bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 2188: Effect of the protein complex formation of a novel tumor suppressor HLJ1 and nuclear protein p54nrb on human lung adenocarcinoma cell migration and invasion
Authors: Lin Sheng-Yi , Chen Ching-Hsien , Yu Sung-Liang , Wang Chi-Chung , Chang Gee-Chen , Chen Jeremy J. W.
Published: June 2012

Lung cancer is the most common cause of cancer deaths in the world, and the incidence is increasing each year. In the previous studies, we have identified and characterized a novel tumor/invasion suppressor gene HLJ1 (human liver DnaJ-like protein, also called DNAJB4) in a lung cancer cell line model with different invasion capability. However, the suppressive mechanism of HLJ1 is unclear. Therefore, the objective of this study is to investigate and characterize the proteins interacting with HLJ1. To obtain the novel HLJ1-binding protein, the differentially cultured density of tumor cell is employed to mimic metastatic characteristics in vitro. We identified two proteins, p54nrb and PSF, that can interact with HLJ1 in high cell culture density by immunoprecipitation assay and MALDI-TOF analysis. The complex of HLJ1 and p54nrb was confirmed in vitro and in vivo by co-immunoprecipitation, GST pull-down and mammalian two-hybrid assay. In addition, the results of GST pull-down assay indicated that p54nrb can interact with HLJ1 through HTH motif. We also found that the expressions of both HLJ1 and p54nrb are increased in A549, CL1-0, CL1-5 and CL1-5F4 cancer cell lines while are culturing in high cell density. The results of immunofluorescent staining and cytosolic/nuclear protein fraction revealed that p54nrb is mainly located in the nucleus. Furthermore, we found that over-expression of p54nrb protein in the highly invasive lung cancer cell line, CL1-5, can reduce cancer cell proliferation, colony formation, invasion and migration. On the other hand, p54nrb was decreased when most cells are in DNA synthesis phase. Once over-expression of p54nrb in CL1-5 cell line, the cell cycle would arrest in G1 phase to reduce cancer cell proliferation. Investigating the action mechanism of HLJ1-p54nrb interaction is important, it may not only provide a new insight to understand the pathway of metastasis and growth regulation but also be as the new target molecule for anti-cancer therapy in the future.

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 2188. doi:10.1158/1538-7445.AM2011-2188
Abstract 2193: Application of a novel cell-based dual luciferase assay to identify inducers of the tumor suppressor RhoB
Authors: Saber Barbara , Zhang Chunyu , Wilson WIllie , Gills Joell , Dennis Phillip
Published: June 2012

RhoB is a small GTPase tumor suppressor whose expression is commonly repressed during tumorigenesis by mechanisms such as Akt activation, HDAC1-mediated histone modification and/or specific microRNAs such as miR-21 overexpression. Forced expression of RhoB inhibits cell proliferation, survival, invasion and metastasis. Because RhoB is rarely mutated or deleted in human tumors, its restoration may have therapeutic benefit. Therefore, a dual-luciferase high throughput assay was developed to monitor induction of RhoB transcription. Four different cancer cell lines were stably transfected with a single vector in which firefly luciferase and a fusion protein of renilla luciferase/neomycin resistant gene were driven by RhoB or CMV promoters, respectively. These cell lines were screened in 96-well format with the NCI Diversity Set II library (1364 compounds). Induction of RhoB activity was compared to that caused by a known RhoB inducer (NSC126188), an HDAC inhibitor (TSA), a farnesyltransferase inhibitor (L744832), and an Akt inhibitor (PIA23). Seven compounds were identified, and induction of RhoB transcription and translation was validated under different serum conditions at several time points. They were also screened in secondary assays that included inhibition of Akt, induction of protein acetylation, and screening in a cancer cell line that cannot induce endogenous RhoB due to a homozygous deletion of chromosome region 2p24. One compound caused sustained induction of endogenous RhoB in multiple cell lines under different conditions, and inhibited cellular proliferation and migration. These studies show that this assay system can identify specific RhoB inducers with potential anticancer activity, and suggest that it can be expanded to screen larger compound libraries, or be adapted to monitor transcriptional activation of other tumor suppressor genes.

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 2193. doi:10.1158/1538-7445.AM2011-2193
Abstract 2198: Role of SLC5A8, a plasma membrane transporter and a tumor suppressor, in the antitumor activity of dichloroacetate
Authors: Babu Ellappan , Thangaraju Muthusamy , Ganapathy Vadivel
Published: June 2012

Introduction: There has been growing interest among the public and scientists in dichloroacetate as a potential anticancer drug. Credible evidence exists for the antitumor activity of this compound, but high concentrations are needed for significant therapeutic effect. Unfortunately, these high concentrations produce detrimental side effects involving nervous system, hence precluding its use for cancer treatment. The mechanistic basis of the compound's antitumor activity is its ability to activate pyruvate dehydrogenase complex through inhibition of pyruvate dehydrogenase kinase. Since the compound inhibits the kinase at micromolar concentrations, it is not known why therapeutically prohibitive high doses are needed for suppression of tumor growth. We hypothesized that lack of effective mechanisms for the entry of dichloroacetate into tumor cells may underlie this phenomenon.

