scholarly journals Antimalarial Activity of Allicin, a Biologically Active Compound from Garlic Cloves

2006 ◽  
Vol 50 (5) ◽  
pp. 1731-1737 ◽  
Author(s):  
Alida Coppi ◽  
Melissa Cabinian ◽  
David Mirelman ◽  
Photini Sinnis
Keyword(s):  
Life Cycle ◽  
Protease Inhibitor ◽  
Cysteine Protease ◽  
Malaria Infection ◽  
Drug Targets ◽  
Host Cells ◽  
Cysteine Protease Inhibitor ◽  
New Drug

ABSTRACT The incidence of malaria is increasing, and there is an urgent need to identify new drug targets for both prophylaxis and chemotherapy. Potential new drug targets include Plasmodium proteases that play critical roles in the parasite life cycle. We have previously shown that the major surface protein of Plasmodium sporozoites, the circumsporozoite protein (CSP), is proteolytically processed by a parasite-derived cysteine protease, and this processing event is temporally associated with sporozoite invasion of host cells. E-64, a cysteine protease inhibitor, inhibits CSP processing and prevents invasion of host cells in vitro and in vivo. Here we tested allicin, a cysteine protease inhibitor found in garlic extracts, for its ability to inhibit malaria infection. At low concentrations, allicin was not toxic to either sporozoites or mammalian cells. At these concentrations, allicin inhibited CSP processing and prevented sporozoite invasion of host cells in vitro. In vivo, mice injected with allicin had decreased Plasmodium infections compared to controls. When sporozoites were treated with allicin before injection into mice, malaria infection was completely prevented. We also tested allicin on erythrocytic stages and found that a 4-day regimen of allicin administered either orally or intravenously significantly decreased parasitemias and increased the survival of infected mice by 10 days. Together, these experiments demonstrate that the same cysteine protease inhibitor can target two different life cycle stages in the vertebrate host.

scholarly journals Effect of phenyl vinyl sulphone cysteine protease inhibitor on Schistosoma mansoni: in vitro and in vivo experimental studies

2017 ◽  
Vol 41 (4) ◽  
pp. 1049-1058
Author(s):  
Manal Salah El-Din Mahmoud ◽  
Ayman Nabil Ibrahim ◽  
Abeer Fathy Badawy ◽  
Nourhan Mohamed Abdelmoniem
Keyword(s):  
Protease Inhibitor ◽  
Schistosoma Mansoni ◽  
Cysteine Protease ◽  
Experimental Studies ◽  
Cysteine Protease Inhibitor ◽  
Phenyl Vinyl

STEM-15. SerpinB3 DRIVES CANCER STEM CELL SURVIVAL IN GLIOBLASTOMA

2021 ◽  
Vol 23 (Supplement_6) ◽  
pp. vi24-vi24
Author(s):  
Adam Lauko ◽  
Soumya M Turaga ◽  
Josephine Volovetz ◽  
Defne Bayik ◽  
Shideng Bao ◽  
...  
Keyword(s):  
Cell Death ◽  
Protease Inhibitor ◽  
Cysteine Protease ◽  
Xenograft Model ◽  
Standard Of Care ◽  
Therapeutic Interventions ◽  
Cysteine Protease Inhibitor ◽  
Orthotopic Xenograft Model

Abstract Despite therapeutic interventions for glioblastoma (GBM), self-renewing, therapy-resistant populations of cells referred to as cancer stem cells (CSCs) drive recurrence. Previously, we identified the unique expression of junctional adhesion molecule-A (JAM-A) on CSCs and demonstrated that JAM-A is both necessary and sufficient for self-renewal and tumor growth. Moreover, we determined that JAM-A signals via Akt in GBM CSCs to sustain pluripotency transcription factor activity; however, the entire signaling network has yet to be fully elucidated. To further delineate this pathway, we immunoprecipitated JAM-A from patient-derived GBM CSCs and performed mass spectrometry to determine JAM-A binding proteins. This led to the identification of the cysteine protease inhibitor SerpinB3 as a putative JAM-A binding partner. Using in vitro CSC functional assays, we show that SerpinB3 is necessary for CSC maintenance and survival. In an in vivo orthotopic xenograft model, knockdown of SerpinB3 extended survival. Mechanistically, knockdown of SerpinB3 led to decreased expression of TGF-β, Myc, WNT, and Notch signaling, known regulators of the CSC state. Additionally, knockdown of SerpinB3 increases susceptibility to radiation therapy. SerpinB3 is essential for buffering cells against cathepsin-mediated cell death, and we found that elevated lysosomal membrane permeability after radiation leads to cathepsin release into the cytoplasm. As a result, SerpinB3 knockdown cells have a diminished capacity to inhibit cathepsin-driven cell death after radiation. The addition of the cathepsin inhibitor E64D partially rescues the SerpinB3 knockdown, however, SerpinB3 mutants that are unable to inhibit cathepsins fail to do the same. Taken together, our findings, identify a novel GBM CSC-specific survival mechanism involving a previously uninvestigated cysteine protease inhibitor, SerpinB3, and provide a potential target to increase the efficacy of standard of care GBM therapies against therapy-resistant CSCs.

