scholarly journals An aromatic imidazoline derived from chloroquinoline triggers cell cycle arrest and inhibits with high selectivity the Trypanosoma cruzi mammalian host-cells infection

2021 ◽  
Vol 15 (11) ◽  
pp. e0009994
Author(s):  
Roberto I. Cuevas-Hernández ◽  
Richard M. B. M. Girard ◽  
Luka Krstulović ◽  
Miroslav Bajić ◽  
Ariel Mariano Silber
Keyword(s):  
Cell Cycle ◽  
Trypanosoma Cruzi ◽  
Host Cells ◽  
Mammalian Host ◽  
Cycle Arrest ◽  
Mitochondrial Inner Membrane ◽  
Intracellular Ca ◽  
Low Toxicity ◽  
Plasma Membrane Permeabilization ◽  
Chimeric Molecules

Trypanosoma cruzi is a hemoflagellated parasite causing Chagas disease, which affects 6–8 million people in the Americas. More than one hundred years after the description of this disease, the available drugs for treating the T. cruzi infection remain largely unsatisfactory. Chloroquinoline and arylamidine moieties are separately found in various compounds reported for their anti-trypanosoma activities. In this work we evaluate the anti-T. cruzi activity of a collection of 26 “chimeric” molecules combining choroquinoline and amidine structures. In a first screening using epimastigote forms of the parasite as a proxy for the clinically relevant stages, we selected the compound 7-chloro-4-[4-(4,5-dihydro-1H-imidazol-2-yl)phenoxy]quinoline (named here as A6) that performed better as an anti-T. cruzi compound (IC50 of 2.2 ± 0.3 μM) and showed a low toxicity for the mammalian cell CHO-K1 (CC50 of 137.9 ± 17.3 μM). We initially investigated the mechanism of death associated to the selected compound. The A6 did not trigger phosphatidylserine exposure or plasma membrane permeabilization. Further investigation led us to observe that under short-term incubations (until 6 hours), no alterations of mitochondrial function were observed. However, at longer incubation times (4 days), A6 was able to decrease the intracellular Ca2+, to diminish the intracellular ATP levels, and to collapse mitochondrial inner membrane potential. After analysing the cell cycle, we found as well that A6 produced an arrest in the S phase that impairs the parasite proliferation. Finally, A6 was effective against the infective forms of the parasite during the infection of the mammalian host cells at a nanomolar concentration (IC50(tryps) = 26.7 ± 3.7 nM), exhibiting a selectivity index (SI) of 5,170. Our data suggest that A6 is a promising hit against T. cruzi.

scholarly journals An Aromatic Diamidine That Targets Kinetoplast DNA, Impairs the Cell Cycle in Trypanosoma cruzi, and Diminishes Trypomastigote Release from Infected Mammalian Host Cells

2016 ◽  
Vol 60 (10) ◽  
pp. 5867-5877 ◽  
Author(s):  
Richard M. B. M. Girard ◽  
Marcell Crispim ◽  
Ivana Stolić ◽  
Flávia Silva Damasceno ◽  
Marcelo Santos da Silva ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Trypanosoma Cruzi ◽  
Nuclear Dna ◽  
Host Cells ◽  
Mammalian Host ◽  
Kinetoplast Dna ◽  
Content Type ◽  
Parasite Cell ◽  
Infection Index ◽  
Dependent Inhibition

ABSTRACTTrypanosoma cruziis the etiological agent of Chagas disease, affecting approximately 10 million people in the Americas and with some 40 million people at risk. The objective of this study was to evaluate the anti-T. cruziactivity of three new diamidines that have a 3,4-ethylenedioxy extension of the thiophene core, designated MB17, MB19, and MB38. All three diamidines exhibited dose-dependent inhibition of epimastigote replication. The mechanisms of action of these diamidines were investigated. Unlike MB17 and MB19, MB38 exhibited a significant increase in the number of annexin-propidium iodide double-labeled cells compared to levels in control parasites. As MB17 had shown a lower 50% inhibitory concentration (IC50) against epimastigote growth, the mechanism of action of this drug was studied in more detail. MB17 triggered a decrease in the intracellular ATP levels. As a consequence, MB17 affected the genomic DNA and kinetoplast DNA (kDNA) and impaired the parasite cell cycle. Moreover, MB17 caused DNA fragmentation, with a more severe effect on kDNA than on nuclear DNA, resulting in dyskinetoplastic cells. MB17 was tested for toxicity and effectiveness for the treatment of infected CHO-K1cells, exhibiting a 50% cytotoxic concentration (CC50) of 13.47 ± 0.37 μM and an IC50of 0.14 ± 0.12 μM against trypomastigote release. MB17 also diminished the infection index by 60% at 0.5 μM. In conclusion, despite belonging to the same family, these diamidines have different efficiencies. To summarize, MB17 was the most potent of these diamidines against epimastigotes, producing DNA damage preferentially in kDNA, impairing the parasite cell cycle, and decreasing the infection index and trypomastigote release from infected mammalian host cells, with a high selectivity index (SI) (<90). These data suggest that MB17 could be an interesting lead compound againstT. cruzi.

