(2-amino-phenyl)-amides of omega-substituted alkanoic acids as new histone deacetylase inhibitors

A variety of omega-substituted alkanoic acid (2-amino-phenyl)-amides were designed and synthesized. These compounds were shown to inhibit recombinant human histone deacetylases (HDACs) with IC(50) values in the low micromolar range and induce hyperacetylation of histones in whole cells. They induced expression of p21WAF1/Cip1 and caused cell-cycle arrest in human cancer cells. Compounds in this class showed efficacy in human tumor xenograft models.     

Increased reliability of selective PCR by using additionally mutated primers and a commercialTaq DNA polymerase enhancer

A reliable selective PCR procedure that combines the use of additionally mutated primers with the specificity-enhancing properties of a commercial preparation (Perfect Match, Stratagene) is described. The human immunodeficiency virus type 1pol gene point mutations known to confer in vitro resistance to azidothymidine were examined as a model for optimization of the assay. The usual strategy of deliberately introducing an additional mismatch 1 residue from the 3′ end in the wild-type and mutant primers did not allow reproducible discrimination between wild-type and mutant target sequences. Addition of minimal amounts of Perfect Match to the same PCR mixtures resulted in a significantly enlarged range of selective annealing temperatures, providing a valuable and cost-effective means for reliable detection of known mutations by selectivePCR.     

Effects of leptin on in vitro maturation, fertilization and embryonic cleavage after ICSI and early developmental expression of leptin (Ob) and leptin receptor (ObR) proteins in the horse

Background  

The identification of the adipocyte-derived obesity gene product, leptin (Ob), and subsequently its association with reproduction in rodents and humans led to speculations that leptin may be involved in the regulation of oocyte and preimplantation embryo development. In mice and pigs, in vitro leptin addition significantly increased meiotic resumption and promoted preimplantation embryo development in a dose-dependent manner. This study was conducted to determine whether leptin supplementation during in vitro maturation (IVM) to horse oocytes could have effects on their developmental capacity after fertilization by IntraCytoplasmic Sperm Injection (ICSI).     

Extracellular <Emphasis Type="Italic">Paracoccidioides brasiliensis</Emphasis> phospholipase B involvement in alveolar macrophage interaction

Background  

Phospholipase B (PLB) has been reported to be one of the virulence factors for human pathogenic fungi and has also been described as necessary for the early events in infection. Based on these data, we investigated the role of PLB in virulence and modulation of the alveolar pulmonary immune response during infection using an in-vitro model of host-pathogen interaction, i.e. Paracoccidioides brasiliensis yeast cells infecting alveolar macrophage (MH-S) cells.     

Phenotypic Features of Circulating Leukocytes from Non-human Primates Naturally Infected with Trypanosoma cruzi Resemble the Major Immunological Findings Observed in Human Chagas Disease

Background

Cynomolgus macaques (Macaca fascicularis) represent a feasible model for research on Chagas disease since natural T. cruzi infection in these primates leads to clinical outcomes similar to those observed in humans. However, it is still unknown whether these clinical similarities are accompanied by equivalent immunological characteristics in the two species. We have performed a detailed immunophenotypic analysis of circulating leukocytes together with systems biology approaches from 15 cynomolgus macaques naturally infected with T. cruzi (CH) presenting the chronic phase of Chagas disease to identify biomarkers that might be useful for clinical investigations.

Methods and Findings

Our data established that CH displayed increased expression of CD32⁺ and CD56⁺ in monocytes and enhanced frequency of NK Granzyme A⁺ cells as compared to non-infected controls (NI). Moreover, higher expression of CD54 and HLA-DR by T-cells, especially within the CD8⁺ subset, was the hallmark of CH. A high level of expression of Granzyme A and Perforin underscored the enhanced cytotoxicity-linked pattern of CD8⁺ T-lymphocytes from CH. Increased frequency of B-cells with up-regulated expression of Fc-γRII was also observed in CH. Complex and imbricate biomarker networks demonstrated that CH showed a shift towards cross-talk among cells of the adaptive immune system. Systems biology analysis further established monocytes and NK-cell phenotypes and the T-cell activation status, along with the Granzyme A expression by CD8⁺ T-cells, as the most reliable biomarkers of potential use for clinical applications.

Conclusions

Altogether, these findings demonstrated that the similarities in phenotypic features of circulating leukocytes observed in cynomolgus macaques and humans infected with T. cruzi further supports the use of these monkeys in preclinical toxicology and pharmacology studies applied to development and testing of new drugs for Chagas disease.     

Identification of a streptococcal octapeptide motif involved in acute rheumatic fever

Acute rheumatic fever is a serious autoimmune sequela of pharyngitis caused by certain group A streptococci. One mechanism applied by streptococcal strains capable of causing acute rheumatic fever is formation of an autoantigenic complex with human collagen IV. In some geographic regions with a high incidence of acute rheumatic fever pharyngeal carriage of group C and group G streptococci prevails. Examination of such strains revealed the presence of M-like surface proteins that bind human collagen. Using a peptide array and recombinant proteins with targeted amino acid substitutions, we could demonstrate that formation of collagen complexes during streptococcal infections depends on an octapeptide motif, which is present in collagen binding M and M-like proteins of different beta-hemolytic streptococcal species. Mice immunized with streptococcal proteins that contain the collagen binding octapeptide motif developed high serum titers of anti-collagen antibodies. In sera of rheumatic fever patients such a collagen autoimmune response was accompanied by specific reactivity against the collagen-binding proteins, linking the observed effect to clinical cases. Taken together, the data demonstrate that the identified octapeptide motif through its action on collagen plays a crucial role in the pathogenesis of rheumatic fever. Eradication of streptococci that express proteins with the collagen binding motif appears advisable for controlling rheumatic fever.     

