Publications by authors named "Olga Martinho"

44 Publications

Comprehensive Molecular Landscape of Cetuximab Resistance in Head and Neck Cancer Cell Lines.

Cells 2022 Jan 4;11(1). Epub 2022 Jan 4.

Molecular Oncology Research Center, Barretos Cancer Hospital, Barretos 14784-400, Brazil.

Cetuximab is the sole anti-EGFR monoclonal antibody that is FDA approved to treat head and neck squamous cell carcinoma (HNSCC). However, no predictive biomarkers of cetuximab response are known for HNSCC. Herein, we address the molecular mechanisms underlying cetuximab resistance in an in vitro model. We established a cetuximab resistant model (FaDu), using increased cetuximab concentrations for more than eight months. The resistance and parental cells were evaluated for cell viability and functional assays. Protein expression was analyzed by Western blot and human cell surface panel by lyoplate. The mutational profile and copy number alterations (CNA) were analyzed using whole-exome sequencing (WES) and the NanoString platform. FaDu resistant clones exhibited at least two-fold higher IC compared to the parental cell line. WES showed relevant mutations in several cancer-related genes, and the comparative mRNA expression analysis showed 36 differentially expressed genes associated with EGFR tyrosine kinase inhibitors resistance, RAS, MAPK, and mTOR signaling. Importantly, we observed that overexpression of KRAS, RhoA, and CD44 was associated with cetuximab resistance. Protein analysis revealed EGFR phosphorylation inhibition and mTOR increase in resistant cells. Moreover, the resistant cell line demonstrated an aggressive phenotype with a significant increase in adhesion, the number of colonies, and migration rates. Overall, we identified several molecular alterations in the cetuximab resistant cell line that may constitute novel biomarkers of cetuximab response such as mTOR and RhoA overexpression. These findings indicate new strategies to overcome anti-EGFR resistance in HNSCC.
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http://dx.doi.org/10.3390/cells11010154DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8750399PMC
January 2022

Increased CD3, CD8, or FoxP3 T Lymphocyte Infiltrations Are Associated with the Pathogenesis of Colorectal Cancer but Not with the Overall Survival of Patients.

Biology (Basel) 2021 Aug 20;10(8). Epub 2021 Aug 20.

Life and Health Sciences Research Institute (ICVS), School of Medicine, University of Minho, 4710-057 Braga, Portugal.

Tumor-infiltrating lymphocytes include heterogeneous populations of T lymphocytes that play crucial roles in the tumor immune response; importantly, their presence in the tumor tissue may predict clinical outcomes. Therefore, we herein studied the prognostic significance of the presence and location of CD3, CD8, and FoxP3 T lymphocytes in colorectal cancer samples. In the intratumor analysis, our data did not reveal any association between lymphocyte infiltrations with clinical or pathological data. However, in the tumor margins, we found that the presence of high infiltrations of CD3, CD8, or FoxP3 T lymphocytes were associated with TNM stages I-II ( = 0.021, = 0.022, and = 0.012, respectively) and absence of lymph node metastases ( = 0.010, = 0.003, and = 0.004, respectively). Despite these associations with good prognostic indicators, we were not able to find any statistically significant alterations in the overall survival of the patients, even though high infiltrations of FoxP3 T lymphocytes in the tumor margins resulted in an increased overall survival of 14 months. Taken together, these data show that the presence of CD3, CD8, or FoxP3T lymphocyte infiltrates in the tumor margins are associated with the pathogenesis of CRC, but only high Foxp3 T lymphocyte infiltrations in the tumor invasive margins are inclined to indicate favorable prognosis.
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http://dx.doi.org/10.3390/biology10080808DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8389643PMC
August 2021

A 30-Year Long-Term Experience in Appendix Neuroendocrine Neoplasms-Granting a Positive Outcome.

Cancers (Basel) 2020 May 26;12(6). Epub 2020 May 26.

Instituto de Investigação e Inovação em Saúde (i3S), 4200-135 Porto, Portugal.

Neuroendocrine neoplasms (NENs) are the most common tumor of the appendix and have an excellent prognosis. Appendiceal tumors diagnosed between 1989 and 2019 were reviewed, and clinical data were collected from patient files. Part of the series was immuno-profiled for markers related to cell cycle proliferation and/or senescence-type, apoptotic, and metastatic potential. Appendix NENs were detected in 74 patients, with 0.47% of incidence per appendectomy. The median age of the patients was 21.5 years, with two age peaks of incidence at 17.0 and 55.2 years. The median tumors size was 5.8 mm, and most were smaller than 10 mm. Lymphovascular and perineural invasion, as well as necrosis, was associated with larger tumor size. G1 tumors composed 96.0% of the cohort. The presence of moderate/strong p16 and the absent/low Bcl-2 expression was frequently observed and associated with a smaller size. This study represents one of the largest cohorts and with a long follow-up. For tumors smaller than 10 mm appendicectomy was sufficient as a curative procedure, as revealed by the good outcome. This series presented a 100% disease-free survival. The indolent phenotype of appendix NENs is supported by the expression of markers that point towards a strong inhibition of cell replication and growth inhibition.
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http://dx.doi.org/10.3390/cancers12061357DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7353034PMC
May 2020

WNK2 Inhibits Autophagic Flux in Human Glioblastoma Cell Line.

Cells 2020 02 20;9(2). Epub 2020 Feb 20.

Molecular Oncology Research Center, Barretos Cancer Hospital, 14784 400 Barretos, Brazil.

Autophagy is a cell-survival pathway with dual role in tumorigenesis, promoting either tumor survival or tumor death. gene, a member of the WNK (with no lysine (K)) subfamily, acts as a tumor suppressor gene in gliomas, regulating cell migration and invasion; however, its role in autophagy process is poorly explored. The -methylated human glioblastoma cell line A172 WT (wild type) was compared to transfected clones A172 EV (empty vector), and A172 WNK2 (WNK2 overexpression) for the evaluation of autophagy using an inhibitor (bafilomycin A1-baf A1) and an inducer (everolimus) of autophagic flux. Western blot and immunofluorescence approaches were used to monitor autophagic markers, LC3A/B and SQSTM1/p62. A172 WNK2 cells presented a significant decrease in LC3B and p62 protein levels, and in LC3A/B ratio when compared with control cells, after treatment with baf A1 + everolimus, suggesting that overexpression inhibits the autophagic flux in gliomas. The mTOR pathway was also evaluated under the same conditions, and the observed results suggest that the inhibition of autophagy mediated by WNK2 occurs through a mTOR-independent pathway. In conclusion, the evaluation of the autophagic process demonstrated that WNK2 inhibits the autophagic flux in glioblastoma cell line.
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http://dx.doi.org/10.3390/cells9020485DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7072831PMC
February 2020

Semi-Synthetic Ingenol Derivative from Euphorbia tirucalli Inhibits Protein Kinase C Isotypes and Promotes Autophagy and S-phase Arrest on Glioma Cell Lines.