Methods: The transport of acetate and its chloro derivatives by human SLC5A8 was studied using the X. laevis oocytes expression system. The human transporter was expressed in oocytes heterologously by injection of SLC5A8 cRNA. The transport function was monitored electrophysiologically by the two-microelectrode voltage-clamp technique. Breast, colon and prostate cancer cell lines were transfected with SLC5A8 and expression was monitored by RT-PCR analysis. Intracellular levels of pyruvate and HDAC activity was also monitored in these cell lines.

Results: SLC5A8 transports dichloroacetate very effectively and with high affinity. This transporter is expressed in normal cells, but the expression is silenced in tumor cells via epigenetic mechanisms. The lack of the transporter makes tumor cells resistant to the antitumor activity of dichloroacetate. However, if the transporter is expressed in tumor cells ectopically, the cells become sensitive to the drug at low concentrations. This is evident in breast cancer cells, colon cancer cells, and prostate cancer cells. Normal cells, which constitutively express the transporter, are however not affected by the compound, indicating the tumor cell-selective therapeutic activity. The mechanism of the antitumor activity of the compound depends on its ability to inhibit pyruvate dehydrogenase kinase and force mitochondrial oxidation of pyruvate.

Conclusions: The silencing of SLC5A8 in tumors involves DNA methylation and its expression can be induced by treatment with DNA methylation inhibitors. Our findings suggest that combining dichloroacetate with a DNA methylation inhibitor would offer a means to reduce the doses of dichloroacetate to avoid detrimental effects associated with high doses but without compromising antitumor activity.

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 2198. doi:10.1158/1538-7445.AM2011-2198
Abstract 2224: EGFR mutation screening in non-small cell lung cancer: Results from 4 German molecular pathology laboratories
Authors: Hartmann Arndt , Stöhr Robert , Gschwendtner Andreas , Geissinger Eva , Gattenlöhner Stefan , Glück Thomas , Rümmele Petra , Dietmaier Wolfgang
Published: June 2012

Aims: Somatic mutations in the epidermal growth factor receptor gene (EGFR) have evolved as predictors for sensitivity to EGFR tyrosine kinase inhibitors (TKI) in patients with non-small-cell lung cancer (NSCLC). To date, molecular data underlining the role of EGFR mutations in the tumor as a strong predictor for a better treatment outcome were mostly collected from Asian patients (mutation frequency up to 59.7%). Only one European study (Rosell et al., 2009) verified the predicting power of EGFR mutations in Caucasians living in Europe so far, and reported a notably lower mutation frequency (16.6%) compared to Asian collectives. For Germany, only sparse data are available about the mutation frequency in NSCLC. Here we report the intermediate results from 4 pathology institutes performing quality approved approved EGFR testing certified by the Quality Initiative of the German Society of pathology (QuiP) in non-small cell lung cancer.

Material and Methods: Overall, 787 (males: 474, females: 312, n.a.: 1) cases were analysed. The cohort consisted of 782 NSCLC (99.4%) and 5 SCLC (0.6%) selected by attending physicians for EGFR mutation analysis considering TKI treatment. After microdissection, DNA was isolated from serial sections of formalin-fixed, paraffin-embedded tumor tissue after microdissection to obtain at least 70% tumor cells. Exons 18, 19 and 21 of the EGFR gene were analysed using Sanger sequencing.

Results: In 72/787 cases (9.2%) an EGFR mutation was detected. Deletions in exon 19 were the most frequent alterations detected (46/72 mutations; 63.9%), followed by point mutations in exon 21 (23/72 mutations; 31.9%). Mutation in exon 18 was a rare event (2/72 mutations; 2.8%). EGFR mutations were significantly associated with adenocarcinomas (p<0.001) (mutation frequency: 11.4%) and female gender (p<0.001). Overall, there was a trend (p=0.052) towards higher age in patients with adenocarcinoma and EGFR mutation compared to cases with EGFR wildtype. This increased age was significantly (p=0.009) associated with an EGFR mutation in female patients.

Conclusion: EGFR mutation frequency in this German cohort is lower than reported in Asian or Spanish studies. Mutations in exon 19 were detected more frequently than described in the literature so far. The association with adenocarcinoma histology and female gender could be verified. EGFR mutations seem to be more frequent in patients with higher age. Screening of patients with advanced NSCLC for EGFR mutations is feasible and should be used for improvement of therapy options.

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 2224. doi:10.1158/1538-7445.AM2011-2224
Abstract 2239: Buthionine sulfoximine enhanced melphalan cytotoxic activity against multiple myeloma cell lines
Authors: Tagde Ashujit , Reynolds C Patrick
Published: June 2012

Multiple myeloma (MM) is a neoplasm of terminally differentiated antibody secreting B cells commonly treated with myeloablative doses of the alkylating agent melphalan (L-PAM) and supported by stem cell transplantation (SCT). Relapse after SCT is common and a second SCT with L-PAM for patients achieving less than a complete response or at time of relapse is usually only modestly effective. Glutathione (GSH) is ubiquitous intracellular tripeptide that acts as antioxidant and protects cells against reactive oxygen species (ROS) and xenobiotics. Conjugation of GSH to alkylating agents is known to be one mechanism of melphalan resistance. Buthionine sulfoximine (BSO) inhibits synthesis of GSH and enhances the activity of melphalan in preclinical and clinical studies of neuroblastoma. The potent hematopoietic toxicity of BSO + L-PAM led us to conduct an ongoing phase I trial in recurrent neuroblastoma of escalating L-PAM + BSO given with SCT. To explore the potential for a similar trial in MM, we have undertaken preclinical studies of BSO + L-PAM in MM.

