bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 5655: Infectivity selective oncolytic adenovirus for pancreatic cancer by redesigning the AB-loop via adenovirus library screening
Authors: Miura Yoshiaki , Davydova Julia , Yamamoto Masato
Published: August 2012

A number of cancers, including pancreatic cancer, are resistant to adenovirus (Ad) infection. Genetically coded viral capsid modification has achieved increased infectivity but not much target specificity. Direct selection of targeted vector from Ad library is a promising approach to overcome this issue. We have established a novel Ad vector generating system enabling a huge-size Ad library. We hypothesize that Ad with redesigned AB-loop isolated by the screening of the new library system would result in a new class of oncolytic adenovirus with selective infectivity for the therapy of pancreatic cancer. Ad AB-loop library was screened with mesothelin-expressing 293 cells (293-MSLN) to target mesothelin (MSLN) which is highly over-expressed in pancreatic cancer, and the clones converged to one dominant clone. The binding of Ad with selected clone was analyzed with 293, Panc-1, MiaPaCa-2, A549 and 293-MSLN. The binding was significantly higher in 293-MSLN than any other cells and corresponded to the order of MSLN-expression by FACS analysis. The anti-MSLN siRNA and antibody significantly reduced the binding to MSLN expressing cells. The binding ability of the virus with this motif in Panc-1 was higher than that of Ad with 5/3 modified fiber which is known to exhibit remarkably enhanced infectivity pancreatic cancer cells. The anti-tumor effect was analyzed in vivo using Panc-1 as a MSLN-positive and MiaPaCa-2 as a MSLN-negative subcutaneous tumors. The injection of the selected clone showed significant anti-tumor effect against Panc-1 tumors, and almost complete regression of some Panc-1 tumors was observed, while it didn't demonstrate anti-tumor effect against MiaPaCa-2 tumors. The hexon immunostaining and the intratumoral viral copy number also indicated that the selected clone replicated in MSLN-positive pancreatic cancer, but not in MSLN-negative cancer. In this study, we selected MSLN targeting Ad vector from the high throughput screening of Ad library by infectivity. The selected clone exhibited significant infectivity to MSLN-positive pancreatic cancer cells in vitro and in vivo. This new genetically modified oncolytic Ad with selective infectivity to pancreatic cancer may embody next generation targeted therapeutics for this devastating disease.

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 5655. doi:1538-7445.AM2012-5655
Abstract 5656: Preclinical evaluation of cytotoxic effect of photosensitive fluorescent protein in human cancer cells
Authors: Tazawa Hiroshi , Sasaki Tsuyoshi , Hashimoto Yuuri , Kikuchi Satoru , Kishimoto Hiroyuki , Mizuguchi Hiroyuki , Fujiwara Toshiyoshi
Published: August 2012

Photodynamic therapy (PDT) is an anticancer therapy to induce tumor-specific cytotoxicity by accumulation of photosensitizers (PSs) and following non-toxic light irradiation to tumor tissues. Non-toxic light irradiation induces the PS-mediated generation of reactive oxygen species (ROS) and subsequent cytotoxicity in tumor cells. For clinical application, some kinds of PSs, such as photofrin and laserphyrin, are intravenously injected before light irradiation in anticancer PDT. However, the lack of sufficient and continuous accumulation of PSs in tumor tissues makes difficult to eradicate tumor cells by PDT. Therefore, the development of novel PSs that accumulate to tumor tissues more efficiently and persistently is required to improve the antitumor effect of PDT. Recently, a bioengineered fluorescent protein KillerRed has been reported to generate ROS upon green light irradiation, suggesting a potential of KillerRed fluorescent protein as a novel PS. In this study, we investigated the antitumor effect of KillerRed fluorescent protein in human cancer cells in combination with PDT. Transient transfection with KillerRed expression vector induced the photosensitive cytotoxicity upon green light irradiation within a few hours in human cancer HeLa and OST cells. To further evaluate the cytotoxic effect of KillerRed fluorescent protein in human cancer cells, we obtained the stable transfectants expressing cytotoxic KillerRed or non-cytotoxic Katushka fluorescent protein from human lung cancer H1299 cells. Green light irradiation induced the cytotoxic effect within one hour in H1299-KillerRed cells, but not in H1299-Katushka or H1299 cells. H1299-KillerRed cells irradiated with green light showed significantly higher levels of ROS, which was confirmed by intracellular fluorescence level of 2′,7′-Dichlorodihydrofluorescein Diacetate, than H1299-Katushka or H1299 cells. Immunocytochemistry further revealed that green light-irradiated H1299-KillerRed cells showed the apoptotic cell death with the cleavage of caspase-3 24 hours after irradiation. These results suggest that KillerRed fluorescent protein is a potent novel PS inducing ROS generation and apoptotic cell death in tumor cells in anticancer PDT. The development of virus-mediated delivery system for tumor-specific induction of KillerRed fluorescent protein is under way.