Molecules 2019 Nov 22;24(23). Epub 2019 Nov 22.

Molecular Oncology Research Center, Barretos Cancer Hospital, Barretos, São Paulo 14784-400, Brazil.

The identification of signaling pathways that are involved in gliomagenesis is crucial for targeted therapy design. In this study we assessed the biological and therapeutic effect of ingenol-3-dodecanoate (IngC) on glioma. IngC exhibited dose-time-dependent cytotoxic effects on large panel of glioma cell lines (adult, pediatric cancer cells, and primary cultures), as well as, effectively reduced colonies formation. Nevertheless, it was not been able to attenuate cell migration, invasion, and promote apoptotic effects when administered alone. IngC exposure promoted S-phase arrest associated with p21CIP/WAF1 overexpression and regulated a broad range of signaling effectors related to survival and cell cycle regulation. Moreover, IngC led glioma cells to autophagy by LC3B-II accumulation and exhibited increased cytotoxic sensitivity when combined to a specific autophagic inhibitor, bafilomycin A1. In comparison with temozolomide, IngC showed a mean increase of 106-fold in efficacy, with no synergistic effect when they were both combined. When compared with a known compound of the same class, namely ingenol-3-angelate (I3A, Picato®), IngC showed a mean 9.46-fold higher efficacy. Furthermore, IngC acted as a potent inhibitor of protein kinase C (PKC) activity, an emerging therapeutic target in glioma cells, showing differential actions against various PKC isotypes. These findings identify IngC as a promising lead compound for the development of new cancer therapy and they may guide the search for additional PKC inhibitors.
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http://dx.doi.org/10.3390/molecules24234265DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6930609PMC
November 2019

RKIP as an Inflammatory and Immune System Modulator: Implications in Cancer.

Biomolecules 2019 11 22;9(12). Epub 2019 Nov 22.

Life and Health Sciences Research Institute (ICVS), School of Medicine, University of Minho, Campus de Gualtar, 4710-057 Braga, Portugal.

Raf kinase inhibitor protein (RKIP), an important modulator of intracellular signalling pathways, is commonly downregulated in multiple cancers. This reduction, or loss of expression, is correlated not only with the presence of metastasis, contributing to RKIP's classification as a metastasis suppressor, but also with tumour aggressiveness and poor prognosis. Recent findings suggest a strong involvement of RKIP in the modulation of tumour microenvironment components, particularly by controlling the infiltration of specific immune cells and secretion of pro-metastatic factors. Additionally, RKIP interaction with multiple signalling molecules seems to potentiate its function as a regulator of inflammatory processes, mainly through stimulation of anti- or pro-inflammatory cytokines. Furthermore, RKIP is involved in the modulation of immunotherapeutic drugs response, through diverse mechanisms that sensitize cells to apoptosis. In the present review, we will provide updated information about the role of RKIP as an inflammatory and immune modulator and its potential implications in cancer will be addressed.
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http://dx.doi.org/10.3390/biom9120769DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6995551PMC
November 2019

Magnetoliposomes Containing Calcium Ferrite Nanoparticles for Applications in Breast Cancer Therapy.

Pharmaceutics 2019 Sep 14;11(9). Epub 2019 Sep 14.

Centre of Physics (CFUM), University of Minho, Campus de Gualtar, 4710-057 Braga, Portugal.

Magnetoliposomes containing calcium ferrite (CaFeO) nanoparticles were developed and characterized for the first time. CaFeO nanoparticles were covered by a lipid bilayer or entrapped in liposomes forming, respectively, solid or aqueous magnetoliposomes as nanocarriers for new antitumor drugs. The magnetic nanoparticles were characterized by UV/Visible absorption, XRD, HR-TEM, and SQUID, exhibiting sizes of 5.2 ± 1.2 nm (from TEM) and a superparamagnetic behavior. The magnetoliposomes were characterized by DLS and TEM. The incorporation of two new potential antitumor drugs (thienopyridine derivatives) specifically active against breast cancer in these nanosystems was investigated by fluorescence emission and anisotropy. Aqueous magnetoliposomes, with hydrodynamic diameters around 130 nm, and solid magnetoliposomes with sizes of ca. 170 nm, interact with biomembranes by fusion and are able to transport the antitumor drugs with generally high encapsulation efficiencies (70%). These fully biocompatible drug-loaded magnetoliposomes can be promising as therapeutic agents in future applications of combined breast cancer therapy.
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http://dx.doi.org/10.3390/pharmaceutics11090477DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6781553PMC
September 2019

Current Status of Raf Kinase Inhibitor Protein (RKIP) in Lung Cancer: Behind RTK Signaling.

Cells 2019 05 10;8(5). Epub 2019 May 10.

Life and Health Sciences Research Institute (ICVS), School of Medicine, University of Minho, Campus de Gualtar, 4710-057 Braga, Portugal.

Lung cancer is the most deadly neoplasm with the highest incidence in both genders, with non-small cell lung cancer (NSCLC) being the most frequent subtype. Somatic mutations within the tyrosine kinase domain of the epidermal growth factor receptor (EGFR) gene are key drivers of NSCLC progression, with EGFR inhibitors being particularly beneficial for patients carrying the so-called "EGFR-sensitizing mutations". However, patients eventually acquire resistance to these EGFR inhibitors, and a better knowledge of other driven and targetable proteins will allow the design of increasingly accurate drugs against patients' specific molecular aberrations. Raf kinase inhibitory protein (RKIP) is an important modulator of relevant intracellular signaling pathways, including those controlled by EGFR, such as MAPK. It has been reported that it has metastasis suppressor activity and a prognostic role in several solid tumors, including lung cancer. In the present review, the potential use of RKIP in the clinic as a prognostic biomarker and predictor of therapy response in lung cancer is addressed.
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http://dx.doi.org/10.3390/cells8050442DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6562953PMC
May 2019

Modified ingenol semi-synthetic derivatives from Euphorbia tirucalli induce cytotoxicity on a large panel of human cancer cell lines.

Invest New Drugs 2019 10 1;37(5):1029-1035. Epub 2019 Feb 1.

