bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 2427: Endothelioma of bone revisited: G-Protein coupled receptor 64 promotes invasiveness and metastasis in Ewing Tumors
Authors: Richter Günther H. , Fasan Annette , Roessler Sabine , Burdach Stefan
Published: June 2012

Metastatic spread in Ewing Tumors (ET) is frequent and hematogenous. Microarray analysis revealed the G-protein coupled receptor 64 (GPR64), an orphan receptor with normal expression restricted to human epididymis, to be over-expressed in ET. Members of this family of receptors have been implicated with vascularization and neuronal development. Inhibition of GPR64 expression by RNA interference impaired endothelial differentiation in vitro and suppressed local tumor growth and metastasis in Rag2-/-γC-/- mice. Microarray analysis of ET after GPR64 knock down revealed a GPR64-mediated suppression of genes involved in differentiation like v-erb-a erythroblastic leukemia viral oncogene homolog 4 (ERBB4), and genes stimulating transcriptional activation including pre-B cell leukemia homeobox 2 (PBX2) and SWI/SNF related, matrix associated, actin dependent regulator of chromatin, subfamily a, member 4 (SMARCA4). Similarly, a GPR64-mediated induction of VEGF receptor 1 ligand placental growth factor (PGF) in ET was observed. PGF itself was induced by EWS-FLI1 in mesenchymal stem cells. The GPR64-mediated induction of placental growth factor (PGF) and repression of PGF resulted in a phenotype similar observed after GPR64 knockdown. GPR64 as well as PGF inhibition was associated with reduced proteolytic activity of Matrix Metalloproteinase (MMP) 1 and invasiveness in vitro. MMP1 knock down abrogated lung metastasis in Rag2-/-γC-/- mice. GPR64 and subsequent up-regulation of PGF and MMP1 orchestrate and promote invasiveness and metastatic spread of this tumor, and provide significant conceptual progress in understanding both the histogenesis of ET in particular and the pathogenesis of metastatic dispersal in general.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 2427. doi:10.1158/1538-7445.AM2011-2427
Abstract 2433: Hepatitis B virus X protein induces metastasis-associated protein 1 gene expression through promoter methylation
Authors: Lee Min-Ho , Na Hyelin , Lee Mi-Ock
Published: June 2012

Expression of metastasis-associated protein 1 (mta1) gene is well correlated with the degree of invasion and lymphatic metastasis in colorectal, hepatocellular and breast carcinomas. Expression of MTA1 was induced by hepatitis B virus X protein (HBx), however, little is known about transcriptional regulation of mta1 gene expression. Here, we reported that 5′-flanking region of MTA1 promoter contains two CpG islands. CpG island 1 and CpG island 2. The CpG island 1 was highly methylated (32.5%) but CpG island 2 (<1%) was not. Using bisulfite-modified direct sequencing, transient expression of HBx led to an increase in methylation of the CpG island 1 up to 48.8 %. In addition, chromatin immunoprecipitation revealed that HBx recruited DNMT3A to CpG island 1 region of MTA1 promoter. In silico analysis of the CpG island 1 predicted an existence of putative p53 binding sequences. We demonstrated that p53 bound to the CpG island 1 by chromatin immunoprecipitation using specific anti-p53 antibody. Further, p53 was pull-downed by DNA probe encoding the p53 binding sequences. However p53 was not pull-downed by the methylated DNA probe using SssI DNA methyltransferase. These data showed that HBx induced methylation of CpG island 1, which interferes the DNA binding of p53 on the specific DNA region. This result may provide the molecular mechanism of how HBx induces the expression of MTA1 gene.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 2433. doi:10.1158/1538-7445.AM2011-2433
Abstract 2439: Association of antigen processing machinery components and CD8+ T-cell infiltration in breast cancer brain metastasis
Authors: Liu Yan , Komohara Yoshihiro , Domenick Natalie , Horbinski Craig , Hamilton Ronald , Ferrone Soldano , Okada Hideho
Published: June 2012

Purpose: Defects in human leukocyte antigen (HLA) class I antigen processing machinery (APM) component expression can have a negative impact on the clinical course of tumors and the response to T cell-based immunotherapy. Since brain metastases of breast cancer are of increasing clinical significance and only scant information is available about the expression of APM component abnormalities in these lesions, the APM component expression and CD8+ T cell infiltration pattern was analyzed in primary breast and metastatic brain lesions of breast cancer.

Experimental design: Immunohistochemistry was performed to evaluate the expression of APM components HLA class I heavy chain, β2-microglobulin, tapasin, antigen peptide transporter (TAP)1, TAP2, LMP2 and calnexin.

Results: Comparison of unpaired 50 primary and 33 brain metastases showed lower expression of β2-microglobulin, TAP1, TAP2 and calnexin in the brain lesions. Although no significant differences were found in APM component scores between primary breast and brain lesions in 15 paired cases, primary breast lesions of which patients eventually developed brain metastases showed lower levels of β2-microglobulin, TAP1 and calnexin compared with newly diagnosed breast lesions without brain metastases. Furthermore, the extent of CD8+ T cell infiltration was significantly higher in the newly diagnosed breast lesions compared with the ones of which the patients developed brain metastases, and was positively associated with the expression of TAP1 and calnexin.

