bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 745: Development of gamma-H2AX assay to assess toxicity and efficacy from chemotherapy in circulating tumor cells (CTCs) and peripheral blood lymphocytes
Authors: Sheikh Hassan S. , Matthew Elizabeth , El-Deiry Wafik S.
Published: August 2012

CTC methodology has been recently adopted following FDA approval of the Veridex CellSearch for the detection of CTCs and offers prognostic value in various solid tumors including breast, colorectal, and prostate cancer. gamma-H2AX is an established marker for double strand breaks from DNA damage, and this is particularly evident and has been seen after exposure to ionizing radiation and exposure to topoisomerase inhibitors. Moreover prolonged gamma-H2AX expression has been observed in repair deficiencies after DNA damage where there may be delays in DNA repair and persistence of H2AX. H2AX may therefore serve as an excellent marker not only of toxicity but the efficacy of the chemotherapy. We have been using human tumor cell lines to expose them to chemotherapeutic agents in the laboratory to detect gamma-H2AX levels and the dynamic changes thereof following exposure to cytotoxic chemotherapy. Our results reveal clear time-dependent upregulation of gamma-H2AX in treated cells. This is being adopted for use on CTCs as well as blood lymphocytes with a goal of making correlations with toxicity to chemotherapy in treated patients as well as use of this assay to mark the efficacy of chemotherapy in CTCs. We are applying quantum dot technology to detection of gamma-H2AX in cells recovered from blood and this is being incorporated into a clinical protocol. This assay has potential to impact on monitoring of chemotherapy with regard to both toxicity and efficacy in the clinic.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 745. doi:1538-7445.AM2012-745
Abstract 762: Therapeutic drug monitoring of Anthracyclines and Taxanes in chemotherapy of breast cancer patients
Authors: Valenzuela Belén , Escudero Vanesa , Rebollo Joseba , Sureda Manuel , Brugarolas Antonio
Published: August 2012

Background: Anthracyclines (A) and Taxanes (T) remain amongst the most active drugs in breast cancer chemotherapy. However, significant toxicities are associated when administered at full doses in a number of patients (pts), so careful dose adjustments should be made in order to optimize their clinical effectiveness. A and T have high inter- and intra-patient variability, and a correlation between exposure and efficacy and toxicity has been demonstrated. Therefore, therapeutic drug monitoring (TDM) of these drugs, based on the individual pharmacokinetic (PK) parameters could personalize doses in order to diminish its toxicity and/or to improve the clinical benefit. Methods: A total of 24 pts (92 cycles) were treated with either single drugs, or combinations of two of these agents, usually with four initial cycles of A and 4 subsequent cycles of T. The initial dose was calculated using body-surface algorithm. Pts were monitored since the first cycle and subsequent doses pharmacokinetically adjusted until the target plasma concentration was reached. Bayesian estimates of individual PK parameters were calculated using a population PK models previously published and used for PK-based dose adjustment. Results: The main results are shown in the following table: 43% of pts treated with A had the initial dose increased by an average of 22% without toxicities grade II, except one pt which developed mucositis grade II. For paclitaxel, initial dose was increased in 25% of patients by an average of 22% with only one case of grade 2 asthenia. However, docetaxel initial dose of 60-75 mg/m2 was more toxic than the other drugs evaluated. The doses were reduced by an average of 22% due to toxicities such as grade 3 neutropenia, grade 2 asthenia or grade 2 leucopenia. Conclusion: Standard dosing regimen of A and T in breast cancer pts could be optimized by TDM in order to achieve the target plasma concentration during the chemotherapy treatment and ensure that pts receive the right dose.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 762. doi:1538-7445.AM2012-762
Abstract 852: ARRY-380, a potent, small molecule inhibitor of ErbB2, increases survival in intracranial ErbB2+ xenograft models in mice
Authors: Dinkel Victoria , Anderson Deborah , Winski Shannon , Winkler James , Koch Kevin , Lee Patrice A.
Published: August 2012