Differentiation of African trypanosomes is controlled by a density sensing mechanism which signals cell cycle arrest via the cAMP pathway

1997 ◽  
Vol 110 (21) ◽  
pp. 2661-2671 ◽  
Author(s):  
E. Vassella ◽  
B. Reuner ◽  
B. Yutzy ◽  
M. Boshart
Keyword(s):  
Cell Cycle ◽  
Cell Cycle Arrest ◽  
Mammalian Host ◽  
Intracellular Camp ◽  
Quiescent State ◽  
Camp Content ◽  
African Trypanosomes ◽  
Sensing Mechanism ◽  
Camp Signalling ◽  
Cycle Arrest

Differentiation of African trypanosomes from replicating slender bloodstream forms to nondividing stumpy forms limits the parasite population size, allowing survival of the mammalian host and establishment of a stable host-parasite relationship. Using a novel in vitro culture system we have shown that slender to stumpy differentiation is induced by parasite density alone and thus is independent of host cues. Here we investigate the density sensing mechanism and show that trypanosomes release a soluble activity of low relative molecular mass, termed stumpy induction factor (SIF), which accumulates in conditioned medium. SIF activity triggers cell cycle arrest in G1/G0 phase and induces differentiation with high efficiency and rapid kinetics. Membrane-permeable derivates of cAMP or the phosphodiesterase inhibitor etazolate perfectly mimic SIF activity. Furthermore, SIF activity elicits an immediate two- to threefold elevation of intracellular cAMP content upon addition to slender forms. We conclude that SIF and hence density sensing operate through the cAMP signalling pathway. Temporal correlation of markers indicates that cell cycle arrest invariably precedes differentiation. Thus, our results indicate that the cell cycle regulation of bloodstream forms is under dominant control of cAMP signalling. Irreversible commitment to the quiescent state is elicited by a cAMP agonist within a period shorter than one complete cell cycle.

scholarly journals Mammalian cell invasion and intracellular trafficking by Trypanosoma cruzi infective forms

2005 ◽  
Vol 77 (1) ◽  
pp. 77-94 ◽  
Author(s):  
Renato A. Mortara ◽  
Walter K. Andreoli ◽  
Noemi N. Taniwaki ◽  
Adriana B. Fernandes ◽  
Claudio V. da Silva ◽  
...  
Keyword(s):  
Trypanosoma Cruzi ◽  
Host Cell ◽  
Mammalian Cells ◽  
Parasitophorous Vacuole ◽  
Host Cells ◽  
Mammalian Host ◽  
Target Cells ◽  
Sylvatic Cycle ◽  
Final Destination ◽  
Different Strains

Trypanosoma cruzi, the etiological agent of Chagas’ disease, occurs as different strains or isolates that may be grouped in two major phylogenetic lineages: T. cruzi I, associated with the sylvatic cycle and T. cruzi II, linked to the human disease. In the mammalian host the parasite has to invade cells and many studies implicated the flagellated trypomastigotes in this process. Several parasite surface components and some of host cell receptors with which they interact have been identified. Our work focused on how amastigotes, usually found growing in the cytoplasm, can invade mammalian cells with infectivities comparable to that of trypomastigotes. We found differences in cellular responses induced by amastigotes and trypomastigotes regarding cytoskeletal components and actin-rich projections. Extracellularly generated amastigotes of T. cruzi I strains may display greater infectivity than metacyclic trypomastigotes towards cultured cell lines as well as target cells that have modified expression of different classes of cellular components. Cultured host cells harboring the bacterium Coxiella burnetii allowed us to gain new insights into the trafficking properties of the different infective forms of T. cruzi, disclosing unexpected requirements for the parasite to transit between the parasitophorous vacuole to its final destination in the host cell cytoplasm.

