16 research outputs found
Dopaminergic control of ADAMTS2 expression through cAMP/CREB and ERK: molecular effects of antipsychotics
A better understanding of the molecular mechanisms that participate in the development and clinical manifestations of schizophrenia can lead to improve our ability to diagnose and treat this disease. Previous data strongly associated the levels of deregulated ADAMTS2 expression in peripheral blood mononuclear cells (PBMCs) from patients at first episode of psychosis (up) as well as in clinical responders to treatment with antipsychotic drugs (down). In this current work, we performed an independent validation of such data and studied the mechanisms implicated in the control of ADAMTS2 gene expression. Using a new cohort of drug-naïve schizophrenia patients with clinical follow-up, we confirmed that the expression of ADAMTS2 was highly upregulated in PBMCs at the onset (drug-naïve patients) and downregulated, in clinical responders, after treatment with antipsychotics. Mechanistically, ADAMTS2 expression was activated by dopaminergic signalling (D1-class receptors) and downstream by cAMP/CREB and mitogen-activated protein kinase (MAPK)/ERK signalling. Incubation with antipsychotic drugs and selective PKA and MEK inhibitors abrogated D1-mediated activation of ADAMTS2 in neuronal-like cells. Thus, D1 receptors signalling towards CREB activation might participate in the onset and clinical responses to therapy in schizophrenia patients, by controlling ADAMTS2 expression and activity. The unbiased investigation of molecular mechanisms triggered by antipsychotic drugs may provide a new landscape of novel targets potentially associated with clinical efficacy.Acknowledgements: We are highly indebted to the participants and their families for their cooperation in this study. We also thank IDIVAL biobank (Inés Santiuste and Jana Arozamena) for clinical samples and data as well as the PAFIP members (Marga Corredera) for the data collection. This work was supported by: SAF2016-76046-R and SAF2013-46292-R (MINECO and FEDER) to B.C.F., PI16/00156 (isciii and FEDER) to J.P.V., LUCHAMOS POR LA VIDA project to F.R.J. and J.P.V., SAF2017-83702-R (MINECO and FEDER), Red TERCEL RD12/0019/0024 (ISCIII) and GVA-PROMETEO 2018/041 (Generalitat Valenciana) to S.M. J.P.V. is supported by the RyC research programme (RYC-2013-14097) and F.R.J. by the predoctoral research programme (BES-2014-070615), from MINECO and FEDER
Dopaminergic control of ADAMTS2 expression through cAMP/CREB and ERK: molecular effects of antipsychotics
© The Author(s) 2019.A better understanding of the molecular mechanisms that participate in the development and clinical manifestations of schizophrenia can lead to improve our ability to diagnose and treat this disease. Previous data strongly associated the levels of deregulated ADAMTS2 expression in peripheral blood mononuclear cells (PBMCs) from patients at first episode of psychosis (up) as well as in clinical responders to treatment with antipsychotic drugs (down). In this current work, we performed an independent validation of such data and studied the mechanisms implicated in the control of ADAMTS2 gene expression. Using a new cohort of drug-naïve schizophrenia patients with clinical follow-up, we confirmed that the expression of ADAMTS2 was highly upregulated in PBMCs at the onset (drug-naïve patients) and downregulated, in clinical responders, after treatment with antipsychotics. Mechanistically, ADAMTS2 expression was activated by dopaminergic signalling (D1-class receptors) and downstream by cAMP/CREB and mitogen-activated protein kinase (MAPK)/ERK signalling. Incubation with antipsychotic drugs and selective PKA and MEK inhibitors abrogated D1-mediated activation of ADAMTS2 in neuronal-like cells. Thus, D1 receptors signalling towards CREB activation might participate in the onset and clinical responses to therapy in schizophrenia patients, by controlling ADAMTS2 expression and activity. The unbiased investigation of molecular mechanisms triggered by antipsychotic drugs may provide a new landscape of novel targets potentially associated with clinical efficacy.This work was supported by: SAF2016-76046-R and SAF2013-46292-R (MINECO and FEDER) to B.C.F., PI16/00156 (isciii and FEDER) to J.P.V., LUCHAMOS POR LA VIDA project to F.R.J. and J.P.V., SAF2017-83702-R (MINECO and FEDER), Red TERCEL RD12/0019/0024 (ISCIII) and GVA-PROMETEO 2018/041 (Generalitat Valenciana) to S.M. J.P.V. is supported by the RyC research programme (RYC-2013-14097) and F.R.J. by the predoctoral research programme (BES-2014-070615), from MINECO and FEDER
