ABSTRACT
Genitourinary cancers encompass some of the most common solid tumours and have high rates of morbidity and mortality. Inflammation is associated with enhanced tumorigenesis, and a number of pro-inflammatory mediators, such as macrophage migration inhibitory factor (MIF), also promote tumorigenesis. Studies of the role of MIF (which largely functions via the type II transmembrane receptor CD74) in prostate, bladder and kidney cancers suggest that it is a pro-tumorigenic factor in genitourinary malignancy. Inhibiting MIF activity in cell culture and in preclinical animal models of genitourinary cancers reduces the phenotypic hallmarks of cancer, such as proliferation, angiogenesis and tumour aggressiveness, by downregulating signalling pathways such as those regulated by extracellular signal-regulated kinase (ERK), protein kinase B and p53, and MIF may also reverse immunosuppression. Progress has been made in our understanding of the role of MIF (and its family member D-dopachrome tautomerase (DDT)) in genitourinary cancers and how it can be therapeutically targeted.
Subject(s)
Carcinogenesis , Macrophage Migration-Inhibitory Factors/physiology , Urogenital Neoplasms/pathology , Urogenital Neoplasms/therapy , HumansABSTRACT
Macrophage migratory inhibitory factor (MIF) is a proinflammatory cytokine shown to promote tumorigenesis. Using the N-butyl-N-(4-hydroxybutyl)-nitrosamine (BBN) model of bladder cancer, we previously showed that MIF knockout mice display decreased angiogenesis and invasion compared with wild-type. This study examines the role of MIF in bladder cancer via use of oral inhibitors of MIF. In vitro, high-grade bladder cancer cells were treated with recombinant human MIF +/- (rhMIF+/-) inhibitor. Measurements included cell counts, proliferation by (3)H-thymidine incorporation (TdR), extracellular signal-regulated kinase (ERK) phosphorylation by western blot analysis, messenger RNA (mRNA) expression by quantitative PCR and protein secretion by enzyme-linked immunosorbent assay. Treatment with rhMIF increased ERK phosphorylation, cell counts, TdR and mRNA expression and protein secretion of vascular endothelial growth factor, which were blocked by specific inhibitors of ERK and MIF. In vivo, 3-month-old male C57Bl/6 mice were given BBN for 22 and 16 weeks in study 1 and study 2, respectively. Mice (n = 8-10 per group) were gavaged with vehicle or doses of MIF inhibitors daily from weeks 16-22 in both studies. Average bladder weights, reflecting tumor mass, tumor stage/burden, mitotic rate and proliferation indices, and microvessel densities were reduced in inhibitor groups versus controls. In summary, MIF promotes bladder cancer via increasing cell proliferation and angiogenesis and oral inhibitors of MIF may prove useful in treatment of this disease.
Subject(s)
Macrophage Migration-Inhibitory Factors/genetics , Macrophage Migration-Inhibitory Factors/metabolism , Neovascularization, Pathologic/pathology , Urinary Bladder Neoplasms/pathology , Animals , Cell Line, Tumor , Cell Proliferation , Disease Progression , Hep G2 Cells , Humans , MAP Kinase Signaling System/genetics , Macrophage Migration-Inhibitory Factors/antagonists & inhibitors , Male , Mice , Mice, Inbred C57BL , Neovascularization, Pathologic/genetics , Phosphorylation/genetics , Proto-Oncogene Proteins c-akt/genetics , Proto-Oncogene Proteins c-akt/metabolism , RNA, Messenger , Urinary Bladder/metabolism , Urinary Bladder/pathology , Urinary Bladder Neoplasms/genetics , Urinary Bladder Neoplasms/metabolism , Vascular Endothelial Growth Factor A/genetics , Vascular Endothelial Growth Factor A/metabolismABSTRACT
Pyrido[3,2-b]pyrazin-3(4H)-ones and pteridin-7(8H)-ones were evaluated as corticotropin-releasing factor-1 receptor antagonists. The synthesis, SAR studies and pharmacokinetic evaluation of these analogs are described herein.
Subject(s)
Pteridines/chemistry , Pyrazines/chemistry , Receptors, Corticotropin-Releasing Hormone/antagonists & inhibitors , Animals , Anti-Anxiety Agents/chemical synthesis , Anti-Anxiety Agents/chemistry , Anti-Anxiety Agents/pharmacokinetics , Half-Life , Pteridines/chemical synthesis , Pteridines/pharmacokinetics , Pyrazines/chemical synthesis , Pyrazines/pharmacokinetics , Rats , Receptors, Corticotropin-Releasing Hormone/metabolism , Structure-Activity RelationshipABSTRACT
The oncoprotein hdm2 ubiquitinates p53, resulting in the rapid degradation of p53 through the ubiquitin (Ub)-proteasome pathway. Hdm2-mediated destabilization and inactivation of p53 are thought to play a critical role in a number of human cancers. We have used an in vitro enzyme assay, monitoring hdm2-catalyzed Ub transfer from preconjugated Ub-Ubc4 to p53, to identify small molecule inhibitors of this enzyme. Three chemically distinct types of inhibitors were identified this way, each with potency in the micromolar range. All three types of compounds display selective inhibition of hdm2 E3 ligase activity, with little or no effect on other Ub-using enzymes. Most strikingly, these compounds do not inhibit the autoubiquitination activity of hdm2. Steady-state analysis reveals that all three classes behave as simple reversible inhibitors of the enzyme and that they are noncompetitive with respect to both substrates, Ub-Ubc4 and p53. Studies of the effects of combinations of two inhibitory molecules on hdm2 activity indicate that the three types of compounds bind in a mutually exclusive fashion, suggesting a common binding site on hdm2 for all of these inhibitors. These compounds establish the feasibility of selectively blocking hdm2-mediated ubiquitination of p53 by small molecule inhibitors. Selective inhibitors of hdm2 E3 ligase activity could provide a novel mechanism for the development of new chemotherapeutics for the treatment of human cancers.
