bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 2398: Activity of the PARP inhibitor olaparib in ATM-deficient gastric cancer: from preclinical models to the clinic
Authors: Hodgson Darren , Mason Helen , Oplustilova Lenka , Harbron Chris , Yin Xiaolu , Im Seock-Ah , Jones Helen , Zhongwu Lai , Dougherty Brian , McLoughlin Matthew , Bradford James , Dickinson Andrew , Fielding Anitra , Robertson Jane , Kim Woo-Ho , Womack Chris , Gu Yi , Bang Yung-Jue , Lau Alan , Barrett J. Carl , O'Connor Mark J.
Published: May 2015

Olaparib is an oral inhibitor of the Poly-(ADP-ribose)-polymerases (PARP-1, -2 and -3), and has demonstrated clinical activity in trials of patients with BRCA-deficient tumors. The BRCA1 and BRCA2 proteins are both important for the repair of DNA double strand breaks (DSBs) by homologous recombination repair (HRR) and sensitivity to olaparib has further been shown preclinically to extend to other DNA DSB repair factor deficiencies, including those in the ataxia telangiectasia mutated (ATM) protein.

We have demonstrated that a high proportion of gastric cancer (GC) cell lines (∼50%) are responsive (IC50 <500 nM) to single agent olaparib and that characterization of their DNA repair status reveals that, as in breast and ovarian cancer, HRR deficiencies (HRD) can account at least in part for this. However, unlike breast and ovarian cancer cell lines, there is no strong correlation with platinum sensitivity, suggesting alternative drivers of response in the GC panel to HRD. Here, we demonstrate that low levels of ATM protein expression correlate with sensitivity of GC cell lines to olaparib, and moreover can be observed in approximately 15-20% of clinical gastric cancer samples when evaluated using ATM immunohistochemistry (IHC).

Based on these data, a Phase II GC trial has been carried out that enriches for patients with tumors having low or undetectable ATM IHC scores (H-score equivalent less than or equal to 10). Data from this clinical trial confirm activity (overall survival [OS] benefit HR=0.56, 95% CI: 0.35-0.87; P=0.010) in the overall population, which includes ATM tumor positive patients, and importantly, a substantially better OS benefit in the ATM-negative population (HR=0.35, 95% CI: 0.17-0.71; P=0.003). Together, these data provide an important example of a PARP inhibitor demonstrating both preclinical and clinical activity in a well defined but non-BRCA DNA repair deficient background.

Citation Format: Darren Hodgson, Helen Mason, Lenka Oplustilova, Chris Harbron, Xiaolu Yin, Seock-Ah Im, Helen Jones, Lai Zhongwu, Brian Dougherty, Matthew McLoughlin, James Bradford, Andrew Dickinson, Anitra Fielding, Jane Robertson, Woo-Ho Kim, Chris Womack, Yi Gu, Yung-Jue Bang, Alan Lau, J. Carl Barrett, Mark J. O'Connor. Activity of the PARP inhibitor olaparib in ATM-deficient gastric cancer: from preclinical models to the clinic. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 2398. doi:10.1158/1538-7445.AM2014-2398
Abstract 24: HFE genotype affects tumor progression and macrophage behavior in a mouse model
Authors: Weston Cody L. , Alkhateeb Ahmed , Hund William , Connor James R.
Published: May 2015

Introduction: Studies suggest that iron metabolism is associated with tumor progression, but a mechanism and the relative importance of different iron handling proteins still remain to be elucidated. We investigated the effects of the frequently polymorphic high iron gene (HFE), on tumor growth. This study is significant because of the ability of HFE mutant protein to alter iron handling and because our previous studies have shown the presence of this mutation is associated with altered cancer phenotype in cells.

Hypothesis: Mutations in the HFE protein modify tumor progression and metastatic potential.

Methods: C57BL/6 mice possessing either H67D/H67D (H63D in humans) or WT/WT HFE genotype, aged 18 months, were injected subcutaneously with 4x106 B16F10 cells. After 2 weeks, mice were sacrificed and the tumors were weighed and prepared for analysis of RNA, protein, and histology. At the same time, plasma was collected by cardiac puncture. Animal methods were approved by our IACUC, protocol 04-166. RT-PCR analysis of tumors was performed using the Mouse Cancer Pathwayfinder Array (SABiosciences). In parallel, bone marrow derived macrophages were cultured to determine how B16F10 cell cultures react to conditioned media from macrophages of each genotype. ELISAs were performed for ferroportin and ferritin. Secreted cytokine and chemokine content of macrophage media was assessed with an ELISA array (Qiagen).

Results: H67D mice had significantly smaller tumors (t-test, P = 0.0166) after two weeks. The two-week time frame was selected because rapid tumor growth affected animal survival at longer incubations. Exploratory qRT-PCR analysis of tumors revealed that the H67D host may promote epithelial phenotype and suppress angiogenesis and growth. In culture, macrophages derived from H67D mice express higher levels of MCP1 (p = 0.0169). Conditioned media from the H67D macrophages was associated with significantly less B16F10 cell growth compared to WT (p < 0.0001 MTT, p = 0.0008 BrdU). In macrophage cell lysates, H67D is associated with higher levels of Ferroportin (p = 0.0297), and WT macrophages showed higher iron levels in response to FAC (p = 0.0295).

