scholarly journals Guanabenz Repurposed as an Antiparasitic with Activity against Acute and Latent Toxoplasmosis

2015 ◽  
Vol 59 (11) ◽  
pp. 6939-6945 ◽  
Author(s):  
Imaan Benmerzouga ◽  
Lisa A. Checkley ◽  
Michael T. Ferdig ◽  
Gustavo Arrizabalaga ◽  
Ronald C. Wek ◽  
...  
Keyword(s):  
Translational Control ◽  
Protozoan Parasite ◽  
Empty Space ◽  
Antiparasitic Activity ◽  
Content Type ◽  
Life Threatening ◽  
Brain Cysts ◽  
The Brain

ABSTRACTToxoplasma gondiiis a protozoan parasite that persists as a chronic infection.Toxoplasmaevades immunity by forming tissue cysts, which reactivate to cause life-threatening disease during immune suppression. There is an urgent need to identify drugs capable of targeting these latent tissue cysts, which tend to form in the brain. We previously showed that translational control is critical during infections with both replicative and latent forms ofToxoplasma. Here we report that guanabenz, an FDA-approved drug that interferes with translational control, has antiparasitic activity against replicative stages ofToxoplasmaand the related apicomplexan parasitePlasmodium falciparum(a malaria agent). We also found that inhibition of translational control interfered with tissue cyst biologyin vitro.Toxoplasmabradyzoites present in these abnormal cysts were diminished and misconfigured, surrounded by empty space not seen in normal cysts. These findings prompted analysis of the efficacy of guanabenzin vivoby using established mouse models of acute and chronic toxoplasmosis. In addition to protecting mice from lethal doses ofToxoplasma, guanabenz has a remarkable ability to reduce the number of brain cysts in chronically infected mice. Our findings suggest that guanabenz can be repurposed into an effective antiparasitic with a unique ability to reduce tissue cysts in the brain.

scholarly journals Surface-Engineered Dendrimeric Nanoconjugates for Macrophage-Targeted Delivery of Amphotericin B: Formulation Development andIn VitroandIn VivoEvaluation

2015 ◽  
Vol 59 (5) ◽  
pp. 2479-2487 ◽  
Author(s):  
Keerti Jain ◽  
Ashwni Kumar Verma ◽  
Prabhat Ranjan Mishra ◽  
Narendra Kumar Jain
Keyword(s):  
Drug Release ◽  
Amphotericin B ◽  
Human Erythrocytes ◽  
Antileishmanial Activity ◽  
Cell Uptake ◽  
Antiparasitic Activity ◽  
Content Type ◽  
Drug Localization

ABSTRACTThe present study aimed to develop an optimized dendrimeric delivery system for amphotericin B (AmB). Fifth-generation (5.0G) poly(propylene imine) (PPI) dendrimers were synthesized, conjugated with mannose, and characterized by use of various analytical techniques, including Fourier transform infrared spectroscopy (FTIR),1H nuclear magnetic resonance (1H-NMR) spectroscopic analysis, and atomic force microscopy (AFM). Mannose-conjugated 5.0G PPI (MPPI) dendrimers were loaded with AmB and evaluated for drug loading efficiency,in vitrodrug release profile, stability, hemolytic toxicity to human erythrocytes, cytotoxicity to and cell uptake by J774A.1 macrophage cells, antiparasitic activity against intracellularLeishmania donovaniamastigotes,in vivopharmacokinetic and biodistribution profiles, drug localization index, toxicity, and antileishmanial activity. AFM showed the nanometric size of the MPPI dendrimers, with a nearly globular architecture. The conjugate showed a good entrapment efficiency for AmB, along with pH-sensitive drug release. Highly significant reductions in toxicity toward human erythrocytes and macrophage cells, without compromising the antiparasitic activity of AmB, were observed. The dendrimeric formulation of AmB showed a significant enhancement of the parasiticidal activity of AmB toward intramacrophagicL. donovaniamastigotes. In thein vitrocell uptake studies, the formulation showed selectivity toward macrophages, with significant intracellular uptake. Further pharmacokinetic and organ distribution studies elucidated the controlled delivery behavior of the formulation. The drug localization index was found to increase significantly in macrophage-rich organs.In vivostudies showed a biocompatible behavior of MPPIA, with negligible toxicity even at higher doses, and promising antileishmanial activity. From the results, we concluded that surface-engineered dendrimers may serve as optimized delivery vehicles for AmB with enhanced activity and low or negligible toxicity.

