bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 1020: VRC2, a novel bortezomib re-sensitizing compound for the treatment of multiple myeloma.
Authors: Lulla Amriti , Stessman Holly , Dicker David , Van Ness Brian , El-Deiry Wafik , Dolloff Nathan G.
Published: August 2014

The development of novel agents including the proteasome inhibitor bortezomib (Btz) has significantly improved treatment outcomes in multiple myeloma (MM). Despite these advances, resistance to therapy invariably emerges and MM remains an incurable disease. The objective of this study was to identify new therapeutic agents and combinations with the ability to overcome resistance to Btz in MM cells. We developed a high throughput drug screening (HTS) assay system to discover small molecules that selectively target Btz resistant MM cells as single agents and/or re-sensitize resistant cells to Btz. For HTS we used isogenic pairs of Btz sensitive (595 P) and resistant (595 VR) cells derived from iMycCα/Bcl-xL transgenic mouse model of MM. Cell viability was the HTS assay read-out, and 3 treatment groups (595 P, 595 VR, and 595 VR + Btz) were included to identify compounds with selective activity against Btz resistant cells as single agents, and compounds that restore Btz sensitivity in the presence of Btz. Our screening of multiple publically-available compound libraries identified several chemical structures with selective activity against Btz resistant cells. One compound in particular showed modest but significant selectivity for Btz resistant cells as a single agent, and most notably, exhibited potent Btz re-sensitizing activity when the 2 drugs were combined. We named this compound Velcade Re-sensitizing Compound 2, or VRC-2 for short. The Btz re-sensitizing activity of VRC-2 was confirmed using multiple human and mouse cell lines that had been selected for Btz resistance in vitro. IC50s for VRC-2 in normal mouse and human fibroblasts were 100-fold higher than that observed in MM cells, suggesting it may have a low toxicity profile in vivo. We are currently evaluating the activity of VRC-2 in animal models of MM, alone and in combination with Btz, and conducting molecular studies to identify its precise mechanism of action. This study establishes the proof-of-principal for our HTS approach and demonstrates the potential of VRC-2 in Btz refractory MM.

Citation Format: Amriti Lulla, Holly Stessman, David Dicker, Brian Van Ness, Wafik El-Deiry, Nathan G. Dolloff. VRC2, a novel bortezomib re-sensitizing compound for the treatment of multiple myeloma. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 1020. doi:10.1158/1538-7445.AM2013-1020
Abstract 1025: Optimized DZNep exposure presensitizes pancreatic cancer cells to anticancer nucleoside analogues: potential clinical implications.
Authors: Hung Sau Wai , Mody Hardik , Marrache Sean , Bhutia Yangzom D. , Davis Franklin , Cho Jong Hyun , Dhar Shanta , Chu Chung K. , Govindarajan Rajgopal
Published: August 2014

We evaluated the potential of a histone methylation reversal agent 3-deazaneplanocin A (DZNep) in improving the chemosensitivity of pancreatic cancer to nucleoside analogs (i.e., gemcitabine). DZNep brought delayed but selective cytotoxicity to pancreatic cancer cells without affecting normal human pancreatic ductal epithelial (HPDE) cells. Co-exposure of DZNep and gemcitabine induced cytotoxic additivity or synergism in both well- and poorly-differentiated pancreatic cell lines. In contrast, DZNep exerted antagonism with gemcitabine against HPDE cells with significant reduction in cytotoxicity compared with the gemcitabine-alone regimen. DZNep marginally depended on purine nucleoside transporters for its cytotoxicity, but the transport dependence was circumvented by acyl derivatization. Drug exposure studies revealed that a short priming with DZNep followed by gemcitabine treatment rather than co-treatment of both agents produced a maximal chemosensitization response in both gemcitabine-sensitive and gemcitabine-resistant pancreatic cancer cells. DZNep rapidly and reversibly decreased trimethylation of histone H3 lysine 27 but increased trimethylation of lysine 9 in an EZH2- and JMJD1A/2C-dependent manner, respectively. However, DZNep potentiation of nucleoside analog chemosensitization was found to be temporally coupled to trimethylation changes in lysine 27 and not lysine 9. Polymeric nanoparticles engineered to chronologically release DZNep followed by gemcitabine produced pronounced chemosensitization and dose-lowering effects. Together, our results identify that an optimized DZNep exposure can presensitize pancreatic cancer cells to anticancer nucleoside analogs and emphasize the promising clinical utilities of histone methylation reversal agents in future pancreatic cancer combination therapies.

Citation Format: Sau Wai Hung, Hardik Mody, Sean Marrache, Yangzom D. Bhutia, Franklin Davis, Jong Hyun Cho, Shanta Dhar, Chung K. Chu, Rajgopal Govindarajan. Optimized DZNep exposure presensitizes pancreatic cancer cells to anticancer nucleoside analogues: potential clinical implications. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 1025. doi:10.1158/1538-7445.AM2013-1025
Abstract 1029: Involvement of endocytic pathway in vorinostat-induced EGFR degradation.
Authors: Elkenawi Asmaa E. , Cowell John K.
Published: August 2014