Conclusions: Tumors grown in H67D mice are significantly lower in weight than control mice. Macrophages from H67D mice neither load iron nor support tumor cell proliferation as well as WT controls. Our data reveal the importance of HFE genotype on macrophage function and tumor growth. The H63D HFE allele is seen in approximately 15% of the human population so its impact on cancer studies is likely significant.

Citation Format: Cody L. Weston, Ahmed Alkhateeb, William Hund, James R. Connor. HFE genotype affects tumor progression and macrophage behavior in a mouse model. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 24. doi:10.1158/1538-7445.AM2014-24
Abstract 240: Natural antioxidants exhibit chemopreventive characteristics through the regulation of CNC-bZip transcription factors in estrogen-induced breast carcinogenesis
Authors: Chatterjee Anwesha , Ronghe Amruta , Abdalla Fatma , Bhat Hari K.
Published: May 2015

Epidemiological data and studies in rodent models strongly support the role of estrogens in the development of breast cancers. Exact mechanisms underlying the initiation and progression of estrogen-related cancers are not clear. Literature evidence and our studies strongly support the role of estrogen metabolism mediated oxidative stress in estrogen-induced breast carcinogenesis. We have recently demonstrated that antioxidants vitamin C (VC) or butylated hydroxyanisole (BHA) severely inhibit 17β-estradiol (E2)-induced breast tumor development in female ACI rats. The objective of this study was to characterize the role of CNC b-zip transcription factors in antioxidant-mediated prevention of breast cancer.

Human non-neoplastic breast epithelial cell line MCF-10A was treated with E2, VC, VC + E2, Resveratrol (Res) and Res + E2 for up to 72 hrs. mRNA and protein expression levels of phase-II detoxifying enzymes SOD1, SOD2, SOD3, GPx, NQO1 and transcription factors from CNC b-zip family: Nrf1, Nrf2 and Nrf3 were quantified after treatment with E2 and compared with that of cells treated with antioxidants either alone or in combination with E2.

The mRNA as well as protein expression levels of SOD3 and NQO1 were suppressed in MCF-10A cells treated with E2. There was no change in mRNA and protein expression levels of SOD1, SOD2 and GPX in MCF-10A cells treated with E2. Nrf2 mRNA and protein expression levels were also significantly suppressed in E2-treated MCF-10A cells. Vitamin C or Res treatment prevented E2-mediated decrease in SOD3, NQO1 and Nrf2 mRNA and protein expression levels. A significant increase in Nrf1 mRNA and protein expression levels in MCF-10A cells treated with E2 was noted. Further, treatment with antioxidants Res or VC also increased mRNA and protein expression levels of Nrf1. Nrf3 mRNA and protein expression levels were significantly increased in MCF-10A cells treated with E2. This induction was overcome by co-treatment with E2 and VC or Res.

Our studies demonstrate that antioxidant-mediated protection against estrogen-induced breast cancer could be through up regulation of phase-II detoxifying enzymes via an Nrf-dependent pathway involving a complex interplay among Nrfs 1, 2 and 3.

Note: This abstract was not presented at the meeting.

Citation Format: Anwesha Chatterjee, Amruta Ronghe, Fatma Abdalla, Hari K. Bhat. Natural antioxidants exhibit chemopreventive characteristics through the regulation of CNC-bZip transcription factors in estrogen-induced breast carcinogenesis. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 240. doi:10.1158/1538-7445.AM2014-240
Abstract 2402: Antitumor effect of a poly (ADP-ribose) polymerase (PARP) inhibitor in endometrial carcinoma cell lines
Authors: Miyasaka Aki , Oda Katsutoshi , Ikeda Yuji , Kashiyama Tomoko , Koso Takahiro , Inaba Kanako , Fukuda Tomohiko , Makii Chinami , Asada Kayo , Kurikawa Reiko , Hiraike Osamu Wada , Sone Kenbun , Uehara Yuriko , Matsumoto Yoko , Arimoto Takahide , Kuramoto Hiroyuki , Yano Tetsu , Kawana Kei , Osuga Yutaka , Fujii Tomoyuki
Published: May 2015

The loss of PTEN is the most frequent genetic aberration in endometrial cancer. One of the phosphatase-independent roles of PTEN is associated with homologous recombination (HR) in the nucleus. Poly (ADP-ribose) polymerase (PARP) plays a key role in the repair of DNA single-strand breaks, and a PARP inhibitor induces synthetic lethality in cancer cells with HR deficiency. We examined the anti-tumor activity of olaparib, a PARP inhibitor, and its correlation between the sensitivity and status of PTEN in a panel of endometrial cancer cell lines.

Methods

The response to olaparib was evaluated using a clonogenic assay with SF50 values (concentration to inhibit cell survival to 50%) in 16 endometrial cancer cell lines. The effects of PTEN introduction on the sensitivity of cells to olaparib and ionizing radiation (IR) exposure were compared between parental HEC-6 (PTEN-null) and HEC-6 PTEN+ (stably expressing wild-type PTEN) cells by clonogenic assay, foci formation of RAD51 and γH2AX, and induction of cleaved PARP.

Results

The SF50 values were 100 nM or less in six of the 16 (38%: sensitive) cell lines; whereas, SF50 values were 1,000 nM or more in five of the 16 (31%: resistant) cell lines. PTEN mutations were not associated with sensitivity to olaparib (Mutant [n = 12]: 995 ± 1,096 nM; Wild-type [n = 4]: 144 ± 91 nM, p = 0.17 by Student'st test). RAD51 expression was observed broadly and was not associated with PTEN status in the 16 cell lines. The number of colonies in the clonogenic assay, the foci formation of RAD51 and γH2AX, and the induction of apoptosis were not affected by PTEN introduction in the HEC-6 PTEN+ cells. The expression level of nuclear PTEN was not elevated within 24 h following IR in the HEC-6-PTEN+ cells.

