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Radioresistance is a substantial barrier to success in cancer management. A number of molecular mechanisms support radioresistance. We have shown experimentally that the thyroid hormone analogue receptor on the extracellular domain of integrin αvβ3 may modulate the state of radiosensitivity of tumor cells. Specifically, tetraiodothyroacetic acid (tetrac), a derivative of L-thyroxine (T4), can reduce radioresistance in cancer cells. In this review, we list a number of intrinsic signal transduction molecules and other host factors that have been reported to support/induce radioresistance in cancer cells and that are also subject to control by T4 through actions primarily initiated at integrin αvβ3. Additional preclinical evidence is needed to support these radioresistance-relevant actions of thyroid hormone.

The PD-1/PD-L1 immune checkpoint involving tumor cells and host immune defense lymphocytes is a well-studied therapeutic target in oncology. That PD-1 and PD-L1 may have additional functions within tumor cells that are independent of the checkpoint is indicated by actions of a thyroid hormone analogue, L-thyroxine (T4), on these checkpoint components. Acting at a cell surface receptor on plasma membrane integrin αvβ3, T4 stimulates intracellular accumulation of PD-L1 in cancer cells. In these thyroid hormone-treated cells, T4-induced PD-L1 is non-immunologically anti-apoptotic, blocking activation of p53. Several laboratories have also described accumulation of PD-1 in a variety of cancer cells, not just immune defense lymphocytes and macrophages. Preliminary observations indicate that T4 stimulates intracellular accumulation of PD-1 in tumor cells, suggesting that, like PD-L1, PD-1 has non-immunologic roles in the setting of cancer. Where such roles are anti-apoptotic, thyroid hormone-directed cancer cell accumulation of PD-1 and PD-L1 may limit effectiveness of immunologic therapy directed at the immune checkpoint.

// Chia-Wen Ho 1, 4, * , Chi-Yu Lin 2, * , Yi-Wei Liaw 2, 3, * , Hsiao-Ling Chiang 2 , Yu-Tang Chin 4 , Rui-Lan Huang 5 , Hung-Cheng Lai 5, 6, 7 , Yaw-Wen Hsu 6 , Po-Jan Kuo 8 , Chiao-En Chen 4 , Hung-Yun Lin 4 , Jacqueline Whang-Peng 4 , Shin Nieh 6, 9 , Earl Fu 6, 8 , Leroy F. Liu 1, 4, # , Jaulang Hwang 1, 2, 3, 6 1 Center for Cancer Research, Taipei Medical University, Taipei, Taiwan 2 Department of Biochemistry, Medical College, Taipei Medical University, Taipei, Taiwan 3 Institute of Biochemistry and Molecular Biology, National Yang-Ming University, Taipei, Taiwan 4 Graduate Institute of Cancer Biology and Drug Discovery, College of Medical Science and Technology, Taipei Medical University, Taipei, Taiwan 5 Department of Obstetrics and Gynecology, Shuang-Ho Hospital, Taipei Medical University, New Taipei City, Taiwan 6 Graduate Institute of Life Sciences, National Defense Medical Center, Taipei, Taiwan 7 Department of Obstetrics and Gynecology, School of Medicine, College of Medicine, Taipei Medical University, Taipei, Taiwan 8 Department of Periodontology, School of Dentistry, National Defense Medical Center and Tri-Service General Hospital, Taipei, Taiwan 9 Department of Pathology, National Defense Medical Center and Tri-Service General Hospital, Taipei, Taiwan # Co-corresponding author * These authors have contributed equally to this work Correspondence to: Jaulang Hwang, email: jaulang@tmu.edu.tw Keywords: Tn antigen, tumor-associated carbohydrate, cytokines, cosmc, hypermethylation Received: October 22, 2015 Accepted: July 16, 2016 Published: August 17, 2016 ABSTRACT Tn antigen (GalNAc-α- O -Ser/Thr), a mucin-type O -linked glycan, is a well-established cell surface marker for tumors and its elevated levels have been correlated with cancer progression and prognosis. There are also reports that Tn is elevated in inflammatory tissues. However, the molecular mechanism for its elevated levels in cancer and inflammation is unclear. In the current studies, we have explored the possibility that cytokines may be one of the common regulatory molecules for elevated Tn levels in both cancer and inflammation. We showed that the Tn level is elevated by the conditioned media of Hras G12V -transformed-BEAS-2B cells. Similarly, the conditioned media obtained from LPS-stimulated monocytes also elevated Tn levels in primary human gingival fibroblasts, suggesting the involvement of cytokines and/or other soluble factors. Indeed, purified inflammatory cytokines such as TNF-α and IL-6 up-regulated Tn levels in gingival fibroblasts. Furthermore, TNF-α was shown to down-regulate the COSMC gene as evidenced by reduced levels of the COSMC mRNA and protein, as well as hypermethylation of the CpG islands of the COSMC gene promoter. Since Cosmc, a chaperone for T-synthase, is known to negatively regulate Tn levels, our results suggest elevated Tn levels in cancer and inflammation may be commonly regulated by the cytokine-Cosmc signaling axis.