scholarly journals Nippocystatin, a Cysteine Protease Inhibitor fromNippostrongylus brasiliensis, Inhibits Antigen Processing and Modulates Antigen-Specific Immune Response

2001 ◽  
Vol 69 (12) ◽  
pp. 7380-7386 ◽  
Author(s):  
Teruki Dainichi ◽  
Yoichi Maekawa ◽  
Kazunari Ishii ◽  
Tianqian Zhang ◽  
Baher Fawzy Nashed ◽  
...  
Keyword(s):  
Immune System ◽  
Protease Inhibitor ◽  
Cysteine Protease ◽  
Antigen Processing ◽  
Cysteine Proteases ◽  
Cysteine Protease Inhibitor ◽  
Host Immune System ◽  
Nippostrongylus Brasiliensis

ABSTRACT During infection, parasites evade the host immune system by modulating or exploiting the immune system; e.g., they suppress expression of major histocompatibility complex class II molecules or secrete cytokine-like molecules. However, it is not clear whether helminths disturb the immune responses of their hosts by controlling the antigen-processing pathways of the hosts. In this study, we identified a new cysteine protease inhibitor, nippocystatin, derived from excretory-secretory (ES) products of an intestinal nematode,Nippostrongylus brasiliensis. Nippocystatin, which belongs to cystatin family 2, consists of 144 amino acids and is secreted as a 14-kDa mature form. In vivo treatment of ovalbumin (OVA)-immunized mice with recombinant nippocystatin (rNbCys) profoundly suppressed OVA-specific proliferation of splenocytes but not non-antigen-specific proliferation of splenocytes. OVA-specific cytokine production was also greatly suppressed in rNbCys-treated mice. Although the serum levels of both OVA-specific immunoglobulin G1 (IgG1) and IgG2a were not affected by rNbCys treatment, OVA-specific IgE was preferentially downregulated in rNbCys-treated mice. In vitro rNbCys inhibited processing of OVA by lysosomal cysteine proteases from the spleens of mice. Mice with anti-nippocystatin antibodies became partially resistant to infection with N. brasiliensis. Based on these findings, N. brasiliensis appears to skillfully evade host immune systems by secreting nippocystatin, which modulates antigen processing in antigen-presenting cells of hosts.

Identification, characterization and localization of chagasin, a tight-binding cysteine protease inhibitor inTrypanosoma cruzi

2001 ◽  
Vol 114 (21) ◽  
pp. 3933-3942 ◽  
Author(s):  
Ana C. S. Monteiro ◽  
Magnus Abrahamson ◽  
Ana P. C. A. Lima ◽  
Marcos A. Vannier-Santos ◽  
Julio Scharfstein
Keyword(s):  
Cell Surface ◽  
Protease Inhibitor ◽  
Cysteine Protease ◽  
Mammalian Cells ◽  
Tight Binding ◽  
Cysteine Proteases ◽  
Host Cells ◽  
Cysteine Protease Inhibitor ◽  
Reversible Inhibitor ◽  
Mammalian Host