Conclusions

Our results suggest that olaparib, a PARP inhibitor, is effective on certain endometrial cancer cell lines. The inactivation of PTEN might not affect the DNA repair function. Predictive biomarkers are warranted to utilize olaparib in endometrial cancer.

Citation Format: Aki Miyasaka, Katsutoshi Oda, Yuji Ikeda, Tomoko Kashiyama, Takahiro Koso, Kanako Inaba, Tomohiko Fukuda, Chinami Makii, Kayo Asada, Reiko Kurikawa, Osamu Wada Hiraike, Kenbun Sone, Yuriko Uehara, Yoko Matsumoto, Takahide Arimoto, Hiroyuki Kuramoto, Tetsu Yano, Kei Kawana, Yutaka Osuga, Tomoyuki Fujii. Antitumor effect of a poly (ADP-ribose) polymerase (PARP) inhibitor in endometrial carcinoma cell lines. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 2402. doi:10.1158/1538-7445.AM2014-2402
Abstract 2403: Imaging of UVB and UVC-induced DNA damage repair in cancer cells in Gelfoam histoculture and minimal cancer in mice
Authors: Uehara Fuminari , Miwa Shinji , Tome Yasunori , Maehara Hiroki , Kanaya Fuminori , Hiroshima Yukihiko , Yano Shuya , Yamamoto Mako , Matsumoto Yasunori , Efimova Elena V. , Hoffman Robert M.
Published: May 2015

We compared the DNA damage repair response of cancer cells after UVB or UVC irradiation. The DNA-damage repair response to UV irradiation was imaged in tumors growing in 3D Gelfoam® histoculture and in superficial tumors grown in mice. UV-induced DNA damage repair was imaged with GFP fused to the DNA damage response (DDR)-related chromatin-binding protein 53BP1 in MiaPaCa-2 human pancreatic cancer cells. 53BP1 forms foci during DNA damage repair. A clonogenic assay initially showed that UVC and UVB inhibited MiaPaCa-2 cell proliferation in a dose-dependent manner, with UVC having more efficacy. Gelfoam® histocultures and confocal imaging enabled 53BP1-GFP nuclear foci to be observed within 1 h after UV irradiation, indicating the onset of DNA damage repair response. Induction of UVB-induced 53BP1-GFP focus formation was observed up to a depth of 120 µm in MiaPaCa-2 cells on Gelfoam®, while UVC induced foci only to a depth of 40 µm. The MiaPaCa-2 cells irradiated by both UVB and UVC light in a skin-flap mouse model had a significant decrease in tumor growth compared to untreated controls. Moreover, UVB-treatment had greater inhibition of tumor growth compared to UVC. Our results demonstrate that both UVB and UVC are useful tools for the treatment of residual cancer, and UVB was more effective than UVC in vivo, possibly due to greater tissue penetration of UVB because of its longer wavelength.

Citation Format: Fuminari Uehara, Shinji Miwa, Yasunori Tome, Hiroki Maehara, Fuminori Kanaya, Yukihiko Hiroshima, Shuya Yano, Mako Yamamoto, Yasunori Matsumoto, Elena V. Efimova, Robert M. Hoffman. Imaging of UVB and UVC-induced DNA damage repair in cancer cells in Gelfoam histoculture and minimal cancer in mice. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 2403. doi:10.1158/1538-7445.AM2014-2403
Abstract 2409: BCL10 regulates RNF8/RNF168-mediated ubiquitination in the DNA damage response
Authors: Zhao Hongchang , Zhu Min , Xu Xingzhi
Published: May 2015

Our genome is constantly attacked by a variety of endogenous and environmental agents. Timely and properly cellular response to DNA damage is essential for maintenance of genome stability and integrity, which plays key roles in prevention of tumor initiation and development. B-cell lymphoma/leukemia 10 (BCL10), when translocated or mutated, contributes to mucosa-associated lymphoid tissue lymphomas and other malignancies. However, the mechanisms by which the resulting aberrant expression of BCL10 leads to tumorigenesis are poorly understood. In this report, we uncovered that, in response to DNA damage, BCL10 is recruited to the damage sites and modified sequentially by phosphorylation and ubiquitination. Modified BCL10 in turn facilitates to bring about cascades of ubiquitination and recruitment of DNA damage response (DDR) factors, promoting high-fidelity DNA repair. Furthermore, BCL10 is over-expressed in breast tumors (376 cases of breast cancer vs. 62 cases of normal breast, p=0.000). Taken together, our study reveals a novel function of BCL10 in ubiquitination-driven DDR, contributing to the maintenance of genome stability and integrity.

Note: This abstract was not presented at the meeting.

