Publications by authors named "G Caljon"

110 Publications

Revisiting Pyrazolo[3,4-]pyrimidine Nucleosides as Anti- and Antileishmanial Agents.

J Med Chem 2021 Apr 30;64(7):4206-4238. Epub 2021 Mar 30.

Laboratory for Medicinal Chemistry (Campus Heymans), Ghent University, Ottergemsesteenweg 460, B-9000 Gent, Belgium.

Chagas disease and visceral leishmaniasis are two neglected tropical diseases responsible for numerous deaths around the world. For both, current treatments are largely inadequate, resulting in a continued need for new drug discovery. As both kinetoplastid parasites are incapable of purine synthesis, they depend on purine salvage pathways that allow them to acquire and process purines from the host to meet their demands. Purine nucleoside analogues therefore constitute a logical source of potential antiparasitic agents. Earlier optimization efforts of the natural product tubercidin (7-deazaadenosine) involving modifications to the nucleobase 7-position and the ribofuranose 3'-position led to analogues with potent anti- brucei and anti- activities. In this work, we report the design and synthesis of pyrazolo[3,4-]pyrimidine nucleosides with 3'- and 7-modifications and assess their potential as anti- and antileishmanial agents. One compound was selected for evaluation in an acute Chagas disease mouse model.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1021/acs.jmedchem.1c00135DOI Listing
April 2021

Synthesis, Biological Activity and In Silico Pharmacokinetic Prediction of a New 2-Thioxo-Imidazoldidin-4-One of Primaquine.

Pharmaceuticals (Basel) 2021 Feb 27;14(3). Epub 2021 Feb 27.

OncoPharma Research Group, Center for Health Technology and Services Research (CINTESIS), Rua Dr. Plácido da Costa, 4200-450 Porto, Portugal.

The discovery of novel antiparasitic drugs for neglected tropical diseases (NTDs) constitutes a global urgency and requires a range of innovative strategies to ensure a sustainable pipeline of lead compounds. Thus far, primaquine (PQ) is the only transmission-blocking antimalarial that is clinically available, displaying marked activity against gametocytes of all causative species of human malaria ( spp.). Chagas disease, caused by , is another PQ-sensitive illness besides malaria. One of the major drawbacks of PQ is its metabolism into carboxyprimaquine (CPQ), which is less active than the parent drug. In this study, we developed different synthetic pathways to confer N-protection to PQ through introduction of thioxo-imidazolidin-4-one. The introduction of this group prevents the formation of CPQ, counteracting one major drawback of the parent drug. After that, we evaluated the potential biological activity of the novel 2-thioxo-imidazolidin-4-one derivative of PQ, which showed relevant in vitro activity against (IC 1.4 μM) compared to PQ (IC 1.7 μM) and the reference drug benznidazole (IC 1.6 μM). Noting its acceptable pharmacokinetic profile, this PQ conjugate may be a potential scaffold for novel drug exploration against Chagas disease.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.3390/ph14030196DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7997226PMC
February 2021

Synthesis and evaluation of 3'-fluorinated 7-deazapurine nucleosides as antikinetoplastid agents.

Eur J Med Chem 2021 Feb 16;216:113290. Epub 2021 Feb 16.

Laboratory for Medicinal Chemistry (Campus Heymans), Ghent University, Ottergemsesteenweg 460, B-9000, Gent, Belgium.

Kinetoplastid parasites are the causative agents of neglected tropical diseases with an unmet medical need. These parasites are unable to synthesize the purine ring de novo, and therefore rely on purine salvage to meet their purine demand. Evaluating purine nucleoside analogs is therefore an attractive strategy to identify antikinetoplastid agents. Several anti-Trypanosoma cruzi and anti-Trypanosoma brucei 7-deazapurine nucleosides were previously discovered, with the removal of the 3'-hydroxyl group resulting in a significant boost in activity. In this work we therefore decided to assess the effect of the introduction of a 3'-fluoro substituent in 7-deazapurine nucleosides on the anti-kinetoplastid activities. Hence, we synthesized two series of 3'-deoxy-3'-fluororibofuranosyl and 3'-deoxy-3'-fluoroxylofuranosyl nucleosides comprising 7-deazaadenine and -hypoxanthine bases and assayed these for antiparasitic activity. Several analogs with potent activity against T. cruzi and T. brucei were discovered, indicating that a fluorine atom in the 3'-position is a promising modification for the discovery of antiparasitic nucleosides.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.ejmech.2021.113290DOI Listing
February 2021

Antiplasmodial Oleanane Triterpenoids from Root Bark.

J Nat Prod 2021 Mar 5;84(3):666-675. Epub 2021 Mar 5.

Natural Products & Food Research and Analysis (NatuRA), Department of Pharmaceutical Sciences, University of Antwerp, Universiteitsplein 1, B-2610 Antwerp, Belgium.

Phytochemical investigation of the -BuOH extract of the roots of Sc. Elliot (Combretaceae) led to the isolation and identification of 10 oleanane triterpenoids (-), among which six new compounds, i.e., albidanoside A (), albidic acid A (), albidinolic acid (), albidienic acid (), albidolic acid (), and albidiolic acid (), and two triterpenoid aglycones, i.e., albidic acid B () and albidic acid C (), were isolated here for the first time from a natural source, along with two known compounds. The structures of these constituents were established by means of 1D and 2D NMR spectroscopy and ESI mass spectrometry. The isolated compounds were evaluated for their antiplasmodial and antimicrobial activity against the chloroquine-resistant strain K1, , and . Compounds -, , , and showed moderate antiplasmodial activity with IC values between 5 and 15 μM. None of the tested compounds were active against or . These findings emphasize the potential of as a source for discovery of new antiplasmodial compounds.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1021/acs.jnatprod.0c01119DOI Listing
March 2021

2-aminobenzimidazoles for leishmaniasis: From initial hit discovery to in vivo profiling.

