Search

We report the results of 3D-QSAR/CoMFA investigations of the activity of bisphosphonate drugs, farnesyl pyrophosphate synthase (FPPSase) inhibitors, in the inhibition of bone resorption as well as the growth of Dictyostelium discoideum. In the case of D. discoideum, we find an experimental versus QSAR predicted pIC50 R2 value of 0.94 for 16 bisphosphonates over the 9−1200 μM range of IC50 values, a cross-validated R2 = 0.90, and a bootstrapped R2 = 0.94, and we demonstrate that this approach has predictive utility (a 0.18 pIC50 rms error for three test sets of 3 predictions). In bone resorption, we find an experimental versus predicted pLED (lowest effective dose) R2 = 0.79 for 35 bisphosphonates over the 0.0001−1 mg of P/kg LED range, a cross-validated R2 = 0.75, and a bootstrapped R2 = 0.79. Two sets of 31 compounds were used as training sets for the predicted pLED values for two sets of 4 compounds which have an rms error of 0.44, larger than that found with D. discoideum. However, this can be attribut...

We report the results of a comparative molecular field analysis (CoMFA) investigation of the growth inhibition of the bloodstream form of Trypanosoma brucei rhodesiense trypomastigotes by bisphosphonates. A quantitative three-dimensional structure-activity relationship CoMFA model for a set of 26 bisphosphonates having a range of activity spanning approximately 3 orders of magnitude (minimum IC(50) = 220 nM; maximum IC(50) = 102 microM) yielded an R(2) value of 0.87 with a cross-validated R(2) value of 0.79. The predictive utility of this approach was tested for three sets of three compounds: the average pIC(50) error was 0.23. For the nitrogen-containing bisphosphonates, in general, the activity was aromatic-aliphatic-containing side chains. The activity of aromatic species lacking an alkyl ring substitution decreased from ortho to meta to para substitution; halogen substitutions also reduced activity. For the aliphatic bisphosphonates, the IC(50) values decreased nearly monotonically with increasing chain length (down to IC(50) = 2.0 microM for the n-C(11) alkyl side chain species). We also show, using a "rescue" experiment, that the molecular target of the nitrogen-containing bisphosphonate, risedronate, in T. b. rhodesiense is the enzyme farnesyl pyrophosphate synthase. In addition, we report the LD(50) values of bisphosphonates in a mammalian cell general toxicity screen and present a comparison between the therapeutic indices and the IC(50) values in the T. b. rhodesiense growth inhibition assay. Several bisphosphonates were found to have large therapeutic indices (or =200:1) as well as low IC(50) values, suggesting their further investigation as antiparasitic agents against T. b. rhodesiense.

We report the inhibition of a human recombinant geranylgeranyl diphosphate synthase (GGPPSase) by 23 bisphosphonates and six azaprenyl diphosphates. The IC50 values range from 140 nM to 690 microM. None of the nitrogen-containing bisphosphonates that inhibit farnesyl diphosphate synthase were effective in inhibiting the GGPPSase enzyme. Using three-dimensional quantitative structure-activity relationship/comparative molecular field analysis (CoMFA) methods, we find a good correlation between experimental and predicted activity: R2 = 0.938, R(cv)2 = 0.900, R(bs)2 = 0.938, and F-test = 86.8. To test the predictive utility of the CoMFA approach, we used three training sets of 25 compounds each to generate models to predict three test sets of three compounds. The rms pIC50 error for the nine predictions was 0.39. We also investigated the pharmacophore of these GGPPSase inhibitors using the Catalyst method. The results demonstrated that Catalyst predicted the pIC50 values for the nine test set compounds with an rms error of 0.28 (R2 between experimental and predicted activity of 0.948).