Conclusions: Our data suggest that patients with low or defective TAP1 or calnexin in the primary breast cancers may be at higher risks for developing brain metastases due to the reduced immunosurveillance.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 2439. doi:10.1158/1538-7445.AM2011-2439
Abstract 2458: MicroRNA miR-34 as a natural regulator and potential therapeutic for pancreatic cancer stem cells
Authors: Meng Xiaojie , Ji Qing , Hao Xinbao , Tang Wenhua , Xu Liang
Published: June 2012

MicroRNAs (miRNAs) have been implicated in cancer initiation and progression and are involved in cancer stem cell dysregulation. We show that miR-34 directly regulates Notch and Bcl-2, suggesting the possibility of a role for miR-34 in the maintenance and survival of cancer stem cells. We examined the roles of miR-34 in p53-mutant human pancreatic cancer cell lines and the potential link to cancer stem cells. Restoration of miR-34 expression in the cancer cells downregulated Bcl-2 and Notch1/2. miR-34 restoration significantly inhibited clonogenic cell growth and invasion, induced apoptosis and G1 and G2/M arrest in cell cycle, and sensitized the cancer cells to chemotherapy and radiation. We identified that CD44+/CD133+ cancer cells are enriched with tumorsphere-forming and tumor-initiating cells or cancer stem/progenitor cells with high levels of Notch/Bcl-2 and loss of miR-34. More significantly, miR-34 restoration led to significant reduction of the tumor-initiating cell population, accompanied by inhibition of tumorsphere growth in vitro and tumor-initiation in vivo. Our results demonstrate that miR-34 may restore, at least in part, the tumor suppressing function of the p53 in p53-deficient human pancreatic cancer. Our data support that miR-34 may be involved in cancer stem cell self-renewal, potentially via the direct modulation of downstream targets Bcl-2 and Notch, implying that miR-34 may play an important role in cancer stem cell self-renewal and/or cell fate determination. Currently, we are exploring the nanoparticle-targeted delivery of miR-34 to cancer stem cells as a novel molecular therapy for human pancreatic cancer, based on our patented nanobiotechnology which has been approved by FDA for clinical trials. Taken together, our results demonstrate that restoration of miR-34 may hold significant promise as a novel molecular therapy for human pancreatic cancer with loss of p53-miR34, potentially via inhibiting cancer stem cells. Molecularly targeted miRNA therapy could be a powerful tool to correct the cancer stem cell dysregulation. Supported in part by NIH grants CA121830(S1) and CA134655.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 2458. doi:10.1158/1538-7445.AM2011-2458
Abstract 2465: Microenvironmental sugars regulate pancreatic cancer stem cell transcription to promote an invasive phenotype
Authors: Liu Haibo , Wang Charles , Du Li , Heaney Anthony
Published: June 2012

Pancreatic cancer is a lethal disease and there are few effective therapies. Increased refined carbohydrate and specifically fructose consumption have been associated with increased pancreatic cancer risk in some studies. We recently demonstrated that fructose-treated pancreatic cancer cells grow in vitro at equivalent rates to glucose and observed that cancer cell metabolism of glucose and fructose are quite different whereby fructose increased nucleic acid synthesis, a feature of a highly aggressive cancer phenotype.

A subset of cells within a tumor, termed cancer stem cells (CSC) that highly tumorigenic and resistant to chemotherapy. We recently demonstrated that the metabolism of fructose and glucose in pancreatic cancer is very different with fructose preferentially providing carbons for nucleic acid synthesis. To gain insight into how nutrients such as fructose might differentially regulate cancer cell metabolic pathways to promote an aggressive phenotype, we performed mRNA expression profiling followed by QT-PCR in fructose-treated pancreatic CSCs and compared gene expression to pancreatic CSCs maintained in normal medium. Following fructose-treatment (0.55-5.5mM), we demonstrated increased chemokine receptor4 (CXCR4), epidermal growth factor receptor (EGFR) and glutathione peroxidase (GPX1) mRNA expression in comparison to glucose- (0.55-5.5mM) or normal medium (22mM glucose) -treated pancreatic CSCs and in comparison to fructose- or glucose-treated pancreatic cancer non stem cells (NSC), whole Panc-1 cells and normal pancreatic ductal HPDE6 cells.

In additional studies we observed that the nutrient sugar fructose selectively activates distinct tumorigenic signaling cascades in pancreatic CSCs including phosphorylated Akt and GSK-3α/β to increase CXCR4 to potentially promote growth, survival and metastases of pancreatic cancer stem cells. We also observed that fructose potently induced pancreatic CSC mRNA expression of the selective fructose transporter GLUT5, providing further insight into fructose-mediated actions in pancreatic CSCs. In summary, we demonstrate that sugars mediate effects beyond that of a simple substrate and that environmental dietary nutrients can alter gene expression to potentially modify the cancer phenotype.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 2465. doi:10.1158/1538-7445.AM2011-2465
Abstract 2471: The tumor suppressor gene Rb1 controls telomeric length and genomic instability that predisposes to osteosarcoma development in irradiated mice
Authors: Gonzalez-Vasconcellos Iria , Sanli-Bonazzi Bahar , Anastasov Natasa , Atkinson Michael J. , Rosemann Michael
Published: June 2012

Osteosarcomas are capable of exhibiting both alternative lengthening of telomeres (ALT) and genomic instability, and are commonly associated with inactivation of the Rb1 gene implicated in both, cell cycle checkpoint regulation and telomere maintainance. We demonstrate a functional relationship between the loss of Rb1, genomic instability and the shortening of telomeres in radiation-induced osteosarcoma. Conditional somatic deletion of a floxed-Rb1 allele in the osteoblastic lineage produces an increased sensitivity to the osteosarcomagenic effects of radiation (Gonzalez-Vasconcellos et al, 2010).