PLoS Negl Trop Dis 2021 Feb 22;15(2):e0009196. Epub 2021 Feb 22.

Institute of Chemistry, University of Campinas (UNICAMP), Campinas-SP, Brazil.

Leishmaniasis is a major infectious disease with hundreds of thousands of new cases and over 20,000 deaths each year. The current drugs to treat this life-threatening infection have several drawbacks such as toxicity and long treatment regimens. A library of 1.8 million compounds, from which the hits reported here are publicly available, was screened against Leishmania infantum as part of an optimization program; a compound was found with a 2-aminobenzimidazole functionality presenting moderate potency, low metabolic stability and high lipophilicity. Several rounds of synthesis were performed to incorporate chemical groups capable of reducing lipophilicity and clearance, leading to the identification of compounds that are active against different parasite strains and have improved in vitro properties. As a result of this optimization program, a group of compounds was further tested in anticipation of in vivo evaluation. In vivo tests were carried out with compounds 29 (L. infantum IC50: 4.1 μM) and 39 (L. infantum IC50: 0.5 μM) in an acute L. infantum VL mouse model, which showed problems of poor exposure and lack of efficacy, despite the good in vitro potency.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1371/journal.pntd.0009196DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7932521PMC
February 2021

Discovery of Diaryl Ether Substituted Tetrahydrophthalazinones as TbrPDEB1 Inhibitors Following Structure-Based Virtual Screening.

Front Chem 2020 21;8:608030. Epub 2021 Jan 21.

Division of Medicinal Chemistry, Amsterdam Institute of Molecular and Life Sciences, Vrije Universiteit Amsterdam, Amsterdam, Netherlands.

Several members of the 3',5'-cyclic nucleotide phosphodiesterase (PDE) family play an essential role in cellular processes, which has labeled them as interesting targets for various diseases. The parasitic protozoan , causative agent of human African trypanosomiasis, contains several cyclic AMP specific PDEs from which TbrPDEB1 is validated as a drug target. The recent discovery of selective TbrPDEB1 inhibitors has increased their potential for a novel treatment for this disease. Compounds characterized by a rigid biphenyl tetrahydrophthalazinone core structure were used as starting point for the exploration of novel TbrPDEB1 inhibitors. Using a virtual screening campaign and structure-guided design, diaryl ether substituted phthalazinones were identified as novel TbrPDEB1 inhibitors with IC values around 1 μM against . This study provides important structure-activity relationship (SAR) information for the future design of effective parasite-specific PDE inhibitors.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.3389/fchem.2020.608030DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7859335PMC
January 2021

Tsetse salivary glycoproteins are modified with paucimannosidic N-glycans, are recognised by C-type lectins and bind to trypanosomes.

PLoS Negl Trop Dis 2021 Feb 2;15(2):e0009071. Epub 2021 Feb 2.

Department of Vector Biology, Liverpool School of Tropical Medicine, Liverpool, United Kingdom.

African sleeping sickness is caused by Trypanosoma brucei, a parasite transmitted by the bite of a tsetse fly. Trypanosome infection induces a severe transcriptional downregulation of tsetse genes encoding for salivary proteins, which reduces its anti-hemostatic and anti-clotting properties. To better understand trypanosome transmission and the possible role of glycans in insect bloodfeeding, we characterized the N-glycome of tsetse saliva glycoproteins. Tsetse salivary N-glycans were enzymatically released, tagged with either 2-aminobenzamide (2-AB) or procainamide, and analyzed by HILIC-UHPLC-FLR coupled online with positive-ion ESI-LC-MS/MS. We found that the N-glycan profiles of T. brucei-infected and naïve tsetse salivary glycoproteins are almost identical, consisting mainly (>50%) of highly processed Man3GlcNAc2 in addition to several other paucimannose, high mannose, and few hybrid-type N-glycans. In overlay assays, these sugars were differentially recognized by the mannose receptor and DC-SIGN C-type lectins. We also show that salivary glycoproteins bind strongly to the surface of transmissible metacyclic trypanosomes. We suggest that although the repertoire of tsetse salivary N-glycans does not change during a trypanosome infection, the interactions with mannosylated glycoproteins may influence parasite transmission into the vertebrate host.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1371/journal.pntd.0009071DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7880456PMC
February 2021

Evaluation of conventional and four real-time PCR methods for the detection of Leishmania on field-collected samples in Ethiopia.

PLoS Negl Trop Dis 2021 Jan 12;15(1):e0008903. Epub 2021 Jan 12.

Department of Clinical Sciences, Institute of Tropical Medicine, Antwerp, Belgium.

In most low-resource settings, microscopy still is the standard method for diagnosis of cutaneous leishmaniasis, despite its limited sensitivity. In Ethiopia, the more sensitive molecular methods are not yet routinely used. This study compared five PCR methods with microscopy on two sample types collected from patients with a suspected lesion to advise on optimal diagnosis of Leishmania aethiopica. Between May and July 2018, skin scrapings (SS) and blood exudate from the lesion spotted on filter paper (dry blood spot, DBS) were collected for PCR from 111 patients of four zones in Southern Ethiopia. DNA and RNA were simultaneously extracted from both sample types. DNA was evaluated by a conventional PCR targeting ITS-1 and three probe-based real-time PCRs: one targeting the SSU 18S rRNA and two targeting the kDNA minicircle sequence (the 'Mary kDNA PCR' and a newly designed 'LC kDNA PCR' for improved L. aethiopica detection). RNAs were tested with a SYBR Green-based RT-PCR targeting spliced leader (SL) RNA. Giemsa-stained SS smears were examined by microscopy. Of the 111 SS, 100 were positive with at least two methods. Sensitivity of microscopy, ITS PCR, SSU PCR, Mary kDNA PCR, LC kDNA PCR and SL RNA PCR were respectively 52%, 22%, 64%, 99%, 100% and 94%. Microscopy-based parasite load correlated well with real-time PCR Ct-values. Despite suboptimal sample storage for RNA detection, the SL RNA PCR resulted in congruent results with low Ct-values. DBS collected from the same lesion showed lower PCR positivity rates compared to SS. The kDNA PCRs showed excellent performance for diagnosis of L. aethiopica on SS. Lower-cost SL RNA detection can be a complementary high-throughput tool. DBS can be used for PCR in case microscopy is negative, the SS sample can be sent to the referral health facility where kDNA PCR method is available.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1371/journal.pntd.0008903DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7802924PMC
January 2021

Synthesis and evaluation of a collection of purine-like C-nucleosides as antikinetoplastid agents.

