Intracellular Forms Research Articles

Change in inorganic phosphate physical state can regulate transcription

ABSTRACTInorganic phosphate (Pi), a ubiquitous metabolite, is involved in all major biochemical pathways. We demonstrate that, in vitro, MgHPO4 (the intracellular Pi form) at physiological concentrations can exist in a metastable supersaturated dissolved state or as a precipitate. We have shown that in solution, MgHPO4 strongly stimulates exonuclease nascent transcript cleavage by RNA polymerase. We report here that MgHPO4 precipitate selectively and efficiently inhibits transcription initiation in vitro. In view of the MgHPO4 solubility and in vitro sensitivity of RNA synthesis to MgHPO4 precipitate, at physiological concentrations, MgHPO4 should cause a 50–98% inhibition of cellular RNA synthesis, thus exerting a strong regulatory action. The effects of Pi on transcription in vivo will, therefore, reflect the physical state of intracellular Pi.

Novel functionalized 1,2,3-triazole derivatives exhibit antileishmanial activity, increase in total and mitochondrial-ROS and depolarization of mitochondrial membrane potential of Leishmania amazonensis

1,2,3-triazolium salts are poorly understood regarding their antileishmanial activity. Hence, as an effort to identify novel chemical scaffolds as antileishmanial agents, a series of 1,2,3-triazolium salts (TS) and corresponding 1,2,3-triazole (T) precursors including new epoxide derivatives were synthesized and assayed against Leishmania amazonensis promastigote and intracellular amastigote forms. Among them, the compound TS-6 exhibited promising activity on promastigotes (IC50 = 3.61 μM) and intracellular amastigotes (IC50 = 7.61 μM) of L. amazonensis, superior to miltefosine (IC50 > 10.0 μM), used as reference drug. In addition, TS-6 showed negligible cytotoxicity on murine peritoneal macrophages with a SI of about 10. Studies on the mode of action of TS-6 indicate mitochondrial dysfunction through an increase in ‘total’ and mitochondrial-ROS as well as depolarization of mitochondrial membrane potential of L. amazonensis promastigotes. In silico physicochemical studies indicate that the TS-6 could potentially be used as an oral drug.

Remodeling of the Actin Network Associated with the Non-Structural Protein 1 (NS1) of West Nile Virus and Formation of NS1-Containing Tunneling Nanotubes.

The cellular response to the recombinant NS1 protein of West Nile virus (NS1WNV) was studied using three different cell types: Vero E6 simian epithelial cells, SH-SY5Y human neuroblastoma cells, and U-87MG human astrocytoma cells. Cells were exposed to two different forms of NS1WNV: (i) the exogenous secreted form, sNS1WNV, added to the extracellular milieu; and (ii) the endogenous NS1WNV, the intracellular form expressed in plasmid-transfected cells. The cell attachment and uptake of sNS1WNV varied with the cell type and were only detectable in Vero E6 and SH-SY5Y cells. Addition of sNS1WNV to the cell culture medium resulted in significant remodeling of the actin filament network in Vero E6 cells. This effect was not observed in SH-SY5Y and U-87MG cells, implying that the cellular uptake of sNS1WNV and actin network remodeling were dependent on cell type. In the three cell types, NS1WNV-expressing cells formed filamentous projections reminiscent of tunneling nanotubes (TNTs). These TNT-like projections were found to contain actin and NS1WNV proteins. Interestingly, similar actin-rich, TNT-like filaments containing NS1WNV and the viral envelope glycoprotein EWNV were also observed in WNV-infected Vero E6 cells.

Trypanocidal Trixikingolides From Trixis Vauthieri

Chagas disease is an illness caused by the protozoan parasite Trypanosoma cruzi. Only two drugs are available, with the drawback of low rate of cure in the chronic phase of the disease and undesirable side effects. These facts highlight the need to find new compounds for Chagas disease chemotherapy. We describe the isolation and identification of an inseparable mixture of two new trixikingolides from Trixis vauthieri, a plant from family Asteraceae, which present outstanding in vitro trypanocidal activity, with IC50 value of 0.053 µM against the intracellular trypomastigotes and amastigotes forms of T. cruzi infecting L929 cells. The IC50 of the mixture against the host cells is 68 times higher and about 70 times more potent than benznidazole, the reference drug used as control at the experiments. The next step, which depends on obtaining larger quantities of the mixture, is to test it on mice infected with T. cruzi.

