| 1. |
- Don-Doncow, Nicholas, et al.
(author)
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Galiellalactone is a Direct Inhibitor of STAT3 in Prostate Cancer Cells.
- 2014
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In: Journal of Biological Chemistry. - 1083-351X. ; 289:23, s. 15969-15978
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Journal article (peer-reviewed)abstract
- The transcription factor STAT3 is constitutively active in several malignancies including castration-resistant prostate cancer and has been identified as a promising therapeutic target. The fungal metabolite galiellalactone, a STAT3 signaling inhibitor, inhibits the growth, both in vitro and in vivo, of prostate cancer cells expressing active STAT3 and induces apoptosis of prostate cancer stem cell-like cells expressing pSTAT3. However, the molecular mechanism of this STAT3 inhibiting effect by galiellalactone has not been clarified. A biotinylated analogue of galiellalactone (GL-biot) was synthesized to be used for identification of galiellalactone target proteins. By adding streptavidin-sepharose beads to GL-biot treated DU145 cell lysates, STAT3 was isolated and identified as a target protein. Confocal microscopy revealed GL-biot in both the cytoplasm and nucleus of DU145 cells treated with GL-biot, appearing to co-localize with STAT3 in the nucleus. Galiellalactone inhibited STAT3 binding to DNA in DU145 cell lysates without affecting phosphorylation status of STAT3. Mass spectrometry analysis of recombinant STAT3 protein pretreated with galiellalactone revealed three modified cysteines (cys-367, cys-468 and cys-542). We here demonstrate with chemical and molecular pharmacological methods that galiellalactone is a cysteine reactive inhibitor that covalently binds to one or more cysteines in STAT3 and that this leads to inhibition of STAT3 binding to DNA and thus blocks STAT3 signaling without affecting phosphorylation. This further validates galiellalactone as a promising direct STAT3 inhibitor for treatment of castration-resistant prostate cancer.
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| 2. |
- Hägglund, Maria G A, et al.
(author)
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B(0)AT2 (SLC6A15) is localized to neurons and astrocytes, and is involved in mediating the effect of leucine in the brain
- 2013
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In: PLOS ONE. - : Public Library of Science (PLoS). - 1932-6203. ; 8:3, s. e58651-
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Journal article (peer-reviewed)abstract
- The B(0)AT2 protein is a product of the SLC6A15 gene belonging to the SLC6 subfamily and has been shown to be a transporter of essential branched-chain amino acids. We aimed to further characterize the B(0)AT2 transporter in CNS, and to use Slc6a15 knock out (KO) mice to investigate whether B(0)AT2 is important for mediating the anorexigenic effect of leucine. We used the Slc6a15 KO mice to investigate the role of B(0)AT2 in brain in response to leucine and in particular the effect on food intake. Slc6a15 KO mice show lower reduction of food intake as well as lower neuronal activation in the ventromedial hypothalamic nucleus (VMH) in response to leucine injections compared to wild type mice. We also used RT-PCR on rat tissues, in situ hybridization and immunohistochemistry on mouse CNS tissues to document in detail the distribution of SLC6A15 on gene and protein levels. We showed that B(0)AT2 immunoreactivity is mainly neuronal, including localization in many GABAergic neurons and spinal cord motor neurons. B(0)AT2 immunoreactivity was also found in astrocytes close to ventricles, and co-localized with cytokeratin and diazepam binding inhibitor (DBI) in epithelial cells of the choroid plexus. The data suggest that B(0)AT2 play a role in leucine homeostasis in the brain.
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| 3. |
- Hägglund, Maria G. A., et al.