Eur J Med Chem 2021 Feb 29;212:113101. Epub 2020 Dec 29.

Laboratory for Medicinal Chemistry (Campus Heymans), Ghent University, Ottergemsesteenweg 460, B-9000, Gent, Belgium. Electronic address:

The kinetoplastid parasites Trypanosoma brucei, Trypanosoma cruzi and Leishmania spp. are the causative agents of neglected tropical diseases with a serious burden in several parts of the world. These parasites are incapable of synthesizing purines de novo, and therefore rely on ingenious purine salvage pathways to acquire and process purines from their host. Purine nucleoside analogs that may interfere with these pathways therefore constitute a privileged source of new antikinetoplastid agents. In this study, we synthetized a collection of C-nucleosides employing five different heterocyclic nucleobase surrogates. C-nucleosides are chemically and enzymatically stable and allow for extensive structural modification. Inspired by earlier 7-deazaadenosine nucleosides and known antileishmanial C-nucleosides, we introduced different modifications tailored towards antikinetoplastid activity. Both adenosine and inosine analogs were synthesized with the aim of discovering new antikinetoplastid hits and expanding knowledge of structure-activity relationships. Several promising hits with potent activity against Trypanosoma brucei, Trypanosoma cruzi and Leishmania infantum were discovered, and the nature of the nucleobase surrogate was found to have a profound influence on the selectivity profile of the compounds.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.ejmech.2020.113101DOI Listing
February 2021

Tetrahydrophthalazinone Inhibitor of Phosphodiesterase with Activity against Intracellular Trypanosomatids.

Antimicrob Agents Chemother 2021 02 17;65(3). Epub 2021 Feb 17.

Laboratório de Biologia Celular, Instituto Oswaldo Cruz, Fundação Oswaldo Cruz, Rio de Janeiro, Brazil

The phosphodiesterase inhibitor tetrahydrophthalazinone NPD-008 was explored by phenotypic screening, target validation, and ultrastructural approaches against NPD-008 displayed activity against different forms and strains of (50% effective concentration [EC], 6.6 to 39.5 μM). NPD-008 increased cAMP levels of and its combination with benznidazole gave synergistic interaction. It was also moderately active against intracellular amastigotes of and , confirming a potential activity profile as an antitrypanosomatid drug candidate.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1128/AAC.00960-20DOI Listing
February 2021

2-((3,5-Dinitrobenzyl)thio)quinazolinones: Potent Antimycobacterial Agents Activated by Deazaflavin (F)-Dependent Nitroreductase (Ddn).

J Med Chem 2021 01 21;64(1):440-457. Epub 2020 Dec 21.

Laboratory for Medicinal Chemistry (FFW), Ghent University, Ottergemsesteenweg 460, B-9000 Gent, Belgium.

Swapping the substituents in positions 2 and 4 of the previously synthesized but yet undisclosed 5-cyano-4-(methylthio)-2-arylpyrimidin-6-ones , ring closure, and further optimization led to the identification of the potent antitubercular 2-thio-substituted quinazolinone . Structure-activity relationship (SAR) studies indicated a crucial role for both -nitro substituents for antitubercular activity, while the introduction of polar substituents on the quinazolinone core allowed reduction of bovine serum albumin (BSA) binding (, ). While most of the tested quinazolinones exhibited no cytotoxicity against MRC-5, the most potent compound was found to be mutagenic via the Ames test. This analogue exhibited moderate inhibitory potency against thymidylate kinase, the target of the 3-cyanopyridones that lies at the basis of the current analogues, indicating that the whole-cell antimycobacterial activity of the present -substituted thioquinazolinones is likely due to modulation of alternative or additional targets. Diminished antimycobacterial activity was observed against mutants affected in cofactor F biosynthesis (), cofactor reduction (), or deazaflavin-dependent nitroreductase activity (), indicating that reductive activation of the 3,5-dinitrobenzyl analogues is key to antimycobacterial activity.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1021/acs.jmedchem.0c01374DOI Listing
January 2021

Synthesis and Structure-Activity Relationships of Imidazopyridine/Pyrimidine- and Furopyridine-Based Anti-infective Agents against Trypanosomiases.

ChemMedChem 2021 Mar 11;16(6):966-975. Epub 2020 Nov 11.

QHeteM - Laboratório de Química Heterocíclica e Medicinal, School of Pharmaceutical Sciences of Ribeirão Preto - University of São Paulo, Ribeirão Preto, São Paulo, 14040-903, Brazil.

Neglected tropical diseases remain among the most critical public health concerns in Africa and South America. The drug treatments for these diseases are limited, which invariably leads to fatal cases. Hence, there is an urgent need for new antitrypanosomal drugs. To address this issue, a large number of diverse heterocyclic compounds were prepared. Straightforward synthetic approaches tolerated pre-functionalized structures, giving rise to a structurally diverse set of analogs. We report on a set of 57 heterocyclic compounds with selective activity potential against kinetoplastid parasites. In general, 29 and 19 compounds of the total set could be defined as active against Trypanosoma cruzi and T. brucei brucei, respectively (antitrypanosomal activities <10 μM). The present work discusses the structure-activity relationships of new fused-ring scaffolds based on imidazopyridine/pyrimidine and furopyridine cores. This library of compounds shows significant potential for anti-trypanosomiases drug discovery.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1002/cmdc.202000616DOI Listing
March 2021

A novel serine protease inhibitor as potential treatment for dry eye syndrome and ocular inflammation.