scholarly journals Efficacy of Guanabenz Combination Therapy against Chronic Toxoplasmosis across Multiple Mouse Strains

2020 ◽  
Vol 64 (9) ◽  
Author(s):  
Jennifer Martynowicz ◽  
J. Stone Doggett ◽  
William J. Sullivan
Keyword(s):  
Limit Of Detection ◽  
Acute Infection ◽  
Antagonistic Effect ◽  
Mouse Strains ◽  
Chronic Infections ◽  
Content Type ◽  
Brain Cyst ◽  
The Brain

ABSTRACT Toxoplasma gondii, an obligate intracellular parasite that can cause life-threatening acute disease, differentiates into a quiescent cyst stage to establish lifelong chronic infections in animal hosts, including humans. This tissue cyst reservoir, which can reactivate into an acute infection, is currently refractory to clinically available therapeutics. Recently, we and others have discovered drugs capable of significantly reducing the brain cyst burden in latently infected mice, but not to undetectable levels. In this study, we examined the use of novel combination therapies possessing multiple mechanisms of action in mouse models of latent toxoplasmosis. Our drug regimens included combinations of pyrimethamine, clindamycin, guanabenz, and endochin-like quinolones (ELQs) and were administered to two different mouse strains in an attempt to eradicate brain tissue cysts. We observed mouse strain-dependent effects with these drug treatments: pyrimethamine-guanabenz showed synergistic efficacy in C57BL/6 mice yet did not improve upon guanabenz monotherapy in BALB/c mice. Contrary to promising in vitro results demonstrating toxicity to bradyzoites, we observed an antagonistic effect between guanabenz and ELQ-334 in vivo. While we were unable to completely eliminate the brain cyst burden, we found that a combination treatment with ELQ-334 and pyrimethamine impressively reduced the brain cyst burden by 95% in C57BL/6 mice, which approached the limit of detection. These analyses highlight the importance of evaluating anti-infective drugs in multiple mouse strains and will help inform further preclinical studies of cocktail therapies designed to treat chronic toxoplasmosis.

scholarly journals Characterization of the pathogenesis and immune response to Listeria monocytogenes strains isolated from a sustained national outbreak

2019 ◽  
Vol 9 (1) ◽  
Author(s):  
Pallab Ghosh ◽  
Yan Zhou ◽  
Quentin Richardson ◽  
Darren E. Higgins
Keyword(s):  
Listeria Monocytogenes ◽  
Laboratory Strain ◽  
The United States ◽  
Clinical Isolates ◽  
T Cell Antigens ◽  
Life Threatening ◽  
Cellular Immune ◽  
The Brain

AbstractListeria monocytogenes is an intracellular pathogen responsible for listeriosis, a foodborne disease that can lead to life-threatening meningitis. The 2011 L. monocytogenes cantaloupe outbreak was among the deadliest foodborne outbreaks in the United States. We conducted in vitro and in vivo infection analyses to determine whether strains LS741 and LS743, two clinical isolates from the cantaloupe outbreak, differ significantly from the common laboratory strain 10403S. We showed that LS741 and LS743 exhibited increased virulence, characterized by higher colonization of the brain and other organs in mice. Assessment of cellular immune responses to known CD8+ T cell antigens was comparable between all strains. However, pre-existing immunity to 10403S did not confer protection in the brain against challenge with LS741. These studies provide insights into the pathogenesis of clinical isolates linked to the 2011 cantaloupe outbreak and also indicate that currently utilized laboratory strains are imperfect models for studying L. monocytogenes pathogenesis.

