Heterocyclic methylsulfone hydroxamic acid LpxC inhibitors as Gram-negative antibacterial agents

The synthesis and antibacterial activity of heterocyclic methylsulfone hydroxamates is presented. Compounds in this series are potent inhibitors of the LpxC enzyme, a key enzyme involved in the production of lipopolysaccharide (LPS) found in the outer membrane of Gram-negative bacteria. SAR evaluation of compounds in this series revealed analogs with potent antibacterial activity against challenging Gram-negative species such as Pseudomonas aeruginosa and Klebsiella pneumoniae.     

The role of short-chain dehydrogenase/oxidoreductase, induced by salt stress, on host interaction of B. pseudomallei

Background

Burkholderia pseudomallei is the causative agent of melioidosis, a frequently occurring disease in northeastern Thailand, where soil and water high in salt content are common. Using microarray analysis, we previously showed that B. pseudomallei up-regulated a short-chain dehydrogenase/oxidoreductase (SDO) under salt stress. However, the importance of SDO in B. pseudomallei infection is unknown. This study aimed to explore the function of B. pseudomallei SDO, and to investigate its role in interactions between B. pseudomallei and host cells.

Results

Bioinformatics analysis of B. pseudomallei SDO structure, based on homology modeling, revealed a NAD⁺ cofactor domain and a catalytic triad containing Ser149, Tyr162, and Lys166. This is similar to Bacillus megaterium glucose 1-dehydrogenase. To investigate the role of this protein, we constructed a B. pseudomallei SDO defective mutant, measured glucose dehydrogenase (GDH) activity, and tested the interactions with host cells. The B. pseudomallei K96243 wild type exhibited potent GDH activity under condition containing 300 mM NaCl, while the mutant showed activity levels 15 times lower. Both invasion into the A549 cell line and early intracellular survival within the J774A.1 macrophage cell were impaired in the mutant. Complementation of SDO was able to restore the mutant ability to produce GDH activity, invade epithelial cells, and survive in macrophages.

Conclusions

Our data suggest that induced SDO activity during salt stress may facilitate B. pseudomallei invasion and affect initiation of successful intracellular infection. Identifying the role of B. pseudomallei SDO provides a better understanding of the association between bacterial adaptation and pathogenesis in melioidosis.     

World Dengue Day: A call for action

Commemorating the 2021 ASEAN Dengue Day and advocacy for World Dengue Day, the International Society for Neglected Tropical Diseases (ISNTD) and Asian Dengue Voice and Action (ADVA) Group jointly hosted the ISNTD-ADVA World Dengue Day Forum–Cross Sector Synergies in June 2021. The forum aimed to achieve international and multisectoral coordination to consolidate global dengue control and prevention efforts, share best practices and resources, and improve global preparedness. The forum featured experts around the world who shared their insight, research experience, and strategies to tackle the growing threat of dengue. Over 2,000 healthcare care professionals, researchers, epidemiologists, and policy makers from 59 countries attended the forum, highlighting the urgency for integrated, multisectoral collaboration between health, environment, education, and policy to continue the march against dengue. Sustained vector control, environmental management, surveillance improved case management, continuous vaccine advocacy and research, capacity building, political commitment, and community engagement are crucial components of dengue control. A coordinated strategy based on science, transparency, timely and credible communication, and understanding of human behavior is needed to overcome vaccine hesitancy, a major health risk further magnified by the COVID-19 pandemic. The forum announced a strong call to action to establish World Dengue Day to improve global awareness, share best practices, and prioritize preparedness in the fight against dengue.     

Differences between adrenal adenoma causing primary aldosteronism and other adrenal tissues in the incorporation of labeled steroid precursors into their products

The incorporation and conversion of several labeled steroid precursors into their products were examined in slices of adrenal tissue from two patients with primary aldosteronism and compared with that in “normal” adrenal tissue and adrenal tissues from a patient with Gushing's syndrome. The products of the incorporation were separated by Sephadex LH-20 column chromatography. The major products of conversion in the adenomatous tissue of primary aldosteronism were 18-hydroxycorticosterone and lesser amounts of aldosterone. Smaller amounts of 18-hydroxycorticosterone were isolated from all other adrenal tissues studied. No aldosterone could be recovered after incubating any of the adrenal tissue studied with labeled 18-hydroxy-11-deoxycorticosterone or 18-hydroxycorticosterone as precursor steroid. These $\text{in vitro}$ results seem to suggest that there is increased 18-hydroxylation in the adenoma of primary aldosteronism compared with other tissues and that relatively more 18-hydroxycorticosterone is produced in such tissue than aldosterone.     

Impairment of the TSH signal transduction system in human thyroid carcinoma cells

In order to further evaluate the role of TSH in the proliferation and the differentiation of human thyroid carcinoma cells, we have analyzed the function of the TSH receptor in the established thyroid carcinoma cell lines NPA and WRO. The TSH signal transduction system in the carcinoma cells was also compared with that in normal thyroid cells. Although unresponsiveness to bovine and human TSH was demonstrated by measurement of cAMP production and [3H]thymidine incorporation after treatment of TSH, cAMP production was induced after stimulation of these cells by forskolin, cholera toxin, and isoproterenol. Specific binding to 125I-TSH was demonstrated in both NPA and WRO cells in addition to the existence of a TSH receptor mRNA and thyroglobulin mRNA species, although thyroid-specific gene expression in these cells was not regulated by TSH. These findings suggest that the unresponsiveness to TSH in these cells may be due to an abnormality of TSH receptor-G protein coupling rather than to a decreased level of TSH-receptor expression or a Gs protein abnormality.     

