Neurodegenerative processes in Huntington's disease

Huntington's disease (HD) is a complex and severe disorder characterized by the gradual and the progressive loss of neurons, predominantly in the striatum, which leads to the typical motor and cognitive impairments associated with this pathology. HD is caused by a highly polymorphic CAG trinucleotide repeat expansion in the exon-1 of the gene encoding for huntingtin protein. Since the first discovery of the huntingtin gene, investigations with a consistent number of in-vitro and in-vivo models have provided insights into the toxic events related to the expression of the mutant protein. In this review, we will summarize the progress made in characterizing the signaling pathways that contribute to neuronal degeneration in HD. We will highlight the age-dependent loss of proteostasis that is primarily responsible for the formation of aggregates observed in HD patients. The most promising molecular targets for the development of pharmacological interventions will also be discussed

Reversible phosphorylation in haematological malignancies: Potential role for protein tyrosine phosphatases in treatment?

Most aspects of leukocyte physiology are under the control of reversible tyrosine phosphorylation. It is clear that excessive phosphorylation of signal transduction elements is a pivotal element of many different pathologies including haematological malignancies and accordingly, strategies that target such phosphorylation have clinically been proven highly successful for treatment of multiple types of leukemias and lymphomas. Cellular phosphorylation status is dependent on the resultant activity of kinases and phosphatases. The cell biology of the former is now well understood: for most cellular phosphoproteins we now know the kinases responsible for their phosphorylation and we understand the principles of their aberrant activity in disease. With respect to phosphatases, however, our knowledge is much patchier. Although the sequences of whole genomes allow us to identify phosphatases using in silico methodology, whereas transcription profiling allows us to understand how phosphatase expression is regulated during disease, most functional questions as to substrate specificity, dynamic regulation of phosphatase activity and potential for therapeutic intervention are still to a large degree open. Nevertheless, recent studies have allowed us to make meaningful statements on the role of tyrosine phosphatase activity in the three major signaling pathways that are commonly affected in leukemias, i.e. the Ras-Raf-ERK1/2, the Jak-STAT and the PI3K-PKB-mTOR pathways. Lessons learned from these pathways may well be applicable elsewhere in leukocyte biology as well. (C) 2010 Elsevier B.V. All rights reserved

Low-molecular-weight Protein Tyrosine Phosphatase Predicts Prostate Cancer Outcome By Increasing The Metastatic Potential

Low-risk patients suffering from prostate cancer (PCa) are currently placed under active surveillance rather than undergoing radical prostatectomy. However, clear parameters for selecting the right patient for each strategy are not available, and new biomarkers and treatment modalities are needed. Low-molecular-weight protein tyrosine phosphatase (LMWPTP) could present such a target. Objective: To correlate expression levels of LMWPTP in primary PCa to clinical outcome, and determine the role of LMWPTP in prostate tumor cell biology. Design, setting, and participants: Acid phosphatase 1, soluble (ACP1) expression was analyzed on microarray data sets, which were subsequently used in Ingenuity Pathway Analysis. Immunohistochemistry was performed on a tissue microarray containing material of 481 PCa patients whose clinicopathologic data were recorded. PCa cell line models were used to investigate the role of LMWPTP in cell proliferation, migration, adhesion, and anoikis resistance. Outcome measurements and statistical analysis: The association between LMWPTP expression and clinical and pathologic outcomes was calculated using chi-square correlations and multivariable Cox regression analysis. Functional consequences of LMWPTP overexpression or downregulation were determined using migration and adhesion assays, confocal microscopy, Western blotting, and proliferation assays. Results and limitations: LMWPTP expression was significantly increased in human PCa and correlated with earlier recurrence of disease (hazard ratio [HR]: 1.99; p < 0.001) and reduced patient survival (HR: 1.53; p = 0.04). Unbiased Ingenuity analysis comparing cancer and normal prostate suggests migratory propensities in PCa. Indeed, overexpression of LMWPTP increases PCa cell migration, anoikis resistance, and reduces activation of focal adhesion kinase/paxillin, corresponding to decreased adherence. Conclusions: Overexpression of LMWPTP in PCa confers a malignant phenotype with worse clinical outcome. Prospective follow-up should determine the clinical potential of LMWPTP overexpression. Patient summary: These findings implicate low-molecular-weight protein tyrosine phosphatase as a novel oncogene in prostate cancer and could offer the possibility of using this protein as biomarker or target for treatment of this disease. (C) 2015 European Association of Urology. Published by Elsevier B.V. All rights reserved.69710719Dutch Cancer Society [EMCR2010-4737

