Probing the B- & C-rings of the antimalarial tetrahydro-ß-carboline MMV008138 for steric and conformational constraints.

The antimalarial candidate MMV008138 (1a) is of particular interest because its target enzyme (IspD) is absent in human. To achieve higher potency, and to probe for steric demand, a series of analogs of 1a were prepared that featured methyl-substitution of the B- and C-rings, as well as ring-chain transformations. X-ray crystallography, NMR spectroscopy and calculation were used to study the effects of these modifications on the conformation of the C-ring and orientation of the D-ring. Unfortunately, all the B- and C-ring analogs explored lost in vitro antimalarial activity. The possible role of steric effects and conformational changes on target engagement are discussed.

Molecular Characterization of Aquaglyceroporine: A Novel Mutation in LmAQP1 from Leishmania major (MRHO/IR/75/ER).

BACKGROUND: The first line treatment for cutaneous leishmaniasis is pentavalent antimony such as sodium stibogluconate (pentostam) and meglumine antimonite (glucantime). One of the most important ways to uptake the drug is by a trans-membrane protein, called aquaglyceroporin encoded by Aquaglyceroprotein1 (LmAQP1). In this study, molecular characterization of LmAQP1 was reported. METHODS: Leishmania major (MRHO/IR/75/ER) promastigotes were cultured, and then DNA extraction and RNA extraction were done and followed by cDNA synthesis. Amplicons resulted from PCR and RT-PCR using specific primers were purified and sequenced. Molecular characterization was done by bioinformatically software such as BLST, ClustalW2, and RMSD. RESULTS: Amplicons resulted from PCR and RT-PCR showed equal size in length. BLASTn analysis showed a point nucleotide change in LmAQP1 gene that encoded 282-amino-acid long protein with a mutation at position 154 including replacement of alanine by threonine. The observed mutation in the interested gene was assessed using the above-mentioned software. The mentioned gene was submitted at GenBank, NCBI with accession number of KU514052. CONCLUSION: The functional prediction of the protein encoded from LmAQP1 showed that the mentioned mutation could not affect the three-dimension structure, but it may modify the drug uptake potential of this important channel. Based on from LmAQP1 role, it seems to be an appropriate candidate for drug development. According to search through internet, this is the first report of LmAQP1 from L. major (MRHO/IR/75/ER).

Questioning the γ-gauche effect: stereoassignment of 1,3-disubstituted-tetrahydro-ß-carbolines using 1H-1H coupling constants.

The Pictet-Spengler reaction of tryptophan esters and aldehydes has been widely applied in natural product synthesis and medicinal chemistry. To date, the trans- or cis-configuration of 1,3-disubstituted tetrahydro-ß-carbolines (THßCs) formed in this reaction has most often been assigned based on the relative 13C chemical shifts of C1 and C3 in the diastereomers. Although the upfield shifts of C1 and C3 in trans-THßCs relative to cis-THßCs has been attributed to steric compression associated with the "γ-gauche" effect, we show that this effect is not borne out experimentally for other carbons that should suffer this same compression. Thus we developed a robust alternative method for stereochemical assignment based on 1H NMR coupling constants (31 examples) and supported by extensive DFT-based conformational analysis and calculation of 1H-1H coupling constants. DFT calculations of 13C NMR chemical shifts also cast doubt upon the role of the "γ-gauche" effect on C1 and C3 chemical shifts in trans-THßCs.

Biological Studies and Target Engagement of the 2- C-Methyl-d-Erythritol 4-Phosphate Cytidylyltransferase (IspD)-Targeting Antimalarial Agent (1 R,3 S)-MMV008138 and Analogs.

Malaria continues to be one of the deadliest diseases worldwide, and the emergence of drug resistance parasites is a constant threat. Plasmodium parasites utilize the methylerythritol phosphate (MEP) pathway to synthesize isopentenyl pyrophosphate (IPP) and dimethylallyl pyrophosphate (DMAPP), which are essential for parasite growth. Previously, we and others identified that the Malaria Box compound MMV008138 targets the apicoplast and that parasite growth inhibition by this compound can be reversed by supplementation of IPP. Further work has revealed that MMV008138 targets the enzyme 2- C-methyl-d-erythritol 4-phosphate cytidylyltransferase (IspD) in the MEP pathway, which converts MEP and cytidine triphosphate (CTP) to cytidinediphosphate methylerythritol (CDP-ME) and pyrophosphate. In this work, we sought to gain insight into the structure-activity relationships by probing the ability of MMV008138 analogs to inhibit PfIspD recombinant enzyme. Here, we report PfIspD inhibition data for fosmidomycin (FOS) and 19 previously disclosed analogs and report parasite growth and PfIspD inhibition data for 27 new analogs of MMV008138. In addition, we show that MMV008138 does not target the recently characterized human IspD, reinforcing MMV008138 as a prototype of a new class of species-selective IspD-targeting antimalarial agents.

The Impact of Demographic Factors and Blood Sugar Control on the Incidence of Urinary Tract Infections in Khorramabad in 2013.