In Rb1 +/- osteoblast cell lines established from non-irradiated Rb1fl/+ Col1a1-Cre mice we show that after irradiation there was the expected impaired cell cycle arrest accompanied by an increase in genomic instability and reduced radiation induced senescence. The DNAmicronucleus assay showed an increase of 1.53 fold in Rb +/- primary cells as compared to Rb+/+ after 2 Gy irradiation (p=0.0032).

We could prove that the reduced Rb1 gene expression in primary osteoblasts leads to a reduction in telomeric length of up to 50% that increases in a passage dependent manner and was shown to be independent of radiation exposure. These mutant cells showed an inclination towards tetraploidy and radiation induced anaphase bridges formation was increased in 2.6 fold in the mutant cells after 2 Gy irradiation (p = 0.001).

These studies suggest the tumor suppressor gene Rb1 functioning not only as a gatekeeper if not also as a caretaker of the genome controlling the telomere dynamics and genomic stability. This maintenance of the genome by Rb1 was proven to be driven by epigenetic changes on the Rb+/- primary osteoblasts involving histone methylation. Lower levels of 3metLys20-H4 fluorescence intensity (p = 0.0079) were detected in Rb1 mutant osteoblasts as compared to wt.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 2471. doi:10.1158/1538-7445.AM2011-2471
Abstract 2492: Protein phosphatase PP6 regulates the sensitivity of glioblastoma cells to radiation
Authors: Shen Ying , Fei Xin , Wang Yongbin , Kong Xianming , Qiu Yongming , Mi Jun
Published: June 2012

Glioblastoma is one of most aggressive primary tumors. The average survival time of GBM patients is 12 months; moreover, GBM is highly resistant to radiotherapy and chemotherapy, so, how to increase the sensitivity of GBM cells to radiotherapy is still a challenging to all oncologists. Our study demonstrate serine/threonine protein phosphatase PP6 is associated with the radioresistance of glioblastoma (GBM) cells, and the expression level of PP6c (PP6 catalytic subunit) is highly correlated with the survival time of GBM patients (R2 = -0.7). Lentiviruses-mediated PP6c knocking down increases the radiosensitivity of glioblastoma in xenografted mouse model, and prolong the survival time of tumor-bearing mouse; while the overexpression of PP6c decreases the radiosensitivity of glioblastoma and shortens the survival time of tumor-bearing mouse. Our data also show the PP6 is complexed with DNA-PK by the co-immunoprecipitation assay, the siRNA knock-down of PP6c inhibits DNA damage-induced DNA-PK activation and DNA-repair capacity, and sensitized the U251, T98G, U87 three cell lines to radiation in vitro. These results strongly suggest PP6 is a therapeutic target for radiosensitization.

Key Words: DNA-PK, PP6, ubiquitination, radiosensitivity, GBM

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 2492. doi:10.1158/1538-7445.AM2011-2492
Abstract 2503: Distinct radiosensitivities of human FancG-/- and FancD2-/- cell lines
Authors: Bernard Mark E. , Epperly Michael W. , Franicola Darcy , Zhang Xichen , Shields Donna , Houghton Frank , Bakkenist Christopher J. , Guinan Eva C. , Kannan Neeta , Greenberger Joel S.
Published: June 2012

Background: Fanconi Anemia is an autosomal recessive disorder marked by a dysfunction of DNA repair, an increased sensitivity to DNA crosslinking agents, and some patients poorly tolerating ionizing irradiation. We determined the radiosensitivity of two Fanconi patient cell lines and we determined if the nitroxide radioprotector Tempo or a novel mitochondrial targeted nitroxide JP4-039 could alter their radiosensitivity.

Materials and Methods: Fanconi patient derived fibroblastic cell lines, PD326 and PD20F, and their respective transgene restored cell lines FancG and FancD2 were irradiated to doses ranging from 0 to 8 Gy in the presence or absence of Tempo or JP4-039 (10 µm), plated, and colonies greater than 50 cells were counted eight days later.

Results: The PD20F cell line was more radiosensitive compared to the restored FancD2 cell line (n = 1.6 + 0.2 and 10.7 + 3.1, respectively, p = 0.0423). In contrast the PD326 cell line was more radioresistant than its restored FancG cell line as seen by an increased shoulder on the survival curve (n = 8.1 + 1.4 and 2.2 + 0.3, respectively, p = 0.0174). JP4-039 but not Tempo increased radioresistance in the PD326 (n = 15.7 + 2.5, p = 0.0373), and FancG cell lines (10.2 + 2.1, p = 0.0151).

