Jong-Whan Choi

Repression of human telomerase reverse transcriptase using artificial zinc finger transcription factors.

Telomerase activation is a key step in the development of human cancers. Expression of the catalytic subunit, human telomerase reverse transcriptase (hTERT), represents the limiting factor for telomerase activity. In this study, we have used artificial zinc finger protein (ZFP) transcription factors (TF) to repress the expression of hTERT in human cancer cell lines at the transcriptional level. We have constructed four-fingered ZFPs derived from the human genome which binds 12-bp recognition sequences within the promoter of the hTERT gene and fused them with a KRAB repressor domain to create a potent transcriptional repressor. Luciferase activity was decreased by >80% in all of the transcriptional repressors with luciferase reporter assay. When they were transfected into the telomerase-positive HEK293 cell line, a decrease of mRNA level and telomerase activity together with shortening of telomere length was observed. Actual growth of HEK293 cells was also inhibited by transfection of artificial ZFP-TFs. The repression was maintained for 100 days of culture. The repression of telomerase expression by artificial ZFP-TFs targeting the promoter region of the hTERT presents a new promising strategy for inhibiting the growth of human cancer cells. Mol Cancer Res; 8(2); 246–53

Brain mechanisms of pain relief by transcutaneous electrical nerve stimulation: A functional magnetic resonance imaging study

Although the exact mechanism of TENS pain relief is unknown, it is believed that TENS impulses interrupt nociceptive signals at the dorsal horn of the spinal cord.

F508 amino acid deletion mutation of CFTR gene in Korean lung cancer patients.

Mutations of the transmembrane conductance regulator (CFTR) gene in cystic fibrosis lead to dysfunction of the lung, pancreas, and sweat glands, etc. To investigate the possibility of the relationship between lung cancer and the mutations of CFTR gene, we determined amino acid sequences using reverse transcription-polymerase chain reaction (RT-PCR) and DNA sequencing. In this study, the deletion mutation of 508th amino acid in one of nine lung caner patients was found confirming that CFTR gene mutation exists in a Korean lung cancer patient.

Insulin priming effect on estradiol-induced breast cancer metabolism and growth

Diabetes is a risk factor for breast cancer development and is associated with poor prognosis for breast cancer patients. However, the molecular and biochemical mechanisms underlying the association between diabetes and breast cancer have not been fully elucidated. Here, we investigated estradiol response in MCF-7 breast cancer cells with or without chronic exposure to insulin. We found that insulin priming is necessary and specific for estradiol-induced cancer cell growth, and induces anaplerotic shunting of glucose into macromolecule biosynthesis in the estradiol treated cells. Treatment with ERK or Akt specific inhibitors, U0126 or LY294002, respectively, suppressed estradiol-induced growth. Interestingly, molecular analysis revealed that estradiol treatment markedly increases expression of cyclin A and B, and decreases p21 and p27 in the insulin-primed cells. In addition, estradiol treatment activated metabolic genes in pentose phosphate (PPP) and serine biosynthesis pathways in the insulin-primed cells while insulin priming decreased metabolic gene expression associated with glucose catabolism in the breast cancer cells. Finally, we found that anti-diabetic drug metformin and AMPK ligand AICAR, but not thiazolidinediones (TZDs), specifically suppress the estradiol-induced cellular growth in the insulin-primed cells. These findings suggest that estrogen receptor (ER) activation under chronic hyperinsulinemic condition increases breast cancer growth through the modulation of cell cycle and apoptotic factors and nutrient metabolism, and further provide a mechanistic evidence for the clinical benefit of metformin use for ER-positive breast cancer patients with diabetes.

Anti-cancer effects of polyphenolic compounds in epidermal growth factor receptor tyrosine kinase inhibitor-resistant non-small cell lung cancer

Background: Polyphenolic phytochemicals are natural compounds, easily found in fruits and vegetables. Importantly, polyphenols have been intensively studied as excellent antioxidant activity which contributes to anticancer function of the natural compounds. Lung cancer has been reported to mainly account for cancer-related deaths in the world. Moreover, epidermal growth factor receptor tyrosine kinase inhibitor (TKI) resistance is one of the biggest issues in cancer treatment, especially in nonsmall cell lung cancer (NSCLC). Even though several studies both in preclinical and clinical trials have showed promising therapeutic effects of polyphenolic compounds in anticancer therapy, the function of the natural compounds in TKI-resistant (TKIR) lung cancer remains poorly studied. Objective: The aim of this study is to screen polyphenolic compounds as potential anticancer adjuvants which suppress TKIR lung cancer. Materials and Methods: Colony formation and thiazolyl blue tetrazolium blue assay were performed in the pair-matched TKI-sensitive (TKIS) versus TKIR tumor cell lines to investigate the therapeutic effect of polyphenolic compounds in TKIR NSCLC. Results: Our data show that equol, kaempferol, resveratrol, and ellagic acid exhibit strong anticancer effect in HCC827 panel. Moreover, the inhibitory effect of most of tested polyphenolic compounds was highly selective for TKIR lung cancer cell line H1993 while sparing the TKIS one H2073. Conclusion: This study provides an important screening of potential polyphenolic compounds for drug development to overcome TKI resistance in advanced lung cancer. Abbreviations used: EGFR: Epidermal growth factor receptor, EMT: Epithelial-to-mesenchymal transition, GTP: Green tea polyphenols, IGF1R: Insulin-like growth factor 1 receptor, MET: Met proto-oncogene, MTT: Thiazolyl blue tetrazolium blue, NSCLC: Non-small cell lung cancer, ROS: Reactive oxygen species, RTK: Receptor tyrosine kinase, STAT3: Signal transducer and activator of transcription 3, TKIR: TKI-resistant, TKIs: Tyrosine kinase inhibitors, TKIS: TKI-sensitive.

