Multiscale interactome analysis coupled with off-target drug predictions reveals drug repurposing candidates for human coronavirus disease.

The COVID-19 pandemic has highlighted the urgent need for the identification of new antiviral drug therapies for a variety of diseases. COVID-19 is caused by infection with the human coronavirus SARS-CoV-2, while other related human coronaviruses cause diseases ranging from severe respiratory infections to the common cold. We developed a computational approach to identify new antiviral drug targets and repurpose clinically-relevant drug compounds for the treatment of a range of human coronavirus diseases. Our approach is based on graph convolutional networks (GCN) and involves multiscale host-virus interactome analysis coupled to off-target drug predictions. Cell-based experimental assessment reveals several clinically-relevant drug repurposing candidates predicted by the in silico analyses to have antiviral activity against human coronavirus infection. In particular, we identify the MET inhibitor capmatinib as having potent and broad antiviral activity against several coronaviruses in a MET-independent manner, as well as novel roles for host cell proteins such as IRAK1/4 in supporting human coronavirus infection, which can inform further drug discovery studies.

Interplay of self-association and conformational flexibility in regulating protein function.

Many functional roles have been attributed to homodimers, the most common mode of protein self-association, notably in the regulation of enzymes, ion channels, transporters and transcription factors. Here we review findings that offer new insights into the different roles conformational flexibility plays in regulating homodimer function. Intertwined homodimers of two-domain proteins and their related family members display significant conformational flexibility, which translates into concerted motion between structural domains. This flexibility enables the corresponding proteins to regulate function across family members by modulating the spatial positions of key recognition surfaces of individual domains, to either maintain subunit interfaces, alter them or break them altogether, leading to a variety of functional consequences. Many proteins may exist as monomers but carry out their biological function as homodimers or higher-order oligomers. We present early evidence that in such systems homodimer formation primes the protein for its functional role. It does so by inducing elevated mobility in protein regions corresponding to the binding epitopes of functionally important ligands. In some systems this process acts as an allosteric response elicited by the self-association reaction itself. Our analysis furthermore suggests that the induced extra mobility likely facilitates ligand binding through the mechanism of conformational selection.This article is part of a discussion meeting issue 'Allostery and molecular machines'.

Comprehensive mapping of cystic fibrosis mutations to CFTR protein identifies mutation clusters and molecular docking predicts corrector binding site.

Cystic Fibrosis (CF) is caused by mutations in the CFTR gene, of which over 2000 have been reported to date. Mutations have yet to be analyzed in aggregate to assess their distribution across the tertiary structure of the CFTR protein, an approach that could provide valuable insights into the structure-function relationship of CFTR. In addition, the binding site of Class I correctors (VX-809, VX-661, and C18) is not well understood. In this study, exonic CFTR mutations and mutant allele frequencies described in 3 curated databases (ABCMdb, CFTR1, and CFTR2, comprising >130 000 data points) were mapped to 2 different structural models: a homology model of full-length CFTR protein in the open-channel state, and a cryo-electron microscopy core-structure of CFTR in the closed-channel state. Accordingly, residue positions of 6 high-frequency mutant CFTR alleles were found to spatially co-localize in CFTR protein, and a significant cluster was identified at the NBD1:ICL4 interdomain interface. In addition, immunoblotting confirmed the approximate binding site of Class I correctors, demonstrating that these small molecules act via a similar mechanism in vitro, and in silico molecular docking generated binding poses for their complex with the cryo-electron microscopy structure to suggest the putative corrector binding site is a multi-domain pocket near residues F374-L375. These results confirm the significance of interdomain interfaces as susceptible to disruptive mutation, and identify a putative corrector binding site. The structural pharmacogenomics approach of mapping mutation databases to protein models shows promise for facilitating drug discovery and personalized medicine for monogenetic diseases.

The Antiretroviral Agent Nelfinavir Mesylate: A Potential Therapy for Systemic Sclerosis.

OBJECTIVE: Transforming growth factor ß1 (TGFß1) is considered a key factor in fibrogenesis, and blocking TGFß1 signaling pathways diminishes fibrogenesis in animal models. The objective of this study was to determine whether nelfinavir mesylate (NFV), a drug approved by the Food and Drug Administration (FDA) for treating HIV infection, could be repurposed to treat pulmonary fibrosis in patients with systemic sclerosis (SSc). METHODS: Normal human lung, ventricular, and skin fibroblasts as well as lung fibroblasts from SSc patients were used to determine the effects of NFV on fibroblast-to-myofibroblast differentiation mediated by TGFß1. The efficacy of NFV was also evaluated in an animal model of SSc (bleomycin-induced pulmonary fibrosis). In addition, in silico analysis was performed to determine novel off-target effects of NFV. RESULTS: NFV inhibited TGFß1-mediated fibroblast-to-myofibroblast differentiation in lung fibroblasts through inhibition of the TGFß1 canonical pathway. NFV also inhibited differentiation of skin and ventricular fibroblasts and adipocyte precursors into myofibroblasts. Activation of the TGFß1/mechanistic target of rapamycin pathway inhibited autophagy in lung fibroblasts, favoring collagen deposition, and NFV counteracted this effect in a dose-dependent manner. Moreover, NFV significantly reduced lung injury and collagen deposition in an animal model of SSc. In silico analysis of NFV binding proteins revealed new putative beneficial mechanisms of action, consistent with known common pathways in fibrogenesis. CONCLUSION: NFV abrogates TGFß1-mediated fibroblast-to-myofibroblast differentiation and pulmonary fibrosis through off-target protein binding, a finding that supports consideration of this FDA-approved medication as an antifibrotic agent.