Molecular Oncology Research Center, Barretos Cancer Hospital, Rua Antenor Duarte Villela, 1331, CEP 14784 400, Barretos, São Paulo, Brazil.

The latex from Euphorbia tirucalli is used in Brazil as a folk medicine for several diseases, including cancer. Recently, we showed a cytotoxic activity of E. tirucalli euphol in a wide range of cancer cell lines. Moreover, we showed that euphol inhibits proliferation, motility and colony formation in pancreatic cancer cells, induces autophagy and sensitizes glioblastoma cells to temozolomide cytotoxicity. Herein, we report in vitro activity of three semi-synthetic ingenol compounds derived from E. tirucalli, IngA (ingenol-3-trans-cinnamate), IngB (ingenol-3-hexanoate) and IngC (ingenol-3-dodecanoate), against a large panel of human cancer cell lines. Antineoplastic effects of the three semi-synthetic compounds were assessed using MTS assays on 70 cancer cell lines from a wide array of solid tumors. Additionally, their antitumor potential was compared with known compounds of the same class, namely ingenol-3-angelate (Picato®) and ingenol 3,20-dibenzoate and in combination with standard chemotherapeutic agents. We observed that IngA, B, and C exhibited dose-dependent cytotoxic effects. Amongst the semi-synthetic compounds, IngC displayed the best activity across the tumor cell lines. In comparison with ingenol-3-angelate and ingenol 3,20-dibenzoate, IngC showed a mean of 6.6 and 3.6-fold higher efficacy, respectively, against esophageal cancer cell lines. Besides, IngC sensitized esophageal cancer cells to paclitaxel treatment. In conclusion, the semi-synthetic ingenol compounds, in particular, IngC, demonstrated a potent antitumor activity on all cancer cell lines evaluated. Although the underlying mechanisms of action of IngC are not elucidated, our results provide insights for further studies suggesting IngC as a putative therapy for cancer treatment.
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http://dx.doi.org/10.1007/s10637-019-00728-0DOI Listing
October 2019

Hexane partition from Annona crassiflora Mart. promotes cytotoxity and apoptosis on human cervical cancer cell lines.

Invest New Drugs 2019 08 29;37(4):602-615. Epub 2018 Aug 29.

Molecular Oncology Research Center, Barretos Cancer Hospital, Barretos, 14784400, São Paulo, Brazil.

Cervical cancer is the third most commonly diagnosed tumor type and the fourth cause of cancer-related death in females. Therapeutic options for cervical cancer patients remain very limited. Annona crassiflora Mart. is used in traditional medicine as antimicrobial and antineoplastic agent. However, little is known about its antitumoral properties. In this study the antineoplastic effect of crude extract and derived partitions from A. crassiflora Mart in cervical cancer cell lines was evaluated. The crude extract significantly alters cell viability of cervical cancer cell lines as well as proliferation and migration, and induces cell death in SiHa cells. Yet, the combination of the crude extract with cisplatin leads to antagonistic effect. Importantly, the hexane partition derived from the crude extract presented cytotoxic effect both in vitro and in vivo, and initiates cell responses, such as DNA damage (H2AX activity), apoptosis via intrinsic pathway (cleavage of caspase-9, caspase-3, poly (ADP-ribose) polymerase (PARP) and mitochondrial membrane depolarization) and decreased p21 expression by ubiquitin proteasome pathway. Concluding, this work shows that hexane partition triggers several biological responses such as DNA damage and apoptosis, by intrinsic pathways, and was also able to promote a direct decrease in tumor perimeter in vivo providing a basis for further investigation on its antineoplastic activity on cervical cancer.
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http://dx.doi.org/10.1007/s10637-018-0657-yDOI Listing
August 2019

In vitro screening of cytotoxic activity of euphol from Euphorbia tirucalli on a large panel of human cancer-derived cell lines.

Exp Ther Med 2018 Aug 31;16(2):557-566. Epub 2018 May 31.

Molecular Oncology Research Center, Barretos Cancer Hospital, Barretos, SP 14784 400, Brazil.

A large number of classic antineoplastic agents are derived from plants. L. (Euphorbiaceae) is a subtropical and tropical plant, used in Brazilian folk medicine against many diseases, including cancer, yet little is known about its true anticancer properties. The present study evaluated the antitumor effect of the tetracyclic triterpene alcohol, euphol, the main constituent of in a panel of 73 human cancer lines from 15 tumor types. The biological effect of euphol in pancreatic cells was also assessed. The combination index was further used to explore euphol interactions with standard drugs. Euphol showed a cytotoxicity effect against several cancer cell lines (IC range, 1.41-38.89 µM), particularly in esophageal squamous cell (11.08 µM) and pancreatic carcinoma cells (6.84 µM), followed by prostate, melanoma, and colon cancer. Cytotoxicity effects were seen in all cancer cell lines, with more than half deemed highly sensitive. Euphol inhibited proliferation, motility and colony formation in pancreatic cancer cells. Importantly, euphol exhibited synergistic interactions with gemcitabine and paclitaxel in pancreatic and esophageal cell lines, respectively. To the best of our knowledge, this study constitutes the largest screening of euphol efficacy on cancer cell lines and revealed its anti-cancer properties, particularly in pancreatic and esophageal cell lines, suggesting that euphol, either as a single agent or in combination with conventional chemotherapy, is a potential anti-cancer drug.
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http://dx.doi.org/10.3892/etm.2018.6244DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6090420PMC
August 2018

Euphol, a tetracyclic triterpene, from Euphorbia tirucalli induces autophagy and sensitizes temozolomide cytotoxicity on glioblastoma cells.

Invest New Drugs 2019 04 22;37(2):223-237. Epub 2018 Jun 22.

Molecular Oncology Research Center, Barretos Cancer Hospital, Barretos, São Paulo, 14784 400, Brazil.