Citation Format: Hongchang Zhao, Min Zhu, Xingzhi Xu. BCL10 regulates RNF8/RNF168-mediated ubiquitination in the DNA damage response. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 2409. doi:10.1158/1538-7445.AM2014-2409
Abstract 2418: Investigating p53 and other potential determinants of cell sensitivity to ATR inhibition by VE-821
Authors: Middleton Fiona K. , Chen Tao , Pollard John R. , Curtin Nicola J.
Published: May 2015

Background: Defects in the multifactorial DNA damage response (DDR) are common in cancer, making them vulnerable to inhibitors of other components of the DDR. ATR is a key component of the DDR, signalling ssDNA, arising from stalled replication forks, resected double strand breaks (DSBs) and NER intermediates to activate cell cycle arrest at the S/G2 checkpoints and initiate homologous recombination repair (HRR). Tumors often have a dysfunctional G1 checkpoint e.g. due to p53 mutations or other DDR defects. ATR inhibition of the S/G2 checkpoints and HRR has the potential to exploit these defects and selectively target cancer. VE-821 is a potent ATR inhibitor under advanced pre-clinical investigation

Hypotheses: 1) Cells with dysfunctional p53 will be more sensitive to ATR inhibition. 2) Defects in other DDR components will sensitise cells to ATR inhibition.

Methods: The survival of isogenic p53 mutant/null and wt/corrected human cell lines (HCT116 p53+/+ and p53-/- or U2OS p53WT and p53DN (dominant negative)) and a panel of repair defective Chinese hamster ovary and lung fibroblast cells was determined using colony formation after 24 hours exposure VE-821 alone or in the presence of gemcitabine or ionising radiation.

Results: There was no significant difference in sensitivity to VE-821 between p53 wt or mutant/null cells in either HCT116 (LC50 = 2.13 and 4.56 µM, respectively) or U2OS pairs (LC50 = 2.54 and 3.34 µM, respectively). However, 1 µM VE-821 significantly potentiated the antimetabolite, gemcitabine, in HCT116 p53-/- (4.3-fold) but not p53+/+ cells (1.5-fold), and enhanced cell kill by 100 nM gemcitabine 40-fold in U2OS p53DN cells but only 3-fold in U2OS p53WT cells. Chinese hamster ovary cells defective in XRCC1 (single stranded break repair) were more sensitive to single agent VE-821 than the parental wt cells (35% and 55% survival at 10 µM, respectively). Data on other repair-defective cell lines will be presented.

Conclusions: p53 status is not a determinant of sensitivity to VE-821 as a single agent but VE-821 preferentially sensitises cells lacking functional p53 to gemcitabine. This supports the hypothesis that p53 status may be predictive of cellular response to ATR inhibition in combination with certain chemotherapies. Furthermore, data using Chinese hamster cell lines indicate that DDR defects may be exploited by ATR inhibition.

Citation Format: Fiona K. Middleton, Tao Chen, John R. Pollard, Nicola J. Curtin. Investigating p53 and other potential determinants of cell sensitivity to ATR inhibition by VE-821. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 2418. doi:10.1158/1538-7445.AM2014-2418
Abstract 2424: DNA mismatch repair protein, PMS2, induces apoptosis in human prostate cancer cells
Authors: Fukuhara Shinichiro , Chang Inik , shahryari varahram , gill ankurpreet , wong darryn K. , yamamura soichiro , majid shahana , saini sharanjot , hirata hiroshi , ueno koji , deng gouren , tabatabai laura Z. , nonomura norio , dahiya rajvir , tanaka yuichiro
Published: May 2015

Prostate cancer is one of the most common malignancies and ranks the second most common cause of cancer-related deaths in men. DNA mismatch repair (MMR) enzymes act as proofreading complexes that maintain the genomic integrity and MMR-deficient cells show an increased mutation rate. Recent studies have shown that MMR influences the regulation of tumor development in various cancers such as colon, ovarian and others, but their effect on prostate cancer growth has not been investigated thus far. The aim of the present study was to investigate the effect of the post-meiotic segregation (PMS) 2 MMR gene on growth properties of prostate cancer cells. Expression of PMS2 was initially measured in normal and cancerous prostate cell lines by real-time PCR and western blotting. Cancerous cells displaying the lowest levels compared to normal cells was then transfected with PMS2. Gene effect on various cellular properties (cell proliferation, migration, invasion, apoptosis and cell cycle assays) as well as growth in nude mice were determined. Of the various prostate cell lines, DU145 had dramatically reduced expression levels of PMS2 with little protein detected, whereas levels were high in normal prostate cells. Interestingly, over-expressing the PMS2 gene in DU145 cells decreased cell proliferation, migration and invasion, and increased apoptosis and cell cycle arrest compared to vector control. PMS2 also inhibited DU145 cell tumor formation in animal models. These results demonstrate PMS2 to protect against prostate cancer progression and to have a functional role by affecting apoptosis and cell cycle of prostate cancer cells.

Citation Format: Shinichiro Fukuhara, Inik Chang, varahram shahryari, ankurpreet gill, darryn K. wong, soichiro yamamura, shahana majid, sharanjot saini, hiroshi hirata, koji ueno, gouren deng, laura Z. tabatabai, norio nonomura, rajvir dahiya, yuichiro tanaka. DNA mismatch repair protein, PMS2, induces apoptosis in human prostate cancer cells. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 2424. doi:10.1158/1538-7445.AM2014-2424
Abstract 2425: Exploiting DNA repair defects in breast cancer
Authors: Naipal Kishan A.T. , Verkaik Nicole S. , Brugge Petra ter , Ameziane Najim , Deurzen Carolien H.M. van , Martens John W. , Winter Johan P. de , Jonkers Jos , Vreeswijk Maaike P. , Jager Agnes , Hoeijmakers Jan , Kanaar Roland , Gent Dik C. van
Published: May 2015