Sci Rep 2020 10 14;10(1):17268. Epub 2020 Oct 14.

Laboratory of Microbiology, Parasitology and Hygiene (LMPH), Faculty of Pharmaceutical, Biomedical and Veterinary Sciences, University of Antwerp, Universiteitsplein 1, 2610, Wilrijk, Belgium.

Dry eye syndrome (DES), a multifactorial disorder which leads to ocular discomfort, visual disturbance and tear film instability, has a rising prevalence and limited treatment options. In this study, a newly developed trypsin-like serine protease inhibitor (UAMC-00050) in a tear drop formulation was evaluated to treat ocular inflammation. A surgical animal model of dry eye was employed to investigate the potential of UAMC-00050 on dry eye pathology. Animals treated with UAMC-00050 displayed a significant reduction in ocular surface damage after evaluation with sodium fluorescein, compared to untreated, vehicle treated and cyclosporine-treated animals. The concentrations of IL-1α and TNF-α were also significantly reduced in tear fluid from UAMC-00050-treated rats. Additionally, inflammatory cell infiltration in the palpebral conjunctiva (CD3 and CD45), was substantially reduced. An accumulation of pro-MMP-9 and a decrease in active MMP-9 were found in tear fluid from animals treated with UAMC-00050, suggesting that trypsin-like serine proteases play a role in activating MMP-9 in ocular inflammation in this animal model. Comparative qRT-PCR analyses on ocular tissue indicated the upregulation of tryptase, urokinase plasminogen activator receptor (uPAR) and protease-activated receptor 2 (PAR2). The developed UAMC-00050 formulation was stable up to 6 months at room temperature in the absence of light, non-irritating and sterile with compatible pH and osmolarity. These results provide a proof-of-concept for the in vivo modifying potential of UAMC-00050 on dry eye pathology and suggest a central role of trypsin-like serine proteases and PAR2 in dry eye derived ocular inflammation.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1038/s41598-020-74159-wDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7560718PMC
October 2020

Correction to: Preparation and Characterization of Nanostructured Lipid Carriers for Improved Topical Drug Delivery: Evaluation in Cutaneous Leishmaniasis and Vaginal Candidiasis Animal Models.

AAPS PharmSciTech 2020 Oct 8;21(7):275. Epub 2020 Oct 8.

Department of Pharmacy, Quaid-i-Azam University, Islamabad, 45320, Pakistan.

In the published manuscript, co-author Sarah Hendrickx name was misspelled and co-author Guy Caljon's last and first names were inadvertently switched.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1208/s12249-020-01819-5DOI Listing
October 2020

An Unbiased Immunization Strategy Results in the Identification of Enolase as a Potential Marker for Nanobody-Based Detection of .

Vaccines (Basel) 2020 Jul 24;8(3). Epub 2020 Jul 24.

Laboratory for Cellular and Molecular Immunology (CMIM), Department of Bioengineering Sciences, Vrije Universiteit Brussel, B-1050 Brussels, Belgium.

is a widely spread parasite that causes the debilitating disease "surra" in several types of ungulates. This severely challenges livestock rearing and heavily weighs on the socio-economic development in the affected areas, which include countries on five continents. Active case finding requires a sensitive and specific diagnostic test. In this paper, we describe the application of an unbiased immunization strategy to identify potential biomarkers for Nanobody (Nb)-based detection of infections. Alpaca immunization with soluble lysates from different strains followed by panning against secretome resulted in the selection of a single Nb (Nb11). By combining Nb11-mediated immuno-capturing with mass spectrometry, the target antigen was identified as the glycolytic enzyme enolase. Four additional anti-enolase binders were subsequently generated by immunizing another alpaca with the recombinant target enzyme. Together with Nb11, these binders were evaluated for their potential use in a heterologous sandwich detection format. Three Nb pairs were identified as candidates for the further development of an antigen-based assay for Nb-mediated diagnosis of infection.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.3390/vaccines8030415DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7565430PMC
July 2020

Sand Fly Studies Predict Transmission Potential of Drug-resistant Leishmania.

Trends Parasitol 2020 09 23;36(9):785-795. Epub 2020 Jul 23.

Laboratory of Microbiology, Parasitology and Hygiene (LMPH), University of Antwerp, Universiteitsplein 1, B-2610 Wilrijk, Belgium. Electronic address:

Leishmania parasites have the capacity to rapidly adapt to changing environments in their digenetic life cycle which alternates between a vertebrate and an invertebrate host. Emergence of resistance following drug exposure can evoke phenotypic alterations that affect several aspects of parasite fitness in both hosts. Current studies of the impact of resistance are mostly limited to interactions with the mammalian host and characterization of in vitro parasite growth and differentiation. Development in the vector and transmission capacity have been largely ignored. This review reflects on the impact of drug resistance on its spreading potential with specific focus on the use of the sand fly infection model to evaluate parasite development in the vector and the ensuing transmission potential of drug-resistant phenotypes.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.pt.2020.06.006DOI Listing
September 2020

A Critical Blimp-1-Dependent IL-10 Regulatory Pathway in T Cells Protects From a Lethal Pro-inflammatory Cytokine Storm During Acute Experimental Infection.

Front Immunol 2020 4;11:1085. Epub 2020 Jun 4.

Research Unit of Cellular and Molecular Immunology, Vrije Universiteit Brussel (VUB), Brussels, Belgium.

