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Journal of Neuropathology and... Sep 2022Facial nerve injury results in degradation of the neuromuscular junction (NMJ) and blocks neurotransmission between the pre- and postsynaptic structures, which are...
Facial nerve injury results in degradation of the neuromuscular junction (NMJ) and blocks neurotransmission between the pre- and postsynaptic structures, which are separated by a synaptic cleft. Matrix metalloproteinases (MMPs), enzymes that degrade and modify the extracellular matrix, play critical roles in regulating NMJ remodeling. We previously demonstrated that MMP1, MMP2, MMP3, MMP7, and MMP9 are overexpressed in facial nerve-innervated orbicularis oris muscle after facial nerve injury in a rat model. In the present study, the MMP inhibitor prinomastat was administered to rats after facial nerve injury. The MMP levels, agrin expression, and muscle-specific kinase (MuSK) phosphorylation were evaluated. Variations in evoked electromyography (EEMG) amplitude were also recorded. Compared with the control group, MMP expression in the orbicularis oris after facial nerve injury was significantly reduced in the prinomastat group. Inhibition of MMP expression maintained agrin expression and MuSK phosphorylation; the NMJ morphology was also protected after the injury. Moreover, prinomastat treatment sustained EEMG amplitude and muscle tension after the injury. These findings indicate that inhibiting MMPs can protect the function and morphology of the NMJ and demonstrate the need for protection of the NMJ at early stages after facial nerve injury.
Topics: Agrin; Animals; Electromyography; Facial Muscles; Facial Nerve Injuries; Matrix Metalloproteinase 1; Matrix Metalloproteinase 2; Matrix Metalloproteinase 3; Matrix Metalloproteinase 7; Matrix Metalloproteinase 9; Matrix Metalloproteinase Inhibitors; Muscle Tonus; Organic Chemicals; Rats
PubMed: 35656867
DOI: 10.1093/jnen/nlac041 -
Toxicon: X Jun 2022Snakebite envenoming affects more than 250,000 people annually in sub-Saharan Africa. Envenoming by (boomslang) results in venom-induced consumption coagulopathy...
Snakebite envenoming affects more than 250,000 people annually in sub-Saharan Africa. Envenoming by (boomslang) results in venom-induced consumption coagulopathy (VICC), whereby highly abundant prothrombin-activating snake venom metalloproteinases (SVMPs) consume clotting factors and deplete fibrinogen. The only available treatment for envenoming is the monovalent SAIMR Boomslang antivenom. Treatment options are urgently required because this antivenom is often difficult to source and, at US$6000/vial, typically unaffordable for most snakebite patients. We therefore investigated the and preclinical efficacy of four SVMP inhibitors to neutralise the effects of venom; the matrix metalloproteinase inhibitors marimastat and prinomastat, and the metal chelators dimercaprol and DMPS The venom of exhibited an SVMP-driven procoagulant phenotype . Marimastat and prinomastat demonstrated equipotent inhibition of the SVMP-mediated procoagulant activity of the venom , whereas dimercaprol and DMPS showed considerably lower potency. However, when tested in preclinical murine models of envenoming using mixed sex CD1 mice, DMPS and marimastat demonstrated partial protection against venom lethality, demonstrated by prolonged survival times of experimental animals, whereas dimercaprol and prinomastat failed to confer any protection at the doses tested. The preclinical results presented here demonstrate that DMPS and marimastat show potential as novel small molecule-based therapeutics for snakebite envenoming. These two drugs have been previously shown to be effective against VICC in preclinical models, and thus we conclude that marimastat and DMPS should be further explored as potentially valuable early intervention therapeutics to broadly treat VICC following snakebite envenoming in sub-Saharan Africa.
PubMed: 35321116
DOI: 10.1016/j.toxcx.2022.100118 -
MBio Aug 2022The ongoing global vaccination program to prevent SARS-CoV-2 infection, the causative agent of COVID-19, has had significant success. However, recently, virus variants...