ARRY-380 is an orally active, potent small molecule targeting ErbB2 inhibitor currently in clinical development in patients with ErbB2+ metastatic breast cancer (MBC). This compound has shown excellent activity in numerous SC mouse tumor models including breast (BT-474, MDA-MB-453), ovarian (SK-OV-3) and gastric (N87) carcinoma models. In breast cancer patients, brain metastases are a serious unmet medical need. Patients with ErbB2+ breast cancer have a significantly increased incidence of brain metastases following trastuzumab therapy. Here we demonstrate significant single agent activity of ARRY-380 in two ErbB2+ intracranial mouse xenograft models. For these studies, female nude mice received intracranial implantations of tumor cells (either NCI-N87 or BT-474) by direct injection into the brain parenchyma (via the sagittal suture). In pilot studies, we demonstrated that the blood brain barrier was not disrupted by mechanical injections and that increasing tumor burden correlates negatively with neurologic outcome, body weight and survival. In the N87 studies, animals received treatments beginning on Day 2 post-implantation and continuing for up to 6 weeks. Dose groups (n=12/group) were vehicle, ARRY-380 at MTD (75 mg/kg, PO, BID) or lapatinib at MTD (50 mg/kg, PO, BID). All animals in the vehicle- or lapatinib-treated groups did not survive beyond Day 22. In the ARRY-380-treated-group, 75% of the animals were alive on Day 43. Brain PK/PD was also evaluated in the N87 model. ARRY-380 and its active metabolite caused a significant reduction in brain pErbB2 (80%). In the BT-474 model, animals received treatments beginning on Day 2 post-implantation and continuing for up to 8 weeks. Dose groups (n=13/group) were vehicle, ARRY-380 at MTD (75 mg/kg, PO, BID), lapatinib at MTD (50 mg/kg, PO, BID) or neratinib at MTD (40 mg/kg, PO, QD). On Day 56, survival in the ARRY-380 group was 69% while survival rates in the vehicle, lapatinib or neratinib-treated groups were 23%, 8% and 23%, respectively. Thus ARRY-380 treatment significantly enhances survival in two ErbB2 driven intracranial tumor xenograft models, with superior activity compared to other ErbB2 agents in these studies. Additionally, ARRY-380 has demonstrated durable clinical activity in heavily pre-treated patients with ErbB2+ MBC. These preclinical and clinical data suggest that ARRY-380 may provide benefit to patients with ErbB2+ MBC with brain metastases.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 852. doi:1538-7445.AM2012-852
Abstract 871: Dual inhibition of PI3K and Erk1/2 shows synergy and efficacy in human tumor cells, either by using drug combinations or novel dual PI3K/Erk inhibitors
Authors: Seipelt Irene , Gerlach Matthias , Blumenstein Lars , Mueller Gilbert , Guenther Eckhard , Engel Jürgen , Teifel Michael
Published: August 2012

The Ras/Raf/Mek/Erk and the PI3K/Akt signaling pathways are frequently deregulated in cancer due to activation of upstream receptors, decreased expression of tumor suppressors like PTEN or activating mutations of Ras, B-Raf or PI3K. Several clinical studies with kinase inhibitors targeting single members of the Ras/Raf/Mek/Erk or the PI3K/Akt pathway are ongoing. However, preclinical and clinical trials indicated limited success by single pathway inhibition and different resistance mechanisms were defined. Further clinical trials with drug combinations of Ras/Raf/Mek/Erk and PI3K/Akt suppressors have been initiated, suggesting a more favorable outcome than targeting only one single pathway, Ras/Raf/Mek/Erk or PI3K/Akt. We performed in-vitro combination experiments with Raf, Mek or Erk inhibitors and PI3K or Akt inhibitors in several human tumor cell lines. Strong synergy was achieved with various combinations including the Mek inhibitor CI-1040 and the PI3K inhibitor GDC-0941 (CI = 0.17) or the Erk inhibitor AEZS-131 and the PI3K inhibitor D-117073 (CI = 0.23) in A549 cells. Also Raf inhibitors like Sorafenib and Zelboraf were combined with different PI3K inhibitors, resulting in synergistic anti-proliferative activity. Due to the attractiveness of parallel inhibition of the Ras/Raf/Mek/Erk and PI3K/Akt pathways, we developed AEZS-136 that concurrently inhibits Erk1/2 (IC50 ∼ 50nM) and PI3K (IC50 ∼ 100nM) by an ATP competitive mode of action. Derivatives of the dual PI3K/Erk inhibitor were co-crystallized with Erk2 and PI3Kα enabling an optimization process by SAR driven medicinal chemistry. The anti-proliferative efficacy of AEZS-136 was evaluated in more than 40 human tumor cell lines and physico-chemical as well as in-vitro ADMET properties were widely assessed. Furthermore, the in-vivo pharmacokinetics and anti-tumor efficacy was explored. AEZS-136 was well tolerated and showed dose dependent inhibition of human colon tumor growth of up to 74% in a Hct116 mouse model. Here we present the concept of dual targeting of the Ras/Raf/Mek/Erk and the PI3K/Akt pathways, either by using drug combinations or our novel dual PI3K/Erk inhibitor. AEZS-136 is a small molecule in preclinical development showing a uniquely advantageous kinase inhibition profile. Broad clinical anti-tumor activity is expected for AEZS-136 in tumors with deregulated Ras/Raf/Mek/Erk and PI3K-Akt signaling.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 871. doi:1538-7445.AM2012-871
Abstract 93: Defining the roles of COIL and WIPI1 in breast cancer metastasis
Authors: Lee Misan , Cheung Grace , Nair Ranju , Done Susan
Published: August 2012