(author)
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Characterization of the transporterB0AT3 (Slc6a17) in the rodent central nervous system
- 2013
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In: BMC Neuroscience. - : Springer Science and Business Media LLC. - 1471-2202. ; 14, s. 54-
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Journal article (peer-reviewed)abstract
- Background: The vesicular B(0)AT3 transporter (SLC6A17), one of the members of the SLC6 family, is a transporter for neutral amino acids and is exclusively expressed in brain. Here we provide a comprehensive expression profile of B(0)AT3 in mouse brain using in situ hybridization and immunohistochemistry. Results: We confirmed previous expression data from rat brain and used a novel custom made antibody to obtain detailed co-labelling with several cell type specific markers. B(0)AT3 was highly expressed in both inhibitory and excitatory neurons. The B(0)AT3 expression was highly overlapping with those of vesicular glutamate transporter 2 (VGLUT2) and vesicular glutamate transporter 1 (VGLUT1). We also show here that Slc6a17mRNA is up-regulated in animals subjected to short term food deprivation as well as animals treated with the serotonin reuptake inhibitor fluoxetine and the dopamine/noradrenaline reuptake inhibitor bupropion. Conclusions: This suggests that the B(0)AT3 transporter have a role in regulation of monoaminergic as well as glutamatergic synapses.
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| 4. |
- Hägglund, Maria G A, et al.
(author)
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Transport of L-glutamine, L-alanine, L-arginine and L-histidine by the neuron-specific Slc38a8 (SNAT8) in CNS
- 2015
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In: Journal of Molecular Biology. - : Elsevier BV. - 0022-2836 .- 1089-8638. ; 427:6, s. 1495-1512
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Journal article (peer-reviewed)abstract
- Glutamine transporters are important for regulating levels of glutamate and GABA in the brain. To date, six members of the SLC38 family (SNATs) have been characterized and functionally subdivided into System A (SNAT1, SNAT2 and SNAT4) and System N (SNAT3, SNAT5 and SNAT7). Here we present a first functional characterization of SLC38A8, one of the previous orphan transporters from the family and we suggest that the encoded protein should be named SNAT8 to adhere with the SNAT nomenclature. We show that SLC38A8 have preference for transporting L-glutamine, L-alanine, L-arginine, L-histidine, and L-aspartate using a Na(+)-dependent transport mechanism and that the functional characteristics of SNAT8 has highest similarity to the known System A transporters. We also provide a comprehensive CNS expression profile in mouse brain for the Slc38a8 gene and the SNAT8 protein. We show that Slc38a8 (SNAT8) is expressed in all neurons, both excitatory and inhibitory, in mouse brain using in situ hybridization and immunohistochemistry. Furthermore, proximity ligation assay show highly similar subcellular expression of SNAT7 and SNAT8. In conclusion, the neuronal SLC38A8 have a broad amino acid transport profile and is the first identified neuronal System A transporter. This suggests a key role of SNAT8 in the glutamine/glutamate(GABA) cycle in the brain.
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| 5. |
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| 6. |
- Lindgren, Anders E. G., et al.
(author)
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Chemical Probes to Study ADP-Ribosylation : Synthesis and Biochemical Evaluation of Inhibitors of the Human ADP-Ribosyltransferase ARTD3/PARP3
- 2013
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In: Journal of Medicinal Chemistry. - : American Chemical Society (ACS). - 0022-2623 .- 1520-4804. ; 56:23, s. 9556-9568
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Journal article (peer-reviewed)abstract
- The racemic 3-(4-oxo-3,4-dihydroquinazolin-2-yl)-N-[1-(pyridin-2-yl)ethyl]propanamide, 1, has previously been identified as a potent but unselective inhibitor of diphtheria toxin-like ADP-ribosyltransferase 3 (ARTD3). Herein we describe synthesis and evaluation of SS compounds in this class. It was found that the stereochemistry is of great importance for both selectivity and potency and that substituents on the phenyl ring resulted in poor solubility. Certain variations at the meso position were tolerated and caused a large shift in the binding pose. Changes to the ethylene linker that connects the quinazolinone to the amide were also investigated but proved detrimental to binding. By combination of synthetic organic chemistry and structure-based design, two selective inhibitors of ARTD3 were discovered.
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