In vitro assessment of 3-alkoxy-5-nitroindazole-derived ethylamines and related compounds as potential antileishmanial drugs

Leishmaniasis is a widespread neglected tropical disease complex that is responsible of one million new cases per year. Current treatments are outdated and pose many problems that new drugs need to overcome. With the goal of developing new, safe, and affordable drugs, we have studied the in vitro activity of 12 different 5-nitroindazole derivatives that showed previous activity against different strains of Trypanosoma cruzi in a previous work. T. cruzi belongs to the same family as Leishmania spp., and treatments for the disease it produces also needs renewal. Among the derivatives tested, compounds 1, 2, 9, 10, 11, and 12 showed low J774.2 macrophage toxicity, while their effect against both intracellular and extracellular forms of the studied parasites was higher than the ones found for the reference drug Meglumine Antimoniate (Glucantime®). In addition, their Fe-SOD inhibitory effect, the infection rates, metabolite alteration, and mitochondrial membrane potential of the parasites treated with the selected drugs were studied in order to gain insights into the action mechanism, and the results of these tests were more promising than those found with glucantime, as the leishmanicidal effect of these new drug candidates was higher. The promising results are encouraging to test these derivatives in more complex studies, such as in vivo studies and other experiments that could find out the exact mechanism of action.

The effects of Fusarium graminarum silver nanoparticles on leishmania tropica

The present study aims to evaluate the in vitro antileishmanial activities of biosynthesis silver nanoparticles from Fusarium graminarum and to compare its efficacy with a pentostam drug against Leishmania Tropica.This nanaoparticles (20-40-80-100 μg/mL) was evaluated in vitro against promastigote and intracellular amastigote forms of L. tropica also measured the viability by use MTTassy. Its significantly ( < 0.05) inhibited the growth rate of promastigote the results showed decrease in number of parasite after 24hr was(1816.6,1300,541.6, ×104) cell\\ml respectively, compared with pentostam and control was(1558.3 and 2441.6,308,3×104) cell\\ml respectively. whereas after 48hr the number of parasite became(1583.3,1166.6,450,225×104) cell\\ml respectively compared with pentostam and control was(1066.6 and 3166.6×104) cell\\ml respectively, also observed (100 μg/mL)more effects on multiplication ofamastigote after 24hr become only 1.6parasite compared with pentostam and control was(3.3,7 parasites).The results of viability also revealed that the nanoparticles Cytotoxicty effect dependent on concentration and reduce to reach (23% ). Obtained findings also provide the scientific evidences that fungi could be used in the biosynthesis nanoparticles and treatment of CL.

Proteolytic and nonproteolytic activation mechanisms result in conformationally and functionally different forms of coagulation factor XIII A.

Factor XIIIA (FXIIIA) is a transglutaminase that cross-links intra- and extracellular protein substrates. FXIIIA is expressed as an inactive zymogen, and during blood coagulation, it is activated by removal of an activation peptide by the protease thrombin. No such proteolytic FXIIIA activation is known to occur in other tissues or the intracellular form of FXIIIA. For those locations, FXIIIA is assumed instead to undergo activation by Ca2+ ions. Previously, we demonstrated a monomeric state for active FXIIIA. Current analytical ultracentrifugation and kinetic experiments revealed that thrombin-activated FXIIIA has a higher conformational flexibility and a stronger affinity toward glutamine substrate than does nonproteolytically activated FXIIIA. The proteolytic activation of FXIIIA was further investigated in a context of fibrin clotting. In a series of fibrin cross-linking assays and scanning electron microscopy studies of plasma clots, the activation rates of FXIIIA V34X variants were correlated with the extent of fibrin cross-linking and incorporation of nonfibrous protein into the clot. Overall, the results suggest conformational and functional differences between active FXIIIA forms, thus expanding the understanding of FXIIIA function. Those differences may serve as a basis for developing therapeutic strategies to target FXIIIA in different physiological environments. ENZYMES: Factor XIIIA ( EC 2.3.2.13).

