scholarly journals Effect of melanin biosynthesis inhibition in the antifungal susceptibility of chromoblastomycosis agents

2021 ◽  
Author(s):  
Daiane Heidrich ◽  
Danielle Machado Pagani ◽  
Alessandra Koehler ◽  
Karine de Oliveira Alves ◽  
Maria Lúcia Scroferneker
Keyword(s):  
Statistical Tests ◽  
Antifungal Susceptibility ◽  
Specific Inhibitor ◽  
Disease Treatment ◽  
Melanin Biosynthesis ◽  
Inoculum Preparation ◽  
Melanin Inhibition ◽  
Susceptibility Assay ◽  
Increased Susceptibility

Chromoblastomycosis (CBM) is a chronic subcutaneous infection caused by genera of melanized fungi: Fonsecaea, Cladophialophora, Phialophora, Exophiala and Rhinocladiella. Melanin is a virulence factor known to influence antifungal susceptibility. A specific inhibitor of melanin biosynthesis is tricyclazole. The aim of this study was to evaluate the effect of melanin inhibition on antifungal susceptibility of chromoblastomycosis agents and describe the susceptibility profile of some unusual CBM agents. Seventy-six clinical isolates, representing 13 species of the five main CBM agents genera, were studied. The antifungal susceptibility was performed according to the M38-A2 protocol of CLSI. In the melanin inhibition test, 16 mg/L of tricyclazole was added to the medium used in the inoculum preparation and the susceptibility assay. CBM agents were less susceptible to amphotericin B in comparison with azoles and terbinafine. The unusual species showed similar susceptibilities profiles to those of other species of the same genera. With tricyclazole exposition, MICs of terbinafine, posaconazole and itraconazole for Fonsecaea spp. significantly decreased (p<0,05). For Phialophora spp., this reduction was significative for posaconazole and itraconazole. For the other genera, there was a reduction in MICs of terbinafine and itraconazole, however, the statistical tests were not significant. Melanin inhibition can increase the antifungal susceptibility of most CBM agents to itraconazole and terbinafine, the main used drugs in the disease treatment. This increased susceptibility may open up new possibilities for therapy in refractory cases of CBM and/or caused by resistant fungal strains. Further studies are needed to confirm the same results in vivo.

SAFETY AND ACTIVITY OF SAQUINAVIR IN HIV INFECTION

PEDIATRICS ◽  
1996 ◽  
Vol 98 (2) ◽  
pp. 346-347
Author(s):  
Davod E. Parry ◽  
Robert F. Lemanske
Keyword(s):  
Hiv Infection ◽  
Statistical Tests ◽  
Specific Inhibitor ◽  
Hiv Replication ◽  
Cd4 Counts ◽  
Hiv Rna ◽  
Final Assembly ◽  
Drug Tolerability ◽  
Dose Response Study

Purpose of the Study. To assess the in vivo antiviral activity and tolerability of saquinavir in a dose-response study design. Study Population. Forty-nine adult male, zidovudinenaive, human immunodeficiency virus (HIV)-positive patients with few or no symptoms of HIV infection and CD4 counts less than 500/mm3. Methods. Patients were stratified according to symptoms or no symptoms of HIV infection and were randomized to receive saquinavir at 25 mg (10 patients), 75 mg (12 patients), 200 mg (12 patients), or 600 mg (10 patients) all orally tid for 16 weeks. Five patients in each group had symptoms of HIV infection. Patients were evaluated at baseline, weekly for the first 4 weeks and then at 6, 8, 12 and 16 weeks. Virus isolation and quantification was performed at baseline and 4-weekly intervals. HIV RNA was measured by polymerase chain reaction (PCR). No statistical tests were performed. Results. Trends were reported and depicted graphically in the article. Patients in the higher dosage groups (particularly 600 mg tid) showed increasing CD4 counts and a reduction in viral load as measured by quantitative PCR for HIV RNA over the treatment period. Little difference was seen between the two lower dosage treatment groups. Data from all patients (including 8 who did not complete the 16 weeks of study) were analyzed for drug tolerability; reported adverse events were mild and considered by investigators mostly to be unrelated to the treatment. Conclusions and Reviewers' Comments. Saquinavir is a highly specific inhibitor of HIV proteases that are essential for the final assembly of viable viral particles in HIV replication.

scholarly journals A Yes-Associated Protein (YAP) and Insulin-Like Growth Factor 1 Receptor (IGF-1R) Signaling Loop Is Involved in Sorafenib Resistance in Hepatocellular Carcinoma

