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Autotaxin (ATX) is a secreted enzyme responsible for producing lysophosphatidic acid (LPA). The ATX/LPA signaling axis is typically activated in wound healing and tissue repair processes. The ATX/LPA axis is highjacked and upregulated in the progression and persistence of several chronic inflammatory diseases, including cancer. As ATX inhibitors are now progressing to clinical testing, innovative diagnostic tools such as positron emission tomography (PET) are needed to measure ATX expression in vivo accurately. The radiotracer, [18F]PRIMATX, was recently developed and tested for PET imaging of ATX in vivo in a murine melanoma model. The goal of the present work was to further validate [18F]PRIMATX as a PET imaging agent by analyzing its in vivo metabolic stability and suitability for PET imaging of ATX in models of human 8305C thyroid tumor and murine 4T1 breast cancer. [18F]PRIMATX displayed favorable metabolic stability in vivo (65% of intact radiotracer after 60 min p.i.) and provided sufficient tumor uptake profiles in both tumor models. Radiotracer uptake could be blocked by 8-12% in 8305C thyroid tumors in the presence of ATX inhibitor AE-32-NZ70 as determined by PET and ex vivo biodistribution analyses. [18F]PRIMATX also showed high brain uptake, which was reduced by 50% through the administration of ATX inhibitor AE-32-NZ70. [18F]PRIMATX is a suitable radiotracer for PET imaging of ATX in the brain and peripheral tumor tissues.

Emerging studies are reporting associations between skeletal muscle abnormalities and survival in cancer patients. Cancer prognosis is associated with depletion of essential fatty acids in erythrocytes and plasma in humans. However the relationship between skeletal muscle membrane fatty acid composition and survival is unknown. This study investigates the relationship between fatty acid content of phospholipids in skeletal muscle and survival in cancer patients. Rectus abdominis biopsies were collected during cancer surgery from 35 patients diagnosed with cancer. Thin-layer and gas chromatography were used for quantification of phospholipid fatty acids. Cutpoints for survival were defined using optimal stratification. Median survival was between 450 and 500 days when patients had arachidonic acid (AA) eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) in muscle phospholipid below the cut-point compared to 720–800 days for patients above. Cox regression analysis revealed that low amounts of AA, EPA and DHA are risk factors for death. The risk of death remained significant for AA [HR 3.5 (1.11–10.87), p = 0.03], EPA [HR 3.92 (1.1–14.0), p = 0.04] and DHA [HR 4.08 (1.1–14.6), p = 0.03] when adjusted for sex. Lower amounts of essential fatty acids in skeletal muscle membrane is a predictor of survival in cancer patients. These results warrant investigation to restore bioactive fatty acids in people with cancer.

Human cytomegalovirus (HCMV) infects 40–70% of adults in developed countries. HCMV proteins and DNA are detected in tumors and metastases, suggesting an association with increased invasion. We investigated HCMV infection in human breast cancer cell lines compared to fibroblasts, a component of tumors, and the role of platelet-derived growth factor receptor-α (PDGFRα). HCMV productively infected HEL299 fibroblasts and, to a lesser extent, Hs578T breast cancer cells. Infection of another triple-negative cell line, MDA-MB-231, and also MCF-7 cells, was extremely low. These disparate infection rates correlated with expression of PDGFRA, which facilitates HCMV uptake. Increasing PDGFRA expression in T-47D breast cancer and BCPAP thyroid cancer cells markedly increased HCMV infection. Conversely, HCMV infection decreased PDGFRA expression, potentially attenuating signaling through this receptor. HCMV infection of fibroblasts promoted the secretion of proinflammatory factors, whereas an overall decreased secretion of inflammatory factors was observed in infected Hs578T cells. We conclude that HCMV infection in tumors will preferentially target tumor-associated fibroblasts and breast cancer cells expressing PDGFRα. HCMV infection in the tumor microenvironment, rather than cancer cells, will increase the inflammatory milieu that could enhance metastasis involving lysophosphatidate.

Background: Emerging studies are reporting associations between skeletal muscle abnormalities and survival in cancer patients. Cancer prognosis is associated with depletion of essential fatty acids in membrane of blood plasma and serum in humans, however the relationship between skeletal muscle membrane fatty acid composition and survival is unknown. This study investigates the relationship between fatty acid content of phospholipids in skeletal muscle and survival in cancer patients.Methods: Rectus abdominis biopsies were collected during cancer surgery from 35 patients diagnosed with cancer. Thin-layer and gas chromatography were used for quantification of phospholipid fatty acid content of the muscle. Cut-points for fatty acids were established using optimal stratification.Results: Median survival was between 450-500 days when patients had arachidonic acid (AA) eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) in muscle phospholipid above the cut-point compared to 720-800 days for patients below. Cox regression analysis revealed that low amounts of AA, EPA and DHA are risk factors for death. The risk of death remained significant for AA [HR 3.5 (1.11-10.87), p=0.03], EPA [HR 3.92 (1.1-14.0), p=0.04] and DHA [HR 4.08 (1.1-14.6), p=0.03] when adjusted for sex. Conclusion: Lower amount of essential fatty acids in skeletal muscle membrane is predictor of survival in cancer patients. These results warrant investigation to restore depletion of bioactive fatty acids in cancer.

Fahed A Elian,1 Ubah Are,1 Sunita Ghosh,2,3 Paulo Nuin,1 Tim Footz,1 Todd PW McMullen,4 David N Brindley,5,6 Michael A Walter1 1Department of Medical Genetics, Faculty of Medicine and Dentistry, University of Alberta, Edmonton, AB, Canada; 2Department of Medical Oncology, Faculty of Medicine and Dentistry, University of Alberta, Edmonton, AB, Canada; 3Department of Mathematical and Statistical Sciences, Faculty of Science, University of Alberta, Edmonton, AB, Canada; 4Department of Surgery, Faculty of Medicine and Dentistry, University of Alberta, Edmonton, AB, Canada; 5Department of Biochemistry, Faculty of Medicine and Dentistry, University of Alberta, Edmonton, AB, Canada; 6Cancer Research Institute of Northern Alberta, University of Alberta, Edmonton, AB, CanadaCorrespondence: Michael A WalterDepartment of Medical Genetics, Faculty of Medicine & Dentistry, University of Alberta, 8-32 Medical Sciences Building, Edmonton, AB, T6G 2H7, CanadaTel +1 (780) 492-3028Email mwalter@ualberta.caPurpose: Forkhead box Q1 (FOXQ1) has been shown to contribute to the development and progression of cancers, including ovarian and breast cancer (BC). However, research exploring FOXQ1 expression, copy number variation (CNV), and prognostic value across different BC subtypes is limited. Our purpose was to evaluate FOXQ1 mRNA expression, CNV, and prognostic value across BC subtypes.Materials and Methods: We determined FOXQ1 expression and CNV in BC patient tumors using RT-qPCR and qPCR, respectively. We also analyzed FOXQ1 expression and CNV in BC cell lines in the CCLE database using K-means clustering. The prognostic value of FOXQ1 expression in the TCGA-BRCA database was assessed using univariate and multivariate Cox’s regression analysis as well as using the online tools OncoLnc, GEPIA, and UALCAN.Results: Our analyses reveal that FOXQ1 mRNA is differentially expressed between different subtypes of BC and is significantly decreased in luminal BC and HER2 patients when compared to normal breast tissue samples. Furthermore, analysis of BC cell lines showed that FOXQ1 mRNA expression was independent of CNV. Moreover, patients with low FOXQ1 mRNA expression had significantly poorer overall survival compared to those with high FOXQ1 mRNA expression. Finally, low FOXQ1 expression had a critical impact on the prognostic values of BC patients and was an independent predictor of overall survival when it was adjusted for BC subtypes and to two other FOX genes, FOXF2 and FOXM1.Conclusion: Our study reveals for the first time that FOXQ1 is differentially expressed across BC subtypes and that low expression of FOXQ1 is indicative of poor prognosis in patients with BC.Keywords: transcription factors, mRNA expression, copy number variation, predictive marker