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 5656. doi:1538-7445.AM2012-5656
Abstract 988: Oncogenic PI3K mutations lead to NF-κB-dependent cytokine expression following growth factor deprivation
Authors: Hutti Jessica , Pfefferle Adam , Perou Charles , Baldwin Albert
Published: August 2012

While growth factor-independent signaling and proliferation are well-established hallmarks of cancer, little is known regarding growth factor-independent changes in gene expression which occur downstream from oncogenes. The PI3K pathway is one of the most commonly misregulated signaling pathways in human cancers. Here, MCF10A cells expressing the two most common PI3K mutations, PIK3CA E545K and H1047R, were used to identify the repertoire of genes altered by oncogenic PI3K mutations following growth factor deprivation. This gene set most closely correlated with gene signatures from claudin-low and basal-like breast tumors, and categorical enrichment analyses suggested that NF-κB target genes were dramatically upregulated by these mutations. An IKKα inhibitor was used to identify the subset of PI3K-driven genes that is NF-κB dependent. Interestingly, virtually all of these NF-κB dependent genes were secreted proteins, suggesting a paracrine role for this gene set. Among these genes was IL-6, a cytokine frequently expressed in tumors which plays a critical role in generating a tumor-promoting microenvironment. Consistent with this, conditioned media from cells expressing the E545K or H1047R mutations led to increased STAT3 activation in recipient THP-1 monocytes or normal epithelial cells in a NF-κB and IL-6-dependent manner. Together, these data define a PI3K-driven, NF-κB-dependent gene expression profile which may play a critical role in promoting a microenvironment amenable to tumor progression.

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 988. doi:1538-7445.AM2012-988
Abstract 990: Nucleosome occupancy at unmethylated promoter CpG islands represents a novel mechanism of epigenetic gene silencing in cancer
Authors: Hesson Luke B. , Wong Jason W.H. , Sloane Mathew A. , Pimanda John E. , Bourke Michael J. , Hawkins Nicholas J. , Ward Robyn L.
Published: August 2012

Promoter CpG island (CGI) hypermethylation is an established mechanism of silencing gene expression in cancer. Although the silenced state is thought to be consolidated by the recruitment of nucleosomes across the promoter, this has been observed in only a limited number of genes. Here we performed genome-wide MBD-Seq, MNase-Seq and RNA-Seq to investigate the relationship between CGI methylation, nucleosome occupancy around the transcription start site (TSS) and gene expression in a premalignant colorectal adenoma and matched normal mucosa. Analysis of RNA-seq data identified 1987 silent genes that were not expressed in the normal or adenoma tissues. Of these 1987 genes, 1401 (70.51%) showed either methylation in both the normal and adenoma tissues or nucleosome occupancy across the TSS, with 524 (26.37%) of genes showing both, suggestive of a consolidated silenced state. Our RNA-Seq data also revealed that 2010 genes were downregulated >2-fold in the adenoma when compared to paired normal mucosa. Of these 2010 genes, only 67 (3.33%) genes showed aberrant CGI hypermethylation, and of these 67 genes, only six (8.96%) showed nucleosome occupancy across the TSS. Therefore, the presence of both CGI hypermethylation and nucleosome occupancy across the TSS of downregulated genes is significantly lower than at silent genes (p<0.0001). Interestingly, 272 of the remaining 1966 (13.84%) genes that were downregulated in the absence of aberrant CGI hypermethylation showed clear evidence of nucleosome occupancy across the TSS, including the important cancer-related genes CDH1 and CDKN2B. Only seven of these 272 genes (2.57%) have previously been described as frequent targets of hypermethylation in cancer, suggesting nucleosome recruitment represents an alternative mechanism of inactivation of these genes. In summary, we identified some of the earliest epigenetic events in colorectal neoplasia by profiling the transcriptome and epigenetic landscape of a premalignant colorectal adenoma. Our data show that for the majority of genes showing adenoma-specific downregulation and aberrant CGI hypermethylation, nucleosomes are not recruited to the promoter. Moreover, our data reveals a potential novel mechanism of epigenetic gene inactivation in the absence of promoter hypermethylation, through physical occlusion of transcription by nucleosome occupancy at the TSS.

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 990. doi:1538-7445.AM2012-990
Abstract 991: The DNA methylome of castration-resistant prostate cancer
Authors: Kleb Brittney N. , Estecio Marcos , Zhang Jiexin , Tzelepi Vasso , Chung Woonbok , Maity Sankar , Logothetis Christopher , Troncoso Patricia , Navone Nora , Jelinek Jaroslav , Liang Shoudan , Issa Jean-Pierre , Aparicio Ana
Published: August 2012

