bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 4452: Regulation of histone deacethylase 8 expression by inhibitory GTP binding proteins signaling via JNK pathway in H1299 non-small-cell lung cancer cells
Authors: Hahn Hwa-jeong , Juhnn Yong-Sung
Published: May 2015

Histone deacetylases (HDACs) have been reported to regulate the cell growth, proliferation, and apoptosis, and HDAC8 seems to be involved in tumor growth and progression. Inhibitory heterotrimeric GTP-binding proteins (Gαi) are activated by seven transmembrane receptors, and regulate cell proliferation by regulating the activity of adenylate cyclase, phospholipase C beta, and ion channels. This study investigated the effects of Gαi signaling system on the expression of HDAC8 and the underlying mechanism in lung cancer cells. When Gi signaling system was activated by expressing Gαi1Q204L, a constitutively active mutant Gαi1, the expression of HDAC8 protein and mRNA was increased in H1299 non-small-cell lung cancer (NSCLC) cells. Inhibition of transcription by treatment with Actinomycin D did not block the increase in mRNA, and the degradation of HDAC8 mRNA was facilitated in Gαi1Q204L -expressing cells, suggesting Gαi1 signaling system increases HDAC8 expression by stabilizing mRNA. In addition, treatment with adenosine or an adenosine receptor agonist, CCPA (2-chloro-N(6)-cyclopentyladenosine), which activate Gαi-coupled receptors, also increased HDAC8 mRNA and protein levels. The effect of adenosine was blocked by treatment with DPCPX (8-Cyclopentyl-1,3-dipropylxanthine), an adenosine A1 receptor antagonist. JNK phosphorylation was inhibited by treatment with the adenosine and CCPA. Treatment with a JNK inhibitor, SP600125 increased HDAC8 expression. Adenosine inhibited the activation of MKK4 and CDC 42, which act upstream of JNK. Activation of Gαi signaling stimulated proliferation of H1299 cells, and the stimulating effect was abolished by knockdown of Gαi1. Thus, we conclude that Gαi signaling increases HDAC8 expression via JNK pathway, resulting in stimulation of cell proliferation of H1299 lung cancer cells. This study suggests that Gαi signaling may be involved in epigenetic control of gene expression by regulating HDAC8 expression.

Citation Format: Hwa-jeong Hahn, Yong-Sung Juhnn. Regulation of histone deacethylase 8 expression by inhibitory GTP binding proteins signaling via JNK pathway in H1299 non-small-cell lung cancer cells. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 4452. doi:10.1158/1538-7445.AM2014-4452
Abstract 446: BAP1 mutation in mesothelioma and “BAP1 Cancer Syndrome”
Authors: Nasu Masaki , Napolitano Andrea , Pastorino Sandra , Tanji Mika , Flores Erin , Baumann Francine , Powers Amy , Gaudino Giovanni , Pass Harvey I. , Yang Haining , Carbone Michele
Published: May 2015

Malignant mesothelioma (MM) is a lethal cancer whose pathogenesis results from complex interactions between host genetics and environmental carcinogens, such as asbestos and erionite fibers. Recently, BAP1 (BRCA associated protein 1) has been identified as a novel MM tumor suppressor gene located at 3p21, a region frequently deleted in MM, which encodes for a deubiquitinase enzyme known to target histones and other proteins.

We discovered that germline BAP1 mutations cause a novel cancer syndrome characterized by a significant excess of both pleural and peritoneal MM, uveal and cutaneous melanoma and possibly other tumors. In the same study, we reported that 22% sporadic MM tumors, among the Caucasian population, harbored somatic BAP1 mutations. Several other studies supported a relevant role of BAP1 in MM. However, a significant variation in the frequency of BAP1 mutation was found across different studies and populations. In fact, one research group found BAP1 gene altered in 61% of tumor samples from a Japanese cohort, pointing out to a possible influence of ethnicity on the prevalence of BAP-1 alterations among MM patients. However, the limitations in tumor sample sizes and methodological differences across studies do not allow for conclusive associations. We are now further analyzing BAP1 status and possible clinicopathological associations using different and more sensitive methods, such as MPLA, DNA and RNA sequencing, DNA copy number and methylation. Our preliminary results indicate that, regardless of ethnicity, BAP1 plays a crucial role in MM pathogenesis. More experiments are urgently needed to see whether BAP1 expression could be used in diagnostic, prognostic, or therapeutic settings. The impact of this work will extend to other cancers with BAP1 mutations.

