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.

Mechanism of action of and mechanism of reduced susceptibility to the novel anti-Clostridium difficile compound LFF571.

LFF571 is a novel semisynthetic thiopeptide and potent inhibitor of Gram-positive bacteria. We report that the antibacterial activity of LFF571 against Clostridium difficile is due to inhibition of translation. Single-step mutants of C. difficile with reduced susceptibility to LFF571 were selected at frequencies of <4.5 × 10(-11) to 1.2 × 10(-9). Sequencing revealed a G260E substitution in the thiopeptide-binding pocket of elongation factor Tu. Importantly, this mutation did not confer cross-resistance to clinically used antimicrobials. These results support the development of LFF571 as a treatment for C. difficile infection.

In vitro and in vivo activities of novel, semisynthetic thiopeptide inhibitors of bacterial elongation factor Tu.

Recently, we identified aminothiazole derivatives of GE2270 A. These novel semisynthetic congeners, like GE2270 A, target the essential bacterial protein elongation factor Tu (EF-Tu). Medicinal chemistry optimization of lead molecules led to the identification of preclinical development candidates 1 and 2. These cycloalklycarboxylic acid derivatives show activity against difficult to treat Gram-positive pathogens and demonstrate increased aqueous solubility compared to GE2270 A. We describe here the in vitro and in vivo activities of compounds 1 and 2 compared to marketed antibiotics. Compounds 1 and 2 were potent against clinical isolates of methicillin-resistant Staphylococcus aureus and vancomycin-resistant enterococci (MIC(90) ≤ 0.25 µg/ml) but weaker against the streptococci (MIC(90) ≥ 4 µg/ml). Like GE2270 A, the derivatives inhibited bacterial protein synthesis and selected for spontaneous loss of susceptibility via mutations in the tuf gene, encoding EF-Tu. The mutants were not cross-resistant to other antibiotic classes. In a mouse systemic infection model, compounds 1 and 2 protected mice from lethal S. aureus infections with 50% effective doses (ED(50)) of 5.2 and 4.3 mg/kg, respectively. Similarly, compounds 1 and 2 protected mice from lethal systemic E. faecalis infections with ED(50) of 0.56 and 0.23 mg/kg, respectively. In summary, compounds 1 and 2 are active in vitro and in vivo activity against difficult-to-treat Gram-positive bacterial infections and represent a promising new class of antibacterials for use in human therapy.

The relationship between Taxol and (+)-discodermolide: synthetic analogs and modeling studies.

BACKGROUND: During the past decade, Taxol has assumed an important role in cancer chemotherapy. The search for novel compounds with a mechanism of action similar to that of Taxol, but with greater efficacy particularly in Taxol-resistant cells, has led to the isolation of new natural products. One such compound, (+)-discodermolide, although structurally distinct from Taxol, has a similar ability to stabilize microtubules. In addition, (+)-discodermolide is active in Taxol-resistant cell lines that overexpress P-glycoprotein, the multidrug-resistant transporter. Interestingly, (+)-discodermolide demonstrates a profound enhancement of the initiation process of microtubule polymerization compared to Taxol. RESULTS: The synthesis of (+)-discodermolide analogs exploiting our highly efficient, triply convergent approach has permitted structure-activity relationship (SAR) studies. Small changes to the (+)-discodermolide structure resulted in a dramatic decrease in the ability of all four discodermolide analogs to initiate tubulin polymerization. Two of the analogs also demonstrated a decrease in total tubulin polymerization, while a change in the olefin geometry at the C8 position produced a significant decrease in cytotoxic activity. CONCLUSIONS: The availability of (+)-discodermolide and the analogs, and the resultant SAR analysis, have permitted an exploration of the similarities and differences between (+)-discodermolide and Taxol. Docking of the X-ray/solution structure of (+)-discodermolide into the Taxol binding site of beta-tubulin revealed two possible binding modes (models I and II). The preferred pharmacophore model (I), in which the C19 side chain of (+)-discodermolide matches with the C2 benzoyl group of Taxol and the delta-lactone ring of (+)-discodermolide overlays with the C13 side chain of Taxol, concurred with the results of the SAR analysis.

Solution structure of (+)-discodermolide.

[structure: see text]. The solution structure of (+)-discodermolide (1) has been determined via 1- and 2-D NMR techniques in conjunction with Monte Carlo conformational analysis. Taken together, the results demonstrate that in solution (+)-discodermolide occupies a helical conformation remarkably similar to the solid state conformation.

Gram-scale synthesis of (+)-discodermolide.

[formula: see text] A triply convergent, highly efficient second-generation synthesis of the potent antimitotic agent (+)-discodermolide (1) has been achieved on a 1-g scale.