Use of a selective inhibitor to define the chemotherapeutic potential of the plasmodial hexose transporter in different stages of the parasite's life cycle

During blood infection, malarial parasites use d-glucose as their main energy source. The Plasmodium falciparum hexose transporter (PfHT), which mediates the uptake of d-glucose into parasites, is essential for survival of asexual blood-stage parasites. Recently, genetic studies in the rodent malaria model, Plasmodium berghei, found that the orthologous hexose transporter (PbHT) is expressed throughout the parasite''s development within the mosquito vector, in addition to being essential during intraerythrocytic development. Here, using a d-glucose-derived specific inhibitor of plasmodial hexose transporters, compound 3361, we have investigated the importance of d-glucose uptake during liver and transmission stages of P. berghei. Initially, we confirmed the expression of PbHT during liver stage development, using a green fluorescent protein (GFP) tagging strategy. Compound 3361 inhibited liver-stage parasite development, with a 50% inhibitory concentration (IC₅₀) of 11 μM. This process was insensitive to the external d-glucose concentration. In addition, compound 3361 inhibited ookinete development and microgametogenesis, with IC₅₀s in the region of 250 μM (the latter in a d-glucose-sensitive manner). Consistent with our findings for the effect of compound 3361 on vector parasite stages, 1 mM compound 3361 demonstrated transmission blocking activity. These data indicate that novel chemotherapeutic interventions that target PfHT may be active against liver and, to a lesser extent, transmission stages, in addition to blood stages.     

Activity of a Trisubstituted Pyrrole in Inhibiting Sporozoite Invasion and Blocking Malaria Infection

Malaria infection is initiated by Plasmodium sporozoites infecting the liver. Preventing sporozoite infection would block the obligatory first step of the infection and perhaps reduce disease severity. In addition, such an approach would decrease Plasmodium vivax hypnozoite formation and therefore disease relapses. Here we describe the activity of a trisubstituted pyrrole, 4-[2-(4-fluorophenyl)-5-(1-methylpiperidine-4-yl)-1H-pyrrol-3-yl] pyridine, in inhibiting motility, invasion, and consequently infection by P. berghei sporozoites. In tissue culture, the compound was effective within the first 3 h of sporozoite addition to HepG2 cells. In vivo, intraperitoneal administration of the compound significantly inhibited liver-stage parasitemia in P. yoelii sporozoite-infected mice and prevented the appearance of blood-stage parasites. P. berghei sporozoites lacking the parasite cGMP-dependent protein kinase, the primary target of the compound in erythrocyte-stage parasites, remained infectious to HepG2 cells and sensitive to the drug. These results suggest that the drug has an additional target(s) in sporozoites. We propose that drugs that inhibit sporozoite infection offer a feasible approach to malaria prophylaxis.The malaria parasite, Plasmodium spp., infects 500 million people a year, killing over 1 million globally. Plasmodium parasites are introduced into humans via mosquito bites, in the form of sporozoites that infect the liver to form liver-stage parasites (LS). LS are eventually released into the bloodstream, where they infect erythrocytes and initiate the symptomatic phase of malaria. Although the liver stage of infection by Plasmodium falciparum and P. vivax, the two major human parasites, is asymptomatic, P. vivax infections often result in the formation of dormant LS called “hypnozoites.” When reactivated, hypnozoites cause disease relapses up to 1 year after the initial infection. Therefore, achieving global eradication of malaria will require prevention or treatment of P. vivax hypnozoites in addition to treatment of erythrocyte stages of both species.The spread of drug-resistant parasites, limited vector control measures, and lack of an effective vaccine make novel drug discovery vital. Drugs that prevent sporozoite infection of the liver would block an obligatory step in the parasite''s life cycle. They would also prevent the formation of hypnozoites by P. vivax and the pathology caused by the erythrocyte stages of all human Plasmodium species. Although the liver stages are asymptomatic, reducing the parasite burden in the liver is likely to significantly diminish blood-stage infection and reduce disease severity in a subpopulation of patients (1). Furthermore, the small number of parasites and the limited number of replications during the liver phase of malaria infection compared to those in the erythrocytic phase reduces the potential for the emergence of drug-resistant parasites (13). However, of the currently licensed drugs, only primaquine has been formally demonstrated to act against sporozoites (13, 22).Here, we investigate the efficacy of a trisubstituted pyrrole, 4-[2-(4-fluorophenyl)-5-(1-methylpiperidine-4-yl)-1H-pyrrol-3-yl] pyridine, against Plasmodium sporozoites. This compound was previously shown to inhibit the in vitro development of several apicomplexan parasites (11). In vitro, the trisubstituted pyrrole blocks the development of late-erythrocyte-stage schizonts of P. falciparum (23). In vivo, it is partially effective against erythrocyte stages, lengthening the survival of mice infected with lethal strains of P. berghei (8).Here, we tested the effect of the trisubstituted pyrrole on sporozoites from the rodent parasite species P. berghei and P. yoelii. The compound significantly inhibited the infection of hepatocytes by Plasmodium sporozoites. In vitro, this inhibition was dose dependent and was observed within the first 3 h of sporozoite addition to HepG2 cells. Neither the treatment of HepG2 cells with the trisubstituted pyrrole prior to the addition of sporozoites nor the addition of the trisubstituted pyrrole to cell culture after infection by sporozoites decreased infection. While sporozoite motility was partially inhibited by the trisubstituted pyrrole, sporozoite invasion of HepG2 cells could be completely blocked in a dose-dependent manner. In vivo, administration of a single dose of the trisubstituted pyrrole caused a significant reduction in the liver-stage parasitemia of mice infected with P. yoelii sporozoites and led to partial protection against the appearance of blood-stage parasites. A higher dose of the trisubstituted pyrrole completely prevented the appearance of blood-stage parasites in a sporozoite-initiated infection. The compound''s inhibition of sporozoite infection was not mediated solely through the drug''s effect on P. berghei cGMP-dependent protein kinase (PbPKG). Sporozoites lacking PbPKG remained sensitive to the trisubstituted pyrrole. Our data suggest that the trisubstituted pyrrole might have additional targets in sporozoites.     

