Is there still room to explore cyclodextrin glycosyltransferase-producers in Brazilian biodiversity?

In the present work, different Brazilian biomes aiming to identify and select cyclodextrin glycosyltransferase-producer bacteria are explored. This enzyme is responsible for converting starch to cyclodextrin, which are interesting molecules to carry other substances of economic interest applied by textile, pharmaceutical, food, and other industries. Based on the enzymatic index, 12 bacteria were selected and evaluated, considering their capacity to produce the enzyme in culture media containing different starch sources. It was observed that the highest yields were presented by the bacteria when grown in cornstarch. These bacteria were also characterized by sequencing of the 16S rRNA region and were classified as Bacillus, Paenibacillus, Gracilibacillus and Solibacillus.

QSAR-Driven Design and Discovery of Novel Compounds With Antiplasmodial and Transmission Blocking Activities.

Malaria is a life-threatening infectious disease caused by parasites of the genus Plasmodium, affecting more than 200 million people worldwide every year and leading to about a half million deaths. Malaria parasites of humans have evolved resistance to all current antimalarial drugs, urging for the discovery of new effective compounds. Given that the inhibition of deoxyuridine triphosphatase of Plasmodium falciparum (PfdUTPase) induces wrong insertions in plasmodial DNA and consequently leading the parasite to death, this enzyme is considered an attractive antimalarial drug target. Using a combi-QSAR (quantitative structure-activity relationship) approach followed by virtual screening and in vitro experimental evaluation, we report herein the discovery of novel chemical scaffolds with in vitro potency against asexual blood stages of both P. falciparum multidrug-resistant and sensitive strains and against sporogonic development of P. berghei. We developed 2D- and 3D-QSAR models using a series of nucleosides reported in the literature as PfdUTPase inhibitors. The best models were combined in a consensus approach and used for virtual screening of the ChemBridge database, leading to the identification of five new virtual PfdUTPase inhibitors. Further in vitro testing on P. falciparum multidrug-resistant (W2) and sensitive (3D7) parasites showed that compounds LabMol-144 and LabMol-146 demonstrated fair activity against both strains and presented good selectivity versus mammalian cells. In addition, LabMol-144 showed good in vitro inhibition of P. berghei ookinete formation, demonstrating that hit-to-lead optimization based on this compound may also lead to new antimalarials with transmission blocking activity.

Chalcone Derivatives: Promising Starting Points for Drug Design.

Medicinal chemists continue to be fascinated by chalcone derivatives because of their simple chemistry, ease of hydrogen atom manipulation, straightforward synthesis, and a variety of promising biological activities. However, chalcones have still not garnered deserved attention, especially considering their high potential as chemical sources for designing and developing new effective drugs. In this review, we summarize current methodological developments towards the design and synthesis of new chalcone derivatives and state-of-the-art medicinal chemistry strategies (bioisosterism, molecular hybridization, and pro-drug design). We also highlight the applicability of computer-assisted drug design approaches to chalcones and address how this may contribute to optimizing research outputs and lead to more successful and cost-effective drug discovery endeavors. Lastly, we present successful examples of the use of chalcones and suggest possible solutions to existing limitations.

In silico repositioning-chemogenomics strategy identifies new drugs with potential activity against multiple life stages of Schistosoma mansoni.

Morbidity and mortality caused by schistosomiasis are serious public health problems in developing countries. Because praziquantel is the only drug in therapeutic use, the risk of drug resistance is a concern. In the search for new schistosomicidal drugs, we performed a target-based chemogenomics screen of a dataset of 2,114 proteins to identify drugs that are approved for clinical use in humans that may be active against multiple life stages of Schistosoma mansoni. Each of these proteins was treated as a potential drug target, and its amino acid sequence was used to interrogate three databases: Therapeutic Target Database (TTD), DrugBank and STITCH. Predicted drug-target interactions were refined using a combination of approaches, including pairwise alignment, conservation state of functional regions and chemical space analysis. To validate our strategy, several drugs previously shown to be active against Schistosoma species were correctly predicted, such as clonazepam, auranofin, nifedipine, and artesunate. We were also able to identify 115 drugs that have not yet been experimentally tested against schistosomes and that require further assessment. Some examples are aprindine, gentamicin, clotrimazole, tetrabenazine, griseofulvin, and cinnarizine. In conclusion, we have developed a systematic and focused computer-aided approach to propose approved drugs that may warrant testing and/or serve as lead compounds for the design of new drugs against schistosomes.