Cancers ◽  
2021 ◽  
Vol 13 (15) ◽  
pp. 3812
Author(s):  
Mai-Huong T. Ngo ◽  
Sue-Wei Peng ◽  
Yung-Che Kuo ◽  
Chun-Yen Lin ◽  
Ming-Heng Wu ◽  
...  
Keyword(s):  
Nuclear Translocation ◽  
Combination Index ◽  
Specific Inhibitor ◽  
In Vivo Model ◽  
Mrna Levels ◽  
Margin Distribution ◽  
Related Proteins ◽  
Emt Markers

The role of a YAP-IGF-1R signaling loop in HCC resistance to sorafenib remains unknown. Method: Sorafenib-resistant cells were generated by treating naïve cells (HepG2215 and Hep3B) with sorafenib. Different cancer cell lines from databases were analyzed through the ONCOMINE web server. BIOSTORM–LIHC patient tissues (46 nonresponders and 21 responders to sorafenib) were used to compare YAP mRNA levels. The HepG2215_R-derived xenograft in SCID mice was used as an in vivo model. HCC tissues from a patient with sorafenib failure were used to examine differences in YAP and IGF-R signaling. Results: Positive associations exist among the levels of YAP, IGF-1R, and EMT markers in HCC tissues and the levels of these proteins increased with sorafenib failure, with a trend of tumor-margin distribution in vivo. Blocking YAP downregulated IGF-1R signaling-related proteins, while IGF-1/2 treatment enhanced the nuclear translocation of YAP in HCC cells through PI3K-mTOR regulation. The combination of YAP-specific inhibitor verteporfin (VP) and sorafenib effectively decreased cell viability in a synergistic manner, evidenced by the combination index (CI). Conclusion: A YAP-IGF-1R signaling loop may play a role in HCC sorafenib resistance and could provide novel potential targets for combination therapy with sorafenib to overcome drug resistance in HCC.

RPR120844, a Novel, Specific Inhibitor of Coagulation Factor Xa Inhibits Venous Thrombosis in the Rabbit

1999 ◽  
Vol 81 (01) ◽  
pp. 157-160 ◽  
Author(s):  
Ross Bentley ◽  
Suzanne Morgan ◽  
Karen Brown ◽  
Valeria Chu ◽  
Richard Ewing ◽  
...  
Keyword(s):  
Jugular Vein ◽  
Drug Effect ◽  
Ex Vivo ◽  
Thrombus Formation ◽  
Specific Inhibitor ◽  
Factor Xa ◽  
Coagulation Factor ◽  
Antithrombotic Activity ◽  
Fxa Inhibitor

SummaryThe in vivo antithrombotic activity of RPR120844, a novel synthetic coagulation factor Xa (fXa) inhibitor (Ki = 7 nM), was assessed by its ability to inhibit thrombus formation in a damaged segment of the rabbit jugular vein. Intravenous dose-response studies were performed and thrombus mass (TM), activated partial thromboplastin time (APTT), prothrombin time (PT), inhibition of ex vivo fXa activity and plasma drug levels (PDL) were determined. TM, measured at the end of a 50 min infusion, was significantly reduced (p <0.05 vs saline-treated animals) by RPR120844 at 30 and 100 μg/kg/min. At doses of 10, 30 and 100 μg/kg/min, APTT was prolonged by 2.1, 4.2 and 6.1-fold, and PT was prolonged by 1.4, 2.2 and 3.5-fold, respectively. PDL were determined by measuring anti-fXa activity using an amidolytic assay. Peak PDL were 0.8 ± 0.3, 1.5 ± 0.9 and 2.4 ± 0.6 μM, respectively. The drug effect was reversible with APTT, PT and PDL returning toward pretreatment values 30 min after termination of treatment. The results suggest that RPR120844, or similar compounds, may provide an efficacious, yet easily reversible, means of inhibiting thrombus formation.

scholarly journals Activation of transmembrane receptor tyrosine kinase DDR1-STAT3 cascade by extracellular matrix remodeling promotes liver metastatic colonization in uveal melanoma

2021 ◽  
Vol 6 (1) ◽  
Author(s):  
Wei Dai ◽  
Shenglan Liu ◽  
Shubo Wang ◽  
Li Zhao ◽  
Xiao Yang ◽  
...  
Keyword(s):  
Extracellular Matrix ◽  
Cancer Cells ◽  
Uveal Melanoma ◽  
Specific Inhibitor ◽  
Type I ◽  
Matrix Remodeling ◽  
Metastatic Colonization ◽  
Tgf Β1