Conclusions: The human Fanconi Anemia cell line PD326 is radioresistant when compared to its transgene restored FancG cell line, and this radioresistance is increased by treatment with the mitochondrial targeted nitroxide JP4-039. In contrast, PD20F is more radiosensitive when compared to its transgene restored FancD2 cell line. Testing the efficacy of JP4-039 and Tempo on mitigating the effects of mitomycin C treatment is being evaluated.

Supported by 2U19A1068021-06

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 2503. doi:10.1158/1538-7445.AM2011-2503
Abstract 2537: Novel 2′-deoxycytidine analogs as DNA demethylation agents
Authors: Thottassery Jaideep V. , Tiwari Kamal , Westbrook Louise , Secrist John A. , Parker William B.
Published: June 2012

Epigenetic therapies using cytidine/deoxycytidine analogs are proving to be effective as indicated by the recent approvals of 5-azacytidine (5-azaCyd) and 5-aza-2’-deoxycytidine (5-azadCyd) in myelodysplastic syndromes (MDS) and certain leukemias. Studies suggest that the inhibition of DNA cytosine-5 methylation and the re-expression of silenced tumor suppressors contribute to the beneficial effects of these drugs. However inhibition of DNA synthesis and other toxicities of these compounds represent major drawbacks in the clinic. Recently another analog previously examined in our anticancer discovery program, 4’-thio-2’-deoxycytidine (T-dCyd), had been shown to be an inhibitor of methyl transfer by the M. HhaI DNA cytosine-5 methyltransferase. In the current studies we demonstrate that T-dCyd and its 5-aza analogue 4’-thio-5-aza-2’-deoxycytidine (5-aza-T-dCyd), can deplete human DNMT1 protein in NCI-H23 lung carcinoma and the myeloid leukemia lines THP-1 and KG1a. Consistent with this we also found that T-dCyd and 5-aza-T-dCyd were also effective in decreasing DNMT activities in cancer cells. Methylation-specific PCR (MSP) analysis also demonstrated that both T-dCyd and 5-aza-T-dCyd induced CpG demethylation and re-expression of the tumor suppressor p15 in KG1a cells. We have also found that T-dCyd is inserted into replicating DNA at nanomolar doses as readily if not better than the natural 2’-deoxycytidine (dCyd) and exhibited very little toxicity at these doses. T-dCyd is therefore readily activated to its triphosphate T-dCTP, which is a good substrate for DNA polymerase mediated incorporation, and DNA polymerases also readily extend the chain after incorporation. Furthermore we also observed that T-dCyd and 5-aza-T-dCyd are efficacious in in vivo tumor models. Collectively our data suggest that these compounds or their analogs could be developed as novel DNA methylation inhibitors with better properties for cancer therapy.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 2537. doi:10.1158/1538-7445.AM2011-2537
Abstract 256: Downregulation of Pim-3 kinase inhibits cell growth and chemosensitizes pancreatic cancer cells to gemcitabine
Authors: Xu Dapeng , Gavilano Lily , Witherspoon Sam M. , Bednarski Brian , Kim Hongjin , Baldwin Albert S. , Baines Antonio T.
Published: June 2012

Pancreatic ductal adenocarcinoma (PDAC) is the most common cancer of the pancreas, comprising over 85% of all cases. PDAC has a relative 1-year survival rate of 24% and a 5-year survival rate of 5%. Recent clinical studies indicate that patients with advanced pancreatic cancer show a 12% tumor response rate after treatment with the chemotherapeutic drug gemcitabine. Even in combination with other chemotherapeutic agents such as erlotinib, survival rates are only increased by a few weeks. Past studies have shown that members of the oncogenic Pim kinase family are aberrantly expressed in a variety of solid tumors, including prostate and pancreatic cancers. Most recently, Pim kinase has been correlated with chemoresistance in prostate cancer cells. Our study aimed to investigate the role of Pim-3 kinase in the chemoresistance of PDAC cancer cells to gemcitabine. We hypothesize that inhibition of Pim-3 will decrease cell growth and sensitize PDAC cells to gemcitabine treatment. Initially, we confirmed by immunohistochemistry and western blot analysis that Pim-3 is aberrantly expressed in PDAC tissues and cell lines, respectively. We found that inhibition of Pim-3 expression by shRNA decreased PDAC cell growth and resulted in an accumulation of cells in G1 or S phase of the cell cycle. Additionally, we demonstrated that PDAC cells expressing Pim-3 shRNA had increased apoptosis after gemcitabine treatment compared to controls. To determine a possible mechanism for this resensitization of PDAC cells to gemcitabine, we measured multidrug resistance (MDR)1 activity in the Pim-3 shRNA cells. We found that inhibition of Pim-3 protein expression in PDAC cells promoted gemcitabine sensitivity, in part, by lowering MDR1 activity. Our results suggest that downregulation of Pim-3 potentiates the efficacy of gemcitabine in combating pancreatic cancer. These findings support the idea that inhibiting Pim kinases, in combination with chemotherapeutic agents, could play an important role in pancreatic cancer treatment by targeting the clinical problem of chemoresistance.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 256. doi:10.1158/1538-7445.AM2011-256
Abstract 2561: Inhibiting the mTOR pathway synergistically enhances etoposide-induced cytotoxicity by upregulating c-Jun in ovarian cancer cells
Authors: Itamochi Hiroaki , Nonaka Michiko , Sato Seiya , Uegaki Kazunori , Naniwa Jun , Sato Shinya , Shimada Muneaki , Tetsuro Oishi , Terakawa Naoki , Kigawa Junzo , Harada Tasuku
Published: June 2012

Objective: The mammalian target of rapamycin (mTOR) pathway is considered to be a central regulator of proliferation and survival of cells. Rapamycin and its analogs are undergoing clinical trials in patients with epithelial ovarian cancer. Rapamycin has been reported to enhance the effectiveness of several chemotherapeutic agents. The present study aimed to assess the potential of combination effects of rapamycin and anticancer drugs used as first- or second-line chemotherapy to treat ovarian cancer.