Antiprotozoal Activity of 1-Phenethyl-4-Aminopiperidine Derivatives

A series of 44 4-aminopiperidine derivatives was screened in vitro against four protozoan parasites (Trypanosoma brucei rhodesiense, Trypanosoma cruzi, Leishmania donovani, and Plasmodium falciparum). This screening identified 29 molecules selectively active against bloodstream-form T. b. rhodesiense trypomastigotes, with 50% inhibitory concentrations (IC₅₀) ranging from 0.12 to 10 μM, and 33 compounds active against the chloroquine- and pyrimethamine-resistant K1 strain of P. falciparum (IC₅₀ range, 0.17 to 5 μM). In addition, seven compounds displayed activity against intracellular T. cruzi amastigotes in the same range as the reference drug benznidazole (IC₅₀, 1.97 μM) but were also cytotoxic to L-6 cells, showing little selectivity for T. cruzi. None of the molecules tested showed interesting antileishmanial activity against axenic amastigotes of L. donovani. To our knowledge, this is the first report of the antitrypanosomal activity of molecules bearing the 4-aminopiperidine skeleton.Tropical diseases due to parasitic protozoa cause great mortality and disability in the less developed world. Malaria and kinetoplastid diseases such as human African trypanosomiasis (HAT, or sleeping sickness), Chagas'' disease, and leishmaniases are among the main neglected parasitic diseases, affecting hundreds of millions of people worldwide (http://www.dndi.org/index.php/diseases.html?ids=2). However, the low income of the affected populations does not make it financially attractive for pharmaceutical companies to invest in the development of new drugs against these diseases. The number of antiprotozoal drugs available is limited, and they are old, toxic, and losing efficacy due to the emergence of resistant parasites. New drugs are thus urgently needed (37).The landscape of drug discovery and development for new antiparasitic drugs has changed in the past few years thanks to financial backing from not-for-profit organizations and the involvement of public-private partnerships. The collaboration of various pharmaceutical companies with the Special Programme for Research and Training in Tropical Diseases (TDR), giving access to their compound libraries to help scientists search for new antiparasitic drugs, is a good example of the pragmatic approach required for the development of new medicaments for those neglected diseases (15, 32, 40). The screening of compound libraries to discover new hit and lead compounds against protozoan parasites has been used very efficiently by our group for the past few years. Very potent antiprotozoal compounds, active in vitro and in vivo against Trypanosoma brucei rhodesiense and Plasmodium falciparum, were identified by our group (16, 17, 38). These lead compounds were discovered by screening against T. brucei and P. falciparum a small library (<100 molecules) of dicationic compounds structurally related to known antiparasitic drugs (e.g., pentamidine, diminazene) but primarily synthesized by us for different medicinal targets.Following this successful approach, we have now selected from our in-house compound collection another series of polyamine molecules that possess the 1-phenethyl-4-aminopiperidine skeleton (Fig. ​(Fig.1).1). These molecules were anticipated to display antiprotozoal activity because they are structurally related to the known antitrypanosomal alkylpolyamines MDL27695 (7, 22), CHE-3-7-3 (5), and BW-1 (43) and particularly to the 4-aminopiperidine-based antimalarial agents described recently by Ellman and coworkers (8, 9). To test this hypothesis, 44 1-phenethyl-4-aminopiperidine derivatives were tested in vitro against four protozoan parasites responsible for HAT (T. brucei rhodesiense), Chagas'' disease (Trypanosoma cruzi), visceral leishmaniasis (Leishmania donovani), and malaria (P. falciparum). Two compounds found to be active in vitro (50% inhibitory concentrations [IC₅₀], <0.2 μM) and displaying acceptable selectivity toward the parasites (selectivity index [SI], >100) were subjected to in vivo assays in mouse models of acute HAT (T. b. brucei) or malaria (Plasmodium berghei). SI is defined as (50% cytotoxic concentration [CC₅₀] for L-6 cells)/(IC₅₀ for the parasite).Open in a separate windowFIG. 1.General structures of 1-phenethyl-4-aminopiperidine derivatives screened in this study and of known alkylpolyamine analogues with antiprotozoal activity.     