We determined if buthionine sulfoximine (BSO) enhanced the activity of melphalan (L-PAM) against MM cell lines. In-vitro cytotoxicity mediated by BSO (0-400 micro molar) alone and in combination with L-PAM (0-50 micro molar) was assessed in a panel of six MM cell lines in both “standard cell culture conditions” (20% O2) and “bone marrow level hypoxia” (5% O2) using the DIMSCAN digital image fluorescence assay. Single-strand DNA (ssDNA) damage by F7-26 antibody, apoptosis by TdT labeling (TUNEL) and mitochondrial depolarization by JC-1 were measured by flow cytometry.

At clinically achievable concentrations, BSO consistently caused two to four logs synergistic enhancement of L-PAM cytotoxicity (combination index < 1) in all six multiple myeloma cell lines at both 5% and 20% O2. BSO + L-PAM caused a significant increase in single strand DNA damage (p<0.001), mitochondrial depolarization (p<0.001), and apoptosis (p<0.001) compared with L-PAM alone. Thus BSO can enhance L-PAM activity and overcome L-PAM resistance in multiple myeloma cell lines and BSO/L-PAM synergy is also achieved in bone marrow level hypoxia. Obtaining additional data on this novel combination approach to therapy of multiple myeloma is warranted.

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 2239. doi:10.1158/1538-7445.AM2011-2239
Abstract 2257: The survival impact of XPA and XPC genetic polymorphisms on patients with esophageal cancer
Authors: Lee Jang-Ming , Yang Pei-Wen , Hsieh Ching-Yueh , Tsai Pey-Yu , Chen Chun-Hui , Chen Jin-Shing , Huang Pei-Ming , Hsu Hsao-Hsun , Kuo Sunn-Wen , Lee Yung-Chie
Published: June 2012

Purpose: To investigate the association of the genetic variants in xeroderma pigmentosum groups A (XPA) and C (XPC) with the overall survival duration of the patients with esophageal cancer.

Patients and Methods: A total of 531 patients with a diagnosis of esophageal cancer were enrolled in the study. The genetic variants of XPA in 5’UTR (rs1800975) as well as XPC at exon 15 Lys939Gln (rs2228001) were analyzed with the TaqMan assay from the genomic DNA of peripheral leukocytes. The association of the genotypes and the survival of the patients were analyzed with Cox's multivariate analysis.

Results: There was no significant difference of XPA and XPC genotype distribution among patients with various clinical profiles. The overall survival of the patients was significantly influenced by the tumor stage, cell type (squamous cell carcinoma vs. adenocarcinoma) and curative surgical resection (p<0.05 respectively). There was an insignificant unfavorable trend for survival outcome of the patients with XPA 5’UTR (rs1800975) C/T or XPC Lys939Gln (rs2228001) G/G genotypes, compared with those who with other genotypes. Cox's multivariate analysis revealed that accumulation with any one this two unfavorable genotype would associate with a significantly increased risk of death (adjusted HR [95% CI]: 1.27[1.07-1.50]; p=0.005). Compared to those patients without any of these unfavorable genotype, simultaneously having both of these genotype would impose a significantly higher risk of death on the patients after treatment (adjusted HR [95% CI]: 1.86[1.24-2.81]; p=0.003).

Conclusion: Our study demonstrates that the hereditary genetic variants in XPA and XPC may play a role for the clinical outcome of the patients with esophageal 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 2257. doi:10.1158/1538-7445.AM2011-2257
Abstract 2279: The response of breast cancer cell lines to HET0016 alone or in combination with other chemotherapeutics
Authors: Slaughter Tiffani , Gunter Barak W. , Fan Fan , Huang Chenghui , Pannuti Antonio , Baker Rodney C. , Falck John R. , Miele Lucio , Roman Richard J. , Duhe Roy J.
Published: June 2012