The ongoing global vaccination program to prevent SARS-CoV-2 infection, the causative agent of COVID-19, has had significant success. However, recently, virus variants that can evade the immunity in a host achieved through vaccination have emerged. Consequently, new therapeutic agents that can efficiently prevent infection from these new variants, and hence COVID-19 spread, are urgently required. To achieve this, extensive characterization of virus-host cell interactions to identify effective therapeutic targets is warranted. Here, we report a cell surface entry pathway of SARS-CoV-2 that exists in a cell type-dependent manner and is TMPRSS2 independent but sensitive to various broad-spectrum metalloproteinase inhibitors such as marimastat and prinomastat. Experiments with selective metalloproteinase inhibitors and gene-specific small interfering RNAS (siRNAs) revealed that a disintegrin and metalloproteinase 10 (ADAM10) is partially involved in the metalloproteinase pathway. Consistent with our finding that the pathway is unique to SARS-CoV-2 among highly pathogenic human coronaviruses, both the furin cleavage motif in the S1/S2 boundary and the S2 domain of SARS-CoV-2 spike protein are essential for metalloproteinase-dependent entry. In contrast, the two elements of SARS-CoV-2 independently contributed to TMPRSS2-dependent S2 priming. The metalloproteinase pathway is involved in SARS-CoV-2-induced syncytium formation and cytopathicity, leading us to theorize that it is also involved in the rapid spread of SARS-CoV-2 and the pathogenesis of COVID-19. Thus, targeting the metalloproteinase pathway in addition to the TMPRSS2 and endosomal pathways could be an effective strategy by which to cure COVID-19 in the future. To develop effective therapeutics against COVID-19, it is necessary to elucidate in detail the infection mechanism of the causative agent, SARS-CoV-2. SARS-CoV-2 binds to the cell surface receptor ACE2 via the spike protein, and then the spike protein is cleaved by host proteases to enable entry. Here, we found that the metalloproteinase-mediated pathway is important for SARS-CoV-2 infection in addition to the TMPRSS2-mediated pathway and the endosomal pathway. The metalloproteinase-mediated pathway requires both the prior cleavage of spike into two domains and a specific sequence in the second domain, S2, conditions met by SARS-CoV-2 but lacking in the related human coronavirus SARS-CoV. Besides the contribution of metalloproteinases to SARS-CoV-2 infection, inhibition of metalloproteinases was important in preventing cell death, which may cause organ damage. Our study provides new insights into the complex pathogenesis unique to COVID-19 and relevant to the development of effective therapies.
Topics: COVID-19; Humans; Metalloproteases; SARS-CoV-2; Serine Endopeptidases; Spike Glycoprotein, Coronavirus; Virus Internalization
PubMed: 35708281
DOI: 10.1128/mbio.00519-22 -
Frontiers in Immunology 2021African spitting cobras are unique among cobras for their potent anticoagulant venom activity arising from strong inhibition of Factor Xa. This anticoagulant effect is...
African spitting cobras are unique among cobras for their potent anticoagulant venom activity arising from strong inhibition of Factor Xa. This anticoagulant effect is exerted by venom phospholipase A (Group I PLA) toxins whose activity contributes to the lethality of these species. This anticoagulant toxicity is particularly problematic as it is not neutralized by current antivenoms. Previous work demonstrated this trait for , , and The present work builds upon previous research by testing across the full taxonomical range of African spitting cobras, demonstrating that , , and are also potently anticoagulant through the inhibition of Factor Xa, and therefore the amplification of potent anticoagulant activity occurred at the base of the African spitting cobra radiation. Previous work demonstrated that the enzyme-inhibitor varespladib was able to neutralize this toxic action for , , and venoms. The current work demonstrates that varespladib was also able to neutralize , , and . Thus varespladib is shown to have broad utility across the full range of African spitting cobras. In addition, we examined the cross-reactivity of the metalloprotease inhibitor prinomastat, which had been previously intriguingly indicated as being capable of neutralizing viperid venom PLA (Group II PLA). In this study prinomastat inhibited the FXa-inhibiting PLA toxins of all the African spitting cobras at the same concentration at which it has been shown to inhibit metalloproteases, and thus was comparably effective in its cross-reactivity. In addition we showed that the metalloprotease-inhibitor marimastat was also able to cross-neutralize PLA but less effectively than prinomastat. Due to logistical (cold-chain requirement) and efficacy (cross-reactivity across snake species) limitations of traditional antivenoms, particularly in developing countries where snakebite is most common, these small molecule inhibitors (SMIs) might hold great promise as initial, field-based, treatments for snakebite envenoming as well as addressing fundamental limitations of antivenom in the clinical setting where certain toxin effects are unneutralized.