Applied diagnostics in liver cancer. Efficient combinations of sorafenib with targeted inhibitors blocking AKT/mTOR
Hepatocellular carcinoma (HCC) is the third most common cause of cancer-related deaths worldwide. There is increasing interest in developing specific markers to serve as predictors of response to sorafenib and to guide targeted therapy. Using a sequencing platform designed to study somatic mutations in a selection of 112 genes (HepatoExome), we aimed to characterize lesions from HCC patients and cell lines, and to use the data to study the biological and mechanistic effects of case-specific targeted therapies used alone or in combination with sorafenib. We characterized 331 HCC cases in silico and 32 paired samples obtained prospectively from primary tumors of HCC patients. Each case was analyzed in a time compatible with the requirements of the clinic (within 15 days). In 53% of the discovery cohort cases, we detected unique mutational signatures, with up to 34% of them carrying mutated genes with the potential to guide therapy. In a panel of HCC cell lines, each characterized by a specific mutational signature, sorafenib elicited heterogeneous mechanistic and biological responses, whereas targeted therapy provoked the robust inhibition of cell proliferation and DNA synthesis along with the blockage of AKT/mTOR signaling. The combination of sorafenib with targeted therapies exhibited synergistic anti-HCC biological activity concomitantly with highly effective inhibition of MAPK and AKT/mTOR signaling. Thus, somatic mutations may lead to identify case-specific mechanisms of disease in HCC lesions arising from multiple etiologies. Moreover, targeted therapies guided by molecular characterization, used alone or in combination with sorafenib, can effectively block important HCC disease mechanisms.FUNDING: Grants from ISCIII, co-financed by the European Union (FEDER) (PI16/00156), Ramón and Cajal research program from MINECO (RYC-2013-14097) and FUNDACIÓN LUCHAMOS POR LA VIDA to JPV. Grants from ISCIII (RD06/0020/0107-RD012/0036/0060) to MAP. Grant from ISCIII (Ref. PIE15/00079) to JC & JPV. NGD is a recipient of a UC-IDIVAL pre-doctoral fellow. I.V. was also supported by the Ramón and Cajal research program
Molecular basis of targeted therapy in T/NKcell lymphoma/leukemia: A comprehensive genomic and immunohistochemical analysis of a panel of 33 cell lines
T and NK-cell lymphoma is a collection of aggressive disorders with unfavorable outcome, in which targeted treatments are still at a preliminary phase. To gain deeper insights into the deregulated mechanisms promoting this disease, we searched a panel of 31 representative T-cell and 2 NK-cell lymphoma/leukemia cell lines for predictive markers of response to targeted therapy. To this end, targeted sequencing was performed alongside the expression of specific biomarkers corresponding to potentially activated survival pathways. The study identified TP53, NOTCH1 and DNMT3A as the most frequently mutated genes. We also found common alterations in JAK/STAT and epigenetic pathways. Immunohistochemical analysis showed nuclear accumulation of MYC (in 85% of the cases), NFKB (62%), p-STAT (44%) and p-MAPK (30%). This panel of cell lines captures the complexity of T/NK-cell lymphoproliferative processes samples, with the partial exception of AITL cases. Integrated mutational and immunohistochemical analysis shows that mutational changes cannot fully explain the activation of key survival pathways and the resulting phenotypes. The combined integration of mutational/expression changes forms a useful tool with which new compounds may be assayed
Dynamic etching of soluble surface layers with on-line inductively coupled plasma mass spectrometry detection – a novel approach for determination of complex metal oxide surface cation stoichiometry
Mutated JAK kinases and deregulated STAT activity are potential therapeutic targets in cutaneous T-cell lymphoma
Mutational landscape of TCL cell lines.