Metastasis of cancerous cells to distant sites is the major cause of death from breast cancer. In a previous study, we performed aCGH on breast invasive duct carcinoma samples with and without lymph node metastases and identified regions of genomic amplification that are associated with lymph node metastasis. These were subsequently validated by q-PCR. Within the 17q22-24 region, we have identified four genes likely to contribute to metastasis based on published data: COIL, WIPI1, TMEM100, and SLC16A6. We are using RNAi and ectopic over-expression to characterize these genes with in vitro functional assays for proliferation, migration, invasion, and survival. We used qRT-PCR analysis on a panel of seven breast cancer cell lines of varying invasiveness: MDA-MB-157, MDA-MB-231, MDA-MB-468, BT-549, CAMA-1, HS578T, MCF-7, and SKBR-3. We identified that WIPI1 and COIL were expressed in the majority. WIPI1 is up-regulated in a variety of tumor cells; studies suggest WIPI1 can activate autophagy by suppressing TORC1. Autophagy can inhibit cancer by maintaining genome stability via disposing of damaged mitochondria and peroxisomes. In clinical data obtained from 298 breast cancer patients, we observed high expression of WIPI1 to be associated with higher survival and lower breast cancer relapse. Alternatively, autophagy can also provide a survival mechanism for tumors with restricted blood supply and resistance to chemotherapy. This is in agreement with our observation of high WIPI1 protein expression in highly invasive breast cancer cell lines. We are over-expressing WIPI1 in SKBR-3 and knocking it down in BT-549 in vitro. WIPI1 could help determine the role of autophagy in tumor initiation processes such as proliferation and invasion, and in maintenance of tumor cells following therapy. COIL is found in a diffuse nucleoplasmic pool and in Cajal bodies (CBs). The role of diffuse COIL is still unclear, but CBs, involved in the formation of macromolecular complexes and found more in cells with high transcriptional activity, require COIL for proper formation and localization. We observed high expression of COIL protein in cell lines with higher proliferation index, noticeably in ER positive cell lines such as CAMA1 and absence in a non-tumorogenic cell line MCF10A. We are investigating the association of COIL with proliferation, invasion and migration abilities by over-expression and knockdown of the COIL protein in MDA231 and MCF7 breast cancer cells. We found from clinical data obtained from 245 breast cancer patients that high expression of COIL resulted in lower survival and a higher risk of relapse. This suggests that COIL may be a potential therapeutic target.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 93. doi:1538-7445.AM2012-93
Abstract LB-375: Inactivation of the deubiquitinase USP44 is an early event in the formation of colorectal cancer
Authors: Sloane Mathew , Hesson Luke , Wong Jason , Pimanda John , Bourke Michael , Hawkins Nick , Ward Robyn
Published: August 2012

Colorectal cancer (CRC) develops from intraepithelial neoplasms known as adenomas. Genomic profiling of these premalignant lesions presents an opportunity to discover molecular changes that can lead to colorectal cancer. Some of the earliest changes are thought to be epigenetic, including hypermethylation of the promoters of key tumour suppressor genes. To identify such changes in colon cancer we performed whole genome DNA methylation profiling (MBD-Seq) on three adenomas and matched normal mucosa. We then validated epigenetic changes in a cohort of 105 adenomas using COBRA, bisulphite sequencing and quantitative RT-PCR. In each adenoma, we identified 1598, 556 and 331 promoter CpG islands respectively that were hypermethylated when compared with matched normal mucosa. Two of the three adenomas showed CpG island hypermethylation of the USP44 gene, a deubiquitinase previously shown to affect mitotic progression by deubiquitinating Cdc20, a regulator of the Anaphase Promoting Complex. In the absence of USP44, cells prematurely enter anaphase resulting in accelerated mitotic progression. Using COBRA and bisulphite sequencing we show that hypermethylation of USP44 occurred in 100% of 18 CRC cell lines, 79% of 105 adenomas, but in only 10% of 45 matched normal mucosa. In adenomas, USP44 hypermethylation correlated with a >2-fold decrease in gene expression. Reduced expression has also been reported in glioblastoma, B-ALL, breast, colorectal, esophageal, kidney and testicular cancer. We show that treatment of four CRC cell lines with the DNA demethylating agent 5-Aza-2′-deoxycytidine (5azaDC) resulted in a 10-250 fold increase in USP44 expression. In summary, we show that epigenetic inactivation of the deubiquitinase USP44 is a frequent and early event in the formation of colon cancer. Taken together with reports that USP44 expression is reduced in a range of solid organ and haematological malignancies, we propose that epigenetic inactivation of this gene may represent an important yet hitherto unrecognized event in tumour formation.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr LB-375. doi:1538-7445.AM2012-LB-375
Abstract LB-385: The role of tumor suppressing microRNA let-7 in undifferentiated nasopharyngeal carcinoma
Authors: Wong TS , Gao W , Li JZH , Chan JYW
Published: August 2012

AIMS:This study aimed at evaluating the potential anti-proliferative effects of the microRNA let-7 family in nasopharyngeal carcinoma (NPC) cells. In addition, the association between let-7 suppression and DNA hypermethylation is examined. MATERIALS AND METHODS:Levels of mature let-7 family members (-a, -b, -d, -e, -g, and -i) in normal nasopharyngeal cells (NP69 and NP460) and nasopharyngeal carcinoma cells (HK1 and HONE1) were measured by real-time quantitative PCR. Cell-proliferation assay and c-Myc immunohistochemical staining were performed on NPC cells transfected with let-7 precursor molecules. In addition, expression changes in let-7 family members in response to demethylating agents (5-azacytidine and zebularine) were also examined. RESULTS:In comparison with the normal nasopharyngeal cells, let-7 (-a, -b, -d, -e, -g, and -i) levels were reduced in nasopharyngeal carcinoma cells. Ectopic expression of the let-7 family in nasopharyngeal carcinoma cells resulted in inhibition of cell proliferation through downregulation of c-Myc expression. Demethylation treatment of nasopharyngeal carcinoma cells caused activation of let-7 expression in poorly differentiated nasopharyngeal carcinoma cells only. CONCLUSION:Our results suggested that miRNA let-7 might play a role in the proliferation of NPC. DNA methylation is a potential regulatory pathway, which is affected when let-7 is suppressed in NPC cells. However, the extent of DNA hypermethylation/hypomethylation in regulating let-7 expression requires further elucidation.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr LB-385. doi:1538-7445.AM2012-LB-385
Abstract LB-392: Loss of Rev7 causes PARP inhibitor resistance in BRCA1;p53-deficient mouse mammary tumor cells
Authors: Xu Guotai , Bouwman Peter , Jaspers Janneke , Pieterse Mark , Kersbergen Ariena , Jonkers Jos , Borst Piet , Rottenberg Sven
Published: August 2012