Targeted therapy is increasingly used to manage metastatic papillary thyroid cancer. The focus of the present study was to examine glucose metabolism and tumor responses for thyroid cancer xenografts expressing the glycolytic pathway modulators platelet-derived growth factor receptor (PDGFR) and BRAFV600E. Radiolabelled glucose derivative [18F]FDG was used to analyze the effects of PDGFR blockade with imatinib, BRAF blockade with vemurafenib, as well as combined PDGFR and BRAF blockade in vitro and in vivo with PET. Dynamic PET data was correlated with immunohistochemistry staining and kinetic analysis for facilitative glucose transporter 1 (GLUT1) and hexokinase-II (HK2). Vemurafenib decreased [18F]FDG uptake in BCPAP cells in vitro; however, it was increased by ~70% with imatinib application to BCPAP cells. This metabolic response to tyrosine kinase inhibition required BRAFV600E as it was not seen in cell lines lacking mutated BRAF (TPC1). In xenografts, imatinib therapy in BCPAP thyroid tumour-bearing mice significantly increased [18F]FDG uptake and retention (>30%) in BCPAP tumours with PDGFRβ or both (α+β) isoforms. Kinetic analysis revealed that the increased glucose uptake is a consequence of increased phosphorylation and intracellular trapping of [18F]FDG confirmed by an increase in HK2 protein expression and activity, but not GLUT1 activity. BRAF inhibition alone, or combined PDGFR and BRAF inhibition, reduced (~60%) [18F]FDG uptake in both types of BCPAP (β or α+β) tumours. In terms of tumour growth, combination therapy with imatinib and vemurafenib led to a near abolition of the tumors (~90% reduction), but single therapy for BCPAP with PDGFRα expression was much less effective. In summary, imatinib led to a paradoxical increase of [18F]FDG uptake in xenografts that was reversed through BRAFV600E inhibition. The present data show that metabolic reprogramming in thyroid cancer occurs as a consequence of BRAF-mediated upregulation of HK2 expression that may permit tumour growth with isolated blockade of upstream tyrosine kinase receptors.

Lysophosphatidate (LPA) signaling through 6 receptors is regulated by the balance of LPA production by autotaxin (ATX) vs. LPA degradation by lipid phosphate phosphatases (LPPs). LPA promotes an inflammatory cycle by increasing the synthesis of cyclooxygenase-2 and multiple inflammatory cytokines that stimulate further ATX production. We aimed to determine whether the anti-inflammatory glucocorticoid (GC) dexamethasone (Dex) functions partly by decreasing the ATX-LPA inflammatory cycle in adipose tissue, a major site of ATX secretion. Treatment of human adipose tissue with 10-1000 nM Dex decreased ATX secretion, increased LPP1 expression, and decreased mRNA expressions of IL-6, TNF-α, peroxisome proliferator-activated receptor (PPAR)-γ, and adiponectin. Cotreatment with rosiglitazone (an insulin sensitizer), insulin, or both abolished Dex-induced decreases in ATX and adiponectin secretion, but did not reverse Dex-induced decreases in secretions of 20 inflammatory cytokines and chemokines. Dex-treated mice exhibited lower ATX activity in plasma, brain, and adipose tissue; decreased mRNA levels for LPA and sphingosine 1-phosphate (S1P) receptors in brain; and decreased plasma concentrations of LPA and S1P. Our results establish a novel mechanism for the anti-inflammatory effects of Dex through decreased signaling by the ATX-LPA-inflammatory axis. The GC action in adipose tissue has implications for the pathogenesis of insulin resistance and obesity in metabolic syndrome and breast cancer treatment.-Meng, G., Tang, X., Yang, Z., Zhao, Y., Curtis, J. M., McMullen, T. P. W., Brindley, D. N. Dexamethasone decreases the autotaxin-lysophosphatidate-inflammatory axis in adipose tissue: implications for the metabolic syndrome and breast cancer.

The application of radioactive iodine in differentiated thyroid carcinomas has become more selective in an attempt to decrease morbidity. While ablative success has been documented, it is less clear how changes in radioactive iodine treatment strategies will influence long-term recurrence rates for patients with larger tumors and adverse pathological features, including extrathyroidal extension and nodal metastases.Patients diagnosed between 1995 and 2008 with differentiated thyroid carcinoma treated with thyroidectomy followed by radioactive iodine treatment were eligible. All patients were followed for a minimum of five years using a standardized follow-up protocol requiring both biochemical and imaging assessments for recurrent disease (n = 219). Patients were stratified by initial radioactive iodine activity, and disease-free survival was calculated using the Kaplan-Meier method, with significant differences defined by the log-rank test.In this cohort, 46% of patients had clinical metastases and 74% had primary tumors1.5 cm. Patients who had recurrences were more likely to present with extrathyroidal extension (p = 0.002) and lymph node metastases at diagnosis (p 0.001). Patients presenting with both extrathyroidal extension and lymph node metastases had a significantly worse time to progression if treated with1850 MBq radioactive iodine compared to those patients treated with1850 MBq (25 months vs. 121 months; p = 0.004). The use of lower-activity radioactive iodine ablative therapy was associated with more early recurrences (p = 0.003). Being aged younger or older than 45 years did not impact the time to recurrence nor did the use of level 6 dissection. On multivariate analysis, lymph node metastases at diagnosis and multiple applications of radioactive iodine were linked to increased risk of recurrence. Patients with neither, or only one, adverse pathologic feature had excellent outcomes, regardless of initial ablative activity, with10% of patients recurring over a 10-year time span.Recurrent disease in differentiated thyroid carcinoma is more common in patients treated with low-activity radioactive iodine in patients with lymph node metastases and extrathyroidal extension. These recurrences typically occur within four years of initial treatment. Patients lacking both of these risk factors treated with low radioactive iodine activity (1850 MBq) have excellent outcomes, even after 10 years.

Introduction Receptor tyrosine kinase (RTK) platelet-derived growth factor receptor-alpha (PDGFRα) was recently identified as a molecular switch for dedifferentiation in thyroid cancer that predicts resistance to therapy as well as recurrence of disease in papillary thyroid cancer. Here we describe the radiolabeling and functional characterization of an imaging probe based on a PDGFRα-specific monoclonal antibody (mAb) for immuno-PET imaging of PDGFRα in papillary thyroid cancer. Methods Antibody D13C6 (Cell Signaling) was decorated with chelator NOTA using bioconjugation reaction with 2-(p-NCS-Bz)-NOTA. Radiolabeling was carried out using 40 μg of antibody-NOTA conjugate with 143–223 MBq of [64Cu]CuCl2 in 0.25 M NaOAc (pH 5.5) at 30 °C for 1 h. The reaction mixture was purified with size-exclusion chromatography (PD-10 column). PDGFRα and mock transfected B-CPAP thyroid cancer cells lines for validation of 64Cu-labeled immuno-conjugates were generated using LVX-Tet-On technology. PET imaging was performed in NSG mice bearing bilaterally-induced PDGFRα (+/−) B-CPAP tumors. Results Bioconjugation of NOTA chelator to monoclonal antibody D13C6 resulted in 2.8 ± 1.3 chelator molecules per antibody as determined by radiometric titration with 64Cu. [64Cu]Cu-NOTA-D13C6 was isolated in high radiochemical purity (>98%) and good radiochemical yields (19–61%). The specific activity was 0.9–5.1 MBq/μg. Cellular uptake studies revealed a specific radiotracer uptake in PDGFRα expressing cells compared to control cells. PET imaging resulted in SUVmean values of ~5.5 for PDGFRα (+) and ~2 for PDGFRα (−) tumors, after 48 h p.i.. After 1 h, radiotracer uptake was also observed in the bone marrow (SUVmean ~5) and spleen (SUVmean ~8.5). Conclusion Radiolabeled antibody [64Cu]Cu-NOTA-D13C6 represents a novel and promising radiotracer for immuno-PET imaging of PDGFRα in metastatic papillary thyroid cancer. Advances in knowledge and implications for patient care The presented work has the potential to allow physicians to identify papillary thyroid cancer patients at risk of metastases by using the novel immuno-PET imaging assay based on PDGFRα-targeting antibody [64Cu]Cu-NOTA-D13C6.