TaqMan real-time PCR assay for specific detection of Opisthorchis viverrini DNA in Thai patients with hepatocellular carcinoma and cholangiocarcinoma

The aim of this study was to develop TaqMan real-time PCR assay that detected Opisthorchis viverrini DNA from 18 normal and 18 tumor tissue specimens from Thai patients with hepatocellular carcinoma (HCC) and cholangiocarcinoma (CCA), who underwent liver resection from October 2005 to May 2006. Control liver specimens were seven non-primary liver cancers. A conserved probe representing 100% sequence homology was used as a reference for O. viverrini-specific probe. Five of six tumors (83%) and all six normal tissues from CCA group; and seven of twelve tumors (58%) and ten of twelve normal tissues (83%) from HCC group were found to have O. viverrini DNA. The O. viverrini DNA detection among HCC and CCA patients were not associated (p = 0.193; 90%CI). This RT-PCR will be a useful tool for investigating the relationship between cancer type and presence of the parasite and also for conducting epidemiological surveys.     

Novel 3-fluoro-6-methoxyquinoline derivatives as inhibitors of bacterial DNA gyrase and topoisomerase IV

Novel (non-fluoroquinolone) inhibitors of bacterial type II topoisomerases (NBTIs) are an emerging class of antibacterial agents. We report an optimized series of cyclobutylaryl-substituted NBTIs. Compound 14 demonstrated excellent activity both in vitro (S. aureus MIC₉₀ = 0.125 μg/mL) and in vivo (systemic and tissue infections). Enhanced inhibition of Topoisomerase IV correlated with improved activity in S. aureus strains with mutations conferring resistance to NBTIs. Compound 14 also displayed an improved hERG IC₅₀ of 85.9 μM and a favorable profile in the anesthetized guinea pig model.     

Ultrastructural and immunohistochemical studies on non-ciliated cells of the tracheal epithelium of normal,phenobarbital-treated,and 3-methylcholanthrene-treated mice

Summary Non-ciliated SER-rich cells of the tracheal epithelium of normal, phenobarbital-treated and 3-methylcholanthrene-treated mice were studied ultrastructurally and immunohistochemically. The apical portion of these cells protrudes into the tracheal lumen, especially in the mice treated with the two compounds, and the apical cytoplasm is filled with numerous tubular elements of SER. Besides, the non-ciliated cells of 3-methylcholanthrene-treated mice show a strong positive reaction to the antiserum against microsomal cytochrome P-450 of liver. These findings support the concept that the non-ciliated tracheal cell may be involved in the metabolism of endogeneous and exogeneous chemical compounds.This study was supported by grants from the Ministry of Education, Science and Culture, Japan     

Dihydrofolate-Reductase Mutations in Plasmodium knowlesi Appear Unrelated to Selective Drug Pressure from Putative Human-To-Human Transmission in Sabah,Malaysia

Background

Malaria caused by zoonotic Plasmodium knowlesi is an emerging threat in Eastern Malaysia. Despite demonstrated vector competency, it is unknown whether human-to-human (H-H) transmission is occurring naturally. We sought evidence of drug selection pressure from the antimalarial sulfadoxine-pyrimethamine (SP) as a potential marker of H-H transmission.

Methods

The P. knowlesi dihdyrofolate-reductase (pkdhfr) gene was sequenced from 449 P. knowlesi malaria cases from Sabah (Malaysian Borneo) and genotypes evaluated for association with clinical and epidemiological factors. Homology modelling using the pvdhfr template was used to assess the effect of pkdhfr mutations on the pyrimethamine binding pocket.

Results

Fourteen non-synonymous mutations were detected, with the most common being at codon T91P (10.2%) and R34L (10.0%), resulting in 21 different genotypes, including the wild-type, 14 single mutants, and six double mutants. One third of the P. knowlesi infections were with pkdhfr mutants; 145 (32%) patients had single mutants and 14 (3%) had double-mutants. In contrast, among the 47 P. falciparum isolates sequenced, three pfdhfr genotypes were found, with the double mutant 108N+59R being fixed and the triple mutants 108N+59R+51I and 108N+59R+164L occurring with frequencies of 4% and 8%, respectively. Two non-random spatio-temporal clusters were identified with pkdhfr genotypes. There was no association between pkdhfr mutations and hyperparasitaemia or malaria severity, both hypothesized to be indicators of H-H transmission. The orthologous loci associated with resistance in P. falciparum were not mutated in pkdhfr. Subsequent homology modelling of pkdhfr revealed gene loci 13, 53, 120, and 173 as being critical for pyrimethamine binding, however, there were no mutations at these sites among the 449 P. knowlesi isolates.

Conclusion

Although moderate diversity was observed in pkdhfr in Sabah, there was no evidence this reflected selective antifolate drug pressure in humans.     

Plasmodium vivax genetic diversity: microsatellite length matters

The Plasmodium vivax genome is very diverse but has a relatively low abundance of microsatellites. Leclerc et al. had shown that these di-nucleotide repeats have a low level of polymorphism, suggesting a recent bottleneck event in the evolutionary history of P. vivax. By contrast, in a recent paper, Imwong et al. show that there is a very high level of microsatellite diversity. The difference in these results is probably due to the set array lengths chosen by each group. Longer arrays are more diverse than are shorter ones because slippage mutations become exponentially more common with an increase in array length. These studies highlight the need to consider carefully the application and design of studies involving microsatellites.