Natural products as leads in schistosome drug discovery.

Schistosomiasis is a neglected parasitic tropical disease that claims around 200,000 human lives every year. Praziquantel (PZQ), the only drug recommended by the World Health Organization for the treatment and control of human schistosomiasis, is now facing the threat of drug resistance, indicating the urgent need for new effective compounds to treat this disease. Therefore, globally, there is renewed interest in natural products (NPs) as a starting point for drug discovery and development for schistosomiasis. Recent advances in genomics, proteomics, bioinformatics, and cheminformatics have brought about unprecedented opportunities for the rapid and more cost-effective discovery of new bioactive compounds against neglected tropical diseases. This review highlights the main contributions that NP drug discovery and development have made in the treatment of schistosomiasis and it discusses how integration with virtual screening (VS) strategies may contribute to accelerating the development of new schistosomidal leads, especially through the identification of unexplored, biologically active chemical scaffolds and structural optimization of NPs with previously established activity.

Failure to detect Plasmodium vivax in West and Central Africa by PCR species typing.

BACKGROUND: Plasmodium vivax is estimated to affect 75 million people annually. It is reportedly absent, however, from west and central Africa due to the high prevalence of the Duffy negative phenotype in the indigenous populations. Despite this, non-African travellers consistently return to their own countries with P. vivax malaria after visiting this region. An attempt was made, therefore, to detect the presence of P. vivax parasites in blood samples collected from the indigenous populations of west and central Africa. METHODS: Parasite species typing (for all four human malaria parasites) was carried out by PCR on 2,588 blood samples collected from individuals from nine African malaria-endemic countries. RESULTS: Most infections (98.5%) were Plasmodium falciparum, Plasmodium malariae was identified in 8.5% of all infections, and Plasmodium ovale in 3.9%. The prevalence of both parasites varied greatly by country. Only one case of P. vivax was detected from Sao Tome, an island off the west coast of Africa, confirming the scarcity of this parasite in Africa. CONCLUSION: The prevalence of P. vivax in local populations in sub-Saharan Africa is very low, despite the frequent identification of this parasite in non-African travellers.

Real-time quantitative PCR with SYBR Green I detection for estimating copy numbers of nine drug resistance candidate genes in Plasmodium falciparum.

BACKGROUND: Evaluating copy numbers of given genes in Plasmodium falciparum parasites is of major importance for laboratory-based studies or epidemiological surveys. For instance, pfmdr1 gene amplification has been associated with resistance to quinine derivatives and several genes involved in anti-oxidant defence may play an important role in resistance to antimalarial drugs, although their potential involvement has been overlooked. METHODS: The DeltaDeltaCt method of relative quantification using real-time quantitative PCR with SYBR Green I detection was adapted and optimized to estimate copy numbers of three genes previously indicated as putative candidates of resistance to quinolines and artemisinin derivatives: pfmdr1, pfatp6 (SERCA) and pftctp, and in six further genes involved in oxidative stress responses. RESULTS: Using carefully designed specific RT-qPCR oligonucleotides, the methods were optimized for each gene and validated by the accurate measure of previously known number of copies of the pfmdr1 gene in the laboratory reference strains P. falciparum 3D7 and Dd2. Subsequently, Standard Operating Procedures (SOPs) were developed to the remaining genes under study and successfully applied to DNA obtained from dried filter blood spots of field isolates of P. falciparum collected in São Tomé & Principe, West Africa. CONCLUSION: The SOPs reported here may be used as a high throughput tool to investigate the role of these drug resistance gene candidates in laboratory studies or large scale epidemiological surveys.

An AFLP-based genetic linkage map of Plasmodium chabaudi chabaudi.