We have previously established that adipose tissue adjacent to breast tumors becomes inflamed by tumor-derived cytokines. This stimulates autotaxin (ATX) secretion from adipocytes, whereas breast cancer cells produce insignificant ATX. Lysophosphatidate produced by ATX promotes inflammatory cytokine secretion in a vicious inflammatory cycle, which increases tumor growth and metastasis and decreases response to chemotherapy. We hypothesized that damage to adipose tissue during radiotherapy for breast cancer should promote lysophosphatidic acid (LPA) signaling and further inflammatory signaling, which could potentially protect cancer cells from subsequent fractions of radiation therapy. To test this hypothesis, we exposed rat and human adipose tissue to radiation doses (0.25-5 Gy) that were expected during radiotherapy. This exposure increased mRNA levels for ATX, cyclooxygenase-2, IL-1β, IL-6, IL-10, TNF-α, and LPA1 and LPA2 receptors by 1.8- to 5.1-fold after 4 to 48 h. There were also 1.5- to 2.5-fold increases in the secretion of ATX and 14 inflammatory mediators after irradiating at 1 Gy. Inhibition of the radiation-induced activation of NF-κB, cyclooxygenase-2, poly (ADP-ribose) polymerase-1, or ataxia telangiectasia and Rad3-related protein blocked inflammatory responses to γ-radiation. Consequently, collateral damage to adipose tissue during radiotherapy could establish a comprehensive wound-healing response that involves increased signaling by LPA, cyclooxygenase-2, and other inflammatory mediators that could decrease the efficacy of further radiotherapy or chemotherapy.-Meng, G., Tang, X., Yang, Z., Benesch, M. G. K., Marshall, A., Murray, D., Hemmings, D. G., Wuest, F., McMullen, T. P. W., Brindley, D. N. Implications for breast cancer treatment from increased autotaxin production in adipose tissue after radiotherapy.

Neuroendocrine tumours (NETs) are heterogeneous, with varying presentations and treatment options. To our knowledge, there are no randomized and few long-term studies of patient outcomes. The role of surgical and medical therapy for local, regional and metastatic disease continues to be evaluated in the literature.We conducted a population-based search of the provincial cancer registry to identify patients with gastrointestinal NETs from the stomach, small intestine, colon and rectum diagnosed between 1990 and 2005 and assessed their outcomes.We examined clinicopathological information on the outcomes of 530 patients with gastrointestinal NETs. The overall incidence of NETs increased from 11 per million to 19 per million during the study period. Advancing stage and patient age were associated with poor overall or disease-specific outcomes. Surgery, both curative and palliative, was associated with decreased risk of overall (hazard ratio [HR] 0.5,There has been a significant increase in incidence of gastrointestinal NETs, and advancing patient age, but not sex, is linked to poor outcomes in terms of overall and disease-specific survival. Surgery, both curative and palliative, was associated with decreased risk of overall and disease-specific death. Compared with patients with residual macroscopic disease, patients with distant disease were nearly twice as likely to survive 5 years if they had R0 resections. The use of radioisotope therapy and long-acting octreotide therapy was also associated with improved outcomes overall.Les tumeurs neuroendocrines (TNE) sont hétérogènes, et les tableaux et options thérapeutiques sont variables. À notre connaissance, il n'existe pas d'études randomisées et il y a peu d'études à long terme sur les résultats chez les patients. Le rôle du traitement chirurgical et médicamenteux de la maladie locale, régionale et métastatique continue d'être évalué dans la littérature.Nous avons procédé à une interrogation démographique du Registre provincial du cancer pour recenser les patients atteints de TNE gastro-intestinales provenant de l'estomac, de l'intestin grêle, du côlon et du rectum, diagnostiqués entre 1990 et 2005 et nous avons évalué les résultats.Nous avons examiné les données clinico-pathologiques des résultats enregistrés chez 530 patients atteints de TNE gastro-intestinales. L'incidence globale des TNE a augmenté de 11 par million à 19 par million pendant la période de l'étude. Le stade de la maladie et l'âge avancés ont été associés à des résultats globaux ou spécifiques à la maladie moins favorables. La chirurgie, curative et palliative, a été associée à un risque moindre de décès global (risque relatif [RR] 0.5,L'incidence des TNE gastro-intestinales a significativement augmenté, et l'âge avancé des patients, mais non le sexe, est lié à des résultats défavorables aux plans de la survie globale et spécifique à la maladie. La chirurgie, curative et palliative, a été associée à un risque moindre de décès global et spécifique à la maladie. Comparativement aux patients ayant une maladie macroscopique résiduelle, ceux qui avaient une maladie distale étaient près de 2 fois plus susceptibles de survivre 5 ans s'ils avaient des résections R0. Les traitements par radio-isotopes et octréotide à longue durée d'action ont aussi été associés à une amélioration globale des résultats.

Platelet derived growth receptor alpha (PDGFRA) promotes the epithelial-mesenchymal transition (EMT) in thyroid follicular cells and is linked to lymphatic metastases in papillary thyroid cancer (PTC). We probed the regulatory network of genes linked to PDGFRA and EMT, comparing matched patient primary tumor and metastatic specimens, as well as engineered cell lines and ex vivo primary cultures with and without PDGFRA. Freshly isolated thyroid tumors with or without metastases, with matching neighboring benign or normal tissue, was isolated for comparative transcriptional analysis using a TaqMan Low Density array (TLDA) assay with genes representing important markers of EMT, cellular adhesion, apoptosis, differentiation, senescence, and signal transduction pathways in thyroid cancer. Transfected primary cultures and immortalized cell lines were also analyzed with respect to PDGFRA expression and cell phenotype. We reveal the consistent upregulation of serine protease DPP4 and structural protein SPP1 with the progression of PTC to metastatic disease, as well as with PDGFRA expression. Conversely, epithelial integrity gene TFF3 and transcription factor SOX10 were strongly down-regulated. This gene network also includes important mediators of EMT including DSG1, MMP3, MMP9, and BECN. We observed similar genomic changes in ex vivo normal thyroid cells transfected with PDGFRA that also exhibited a partially dedifferentiated phenotype. In particular, we observed lamellopodia with induction of PDGFRA and illustrate that DPP4 and SPP1 were upregulated in this process, with decreased TFF3 and SOX10 as seen in tissue specimens. PDGFRA did decrease nuclear protein levels of differentiation factor TTF1, but not the transcription of TTF1 and PAX8. We demonstrate that PDGFRA activates EMT pathways and decreases expression of genes favoring epithelial integrity, pushing follicular cells toward a dedifferentiated phenotype. SPP1 and DPP4, previously linked with adverse outcomes in thyroid cancer, appear to be regulated by PDGFRA. PDGFRA expression promotes metastatic disease through multiple EMT levers that favor formation of an invasive phenotype and increased metalloproteinase expression.

// Esther Ekpe-Adewuyi 1 , Ana Lopez-Campistrous 1 , Xiaoyun Tang 2 , David N. Brindley 2 , Todd P. W. McMullen 1, 3 1 Department of Surgery, University of Alberta, Edmonton, Canada 2 Signal Transduction Research Group, Department of Biochemistry, University of Alberta, Edmonton, Canada 3 Division of Surgical Oncology, Cross Cancer Institute, University of Alberta, Edmonton, Canada Correspondence to: Todd P. W. McMullen, email: todd.mcmullen@albertahealthservices.ca Keywords: papillary thyroid cancer (PTC), platelet derived growth factor receptor-alpha (PDGFRα), epithelial to mesenchymal transition (EMT), invadopodia, crenolanib Received: June 30, 2016 Accepted: October 22, 2016 Published: November 11, 2016 ABSTRACT Recently platelet derived growth factor receptor-alpha (PDGFRα) was recognized as a potential target to treat aggressive papillary thyroid cancer given its strong association with lymph node metastases. However, it is unclear how PDGFRα potentiates metastases and if it works through the canonical MAPK pathway traditionally linked to PTC oncogenesis. We explored the phenotypic changes driven by PDGFRα activation in human papillary thyroid cancer (PTC) cells and the downstream signalling cascades through which they are effected. We demonstrate that PDGFRα drives an impressive phenotypic change in PTC cell lines as documented by significant cytoskeletal rearrangement, increased migratory potential, and the formation of invadopodia. Cells lacking PDGFRα formed compact and dense spheroids, whereas cells expressing active PDGFRα exhibited invadopodia in three-dimensional culture. To achieve this, active PDGFRα provoked downstream activation of the MAPK/Erk, PI3K/Akt and STAT3 pathways. We further confirmed the role of PDGFRα as a transformative agent promoting the epithelial to mesenchymal transition of PTC cells, through the augmentation of Snail and Slug expression. Crenolanib, a small molecule inhibitor of PDGFRα, suppressed the levels of Snail and Slug and almost completely reversed all the phenotypic changes. We demonstrate that PDGFRα activation is an essential component that drives aggressiveness in PTC cells, and that the signaling pathways are complex, involving not only the MAPK/Erk but also the PI3K/Akt and STAT3 pathways. This argues for upstream targeting of the PDGFRα given the redundancy of oncogenic pathways in PTC, especially in patients whose tumors over-express this tyrosine kinase receptor.