Methods: Six cell lines of ovarian serous adenocarcinoma (KF, KOC-2S, SHIN-3, SK-OV-3, TU-OS-3, TU-OS-4) were used in this study. We treated the cells with rapamycin combined with anticancer agents, then assessed cell viability, apoptosis, and the expression of proteins in apoptotic pathways and molecules downstream of the mTOR signaling pathways. We also investigated the effect of combined treatment on survival in xenograft models with nude mice.

Results: The protein expression levels of PTEN, phosphorylated (p) Akt, pmTOR, p4E-BP1, and pS6K1 were confirmed in all cell lines. Synergistic effects with rapamycin were observed in combination with etoposide in five cell lines, with doxorubicin in four lines, and with SN38, which is an active metabolite of camptothecin-11, in two lines. However, we observed antagonistic effects when rapamycin was combined with gemcitabine, cisplatin, or paclitaxel on more than two cell lines. Rapamycin dramatically enhanced apoptosis induced by etoposide and the expression of cleaved caspase 9. This effect was associated with up-regulation of phosphorylated c-Jun and down-regulation of Bcl-xL. The synergistic interaction of rapamycin and etoposide was lower when the c-Jun pathway was suppressed by a c-Jun N-terminal kinase inhibitor (SP600125). Finally, treating nude mice with rapamycin and etoposide significantly prolonged survival in model mice with ovarian cancer xenografts (P<0.001).

Conclusion: Combination chemotherapy consisting of rapamycin and etoposide showed synergistic effect against epithelial ovarian cancer. This treatment modality is worth exploring further for patients with ovarian cancer.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 2561. doi:10.1158/1538-7445.AM2011-2561
Abstract 2566: Beneficial impact of valproate on the response of small cell lung cancer to cisplatin – etoposide therapy
Authors: Vandermeers Fabian , Hubaux Roland , Crisanti Maria Cecilia , Kapoor Veena , Albelda Steven M. , Willems Luc
Published: June 2012

Prognosis of small cell lung carcinoma (SCLC) is particularly poor, less than 5% of patients with extensive stage being alive after two years. We hypothesized that SCLC chemotherapy could be improved by using histone deacetylase (HDAC) inhibitors based on their ability to interfere with lysine acetylation and to alter gene expression. The goal of this study was to evaluate the anticancer efficacy of a HDAC inhibitor (valproate: VPA) on SCLC cells in combination with the standard chemotherapeutic first-line regimen (cisplatin + etoposide). We show that VPA induces apoptosis of small cell lung cancer cell lines and improves efficacy of cisplatin combined with etoposide. Both mitochondrial and death receptor pathways are involved in VPA-induced apoptosis. As expected for an HDAC inhibitor, VPA hyperacetylates histone H3. The mechanism of VPA pro-apoptotic activity involves induction of p21, inhibition of Bcl-XL, cleavage of Bid and phosphorylation of Erk and H2AX. In the presence of VPA, Bax is translocated from the cytoplasm to the mitochondria and cleaved in an 18 kDa isoform. Cytochrome c is released from the mitochondria into the cytosol. Transcriptomic analyses by microarray show that VPA modulates transcription of genes (Na+/ K+ ATPase, Bcl-XL) involved in chemoresistance to cisplatin and etoposide. Finally, the efficacy of VPA combined with cisplatin and etoposide is supported by preclinical models of SCLC cells engrafted into SCID mice. Together, these data demonstrate that VPA augments anticancer activity of cisplatin and etoposide, two components of the standard first-line chemotherapy of small cell lung cancer.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 2566. doi:10.1158/1538-7445.AM2011-2566
Abstract 2588: Targeting overexpressed PLK1 and Aurora kinases in childhood medulloblastoma
Authors: Holst Martin I. , Westerhout Ellen , Kool Marcel , Caron Huib , Versteeg Rogier , Clifford Steve , Rutkowski Stefan , Pietsch Torsten
Published: June 2012

The European research project Kids Cancer Kinome (KCK) is focusing on the investigation of aggressive childhood cancers. The competence of nine research centers is combined to explore the human protein kinase family in different highly malignant pediatric tumors (medulloblastoma, Ewing sarcoma, osteosarcoma, rhabdomyosarcoma, neuroblastoma, ALL) to validate novel drug targets and develop novel therapeutic options. Medulloblastomas belong to the most frequent malignant brain tumors in children and represent about 20% of all pediatric CNS cancers. Analyzing mRNA expression profiles (Affymetrix microarrays) we discovered a series of kinases considerably overexpressed in primary medulloblastoma tumors samples and medulloblastoma cell lines compared to non-neoplastic tissues.