In many infectious diseases, the immune response operates as a double-edged sword. While required for protective immunity, infection-induced inflammation can be detrimental if it is not properly controlled, causing collateral body damage and potentially leading to death. It is in this context that the potent anti-inflammatory cytokine interleukin-10 (IL-10) is required to dampen the pro-inflammatory immune response that hallmarks trypanosomosis. Effective control of this infection requires not just the action of antibodies specific for the parasite's variable surface glycoprotein (VSG) coat antigens, but also a pro-inflammatory immune response mediated mainly by IFNγ, TNF, and NO. However, strict control of inflammation is mandatory, as IL-10-deficient mice succumb from an unrestrained cytokine storm within 10 days of a infection. The relevant cellular source of IL-10 and the associated molecular mechanisms implicated in its trypanosomosis associated production are poorly understood. Using an IL-10 reporter mouse strain (Vert-X), we demonstrate here that NK cells, CD8 T cells and CD4 T cells as well as B cells and plasma cells constitute potential cellular sources of IL-10 within the spleen and liver during acute infection. The IL-10 wave follows peak pro-inflammatory cytokine production, which accompanied the control of peak parasitemia. Similar results were observed following conventional experimental needle infection and physiological infections via -infected tsetse flies. Our results show that conditional T cell-specific ablation of the IL-10 regulating gene (encoding for the Blimp-1 transcription factor), leads to an uncontrolled trypanosome-induced pro-inflammatory syndrome like the one observed in infected IL-10-deficient mice. This result indicates that the biological role of IL-10-derived from non-T cells, including NK cells, is of minor importance when considering host survival. The cytokine IL-27 that is also considered to be an IL-10 regulator, did not affect IL-10 production during infection. Together, these data suggest that activates a Blimp-1-dependent IL-10 regulatory pathway in T cells that acts as a critical anti-inflammatory rheostat, mandatory for host survival during the acute phase of parasitemia.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.3389/fimmu.2020.01085DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7325990PMC
March 2021

Synthesis and structure activity relationships of cyanopyridone based anti-tuberculosis agents.

Eur J Med Chem 2020 Sep 12;201:112450. Epub 2020 Jun 12.

Laboratory for Medicinal Chemistry (FFW), Ghent University, Ottergemsesteenweg 460, B9000, Gent, Belgium. Electronic address:

Mycobacterium tuberculosis, the causative agent of tuberculosis, relies on thymidylate kinase (MtbTMPK) for the synthesis of thymidine triphosphates and thus also DNA synthesis. Therefore, this enzyme constitutes a potential Achilles heel of the pathogen. Based on a previously reported MtbTMPK 6-aryl-substituted pyridone inhibitor and guided by two co-crystal structures of MtbTMPK with pyridone- and thymine-based inhibitors, we report the synthesis of a series of aryl-shifted cyanopyridone analogues. These compounds generally lacked significant MtbTMPK inhibitory potency, but some analogues did exhibit promising antitubercular activity. Analogue 11i demonstrated a 10-fold increased antitubercular activity (MIC H37Rv, 1.2 μM) compared to literature compound 5. Many analogues with whole-cell antimycobacterial activity were devoid of significant cytotoxicity.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.ejmech.2020.112450DOI Listing
September 2020

Antileishmanial Aminopyrazoles: Studies into Mechanisms and Stability of Experimental Drug Resistance.

Antimicrob Agents Chemother 2020 08 20;64(9). Epub 2020 Aug 20.

Laboratory of Microbiology, Parasitology and Hygiene (LMPH), University of Antwerp, Antwerp, Belgium

Current antileishmanial treatment is hampered by limitations, such as drug toxicity and the risk of treatment failure, which may be related to parasitic drug resistance. Given the urgent need for novel drugs, the Drugs for Neglected Diseases (DND) has undertaken a drug discovery program, which has resulted in the identification of aminopyrazoles, a highly promising antileishmanial chemical series. Multiple experiments have been performed to anticipate the propensity for resistance development. Resistance selection was performed by successive exposure of promastigotes () and intracellular amastigotes (both and in golden Syrian hamsters). The stability of the resistant phenotypes was assessed after passage in mice and sandflies. Whole-genome sequencing (WGS) was performed to identify mutated genes, copy number variations (CNVs), and somy changes. The potential role of efflux pumps (the MDR and MRP efflux pumps) in the development of resistance was assessed by coincubation of aminopyrazoles with specific efflux pump inhibitors (verapamil, cyclosporine, and probenecid). Repeated drug exposure of amastigotes did not result in the emergence of drug resistance either or Selection at the promastigote stage, however, was able to select for parasites with reduced susceptibility (resistance index, 5.8 to 24.5). This phenotype proved to be unstable after passage in mice and sandflies, suggesting that nonfixed alterations are responsible for the elevated resistance. In line with this, single nucleotide polymorphisms and indels identified by whole-genome sequencing could not be directly linked to the decreased drug susceptibility. Copy number variations were absent, whereas somy changes were detected, which may have accounted for the transient acquisition of resistance. Finally, aminopyrazole activity was not influenced by the MDR and MRP efflux pump inhibitors tested. The selection performed does not suggest the rapid development of resistance against aminopyrazoles in the field. Karyotype changes may confer elevated levels of resistance, but these do not seem to be stable in the vertebrate and invertebrate hosts. MDR/MRP efflux pumps are not likely to significantly impact the activity of the aminopyrazole leads.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1128/AAC.00152-20DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7449183PMC
August 2020

Experimental Strategies to Explore Drug Action and Resistance in Kinetoplastid Parasites.

Microorganisms 2020 Jun 24;8(6). Epub 2020 Jun 24.

Laboratory of Microbiology, Parasitology and Hygiene (LMPH), University of Antwerp, 2610 Wilrijk, Belgium.