One of the vexing problems in controlling breast cancer is the emergence of acquired endocrine-resistance in estrogen receptor positive breast cancers that initially respond to endocrine therapy. Published reports suggest there is a close overlap between the signal transduction pathways affected by 20-hydroxyeicosatetraenoic acid (20-HETE) and those which may confer endocrine-resistance. Preliminary data indicate that 20-HETE-targeted drugs have direct cytotoxic and indirect anti-angiogenic effects on multiple carcinomas, and they therefore might be extended to unsolved problems in breast cancer control. Yet almost nothing is known about the specific role(s) of 20-HETE in either healthy breast tissue or in breast cancer. We observed that treatment of certain tamoxifen-resistant human breast cancer cell lines with HET0016, a compound which inhibits the biosynthesis of 20-HETE, restores sensitivity to tamoxifen in cell culture. We are elucidating the signal transduction pathways involved in this phenomenon. We also observed that certain human breast cancer cell lines with intrinsic tamoxifen resistance (such as the “triple-negative” MDA-MB-468 cell line) express cytochrome P450 isoforms essential for the synthesis of 20-HETE, and we are evaluating whether we can exploit a synergistic effect of HET0016 with anti-cancer drugs currently used to treat aggressive “triple-negative” breast cancers which have an intrinsic resistance to endocrine therapy. We hypothesize that a 20-HETE-targeted agent such as HET0016 will provide an additional anti-cancer benefit due to its anti-angiogenic activities, and we are further exploring the characteristics of breast cancers that may be susceptible to combination therapy involving HET0016 and related compounds.

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 2279. doi:10.1158/1538-7445.AM2011-2279
Abstract 2292: Absent E2 liganded AhR inhibits tumor inhibition by ERα in Min mice
Authors: Carothers Adelaide M. , Rizvi Hira , Lubet Ronald , Bertagnolli Monica
Published: June 2012

Previously, we showed that E2 and ERα prevent intestinal tumor formation in the Min mouse. Others showed that Aryl hydrocarbon Receptor (AhR) associates with ubiquitin ligase to down-regulate β-catenin and Wnt signaling. AhR activity also may affect ERα. AhR has endogenous, dietary, and xenobiotic ligands. We hypothesized that the tumor inhibiting function of ERα promotes Wnt signaling in vivo and is antagonized by liganded-AhR in the absence of E2. In groups of 10, ovariectomized Min (OvxMin) mice were treated w/wo AhR ligand 5, 6-benzoflavone (BF; 2,000 ppm) by diet for 17 weeks. Treated mice developed more tumors in the small intestine (54.7±11.1) than controls (38.3±4.5 P<0.0001). Protein expression was evaluated in non-tumor ileum of study mice by immunohistochemistry, immunoprecipitation (IP), and immunoblotting (IB). ERα and β-catenin nuclear expression in crypt base cells, total β-catenin and ERα protein amounts, and Wnt signaling-dependent Paneth cell markers (Sox9 and Lysozyme) were significantly reduced in BF-treated OvxMin compared to control mice. Reciprocal IP/IBs using total cell lysates from study mice and antibodies for β-catenin and ubiquitin (Ub) showed increased expression of ubiquitylated β-catenin in BF-treated OvxMin mice relative to controls. Wnt signaling-dependent cell proliferation (Ki67+ nuclei/crypt) was reduced in BF-treated OvxMin (11.4±2.5) relative to controls (16.2±3.0) and untreated intact Min (20.7±3.2) mice (P<0.01). AhR was expressed throughout the crypt-villus unit of OvxMin ileum; its activity was assessed by induction of Cyp1A1. A minimal amount of Cyp1A1, but elevated levels were found in untreated mucosa of intact Min vs. OvxMin mice w/wo BF, respectively. Lower expression of the AhR negative regulator, AhR Repressor, was present in ileum of OvxMin mice regardless of treatment relative to untreated intact Min mice. In study mice mucosa, other xenobiotic responsive regulators (Pxr, Nrf2, Car1/2) were minimally expressed, not affected by BF, hence not implicated in Cyp1A1 induction. In conclusion, these results indicate that E2 loss reduces AhR feedback inhibition and Wnt signaling in Min intestine. Since liganded-AhR promoted β-catenin turnover and reduced ERα expression absent E2, our data suggest that deficient Wnt signaling in the non-tumor mucosa of Min mice provides a selective pressure for Apc functional loss in intestinal stem cells and tumor initiation.

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 2292. doi:10.1158/1538-7445.AM2011-2292
Abstract 2300: Insulin receptor kinase inhibition is better tolerated than hypoinsulinemia and more effective than metformin in treating breast cancer
Authors: Dool Carly , Mashhedi Haider , Zakikhani Mahvash , David Stephanie , Zhao Yunhua , Birman Elena , Carboni Joan M. , Gottardis Marco , Blouin Marie-Jose , Pollak Michael N.
Published: June 2012

Epidemiologic and experimental evidence suggest that a subset of breast cancer is insulin-responsive, but it is unclear if safe and effective therapies that target the insulin receptor can be developed. We demonstrate that both insulin receptor family tyrosine kinase inhibition and insulin deficiency have anti-neoplastic activity in a model of insulin-responsive breast cancer in mice metabolically normal at baseline. In contrast to insulin deficiency, insulin receptor inhibition does not lead to hyperglycemia and is well-tolerated. We show that pharmacokinetic factors explain the safety of receptor inhibition relative to ligand deficiency, as BMS-536924 does not accumulate in muscle at levels sufficient to block insulin-stimulated glucose uptake. Metformin, which lowers the elevated insulin levels present in settings of insulin resistance, had minimal activity in this model. The findings highlight the importance of tissue-specific drug accumulation as a determinant of efficacy and toxicity of tyrosine kinase inhibitors, and suggest that therapeutic targeting of the insulin receptor family for cancer treatment is practical.

