bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 4423: Chemoresistance regulation by COL11A1 in ovarian cancer
Authors: Wu Yi-Hui
Published: August 2015

Epithelial ovarian carcinoma remains the most lethal gynecologic malignancy. While initial response to first-line therapy (consisting of surgical cytoreduction and combination platinum/taxane therapy) is usually effective, the majority of patients will ultimately recur with chemotherapy-resistant cancer and succumb to disease. Biomarkers that predict patient's responsiveness to chemotherapy might assist the development of better therapeutic strategies for aggressive cancers, such as ovarian cancer. Here, we investigated the molecular mechanisms of collagen type XI alpha 1 (COL11A1) in chemo-resistance of ovarian cancer. We analyzed the expression profiles of 60 epithelial ovarian cancer tissue samples obtained at first cytoreductive surgery to identify genes linked with chemotherapy response. Among the 47 selected genes, collagen type XI alpha 1 (COL11A1) was the most highly elevated in chemo-resistance tumors. Small interfering RNA-mediated specific reduction of COL11A1 protein levels increased the sensitivity of ovarian chemoresistant cells to cisplatin and paclitaxel. A combination of experimental approaches, including real-time RT-PCR, western blotting, luciferase assay and chromatin immunoprecipitation (ChIP) assays, showed that the c/EBPβ binding site on the COL11A1 promoter was identified as the major determinant of cisplatin and paclitaxel-dependent COL11A1 activation. Furthermore, COL11A1 silencing suppressed phosphorylated Akt expression, which was in concert with increased PDK1 degradation in chemoresistant cells. Analysis of 104 EOC patients showed that high COL11A1 mRNA levels are significantly associated with poor chemoresponse and poor clinical outcome. In conclusion, activation of COL11A1 by anticancer drugs confers chemoresistance on ovarian cancer cells through the activation of Akt/c/EBPβ pathway and PDK1 stabilization.

Note: This abstract was not presented at the meeting.

Citation Format: Yi-Hui Wu. Chemoresistance regulation by COL11A1 in ovarian cancer. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 4423. doi:10.1158/1538-7445.AM2015-4423
Abstract 4425: Multiple -omic analyses of a pair of primary HCC tumor cell lines with different drug response revealed the mechanisms of drug resistance
Authors: Hu Gang , Du Alicia , Huang Yong , Liu Kunyan , Xie Fubo , Tang Xuzhen , Yang Xueyan , Gu Qi , Zhang Yixin , Tao Weikang , Zhang Yingjia , Tang Wei , Zhou He
Published: August 2015

Previously we reported the establishment of a pair of primary HCC cell lines from LIX-004, a patient-derived HCC xenograft model. LIXC004-NA was developed from an untreated PDX tumor; LIXC004-SR was generated from a PDX tumor progressed despite long term in vivo treatment with sorafenib. The two cell lines displayed similar response to sorafenib treatment in vitro. However, LIXC004-NA is sensitive and LIXC004-SR is resistant to sorafenib treatment in vivo. IHC and in vitro functional studies revealed activation of alternative angiogenic pathways as one of the potential reasons for the sorafenib resistance of LIXC004-SR. To fully explore different mechanisms/pathways, we performed a) micro-array analyses; b) whole exome sequencing; c) RNA-seq analyses of the two cell lines; d) metabolomic analyses of the cell lines in vitro culture in the presence and absence of different therapeutics; e) RNAseq analyses of the tumors derived from these two cell lines. The combination of different types of -omic analyses is our systematic approach to address the potential mechanisms of drug resistance without bias. The two cell lines shared a lot of the genetic mutation and expression profile for most genes. Differentially mutated or expressed genes were also identified at basal level. More changes were observed with the treatment of different therapeutics as well as analyses of the xenograft derived from the cell lines. Pathway analyses are performed to explore the underlying mechanisms for the development of drug resistance.

Citation Format: Gang Hu, Alicia Du, Yong Huang, Kunyan Liu, Fubo Xie, Xuzhen Tang, Xueyan Yang, Qi Gu, Yixin Zhang, Weikang Tao, Yingjia Zhang, Wei Tang, He Zhou. Multiple -omic analyses of a pair of primary HCC tumor cell lines with different drug response revealed the mechanisms of drug resistance. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 4425. doi:10.1158/1538-7445.AM2015-4425
Abstract 4427: Apoptosis, autophagy and ATP: The dynamic interaction of cellular ATP and drug resistance in MET amplified gastric cancer
Authors: Dunbar Schroeder Rebecca , Hong David , McConkey David J.
Published: August 2015

Background: MET amplification occurs in 20% of gastric tumors, MET protein overexpression is present in approximately 50% of advanced gastric cancers and activating MET mutations are exceedingly rare in gastric cancer. Several ongoing trials have reported responses in gastric and esophageal patients with MET amplification .Methods: To investigate the effects, of MET inhibition on proliferation and cell death in gastric cancer I used two complementary approaches 1) siRNA mediated knockdown of MET; 2) incubation with crizotinib, a clinically approved small molecule inhibitor of MET. Both resulted in decreased proliferation and increased cell death only in the MET amplified gastric cancer cell lines. Results: The effects on growth arrest were stronger than the effects on cell death. To look for possible resistance mechanisms we performed whole genome mRNA expression profiling. Molecular changes related to autophagy were among the top alterations observed. Consistent with these findings, autophagy levels increased in a concentration dependent manner when exposed to crizotinib. In order to investigate the role of autophagy on cell death, I inhibited autophagy using two approaches 1) siRNA mediated knockdown of ATG5 and ATG7; 2) exposure to chloroquine, a clinically approved agent that inhibits autophagy by raising lysosomal pH. I then measured apoptosis levels with the autophagy inhibitor alone or in combination with MET knockdown or crizotinib. Autophagy inhibition caused a dramatic decrease in apoptosis in one of the MET amplified cell lines (MKN45) but not in the other (SNU-5). Because autophagy provides energy for cells, we explored the effects of MET or autophagy inhibition on cellular ATP levels using the Cell Titer-glo assay. The MKN45 cells had the lowest basal levels of ATP and these levels were further diminished in combination with autophagy inhibition. Conclusions: Overall, our data reveal a novel relationship between low ATP levels and resistance to MET inhibitor-induced cell death. Clinically this is significant because it may discourage the use of combination therapies with autophagy inhibitors in gastric cancer patients whose tumors contain MET amplification.