// Yu-Chen SH Yang 1, * , Yu-Tang Chin 2, * , Meng-Ti Hsieh 2, 3 , Hsuan-Yu Lai 2 , Chien-Chih Ke 4 , Dana R. Crawford 5 , Oscar K. Lee 6 , Earl Fu 7 , Shaker A. Mousa 8 , Patricia Grasso 9 , Leroy F. Liu 2 , Heng-Yu Chang 10, 11 , Heng-Yuan Tang 8 , Hung-Yun Lin 2, 3 , Paul J. Davis 8, 9 1 Joint Biobank, Office of Human Research, Taipei Medical University, Taipei, Taiwan 2 Taipei Cancer Center, Taipei Medical University, Taipei, Taiwan 3 PhD Program for Cancer Biology and Drug Discovery, College of Medical Science and Technology, Taipei Medical University, Taipei, Taiwan 4 Biomedical Imaging Research Center, National Yang-Ming University, Taipei, Taiwan 5 Center for Immunology and Microbial Disease, Albany Medical College, Albany, New York, USA 6 Stem Cell Research Center, National Yang-Ming University, Taipei, Taiwan 7 Department of Periodontology, School of Dentistry, National Defense Medical College, Taipei, Taiwan 8 Pharmaceutical Research Institute, Albany College of Pharmacy and Health Sciences, Albany, New York, USA 9 Department of Medicine, Albany Medical College, Albany, New York, USA 10 Department of Biochemistry and Molecular Cell Biology, College of Medicine, Taipei Medical University, Taipei, Taiwan 11 Graduate Institute of Medical Sciences, College of Medicine, Taipei Medical University, Taipei, Taiwan * These authors have contributed equally to this work Correspondence to: Heng-Yu Chang, e-mail: hychang@tmu.edu.tw Hung-Yun Lin, e-mail: linhy@tmu.edu.tw Keywords: obesity, leptin, OB3-leptin peptide, cancer cell invasion Received: September 15, 2015 Accepted: March 18, 2016 Published: March 30, 2016 ABSTRACT Obesity results in increased secretion of cytokines from adipose tissue and is a risk factor for various cancers. Leptin is largely produced by adipose tissue and cancer cells. It induces cell proliferation and may serve to induce various cancers. OB3-leptin peptide (OB3) is a new class of functional leptin peptide. However, its mitogenic effect has not been determined. In the present study, because of a close link between leptin and the hypothalamic-pituitary-thyroid axis, OB3 was compared with leptin in different thyroid cancer cells for gene expression, proliferation and invasion. Neither agent stimulated cell proliferation. Leptin stimulated cell invasion, but reduced adhesion in anaplastic thyroid cancer cells. Activated ERK1/2 and STAT3 contributed to leptin-induced invasion. In contrast, OB3 did not affect expression of genes involved in proliferation and invasion. In vivo studies in the mouse showed that leptin, but not OB3, significantly increased circulating levels of thyrotropin (TSH), a growth factor for thyroid cancer. In summary, OB3 is a derivative of leptin that importantly lacks the mitogenic effects of leptin on thyroid cancer cells.