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 2300. doi:10.1158/1538-7445.AM2011-2300
Abstract 2317: Mild hyperthermia and thermosensitive liposomes to improve chemotherapy delivery to solid tumors
Authors: Li Li , ten Hagen Timo L.M. , Schipper Debby , Seynhaeve Ann L.B. , van Rhoon Gerard , Eggermont Alexander M.M. , Lindner Lars H. , Haemmerich Dieter , Koning Gerben A.
Published: June 2012

Long circulating liposomes are potent nanocarriers for the delivery of chemotherapeutic drugs to tumors. These liposomes can extravasate through leaky tumor microvasculature, and accumulate in the intratumoral space. However, inefficient drug release impairs efficacy, and hinders a successful application. Local hyperthermia (HT) can augment drug bioavailability and treatment efficacy by enhancing tumor vessel leakage for intratumoral liposomal drug accumulation, and triggering drug release from thermosensitive liposomes (TSL). Our study explored the thermal dose to maximize the permeability of tumor microvasculature for liposome extravasation and sequentially trigger drug release from the TSL in a two-step approach.

We have optimized long-circulating TSL to improve content release efficiency under mild HT (Li et al. J Control Release 143(2):274-9, 2010). Doxorubicin (Dox) at a quenching concentration was encapsulated in TSL. In vitro temperature/time-dependent content release and TSL stability in serum were quantified by fluorometry. In vivo liposome extravasation was studied in dorsal skin flap window chamber models implanted with murine B16 melanoma, LLC carcinoma, BFS-1 sarcoma, and human BLM melanoma tumors in mice with constitutive vascular endothelial cell expression of an eNOS-Tag-GFP fusion protein using confocal microscopy. In vivo Dox release was observed in the same window chamber models.

Mild HT induced a significant amount of liposome extravasation through tumor microvasculature, and intratumoral accumulation in the interstitial space in all tumor models. Minimum thermal dose of 41 °C for 30 min is required to open up tumor vessels. The permeability of tumor vessels increased over time with increasing thermal dose to 41 °C for 1 hr, and vasculature remained leaky for up to 8 hr post-HT. Increased permeability was not observed in heated normal vasculature or in non-heated tumor vasculature. Optimized Dox-TSL were stable at 37 °C and demonstrated efficient in vitro and in vivo drug release at 42 °C. Rapid intratumoral distribution of released Dox and delivery into tumor cells was observed, demonstrating strongly improved drug bioavailability. The superior application of TSL and mild HT will achieve both maximal intratumoral liposomal drug accumulation and rapid triggered drug release to aid liposomal chemotherapy in clinical practice.

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 2317. doi:10.1158/1538-7445.AM2011-2317
Abstract 234: Fibroblast growth factor receptor (FGFR)-3 as a suitable target in bladder cancer
Authors: Gust Kilian M. , McConkey David J. , Hegarty Paul K. , Bondaruk Jolanta E. , Czerniak Bogdan A. , Dinney Colin P. , Black Peter C.
Published: June 2012

Introduction and Objective: Mutations in the FGFR3 gene, which can confer constitutive activity, are found in up to 80% of low grade papillary bladder cancers (BC). Overexpression of FGFR3 has been detected in approximately half of high grade and invasive tumors. In this study we evaluate FGFR3 in a cohort of American patients and investigate the effect of targeting FGFR3 in pre-clinical models of bladder cancer.

Materials and Methods: DNA was extracted from 170 fresh frozen BC specimens using a commercial kit and PCR of exons 7 and 10 was performed. Mutation analysis was performed by ABI Big Dye Terminator Cycle Sequencing. FGFR3 expression was analyzed by immunohistochemistry in an overlapping cohort of 143 patients with BC. FGFR3 expression, phosphorylation and downstream signaling were measured in a panel of BC cell lines, and the efficacy of two small molecule inhibitors (TKI258 and BGJ398) was tested on signaling and 3H-thymidine incorporation. A specific inhibitory monoclonal antibody targeting FGFR3 was evaluated in an orthotopic xenograft model of BC. Tumor burden was measured with bioluminescent imaging.

Results: FGFR3 mutations were identified in 44 (26%) BC samples, including 28/50 (56%) of low grade papillary lesions. Overexpression of FGFR3 was noted in 53% of all BC specimens regardless of stage and grade, but not in normal urothelium. FGFR3 inhibition effectively abrogated constitutive phosphorylation of FGFR3 and p42/44MAPK in a subset of bladder cancer cell lines. A growth inhibitory effect was observed in the same cell lines. The growth of UC14 xenografts in the bladder of nude mice was inhibited by 40%.

