manumycin and Disease-Models--Animal

No effective treatment was available for myotonic dystrophy, even in animal model. We have established a new antisense oligonucleotide delivery to skeletal muscle of mice with bubble liposomes, and led to increased expression of chloride channel (CLCN1) protein and the amelioration of myotonia. In other experiments, we also identified small molecule compounds that correct aberrant splicing of Clcn1 gene. Manumycin A corrected aberrant splicing of Clcn1 in mouse model.

Topics: Animals; Chloride Channels; Disease Models, Animal; Gene Transfer Techniques; Genetic Therapy; Humans; Liposomes; Mice; Molecular Targeted Therapy; Muscle, Skeletal; Myotonic Dystrophy; Oligonucleotides, Antisense; Polyenes; Polyunsaturated Alkamides; RNA Splicing

The chemopreventive effect of RAS inhibitors on colorectal cancer is unknown. Because aberrant crypt foci (ACF), earliest preneoplastic lesions, are highly positive for K-RAS mutation, RAS inhibitors are likely to be effective for chemoprevention. Therefore, in the present study, the suppressive effect of a RAS inhibitor, manumycin A, on ACF formation in an azoxymethane (AOM)-induced rat colorectal carcinogenesis model was investigated.. Rats injected with AOM were administered manumycin A (30 mg/kg) subcutaneously thrice weekly for 8 weeks or for 4 weeks (latter half), sacrificed at 8 weeks, and examined for ACF in the colorectum. Phosphorylated ERK and Ki-67 expression was evaluated by immunohistochemistry. Apoptosis was assessed by TUNEL staining.. The mean number of ACF in the 8-week manumycin A group (72.9 ± 20.1) was significantly lower than in the vehicle group (155.6 ± 56.7, P < 0.01), and it was significantly lower even in the 4-week manumycin A group than in the vehicle group (92.2 ± 13.0 vs 222.3 ± 83.3, P < 0.01). The positive rate for phosphorylated ERK in the manumycin A group (13.5 ± 19.2%) was significantly lower than in the vehicle group (50.2 ± 19.8%, P < 0.01). The positive rate for Ki-67 in the manumycin A group (2.2 ± 3.4%) was significantly lower than in the vehicle group (14.7 ± 8.2%, P < 0.01). There were significantly more terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate nick-end labeling-positive cells in tissue samples from the manumycin A group versus the vehicle group (8.6 ± 9.7% vs 2.9 ± 2.0%, P < 0.05).. Manumycin A suppressed ACF formation in the AOM-induced colorectal carcinogenesis model, demonstrating that RAS inhibitors may be very effective for chemoprevention of colorectal cancers.

Topics: Aberrant Crypt Foci; Animals; Apoptosis; Azoxymethane; Colorectal Neoplasms; Disease Models, Animal; Enzyme Inhibitors; Extracellular Signal-Regulated MAP Kinases; Farnesyltranstransferase; Genes, ras; Injections, Subcutaneous; Ki-67 Antigen; Mutation; Phosphorylation; Polyenes; Polyunsaturated Alkamides; ras Proteins; Rats; Rats, Inbred F344

RhoA and Rho kinase contribute to pulmonary vasoconstriction and vascular remodeling in pulmonary hypertension. RhoB, a protein homologous to RhoA and activated by hypoxia, regulates neoplastic growth and vasoconstriction but its role in the regulation of pulmonary vascular function is not known.. To determine the role of RhoB in pulmonary endothelial and smooth muscle cell responses to hypoxia and in pulmonary vascular remodeling in chronic hypoxia-induced pulmonary hypertension.. Hypoxia increased expression and activity of RhoB in human pulmonary artery endothelial and smooth muscle cells, coincidental with activation of RhoA. Hypoxia or adenoviral overexpression of constitutively activated RhoB increased actomyosin contractility, induced endothelial permeability, and promoted cell growth; dominant negative RhoB or manumycin, a farnesyltransferase inhibitor that targets the vascular function of RhoB, inhibited the effects of hypoxia. Coordinated activation of RhoA and RhoB maximized the hypoxia-induced stress fiber formation caused by RhoB/mammalian homolog of Drosophila diaphanous-induced actin polymerization and RhoA/Rho kinase-induced phosphorylation of myosin light chain on Ser19. Notably, RhoB was specifically required for hypoxia-induced factor-1α stabilization and for hypoxia- and platelet-derived growth factor-induced cell proliferation and migration. RhoB deficiency in mice markedly attenuated development of chronic hypoxia-induced pulmonary hypertension, despite compensatory expression of RhoA in the lung.. RhoB mediates adaptational changes to acute hypoxia in the vasculature, but its continual activation by chronic hypoxia can accentuate vascular remodeling to promote development of pulmonary hypertension. RhoB is a potential target for novel approaches (eg, farnesyltransferase inhibitors) aimed at regulating pulmonary vascular tone and structure.