Epidermal growth factor receptor (EGFR) is frequently overexpressed or aberrantly activated in various malignancies. As a result, EGFR is considered an important target in cancer therapeutics. Strategies of EGFR targeting include blockade of its activation, inhibition of downstream signaling or induction of EGFR downregulation and/or degradation. Histone deacetylases (HDACs) inhibitors were previously shown to deplete EGFR through numerous mechanisms varying from suppression of EGFR transcription, attenuation of its mRNA stability and disruption of chaperone activity of HSP90. In this work, we are studying alteration of EGFR vesicular trafficking as another mechanism that may be involved in HDAC inhibitors-induced EGFR depletion. Cytotoxic and EGFR-degrading effects of vorinostat, a pan-HDAC inhibitor, were investigated in the EGFR-overexpressing cell lines, DU-145 (prostate cancer cells) and MDA-MB-231 (breast cancer cells). Using different experimental designs, dose- and time- effects of vorinostat were studied in both cell lines. Proliferation assay, annexin V staining, PARP and caspase 7 cleavage were used to evaluate overall cellular toxicity. Effects on total EGFR protein level and relevant ERK phosphorylation were also investigated by immunoblotting. Moreover, accumulation of acetyl-α-tubulin was verified through immunoblotting and immunofluorescence imaging. Finally, serum-starved vorinostat-treated cells were stimulated with EGF in order to elucidate effect of vorinostat on EGF-induced EGFR degradation, an endocytic trafficking-dependent process. As presented in this study, vorinostat exerted potent cytotoxic effects on DU-145 and MDA-MB-231 cells manifested by a dose-dependent decrease in proliferation and increase in percentage of apoptotic cells. In addition, vorinostat caused a dose-dependent cleavage of both PARP and caspase 7, two markers of apoptotic cascade activation. Furthermore, vorinostat elicited a dose- and a time-dependent depletion of EGFR total protein in both cell lines with a corresponding decrease in ERK activation. Add to that, vorinostat caused accumulation of acetyl-α-tubulin and promoted EGFR degradation upon EGF stimulation denoting accelerated endocytic trafficking. In conclusion, vorinostat demonstrated a potent anti-tumor activity against EGFR-overexpressing cell lines. Such activity was accompanied by EGFR depletion that may be contributed in part by vorinostat-induced deregulated vesicular trafficking. 

Citation Format: Asmaa E. Elkenawi, John K. Cowell. Involvement of endocytic pathway in vorinostat-induced EGFR degradation. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 1029. doi:10.1158/1538-7445.AM2013-1029
Abstract 1081: Rapamycin is a chemopreventive and chemotherapeutic agent for ras-driven epidermal squamous cell carcinoma: evidence from mouse models.
Authors: Kitagawa Hiroshi , Cataisson Christophe , Handisurya Alessandra , Day Patricia M. , Yuspa Stuart H. , Dennis Phillip A.
Published: August 2014

Braf inhibitors are clinically important agents for the treatment of advanced melanoma however secondary cutaneous tumors are a common side effect; non-melanoma skin cancers (NMSCs) arise in 15-30% of patients on Braf inhibitor therapy and 60% of these harbor ras mutations. NMSC also occurs in 40% of organ transplant patients receiving immunosuppressive therapy, and switching from a calcineurin inhibitor-containing regimen to rapamycin reduces the incidence. To examine the effect of rapamycin on ras-driven epidermal squamous cell tumors, we treated mutant K-RasLA2 mice with rapamycin or vehicle by intraperitoneal injection and found that rapamycin prevented the development of squamous skin tumors (0/21 vs 6/24; p=0.04), and also rapidly reduced the tumor size. To explore this effect we employed a syngeneic orthotopic grafting model using H-Ras mutant primary murine keratinocytes to determine if rapamycin could hamper the ability of a Braf inhibitor to enhance squamous skin tumor growth. Similar to results in the K-RasLA2 model, rapamycin alone diminished the growth of tumors (size difference; 334.5 vs 22.3mm3; p=0.03), and decreased the volume even more in the presence of a Braf inhibitor (size difference; 699.6 vs 17.6 mm3; p=0.02). Treatment of established tumors with rapamycin resulted in significant tumor shrinkage even in the continuous presence of a Braf inhibitor. Size reduction was 78.7% and 66.7% in Braf inhibitor untreated and treated groups respectively. In vitro, Braf inhibition enhanced mutant H-Ras-induced activation of the Raf-ERK and mTOR pathways in keratinocytes, while rapamycin addition blocked the activation of signaling pathways and decreased cell proliferation. Taken together, rapamycin prevents murine skin tumor development arising from oncogenic mutations in two distinct types of ras gene alleles, and reduced the tumor size by inhibiting downstream oncogenic pathways. Rapamycin may have clinical application as a chemopreventive and therapeutic agent for patients at high risk to develop ras-driven NMSC, including those receiving Braf inhibitors.