Lysosomal cysteine proteases from mammalian cells and plants are regulated by endogenous tight-binding inhibitors from the cystatin superfamily. The presence of cystatin-like inhibitors in lower eukaryotes such as protozoan parasites has not yet been demonstrated, although these cells express large quantities of cysteine proteases and may also count on endogenous inhibitors to regulate cellular proteolysis. Trypanosoma cruzi, the causative agent of Chagas’ heart disease, is a relevant model to explore this possibility because these intracellular parasites rely on their major lysosomal cysteine protease (cruzipain) to invade and multiply in mammalian host cells. Here we report the isolation, biochemical characterization, developmental stage distribution and subcellular localization of chagasin, an endogenous cysteine protease inhibitor in T. cruzi. We used high temperature induced denaturation to isolate a heat-stable cruzipain-binding protein (apparent molecular mass, 12 kDa) from epimastigote lysates. This protein was subsequently characterized as a tight-binding and reversible inhibitor of papain-like cysteine proteases. Immunoblotting indicated that the expression of chagasin is developmentally regulated and inversely correlated with that of cruzipain. Gold-labeled antibodies localized chagasin to the flagellar pocket and cytoplasmic vesicles of trypomastigotes and to the cell surface of amastigotes. Binding assays performed by probing living parasites with fluorescein (FITC)-cruzipain or FITC-chagasin revealed the presence of both inhibitor and protease at the cell surface of amastigotes. The intersection of chagasin and cruzipain trafficking pathways may represent a checkpoint for downstream regulation of proteolysis in trypanosomatid protozoa.

scholarly journals Organoids and Bioengineered Intestinal Models: Potential Solutions to the Cryptosporidium Culturing Dilemma

2020 ◽  
Vol 8 (5) ◽  
pp. 715 ◽  
Author(s):  
Samantha Gunasekera ◽  
Alireza Zahedi ◽  
Mark O’Dea ◽  
Brendon King ◽  
Paul Monis ◽  
...  
Keyword(s):  
Life Cycle ◽  
Host Cells ◽  
Protozoan Parasites ◽  
Future Directions ◽  
In Vivo Models ◽  
Severe Diarrhea ◽  
Transformed Cell Lines ◽  
Gnotobiotic Piglets

Cryptosporidium is a major cause of severe diarrhea-related disease in children in developing countries, but currently no vaccine or effective treatment exists for those who are most at risk of serious illness. This is partly due to the lack of in vitro culturing methods that are able to support the entire Cryptosporidium life cycle, which has led to research in Cryptosporidium biology lagging behind other protozoan parasites. In vivo models such as gnotobiotic piglets are complex, and standard in vitro culturing methods in transformed cell lines, such as HCT-8 cells, have not been able to fully support fertilization occurring in vitro. Additionally, the Cryptosporidium life cycle has also been reported to occur in the absence of host cells. Recently developed bioengineered intestinal models, however, have shown more promising results and are able to reproduce a whole cycle of infectivity in one model system. This review evaluates the recent advances in Cryptosporidium culturing techniques and proposes future directions for research that may build upon these successes.

Molecular characterization of neuroendocrine prostate cancer (NEPC) and identification of new drug targets.

2011 ◽  
Vol 29 (7_suppl) ◽  
pp. 19-19 ◽  
Author(s):  
H. Beltran ◽  
D. Rickman ◽  
K. Park ◽  
A. Sboner ◽  
T. Macdonald ◽  
...  
Keyword(s):  
Prostate Cancer ◽  
Drug Targets ◽  
Kinase Activity ◽  
Aurora Kinase ◽  
Large Cohort ◽  
Small Subset ◽  
Aurora Kinases ◽  
New Drug

19 Background: NEPC is an aggressive variant of prostate cancer that can arise de novo or from existing prostate adenocarcinoma (PCA). We sought to better understand the molecular transformation of NEPC and identify new drug targets. Methods: We used Next Generation RNA sequencing and oligonucleotide arrays to profile 7 NEPC, 30 PCA, 5 benign prostate (BEN), and validated findings on tumors from a large cohort of patients (30 NEPC, 118 PCA, 30 BEN) using IHC and FISH. Functional studies were performed using NCI-H660 (NEPC), VCaP and LnCaP (PCA), RWPE (BEN). Results: ERG rearrangement was present in 47% of NEPC, but ERG protein expression was absent and corresponded directly with lack of AR expression. 936/25932 genes were differentially expressed in NEPC versus PCA (P<0.001). Aurora kinases (AURKA, AURKB) and N-myc (MYCN) were overexpressed in NEPC (P<0.001) and AURKA and MYCN amplified. Using IHC and FISH, we validated these findings on a large cohort and found majority (>80%) of NEPC showed Aurora overexpression, 35% had AURKA and MYCN amplification. A small subset of PCA (5%) and no BEN were positive. Transfection of MYCN induced AURKA expression and kinase activity in vitro, and MYCN or AURKA could induce expression of neuroendocrine (NE) markers (SYP, NSE). After validating NCI-H660 as model of NEPC, we observed dramatic and enhanced in vitro and in vivo sensitivity to the Aurora kinase inhibitor PHA-739358 in NCI-H660 compared to minimal to no effect in LnCaP and VCaP. Phospho-H3 expression, a downstream marker of Aurora kinase activity, was inhibited in the treated NCI-H660 and not in PCA. Notably, NE marker expression was also suppressed in the treated NCI-H660 xenografts, again supporting a role of Aurora kinase in modulating the NE phenotype. Conclusions: There is likely clonal origin of NEPC from PCA (with ERG fusion positivity seen in both), but ERG expression is limited to PCA and driven by AR signaling. We discovered significant overexpression and gene amplification of Aurora kinases and N-myc in NEPC and a small subset of PCA, and evidence that that they cooperate and induce a NE phenotype in prostate cells. In vitro and in vivo data confirms that these are novel drug targets for NEPC. No significant financial relationships to disclose.