<p>The results of targeted deep sequencing of 16 genes in 20 T-ALL (black), 5 ALCL (dark grey), 3 CTCL (medium grey), 2 NK (light grey), 2 ATLL (diagonal lines) and one T-LGL (dots) cell lines. Mutated genes (rows) are arranged in decreasing order of mutation frequency. Cell lines (columns) are arranged from left to right on the basis of their mutational frequency following gene ranking. HTLV-1-positive cell lines (green) and translocation t(2;5)(p23;q35) (ALK +, dark blue) are showed.</p
Mapping of variants in a TCL gene panel.
<p>Schematic of the alterations encoded by SNVs in <i>TP53</i>, <i>NOTCH1</i>, <i>DNMT3A</i>, <i>JAK1</i>, <i>JAK3</i>, <i>STAT3</i> and <i>STAT5B</i>. Type of variation and disease are represented by color and shape, respectively. TAD: transactivation domain; PRD: proline-rich domain; TD: tetramerization domain; C-term: C-terminal domain; HD: heterodimerization domain; TM: transmembrane domain; RAM: Rbp-associated molecule domain; ANK: ankyrin domain; PEST: proline (P), glutamic acid (E), serine (S), threonine (T) degradation domain; ZNF: zinc-finger domain; Mtase: methyltransferase domain.</p
Unsupervised hierarchical clustering analysis with 26 immunomarkers.
<p>Each row represents a single cell line; each column represents a single immunomarker. Blue (score 0); white, weak immunostaining (score 1); light red (score 2); red, strong immunoreactivity (score 3); grey, missing data.</p
PIM kinases as potential therapeutic targets in PTCL.
<p>(A) Gene expression profiling of tumoral samples from 38 human PTCL patients compared with 6 reactive lymph nodes (LN) by microarrays revealed a significantly increased expression of <i>PIM1</i> and <i>PIM2</i> genes (FDR<0.05), but not <i>PIM3</i>. The heatmap is shown in the upper panel, and the relative quantification (Log<sub>2</sub> ratio) comparing PIM expression in PTCL <i>versus</i> LN is shown in the lower panel. (B) GSEA ranked all significantly altered genes between PTCL and LN according to its correlation with <i>PIM1</i> or <i>PIM2</i> expression and displayed them in the red-to-blue bar. Each gene belonging to every pathway was interrogated whether it appeared positively (in the red region of the bar) or negatively (in the blue side) correlated. Using this approach GSEA identified a positive and significant correlation between <i>PIM1</i> and <i>PIM2</i> expression and Jak/STAT, NF-κB and IL-2 signaling pathways in the PTCL molecular signature (FDR<0.25). (C) PIM family genes mRNA level was measured by RT-qPCR in eight PTCL cell lines and (D) primary tumoral T cells from 5 Sézary Syndrome patients (SS #1–5), and compared with normal T cells isolated from 3 healthy donors (Control #1–3). The relative RNA amount of PIM has been calculated as a relative quantification, as described in the <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0112148#s2" target="_blank">Methods</a> section (RQ = 2<sup>−ΔCt</sup>), normalized with non-tumoral cells: RQ in PTCL/RQ in healthy #3. In both settings, <i>PIM1</i>, and especially <i>PIM2</i>, but not <i>PIM3</i> expression was found to be increased in PTCL. (E) PIM kinase protein basal levels in PTCL cell lines measured by Western blot. PIM1 and PIM2 isoforms are also shown. (F) Distribution of PIM2 protein in a series of tumoral samples from 136 PTCL patients measured by immunohistochemistry. Negative, weakly positive and strongly positive samples were defined by the presence of <5%, 5–20% and >20% positive cells. (G) Distribution of PIM2 protein in the most common PTCL subtypes measured by immunohistochemistry.</p