Citation Format: Hiroshi Kitagawa, Christophe Cataisson, Alessandra Handisurya, Patricia M. Day, Stuart H. Yuspa, Phillip A. Dennis. Rapamycin is a chemopreventive and chemotherapeutic agent for ras-driven epidermal squamous cell carcinoma: evidence from mouse models. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 1081. doi:10.1158/1538-7445.AM2013-1081
Abstract 1094: Breast cancer metastatic competency in the hepatic niche is driven by E-cadherin re-expression.
Authors: Taylor Donald P.
Published: August 2014

Nearly half of breast cancer metastases will become clinically evident 5 or more years after the cancer was seemingly ablated. This implies that metastatic cancer cells survived over this extended timeframe without emerging as detectable nodules. Breast cancer metastatic dormancy has yet to be universally defined, and is not well understood as a potential means by which clinically evident metastases emerge after being undetected for years (or decades) from the primary tumor diagnosis. Various mechanistic and microenvironmental factors may work independently, collaboratively (or both) in order to allow a dormant metastatic breast cancer cell (or small cluster) to survive long term. At least one tumor dormancy hallmark is likely required: that of E-cadherin re-expression in order to form heterotypic connections between the carcinoma cells and the liver parenchyma. This re-expression is known as the mesenchymal-to-epithelial-reverting-transition (MErT). This re-expression provides not only survival signals to the breast cancer cells but may also establish cell polarity and protection from autocrine signaling (or extracellular factors) that otherwise may prompt the breast cancer cells to initiate metastatic outgrowth.

The purpose of this study is to determine E-cadherin's function in the establishment of breast cancer metastases to the liver. The MDA-MB-231 human breast cancer cell line, that can undergo induced MErT ex vivo, was transfected with either exogenously expressing E-cadherin or shRNA to E-cadherin. Xenograft splenic injections of these cell line variants into mice were performed and tumors allowed to grow for 30-60 days until sacrifice. Immunohistochemistry was performed on the liver and splenic tissue sections in order to assess the number and size of metastatic depostis. Preliminary data demonstrate that cells expressing exogenously encoded E-cadherin developed large splenic tumors with fewer to no liver metastases. MDA-MB-231 wild type cells spawned liver metastases about half the time. In order to more fully probe E-cadherin's role in metastatic competency experiments are underway to develop a doxycycline-inducible E-cadherin bicistronic vector allowing for the down-regulation of E-cadherin in the primary tumor and the re-expression in the metastatic niche. Subsequent immunohistochemistry will determine the metastatic E-cadherin status across these MDA-MB-231 cell variants. These results will provide insights into the role of MErT in metastatic establishment.

Citation Format: Donald P. Taylor. Breast cancer metastatic competency in the hepatic niche is driven by E-cadherin re-expression. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 1094. doi:10.1158/1538-7445.AM2013-1094
Abstract 1124: Site-specific DICER and DROSHA RNA products control the DNA-damage response.
Authors: d'Adda di Fagagna Fabrizio
Published: August 2014

The DNA damage response (DDR) is a signaling pathway that arrests the proliferation of cells undergoing genotoxic events to preserve genome stability and as such plays important tumor suppressing functions. DICER and DROSHA are crucial ribonucleases involved in RNA interference (RNAi). Components of RNAi are thought to have evolved to preserve genome stability from the attacks of viruses and mobile genetic elements. RNA products generated by DICER and DROSHA are involved in chromatin assembly, gene silencing and cancer. So far, RNAi and DDR pathways have not been demonstrated to directly interact.

We have recently shown in human, mouse and zebrafish that DICER and DROSHA, but not downstream elements of the RNAi pathway, are necessary to activate the DDR upon exogenous DNA damage and oncogene-induced genotoxic stress, as studied by DDR foci formation and by checkpoint assays. Indeed, oncogene-induced senescent cells, upon DICER or DROSHA inactivation, escape DDR-imposed arrest and proliferate again.

DDR foci are sensitive to RNase A treatment and DICER- and DROSHA-dependent RNA products are required to restore DDR foci in RNase-A-treated cells. Through RNA deep sequencing and the study of DDR activation at a single inducible DNA double-strand break, we demonstrate that DDR foci formation requires site-specific DICER- and DROSHA-dependent small RNAs, named DDRNAs, which act in a MRE11-RAD50-NBS1- complex-dependent manner to fuel DDR. DDRNAs, either chemically synthesized or in vitro generated by DICER cleavage, are sufficient to restore the DDR in RNase-A-treated cells, also in the absence of other cellular RNAs.

We propose an unanticipated direct role of a novel class of ncRNAs in the control of DDR activation at sites of DNA damage. The relevance of these findings in the context of tumor suppression will be discussed.

Citation Format: Fabrizio d'Adda di Fagagna. Site-specific DICER and DROSHA RNA products control the DNA-damage response. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 1124. doi:10.1158/1538-7445.AM2013-1124
Abstract 1138: Rapid validation of a novel kinase target using a large scale genomic database and matched patient derived tumor models.
Authors: Broudy Thomas , Sanga Sandeep , Ricono Jill , Trikha Mohit , Mirsaidi Cyrus , Praveen Kesavan Nair
Published: August 2014

Target discovery and validation in oncology has largely relied on molecular and functional studies performed in cell lines. Recent advances in genomics have now afforded large databases based on well-characterized tumor tissue, which has enabled direct investigation of patient tumors for novel targets. Following these discoveries, it is routine to perform functional studies in cell line-based systems; however, it is often difficult to find relevant cell line model and if identified, they can be confounded with artifacts created from decades of culture. The result can be a target validation process which takes considerable time and does not readily translate to clinical relevance.

We report here the creation and use of a target validation platform based on a large-scale genomic database matched to patient-derived tumor models. The platform relies on Molecular Response's proprietary bank of more than 144,000 patient derived tumors, of which nearly 400 tumors have been genomically characterized and databased for target discovery studies. The database is growing, but currently features the following cancer indications: colon carcinoma, NSCLC, melanoma, ovarian carcinoma, prostate and non-hodgkins lymphoma. Upon discovery of a novel target, tumors of interest are immediately implanted into mice to perform functional studies in direct patient derived models–either in vivo or using mouse-passaged cells for broader ex vivo studies.