Glioblastoma (GBM) is the most frequent and aggressive type of brain tumor. There are limited therapeutic options for GBM so that new and effective agents are urgently needed. Euphol is a tetracyclic triterpene alcohol, and it is the main constituent of the sap of the medicinal plant Euphorbia tirucalli. We previously identified anti-cancer activity in euphol based on the cytotoxicity screening of 73 human cancer cells. We now expand the toxicological screening of the inhibitory effect and bioactivity of euphol using two additional glioma primary cultures. Euphol exposure showed similar cytotoxicity against primary glioma cultures compared to commercial glioma cells. Euphol has concentration-dependent cytotoxic effects on cancer cell lines, with more than a five-fold difference in the IC values in some cell lines. Euphol treatment had a higher selective cytotoxicity index (0.64-3.36) than temozolomide (0.11-1.13) and reduced both proliferation and cell motility. However, no effect was found on cell cycle distribution, invasion and colony formation. Importantly, the expression of the autophagy-associated protein LC3-II and acidic vesicular organelle formation were markedly increased, with Bafilomycin A1 potentiating cytotoxicity. Finally, euphol also exhibited antitumoral and antiangiogenic activity in vivo, using the chicken chorioallantoic membrane assay, with synergistic temozolomide interactions in most cell lines. In conclusion, euphol exerted in vitro and in vivo cytotoxicity against glioma cells, through several cancer pathways, including the activation of autophagy-associated cell death. These findings provide experimental support for further development of euphol as a novel therapeutic agent for GBM, either alone or in combination chemotherapy.
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http://dx.doi.org/10.1007/s10637-018-0620-yDOI Listing
April 2019

Metabolic alterations underlying Bevacizumab therapy in glioblastoma cells.

Oncotarget 2017 Nov 10;8(61):103657-103670. Epub 2017 Oct 10.

Life and Health Sciences Research Institute (ICVS), School of Medicine, University of Minho, Campus de Gualtar, Braga, Portugal.

Anti-VEGF therapy with Bevacizumab is approved for glioblastoma treatment, however, it is known that tumors acquired resistance and eventually became even more aggressive and infiltrative after treatment. In the present study we aimed to unravel the potential cellular mechanisms of resistance to Bevacizumab in glioblastoma models. Using a panel of glioblastoma cell lines we found that Bevacizumab is able to block the secreted VEGF by the tumor cells and be internalized to the cytoplasm, inducing cytotoxicity . We further found that Bevacizumab increases the expression of hypoxic (HIF-1α and CAIX) and glycolytic markers (GLUT1 and MCT1), leading to higher glucose uptake and lactate production. Furthermore, we showed that part of the consumed glucose by the tumor cells can be stored as glycogen, hampering cell dead following Bevacizumab treatment. Importantly, we found that this change on the glycolytic metabolism occurs independently of hypoxia and before mitochondrial impairment or autophagy induction. Finally, the combination of Bevacizumab with glucose uptake inhibitors decreased tumor growth and angiogenesis and shift the expression of glycolytic proteins. In conclusion, we reported that Bevacizumab is able to increase the glucose metabolism on cancer cells by abrogating autocrine VEGF . Define the effects of anti-angiogenic drugs at the cellular level can allow us to discover ways to revert acquired resistance to this therapeutic approaches in the future.
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http://dx.doi.org/10.18632/oncotarget.21761DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5732757PMC
November 2017

Serotonin regulates prostate growth through androgen receptor modulation.

Sci Rep 2017 11 13;7(1):15428. Epub 2017 Nov 13.

Life and Health Sciences Research Institute (ICVS), School of Medicine, University of Minho, 4710-057, Braga, Portugal.

Aging and testosterone almost inexorably cause benign prostatic hyperplasia (BPH) in Human males. However, etiology of BPH is largely unknown. Serotonin (5-HT) is produced by neuroendocrine prostatic cells and presents in high concentration in normal prostatic transition zone, but its function in prostate physiology is unknown. Previous evidence demonstrated that neuroendocrine cells and 5-HT are decreased in BPH compared to normal prostate. Here, we show that 5-HT is a strong negative regulator of prostate growth. In vitro, 5-HT inhibits rat prostate branching through down-regulation of androgen receptor (AR). This 5-HT's inhibitory mechanism is also present in human cells of normal prostate and BPH, namely in cell lines expressing AR when treated with testosterone. In both models, 5-HT's inhibitory mechanism was replicated by specific agonists of 5-Htr1a and 5-Htr1b. Since peripheral 5-HT production is specifically regulated by tryptophan hydroxylase 1(Tph1), we showed that Tph1 knockout mice present higher prostate mass and up-regulation of AR when compared to wild-type, whereas 5-HT treatment restored the prostate weight and AR levels. As 5-HT is decreased in BPH, we present here evidence that links 5-HT depletion to BPH etiology through modulation of AR. Serotoninergic prostate pathway should be explored as a new therapeutic target for BPH.
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http://dx.doi.org/10.1038/s41598-017-15832-5DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5684231PMC
November 2017

AKT can modulate the response of HNSCC cells to irreversible EGFR inhibitors.

Oncotarget 2017 Aug 7;8(32):53288-53301. Epub 2017 Jun 7.

Molecular Oncology Research Center, Barretos Cancer Hospital, Barretos, Brazil.

Epidermal growth factor receptor (EGFR) is overexpressed in up to 90% of head and neck squamous cell carcinoma (HNSCC) tumors. Cetuximab is the first targeted (anti-EGFR) therapy approved for the treatment of HNSCC patients. However, its efficacy is limited due to primary and secondary resistance, and there is no predict biomarkers of response. New generation of EGFR inhibitors with pan HER targeting and irreversible action, such as afatinib and allitinib, represents a significant therapeutic promise. In this study, we intend to compare the potential cytotoxicity of two anti-EGFR inhibitors (afatinib and allitinib) with cetuximab and to identify potential predictive biomarkers of response in a panel of HNSCC cell lines. The mutational analysis in the eight HNSCC cell lines revealed an mutation (p.H773Y) and gene amplification in the HN13 cells. According to the growth inhibition score (GI), allitinib was the most cytotoxic drug, followed by afatinib and finally cetuximab. The higher AKT phosphorylation level was associated with resistance to anti-EGFR agents. Therefore, we further performed drug combinations with anti-AKT agent (MK2206) and gene editing, which demonstrated afatinib and allitinib sensitivity restored. Additionally, analysis of TCGA database showed that AKT1 overexpression was present in 14.7% (41/279) of HNSCC cases, and was associated with perineural invasion in advanced stage. In conclusion, allitinib presented a greater cytotoxic profile when compared to afatinib and cetuximab. AKT pathway constitutes a predictive marker of allitinib response and combination with AKT inhibitors could restore response and increase treatment success.
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http://dx.doi.org/10.18632/oncotarget.18395DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5581110PMC
August 2017

Monocarboxylate transporter 1 is a key player in glioma-endothelial cell crosstalk.

Mol Carcinog 2017 Dec 25;56(12):2630-2642. Epub 2017 Aug 25.

Life and Health Sciences Research Institute (ICVS), School of Health Sciences, University of Minho, Campus Gualtar, Braga, Portugal.