We report the cloning and sequencing of a gene encoding the farnesyl pyrophosphate synthase of Trypanosoma cruzi. The protein (T. cruzi farnesyl pyrophosphate synthase, TcFPPS) is an attractive target for drug development, since the growth of T. cruzi is inhibited by carbocation transition state/reactive intermediate analogs of its substrates, the nitrogen-containing bisphosphonates currently in use in bone resorption therapy. The protein predicted from the nucleotide sequence of the gene has 362 amino acids and a molecular mass of 41.2 kDa. Several sequence motifs found in other FPPSs are present in TcFPPS. Heterologous expression of TcFPPS in Escherichia coli produced a functional enzyme that was inhibited by the nitrogen-containing bisphosphonates alendronate, pamidronate, homorisedronate, and risedronate but was less sensitive to the non-nitrogen-containing bisphosphonate etidronate, which, unlike the nitrogen-containing bisphosphonates, does not affect parasite growth. The protein contains a unique 11-mer insertion located near the active site, together with other sequence differences that may facilitate the development of novel anti-Chagasic agents.

We have investigated the effects in vitro of a series of bisphosphonates on the proliferation of Trypanosoma cruzi, Trypanosoma brucei rhodesiense, Leishmania donovani, Toxoplasma gondii, and Plasmodium falciparum. The results show that nitrogen-containing bisphosphonates of the type used in bone resorption therapy have significant activity against parasites, with the aromatic species having in some cases nanomolar or low-micromolar IC(50) activity values against parasite replication (e.g. o-risedronate, IC(50) = 220 nM for T. brucei rhodesiense; risedronate, IC(50) = 490 nM for T. gondii). In T. cruzi, the nitrogen-containing bisphosphonate risedronate is shown to inhibit sterol biosynthesis at a pre-squalene level, most likely by inhibiting farnesylpyrophosphate synthase. Bisphosphonates therefore appear to have potential in treating parasitic protozoan diseases.

Capillary flow techniques have been used to measure D T , the translational diffusion constant at temperature T , for three transition metal complexes in several low-viscosity liquids. The translational radius, r t , for each of the complexes was obtained from D T using the Stokes–Einstein relation. The common value of r t of Ni(mnt) 2 − in n -butyl alcohol, acetonitrile (ACN), acetone, and ethyl alcohol (EtOH) indicates that the apparently different rotational radii (from ESR) in a series of polar solvents are due to solute–solvent interactions and not to solvation or ion pairing. Another complex, Ni(mnt) 2 2− , was also studied in ACN; our values of D T for Ni(mnt) 2 2− and Ni(mnt) 2 − in ACN are in agreement with electrochemical values. The reproducibility of our techniques was checked by using three different capillaries to make four separate determinations of D T for Ni(mnt) 2 − in EtOH; the values are in good agreement. Values of D T were measured for Mn(Cat–N–SQ) 2 , MnR 2 , in tetrahydrofuran, 2-methyltetrahydrofuran, diethyl ether, and acetone. This complex is of interest because of its screw-propeller geometry. The values of r t in our solvents are in agreement with each other and are consistent with ESR studies of the reorientational motion of MnR 2 .

Class IA phosphoinositide 3-kinase (PI3K) is essential for clonal expansion, differentiation, and effector function of B and T lymphocytes. The p110δ catalytic isoform of PI3K is highly expressed in lymphocytes and plays a prominent role in B and T cell responses. Another class IA PI3K catalytic isoform, p110α, is a promising drug target in cancer but little is known about its function in lymphocytes. Here we used highly selective inhibitors to probe the function of p110α in lymphocyte responses in vitro and in vivo. p110α inhibition partially reduced B cell receptor (BCR)-dependent AKT activation and proliferation, and diminished survival supported by the cytokines BAFF and IL-4. Selective p110δ inhibition suppressed B cell responses much more strongly, yet maximal suppression was achieved by targeting multiple PI3K isoforms. In mouse and human T cells, inhibition of single class IA isoforms had little effect on proliferation, whereas pan-class I inhibition did suppress T cell expansion. In mice, selective p110α inhibition using the investigational agent MLN1117 (previously known as INK1117) did not disrupt the marginal zone B cell compartment and did not block T cell-dependent germinal center formation. In contrast, the selective p110δ inhibitor IC87114 strongly suppressed germinal center formation and reduced marginal zone B cell numbers, similar to a pan-class I inhibitor. These findings show that although acute p110α inhibition partially diminishes AKT activation, selective p110α inhibitors are likely to be less immunosuppressive in vivo compared with p110δ or pan-class I inhibitors.