A Chemical Rescue Screen Identifies a Plasmodium falciparum Apicoplast Inhibitor Targeting MEP Isoprenoid Precursor Biosynthesis

The apicoplast is an essential plastid organelle found in Plasmodium parasites which contains several clinically validated antimalarial-drug targets. A chemical rescue screen identified MMV-08138 from the “Malaria Box” library of growth-inhibitory antimalarial compounds as having specific activity against the apicoplast. MMV-08138 inhibition of blood-stage Plasmodium falciparum growth is stereospecific and potent, with the most active diastereomer demonstrating a 50% effective concentration (EC₅₀) of 110 nM. Whole-genome sequencing of 3 drug-resistant parasite populations from two independent selections revealed E688Q and L244I mutations in P. falciparum IspD, an enzyme in the MEP (methyl-d-erythritol-4-phosphate) isoprenoid precursor biosynthesis pathway in the apicoplast. The active diastereomer of MMV-08138 directly inhibited PfIspD activity in vitro with a 50% inhibitory concentration (IC₅₀) of 7.0 nM. MMV-08138 is the first PfIspD inhibitor to be identified and, together with heterologously expressed PfIspD, provides the foundation for further development of this promising antimalarial drug candidate lead. Furthermore, this report validates the use of the apicoplast chemical rescue screen coupled with target elucidation as a discovery tool to identify specific apicoplast-targeting compounds with new mechanisms of action.     

Antimalarial activity of phenylthiazolyl-bearing hydroxamate-based histone deacetylase inhibitors

The antimalarial activity and pharmacology of a series of phenylthiazolyl-bearing hydroxamate-based histone deacetylase inhibitors (HDACIs) was evaluated. In in vitro growth inhibition assays approximately 50 analogs were evaluated against four drug resistant strains of Plasmodium falciparum. The range of 50% inhibitory concentrations (IC₅₀s) was 0.0005 to >1 μM. Five analogs exhibited IC₅₀s of <3 nM, and three of these exhibited selectivity indices of >600. The most potent compound, WR301801 (YC-2-88) was shown to cause hyperacetylation of P. falciparum histones, which is a marker for HDAC inhibition in eukaryotic cells. The compound also inhibited malarial and mammalian HDAC activity in functional assays at low nanomolar concentrations. WR301801 did not exhibit cures in P. berghei-infected mice at oral doses as high as 640 mg/kg/day for 3 days or in P. falciparum-infected Aotus lemurinus lemurinus monkeys at oral doses of 32 mg/kg/day for 3 days, despite high relative bioavailability. The failure of monotherapy in mice may be due to a short half-life, since the compound was rapidly hydrolyzed to an inactive acid metabolite by loss of its hydroxamate group in vitro (half-life of 11 min in mouse microsomes) and in vivo (half-life in mice of 3.5 h after a single oral dose of 50 mg/kg). However, WR301801 exhibited cures in P. berghei-infected mice when combined at doses of 52 mg/kg/day orally with subcurative doses of chloroquine. Next-generation HDACIs with greater metabolic stability than WR301801 may be useful as antimalarials if combined appropriately with conventional antimalarial drugs.     

Repurposing the Open Access Malaria Box To Discover Potent Inhibitors of Toxoplasma gondii and Entamoeba histolytica

Toxoplasmosis and amebiasis are important public health concerns worldwide. The drugs currently available to control these diseases have proven limitations. Therefore, innovative approaches should be adopted to identify and develop new leads from novel scaffolds exhibiting novel modes of action. In this paper, we describe results from the screening of compounds in the Medicines for Malaria Venture (MMV) open access Malaria Box in a search for new anti-Toxoplasma and anti-Entamoeba agents. Standard in vitro phenotypic screening procedures were adopted to assess their biological activities. Seven anti-Toxoplasma compounds with a 50% inhibitory concentration (IC₅₀) of <5 μM and selectivity indexes (SI) of >6 were identified. The most interesting compound was MMV007791, a piperazine acetamide, which has an IC₅₀ of 0.19 μM and a selectivity index of >157. Also, we identified two compounds, MMV666600 and MMV006861, with modest activities against Entamoeba histolytica, with IC₅₀s of 10.66 μM and 15.58 μM, respectively. The anti-Toxoplasma compounds identified in this study belong to scaffold types different from those of currently used drugs, underscoring their novelty and potential as starting points for the development of new antitoxoplasmosis drugs with novel modes of action.     