Human intrahepatic cholangiocarcinomas are one of the most difficult cancers to treat. In our study, Lovastatin, a 3-hydroxy-3-methylglutaryl-coenzyme-CoA (HMG-CoA) reductase inhibitor, demonstrated anticancer properties by inhibiting cancer cell proliferation, cell migration and cell adhesion. Lovastatin inhibited the expressions of transforming growth factor (TGF)-β1, cyclooxygenase (COX)-2, and intercellular adhesion molecule (ICAM)-1. Furthermore, lovastatin inhibited the expressions of integrin β1 and integrin β3 but not integrin αv or integrin β5. While Lovastatin's inhibitory effects on TGFβ1, COX2, and ICAM-1 expression were independently controlled by the tumor suppressor LKB1, integrin β3 expression was not affected. Lovastatin's inhibitory effect on cell adhesion was associated with the decreased expression of integrin β3 and cell surface heterodimer integrin αvβ3. Quantitative real time PCR, fluorescent microscopy, and cell migration assays all confirmed that Lovastatin inhibits integrin αvβ3 downstream signaling including FAK activation, and β-catenin, vimentin, ZO-1, and β-actin. Overall, Lovastatin reduced tumor cell proliferation and migration by modifying the expression of genes involved in cell adhesion and other critical cellular processes. Our study highlights novel anti-cancer properties of Lovastatin and supports further exploration of statins in the context of cholangiocarcinoma therapy.

Dihydrotestosterone (DHT) has been shown to promote breast cancer growth via different mechanisms. In addition to binding to ERα, the DHT membrane receptor exists on integrin αvβ3. Resveratrol induces p53-dependent apoptosis via plasma membrane integrin αvβ3. Resveratrol and DHT signals are both transduced by activated ERK1/2; however, DHT promotes cell proliferation in cancer cells, whereas resveratrol is pro-apoptotic. In this study, we examined the mechanism by which DHT inhibits resveratrol-induced apoptosis in human ERα positive (MCF-7) and negative (MDA-MB-231) breast cancer cells. DHT inhibited resveratrol-stimulated phosphorylation of Ser-15 of p53 in a concentration-dependent manner. These effects of DHT on resveratrol action were blocked by an ERα antagonist, ICI 182,780, in MCF-7 breast cancer cells. DHT inhibited resveratrol-induced nuclear complex of p53-COX-2 formation which is required p53-dependent apoptosis. ChIP studies of COX-2/p53 binding to DNA and expression of p53-responsive genes indicated that DHT inhibited resveratrol-induced p53-directed transcriptional activity. In addition, DHT did inhibit resveratrol-induced COX-2/p53-dependent gene expression. These results suggest that DHT inhibits p53-dependent apoptosis in breast cancer cells by interfering with nuclear COX-2 accumulation which is essential for stimulation of apoptotic pathways. Thus, the surface receptor sites for resveratrol and DHT are discrete and activate ERK1/2-dependent downstream effects on apoptosis that are distinctive. These studies provide new insights into the antagonizing effects of resveratrol versus DHT, an important step toward better understanding and eventually treating breast cancer. It also indicates the complex pathways by which apoptosis is induced by resveratrol in DHT-depleted and -repleted environments.

