Discovery of AZD3514, a small-molecule androgen receptor downregulator for treatment of advanced prostate cancer

Removal of the basic piperazine nitrogen atom, introduction of a solubilising end group and partial reduction of the triazolopyridazine moiety in the previously-described lead androgen receptor downregulator 6-[4-(4-cyanobenzyl)piperazin-1-yl]-3-(trifluoromethyl)[1,2,4]triazolo[4,3-b]pyridazine (1) addressed hERG and physical property issues, and led to clinical candidate 6-(4-{4-[2-(4-acetylpiperazin-1-yl)ethoxy]phenyl}piperidin-1-yl)-3-(trifluoromethyl)-7,8-dihydro[1,2,4]triazolo[4,3-b]pyridazine (12), designated AZD3514, that is being evaluated in a Phase I clinical trial in patients with castrate-resistant prostate cancer.     

Protoplast fusion for production of tetraploids and triploids: applications for scion and rootstock breeding in citrus

Protoplast fusion technology has been utilized in many crops to generate allotetraploid somatic hybrids, and sometimes autotetraploids as a byproduct of the process. A brief history of this technology development is provided, along with a simple protocol developed for citrus, which can be easily adapted to other plants. Protoplast fusion has become a significant tool in ploidy manipulation that can be applied in various cultivar improvement schemes. In rare cases, a new somatic hybrid may have direct utility as an improved cultivar; however, the most important application of somatic hybridization is the building of novel germplasm as a source of elite breeding parents for various types of conventional crosses for both scion and rootstock improvement. Somatic hybridization is generating superior allotetraploid breeding parents for use in interploid crosses to generate seedless triploids. Seedlessness is a primary breeding objective for new fresh fruit citrus varieties, and several thousand triploid hybrids have been produced using somatic hybrids as the tetraploid parent. Protoplast fusion is also being utilized to produce somatic hybrids that combine complementary diploid rootstocks, which have shown good potential for tree size control. Tree size control has gained importance as a means of reducing harvesting costs, maximizing the efficiency of modern cold protection methodology, and facilitating the adaptation of new fruit production systems. Successful somatic hybridization in citrus rootstock improvement has enabled rootstock breeding at the tetraploid level via sexual hybridization, which can yield maximum genetic diversity in zygotic progeny upon which to impose selection for the many traits required in improved rootstock cultivars, including disease and insect resistance, broad adaptation, tree size control, and the ability to consistently produce high yields of quality fruit. Recent progress and successful examples of these applications are discussed. Finally, a discussion of the genetic potential of somatic hybrids as breeding parents, including meiotic behavior and inheritance is provided.     

Small-molecule androgen receptor downregulators as an approach to treatment of advanced prostate cancer

Chemical starting points were investigated for downregulation of the androgen receptor as an approach to treatment of advanced prostate cancer. Although prototypic steroidal downregulators such as 6a designed for intramuscular administration showed insufficient cellular potency, a medicinal chemistry program derived from a novel androgen receptor ligand 8a led to 6-[4-(4-cyanobenzyl)piperazin-1-yl]-3-(trifluoromethyl)[1,2,4]triazolo[4,3-b]pyridazine (10b), for which high plasma levels following oral administration in a preclinical model compensate for moderate cellular potency.     

Alkaline subcritical water gasification of dairy industry waste (Whey)

The near-critical water gasification of dairy industry waste in the form of Whey, a product composed of mixtures of carbohydrates (mainly lactose) and amino acids such as glycine and glutamic acid, has been studied. The gasification process involved partial oxidation with hydrogen peroxide in the presence of NaOH. The reactions were studied over the temperature range from 300°C to 390°C, corresponding pressures of 9.5-24.5 MPa and reaction times from 0 min to 120 min. Hydrogen production was affected by the presence of NaOH, the concentration of H(2)O(2), temperature, reaction time and feed concentration. Up to 40% of the theoretical hydrogen gas production was achieved at 390°C. Over 80% of the Whey nitrogen content was found as ammonia, mainly in the liquid effluent.     

Intracellular protozoan parasites of humans: the role of molecular chaperones in development and pathogenesis

Certain kinetoplastid (Leishmania spp. and Tryapnosoma cruzi) and apicomplexan parasites (Plasmodium falciparum and Toxoplasma gondii) are capable of invading human cells as part of their pathology. These parasites appear to have evolved a relatively expanded or diverse complement of genes encoding molecular chaperones. The gene families encoding heat shock protein 90 (Hsp90) and heat shock protein 70 (Hsp70) chaperones show significant expansion and diversity (especially for Leishmania spp. and T. cruzi), and in particular the Hsp40 family appears to be an extreme example of phylogenetic radiation. In general, Hsp40 proteins act as co-chaperones of Hsp70 chaperones, forming protein folding pathways that integrate with Hsp90 to ensure proteostasis in the cell. It is tempting to speculate that the diverse environmental insults that these parasites endure have resulted in the evolutionary selection of a diverse and expanded chaperone network. Hsp90 is involved in development and growth of all of these intracellular parasites, and so far represents the strongest candidate as a target for chemotherapeutic interventions. While there have been some excellent studies on the molecular and cell biology of Hsp70 proteins, relatively little is known about the biological function of Hsp70-Hsp40 interactions in these intracellular parasites. This review focuses on intracellular protozoan parasites of humans, and provides a critique of the role of heat shock proteins in development and pathogenesis, especially the molecular chaperones Hsp90, Hsp70 and Hsp40.     