Citation Format: Rebecca Dunbar Schroeder, David Hong, David J. McConkey. Apoptosis, autophagy and ATP: The dynamic interaction of cellular ATP and drug resistance in MET amplified gastric cancer. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 4427. doi:10.1158/1538-7445.AM2015-4427
Abstract 4429: EGR1-MIR152 pathway overcomes acquired cisplatin resistance in ovarian cancer cells by inhibiting cyto-protective autophagy via ATG14
Authors: He Jun , Yu Jing-Jie , Jiang Bing-Hua
Published: August 2015

The cytotoxic-based platinum compound cisplatin has been commonly used in ovarian cancer treatment for almost three decades. However, the intrinsic and acquired resistance to cisplatin in cancer cells remains a big challenge for overall survival. A growing body of evidence indicates a role of autophagy as a prosurvival and resistance mechanism against chemotherapy. Here, we reported MIR152 as a new autophagy-regulating miRNA that plays a role in cisplatin resistance. We showed that MIR152 expression was dramatically downregulated in the cisplatin-resistant cell lines A2780/CP70, SKOV3/DDP compared with their respective parental cells, and in ovarian cancer tissues associated with cisplatin resistance. Overexpression of MIR152 sensitized cisplatin-resistant ovarian cancer cells by reducing cisplatin-induced autophagy, enhancing cisplatin-induced apoptosis and inhibition of cell proliferation. A mouse subcutaneous xenograft tumor model using A2780/CP70 cells with overexpressing MIR152 was established and displayed decreased tumor growth in response to cisplatin. We also identified that ATG14 is a functional target of MIR152 in regulating autophagy inhibition. Furthermore, we found that EGR1 (early growth response 1) regulated the MIR152 gene at the transcriptional level. Ectopic expression of EGR1 enhanced efficacy of chemotherapy in A2780/CP70 cells. More importantly, these findings were relevant to clinical cases. Both EGR1 and MIR152 expression levels were significantly lower in human ovarian cancer tissues with high levels of ERCC1 (excision repair cross-complementation group 1), a marker for cisplatin resistance. Collectively, these data provide insights into novel mechanisms for acquired cisplatin resistance. Activation of EGR1 and MIR152 may be a useful therapeutic strategy to overcome cisplatin resistance by preventing cyto-protective autophagy in ovarian cancer.

Note: This abstract was not presented at the meeting.

Citation Format: Jun He, Jing-Jie Yu, Bing-Hua Jiang. EGR1-MIR152 pathway overcomes acquired cisplatin resistance in ovarian cancer cells by inhibiting cyto-protective autophagy via ATG14. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 4429. doi:10.1158/1538-7445.AM2015-4429
Abstract 4455: Relapsed/refractory AML responds robustly to IACS-10759, a novel OXPHOS inhibitor
Authors: Protopopova Marina , Bandi Madhavi , Bardenhagen Jennifer , Bristow Christophor , Carroll Christopher , Chang Edward , Feng Ningping , Gay Jason , Geck Do Mary , Greer Jennifer , Konopleva Marina , Matre Polina , Kang Zhijun , Liu Gang , Muller Florian , Lofton Timothy , McAfoos Timothy , Theroff Jay , Sun Yuting , Wu Yuanqiang , Smith Melinda , Chin Lynda , Draetta Giulio , Jones Philip , Toniatti Carlo , Di Francesco M. Emilia , Marszalek Joseph R.
Published: August 2015

Acute myeloid leukemia (AML) is a highly aggressive disease that is made up of several genetically and clinically diverse hematological malignancies that are characterized by clonal expansion of malignant stem/progenitor cells with limited ability to differentiate into mature blood cells. Standard of care for AML has progressed minimally in the past 30 years for relapse/refractory AML, with survival rates of <12% for those aged >65 years. Therefore, novel, highly effective therapeutics are needed for this population. Growing evidence suggests that in AML, metabolism is altered and that the tumor cells become highly dependent of mitochondria oxidative phosphorylation (OXPHOS) for their survival. We developed IACS-10759 as a novel small molecule inhibitor of complex I that potently inhibits oxygen consumption, eliminates hypoxia, and strongly inhibits the proliferation of cells grown in galactose medium with EC50 values between 1-10 nM. When AML cell lines as well as primary AML cells from relapsed/refractory patients were treated ex vivo with IACS-10759, apoptosis was robustly induced with EC50 values also ranging between 1-10 nM. It is noteworthy, that while apoptosis was induced in primary AML cells, normal patient-derived bone marrow cells were not sensitive. In AML orthotopic xenografts, daily oral dosing with 15 mg/kg IACS-10759 extended median survival to 70 days from 18 days in control animals. Taken together, the robust response in AML cell lines, primary AML samples ex vivo, and efficacy in orthotopic xenografts, IACS-10759 has entered IND enabling studies with first-in-human studies targeted for third quarter of 2015.

