An amphiphilic graft copolymer-based nanoparticle platform for reduction-responsive anticancer and antimalarial drug delivery.

Medical applications of anticancer and antimalarial drugs often suffer from low aqueous solubility, high systemic toxicity, and metabolic instability. Smart nanocarrier-based drug delivery systems provide means of solving these problems at once. Herein, we present such a smart nanoparticle platform based on self-assembled, reduction-responsive amphiphilic graft copolymers, which were successfully synthesized through thiol-disulfide exchange reaction between thiolated hydrophilic block and pyridyl disulfide functionalized hydrophobic block. These amphiphilic graft copolymers self-assembled into nanoparticles with mean diameters of about 30-50 nm and readily incorporated hydrophobic guest molecules. Fluorescence correlation spectroscopy (FCS) was used to study nanoparticle stability and triggered release of a model compound in detail. Long-term colloidal stability and model compound retention within the nanoparticles was found when analyzed in cell media at body temperature. In contrast, rapid, complete reduction-triggered disassembly and model compound release was achieved within a physiological reducing environment. The synthesized copolymers revealed no intrinsic cellular toxicity up to 1 mg mL(-1). Drug-loaded reduction-sensitive nanoparticles delivered a hydrophobic model anticancer drug (doxorubicin, DOX) to cancer cells (HeLa cells) and an experimental, metabolically unstable antimalarial drug (the serine hydroxymethyltransferase (SHMT) inhibitor (±)-1) to Plasmodium falciparum-infected red blood cells (iRBCs), with higher efficacy compared to similar, non-sensitive drug-loaded nanoparticles. These responsive copolymer-based nanoparticles represent a promising candidate as smart nanocarrier platform for various drugs to be applied to different diseases, due to the biocompatibility and biodegradability of the hydrophobic block, and the protein-repellent hydrophilic block.

Evaluation of Xpert® MTB/RIF and Ustar EasyNAT™ TB IAD for diagnosis of tuberculous lymphadenitis of children in Tanzania: a prospective descriptive study.

BACKGROUND: Fine needle aspiration biopsy has become a standard approach for diagnosis of peripheral tuberculous lymphadenitis. The aim of this study was to compare the performance of Xpert MTB/RIF and Ustar EasyNAT TB IAD nucleic acid amplification assays, against acid-fast bacilli microscopy, cytology and mycobacterial culture for the diagnosis of TB lymphadenitis in children from a TB-endemic setting in Tanzania. METHODS: Children of 8 weeks to 16 years of age, suspected of having TB lymphadenitis, were recruited at a district hospital in Tanzania. Fine needle aspirates of lymph nodes were analysed using acid-fast bacilli microscopy, liquid TB culture, cytology, Xpert MTB/RIF and EasyNAT. Latent class analysis and comparison against a composite reference standard comprising "culture and/or cytology" was done, to assess the performance of Xpert MTB/RIF and EasyNAT for the diagnosis of TB lymphadenitis. RESULTS: Seventy-nine children were recruited; 4 were excluded from analysis. Against a composite reference standard of culture and/or cytology, Xpert MTB/RIF and EasyNAT had a sensitivity and specificity of 58 % and 93 %; and 19 % and 100 % respectively. Relative to latent class definitions, cytology had a sensitivity of 100 % and specificity of 94.7 %. CONCLUSIONS: Combining clinical assessment, cytology and Xpert MTB/RIF may allow for a rapid and accurate diagnosis of childhood TB lymphadenitis. Larger diagnostic evaluation studies are recommended to validate these findings and on Xpert MTB/RIF to assess its use as a solitary initial test for TB lymphadenitis in children.

Spiroindolones, a potent compound class for the treatment of malaria.

Recent reports of increased tolerance to artemisinin derivatives--the most recently adopted class of antimalarials--have prompted a need for new treatments. The spirotetrahydro-beta-carbolines, or spiroindolones, are potent drugs that kill the blood stages of Plasmodium falciparum and Plasmodium vivax clinical isolates at low nanomolar concentration. Spiroindolones rapidly inhibit protein synthesis in P. falciparum, an effect that is ablated in parasites bearing nonsynonymous mutations in the gene encoding the P-type cation-transporter ATPase4 (PfATP4). The optimized spiroindolone NITD609 shows pharmacokinetic properties compatible with once-daily oral dosing and has single-dose efficacy in a rodent malaria model.

Differential var gene expression in children with malaria and antidromic effects on host gene expression.