Lethal prostate cancer is predominantly a castrate-resistant disease. However, castrate-resistant prostate cancer (CRPC) is clinically heterogeneous and a classification that is predictive of outcome and response to therapies is lacking. Previous epigenetic profiling studies in CRPC have been limited to a small number of genes or sequences and therefore we explored the DNA methylome of CRPC and its capacity to classify the disease. We performed methylated CpG island amplification microarrays (MCAMs) using a panel of xenografts derived from 24 patients with CRPC tumors, 8 prostate tumors, 1 local recurrence, 5 liver metastases, 3 pleural fluid samples, 2 adrenal gland metastases and 1 each of the following: bone, subcutaneous tissue, retroperitoneal lymph node, ascitic fluid and brain. The xenograft morphology consisted of 15 adenocarcinomas, 11 small cell carcinomas, 3 large cell neuroendocrine carcinomas, 1 squamous cell carcinoma, 2 mixed morphologies and 2 unknown. The Lowess-normalized M values were used for analysis and the MCAM results were validated by pyrosequencing of 20 sequences associated with 12 genes with a sensitivity of 61.9%, a specificity of 84.3%, a positive predictive value of 73.6% and a negative predictive value of 75.7%. An unsupervised hierarchal clustering analysis using M values for probes located in CpG islands was performed and these results were compared with the clinicopathological features of the tissues. We observed that, different xenograft sublines derived from the same donor tumor clustered together suggesting that the DNA methylome is homogenous throughout a given tumor mass, despite the presence of histological heterogeneity, and that it is maintained through serial passages in mice. Unsupervised hierarchal clustering classified the xenografts into 3 groups, which showed trend towards distinct clinicopathological features including overall survival and androgen receptor (AR) status. In summary, we found that DNA methylation is a frequent event in CRPC and well preserved in xenograft tumors. Furthermore, the methylation profile of CRPC may represent a marker of clinicopathological subgroups with differential survival and AR status.

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 991. doi:1538-7445.AM2012-991
Abstract 993: A high-throughput peptide array screen of H3K9 methyl effectors identifies UHRF1 as an exception to the “phospho/methyl switch.”
Authors: Rothbart Scott B. , Fuchs Stephen M. , Nady Nataliya , Krajewski Krzysztof , Tempel Wolfram , Xue Sheng , Iberg Aimee N. , Barsyte-Lovejoy Dalia , Martinez Jorge Y. , Bedford Mark T. , Arrowsmith Cheryl H. , Strahl Brian D.
Published: August 2012

Increasing evidence suggests that histone post-translational modifications (PTMs) function in combination to regulate the diverse activities associated with chromatin. However, the mechanisms through which multiple histone PTMs influence the association of effector proteins with chromatin is poorly understood. To address this problem, we recently developed a histone peptide array platform containing singly- and multiply-modified histone peptides to examine the binding of chromatin-associated proteins. Using these arrays, we surveyed protein domains that bound methylated H3K9, and uncovered a number of PTM “patterns” that influenced the association with this mark. Interestingly, while most H3K9 methyl binders were ejected by neighboring H3S10 phosphorylation (a PTM enriched in G2/M-phase chromatin), the tandem Tudor domain (TTD) of UHRF1 was insensitive to this known “phospho/methyl switch”. In agreement, structural and biophysical studies confirmed the insensitivity of the UHRF1 TTD to H3S10 phosphorylation. Furthermore, we found that UHRF1 remains bound to chromatin throughout the cell cycle in contrast to HP1, which is ejected from chromatin during mitosis. These results suggest an important role for this ubiquitin ligase during mitosis. Significantly, we further find that the TTD of UHRF1 regulates global DNA methylation, thus definitively linking H3K9me3 binding by UHRF1 to the maintenance of DNA methylation. Taken together, our data suggest that the regulation of DNA methylation by UHRF1 extends beyond S-phase and requires H3K9me3 recognition during mitosis.

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 993. doi:1538-7445.AM2012-993
Abstract 997: Oncogenic tyrosine kinases are localized to mitochondria and regulate cancer metabolism by phosphorylating key components of pyruvate dehydrogenase kinase complex
Authors: Chen Jing , Hitosugi Taro , Fan Jun , Kang Sumin , Gu Ting-Lei , Boggon Titus
Published: August 2012

The Warburg effect describes the observation that cancer cells take up more glucose than normal tissue and favor aerobic glycolysis. Many tumor cells rely on aerobic glycolysis instead of mitochondrial oxidative phosphorylation for their continued proliferation and survival. This may be, in part, due to attenuated mitochondrial function, which is suggested to be achieved through inhibition of pyruvate dehydrogenase complex (PDC) in cancer cells. Pyruvate dehydrogenase (PDH) is the first and most important enzyme component of PDC, which catalyzes the conversion of pyruvate to acetyl-CoA that subsequently enters into the Krebs cycle to produce ATP and electron donors including NADH. PDH activity is negatively regulated by phosphorylation at several serine sites. Phosphorylation of PDH by PDH kinase (PDHK) results in the inactivation of PDC, while dephosphorylation by pyruvate dehydrogenase phosphatase (PDP) restores PDC activity. Thus, PDHK and PDP coordinate to regulate the phosphorylation and consequently activation status of PDH. In cancer cells, c-Myc and HIF-1alpha are believed to promote inhibition of PDC by, in part, upregulating gene expression of PDHK. However, how oncogenic signals regulate PDH, and/or coordinate PDHK and PDP activity to inhibit PDH and ultimately attenuate PDC activity in cancer metabolism still remains unclear. We recently reported that, surprisingly, functional PDC can form in mitochondria outside of matrix in some cancer cells. Moreover, several oncogenic tyrosine kinases including FGFR1, FGFR1 fusion tyrosine kinase, and FLT3-ITD mutant, as well as BCR-ABL and JAK2 V617F mutants, are localized to different mitochondrial compartments including outer membrane and matrix, respectively, in cancer cells, where they phosphorylate PDHK1, PDP1 and PDHA. Our studies demonstrate that tyrosine phosphorylation enhances PDHK1 kinase activity by promoting ATP and PDC binding, which promotes cancer cell metabolism, proliferation and tumor growth. In consonance with these findings, we found that tyrosine phosphorylation attenuates PDP1 phosphatase activity but inhibits PDHA enzyme activity. These data suggest that mitochondrial oncogenic tyrosine kinases regulate PDC activity by inhibiting PDHA1 enzyme activity directly through tyrosine phosphorylation and indirectly by enhancing and attenuating PDHK1 and PDP1 activity, respectively. These findings represent an acute mechanism underlying the Warburg effect, in addition to the long-term regulation that is believed to be regulated by c-Myc and HIF-1alpha.