We have previously shown that mammary tumors spontaneously arising in a genetically engineered mouse model for human BRCA1-associated breast cancer (K14Cre;Brca1F/F;p53F/F) are highly sensitive to the poly(ADP-ribose) polymerase (PARP) inhibitor olaparib due to their deficiency in homology-directed DNA repair (Rottenberg et al. PNAS 2008;105:17079-84). Despite the initial sensitivity, tumors are not eradicated and eventually develop drug resistance. In this project we aim to identify mechanisms that explain this resistance. In particular we are interested in alterations that result in restoration of DNA repair by homologous recombination (HR). For this purpose we performed a loss-of-function shRNA screen using Brca1; p53-deficient cell lines derived from a mouse mammary tumor. The CGH profile of these cell lines is highly similar to that of the original tumor. Moreover, these cells have successfully been transplanted orthotopically into syngeneic mice and they recapitulate the morphology and olaparib response of the donor tumor. In our screen we included 2000 hairpins from the Sigma Mission library which target 409 mouse genes involved in the DNA damage response. As a result we found enrichments for hairpins targeting Rev7 in olaparib-surviving colonies. This finding was validated with 3 individual hairpins which caused substantially decreased Rev7 mRNA levels. Moreover, Rev7-deficient Brca1−/−;p53−/− tumor cells are less sensitive to doxorubicin, carboplatin and cisplatin. Loss of Rev7 rescued mES cells from Brca1 deficiency and also correlated with the formation of Rad51 foci in both Brca1-deficient mES cells and Brca1−/−;p53−/− tumor cells after irradiation. We therefore think that homology-directed DNA repair is restored in Rev7-depleted Brca1−/−;p53−/− cells. We are currently confirming this finding using HR reporter assays and we are investigating whether the loss of Rev7 causes olaparib resistance in mouse tumors. Our data suggest that absence of REV7 function, like lack of 53BP1, contributes to PARP inhibitor resistance in vivo.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr LB-392. doi:1538-7445.AM2012-LB-392
Abstract LB-394: Type II topoisomerases contribute to nitric oxide-induced DNA breakage during cancer-related inflammation
Authors: Li Tsai-Kun , Yang Yu-Chen , Shen Tang-Long
Published: August 2012

Activated inflammatory leukocytes generate a variety of reactive nitrogen and species (RNOS) which have been proposed to contribute to genome instability and cancer development. We previously reported that oxidative stress induces topoisomerase II (TOP2)-mediated DNA breaks via TOP2 cleavable complex (TOP2cc) formation. Moreover, drug-induced TOP2αcc and TOP2βcc are suggested to be anti-cancer and carcinogenic, respectively. Here we show that either inflammation-produced or chemical-released nitric oxide (NO), like TOP2-targeting etoposide (VP-16), induce TOP2cc-meidated DNA breakage, mutagenesis and carcinogenesis: (i) NO-releaser S-nitrosoglutathione (GSNO) induces formation of skin melanomas in 9,10-dimethyl-1,2-benzanthracene (DMBA)-initiated mice; (ii) the co-culture experiments revealed that inflamed macrophages causes activation of DNA damage response and chromosome DNA breakage in target cells (iii) NO production and two TOP2 isozymes are responsible for the generation of chromosome DNA breaks; (iv) consistently, treatment of a NO releaser GSNO induces hTOP2αcc and hTOP2βcc formation; (v) GSNO also increases mutation rate and kill cells in TOP2-dependent manner; and (vi) the incidences of GSNO- and VP-16-induced melanomas are found to be lower in skin-specific top2β-knockout mice. Thus, our results provide the first demonstration for the functional role of TOP2 in the NO-caused DNA damage, mutagenesis and carcinogenesis. The TOP2 involvement in pathological settings such as chronic inflammation-associated mutagenesis and subsequent cancer development were discussed.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr LB-394. doi:1538-7445.AM2012-LB-394
Abstract LB-404: Genomic analysis reveals roles for chromatin modification and axon guidance in pancreatic cancer
Authors: Biankin Andrew V. , McPherson John D. , Gibbs Richard A. , Grimmond Sean M. ,   Australian Pancreatic Cancer Genome Initiative.
Published: August 2012