Citation Format: Marina Protopopova, Madhavi Bandi, Jennifer Bardenhagen, Christophor Bristow, Christopher Carroll, Edward Chang, Ningping Feng, Jason Gay, Mary Geck Do, Jennifer Greer, Marina Konopleva, Polina Matre, Zhijun Kang, Gang Liu, Florian Muller, Timothy Lofton, Timothy McAfoos, Jay Theroff, Yuting Sun, Yuanqiang Wu, Melinda Smith, Lynda Chin, Giulio Draetta, Philip Jones, Carlo Toniatti, M. Emilia Di Francesco, Joseph R. Marszalek. Relapsed/refractory AML responds robustly to IACS-10759, a novel OXPHOS inhibitor. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 4455. doi:10.1158/1538-7445.AM2015-4455
Abstract 4456: Prevention of MICA shedding from HBV infected hepatocytes to activate NK cells for better clearance of malignant foci and virus-infected cells
Authors: Ohno Motoko , Otsuka Motoyuki , Kishikawa Takahiro , Yoshikawa Takeshi , Takata Akemi , Koike Kazuhiko
Published: August 2015

Introduction: Hepatocellular carcinoma (HCC) is an important issue to public health of the world. By a genome-wide association study, we previously identified that a single nucleotide polymorphism (SNP) in the promoter region of the MHC class I polypeptide-related sequence A (MICA) gene is significantly associated with the risk of hepatitis-virus-related HCC. MICA is a ligand for NKG2D, a cell receptor of NK cells, which activates NK cells to clear the pathogenic cells expressing MICA such as infected cells or malignant cells as a “danger” signal. However, unexpectedly, the Hepatitis B virus (HBV)-infected patients with risk allele of MICA gene showed higher levels of serum MICA. This suggested that the serum MICA, which is excreted from the infected hepatocytes by a shedding mechanism, may work as a decoy for NK cells and inhibit their activities. Therefore, increasing cellular MICA levels by preventing those shedding and release may contribute to the clearance of the pathogenic cells and the prevention of HCC. In this study, we constructed a convenient MICA shedding assay system and, by a comprehensive screening, we identified several compounds which may prevent MICA shedding from the cells.

Methods: We constructed lentivirual vector expressing MICA protein tagged with nano-luciferase (nanoLuc) in its N terminal because N terminal of MICA is excreted from the cells by shedding. We expressed this construct in HBV replicating cells, HepG2.2.15, and after adding compounds, we determined the luciferase activities in the supernatant and cell lysates.

Results: We picked up the samples which showed low activities in supernatant with high activities in the cell lysates, indicating low levels of shedding and high levels of remaining cellular MICA. Out of over 1,000 compounds screened, we determined four candidate compounds which efficiently prevented MICA shedding. We confirmed the effects of these compounds in other cell systems infected with HBV.

Conclusion: Because these compounds enhanced the expression levels of cellular MICA and decreased its shedding in HBV-replicating human hepatocytes, using these compounds may be favorable during HBV chronic infection to enhance the NK cell-mediated clearance of the pathogenic hepatocytes and to prevent HCC.

Citation Format: Motoko Ohno, Motoyuki Otsuka, Takahiro Kishikawa, Takeshi Yoshikawa, Akemi Takata, Kazuhiko Koike. Prevention of MICA shedding from HBV infected hepatocytes to activate NK cells for better clearance of malignant foci and virus-infected cells. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 4456. doi:10.1158/1538-7445.AM2015-4456
Abstract 4464: Effect of bacterial quorum sensing lactones on human pancreatic carcinoma cells
Authors: Kumar Senthil R. , Bryan Jeffrey
Published: August 2015

INTRODUCTION

Pancreatic cancer (PC) is the fourth leading cause of cancer related mortality in the United States. In spite of the discovery of gemcitabine, administered either alone or in combination with other drugs, the survival rate of 5% has remained unchanged for several decades now. In this study, we hypothesize that bacterial quorum sensing molecules such as N-3-oxo-dodecanoyl-L-homoserine lactone (O-DDHSL) could affect pancreatic carcinoma cell phenotype and characteristics. Elucidation of the mechanism of action of O-DDHSL could lead to the development of more effective analogs and novel therapeutic targets in PC.

MATERIALS AND METHODS

Human pancreatic carcinoma cells (Panc-1 and Aspc-1) was grown in DMEM-F12 media with 10% FBS at 37oC. Normal pancreatic ductal epithelial cells (HPDE) were maintained in keratinocyte serum free media. Total RNA was extracted from the cells and converted to cDNA which served as a template for qRT-PCR amplification in the presence of various gene primers. The effect of O-DDHSL on pancreatic carcinoma cell phenotype, apoptosis, viability, migration, proliferation and colony forming ability was investigated.

RESULTS

Both Panc-1 and Aspc-1 cell viability decreased in a concentration dependent manner in the presence of O-DDHSL (IC50 = 120 ± 5 μM; P ≤ 0.02) and (IC50 = 48 ± 8 μM; P ≤ 0.02), respectively. O-DDHSL had only minimal effect on HPDE cell viability. Apoptosis and inhibition of cell migration in Panc-1 and Aspc-1 cells was observed in the presence of O-DDHSL. Variable effect on migration associated genes such as IQGAP-1, cofilin and rhoC was noted in these cells upon O-DDHSL treatment as determined by qRT-PCR as well as protein expression by Western analysis.