One of the hallmarks of cancer is genetic instability. Many chemotherapeutic strategies make use of DNA damaging agents, that preferentially target dividing cells. Recently, DNA repair defects have been identified in many different types of tumors, suggesting that the effectiveness of such treatments may be modulated by the precise genetic make-up of the tumor; defects in specific DNA repair pathways can be used as the Achilles' Heel of the tumor. An important new concept is synthetic lethality, the observation that combinations of mutations can be lethal, while both single mutations are viable. In selected tumors, the specific DNA repair defect can be targeted in such a way that the tumor cells can be killed without causing severe side effect on normal tissues. The combination of PARP inhibitors and homologous recombination (HR) deficient hereditary breast cancer is on its way to the clinic and more combinations are currently in the preclinical phase. Selection of breast cancer patients for targeted treatment with PARP inhibitors requires a validated test to identify HR deficient tumors. For this purpose, we have assessed ionizing radiation induced RAD51 foci (IRIF) after ex vivo irradiation of individual breast tumor slices as a marker for HR defects. We validated this approach in known HR- deficient tumors, where we found a severe defect in RAD51 IRIF formation. Subsequently, we used this assay to identify a subgroup of HR deficient primary breast cancers. Out of 42 tumors, we found five HR deficient breast cancers, caused by two distinct modes of BRCA gene inactivation, mutations and promoter hypermethylation. HR deficiency was significantly associated with high grade, triple-negative breast cancer (TNBC). A large fraction of TNBC showed HR defects, suggesting that PARP inhibitors may be particularly promising for this group of tumors that currently have a poor prognosis. We conclude that the RAD51 IRIF assay is a powerful tool to select patients with HR-deficient primary breast cancers eligible for PARP-inhibitor treatment in the clinic. These approaches may change cancer medicine in a fundamental way, from a one-size-fits-all concept to an individualized treatment strategy based on the functional molecular make-up of the tumor for each patient.

Citation Format: Kishan A.T. Naipal, Nicole S. Verkaik, Petra ter Brugge, Najim Ameziane, Carolien H.M. van Deurzen, John W. Martens, Johan P. de Winter, Jos Jonkers, Maaike P. Vreeswijk, Agnes Jager, Jan Hoeijmakers, Roland Kanaar, Dik C. van Gent. Exploiting DNA repair defects in breast cancer. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 2425. doi:10.1158/1538-7445.AM2014-2425
Abstract 2429: Temporally and spatially inducible mouse model of aneuploidy
Authors: Eichberg Daniel , Knouse Kristin , Amon Angelika
Published: May 2015

Aneuploidy, the state in which cells have an abnormal number of chromosomes, is a hallmark of most human cancers. Despite aneuploidy's near-ubiquitous presence in tumors, it is unknown whether aneuploidy is a driver or passenger in cancer progression. Aneuploidy is normally prevented by the spindle assembly checkpoint (SAC), a critical cellular mechanism which blocks chromosomal missegregation by inhibiting premature sister chromatid separation during mitosis. Although transgenic mouse models with defective SAC proteins exist, the dysfunctional proteins are expressed constitutively from birth in all tissues, a milieu that differs from that of an in vivo human tumor in which cancer cells develop aneuploidy de novo in the context of a euploid organism. In order to generate a mouse model that more closely approximates this tumor environment, we will insert single copies of shRNA hairpins that target SAC proteins BubR1 or Mad2 into the mouse genome at the Col1A1 locus using recombination mediated cassette exchange. shRNA expression will be under the control of a tetracycline responsive element in tissues that co-express rtTA, leading to temporally and spatially inducible disruption of the SAC, and thus aneuploidy. In vitro shRNA construct validation experiments indicate our shRNAs successfully knockdown target RNA and protein levels, as well as functionally promote mitotic slippage after nocodazole arrest, which is indicative of an impaired SAC. We will use this mouse model to investigate the role of increased chromosomal missegregation on tumor initiation and evolution, as well as the impact of increased aneuploidy on normal organ function.

[Daniel Eichberg is a Howard Hughes Medical Institute Medical Research Fellow.]

Citation Format: Daniel Eichberg, Kristin Knouse, Angelika Amon. Temporally and spatially inducible mouse model of aneuploidy. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 2429. doi:10.1158/1538-7445.AM2014-2429
Abstract 2431: Downregulation of PHLPP promotes glycolysis in colon cancer cells
Authors: Wen Yang-an , Xiong Xiaopeng , Gao Tianyan
Published: May 2015

PHLPP belongs to a novel family of Ser/Thr protein phosphatases. Our previous studies have shown that PHLPP can antagonize multiple signaling pathways by negatively regulating signaling molecules such as Akt, S6K, and PKC. Functionally, PHLPP serves as a tumor suppressor to inhibit cell proliferation, promote apoptosis and increase therapeutic sensitivity in colon cancer cells. Here, we further examined the role of PHLPP in regulating metabolic pathways in colon cancer. Control and PHLPP knockdown cells colon cancer cells, as well as wild-type and Phlpp knockout MEF cells, were subjected to bioenergetic phenotype analysis using Seahorse XF96 analyzer. The mitochondrial respiration measurements revealed that decreased PHLPP expression resulted in an increase in oxygen consumption rate (OCR) suggesting enhanced cellular respiration. In addition, PHLPP depletion significantly stimulated glycolysis as indicated by increased extracellular acidification rate (ECAR). Consistently, the rates of glucose consumption and lactate production were significantly increased in PHLPP knockdown cells. Furthermore, the numbers of apoptotic cells were increased in PHLPP knockdown cells compared to control cells when cultured in glucose-free medium, suggesting that loss of PHLPP expression render cells more sensitive to glucose deprivation as they became relied on glycolysis. To begin determining the underlying molecular mechanism, we found that the activity of both Akt and S6K was largely induced in PHLPP-knockdown cancer cells and PHLPP-knockout MEF cells. Since both kinases are known to play important roles in re-programming metabolic networks to favor tumor growth, it is likely that PHLPP inhibits glycolysis by negatively regulating Akt- and S6K-mediated metabolic effects. Taken together, our studies identified PHLPP as a novel regulator of glucose metabolism in colon cancer cells. Downregulation of PHLPP may promote tumorigenesis by providing metabolic advantages in colon cancer