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 997. doi:1538-7445.AM2012-997
Abstract 999: Potential role of reactive oxygen species in mediating glioblastoma stem cell differentiation
Authors: Wang Feng , Yuan Shuqiang , Chen Gan , Kim Sangbae , Hammoudi Naima , Feng Li , Lu Weiqin , Lee Ju-Seog , Huang Peng
Published: August 2012

Glioblastoma multiforme (GBM) is the most common and aggressive primary brain tumor with poor prognosis due in part to drug resistance and high incidence of tumor recurrence. The drug resistant and cancer recurrence phenotype may be ascribed to the presence of cancer stem cells, which seem to reside in special stem-cell niches in vivo and require special culture conditions including certain growth factors and serum-free medium to maintain their stemness in vitro. In this study, we used several stem-like glioblastoma cell lines (GSC11, GSC23 and GBM3752) derived from patients’ tissues by typical neurosphere culture system, and showed that addition of fetal bovine serum (FBS) to the medium induced an increase of ROS, leading to aberrant differentiation and decreases of stem cell markers such as CD133. We found that exposure of glioblastoma stem cells (GSCs) to serum induces their differentiation through activation of mitochondrial respiration, leading to an increase in superoxide (O2−) generation and a profound reactive oxygen species (ROS) stress response manifested by upregulation of oxidative stress response pathway. This increase in mitochondrial ROS leads to a down-regulation of molecules including SOX2, and Olig2, and Notch1 that are important for stem cell function and an upregulation of mitochondrial superoxide dismutase SOD2 that converts O2− to H2O2. Neutralization of ROS by antioxidant N-acetylcysteine in the serum-treated GSCs suppresses the increase of superoxide and largely rescues the expression of SOX2, Olig2, and Notch1, and prevents the serum-induced differentiation phenotype. Upregulation of ROS by chemical disruption of mitochondrial electron respiration chain induced further decrease of CD133. Our study suggests that mitochondrial function and cellular redox status may profoundly affect the fates of glioblastoma stem cells, and that modulation of ROS generation may be an effective way to impact CSCs, which may provide a new basis for developing novel therapeutic approaches.

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 999. doi:1538-7445.AM2012-999
Abstract LB-117: Phenotype and function of tumor-infiltrating macrophages and B cells in a novel humanized mouse model (NSG-HIS) for ovarian cancer
Authors: Nunez-Cruz Selene , Walter Marc , Danet-Desnoyers Gwenn , Scholler Nathalie
Published: August 2012

Anti-cancer treatments are often highly effective in human tumor xenografts grown subcutaneously in immune deficient hosts such as athymic (nude) or severe combined immune deficient (SCID) mice. However, such preclinical results are frequently followed by failure or limited activity of the drug/therapy in clinical trial, prompting the development of more sophisticated animal models engrafted with fully human immune system (HIS). We reconstituted NOD-Scid-IL2Rγcnull (NSG) mice with cord blood-derived CD34+ cells and 12 to 16 weeks after transplantation NSG-HIS mice were injected with human ovarian tumor cells (OVCAR5) orthotopically (io), subcutaneously (sc), or both (io/sc). Although sc tumor growth was the fastest in the absence of human immune system, four weeks after tumor implantation the average sizes of sc, io and sc/io tumors were similar across all NSG-HIS groups (0.3cm3); regardless of the tumor site, all tumors were infiltrated with human B cells, T cells and macrophages. However, sc tumors contained proportionally more human B cells and macrophages than io tumors. Macrophages infiltrating sc and io, but not sc or sc/io tumors, exhibited an IL10high immunosuppressive phenotype. Io tumor-infiltrating macrophages expressed the highest levels of CD32 (FcγRII) and mannose receptor (CD206), and the lowest level of CD64 (FcγRI) compared to macrophages infiltrating sc or io/sc tumors. Finally, io tumors were infiltrated with less human B cells than all the other tumors; B cells from tumor-bearing mice presented an altered phenotype compared to non tumor-bearing animals, characterized by the upregulation of plasma and activation markers and the production of IgM and IgG. Our results support the hypothesis that the location of the tumor implantation impacts the phenotype and function of human tumor cell infiltrate. The novel NSG-HIS mouse model of ovarian cancer may help understanding the role of tumor microenvironment during tumor development.