Two experiments were conducted to examine the effect of self-monitoring (SM) strategies on motor performance of varied difficulty. In a pilot test, participants’ perceptions of task difficulty agreed with performance on the easy task. Participants perceived the hard task to be significantly more difficult than indicated by the performance scores and perceived the easy task to be significantly less difficult than their performance on the complex task (p < .05). In the subsequent experiment, subjects performed 90 trials on either the difficult or easy motor task using either positive self-monitoring (PSM), negative self-monitoring (NSM), or no self-monitoring. MANOVAs indicated that PSM resulted in superior performance in comparison to NSM across trials while performing the difficult task (p < .05). In the easy task, PSM was inferior to NSM on motor performance across trials (p < .01). Further results also indicated that negative affect significantly decreased for PSM performing the difficult task, and for NSM performing the easy task.

The mechanistic target of rapamycin (mTOR) is a serine/threonine kinase whose activity contributes to leukemia proliferation and survival. Compounds targeting the mTOR active site inhibit rapamycin-resistant functions and have enhanced anti-cancer activity in mouse models. MLN0128 (formerly known as INK128) is a novel, orally active mTOR kinase inhibitor currently in clinical development. Here we evaluated MLN0128 in preclinical models of B-cell acute lymphoblastic leukemia (B-ALL). MLN0128 suppressed proliferation of B-ALL cell lines in vitro and reduced colony formation by primary human leukemia cells from adult and pediatric B-ALL patients. MLN0128 also boosted the efficacy of dasatinib in Philadelphia Chromosome-positive (Ph+) specimens. In a syngeneic mouse model of lymphoid BCR-ABL+ disease, daily oral dosing of MLN0128 rapidly cleared leukemic outgrowth. In primary xenografts of Ph+ B-ALL specimens, MLN0128 significantly enhanced the efficacy of dasatinib. In non-Ph B-ALL xenografts, single agent MLN0128 had a cytostatic effect that was most pronounced in mice with low disease burden. In all in vivo models, MLN0128 was well tolerated and did not suppress endogenous bone marrow proliferation. These findings support the rationale for clinical testing of MLN0128 in both adult and pediatric B-ALL and provide insight towards optimizing therapeutic efficacy of mTOR kinase inhibitors.

The effects in vivo of the bisphosphonate drug pamidronate, used in bone resorption therapy, were investigated in an experimental model of cutaneous leishmaniasis. Pamidronate at an intraperitoneal dose of 10 mg/kg/day for 5 days effects a radical cure of cutaneous leishmaniasis in Balb/c mice, as evidenced by long-term disappearance of lesions; disappearance of amastigotes in lesion sites, as determined by histopathological analysis and cultivation of material obtained from lesions; and polymerase chain reaction analysis of necropsy material, using probes specific for kinetoplast DNA. Pamidronate is, therefore, a new lead compound for the synthesis of drugs effective against cutaneous leishmaniasis.

The discovery of second generation N-hydroxyurea 5-lipoxygenase inhibitors was accomplished through the development of a broad structure−activity relationship (SAR) study. This study identified requirements for improving potency and also extending duration by limiting metabolism. Potency could be maintained by the incorporation of heterocyclic templates substituted with selected lipophilic substituents. Duration of inhibition after oral administration was optimized by identification of structural features in the proximity of the N-hydroxyurea which correlated to low in vitro glucuronidation rates. Furthermore, the rate of in vitro glucuronidation was shown to be stereoselective for certain analogs. (R)-N-[3-[5-(4-Fluorophenoxy)-2-furyl]-1-methyl-2-propynyl]-N-hydroxyurea (17c) was identified and selected for clinical development.