Citation Format: Yang-an Wen, Xiaopeng Xiong, Tianyan Gao. Downregulation of PHLPP promotes glycolysis in colon cancer cells. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 2431. doi:10.1158/1538-7445.AM2014-2431
Abstract 2433: Metabolic reprogramming by an epigenetic mechanism in endocrine therapy resistance of breast cancer
Authors: Zou June X. , Wang Junjian , Duan Zhijian , Chen Hongwu , Kung Hsing-Jien , Chen Xinbin , Murphy Leigh C. , Borowsky Alexander
Published: May 2015

Metabolic reprogramming is a major mechanism of fueling cancer cell growth and proliferation. However, how tumor metabolism is altered to confer therapeutic resistance is poorly understood. Gene expression profiling of tamoxifen sensitive and resistant breast cancer cells revealed that transition to the resistance was associated with a specific expression reprogramming of genes involved in glucose/energy metabolism, which was accompanied by aberration of specific epigenetic regulators such as the histone methyltransferase NSD2/MMSET/WHSC1. In cell and xenograft tumor models, NSD2 overexpression alone was sufficient to confer tamoxifen resistance. Analysis of clinical specimens indicates that NSD2 was a strong predictive factor for early relapse of tamoxifen therapy. NSD2 wild type, but not its methylase-defective mutant form, coordinately stimulated the expression and enzymatic activity of HK2, TIGAR and G6PD and strongly augmented the pentose phosphate pathway (PPP) production of NADPH for ROS reduction and survival of tamoxifen treated cells and tumors. Further mechanistic studies demonstrated that elevated NSD2 stimulates the metabolic gene expression through H3K36 methylation at the target chromatin. Pharmacological targeting aberrant NSD2 by DZNeP, an S-adenosylhomocysteine (AdoHcy) hydrolase inhibitor, effectively blocked the growth of tamoxifen-resistant tumor. The study reveals a novel mechanism of cancer metabolic reprogramming and suggests targeting the key metabolic re-programmer as a viable option for effective treatment of endocrine resistant tumors.

Note: This abstract was not presented at the meeting.

Citation Format: June X. Zou, Junjian Wang, Zhijian Duan, Hongwu Chen, Hsing-Jien Kung, Xinbin Chen, Leigh C. Murphy, Alexander Borowsky. Metabolic reprogramming by an epigenetic mechanism in endocrine therapy resistance of breast cancer. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 2433. doi:10.1158/1538-7445.AM2014-2433
Abstract 2441: mTOR complex 2 modulates glycosylation of CD147 via the hexosamine biosynthetic pathway
Authors: Wu Chang-Chih , Lynch Thomas , Moloughney Joseph , Navia Aixa , Ibironke Olufunmilola , Chou Po-Chien , Vega-Cotto Nicole M. , Zhang Sisi , Rabinowitz Joshua , Werlen Guy , Jacinto Estela
Published: May 2015

Cell surface proteins transduce extracellular signals into the cell to control metabolism and growth. In turn, their expression is linked to nutrient availability and other growth signals by mechanisms that are poorly understood. The mammalian target of rapamycin (mTOR) regulates cell growth and metabolism and is part of two distinct protein complexes, mTOR complex 1 (mTORC1) and mTORC2. We found that mTORC2 is involved in the processing and maturation of cell surface receptors such as CD147. CD147 is a highly glycosylated receptor that has been linked to tumor progression via its role in activating matrix metalloproteinases and maturation of lactate transporters. In breast cancer cells, CD147 glycosylation is highly sensitive to glucose starvation and mTOR inhibition. In mTORC2-disrupted cells CD147 is misprocessed and occurs predominantly in a low glycosylated form. CD147 misprocessing can be partly rescued by addition of exogenous UDP-GlcNAc, the end product of the hexosamine biosynthetic pathway (HBP). However, UDP-GlcNAc cannot restore the abnormal growth and metabolism in mTORC2-disrupted cells due to defects in expression of other key metabolic enzymes in these cells. Our findings define a role for mTORC2 in regulating receptor glycosylation via the HBP and reveal a broader role for mTORC2 in controlling other biosynthetic pathways that become deregulated in cancer.