We found a significant mRNA overexpression of PLk1 and Aurora kinases A and B in medulloblastoma tumor samples and cell lines. For further investigation we selected these kinases which are all involved in cell cycle control. Defects in the process of cell division are believed to be associated with tumorigenesis.

Using the small molecules inhibitors ZM447439 and VX680 for Aurora kinases and BI2536 and GW843682 for PLK1 inhibition we evaluated the effects on medulloblastoma cell proliferation and apoptosis. Incubation of medulloblastoma cells with the inhibitors for 72h resulted in a concentration dependent decrease of the proliferative activity as measured by MTS proliferation assays and an increased apoptosis determined by Caspase-Glo 3/7 assay.

In a second approach, we transduced cell lines with lentiviral particles carrying shRNA targeting PLK1 or Aurora kinases A and B. The specific downregulation of the kinases mediated by the lentiviral shRNA was confirmed by Western blotting and had a negative effect on the viability of the medulloblastoma cells. We observed that the effect of PLK1 inhibition on the proliferation of the medulloblastoma cell lines was more effective compared to the aurora kinases.

In conclusion, these data suggest that the inhibition of PLK1 or Aurora kinases may represent a promising approach for novel targeted strategies in the treatment of medulloblastoma.

http://www.kidscancerkinome.org

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 2588. doi:10.1158/1538-7445.AM2011-2588
Abstract 2589: Sulfated hyaluronic acid: A possible targeted therapy for prostate cancer
Authors: Yates Travis , Benitez Anaid , Lopez Luis E. , Lokeshwar Vinata B.
Published: June 2012

Tumor cell-derived hyaluronidase HYAL1, which degrades hyaluronic acid (HA), promotes tumor growth and metastasis. In prostate cancer (CaP), HYAL1 expression in biopsy and prostatectomy specimens is an independent prognostic marker for predicting disease progression. Sulfated hyaluronic acid (sHA) inhibits HYAL1 activity through a mixed inhibition mechanism. Antitumor activity of sHA has not been evaluated. In this study we evaluated effects of sHA on CaP cells. The effect of sHA (0-40 ug/ml) on cell proliferation and apoptosis was examined in CaP cells (PC3-ML, LNCaP, LNAI, C4-2B, LAPC-4, DU145, RWPE1) by cell counting and Cell Death ELISA kit. Matrigel invasion and Boyden chamber assays were used to test the effect of sHA on invasive activity. Effect of sHA on signaling, apoptosis cascade, androgen receptor activity, VEGF, NFkB activity, and HA receptor levels, was evaluated by immunoblotting, Q-PCR and reporter assays. Athymic mice bearing LNAI xenografts were treated with sHA (25 and 50 mg/kg) by i.p. injection. sHA inhibited proliferation, motility and invasion in CaP cells. At IC50 for HAase activity inhibition (∼ 10-μg/ml), sHA induced > 3-fold apoptosis and inhibited invasive activity of CaP cells, which could be partially reversed by HA-fragments. sHA induced caspase-3, -8, -9 activation, up-regulation of Fas, Fas-L, FADD, and DR4 and down regulated bcl-2, phospho-bad, bcl-XL, phospho-Akt, phospho-IkB, and phospho-AR. At IC50, sHA caused > 90% inhibition of NFKB-reporter activity, PSA-promoter activity and transcriptionally down regulated HA-receptors (CD44 and RHAMM) and VEGF. Effect of sHA could be reversed by overexpression of m-Akt. Down regulation of CD44 and RHAMM, together with sHA synergistically inhibited cell proliferation and Akt activation. sHA significantly inhibited LNAI tumor growth. The majority of the animals did not form palpable tumors at 50-mg/kg dose even after 70 days when the treatment was stopped on 42nd day. No weight loss or serum and organ toxicity was observed in sHA treated animals. Tumors showed reduced microvessel density (∼3-fold) and increased TUNEL positive cells (> 5-fold). This is the first study that shows targeting HYAL1 (by sHA – a novel non-toxic agent), may control CaP growth and progression.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 2589. doi:10.1158/1538-7445.AM2011-2589
Abstract 2603: Inhibition of Notch and mTOR by nelfinavir as a novel approach for T cell acute lymphoblastic leukemia (T-ALL)
Authors: Chang Yoon Soo , Meza Leah M. , Onozawa Masahiro , Gills Joell J. , Aplan Peter D. , Dennis Phillip A.
Published: June 2012

Background: T-ALL accounts for 10-15% of childhood ALL and 25% of adult ALL. Although intensive chemotherapy improves disease free survival, patients who relapse or progress have a dismal prognosis. Notch1 is a promising target in T-ALL because it is commonly activated, and inhibitors of γ-secretase (GSI), an aspartic protease that processes Notch1, are in clinical trials. Nelfinavir is an HIV protease inhibitor with anti-cancer activity through its inhibition of the Akt/mTOR pathway as well as unknown mechanisms. Because the HIV protease is an aspartic protease, we hypothesized that a target for nelfinavir in cancer cells could be γ-secretase.

