ABSTRACT
Mutations resulting in progranulin haploinsufficiency cause disease in patients with a subset of frontotemporal lobar degeneration; however, the biological functions of progranulin in the brain remain unknown. To address this subject, the present study initially assessed changes in gene expression and cytokine secretion in rat primary cortical neurons treated with progranulin. Molecular pathways enriched in the progranulin gene set included cell adhesion and cell motility pathways and pathways involved in growth and development. Secretion of cytokines and several chemokines linked to chemoattraction but not inflammation were also increased from progranulin-treated primary neurons. Therefore, whether progranulin is involved in recruitment of immune cells in the brain was investigated. Localized lentiviral expression of progranulin in C57BL/6 mice resulted in an increase of Iba1-positive microglia around the injection site. Moreover, progranulin alone was sufficient to promote migration of primary mouse microglia in vitro. Primary microglia and C4B8 cells demonstrated more endocytosis of amyloid ß1-42 when treated with progranulin. These data demonstrate that progranulin acts as a chemoattractant in the brain to recruit or activate microglia and can increase endocytosis of extracellular peptides such as amyloid ß.
Subject(s)
Brain/physiology , Chemotactic Factors/physiology , Endocytosis , Intercellular Signaling Peptides and Proteins/physiology , Microglia/physiology , Animals , Brain/drug effects , Brain/metabolism , Calcium-Binding Proteins/metabolism , Cell Line, Tumor , Cell Movement , Chemotactic Factors/genetics , Chemotactic Factors/pharmacology , Cytokines/metabolism , Endocytosis/drug effects , Humans , Intercellular Signaling Peptides and Proteins/genetics , Intercellular Signaling Peptides and Proteins/pharmacology , Mice , Mice, Mutant Strains , Microfilament Proteins , Microglia/drug effects , Microglia/metabolism , Neurons/drug effects , Neurons/metabolism , Neurons/physiology , Progranulins , RatsABSTRACT
Synthesis, SAR and evaluation of styrenyl quinazolinones as novel gamma secretase modulators are presented in this communication. Starting from literature and in-house leads we evaluated a range of quinazolinones which showed good modulation of γ-secretase activity.
Subject(s)
Amyloid Precursor Protein Secretases/metabolism , Amyloid beta-Peptides/antagonists & inhibitors , Quinazolinones/chemistry , Quinazolinones/pharmacology , Alzheimer Disease/drug therapy , Alzheimer Disease/enzymology , Alzheimer Disease/metabolism , Amyloid beta-Peptides/metabolism , Animals , Cell Line , Humans , Inhibitory Concentration 50 , Peptide Fragments/antagonists & inhibitors , Peptide Fragments/metabolism , Protein Binding , Quinazolinones/pharmacokinetics , Rats , Rats, Sprague-DawleyABSTRACT
The development of a novel series of purines as gamma-secretase modulators for potential use in the treatment of Alzheimer's disease is disclosed herein. Optimization of a previously disclosed pyrimidine series afforded a series of potent purine-based gamma-secretase modulators with 300- to 2000-fold in vitro selectivity over inhibition of Notch cleavage and that selectively reduces Alphabeta42 in an APP-YAC transgenic mouse model.
Subject(s)
Alzheimer Disease/drug therapy , Amyloid Precursor Protein Secretases/metabolism , Amyloid beta-Peptides/antagonists & inhibitors , Peptide Fragments/antagonists & inhibitors , Purines/chemistry , Purines/therapeutic use , Amyloid Precursor Protein Secretases/genetics , Amyloid beta-Peptides/metabolism , Animals , Humans , Mice , Mice, Transgenic , Peptide Fragments/metabolism , Purines/pharmacology , Receptors, Notch/metabolism , Structure-Activity RelationshipABSTRACT
We report herein a novel series of difluoropiperidine acetic acids as modulators of gamma-secretase. Synthesis of 2-aryl-3,3-difluoropiperidine analogs was facilitated by a unique and selective beta-difluorination with Selectfluor. Compounds 1f and 2c were selected for in vivo assessment and demonstrated selective lowering of Abeta42 in a genetically engineered mouse model of APP processing. Moreover, in a 7-day safety study, rats treated orally with compound 1f (250mg/kg per day, AUC(0-24)=2100microMh) did not exhibit Notch-related effects.
Subject(s)
Acetates/chemistry , Amyloid Precursor Protein Secretases/metabolism , Fluorine/chemistry , Piperidines/chemistry , Acetates/chemical synthesis , Acetates/pharmacokinetics , Amyloid beta-Peptides/genetics , Amyloid beta-Peptides/metabolism , Animals , Diazonium Compounds/chemistry , Disease Models, Animal , Mice , Mice, Transgenic , Peptide Fragments/genetics , Peptide Fragments/metabolism , Piperidines/chemical synthesis , Piperidines/pharmacokinetics , Rats , Receptors, Notch/metabolismABSTRACT
The development of a novel series of piperazinyl pyrimidines as gamma-secretase modulators for potential use in the treatment of Alzheimer's disease is disclosed herein. Optimization of a screening hit provided a series of potent gamma-secretase modulators with >180-fold in vitro selectivity over inhibition of Notch cleavage.