Through use of this platform, we have identified a novel kinase target for potential therapeutic development. We investigated prevalence of target overexpression across 7 cancer indications, and identified melanoma as a clinical indication of high interest. We examined growth characteristics from patient tumors featuring high kinase gene expression vs. low expression to help characterize the role of this target in oncology disease progression. Finally, we performed functional knockdown studies in patient derived models to further validate this novel kinase as a druggable target of pharmaceutical interest. Studies are ongoing to develop small molecule and antibody-based therapeutics that will serve as drug candidates for further development.

Citation Format: Thomas Broudy, Sandeep Sanga, Jill Ricono, Mohit Trikha, Cyrus Mirsaidi, Kesavan Nair Praveen. Rapid validation of a novel kinase target using a large scale genomic database and matched patient derived tumor models. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 1138. doi:10.1158/1538-7445.AM2013-1138
Abstract 1165: Measuring telomere length for the early detection of precursor lesions of esophageal squamous cell carcinoma.
Authors: Lin Shih-Wen , Abnet Christian C. , Freedman Neal D. , Risques Rosana , Prunkard Donna , Rabinovitch Peter , Lu Ning , Qiao You-Lin , Dawsey Sanford M.
Published: August 2014

Objectives: Esophageal cancer is the fifth leading cause of cancer death worldwide. The five-year survival is approximately 16%. Patients can be successfully treated if diagnosed early, but current early detection screening tests are inadequate. Balloon cytology is a simple and inexpensive method of retrieving esophageal cells, but traditional cytologic examination of these cells has poor sensitivity and specificity. We examined whether telomere length, a marker for chromosomal instability, in DNA extracted from balloon cytology-collected esophageal cells is associated with precursor lesions of esophageal squamous cell carcinoma.

Methods: The Cytology Sampling Study 2, a population-based esophageal cancer early detection screening study, recruited over 700 healthy subjects from a high-risk population in Linxian, China. The screening study used a mesh-covered balloon to collect esophageal cells. All subjects underwent endoscopy, and precursor lesions were diagnosed by biopsy. Using a nested case-control study design, we examined the association between telomere length and dysplasia in 37 low-grade dysplasia cases, 52 high-grade dysplasia cases, and 92 matched controls. DNA was extracted from cells collected by balloon cytology, and telomere length was measured in triplicate by a quantitative PCR assay. Logistic regression models adjusted for age, sex, smoking tobacco, alcohol intake, and hypertension were used to examine the association between telomere length and risk of low-grade dysplasia and high-grade dysplasia, compared with normal controls.

Results: Telomere lengths were comparable among the low-grade dysplasia cases, high-grade dysplasia cases, and controls, with a median of 0.93 [interquartile range (IQR), 0.84-1.06], 0.94 (IQR, 0.82-1.09), and 0.93 (IQR, 0.78-1.05), respectively. Overall, telomere length was not associated with risk of low-grade dysplasia or high-grade dysplasia, with an odds ratio [OR, 95% confidence interval (CI)] of 1.16 (0.87-1.54) and 1.20 (0.92-1.56), respectively.

Conclusions: Given that esophageal cancers have very poor prognoses, it is of utmost importance to improve early detection. In this study, esophageal cell telomere length was not associated with esophageal precursor lesions and cannot serve as an early detection marker.

Citation Format: Shih-Wen Lin, Christian C. Abnet, Neal D. Freedman, Rosana Risques, Donna Prunkard, Peter Rabinovitch, Ning Lu, You-Lin Qiao, Sanford M. Dawsey. Measuring telomere length for the early detection of precursor lesions of esophageal squamous cell carcinoma. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 1165. doi:10.1158/1538-7445.AM2013-1165
Abstract 1182: Ocimum gratissimum aqueous extract induces apoptotic signalling in human osteosarcoma cells.
Authors: Lin Chien Chung
Published: August 2014

Osteosarcoma (Osteosarcoma) is a kind of bone cancer. Eighty percent of this tumor will be metastatic to the lungs or liver. Therefore, these patients generally need to use chemotherapy to improve disease and increase survival possibility. Recently, it has been reported that Ocimum gratissimum (OG) aqueous extract has anti-tumor activity. In this study, we detected the effect of OG extract on human bone cancer cell growth and metastasis.

The results showed that the OG extract can inhibit cell growth and induce cell apoptosis in human bone cancer cells, but no effect in normal nerve cell. Therefore, we suggest that OG extract s may have anti-cancer ability, and can provide the use in the development of new anti-cancer drugs

Figure . OGE effect on the cell growth in two osteosarcoma cells. Cell growth was determined 0∼3 or 0∼5 days after subculture using the MTT assay as described in Materials and Methods. Absorbance values obtained from the cells on day 0 after subculture were taken as 100%.