BACKGROUND: Diabetes is one of the most serious metabolic disorders to affect different organs in the body. Patients with diabetes are hence at risk of developing other problems. Urinary tract infections (UTI) are one of the problems that occur more commonly in diabetic patients than in others. There are several risk factors that lead to the incidence of UTI in diabetic patients. OBJECTIVES: We conducted this study to determine the effect of different factors on the incidence of UTI in diabetic patients in order to decrease the incidence of this problem. PATIENTS AND METHODS: In this cross-sectional study, data concerning 233 patients who were referred to the largest diabetes center in Khorramabad, Lorestan province, Iran, were recorded. The relationships between various factors and the incidence of UTI in these patients were evaluated using SPSS version 16. We used chi-square and logistic regression to analyze the data. A P < 0.05 was considered to be statistically significant. RESULTS: The incidence of UTI increased significantly with increasing age (P = 0.009). The incidence of UTI was higher among females than among males, and it was higher among unemployed people than among those in other occupations. The prevalence of UTI among patients who control their diabetes through diet and exercise was lower than that among those undergoing other types of treatment; however, no significant relationship was found between the different types of oral medication and the incidence of UTI. We did not find any relationship between the incidence of UTI in diabetic patients and other factors such as literacy, marital status, BMI, addiction, history of hospitalization, and level of HbA1c. CONCLUSIONS: We found that age, gender, occupation, and the type of medication used are all risk factors for the incidence of UTI in diabetic patients. Other factors such as literacy, marital status, addiction, and history of hospitalization did not have any significant association with the incidence of UTI in diabetic patients.

Parallel inhibition of amino acid efflux and growth of erythrocytic Plasmodium falciparum by mefloquine and non-piperidine analogs: Implication for the mechanism of antimalarial action.

Despite the troubling psychiatric side-effects it causes in some patients, mefloquine (MQ) has been used for malaria prophylaxis and therapy, due to its activity against all Plasmodium species, its ease of dosing, and its relative safety in children and pregnant women. Yet at present there is no consensus on the mechanism of antimalarial action of MQ. Two leading hypotheses for the mechanism of MQ are inhibition of heme crystallization and inhibition of host cell hemoglobin endocytosis. In this report we show that MQ is a potent and rapid inhibitor of amino acid efflux from intact parasitized erythrocytes, which is a measure of the in vivo rate of host hemoglobin endocytosis and catabolism. To further explore the mechanism of action of MQ, we have compared the effects of MQ and 18 non-piperidine analogs on amino acid efflux and parasite growth. Among these closely related compounds, an excellent correlation over nearly 4 log units is seen for 50% inhibition concentration (IC50) values for parasite growth and leucine efflux. These data and other observations are consistent with the hypothesis that the antimalarial action of these compounds derives from inhibition of hemoglobin endocytosis.

3-Oxoisoxazole-2(3H)-carboxamides and isoxazol-3-yl carbamates: Resistance-breaking acetylcholinesterase inhibitors targeting the malaria mosquito, Anopheles gambiae.

To identify potential selective and resistance-breaking mosquitocides against the African malaria vector Anopheles gambiae, we investigated the acetylcholinesterase (AChE) inhibitory and mosquitocidal properties of isoxazol-3-yl dimethylcarbamates (15), and the corresponding 3-oxoisoxazole-2(3H)-dimethylcarboxamide isomers (14). In both series, compounds were found with excellent contact toxicity to wild-type susceptible (G3) strain and multiply resistant (Akron) strain mosquitoes that carry the G119S resistance mutation of AChE. Compounds possessing good to excellent toxicity to Akron strain mosquitoes inhibit the G119S mutant of An. gambiae AChE (AgAChE) with ki values at least 10- to 600-fold higher than that of propoxur, a compound that does not kill Akron mosquitoes at the highest concentration tested. On average, inactivation of WT AgAChE by dimethylcarboxamides 14 was 10-20 fold faster than that of the corresponding isoxazol-3-yl dimethylcarbamates 15. X-ray crystallography of dimethylcarboxamide 14d provided insight into that reactivity, a finding that may explain the inhibitory power of structurally-related inhibitors of hormone-sensitive lipase. Finally, human/An. gambiae AChE inhibition selectivities of these compounds were low, suggesting the need for additional structural modification.

Ultrastructural and Morphalogical Changes of Mouse Ovarian Tissues Following Direct Cover Vitrification with Different Cryoprotectants.

BACKGROUND: Cryopreservation of mammalian ovaries has been reported with different levels of success. Cryopreservation of ovarian tissue may be a potential alternative for treatment of infertility and many attempts have been done to improve the efficiency of ovarian cryopreservation. The objective of the present study was to compare the direct cover vitrification (DCV) with ethylene glycol (EG), dimethyl sulfoxide (DMSO) and EG plus DMSO. METHODS: Eighty five mice were sacrificed by cervical dislocation and their ovaries were cryopreserved in the presence of 5% EG or DMSO alone or as mixture, 10% EG or DMSO alone or as mixture and a group with ascending concentrations of cryoprotectants. After toxicity testing and vitrification warming, the ovaries were fixed for histological and ultrastructural studies. In addition, the viability of mechanically isolated follicles was studied by trypan blue staining. All data were compared by ANOVA (p<0.05). RESULTS: Ovarian tissues frozen in EG plus DMSO in ascending concentrations retained a higher percentage of morphologically normal and or viable follicles than tissues frozen in 10 M EG plus DMSO or in either concentration of EG and DMSO alone (p<0.001). Ultrastructural analysis of ovarian tissues frozen in ascending concentrations of EG plus DMSO showed that these follicles were well preserved and it was very similar to the control group. CONCLUSION: Cryopreservation of ovarian tissue in EG plus DMSO is the most effective method for preserving the structural integrity of follicles within the ovary.