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-117. doi:1538-7445.AM2012-LB-117
Abstract LB-128: Targeted RNAi screening of an EGFR network reveals molecular determinants of tumor cell sensitivity to antibody-dependent cellular cytotoxicity (ADCC)
Authors: Murray Joseph C. , Nasto Rochelle E. , Jablonski Sandra A. , Tang Yong , Weiner Louis M.
Published: August 2012

Monoclonal antibodies can modulate signal transduction and survival of tumor cells, and can induce antibody-dependent cellular cytotoxicity (ADCC) through immune effectors, such as natural killer (NK) cells. Although clinical evidence suggests a role for ADCC in effective antibody therapy, molecular determinants of ADCC responsiveness—in oncogenic signaling networks, for example—have not been identified. Using a novel high-throughput screening platform, siRNAs targeting 60 genes of an EGFR network were reverse transfected into A431 cells, and cetuximab and an NK-like cell line (NK92-CD16V) were used to induce ADCC. Primary and validation screens identified four genes whose knockdown enhanced ADCC: GRB7, PRKCE, RET and ABL1. Pharmacological inhibition of Abl kinase activity enhanced ADCC, recapitulating the effects of ABL1 gene knockdown. Additional studies revealed distinct mechanisms associated with enhanced ADCC, including increased EGFR surface expression and enhanced susceptibility to apoptosis. This screening strategy has revealed putative molecular determinants of tumor cell sensitivity to ADCC that may translate to increased monoclonal antibody 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 LB-128. doi:1538-7445.AM2012-LB-128
Abstract LB-129: CD8+ T cell response mediates the therapeutic effects of oncolytic adenovirus in an immunocompetent mouse model
Authors: Yang Yajun , Li Xiaozhu , Wang Yaohe , Wang Shengdian
Published: August 2012

The role of anti-tumor immune responses in oncolytic adenoviral therapy has not been well studied due to lack of efficacious tumor model in immunocompetent mice. Here, we evaluated the contributions of immune components to the therapeutic effects of oncolytic adenoviruse in an immunocompetent murine tumor model permissive for infection and replication of adenovirus. We found that CD8+ T cells were critical mediator for antitumor efficacy by oncolytic adenovirus. Intratumoral viral therapy induced intensive infiltration of CD8+ T cells in tumor, increased tumor-specific IFN-γ (interferon-γ) production and CTL (cytotoxic T lymphocyte) activity of lymphocytes, and generated a long-term tumor-specific immune memory. Boosting CD8+ T cell responses by agonistic anti-4-1BB (cluster differentiation 137, CD137) antibody showed synergistic anticancer effects with oncolytic viro-therapy. Our results provide insight into antitumor mechanisms of oncolytic adenovirus in addition to their direct oncolytic effect.

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-129. doi:1538-7445.AM2012-LB-129
Abstract LB-133: Vaccination with dendritic cells pulsed with autologous oxidized whole tumor lysate induced strong and long-lasting anti-tumor immunity in recurrent ovarian cancer patients
Authors: Chiang Cheryl L-L , Kandalaft Lana , Tanyi Janos , Mantina Gina , Zsiros Emese , Powell Daniel J. , Coukos George
Published: August 2012

Background Despite surgical and chemotherapeutic advances, the death rate from ovarian cancer has not changed over the past two decades. We report here an enhanced, more rapid and more immunogenic dendritic cell vaccine platform developed for clinical testing. Furthermore we report the application of this novel immunotherapy platform comprising of dendritic cell (DC)-based autologous whole tumor antigen vaccination in a pilot study of patients with recurrent ovarian cancer. Methods To determine the optimal tumor lysate preparation for loading DCs, ovarian tumor lines (SKOV3, OVCAR5 and A1847) were prepared by HOCl-oxidation, UVB-irradiation or freeze and thaw. Normal donor DCs were evaluated for tumor lysate uptake, cytokine and chemokine productions and phenotype after maturation with lipopolysaccharides (LPS) and interferon (IFN)-gamma. The optimal lysate preparation, was used in a phase I study where five patients with recurrent, progressive stage III and IV ovarian cancer with available tumor lysate from secondary debulking surgery underwent intranodal vaccination with OC-DC, an autologous DC preparation pulsed with HOCl-oxidized autologous tumor cells (5-10 x 106 DC per dose, 5 doses). Feasibility, safety, and biological and clinical efficacy were evaluated. Results Normal donor DCs pulsed with HOCl-oxidized tumor lines demonstrated the highest tumor lysate uptake, matured efficiently after LPS and IFN-gamma stimulation, and produced higher levels of proinflammatory cytokines and chemokines including IL-1, IL-6, IL-8, IL-12, IP-10, MIP-1alpha and MIG. In vitro, these lysates loaded DCs primed T cell responses against ovarian tumor associated antigens and effectively expanded against tumor specific T cells from donors and patients. Therapy was feasible and well tolerated in all five subjects. Vaccination with OC-DC produced limited grade 1 toxicities and elicited tumor-specific T cell responses. Moreover specific HLA-A2-restricted responses to HER-2/neu, MUC1, NYESO-1, mesothelin and WT1 were documented following vaccination and HER-2/neu specific T cells were expanded following 10 days of in vitro culture. Patients exhibiting immune response demonstrated clinical benefit including two patients who demonstrated remission inversion on vaccine maintenance. Conclusions We developed a DC-HOCl oxidized whole tumor lysate vaccine which was safe and well-tolerated by all patients. The DC vaccine was highly proinflammatory and elicited cellular and humoral anti-tumor responses establishing a platform for immune-combinatorial 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 LB-133. doi:1538-7445.AM2012-LB-133
Abstract LB-140: Targeting brain tumour initiating cells using a dual-pronged approach of oncolytic virotherapy and chemotherapeutics
Authors: McKenzie Brienne , Zemp Franz J. , Lun Xueqing , Narendran Aru , McFadden Grant , Kurz Ebba , Forsyth Peter
Published: August 2012