Subject(s)
Amyloid Precursor Protein Secretases/metabolism , Piperazines/chemistry , Pyrimidines/chemistry , Amyloid Precursor Protein Secretases/antagonists & inhibitors , Amyloid beta-Peptides/antagonists & inhibitors , Amyloid beta-Peptides/metabolism , Amyloid beta-Peptides/physiology , Cell Line, Tumor , HeLa Cells , Humans , Peptide Fragments/antagonists & inhibitors , Peptide Fragments/metabolism , Piperazines/pharmacology , Pyrimidines/pharmacologyABSTRACT
In a previous study we demonstrated a requirement for activation of mTORC1 in the stimulation of eIF2Bepsilon mRNA translation in skeletal muscle in response to resistance exercise. Although that study established the necessity of mTORC1 activation, the experimental model used did not lend itself readily to address the question of whether or not mTORC1 activation was sufficient to produce the response. Therefore, the present study was designed to address the sufficiency of mTORC1 activation, using cultures of Rat2 fibroblasts in which mTORC1 signaling was repressed by serum/leucine-depletion and stimulated by repletion of leucine and/or IGF-1. Repletion with leucine and IGF-1 caused a shift of eIF2Bepsilon mRNA into actively translating polysomes and a stimulation of new eIF2Bepsilon protein synthesis, but had no effect on mRNAs encoding the other four eIF2B subunits. Stimulation of eIF2Bepsilon translation was reversed by pre-treatment with the mTORC1 inhibitor rapamycin. Exogenous overexpression of FLAG-Rheb, a proximal activator of mTORC1, also caused a re-distribution of eIF2Bepsilon mRNA into polysomes and a stimulation of eIF2Bepsilon protein synthesis. The stimulation of eIF2Bepsilon mRNA translation occurred in the absence of any effect on eIF2Bepsilon mRNA abundance. RNAi-mediated knockdown of eIF2Bepsilon resulted in reduced cellular proliferation, a result that phenocopied the known cytostatic effect of mTORC1 repression. Overall the results demonstrate that activation of mTORC1 is both necessary and sufficient to stimulate eIF2Bepsilon mRNA translation and that this response may represent a novel mechanism through which mTORC1 can affect mRNA translation initiation, rates of protein synthesis, and cellular growth/proliferation.
Subject(s)
Eukaryotic Initiation Factor-2B/metabolism , Protein Biosynthesis/physiology , RNA, Messenger/metabolism , Transcription Factors/metabolism , Animals , Antibiotics, Antineoplastic/pharmacology , Cell Proliferation , Cells, Cultured , Eukaryotic Initiation Factor-2B/genetics , Fibroblasts/cytology , Fibroblasts/drug effects , Fibroblasts/physiology , Insulin-Like Growth Factor I/metabolism , Leucine/metabolism , Monomeric GTP-Binding Proteins/genetics , Monomeric GTP-Binding Proteins/metabolism , Neuropeptides/genetics , Neuropeptides/metabolism , Polyribosomes/metabolism , RNA, Messenger/genetics , RNA, Small Interfering/genetics , RNA, Small Interfering/metabolism , Ras Homolog Enriched in Brain Protein , Rats , Sirolimus/pharmacology , Transcription Factors/geneticsABSTRACT
Current trends in the treatment of human tumors are with drug combinations that result in improved responses as well as the ability to use less toxic concentrations of the drugs. Recent reports have shown that COX-2 inhibitors and taxanes are effective in the suppression of human tumor growth. The bisphosphonate, zoledronic acid, primarily used in the treatment of bone metastases, also inhibits proliferation and induces apoptosis in human breast and prostate carcinoma and multiple myeloma. HER-2/neu overexpression has been suggested as a mechanism for resistance to both hormonal therapy as well as chemotherapy. This study examines the effects of combining a cyclooxygenase-2 inhibitor with zoledronic acid and/or docetaxel in a HER-2/neu transfected and control human breast cancer cell line. All three compounds produced dose-dependent growth inhibition in both cell lines. The HER-2/neu transfected MCF/18 cells, however, were less sensitive to zoledronic acid than the control MCF/neo cells, 9% to 53% inhibition and 18% to 67%, respectively. Enhanced growth inhibition was observed in both cell lines with the combination of docetaxel and SC236 and the combination of docetaxel and zoledronic acid. The combination of SC236 with zoledronic acid also gave an enhanced inhibitory effect in the MCF/neo line. This combination, however, was additive in the HER-2/neu transfected MCF/18 cell line. The triple combination of SC236, zoledronic acid and/or docetaxel resulted in a small increase in growth inhibition compared to that seen with the double combinations.