scholarly journals Palmitic Acid Methyl Ester Induces G2/M Arrest in Human Bone Marrow-Derived Mesenchymal Stem Cells via the p53/p21 Pathway

2019 ◽  
Vol 2019 ◽  
pp. 1-15
Author(s):  
Jian-Hong Lin ◽  
Pei-Ching Ting ◽  
Wen-Sen Lee ◽  
Hung-Wen Chiu ◽  
Chun-An Chien ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Bone Marrow ◽  
P53 Protein ◽  
Acid Methyl Ester ◽  
Cyclin B1 ◽  
Cycle Arrest ◽  
Acid Methyl ◽  
M Phase ◽  
Intracellular Ca ◽  
Palmitic Acid Methyl Ester

Bone marrow-derived mesenchymal cells (BM-MSCs) are able to differentiate into adipocytes, which can secrete adipokines to affect BM-MSC proliferation and differentiation. Recent evidences indicated that adipocytes can secrete fatty acid metabolites, such as palmitic acid methyl ester (PAME), which is able to cause vasorelaxation and exerts anti-inflammatory effects. However, effects of PAME on BM-MSC proliferation remain unclear. The aim of this study was to investigate the effect of PAME on human BM-MSC (hBM-MSC) proliferation and its underlying molecular mechanisms. hBM-MSCs were treated with PAME for 48 h and then subjected to various analyses. The results from the present study show that PAME significantly reduced the levels of G2/M phase regulatory proteins, cyclin-dependent kinase 1 (Cdk1), and cyclin B1 and inhibited proliferation in hBM-MSCs. Moreover, the level of Mdm2 protein decreased, while the levels of p21 and p53 protein increased in the PAME-treated hBM-MSCs. However, PAME treatment did not significantly affect apoptosis/necrosis, ROS generation, and the level of Cdc25C protein. PAME also induced intracellular acidosis and increased intracellular Ca2+ levels. Cotreatment with PAME and Na+/H+ exchanger inhibitors together further reduced the intracellular pH but did not affect the PAME-induced decreases of cell proliferation and increases of the cell population at the G2/M phase. Cotreatment with PAME and a calcium chelator together inhibited the PAME-increased intracellular Ca2+ levels but did not affect the PAME-induced cell proliferation inhibition and G2/M cell cycle arrest. Moreover, the half-life of p53 protein was prolonged in the PAME-treated hBM-MSCs. Taken together, these results suggest that PAME induced p53 stabilization, which in turn increased the levels of p53/p21 proteins and decreased the levels of Cdk1/cyclin B1 proteins, thereby preventing the activation of Cdk1, and eventually caused cell cycle arrest at the G2/M phase. The findings from the present study might help get insight into the physiological roles of PAME in regulating hBM-MSC proliferation.

scholarly journals Inhibition of Trypanosoma cruzi growth in mammalian cells by purine and pyrimidine analogs.

1996 ◽  
Vol 40 (11) ◽  
pp. 2455-2458 ◽  
Author(s):  
J Nakajima-Shimada ◽  
Y Hirota ◽  
T Aoki
Keyword(s):  
Trypanosoma Cruzi ◽  
Growth Inhibition ◽  
Host Cell ◽  
Culture System ◽  
Mammalian Cells ◽  
Screening Method ◽  
Host Cells ◽  
Mammalian Host ◽  
Dependent Manner ◽  
Pyrimidine Analogs