Pancreatic cancer (PC) is the fourth leading cause of cancer death with an overall 5-year survival rate of < 5%, a statistic that has not changed in almost 50 years. A deeper understanding of the underlying molecular pathophysiology is expected to advance the urgent need to develop effective therapeutic and early detection strategies for this disease. Genomic characterisation of Pancreatic Ductal Adenocarcinoma (PDAC), which accounts for over 90% of PC has to date relied on targeted PCR based exome sequencing of primary and metastatic lesions propagated as xenografts or cell lines. Here we use exome sequencing and copy number analysis to define genomic aberrations in a prospectively accrued clinical cohort (n = 142) of early (Stage I and II) PDAC. Detailed analysis of 99 informative tumours identified 1984 non-silent mutations and 1628 significant CNV events, and defined 439 significantly mutated genes based on stringent Significant Mutated Gene and GISTIC analysis. Integration with functional data from in vitro shRNA and in vivo Sleeping Beauty-mediated somatic mutagenesis screens provided supportive evidence for 182 of these as candidate driver mutations. Pathway based analysis recapitulated clustering of mutations in core signalling pathways in PDAC, and identified multiple new components in each, particularly in DNA damage repair mechanisms (ATM, TOP2A, RPA1, BRCC3). We also identified frequent and diverse somatic aberrations in genes involved in chromatin modification (SWI/SNF complex members, SETD2, EPC1), and novel mechanisms such as axon guidance (Semaphorin, Slit, Netrin and Ephrin signalling), extending the number of core perturbed pathways in PDAC. Aberrant expression of axon guidance genes co-segregated with poor patient survival, and in animal models was associated with disease development and progression, further implicating perturbation of the axon guidance pathway as a novel mechanism important in PDAC.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr LB-404. doi:1538-7445.AM2012-LB-404
Abstract LB-75: Glucocorticoid receptor activation inhibits p53-induced apoptosis of MCF10A myc cells via induction of protein kinase C epsilon
Authors: Aziz Moammir H. , Shen Hong , Maki Carl G.
Published: August 2012

Glucocorticoid receptor (GR) is a ligand dependent transcription factor that can promote apoptosis or survival in a cell specific manner. Activated GR has been reported to inhibit apoptosis in mammary epithelial cells (MECs) and breast cancer cells by increasing expression of pro-survival genes. In the current report, activated GR inhibited p53-dependent apoptosis in MCF10Amyc cells, immortalized MECs that are p53 wild-type and overexpress the MYC oncogene. Specifically, GR agonists hydrocortisone or dexamethasone (Dex) inhibited p53- dependent apoptosis induced in these cells by Cisplatin (Cis), ionizing radiation (IR), or the MDM2 antagonist Nutlin-3. In contrast, the GR antagonist RU486 sensitized the cells to apoptosis by these agents. Apoptosis inhibition was associated with maintenance of mitochondria membrane potential, diminished caspase 3 and 7 activation, and increased expression at both the mRNA and protein level of the antiapoptotic PKC family member, PKCepsilon (PKCε). Knockdown of PKCε via siRNA targeting reversed the protective effect of Dex and restored sensitivity to Cis, IR, and Nutlin-3 (Nutlin)-induced apoptosis. These data provide strong evidence that activated GR can inhibit p53-dependent apoptosis through induction of the anti-apoptotic factor PKCε.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr LB-75. doi:1538-7445.AM2012-LB-75
Abstract LB-77: Crm1-mediated nucleocytoplasmic transport inhibition perturbs multiple cellular pathways to trigger melanoma cell apoptosis
Authors: Pathria Gaurav , Wagner Christine , Wagner Stephan
Published: August 2012

Development of multiple resistance mechanisms in melanomas has necessitated the identification of novel drug targets, which when inhibited could impact multiple cellular pathways thus circumventing potential resistance mechanisms. By performing cDNA-microarray analysis, we identified four key components of the nucleocytoplasmic transport machinery, Chromosome Region Maintenance 1 (Crm1), Ran-GTPase, Ran-GTPase Activating Protein 1 (RanGAP1) and Ran Binding Protein 1 (RanBP1) to be overexpressed in human melanoma metastases. Crm1 inhibition, employing the highly specific Crm1 inhibitor Leptomycin B (LMB) induced a dramatic depletion of pro-survival/cytoplasmic Erk1/2 and p90RSK1 and mediated persistent Erk signaling hyperactivation. Consistently, Crm1 inhibition inflicted extensive apoptosis in melanoma cells while sparing immortalized melanocytes and primary lung fibroblasts. Apoptosis required both the intrinsic and extrinsic apoptotic pathways and was associated with a nuclear entrapment and down-regulation of anti-apoptotic Survivin. Apoptosis was preceded by a G1 cell-cycle arrest and even though Crm1 inhibition mediated dramatic p53 and p21 induction exclusively in wild-type p53 melanoma cells, latter's silencing or inactivation failed to alleviate ensuing apoptosis. Notably, Crm1 inhibition elicited cell line specific, G1 to S progression-retarding changes in the expression of multiple cell-cycle regulatory proteins (Cdk2, Cdk4, p27, p16 and p18) thus potentially explaining p53 dispensability. Based on our results, we propose Crm1 as a potential therapeutic target in human melanoma, whose inhibition initiates a multitude of cell context-dependent molecular events to trigger G1-arrest followed by massive apoptosis.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr LB-77. doi:1538-7445.AM2012-LB-77
Abstract LB-81: Autophagic degradation of BCR/ABL by arsenic trioxide and the role of cysteine cathepsins
Authors: Goussetis Dennis , Gounaris Elias , Wu Edward J. , Vakana Eliza , Sharma Bhumika , Altman Jessica K. , Platanais Leonidas C. , Bogyo Matthew
Published: August 2012

