Search

Protein-protein interactions represent targets for drug discovery that are highly relevant in a biological sense, but have proven difficult in a practical sense. Nevertheless, there have been recent successes in discovering drug-like small molecule inhibitors of protein-protein systems. To build on this progress, it is worth analyzing successful cases to understand at a molecular level the strategies by which these compounds effectively interfere with protein-protein pairing. A commonly observed situation is one wherein the small molecule acts as a direct mimic of one of the protein partners. This review focuses exclusively on cases where this strategy is employed, and examines the structural characteristics of the binding sites and the conformational attributes of the small molecule ligands. Common traits shared among these successful examples are identified, and formulated into potentially useful guidance for drug discovery efforts within this target class.

4-(1-Benzo[1,3]dioxol-5-ylmethylpiperidine-4-ylmethyl)-6-chlorochromen-2-one (7) is a potent, orally bioavailable melanin concentrating hormone receptor 1 (MCHr1) antagonist that causes dose-dependent weight loss in diet-induced obese mice. Further evaluation of 7 in an anesthetized dog model of cardiovascular safety revealed adverse hemodynamic effects at a plasma concentration comparable to the minimally effective therapeutic concentration. These results highlight the need for scrutiny of the cardiovascular safety profile of MCHr1 antagonists.

A pharmacological approach to inhibition of cyclin-dependent kinases 4 and 6 (Cdk4/6) using highly selective small molecule inhibitors has the potential to provide novel cancer therapies for clinical use. Achieving high levels of selectivity for Cdk4/6, versus other ATP-dependent kinases, presents a significant challenge. The pyrido[2,3-d]pyrimidin-7-one template provides an effective platform for the inhibition of a broad cross-section of kinases, including Cdks. It is now demonstrated that the modification of pyrido[2,3-d]pyrimidin-7-ones to include a 2-aminopyridine side chain at the C2-position provides inhibitors with exquisite selectivity for Cdk4/6 in vitro. This selectivity profile is recapitulated in cells where the most selective inhibitors create a G1 block at concentrations up to 100-fold the IC50 for cell proliferation. On the basis of its selectivity profile and pharmacokinetic profile, compound 43 (PD 0332991) was identified as a drug candidate for the treatment of cancer.

Inhibition of the cell cycle kinase, cyclin-dependent kinase-4 (Cdk4), is expected to provide an effective method for the treatment of proliferative diseases such as cancer. The pyrido[2,3-d]pyrimidin-7-one template has been identified previously as a privileged structure for the inhibition of ATP-dependent kinases, and good potency against Cdks has been reported for representative examples. Obtaining selectivity for individual Cdk enzymes, particularly Cdk4, has been challenging. Here, we report that the introduction of a methyl substituent at the C-5 position of the pyrido[2,3-d]pyrimidin-7-one template is sufficient to confer excellent selectivity for Cdk4 vs other Cdks and representative tyrosine kinases. Further optimization led to the identification of highly potent and selective inhibitors of Cdk4 that exhibit potent antiproliferative activity against human tumor cells in vitro. The most selective Cdk4 inhibitors were evaluated for antitumor activity against MDA-MB-435 human breast carcinoma xenografts in mice.

The results for cluster shape anisotropy over a broad range (10^-3−10^-1) of monomer volume fractions, fv values, are presented for both two- (2d) and three-dimensional (3d) simulations of diffusion-limited (DLCA), ballistic-limited (BLCA), and reaction-limited (RLCA) cluster−cluster aggregation classes. We find that all three aggregation classes have different dilute-limit shape anisotropies, with the diffusion-limited model having the largest value of anisotropy and the reaction-limited model having the smallest. The simulation result for the cluster shape anisotropy for each of the three aggregation classes is slightly less than the corresponding prediction of the hierarchial model. In addition, we find excellent agreement between the 2d DLCA simulation results and experimental measurements of shape anisotropy. At late times, shape anisotropy decreases from the dilute-limit value.

The morphology of clusters formed by selective aggregation of binary colloids is studied in a two-dimensional Monte Carlo simulation for a large range of number fractions (200:1, 100:1, 10:1, 2:1). We find remarkable similarity in morphology to those observed in experiments, from the formation of closed “micelles” to large branched clusters. Quantitative studies of the fractal dimension, kinetics, and cluster size distribution are also carried out and compared with diffusion-limited cluster aggregation and reaction-limited cluster aggregation models.

We report the observation of a novel phase of soot in an acetylene/air laminar diffusion flame. As was measured by a small-angle light scattering apparatus, the phase consists of about 10-μm superaggregates with a fractal dimension of D ≃ 2.6. This phase coexists with submicrometer D ≃ 1.8 soot fractal aggregates typical for most flames. Both percolation and shear-induced restructuring are considered as possible mechanisms of formation for those superaggregates.