Kinetoplastids are the causative agents of leishmaniasis, human African trypanosomiasis, and American trypanosomiasis. They are responsible for high mortality and morbidity in (sub)tropical regions. Adequate treatment options are limited and have several drawbacks, such as toxicity, need for parenteral administration, and occurrence of treatment failure and drug resistance. Therefore, there is an urgency for the development of new drugs. Phenotypic screening already allowed the identification of promising new chemical entities with anti-kinetoplastid activity potential, but knowledge on their mode-of-action (MoA) is lacking due to the generally applied whole-cell based approach. However, identification of the drug target is essential to steer further drug discovery and development. Multiple complementary techniques have indeed been used for MoA elucidation. In this review, the different 'omics' approaches employed to define the MoA or mode-of-resistance of current reference drugs and some new anti-kinetoplastid compounds are discussed.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.3390/microorganisms8060950DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7356981PMC
June 2020

Interferon Alpha Favors Macrophage Infection by Visceral Species Through Upregulation of Sialoadhesin Expression.

Front Immunol 2020 9;11:1113. Epub 2020 Jun 9.

Laboratory of Microbiology, Parasitology and Hygiene (LMPH), Faculty of Pharmaceutical, Biomedical and Veterinary Sciences, University of Antwerp, Wilrijk, Belgium.

Type I interferons (IFNs) induced by an endogenous RNA virus or exogenous viral infections have been shown to exacerbate infections with New World Cutaneous parasites, however, the impact of type I IFNs in visceral infections and implicated mechanisms remain to be unraveled. This study assessed the impact of type I IFN on macrophage infection with and and the implication of sialoadhesin (Siglec-1/CD169, Sn) as an IFN-inducible surface receptor. Stimulation of bone marrow-derived macrophages with type I IFN (IFN-α) significantly enhanced susceptibility to infection of reference laboratory strains and a set of recent clinical isolates. IFN-α particularly enhanced promastigote uptake. Enhanced macrophage susceptibility was linked to upregulated Sn surface expression as a major contributing factor to the infection exacerbating effect of IFN-α. Stimulation experiments in Sn-deficient macrophages, macrophage pretreatment with a monoclonal anti-Sn antibody or a novel bivalent anti-Sn nanobody and blocking of parasites with soluble Sn restored normal susceptibility levels. Infection of Sn-deficient mice with bioluminescent promastigotes revealed a moderate, strain-dependent role for Sn during visceral infection under the used experimental conditions. These data indicate that IFN-responsive Sn expression can enhance the susceptibility of macrophages to infection with visceral promastigotes and that targeting of Sn may have some protective effects in early infection.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.3389/fimmu.2020.01113DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7296180PMC
April 2021

Structure-Activity Relationship Exploration of 3'-Deoxy-7-deazapurine Nucleoside Analogues as Anti- Agents.

ACS Infect Dis 2020 08 8;6(8):2045-2056. Epub 2020 Jul 8.

Laboratory for Medicinal Chemistry (Campus Heymans), Ghent University, Ottergemsesteenweg 460, B-9000 Gent, Belgium.

Human African trypanosomiasis is a neglected tropical disease caused by parasites. These protists are unable to produce the purine ring, making them vulnerable to the effects of purine nucleoside analogues. Starting from 3'-deoxytubercidin (), a lead compound with activity against central-nervous-stage human African trypanosomiasis, we investigate the structure-activity relationships of the purine and ribofuranose rings. The purine ring tolerated only modifications at C7, while from the many alterations of the 3'-deoxyribofuranosyl moiety only the arabino analogue showed pronounced antitrypanosomal activity. Profiling of the most potent analogues against resistant strains (resistant to pentamidine, diminazene, and isometamidium) showed reduced dependence on uptake mediated by the P2 aminopurine transporter relative to . The introduction of a 7-substituent confers up to 10-fold increased affinity for the P1 nucleoside transporter while generally retaining high affinity for P2. Four of the most promising analogues were found to be metabolically stable, earmarking them as suitable backup analogues for lead .
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1021/acsinfecdis.0c00105DOI Listing
August 2020

1-(1-Arylethylpiperidin-4-yl)thymine Analogs as Antimycobacterial TMPK Inhibitors.

Molecules 2020 Jun 17;25(12). Epub 2020 Jun 17.

Laboratory for Medicinal Chemistry (FFW), Ghent University, Ottergemsesteenweg 460, B-9000 Gent, Belgium.

A series of TMPK (TMPK) inhibitors based on a reported compound were synthesized and evaluated for their capacity to inhibit TMPK catalytic activity and the growth of a virulent strain (H37Rv). Modifications of the scaffold of failed to afford substantial improvements in TMPK inhibitory activity and antimycobacterial activity. Optimization of the substitution pattern of the D ring of resulted in compound with improved TMPK inhibitory potency (three-fold) and H37Rv growth inhibitory activity (two-fold). Moving the 3-chloro substituent of to the -position afforded isomer , which, despite a 10-fold increase in IC-value, displayed promising whole cell activity (minimum inhibitory concentration (MIC) = 12.5 μM).
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.3390/molecules25122805DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7356956PMC
June 2020

Evaluation of a pan-Leishmania SL RNA qPCR assay for parasite detection in laboratory-reared and field-collected sand flies and reservoir hosts.

Parasit Vectors 2020 Jun 1;13(1):276. Epub 2020 Jun 1.

Laboratory of Microbiology, Parasitology and Hygiene, University of Antwerp, Wilrijk, Belgium.

Background: In eco-epidemiological studies, Leishmania detection in vectors and reservoirs is frequently accomplished by high-throughput and sensitive molecular methods that target minicircle kinetoplast DNA (kDNA). A pan-Leishmania SYBR green quantitative PCR (qPCR) assay which detects the conserved spliced-leader RNA (SL RNA) sequence was developed recently. This study assessed the SL RNA assay performance combined with a crude extraction method for the detection of Leishmania in field-collected and laboratory-reared sand flies and in tissue samples from hyraxes as reservoir hosts.