// Sheng-Huei Yang 1 , Hung-Yun Lin 1, 2 , Vincent H.S. Chang 3 , Chien-Chung Chen 4 , Yun-Ru Liu 5 , Jinghan Wang 6 , Keqiang Zhang 7 , Xiaoqing Jiang 6 , Yun Yen 1 1 PhD Program for Cancer Biology and Drug Discovery, College of Medical Science and Technology, Taipei Medical University, Taipei, Taiwan 2 Taipei Cancer Center, Taipei Medical University, Taipei, Taiwan 3 Institute of Translational Medicine, Taipei Medical University, Taipei, Taiwan 4 Graduate Institute of Biomedical Materials and Engineering, Taipei Medical University, Taipei, Taiwan 5 Office of Human Research, Taipei Medical University, Taipei, Taiwan 6 The First Department of Biliary Surgery, Eastern Hepatobiliary Surgical Hospital, The Second Military Medical University, Shanghai, China 7 Department of Molecular Pharmacology, City of Hope National Medical Center and Beckman Research Center, Duarte, California, USA Correspondence to: Yun Yen, e-mail: yyen@tmu.edu.tw Keywords: cholangiocarcinomas, combination therapy, lovastatin, gefitinib Received: January 29, 2015 Accepted: June 12, 2015 Published: June 23, 2015 ABSTRACT Gefitinib resistance has been shown to complicate cancer therapy. Lovastatin is a proteasome inhibitor that enhances gefitinib-induced antiproliferation in non-small cell lung cancer. The objective of this study is to investigate the mechanism of lovastatin-induced antiproliferation in gefitinib-resistant human cholangiocarcinoma. Two gefitinib-resistant cholangiocarcinoma cell lines, SSP-25 and HuH-28, were used in this study to determine how to compensate gefitinib resistance. The combined effect of these two drugs was examined using the MTT assay, qPCR, immunoblotting, flow cytometry, and in vivo xenograft. Results indicated that lovastatin enhanced TNF-α-induced cell death in vitro . In addition, the combination of lovastatin with gefitinib enhanced accumulation of TNF-α. Furthermore, the treatment induced a synergistic cytotoxic effect and antiproliferation through apoptosis in SSP-25 cells and cell cycle arrest in HuH-28 cells. Reproductive results were also observed in in vivo xenografts. These observations suggest that the combination of gefitinib and lovastatin might have additive antiproliferative effects against gefitinib-resistant cholangiocarcinoma cells. Based on these observations, we concluded that the combination of gefitinib and lovastatin could be used to overcome gefitinib resistance in cholangiocarcinoma cells.

The principal secretory product of the thyroid gland, L-thyroxine (T4), is anti-apoptotic at physiological concentrations in a number of cancer cell lines. Among the mechanisms of anti-apoptosis activated by the hormone are interference with the Ser-15 phosphorylation (activation) of p53 and with TNFα/Fas-induced apoptosis. The hormone also decreases cellular abundance and activation of proteolytic caspases and of BAX and causes increased expression of X-linked inhibitor of apoptosis (XIAP). The anti-apoptotic effects of thyroid hormone largely are initiated at a cell surface thyroid hormone receptor on the extracellular domain of integrin αvβ3 that is amply expressed and activated in cancer cells. Tetraiodothyroacetic acid (tetrac) is a T4 derivative that, in a model of resveratrol-induced p53-dependent apoptosis in glioma cells, blocks the anti-apoptotic action of thyroid hormone, permitting specific serine phosphorylation of p53 and apoptosis to proceed. In a nanoparticulate formulation limiting its action to αvβ3, tetrac modulates integrin-dependent effects on gene expression in human cancer cell lines that include increased expression of a panel of pro-apoptotic genes and decreased transcription of defensive anti-apoptotic XIAP and MCL1 genes. By a variety of mechanisms, thyroid hormone (T4) is an endogenous anti-apoptotic factor that may oppose chemotherapy-induced apoptosis in αvβ3-expressing cancer cells. It is possible to decrease this anti-apoptotic activity pharmacologically by reducing circulating levels of T4 or by blocking effects of T4 that are initiated at αvβ3.