Antimalarial Efficacy of Hydroxyethylapoquinine (SN-119) and Its Derivatives

Quinine and other cinchona-derived alkaloids, although recently supplanted by the artemisinins (ARTs), continue to be important for treatment of severe malaria. Quinine and quinidine have narrow therapeutic indices, and a safer quinine analog is desirable, particularly with the continued threat of antimalarial drug resistance. Hydroxyethylapoquinine (HEAQ), used at 8 g a day for dosing in humans in the 1930s and halving mortality from bacterial pneumonias, was shown to cure bird malaria in the 1940s and was also reported as treatment for human malaria cases. Here we describe synthesis of HEAQ and its novel stereoisomer hydroxyethylapoquinidine (HEAQD) along with two intermediates, hydroxyethylquinine (HEQ) and hydroxyethylquinidine (HEQD), and demonstrate comparable but elevated antimalarial 50% inhibitory concentrations (IC₅₀) of 100 to 200 nM against Plasmodium falciparum quinine-sensitive strain 3D7 (IC₅₀, 56 nM). Only HEAQD demonstrated activity against quinine-tolerant P. falciparum strains Dd2 and INDO with IC₅₀s of 300 to 700 nM. HEQD had activity only against Dd2 with an IC₅₀ of 313 nM. In the lethal mouse malaria model Plasmodium berghei ANKA, only HEQD had activity at 20 mg/kg of body weight comparable to that of the parent quinine or quinidine drugs measured by parasite inhibition and 30-day survival. In addition, HEQ, HEQD, and HEAQ (IC₅₀ ≥ 90 μM) have little to no human ether-à-go-go-related gene (hERG) channel inhibition expressed in CHO cells compared to HEAQD, quinine, and quinidine (hERG IC₅₀s of 27, 42, and 4 μM, respectively). HEQD more closely resembled quinine in vitro and in vivo for Plasmodium inhibition and demonstrated little hERG channel inhibition, suggesting that further optimization and preclinical studies are warranted for this molecule.     

Seeking potent anti-tubercular agents: design and synthesis of substituted-N-(6-(4-(pyrazine-2-carbonyl)piperazine/homopiperazine-1-yl)pyridin-3-yl)benzamide derivatives as anti-tubercular agents

Pyrazinamide is an important first-line drug used in shortening TB therapy. In our current work, a series of novel substituted-N-(6-(4-(pyrazine-2-carbonyl)piperazine/homopiperazine-1-yl)pyridin-3-yl)benzamide derivatives were designed, synthesized, and evaluated for their anti-tubercular activity against Mycobacterium tuberculosis H37Ra. Among the tested compounds, five compounds (6a, 6e, 6h, 6j and 6k) from Series-I and one compound (7e) from Series-II exhibited significant activity against Mycobacterium tuberculosis H37Ra with 50% inhibitory concentrations (IC₅₀) ranging from 1.35 to 2.18 μM. To evaluate the efficacy of these compounds, we examined their IC₉₀ values. Five of the most active compounds were found to be more active with IC₉₀s ranging from 3.73 to 4.00 μM and one compound (6e) showed an IC₉₀ of 40.32 μM. Moreover, single crystals were developed for 6d, 6f and 6n. In addition, most active compounds were evaluated for their cytotoxicity on HEK-293 (human embryonic kidney) cells. Our results indicate that the compounds are nontoxic to human cells. The molecular interactions of the derivatised conjugates in docking studies reveal their suitability for further development.

We herein report 27 pyrazinamide analogues as anti-tubercular agents, of which six exhibited excellent activity with IC₅₀ ≤ 2.18 μM and these were less toxic against HEK 293 cells.     

Contrasting Ex Vivo Efficacies of “Reversed Chloroquine” Compounds in Chloroquine-Resistant Plasmodium falciparum and P. vivax Isolates

Chloroquine (CQ) has been the mainstay of malaria treatment for more than 60 years. However, the emergence and spread of CQ resistance now restrict its use to only a few areas where malaria is endemic. The aim of the present study was to investigate whether a novel combination of a CQ-like moiety and an imipramine-like pharmacophore can reverse CQ resistance ex vivo. Between March to October 2011 and January to September 2013, two “reversed chloroquine” (RCQ) compounds (PL69 and PL106) were tested against multidrug-resistant field isolates of Plasmodium falciparum (n = 41) and Plasmodium vivax (n = 45) in Papua, Indonesia, using a modified ex vivo schizont maturation assay. The RCQ compounds showed high efficacy against both CQ-resistant P. falciparum and P. vivax field isolates. For P. falciparum, the median 50% inhibitory concentrations (IC₅₀s) were 23.2 nM for PL69 and 26.6 nM for PL106, compared to 79.4 nM for unmodified CQ (P < 0.001 and P = 0.036, respectively). The corresponding values for P. vivax were 19.0, 60.0, and 60.9 nM (P < 0.001 and P = 0.018, respectively). There was a significant correlation between IC₅₀s of CQ and PL69 (Spearman''s rank correlation coefficient [r_s] = 0.727, P < 0.001) and PL106 (r_s = 0.830, P < 0.001) in P. vivax but not in P. falciparum. Both RCQs were equally active against the ring and trophozoite stages of P. falciparum, but in P. vivax, PL69 and PL106 showed less potent activity against trophozoite stages (median IC₅₀s, 130.2 and 172.5 nM) compared to ring stages (median IC₅₀s, 17.6 and 91.3 nM). RCQ compounds have enhanced ex vivo activity against CQ-resistant clinical isolates of P. falciparum and P. vivax, suggesting the potential use of reversal agents in antimalarial drug development. Interspecies differences in RCQ compound activity may indicate differences in CQ pharmacokinetics between the two Plasmodium species.     