The elevated expression of cell adhesion molecules (CAMs) on the lumenal surface of vascular endothelial cells is a critical early event in the complex inflammatory process. The adhesive interactions of these CAMs that include E-selectin, ICAM-1, and VCAM-1 with their counter-receptors on leukocytes, such as integrins of the αLβ2 family, result in migration of the leukocytes to the site of inflammation and cause tissue injury. Pharmaceutical agents that could suppress the induced expression of one or more of these cell adhesion molecules would provide a novel mechanism to attenuate the inflammatory responses associated with chronic inflammatory diseases. A-205804 (1), a potent and selective inhibitor of the induced expression of E-selectin and ICAM-1 over VCAM-1, was further modified with emphasis at the C-4 and C-2 positions to identify a more potent drug candidate with a good pharmacokinetic profile and physical properties. Replacement of the C-4 sulfur linkage in 1 with an oxygen atom eliminated one of the two major metabolites for this lead molecule. The para-position of the 4-phenoxy group of the thieno[2,3-c]pyridine lead is found to be very critical for a higher in vitro potency and selectivity of E-selectin and ICAM-1 over VCAM-1 expression. This position is presumably close to the solvent-accessible region of the target protein−inhibitor complex. An attempt to install a water-solubilizing group at the para-position of the phenoxy group to increase the aqueous solubility of this lead series through various linkages failed to provide an ideal inhibitor. Only small substituents such as fluorine are tolerated at the meta- and ortho-positions of the 4-phenoxy to retain a good in vitro potency. Bromo, trifluoromethyl, pyrazol-1-yl, and imidazol-1-yl are among the better substituents at the para-position. With fine-tuning at the C-2 position we discovered a series of very potent (IC50 < 5 nM for ICAM-1) and selective (>200-fold vs VCAM-1) inhibitors with a good pharmacokinetic profile. Demonstrated efficacy in a rat rheumatoid arthritis model and in a mice asthma model with selected compounds is also reported.

A critical early event in the inflammatory cascade is the induced expression of cell adhesion molecules on the lumenal surface of vascular endothelial cells. These adhesion molecules include E-selectin, ICAM-1, and VCAM-1, which serve to recruit circulating leukocytes to the site of the inflammation. These adhesive interactions allow the leukocytes to firmly adhere to and cross the vascular endothelium and migrate to the site of tissue injury. Pharmaceutical agents which would prevent the induced expression of one or more of the cell adhesion molecules on the endothelium might be expected to provide a novel mechanism to attenuate the inflammatory responses associated with chronic inflammatory diseases. A thieno[2,3-d]pyrimidine, A-155918, was identified from a whole-cell high-throughput assay for compounds which inhibited the tumor necrosis factor-α (TNFα)-induced expression of E-selectin, ICAM-1, or VCAM-1 on human vascular endothelial cells. Traditional medicinal chemistry methods were applied to this low-micromolar inhibitor, resulting in the 2,4-disubstituted thieno[2,3-c]pyridine A-205804, a potent and selective lead inhibitor of E-selectin and ICAM-1 expression (IC50 = 20 and 25 nM, respectively). The relative position of the nitrogen atom in the thienopyridine isomer was shown to be critical for activity, as was a small amide 2-substituent.

4-Anilinoquinazoline- and 4-anilinopyrido[3,4-d]pyrimidine-6-acrylamides are potent pan-erbB tyrosine kinase inactivators, and one example (CI-1033) is in clinical trial. A series of analogues with a variety of Michael acceptor units at the 6-position were prepared to define the structural requirements for irreversible inhibition. A particular goal was to determine whether additional functions to increase solubility could be appended to the Michael acceptor. Substituted acrylamides were prepared by direct acylation of the corresponding 6-amines with the requisite acid or acid chloride. Vinylsulfonamide derivatives were obtained by acylation of the amines with chloroethylsulfonyl chloride followed by base-promoted elimination. Vinylsulfone and vinylsulfine derivatives were prepared by oxidation and base elimination of a hydroxyethylthio intermediate. The compounds were evaluated for their inhibition of phosphorylation of the isolated EGFR enzyme and for inhibition of EGF-stimulated autophosphorylation of EGFR in A431 cells and of heregulin-stimulated autophosphorylation of erbB2 in MDA-MB 453 cells. Substitution at the nitrogen of the acrylamide was tolerated only with a methyl group; larger substituents were dystherapeutic, and no substitution at all was tolerated at the acrylamide α-carbon. In contrast, while electron-donating groups at the acrylamide β-carbon were not useful, even quite large electron-withdrawing groups (which increase its electrophilicity) were tolerated. A series of derivatives with solubility-enhancing substituents linked to the acrylamide β-carbon via amides were potent irreversible inhibitors of isolated EGFR (IC50s = 0.4−1.1 nM), with weakly basic morpholine and imidazole derivatives being the best. Vinylsulfonamides were also potent and irreversible inhibitors, but vinylsulfones and vinylsulfines were reversible and only poorly active. Two compounds were evaluated against A431, H125, and MCF-7 xenografts in nude mice but were inferior in these assays to the clinical trial compound CI-1033.