AbstractColonization is believed a rate-limiting step of metastasis cascade. However, its underlying mechanism is not well understood. Uveal melanoma (UM), which is featured with single organ liver metastasis, may provide a simplified model for realizing the complicated colonization process. Because DDR1 was identified to be overexpressed in UM cell lines and specimens, and abundant pathological deposition of extracellular matrix collagen, a type of DDR1 ligand, was noted in the microenvironment of liver in metastatic patients with UM, we postulated the hypothesis that DDR1 and its ligand might ignite the interaction between UM cells and their surrounding niche of liver thereby conferring strengthened survival, proliferation, stemness and eventually promoting metastatic colonization in liver. We tested this hypothesis and found that DDR1 promoted these malignant cellular phenotypes and facilitated metastatic colonization of UM in liver. Mechanistically, UM cells secreted TGF-β1 which induced quiescent hepatic stellate cells (qHSCs) into activated HSCs (aHSCs) which secreted collagen type I. Such a remodeling of extracellular matrix, in turn, activated DDR1, strengthening survival through upregulating STAT3-dependent Mcl-1 expression, enhancing stemness via upregulating STAT3-dependent SOX2, and promoting clonogenicity in cancer cells. Targeting DDR1 by using 7rh, a specific inhibitor, repressed proliferation and survival in vitro and in vivo outgrowth. More importantly, targeting cancer cells by pharmacological inactivation of DDR1 or targeting microenvironmental TGF-β1-collagen I loop exhibited a prominent anti-metastasis effect in mice. In conclusion, targeting DDR1 signaling and TGF-β signaling may be a novel approach to diminish hepatic metastasis in UM.

scholarly journals Cwp84, a Surface-Associated Protein of Clostridium difficile, Is a Cysteine Protease with Degrading Activity on Extracellular Matrix Proteins

2007 ◽  
Vol 189 (20) ◽  
pp. 7174-7180 ◽  
Author(s):  
Claire Janoir ◽  
Séverine Péchiné ◽  
Charlotte Grosdidier ◽  
Anne Collignon
Keyword(s):  
Extracellular Matrix ◽  
Clostridium Difficile ◽  
Cysteine Protease ◽  
Specific Inhibitor ◽  
Extracellular Matrix Proteins ◽  
Maximum Activity ◽  
Matrix Proteins ◽  
Specific Antibodies ◽  
Type Iv

ABSTRACT Clostridium difficile pathogenicity is mediated mainly by its A and B toxins, but the colonization process is thought to be a necessary preliminary step in the course of infection. The aim of this study was to characterize the Cwp84 protease of C. difficile, which is highly immunogenic in patients with C. difficile-associated disease and is potentially involved in the pathogenic process. Cwp84 was purified as a recombinant His-tagged protein, and specific antibodies were generated in rabbits. Treatment of multiple-band-containing eluted fractions with a reducing agent or with trypsin led to accumulation of a unique protein species with an estimated molecular mass of 61 kDa, corresponding most likely to mature autoprocessed Cwp84 (mCwp84). mCwp84 showed concentration-dependent caseinolytic activity, with maximum activity at pH 7.5. The Cwp84 activity was inhibited by various cysteine protease inhibitors, such as the specific inhibitor E64, and the anti-Cwp84-specific antibodies. Using fractionation experiments followed by immunoblot detection, the protease was found to be associated with the S-layer proteins, mostly as a nonmature species. Proteolytic assays were performed with extracellular matrix proteins to assess the putative role of Cwp84 in the pathogenicity of C. difficile. No degrading activity was detected with type IV collagen. In contrast, Cwp84 exhibited degrading activity with fibronectin, laminin, and vitronectin, which was neutralized by the E64 inhibitor and specific antibodies. In vivo, this proteolytic activity could contribute to the degradation of the host tissue integrity and to the dissemination of the infection.

scholarly journals Activation of the oxygen-sensing signal cascade prevents mitochondrial injury after mouse liver ischemia-reperfusion

2008 ◽  
Vol 295 (4) ◽  
pp. G823-G832 ◽  
Author(s):  
Zhi Zhong ◽  
Venkat K. Ramshesh ◽  
Hasibur Rehman ◽  
Robert T. Currin ◽  
Vijayalakshmi Sridharan ◽  
...  
Keyword(s):  
Mitochondrial Permeability Transition ◽  
Oxygen Sensing ◽  
Ischemia Reperfusion ◽  
Multiphoton Microscopy ◽  
Specific Inhibitor ◽  
Permeability Transition ◽  
Heme Oxygenase 1 ◽  
Mitochondrial Depolarization ◽  
Signal Cascade