// Yu-Tang Chin 1, * , Meng-Ti Hsieh 1, * , Sheng-Huei Yang 2 , Po-Wei Tsai 3 , Shwu-Huey Wang 4 , Ching-Chiung Wang 3 , Yee-Shin Lee 2 , Guei-Yun Cheng 1 , Wei-Chun HuangFu 1, 2 , David London 5 , Heng-Yuan Tang 5 , Earl Fu 6 , Yun Yen 2, 7 , Leroy F. Liu 1 , Hung-Yun Lin 1, 2 , Paul J. Davis 5, 8 1 Taipei Cancer Center, Taipei Medical University, Taipei, Taiwan 2 PhD Program for Cancer Biology and Drug Discovery, College of Medical Science and Technology, Taipei Medical University, Taipei, Taiwan 3 School of Pharmacy, College of Pharmacy, Taipei Medical University, Taipei, Taiwan 4 Core Facility, Taipei Medical University, Taipei, Taiwan 5 Pharmaceutical Research Institute, Albany College of Pharmacy and Health Sciences, Albany, New York, USA 6 Department of Periodontology, School of Dentistry, National Defense Medical Center and Tri-Service General Hospital, Taipei, Taiwan 7 Department of Molecular Pharmacology, City of Hope National Medical Center and Beckman Research Center, Duarte, California, USA 8 Albany Medical College, Albany, New York, USA * These authors contributed equally to this work Correspondence to: Hung-Yun Lin, e-mail: linhy@tmu.edu.tw Keywords: stilbene, integrin αvβ3, apoptosis, anti-proliferation, breast cancer Received: July 21, 2014 Accepted: October 23, 2014 Published: November 08, 2014 ABSTRACT We have used a perfusion bellows cell culture system to investigate resveratrolinduced anti-proliferation/apoptosis in a human estrogen receptor (ER)-negative breast cancer cell line (MDA-MB-231). Using an injection system to perfuse media with stilbene, we showed resveratrol (0.5 – 100 μM) to decrease cell proliferation in a concentration-dependent manner. Comparison of influx and medium efflux resveratrol concentrations revealed rapid disappearance of the stilbene, consistent with cell uptake and metabolism of the agent reported by others. Exposure of cells to 10 μM resveratrol for 4 h daily × 6 d inhibited cell proliferation by more than 60%. Variable extracellular acid-alkaline conditions (pH 6.8 – 8.6) affected basal cell proliferation rate, but did not alter anti-proliferation induced by resveratrol. Resveratrol-induced gene expression, including transcription of the most up-regulated genes and pro-apoptotic p53-dependent genes, was not affected by culture pH changes. The microarray findings in the context of induction of anti-proliferation with brief daily exposure of cells to resveratrol—and rapid disappearance of the compound in the perfusion system—are consistent with existence of an accessible initiation site for resveratrol actions on tumor cells, e.g., the cell surface receptor for resveratrol described on integrin αvβ3.

// Meng-Ti Hsieh 1, 2, * , Le-Ming Wang 3, * , Chun A. Changou 2, 4, 5 , Yu-Tang Chin 1, 6, 7, 8 , Yu-Chen S.H. Yang 9 , Hsuan-Yu Lai 1 , Sheng-Yang Lee 6, 7, 8 , Yung-Ning Yang 10 , Jacqueline Whang-Peng 1 , Leroy F. Liu 1 , Hung-Yun Lin 1, 2 , Shaker A. Mousa 11 , Paul J. Davis 11, 12 1 Taipei Cancer Center, Taipei Medical University, Taipei, Taiwan 2 The PhD Program for Cancer Biology and Drug Discovery, College of Medical Science and Technology, Taipei Medical University, Taipei, Taiwan 3 Department of Obstetrics and Gynecology, Wan Fang Hospital, Taipei Medical University, Taipei, Taiwan 4 Integrated Laboratory, Center of Translational Medicine, Taipei Medical University, Taipei, Taiwan 5 Core Facility, Taipei Medical University, Taipei, Taiwan 6 Department of Dentistry, Wan-Fang Medical Center, Taipei Medical University, Taipei, Taiwan 7 School of Dentistry, Taipei Medical University, Taipei, Taiwan 8 Center for Teeth Bank and Dental Stem Cell Technology, Taipei Medical University, Taipei, Taiwan 9 Joint Biobank, Office of Human Research, Taipei Medical University, Taipei, Taiwan 10 Department of Pediatrics, E-DA Hospital, I-Shou University, Kaohsiung, Taiwan 11 Pharmaceutical Research Institute, Albany College of Pharmacy and Health Sciences, Albany, New York, USA 12 Department of Medicine, Albany Medical College, Albany, New York, USA * These authors contributed equally to this work Correspondence to: Yung-Ning Yang, email: ancaly@yahoo.com.tw Hung-Yun Lin, email: linhy@tmu.edu.tw Keywords: thyroid hormone, integrin αvβ3, ERα crosstalk, ovarian cancer Received: April 23, 2016 Accepted: July 10, 2016 Published: July 21, 2016 ABSTRACT Ovarian cancer is the leading cause of death in