Firm breast compression during film-screen mammography is necessary to achieve optimum image quality while minimizing radiation dose. Of 374 women who fully completed a questionnaire following mammography, 225 (60%) reported no pain, 115 (31%) moderate pain and only 3 (1%) reported severe pain. Only one patient stated that the pain from the procedure would prevent her from having a further mammogram. Underlying breast disease (usually fibrocystic disease) is associated with a greater incidence and severity of breast pain, but, no relationship has been demonstrated with regards to the patient's age, hormonal status, menstruation or caffeine intake. The high level of acceptance of firm compression by women in our study indicates that undue concern regarding patient discomfort should not deter people from referral for mammography or from the application of firm compression.

Targeting the mammalian target of rapamycin (mTOR) protein is a promising strategy for cancer therapy. The mTOR kinase functions in two complexes, TORC1 (target of rapamycin complex-1) and TORC2 (target of rapamycin complex-2); however, neither of these complexes is fully inhibited by the allosteric inhibitor rapamycin or its analogs. We compared rapamycin with PP242, an inhibitor of the active site of mTOR in both TORC1 and TORC2 (hereafter referred to as TORC1/2), in models of acute leukemia harboring the Philadelphia chromosome (Ph) translocation. We demonstrate that PP242, but not rapamycin, causes death of mouse and human leukemia cells. In vivo, PP242 delays leukemia onset and augments the effects of the current front-line tyrosine kinase inhibitors more effectively than does rapamycin. Unexpectedly, PP242 has much weaker effects than rapamycin on the proliferation and function of normal lymphocytes. PI-103, a less selective TORC1/2 inhibitor that also targets phosphoinositide 3-kinase (PI3K), is more immunosuppressive than PP242. These findings establish that Ph(+) transformed cells are more sensitive than normal lymphocytes to selective TORC1/2 inhibitors and support the development of such inhibitors for leukemia therapy.

High-resolution 303.6 MHz 31 P NMR spectra have been obtained of perchloric acid extracts of Plasmodium berghei trophozoites, Toxoplasma gondii tachyzoites, and Cryptosporidium parvum oocysts. Essentially complete resonance assignments have been made based on chemical shifts and by coaddition of authentic reference compounds. Signals corresponding to inorganic pyrophosphate were detected in all three species. In T. gondii and C. parvum, additional resonances were observed corresponding to linear triphosphate as well as longer chain polyphosphates. Spectra of P. berghei and T. gondii also indicated the presence of phosphomonoesters and nucleotide phosphates. We also report that the pyrophosphate analog drug, risedronate (used in bone resorption therapy), inhibits the growth of C. parvum in a mouse xenograft model. When taken together, our results indicate that all the major disease-causing apicomplexan parasites contain extensive stores of condensed phosphates and that as with Plasmodium falciparum and T. gondii, the pyrophosphate analog drug risedronate is an inhibitor of C. parvum cell growth.

Nitrogen-containing bisphosphonates are potent bone antiresorptive agents as well as having herbicidal and antiparasitic activity, and are thought to act by inhibiting enzymes of the mevalonate pathway. Using molecular modeling and ab initio quantum chemical calculations, we show that bisphosphonates can act as aza-isoprenoid transition state analogs, thereby inhibiting isoprenoid biosynthesis. The two phosphonate groups of the 1,1-bisphosphonates readily dock into the diphosphate-Mg(2+) binding site in farnesyl diphosphate synthase, while the charged ammonium (or pyridinium or imidazolium) groups act as carbocation transition state analogs, whose binding is stabilized by a cluster of oxygen atoms in the active site cleft, and an overall negative electrostatic potential in this region. Enhanced activity is shown to correlate with increasing van der Waals stabilization due to N-alkylation, or the presence of a charged, planar (sp(2)-hybridized) aromatic residue in the carbocation binding site. These results are of general interest since they suggest a rational approach to bisphosphonate drug design.