Citation Format: Chien Chung Lin. Ocimum gratissimum aqueous extract induces apoptotic signalling in human osteosarcoma cells. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 1182. doi:10.1158/1538-7445.AM2013-1182

Note: This abstract was not presented at the AACR Annual Meeting 2013 because the presenter was unable to attend.
Abstract 1224: Intratumoral chitosan/IL-12 neoadjuvant to tumor resection is safe and generates tumor specific immunity.
Authors: Vo Jimmy Loc Nhu , Yang Lirong , Zaharoff David
Published: August 2014

Interleukin-12 (IL-12) is a potent antitumor cytokine that exhibits significant clinical toxicities after systemic administration. Local delivery strategies capable of maintaining high concentrations of IL-12 in the tumor microenvironment while minimizing systemic exposure are under investigation. We have previously shown that intratumoral injections of IL-12 co-formulated with a solution of chitosan (chitosan/IL-12) can eliminate established primary tumors. Chitosan is an abundant, natural polysaccharide derived primarily from the exoskeletons of crustaceans. Chitosan has been shown to maintain high local concentrations of protein antigens and cytokines through viscous, electrostatic and bioadhesive interactions.

Because of IL-12’s well documented ability to generate tumor-specific cell-mediated immunity, we are exploring the anti-metastatic potential of intratumoral chitosan/IL-12 immunotherapy in a highly metastatic model of breast cancer (4T1). Previously, we have shown that intratumoral chitosan/IL-12 neoadjuvant prior to tumor resection confers long-term survival benefits and reduces metastasis. Recent studies reveal that chitosan/IL-12 is safe and generates tumor-specific systemic immunity to confer long-term survival benefits. Specifically, splenocytes from long term survivors demonstrated anti-tumor immunity in a cytotoxic T-lymphocyte (CTL) killing assay. After 1 week of in vitro stimulation, splenocytes were found to lyse approximately 40% of target tumor cells at an effector: target ratio of 50:1. In addition, toxicological studies reveal that chitosan/IL-12 does not adversely affect leukocyte levels or hematocrit. Based on data obtained thus far, intratumoral chitosan/IL-12 shows promising potential as a neoadjuvant immunotherapy to reduce metastatic disease while minimizing the toxicities that hindered the progress of IL-12 based cancer immunotherapy.

Citation Format: Jimmy Loc Nhu Vo, Lirong Yang, David Zaharoff. Intratumoral chitosan/IL-12 neoadjuvant to tumor resection is safe and generates tumor specific immunity. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 1224. doi:10.1158/1538-7445.AM2013-1224
Abstract 1256: Humoral autoimmune response to a RNA-binding protein HCC1/CAPER in hepatocellular carcinoma (HCC).
Authors: Dai Liping , Ren Pengfei , Liu Xinxin , Ortega Rosalia , Yao Wu , Wang Kaijuan , Wang Mingan , Tan Eng M. , Zhang Jianying
Published: August 2014

HCC1 was originally cloned as a cell nuclear autoantigen in a patient with liver cirrhosis that progressed to HCC. The full-length sequence of HCC1encodes a protein of 530 amino acids and migrating at 64 kDa in SDS-PAGE. HCC1 has two isoforms of HCC1.4 and HCC1.3. The difference is that HCC1.3 has 6-amino acid deletion located in the RNP-1-like region of the putative third RNP-CS domain. The nucleotides and protein sequences of HCC1 were not found at that time in searching available databases and, therefore, it was considered as a novel gene with unknown function. Several years later, a group showed that HCC1 was a co-activator of activating protein-1 and estrogen receptors and proposed naming it CAPER. Further study by other group has demonstrated that HCC1/CAPER is a novel transcriptional co-activator for v-Rel that strongly suppresses its transforming activity, uncovering a tumor suppressor role for HCC1/CAPER in regulating Rel's oncogenic activity. Our previous studies have demonstrated that autoantibodies against tumor-associated antigens (TAAs) can be regarded as serological biomarkers in cancer immunodiagnosis. Whether autoantibodies to HCC1/CAPER can be also used as one of the markers in HCC remains to be investigated. In the present study, autoantibody responses to HCC1.3 and HCC1.4 were firstly evaluated by ELISA, western blotting and indirect immunofluorescence assay in sera from patients with HCC, liver cirrhosis and chronic hepatitis, as well as from normal human individuals. Immunohistochemistry (IHC) with HCC tissue array slides was also performed to analyze protein expression profiles of HCC1. The prevalence of anti-HCC1.4 antibodies was 13.5% (23/171) in HCC, which was significantly higher than that in normal human sera (1.1%). Whereas the significant difference regarding frequency of anti-HCC1.3 antibodies between HCC patients and normal individuals was not found. The frequency of HCC1 expression in HCC tissues was also significantly higher than that in normal liver tissues (57.5% vs. 0%; P<0.001). The data suggests that anti-HCC1.4 autoantibody may be a potential biomarker for HCC, but not HCC1.3. Further functional study of HCC1 may provide more evidence to address the question why these two isoforms of HCC1 can induce different humoral immune response in HCC patients.

Citation Format: Liping Dai, Pengfei Ren, Xinxin Liu, Rosalia Ortega, Wu Yao, Kaijuan Wang, Mingan Wang, Eng M. Tan, Jianying Zhang. Humoral autoimmune response to a RNA-binding protein HCC1/CAPER in hepatocellular carcinoma (HCC). [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 1256. doi:10.1158/1538-7445.AM2013-1256
Abstract 1258: The frequency of CD11c+ cells within the gastric cancer tissue is associated with favorable prognosis.
Authors: Jiang Jingting , Wu Changping , Lu Binfeng
Published: August 2014

Aim: CD11c is a marker for dendritic cells which help initiating adaptive antitumor immunity. The study was designed to investigate correlation between the frequency of CD11c+ cells in the gastric cancer tissue and the clinical and pathological parameters of gastric cancer patients.