BRD4 Short Isoform Interacts with RRP1B,SIPA1 and Components of the LINC Complex at the Inner Face of the Nuclear Membrane

Recent studies suggest that BET inhibitors are effective anti-cancer therapeutics. Here we show that BET inhibitors are effective against murine primary mammary tumors, but not pulmonary metastases. BRD4, a target of BET inhibitors, encodes two isoforms with opposite effects on tumor progression. To gain insights into why BET inhibition was ineffective against metastases the pro-metastatic short isoform of BRD4 was characterized using mass spectrometry and cellular fractionation. Our data show that the pro-metastatic short isoform interacts with the LINC complex and the metastasis-associated proteins RRP1B and SIPA1 at the inner face of the nuclear membrane. Furthermore, histone binding arrays revealed that the short isoform has a broader acetylated histone binding pattern relative to the long isoform. These differential biochemical and nuclear localization properties revealed in our study provide novel insights into the opposing roles of BRD4 isoforms in metastatic breast cancer progression.     

Enhancement of the Anti-inflammatory Effect of Bromelain by Its Immobilization on Probiotic Spore of Bacillus cereus

The therapeutic application of bromelain is limited due to its sensitivity to operating conditions such as high acidity, gastric proteases in the stomach juice, chemicals, organic solvents and elevated temperature. We hypothesized that bromelain immobilized on probiotic bacterial spores would show enhanced therapeutic activity through possible synergistic or additive effects. In this study, the oedema inhibition potential of bromelain immobilized on probiotic Bacillus spores was compared to the free enzyme using the carrageenan paw oedema model with Wistar rats. In batch A rats (carrageenan-induced inflammation 30 min after receiving oral treatments), group 7 rats treated with a lower dose of spore-immobilized bromelain suspension showed the highest oedema inhibition, 89.20 ± 15.30%, while group 4 treated with a lower dose of free bromelain had oedema inhibition of 60.25 ± 13.00%. For batch B rats (carrageenan-induced inflammation after receiving oral treatment for three days), group 7 rats treated with a lower dose of spore-immobilized bromelain suspension showed higher inhibition percentage (81.94 ± 8.86) than group 4 treated with a lower dose of free bromelain (78.45 ± 4.46) after 24 h. Our results showed that used alone, the enzyme and the spores produced oedema inhibition and improved the motility of the rats. The spore-immobilized bromelain formulation performed approximately 0.9-fold better than the free bromelain and the free spores at the lower evaluated dose.

     

Characterization of the Nuclear Import Mechanism of the CCAAT-Regulatory Subunit Php4

Php4 is a nucleo-cytoplasmic shuttling protein that accumulates in the nucleus during iron deficiency. When present in the nucleus, Php4 associates with the CCAAT-binding protein complex and represses genes encoding iron-using proteins. Here, we show that nuclear import of Php4 is independent of the other subunits of the CCAAT-binding complex. Php4 nuclear import relies on two functionally independent nuclear localization sequences (NLSs) that are located between amino acid residues 171 to 174 (KRIR) and 234 to 240 (KSVKRVR). Specific substitutions of basic amino acid residues to alanines within these sequences are sufficient to abrogate nuclear targeting of Php4. The two NLSs are biologically redundant and are sufficient to target a heterologous reporter protein to the nucleus. Under low-iron conditions, a functional GFP-Php4 protein is only partly targeted to the nucleus in imp1Δ and sal3Δ mutant cells. We further found that cells expressing a temperature-sensitive mutation in cut15 exhibit increased cytosolic accumulation of Php4 at the nonpermissive temperature. Further analysis by pull-down experiments revealed that Php4 is a cargo of the karyopherins Imp1, Cut15 and Sal3. Collectively, these results indicate that Php4 can be bound by distinct karyopherins, connecting it into more than one nuclear import pathway.     

Co-infections of Malaria and Geohelminthiasis in Two Rural Communities of Nkassomo and Vian in the Mfou Health District,Cameroon

Background

Human co-infection with malaria and helmimths is ubiquitous throughout Africa. Nevertheless, its public health significance on malaria severity remains poorly understood.

Methodology/Principal Findings

To contribute to a better understanding of epidemiology and control of this co-infection in Cameroon, a cross-sectional study was carried out to assess the prevalence of concomitant intestinal geohelminthiasis and malaria, and to evaluate its association with malaria and anaemia in Nkassomo and Vian. Finger prick blood specimens from a total of 263 participants aged 1–95 years were collected for malaria microscopy, assessment of haemoglobin levels, and molecular identification of Plasmodium species by PCR. Fresh stool specimens were also collected for the identification and quantification of geohelminths by the Kato-Katz method. The prevalence of malaria, geohelminths, and co-infections were 77.2%, 28.6%, and 22.1%, respectively. Plasmodium falciparum was the only malaria parasite species identified with mean parasite density of 111 (40; 18,800) parasites/µl of blood. The geohelminths found were Ascaris lumbricoides (21.6%) and Trichuris trichiura (10.8%), with mean parasite densities of 243 (24; 3,552) and 36 (24; 96) eggs/gram of faeces, respectively. Co-infections of A. lumbricoides and P. falciparum were the most frequent and correlated positively. While no significant difference was observed on the prevalences of single and co-infections between the two localities, there was a significant difference in the density of A. lumbricoides infection between the two localities. The overall prevalence of anaemia was 42%, with individuals co-infected with T. trichiura and P. falciparum (60%) being the most at risk. While the prevalence of malaria and anaemia were inversely related to age, children aged 5–14 years were more susceptible to geohelminthiasis and their co-infections with malaria.

Conclusion/Significance

Co-existence of geohelminths and malaria parasites in Nkassomo and Vian enhances the occurrence of co-infections, and consequently, increases the risk for anaemia.