BACKGROUND: Brain tumour initiating cells (BTICs) are stem-like cells hypothesized to mediate tumorigenesis and recurrence in glioblastoma multiforme (GBM). Patient-derived BTICs represent highly relevant preclinical GBM models, forming aggressive, infiltrative tumours in nude mice that closely recapitulate the phenotypic heterogeneity of patient GBMs. BTICs have also been shown to possess mechanisms mediating both chemo- and radio-resistance. Recently, success has been demonstrated in treating BTICs with oncolytic virotherapy (OV), which uses replication-competent viruses to specifically target and kill malignant cells. Myxoma virus (MyxV) is a promising oncolytic candidate, which our lab has shown to be highly efficacious in preclinical GBM models, effectively curing GBM xenografts with a single intra-cranial injection. By contrast, long-term survival is not obtained in MyxV-treated, BTIC-tumour-bearing mice, though survival is prolonged. HYPOTHESIS: We hypothesize that MyxV treatment of BTICs can be improved by utilizing clinically relevant chemotherapeutics identified via high-throughput pharmacoviral screens. RESULTS: We utilized a diverse, comprehensive panel of 80 small molecule inhibitors with preclinical and clinical anti-cancer efficacy to screen for synergy with MyxV treatment in vitro. Multiple candidates have been identified and are currently being validated for synergy (Chou-Talalay method) using a panel of genetically distinct, patient-derived BTICs. Our results implicate multiple potential targets for OV combination therapy, including topoisomerase I and the PI-3K/Akt/mTOR pathway. Target validation is currently underway using shRNA knockdowns and additional targeted inhibitors of these pathways. Further, the mechanism of synergistic cell death during these combination treatments is being explored. In vivo experiments utilizing the dual-pronged pharmacoviral approach are also underway. SIGNIFICANCE: We describe the nature and identity of compounds that carry the potential to sensitize BTICs to MyxV infection. Our findings offer an effective avenue to elucidate resistance mechanisms and develop efficacious combination therapies for targeting disease reservoirs within highly refractory GBMs.

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-140. doi:1538-7445.AM2012-LB-140
Abstract LB-144: A p53-dependent G2/M checkpoint governed by the cell-fate factor dachshund in non-small cell lung cancer
Authors: Chen Ke , Wu Kongming , Zhang Wei , Zhou Jie , Stanek Timothy , Li Zhiping , Wang Chenguang , Shirley L. Andrew , Rui Hallgeir , Mcmahon Steven , Pestell Richard G.
Published: August 2012

Lung cancer is the leading cause of cancer mortality worldwide with more than 170,000 cases diagnosed each year in the United States. Genetic studies have identified several oncogenes activated in lung cancer including K-Ras and EGFR. Tumor suppressor inactivation occurs including p53 in which LOH is observed in - 60% and mutations arise in 50%. Initially cloned as a dominant inhibitor of the hyperactive EGFR, Ellipse in Drosophila, the mammalian DACH1 gene regulates gene expression of target genes in part through interacting with DNA-binding transcription factors and in part through intrinsic DNA-sequence specific binding properties. Clinical studies have demonstrated a reduced expression of the DACH1 in breast, prostate and endometrial cancer. Given the importance of the EGFR in human lung cancer, we examined the role of DACH1 in lung cancer cellular growth, migration and DNA damage response. Herein, endogenous DACH1 was reduced in human non-small cell lung cancer (NSCLS). Expression levels of DACH1 correlated inversely with clinical stage and pathological grade. Re-expression of DACH1 reduced lung cancer cell colony formation and cellular migration. Cell cycle analyses demonstrated G2/M block by ectopic expression of DACH1 occurs synergistically with p53. DACH1 band p53, requiring p53 R248 and R273, residues commonly mutated in lung cancer. Fluorescent microscopy demonstrated co-localization of DACH1 with p53. Immunoprecipitation and western blot assay showed DACH1 association with p53 required the C-terminus of DACH1. DACH1 enhanced p53 transcriptional induction of p21 promoter, p53 and DACH1 repressed Rad51 expression. DACH1 enhanced the cytotoxcity of cisplatin and doxorubicin, two commonly used drugs for NSCLS. Together, our studies demonstrate that p53 is DACH1 binding partner that inhibits NSCLS cellular proliferation.