CONCLUSIONS

In conclusion, the combined findings from our study demonstrate that O-DDHSL decreases pancreatic carcinoma cell viability, promotes apoptosis by activating caspases, and inhibits cell migration. O-DDHSL also modulates genes responsible for migration such as RhoC, cofilin and IQGAP-1 in these cells.

Citation Format: Senthil R. Kumar, Jeffrey Bryan. Effect of bacterial quorum sensing lactones on human pancreatic carcinoma cells. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 4464. doi:10.1158/1538-7445.AM2015-4464
Abstract 4466: New biomarkers to optimize preclinical development of the PDI inhibitor XCE853
Authors: Prevost Gregoire Pierre , Garrido Marine , Xu Shili , Lefebvre Celine , Chauchereau Anne , Carniato Denis , Briand Jean François , Gutmann Mathieu , Serova Maria , Tijeras-Raballand Annemilai , De Grammont Armand , Raymond Eric , Gespach Christian , Sabbah Michèle , Neamati Nouri , Pitty Marc-Henry , Foster Paul
Published: August 2015

Protein disulfide isomerase (PDI) is a chaperone protein that regulates oxidative protein folding as well as cell viability. Increased PDI levels have been documented in a variety of human cancers, including ovarian, prostate, and lung cancers. Inhibition of PDI activity leads to apoptosis in cancer, suggesting that PDI is a promising druggable target. XCE853 was originally identified within a family of synthetic compounds displaying a preferential antiproliferative activity on drug-resistant human cancer cells (Gutmann et al, AACR annual meeting, 2013). XCE853 inhibits in vitro recombinant PDI activity and blocks in vitro proliferation of human tumor cells with IC50s in the nanomolar range through an irreversible cytolysis (Prevost et al, AACR annual meeting, 2014). By contrast, XCE853 has no or limited antiproliferative activity on several human non-tumoral immortalized cells (MCF10, RPE1 or RWPE1) even at high concentrations. The ex-vivo approach using fresh human hepatocellular carcinoma explants incubated 48h with XCE853 (100nM) has shown the decrease of the proliferation (KI-67 labeling) with an apoptosis induction (Caspase-3 labeling). In addition, XCE853 which displayed an excellent oral bioavailability in mouse blocks growth of human ovarian cancer using the in vivo OVCAR-3 xenograft model leading to complete tumor growth arrest even after the cessation of treatment.

To optimize the preclinical and clinical development, we are now characterizing new biomarkers to 1) stratify the tumors which are sensitive and non-sensitive to XCE853 and to 2) identify the optimal dose regimen.

Altogether, these data support further efforts on XCE853 preclinical studies to prepare the Phase-I clinical study entry.

Citation Format: Gregoire Pierre Prevost, Marine Garrido, Shili Xu, Celine Lefebvre, Anne Chauchereau, Denis Carniato, Jean François Briand, Mathieu Gutmann, Maria Serova, Annemilai Tijeras-Raballand, Armand De Grammont, Eric Raymond, Christian Gespach, Michèle Sabbah, Nouri Neamati, Marc-Henry Pitty, Paul Foster. New biomarkers to optimize preclinical development of the PDI inhibitor XCE853. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 4466. doi:10.1158/1538-7445.AM2015-4466
Abstract 4467: Targeting of chemokine receptor CXCR4 inhibits invasion and sensitizes melanoma cells to chemotherapeutic agents
Authors: Pandey Manoj K. , Karelia Deepkamal , Kale Vijay P. , Gowda Krishne , Sharma Arun K. , Neves Rogerio I. , Amin Shantu G.
Published: August 2015

Malignant melanoma is highly metastatic and resistant to conventional therapies. Although numerous clinical approaches for treating melanoma have been adopted, none led to a remarkable therapeutic effect. Recently new drugs have been developed for treatment of metastatic melanoma, and hold great promise; however complete effects are still distant dream because of resistance. We have learnt that the most aggressive melanomas are generally resistant to strategies which target one signaling pathway. Thus an agent targeting multiple signaling pathways would maximize therapeutic outcome. We have identified one such agent named as gambogic acid (GA) that is natural and derived from medicinal plant Garcinia hanburyi (also called mangosteen or kokum). Moreover this agent has a potential to target several pathways responsible for survival and chemoresistance of several cancers including melanoma. Stromal cell-derived factor 1α (SDF-1α) and its receptor, CXC chemokine receptor 4 (CXCR4), has been associated with inferior survival, poor prognosis, chemoresistance and metastasis of malignant melanoma. We, therefore, investigated whether GA could inhibit CXCR4 signaling and suppress invasion of malignant melanoma cells. Through docking studies we predicted that GA directly interacts with CXCR4. This xanthone down-regulates the expression of CXCR4 on melanoma cells in a dose- and time-dependent manner. Suppression of CXCR4 expression by GA correlated with both inhibition of SDF-1α-induced invasion of melanoma cells. GA also inhibited expression of MMP-9, VEGF, c-Myc and phosphorylation of Erk1/2 in melanoma cells. Importantly, GA markedly enhances the efficacy of PLX-4032 and dacarbazine against melanoma cells.