scholarly journals Hydroxylamine and Carboxymethoxylamine Can Inhibit Toxoplasma gondii Growth through an Aspartate Aminotransferase-Independent Pathway

2020 ◽  
Vol 64 (3) ◽  
Author(s):  
Jixu Li ◽  
Huanping Guo ◽  
Eloiza May Galon ◽  
Yang Gao ◽  
Seung-Hun Lee ◽  
...  
Keyword(s):  
Toxoplasma Gondii ◽  
Drug Targets ◽  
Protozoan Parasite ◽  
Cell Types ◽  
Lytic Cycle ◽  
Type I ◽  
Content Type ◽  
Mechanism Of Inhibition

ABSTRACT Toxoplasma gondii is an obligate intracellular protozoan parasite and a successful parasitic pathogen in diverse organisms and host cell types. Hydroxylamine (HYD) and carboxymethoxylamine (CAR) have been reported as inhibitors of aspartate aminotransferases (AATs) and interfere with the proliferation in Plasmodium falciparum. Therefore, AATs are suggested as drug targets against Plasmodium. The T. gondii genome encodes only one predicted AAT in both T. gondii type I strain RH and type II strain PLK. However, the effects of HYD and CAR, as well as their relationship with AAT, on T. gondii remain unclear. In this study, we found that HYD and CAR impaired the lytic cycle of T. gondii in vitro, including the inhibition of invasion or reinvasion, intracellular replication, and egress. Importantly, HYD and CAR could control acute toxoplasmosis in vivo. Further studies showed that HYD and CAR could inhibit the transamination activity of rTgAAT in vitro. However, our results confirmed that deficiency of AAT in both RH and PLK did not reduce the virulence in mice, although the growth ability of the parasites was affected in vitro. HYD and CAR could still inhibit the growth of AAT-deficient parasites. These findings indicated that HYD and CAR inhibition of T. gondii growth and control of toxoplasmosis can occur in an AAT-independent pathway. Overall, further studies focusing on the elucidation of the mechanism of inhibition are warranted. Our study hints at new substrates of HYD and CAR as potential drug targets to inhibit T. gondii growth.

scholarly journals Interferon-Stimulated Gene (ISG)-Expression Screening Reveals the Specific Antibunyaviral Activity of ISG20

2018 ◽  
Vol 92 (13) ◽  
Author(s):  
Junjie Feng ◽  
Arthur Wickenhagen ◽  
Matthew L. Turnbull ◽  
Veronica V. Rezelj ◽  
Felix Kreher ◽  
...  
Keyword(s):  
Viral Infections ◽  
Rhesus Macaques ◽  
Type I ◽  
Emerging Viruses ◽  
Content Type ◽  
Expression Screening ◽  
Interferon Stimulated Genes ◽  
Life Threatening

