock in the active site of SARS-CoV-2 Mpro, with greater strength than LPV. ATV blocked Mpro activity. We confirmed that ATV inhibits SARS-CoV-2 replication, alone or in combination with ritonavir (RTV) in Vero cells, human pulmonary epithelial cell line and primary monocytes, impairing virus-induced enhancement of IL-6 and TNF- levels. Together, our data strongly suggest that ATV and ATV/RTV should be considered among the candidate repurposed drugs undergoing clinical trials in the fight against COVID-19.

microbiology↗

bioRxiv2020

Multi-organ transcriptomic profiling reveals differential regulation of the immune system among the brain-liver-fats organ network of short-term fasted mice

Different fasting regimens are known to promote health, mitigate chronic immunological disorders, and improve age-related pathophysiological parameters in animals and humans. Indeed, several clinical trials are currently ongoing using fasting as a potential therapy for a wide range of conditions. Fasting alters metabolism by acting as a reset for energy homeostasis. However, the molecular mechanisms underlying the beneficial effects of short-term fasting (STF) are still not well understood, particularly at the systems or multi-organ level. Here, we investigated the dynamic gene expression patterns associated with six periods of STF in nine different mouse organs. We cataloged the transcriptional dynamics within and between organs during STF and discovered differential temporal effects of STF among organs. Using gene ontology enrichment analysis, we identified an organ network sharing 37 common biological pathways perturbed by STF. This network incorporates the brain, liver, interscapular brown adipose tissue, and posterior-subcutaneous white adipose tissue, hence we named it the brain-liver-fats organ network. Using Reactome pathways analysis, we identified the immune system, dominated by T cell regulation processes, as a central and prominent target of systemic modulations during STF in this organ network. The changes we identified in specific immune components point to the priming of adaptive immunity and parallel the fine-tuning of innate immune signaling. Our study provides a comprehensive multi-organ transcriptomic profiling of mice subjected to multiple periods of STF, and adds new insights into the molecular modulators involved in the systemic immuno-transcriptomic changes that occur during short-term energy loss.

systems biology↗

bioRxiv2020

Identifying the spatial and temporal dynamics of molecularly-distinct glioblastoma sub-populations

Glioblastomas (GBMs) are the most aggressive primary brain tumours and have no known cure. Each individual tumour comprises multiple sub-populations of genetically-distinct cells that may respond differently to targeted therapies and may contribute to disappointing clinical trial results. Image-localized biopsy techniques allow multiple biopsies to be taken during surgery and provide information that identifies regions where particular sub-populations occur within an individual GBM, thus providing insight into their regional genetic variability. These sub-populations may also interact with one another through a competitive or cooperative nature; it is important to ascertain the nature of these interactions, as they may have implications for responses to targeted therapies. We combine genetic information from biopsies with a mechanistic model of interacting GBM sub-populations to characterise the nature of interactions between two commonly occurring GBM sub-populations, those with EGFR and PDGFRA genes amplified. We study population levels found across image-localized biopsy data from a cohort of 25 patients and compare this to model outputs under competitive, cooperative and neutral interaction assumptions. We explore other factors affecting the observed simulated sub-populations, such as selection advantages and phylogenetic ordering of mutations, which may also contribute to the levels of EGFR and PDGFRA amplified populations observed in biopsy data.

cancer biology↗

bioRxiv2020

Dual genome-wide CRISPR knockout and CRISPR activation screens identify common mechanisms that regulate the resistance to multiple ATR inhibitors

The ataxia telangiectasia and Rad3-related (ATR) protein kinase is a key regulator of the cellular response to DNA damage. Due to increased amount of replication stress, cancer cells heavily rely on ATR to complete DNA replication and cell cycle progression. Thus, ATR inhibition is an emerging target in cancer therapy, with multiple ATR inhibitors currently undergoing clinical trials. Here, we describe dual genome-wide CRISPR knockout and CRISPR activation screens employed to comprehensively identify genes that regulate the cellular resistance to ATR inhibitors. Specifically, we investigated two different ATR inhibitors, namely VE822 and AZD6738, in both HeLa and MCF10A cells. We identified and validated multiple genes that alter the resistance to ATR inhibitors. Importantly, we show that the mechanisms of resistance employed by these genes are varied, and include restoring DNA replication tract progression, and prevention of ATR inhibitor-induced apoptosis. Our dual genome-wide screen findings pave the way for personalized medicine by identifying potential biomarkers for ATR inhibitor resistance.

cancer biology↗

bioRxiv2020

Testosterone use in female mice does not impair fertilizability of eggs: Implications for the fertility care of transgender males