The mitochondrial permeability transition (MPT) plays an important role in hepatocyte death caused by ischemia-reperfusion (IR). This study investigated whether activation of the cellular oxygen-sensing signal cascade by prolyl hydroxylase inhibitors (PHI) protects against the MPT after hepatic IR. Ethyl 3,4-dihyroxybenzoate (EDHB, 100 mg/kg ip), a PHI, increased mouse hepatic hypoxia-inducible factor-1α and heme oxygenase-1 (HO-1). EDHB-treated and untreated mice were subjected to 1 h of warm ischemia to ∼70% of the liver followed by reperfusion. Mitochondrial polarization, cell death, and the MPT were assessed by intravital confocal/multiphoton microscopy of rhodamine 123, propidium iodide, and calcein. EDHB largely blunted alanine aminotransferase (ALT) release and necrosis after reperfusion. In vehicle-treated mice at 2 h after reperfusion, viable cells with depolarized mitochondria were 72%, and dead cells were 2%, indicating that depolarization preceded necrosis. Mitochondrial voids excluding calcein disappeared, indicating MPT onset in vivo. NIM811, a specific inhibitor of the MPT, blocked mitochondrial depolarization after IR, further confirming that mitochondrial depolarization was due to MPT onset. EDHB decreased mitochondrial depolarization to 16% and prevented the MPT. Tin protoporphyrin (10 μmol/kg sc), an HO-1 inhibitor, partially abrogated protection by EDHB against ALT release, necrosis, and mitochondrial depolarization. In conclusion, IR causes the MPT and mitochondrial dysfunction, leading to hepatocellular death. PHI prevents MPT onset and liver damage through an effect mediated partially by HO-1.

scholarly journals Quantitation of Candida albicans Ergosterol Content Improves the Correlation between In Vitro Antifungal Susceptibility Test Results and In Vivo Outcome after Fluconazole Treatment in a Murine Model of Invasive Candidiasis

2000 ◽  
Vol 44 (8) ◽  
pp. 2081-2085 ◽  
Author(s):  
Beth A. Arthington-Skaggs ◽  
David W. Warnock ◽  
Christine J. Morrison
Keyword(s):  
Murine Model ◽  
Invasive Candidiasis ◽  
Clinical Laboratory ◽  
Antifungal Susceptibility ◽  
Resistant Isolate ◽  
National Committee ◽  
Ergosterol Content ◽  
In Vitro Antifungal Susceptibility

ABSTRACT MIC end point determination for the most commonly prescribed azole antifungal drug, fluconazole, can be complicated by “trailing” growth of the organism during susceptibility testing by the National Committee for Clinical Laboratory Standards approved M27-A broth macrodilution method and its modified broth microdilution format. To address this problem, we previously developed the sterol quantitation method (SQM) for in vitro determination of fluconazole susceptibility, which measures cellular ergosterol content rather than growth inhibition after exposure to fluconazole. To determine if SQM MICs of fluconazole correlated better with in vivo outcome than M27-A MICs, we used a murine model of invasive candidiasis and analyzed the capacity of fluconazole to treat infections caused by C. albicansisolates which were trailers (M27-A MICs at 24 and 48 h, ≤1.0 and ≥64 μg/ml, respectively; SQM MIC, ≤1.0 μg/ml), as well as those which were fluconazole sensitive (M27-A and SQM MIC, ≤1.0 μg/ml) and fluconazole resistant (M27-A MIC, ≥64 μg/ml; SQM MIC, 54 μg/ml). Compared with the untreated controls, fluconazole therapy increased the survival of mice infected with a sensitive isolate and both trailing isolates but did not increase the survival of mice infected with a resistant isolate. These results indicate that the SQM is more predictive of in vivo outcome than the M27-A method for isolates that give unclear MIC end points due to trailing growth in fluconazole.

scholarly journals The consequences of deglycosylation of recombinant intra-melanosomal domain of human tyrosinase

2017 ◽  
Vol 399 (1) ◽  
pp. 73-77 ◽  
Author(s):  
Monika B. Dolinska ◽  
Yuri V. Sergeev
Keyword(s):  
Oculocutaneous Albinism ◽  
Site Directed Mutagenesis ◽  
Directed Mutagenesis ◽  
Multiple Site ◽  
Insect Larvae ◽  
Melanin Biosynthesis ◽  
Human Tyrosinase

AbstractTyrosinase, a melanosomal glycoenzyme, catalyzes initial steps of the melanin biosynthesis. While glycosylation was previously studiedin vivo, we present three recombinant mutant variants of human tyrosinase, which were obtained using multiple site-directed mutagenesis, expressed in insect larvae, purified and characterized biochemically. The mutagenesis demonstrated the reduced protein expression and enzymatic activity due to possible loss of protein stability and protein degradation. However, the complete deglycosylation of asparagine residuesin vitro, including the residue in position 371, interrupts tyrosinase function, which is consistent with a melanin loss in oculocutaneous albinism type 1 (OCA1) patients.

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