Methods: CD11c expression in gastric cancer tissues was examined by immunohistochemistry. Surface CD11c expression on the leukocytes within cancer tissues was examined by flow cytometry.

Results: The number of CD11c+ cells was higher in gastric cancer tissues than that in gastritis or gastric polyps tissues. The number of CD11c+ cells in tumor specimens from the early-stage gastric cancer was significantly higher than that in specimens from the late-stage gastric cancer (t=2.77,P=0.0064). Flow cytometry found that CD11c surface expression was stronger in specimens from the early stages than those from the late stages of gastric cancer. The Univariate analysis revealed that the frequency of intratumoral CD11c+ cells showed association with gender and tumor location, and was reversely correlated with tumor size, depth of infiltration, nodal metastasis, tumor histological grade and cancer stages. The median survival time (MST) for the subgroups of patients with stronger and weaker CD11c expression were 67.0 months (95%CI, 39.0-112.0 months) and 29.0 months (95%CI, 21.0-39.0 months), respectively. And MSTs were significantly different between the CD11c higher and lower subgroups (χ2=11.6425,P=0.0006). Among patients who suffered from late-stage diseases, those with higher numbers of intratumoral CD11c cells had a significantly lower risk of death than those with lower numbers of CD11c cells (χ2=8.549,P=0.0035), and the lowered risk of death was statistically significant after adjusting gender, age, tumor size and infiltrating depth (RR=0.52,95%CI=0.29-0.94).

Conclusions: The number of CD11c+ cells within gastric cancer tissue is a favorable prognostic marker for gastric cancer. Our study suggests gradual elimination of dendritic cells is characteristic of gastric cancer progression.

Citation Format: Jingting Jiang, Changping Wu, Binfeng Lu. The frequency of CD11c+ cells within the gastric cancer tissue is associated with favorable prognosis. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 1258. doi:10.1158/1538-7445.AM2013-1258

Note: This abstract was not presented at the AACR Annual Meeting 2013 because the presenter was unable to attend.
Abstract 1286: FANCA binds to nucleic acids and stimulates endonuclease activity of FEN1.
Authors: Qian Liangyue , Yuan Fenghua , Rodriguez-Tello Paola , Zhao Xinliang , Song Limin , D'Urso Gennaro , Jain Chaitanya , Zhang Yanbin
Published: August 2014

Fanconi anemia is a rare autosomal recessive or X-linked genetic disease characterized by progressive bone marrow failure, various developmental anomalies, and cancer predisposition. Fanconi anemia complementation group A (FANCA) gene is one of the 15 disease-causing genes and has been found to be mutated in ∼60% of Fanconi anemia patients. Except for its constituent role in the ubiquitin ligase of the Fanconi anemia core complex, no other biochemical activity has been identified thus far.

Using purified protein, we observed that human FANCA has intrinsic affinity to nucleic acids. FANCA binds to both single-stranded and double-stranded DNA; however, its affinity to single-stranded DNA is dramatically higher. FANCA also binds to RNA with an intriguingly higher affinity than its DNA counterpart. A patient-derived FANCA truncation mutant, Q772X, has diminished affinity to both DNA and RNA. On the contrary, the complementing C-terminal fragment of Q772X, C772-1455, retains the differentiated nucleic acids binding activity (RNA>ssDNA>dsDNA), indicating that the nucleic acids binding domain of FANCA is primarily located at its C-terminus. Using DNA and RNA ladders, we determined that the minimum numbers of nucleotides required for FANCA recognition are ∼30 for both DNA and RNA. By testing the affinity between FANCA and a variety of DNA structures, we found that a 5’ flap or tail on DNA facilitates its interaction with FANCA. In addition, we observed that FANCA greatly stimulates the endonuclease activity of flap endonuclease 1 (FEN1), which processes 5’ flap structures.

Our study reported that purified human FANCA binds to nucleic acids with strong preference for single-stranded forms. Our finding suggests that FANCA FANCA may participate in maintenance of replication forks by regulating removal of Okazaki fragment during maturation of lagging strand replication.

Citation Format: Liangyue Qian, Fenghua Yuan, Paola Rodriguez-Tello, Xinliang Zhao, Limin Song, Gennaro D'Urso, Chaitanya Jain, Yanbin Zhang. FANCA binds to nucleic acids and stimulates endonuclease activity of FEN1. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 1286. doi:10.1158/1538-7445.AM2013-1286
Abstract 1287: Epigenetic deregulation of breast cancer is associated with DNA Repair Capacity.
Authors: Matta Jaime , Hadar Tal , Morales Luisa , Ostrow Kim , Sidransky David , Guerrero-Preston Rafael E.
Published: August 2014

Introduction 226,870 women are estimated to be diagnosed with breast cancer in 2012. Search for accurate methods for screening, diagnosis and treatment follow up, that will be minimally invasive are ongoing.

Hypermethylation of gene promoters occurs frequently in the pathogenesis of breast cancer, and causes silencing of tumor suppressor genes. Hypermethylation of Mal, Kif1a and OGDHL in breast cancer cell lines and tissue samples was previously reported.

Aberrant DNA repair capacity (DRC) has been studied and shown to elevate the risk for breast cancer.