Glioblastoma (GBM) is one of the most glycolytic and angiogenic human tumors, characteristics that contribute to the poor prognosis associated with this type of tumor. A lactate shuttle has been described between tumor cells and endothelial cells (ECs), with the monocarboxylate transporters (MCTs) acting as important players in this tumor-EC communication. In this study, we aimed to understand how the tumor microenvironment modulates EC metabolism, and to characterize the role of MCTs in the glioma-brain EC crosstalk. Exposure of human brain microvascular ECs (HBMEC) to GBM cell-conditioned media increased the expression of MCT1, which corresponded to activation of oxidative metabolism and an increase in angiogenic capacity, as determined by increased proliferation, migration, and vessel assembly. Lactate depletion from the microenvironment or inhibition of lactate uptake in HBMEC induced an increase in lactate production and a decrease in proliferation, migration, and vessel assembly. Moreover, addition of lactate to HBMEC media promoted activation of AKT and AMPK pathways and increased expression in NFκB, HIF-1α, and the lactate receptor GPR81. Here, we demonstrate a role for MCT1 as a mediator of lactate signaling between glioma cells and brain ECs. Our results suggest that MCT1 can mediate EC metabolic reprograming, proliferation, and vessel sprouting in response to tumor signaling. Thus, targeting MCT1 in both tumor cells and brain EC may be a promising therapeutic strategy for the treatment of GBM.
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http://dx.doi.org/10.1002/mc.22707DOI Listing
December 2017

HER Family Receptors are Important Theranostic Biomarkers for Cervical Cancer: Blocking Glucose Metabolism Enhances the Therapeutic Effect of HER Inhibitors.

Theranostics 2017 15;7(3):717-732. Epub 2017 Jan 15.

Life and Health Sciences Research Institute (ICVS), School of Medicine, University of Minho, Braga, Portugal;; ICVS/3B's - PT Government Associate Laboratory, Braga/Guimarães, Portugal;; Molecular Oncology Research Center (CPOM), Barretos Cancer Hospital, Barretos, São Paulo, Brazil.

Persistent HPV infection alone is not sufficient for cervical cancer development, which requires additional molecular alterations for tumor progression and metastasis ultimately leading to a lethal disease. In this study, we performed a comprehensive analysis of HER family receptor alterations in cervical adenocarcinoma. We detected overexpression of HER protein, mainly HER2, which was an independent prognostic marker for these patients. By using and approaches, we provided evidence that HER inhibitors, allitinib and lapatinib, were effective in reducing cervical cancer aggressiveness. Furthermore, combination of these drugs with glucose uptake blockers could overcome the putative HIF1-α-mediated resistance to HER-targeted therapies. Thus, we propose that the use of HER inhibitors in association with glycolysis blockers can be a potentially effective treatment option for HER-positive cervical cancer patients.
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http://dx.doi.org/10.7150/thno.17154DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5327645PMC
October 2017

Hypoxia-mediated upregulation of MCT1 expression supports the glycolytic phenotype of glioblastomas.

Oncotarget 2016 Jul;7(29):46335-46353

Life and Health Sciences Research Institute (ICVS), School of Health Sciences, University of Minho, Braga, Portugal.

Background: Glioblastomas (GBM) present a high cellular heterogeneity with conspicuous necrotic regions associated with hypoxia, which is related to tumor aggressiveness. GBM tumors exhibit high glycolytic metabolism with increased lactate production that is extruded to the tumor microenvironment through monocarboxylate transporters (MCTs). While hypoxia-mediated regulation of MCT4 has been characterized, the role of MCT1 is still controversial. Thus, we aimed to understand the role of hypoxia in the regulation of MCT expression and function in GBM, MCT1 in particular.

Methods: Expression of hypoxia- and glycolytic-related markers, as well as MCT1 and MCT4 isoforms was assessed in in vitro and in vivo orthotopic glioma models, and also in human GBM tissues by immunofluorescence/immunohistochemistry and Western blot. Following MCT1 inhibition, either pharmacologically with CHC (α-cyano-4-hydroxynnamic acid) or genetically with siRNAs, we assessed GBM cell viability, proliferation, metabolism, migration and invasion, under normoxia and hypoxia conditions.

Results: Hypoxia induced an increase in MCT1 plasma membrane expression in glioma cells, both in in vitro and in vivo models. Additionally, treatment with CHC and downregulation of MCT1 in glioma cells decreased lactate production, cell proliferation and invasion under hypoxia. Moreover, in the in vivo orthotopic model and in human GBM tissues, there was extensive co-expression of MCT1, but not MCT4, with the GBM hypoxia marker CAIX.

Conclusion: Hypoxia-induced MCT1 supports GBM glycolytic phenotype, being responsible for lactate efflux and an important mediator of cell survival and aggressiveness. Therefore, MCT1 constitutes a promising therapeutic target in GBM.
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http://dx.doi.org/10.18632/oncotarget.10114DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5216802PMC
July 2016

Cytotoxicity of allitinib, an irreversible anti-EGFR agent, in a large panel of human cancer-derived cell lines: KRAS mutation status as a predictive biomarker.

Cell Oncol (Dordr) 2016 Jun 26;39(3):253-63. Epub 2016 Feb 26.

Molecular Oncology Research Center, Barretos Cancer Hospital, Rua Antenor Duarte Villela, 1331, CEP 14784 400, Barretos, São Paulo, Brazil.

Background: The epidermal growth factor receptor (EGFR) is a member of the HER family of growth factors that activates several intracellular signaling pathways promoting proliferation and survival. EGFR over-expression is frequently associated with gene mutation or amplification, thereby constituting a major target for molecular therapies. Recently, a new generation of EGFR inhibitors has been developed with pan-HER properties and irreversible actions. Allitinib® (AST1306) is an orally active, highly selective irreversible inhibitor of the HER family of receptor tyrosine kinases with promising efficacies. In the present study we aimed to investigate the cytotoxicity of allitinib in a large panel of human cancer-derived cell lines and to correlate its efficacy to the mutational status of the EGFR, KRAS, BRAF, PI3KCA and PTEN genes. In addition, we aimed to evaluate the functional role of KRAS mutations in the response to this new inhibitor.

Results: In total 76 different cancer-derived cell lines, representing 11 distinct histological types, were analyzed and classified into three groups: highly sensitive (HS), moderately sensitive (MS) and resistant (R). We found that 28 (36.8 %) cancer-derived cell lines exhibited a HS phenotype, 19 (25.0 %) a MS phenotype and 29 (38.1 %) a R phenotype. Allitinib showed a stronger cytotoxicity in head and neck, esophageal, melanoma and lung cancer-derived cell lines. We found that KRAS mutations were significantly associated with the R phenotype. To substantiate these results, an allitinib-sensitive lung cancer-derived cell line (H292) was transfected with plasmids carrying the two most common activating KRAS mutations (p.G12D and p.G12S). We found that both mutations reverted the allitinib-sensitive phenotype in these cells.

