S/GSK1349572 is a next-generation HIV integrase (IN) inhibitor designed to deliver potent antiviral activity with a low-milligram once-daily dose requiring no pharmacokinetic (PK) booster. In addition, S/GSK1349572 demonstrates activity against clinically relevant IN mutant viruses and has potential for a high genetic barrier to resistance. S/GSK1349572 is a two-metal-binding HIV integrase strand transfer inhibitor whose mechanism of action was established through in vitro integrase enzyme assays, resistance passage experiments, activity against viral strains resistant to other classes of anti-HIV agents, and mechanistic cellular assays. In a variety of cellular antiviral assays, S/GSK1349572 inhibited HIV replication with low-nanomolar or subnanomolar potency and with a selectivity index of 9,400. The protein-adjusted half-maximal effective concentration (PA-EC 50 ) extrapolated to 100% human serum was 38 nM. When virus was passaged in the presence of S/GSK1349572, highly resistant mutants were not selected, but mutations that effected a low fold change (FC) in the EC 50 (up to 4.1 fold) were identified in the vicinity of the integrase active site. S/GSK1349572 demonstrated activity against site-directed molecular clones containing the raltegravir-resistant signature mutations Y143R, Q148K, N155H, and G140S/Q148H (FCs, 1.4, 1.1, 1.2, and 2.6, respectively), while these mutants led to a high FC in the EC 50 of raltegravir (11-to >130-fold). Either additive or synergistic effects were observed when S/GSK1349572 was tested in combination with representative approved antiretroviral agents; no antagonistic effects were seen. These findings demonstrate that S/GSK1349572 would be classified as a nextgeneration drug in the integrase inhibitor class, with a resistance profile markedly different from that of first-generation integrase inhibitors.Twenty-three compounds are currently approved for the treatment of HIV infection. These drugs can be assigned to six classes: nucleoside (nucleotide) reverse transcriptase inhibitors [N(t)RTIs], nonnucleoside reverse transcriptase inhibitors [NNRTIs], protease inhibitors [PIs], integrase inhibitors [INIs], CCR5 antagonists, and fusion inhibitors. The development of resistance to all currently marketed drugs has been observed and is a major reason for failure of therapy. Thus, the development of new, potent antiretroviral compounds with different resistance profiles and mechanisms of action is urgently needed for patients who have multidrugresistant HIV. In addition to these characteristics, an improved side effect profile and improved dosing convenience (oncedaily dosing, fixed-dose combination pills) are desirable, because they would promote high compliance, decrease the emergence of drug-resistant variants, and thus enhance the length and quality of life.After an initial period of false starts, advances in the field of HIV integrase drug discovery since the late 1990s have been outstanding. Beginning with the discovery that molecules capable of binding two metals within the int...

The integrase inhibitor (INI) dolutegravir (DTG; S/GSK1349572

Spinocerebellar ataxia type 14 (SCA14) is an autosomal dominant neurodegenerative disease caused by mutations in protein kinase C␥ (PKC␥). Interestingly, 18 of 22 mutations are concentrated in the C1 domain, which binds diacylglycerol and is necessary for translocation and regulation of PKC␥ kinase activity. To determine the effect of these mutations on PKC␥ function and the pathology of SCA14, we investigated the enzymological properties of the mutant PKC␥s. We found that wild-type PKC␥, but not C1 domain mutants, inhibits Ca 2؉ influx in response to muscarinic receptor stimulation. The sustained Ca 2؉ influx induced by muscarinic receptor ligation caused prolonged membrane localization of mutant PKC␥. Pharmacological experiments showed that canonical transient receptor potential (TRPC) channels are responsible for the Ca 2؉ influx regulated by PKC␥. Although in vitro kinase assays revealed that most C1 domain mutants are constitutively active, they could not phosphorylate TRPC3 channels in vivo. Single molecule observation by the total internal reflection fluorescence microscopy revealed that the membrane residence time of mutant PKC␥s was significantly shorter than that of the wild-type. This fact indicated that, although membrane association of the C1 domain mutants was apparently prolonged, these mutants have a reduced ability to bind diacylglycerol and be retained on the plasma membrane. As a result, they fail to phosphorylate TRPC channels, resulting in sustained Ca 2؉ entry. Such an alteration in Ca 2؉ homeostasis and Ca 2؉ -mediated signaling in Purkinje cells may contribute to the neurodegeneration characteristic of SCA14.Autosomal dominant SCA14 is a genetically heterogenous group of neurodegenerative disorders characterized by progressive motor incoordination affecting the gait and limbs, cerebellar dysarthria, and nystagmus due to degeneration of cerebellar Purkinje cells. SCA14 is caused by missense or in-frame deletion mutations in the PRKCG gene encoding protein kinase C␥ (PKC␥) 2 (1). PKC␥ is a member of the PKC family that plays critical roles in many cellular functions, affecting diverse signal transduction pathways (2). PKC␥ is selectively expressed in neurons throughout the brain and is most abundant in cerebellar Purkinje cells (3), which specifically degenerate in SCA14 patients.One of the characteristic features of PKC␥ is its translocation from the cytoplasm to the plasma membrane (4). Translocation is a hallmark of enzyme activation and is triggered by the stimulation of G protein-coupled receptors. It is well known that activation of such receptors causes elevations of DAG and intracellular Ca 2ϩ (5). PKC␥ contains C1 and C2 domains in its regulatory domain (6). The C1 domain has two zinc-finger motifs, C1A and C1B, that contain highly conserved Cys residues that bind to diacylglycerol (DAG) and tumor promoting phorbol esters. The C2 domain is a Ca 2ϩ sensor that binds phosphatidylserine (PS) in the presence of elevated Ca 2ϩ . The C1 and C2 domains play crucial roles in PKC␥ transloca...