Methods: Field-collected and laboratory-infected sand fly and hyrax extracts were subjected to three different qPCR approaches to assess the suitability of the SL RNA target for Leishmania detection. Nucleic acids of experimentally infected sand flies were isolated with a crude extraction buffer with ethanol precipitation and a commercial kit and tested for downstream DNA and RNA detection. Promastigotes were isolated from culture and sand fly midguts to assess whether there was difference in SL RNA and kDNA copy numbers. Naive sand flies were spiked with a serial dilution of promastigotes to make a standard curve.

Results: The qPCR targeting SL RNA performed well on infected sand fly samples, despite preservation and extraction under presumed unfavorable conditions for downstream RNA detection. Nucleic acid extraction by a crude extraction buffer combined with a precipitation step was highly compatible with downstream SL RNA and kDNA detection. Copy numbers of kDNA were found to be identical in culture-derived parasites and promastigotes isolated from sand fly midguts. SL RNA levels were slightly lower in sand fly promastigotes (ΔCq 1.7). The theoretical limit of detection and quantification of the SL RNA qPCR respectively reached down to 10 and 10 parasite equivalents. SL RNA detection in stored hyrax samples was less efficient with some false-negative assay results, most likely due to the long-term tissue storage in absence of RNA stabilizing reagents.

Conclusions: This study shows that a crude extraction method in combination with the SL RNA qPCR assay is suitable for the detection and quantification of Leishmania in sand flies. The assay is inexpensive, sensitive and pan-Leishmania specific, and accordingly an excellent assay for high-throughput screening in entomological research.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1186/s13071-020-04141-yDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7268266PMC
June 2020

Inflammation following trypanosome infection and persistence in the skin.

Curr Opin Immunol 2020 10 20;66:65-73. Epub 2020 May 20.

Laboratory of Microbiology, Parasitology and Hygiene (LMPH), University of Antwerp, Wilrijk, Belgium. Electronic address:

Human African trypanosomes rely for their transmission on tsetse flies (Glossina sp.) that inoculate parasites into the skin during blood feeding. The absence of a protective vaccine, limited knowledge about the infection immunology, and the existence of asymptomatic carriers sustaining transmission are major outstanding challenges towards elimination. All these relate to the skin where (i) parasites persist and transmit to tsetse flies and (ii) a successful vaccination strategy should ideally be effective. Host immune processes and parasite strategies that underlie early infection and skin tropism are essential aspects to comprehend the transmission-success of trypanosomes and the failure in vaccine development. Recent insights into the early infection establishment may pave the way to novel strategies aimed at blocking transmission.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.coi.2020.04.006DOI Listing
October 2020

Impact of clinically acquired miltefosine resistance by Leishmania infantum on mouse and sand fly infection.

Int J Parasitol Drugs Drug Resist 2020 08 1;13:16-21. Epub 2020 May 1.

Laboratory of Microbiology, Parasitology and Hygiene (LMPH), University of Antwerp, Universiteitsplein 1, B-2610, Wilrijk, Belgium. Electronic address:

Objectives: This study evaluated the implications of clinically acquired miltefosine resistance (MIL-R) by assessing virulence in mice and sand flies to reveal the potential of MIL-R strains to circulate.

Methods: Experimental infections with the MIL-R clinical Leishmania infantum isolate MHOM/FR/2005/LEM5159, having a defect in the LiROS3 subunit of the MIL-transporter, and its syngeneic experimentally reconstituted MIL-S counterpart (LEM5159) were performed in BALB/c mice and Lutzomyia longipalpis and Phlebotomus perniciosus sand flies. In mice, the amastigote burdens in liver and spleen were compared microscopically using Giemsa smears and by bioluminescent imaging. During the sand fly infections, the percentage of infected flies, parasite load, colonization of the stomodeal valve and metacyclogenesis were evaluated. The stability of the MIL-R phenotype after sand fly and mouse passage was determined as well.

Results: The fitness of the MIL-R strain differed between the mouse and sand fly infection model. In mice, a clear fitness loss was observed compared to the LiROS3-reconstituted susceptible strain. This defect could be rescued by episomal reconstitution with a wildtype LiROS3 copy. However, this fitness loss was not apparent in the sand fly vector, resulting in metacyclogenesis and efficient colonization of the stomodeal valve. Resistance was stable after passage in both sand fly and mouse.

Conclusion: The natural MIL-R strain is significantly hampered in its ability to multiply and cause a typical visceral infection pattern in BALB/c mice. However, this LiROS3-deficient strain efficiently produced mature infections and metacyclic promastigotes in the sand fly vector highlighting the transmission potential of this particular MIL-R clinical Leishmania strain.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.ijpddr.2020.04.004DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7215113PMC
August 2020

Comparative evaluation of nucleic acid stabilizing reagents for RNA- and DNA-based Leishmania detection in blood as proxy for visceral burdens.

J Microbiol Methods 2020 06 4;173:105935. Epub 2020 May 4.

Laboratory of Microbiology, Parasitology and Hygiene (LMPH), University of Antwerp, Wilrijk, Belgium. Electronic address:

Background: Molecular detection techniques using peripheral blood are preferred over invasive tissue aspiration for the diagnosis and post-treatment follow-up of visceral leishmaniasis (VL) patients. This study aims to identify suitable stabilizing reagents to prevent DNA and RNA degradation during storage and transport to specialized laboratories where molecular diagnosis is performed.

Methodology: The stabilizing capacities of different commercially available reagents were compared using promastigote-spiked human blood and peripheral blood of Syrian golden hamsters subjected to experimental infection, treatment (miltefosine or aminopyrazole DNDi-1044) and immunosuppression. The impact of various storage temperature conditions was tested in combination with an established kinetoplast DNA (kDNA) qPCR and a recently developed spliced leader RNA (SL-RNA) assay for Leishmania detection.

