Inhibition of host cell invasion and intracellular replication of Trypanosoma cruzi by N,N'-bis(benzyl)-substituted polyamine analogs

Abstract

We studied the effects of two N,N'-bis(benzyl)-substituted polyamine analogs on the capacities of Trypanosoma cruzi to invade and multiply within a mammalian host cell. At concentrations as low as 1 ,uM, these compounds reduced significantly the infectivity of the parasite for rat heart myoblasts in a time-dependent manner. Pretreatment of virulent T. cruzi trypomastigotes, but not myoblast pretreatment, reduced the level of infectivity. The inhibitory effects started to subside 3 h after removal of the drugs and were no longer detectable after 4 h. A significant decrease in the rate of intracellular amastigote multiplication was also seen when the drugs were added to myoblast cultures which had been previously infected with untreated T. cruzi.These results show that N,N'-bis(benzyl)-substituted polyamine analogs meet the two most important criteria for potential chemotherapeutic agents against T. cruzi infection, namely, inhibition of both host cell invasion and intracellular replication by this parasite.The unicellular parasite Trypanosoma cnrzi, the causative agent of Chagas' disease, requires an intracellular localization to multiply in mammalian hosts. This stresses the importance of developing drugs that can inhibit host cell invasion and subsequent intracellular parasite replication. Such drugs are badly needed given the lack of a consistently effective, nontoxic drug to cure Chagas' disease (9). There is evidence that both host cell invasion by T. cruzi trypomastigotes and intracellular multiplication of the amastigote form are impaired when production of the polyamines agmatine and putrescine is decreased by the specific arginine decarboxylase inhibitor DL-a-dlfluoromethylarginine (7,13).Polyamines play key roles in cell growth regulation, and polyamine synthesis inhibitors have been shown to affect some parasitic organisms (1,8,16). N,N'-Di-substituted tetraamines with the general formula RNH(CH2)XNH (CH2)YNH(CH2)NINHR have been reported to suppress the growth of several pathogenic protozoans (e.g., Leishmania donovani, Plasmodium falciparum, and Plasmodium berghei) in vitro and in infected animals (2-4) and could be suitable for exploring the role that polyamines and polyamine biosynthesis plays in T. cnuzi infection. In this work, we used two N,N'-bis(benzyl)-substituted polyamine analogs and studied their effects on T. cnuzi infectivity and replication using an in vitro model system of mammalian host cell infection.Stock CD1(ICR) Swiss mice (Charles River Laboratory, Portage, Mich.) were infected subcutaneously with 106 T. cruzi (Tulahen isolate) organisms. Trypomastigotes were purified 10 to 12 days later from the blood by centrifugation through Isolymph (Gallard-Schlesinger, Carle Place, N.Y.) (5) followed by column chromatography through DEAEcellulose (15). The eluted trypanosomes were washed twice with Dulbecco's modified minimal essential medium (GIBCO, Grand Island, N.Y.) containing 100 IU of penicillin and 100 tg of streptomycin per ml and supplemented with 10% fetal bovine serum (Sigma Chemi...

CD4+ T Cell-Dependent Macrophage Activation Modulates Sustained PS Exposure on Intracellular Amastigotes of Leishmania amazonensis

“…However, when we quantified parasite loads on infected-MΦs activated with 25% of iSN, we observed that DFMO treatment was detrimental for the proliferation of intracellular parasites at 72 h post-infection (Figure 4F). This finding may indicate that polyamine synthesis is a requirement for parasite proliferation and maintenance in these conditions, as described in other models (Majumder and Kierszenbaum, 1993; Vannier-Santos et al, 2008). Nevertheless, our data imply that PS exposure on intracellular amastigotes is modulated by cytokine-mediated, iNOS-dependent MΦ activation, rather than by arginase I expression.…”

A minimalistic approach to develop new anti-apicomplexa polyamines analogs

“…Future studies with the best candidates will focus on elucidating mode of action. The fact that similar compounds act by different mechanisms in trypanosomatids [22,47] and the pathogenic fungus Pneumocystis carinii [48] support this hypothesis. Polyamine oxidase have been studied as a possible target of MDL-27695 on P. falciparum [49].…”

Effect of a Bis-Benzyl Polyamine Analogue on Pneumocystis carinii

“…An in vitro MIC 50 of 3.0 M was reported for the human malaria parasite (18). Exposure of mammal-infective T. cruzi trypomastigotes to 1 M BBS prior to adding to mammalian cells caused a 61% reduction in the number of infected mammalian cells and a 69% reduction in the mean number of parasites per infected cell compared to untreated control trypomastigotes (11). When T. cruzi was already growing in mammalian cells, a 24-h exposure to 1 M BBS caused a 41% reduction in parasite multiplication, compared to untreated controls.…”

“…The results reported here show that a polyamine analogue, BBS, is also active against PCP and provide information about the interaction of this analogue and P. carinii. The choice of BBS for the study was based on previous reports of activity against Plasmodium, the malaria parasite, about 10 years ago (7) and subsequent reports of activity against the protozoans Leishmania donovani (1) and Trypanosoma cruzi (11), as well as the filarial parasite Brugia pahangi (19). A summary of previous results with BBS and other parasites is presented below to provide perspective for the results presented here.…”