PPARγ sumoylation-mediated lipid accumulation in lung cancer

Metabolic reprogramming as a crucial emerging hallmark of cancer is critical for tumor cells to maintain cellular bioenergetics, biosynthesis and reduction/oxidation (REDOX) balance. Peroxisome proliferator-activated receptor gamma (PPARγ) is a nuclear hormone receptor regulating transcription of diverse gene sets involved in inflammation, metabolism, and suppressing tumor growth. Thiazolidinediones (TZDs), as selective PPARγ ligands, are insulin-sensitizing drugs widely prescribed for type 2 diabetic patients in the clinic. Here, we report that sumoylation of PPARγ couples lipid metabolism to tumor suppressive function of the receptor in lung cancer. We found that ligand activation of PPARγ dramatically induced de novo lipid synthesis as well as fatty acid beta (β)-oxidation in lung cancer both in vitro and in vivo. More importantly, it turns out that PPARγ regulation of lipid metabolism was dependent on sumoylation of PPARγ. Further biochemical analysis revealed that PPARγ-mediated lipid synthesis depletes nicotinamide adenine dinucleotide phosphate (NADPH), consequently resulting in increased mitochondrial reactive oxygen species (ROS) level that subsequently disrupted REDOX balance in lung cancer. Therefore, liganded PPARγ sumoylation is not only critical for cellular lipid metabolism but also induces oxidative stress that contributes to tumor suppressive function of PPARγ. This study provides an important insight of future translational and clinical research into targeting PPARγ regulation of lipid metabolism in lung cancer patients accompanying type 2 diabetes.

Combined therapeutic potential of nuclear receptors with receptor tyrosine kinase inhibitors in lung cancer

Cancer heterogeneity is a big hurdle in achieving complete cancer treatment, which has led to the emergence of combinational therapy. In this study, we investigated the potential use of nuclear receptor (NR) ligands for combinational therapy with other anti-cancer drugs. We first profiled all 48 NRs and 48 biological anti-cancer targets in four pairs of lung cell lines, where each pair was obtained from the same patient. Two sets of cell lines were normal and the corresponding tumor cell lines while the other two sets consisted of primary versus metastatic tumor cell lines. Analysis of the expression profile revealed 11 NRs and 15 cancer targets from the two pairs of normal versus tumor cell lines, and 9 NRs and 9 cancer targets from the primary versus metastatic tumor cell lines had distinct expression patterns in each category. Finally, the evaluation of nuclear receptor ligand T0901317 for liver X receptor (LXR) demonstrated its combined therapeutic potential with tyrosine kinase inhibitors. The combined treatment of cMET inhibitor PHA665752 or EGFR inhibitor gefitinib with T0901317 showed additive growth inhibition in both H2073 and H1993 cells. Mechanistically, the combined treatment suppressed cell cycle progression by inhibiting cyclinD1 and cyclinB expression. Taken together, this study provides insight into the potential use of NR ligands in combined therapeutics with other biological anti-cancer drugs.

PPARgamma-mediated ALDH1A3 suppression exerts anti-proliferative effects in lung cancer by inducing lipid peroxidation