New protein tyrosine phosphatase inhibitors from fungus Aspergillus gorakhpurensis F07ZB1707

Twelve new compounds, aspergorakhins A–L (1–12) coupled with one known xanthone leptosphaerin D (13), were isolated from the extract of soil-derived fungus Aspergillus gorakhpurensis F07ZB1707. Their structures were elucidated by spectroscopic data analysis including UV, IR, NMR, and HRESIMS. The absolute configurations of 5 and 8–11 were identified using ECD and OR calculations. All compounds were tested by enzyme inhibitory activity assay in vitro. Aspergorakhin A (1) showed selective activities against PTP1B and SHP1 over TCPTP with IC₅₀ values 0.57, 1.19, and 22.97 μM, respectively. Compounds 1 and 2 exhibited modest cytotoxicity against tumor cell lines A549, HeLa, Bel-7402, and SMMC-7721 with IC₅₀ values in the range of 6.75–83.4 μM.

Twelve novel metabolites were isolated from Aspergillus gorakhpurensis F07ZB1707. Aspergorakhin A (1) showed selective activities against PTP1B and SHP1 over TCPTP with IC₅₀ values of 0.57, 1.19, and 22.97 μM, respectively.     

Cembrane-type diterpenoids from the gum resin of Boswellia carterii and their biological activities

Eight new cembrane-type diterpenoids, boscartins AH–AK (1–8), along with two known ones (9-10), were isolated from the gum resin of Boswellia carterii. Compounds 1–3 were characteristic of high oxidation assignable to three epoxy groups, while compounds 4–8 were characteristic of two epoxy groups. Spectroscopic examination was used to elucidate their structures. All isolates were evaluated for antiproliferative activity against HCT-116 human colon cancer cells, anti-inflammatory activity against nitric oxide (NO) production, and hepatoprotective activity in vitro. All of them showed weak antiproliferative activity (IC₅₀ > 100 μM), 8 exhibited potent inhibitory effects on NO production (IC₅₀ of 14.8 μM), with the others showing weak anti-inflammatory activity (IC₅₀ > 30 μM), and 1 exhibited more potent hepatoprotective activity than the positive control, bicyclol, at 10 μM against the damage induced by paracetamol in HepG2 cells.

Cembrane-type diterpenoids from the gum resin of Boswellia carterii.     

Photo-induced synthesis,structure and in vitro bioactivity of a natural cyclic peptide Yunnanin A analog

A cyclic analog of natural peptide Yunnanin A was synthesized via photoinduced single electron transfer reaction (SET) in the paper. The resulting compound exhibited potent bioactivity (with IC₅₀ values 29.25 μg mL⁻¹ against HepG-2 cell lines and 65.01 μg mL⁻¹ against HeLa cell lines), but almost have no toxicity to normal cells (with IC₅₀ values 203.25 μg mL⁻¹ against L929 cell lines), which may be served as a potential antitumor drug for medical treatment. The spatial structure was examined by experimental electronic circular dichroism (ECD) and quantum chemistry calculations. Moreover, the theoretical study suggested that special intramolecular hydrogen bonds and γ, β-turn secondary structures may be possible sources affecting cyclic peptide''s bioactivity.

The photo-induced synthesis, structure and in vitro bioactivity study of a Yunnanin A cyclopeptide analog was presented.     