Topics: Actomyosin; Animals; Capillary Permeability; Cell Hypoxia; Cell Movement; Cell Proliferation; Cells, Cultured; Chronic Disease; Disease Models, Animal; Endothelial Cells; Enzyme Activation; Enzyme Inhibitors; Familial Primary Pulmonary Hypertension; Farnesyltranstransferase; Humans; Hypertension, Pulmonary; Hypoxia; Hypoxia-Inducible Factor 1, alpha Subunit; Male; Mice; Mice, Inbred C57BL; Mice, Knockout; Muscle, Smooth, Vascular; Myocytes, Smooth Muscle; Myosin Light Chains; Phosphorylation; Polyenes; Polyunsaturated Alkamides; Pulmonary Artery; rhoA GTP-Binding Protein; rhoB GTP-Binding Protein; RNA Interference; Serine; Stress Fibers; Time Factors; Transfection; Vasoconstriction

Hepatotoxicity due to overdose of the analgesic and antipyretic acetaminophen (APAP) is a major cause of liver failure in adults. To better understand the contributions of different signaling pathways, the expression and role of Ras activation was evaluated after oral dosing of mice with APAP (400-500 mg/kg). Ras-guanosine triphosphate (GTP) is induced early and in an oxidative stress-dependent manner. The functional role of Ras activation was studied by a single intraperitoneal injection of the neutral sphingomyelinase and farnesyltransferase inhibitor (FTI) manumycin A (1 mg/kg), which lowers induction of Ras-GTP and serum amounts of alanine aminotransferase (ALT). APAP dosing decreases hepatic glutathione amounts, which are not affected by manumycin A treatment. However, APAP-induced activation of c-Jun N-terminal kinase, which plays an important role, is reduced by manumycin A. Also, APAP-induced mitochondrial reactive oxygen species are reduced by manumycin A at a later time point during liver injury. Importantly, the induction of genes involved in the inflammatory response (including iNos, gp91phox, and Fasl) and serum amounts of proinflammatory cytokines interferon-gamma (IFNgamma) and tumor necrosis factor alpha, which increase greatly with APAP challenge, are suppressed with manumycin A. The FTI activity of manumycin A is most likely involved in reducing APAP-induced liver injury, because a specific neutral sphingomyelinase inhibitor, GW4869 (1 mg/kg), did not show any hepatoprotective effect. Notably, a structurally distinct FTI, gliotoxin (1 mg/kg), also inhibits Ras activation and reduces serum amounts of ALT and IFN-gamma after APAP dosing. Finally, histological analysis confirmed the hepatoprotective effect of manumycin A and gliotoxin during APAP-induced liver damage.. This study identifies a key role for Ras activation and demonstrates the therapeutic efficacy of FTIs during APAP-induced liver injury.

Topics: Acetaminophen; Alanine Transaminase; Analgesics, Non-Narcotic; Animals; Chemical and Drug Induced Liver Injury; Disease Models, Animal; Dose-Response Relationship, Drug; Enzyme Inhibitors; Farnesyltranstransferase; Gliotoxin; Guanosine Triphosphate; Interferon-gamma; MAP Kinase Kinase 4; Mice; Mice, Inbred BALB C; Oxidative Stress; Polyenes; Polyunsaturated Alkamides; ras Proteins; Tumor Necrosis Factor-alpha