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-144. doi:1538-7445.AM2012-LB-144
Abstract LB-200: IL-1beta promotes the development of intestinal epithelial stem cells
Authors: Shi Jishu , Wang Lei , Liu Ziyan , Li Yijing , Pappan Loretta
Published: August 2012

Although iflammation has been linked to stem cell-mediated tumorigenesis, the inflammatory factors responsible for the induction of abnormal proliferation of stem cells remain to be identified. Up-regulation of IL-1β has been associated with colon tumor initiation and progression. The objective of this study is to determine whether IL-1β alone may contribute to intestinal stem cells (ISC)-mediated tumorigenesis by promoting the development of ISCs. In a preliminary study, it was shown that rat normal intestinal epithelia cell line IEC-18 cells proliferated as a monolayer culture under serum-free conditions, demonstrating that either IEC-18-ISCs are not active in these conditions or ISCs are absent in IEC-18 cell line. However, we have found that addition of IL-1β to the cell culture medium caused monolayer IEC-18 cells to form spheres with up-regulated expression of stem cell markers including Bmi-1, β-catenin, Lgr5, c-Myc, and Nanog. This shows that IL-1β can induce the development and/or self-renewal of ISCs to form spheres. IL-1β-mediated ISC self-renewal is further demonstrated by a colony-forming assay when IEC-18 cells were cultured in soft agar under serum-free conditions. Interestingly, IL-1β promoted only a relatively small population (about 0.03%) of IEC-18 cells to form colonies, suggesting that the majority of IEC-18 cells are terminally differentiated cells. Furthermore, we have found that, when cultured in collagen-coated plates in serum-free medium, IL-1β-treated IEC-18 cells demonstrated a loss of cell-cell contact inhibition, growth to relatively high cell densities as evidenced by the presence of foci, and up-regulated expression of stem cell markers. Overall, results from our studies provide the first evidence that IL-1β alone can promote the development and self-renewal of ISCs. IL-1β-activated ISCs possess typical characteristics of malignant cells including colony formation in soft agar and loss of cell-cell contact inhibition of growth. Thus, aberrant production of IL-1β in intestinal inflammation may contribute to the initiation and progression of colon cancer by inducing the abnormal development of intestinal epithelial 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-200. doi:1538-7445.AM2012-LB-200
Abstract LB-202: The significance of stress-responsive molecules in cancer stem cells derived from ovarian and endometrial cancer
Authors: Yasuda Kazuyo
Published: August 2012

Cancer stem cells (CSC) are characterized by their high tumor-initiating capacity, self-renewal and differentiation. Recent studies have been reported that higher aldehyde dehydrogenase1 (ALDH1) activity represent the characters of CSCs. By using ALDEFLUOR assay, we could successfully isolate stem-like cells from endometrial and ovarian cancer cell lines. ALDH1-high cells expressed stem cell markers, such as SOX-2, Oct-3/4 and Nanog. They showed significantly higher tumorigenicity than ALDH1-low cells in the transplantation test using NOD/SCID mice. Comparative gene expression analysis revealed several characteristic molecules that were expressed preferentially in ALDH1-high cells, including stress-responsive molecules, oxidoreductase and heat shock proteins. It is suggested that high levels of stress-responsive genes may explain at least in part highly stress resistance properties of CSCs. In addition, stress-responsive pathway might become a therapeutic molecular target of CSCs.

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-202. doi:1538-7445.AM2012-LB-202
Abstract LB-207: Tissue transglutaminase (TG2) - the key driver in inflammation-induced progression of cancer*
Authors: Mehta Kapil , Kumar Santosh , Kumar Anupam
Published: August 2012

It is becoming evident that chronic inflammation plays important roles during carcinogenesis, tumor invasion and metastasis. However, molecular mechanisms linking inflammation to cancer progression remains elusive. Here we report that aberrant expression of tissue transglutaminase (TG2), a stress-responsive gene whose expression is modulated by multiple inflammatory signals, plays a key role in reprogramming epithelial cancer cells to dedifferentiation pathway. TG2 expression initiated the developmental program of epithelial to mesenchymal transition (EMT) and conferred stem cell traits in non-transformed and transformed epithelial cells. At molecular level, TG2 expression resulted in constitutive activation of nuclear transcription factor, NF-κB via a non-canonical pathways. Thus, TG2 associates in complex with the inhibitory protein IκBα, resulting in its rapid degradation via non-proteasomal pathway. Hypoxia-induced factor (HIF-1α) was identified as one of the direct downstream targets for TG2-induced NF-κB. TG2/ NF-κB-induced increase in HIF-1α resulted in regulation of transcription repressors like Snail, Zeb-1, and Twist, the known inducers of EMT/CSC. Importantly, our studies suggested that TG2 in closed or compact conformation could promote the oncogenic functions while open or extended conformation is completely inactive in this regard. TG2-induced programming rendered epithelial cells resistant to stress-induced death (including chemo- and radiation-therapies) and increased their propensity to metastasize. Indeed, previous studies support this contention and suggested that inhibition of TG2 could increase the sensitivity of cancer cells to chemotherapy and inhibit their metastasis, both in vitro and in animal models. Moreover, high basal expression of TG2 in breast, ovarian and pancreatic tumor samples is associated with poor disease outcome. These data warrants further studies to develop selective inhibitors/modulators of TG2 to increase the sensitivity of cancer cells to chemotherapy and to prevent their progression to metastatic disease. *Supported by a grant from Susan G. Komen Foundation for Cure.

