Anti-hepatitis C virus activity and toxicity of type III phosphatidylinositol-4-kinase beta inhibitors.

Type III phosphatidylinositol-4-kinase beta (PI4KIIIß) was previously implicated in hepatitis C virus (HCV) replication by small interfering RNA (siRNA) depletion and was therefore proposed as a novel cellular target for the treatment of hepatitis C. Medicinal chemistry efforts identified highly selective PI4KIIIß inhibitors that potently inhibited the replication of genotype 1a and 1b HCV replicons and genotype 2a virus in vitro. Replicon cells required more than 5 weeks to reach low levels of 3- to 5-fold resistance, suggesting a high resistance barrier to these cellular targets. Extensive in vitro profiling of the compounds revealed a role of PI4KIIIß in lymphocyte proliferation. Previously proposed functions of PI4KIIIß in insulin secretion and the regulation of several ion channels were not perturbed with these inhibitors. Moreover, PI4KIIIß inhibitors were not generally cytotoxic as demonstrated across hundreds of cell lines and primary cells. However, an unexpected antiproliferative effect in lymphocytes precluded their further development for the treatment of hepatitis C.

TiO(2) nanorods/PMMA copolymer-based nanocomposites: highly homogeneous linear and nonlinear optical material.

Original nanocomposites have been obtained by direct incorporation of pre-synthesized oleic acid capped TiO(2) nanorods into properly functionalized poly(methyl methacrylate) copolymers, carrying carboxylic acid groups on the repeating polymer unit. The presence of carboxylic groups on the alkyl chain of the host functionalized copolymer allows an highly homogeneous dispersion of the nanorods in the organic matrix. The prepared TiO(2)/PMMA-co-MA nanocomposites show high optical transparency in the visible region, even at high TiO(2) nanorod content, and tunable linear refractive index depending on the nanoparticle concentration. Finally measurements of nonlinear optical properties of TiO(2) polymer nanocomposites demonstrate a negligible two-photon absorption and a negative value of nonlinear refractive index, highlighting the potential of the nanocomposite for efficient optical devices operating in the visible region.

Granulocyte/macrophage colony-stimulating factor treatment of human chronic ulcers promotes angiogenesis associated with de novo vascular endothelial growth factor transcription in the ulcer bed.

Summary Background Granulocyte/macrophage colony-stimulating factor (GM-CSF), a cytokine with pleiotropic functions, has been successfully employed in the treatment of chronic skin ulcers. The biological effects underlying GM-CSF action in impaired wound healing have been only partly clarified. Objectives To investigate the effects of GM-CSF treatment of chronic venous ulcers on lesion vascularization and on the local synthesis of the angiogenic factors vascular endothelial growth factor (VEGF) and placenta growth factor (PlGF). Methods Patients with nonhealing venous leg ulcers were treated with intradermal injection of recombinant human GM-CSF, and biopsies were taken at the ulcer margin before and 5 days after administration. Wound vascularization was analysed by immunohistochemistry using antiplatelet endothelial cell adhesion molecule-1/CD31 and anti-alpha-smooth muscle actin antibodies. VEGF and PlGF transcription was assessed by in situ hybridization. To identify the cell populations transcribing VEGF within the ulcer bed, the VEGF hybridization signal was correlated with the immunostaining for different cell type markers on serial sections. Direct induction of VEGF transcription by GM-CSF was investigated in GM-CSF-treated cultured macrophages and keratinocytes. Results Blood vessel density was significantly increased in the ulcer bed following GM-CSF treatment. VEGF transcripts were localized in keratinocytes at the ulcer margin both before and after GM-CSF treatment, whereas a VEGF hybridization signal was evident within the ulcer bed only following administration. PlGF mRNA was barely detectable in keratinocytes at the ulcer margin and was not visibly increased after treatment. Unlike VEGF, a specific PlGF hybridization signal could not be detected in cells within the ulcer following GM-CSF administration. Monocytes/macrophages were the main cell population transcribing VEGF after GM-CSF treatment. In vitro analysis demonstrated that VEGF transcription can be directly stimulated by GM-CSF in a differentiated monocytic cell line, but not in keratinocytes. Conclusions Our data show that increased vascularization is associated with GM-CSF treatment of chronic venous ulcers and indicate that inflammatory cell-derived VEGF may act as an angiogenic mediator of the healing effect of GM-CSF in chronic ulcers.

High third-order nonlinear optical susceptibility in new fluorinated poly(p-phenylenevinylene) copolymers measured with the Z-scan technique.

The third-order nonlinear optical properties of a series of copoly(2, 3, 5, 6-tetrafluoro-1, 4-phenylenevinylene-2, 5-dioctyloxy-1, 4-phenylenevinylene) that contain variable ratios of two differently substituted monomers have been studied in chloroform solutions at l=1064 nm by the picosecond Z-scan technique. Nonlinear refractive index n(2) of the samples investigated has been found to be negative, and a strong dependence of its magnitude on the copolymer's composition has been observed. The highest third-order nonlinear optical susceptibility, |x((3))|=(6 +/- 2)x 10(-10) esu, was measured for a copolymer obtained by reaction of equimolar quantitites of the parent monomers.

Identification of dipeptidyl nitriles as potent and selective inhibitors of cathepsin B through structure-based drug design.

