tyrosine phosphatase PTP-1B and the kinase Akt2 are oxidized in the cancer cells after stimulation with LPA, and these oxidation events are blocked by VPC32183 (an LPA receptor antagonist), N-acetyl cysteine, NADPH oxidase inhibitors, and intracellular catalase (which specifically scavenges H2O2). Supported by R01 CA142838 to LWD and R33 CA177461 to LBP.

DUOX1 Silencing in Lung Cancer Is Associated with Acquisition of Cancer Stem Cell Characteristics and Drug Resistance Andrew Little1, Derek Sham1, Milena Hristova1, Aida Habibovic1, and Albert Van der Vliet1 1University of Vermont, United States The NADPH oxidase homolog DUOX1 is a highly expressed enzyme within the airway epithelium. H2O2 produced via DUOX1 is known to be a potent signaling molecule in host defense and in response to inflammation. It is well established that elevated levels of reactive oxygen species (ROS) is a hallmark of cancer, paradoxically, DUOX1 has been found to be silenced in lung cancer. in order to further investigate the consequences of DUOX1 suppression, stably transfected DUOX1 silenced (shDUOX1) human pulmonary mucoepidermoid carcinoma cells (NCI-H292) were utilized. DUOX1 silencing in vitro was found to be associated with various features of epithelial-mesenchymal transition (EMT) such as: loss of E-cadherin, increased migratory potential and anchorage-independent growth. EMT in lung cancer is often linked with increased resistance to EGFR tyrosine kinase inhibitors, such as erlotinib, as well as the acquisition of cancer stem cell (CSC) phenotypes. Thus, we explored the relationship of DUOX1 silencing and these outcomes. It was found that erlotinib-resistant cells lack DUOX1 expression and congruently shDUOX1 cells display resistance to the drug erlotinib, implying that lack of DUOX1 could potentially drive EMT and the acquisition of CSC phenotypes. Flow cytometric analysis (FACS) was utilized to evaluate several proposed markers of CSCs such as: marker pair CD24Low/CD44High as well as markers CD133 and ALDH1. shDUOX1 cells displayed a greater ratio of CD24Low/CD44High as well as enhanced levels of CD133+ cells as compared to its control. Similar FACS results were observed in erlotinib-resistant cells. the relationship between DUOX1 suppression and lung cancer needs further characterization, although the downregulation of DUOX1 could prove to be an invaluable, novel marker of CSCs.

Involvement of Reactive Oxygen Species in Apoptosis Induced by Sonodynamic Therapy with Chloroaluminum Phthalocyanine Tetrasulfonate Nagahiko Yumita1, Saeko Nakamura1, Yumiko Iwase1, Toshio Fukai1, Toshihiko Ikeda1, and Shin-ichiro Umemura2 1Yokohama College of Pharmacy, Japan, 2Tohoku University, Japan Recently, we reported that some porphyrins can induce significant antitumor effect when activated by ultrasound. the present study

aims to investigate sonodynamically inducted apoptosis by chloroaluminum phthalocyanine tetrasulfonate (AlPcTS). HL-60 cells were exposed to ultrasound for up to 3 min in the presence and absence of AlPcTS. Apoptosis was analyzed by cell morphology, DNA fragmentation, and the measurement of caspase-3 activity. ESR spectrometry and spin trapping technique were used to measure reactive oxygen species. Cells treated with

and ultrasound clearly showed membrane blebbing and cell shrinkage. the number of apoptotic cells after combined treatment was significantly higher than those of other treatments. Also, DNA ladder formation and caspase-3 activation were observed in cells treated with ultrasound and AlPcTS, but not in cells treated with ultrasound or AlPcTS alone. in addition, the combination of AlPcTS and ultrasound enhanced nitroxide generation. Sonodynamically induced apoptosis, caspase-3 activation, and nitroxide generation were significantly suppressed by histidine. This significant inhibition suggests that ultrasonically generated reactive oxygen species such as singlet oxygen is an important mediator of sonodynamically induced apoptosis.

MnTE-2-PyP Inhibits Prostate Tumor Growth and Metastasis by Inhibiting p300 Activity Michael R Weaver1, Sean Smith1, Sujatha Venkataraman2, and Rebecca E. Oberley-Deegan1 1National Jewish Health, United States, 2University of Colorado, United States Roughly 1.5 million US men are prostate cancer survivors due to improvements in radiation therapy. However, the majority of these survivors will suffer significant reduction in quality of life stemming from the long term side effect(s) of pelvic irradiation exposure, including erectile dysfunction. Currently there are no effective clinical treatments used to protect normal tissues from pelvic radiation induced damage. Our laboratory has shown that the superoxide dismutase (SOD) mimetic, MnTE-2-PyP, is a potent radio-protector against radiation induced damage to urogenital tissues. We have shown that the administration of MnTE-2-PyP during radiation therapy protects from developing erectile dysfunction in rats. However, in order for MnTE-2-PyP to work as a radio-protector, MnTE-2-PyP cannot protect the prostate tumor from radiation damage. the purpose of this study was to determine if MnTE-2-PyP has anti-tumor properties. MnTE-2-PyP (1-30 μM), in a dose dependant fashion, significantly inhibited growth of three prostate cancer cells: PC3, Du145, and LnCAP cells using both colony focus assays and soft agar assays. MnTE-2-PyP further reduced the growth of tumor cells when added in combination with radiation (2 or 5 Gy). MnTE-2-PyP also significantly inhibited the invasiveness of PC3, Du145 and LnCAP cells on matrigel and significantly inhibited LnCAP and Du145 cells ability to migrate across a filter. in nude mice, MnTE-2-PyP inhibited growth of PC3 cells in the absence or presence of radiation. We next wanted to better understand how MnTE-2-PyP was inhibiting growth of the prostate tumors. p300 is enhanced in prostate tumors and acts as a co-factor to enhance transcription of pro-survival genes in tumors. We made the novel discovery that MnTE-2-PyP inhibits the activity of the histone acetyltransferase (HAT) enzyme, p300. MnTE-2-PyP also inhibits one of the downstream pro-survival proteins regulated by p300, cIAP2. Thus, we conclude that MnTE-2-PyP may work as an anti-tumor agent by inhibiting p300 activity in prostate cancer cells.

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