Conclusion: These studies underline the potential clinical relevance of FGFR3 mutation and expression in bladder cancer. Furthermore, the preclinical investigations offer proof of principle for moving forward with clinical trials studying the administration of specific FGFR3 inhibitors in patients with bladder 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 234. doi:10.1158/1538-7445.AM2011-234
Abstract 2346: Switching from an ROR1-positive phenotype to an ROR2-positive phenotype promotes melanoma progression
Authors: O'Connell Michael P. , Nelson Janelle L. , Fiori Jennifer L. , Subaran Sarah S. , Indig Fred E. , Weeraratna Ashani T.
Published: June 2012

The orphan tyrosine kinase receptors, ROR1 and ROR2, make up a superfamily of receptors that, until recently, had no known signal pathway association or ligand identification. ROR2 has since been shown to play a role in Wnt signal transduction and studies from our laboratory have demonstrated an important role for ROR2 in increasing the metastatic potential of melanoma cells. Here, we demonstrate, that, despite significant homology, ROR1 expression is inversely correlated to that of ROR2 in melanoma and decreasing ROR1 expression increases overall metastatic potential. ROR1 is expressed on the cell surface in the hydrophobic region of the cell membrane and in the cytoplasm of less metastatic melanoma cells. Treatment of less metastatic melanoma cells with recombinant Wnt5A decreased the levels of ROR1 in these cells. We have previously demonstrated that increases in Wnt5A result in an increase in metastatic capacity of melanoma cells, and a decrease in the melanoma differentiation antigens MART1 and gp100. These data have been supported by several other laboratories, giving rise to the phenotype switching model of melanoma progression that suggests that melanomas express a “proliferative” gene expression signature governed by the transcription factor MITF, and its targets MART1, gp100, DCT, etc., during the initial stages of melanoma development, and an “invasive” gene expression signature governed by Wnt5A in the later stages of melanoma progression. In keeping with these data, reduction of ROR1 by siRNA demonstrates a decrease in the melanoma differentiation markers MART1 and GP100, and motility, and an increase in the invasive capability of melanoma cells. In addition, siRNA knockdown of ROR1 expression increased WNT5A expression, implying that downregulation of ROR1 may be a step in the progression towards melanoma metastasis. Further, WNT5A knockdown in cells that have high levels of WNT5A, but low levels of ROR1, increases ROR1 expression and knockdown of ROR2 increases ROR1 expression. These data suggest that melanoma receptors in the ROR superfamily can influence the expression of one another and the switch from ROR1 positivity to ROR2 positivity can contribute to metastatic potential. The search for the micro-environmental factors that can initiate this switch is currently under investigation in our laboratory.

(This work was supported in part by the Intramural Research Program of the National Institute on Aging, Baltimore, MD).

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 2346. doi:10.1158/1538-7445.AM2011-2346
Abstract 2397: Fat soluble vitamin B1 analogue, benfotiamine, prevents colon cancer cell growth and aberrant crypt foci formation in murine models of colon cancer
Authors: Ramana Kota V. , Saxena Ashish , Tammali Ravinder , Reddy Aramati BM , Srivastava Satish K.
Published: June 2012

Chronic inflammatory diseases and oxidative stress are major risk factors of colorectal cancer (CRC), the third most common cause of death among cancer patients. Although several agents such as antioxidants, plant products and nutritional supplements have been shown to have some chemopreventive effects on colon cancer, none are very effective to completely. Recent studies indicate that a fat soluble analogue of vitamin B1, benfotiamine is an excellent antioxidant and anti-inflammatory. However, the chemo-preventive or -therapeutic efficacy of benfotiamine in preventing CRC is not known. Our hypothesis is that benfotiamine through its anti-oxidant and anti-inflammatory properties could be very effective in preventing CRC. We therefore investigated the role of benfotiamine in the prevention of carcinogenic signals leading to CRC in cell culture as well as murine models of CRC. Our results show that treatment of human colon cancer cells (HT-29 and Caco-2) in culture with benfotiamine prevented the cancer cell proliferation. Further, benfotiamine also prevented the growth of human adenocarcinoma cells (SW480) -induced tumor growth in nude mice xenografts. Our studies also indicate that benfotiamine prevented NF-kB-dependent survival signals and activated caspase-3 dependent apoptotic signals in colon cancer cells. Further, benfotiamine supplementation also suppressed azoxymethane (AOM)-induced aberrant crypt foci (ACF) formation in mice and AOM-induced inflammatory changes such as activation of Cox-2 and iNOS, and carcinogenic changes such as expression of cyclin D1 and β-catenin in mice colons. In conclusion, our results indicate that benfotiamine prevents colon cancer cell growth in culture and nude mice xenografts as well as formation of ACF in AOM-treated mice. Thus benfotiamine supplementation could be used as an excellent chemopreventive agent for the treatment of CRC.