BACKGROUND: Plasmodium chabaudi chabaudi can be considered as a rodent model of human malaria parasites in the genetic analysis of important characters such as drug resistance and immunity. Despite the availability of some genome sequence data, an extensive genetic linkage map is needed for mapping the genes involved in certain traits. METHODS: The inheritance of 672 Amplified Fragment Length Polymorphism (AFLP) markers from two parental clones (AS and AJ) of P. c. chabaudi was determined in 28 independent recombinant progeny clones. These, AFLP markers and 42 previously mapped Restriction Fragment Length Polymorphism (RFLP) markers (used as chromosomal anchors) were organized into linkage groups using Map Manager software. RESULTS: 614 AFLP markers formed linkage groups assigned to 10 of 14 chromosomes, and 12 other linkage groups not assigned to known chromosomes. The genetic length of the genome was estimated to be about 1676 centiMorgans (cM). The mean map unit size was estimated to be 13.7 kb/cM. This was slightly less then previous estimates for the human malaria parasite, Plasmodium falciparum CONCLUSION: The P. c. chabaudi genetic linkage map presented here is the most extensive and highly resolved so far available for this species. It can be used in conjunction with the genome databases of P. c chabaudi, P. falciparum and Plasmodium yoelii to identify genes underlying important phenotypes such as drug resistance and strain-specific immunity.

Chloroquine resistance in Plasmodium chabaudi: are chloroquine-resistance transporter (crt) and multi-drug resistance (mdr1) orthologues involved?

We have identified in the rodent malaria parasite Plasmodium chabaudi orthologues of two Plasmodium falciparum genes, pfcrt and pfmdr1 which have been implicated as determinants of chloroquine resistance in the latter species. The sequences of the P. chabaudi genes, denoted, respectively, pccg10 and pcmdr1, were first determined in the chloroquine-sensitive clone AS, and found to be highly similar to those of P. falciparum. For pccg10, there was a nucleotide sequence identity of 68.6% and amino acid sequence identity of 75.1% within the predicted coding region. For pcmdr1, the sequence identities were 75.0% (nucleotide) and 78.1% (amino acid). The sequences of the genes were then determined in three P. chabaudi clones selected from clone AS which possessed three different levels of resistance to chloroquine. The sequences of both genes in all mutants were found to be identical to those of the sensitive AS from which they had been derived. Polymorphic sites were found in both genes between the AS clones and a genetically unrelated sensitive clone AJ. Analysis of genetic crosses between AJ and resistant AS clones showed no linkage between inherited parental alleles of pccrt and pcmdr1 and drug responses of the cloned progeny. This showed that neither of these genes, nor genes closely linked to them, were determinants of the chloroquine resistance in the P. chabaudi mutants.

Genetics of mefloquine resistance in the rodent malaria parasite Plasmodium chabaudi.

The genetic determinants of resistance to mefloquine in malaria parasites are unclear. Some studies have implied that amplification of, or mutations in, the multidrug resistance gene pfmdr1 in Plasmodium falciparum may be involved. Using the rodent malaria model Plasmodium chabaudi, we investigated the role of the orthologue of this gene, pcmdr1, in a stable mefloquine-resistant mutant, AS(15MF/3), selected from a sensitive clone. pcmdr1 exists as a single copy gene on chromosome 12 of the sensitive clone. In AS(15MF/3), the gene was found to have undergone duplication, with one copy translocating to chromosome 4. mRNA levels of pcmdr1 were higher in the mutant than in the parent sensitive clone. A partial genetic map of the translocation showed that other genes in addition to pcmdr1 had been cotranslocated. The sequences of both copies of pcmdr1 of AS(15MF/3) were identical to that of the parent sensitive clone. A cross was made between AS(15MF/3) and an unrelated mefloquine-sensitive clone, AJ. Phenotypic and molecular analysis of progeny clones showed that duplication and overexpression of the pcmdr1 gene was an important determinant of resistance. However, not all mefloquine-resistant progeny contained the duplicated gene, showing that at least one other gene was involved in resistance.