Dedifferentiation of follicular cells is a central event in resistance to radioactive iodine and patient mortality in papillary thyroid carcinoma (PTC). We reveal that platelet derived growth factor receptor alpha (PDGFRα) specifically drives dedifferentiation in PTC by disrupting the transcriptional activity of thyroid transcription factor-1 (TTF1). PDGFRα activation dephosphorylates TTF1 consequently shifting the localization of this transcription factor from the nucleus to the cytoplasm. TTF1 is required for follicular cell development and disrupting its function abrogates thyroglobulin production and sodium iodide transport. PDGFRα also promotes a more invasive and migratory cell phenotype with a dramatic increase in xenograft tumor formation. In patient tumors we confirm that nuclear TTF1 expression is inversely proportional to PDGFRα levels. Patients exhibiting PDGFRα at time of diagnosis are three times more likely to exhibit nodal metastases and are 18 times more likely to recur within 5 years than those patients lacking PDGFRα expression. Moreover, high levels of PDGFRα and low levels of nuclear TTF1 predict resistance to radioactive iodine therapy. We demonstrate in SCID xenografts that focused PDGFRα blockade restores iodide transport and decreases tumor burden by > 50%. Focused PDGFRα inhibitors, combined with radioactive iodine, represent an additional avenue for treating patients with aggressive variants of PTC., Graphical Abstract Image 1, Highlights • PDGFRα induces dedifferentiation of papillary thyroid cancer cells. • This depends on decreased phosphorylation and decreased nuclear targeting of TTF1. • Loss of nuclear TTF1 decreases thyroglobulin production and NaI transport. • PDGFRα expression is prognostic of PTC recurrence and treatment resistance. • Blocking PDGFRα activation is a potential therapeutic target. Treatment of papillary thyroid cancer historically relied upon a combination of surgery and radioactive iodine ablation with few alternatives if the disease progresses. We found that platelet derived growth factor receptor alpha (PDGFRα) is a key driver of metastatic disease and resistance to radioactive iodine therapy. PDGFRα expression can be tested in tumor specimens to predict aggressive disease. In addition, we show that targeting PDGFRα could restore sensitivity to radioactive iodine treatment that might slow disease growth and spread.

We recently showed that radiation-induced DNA damage in breast adipose tissue increases autotaxin secretion, production of lysophosphatidate (LPA) and expression of LPA1/2 receptors. We also established that dexamethasone decreases autotaxin production and LPA signaling in non-irradiated adipose tissue. In the present study, we showed that dexamethasone attenuated the radiation-induced increases in autotaxin activity and the concentrations of inflammatory mediators in cultured human adipose tissue. We also exposed a breast fat pad in mice to three daily 7.5 Gy fractions of X-rays. Dexamethasone attenuated radiation-induced increases in autotaxin activity in plasma and mammary adipose tissue and LPA1 receptor levels in adipose tissue after 48 h. DEX treatment during five daily fractions of 7.5 Gy attenuated fibrosis by ~70% in the mammary fat pad and underlying lungs at 7 weeks after radiotherapy. This was accompanied by decreases in CXCL2, active TGF-&beta, 1, CTGF and Nrf2 at 7 weeks in adipose tissue of dexamethasone-treated mice. Autotaxin was located at the sites of fibrosis in breast tissue and in the underlying lungs. Consequently, our work supports the premise that increased autotaxin production and lysophosphatidate signaling contribute to radiotherapy-induced breast fibrosis and that dexamethasone attenuated the development of fibrosis in part by blocking this process.

Rapidly accumulating evidence implicates forkhead box C1 (FOXC1) in Triple Negative Breast Cancer (TNBC), particularly in Basal-Like Breast cancer (BLBC). Recently additional studies have demonstra...

A quarter-century after the discovery of autotaxin in cell culture, the autotaxin-lysophosphatidate (LPA)-lipid phosphate phosphatase axis is now a promising clinical target for treating chronic inflammatory conditions, mitigating fibrosis progression, and improving the efficacy of existing cancer chemotherapies and radiotherapy. Nearly half of the literature on this axis has been published during the last five years. In cancer biology, LPA signaling is increasingly being recognized as a central mediator of the progression of chronic inflammation in the establishment of a tumor microenvironment which promotes cancer growth, immune evasion, metastasis, and treatment resistance. In this review, we will summarize recent advances made in understanding LPA signaling with respect to chronic inflammation and cancer. We will also provide perspectives on the applications of inhibitors of LPA signaling in preventing cancer initiation, as adjuncts extending the efficacy of current cancer treatments by blocking inflammation caused by either the cancer or the cancer therapy itself, and by disruption of the tumor microenvironment. Overall, LPA, a simple molecule that mediates a plethora of biological effects, can be targeted at its levels of production by autotaxin, LPA receptors or through LPA degradation by lipid phosphate phosphatases. Drugs for these applications will soon be entering clinical practice.

Autotaxin is a secreted enzyme that converts extracellular lysophosphatidylcholine to lysophosphatidate (LPA). In cancers, LPA increases tumour growth, metastasis and chemoresistance by activating six G-protein coupled receptors. We examined >200 human thyroid biopsies. Autotaxin expression in metastatic deposits and primary carcinomas was four- to tenfold higher than in benign neoplasms or normal thyroid tissue. Autotaxin immunohistochemical staining was also increased in benign neoplasms with leukocytic infiltrations. Malignant tumours were distinguished from benign tumours by high tumour autotaxin, LPA levels and inflammatory mediators including IL1β, IL6, IL8, GMCSF, TNFα, CCL2, CXCL10 and platelet-derived growth factor (PDGF)-AA. We determined the mechanistic explanation for these results and revealed a vicious regulatory cycle in which LPA increased the secretion of 16 inflammatory modulators in papillary thyroid cancer cultures. Conversely, treating cancer cells with ten inflammatory cytokines and chemokines or PDGF-AA and PDGF-BB increased autotaxin secretion. We confirmed that this autotaxin/inflammatory cycle occurs in two SCID mouse models of papillary thyroid cancer by blocking LPA signalling using the autotaxin inhibitor ONO-8430506. This decreased the levels of 16 inflammatory mediators in the tumours and was accompanied by a 50–60% decrease in tumour volume. This resulted from a decreased mitotic index for the cancer cells and decreased levels of vascular endothelial growth factor and angiogenesis in the tumours. Our results demonstrate that the autotaxin/inflammatory cycle is a focal point for driving malignant thyroid tumour progression and possibly treatment resistance. Inhibiting autotaxin activity provides an effective and novel strategy for decreasing the inflammatory phenotype in thyroid carcinomas, which should complement other treatment modalities.

Autotaxin (ATX) is a secreted enzyme, which produces extracellular lysophosphatidate (LPA) from lysophosphatidylcholine (LPC). LPA activates six G protein-coupled receptors and this is essential for vasculogenesis during embryonic development. ATX is also involved in wound healing and inflammation, and in tumor growth, metastasis, and chemo-resistance. It is, therefore, important to understand how ATX is regulated. It was proposed that ATX activity is inhibited by its product LPA, or a related lipid called sphingosine 1-phosphate (S1P). We now show that this apparent inhibition is ineffective at the high concentrations of LPC that occur in vivo. Instead, feedback regulation by LPA and S1P is mediated by inhibition of ATX expression resulting from phosphatidylinositol-3-kinase activation. Inhibiting ATX activity in mice with ONO-8430506 severely decreased plasma LPA concentrations and increased ATX mRNA in adipose tissue, which is a major site of ATX production. Consequently, the amount of inhibitor-bound ATX protein in the plasma increased. We, therefore, demonstrate the concept that accumulation of LPA in the circulation decreases ATX production. However, this feedback regulation can be overcome by the inflammatory cytokines, TNF-α or interleukin 1β. This enables high LPA and ATX levels to coexist in inflammatory conditions. The results are discussed in terms of ATX regulation in wound healing and cancer.