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 2397. doi:10.1158/1538-7445.AM2011-2397
Abstract 2405: Identification of PHLPP as a tumour suppressor reveals the role of pathway feedback compensation in PTEN-mutant prostate cancer progression
Authors: Chen Muhan , Pratt Christopher P. , Zeeman Martha E. , Schultz Nicholaus , Taylor Barry S. , O'Neill Audrey , Castillo-Martin Mireia , Nowak Dawid G. , Naguib Adam , Grace Danielle M. , Murn Jernej , Sander Chris , Gerald William L. , Cordon-Cardo Carlos , Newton Alexandra C. , Carver Brett S. , Trotman Lloyd C.
Published: June 2012

Hyper-activation of the PI 3-Kinase/ AKT pathway is common in many cancer types. Tumourigenesis through this pathway is prevented by concerted action of multiple tumour suppressor genes. Most notably, PTEN reverts PI 3-Kinase activity whereas excessive pathway activation triggers the p53-mediated senescence arrest. However, it remains ill defined if and at what stage this response acts in human prostate cancer. Here we identify the AKT-inactivating phosphatase PHLPP as a tumour suppressor and demonstrate how the p53-response can antagonise co-deletion of PTEN and PHLPP to form a barrier against prostate cancer progression. We show that Phlpp-loss causes neoplasia and upon partial Pten-loss, carcinoma in mouse prostate. In this setting, Phlpp-deficiency triggers growth arrest via mTorC1-dependent activation of p53 and we find that co-deletion of Pten and Phlpp selects for spontaneous inactivation of p53 in prostate. Validating this conditional gene inactivation scheme in a comprehensive genomic patient data set we find that co-deletion of PTEN and PHLPP is almost exclusively observed in metastatic prostate cancer and tightly correlated to deletion of TP53. Furthermore, PTEN/ PHLPP expression can be used to predict disease outcome in these patients, comparable to the standard histology based method, but adding actionable information on pathway status. Finally, we show that both known PHLPP isoforms compensate for PTEN-suppression in a novel pathway feedback explaining their co-deletion with PTEN in the metastatic samples. Surprisingly, we find that the feedback surge of these genes is sensitive to some pharmacological inhibitors of the PI 3-Kinase pathway. Collectively, our findings emphasise the need for careful evaluation of PI 3-Kinase target therapy effects in prostate cancer and highlight the value of genetically engineered mouse models in this process.

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 2405. doi:10.1158/1538-7445.AM2011-2405
Abstract 2411: Hurp deficiency causes mitotic infidelity and impairs liver regeneration in mice
Authors: Tsai Ting-Fen , Huang Chiao-Ming , Li Chao-Chin , Kao Cheng-Heng
Published: June 2012

Aberrant expression of hepatoma up-regulated protein, HURP, was found in hepatocellular carcinoma (HCC). Recent studies revealed that HURP is an essential factor during mitosis. However, the function of HURP in an in vivo context has not been well studied. In this study, we have concentrated our efforts on the pathological effects of Hurp deficiency during liver regeneration. We generated Hurp knockout (KO) mice to investigate the function of Hurp involved in hepatocyte cell cycle progression; we used the in vivo model of two-third partial-hepatectomy (PH) to induce synchronous cell cycle entry in liver. Our results revealed striking phenotypes of mitotic errors and chromosome instability in the Hurp KO livers. These phenotypes include impaired cell cycle progression as revealed by delayed entry of S-phase and M-phase of hepatocytes post-PH; mitotic infidelity as revealed by lagging chromosomes, multi-polar segregation and unequal segregation; aberrant nuclei of micro-nucleus, multi-nucleus and atypical nuclear morphology were frequently observed in the Hurp KO livers. Mechanistically, these abnormalities caused by Hurp deficiency did not turn on the spindle checkpoint machinery, such as BubR1 and Mad2, but up-regulated p21 expression to delay the cell cycle progression in liver. As an evidence of chronic liver damage, we observed continuously elevated serum ALT levels in Hurp KO mice post-PH. Together, these phenotypes in liver indicated that Hurp deficiency interferes spindle checkpoint (directly or indirectly), leading to mitotic infidelities, and possibly causes DNA breakage during chromatin segregation; this DNA breakage can further induce p21 to delay cell cycle during liver regeneration which is accompanying by chronic liver damages. All of these abnormalities may further lead to enhanced carcinogenesis in liver, which is currently under investigation.

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 2411. doi:10.1158/1538-7445.AM2011-2411
Abstract 2412: Molecular characterization of hepatocellular carcinoma (HCC) patient-derived explant models
Authors: Zhang Lan , Zhang Jingchuan , Xie Liang , Xie Xiaoying , Guo Qiuli , Lv Jing , Gao Zeren , Qian Ziliang , Yin Xiaolu , Zheng Li , Zhu Guanshan , Ji Qunsheng , Ren Zhenggang
Published: June 2012

Recent evidence suggested that cancer patient-derived explant mouse models can maintain certain pathological and molecular features of the original diseases/patients, which offers potential opportunities for testing “personalized medicine” preclinically. We have established over 20 HCC explant models by directly implanting fresh tumor tissues derived from HCC patients into immunodeficient mice. To assess whether these HCC models would represent HCC clinical characteristics and individual patient diversity, we have compared the molecular profiles of these xenografts with their corresponding patient tissues (F0). This included gene copy variation by aCGH, mRNA expression by Affymetrix and gene mutations as well as protein expressions by immunohistochemistry. In addition, we also assessed the stability of the molecular profiles of various models by comparing different passages (F1-F10). Analysis of the data revealed that the xenograft tumors maintain fundamental genotypic features of the original patients, which recapitulate the genetic diversity of the HCC. The assessment on the stability of molecular profiles of individual model based on Pearson correlation coefficient (r) of aCGH data from different passages (F1-F10) suggests that those HCC models are very stable across passages (r > 0.8). In addition, no dramatic changes of the known molecular and histological features were observed throughout serial passages. AFP mRNA expressions of various models by Affymetrix are very different and mirror patient clinical data very well. Amplification of myc and cycline D1 and deletion of DLC1 and FHIT were detected in different individual models, as well as in corresponding patient HCC tissues, and those aberrations are consistent throughout the passages within various individual model. Furthermore, passages of the tumors in successive cohorts of mice did not change their sensitivity to sorafenib. In summary, we have developed preclinical HCC patient-derived explant models that can be used to further understand the disease and develop mechanism-based anti-cancer drugs for treatment of HCC patients.