Citation Format: Manoj K. Pandey, Deepkamal Karelia, Vijay P. Kale, Krishne Gowda, Arun K. Sharma, Rogerio I. Neves, Shantu G. Amin. Targeting of chemokine receptor CXCR4 inhibits invasion and sensitizes melanoma cells to chemotherapeutic agents. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 4467. doi:10.1158/1538-7445.AM2015-4467
Abstract 447: Tumor microenvironment in pancreatic cancer (PDAC): interplay between tumor cells, stromal cells and immune cells
Authors: Karamitopoulou Eva , Wartenberg Martin , Galván José A. , Zlobec Inti , Lugli Alessandro , Perren Aurel
Published: August 2015

Introduction: Tumor budding is considered to be the morphologic correlate of epithelial-mesenchymal-transition (EMT) and is an independent prognostic factor in pancreatic ductal adenocarcinoma (PDAC). Here we explore the interactions between tumor budding cells, stromal cells and immune cells in the microenvironment of the tumor buds in PDAC.

Methods: Multi-punch tissue-microarrays (TMAs) containing punches from the tumor center and the tumor front, including tumor buds, from a well characterized cohort of 120 PDAC patients, were stained immunohistochemically and by mRNA-in-situ-Hybridization (mRNA-ISH)for the EMT markers E-cadherin, β-catenin, Snail, ZEB1, ZEB2, N-cadherin and Twist and for the immune cell markers CD8, CD4, Foxp3, M1- and M2-macrophages. Expression in budding- and stromal cells and immune-cell counts in the microenvironment of the tumor buds were compared with findings in the main tumor.

Results: Tumor buds showed loss of E-cadherin and b-catenin and overexpressed ZEB1 and ZEB2 compared with the neoplastic cells of the main tumor both at protein and mRNA level (p<0.0001 respectively). Stromal cells surrounding tumor buds showed increased protein and mRNA levels of the transcription factors Snail, ZEB1 and ZEB2. The immune microenvironment of the tumor buds composed of numerous Foxp3+cells and M1-macrophages while M2-macrophages were significantly reduced and CD8+cells were almost absent, in contrast to the main tumor.

Conclusions: Tumor budding cells show a shift towards EMT-promoting profiles at protein and mRNA level, compared with the neoplastic cells of the main tumor. Stromal cells surrounding tumor buds express high levels of Snail, ZEB1 and ZEB2 suggesting that some stromal cells may represent complete mesenchymally transformed tumor cells or alternatively that there is a special phenotype of cancer associated fibroblasts supporting EMT-Type tumor-budding through cellular crosstalk in the tumor microenvironment of PDAC. Moreover, there is a tumor favoring immune cell composition in the immediate microenvironment of the tumor buds. Our findings suggest a close interaction of the stromal and immune response with the EMT process in PDAC. The combined assessment of host-associated factors such as stromal and immune response and tumor-associated factors such as EMT-type tumor-budding could help us to achieve superior prognostication and patient stratification than either factor alone.

Citation Format: Eva Karamitopoulou, Martin Wartenberg, José A. Galván, Inti Zlobec, Alessandro Lugli, Aurel Perren. Tumor microenvironment in pancreatic cancer (PDAC): interplay between tumor cells, stromal cells and immune cells. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 447. doi:10.1158/1538-7445.AM2015-447
Abstract 4470: Targeting autophagy and apoptosis to enhance anticancer effect on esophageal carcinoma in vitro
Authors: Cai Yan , Cai Jiajing , Ma Qiang , Xu Yuan , Wang Dongsheng , Guo Xiaolan
Published: August 2015

Esophageal cancer is one of the malignancies threatening human health with high incidence and a very low 5-year survival rate with poor outcomes in the developing countries. As one of the main treatment for esophageal cancer, chemotherapy has been played indispensable roles along with more and more chemoresistance and side effects arised though. Recent studies showed that induction of autophagy has emerged as a drug resistance mechanism that promotes cancer cell survival via self-digestion. Therefore targeting autophagy in cancer might be another promising approach of therapeutics. In this study, chloroquine (CQ) was used as the autophagy inhibitor, which has been the anti-malarial drug with 60 years history and safety performance, to study esophageal carcinoma cells EC109 in vivo and explore the potential molecular mechanisms. Chloroquine was employed to treat EC109 cells at different dosages and time points, and MTT results showed that the viability of EC109 was reduced dramatically in a dose dependent manner after 24 hours treatment. Moreover, we found that CQ could inhibit the migration and invasion abilities through scratch test and cell invasion Transwell chamber test, also significantly suppressed the colony formation using soft agar assay. Further investigation showed that mTOR pathway was activated with CQ exposure, and autophagy marker LC3I/II conversion decreased at early-stage, with obvious elevation eventually. More interestingly, we observed that CQ promoted the EC109 cells apoptosis by flow cytometry, also the mRNA expression of apoptosis associated proteins, such as Fas and Bax were increased. According to our findings, chloroquine showed its anticancer effect through modulating autophagy and promoting cell apoptosis, which might have promising therapeutic benefits for esophageal carcinoma.

Citation Format: Yan Cai, Jiajing Cai, Qiang Ma, Yuan Xu, Dongsheng Wang, Xiaolan Guo. Targeting autophagy and apoptosis to enhance anticancer effect on esophageal carcinoma in vitro. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 4470. doi:10.1158/1538-7445.AM2015-4470
Abstract 4479: ONC201 is non-toxic at efficacious doses in vitro and in vivo
Authors: Allen Joshua E. , El-Deiry Wafik S.
Published: August 2015

ONC201 is a small molecule that is entering clinical trials based on its favorable preclinical profile that includes robust anti-tumor activity in several preclinical models of advanced cancers. Despite inducing a strong cytotoxic response in tumor cells, ONC201 does not induce cell death in normal cells under the same conditions. In addition to a lack of cytoxicity towards normal cells, ONC201 also does not appear to be genotoxic based on gamma-H2AX assessment. Recent mechanistic studies have implicated the early stage induction of the ER stress response by ONC201, which includes induction of DR5 as part of its pro-apoptotic response. Biochemical studies reveal that ONC201 does not induce DR5 in normal cells at doses that cause strong DR5 induction in tumor cells.