Cathepsin B is a member of the papain superfamily of cysteine proteases and has been implicated in the pathology of numerous diseases, including arthritis and cancer. As part of an effort to identify potent, reversible inhibitors of this protease, we examined a series of dipeptidyl nitriles, starting with the previously reported Cbz-Phe-NH-CH(2)CN (19, IC(50) = 62 microM). High-resolution X-ray crystallographic data and molecular modeling were used to optimize the P(1), P(2), and P(3) substituents of this template. Cathepsin B is unique in its class in that it contains a carboxylate recognition site in the S(2)' pocket of the active site. Inhibitor potency and selectivity were enhanced by tethering a carboxylate functionality from the carbon alpha to the nitrile to interact with this region of the enzyme. This resulted in the identification of compound 10, a 7 nM inhibitor of cathepsin B, with excellent selectivity over other cysteine cathepsins.

Tipranavir (PNU-140690): a potent, orally bioavailable nonpeptidic HIV protease inhibitor of the 5,6-dihydro-4-hydroxy-2-pyrone sulfonamide class.

A broad screening program previously identified phenprocoumon (1) as a small molecule template for inhibition of HIV protease. Subsequent modification of this lead through iterative cycles of structure-based design led to the activity enhancements of pyrone and dihydropyrone ring systems (II and V) and amide-based substitution (III). Incorporation of sulfonamide substitution within the dihydropyrone template provided a series of highly potent HIV protease inhibitors, with structure-activity relationships described in this paper. Crystallographic studies provided further information on important binding interactions responsible for high enzymatic binding. These studies culminated in compound VI, which inhibits HIV protease with a Ki value of 8 pM and shows an IC90 value of 100 nM in antiviral cell culture. Clinical trials of this compound (PNU-140690, Tipranavir) for treatment of HIV infection are currently underway.

Non-peptidic HIV protease inhibitors: C2-symmetry-based design of bis-sulfonamide dihydropyrones.

Potent, non-peptidic, dihydropyrone sulfonamide HIV protease inhibitors have been previously described. Crystallographic analysis of dihydropyrone sulfonamide inhibitor/HIV protease complexes suggested incorporation of a second, C2 symmetry-related sulfonamide group. Selected bis-sulfonamide dihydropyrone analogues display high HIV protease inhibitory activity.

Structure-based design of HIV protease inhibitors: 5,6-dihydro-4-hydroxy-2-pyrones as effective, nonpeptidic inhibitors.

From a broad screening program, the 4-hydroxycoumarin phenprocoumon (I) was previously identified as a lead template with HIV protease inhibitory activity. The crystal structure of phenprocoumon/HIV protease complex initiated a structure-based design effort that initially identified the 4-hydroxy-2-pyrone U-96988 (II) as a first-generation clinical candidate for the potential treatment of HIV infection. Based upon the crystal structure of the 4-hydroxy-2-pyrone III/HIV protease complex, a series of analogues incorporating a 5,6-dihydro-4-hydroxy-2-pyrone template were studied. It was recognized that in addition to having the required pharmacophore (the 4-hydroxy group with hydrogen-bonding interaction with the two catalytic aspartic acid residues and the lactone moiety replacing the ubiquitous water molecule in the active site), these 5,6-dihydro-4-hydroxy-2-pyrones incorporated side chains at the C-6 position that appropriately extended into the S1' and S2' subsites of the enzyme active site. The crystal structures of a number of representative 5,6-dihydro-4-hydroxy-2-pyrones complexed with the HIV protease were also determined to provide better understanding of the interaction between the enzyme and these inhibitors to aid the structure-based drug design effort. The crystal structures of the ligands in the enzyme active site did not always agree with the conformations expected from experience with previous pyrone inhibitors. This is likely due to the increased flexibility of the dihydropyrone ring. From this study, compound XIX exhibited reasonably high enzyme inhibitory activity (Ki = 15 nM) and showed antiviral activity (IC50 = 5 microM) in the cell-culture assay. This result provided a research direction which led to the discovery of active 5,6-dihydro-4-hydroxy-2-pyrones as potential agents for the treatment of HIV infection.

[Metastatic pancreatic endocrine tumor with elevation of alpha-fetoprotein]. / Tumeur endocrine pancréatique métastatique avec élévation de l'alpha-foetoprotéine.

Hepatoma was diagnosed in a 47 year old man presenting with multiple liver tumors and elevated alpha-fetoprotein concentration (460 micrograms/l). Liver biopsy showed a poorly differentiated carcinoma. The discovery of 2 hypervascular tumors in the splenic area at celiac arteriography led to challenge the diagnosis: ultrasound guided-aspiration biopsy showed endocrine cells at the level of the hepatic and pancreatic tumors. The diagnosis of pancreatic islet cell tumor with liver secondaries was confirmed by the pathology of the operative specimen. During the following months, alpha-fetoprotein concentration steadily increased. The patient died 5 months later from widespread metastases. At necropsy, no evidence of hepatoma was found. Immunocytochemical study with antibody raised against alpha-fetoprotein was negative both at the level of the tumoral cells and of the liver cells around the tumor. This case adds a new cause to abnormally high plasma alpha-fetoprotein. As hepatoma and endocrine tumor metastatic to the liver could be difficult to differentiate, this new cause could be clinically relevant.

Periaqueductal gray matter involvement in the muscimol-induced decrease of morphine antinociception.

Microinjections of muscimol, a GABA receptor agonist, into the periaqueductal gray matter (PAG) counter-acted the antinociceptive effect of morphine in rats, as measured by the "tail-flick" method. Muscimol's effect was partially reversed by bicuculline.