Metastatic dissemination of papillary thyroid cancer has been reported to be strongly associated with expression of platelet-derived growth factor (PDGFR) α and altered TTF1 function. However, the status of PDGF ligands in papillary thyroid cancer and the potential role of these ligands in metastatic disease are obscure. We assessed the prevalence of PDGF ligands in benign and malignant thyroid tumors to determine if ligand upregulation is associated with α-isoform (PDGF-AA or PDGF-BB) or the β-isoform (PDGF-BB or PDGF-DD) of PDGFR in individual tumors. The immunohistochemical expression of PDGFRα, PDGF-AA, PDGF-BB, and PDGF-DD was surveyed in follicular adenomas (n=55), papillary thyroid carcinomas (103 with and 59 without nodal metastases), and lymph node metastasis (n=12). There is an augmented tendency for PDGF-AA expression in node-positive papillary thyroid cancer metastases (P

Ultrasonography for thyroid nodules is one of the most common imaging tests performed in the general population. Details from ultrasound reports guide biopsies and surgery. This study quantifies the completeness of these reports based on Thyroid Imaging and Reporting System (TI-RADS) criteria and considers their utility in predicting malignant disease.We retrospectively reviewed ultrasound reports for 329 thyroidectomy patients and extracted data elements using the TI-RADS criteria: nodule size, echogenicity, margins, vascularity, solid/cystic composition and the presence or absence of microcalcifications and the halo sign. We assessed the reports to determine whether individual or multiple criteria were associated with malignancy.More than 97% of reports document nodule size; however, more than 90% of the reports noted only 3 or fewer of the 6 remaining TI-RADS criteria. The presence of microcalcifications was the most sensitive marker of malignancy (90%), whereas the documentation of irregular margins was the most specific indicator of malignancy (88%). Overall it was clear that microcalcifications, hypoechogenicity, irregular margins and solid nodules were significantly more likely to be found in malignant neoplasms; their absence predicted benign disease. Because so few reports consistently documented all criteria, the overall ability of thyroid ultrasonography to discriminate between lowerand higher-risk nodules is limited.Although the accuracy of thyroid ultrasonography is good, few ultrasound reports contain the necessary information, as defined by TI-RADS, to predict malignancy and guide management. When reported, microcalcifications and/or irregular margins are the best predictors of malignancy.L’échographie des nodules thyroïdiens est l’une des épreuves d’imagerie les plus souvent effectuées dans la population générale. Les détails fournis par l’échographie guident les biopsies et la chirurgie. Cette étude quantifie l’exhaustivité des rapports d’échographie selon les critères TI-RADS (Thyroid Imaging and Reporting System) et en mesure l’utilité pour prédire les cancers.Nous avons passé en revue de façon rétrospective les rapports d’échographie de 329 patients ayant subi une thyroïdectomie et nous en avons extrait les éléments sous l’angle des critères TI-RADS : taille des nodules, échogénicité, marges, vascularité, composition solide c. kystique, présence ou absence de microcalcifications et signe du halo. Nous avons évalué les rapports afin de déterminer si certains critères individuels ou multiples pouvaient être associés au cancer.Plus de 97 % des rapports mentionnent la taille des nodules; mais, plus de 90 % des rapports ne font état que de 3 critères ou moins sur les 6 autres critères TI-RADS. La présence de microcalcifications a été le marqueur tumoral le plus sensible (90 %), tandis que la présence de marges irrégulières a été le marqueur tumoral le plus spécifique (88 %). Dans l’ensemble, les microcalcifications, l’hypoéchogénicité, les marges irrégulières et les nodules solides ont sans contredit été significativement plus susceptibles d’être observés en présence de malignité; et en revanche, leur absence permettait de prédire une maladie bénigne. Étant donné que si peu de rapports ont documenté avec constance tous les critères, la capacité globale de l’échographie de la thyroïde à distinguer entre nodules de risque faible et élevé est limitée.Même si la précision de l’échographie thyroïdienne est bonne, peu de rapports d’échographie renferment les renseignements nécessaires, selon les critères TI-RADS, pour prédire un cancer et orienter sa prise en charge. Lorsqu’elles sont signalées, les microcalcifications ou les marges irrégulières sont les meilleurs prédicteurs de cancer.

International audience; Lipin-1 is a Mg2+-dependent phosphatidic acid phosphatase (PAP) that in mice is necessary for normal glycerolipid biosynthesis, controlling adipocytes metabolism and adipogenic differentiation. Mice carrying inactivating mutations in the Lpin1 gene display the characteristic features of human familial lipodystrophy. Very little is known on the roles of lipin-1 in human adipocyte physiology. Apparently fat distribution and weight is normal in humans carrying LPIN1 inactivating mutations, but a detailed analysis of adipose tissue appearance and functions in these patients has not been available so far. In this study, we performed a systematic histopathological, biochemical and gene expression analysis of adipose tissue biopsies from human patients harbouring LPIN1 biallelic inactivating mutations and affected by recurrent episodes of severe rhabdomyolysis. We also explored the adipogenic differentiation potential of human mesenchymal cell populations derived from lipin-1 defective patients. White adipose tissue from human LPIN1 mutant patients displayed a dramatic decrease in lipin-1 protein levels and PAP activity, with a concomitant moderate reduction of the adipocyte size. Nevertheless the adipose tissue develops without obvious histological signs of lipodystrophy and with a normal qualitative composition of the storage lipids. The increased expression of key adipogenic determinants such as SREBP1, PPARG and PGC1A shows that specific compensatory phenomena can be activated in vivo in human adipocytes under deficiency of a functional lipin-1.

We report a case involving a female patient with primary hyperparathyroidism arising from a middle mediastinal parathyroid adenoma. Initial diagnosis was made based on a history of polyuria and recurrent renal colic. At the age of 77 she underwent videoscopic assisted mediastinoscopy (VAM) for resection of the adenoma which was localized using single-photon emission computed tomography (SPECT/CT). This case illustrates the importance for accurate pre-operative localization and intra-operative monitoring to ensure complete surgical removal in order to improve postoperative normocalcemia success rates.

Epigenetic silencing of RASSF1A is frequently observed in numerous cancers and has been previously reported. The promoter region of RASSF1A is predicted to have 75 CpG sites, and very few studies demonstrate how the methylation of these sites affects expression. In addition, the expression relationship between RASSF1A and its downstream target, modulator of apoptosis 1 (MOAP-1), is poorly understood. In this study, we have explored the mRNA expression of RASSF1A, MOAP-1 and the well-characterized splice variant of RASSF1, RASSF1C, in cancer cell lines and primary tumors. We confirmed that the RASSF1A promoter is robustly methylated within a 32-CpG region in solid tumors and results in lower mRNA expression. The MOAP-1 promoter contains ~110 CpG sites, but was not found to be methylated in cancer cell lines when 19 predicted CpG sites were explored. Interestingly, MOAP-1 mRNA expression positively correlated with RASSF1A expression in numerous cancers, whereas RASSF1C expression remained the same or was increased in cell lines or tissues with epigenetic loss of RASSF1A. We speculate that MOAP-1 and RASSF1A may be more intimately connected than originally thought, and the expression of both are warranted in experimental designs exploring the biology of the RASSF1A/MOAP-1 molecular pathway.