Topics: Acetates; Africa; Animals; Antivenins; Enzyme Inhibitors; Hydroxamic Acids; In Vitro Techniques; Indoles; Keto Acids; Naja; Organic Chemicals; Snake Bites; Snake Venoms
PubMed: 34691069
DOI: 10.3389/fimmu.2021.752442 -
Blood Advances May 2022Acute myeloid leukemia (AML) is a blood cancer of the myeloid lineage. Its prognosis remains poor, highlighting the need for new therapeutic and precision medicine...
Acute myeloid leukemia (AML) is a blood cancer of the myeloid lineage. Its prognosis remains poor, highlighting the need for new therapeutic and precision medicine approaches. AML symptoms often include cytopenias linked to loss of healthy hematopoietic stem and progenitor cells (HSPCs). The mechanisms behind HSPC decline are complex and still poorly understood. Here, intravital microscopy (IVM) of a well-established experimental model of AML allows direct observation of the interactions between healthy and malignant cells in the bone marrow (BM), suggesting that physical dislodgment of healthy cells by AML through damaged vasculature may play an important role. Multiple matrix metalloproteinases (MMPs), known to remodel extracellular matrix, are expressed by AML cells and the BM microenvironment. We reason MMPs could be involved in cell displacement and vascular leakiness; therefore, we evaluate the therapeutic potential of MMP pharmacological inhibition using the broad-spectrum inhibitor prinomastat. IVM analyses of prinomastat-treated mice reveal reduced vascular permeability and healthy cell clusters in circulation and lower AML infiltration, proliferation, and cell migration. Furthermore, treated mice have increased retention of healthy HSPCs in the BM and increased survival following chemotherapy. Analysis of a human AML transcriptomic database reveals widespread MMP deregulation, and human AML cells show susceptibility to MMP inhibition. Overall, our results suggest that MMP inhibition could be a promising complementary therapy to reduce AML growth and limit HSPC loss and BM vascular damage caused by MLL-AF9 and possibly other AML subtypes.
Topics: Animals; Bone Marrow; Hematopoietic Stem Cells; Leukemia, Myeloid, Acute; Metalloproteases; Mice; Prognosis; Tumor Microenvironment
PubMed: 35157757
DOI: 10.1182/bloodadvances.2021004321 -
Toxins Jul 2022Within Neotropical pit-vipers, the Mexican/Central-American clade consisting of , , , and is a wide-ranging, morphologically and ecologically diverse group of snakes....
Within Neotropical pit-vipers, the Mexican/Central-American clade consisting of , , , and is a wide-ranging, morphologically and ecologically diverse group of snakes. Despite their prevalence, little is known of the functional aspects of their venoms. This study aimed to fill the knowledge gap regarding coagulotoxic effects and to examine the potential of different therapeutic approaches. As a general trait, the venoms were shown to be anticoagulant but were underpinned by diverse biochemical actions. Pseudo-procoagulant activity (i.e., thrombin-like), characterized by the direct cleavage of fibrinogen to form weak fibrin clots, was evident for , , and In contrast, other venoms cleaved fibrinogen in a destructive (non-clotting) manner, with and being the most potent. In addition to actions on fibrinogen, clotting enzymes were also inhibited. FXa was only weakly inhibited by most species, but and were extremely strong in their inhibitory action. Other clotting enzymes were more widely inhibited by diverse species spanning the full taxonomical range, but in each case, there were species that had these traits notably amplified relatively to the others. and were the most potent amongst those that inhibited the formation of the prothrombinase complex and were also amongst the most potent inhibitors of Factor XIa. While most species displayed only low levels of thrombin inhibition, potently inhibited this clotting factor. The regional polyvalent antivenom produced by Instituto Picado Clodomiro was tested and was shown to be effective against the diverse anticoagulant pathophysiological effects. In contrast to the anticoagulant activities of the other species, was uniquely procoagulant through the activation of Factor VII and Factor XII. This viperid species is the first snake outside of the elapid snake clade to be shown to activate FVII and the first snake venom of any kind to activate FXII. Interestingly, while small-molecule metalloprotease inhibitors prinomastat and marimastat demonstrated the ability to prevent the procoagulant toxicity of , neither ICP antivenom nor inhibitor DMPS showed this effect. The extreme variation among the snakes here studied underscores how venom is a dynamic trait and how this can shape clinical outcomes and influence evolving treatment strategies.