ABSTRACT Bunyaviruses pose a significant threat to human health, prosperity, and food security. In response to viral infections, interferons (IFNs) upregulate the expression of hundreds of interferon-stimulated genes (ISGs), whose cumulative action can potently inhibit the replication of bunyaviruses. We used a flow cytometry-based method to screen the ability of ∼500 unique ISGs from humans and rhesus macaques to inhibit the replication of Bunyamwera orthobunyavirus (BUNV), the prototype of both the Peribunyaviridae family and the Bunyavirales order. Candidates possessing antibunyaviral activity were further examined using a panel of divergent bunyaviruses. Interestingly, one candidate, ISG20, exhibited potent antibunyaviral activity against most viruses examined from the Peribunyaviridae , Hantaviridae , and Nairoviridae families, whereas phleboviruses ( Phenuiviridae ) largely escaped inhibition. Similar to the case against other viruses known to be targeted by ISG20, the antibunyaviral activity of ISG20 is dependent upon its functional RNase activity. Through use of an infectious virus-like particle (VLP) assay (based on the BUNV minigenome system), we confirmed that gene expression from all 3 viral segments is strongly inhibited by ISG20. Using in vitro evolution, we generated a substantially ISG20-resistant BUNV and mapped the determinants of ISG20 sensitivity/resistance. Taking all the data together, we report that ISG20 is a broad and potent antibunyaviral factor but that some bunyaviruses are remarkably ISG20 resistant. Thus, ISG20 sensitivity/resistance may influence the pathogenesis of bunyaviruses, many of which are emerging viruses of clinical or veterinary significance. IMPORTANCE There are hundreds of bunyaviruses, many of which cause life-threatening acute diseases in humans and livestock. The interferon (IFN) system is a key component of innate immunity, and type I IFNs limit bunyaviral propagation both in vitro and in vivo . Type I IFN signaling results in the upregulation of hundreds of IFN-stimulated genes (ISGs), whose concerted action generates an “antiviral state.” Although IFNs are critical in limiting bunyaviral replication and pathogenesis, much is still unknown about which ISGs inhibit bunyaviruses. Using ISG-expression screening, we examined the ability of ∼500 unique ISGs to inhibit Bunyamwera orthobunyavirus (BUNV), the prototypical bunyavirus. Using this approach, we identified ISG20, an interferon-stimulated exonuclease, as a potent inhibitor of BUNV. Interestingly, ISG20 possesses highly selective antibunyaviral activity, with multiple bunyaviruses being potently inhibited while some largely escape inhibition. We speculate that the ability of some bunyaviruses to escape ISG20 may influence their pathogenesis.

A realistic brain tissue phantom for intraparenchymal infusion studies

2004 ◽  
Vol 101 (2) ◽  
pp. 314-322 ◽  
Author(s):  
Zhi-Jian Chen ◽  
George T. Gillies ◽  
William C. Broaddus ◽  
Sujit S. Prabhu ◽  
Helen Fillmore ◽  
...  
Keyword(s):  
Mr Imaging ◽  
Bromophenol Blue ◽  
Agarose Gel ◽  
Convection Enhanced Delivery ◽  
Content Type ◽  
Force Profile ◽  
The Brain ◽  
Fine Print

Object. The goal of this study was to validate a simple, inexpensive, and robust model system to be used as an in vitro surrogate for in vivo brain tissues in preclinical and exploratory studies of infusion-based intraparenchymal drug and cell delivery. Methods. Agarose gels of varying concentrations and porcine brain were tested to determine the infusion characteristics of several different catheters at flow rates of 0.5 and 1 µl per minute by using bromophenol blue (BPB) dye (molecular weight [MW] ∼690) and gadodiamide (MW ∼573). Magnetic resonance (MR) imaging and videomicroscopy were used to measure the distribution of these infusates, with a simultaneous measurement of infusion pressures. In addition, the forces of catheter penetration and movement through gel and brain were measured. Agarose gel at a 0.6% concentration closely resembles in vivo brain with respect to several critical physical characteristics. The ratio of distribution volume to infusion volume of agarose was 10 compared with 7.1 for brain. The infusion pressure of the gel demonstrated profiles similar in configuration and magnitude to those of the brain (plateau pressures 10–20 mm Hg). Gadodiamide infusion in agarose closely resembled that in the brain, as documented using T1-weighted MR imaging. Gadodiamide distribution in agarose gel was virtually identical to that of BPB dye, as documented by MR imaging and videomicroscopy. The force profile for insertion of a silastic catheter into agarose gel was similar in magnitude and configuration to the force profile for insertion into the brain. Careful insertion of the cannula using a stereotactic guide is critical to minimize irregularity and backflow of infusate distribution. Conclusions. Agarose gel (0.6%) is a useful surrogate for in vivo brain in exploratory studies of convection-enhanced delivery.

scholarly journals Significant Reduction of Brain Cysts Caused by Toxoplasma gondii after Treatment with Spiramycin Coadministered with Metronidazole in a Mouse Model of Chronic Toxoplasmosis