Metabolomics, lipidomics and proteomics profiling of myoblasts infected with Trypanosoma cruzi after treatment with different drugs against Chagas disease.

Chagas disease, the most important parasitic infection in Latin America, is caused by the intracellular protozoan Trypanosoma cruzi. To treat this disease, only two nitroheterocyclic compounds with toxic side effects exist and frequent treatment failures are reported. Hence there is an urgent need to develop new drugs. Recently, metabolomics has become an efficient and cost-effective strategy for dissecting drug mode of action, which has been applied to bacteria as well as parasites, such as different Trypanosome species and forms. We assessed if the metabolomics approach can be applied to study drug action of the intracellular amastigote form of T. cruzi in a parasite-host cell system. We applied a metabolic fingerprinting approach (DI-MS and NMR) to evaluate metabolic changes induced by six different (candidate) drugs in a parasite-host cell system. In a second part of our study, we analyzed the impact of two drugs on polar metabolites, lipid and proteins to evaluate if affected pathways can be identified. Metabolic signatures, obtained by the fingerprinting approach, resulted in three different clusters. Two can be explained by already known of mode actions, whereas the three experimental drugs formed a separate cluster. Significant changes induced by drug action were observed in all the three metabolic fractions (polar metabolites, lipids and proteins). We identified a general impact on the TCA cycle, but no specific pathways could be attributed to drug action, which might be caused by a high percentage of common metabolome between a eukaryotic host cell and a eukaryotic parasite. Additionally, ion suppression effects due to differences in abundance between host cells and parasites may have occurred. We validated the metabolic fingerprinting approach to a complex host-cell parasite system. This technique can potentially be applied in the early stage of drug discovery and could help to prioritize early leads or reconfirmed hits for further development.

Organometallic Gold(III) Complex [Au(Hdamp)(L14)]+ (L1 = SNS-Donating Thiosemicarbazone) as a Candidate to New Formulations against Chagas Disease

Chagas disease remains a serious public health concern with unsatisfactory treatment outcomes due to strain-specific drug resistance and various side effects. To identify new therapeutic drugs against Trypanosoma cruzi, we evaluated both the in vitro and in vivo activity of the organometallic gold(III) complex [Au(III)(Hdamp)(L14)]Cl (L1 = SNS-donating thiosemicarbazone), henceforth denoted 4-Cl. Our results demonstrated that 4-Cl was more effective than benznidazole (Bz) in eliminating both the extracellular trypomastigote and intracellular amastigote forms of the parasite without cytotoxic effects on mammalian cells. In in vivo assays, 4-Cl in PBS solution loses the protonation and becomes the 4-neutral. 4-Neutral reduced parasitaemia and tissue parasitism in addition to protecting the liver and heart from tissue damage at 2.8 mg/kg/day. All these changes resulted in the survival of 100% of the mice treated with the gold complex during the acute phase. Analyzing the surviving animals of the acute infection, the parasite load after 150 days of infection was equivalent to those treated with the standard dose of Bz without demonstrating the hepatotoxicity of the latter. In addition, we identified a modulation of interferon gamma (IFN-γ) levels that may be targeting the disease's positive outcome. To the best of our knowledge, this is the first gold organometallic study that shows promise in an in vivo experimental model against Chagas disease.

