SAR inspired by aldehyde oxidase (AO) metabolism: Discovery of novel,CNS penetrant tricyclic M4 PAMs

This letter describes progress towards an M₄ PAM preclinical candidate inspired by an unexpected aldehyde oxidase (AO) metabolite of a novel, CNS penetrant thieno[2,3-c]pyridine core to an equipotent, non-CNS penetrant thieno[2,3-c]pyrdin-7(6H)-one core. Medicinal chemistry design efforts yielded two novel tricyclic cores that enhanced M₄ PAM potency, regained CNS penetration, displayed favorable DMPK properties and afforded robust in vivo efficacy in reversing amphetamine-induced hyperlocomotion in rats.     

Effects of decadal climate change on zooplankton over the last 50 years in the western subarctic North Pacific

Decadal‐ to multi‐decadal variations have been reported in many regional ecosystems in the North Pacific, resulting in an increasing demand to elucidate the link between long‐term climatic forcing and marine ecosystems. We detected phenological and quantitative changes in the copepod community in response to the decadal climatic variation in the western subarctic North Pacific by analyzing the extensive zooplankton collection taken since the 1950s, the Odate Collection. Copepod species were classified into five seasonal groups depending on the timing of the annual peak in abundance. The abundance of the spring community gradually increased for the period 1960–2002. The spring–summer community also showed an increasing trend in May, but a decadal oscillation pattern of quasi‐30‐year cycles in July. Phenological changes coincided with the climate regime shift in the mid‐1970s, indicated by the Pacific decadal oscillation index (PDO). After the regime shift, the timing of the peak abundance was delayed one month, from March–April to April–May, in the spring community, whereas it peaked earlier, from June–July to May–June, in the spring–summer community, resulting in an overlap of the high productivity period for the two communities in May. Wintertime cooling, followed by rapid summertime warming, was considered to be responsible for delayed initiation and early termination of the productive season after the mid‐1970s. Another phenological shift, quite different from the previous decade, was observed in the mid‐1990s, when warm winters followed by cool summers lengthened the productive season. The results suggest that climatic forcing with different decadal cycles may operate independently during winter–spring and spring–summer to create seasonal and interannual variations in hydrographic conditions; thus, combinations of these seasonal processes may determine the annual biological productivity.     

Roles of basic residues and salt-bridge interaction in a vacuolar H-pumping pyrophosphatase (AVP1) from Arabidopsis thaliana

To investigate the possible role of basic residues in H⁺ translocation through vacuolar-type H⁺-pumping pyrophosphatases (V-PPases), conserved arginine and lysine residues predicted to reside within or close to transmembrane domains of an Arabidopsis thaliana V-PPase (AVP1) were subjected to site-directed mutagenesis. One of these mutants (K461A) exhibited a “decoupled” phenotype in which proton-pumping but not hydrolysis was inhibited. Similar results were reported previously for an E427Q mutant, resulting in the proposal that E427 might be involved in proton translocation. However, the double mutant E427K/K461E has a wild type phenotype, suggesting that E427 and K461 form a stabilising salt bridge, but that neither residue plays a critical role in proton translocation.     

Plumbagin and juglone induce caspase-3-dependent apoptosis involving the mitochondria through ROS generation in human peripheral blood lymphocytes

The phytochemicals plumbagin and juglone have recently been gaining importance because of their various pharmacological activities. In this study, these compounds are shown to induce concentration- and time-dependent toxicity in human peripheral blood lymphocytes via the apoptotic pathway. Flow cytometry data revealed the occurrence of about 28% early apoptotic cells after 6 h exposure to 10 μM plumbagin and 35% late apoptotic cells and about 43% sub-G1 population after 24 h. The cytotoxic effect of plumbagin was at least twofold higher than that of juglone as evidenced by the IC₅₀ value for cytotoxicity. Characteristic apoptotic features such as chromatin condensation and apoptotic body formation were observed through TEM, and membrane blebbing and cell surface smoothening were seen in SEM studies. Generation of ROS was evidenced through the HPLC analysis of superoxide-specific 2-OH-E+ formation. In addition, a decrease in GSH levels parallel to ROS production was observed. Reversal of apoptosis in both NAC- and Tempol-pretreated cells indicates the involvement of both ROS generation and GSH depletion in plumbagin- and juglone-induced apoptosis. The mechanistic pathway involves a decrease in MMP; alterations in the levels of Bcl-2, Bax, and cytosolic cytochrome c; and PARP-1 cleavage subsequent to caspase-3 activation.     

Selective binding of imatinib to the genetic variants of human α1-acid glycoprotein

Imatinib is a selective tyrosine kinase inhibitor, successfully used for the treatment of chronic myelogenous leukaemia. Its strong plasma protein binding referred to α₁-acid glycoprotein (AGP) component was found to inhibit the pharmacological activity. AGP shows genetic polymorphism and the two main genetic variants have different drug binding properties. The binding characteristics of imatinib to AGP genetic variants and the possibility of its binding interactions were investigated by various methods. The results proved that binding of imatinib to the two main genetic variants is very different, the high affinity binding belongs dominantly to the F1-S variant. This interaction is accompanied with specific spectral changes (induced circular dichroism, UV change, intrinsic fluorescence quenching), suggesting that the bound ligand has chiral conformation that would largely overlap with other ligands inside the protein cavity. Binding parameters of K_a = 1.7(± 0.2) × 10⁶ M^− 1 and n = 0.94 could be determined for the binding on the F1-S variant at 37°. Imatinib binding on the A variant is weaker and less specific. The binding affinity of imatinib to human serum albumin (nK_a ≈ 3 × 10⁴ M^− 1) is low. Pharmacologically relevant binding interactions with other drugs can be expected on the F1-S variant of AGP.     

Mutagenesis of acridine orange in mitotic cleavage nuclei of the silkworm, Bombyx mori

The present study was done to determine whether acridine orange (AO) is mutagenic for the mitotic cleavage nuclei in the silkworm. The mutation frequency was estimated by the specific locus method using egg-color genes. AO was injected into the body cavity of marked female pupae (homozygous for pe and re genes) in active vitellogenesis (prophase I oocytes). The moths emerging from the treated pupae were mated to wild type male moths. AO increased the frequency of mosaic type mutations, indicating that AO has a positive mutagenic action on the paternal chromosomes in the mitotic cleavage nuclei in the silkworm.