The discovery of second generation N-hydroxyurea 5-lipoxygenase inhibitors was accomplished through the development of a broad structure-activity relationship (SAR) study. This study identified requirements for improving potency and also extending duration by limiting metabolism. Potency could be maintained by the incorporation of heterocyclic templates substituted with selected lipophilic substituents. Duration of inhibition after oral administration was optimized by identification of structural features in the proximity of the N-hydroxyurea which correlated to low in vitro glucuronidation rates. Furthermore, the rate of in vitro glucuronidation was shown to be stereoselective for certain analogs. (R)-N-[3-[5-(4-Fluorophenoxy)-2-furyl]-1-methyl-2-propynyl]-N-hydroxyure a (17c) was identified and selected for clinical development.

The aim of this study was to determine the effectiveness of implementing different self-monitoring (SM) strategies as a function of varying task complexity on a computer game, 'Shufflepuck'. Self-monitoring theory suggests that positive self-monitoring (PSM) results in better performance for difficult tasks, whereas negative self-monitoring (NSM) is preferable for easy tasks. When SM was tested under easy and complex task conditions, it was found that performance effectiveness was a function of matching appropriate task difficulty with the SM strategy. Unpaired t-tests compared the first set of 100 trials (pre-task) with the second set of 100 trials (post-task) on a computer skills game requiring hand-eye coordination. The results showed that subjects who changed from the difficult task/PSM condition to the easy task/PSM condition (different task, same SM strategy) performed significantly poorer (P < 0.001) than subjects who changed from the difficult task/PSM condition to the easy task/NSM condition (different task, different SM strategy). A second unpaired t-test comparing the two sets of difference performance scores indicated that performance was significantly superior for subjects using the PSM than the NSM strategy while performing the difficult task (P < 0.05). That is, changing from the easy task/NSM condition to the difficult task/NSM condition produced poorer performance than changing from the easy task/NSM to the difficult task/PSM condition. In terms of subjects' emotions, the use of NSM and PSM when performing easy and difficult tasks, respectively, resulted in markedly less negative affect (P < 0.01), but did not influence positive affect, on pre- and post-task comparisons. Finally, correlations between affect and performance, based on pre- and post-performance and affect scores, were moderate to high. Implications of the present results for slumps in sport performance are offered.

Indole compounds substituted at the 3-position by a 7-carbonyl(pyridin-3-yl)pyrrolo[1,2-c]thiazole, 7-carbonyl(pyridin-3-yl)pyrrolo[1,2-c]oxazoline, or 7-carbonyl(pyridin-3-yl)pyrrolo[1,2-c]pyrrole group are potent antagonists of PAF and are useful in the treatment of PAF-related disorders including asthma, shock, respiratory distress syndrome, acute inflammation, transplanted organ rejection, gastrointestinal ulceration, allergic skin diseases, delayed cellular immunity, parturition, fetal lung maturation, and cellular differentiation.

procedures on catheterization, it may be difficult to introduce the poorly tapered 7 French balloon catheter into the artery without damaging the artery or catheter tip. Use of an 8 French sheath (USCI) may facilitate passage of the dilating balloon catheter. Unfortunately, it may also be difficult to pass tht sheath into the artery because the dilator supplied with the sheath is often too short and inadequately tapered to allow the sheath to negotiate the acute entry angle into the artery. We have found that replacing the standard 8 French sheath dilator with an 8 French, 80 cm, Teflon Van Andel dilating catheter (Cook) greatly facilitates placement of the sheath (fig. 1 ). The Van Andel catheter can be easily seated over a guide wire into the artery even in patients who are markedly obese and with significant inguinal scarring. The 8 French sheath fits tightly over the Van Andel dilating catheter and readily passes along with it into the common femonal artery.