Structural coverage of the proteome for pharmaceutical applications.

Structure-based computational drug discovery efforts have traditionally focused on the structure of a single, well-known drug target. Important applications, such as target deconvolution and the analysis of polypharmacology, require proteome-scale molecular docking and have been inaccessible to structure-based in silico approaches. One important reason for this inaccessibility was that the structure of most proteins was not known. Lately, this 'structure gap' has been closing rapidly, and proteome-scale molecular docking seems within reach. Here, we survey the current state of structural coverage of the human genome and find that coverage is truly proteome-wide, both overall and in most pharmaceutically relevant categories of proteins. The time is right for structure-based approaches to target deconvolution and polypharmacology.

Computational proteome-wide screening predicts neurotoxic drug-protein interactome for the investigational analgesic BIA 10-2474.

The investigational compound BIA 10-2474, designed as a long-acting and reversible inhibitor of fatty acid amide hydrolase for the treatment of neuropathic pain, led to the death of one participant and hospitalization of five others due to intracranial hemorrhage in a Phase I clinical trial. Putative off-target activities of BIA 10-2474 have been suggested to be major contributing factors to the observed neurotoxicity in humans, motivating our study's proteome-wide screening approach to investigate its polypharmacology. Accordingly, we performed an in silico screen against 80,923 protein structures reported in the Protein Data Bank. The resulting list of 284 unique human interactors was further refined using target-disease association analyses to a subset of proteins previously linked to neurological, intracranial, inflammatory, hemorrhagic or clotting processes and/or diseases. Eleven proteins were identified as potential targets of BIA 10-2474, and the two highest-scoring proteins, Factor VII and thrombin, both essential blood-clotting factors, were predicted to be inhibited by BIA 10-2474 and suggest a plausible mechanism of toxicity. Once this small molecule becomes commercially available, future studies will be conducted to evaluate the predicted inhibitory effect of BIA 10-2474 on blood clot formation specifically in the brain.

The Landscape of Intertwined Associations in Homooligomeric Proteins.

We present an overview of the full repertoire of intertwined associations in homooligomeric proteins. This overview summarizes recent findings on the different categories of intertwined associations in known protein structures, their assembly modes, the properties of their interfaces, and their structural plasticity. Furthermore, the current body of knowledge on the so-called three-dimensional domain-swapped systems is reexamined in the context of the wider landscape of intertwined homooligomers, with a particular focus on the mechanistic aspects that underpin intertwined self-association processes in proteins. Insights gained from this integrated overview into the physical and biological roles of intertwining are highlighted.

Landscape of intertwined associations in multi-domain homo-oligomeric proteins.

This study charts the landscape of multi-domain protein structures that form intertwined homodimers by exchanging structural domains between subunits. A representative dataset of such homodimers was derived from the Protein Data Bank, and their structural and topological properties were compared to those of a representative set of non-intertwined homodimers. Most of the intertwined dimers form closed assemblies with head-to-tail arrangements, where the subunit interface involves contacts between dissimilar domains. In contrast, the non-intertwined dimers form preferentially head-to-head arrangements, where the subunit interface involves contacts between identical domains. Most of these contacts engage only one structural domain from each subunit, leaving the remaining domains free to form other associations. Remarkably, we find that multi-domain proteins closely related to the intertwined homodimers are significantly more likely than relatives of the non-intertwined versions to adopt alternative intramolecular domain arrangements. In ~40% of the intertwined dimers, the plasticity in domain arrangements among relatives affords maintenance of the head-to-head or head-to-tail topology and conservation of the corresponding subunit interface. This property seems to be exploited in several systems to regulate DNA binding. In ~58%, however, intramolecular domain re-arrangements are associated with changes in oligomeric states and poorly conserved interfaces among relatives. This time, the corresponding structural plasticity appears to be exploited by evolution to modulate function by switching between active and inactive states of the protein. Surprisingly, in total, only three systems were found to undergo the classical monomer to intertwined dimer conversion associated with three-dimensional domain swapping.

Intertwined associations in structures of homooligomeric proteins.

Intertwined homo-oligomers are complexes comprising identical protein subunits, where small segments or compact protein substructures (domains) are exchanged between the subunits. Using a formal definition of intertwined homo-oligomers, we survey the Protein Data Bank for all such complexes. Results show that intertwining occurs in 13,442 (24%) of all surveyed structures. A majority (∼72%) exchanges one contiguous chain segment of varying length. Another ∼10%, exchange structural domains, and the remaining ∼20% display complex intertwining topologies. Smaller proteins are more often intertwined, and intertwining is dominant in solution homodimers. These findings and analyses of various properties of the major category of intertwined complexes, their interfaces and quaternary context, support the physiological role of intertwining in promoting homooligomer stability. Furthermore, the number of different intertwining modes observed in families of related proteins is limited, and likely specific to the protein fold. These findings yield unique insights into the role of intertwining in homomeric association.