Experimental Chemotherapy against Trypanosoma cruzi Infection Using Ruthenium Nitric Oxide Donors

The ruthenium NO donors of the group trans-[Ru(NO)(NH₃)₄L]ⁿ⁺, where the ligand (L) is N-heterocyclic H₂O, SO₃²⁻, or triethyl phosphite, are able to lyse Trypanosoma cruzi in vitro and in vivo. Using half-maximal (50%) inhibitory concentrations against bloodstream trypomastigotes (IC₅₀^try) and cytotoxicity data on mammalian V-79 cells (IC₅₀^V79), the in vitro therapeutic indices (TIs) (IC₅₀^V79/IC₅₀^try) for these compounds were calculated. Compounds that exhibited an in vitro TI of ≥10 and trypanocidal activity against both epimastigotes and trypomastigotes with an IC₅₀^try/epi of ≤100 μM were assayed in a mouse model for acute Chagas'' disease, using two different routes (intraperitoneal and oral) for drug administration. A dose-effect relationship was observed, and from that, the ideal dose of 400 nmol/kg of body weight for both trans-[Ru(NO)(NH₃)₄isn](BF₄)₃ (isn, isonicotinamide) and trans-[Ru(NO)(NH₃)₄imN](BF₄)₃ (imN, imidazole) and median (50%) effective doses (ED₅₀) of 86 and 190 nmol/kg, respectively, were then calculated. Since the 50% lethal doses (LD₅₀) for both compounds are higher than 125 μmol/kg, the in vivo TIs (LD₅₀/ED₅₀) of the compounds are 1,453 for trans-[Ru(NO)(NH₃)₄isn](BF₄)₃ and 658 for trans-[Ru(NO)(NH₃)₄imN](BF₄)₃. Although these compounds exhibit a marked trypanocidal activity and are able to react with cysteine, they exhibit very low activity in T. cruzi-glycosomal glyceraldehyde-3-phosphate dehydrogenase tests, suggesting that this enzyme is not their target. The trans-[Ru(NO)(NH₃)₄isn](BF₄)₃ and trans-[Ru(NO)(NH₃)₄imN](BF₄)₃ compounds are able to eliminate amastigote nests in myocardium tissue at 400-nmol/kg doses and ensure the survival of all infected mice, thus opening a novel set of therapies to try against trypanosomatids.Although American trypanosomiasis, or Chagas'' disease, has existed on the American continent for more than 9,000 years (3), it still is considered incurable, in view of the fact that the available anti-Trypanosoma cruzi drugs exhibit limited efficacy and undesirable side effects (11). Chagas'' disease takes third place, after malaria and leishmaniasis, in mortality and morbidity prevalence due to vector-associated diseases on the American continent (17, 26). Since the parasite can be found in the blood of up to 50% of infected people several years after the primary infection (36), Chagas'' disease is also a cause for government concern in several countries where the disease is not endemic (31). This infection can be contracted by infected-blood transfusion and organ transplant from donors originating from areas of Latin America where the disease is endemic (31).The parasite''s biological cycle includes two multiplicative forms (epimastigote and amastigote), one infective form (trypomastigote), and an obligatory passage through vertebrate and invertebrate hosts (14). The trypomastigotes escape to the cytosol of the macrophage and transform into amastigotes, which are released as trypomastigote forms (26, 30). Under these circumstances, the infection triggers interleukin-12 (2) and tumor necrosis factor alpha production (10), which leads to gamma interferon (IFN-γ) synthesis (2) and to inducible nitric oxide (NO) synthase activation (22). As a consequence, NO is synthesized, and this NO production has been thought to be responsible for the trypanocidal effect (53). At this stage of the infection, the parasite also triggers production of transforming growth factor beta and interleukin-10, which are negative regulators of NO production inhibiting IFN-γ-produced macrophage activation and NO synthesis (22). Transforming growth factor beta production, which is described as an anti-inflammatory response, often promotes permissiveness to T. cruzi infection (53). Furthermore, it has been observed that the NO produced during acute infection also plays an important role in at least two of the processes that facilitate parasite evasion from the cellular immune response (26). In addition to mediating resistance against infection, NO can suppress the immune response to T. cruzi via the induction of apoptosis of T cells (25). Thus, this experimental evidence gives support to the idea that the intracellular NO production by host cells plays an essential role in resistance to the parasite infection (51). Indeed, the pharmacological modulation of the host immune response against T. cruzi through control of the NO levels has been recently accepted as a therapeutic target (26).In this context, the ruthenium species of the group trans-[Ru^II(NO⁺)(NH₃)₄L]ⁿ⁺, where the ligand (L) is N heterocyclic, H₂O, sulfite (SO₃²⁻), or triethyl phosphite [P(OEt)₃], which can deliver NO at a selected rate constant (k_−NO) upon reduction of the nitrosyl ligand, can be useful in chemotherapy against T. cruzi and other pathogens. The NO release specific rate constant and the redox potential for the [Ru^IINO⁺]/[Ru^IINO⁰] couple in these complexes can be controlled through the judicious selection of the trans ligand (44). Additionally, these compounds exhibit low cytotoxicity and good water solubility (up to 10⁻² M) and are stable for days under physiological hydrogen ion conditions (43). Certainly there are many other compounds that could be tested for such purposes (4, 47). However, the main advantages of our compounds are that they are kinetically robust and are activated only by reductors (43). Furthermore, they offer the possibility of tuning the redox potential of the nitrosonium ligand and the lability of the formed NO as a function of the trans ligand in a very efficient manner (41, 43). As far as we know, the other compounds in question do not offer the same flexibility, at least to the same extent. In addition, we are now exploring the possibility that our compounds operate on the catalytic cycle converting NO²⁻ into NO⁺ (28).As a natural extension of a previous work in which we described the antiproliferative activity of ruthenium NO donors against epimastigote forms (38), this work evaluates the trypanocidal activity of multiplicative forms of T. cruzi. In addition, based on in vitro and in vivo therapeutic indices (TIs), this work also demonstrates that trans-[Ru^II(NO⁺)(NH₃)₄imN](BF₄)₃, trans-[Ru(NO)(NH₃)₄py](BF₄)₃, and trans-[Ru^II(NO⁺)(NH₃)₄isn](BF₄) (imN, imidazole; py, pyridine; isn, isonicotinamide) can be successfully used to treat acute murine Chagas'' disease and that at nanomolar doses they are able to lyse in vivo the parasite and to ensure the survival of all infected mice.This work is presently under Brazilian patent of invention deposit request PI 0705849-7 (D. W. Franco et al.).(Part of this work was conducted by J. J. N. Silva in partial fulfillment of the requirements for a Ph.D. from the Instituto de Química de São Carlos, Universidade de São Paulo [USP], Brazil, 2008.)     