The effects of UV radiation on the visual system of the crab Neohelice granulata: a protective role of melatonin

The first and main target-structure of ultraviolet (UV) radiation in animals is the body surface, including the skin and eyes. Here, we investigated cell damage in the visual system of the crab Neohelice granulata acclimated to constant light and exposed to UVA or UVB at 12:00 h for 30 min. The reactive oxygen species (ROS) production, antioxidant capacity against peroxyl radicals (ACAP), lipid peroxidation (LPO) damage, catalase (CAT) activity, and the melatonin immunohistochemical reactivity in the eyestalks were evaluated. The animals that received melatonin and were exposed to UVA and UVB radiation showed a decreased ROS concentration (p < 0.05).The ACAP test showed a decrease (p < 0.05) in their values when the animals received 2 pmol/crab of melatonin (physiological dose) before the exposure to UVA radiation. The animals exposed to UVB radiation after receiving the same dose of melatonin showed an increase (p < 0.05) in the ACAP test compared with the animals exposed to UVB radiation after receiving only crab physiological saline. The CAT activity increased (p < 0.05) in the animals that received melatonin and were exposed to UVA and UVB radiation. Animals exposed to UVA and UVB displayed an increase (p < 0.05) in the LPO levels, whereas animals treated with melatonin showed lower (p < 0.05) LPO levels when irradiated. The results indicate that the specific oxidative parameters altered by UV radiation can be modulated by a physiological dose of melatonin. Moreover, the melatonin regularly produced by virtually all eyestalk cells suggests that it may function to modulate the noxious effects of radiation, at least in the crab N. granulata.     

Multifunctional liposomes for nasal delivery of the anti-Alzheimer drug tacrine hydrochloride

The purpose of this study was the development of multifunctional liposomes for nasal administration of tacrine hydrochloride. Liposomes were prepared using traditional excipients (cholesterol and phosphatidylcholine), partly enriched with α-tocopherol and/or Omega3 fatty acids. This approach was chosen in order to obtain at the same time two positive results: an enhanced drug permeation through nasal mucosa and a concomitant neuroprotective effect. Several liposome formulations were prepared using the Reverse Phase Evaporation technique followed by membrane filter extrusion. In particular, liposome capacity to enhance drug permeation was evaluated by means of membrane permeation and cellular uptake studies. Furthermore, liposome effect on neuronal viability and intracellular ROS production was evaluated as well as their cytoprotective effect against oxidative stress. All liposome formulations showed a mean diameter in the range of 175?nm to 219?nm with polydispersity index lower than 0.22, a lightly negative zeta potential and excellent encapsulation efficiency. Moreover, along with good mucoadhesive properties, multifunctional liposomes showed a markedly increase in tacrine permeability, which can be related to liposome fusion with cellular membrane, a hypothesis, which was also supported by cellular uptake studies. Finally, the addition of α-tocopherol without Omega3 fatty acids, was found to increase the neuroprotective activity and antioxidant properties of liposomes.     

A pilot randomised trial of induced blood-stage Plasmodium falciparum infections in healthy volunteers for testing efficacy of new antimalarial drugs

Background

Critical to the development of new drugs for treatment of malaria is the capacity to safely evaluate their activity in human subjects. The approach that has been most commonly used is testing in subjects with natural malaria infection, a methodology that may expose symptomatic subjects to the risk of ineffective treatment. Here we describe the development and pilot testing of a system to undertake experimental infection using blood stage Plasmodium falciparum parasites (BSP). The objectives of the study were to assess the feasibility and safety of induced BSP infection as a method for assessment of efficacy of new drug candidates for the treatment of P. falciparum infection.

Methods and Findings

A prospective, unblinded, Phase IIa trial was undertaken in 19 healthy, malaria-naïve, male adult volunteers who were infected with BSP and followed with careful clinical and laboratory observation, including a sensitive, quantitative malaria PCR assay. Volunteers were randomly allocated to treatment with either of two licensed antimalarial drug combinations, artemether–lumefantrine (A/L) or atovaquone-proguanil (A/P). In the first cohort (n = 6) where volunteers received ∼360 BSP, none reached the target parasitemia of 1,000 before the day designated for antimalarial treatment (day 6). In the second and third cohorts, 13 volunteers received 1,800 BSP, with all reaching the target parasitemia before receiving treatment (A/L, n = 6; A/P, n = 7) The study demonstrated safety in the 19 volunteers tested, and a significant difference in the clearance kinetics of parasitemia between the drugs in the 13 evaluable subjects, with mean parasite reduction ratios of 759 for A/L and 17 for A/P (95% CI 120–4786 and 7–40 respectively; p<0.01).

Conclusions

This system offers a flexible and safe approach to testing the in vivo activity of novel antimalarials.

Trial Registration:

ClinicalTrials.gov {"type":"clinical-trial","attrs":{"text":"NCT01055002","term_id":"NCT01055002"}}NCT01055002