Autophagy is increasingly an area of high potential therapeutic interest in hematopoietic malignancies, but its precise involvement in the control of leukemic cell growth and survival remain to be defined. We provide evidence that arsenic trioxide (As2O3) targets the BCR-ABL oncoprotein for degradation via autophagy, involving the p62/SQSTM1 and the protease cathepsin B. Utilizing either florescence probes or gold-conjugate antibodies we show co-localization of BCR-ABL and p62/SQSTM1 in autolysosomes. Also, by drug inhibition of either autophagy or cathepsin B, or by molecularly targeting p62/SQSMT1, Atg7 or cathepsin B, we documented reversal of the suppressive effect of As2O3 on BCR-ABL expressing cells, including primitive leukemic precursors from chronic myelogenous leukemia (CML) patients. Altogether, our data indicate that autophagy-induced degradation of BCR-ABL is critical for the generation of As2O3 antileukemic effects. These results raise the potential of targeting the autophagic machinery to enhance the antileukemic properties of arsenic trioxide on leukemia initiating stem cells.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr LB-81. doi:1538-7445.AM2012-LB-81
Abstract LB-82: Modulation of autophagic flux by CPTH6, a Gcn5/pCAF histone acetyltransferase inhibitor with antitumoral activity
Authors: Gabellini Chiara , Ragazzoni Ylenia , Desideri Marianna , Nescatelli Riccardo , Carradori Simone , Secci Daniela , Trisciuoglio Daniela , Bufalo Donatella Del
Published: August 2012

The thiazole-derivative 3-methylcyclopentilidene-[4-(4′-chlorophenyl)thiazol-2-yl]hydrazone (CPTH6) is a Gcn5 and pCAF histone acetyltransferases inhibitor that induces apoptosis and cell cycle arrest in human leukemia cells. The aim of this study was to evaluate whether CPTH6-induced apoptosis is associated with other cell death mechanisms, such as autophagy. Herein, we show that CPTH6 interferes with autophagic flux in several human tumor cell lines of different origin. CPTH6 treatment increases microtubule-associated protein 1 light chain 3-II (LC3-II) levels and induces the appearance of typical LC3-II- associated autophagosomal puncta in a time- and dose-dependent manner. Moreover, this compound decreases the expression of autophagy promoting proteins beclin1, Atg5 and Atg12. Strikingly, combined treatment of CPTH6 with bafilomycin A1, a proton ATPase inhibitor, demonstrates that CPTH6 reduces autophagosomes turnover, through an impairment of their degradation pathway, rather than enhancing autophagosomes formation. According to these results, CPTH6 treatment enhances p62/SQSTM1 protein levels in several tumor cell lines, indicating a blockage of autophagic cargo degradation. CPTH6 also reduces the phosphorylation of several components of transduction signalling pathways, such as Akt, 4E-BP1 and eIF4E, ERK1/2 and GSK-3α/β while it activates p38 MAPK pathway. In vivo CPTH6 exposure does not produce any adverse effects on health in mice as monitored by diet consumption, body-weight loss, postural and behavioral changes. Most importantly, CPTH6 exerts antitumoral effect on U937 human leukemia xenografts. These findings demonstrate that CPTH6 induces apoptosis and interferes with autophagic flux in human cancer cells, supporting further exploration of CPTH6 and related molecules as potential anticancer agents.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr LB-82. doi:1538-7445.AM2012-LB-82
Abstract LB-84: The role of survivin and survivin variants in the mitotic arrest-induced cell death
Authors: Cáceres Rodrigo E. , Andonegui Marco A. , Saavedra Alejandro Lopez , Herrera Luis A.
Published: August 2012

Eukaryotic cells rely on a mitotic checkpoint in order to preserve their proper ploidy. This mechanism stops mitosis progression before anaphase until each kinetochore is adequately bound to mitotic microtubules, and tension is generated which splits up chromatids towards opposite poles of the mitotic spindle. The resulting paused state of the cell cycle is known as mitotic arrest, and has been long known to induce cell death. However, the mechanisms driving mitotic arrest-mediated cell death have not yet been elucidated. It has been hypothesized that an anti-apoptotic/pro-apoptotic protein interplay might define the mitotic arrest outcome. We are interested in defining the role of survivin and its splicing variants, survivin Δex3 and survivin 2b, in the mitotic arrest-induced cell death. Survivin is a multifaceted member of the IAP family of proteins with functions in the chromosome passenger complex that has been shown to be over expressed in nearly every human tumor type. Noteworthy, Survivin Δex3 retains IAP activity but Survivin 2b has been shown to act as a pro-apoptotic factor. In this study, we defined the mitotic index (MI) of HCT116 cells in response to different concentrations of taxol and vinblastine. The results show that vinblastine induces a wide MI oscillation in contrast to taxol, which produces a homogeneous behavior in the cell population. This difference in population behavior might be explained by the different mechanisms of action of taxol and vinblastine. From the maximum taxol-induced MI point on, the percentage of sub-2n cells increased at the expense of mitotic cells, as assessed by flow cytometry. These results suggest that, in our model, cells die by exposure to taxol in a mitotic arrest-dependent manner. We also determined, by RT-PCR, that survivin variants are endogenously expressed in HCT116 cells. Furthermore, in immunoblot experiments, we found that survivin protein level decreases throughout taxol treatment in contrast to Bub1 levels, a member of the spindle assembly checkpoint. These results suggest that the decreased survivin level might be related to mitotic arrest-dependent cell death. The analysis of the intracellular localization of survivin variants throughout the mitotic arrest showed that survivin is localized in mitotic chromosomes throughout mitosis (before anaphase) independently of the time of treatment with taxol and vinblastine. Noteworthy, we also found that survivin Δex3 localized to the cytosol, and its level decreases throughout mitosis while survivin 2b protein was diffusely localized in the cytosol and mitotic chromosomes, and its level and localization remained unchanged.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr LB-84. doi:1538-7445.AM2012-LB-84
Abstract LB-85: Ectopic expression of eukaryotic translation elongation factor eEF1A2 is a hallmark for human lung cancer and is associated with poor prognosis
Authors: Liu Xiao-Yun , Zhang Wei , Girard Luc , Gao Boning , Larsen Jill , Wistuba Ignacio Ivan , Frenkel Eugene , Toombs Jason , Brekken Rolf , Minna John , Gazdar Adi
Published: August 2012