Lymph node metastases are common in papillary thyroid cancer (PTC) and can be resistant to surgical extirpation or radioiodine ablation. We examined the role of platelet-derived growth factor receptor (PDGFR) in mediating lymph node metastases in PTC. Clinical specimens of PTC (n = 137) were surveyed in a tissue array and by western blots to examine the relationship between expression of the α and β subunits of PDGFR and lymph node metastases. PDGFR-α was found at high levels in primary tumours with known lymphatic metastases but not in those tumours lacking nodal involvement (p < 0.0001). However, PDGFR-β expression was not linked to metastatic disease (p = 0.78) as it was found in virtually all PTC specimens. A matching analysis in fresh PTC specimens (n = 13) confirmed that PDGFR-α expression was strongly linked to metastatic spread (p = 0.0047). PDGFR-α and -β were not found in normal thyroid tissue (p < 0.0001). PTC cell lines selectively expressing PDGFR-α or -β were assessed for invasive potential and activation of downstream signal transduction pathways. PTC cell lines expressing PDGFR-α responded to PDGF-BB stimulation with increased invasive potential and this process can be blocked by the tyrosine kinase receptor inhibitor sunitinib (p < 0.009). Cell lines with only PDGFR-β, or no PDGFR, did not show significant changes in invasive potential. Activation of PDGFR-α led to downstream up-regulation of both the MAPK/ERK and PI3K/Akt pathways and disruption of either pathway is sufficient to block PDGFR-mediated increases in invasive potential. Thus, PDGFR-α is associated with lymph node metastases in papillary thyroid carcinoma and PDGFR-α promotes increased invasive potential in PTC cell lines. PDGFR-α is a strong candidate for a diagnostic biomarker to identify patients at risk of nodal metastases. Our results also strengthen the rationale for selection of tyrosine kinase receptor inhibitors that target PDGFR in the treatment of progressive, metastatic PTC. Copyright © 2012 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.

Thyroid nodules occupy a unique position in relation to clinical diagnosis since most patients with a thyroid nodule do not present with overt symptoms. There are, however, no good published data demonstrating the way in which clinically solitary thyroid nodules come to medical attention, with most of the literature being anecdotal. This study aims to evaluate the mode of presentation of solitary thyroid nodules, and to assess whether the presence of a malignancy influences that presentation.In this retrospective cohort study, data were obtained from the University of Sydney Endocrine Surgery Database and through a review of the patient records. The study cohort comprised 419 consecutive patients who presented with a clinically solitary thyroid nodule and who subsequently underwent surgery. Patient files lacking data on mode of presentation and patients in whom thyroid nodules were incidentally discovered during parathyroid surgery were excluded, leaving a total of 299 patients. Data were analyzed using a computer-based statistical software package. Continuous variables were compared using Student's t-test. Categorical variables were analyzed using Fishers exact test. Statistical significance was set at p 0.05.Solitary thyroid nodules are most commonly detected by the patients themselves (40%), followed by the incidental discovery of nodules on imaging studies performed for unrelated reasons (30%), and lastly due to third-party diagnosis by family, friends, acquaintances, or medical practitioners (30%). Nodules detected by medical practitioners were statistically more likely to be malignant (p = 0.02). No other differences between the clinical presentation of benign and malignant nodules were found.Patients themselves are the ones who most commonly first become aware of the thyroid nodules that are eventually seen by thyroid surgeons. Incidental discovery on image studies is also important. Thyroid nodules detected by medical practitioners should be dealt with most urgently because they are most likely to be malignant.

Background: Recent published data has shown that metastatic involvement of the prelaryngeal or Delphian lymph node (DLN), the highest of the central (level VI) cervical lymph nodes, is highly predictive of advanced nodal disease in papillary thyroid cancer (PTC). The aims of this study were to determine the diagnostic accuracy of all the level VI cervical nodes in PTC and to determine which node group, if any, is the most accurate in predicting lateral node (N1b) disease. Methods: This was a retrospective cohort study. Data were obtained from the University of Sydney Endocrine Surgical Unit Database and through a review of the histopathology records. The study cohort was composed of 177 consecutive patients with a final diagnosis of PTC who underwent total thyroidectomy and lymph node dissection, spanning the period from May 2001 to December 2006. Results: Of the 177 patients with PTC, 86 had the DLN removed, 51 had a pretracheal node removed and 76 had the paratracheal group removed. DLN, paratracheal and pretracheal node disease was present in 21%, 39% and 46%, respectively. Lateral node (N1b) disease was present in 35%. Paratracheal node involvement was mildly predictive of further disease with patients 1.7 times more likely to have lateral node involvement (sensitivity = 55%, specificity = 68%). Pretracheal node involvement was moderately predictive of further disease with patients three times more likely to have lateral node involvement (sensitivity = 72%, specificity = 74%). DLN involvement was highly predictive of further node involvement with patients nine times more likely to have lateral node disease (sensitivity = 53%, specificity = 94%) and 40 times more likely to have any nodal disease (sensitivity = 41%, specificity = 100%). Conclusion: This is the first study to examine the diagnostic accuracy of all level VI lymph nodes in PTC. While, metastatic involvement of all central nodal groups is indicative of further disease, the DLN is the most accurate predictor.

SummaryLymphoid nodules are a normal component of the mucosa of the rectum, but little is known about their function and whether they contribute to the host immune response in malignancy. In rectal cancer specimens from patients with local (n = 18), regional (n = 12) and distant (n = 10) disease, we quantified T cell (CD3, CD25) and dendritic cell (CD1a, CD83) levels at the tumour margin as well as within tumour-associated lymphoid nodules. In normal tissue CD3+, but not CD25+, T cells are concentrated at high levels within lymphoid nodules, with significantly fewer cells found in surrounding normal mucosa (P = 0·001). Mature (CD83), but not immature (CD1a), dendritic cells in normal tissue are also found clustered almost exclusively within lymphoid nodules (P =

Background: In thyroid cancer, metastases to intra-abdominal solid organs are uncommon and intra-peritoneal metastases are extremely rare. Here we present the management and outcome of a young patient with a large radioiodine resistant intra-peritoneal metastasis of insular thyroid cancer. Summary: A 28-year-old woman with known radioiodine resistant metastatic insular (poorly differentiated) thyroid carcinoma and multiple previous resected metastases presented with acute onset of bowel obstruction due to a large pelvic mass. The pelvic mass was resected and histologically confirmed to be an insular thyroid carcinoma metastasis. She made an uneventful recovery from her surgery and continues to have a good quality of life despite low volume metastatic disease. Conclusion: As this patient demonstrates, a rational but aggressive surgical approach is warranted for patients with radioiodine-resistant metastatic thyroid disease.

Minimally invasive thyroidectomy techniques are being developed in an effort to minimize pain, shorten the length of hospital stay, and improve cosmesis. Various minimally invasive thyroid surgery (MITS) techniques have been shown to be safe and feasible with some benefits in terms of cosmesis and pain outcomes; however, no single technique has been broadly accepted. This study was designed to review the evidence in relation to MITS and our experience with the direct lateral mini-incision technique.A review of literature published until December 2007 on minimally invasive thyroidectomy techniques was undertaken. Three issues were addressed: 1) Does MITS provide any benefit compared with conventional open thyroidectomy? 2) Is there any advantage to the use of endoscopic or video-assisted techniques compared with the direct mini-incision technique? 3) Is the lateral mini-incision technique safe and efficacious? Additional data in relation to the above issues was derived from a retrospective cohort study of patients undergoing mini-incision thyroid surgery within our unit.Issue 1: Five prospective randomized studies and eight studies at a lower level of evidence have demonstrated consistent advantages of MITS compared with open thyroid surgery in terms of reduced pain and improved cosmesis with equivalent operative safety. Issue 2: In compiling four level III and IV studies that compared open and video-assisted minimally invasive surgery, there do not seem to be significant differences in patient satisfaction with the incision. The video-assisted approaches require significantly longer operative times but also seem to be less painful. Issue 3: Three cohort studies (level IV) have demonstrated that the lateral mini-incision technique is both safe and efficacious compared with open surgery for hemi-thyroidectomy. Data from our cohort study of 1281 patients (open hemi-thyroidectomy 1054 vs. MITS 227) confirmed MITS to be a safe and effective procedure. The rate of postoperative hematoma formation and wound infection was equivalent between groups. The rate of permanent recurrent laryngeal nerve injury was 0.4% for MITS and 0.3% for CHT and not significantly different (p = 0.7).MITS has demonstrated advantages over conventional open approaches for both hemi- and total thyroidectomy and the benefits do not depend on the open or video-assisted approach. For thyroid lobectomies, the lateral mini-incision approach can be performed with an operative time and postoperative complication profile equivalent to conventional hemi-thyroidectomy while providing excellent cosmesis with a 2-3 cm scar.