Synthesis of 2-ethoxycarbonylthieno[2,3-b]quinolines in biomass-derived solvent γ-valerolactone and their biological evaluation against protein tyrosine phosphatase 1B

A series of 2-ethoxycarbonylthieno[2,3-b]quinolines were synthesized in the bio-derived “green” solvent γ-valerolactone (GVL) and evaluated for their inhibitory activities against PTP1B, the representative compound 6a displayed an IC₅₀ value of 8.04 ± 0.71 μM with 4.34-fold preference over TCPTP. These results provided novel lead compounds for the design of inhibitors of PTP1B as well as other PTPs.

A series of 2-ethoxycarbonylthieno[2,3-b]quinolines were synthesized in the bio-derived “green” solvent γ-valerolactone and evaluated for their inhibitory activities against PTP1B, compound 6a displayed an IC₅₀ value of 8.04 ± 0.71 μM with 4.34-fold preference over TCPTP.     

Design,synthesis, and bioactivity evaluation of antitumor sorafenib analogues

Malignant tumors are a serious threat to human health and are generally treated with chemical therapy. This chemical therapy uses agents that act on signal transduction pathway mechanism of tumor with good selectivity and low toxicity. Sorafenib is a multikinase target inhibitor with good tumor inhibitory activity and a protein kinase inhibitor. In this research, a novel series of sorafenib analogues and derivatives were designed, synthesized, and evaluated as tumor inhibitors. These compounds used sorafenib as the lead compound and achieved modifications using bioisosteres and the alkyl principle. The in vitro the results showed that compounds 3c, 3d, 3h, 3n, 3r, and 3z had good inhibitory effects on human cervical cancer cells (Hela), while compounds 3t and 3v had good inhibitory effects on human lung cancer cells (H1975 and A549). Among these, compound 3d had an inhibitory activity (IC₅₀) of 0.56 ± 0.04 μmol L⁻¹ against Hela cells (human cervical cancer), the compound 3t had an IC₅₀ of 2.34 ± 0.07 μmol L⁻¹ against H1975 cells (human lung cancer), and compound 3v had an IC₅₀ of 1.35 ± 0.03 μmol L⁻¹ against A549 cells (human lung cancer). The in vivo results showed that these compounds had good antitumor effects and low acute toxicity.

Malignant tumors are a serious threat to human health and are generally treated with chemical therapy.     

Saccharomonosporine A inspiration; synthesis of potent analogues as potential PIM kinase inhibitors

Saccharomonosporine A was recently reported as a natural anti-cancer agent working through inhibition of a Proviral integration site for Moloney murine leukemia virus-1 (PIM-1) kinase. Structural bioisosteres of this natural product were synthesized and tested against PIM kinase enzymes. They showed potent inhibitory activity against all the known PIM kinases (PIM-1, 2 and 3) with IC₅₀ values ranging from 0.22 to 2.46 μM. Compound 5 was the most potent pan-inhibitor with IC₅₀ values of 0.37, 0.41, and 0.3 μM, against PIM-1, 2, 3 respectively. Compounds 4–6 were tested for their cytotoxic activities against 3 cell lines: H1650, HT-29, and HL-60. Compound 5 exhibited significant cytotoxic activity against human colon adenocarcinoma HT-29 and the human promyelocytic leukemia HL-60, with IC₅₀ μM values of 1.4 and 1.7 respectively. Molecular docking and homology modeling studies were carried out to confirm the affinity of these synthesized compounds to the three different PIM kinases. Additionally, a number of in silico predictions, ADME/Tox, were adopted to evaluate their drug-likeness.