Among 62 children with mild malaria, cerebral malaria, or severe malarial anemia, we analyzed the transcription of different var gene types. There was no difference in parasitemia level or body temperature between groups. However, a significantly different expression pattern was observed in children with cerebral malaria, compared with that in patients in the other 2 groups: children with cerebral malaria had lower expression of the upsA subtype but higher expression of the upsB and upsC subtypes. Furthermore, expression of human genes responsive to tumor necrosis factor and hypoxia correlated with distinct ups types.

Cyclic nucleotide-specific phosphodiesterases of Plasmodium falciparum: PfPDEalpha, a non-essential cGMP-specific PDE that is an integral membrane protein.

Cyclic nucleotide-specific phosphodiesterases (PDEs) have come into focus as interesting potential targets for PDE inhibitor-based anti-parasitic drugs. Genomes of the various agents of human malaria, most notably Plasmodium falciparum, all contain four genes for class 1 PDEs. The catalytic domains of these enzymes are closely related to those of the 11 human PDE families. This presents the possibility that the available vast expertise in developing drugs against human PDEs might now also be applied to developing compounds that are active against malarial PDEs. The current study identifies four Plasmodium genes that code for PfPDEalpha, PfPDEbeta, PfPDEgamma and PfPDEdelta, respectively. It further demonstrates that the PfPDEalpha polypeptide exists in two versions (PfPDEalphaA and PfPDEalphaB) that are generated by alternative splicing of the primary transcript. All malarial PDEs contain several transmembrane helices in their N-terminal regions, indicating that they are integral membrane proteins. In agreement with this prediction, essentially all PDE activity is associated with the cell membranes. PfPDEalpha was characterized as a cGMP-specific PDE that is not sensitive to a number of standard PDE inhibitors. Genetic ablation of the PfPDE1 gene produced no major phenotype in erythrocyte cultures.

Strain-specific humoral response to a polymorphic malaria vaccine.

The 3D7 form of the merozoite surface protein 2 (MSP2) of Plasmodium falciparum was one of three subunits of the malaria vaccine Combination B that were tested in a phase I/IIb double-blind randomized placebo-controlled trial, which was undertaken with 120 Papua New Guinean children of 5 to 9 years of age. Because only one variant of the highly polymorphic MSP2 was used for vaccination, we examined whether the elicited response was directed against conserved or strain-specific epitopes. Postvaccination (week 12) titers of antibody against recombinantly expressed individual domains of MSP2 were measured by enzyme-linked immunosorbent assay and compared to baseline values. We found that vaccination with the 3D7 form of MSP2 induced a significant strain-specific humoral response directed against the repetitive and semiconserved family-specific part. The conserved N- and C-terminal domains were not immunogenic. Titers of antibody against the alternate FC27 family-specific domain showed a tendency to increase in vaccinated children, but there was no increase in antibodies against FC27-type 32-mer repeats. These results indicate that vaccination with one MSP2 variant mainly induced a strain-specific response, which can explain the selective effect of vaccination with combination B on the genotypes of breakthrough parasites. These findings support the inclusion of both family-specific domains (3D7 and FC27) in an improved vaccine formulation.

Safety and immunogenicity of a three-component blood-stage malaria vaccine (MSP1, MSP2, RESA) against Plasmodium falciparum in Papua New Guinean children.

Combination B is a malaria vaccine that comprises recombinant Plasmodium falciparum (P. falciparum) blood-stage proteins MSP1, MSP2 and RESA, formulated with the adjuvant Montanide ISA 720. A phase I-IIb double-blind randomised placebo-controlled trial was undertaken in 120 children aged 5-9 years. Subjects were randomised in four groups: (i) No sulphadoxine-pyrimethamine (SP)+vaccine, (ii) No SP+placebo, (iii) SP+vaccine, (iv) SP+placebo. 15 microg of each protein were given in the thigh, at both first and second injection (4 weeks apart). The placebo was adjuvant emulsified with saline. No serious or severe AEs occurred. Moderate AEs were seen in 3% of the vaccine and 3% of the placebo recipients after first injection and in 12 and 10% after second injection. The vaccine induced significant antibody responses to all three antigens but triggered an IFN-gamma response to MSP1 only. At Week 12, the IFN-gamma response to MSP1 was substantially higher in the vaccine group where No SP had been given. Combination B proved to be safe and immunogenic in children aged 5-9 years. Vaccine immunogenicity was neither impaired by circulating parasites nor increased after pre-treatment with SP and pre-treatment is not advisable in future trials of malaria vaccines, at least for those including blood-stage antigens.