2012 ◽  
Vol 56 (4) ◽  
pp. 1762-1768 ◽  
Author(s):  
Wai Kit Chew ◽  
Ignacio Segarra ◽  
Stephen Ambu ◽  
Joon Wah Mak
Keyword(s):  
Toxoplasma Gondii ◽  
Vero Cells ◽  
Toxoplasmic Encephalitis ◽  
Combination Group ◽  
Control Group ◽  
Time Curve ◽  
Content Type ◽  
Brain Cysts ◽  
The Brain

ABSTRACTToxoplasma gondiiis a parasite that generates latent cysts in the brain; reactivation of these cysts may lead to fatal toxoplasmic encephalitis, for which treatment remains unsuccessful. We assessed spiramycin pharmacokinetics coadministered with metronidazole, the eradication of brain cysts and thein vitroreactivation. Male BALB/c mice were fed 1,000 tachyzoites orally to develop chronic toxoplasmosis. Four weeks later, infected mice underwent different treatments: (i) infected untreated mice (n= 9), which received vehicle only; (ii) a spiramycin-only group (n= 9), 400 mg/kg daily for 7 days; (iii) a metronidazole-only group (n= 9), 500 mg/kg daily for 7 days; and (iv) a combination group (n= 9), which received both spiramycin (400 mg/kg) and metronidazole (500 mg/kg) daily for 7 days. An uninfected control group (n= 10) was administered vehicle only. After treatment, the brain cysts were counted, brain homogenates were cultured in confluent Vero cells, and cysts and tachyzoites were counted after 1 week. Separately, pharmacokinetic profiles (plasma and brain) were assessed after a single dose of spiramycin (400 mg/kg), metronidazole (500 mg/kg), or both. Metronidazole treatment increased the brain spiramycin area under the concentration-time curve from 0 h to ∞ (AUC0–∞) by 67% without affecting its plasma disposition. Metronidazole plasma and brain AUC0–∞values were reduced 9 and 62%, respectively, after spiramycin coadministration. Enhanced spiramycin brain exposure after coadministration reduced brain cysts 15-fold (79 ± 23 for the combination treatment versus 1,198 ± 153 for the untreated control group [P< 0.05]) and 10-fold versus the spiramycin-only group (768 ± 125). Metronidazole alone showed no effect (1,028 ± 149). Tachyzoites were absent in the brain. Spiramycin reducedin vitroreactivation. Metronidazole increased spiramycin brain penetration, causing a significant reduction ofT. gondiibrain cysts, with potential clinical translatability for chronic toxoplasmosis treatment.

scholarly journals Toll-Like Receptor 9 Signaling in Dendritic Cells Regulates Neutrophil Recruitment to Inflammatory Foci following Leishmania infantum Infection

2015 ◽  
Vol 83 (12) ◽  
pp. 4604-4616 ◽  
Author(s):  
Laís Sacramento ◽  
Silvia C. Trevelin ◽  
Manuela S. Nascimento ◽  
Djalma S. Lima-Jùnior ◽  
Diego L. Costa ◽  
...  
Keyword(s):  
Leishmania Infantum ◽  
Critical Role ◽  
Costimulatory Molecule ◽  
Protozoan Parasite ◽  
Neutrophil Recruitment ◽  
Content Type ◽  
Target Organs ◽  
Myd88 Signaling