Statins are presumed to exert their antiatherogenic effects in part via lipid-lowering-independent mechanisms. Inhibition of protein farnesylation and/or geranylgeranylation by statins has been postulated to contribute to the lipid-lowering-independent effects. However, a role for protein farnesylation in atherogenesis has not yet been studied. Therefore, we examined the effects of farnesyltransferase inhibitor, manumycin A, on the development of atherosclerosis in apolipoprotein E (apoE)-deficient mice fed a high-fat diet.. Manumycin A treatment for 22 weeks decreased Ras activity, and reduced fatty streak lesion size at the aortic sinus to 43% of that in vehicle-treated apoE-deficient mice (P<0.05), while plasma total cholesterol was unaltered. Moreover, manumycin A reduced alpha-smooth muscle actin-positive area to 29% of that in vehicle-treated apoE-deficient mice (P<0.01). The prevention of atherogenesis by manumycin A was accompanied by amelioration of oxidative stress, as judged by reduced ex vivo superoxide production and nitrotyrosine immunoreactivity.. These results indicate that the inhibition of farnesyltransferase prevents the development of mature atherosclerosis with concomitant alleviation of oxidative stress in apoE-deficient mice. The present data highlight farnesyltransferase as a potential molecular target for preventive and/or therapeutic intervention against atherosclerosis.

Topics: Animals; Apolipoproteins E; Atherosclerosis; Dietary Fats; Disease Models, Animal; Enzyme Inhibitors; Farnesyltranstransferase; Male; Mice; Mice, Inbred C57BL; Mice, Knockout; Muscle, Smooth, Vascular; Oxidative Stress; Phosphorylation; Polyenes; Polyunsaturated Alkamides; Protein Prenylation; Proto-Oncogene Proteins c-raf; ras Proteins; Sinus of Valsalva; Superoxides; Time Factors; Tyrosine

The studies presented were designed to test the efficacy of farnesyltransferase inhibitors (FTIs) as potential chemopreventive compounds in the mouse lung tumor model, and in tumor cell lines. The compounds included manumycin, gliotoxin, dihydroepiandrosterone (DHEA), perillyl alcohol (POH), and FTI-276. Each of these compounds had the potential, based on in vitro and limited in vivo evidence, to inhibit mouse lung tumorigenesis. In vitro studies were conducted with both K-ras-transformed NIH-3T3 cells and mouse lung tumor epithelial cell lines. We utilized 2 primary mouse lung tumor models that reliably produce lung tumors with an oncogenic K-ras mutation when induded by 4-(methylnitrosoamino)-1-(3-pyridyl)-1-butanone (NNK). Manumycin, gliotoxin, DHEA, and POH were administered 3 times per week peritoneally (i.p.), starting 1 week prior to carcinogen treatment, and throughout the test period (4.5 months). FTI-276 was delivered daily for 4 months by a time-release pellet method. Both the manumycin and gliotoxin treatment groups demonstrated 100% incidence and an increase in tumor multiplicity over control, of 66% and 58% increase respectively (P < .05). Although DHEA showed no significant chemopreventive effect, POH treatment demonstrated a 22% reduction in tumor incidence (P < .05) and a 58% reduction in tumor multiplicity (P < .05). Finally, FTI-276 reduced both the tumor multiplicity by 41.7% (P < .005), and the total tumor volume/burden per mouse by 79.4% (P < .0001). The apoptotic index in FTI-276-treated tumors showed an increase of 77% over control tumors (P < .05). In vitro, all compounds demonstrated growth inhibition at a dose-response manner; however, manumycin, gliotoxin, and DHEA demonstrated an initial increase in growth rate at lower doses. In summary, we have shown that POH and FTI-276 are chemopreventive in a primary mouse lung tumor model. In contrast, DHEA was not significantly chemopreventive at the dosage utilized, and treatment of an immunocompetent host with manumycin or gliotoxin demonstrated a significant increase in tumorigenicity over carcinogen control.

Topics: 3T3 Cells; Adenoma; Alkyl and Aryl Transferases; Animals; Apoptosis; Chemoprevention; Dehydroepiandrosterone; Disease Models, Animal; DNA, Neoplasm; Dose-Response Relationship, Drug; Enzyme Inhibitors; Farnesyltranstransferase; Fluorescent Antibody Technique, Indirect; Gliotoxin; In Situ Nick-End Labeling; Lung Neoplasms; Methionine; Mice; Mice, Inbred A; Mice, Inbred C3H; Monoterpenes; Polyenes; Polymerase Chain Reaction; Polyunsaturated Alkamides; Proliferating Cell Nuclear Antigen; Terpenes