Conclusions: The current study represents the largest in vitro assessment of allitinib cytotoxicity performed to date. Through this study, we identified cancer types that could potentially benefit from this drug. Additionally, our findings suggest that prevalent KRAS mutations constitute potential predictive biomarkers for allitinib response.
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http://dx.doi.org/10.1007/s13402-016-0270-zDOI Listing
June 2016

Raf Kinase Inhibitor Protein Expression and Prognostic Value in Soft Tissue Sarcomas.

Pathobiology 2016 10;83(1):41-6. Epub 2016 Feb 10.

Life and Health Sciences Research Institute (ICVS), School of Health Sciences, University of Minho, Braga, Portugal.

Objective: Soft tissue sarcomas (STSs) are heterogeneous tumors displaying multiple and complex molecular abnormalities with no specific pattern. Despite current therapeutic advances, the patients with STS still have a poor outcome, which makes it necessary to find out new prognostic markers. The Raf kinase inhibitory protein (RKIP) has been associated with prognosis in several human neoplasms; however, its role in STS is unknown.

Methods: In the present study RKIP expression was assessed by immunohistochemistry in a series of 87 STSs, and its expression profile was associated with the patients' pathological parameters.

Results: We found that RKIP is expressed in the cytoplasm of the great majority of cases, and absent in only approximately 18% of cases (16/87). Importantly, we observed that loss of RKIP expression was associated with poor outcome, constituting an independent prognostic marker.

Conclusion: This is the first study assessing RKIP expression levels in STS. We showed that loss of RKIP expression is present in a small subset of cases; however, its absence was associated with poor survival and may be a potential marker for STS prognosis.
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http://dx.doi.org/10.1159/000441227DOI Listing
November 2016

Molecular profiling, including TERT promoter mutations, of acral lentiginous melanomas.

Melanoma Res 2016 Apr;26(2):93-9

aMolecular Oncology Research Center bDepartment of Surgery, Melanoma and Sarcoma Unity cDepartment of Pathology, Barretos Cancer Hospital, Barretos, São Paulo, Brazil dInstitute of Pathology and Molecular Immunology of University of Porto, (IPATIMUP), Porto eLife and Health Sciences Research Institute (ICVS), Health Sciences School, University of Minho fICVS/3B's-PT Government Associate Laboratory, Braga, Portugal.

Acral lentiginous melanoma (ALM) is the less common subtype with singular characterization. TERT (human telomerase reverse transcriptase) promoter mutations have being described as recurrent in melanomas and infrequent in ALM, but their real incidence and clinical relevance is unclear. The objectives of this study were to describe the prevalence of TERT promoter mutations in ALM, and correlate with the molecular profile of other drive genes and clinical features. Sixty-one samples from 48 patients with ALM were analyzed. After DNA isolation, the mutation profiles of the hotspot region of BRAF, NRAS, KIT, PDGFRA, and TERT genes were determined by PCR amplification followed by direct Sanger sequencing. KIT, PDGFRA, and VEGFR2 gene amplification was performed by quantitative PCR. Clinical information such as survival, clinical stage, and Breslow tumor classification were obtained from medical records. TERT promoter mutations were found in 9.3% of the cases, BRAF in 10.3%, NRAS in 7.5%, KIT in 20.7%, and PDGFRA in 14.8% of ALM. None of the cases showed KIT, PDGFRA, or VEGFR2 gene amplification. We found an association between KIT mutations and advanced Clark level (IV and V, P=0.043) and TERT promoter mutations with low mitotic index. No other significant associations were observed between mutation profile and patients' clinical features nor survival rates. Oncogenic TERT promoter mutations are present in a fraction of ALMs. No relevant associations were found between TERT mutation status and clinical/molecular features nor survival. Mutations of KIT and PDGFRA are the most common genetic alterations, and they can be therapeutic targets for these patients.
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http://dx.doi.org/10.1097/CMR.0000000000000222DOI Listing
April 2016

Glucose Addiction in Cancer Therapy: Advances and Drawbacks.

Curr Drug Metab 2015 ;16(3):221-42

While normal differentiated cells primarily use mitochondrial respiration to generate the required energy for cellular processes, most cancer cells rely on glycolysis, even in sufficient oxygen conditions. This phenomenon is known as the "Warburg effect" or aerobic glycolysis and the metabolic reprogramming of cancer cells towards this altered energy metabolism is currently recognized as one of the "hallmarks of cancer". Aerobic glycolysis underlies the rapid growth of tumor cells, with high rates of glucose consumption and lactic acid production, leading to cellular acidosis. Metabolic reprogramming renders cancer cells dependent on specific metabolic enzymes or pathways that could be exploited in cancer therapy. The development of treatments that target tumor glucose metabolism is receiving renewed attention, with several drugs targeting metabolic pathways currently in clinical trials. The search for suitable targets, however, is limited by the high plasticity of the metabolic network that can induce compensatory routes. Deregulated glucose metabolism is a prominent feature associated with resistance to classical chemotherapy or oncogene-targeted therapies, strengthening the clinical potential of combining these therapies with glycolysis inhibitors. The aim of this review is to compare the advances of different therapeutic strategies targeting the glucose "addiction" of tumor cells, highlighting their potential as effective weapons against cancer. We further discuss recent evidence for the involvement of glucose metabolism as a compensatory response to the use of drugs that target different signaling pathways, where the combination with glycolysis inhibitors could prove extraordinarily useful.
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http://dx.doi.org/10.2174/1389200216666150602145145DOI Listing
May 2016

AXL as a modulator of sunitinib response in glioblastoma cell lines.

Exp Cell Res 2015 Mar 28;332(1):1-10. Epub 2015 Jan 28.