In accordance with in vitro profile, efficacious ONC201 dose schedules in murine models are also well tolerated without apparent alterations of normal tissue histology or significant loss of body weight. Similar observations were revealed in GLP toxicology studies in beagle dogs and Sprague-Dawley rats. Dogs received a single dose of 0, 4.2, 42, or 120 mg/kg by oral gavage, which represents a human equivalent of 0, 125 mg, 1.25 g, and 3.57 g, respectively. Rats received 0, 12.5, 125, or 225 mg/kg ONC201 by oral gavage, which represents a human equivalent of 0, 125 mg, 1.25 g, and 2.25 g, respectively. There were no deaths or dose-limiting toxicities. The only findings that were observed in both rats and dogs occurred at the highest doses tested and were mild and reversible: decreased activity (no indications of anemia), decreased food consumption (weight loss only seen in rats), and salivation. The NOAEL was at least 42 mg/kg in dogs and at least 125 mg/kg in rats.

In summary, these in vitro and in vivo studies support that ONC201 has a large and favorable therapeutic index that is compelling given its powerful single agent anti-tumor efficacy. The coupling of the strong efficacy and safety profile of ONC201 is unique among cancer therapies and opens a broad range of clinical applications for its future development, including potential for combinatorial therapeutics.

Citation Format: Joshua E. Allen, Wafik S. El-Deiry. ONC201 is non-toxic at efficacious doses in vitro and in vivo. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 4479. doi:10.1158/1538-7445.AM2015-4479
Abstract 4485: Anticancer effect of platinum(IV) complex against cisplatin-resistant human ovarian cancer
Authors: Okamoto Yoshinori , Tobe Takao , Ueda Koji , Kojima Nakao
Published: August 2015

Background: Platinum(IV) [Pt(IV)] complexes are one of the attractive next-generation anticancer drugs. Beside Pt(II)-based drugs such as cisplatin, carboplatin, and oxaliplatin, Pt(IV) complexes show less adverse effects and can be administrated orally. In addition, because Pt(IV) complexes adopt octahedral coordination geometry, it also have an advantage that can be coordinated with more functional ligands than Pt(II) complexes. These different features prompted us to investigate the effectiveness of Pt(IV) complexes on cisplatin-resistant cancer cells with its mechanisms. This study aims to elucidate the anti-cancer effects of Pt(IV) complexes. Materials and Methods: The cytotoxicity of cis-diammine-tetrachloro-Pt(IV) [cis-Pt(IV)] and cisplatin was determined using human ovarian cancer cell lines (A2780), and its cisplatin-resistant subline (A2780cis), with their influx/efflux patterns using ICP-MS. Pt-DNA-crosslink formation was also determined by agarose gel electrophoresis, in which the crosslink was quantified by decrease of ethidium bromide staining of DNA. Results and Discussion: Cisplatin completely failed to decrease the survival of A2780cis cells at a concentration that killed A2780, whereas cis-Pt(IV) exerted cytotoxicity in both cell lines. Pt-DNA-crosslink formation assay showed cis-Pt(IV) bound to DNA only in the presence of reductant such as glutathione (GSH) or ascorbic acid, indicating cisplatin production. However, it is also known excess amount of GSH suppress DNA binding of cisplatin. Indeed, intracellular GSH level in A2780cis cells was about 3 times higher as compared with A2780 cells; therefore cisplatin would be inactivated in A2780cis cells. These findings suggest that cis-Pt(IV) is actually reduced to cisplatin in GSH-rich environment and then induce cytotoxicity by somehow escaping from GSH-dependent inactivation. Pt(IV)-specific subcellular distribution might contribute this paradoxical advantageous effect of GSH on cis-Pt(IV). Intracellular accumulation of Pt was reduced by competing a related transporter CTR1 by copper in cis-Pt(IV) treatment, whereas cytotoxicity was not suppressed. Therefore CTR1 is involved in the intake of Pt complexes including cis-Pt(IV), but the alternative pathway(s) could be responsible for anti-cancer effect of Pt(IV) complexes.

Citation Format: Yoshinori Okamoto, Takao Tobe, Koji Ueda, Nakao Kojima. Anticancer effect of platinum(IV) complex against cisplatin-resistant human ovarian cancer. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 4485. doi:10.1158/1538-7445.AM2015-4485
Abstract 4488: Folate receptor targeted therapy using small molecule drug conjugates constructed with high affinity antifolate ligands
Authors: Leamon Christopher P. , Reddy Joseph A. , Vlahov Iontcho R. , You Fei , Klein Hanna F. , Kleindl Paul J. , Nelson Melissa , Vetzel Marilynn , Klein Patrick J. , Matherly Larry H. , Gangjee Aleem
Published: August 2015