Influence of pH on the activity of finafloxacin against extracellular and intracellular Burkholderia thailandensis, Yersinia pseudotuberculosis and Francisella philomiragia and on its cellular pharmacokinetics in THP-1 monocytes

ObjectivesBurkholderia pseudomallei, Yersinia pestis and Francisella tularensis are facultative intracellular bacteria causing life-threatening infections. We have (a) compared the activity of finafloxacin (a fluoroquinolone in development showing improved activity at acidic pH) with that of ciprofloxacin, levofloxacin and imipenem against the extracellular and intracellular (THP-1 monocytes) forms of infection by attenuated surrogates of these species (B. thailandensis, Y. pseudotuberculosis, F. philomiragia) and (b) assessed finafloxacin cellular pharmacokinetics (accumulation, distribution, efflux). MethodsBacteria in broth or in infected monocytes were exposed to antibiotics at pH 7.4 or 5.5 for 24 hr. Maximal relative efficacies (Emax) and static concentrations (Cs) were calculated using the Hill equation (concentration–response curves). Finafloxacin pharmacokinetics in cells at pH 7.4 or 5.5 was investigated using 14C-labelled drug. ResultsExtracellularly, all drugs sterilized the cultures, with finafloxacin being two to six times more potent at acidic pH. Intracellularly, Emax reached the limit of detection (4–5 log10 cfu decrease) for finafloxacin against all species, but only against B. thailandensis and F. philomiragia for ciprofloxacin and levofloxacin, while imipenem caused less than 2 log10 cfu decrease for all species. At acid pH, Cs shifted to two to five times lower values for finafloxacin and to one to four times higher values for the other drugs. Finafloxacin accumulated in THP-1 cells by approximately fivefold at pH 7.4 but up to 20-fold at pH 5.5, and distributed in the cytosol. ConclusionsFluoroquinolones have proven to be effective in reducing the intracellular reservoirs of B. thailandensis, Y. pseudotuberculosis and F. philomiragia, with finafloxacin demonstrating an additional advantage in acidic environments.

Mini-Hydrocyclone Separation of Cyanobacterial and Green Algae: Impact on Cell Viability and Chlorine Consumption

The co-occurrence of non-toxic phytoplankton alongside cyanobacteria adds to the challenge of treating source waters with harmful algal blooms. The non-toxic species consume the oxidant and, thereby, reduce the efficacy of oxidation of both the extracellular and intracellular cyanotoxins. In this work, a 3D printed mini-hydrocyclone was used to separate a mixture of non-toxic green algae, Scenedesmus obliquus, from a toxic species of cyanobacteria, Microcystis aeruginosa. When water is pumped through the mini-hydrocyclone, cells exit through an overflow or underflow port depending on their size, shape, and density relative to the other cells and particles in the water matrix. The overflow port contains the cells that are smaller and less dense since these particles move toward the center of the hydrocyclone. In this work, the majority (&gt;93%) of Microcystis cells were found in the overflow while the underflow contained primarily the Scenedesmus (&gt;80%). This level of separation efficiency was maintained over the 30-min experiment and the majority of both cells (&gt;86%) remained viable following the separation, which indicates that the pumping combined with forces exerted within the mini-hydrocyclone were not sufficient to cause cell death. The impact of free chlorine on the cells both pre-separation and post-separation was evaluated at two doses (1 and 2 mg/L). After separation, the overflow, which contained primarily Microcystis, had at least a 24% reduction in the free chlorine decay rate as compared to the feed water, which contained both species. This reduction in chlorine consumption shows that the cells separated via mini-hydrocyclone would likely require lower doses of oxidant to produce a similar level of degradation of the cyanotoxins present in either the extracellular or intracellular form. However, future work should be undertaken to evaluate this effect in natural bloom samples.

Endlicheria bracteolata (Meisn.) Essential Oil as a Weapon Against Leishmania amazonensis: In Vitro Assay.