Topics: Animals; Anticoagulants; Antivenins; Crotalid Venoms; Crotalinae; Elapid Venoms; Elapidae; Fibrinogen; Snake Venoms; Thrombin; Viperidae
PubMed: 35893753
DOI: 10.3390/toxins14080511 -
Toxins Dec 2022The African viperid snake genera , and are closely related, similar in size, but occupy extremely divergent ecological niches (arboreal in tropical rainforests,...
The African viperid snake genera , and are closely related, similar in size, but occupy extremely divergent ecological niches (arboreal in tropical rainforests, fossorial in deserts, and swamp-dwelling, respectively). Their venoms have not previously been subjected to comparative analyses for their action upon the coagulation of blood, most notably with significant data deficiencies from and In contrast, the closely related genus is well-documented as capable of producing potent procoagulant effects. In light of this, we set out to compare the coagulotoxic actions of , , , , , , , , and and explore potential pharmacological interventions to reestablish normal blood coagulation. All venoms displayed extremely potent procoagulant effects, over twice as fast as the most potent reported to date. Although is used in the immunising mixture of two different regionally available antivenoms (Inoserp-MENA with , , and Saudi Arabian polyvalent with ), none of the other species in this study are included in the immunising mixture of any antivenom. Notably, all the species were only neutralised by the Inoserp-MENA antivenom. venom was not neutralised well by the Saudi Arabian antivenom, with the low levels of recognition for any of the venoms suggesting a strong regional variation in the venom of this species, as the venom tested was of African (Tunisian) origin versus Saudi locality used in that antivenom's production. The other antivenoms (Micropharm EchiTAbG, ICP EchiTAb-Plus-ICP, Inosan Inoserp Pan-Africa, Premium Serums PANAF Sub-Sahara Africa, South African Vaccine Producers , South African Vaccine Producers Polyvalent) all displayed trivial-to-no ability to neutralise the procoagulant toxicity of any of the , or venoms. Comparative testing of the enzyme inhibitors DMPS, marimastat, and prinomastat, revealed a very potent neutralising capacity of marimastat, with prinomastat showing lower but still significant potency at the same molar concentration, while a 5× molar concentration of DMPS had no apparent effect on procoagulant venom effects normalized by the other inhibitors. These results and methods contribute to the body of knowledge of potential clinical effects and data necessary for evidence-based advancement of clinical management strategies.
Topics: Animals; Humans; Antivenins; Viperidae; Saudi Arabia; Viper Venoms; Africa South of the Sahara; Enzyme Inhibitors; African People; Snake Bites
PubMed: 36548733
DOI: 10.3390/toxins14120836 -
Frontiers in Microbiology 2024HIV-1 gp120 glycan binding to C-type lectin adhesion receptor L-selectin/CD62L on CD4 T cells facilitates viral attachment and entry. Paradoxically, the adhesion...
HIV-1 gp120 glycan binding to C-type lectin adhesion receptor L-selectin/CD62L on CD4 T cells facilitates viral attachment and entry. Paradoxically, the adhesion receptor impedes HIV-1 budding from infected T cells and the viral release requires the shedding of CD62L. To systematically investigate CD62L-shedding mediated viral release and its potential inhibition, we screened compounds specific for serine-, cysteine-, aspartyl-, and Zn-dependent proteases for CD62L shedding inhibition and found that a subclass of Zn-metalloproteinase inhibitors, including BB-94, TAPI, prinomastat, GM6001, and GI25423X, suppressed CD62L shedding. Their inhibition of HIV-1 infections correlated with enzymatic suppression of both ADAM10 and 17 activities and expressions of these ADAMs were transiently induced during the viral infection. These metalloproteinase inhibitors are distinct from the current antiretroviral drug compounds. Using immunogold labeling of CD62L, we observed association between budding HIV-1 virions and CD62L by transmission electron microscope, and the extent of CD62L-tethering of budding virions increased when the receptor shedding is inhibited. Finally, these CD62L shedding inhibitors suppressed the release of HIV-1 virions by CD4 T cells of infected individuals and their virion release inhibitions correlated with their CD62L shedding inhibitions. Our finding reveals a new therapeutic approach targeted at HIV-1 viral release.