Life and Health Sciences Research Institute (ICVS), Health Sciences School, University of Minho, Braga, Portugal; ICVS/3B's-PT Government Associate Laboratory, Braga/Guimarães, Portugal; Molecular Oncology Research Center, Barretos Cancer Hospital, Barretos, São Paulo, Brazil. Electronic address:

Receptor tyrosine kinase (RTK) targeted therapy has been explored for glioblastoma treatment. However, it is unclear which RTK inhibitors are the most effective and there are no predictive biomarkers available. We recently identified the RTK AXL as a putative target for the pan-RTK inhibitors cediranib and sunitinib, which are under clinical trials for glioblastoma patients. Here, we provide evidence that AXL activity can modulate sunitinib response in glioblastoma cell lines. We found that AXL knockdown conferred lower sensitivity to sunitinib by rescuing migratory defects and inhibiting apoptosis in cells expressing high AXL basal levels. Accordingly, overactivation of AXL by its ligand GAS6 rendered AXL positive glioblastoma cells more sensitive to sunitinib. AXL knockdown induced a cellular rewiring of several growth signaling pathways through activation of RTKs, such as EGFR, as well as intracellular pathways such as MAPK and AKT. The combination of sunitinib with a specific AKT inhibitor reverted the resistance of AXL-silenced cells to sunitinib. Together, our results suggest that sunitinib inhibits AXL and AXL activation status modulates therapy response of glioblastoma cells to sunitinib. Moreover, it indicates that combining sunitinib therapy with AKT pathway inhibitors could overcome sunitinib resistance.
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http://dx.doi.org/10.1016/j.yexcr.2015.01.009DOI Listing
March 2015

Silencing of the tumor suppressor gene WNK2 is associated with upregulation of MMP2 and JNK in gliomas.

Oncotarget 2015 Jan;6(3):1422-34

ICVS-Life and Health Sciences Research Institute, School of Health Sciences, University of Minho, Campus Gualtar, Braga 4710-057, Portugal.

Matrix metalloproteinases (MMPs) are proteolytic enzymes that degrade extracellular matrix (ECM), thus assisting invasion. Upregulation of MMPs, frequently reported in gliomas, is associated with aggressive behavior. WNK2 is a tumor suppressor gene expressed in normal brain, and silenced by promoter methylation in gliomas. Patients without WNK2 exhibited poor prognosis, and its downregulation was associated with increased glioma cell invasion. Here we showed that MMP2 expression and activity are increased in glioma cell lines that do not express WNK2. Also, WNK2 inhibited JNK, a process associated with decreasing levels of MMP2. Thus, WNK2 promoter methylation and silencing in gliomas is associated with increased JNK activation and MMP2 expression and activity, thus explaining in part tumor cell invasion potential.
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http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4359304PMC
http://dx.doi.org/10.18632/oncotarget.2805DOI Listing
January 2015

Analysis of a synchronous gliosarcoma and meningioma with long survival: A case report and review of the literature.

Surg Neurol Int 2013 27;4:151. Epub 2013 Nov 27.

Life and Health Sciences Research Institute (ICVS), School of Health Sciences, University of Minho, Braga, Portugal ; Molecular Oncology Research Center, Barretos Cancer Hospital, Barretos, S. Paulo, Brazil ; ICVS/3B's - PT Government Associate Laboratory, Braga/Guimarães, Portugal.

Background: The simultaneous occurrence of multiple intracranial neoplasms has been reported, especially in genetic familial syndromes and after cranial irradiation. In the absence of these etiologic factors, some reports showed simultaneous occurrence of glioblastoma and meningioma but the association between gliosarcoma and meningioma is unknown.

Case Description: We report a case of a 51-year-old woman with synchronous gliosarcoma and meningioma in whom extensive immunohistochemical characterization and molecular profile was performed. The gliosarcoma recurred 21 months after the first resection, reaching 3 years of overall survival. A molecular characterization of all three lesions was performed. None of the lesions showed the presence of mutations in TP53 and BRAF genes. MGMT analysis showed the presence of loss of expression associated with promoter hypermethylation in both gliosarcoma lesions. EGFR overexpression and gene amplification was found only in the recurrent gliosarcoma.

Conclusion: The immunohistochemistry and molecular data of this unique case, suggest the distinct clonal origin of meningioma and gliosarcoma lesions, and the association of MGMT methylation with the presumable favorable prognosis observed.
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http://dx.doi.org/10.4103/2152-7806.122229DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3872647PMC
January 2014

In Vitro and In Vivo Analysis of RTK Inhibitor Efficacy and Identification of Its Novel Targets in Glioblastomas.

Transl Oncol 2013 Apr 1;6(2):187-96. Epub 2013 Apr 1.

Life and Health Sciences Research Institute (ICVS), Health Sciences School, University of Minho, Braga, Portugal ; ICVS/3B's, PT Government Associate Laboratory, Braga, Guimarães, Portugal ; Molecular Oncology Research Center, Barretos Cancer Hospital, Barretos, São Paulo, Brazil.

Treatment for glioblastoma consists of radiotherapy and temozolomide-based chemotherapy. However, virtually all patients recur, leading to a fatal outcome. Receptor tyrosine kinase (RTK)-targeted therapy has been the focus of attention in novel treatment options for these patients. Here, we compared the efficacy of imatinib, sunitinib, and cediranib in glioblastoma models. In the present work, the biologic effect of the drugs was screened by viability, cell cycle, apoptosis, migration, and invasion in vitro assays or in vivo by chick chorioallantoic membrane assay. Intracellular signaling was assessed by Western blot and the RTK targets were identified using phospho-RTK arrays. The amplified status of KIT, PDGFRA, and VEGFR2 genes was assessed by quantitative polymerase chain reaction. In a panel of 10 glioblastoma cell lines, we showed that cediranib was the most potent. In addition, cediranib and sunitinib synergistically sensitize the cells to temozolomide. Cediranib efficacy was shown to associate with higher cytostatic and unique cytotoxic effects in vitro and both antitumoral and antiangiogenic activity in vivo, which could associate with its great capacity to inhibit mitogen-activated protein kinase (MAPK) and AKT pathways. The molecular status of KIT, PDGFRA, and VEGFR2 did not predict glioblastoma cell responsiveness to any of the RTK inhibitors. Importantly, phospho-RTK arrays revealed novel targets for cediranib and sunitinib therapy. In conclusion, the novel targets found may be of value as future biomarkers for therapy response in glioblastoma and lead to the rational selection of patients for effective molecular targeted treatment.
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http://dx.doi.org/10.1593/tlo.12400DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3610556PMC
April 2013

RKIP inhibition in cervical cancer is associated with higher tumor aggressive behavior and resistance to cisplatin therapy.

PLoS One 2013 19;8(3):e59104. Epub 2013 Mar 19.

Life and Health Sciences Research Institute (ICVS), Health Sciences School, University of Minho, Braga, Portugal.