Background: The elongation factor eEF1A is one of the most abundantly expressed cellular proteins. It exists in two forms, eEF1A2, expressed in neurons and striated muscle, and eEF1A1, expressed in all other cell types. No normal adult cell expresses both forms. We investigated their role in lung cancer. Methods: We studied over 500 NSCLC and SCLC lung cancers and over 170 cell lines, as well as corresponding non- malignant lung tissues and immortalized respiratory epithelial lines. Methods included microarray, qPCR, immunostaining and western blots for eEF1A1 and eEF1A2, genomic sequencing and SNP analyses. Results: All normal and malignant cells highly expressed eEF1A1. EEF1A2 was expressed in 70-80% of all forms of lung cancers and 95% of cell lines, while all non-malignant tissues were negative. Expression of eEF1A2 in neuroendocrine tumors was particular high. The ectopic expression of eEF1A2 was not the consequence of mutations or copy number gains. Use of demethylating agents and HDAC inhibitors resulted in expression of eEF1A2 in respiratory epithelial cells, indicating that epigenetic mechanisms prevented expression in non-malignant cells. Transient knockdown of the eEF1A2 expression inhibited proliferation and colony formation of lung cancer cell lines. Furthermore, the expression of eEF1A2 in resected NSCLC cancers was associated with poor survival. Conclusions: Highly specific ectopic expression of eEF1A2 gene is frequent in all types of lung cancer, plays an important role in tumor pathogenesis and is a potential diagnostic and prognostic factor.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr LB-85. doi:1538-7445.AM2012-LB-85
Abstract SY02-02: Inhibition of methyltransferase EZH2
Authors: Dhanak Dashyant
Published: August 2012

Enhance of Zeste Homolog 2 (EZH2) is the biochemical activity of the Polycomb Repressive Complex 2 (PRC2) that transfers a methyl group from the cofactor S-adenosyl methionine (SAM) to histone H3 on lysine 27 (H3K27), resulting in epigenetic silencing of target gene expression. Activity of the multi-component PRC2 complex is crucial for normal development and differentiation. However, dysregulation of H3K27 methylation is implicated in tumorigenesis and occurs through multiple mechanisms. Elevated levels of EZH2 correlate with poor prognosis in a number of solid tumors including prostate, breast, kidney and lung. The increased EZH2 expression observed in many solid tumors has been linked to loss of one or more miRNAs including mir-101, mir-26A, and mir-214; aberrant E2F activity and chromosomal amplification. Inactivating mutations in UTX, an H3K27 demethylase which acts in opposition to EZH2, have been described in several tumor types including transitional cell bladder carcinoma, esophageal squamous cell carcinoma, renal cell carcinoma and multiple myeloma. Recently, somatic activating mutations in EZH2 have been identified in follicular lymphoma (FL) and GCB diffuse large B cell lymphoma (DLBCL). The frequency of the most prevalent mutation, Y641, is reported to be 7-12% in FL and as high as 22% in DLBCL. In view of the above, biochemical inhibitors of EZH2 may have therapeutic utility and herein we report on the identification and characterization of potent and selective small molecule EZH2 inhibitors with in vitro and in vivo antiproliferative activity in cancer cell lines and mouse xenografts. The studies provide a compelling rationale for the clinical evaluation of EZH2 inhibitors in EZH2 driven human cancers.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr SY02-02. doi:1538-7445.AM2012-SY02-02
Abstract SY05-03: microRNAs, EMT, and cancer stem cells
Authors: Brabletz Thomas
Published: August 2012

We have shown, that in particular tumor cells at the invasive front undergo an epithelial-mesenchymal transition (EMT) and aberrantly express EMT-associated transcriptional repressors, like ZEB1. The amount of such cells strongly correlates with metastasis formation and poor clinical outcome. Strikingly, metastases show again a differentiated phenotype, indicating a mesenchymal-epithelial re-transition (MET) and a support a regulatory role of the tumor environment for malignant tumor progression.