Citation Format: Chang-Chih Wu, Thomas Lynch, Joseph Moloughney, Aixa Navia, Olufunmilola Ibironke, Po-Chien Chou, Nicole M. Vega-Cotto, Sisi Zhang, Joshua Rabinowitz, Guy Werlen, Estela Jacinto. mTOR complex 2 modulates glycosylation of CD147 via the hexosamine biosynthetic pathway. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 2441. doi:10.1158/1538-7445.AM2014-2441
Abstract 2444: Targeting Epstein-Barr virus oncoprotein LMP1-mediated glycolysis sensitizes nasopharyngeal carcinoma to radiation therapy
Authors: Xiao Lanbo , Hu Zhe-yu , Cao Ya
Published: May 2015

Our goal in this work was to illustrate the Epstein-Barr Virus (EBV)-modulated global biochemical profile and provide a novel metabolism-related target to improve the therapeutic regimen of nasopharyngeal carcinoma (NPC). We used a metabolomics approach to investigate EBV-modulated metabolic changes, and found that the exogenous overexpression of the EBV encoded latent membrane protein 1 (LMP1) significantly increased glycolysis. The deregulation of several glycolytic genes, including hexokinase 2 (HK2), was determined to be responsible for the reprogramming of LMP1-mediated glucose metabolism in NPC cells. The up-regulation of HK2 elevated aerobic glycolysis and facilitated proliferation by blocking apoptosis. More importantly, HK2 was positively correlated with LMP1 in NPC biopsies, and high HK2 levels were significantly associated with poor overall survival of NPC patients following radiation therapy. Knockdown of HK2 effectively enhanced the sensitivity of LMP1-overexpressing NPC cells to irradiation. Finally, c-Myc was demonstrated to be required for LMP1-induced up-regulation of HK2. The LMP1-mediated attenuation of the PI3-K/Akt-GSK3beta-FBW7 signaling axis resulted in the stabilization of c-Myc. These findings indicate a close relationship between EBV and glycolysis in NPC. Notably, LMP1 is the key regulator of the reprogramming of EBV-mediated glycolysis in NPC cells. Given the importance of EBV-mediated deregulation of glycolysis, anti-glycolytic therapy might represent a worthwhile avenue of exploration in the treatment of EBV-related cancers.

Note: This abstract was not presented at the meeting.

Citation Format: Lanbo Xiao, Zhe-yu Hu, Ya Cao. Targeting Epstein-Barr virus oncoprotein LMP1-mediated glycolysis sensitizes nasopharyngeal carcinoma to radiation therapy. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 2444. doi:10.1158/1538-7445.AM2014-2444
Abstract 2446: Signaling event of alcohol-induced deregulation of Pol III genes in breast cancer cells
Authors: Zhong Shuping , Zhang Qingsong , Shi Ganggang
Published: May 2015

Alcohol intake is consistently associated with an increased risk of breast cancer. Alcohol consumption is more pronounced in ER+ breast cancer cases than in ER- cases. However, its mechanism remains to be determined. We reported that alcohol induces Pol III gene (RNA polymerase III-dependent genes) transcription in vivo and in vitro. We continue to define novel and unexpected targets, MSK1 (mitogen- stress-activated protein kinase 1) and Brf1 (TFIIIB-related factor 1), in the alcohol-induced response. Our aim is to delineate the mechanisms regulating the processes of deregulation of Pol III genes, cell transformation and tumorgenesis through this alcohol-induced response. Brf1 specifically regulates Pol III gene transcription. Changes in Pol III gene and Brf1 expression tightly link to cell transformation and tumor formation. Alcohol-induced deregulation of Pol III genes may be fundamental to the development of breast cancer. We have reported that a) enhancement of Brf1 and Pol III gene expression is correlated with tumor formation in alcohol-fed mice; b) alcohol increases ERα activity to elevate Brf1 and Pol III gene expression; c) alcohol activates JNK1 to increase ERα and Brf1 expression; d) ethanol induces H3ph (histone H3 phosphorylation), e) H3ph elevates Brf1 expression. However, nothing is known about the role of MSK1 in transcription of Brf1 and Pol III genes. Analysis of signal event has revealed that ethanol markedly stimulates phosphorylation of MSK1 at serine 376 and threonine 581 in MCF-7 cells. Blocking MSK1 singling decreases cellular level of Brf1 expression and Pol III gene transcription. Further analysis indicates that MSK1 mediates ERα expression. Inhibition of MSK signaling decreases alcohol-induced ERE-Luc reporter activity. Repression of ERα expression by its siRNA reduces alcohol-induced anchorage-independent growth of MCF-7 cells. Our results have demonstrated that repression of MSK1 by its siRNA decreases colony formation of MCF-7 cells. Further analysis of ChIP assay indicates that ethanol treatment increases occupancy of ERα to Brf1 promoter. These results support the idea that alcohol activates MSK1 and enhances ERα and Brf1 expression resulting greater phenotypic changes. Together, our studies indicate that ethanol-induced MSK1 activation may play a critical role in alcohol-induced cell transformation and alcohol-associated ER+ breast cancer.

*: The project is supported by NIH grants AA017288 and AA021114 to Shuping Zhong

Citation Format: Shuping Zhong, Qingsong Zhang, Ganggang Shi. Signaling event of alcohol-induced deregulation of Pol III genes in breast cancer cells. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 2446. doi:10.1158/1538-7445.AM2014-2446
Abstract 2457: Functional Study of DCC Gene in multiple myeloma mell lines
Authors: Rodrigues-Junior Dorival Mendes , Biassi Thais Priscilla , Carlin Viviane , Machado-Junior Joel , Vettore André Luiz
Published: May 2015