Tamoxifen is the accepted therapy for patients with estrogen receptor-α (ERα)-positive breast cancer. However, clinical resistance to tamoxifen, as demonstrated by recurrence or progression on therapy, is frequent and precedes death from metastases. To improve breast cancer treatment it is vital to understand the mechanisms that result in tamoxifen resistance. This study shows that concentrations of tamoxifen and its metabolites, which accumulate in tumors of patients, killed both ERα-positive and ERα-negative breast cancer cells. This depended on oxidative damage and anti-oxidants rescued the cancer cells from tamoxifen-induced apoptosis. Breast cancer cells responded to tamoxifen-induced oxidation by increasing Nrf2 expression and subsequent activation of the anti-oxidant response element (ARE). This increased the transcription of anti-oxidant genes and multidrug resistance transporters. As a result, breast cancer cells are able to destroy or export toxic oxidation products leading to increased survival from tamoxifen-induced oxidative damage. These responses in cancer cells also occur in breast tumors of tamoxifen-treated mice. Additionally, high levels of expression of Nrf2, ABCC1, ABCC3 plus NAD(P)H dehydrogenase quinone-1 in breast tumors of patients at the time of diagnosis were prognostic of poor survival after tamoxifen therapy. Therefore, overcoming tamoxifen-induced activation of the ARE could increase the efficacy of tamoxifen in treating breast cancer.

Compared to normal tissues, many cancer cells overexpress autotaxin (ATX). This secreted enzyme produces extracellular lysophosphatidate, which signals through 6 GPCRs to drive cancer progression. Our previous work showed that ATX inhibition decreases 4T1 breast tumor growth in BALB/c mice by 60% for about 11 d. However, 4T1 cells do not produce significant ATX. Instead, the ATX is produced by adjacent mammary adipose tissue. We investigated the molecular basis of this interaction in human and mouse breast tumors. Inflammatory mediators secreted by breast cancer cells increased ATX production in adipose tissue. The increased lysophosphatidate signaling further increased inflammatory mediator production in adipose tissue and tumors. Blocking ATX activity in mice bearing 4T1 tumors with 10 mg/kg/d ONO-8430506 (a competitive ATX inhibitor, IC90 = 100 nM; Ono Pharma Co., Ltd., Osaka, Japan) broke this vicious inflammatory cycle by decreasing 20 inflammatory mediators by 1.5-8-fold in cancer-inflamed adipose tissue. There was no significant decrease in inflammatory mediator levels in fat pads that did not bear tumors. ONO-8430506 also decreased plasma TNF-α and G-CSF cytokine levels by >70% and leukocyte infiltration in breast tumors and adjacent adipose tissue by >50%. Hence, blocking tumor-driven inflammation by ATX inhibition is effective in decreasing tumor growth in breast cancers where the cancer cells express negligible ATX.

Thyroid nodules are a common problem worldwide. The incidence and aetiology of nodule formation may vary by geography, but a common problem faced by all physicians is how to differentiate benign disease from malignancy. Clinical examination and ultrasonography may identify advanced thyroid carcinoma, but the diagnosis of the vast majority of well-differentiated thyroid cancers (WDTC) relies upon fine-needle aspiration biopsy (FNAB). Important advances in timely identification and management of thyroid cancer have been achieved by improvements in biopsy technique and standardization of thyroid cytopathology reporting. However, many patients still require a thyroid lobectomy before malignancy can be diagnosed. Minimizing the morbidity and cost of diagnostic surgery is an important goal given that the number of patients screened for thyroid malignancy continues to increase significantly across the world. In both the northern and southern hemispheres the incidence of thyroid cancer has more than doubled in the past two decades. Many investigators are now focusing their efforts on improving the diagnostic potential of aspiration biopsies through molecular diagnostics. Success in these efforts is reflected by the appearance of commercially available options for molecular diagnostics in thyroid nodule specimens. In the future, immunocytochemical markers combined with mutation analysis and large-scale genetic sequencing are likely to be used to define distinct benign and malignant signatures in thyroid neoplasms.

The present work elucidates novel mechanisms for lysophosphatidate (LPA)-induced chemoresistance using human breast, lung, liver, and thyroid cancer cells. LPA (0.5-10 μM) increased Nrf2 transcription factor stability and nuclear localization by ≤5-fold. This involved lysophosphatidate type 1 (LPA1) receptors as identified with 1 μM wls-31 (LPA1/2 receptor agonist) and blocking this effect with 20 μM Ki16425 (LPA1-3 antagonist, Ki = 0.34 μM). Knockdown of LPA1 by 50% to 60% with siRNA decreased Nrf2 stability and expressing LPA1, but not LPA2/3, in human HepG2 cells increased Nrf2 stabilization. LPA-induced Nrf2 expression increased transcription of multidrug-resistant transporters and antioxidant genes by 2- to 4-fold through the antioxidant response element. This protected cells from doxorubicin-induced death. This pathway was verified in vivo by orthotopic injection of 20,000 mouse 4T1 breast cancer cells into syngeneic mice. Blocking LPA production with 10 mg/kg per d ONO-8430506 (competitive autotaxin inhibitor, IC90 = 100 nM) decreased expression of Nrf2, multidrug-resistant transporters, and antioxidant genes in breast tumors by ≤90%. Combining 4 mg/kg doxorubicin every third day with ONO-8430506 synergistically decreased tumor growth and metastasis to lungs and liver by70%, whereas doxorubicin alone had no significant effect. This study provides the first evidence that LPA increases antioxidant gene and multidrug-resistant transporter expression. Blocking this aspect of LPA signaling provides a novel strategy for improving chemotherapy.

Lipid phosphate phosphatase-1 (LPP1) degrades lysophosphatidate (LPA) and attenuates receptor-mediated signaling. LPP1 expression is low in many cancer cells and tumors compared with normal tissues. It was hypothesized from studies with cultured cells that increasing LPP1 activity would decrease tumor growth and metastasis. This hypothesis has never been tested in vivo. To do this, we inducibly expressed LPP1 or a catalytically inactive mutant in cancer cells. Expressing active LPP1 increased extracellular LPA degradation by 5-fold. It also decreased the stimulation of Ca(2+) transients by LPA, a nondephosphorylatable LPA1/2 receptor agonist and a protease-activated receptor-1 peptide. The latter results demonstrate that LPP1 has effects downstream of receptor activation. Decreased Ca(2+) mobilization and Rho activation contributed to the effects of LPP1 in attenuating the LPA-induced migration of MDA-MB-231 breast cancer cells and their growth in 3D culture. Increasing LPP1 expression in breast and thyroid cancer cells decreased tumor growth and the metastasis by up to 80% compared with expression of inactive LPP1 or green fluorescent protein in syngeneic and xenograft mouse models. The present work demonstrates for the first time that increasing the LPP1 activity in three lines of aggressive cancer cells decreases their abilities to produce tumors and metastases in mice.

Autotaxin is a secreted enzyme that produces most extracellular lysophosphatidate, which stimulates 6 G-protein-coupled receptors. Lysophosphatidate promotes cancer cell survival, growth, migration, invasion, metastasis, and resistance to chemotherapy and radiotherapy. The present work investigated whether inhibiting autotaxin could decrease breast tumor growth and metastasis. We used a new autotaxin inhibitor (ONO-8430506; IC90=100 nM), which decreased plasma autotaxin activity by >60% and concentrations of unsaturated lysophosphatidates by >75% for 24 h compared with vehicle-treated mice. The effects of ONO-8430506 on tumor growth were determined in a syngeneic orthotopic mouse model of breast cancer following injection of 20,000 BALB/c mouse 4T1 or 4T1-12B cancer cells. We show for the first time that inhibiting autotaxin decreases initial tumor growth and subsequent lung metastatic nodules both by 60% compared with vehicle-treated mice. Significantly, 4T1 cells express negligible autotaxin compared with the mammary fat pad. Autotaxin activity in the fat pad of nontreated mice was increased 2-fold by tumor growth. Our results emphasize the importance of tumor interaction with its environment and the role of autotaxin in promoting breast cancer growth and metastasis. We also established that autotaxin inhibition could provide a novel therapeutic approach to blocking the adverse effects of lysophosphatidate in cancer.