We described that the EMT-activator ZEB1 is a crucial promoter of metastasis and demonstrated that ZEB1 inhibits expression of cell polarity factors and the microRNA-200 family, whose members are strong inducers of epithelial differentiation. These results indicate that ZEB1 triggers a microRNA-mediated feedback-loop, which stabilizes EMT and promotes dissemination of cancer cells. Moreover we detected that in addition ZEB1 is necessary for the tumor initiating capacity of pancreatic and colorectal cancer cells. ZEB1 inhibits expression of miR-200c, miR-203 and miR-183, which cooperate to suppress expression of stem cell factors, as demonstrated for the polycomb repressor Bmi1.

We propose that ZEB1 links EMT-activation and stemness-maintenance by suppressing stemness-inhibiting microRNAs and thereby is a promoter of mobile, migrating cancer stem cells. Notably, these cells also acquired a drug-resistance phenotype. Thus, targeting the ZEB1 - miR-200 feedback loop might be a promising treatment option for fatal tumors, such as pancreatic cancer.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr SY05-03. doi:1538-7445.AM2012-SY05-03
Abstract SY07-03: The ubiquitin-based response to DNA double-strand breaks
Authors: Durocher Daniel
Published: August 2012

The chromatin-based response to DNA double-strand breaks (DSBs) is characterized by the accumulation of DNA repair factors and DNA damage signaling molecules within a large domain that surrounds the lesion. This response is largely dependent on ATM-dependent phosphorylation but also on chromatin ubiquitylation. Indeed, the RNF8 and RNF168 E3 ubiquitin ligases are necessary for the accumulation of proteins such as 53BP1, BRCA1 and RAD18 at sites of DNA damage. The critical importance of this pathway is manifested by the observation that biallelic mutations in the RNF168 gene results in an ataxia-telangiectasia-like syndrome referred to as RIDDLE. We are interested in understanding how this pathway is regulated and how ubiquitin-dependent protein interactions orchestrate the hierarchy of protein recruitment at sites of DNA lesions. I will present our current understanding on the regulation and organization of this signaling cascade and I will contend that DNA double-strand break signaling will be a useful system for deciphering the contribution of non-degradative ubiquitylation in cell signaling.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr SY07-03. doi:1538-7445.AM2012-SY07-03
Abstract SY08-02: Gene regulatory network models identify targets for mammary tumor normalization
Authors: Brock Amy
Published: August 2012

The majority of breast cancer therapeutics aim at slowing tumor expansion through selective killing of tumor cells. Given the enormous plasticity of cells (highlighted by recent advances in the iPS field), we have instead sought to induce a cell state transition from a mammary tumor cell phenotype to a normal mammary epithelial cell state. We have tackled the critical problem of identifying target genes for the normalization or differentiation of mammary tumor cells by employing a systems biology approach that integrates computational modeling with in vivo and in vitro studies. In vivo gene expression data of mammary tumor progression in the FVB C3(1) SV40Tag transgenic mouse model were collected at multiple time points as the disease progressed from normal gland to DCIS-like lesions to invasive carcinoma. Gene regulatory network models predicted novel transcription factor targets that appeared critical to the rise of the tumor phenotype. These putative gene targets were then assessed in a 3D spheroid assay. RNAi inhibition of two computationally predicted transcription factors reverted mammary tumor cells to a normalized mammary epithelial cell phenotype, with appropriate apical/basal polarization, lumen formation and reduced proliferative index. Validated targets were then tested in vivo. This integrated strategy has pinpointed transcription factors for RNAi therapeutic targeting in vivo but importantly, the method is not limited to breast cancer and may be useful for many different tumor types. In summary, our major advance has been the demonstration that a gene regulatory network reverse engineering approach can identify genes that are causatively involved in cancer progression and that represent viable therapeutic differentiation targets.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr SY08-02. doi:1538-7445.AM2012-SY08-02
Abstract SY41-01: Tumor-endothelial transdifferentiation
Authors: Verma Inder M. , Soda Yasushi , Friedmann-Morvinski Dinorah , Pastorino Sandra , Kesari Santosh
Published: August 2012

Glioblastoma (GBM) is the most malignant brain tumor and highly resistant to intensive combination therapies and anti-VEGF therapies. To assess the resistance mechanism to anti-VEGF therapy, we examined the vessels of GBMs in tumors that were induced by the transduction of p53+/− heterozygous mice with lentiviral vectors containing oncogenes and the marker GFP in the hippocampus of GFAP-Cre recombinase (Cre) mice. We were surprised to observe GFP+ vascular endothelial cells (ECs). Transplantation of mouse GBM cells revealed that the tumor-derived endothelial cells (TDECs) originated from tumor-initiating cells and did not result from cell fusion of ECs and tumor cells. An in vitro differentiation assay suggested that hypoxia is an important factor in the differentiation of tumor cells to ECs and is independent of VEGF. TDEC formation was not only resistant to an anti-VEGF receptor inhibitor in mouse GBMs but it led to an increase in their frequency. A xenograft model of human GBM spheres from clinical specimens and direct clinical samples from patients with GBM also showed the presence of TDECs. We suggest that the TDEC is an important player in the resistance to anti-VEGF therapy, and hence a potential target for GBM therapy.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr SY41-01. doi:1538-7445.AM2012-SY41-01