Calix[6]arene bypasses human pancreatic cancer aggressiveness: Downregulation of receptor tyrosine kinases and induction of cell death by reticulum stress and autophagy

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)Pancreatic cancer ranks fourth among cancer-related causes of death in North America. Minimal progress has been made in the diagnosis and treatment of patients with late-stage tumors. Moreover, pancreatic cancer aggressiveness is closely related to high levels of pro-survival mediators, which can ultimately lead to rapid disease progression, resistance and metastasis. The main goal of this study was to define the mechanisms by which calix[6]arene, but not other calixarenes, efficiently decreases the aggressiveness of a drug resistant human pancreas carcinoma cell line (Panc-1). Calix[6]arene was more potent in reducing Panc-1 cell viability than gemcitabine and 5-fluorouracil. In relation to the underlying mechanisms of cytotoxic effects, it led to cell cycle arrest in the G0/G1 phase through downregulation of PIM1, CDK2, CDK4 and retinoblastoma proteins. Importantly, calix[6]arene abolished signal transduction of Mer and AXL tyrosine kinase receptors, both of which are usually overexpressed in pancreatic cancer. Accordingly, inhibition of PI3K and mTOR was also observed, and these proteins are positively modulated by Mer and AXL, Despite decreasing the phosphorylation of ART at Thr308, calix[6]arene caused an increase in phosphorylation at Ser473. These findings in conjunction with increased BiP and IRE1-alpha provide a molecular basis explaining the capacity of calix[6[arene to trigger endoplasmic reticulum stress and autophagic cell death. Our findings highlight calix[6]arene as a potential candidate for overcoming pancreatic cancer aggressiveness. Importantly, we provide evidence that calix[6]arene affects a broad array of key targets that are usually dysfunctional in pancreatic cancer, a highly desirable characteristic for chemotherapeutics. (C) 2013 Elsevier B.V. All rights reserved.18331228562865Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)Fundação de Amparo à Pesquisa do Estado de Minas Gerais (FAPEMIG)Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)FAPESP [Proc. 2010/15946-3, Proc. 2012/01038-9, 08/57906-3]CNPq [573913/2008-0

Hedgehog signaling maintains chemoresistance in myeloid leukemic cells

Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)The development of resistance against chemotherapy remains one of the major challenges in the clinical management of leukemia. There is still limited insight into the molecular mechanisms that maintain the chemotherapy-resistant phenotype, despite the obvious clinical relevance that such knowledge would have. In this study, we show that the chemotherapy-resistant phenotype of myeloid leukemia cells correlates with activation of the Hedgehog (Hh) pathway, whereas in chemosensitive cells, such activation is less pronounced. Importantly, the overexpression of Hh pathway components induces chemoprotection and inhibition of the pathway reverts chemoresistance of Lucena-1 cells, apparently by interfering with P-glycoprotein-dependent drug resistance. Our data thus identify the Hh pathway as an essential component of multidrug resistance (MDR) myeloid leukemia and suggest that targeting the Hh pathway might be an interesting therapeutic avenue for overcoming MDR resistance in myeloid leukemia. Oncogene (2010) 29, 6314-6322; doi:10.1038/onc.2010.375; published online 30 August 2010294863146322Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)Top-Institute pharmaMaag-, Lever-, DarmstichtingConselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq

Effect of Phoneutria nigriventer Venom on the Expression of Junctional Protein and P-gp Efflux Pump Function in the Blood-Brain Barrier