Trypanosoma cruzi, the causative agent of Chagas' disease, exhibits two different developmental stages in mammals, the amastigote, an intracellular form that proliferates in the cytoplasm of host cells, and the trypomastigote, an extracellular form that circulates in the bloodstream. We have already established an in vitro culture system using mammalian host cells (HeLa) infected with T. cruzi in which the time course of parasite growth is determined quantitatively. We adopted this system for the screening of anti-T. cruzi agents that would ideally prove to be effective against trypanosomes with no toxicity to the host cell. Of the purine analogs tested, allopurinol markedly inhibited the growth of amastigotes in a dose-dependent manner, with no lethal effect on trypomastigotes. 3'-Deoxyinosine and 3'-deoxyadenosine also suppressed T. cruzi growth inside the host cell, with the concentrations causing 50% growth inhibition being 10 and 5 microM, respectively, in contrast to a concentration causing 50% growth inhibition of 3 microM for allopurinol. Among the pyrimidine analogs examined, 3'-azido-3'-deoxythymidine (zidovudine) significantly reduced the growth of the parasite at concentrations as low as 1 microM. The anti-human immunodeficiency virus agents 2',3'-dideoxyinosine and 2',3'-dideoxyadenosine caused a decrease in amastigote growth, while 2',3'-dideoxycytidine and 2',3'-dideoxyuridine had no inhibitory effect. When Swiss 3T3 fibroblasts were used as host cells, allopurinol, 3'-deoxyinosine, 3'-deoxyadenosine, and 3'-azid-3'-deoxythymidine also markedly inhibited T. cruzi proliferation. These results indicate that our culture system is useful as a primary screening method for candidate compounds against T. cruzi on the basis of two criteria, namely, intracellular replication by the parasite and host-cell infection rate.

How Trypanosoma cruzi handles cell cycle arrest promoted by camptothecin, a topoisomerase I inhibitor

2014 ◽  
Vol 193 (2) ◽  
pp. 93-100 ◽  
Author(s):  
Aline Araujo Zuma ◽  
Isabela Cecília Mendes ◽  
Lissa Catherine Reignault ◽  
Maria Carolina Elias ◽  
Wanderley de Souza ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Trypanosoma Cruzi ◽  
Cell Cycle Arrest ◽  
Topoisomerase I ◽  
Topoisomerase I Inhibitor ◽  
Cycle Arrest

scholarly journals Efficacy of Novel Pyrazolone Phosphodiesterase Inhibitors in Experimental Mouse Models of Trypanosoma cruzi

2020 ◽  
Vol 64 (9) ◽  
Author(s):  
Julianna Siciliano de Araújo ◽  
Cristiane França da Silva ◽  
Denise da Gama Jaén Batista ◽  
Aline Nefertiti ◽  
Ludmila Ferreira de Almeida Fiuza ◽  
...  
Keyword(s):  
Trypanosoma Cruzi ◽  
Chagas Disease ◽  
Phosphodiesterase Inhibitors ◽  
Biological Properties ◽  
Host Cells ◽  
Mammalian Host ◽  
Intracellular Camp ◽  
Additive Interaction ◽  
Content Type

ABSTRACT Pyrazolones are heterocyclic compounds with interesting biological properties. Some derivatives inhibit phosphodiesterases (PDEs) and thereby increase the cellular concentration of cyclic AMP (cAMP), which plays a vital role in the control of metabolism in eukaryotic cells, including the protozoan Trypanosoma cruzi, the etiological agent of Chagas disease (CD), a major neglected tropical disease. In vitro phenotypic screening identified a 4-bromophenyl-dihydropyrazole dimer as an anti-T. cruzi hit and 17 novel pyrazolone analogues with variations on the phenyl ring were investigated in a panel of phenotypic laboratory models. Potent activity against the intracellular forms (Tulahuen and Y strains) was obtained with 50% effective concentration (EC50) values within the 0.17 to 3.3 μM range. Although most were not active against bloodstream trypomastigotes, an altered morphology and loss of infectivity were observed. Pretreatment of the mammalian host cells with pyrazolones did not interfere with infection and proliferation, showing that the drug activity was not the result of changes to host cell metabolism. The pyrazolone NPD-227 increased the intracellular cAMP levels and was able to sterilize T. cruzi-infected cell cultures. Thus, due to its high potency and selectivity in vitro, and its additive interaction with benznidazole (Bz), NPD-227 was next assessed in the acute mouse model. Oral dosing for 5 days of NPD-227 at 10 mg/kg + Bz at 10 mg/kg not only reduced parasitemia (>87%) but also protected against mortality (>83% survival), hence demonstrating superiority to the monotherapy schemes. These data support these pyrazolone molecules as potential novel therapeutic alternatives for Chagas disease.

scholarly journals Anti-parasitic effect of novel amidines against Trypanosoma cruzi: phenotypic and in silico absorption, distribution, metabolism, excretion and toxicity analysis