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 2412. doi:10.1158/1538-7445.AM2011-2412
Abstract 2414: In vitro profiling of gastric and liver cancer cell lines developed from Asian cancer patients
Authors: Kelter Gerhard , Maier Armin , Krumbach Rebekka , Schüler Julia , Fiebig Heinz-Herbert , Beckers Thomas
Published: June 2012

In East Asia stomach and liver cancer are the 2nd and 3rd most common malignancies and there is a high medical need to develop new drugs. However, well characterized in vitro and in vivo models developed from Asian cancer patients are rare. The Oncotest tumor xenograft and cell line repository used for profiling of anticancer compounds comprised mainly models established from patients with a Caucasian origin. Now the Oncotest tumor repository was extended by 8 gastric and 12 liver cancer cell lines of Asian origin obtained from KCLB (Korean Cell Line Bank) and JCRB (Japanese Collection of Research Bioresources). Additionally, 2 head & neck cancer cell lines from KCLB and the gastric cell line GXA 3011L established at Oncotest from a Korean patient-derived tumor xenograft were added to the panel.

Among these 23 cell lines, 6 currently grow subcutaneously as solid tumor xenografts in nude mice. Gene expression profiles (Affymetrix chip HG-U133 plus 2.0 consisting of 38,500 genes) have been determined for all cell lines and respective xenografts. Doubling times as determined for all cell lines were in the range of 23 h (MKN45) to 76 h (SNU-423).

In vitro chemosensitivity to standard of care (e.g. 5-FU, doxorubicin, oxaliplatin, paclitaxel) and selected targeted drugs (everolimus, lapatinib) was assessed by a fluorescence based monolayer cytotoxicity assay as well as by the ex-vivo clonogenic assay. The drugs showed diverse patterns of selectivity and potency in both assays. As an example, the mTor inhibitor everolimus exhibited pronounced activity with IC50 values in the low nanomolar range towards the liver cancer cell lines SNU-398, SNU-475, SNU-878 and SNU-886 in the monolayer as well as in the clonogenic assay. In contrast, towards gastric cancer cell lines no or only marginal activity was detected for everolimus in the monolayer assay. The HER2 / EGFR kinase selective inhibitor lapatinib showed selective activity towards the gastric cancer cell line NUGC-4 (IC50=0.16µM), the liver cancer cell line JHH-4 (IC50=0.56µM) and the H&N cancer cell line SNU-1076 (IC50=0.54µM). Overall, in vitro activity profiles for 12 SOC drugs and correlations with molecular alterations, target expression and pathway activation will be presented.

This collection of cell lines is of high relevance for the development of drugs against the most widely occurring cancers in East Asia, by providing models for the study of the biology and response to novel anticancer drugs.

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 2414. doi:10.1158/1538-7445.AM2011-2414
Abstract 2422: Epithelial interleukin-4 receptor promotes metastasis of mammary tumor cells
Authors: Hwang Daniel G. , Dozier Ashley , Carter Kathy J. , Fingleton Barbara
Published: June 2012

Interleukin-4 (IL4) is known primarily for its immune system functions where it can stimulate lymphocytes and promote macrophage activation. Various cancer cells, including breast, are now known to express functionally active IL4 receptors, which appear to play a role in cell survival. We believe that blocking IL4 signaling specifically in epithelial cells will attenuate metastasis because of reduced cell survival in secondary sites. To test this hypothesis, we have generated multiple clones of two murine mammary tumor cell lines that show efficient knockdown of an essential component of the IL4 receptor, IL4-receptor [IL4R] alpha. Initial characterization of knockdown and control clones showed a significantly slower growth rate in vitro as assessed both by metabolic (MTT) assays and direct cell counting. Flow cytometric analysis using either annexin V staining or sub G1 measurement showed that clones lacking IL4Ralpha had 2-fold higher basal levels of apoptosis. To test effects in vivo, we have performed experimental and spontaneous metastasis assays using clones of the 4T1 cell line. Both pulmonary and hepatic metastatic lesions were significantly decreased [2.4-fold lower] in mice injected with IL4Ralpha knockdown clones compared to controls. There was however no significant effect on the primary mammary tumors. Together these results suggest that IL4 signaling in epithelial tumor cells is pro-metastatic, and blocking it may be a useful therapeutic approach for preventing metastatic spread.

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 2422. doi:10.1158/1538-7445.AM2011-2422