The difficulties encountered and the numerous side effects present in the treatment of cutaneous leishmaniasis have encouraged the research for new compounds that can complement or replace existing treatment. The growing scientific interest in the study of plants, which are already used in folk remedies, has led our group to test Endlicheria bracteolata essential oil against Leishmania amazonensis. Several species of the Lauraceae family, or their compounds, have relevant antiprotozoal activities Therefore, the biological potential on L. amazonensis forms from the essential oil of Endlicheria bracteolata leaves was verified for the first time in that work. The antileishmanial activity was evaluated against promastigotes and intracellular amastigotes, and cytotoxicity were performed with J774.G8, which were incubated with different concentrations of E. bracteolata essential oil. Transmission electron microscopy and flow cytometry were performed with E. bracteolata essential oil IC50. Promastigote forms showed E. bracteolata essential oil IC50 of 7.945 ± 1.285 µg/mL (24 h) and 6.186 ± 1.226 µg/mL (48 h), while for intracellular amastigote forms it was 3.546 ± 1.184 µg/mL (24 h). The CC50 was 15.14 ± 0.090 µg/mL showing that E. bracteolata essential oil is less toxic to macrophages than to parasites. Transmission electron microscopy showed that E. bracteolata essential oil treatment is capable of inducing mitochondrial damage to promastigote and intracellular amastigote forms, while flow cytometry showed ΔѰm disruption in treated parasites. These results could bring about new possibilities to develop products based on E. bracteolata essential oil to treat cutaneous leishmaniasis, especially for people who cannot receive the conventional therapy.

4-Aminoquinoline-based compounds as antileishmanial agents that inhibit the energy metabolism of Leishmania

Among neglected tropical diseases, leishmaniasis is one of the most relevant with an estimated 30,000 deaths annually. Existing therapies have serious drawbacks in safety, drug resistance, field-adapted application and cost; therefore, new safer and shorter treatments are needed for this disease. Here we report on the synthesis of novel 4-amino-7-chloroquinoline-based compounds with leishmanicidal activity, together with deeper insight into the mechanism of action of our previously published hit compound 1. New derivatives showed comparable activity to 1 against both promastigote and intracellular amastigote forms of Leishmania infantum, with IC50 < 1 μM. Furthermore, we have determined that compound 1 induced a decrease of intracellular ATP levels, as well as a mitochondrial depolarization, together with an alteration of plasma membrane permeability and a significant ROS production. The inhibition of the energy metabolism of Leishmania plays an important role in the leishmanicidal mechanism of this compound. In all, these results support the consideration of compound 1 for the future development of new leishmanicidal drugs.

Heterobimetallic nickel(II) and palladium(II) complexes derived from S-benzyl-N- (ferrocenyl)methylenedithiocarbazate: Trypanocidal activity and interaction with Trypanosoma cruzi Old Yellow Enzyme (TcOYE)

Reactions of Ni(II) and Pd(II) precursors with S-benzyl-N-(ferrocenyl)methylenedithiocarbazate (HFedtc) led to the formation of heterobimetallic complexes of the type [MII(Fedtc)2] (M = Ni and Pd). The characterization of the compounds involved the determination of melting point, FTIR, UV–Vis, 1H NMR, elemental analysis and electrochemical experiments. Furthermore, the crystalline structures of HFedtc and [NiII(Fedtc)2] were determined by single crystal X-ray diffraction. The compounds were evaluated against the intracellular form of Trypanosoma cruzi (Tulahuen Lac-Z strain) and the cytotoxicity assays were assessed using LLC-MK2 cells. The results showed that the coordination of HFedtc to Ni(II) or Pd(II) decreases the in vitro trypanocidal activity while the cytotoxicity against LLC-MK2 cells does not change significantly. [PdII(Fedtc)2] showed the greater potential between the two complexes studied, showing an SI value of 8.9. However, this value is not better than that of the free ligand with an SI of 40, a similar value to that of the standard drug benznidazole (SI = 48). Additionally, molecular docking simulations were performed with Trypanosoma cruzi Old Yellow Enzyme (TcOYE), which predicted that HFedtc binds to the protein, almost parallel to the flavin mononucleotide (FMN) prosthetic group, while the [NiII(Fedtc)2] complex was docked into the enzyme binding site in a significantly different manner. In order to confirm the hypothetical interaction, in vitro experiments of fluorescence quenching and enzymatic activity were performed which indicated that, although HFedtc was not processed by the enzyme, it was able to act as a competitive inhibitor, blocking the hydride transfer from the FMN prosthetic group of the enzyme to the menadione substrate.