Abstract Context: The metabolic function of peroxisome proliferator-activated receptor gamma (PPARγ) in lung cancer remains unclear. Objectives: To determine the relationship of PPARγ on ALDH1A3-induced lipid peroxidation to inhibit lung cancer cell growth. Materials and methods: In silico analysis using microarray dataset was performed to screen the positive correlation between PPARγ and all ALDH isoforms. NUBIscan software and ChIP assay were used to identify the binding sites (BSs) of PPARγ on ALDH1A3 promoter. The expression of ALDH1A3 under thiazolidinedione (TZD) treatment was evaluated by QPCR and Western Blot in HBEC and H1993 cell lines. Upon treatment of TZD, colony formation assay was used to check cell growth inhibition and 4-hydroxy-2-nonenal (4HNE) production as lipid peroxidation marker was determined by Western Blot in PPARγ positive cell H1993 and PPARγ negative cell H1299. Results: Compared to other ALDH isoforms, ALDH1A3 showed the highest positive correlation to PPARγ expression. ALDH1A3 upregulated PPARγ expression while PPARγ activation suppressed ALDH1A3. Among 2 potential screened PPARγ response elements, BS 1 and 2 in the promoter of ALDH1A3 gene, PPARγ bound directly to BS2. Ligand activation of PPARγ suppressed mRNA and protein expression of ALDH1A3. Growth inhibition was observed in H1993 (PPARγ positive cell) treated with PPARγ activator and ALDH inhibitor compared to H1299 (PPARγ negative cell). PPARγ activation increased 4HNE which is known to be suppressed by ALDH1A3. Conclusions: ALDH1A3 suppression could be one of PPARγ tumor suppressive function. This study provides a better understanding of the role of PPARγ in lung cancer.

Abstract 4657: LOX and ZFPM2 as novel diagnostic biomarkers for malignant pleural mesothelioma

Malignant pleural mesothelioma (MPM) is a rare, aggressive cancer being developed on outlayer of lung tissues and caused by mostly occupational exposure to asbestos. Poor prognosis needs to develop therapeutic drug as well as early diagnostic biomarker. Currently, mesothelin (MSLN), osteopontin (OPN), and fibulin 3 (FBLN3) have been reported as potential diagnostic biomarkers for MPM. In this study, we first performed bioinformatics analysis for public database to find diagnostic biomarkers for MPM. From the analysis using Cancer Cell Line Encyclopedia (CCLE) and Gene Expression Omnibus (GEO) databases, included were 7 genes involving LOX, LOXL1, LOXL2, ZFPM2, THBS2, SULF1, CDH11 identified as potential diagnostic biomarkers. These genes showed a similar diagnostic ability to FBLN3 or MSLN as MPM biomarker candidates. Further molecular approach using quantitative real-time polymerase chain reaction (QPCR) confirmed the higher mRNA expression of these candidates in MPM cell lines and patient samples. Moreover, two particular genes, LOX and ZFPM, showed MPM specific patterns of mRNA expression that were further confirmed in protein level by western blot assay. Together with biological approach, biostatistical analysis of receiver operating characteristic (ROC) analysis using the GEO database revealed significantly higher diagnostic potential of LOX and ZFPM2 genes compared to the FBLN3, one of the best diagnostic biomarkers currently reported. From this study, we believe these genes together with FBLN3 and MSLN would become novel potential biomarker candidates for MPM diagnosis. Citation Format: Minkyu Kim, Hyun-Won Kim, Soon-Hee Jung, Sung Soo Oh, Yangsik Jeong, Jong-Whan Choi. LOX and ZFPM2 as novel diagnostic biomarkers for malignant pleural mesothelioma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 4657. doi:10.1158/1538-7445.AM2017-4657

Abstract 5434: Tumor necrosis factor receptor-associated protein 1 (TRAP1) as a potential target for glutamine addicted cancer cells

Glutamine, a non-essential amino acid, is an important nutrient which is involved in many biochemical pathways such as energy production, macromolecular synthesis, and oxidative stress scavenging. Glutamine metabolism is dysregulated in many cancers which mostly display glutamine addiction for cell proliferation. Thus, glutamine metabolism has become a potential target for treating cancer. Here we report that glutamine-dependent cancer cells are more susceptible for inhibiting cell proliferation with inhibitor treatment of Tumor Necrosis Factor Receptor-Associated Protein 1 (TRAP1), a downstream factor of oncogenic c-Myc involved in glutamine metabolism. Using cell proliferation and cell viability assays, we examined growth inhibitory effects of TRAP1 inhibitor, gamitrinib-triphenylphosphonium (G-TPP), on two groups of cell lines, glutamine-deprivation sensitive versus resistant cell lines. Included are cell lines for each group: HCC827 acquired gefitinib resistance and A549 for the sensitive; HuH7 for the resistant group. Glutamine-deprivation sensitive cell lines showed significant growth inhibition with TRAP1 inhibitor treatment while the corresponding resistant group of cell lines showed no growth inhibitory effects in the same treatment condition. This suggests a potential mechanistic connection of TRAP1 to c-Myc regulation in glutamine metabolism. Taken together, this finding provides a better understanding of oncogene-driven glutamine metabolism as well as evidence that TRAP1 is a promising therapeutic target for glutamine addicted cancer cells. Citation Format: Vu T. Vo, Ai N. Phan, Tuyen N. Hua, Yangsik Jeong, Byoung Heon Kang, Hyun-Won Kim, Jong-Whan Choi. Tumor necrosis factor receptor-associated protein 1 (TRAP1) as a potential target for glutamine addicted cancer cells [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 5434. doi:10.1158/1538-7445.AM2017-5434