In combination with PSS, the scale has predictive utility to detect suicidal ideation in elderly community residents.

We prepared a mechanically tough, highly transparent elastomer by using a polyrotaxane cross-linker acting as a molecular pulley.

There is increasing evidence that bone marrow stromal cells (BMSC) have the potential to migrate into the injured neural tissue and to differentiate into the CNS cells, indicating the possibility of autograft transplantation therapy. The present study was aimed to clarify whether the mouse BMSC can migrate into the lesion and differentiate into the CNS cells when transplanted into the mice subjected to focal cerebral infarct or spinal cord injury. The BMSC were harvested from mice and characterized by flow cytometry. Then, the BMSC were labeled by bis-benzimide, a nuclear fluorescence dye, over 24 h, and were stereotactically transplanted into the brain or spinal cord of the mice. The cultured BMSC expressed low levels of CD45 and high levels of CD90 and Sca-1 on flow cytometry. A large number of grafted cells survived in the normal brain 4 weeks after transplantation, many of which were located close to the transplanted sites. They expressed the neuronal marker including NeuN, MAP2, and doublecortin on fluorescent immunohistochemistry. However, when the BMSC were transplanted into the ipsilateral striatum of the mice subjected to middle cerebral artery occlusion, many of the grafted cells migrated into the corpus callosum and injured cortex, and also expressed the neuronal markers 4 weeks after transplantation. In particular, NeuN was very useful to validate the differentiation of the grafted cells, because the marker was expressed in the nuclei and was overlapped with bis-benzimide. Similar results were obtained in the mice subjected to spinal cord injury. However, many of the transplanted BMSC expressed GFAP, an astrocytic protein, in injured spinal cord. The present results indicate that the mouse BMSC can migrate into the CNS lesion and differentiate into the neurons or astrocytes, and that bis-benzimide is a simple and useful marker to label the donor cells and to evaluate their migration and differentiation in the host neural tissues over a long period.

Farbecht: Pigmente mit verschiedenen winkelunabhängigen Farben wurden mit einer außergewöhnlich einfachen Sprühmethode hergestellt (siehe Bild). Die Methode basiert auf der Nutzung von Submikrometer‐großen SiO2‐Partikeln und Industrieruß. Ein Polyelektrolyt, das an den Partikeln haftet, kann die Struktur der kolloidalen amorphen Anordnung stabilisieren und damit hochbelastbare, farbechte Pigmente erzeugen.

We report herein the discovery of the human immunodeficiency virus type-1 (HIV-1) integrase inhibitors dolutegravir (S/GSK1349572) (3) and S/GSK1265744 (4). These drugs stem from a series of carbamoyl pyridone analogues designed using a two-metal chelation model of the integrase catalytic active site. Structure-activity studies evolved a tricyclic series of carbamoyl pyridines that demonstrated properties indicative of once-daily dosing and superior potency against resistant viral strains. An inherent hemiaminal ring fusion stereocenter within the tricyclic carbamoyl pyridone scaffold led to a critical substrate controlled diastereoselective synthetic strategy whereby chiral information from small readily available amino alcohols was employed to control relative and absolute stereochemistry of the final drug candidates. Modest to extremely high levels of stereochemical control were observed depending on ring size and position of the stereocenter. This approach resulted in the discovery of 3 and 4, which are currently in clinical development.