STUDY QUESTIONDoes testosterone use in females affect reproductive potential, particularly with regard to the production of fertilizable gametes? SUMMARY ANSWERTestosterone cypionate injections given to post-pubertal female mice caused virilization and ovaries were smaller than control ovaries, but ovaries were still responsive to hormonal stimulation and produced fertilizable eggs when superovulated. WHAT IS KNOWN ALREADYStudies to examine the effects of testosterone on reproductive potential in transgender males are lacking. Recently, a model was developed that simulates many aspects of testosterone use in transgender males in order to look at reproductive effects of testosterone in female mice. This study found masculinizing effects on the mice but did not find significant deficits on the number of ovarian follicles; however, effects of testosterone use on ovarian stimulation and fertilizability of oocytes were not investigated. STUDY DESIGN, SIZE, DURATIONA total of 66, 6-week-old Hsd:NSA(CF-1) female mice and 6 Hsd:ICR (CD-1) mice were used for this study. Mice were injected subcutaneously with 400 g testosterone cypionate or sesame oil once a week for 6 weeks and were either sacrificed a week after the 6th injection (active exposure group), or were sacrificed 6-7 weeks after the final testosterone injection (washout group). PARTICIPANTS/MATERIALS, SETTING, METHODSBoth active exposure and washout groups were further subdivided into 3 groups: unstimulated, eCG-stimulated, or eCG/hCG-stimulated. eCG-stimulated mice were sacrificed 44-48 hrs after eCG injection. eCG/hCG-stimulated mice were injected with eCG, followed 48 hrs later with hCG. Mice were sacrificed [~]13-18 hrs after the hCG injection. Data collected included daily vaginal cytology, terminal hormone levels and ovary weights, ovarian histology, number of oocytes/eggs collected in each group, and cleavage to the 2-cell stage following in vitro fertilization. MAIN RESULTS AND THE ROLE OF CHANCETestosterone cypionate-treated mice had testosterone levels elevated to the level of male mice and ceased cycling. Ovaries were significantly smaller in testosterone-treated mice, but they contained normal cohorts of follicles and responded to gonadotropin stimulation by ovulating similar numbers of eggs that fertilized and cleaved in vitro. LIMITATIONS, REASONS FOR CAUTIONOur model treated female mice for only 6 weeks, whereas many transgender men use testosterone for many years before considering biological children. Importantly, a mouse system may not perfectly simulate human reproductive physiology. WIDER IMPLICATIONS OF THE FINDINGSThe current standard of care for transgender men who desire biological children is to cease testosterone therapy prior to ovarian stimulation, but the necessity for stopping testosterone is not known. Our model demonstrates that it is possible for testosterone-suppressed ovaries to respond to gonadotropic stimulation by producing and ovulating fertilizable eggs, thereby obviating the need for testosterone cessation prior to ovarian stimulation. In time, these results may provide insights for future clinical trials of fertility treatment options for transgender men.

physiology↗

bioRxiv2020

Replication of Equine arteritis virus is efficiently suppressed by purine and pyrimidine biosynthesis inhibitors

RNA viruses are responsible for a large variety of animal infections. Equine Arteritis Virus (EAV) is a positive single-stranded RNA virus member of the family Arteriviridae from the order Nidovirales like the Coronaviridae. EAV causes respiratory and reproductive diseases in equids. Although two vaccines are available, the vaccination coverage of the equine population is largely insufficient to prevent new EAV outbreaks around the world. In this study, we present a high-throughput in vitro assay suitable for testing candidate antiviral molecules on equine dermal cells infected by EAV. Using this assay, we identified three molecules that impair EAV infection in equine cells: the broad-spectrum antiviral and nucleoside analog ribavirin, and two compounds previously described as inhibitors of dihydroorotate dehydrogenase (DHODH), the fourth enzyme of the pyrimidine biosynthesis pathway. These molecules effectively suppressed cytopathic effects associated to EAV infection, and strongly inhibited viral replication and production of infectious particles. Since ribavirin is already approved in human and small animal, and that several DHODH inhibitors are in advanced clinical trials, our results open new perspectives for the management of EAV outbreaks.

microbiology↗

bioRxiv2020

Discovery of baicalin and baicalein as novel, natural product inhibitors of SARS-CoV-2 3CL protease in vitro

Human infections with severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) cause coronavirus disease 19 (COVID-19) and there is currently no cure. The 3C-like protease (3CLpro), a highly conserved protease indispensable for replication of coronaviruses, is a promising target for development of broad-spectrum antiviral drugs. To advance the speed of drug discovery and development, we investigated the inhibition of SARS-CoV-2 3CLpro by natural products derived from Chinese traditional medicines. Baicalin and baicalein were identified as the first non-covalent, non-peptidomimetic inhibitors of SARS-CoV-2 3CLpro and exhibited potent antiviral activities in a cell-based system. Remarkably, the binding mode of baicalein with SARS-CoV-2 3CLpro determined by X-ray protein crystallography is distinctly different from those of known inhibitors. Baicalein is perfectly ensconced in the core of the substrate-binding pocket by interacting with two catalytic residues, the crucial S1/S2 subsites and the oxyanion loop, acting as a "shield" in front of the catalytic dyad to prevent the peptide substrate approaching the active site. The simple chemical structure, unique mode of action, and potent antiviral activities in vitro, coupled with the favorable safety data from clinical trials, emphasize that baicalein provides a great opportunity for the development of critically needed anti-coronaviral drugs.

molecular biology↗

bioRxiv2020

Comparative in vitro transcriptomic analyses of COVID-19 candidate therapy hydroxychloroquine suggest limited immunomodulatory evidence of SARS-CoV-2 host response genes.