Folate receptor (FR)-targeted small molecule drug conjugates (SMDC) have shown promising clinical results. To date, all related SMDCs have been constructed with folic acid. Whereas folic acid displays very high affinity for the FR (kd ∼ 0.1 nM), antifolates with similar high binding affinity and specificity have also been reported. This knowledge prompted our investigation towards substituting the folic acid targeting moiety in our SMDCs with high affinity antifolates, such as CB3717 (a thymidylate synthase inhibitor), and the glycinamide ribonucleotide formyltransferase (GARFTase) inhibitors, AG94 and AG147, thereby testing the hypothesis that greater antitumor activity should result from the combination of inhibitors. Using mice bearing FR-positive human tumor xenografts, we observed curative activity with a CB3717-based vinca alkaloid SMDC under conditions where no cures resulted with vintafolide (EC145) treatment. Similarly, tubulysin SMDCs constructed with AG94 and AG147 were found to produce curative activity using a dosing regimen where a folic acid-tubulysin conjugate did not. Of note, treatment of mice with simple co-mixtures of the antifolates and untargeted drugs was found to be ineffective, thereby confirming the need for linking the two moieties together to produce maximal activity. Taken together, these studies demonstrate that high affinity antifolates can be used in an SMDC format serving as both a high affinity targeting ligand and as a second drug having different, and perhaps a complementary mechanism of action to yield greater antitumor activity.

Citation Format: Christopher P. Leamon, Joseph A. Reddy, Iontcho R. Vlahov, Fei You, Hanna F. Klein, Paul J. Kleindl, Melissa Nelson, Marilynn Vetzel, Patrick J. Klein, Larry H. Matherly, Aleem Gangjee. Folate receptor targeted therapy using small molecule drug conjugates constructed with high affinity antifolate ligands. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 4488. doi:10.1158/1538-7445.AM2015-4488
Abstract 4500: Nisin ZP, a food preservative, has antitumor potential for head and neck cancer and extends survival
Authors: Kamarajan Pachiyappan , Hayami Takayuki , Matte Bibiana , Liu Yang , Ramamoorthy Ayyalusamy , Worden Francis P. , Kapila Sunil , Kapila Yvonne
Published: August 2015

The use of small antimicrobial agents or bacteriocins, like nisin, to treat cancer is a new approach that holds great promise. Nisin exemplifies this new approach because it has been used safely in humans for many years as a food preservative, and recent laboratory studies support its anti-tumor potential in head and neck cancer. Previously, we showed that nisin (2.5%, low content) has antitumor potential in head and neck squamous cell carcinoma (HNSCC) in vitro and in vivo. The current studies explored a naturally occurring variant of nisin (nisin ZP; 95%, high content) for its antitumor effects in vitro and in vivo. Nisin ZP induced the greatest level of apoptosis in HNSCC cells compared to low content nisin. HNSCC cells treated with increasing concentrations of nisin ZP exhibited increasing levels of apoptosis and decreasing levels of cell proliferation and clonogenic capacity. Nisin ZP-induced apoptosis was defined by increased ethidium bromide and acridine orange (EB/AO) staining and by increased levels of calpain-dependent PARP cleavage. Importantly, nisin ZP also induced apoptosis dose-dependently in HUVEC endothelial cells and concomitant decreases in vascular sprout formation. Nisin ZP inhibited sphere formation of HNSCC cells in vitro and tumorigenesis in vivo and long term treatment extended survival. In summary, nisin ZP exhibits greater antitumor effects than low content nisin, and thus has the potential to serve as a novel therapeutic for HNSCC.

Note: This abstract was not presented at the meeting.

Citation Format: Pachiyappan Kamarajan, Takayuki Hayami, Bibiana Matte, Yang Liu, Ayyalusamy Ramamoorthy, Francis P. Worden, Sunil Kapila, Yvonne Kapila. Nisin ZP, a food preservative, has antitumor potential for head and neck cancer and extends survival. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 4500. doi:10.1158/1538-7445.AM2015-4500
Abstract 4501: Development of novel Lithocholic acid carboxamides with antiproliferative and pro-apoptotic effects on human cancer cells
Authors: Ramisetti Srinivasa R. , Karelia Deepkamal , Neagley Melanie , Amin Shantu , Sharma Arun K.
Published: August 2015

Bile acids, the products of cholesterol catabolism, are synthesized in the liver and subsequently excreted into the bile canaliculus and the digestive tract as N-acyl conjugates of glycine or taurine. Among all bile acids, Lithocholic acid (LCA) and Ursodeoxycholic acid (UDCA) conjugates have been reported to inhibit the growth of various human cancer cells both in vitro and in vivo. Most importantly LCA has been shown to inhibit growth of different cancer cell lines including prostate and neuroblastoma cells, while sparing normal cells. In addition, a report has shown LCA-amphiphiles to be effective anti-tumor agents in a colon cancer xenograft model. In contrast, there are reports indicating that LCA may promote dimethyl hydrazine (DMH) induced colon carcinogenesis and may also act as a carcinogen itself. This indicates that in spite of having anti-cancer properties, LCA may pose a risk of developing colon cancer. We hypothesized that LCA structure can be optimized to eliminate the carcinogenesis promoting properties, while enhancing its potency to create effective therapeutics against colon, prostate, and other cancers. Therefore, we designed and synthesized a series of novel cyclic and acyclic LCA-carboxamides by blocking the acid functionality of LCA. The cytotoxicity of the novel analogs was evaluated against colon (HT29) and prostate (DU145) cancer cells. Three analogs ASR-320, ASR-322 and ASR-339 inhibited the viability of these cells with an EC50 value of 5-10 μM, while LCA had no effect on cell viability at highest dose used (50 μM). The results suggests that the new analogs are at least 10 times more potent than the parent compound, LCA. One of the agents, induced apoptosis in both prostate (DU145) and colon (HT29) cancer cell lines, as indicated by Caspase 3 activation and PARP degradation. Currently, we are further optimizing the structure based on the initial leads and investigating efficacy and mechanistic studies in prostate and colon cancer models. Detailed results of these investigations will be presented.