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)Phoneutria nigriventer spider venom (PNV) contains Ca2+, K+ and Na+ channel-acting peptides that affect neurotransmitter release and causes excitotoxicity in PNS and CNS. It has been demonstrated that PNV causes blood-brain barrier (BBB) breakdown of hippocampal microvessels time-dependently through enhanced microtubule-mediated vesicular transport. Herein, it is hypothesized that PNV can cause BBB breakdown in the hippocampus and cerebellum time-dependently through other molecular mechanisms. The BBB integrity was assessed through the analysis of expression of Poly-glycoprotein (P-gp) efflux transporter protein, laminin from basement membrane and endothelial tight junctional and adhesion junctional (TJ/AJ) proteins. Phosphatase and tensin homolog (PTEN) and protein phosphatase 2A (PP2A) expression, which are known to have a role in the phosphorylation of junctional proteins and BBB opening, were also investigated. Astrocytes P-gp activity was determined by flow cytometry. The study demonstrated temporary decreased expression of laminin, TJ and AJ proteins (ZO1//occludin//claudin-5//beta-catenin) and P-gp (more prominently in hippocampus), which was completely or partially resolved between 2 and 5 h (and more quickly for cerebellum). PNV inhibited P-gp activity in astrocytes. PP2A phosphorylation, which inhibits the enzyme activity, was increased in both regions (15-45 min); however the phosphorylation level returned to baseline after 2 h. In conclusion, PNV disrupts paracellular transport in the BBB and possesses substrates for the active P-gp efflux transporter located in the BBB complex. Further studies into cellular mechanisms of astrocyte/endothelial interactions, using PNV as tool, may identify how astrocytes regulate the BBB, a characteristic that may be useful for the temporary opening of the BBB.37919671981Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)FAPESP [98/00341-0, 07/50242-6, 08/55748-1]CNPq [522131/95-6

Low-Molecular-Weight Protein Tyrosine Phosphatase Predicts Prostate Cancer Outcome by Increasing the Metastatic Potential

Background: Low-risk patients suffering from prostate cancer (PCa) are currently placed under active surveillance rather than undergoing radical prostatectomy. However, clear parameters for selecting the right patient for each strategy are not available, and new biomarkers and treatment modalities are needed. Low-molecular-weight protein tyrosine phosphatase (LMWPTP) could present such a target. Objective: To correlate expression levels of LMWPTP in primary PCa to clinical outcome, and determine the role of LMWPTP in prostate tumor cell biology. Design, setting, and participants: Acid phosphatase 1, soluble (ACP1) expression was analyzed on microarray data sets, which were subsequently used in Ingenuity Pathway Analysis. Immunohistochemistry was performed on a tissue microarray containing material of 481 PCa patients whose clinicopathologic data were recorded. PCa cell line models were used to investigate the role of LMWPTP in cell proliferation, migration, adhesion, and anoikis resistance. Outcome measurements and statistical analysis: The association between LMWPTP expression and clinical and pathologic outcomes was calculated using chi-square correlations and multivariable Cox regression analysis. Functional consequences of LMWPTP overexpression or downregulation were determined using migration and adhesion assays, confocal microscopy, Western blotting, and proliferation assays. Results and limitations: LMWPTP expression was significantly increased in human PCa and correlated with earlier recurrence of disease (hazard ratio [HR]: 1.99; p &lt; 0.001) and reduced patient survival (HR: 1.53; p = 0.04). Unbiased Ingenuity analysis comparing cancer and normal prostate suggests migratory propensities in PCa. Indeed, overexpression of LMWPTP increases PCa cell migration, anoikis resistance, and reduces activation of focal adhesion kinase/paxillin, corresponding to decreased adherence. Conclusions: Overexpression of LMWPTP in PCa confers a malignant phenotype with worse clinical outcome. Prospective follow-up should determine the clinical potential of LMWPTP overexpression. Patient summary: These findings implicate low-molecular-weight protein tyrosine phosphatase as a novel oncogene in prostate cancer and could offer the possibility of using this protein as biomarker or target for treatment of this disease. (C) 2015 European Association of Urology. Published by Elsevier B.V. All rights reserved

SGC-AAK1-1: a chemical probe targeting AAK1 and BMP2K

Inhibitors based on a 3-acylaminoindazole scaffold were synthesized to yield potent dual AAK1/BMP2K inhibitors. Optimization furnished a small molecule chemical probe (SGC-AAK1-1, 25) that is potent and selective for AAK1/BMP2K over other NAK family members, demonstrates narrow activity in a kinome-wide screen, and is functionally active in cells. This inhibitor represents one of the best available small molecule tools to study the functions of AAK1 and BMP2K