Methods: Nelfinavir was compared to compound E, a commercially available GSI, in in vitro studies. Notch1 and mTOR signalling was evaluated in human and murine T-ALL cell lines by immunoblotting. γ-secretase activity was assessed in cell-free in vitro assays. Cell death was measured by flow cytometry after annexin V and propidium iodide staining. Nelfinavir was also tested in a transgenic model of T-ALL (established by overexpression of SCL and LMO1 in the mouse thymus) using microCT scans, flow cytometry and IHC.

Results: Among 7 HIV protease inhibitors evaluated, only nelfinavir inhibited expression of the Notch1 transmembrane and intracellular domains as well as Hes1 and survivin, well-established downstream targets of Notch1. At clinically achievable concentrations, nelfinavir rapidly inhibited Notch1, which was sustained to 16 hr when death was evident. Cell-free assays showed that although compound E was a more potent inhibitor of γ-secretase, the maximal enzymatic inhibition by nelfinavir was equivalent at higher doses. Nelfinavir was far more effective than compound E in reducing expression of Notch targets such as phospho-Rb, c-Myc, and Hrb, Hes1, and survivin. Nelfinavir induced apoptosis to levels equivalent to 20 µM etoposide, but compound E was only cytostatic. To explore other mechanisms that could explain the decreased protein expression and higher levels of cell death with nelfinavir, we assessed the Akt/mTOR pathway. Nelfinavir rapidly induced AMPK activation and mTOR inhibition, coincident with Notch1 inhibition. In SCL-LMO1 double transgenic mice, nelfinavir decreased thymic size and markedly increased apoptotic cells in the thymus. It also decreased the CD3+ populations in spleen and bone marrow and decreased CD4+CD8+ double positive leukemic cells.

Conclusion: These studies show that nelfinavir is superior to a GSI and highly active in preclinical studies of T-ALL. Clinical testing of nelfinavir in T-ALL patients is planned.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 2603. doi:10.1158/1538-7445.AM2011-2603
Abstract 2667: L1CAM neural adhesion molecule is overexpressed in human brain metastases of breast cancer and contributes to colony formation efficiency
Authors: Evans Lynda M. , Woditschka Stephan , Fitzgerald Daniel P. , Palmieri Diane , Badve Sunil , Sledge George W. , Steeg Patricia S.
Published: June 2012

To investigate the molecular mechanisms responsible for breast cancer metastasis to the brain, a gene expression analysis was performed using formalin-fixed, paraffin embedded sections of matched primary breast tumors and resected brain metastases. The neuronal cell adhesion molecule L1CAM was upregulated by approximately 1.57 fold in the brain metastases relative to the primary tumors (p=0.037). This trend was most apparent in Her-2+ tumors (p=0.044) and did not extend to ER+ patients (p=0.42). L1CAM has been correlated to decreased patient survival in breast cancer. MDA-MB-231 breast cancer cells selected to grow in the brain (231-BR) expressed 6.4 fold greater L1CAM on western blots than the parental cell line (231-P). Using three shRNA constructs, L1CAM expression was decreased by 1.76, 3.82, and 63.3 fold in the 231-BR cell line. Decreased L1CAM expression inhibited the clonogenic formation of 231-BR cells by 40-50% (p<0.002); however, increased proliferation by MTT assay by 30-60% (p<0.002). The most efficient L1CAM knockdown also showed a 30% decrease in migration via scratch assay relative to the non-targeting control, suggesting that L1CAM may be involved in 231-BR cell migration (p<0.03). Studies are underway to determine the effect of L1CAM down-regulation on the development of breast cancer brain metastases. These results suggest L1CAM may be a potential novel therapeutic target for metastatic breast cancer in the brain.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 2667. doi:10.1158/1538-7445.AM2011-2667
Abstract 2688: Establishment of high avidity CTL clones induced by HLA-A2 restricted glypican-3 (GPC3) peptide vaccine
Authors: Yoshikawa Toshiaki , Suzuki Shiro , Nakatsugawa Munehide , Shirakawa Hirofumi , Nobuoka Daisuke , Sakemura Noriko , Motomura Yutaka , Nakatsura Tetsuya
Published: June 2012

Glypican-3 (GPC3) is an onco-fetal antigen which is overexpressed in human hepatocellular carcinoma (HCC) and that among normal tissues it is slightly expressed in placenta and embryonic liver. Previously, we have identified a HLA-A2-restricted GPC3144-152 (FVGEFFTDV) peptide which can induce GPC3-reactive cytotoxic T lymphocytes (CTLs) without inducing autoimmunity. In the present study, we performed phase I clinical trial of HLA-A2 restricted GPC3-derived peptide vaccine to the 14 patients with advanced HCC. Immunological responses were analyzed by ex vivo IFN-γ Enzyme-linked immunospot (ELISPOT) assay. The frequency of GPC3 peptide specific CTLs after vaccinations (mean, 96; range, 5-441) was statistically significantly larger than that before vaccination (mean, 6.5; range, 0-43) (P < 0.01). An increase of the GPC3 peptide specific CTL frequency was detected in 12 (86%) of 14 patients after vaccination. Additionally, there was a significant correlation between the maximum value of GPC3 peptide specific CTLs after vaccination and dose of peptide injected (P = 0.0166, r = 0.665). Moreover, we established some GPC3 peptide specific CTL clones from PBMCs of patients vaccinated with GPC3 peptide by single cell sort using Dextramer and CD107a antibody. These CTL clones were high avidity CTLs (10−10M and 10−11M) and could recognize HCC cell lines expressing GPC3 in a HLA-class I-restricted manner. These results suggest that tumor-cytolytic T cells are indeed elicited in patients after vaccination. Now we investigate whether the GPC3 based immunotherapy have possibility of the application for other GPC3 expressing cancers including melanoma, clear cell carcinoma of the ovary, pediatric cancer or not, using established GPC3 peptide specific CTL clone.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 2688. doi:10.1158/1538-7445.AM2011-2688
Abstract 2712: Inflammation in the background cirrhotic tissue might accelerate the promotion in the HCC development from HCV-associated cirrhosis
Authors: Tarao Kazuo , Miyakawa Kaoru , Endoh Yuichi , Kobayashi Satoshi , Ueno Makoto , Ohkawa Shinichi , Morinaga Soichiro , Nakamura Yoshiyasu , Miyagi Yohei , Kameda Yoichi , Okudaira Masahiko
Published: June 2012