PubMed: 38572241
DOI: 10.3389/fmicb.2024.1385775 -
Frontiers in Immunology 2021Palearctic vipers are medically significant snakes in the genera , and which occur throughout Europe, Central Asia, Near and Middle East. While the ancestral condition... (Comparative Study)
Comparative Study
Palearctic vipers are medically significant snakes in the genera , and which occur throughout Europe, Central Asia, Near and Middle East. While the ancestral condition is that of a small-bodied, lowland species, extensive diversification has occurred in body size, and niche specialization. Using 27 venom samples and a panel of coagulation assays, we evaluated the relative coagulotoxic potency of Palearctic viper venoms and compared their neutralization by three antivenoms (Insoserp Europe, VIPERFAV and ViperaTAb) and two metalloprotease inhibitors (prinomastat and DMPS). We show that variation in morphology parallels variation in the Factor X activating procoagulant toxicity, with the three convergent evolutions of larger body sizes ( genus, genus, and uniquely within the genus) were each accompanied by a significant increase in procoagulant potency. In contrast, the two convergent evolutions of high altitude specialization (the genus and uniquely within the genus) were each accompanied by a shift away from procoagulant action, with the species being particularly potently anticoagulant. Inoserp Europe and VIPERFAV antivenoms were both effective against a broad range of species, with Inoserp able to neutralize additional species relative to VIPERFAV, reflective of its more complex antivenom immunization mixture. In contrast, ViperaTAb was extremely potent in neutralizing but, reflective of this being a monovalent antivenom, it was not effective against other species. The enzyme inhibitor prinomastat efficiently neutralized the metalloprotease-driven Factor X activation of the procoagulant venoms. In contrast, DMPS (2,3-dimercapto-1-propanesulfonic acid), which as been suggested as another potential treatment option in the absence of antivenom, DMPS failed against all venoms tested. Overall, our results highlight the evolutionary variations within Palearctic vipers and help to inform clinical management of viper envenomation.
Topics: Animals; Antivenins; Blood Coagulation; Blood Coagulation Tests; Evolution, Molecular; Humans; Immunoglobulin Fab Fragments; Matrix Metalloproteinase Inhibitors; Organic Chemicals; Snake Bites; Species Specificity; Time Factors; Unithiol; Viper Venoms; Viperidae
PubMed: 34177943
DOI: 10.3389/fimmu.2021.688802 -
The International Journal of Biological... 1999Matrix metalloproteases (MMPs) are a family of structurally related enzymes that are capable of degrading proteins of the extracellular matrix. These enzymes play a role... (Review)
Review
Matrix metalloproteases (MMPs) are a family of structurally related enzymes that are capable of degrading proteins of the extracellular matrix. These enzymes play a role in tissue remodelling associated with both physiological and pathogenic processes. A high expression of MMPs is associated with cancer malignancy: it is related to the tumor's ability to metastasize and to the process of angiogenesis. Treatment with MMP inhibitors alone or in combination with cytotoxic therapy is an interesting novel approach to control tumor progression. The expected mechanism of action of these compounds and the difference in side effects compared to cytotoxic drugs make the definition of endpoints and the assessment of response difficult. Furthermore, it is not yet clear whether tumor vascularization or, more specifically, MMP expression/activation should be a criterion of eligibility for this kind of treatment. This review provides an overview of the characteristics of MMPs and their role in tumor progression, metastasis and angiogenesis. Preclinical and clinical studies with synthetic MMP inhibitors are described. The presence of MMPs in biological fluids of patients and their use in prognostic evaluation and in determining the efficacy of treatment with MMP inhibitors is discussed.
Topics: Animals; Antineoplastic Agents; Azepines; Humans; Hydroxamic Acids; Matrix Metalloproteinase Inhibitors; Organic Chemicals; Phenylalanine; Protease Inhibitors; Thiophenes
PubMed: 10669951
DOI: 10.1177/172460089901400406