Principal Findings: Irrespective of the blood type and stabilizer used, threshold (cT) values obtained with the SL-RNA qPCR were systematically lower than those obtained with the kDNA assay, confirming the advantage of the SL-RNA assay over the widely used kDNA assay for low-level Leishmania detection. Peripheral blood parasite levels correlated relatively well with hepatic burdens. RNA protect cell reagent provided the most optimal simultaneous DNA and RNA stabilization in both human and hamster blood. However, this stabilizer requires an erythrocyte lysis step, which can be challenging under field conditions. DNA/RNA shield provides a good alternative for downstream kDNA and SL-RNA assays, especially if sample storage capacity at 4 °C can be guaranteed.

Conclusions/significance: The recommended stabilizing reagents are compatible with RNA- and DNA-based Leishmania detection in peripheral blood in the VL hamster model and spiked human blood. Since molecular detection techniques using peripheral blood are less invasive than microscopic assessment of tissue aspirates, the findings of this study may be applied to human VL clinical studies.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.mimet.2020.105935DOI Listing
June 2020

Structure-Activity Relationship of Phenylpyrazolones against Trypanosoma cruzi.

ChemMedChem 2020 07 27;15(14):1310-1321. Epub 2020 Apr 27.

Division of Medicinal Chemistry, Faculty of Sciences, Amsterdam Institute for Molecules, Medicines and Systems (AIMMS), Vrije Universiteit Amsterdam, De Boelelaan 1108, 1081 HZ, Amsterdam, The Netherlands.

Chagas disease is a neglected parasitic disease caused by the parasitic protozoan Trypanosoma cruzi and currently affects around 8 million people. Previously, 2-isopropyl-5-(4-methoxy-3-(pyridin-3-yl)phenyl)-4,4-dimethyl-2,4-dihydro-3H-pyrazol-3-one (NPD-0227) was discovered to be a sub-micromolar inhibitor (pIC =6.4) of T. cruzi. So far, SAR investigations of this scaffold have focused on the alkoxy substituent, the pyrazolone nitrogen substituent and the aromatic substituent of the core phenylpyrazolone. In this study, modifications of the phenyldihydropyrazolone scaffold are described. Variations were introduced by installing different substituents on the phenyl core, modifying the geminal dimethyl and installing various bio-isosteres of the dihydropyrazolone group. The anti T. cruzi activity of NPD-0227 could not be surpassed as the most potent compounds show pIC values of around 6.3. However, valuable additional SAR data for this interesting scaffold was obtained, and the data suggest that a scaffold hop is feasible as the pyrazolone moiety can be replaced by a oxazole or oxadiazole with minimal loss of activity.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1002/cmdc.202000136DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7496920PMC
July 2020

In Vitro Growth Inhibition Assays of Leishmania spp.

Methods Mol Biol 2020 ;2116:791-800

Laboratory of Microbiology, Parasitology and Hygiene (LMPH), University of Antwerp, Wilrijk-Antwerp, Belgium.

In vitro growth (inhibition) assays have a dual application, either supporting the discovery of novel drugs or as a monitoring tool of drug resistance in patient isolates. From an experimental design point of view, both are quite different with regard to the infecting Leishmania species and strain, the wide variety of permissive host cells (primary cells versus cell lines), drug exposure times, detection methods and endpoint criteria. Recognizing the need for enhanced assay standardization to decrease interlaboratory variation and improve proper interpretation of results, a detailed description is given of the basic fundamental procedures and requirements for routine in vitro growth of Leishmania spp. with specific focus on the intracellular amastigote susceptibility assay. Although the described experimental procedures focus on visceral Leishmania species, the same assay principles may apply for the cutaneous species as well.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1007/978-1-0716-0294-2_47DOI Listing
February 2021

Feeding behavior and activity of Phlebotomus pedifer and potential reservoir hosts of Leishmania aethiopica in southwestern Ethiopia.

PLoS Negl Trop Dis 2020 03 20;14(3):e0007947. Epub 2020 Mar 20.

Biology Department, Arba Minch University, Arba Minch, Ethiopia.

Background: Cutaneous leishmaniasis (CL) is a major public health concern in Ethiopia. However, knowledge about the complex zoonotic transmission cycle is limited, hampering implementation of control strategies. We explored the feeding behavior and activity of the vector (Phlebotomus pedifer) and studied the role of livestock in CL transmission in southwestern Ethiopia.

Methods: Blood meal origins of engorged sand flies were determined by sequencing host DNA. A host choice experiment was performed to assess the feeding preference of P. pedifer when humans and hyraxes are equally accessible. Ear and nose biopsies from livestock were screened for the presence of Leishmania parasites. Sand flies were captured indoor and outdoor with human landing catches and CDC light traps to determine at which time and where P. pedifer is mostly active.

Principal Findings: A total of 180 P. pedifer sand flies were found to bite hosts of 12 genera. Humans were the predominant blood meal source indoors (65.9%, p < 0.001), while no significant differences were determined outdoors and in caves. In caves, hyraxes were represented in blood meals equally as humans (45.5% and 42.4%, respectively), but the host choice experiment revealed that sand flies have a significant preference for feeding on hyraxes (p = 0.009). Only a single goat nose biopsy from 412 animal samples was found with Leishmania RNA. We found that P. pedifer is predominantly endophagic (p = 0.003), but occurs both indoors and outdoors. A substantial number of sand flies was active in the early evening, which increased over time reaching its maximum around midnight.

Conclusion: In contrast to earlier suggestions of exclusive zoonotic Leishmania transmission, we propose that there is also human-to-human transmission of CL in southwestern Ethiopia. Livestock does not play a role in CL transmission and combined indoor and outdoor vector control measures at night are required for efficient vector control.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1371/journal.pntd.0007947DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7112221PMC
March 2020