gynecological diseases. Thyroid hormone promotes proliferation of ovarian cancer cells via cell surface receptor integrin αvβ3 that activates extracellular regulated kinase (ERK1/2). However, the mechanisms are still not fully understood. Thyroxine (T 4 ) at a physiologic total hormone concentration (10 –7 M) significantly increased proliferating cell nuclear antigen (PCNA) abundance in these cell lines, as did 3, 5, 3’-triiodo-L-thyronine (T 3 ) at a supraphysiologic concentration. Thyroid hormone (T 4 and T 3 ) treatment of human ovarian cancer cells resulted in enhanced activation of the Ras/MAPK(ERK1/2) signal transduction pathway. An MEK inhibitor (PD98059) blocked hormone-induced cell proliferation but not ER phosphorylation. Knock-down of either integrin αv or β3 by RNAi blocked thyroid hormone-induced phosphorylation of ERK1/2. We also found that thyroid hormone causes elevated phosphorylation and nuclear enrichment of estrogen receptor α (ERα). Confocal microscopy indicated that both T4 and estradiol (E 2 ) caused nuclear translocation of integrin αv and phosphorylation of ERα. The specific ERα antagonist (ICI 182,780; fulvestrant) blocked T 4 -induced ERK1/2 activation, ERα phosphorylation, PCNA expression and proliferation. The nuclear co-localization of integrin αv and phosphorylated ERα was inhibited by ICI. ICI time-course studies indicated that mechanisms involved in T 4 - and E 2 -induced nuclear co-localization of phosphorylated ERα and integrin αv are dissimilar. Chromatin immunoprecipitation results showed that T 4 -induced binding of integrin αv monomer to ERα promoter and this was reduced by ICI. In summary, thyroid hormone stimulates proliferation of ovarian cancer cells via crosstalk between integrin αv and ERα, mimicking functions of E 2 .

// Ido Didi Fabian 1 , Mordechai Rosner 1 , Ina Fabian 2 , Vicktoria Vishnevskia-Dai 1 , Ofira Zloto 1 , Elena Shinderman Maman 3,4 , Keren Cohen 3,4 , Martin Ellis 4 , Hung-Yun Lin 5,6 , Aleck Hercbergs 7 , Paul J. Davis 6,8 and Osnat Ashur-Fabian 3,4 1 Goldschleger Eye Institute, Sheba Medical Center, Tel Hashomer, affiliated to Sackler Faculty of Medicine, Tel Aviv University, Tel Aviv, Israel 2 Department of Cell and Developmental Biology, the Sackler Faculty of Medicine, Tel Aviv University, Tel Aviv, Israel 3 Department of Human Molecular Genetics and Biochemistry, the Sackler Faculty of Medicine, Tel Aviv University, Tel Aviv, Israel 4 Translational Hemato-Oncology Laboratory, The Hematology Institute and Blood Bank, Meir Medical Center, Kfar-Saba, affiliated to Sackler Faculty of Medicine, Tel Aviv University, Tel Aviv, Israel 5 Institute of Cancer Biology and Drug Discovery, School of Medical Technology, Taipei Medical University, Taipei, Taiwan 6 Pharmaceutical Research Institute, Albany College of Pharmacy and Health Sciences, Albany, NY, USA 7 Department of Radiation Oncology, Cleveland Clinic, Cleveland, OH, USA 8 Department of Medicine, Albany Medical College, Albany, NY, USA Correspondence to: Ido Didi Fabian, email: // Keywords : integrin, uveal melanoma, thyroid Received : Spetember 23, 2014 Accepted : February 22, 2015 Published : March 14, 2015 Abstract Uveal melanoma is highly metastatic, prognosis is poor and there are no effective treatments to extend survival. Accumulating evidence suggests that thyroid hormones have a mitogenic effect via binding to αvβ3 integrin. We aimed to examine the impact of thyroid status on survival in a murine B16F10 model for ocular melanoma, highly expressing the integrin. In two independent experiments oral propylthiouracil (PTU) was used to induce hypothyroidism (n=9), thyroxine to induce hyperthyroidism (n=11) and mice given plain water served as control (n=8). At day 21, the subretinal space was inoculated with 10 2 B16F10 cells. In non-inoculated mice (n=6 of each group) serum free T 4 (FT 4 ) levels were measured and additional non-inoculated mice (3 given PTU and 4 given thyroxine or water) served as internal control to demonstrate the impact of the dissolved substance. The PTU-inoculated mice showed clinical evidence of intraocular tumor growth significantly later than the thyroxine mice ( P =0.003) and survival time was significantly longer ( P