SALL4 is one of the master transcriptional factors that are crucial in maintaining the pluripotency of embryonic stem cells (ESCs). While the expression of SALL4 is normally restricted to ESCs and somatic stem cells, we found that it is aberrantly expressed in ALK-positive anaplastic large cell lymphoma (ALK+ ALCL), a type of lymphoid malignancy carrying a mature T-cell immunophenotype. shRNA knockdown of SALL4 in ALK+ ALCL cell lines resulted in apoptosis and cell-cycle arrest, and significantly decreased colony formation on soft agar. These changes correlated with the downregulation of several anti-apoptotic proteins and facilitators of cell-cycle progression. Based on the differential response to a SALL4 reporter construct, we were able to separate two distinct cell subsets in Karpas 299 (an ALK+ ALCL cell line), namely SALL4(high) and SALL4(low). Importantly, the biological effects induced by SALL4 knock-down in Karpas 299 were restricted to the purified SALL4(high) cells, and this finding further supports the concept that SALL4 is biologically important in ALK+ ALCL. Lastly, the expression of SALL4 was not dependent on the nucleophosmin (NPM)-anaplastic lymphoma kinase (ALK)/STAT3 signaling axis, the key oncogenic driver in ALK+ ALCL. To conclude, for the first time, our study has revealed the oncogenic contributions of an ESC protein in the pathogenesis of ALK+ ALCL.

The transcriptional factor Twist1 has been shown to play a key role in regulating epithelial mesenchymal transition, invasiveness and migratory properties in solid tumors. We found that Twist1 is aberrantly expressed in ALK-positive anaplastic large cell lymphoma (ALK+ALCL), a type of T-cell lymphoid malignancy. Using RT-PCR and Western blots, Twist1 was detectable in all 3 ALK+ALCL cell lines examined but absent in normal T-cells. By immunohistochemistry, Twist1 was detectable in all 10 cases of ALK+ALCL examined; benign lymphoid tissues were consistently negative. Twist1 expression in ALK+ALCL cells can be attributed to the NPM-ALK/STAT3 signaling axis, the key oncogenic driving force in this tumor type. Twist1 is biologically important in ALK+ALCL cells, as Twist1 knockdown resulted in a significant decrease in their invasiveness in an in-vitro assay. Further investigation revealed that this increase in invasiveness is linked to the activation of AKT and down-regulation of p66Shc, two signaling proteins known to be involved in NPM-ALK-mediated oncogenesis. Lastly, knockdown of Twist1 sensitizes ALK+ALCL cells to the growth inhibitory effect of PF-2341066 (Crizotinib®), an ALK inhibitor being used in clinical trials. In conclusion, Twist1 expression, owing to the abnormal NPM-ALK/STAT3 signaling, contributes to its invasiveness and decreased sensitivity to PF-2341066 in ALK+ALCL.

We examined the expression of cyclin D1 in conjunction with β-catenin and the phosphorylated inactive form of glycogen synthase kinase 3β (GSK-3β) in benign, nonneoplastic thyroid tissue as well as papillary thyroid carcinoma primary tumors and nodal metastases. We aim to unravel the regulation of cyclin D1 and determine if this cell cycle protein is a useful biomarker for metastatic disease. It is clear that expression of cyclin D1 (P < .0001), β-catenin (P < .0001), and inactive form of GSK-3β (P < .0001) are significantly higher in papillary thyroid carcinoma primary tumors than in corresponding benign, nonneoplastic tissue thyroid specimens. Interestingly, β-catenin and cyclin D1 expressions in papillary thyroid carcinoma are correlated (P = .025), implying that β-catenin is a factor driving higher levels of cyclin D1 consistent with previous cell models linking Wnt/β-catenin signaling and cyclin D1 expression. Conversely, inactive form of GSK-3β expression does not correlate with cyclin D1 (P = .52) or β-catenin expression (P = .54). We also did not observe any relationship between tumor size and marker expression. Comparing papillary thyroid carcinoma primary tumors with or without nodal metastases, we did not see any differences in expression of inactive form of GSK-3β (P = .95), β-catenin (P = .14), or cyclin D1 (P = .46). However, in papillary thyroid carcinoma lymph node specimens, the up-regulation of cyclin D1 (P = .0083) was highly significant compared with primary tumors. pGSK-3β and β-catenin expression did not vary between primary tumors and nodal specimens. In conclusion, we have demonstrated that expression of cyclin D1 is linked to nodal metastases and that cyclin D1 levels are regulated by Wnt/β-catenin signaling. GSK pathway-mediated regulation of β-catenin or cyclin D1 expression does not appear operative in papillary thyroid carcinoma.

Background Of all thyroid nodules assessed by fine needle aspiration cytology (FNAC), 10–20% are classified as indeterminate/atypical. Traditionally, this group is considered to primarily represent follicular neoplasia. We hypothesize that papillary carcinoma accounts for a significant proportion of lesions classified as “atypical” on FNAC. Methods This retrospective study includes 228 patients who had an atypical FNAC result and who were subsequently found to have a malignancy on histologic examination of the excised thyroid lesion. Patients with papillary microcarcinomas, defined as lesions less than 10-mm diameter, were excluded. The study period was from 1987 to 2005. The patients were divided chronologically into 3 groups ( n = 76) for analysis: group 1, December 1987–March 1997; group 2, July 1997–October 2002; and group 3, October 2002–December 2005. Results Age- and sex-distribution of the 3 groups were not significantly different. Median nodule size of group 3 was significantly smaller. The distributions of histopathology of the 3 time periods were significantly different overall ( P = .0325). Prevalence of papillary carcinoma was not statistically significant (33/76 vs 34/76 vs 46/76; P = .0636), but showed a statistical significant trend to increase over time ( P = .0349). Prevalence of follicular variant papillary carcinoma was also found to be significantly different between the groups (7/76 vs 12/76 vs 19/76; P = .0320; P = .0349). Conclusion Papillary carcinoma accounted for most histopathologically confirmed cancers that had an atypical cytology. Papillary cancer in this group of patients trended up, probably due to a significant increase in the diagnosis of follicular variant of papillary cancer.

BACKGROUND: The role of minimally invasive thyroid surgery (MITS) is currently in evolution. The aim of this study is to compare the outcomes of MITS using the direct approach through a lateral incision with conventional hemithyroidectomy (CHT) for the management of atypical thyroid nodules. METHODS: A prospective, single-blinded, randomized controlled trial involving patients presenting with atypical thyroid nodules of 3-cm diameter or less was performed. Patients were randomized to MITS through a lateral 2.5-cm incision or CHT through a traditional 5- to 6-cm cervicotomy. Pain was measured using a 7-point visual analog scale on the 1st and 10th postoperative days. Serum C-reactive protein was measured on postoperative days 1 and 10. Satisfaction with cosmetic outcome was measured at 3 months. RESULTS: One-hundred patients were randomized to undergo MITS or CHT. The 2 groups were equivalent in terms of age and thyroid nodule size. Mean operative times were longer for the MITS group (56 vs 46 min, P < .001). Mean pain scores were less in the MITS group on the 1st postoperative day (2.67 vs 3.43, P = .032). Pain scores at 10 days were equivalent (1.5 vs 1.8, P = .36). Serum C-reactive protein levels were equivalent postoperatively. At 3 months, patients undergoing MITS reported a greater mean cosmetic satisfaction score (6.3 vs 5.0, P = .002). Incision lengths measured at 3 months were 2.6 cm for MITS and 5.4 cm for CHT group, P < .001. CONCLUSION: In the management of small, atypical thyroid nodules, MITS through a direct lateral approach results in less early postoperative pain and superior cosmetic results when compared with conventional thyroidectomy.

Autotaxin is a secreted enzyme that produces most of the extracellular lysophosphatidate from lysophosphatidylcholine, the most abundant phospholipid in blood plasma. Lysophosphatidate mediates many physiological and pathological processes by signaling through at least six G-protein coupled receptors to promote cell survival, proliferation and migration. The autotaxin/lysophosphatidate signaling axis is involved in wound healing and tissue remodeling, and it drives many chronic inflammatory conditions from fibrosis to colitis, asthma and cancer. In cancer, lysophosphatidate signaling promotes resistance to chemotherapy and radiotherapy, and increases both angiogenesis and metastasis. Research into autotaxin inhibitors is accelerating, both as primary and adjuvant therapy. Historically, autotaxin inhibitors had poor bioavailability profiles and thus had limited efficacy in vivo. This situation is now changing, especially since the recent crystal structure of autotaxin is now enabling rational inhibitor design. In this review, we will summarize current knowledge on autotaxin-mediated disease processes including cancer, and discuss recent advancements in the development of autotaxin-targeting strategies. We will also provide new insights into autotaxin as an inflammatory mediator in the tumor microenvironment that promotes cancer progression and therapy resistance.