Leishmania infantumis a protozoan parasite that causes visceral leishmaniasis (VL). This infection triggers dendritic cell (DC) activation through the recognition of microbial products by Toll-like receptors (TLRs). Among the TLRs, TLR9 is required for DC activation by differentLeishmaniaspecies. We demonstrated that TLR9 is upregulatedin vitroandin vivoduring infection. We show that C57BL/6 mice deficient in TLR9 expression (TLR9−/−mice) are more susceptible to infection and display higher parasite numbers in the spleen and liver. The increased susceptibility of TLR9−/−mice was due to the impaired recruitment of neutrophils to the infection foci associated with reduced levels of neutrophil chemoattractants released by DCs in the target organs. Moreover, both Th1 and Th17 cells were also committed in TLR9−/−mice. TLR9-dependent neutrophil recruitment is mediated via the MyD88 signaling pathway but is TIR domain-containing adapter-inducing interferon beta (TRIF) independent. Furthermore,L. infantumfailed to activate both plasmacytoid and myeloid DCs from TLR9−/−mice, which presented reduced surface costimulatory molecule expression and chemokine release. Interestingly, neutrophil chemotaxis was affected bothin vitroandin vivowhen DCs were derived from TLR9−/−mice. Our results suggest that TLR9 plays a critical role in neutrophil recruitment during the protective response againstL. infantuminfection that could be associated with DC activation.

scholarly journals Listeria monocytogenes Spreads within the Brain by Actin-Based Intra-Axonal Migration

2015 ◽  
Vol 83 (6) ◽  
pp. 2409-2419 ◽  
Author(s):  
Diana Henke ◽  
Sebastian Rupp ◽  
Véronique Gaschen ◽  
Michael H. Stoffel ◽  
Joachim Frey ◽  
...  
Keyword(s):  
Immune Response ◽  
Listeria Monocytogenes ◽  
Brain Cell ◽  
Bovine Brain ◽  
Content Type ◽  
Bacterial Migration ◽  
Bacterial Replication ◽  
The Brain

Listeria monocytogenesrhombencephalitis is a severe progressive disease despite a swift intrathecal immune response. Based on previous observations, we hypothesized that the disease progresses by intra-axonal spread within the central nervous system. To test this hypothesis, neuroanatomical mapping of lesions, immunofluorescence analysis, and electron microscopy were performed on brains of ruminants with naturally occurring rhombencephalitis. In addition, infection assays were performed in bovine brain cell cultures. Mapping of lesions revealed a consistent pattern with a preferential affection of certain nuclear areas and white matter tracts, indicating thatListeria monocytogenesspreads intra-axonally within the brain along interneuronal connections. These results were supported by immunofluorescence and ultrastructural data localizingListeria monocytogenesinside axons and dendrites associated with networks of fibrillary structures consistent with actin tails.In vitroinfection assays confirmed that bacteria were moving within axon-like processes by employing their actin tail machinery. Remarkably,in vivo, neutrophils invaded the axonal space and the axon itself, apparently by moving between split myelin lamellae of intact myelin sheaths. This intra-axonal invasion of neutrophils was associated with various stages of axonal degeneration and bacterial phagocytosis. Paradoxically, the ensuing adaxonal microabscesses appeared to provide new bacterial replication sites, thus supporting further bacterial spread. In conclusion, intra-axonal bacterial migration and possibly also the innate immune response play an important role in the intracerebral spread of the agent and hence the progression of listeric rhombencephalitis.

scholarly journals Azole Heteroresistance in Cryptococcus neoformans: Emergence of Resistant Clones with Chromosomal Disomy in the Mouse Brain during Fluconazole Treatment

2013 ◽  
Vol 57 (10) ◽  
pp. 5127-5130 ◽  
Author(s):  
Edward Sionov ◽  
Yun C. Chang ◽  
Kyung J. Kwon-Chung
Keyword(s):  
Mouse Model ◽  
Cryptococcus Neoformans ◽  
Mouse Brain ◽  
Content Type ◽  
The Brain ◽  
Strain Dependent ◽  
Is Strain

ABSTRACTWe have previously reported thatCryptococcus neoformansstrains are innately heteroresistant to fluconazolein vitro, producing minor, highly resistant subpopulations due to adaptive formation of disomic chromosomes. Using a mouse model, we assessed the emergence of heteroresistant clones in the brain during fluconazole treatment and found that the occurrence of heteroresistant clonesin vivowith chromosomal disomy is strain dependent. Interestingly, emergence of heteroresistant clonesin vivowas unrelated to the strain's MIC to fluconazole.

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