However, information on the association between the epigenetic promoter hypermethylation and DRC is lacking in the literature.

Objective The aim of this study is to assess the association between breast cancer and promoter hypermethylation of Mal, Kif1a and OGDHL in serum of breast cancer patients and controls in association with the DRC.

Methods 240 serum samples from breast cancer patients and controls from Puerto Rico were collected. DNA was extracted and methylation levels were measured after bisulfite treatment, by QMSP. DRC was assessed by host reactivation assay of blood samples. The overall risk for breast cancer was calculated and adjusted for patient characteristics including DRC.

Results Methylation of at least one of the genes was found in 49% of breast cancer samples compared to 20% of controls. ROC curves for individual genes showed values of 0.64 for MAL, 0.51 for Kif1a and 0.53 for OGDHL. In the linear regression unadjusted analysis, mean methylation levels (what are the values?) of the three genes and odds ratio (OR) were higher in breast cancer cases compared to controls, with Kif1a and OGDHL values being statistically significant. These values became non significant after adjusting for DRC. Hypermethylation of Kif1a, but not MAL or OGDHL was found to be significantly associated with DRC.

Conclusion Correlation between promoter hypermethylation of the genes Mal, Kif1a and OGDHL and breast cancer was non significant when adjusted for DRC.

A significant association between hypermethylation of Kif1a and DRC was shown.

The association between promoter hypermethylation and DRC needs to be further studied.

Citation Format: Jaime Matta, Tal Hadar, Luisa Morales, Kim Ostrow, David Sidransky, Rafael E. Guerrero-Preston. Epigenetic deregulation of breast cancer is associated with DNA Repair Capacity. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 1287. doi:10.1158/1538-7445.AM2013-1287
Abstract 1290: Investigations to evaluate the role of Twist1 during tumor promotion and epithelial carcinogenesis.
Authors: Srivastava Jaya , Macias Everardo , Rao Dharanija , Kiguchi Kaoru , DiGiovanni John
Published: August 2014

Recent studies have identified the involvement of receptor tyrosine kinases (RTKs) and their downstream signaling pathways, including MAPK, PI3K/Akt/mTOR, and Stat3, as key regulatory factors of epidermal proliferative capacity during skin carcinogenesis. Through development and study of transgenic mice with overexpression of a constitutively active form of Stat3 (Stat3C) in the basal cell layer, previous studies in the laboratory discovered that constitutive Stat3 activation accelerates tumor progression in the two-stage skin carcinogenesis model, which utilizes the tumor initiator 7,12-dimethylbenz[α]anthracene (DMBA) and tumor promoter 12-O-tetradecanoylphorbol-13-acetate (TPA). Twist1, a well-known inducer of EMT, has been shown to be regulated by Stat3 in a number of systems. Western blot analysis of epidermal protein lysates from Stat3C transgenic mice indicated a two-fold induction of Twist1 expression as compared to wild type mice. In addition, immunofluorescence staining analysis of tumors from wild type mice [papillomas and squamous cell carcinomas (SCCs)] and Stat3C mice (SCCs) showed nuclear co-localization of Twist1 and Stat3. In the present study, we investigated the activation and subsequent downstream signaling of Twist1 during TPA-induced epidermal hyperproliferation. Western blot analysis of epidermal lysates from mice treated topically with multiple applications of TPA showed an initial decrease (4-6 h) followed by a subsequent rebound of Twist1 protein at 18 hours post TPA treatment. This initial decrease and then rebound coincided with that of the cell cycle regulatory proteins cyclin D1, cyclin E1, E2F-1, and c-myc. Interestingly, basal levels of Twist1 were reduced in epidermis specific Stat3 deficient mice. Furthermore, Stat3 deficient mice treated with TPA failed to show a rebound in expression of Twist1 in the epidermis. Immunofluorescence experiments of TPA treated epidermis revealed that nuclear localization of Twist1 was restricted to the proliferative compartment. Collectively, these results, combined with other data to be presented, suggest that Stat3 regulates basal levels of Twist1. This protein is localized in the nucleus of cells found primarily in the proliferative compartment of hyperplastic epidermis and skin tumors. The current results further suggest that Twist1 may play a role in cell cycle progression in keratinocytes during tumor promotion. Research supported by CA76520.

Citation Format: Jaya Srivastava, Everardo Macias, Dharanija Rao, Kaoru Kiguchi, John DiGiovanni. Investigations to evaluate the role of Twist1 during tumor promotion and epithelial carcinogenesis. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 1290. doi:10.1158/1538-7445.AM2013-1290
Abstract 1296: Involvement of RSK2 in human skin cancer.
Authors: Lee Cheol-Jung , Lee Hye Suk , Lee Ji-Young , Cho Yong Yeon
Published: August 2014