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-207. doi:1538-7445.AM2012-LB-207
Abstract LB-212: Ionizing radiation selectively promotes invasion of ErbB2-expressing breast cancer cells via regulation of FoxM1
Authors: Kambach Diane M. , Sodi Valerie , Lelkes Peter I. , Reginato Mauricio , Azizkhan-Clifford Jane
Published: August 2012

ErbB2 is frequently amplified in premalignant breast cancers including ductal carcinoma in situ (DCIS); however, little is known about the signals or pathways it modulates in the progression into the invasive/malignant state. Radiotherapy is often used to treat early premalignant lesions regardless of ErbB2 status. Here we show that clinically relevant doses of ionizing radiation (IR) induce cellular invasion of ErbB2-expressing breast cancer cells, such as SKBR3 and BT-474, as well as MCF10A cells transduced to express ErbB2. This effect is specific to ErbB2 expression, as ErbB2-negative breast cancer cells, such as MCF7 and T47D, do not invade following treatment with IR, nor do parental MCF10A cells or MCF10A cells overexpressing EGF receptor alone. Exposure of cells to X-rays leads to ErbB2 phosphorylation at Y877 in a dose- and time-dependent manner, and activates signaling pathways, including Src and PI3K/Akt. Inhibition of these pathways, as well as inactivation of IR-induced reactive oxygen species with the antioxidant N-acetylcysteine, prevented invasion. Activation of ErbB2-dependent signaling resulted in the upregulation of the transcription factor, FoxM1, a member of the forkhead family which has previously been shown to have a role in invasion, in part through transcriptional regulation of MMP2. Indeed, we have seen upregulation of MMP2 in irradiated ErbB2-expressing breast cancer cells, which is abolished when Src or ErbB2 is inhibited. Inhibition of FoxM1 by shRNA abrogated the increase in MMP2 level and prevented induction of invasion by IR, and overexpression of FoxM1 alone in MCF10A cells was sufficient to promote IR-induced invasion. Our data show that low level IR stimulates invasion of ErbB2-expressing breast cancer cells in a FoxM1-dependent manner.

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-212. doi:1538-7445.AM2012-LB-212
Abstract LB-22: Gemcitabine resistance is associated with EMT phenotype along with expression of cancer stem cell markers and a specific miRNA profile in the human pancreatic cancer cell line BxPC3
Authors: Bera Alakesh , Freeman James W. , VenkataSubbaRao Kolaparthi , Zhao Shujie
Published: August 2012

Despite new therapeutic strategies, human pancreatic ductal adenocarcinomas (PDAC) remain one of the greatest oncological challenges, as the rate of incidence almost equals the rate of mortality with a 5-year survival of less than 5%. Gemcitabine is the standard therapy for PDAC and is mostly used alone or in combination with an EGFR inhibitor. While tumors respond to Gemcitabine treatment initially, they generally develop chemoresistance. A thorough understanding of molecular events that promote resistance against Gemcitabine may identify pathways that can be efficiently targeted to reverse chemoresistance. A cell line model resistant to Gemcitabine was generated by transiently exposing BxPC3 cells to increasing concentrations of Gemcitabine. This Gemcitabine resistant cell line, BxPC3-GzR, was compared with untreated BxPC3 control cells. BxPC3-GzR cells showed properties of undergoing epithelial to mesenchymal transition (EMT) and these included a spindle shaped morphology, an increase in expression of vimentin and a decrease in expression of E-cadherin. BxPC3-GzR cells also showed an increase of expression of CD44 and in expression of phosphorylated STAT3TY705, which are characteristic of cancer stem cells. BxPC3 and BxPC3-GzR where further compared by microarray analyses for expression of micro-RNAs (miRNAs). BxPC3-GzR cells showed a specific miRNA expression profile (9 positively and 8 negatively regulated). Thus far seven of eight miRNAs identified as altered in BxPC3-GzR cells were validated by real-time RT-PCR (miR-15b, miR-21, miR-100, miRNA -125b, miRNA-155, miRNA-205 and miRNA-455-3p). These studies suggest that Gemcitabine treatment of PDAC selects for a resistant population of tumor cells that have a mesenchymal/stem cell-like phenotype and that express a distinct miRNA profile. Targeting these miRNAs may provide novel targets for restoring or enhancing anti-tumor response of Gemcitabine.

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-22. doi:1538-7445.AM2012-LB-22