2017 ◽  
Vol 3 ◽  
Author(s):  
ALINE SILVA DA GAMA NEFERTITI ◽  
MARCOS MEUSER BATISTA ◽  
PATRÍCIA BERNARDINO DA SILVA ◽  
EDUARDO CAIO TORRES-SANTOS ◽  
EDEZIO F. CUNHA-JÚNIOR ◽  
...  
Keyword(s):  
Trypanosoma Cruzi ◽  
In Silico ◽  
Host Cells ◽  
Mammalian Host ◽  
Aromatic Molecules ◽  
Toxicity Analysis ◽  
Parasitic Effect ◽  
The Many ◽  
Discrete Typing Unit

SUMMARYNew more selective and potent drugs are urgently need to treat Chagas disease (CD). Among the many synthetic compounds evaluated againstTrypanosoma cruzi, aromatic amidines (AAs) and especially arylimidamides (AIAs) have potent activity against this parasite. Presently, the effect of four mono-amidines (DB2228, DB2229, DB2292 and DB2294), four diamidines (DB2232, DB2235, DB2251 and DB2253) and one AIA (DB2255) was screenedin vitroagainst different forms (bloodstream trypomastigotes – BT and intracellular forms) and strains from discrete typing unit (DTU) I and VI ofT. cruziand their cytotoxic profile on mammalian host cells. Except for DB2253, all molecules were as active as benznidazole (Bz), resulting in 50% of reduction in the number of alive BT, with EC50ranging from 2·7 to 10·1µmafter 24 h of incubation. DB2255 was also the most potent against amastigotes (Tulahuen strain) showing similar activity to that of Bz (3µm).In silicoabsorption, distribution, metabolism, excretion and toxicity analysis demonstrated probability of human intestinal adsorption, while mutagenicity and inhibition of hERG1 were not predicted, besides giving acceptable predicted volumes of distribution. Our findings contribute for better knowledge regarding the biological effect of this class of aromatic molecules againstT. cruziaiming to identify novel promising agent for CD therapy.

scholarly journals Molecular, functional and structural properties of the prolyl oligopeptidase of Trypanosoma cruzi (POP Tc80), which is required for parasite entry into mammalian cells

2005 ◽  
Vol 388 (1) ◽  
pp. 29-38 ◽  
Author(s):  
Izabela M. D. BASTOS ◽  
Philippe GRELLIER ◽  
Natalia F. MARTINS ◽  
Gloria CADAVID-RESTREPO ◽  
Marian R. de SOUZA-AULT ◽  
...  
Keyword(s):  
Trypanosoma Cruzi ◽  
Mammalian Cells ◽  
Catalytic Domain ◽  
Single Copy ◽  
Catalytic Triad ◽  
Host Cells ◽  
Mammalian Host ◽  
Kinetic Properties ◽  
Prolyl Oligopeptidase ◽  
Single Chromosome

We have demonstrated that the 80 kDa POP Tc80 (prolyl oligopeptidase of Trypanosoma cruzi) is involved in the process of cell invasion, since specific inhibitors block parasite entry into non-phagocytic mammalian host cells. In contrast with other POPs, POP Tc80 is capable of hydrolysing large substrates, such as fibronectin and native collagen. In this study, we present the cloning of the POPTc80 gene, whose deduced amino acid sequence shares considerable identity with other members of the POP family, mainly within its C-terminal portion that forms the catalytic domain. Southern-blot analysis indicated that POPTc80 is present as a single copy in the genome of the parasite. These results are consistent with mapping of POPTc80 to a single chromosome. The active recombinant protein (rPOP Tc80) displayed kinetic properties comparable with those of the native enzyme. Novel inhibitors were assayed with rPOP Tc80, and the most efficient ones presented values of inhibition coefficient Ki≤1.52 nM. Infective parasites treated with these specific POP Tc80 inhibitors attached to the surface of mammalian host cells, but were incapable of infecting them. Structural modelling of POP Tc80, based on the crystallized porcine POP, suggested that POP Tc80 is composed of an α/β-hydrolase domain containing the catalytic triad Ser548–Asp631–His667 and a seven-bladed β-propeller non-catalytic domain. Docking analysis suggests that triple-helical collagen access to the catalytic site of POP Tc80 occurs in the vicinity of the interface between the two domains.

Sign in / Sign up

Export Citation Format

Share Document