Cervical cancer is one of the most common cancers in women worldwide, being high-risk group the HPV infected, the leading etiological factor. The raf kinase inhibitory protein (RKIP) has been associated with tumor progression and metastasis in several human neoplasms, however its role on cervical cancer is unclear. In the present study, 259 uterine cervix tissues, including cervicitis, cervical intraepithelial lesions and carcinomas, were analyzed for RKIP expression by immunohistochemistry. We found that RKIP expression was significantly decreased during malignant progression, being highly expressed in non-neoplastic tissues (54% of the samples; 73/135), and expressed at low levels in the cervix invasive carcinomas (∼15% (19/124). Following in vitro downregulation of RKIP, we observed a viability and proliferative advantage of RKIP-inhibited cells over time, which was associated with an altered cell cycle distribution and higher colony number in a colony formation assay. An in vitro wound healing assay showed that RKIP abrogation is associated with increased migratory capability. RKIP downregulation was also associated with an increased vascularization of the tumors in vivo using a CAM assay. Furthermore, RKIP inhibition induced cervical cancer cells apoptotic resistance to cisplatin treatment. In conclusion, we described that RKIP protein is significantly depleted during the malignant progression of cervical tumors. Despite the lack of association with patient clinical outcome, we demonstrate, in vitro and in vivo, that loss of RKIP expression can be one of the factors that are behind the aggressiveness, malignant progression and chemotherapy resistance of cervical cancer.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0059104PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3602518PMC
September 2013

Low RKIP expression associates with poor prognosis in bladder cancer patients.

Virchows Arch 2013 Apr 6;462(4):445-53. Epub 2013 Mar 6.

Life and Health Sciences Research Institute-ICVS, University of Minho, Braga, Portugal.

Urothelial bladder cancer (UBC) is a heterogeneous type of disease. It is urgent to screen biomarkers of tumour aggressiveness in order to clarify the clinical behaviour and to personalize therapy in UBC patients. Raf kinase inhibitory protein (RKIP) is a metastasis suppressor, and its downregulation is associated with metastatic events in an increasing number of solid tumours. We evaluated the clinical and prognostic significance of RKIP expression in patients with high risk of progression UBC. Using immunohistochemistry, we determined RKIP expression levels in a series of 81 patients with high-grade pT1/pTis or muscle-invasive UBC. Staining of CD31 and D2-40 was used to assess blood and lymphatic vessels, in order to distinguish between blood and lymphatic vessel invasion (LVI). We found that 90 % of pT1/pTis tumours, 94 % of non-muscle invasive papillary tumours and 76 % of the cases without LVI occurrence expressed RKIP in >10 % of cells. In this group, we observed a subgroup of tumours (42 %) in which the tumour centre was significantly more intensely stained than the invasion front. This heterogeneous pattern was observed in 63 % of the cases with LVI. Low RKIP expression was associated with poorer 5-year disease-free and overall survival rates, and remained as an independent prognostic factor for disease-free survival. Loss of RKIP expression may be an important prognostic factor for patients with high risk of progression bladder cancer.
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http://dx.doi.org/10.1007/s00428-013-1388-2DOI Listing
April 2013

Monocarboxylate transporters (MCTs) in gliomas: expression and exploitation as therapeutic targets.

Neuro Oncol 2013 Feb 20;15(2):172-88. Epub 2012 Dec 20.

Life and Health Sciences Research Institute, School of Health Sciences, University of Minho, Campus de Gualtar, 4710-057 Braga, Portugal.

Background: Gliomas exhibit high glycolytic rates, and monocarboxylate transporters (MCTs) play a major role in the maintenance of the glycolytic metabolism through the proton-linked transmembrane transport of lactate. However, their role in gliomas is poorly studied. Thus, we aimed to characterize the expression of MCT1, MCT4, and their chaperone CD147 and to assess the therapeutic impact of MCT inhibition in gliomas.

Methods: MCTs and CD147 expressions were characterized by immunohistochemistry in nonneoplastic brain and glioma samples. The effect of CHC (MCT inhibitor) and MCT1 silencing was assessed in in vitro and in vivo glioblastoma models.

Results: MCT1, MCT4, and CD147 were overexpressed in the plasma membrane of glioblastomas, compared with diffuse astrocytomas and nonneoplastic brain. CHC decreased glycolytic metabolism, migration, and invasion and induced cell death in U251 cells (more glycolytic) but only affected proliferation in SW1088 (more oxidative). The effectiveness of CHC in glioma cells appears to be dependent on MCT membrane expression. MCT1 downregulation showed similar effects on different glioma cells, supporting CHC as an MCT1 inhibitor. There was a synergistic effect when combining CHC with temozolomide treatment in U251 cells. In the CAM in vivo model, CHC decreased the size of tumors and the number of blood vessels formed.

Conclusions: This is the most comprehensive study reporting the expression of MCTs and CD147 in gliomas. The MCT1 inhibitor CHC exhibited anti-tumoral and anti-angiogenic activity in gliomas and, of importance, enhanced the effect of temozolomide. Thus, our results suggest that development of therapeutic approaches targeting MCT1 may be a promising strategy in glioblastoma treatment.
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http://dx.doi.org/10.1093/neuonc/nos298DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3548586PMC
February 2013

Absence of RKIP expression is an independent prognostic biomarker for gastric cancer patients.

Oncol Rep 2013 Feb 10;29(2):690-6. Epub 2012 Dec 10.

Life and Health Sciences Research Institute, Health Sciences School, University of Minho, Braga, Portugal.

Gastric cancer is a leading cause of cancer-related mortality, and the presence of lymph node metastasis an important prognostic factor. Downregulation of RKIP has been associated with tumor progression and metastasis in several types of neoplasms, being currently categorized as a metastasis suppressor gene. Our aim was to determine the expression levels of RKIP in gastric tissues and to evaluate its impact in the clinical outcome of gastric carcinoma patients. RKIP expression levels were studied by immunohistochemistry in a series of gastric tissues. Overall, we analysed 222 non-neoplastic gastric tissues, 152 primary tumors and 42 lymph node metastasis samples. We observed that RKIP was highly expressed in ~83% of non-neoplastic tissues (including normal tissue and metaplasia), was lost in ~56% of primary tumors and in ~90% of lymph node metastasis samples. Loss of RKIP expression was significantly associated with several markers of poor clinical outcome, including the presence of lymph node metastasis. Furthermore, the absence of RKIP protein constitutes an independent prognostic marker for these patients. In conclusion, RKIP expression is significantly lost during gastric carcinoma progression being almost absent in lymph node metastasis samples. Of note, we showed that the absence of RKIP expression is associated with poor outcome features of gastric cancer patients, this being also an independent prognostic marker.
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http://dx.doi.org/10.3892/or.2012.2179DOI Listing
February 2013
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