Citation Format: Srinivasa R. Ramisetti, Deepkamal Karelia, Melanie Neagley, Shantu Amin, Arun K. Sharma. Development of novel Lithocholic acid carboxamides with antiproliferative and pro-apoptotic effects on human cancer cells. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 4501. doi:10.1158/1538-7445.AM2015-4501
Abstract 4521: Evaluation of the relationship between serum exposure, receptor (GITR) availability and tumor suppression following administration of the anti-GITR antibody DX400 in mouse syngeneic tumor models
Authors: Ovacik Ayse Meric , Shinsky-Bjorde Natalie , Hodges Douglas , Antonenko Svetlana , Ueda Roanna , Mauze Smita , Gu Danling , Wiswell Derek , Zhang Shuli , Beebe Amy , Tabrizi Mohammad
Published: August 2015

GITR is a type I transmembrane protein of the tumor necrosis factor receptor superfamily which is expressed primarily on T lymphocytes and natural killer cells. Ligation of GITR on activated T cells provides a costimulatory signal that positively modulates antigen-specific T cell responses, leading to enhanced cellular and humoral immunity. The anti-GITR antibody DX400 is a murinized agonistic monoclonal antibody that targets mouse GITR, and has shown tumor growth inhibition in mouse syngeneic tumor models. In this study we examined the pharmacokinetic/pharmacodynamics (PK/PD) properties of DX400. Studies were conducted to examine the potential relationships between anti-GITR antibody serum exposure (PK) and receptor availability (PD) on the relevant T-cell subsets. The concentrations of DX400 in serum were determined using an electrochemiluminescence (ECL) assay, and the availability of receptor (GITR) was determined using flow cytometry. Non-linear PK properties for DX400 were observed over the dose range examined. In line with changes in serum concentration-time profiles for the antibody, a dose dependent effect in receptor availability was also observed. The relationships between receptor availability, serum concentrations of DX400 and tumor suppression were described by a mechanistic PK/PD model. This analysis provided an estimated potency (EC50) value of 4.2 ng/mL for blood receptor engagement on T-cell subsets.

Citation Format: Ayse Meric Ovacik, Natalie Shinsky-Bjorde, Douglas Hodges, Svetlana Antonenko, Roanna Ueda, Smita Mauze, Danling Gu, Derek Wiswell, Shuli Zhang, Amy Beebe, Mohammad Tabrizi. Evaluation of the relationship between serum exposure, receptor (GITR) availability and tumor suppression following administration of the anti-GITR antibody DX400 in mouse syngeneic tumor models. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 4521. doi:10.1158/1538-7445.AM2015-4521
Abstract 4544: Synthesis and biological evaluation of novel 6-substituted pyrrolo[2,3-d]pyrimidines as targeted antifolates
Authors: Gangjee Aleem , Golani Lalit , Wallaceb Adrianne , Matherly Larry H.
Published: August 2015

Reduced folates are essential cofactors for the biosynthesis of purines and pyrimidines. Since humans do not synthesize folate, it is necessary to obtain these cofactors from dietary sources. In mammals, three specialized systems exist that mediate membrane transport of folates and antifolates across biological membranes. These include the reduced folate carrier (RFC), the primary route for the uptake of folates and antifolates in mammalian cells, folate receptors (FRs) α and β, and the proton-coupled folate transporter (PCFT). Whereas RFC is ubiquitously expressed, FRs and PCFT show a narrower pattern of tissue expression. Toxicity of clinically used antifolates is attributed to their lack of selectivity for tumor cells over normal cells, likely due in part to their RFC transport. Antifolates with tumor-selective FR and/or PCFT drug uptake would circumvent major toxicities of currently used antifolates. We previously described the synthesis and biological activity of AGF23, a 6-substituted pyrrolo[2,3-d]pyrimidine. Against FRα- and PCFT-expressing KB human tumor cells, AGF23 was potently inhibitory (IC50 = 1.9 nM). Two novel isosteres of AGF23, AGF215 and AGF234 with oxygen and nitrogen in place of carbon at C11, respectively, were designed and synthesized. This simple isosteric replacement of carbon with heteroatoms in the bridge affords compounds with different chain lengths, angles, conformations and extra hydrogen bond donors and/or acceptors compared to the parent carbon chain analog. AGF215 and AGF234 were compared to AGF23 as inhibitors of proliferation of engineered Chinese hamster ovary (CHO) cells expressing human PCFT (R2/PCFT4), FRα (RT16), or RFC (PC43-10). AGF215 and AGF234, like AGF23, were inactive toward PC43-10 cells. AGF23 and AGF234 showed low level activity toward R2/PCFT4 cells (IC50∼200 nM), whereas AGF215 was inactive. Toward FRα-expressing RT16 cells, all the analogs were potently active with IC50 values of 1-6 nM. Toward KB tumor cells, AGF215 and AGF234 were highly potent with IC50 values of 3 and 1.3 nM, respectively. AGF215 and AGF234 are potential analogs for further preclinical studies and analog design.

Citation Format: Aleem Gangjee, Lalit Golani, Adrianne Wallaceb, Larry H. Matherly. Synthesis and biological evaluation of novel 6-substituted pyrrolo[2,3-d]pyrimidines as targeted antifolates. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 4544. doi:10.1158/1538-7445.AM2015-4544