The E isomer of compound 5 exhibited a potent inhibitory effect against PIM kinase isoforms of IC₅₀s 0.30–0.41 μM.     

Antiproliferative activity of new pentacyclic triterpene and a saponin from Gladiolus segetum Ker-Gawl corms supported by molecular docking study

A new triterpenoidal saponin identified as 3-O-[β-d-glucopyranosyl-(1 → 2)-β-d-glucopyranosyl-(1 → 4)-β-d-xylopyranosyl]-2β,3β,16α-trihydroxyolean-12-en-23,28-dioic acid-28-O-α-l-rhamnopyranosyl-(1 → 4)-α-l-rhamnopyranosyl-(1 → 2)-β-d-glucopyranosyl-(1 → 2)-α-l-arabinopyranoside 1 together with a new oleanane triterpene identified as 2β,3β,13α,22α-tetrahydroxy olean-23,28-dioic acid 2 and 6 known compounds (3–8) have been isolated from Gladiolus segetum Ker-Gawl corms. The structural elucidation of the isolated compounds was confirmed using different chemical and spectroscopic methods, including 1D and 2D NMR experiments as well as HR-ESI-MS. Moreover, the in vitro cytotoxic activity of the fractions and that of the isolated compounds 1–8 were investigated against five human cancer cell lines (PC-3, A-549, HePG-2, MCF-7 and HCT-116) using doxorubicin as a reference drug. The results showed that the saponin fraction exhibited potent in vitro cytotoxic activity against the five human cancer cell lines, whereas the maximum activity was exhibited against the PC-3 and A-549 cell lines with the IC₅₀ values of 1.13 and 1.98 μg mL⁻¹, respectively. In addition, compound 1 exhibited potent activity against A-549 and PC-3 with the IC₅₀ values of 2.41 μg mL⁻¹ and 3.45 μg mL⁻¹, respectively. Interestingly, compound 2 showed the maximum activity against PC-3 with an IC₅₀ of 2.01 μg mL⁻¹. These biological results were in harmony with that of the molecular modeling study, which showed that the cytotoxic activity of compound 2 might occur through the inhibition of the HER-2 enzyme.

A new triterpenoidal saponin 1, a new oleanane triterpene 2, and 6 known compounds (3–8) have been isolated from Gladiolus segetum Ker-Gawl corms.     

Synthesis and insight into the structure–activity relationships of 2-phenylbenzimidazoles as prospective anticancer agents

In order to explore and develop new anticancer agents, three series of 2-phenylbenzimidazoles, 15–46, were condensed under simple and mild conditions using sodium metabisulfite as an oxidation agent and another series, 47–55, were obtained via a reduction reaction using sodium borohydride. All the compounds synthesized were evaluated for their in vitro anticancer activities against three human cancer cell lines. The novel compound 38 was found to be the most potent multi cancer inhibitor against A549, MDA-MB-231, and PC3 cell lines (IC₅₀ values 4.47, 4.68 and 5.50 μg mL⁻¹, respectively). In addition, compound 40 exhibited the best IC₅₀ value of 3.55 μg mL⁻¹ against the MDA-MB-231 cell line. The results demonstrated that introducing a new substituent to compounds 37–55 could improve their antiproliferative activities.

Three series of 2-phenylbenzimidazoles obtained under simple and convenient pathways, were used to elucidate their SARs against three cancer cell lines: A549, MDA-MB-231 and PC3.     

Synthesis,human topoisomerase IIα inhibitory properties and molecular modeling studies of anti-proliferative curcumin mimics

3,5-Bis(arylidene)-N-substituted-4-oxo-piperidine-1-carboxamides 24–51 were synthesized as curcumin mimics in a facile pathway through reaction of 3,5-bis(arylidene)-4-piperidones with the appropriate isocyanate in the presence of triethylamine. The 3E,5E′-stereochemical configuration was conclusively supported by single crystal X-ray studies of compounds 25 and 34. Most of the synthesized piperidinecarboxamides showed high anti-proliferative properties with potency higher than that of 5-fluorouracil (clinically approved drug against colon, breast and skin cancers) through in vitro MTT bio-assay. Some of them revealed anti-proliferative properties at sub-micromolar values (IC₅₀ = 0.56–0.70 μM for compounds 29, 30 and 34–38 against HCT116; and IC₅₀ = 0.64 μM for compound 30 against A431 cell lines) with promising inhibitory properties against human DNA topoisomerase IIα. The safe profile of the anti-proliferative active agents against the RPE1 normal cell line may prove their selectivity towards carcinoma cells. Robust molecular models (2D-QSAR, 3D-pharmacophore) supported the SAR and validated the observed bio-properties.

Piperidinecarboxamides (curcumin mimics) show promising anti-proliferative properties against HCT116 (colon), MCF7 (breast) and A431 (squamous skin) carcinoma cell lines with potency higher than that of 5-fluorouracil.