scholarly journals Synthetic lethality-based prediction of anti-SARS-CoV-2 targets

2021 ◽  
Author(s):  
Lipika R. Pal ◽  
Kuoyuan Cheng ◽  
Nishanth U Nair ◽  
Laura Martin-Sancho ◽  
Sanju Sinha ◽  
...  
Keyword(s):  
Drug Targets ◽  
Synthetic Lethality ◽  
Host Cells ◽  
Synthetic Lethal ◽  
Viral Propagation ◽  
In Vivo Testing ◽  
Altered Gene ◽  
Host Genes

Novel strategies are needed to identify drug targets and treatments for the COVID-19 pandemic. The altered gene expression of virus-infected host cells provides an opportunity to specifically inhibit viral propagation via targeting the synthetic lethal (SL) partners of such altered host genes. Pursuing this antiviral strategy, here we comprehensively analyzed multiple in vitro and in vivo bulk and single-cell RNA-sequencing datasets of SARS-CoV-2 infection to predict clinically relevant candidate antiviral targets that are SL with altered host genes. The predicted SL-based targets are highly enriched for infected cell inhibiting genes reported in four SARS-CoV-2 CRISPR-Cas9 genome-wide genetic screens. Integrating our predictions with the results of these screens, we further selected a focused subset of 26 genes that we experimentally tested in a targeted siRNA screen using human Caco-2 cells. Notably, as predicted, knocking down these targets reduced viral replication and cell viability only under the infected condition without harming non-infected cells. Our results are made publicly available, to facilitate their in vivo testing and further validation.

scholarly journals A Phytophthora palmivora Extracellular Cystatin-Like Protease Inhibitor Targets Papain to Contribute to Virulence on Papaya

2018 ◽  
Vol 31 (3) ◽  
pp. 363-373 ◽  
Author(s):  
Rebecca Gumtow ◽  
Dongliang Wu ◽  
Janice Uchida ◽  
Miaoying Tian
Keyword(s):  
Protease Inhibitor ◽  
Protease Inhibitors ◽  
Cysteine Protease ◽  
Gene Editing ◽  
Sequence Data ◽  
Cysteine Protease Inhibitor ◽  
Phytophthora Palmivora ◽  
Cysteine Protease Inhibitors ◽  
Successful Infection

Papaya fruits, stems, and leaves are rich in papain, a cysteine protease that has been shown to mediate plant defense against pathogens and insects. Yet the oomycete Phytophthora palmivora is a destructive pathogen that infects all parts of papaya plants, suggesting that it has evolved cysteine protease inhibitors to inhibit papain to enable successful infection. Out of five putative extracellular cystatin-like cysteine protease inhibitors (PpalEPICs) from P. palmivora transcriptomic sequence data, PpalEPIC8 appeared to be unique to P. palmivora and was highly induced during infection of papaya. Purified recombinant PpalEPIC8 strongly inhibited papain enzyme activity, suggesting that it is a functional cysteine protease inhibitor. Homozygous PpalEPIC8 mutants were generated using CRISPR/Cas9-mediated gene editing via Agrobacterium-mediated transformation (AMT). Increased papain sensitivity of in-vitro growth and reduced pathogenicity during infection of papaya fruits were observed for the mutants compared with the wild-type strain, suggesting that PpalEPIC8, indeed, plays a role in P. palmivora virulence by inhibiting papain. This study provided genetic evidence demonstrating that plant-pathogenic oomycetes secrete cystatins as important weapons to invade plants. It also established an effective gene-editing system for P. palmivora by the combined use of CRISPR/Cas9 and AMT, which is expected to be applicable to other oomycetes.

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