Hedyosulide, a novel trypanosomicidal sesterterpene lactone from Hedyosmum brasiliense Mart. ex Miq

A new sesterterpene lactone, Hedyosulide, together with five previously described sesquiterpene lactones were isolated from the leaves of Hedyosmum brasiliense Mart. ex Miq. (Chloranthaceae). Their structures were established by NMR analysis and HRESIMS data. The isolated compounds were evaluated for antiprotozoal activity against trypomastigote and amastigote forms of Trypanosoma cruzi. Hedyosulide displayed the highest activity against the amastigote forms, in which the parasite takes in the vertebrate host, with IC50 of 21.6 μmol/L with a selectivity index (SI) higher than 9. Oxyonoseriolide showed moderate activity against the intracellular form with IC50 of 60.2 μmol/L and a weak SI (2.7), while the four other sesquiterpene lactones were inactive. Therefore, sesterterpene lactones, a relatively rare group of terpenoids, might be considered as a scaffold for developing new trypanosomicidal agents that are effective against T. cruzi. Also, this work brings an important contribution on the chemical/biological aspects of H. brasiliense due to the isolation of a sesterterpene lactone selectively active against both clinical forms of T. cruzi.

Abstract 2096: Resistance to APTO-253 caused by internal deletion and alternate promoter usage of the MYC gene in malignant B cells

Abstract APTO-253 is a clinical stage small molecule that acts in part by down-regulation of MYC at mRNA and protein levels. MYC onco-protein dysregulation, which leads to increased proliferation and survival, is implicated in the etiology of numerous cancer subtypes. In the current study, we explored sensitivity of B-cell cancer cells to APTO-253 and sought to understand how changes in MYC can yield resistance to APTO-253. APTO-253 showed potency against B-cell cancer cell lines (IC50 30-600 nM) and bone marrow samples from patients. Sensitivity of cell lines correlated with MYC over-expression, amplification or translocation. APTO-253 resistant Raji cells (Raji253R) are characterized by upregulation of the ABCG2 transporter. Treatment with ABCG2 inhibitors only partially reversed the resistant phenotype suggesting additional contributing mechanisms. RNA-seq analysis revealed that while levels of MYC mRNA were unchanged in Raji253R, a truncation of exon 2 was present. This internal deletion occurred at the DNA level, not by alternative splicing, and resulted in a truncated MYC protein (45 kDa) lacking the conserved core sequence of MYC box III (MBIII). At early stages in the evolution of resistance residual full length MYC mRNA and protein was detectable; however, this expression was eventually lost in Raji253R cells as selection continued. Deletion of MBIII has been implicated in MYC protein stability, in vitro and in vivo transformation, gene repression, and MYC dependent apoptosis. Cycloheximide treatment of Raji and Raji253R cells demonstrated a modest increase in stability of truncated over full length MYC protein. In addition, Raji253R cells exhibited hypersensitivity to etoposide. The internal truncation is flanked by sites of microhomology suggesting a method for deletion. We hypothesize that DNA damage in response to APTO-253 treatment led to error prone repair by microhomology end joining and resulted in a MYC internal deletion. Interestingly, transcription from the canonical P1/P2 promoter, as seen in WT Raji cells, was systematically lost and transcription initiation shifted to a downstream promoter, P3. APTO-253 inhibits MYC expression via binding and stabilizing the G-quadruplex (G4) sequence in the P1/P2 promoter therefore switching of transcriptional initiation to P3 should relieve APTO-253 mediated repression. Indeed, APTO-253 treatment of Raji253R cells did not repress MYC expression, even at concentrations 50-fold higher than required for MYC repression in Raji WT cells. Presence of the active intra-cellular form of APTO-253, Fe(253)3, was confirmed in Raji253R cells. In summary, MYC driven Raji cells become resistant to APTO-253 by acquisition of a more stable MYC protein and by utilization of an alternate promoter not inhibited by G4 binding and stabilization, thereby confirming the essential role of MYC repression in the mechanism of action of APTO-253. Citation Format: Andrea Local, Cheng-Yu Tsai, Hongying Zhang, Susan Sheng, Khalid Benbatoul, Stephen B. Howell, William G. Rice. Resistance to APTO-253 caused by internal deletion and alternate promoter usage of the MYC gene in malignant B cells [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 2096.