Hydroxychloroquine (HCQ) has emerged as a potential and controversial antiviral candidate therapy for COVID-19. While many clinical trials are underway to test the efficacy of HCQ as a treatment for COVID-19, underlying mechanisms of HCQ in the setting of COVID-19 remain unclear. Hence, we examined differential gene expression signatures of HCQ exposure, in vitro SARS-CoV-2 infection, and host signatures of COVID-19 in blood, bronchoalveolar lavage, and postmortem lung to evaluate whether HCQ transcriptome signatures associate with restoration of SARS-CoV-2-related host transcriptional responses. Here, we show that 24 hours of in vitro treatment of peripheral blood mononuclear cells(PBMC) with HCQ significantly impacted transcription of 16 genes involved in immune regulation and lipid metabolism. Using transcriptome data from in vitro SARS-CoV-2 infected NHBE and A549 cells and PBMC derived from confirmed COVID-19 infected patients, we determined that only 0.24% of the COVID-19 PBMC differentially expressed gene set and 0.39% of the in vitro SARS-CoV-2 cells differentially expressed gene set overlapped with HCQ-related differentially expressed genes. Moreover, we observed that HCQ treatment significantly impacted transcription of 159 genes in human primary monocyte-derived macrophages involved in cholesterol biosynthetic process and chemokine activity. Notably, when we compared the macrophage HCQ-related gene lists with genes transcriptionally altered during SARS-CoV-2 infection and in bronchoalveolar lavage of COVID-19+ patients, the CXCL6 gene was impacted in all three transcriptional signatures revealing evidence in favor of chemokine modulation. HCQ-related transcriptional changes minimally overlapped with host genes altered in postmortem lung biopsies from COVID-19 participants. These results may provide insight into the immunomodulation mechanisms of HCQ treatment in the setting of COVID-19 and suggest HCQ is not a panacea to SARS-CoV-2 infection.

immunology↗

bioRxiv2020

Immunity-and-Matrix-Regulatory Cells Derived from Human Embryonic Stem Cells Safely and Effectively Treat Mouse Lung Injury and Fibrosis

Lung injury and fibrosis represent the most significant outcomes of severe and acute lung disorders, including COVID-19. However, there are still no effective drugs to treat lung injury and fibrosis. In this study, we report the generation of clinical-grade human embryonic stem cells (hESCs)-derived immunity- and matrix-regulatory cells (IMRCs) produced under good manufacturing practice (GMP) requirements, that can treat lung injury and fibrosis in vivo. We generate IMRCs by sequentially differentiating hESCs with serum-free reagents. IMRCs possess a unique gene expression profile distinct from umbilical cord mesenchymal stem cells (UCMSCs), such as higher levels of proliferative, immunomodulatory and anti-fibrotic genes. Moreover, intravenous delivery of IMRCs inhibits both pulmonary inflammation and fibrosis in mouse models of lung injury, and significantly improves the survival rate of the recipient mice in a dose-dependent manner, likely through paracrine regulatory mechanisms. IMRCs are superior to both primary UCMSCs and FDA-approved pirfenidone, with an excellent efficacy and safety profile in mice and monkeys. In light of public health crises involving pneumonia, acute lung injury (ALI) and acute respiratory distress syndrome (ARDS), our findings suggest that IMRCs are ready for clinical trials on lung disorders.

cell biology↗

bioRxiv2020

Cryo-EM structures of human TRPC5 reveal interaction of a xanthine-based TRPC1/4/5 inhibitor with a conserved lipid binding site.

TRPC1/4/5 channels are non-specific cation channels implicated in a wide variety of diseases, and TRPC1/4/5 inhibitors have recently entered the first clinical trials. However, fundamental and translational studies require a better understanding of TRPC1/4/5 channel regulation by endogenous and exogenous factors. Although several potent and selective TRPC1/4/5 modulators have been reported, the paucity of mechanistic insights into their modes-of-action remains a barrier to the development of new chemical probes and drug candidates. The xanthine class of modulators includes the most potent and selective TRPC1/4/5 inhibitors described to date, as well as TRPC5 activators. Our previous studies suggest that xanthines interact with a, so far, elusive pocket of TRPC1/4/5 channels that is essential to channel gating. Targeting this pocket may be a promising strategy for TRPC1/4/5 drug discovery. Here we report the first structure of a small molecule-bound TRPC1/4/5 channel - human TRPC5 in complex with the xanthine Pico145 - to 3.0 [A]. We found that Pico145 binds to a conserved lipid binding site of TRPC5, where it displaces a bound phospholipid. Our findings explain the mode-of-action of xanthine-based TRPC1/4/5 modulators, and suggest a structural basis for TRPC1/4/5 modulation by endogenous factors such as (phospho)lipids and Zn2+ ions. These studies lay the foundations for the structure-based design of new generations of TRPC1/4/5 modulators.

molecular biology↗