Functional Study of DCC Gene in Multiple Myeloma Cell Lines

ABSTRACT

INTRODUCTION: Multiple Myeloma (MM) is an incurable hematological malignancy characterized by multiorgan dysfunction as a result of bone marrow infiltration by malignant cells and the systemic damage of monoclonal circulating protein. Despite considerable progress in understanding MM biology, the molecular basis of this disease still remains open to debate. Previous data showed that DCC gene hypermethylation is significantly correlated with poor prognosis of MM patients. However, the involvement of DCC in the tumorigenesis process of MM remains unclear. PURPOSE: To better understand the role of DCC in this pathology through gene silencing in MM cell lines and evaluation of its interference in different tumorigenic processes. METHODS: The DCC gene expression was evaluated in three MM cell lines (RPMI-8226 and U266) and promoter methylation profile of this gene in the three cell lines was determined. The presence of DCC protein was confirmed by Western blotting (WB). The DCC expression was silenced by esiRNA and transfections were performed by electroporation. RESULTS: We detected the presence of aberrant methylation in DCC promoter in RPMI-8226, a cell line with undetectable expression of DCC. On the other hand, DCC was not methylated in U266, which present expression of DCC at mRNA and protein level. For this reason, DCC was silenced in U266 with esiRNA and its impact in the apoptosis, cell viability, proliferation and cell cycle processes was evaluated in this cell line.

Support by FAPESP

Citation Format: Dorival Mendes Rodrigues-Junior, Thais Priscilla Biassi, Viviane Carlin, Joel Machado-Junior, André Luiz Vettore. Functional Study of DCC Gene in multiple myeloma mell lines. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 2457. doi:10.1158/1538-7445.AM2014-2457
Abstract 246: Mammary cancer preventive activity of an extract of withania somnifera roots
Authors: Khazal kamel F. , Hill Donald , Grubbs Clinton J.
Published: May 2015

Mammary cancer preventive activity of an alcoholic extract of Withania somnifera (WS) roots was examined in female ErbB2+/- (MMTV/Neu) mice. The WS root extract was mixed with feed at a concentration of 750 mg/kg of diet. Mice treated with the root extract developed fewer mammary tumors relative to mice in the untreated group; after 310 days, mice in the treated group had an average of 1.62 tumors, and mice in the control group had 2.48, a reduction of 33% (p=0.1). Representative samples of the mammary tumors were examined histologically after staining with hematoxylin and eosin (H&E) and for proliferation (Ki67 and PCNA). For mammary tumors of control mice, sections stained with H&E showed high mitotic activity, whereas in tumors from treated mice, there was low mitotic activity. Labeling indices for Ki67 and PCNA markers in cancers of treated mice were 35% and 30% lower, respectively, than those of the control group. Expression of the cytokine CCL2 was 50% lower in cancers of the treated group. These results indicate that the root extract of WS reduces mammary tumor formation and tumor cell proliferation. Supported by grant U54 CA118948.

Citation Format: kamel F. Khazal, Donald Hill, Clinton J. Grubbs. Mammary cancer preventive activity of an extract of withania somnifera roots. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 246. doi:10.1158/1538-7445.AM2014-246
Abstract 2460: APC regulates COX-2 independent of Wnt signaling in breast cancer
Authors: VanKlompenberg Monica K. , Bedalov Claire , Prosperi Jenifer R.
Published: May 2015

Breast cancer is the leading cause of cancer-related death in non-smoking women, and the most commonly diagnosed cancer in women. The Adenomatous Polyposis Coli (APC) tumor suppressor gene is mutated or silenced by hypermethylation in 18-70% of sporadic breast cancers depending on subtype; however, the molecular mechanisms downstream of APC loss in the breast remain largely unexplored. While inactivation of APC most commonly leads to increased signaling through the Wnt/β-catenin pathway, there are multiple, less-investigated functions of APC, including regulation of GSK3β-mediated signaling, proliferation, epithelial polarity, cytoskeletal organization, and DNA repair. Given that restoration of tumor suppressors is a hurdle in treating tumors arising from loss of gene function, it is critical to investigate and understand the downstream signaling pathways involved in tumorigenesis. Our laboratory is particularly interested in investigating the effect of APC loss or mutation in breast cancer, and the complex Wnt-independent signaling downstream of APC loss in the breast. We previously demonstrated that Apc mutation accelerates the mouse mammary tumor virus-Polyoma middle T antigen (MMTV-PyMT) transgenic model of breast tumorigenesis independently of Wnt/β-catenin signaling. Interestingly, focal adhesion kinase (FAK)/Src/JNK signaling was up-regulated and required for the enhanced proliferation and squamous phenotype mediated by Apc mutation. Despite the lack of Wnt/β-catenin pathway activation, Apc-mutant mammary tumor cell lines demonstrated enhanced expression of Cyclooxygenase-2 (COX-2) and production of prostaglandin E2 (PGE2), known to be important in triple-negative and inflammatory breast cancers, two aggressive breast cancer subtypes. While treatment with the Src inhibitor (PP2) decreases COX-2 protein expression, there is no effect on the level of phosphorylated JNK, indicating a non-linear signaling pathway. Consistent with this, treatment with an inhibitor of JNK (SP600125) has no effect on COX-2 expression. These data suggest that while Src activation downstream of Apc mutation enhances COX-2 in a Wnt-independent manner, the pathway does not travel through activation of JNK. Therefore, the mechanisms by which Apc mutation initiates this stream of events to enhance mammary tumorigenesis, and the contribution of COX-2 to the phenotypes acquired in Apc-mutant breast cancer remain unclear.

Citation Format: Monica K. VanKlompenberg, Claire Bedalov, Jenifer R. Prosperi. APC regulates COX-2 independent of Wnt signaling in breast cancer. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 2460. doi:10.1158/1538-7445.AM2014-2460