Discovery and characterization of trypanocidal cysteine protease inhibitors from the ‘malaria box’

Chagas disease, Human African Trypanosomiasis, and schistosomiasis are neglected parasitic diseases for which new treatments are urgently needed. To identify new chemical leads, we screened the 400 compounds of the Open Access Malaria Box against the cysteine proteases, cruzain (Trypanosoma cruzi), rhodesain (Trypanosoma brucei) and SmCB1 (Schistosoma mansoni), which are therapeutic targets for these diseases. Whereas just three hits were observed for SmCB1, 70 compounds inhibited cruzain or rhodesain by at least 50% at 5 μM. Among those, 15 commercially available compounds were selected for confirmatory assays, given their potency, time-dependent inhibition profile and reported activity against parasites. Additional assays led to the confirmation of four novel classes of cruzain and rhodesain inhibitors, with potency in the low-to mid-micromolar range against enzymes and T. cruzi. Assays against mammalian cathepsins S and B revealed inhibitor selectivity for parasitic proteases. For the two competitive inhibitors identified (compounds 7 and 12), their binding mode was predicted by docking, providing a basis for structure-based optimization efforts. Compound 12 also acted directly against the trypomastigote and the intracellular amastigote forms of T. cruzi at 3 μM. Therefore, through a combination of experimental and computational approaches, we report promising hits for optimization in the development of new trypanocidal drugs.

GDF-15: A Multifunctional Modulator and Potential Therapeutic Target in Cancer.

Various pathological processes are associated with the aberrant expression and function of cytokines, especially those belonging to the transforming growth factor-β (TGF-β) family. Nevertheless, the functions of members of the TGF-β family in cancer progression and therapy are still uncertain. Growth differentiation factor- 15, which exists in intracellular and extracellular forms, is classified as a divergent member of the TGF-β superfamily. It has been indicated that GDF-15 is also connected to the evolution of cancer both positively and negatively depending upon the cellular state and environment. Under normal physiological conditions, GDF-15 inhibits early tumour promotion. However, its abnormal expression in advanced cancers causes proliferation, invasion, metastasis, cancer stem cell formation, immune escape and a reduced response to therapy. As a clinical indicator, GDF-15 can be used as a tool for the diagnosis and therapy of an extensive scope of cancers. Although some basic functions of GDF-15 are noncontroversial, their mechanisms remain unclear and complicated at the molecular level. Therefore, GDF-15 needs to be further explored and reviewed.

Niosomal formulation of amphotericin B alone and in combination with glucantime: In vitro and in vivo leishmanicidal effects

This study aimed to evaluate the efficacy of glucantime and amphotericin B (AmB) encapsulated in niosome against cutaneous leishmaniasis (CL) using in vitro and in vivo models. The niosomal formulations of the drugs alone and in combination were prepared and characterized. Subsequent to the examination of their cytotoxicity, their efficacy was evaluated using an in vitro MTT assay, macrophage model, flow cytometry, and gene expression profiling. For evaluation of therapeutic effect of niosomal combination on the lesion induced by Leishmania major in inbred BALB/c mice, the size of lesions and number of parasites in spleen was assessed. The niosomal formulations demonstrated significantly greater inhibitory effects compared with the non-niosomal forms when the IC50 was considered. The niosomal combination showed an increase in the apoptotic values and gene expression levels of IL-12 and metacaspase and a decrease in the levels of IL-10 with a dose-response effect. The niosomal combination was also effective in reducing the lesion size and splenic parasite burden in mice. Our findings indicated that there is a synergistic effect between AmB and glucantime in niosomal form in the inhibition of intracellular and extracellular forms of L. tropica. Additionally, the in vivo results on L. major suggest that topical niosomal formulation could be useful in the treatment of CL.

Leishmania interaction with an osteoclast

Visceral leishmaniasis is caused by an intracellular protozoan parasite, Leishmania donovani. Frequently, a bone marrow aspirate can confirm the diagnosis by showing intracellular forms of Leishmania amastigotes in macrophages. Here we present a unusual interaction of Leishmania with an osteoclast in the bone marrow.