Our previous study demonstrated that the total p90 ribosomal S6 kinase 2 (RSK2) protein level was significantly higher in human skin cancer tissues and cancer cells compared with normal skin tissues and premalignant cell lines. However, the role of RSK2 activation and signaling in human skin cancer has not yet been fully elucidated. In this study, we provided evidences that RSK2 plays a key role in human skin cancer. We examined the effect of knocking down RSK2 expression on EGF-induced anchorage-independent transformation in the premalignant HaCaT human skin keratinocyte cell line and on soft agar colony growth of SK-MEL-28 malignant melanoma (MM) cells. We found that UVB stimulation induced increased in not only the total and phosphorylated protein levels of ERKs and RSK2, but also the nuclear localization and gene expression of RSK2. Moreover, RSK2−/− mouse embryonic fibroblasts (MEFs) showed enhanced sub-G1 accumulation induced by UVB stimulation compared with RSK2+/+ MEFs, indicating that RSK2 might play an important role in tolerance against stress associated with UV. RSK2 knockdown inhibited proliferation and anchorage-independent transformation of HaCaT cells and soft agar colony growth of MM cells. We found that the phosphorylated protein levels of RSK2 were enhanced in cancer tissues compared with normal tissues in a human skin cancer tissue array. Importantly, activated RSK2 protein levels were highly abundant in human skin cancer tissues compared with matched skin normal tissues. Taken together, our results demonstrated that RSK2 plays a key role in neoplastic transformation of human skin cells and in skin cancer growth.

Citation Format: Cheol-Jung Lee, Hye Suk Lee, Ji-Young Lee, Yong Yeon Cho. Involvement of RSK2 in human skin cancer. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 1296. doi:10.1158/1538-7445.AM2013-1296
Abstract 1306: SR48692 inhibits EGF receptor transactivation and proliferation of lung cancer cells.
Authors: Moody Terry W. , Jensen Robert T.
Published: August 2014

Neurotensin (NTS) is an autocrine growth factor for some lung cancer cells (Moody et al., 1985;Life Sci 36:1727). NTS binds with high affinity to the GPCR NTSR1 causing phosphatidyl inositol turnover and proliferation of lung cancer cells; SR48692 is a NTSR1 antagonist (Moody et al., 2001; Peptides 22:109). NTSR1 expression is associated with poor survival of non-small cell lung cancer (NSCLC) patients (Alfano et al., 2010; Clin Cancer Res 16:4401). NSCLC cells express the epidermal growth factor receptor (EGFR) whose tyrosine kinase activity is inhibited by gefitinib. Here the abillity of SR48692 to block EGFR transactivation caused by NTS was investigated. NTS and NTS8-13 but not NTS1-8 addition to NSCLC cell lines NCI-H1299 or A549 cells increased phosphorylation of Tyr1068 of the EGFR after 2 min which was blocked by 10 uM SR48692 or 10 ug/ml gefitinib. The ability of NTS to regulate EGF receptor transactivation was blocked by GM6001 (matrix metalloprotease inhibitor), PP2 (Src inhibitor), N-acetylcysteine (anti-oxidant) and transforming growth factor TGF) α antibody. By ELISA, NTS significantly increased secretion of TGFα from lung cancer cells. NTS but not levocabastine (NTSR2 agonist) increased cytosolic Ca2+ within seconds after addition to NCI-H1299 cells which was antagonized by SR48692. NTS addition to NCI-H1299 cells increased tyrosine phosphorylation of beta-catenin, ERK, FAK, PYK-2 and Src which was antagonized by SR48692. SR48692 inhibited the proliferation of NCI-H1299 and A549 cells which have wild type EGFR. Also, SR48692 potentiated the ability of gefitinib to inhibit growth of NCI-H1299 and A549 cells. These results indicate that SR48692 is a NTSR1 antagonist which increases the potency of gefitinib in lung cancer cells.

Citation Format: Terry W. Moody, Robert T. Jensen. SR48692 inhibits EGF receptor transactivation and proliferation of lung cancer cells. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 1306. doi:10.1158/1538-7445.AM2013-1306
Abstract 136: Statins and breast cancer mortality.
Authors: Murtola Teemu J. , Visvanathan Kala , Artama Miia , Vainio Harri , Pukkala Eero
Published: August 2014

Background: Breast cancer is a common malignancy and cause of cancer death. Recent studies have suggested that statins, an established drug group in the prevention of cardiovascular mortality, could have beneficial effects against recurrence and progression of breast cancer. We evaluated breast cancer-specific and overall mortality among statin users in a cohort of breast cancer patients.

Methods: Our study cohort included all newly diagnosed breast cancer patients in Finland during 1995-2003 (31,236 cases), identified from the Finnish Cancer Registry. Information on statin use during these years was obtained from a national prescription database. We used Cox proportional hazards regression method to estimate mortality among statin users separately for pre- and post-diagnostic usage.

Results: A total of 4,169 participants had ever-used statins. During the median follow-up of 3.25 years (range 0.08-9.0 years), a total of 6,011 participants died, of which 3,619 (60.2%) was due to breast cancer. Post-diagnostic statin use was inversely associated with breast cancer mortality (participants with localized tumors: HR 0.33, 95% CI 0.24-0.42; metastatic tumors: HR 0.52, 95% CI 0.31-0.86) and also overall mortality. The association between statin use and mortality was dose-dependent. In stratified analyses, the reduction did not significantly vary by lipophilicity of statins. Additionally, long-term pre-diagnostic statin use was also associated with lower breast cancer mortality.

Conclusion: Lower breast cancer mortality observed for both pre-diagnostic and post-diagnostic statin use suggests that statins could lower risk of death in breast cancer patients. A clinical trial testing statins’ effect on survival in breast cancer patients is feasible.

Citation Format: Teemu J. Murtola, Kala Visvanathan, Miia Artama, Harri Vainio, Eero Pukkala. Statins and breast cancer mortality. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 136. doi:10.1158/1538-7445.AM2013-136