Aim: Hamada et al. succeeded in converting ER-1, a weakly tumorigenic cloned cell line of a rat mammary carcinoma, into a highly tumorigenic and metastatic cell line by interaction with host cell reactive to a foreign body (plastic plates). They attributed these phenomenon to the cytokines such as LL-6 derived from host reactive inflammatory cells crowded around the foreign body. It is possible that the same mechanism is involved in the development of human HCC, and that inflammation in the background cirrhotic tissue might accelerate the process of promotion. Methods: Curative hepatic resection for HCC with LC was performed in 223 patients during the past 8 years in Kanagawa Cancer Center hospital. Of 223 patients 182 patients were anti-HCV-positive or HCV-RNA positive without habitual alcohol intake. Out of 182 patients, 32 patents whose HCC revealed only one small HCC nodule of less than 3 centimeter in diameter were selected for this study. For the evaluation of inflammation in the background LC, modified Knodell's histology activity index (HAI) was used. (for the evaluation of pure inflammation, items of fibrosis was omitted); piece meal necrosis (1-4), intra lobular cellular degeneration and focal necrosis (1-4), portal cellular inflammation (1-4), were estimated by two well experienced pathologist. In each case, the HAI score was estimated in five field and average HAI score was calculated. All the sections of resected HCC tissue was examined pathologically in all patients for determining the grade of malignancy according to Edmondson Steiner's classification; Ed I (well differentiated), Ed II (moderately diff.), Ed III (poorly diff.) Results: The average HAI score in the non-cancerous cirrhotic portion in HCC patients with Ed III was 5.50 ± 0.98 (mean ± SD), and was significantly larger than that in HCC patients without Ed III (4.18 ± 0.70, p < 0.001).

Furthermore, the occurence rate of HCC containing Ed III component in the HCC patients whose HAI score was more than 5.0 was as high as 88.9% (16 out of 18), and was highly significant as compared with those in HCC patients whose HAI score was less than 5.0% (3 out of 14, 21.4%, p < 0.005).Conclusions: It is suggested that inflammation in the surrounding non-cancerous cirrhotic tissue would accelerate the process of promotion in the HCC development.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 2712. doi:10.1158/1538-7445.AM2011-2712
Abstract 2730: Role of Stat1 in skin tumor promotion
Authors: Bozeman Ronald , DiGiovanni John , Abel Erika
Published: June 2012

Inflammation is a key component of cancer development in many tissues. Emerging clinical observations support the idea of a strong association of chronic inflammation and cancer. The JAK-STAT pathway is one of the major signaling pathways involved in modulating both pro- and anti-inflammatory responses. Aberrant expression of STATs has been reported in multiple human cancers and in animal models of tumorigenesis. Previous work performed in our lab demonstrated a role for Stat1 in mediating the promotion stage of epithelial multi-stage carcinogenesis in mouse skin when chrysarobin was used as the promoting agent. To further investigate the role of Stat1 in chrysarobin-mediated skin tumor promotion, Stat1 knockout (KO) mice and wild-type controls were treated with chrysarobin and the inflammatory response was assessed. Stat1 deficiency caused a reduction in the dermal inflammatory response following a single topical application of chrysarobin. In this regard, Stat1 KO mice had reduced dermal immune cell influx following chrysarobin treatment. In particular, Stat1 KO mice exhibited reduced influx of macrophages following a single application. Other results also suggest that Stat1 may regulate various cytokine/chemokines involved in the inflammatory response following chrysarobin treatment. Mice deficient in Stat1 displayed an attenuated induction of Cox-2 mRNA and protein compared to wild-type controls following a single application of chrysarobin (200 nmol per mouse). This latter data suggest that Stat1 may be involved in the regulation of Cox-2 in keratinocytes. Collectively, these findings suggest that Stat1 may contribute to chrysarobin-mediated skin tumor promotion by regulating, in part, levels of Cox-2, PGE2 and ultimately skin inflammation. These and other ongoing studies aimed at identifying the mechanistic basis for the role of Stat1 in chrysarobin-mediated tumor promotion will be presented.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 2730. doi:10.1158/1538-7445.AM2011-2730