The identification of 8-ethyl-2-phenylamino-8H-pyrido[2,3-d]pyrimidin-7-one (1) as an inhibitor of Cdk4 led to the initiation of a program to evaluate related pyrido[2,3-d]pyrimidin-7-ones for inhibition of cyclin-dependent kinases (Cdks). Analysis of more than 60 analogues has identified some clear SAR trends that may be exploited in the design of more potent Cdk inhibitors. The most potent Cdk4 inhibitors reported in this study inhibit Cdk4 with IC50 = 0.004 μM ([ATP] = 25 μM). X-ray crystallographic analysis of representative compounds bound to the related kinase, Cdk2, reveals that they occupy the ATP binding site. Modest selectivity between Cdks is exhibited by some compounds, and Cdk4-selective inhibitors block pRb⁺ cells in the G1-phase of the cell division cycle.

The ion counting as well as the current gain of an electron multiplier type 119EM with BeCu dynodes having a Venetian blind structure was measured in terms of the dependence on the position where ions impact the surface of the first dynode. The dependence of the 119EM's detection efficiency on the impact position exhibited large variations across the Venetian blind of the first dynode. The highest detection efficiency was localized at the first dynode’s surface near its output area into the second dynode region. The lowest one was measured at the input area of the first dynode region that is far from the second dynode. The measurements also show that the 119EM is not very reliable for ion counting. The analog particle gain derived from the mean current gain measured across a single slat increases with increasing charge state as well as with ion energy for Coq+(10⩽q⩽26) and Taq+(14⩽q⩽34) ions with kinetic energy per charge from 33 keV/qto 163 keV/q. The gains were derived from comparison with the Faraday cup measurements.

Background: The lack of an early, sensitive marker for intestinal ischemia has led to delay in diagnosis and worsened outcome for patients with acute onset of this condition. Our preliminary studies revealed that guinea pig cytosolic @b-glucosidase (CBG) is expressed predominantly in the small intestine, with lower levels in the liver and pancreas and undetectable levels in other organs. Cytosolic @b-glucosidase was investigated as a serum marker of small intestinal ischemia in a guinea pig model. Methods: Guinea pigs underwent anesthesia, sham laparotomy, 30 minutes of mesenteric ischemia followed by 6 hours of reperfusion, 6 hours of sustained mesenteric ischemia, or closed-loop small bowel obstruction. Serum samples were assayed for CBG activity. At the conclusion of the ischemia/reperfusion experiments, small bowel samples were assayed for residual enzyme activity, and paraffin sections were graded for the severity of histologic injury. Results: Serum CBG activity rose rapidly after intestinal ischemia with and without reperfusion. Peak enzyme activities were elevated 23-fold for reperfused animals (P < .001) by 4 hours. For nonreperfused animals, peak serum CBG activities reached 29-fold above baseline and were significantly higher than the CBG activities of reperfused animals at 4 hours (P < .01) and at 6 hours (P < .05). Mucosal injury ranged from undetectable to moderate and corresponded in severity with both peak serum enzyme activity and decreased residual activity in the small bowel. In animals subjected to closed-loop obstruction, there was a mean increase of serum CBG of 9.2-fold from 4 to 6 hours after establishment of obstruction (P < .05). Conclusions: In the guinea pig model, CBG is a sensitive marker of ischemic injury caused by arterial occlusion or closed-loop obstruction of the small bowel. (Surgery 1999;125:202-10.)

Several elaborations of the fundamental anilinopyrimidine pharmacophore have been reported as potent and selective inhibitors of the epidermal growth factor receptor (EGFr) tyrosine kinase. This paper reports on a series of inhibitors whereby some 6,5-bicyclic heteroaromatic systems were fused through their C-2 and C-3 positions to this anilinopyrimidine pharmacophore. Although the resulting tricycles did not produce the enormous potency of some of the (5/6),6,6-bicyclic systems, the best of them had IC50s for the EGFr TK around 1 nM. Investigation of 4-position side chains in the indolopyrimidines confirmed that m-bromoaniline was an optimal substituent for potency. Investigation of substitution within the C-(benzo)ring of benzothienopyrimidines confirmed that introduction of an extra ring can change sharply the effects of substituents when compared to similar bicyclic nuclei, and only two substituents were found which even moderately enhanced inhibitory activity over the parent compound for this series.