Citation Format: Masaki Nasu, Andrea Napolitano, Sandra Pastorino, Mika Tanji, Erin Flores, Francine Baumann, Amy Powers, Giovanni Gaudino, Harvey I. Pass, Haining Yang, Michele Carbone. BAP1 mutation in mesothelioma and “BAP1 Cancer Syndrome”. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 446. doi:10.1158/1538-7445.AM2014-446
Abstract 4462: Structure-activity relationship studies for PSMA-targeted tubulysin conjugates
Authors: Wang Kevin , Hahn Spencer , Santhapuram Hari , Qi Longwu , Kleindl Paul , Vaughn Jeremy , You Fei , Reddy Joe , Dorton Ryan , Leamon Christopher , Vlahov Iontcho
Published: May 2015

Prostate-specific membrane antigen (PSMA) is a cell-surface receptor expressed on prostate cancer cells. Recent findings suggest that PSMA is also abundant on newly formed blood vessels which supply most non-prostatic solid tumors; including lung, colon, breast, renal, liver, and pancreatic carcinomas. These reports have provided solid support for the concept of designing drug conjugates for PSMA-targeted cancer therapy. Here we describe our structure-activity relationship analyses resulting in the discovery of highly potent PSMA-targeted tubulysin conjugates. In our study, more than thirty ligand-drug constructs were designed and synthesized based on strategically placing a variety of chemical groups responsible for evoking a targeted biological effect in vitro and in vivo. The optimized molecular architecture is comprised of a high affinity PSMA-binding urea-type ligand, a water soluble peptide- or carbohydrate-based spacer unit, a cleavable disulfide-containing self-immolative linker system, and the highly potent chemotherapy drug tubulysin which inhibits microtubule activity . One of the conjugates possessed superior targeted activity and was selected for further preclinical development.

Citation Format: Kevin Wang, Spencer Hahn, Hari Santhapuram, Longwu Qi, Paul Kleindl, Jeremy Vaughn, Fei You, Joe Reddy, Ryan Dorton, Christopher Leamon, Iontcho Vlahov. Structure-activity relationship studies for PSMA-targeted tubulysin conjugates. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 4462. doi:10.1158/1538-7445.AM2014-4462
Abstract 4477: pH-responsive PEG-HPAH micelles for drug delivery and their potential in cancer therapy
Authors: Xu Qin , Yu Jingshuang , Zhu Bangshang
Published: May 2015

pH-responsive hyperbranched polymers have attracted much attention due to their unique properties for tumor-targeted drug delivery. In this study, we describe a pH-responsive drug carrier, poly (ethylene glycol) (PEG)-hyperbranched polyacylhydrazone (HPAH), which can form nanoscale micelles to be used as anti-cancer drug carriers with pH-controlled drug release. The molecular structure of PEG-HPAH was confirmed by nuclear magnetic resonance spectroscopy (NMR) and Fourier transform infrared spectroscopy (FTIR). The drug-loaded micelles with a diameter of approximately 190 nm, were prepared using a dialysis method against PBS with a pH of 8.0. The drug-loaded micelles showed the desired pH-dependent drug release properties. The drug release levels were low at neutral and alkaline pH, but increased significantly with a decrease in the pH of the medium. Intracellular uptake results indicated that the PEG-HPAH-drug micelles could efficiently deliver chemotherapeutic drugs into the cells. In addition, it was found that the subcellular localization of the drug-loaded micelles was different from that of free drugs, in which the drug-loaded micelles were mainly in the cytoplasm. The docetaxel (DTX)-loaded PEG-HPAH micelles presented a high cytotoxic activity against tumor cells in vitro. When combined with the administration of glucose, the PEG-HPAH-DTX micelles exhibited a superior anti-tumor efficacy and a lower systemic toxicity in vivo. The biodistribution profile showed increased accumulated drug levels in tumor tissue and plasma in micelles treated group. The results indicate that the nanoscale PEG-HPAH-DTX micelles may serve as a novel selective tumor-targeting drug delivery system.

Citation Format: Qin Xu, Jingshuang Yu, Bangshang Zhu. pH-responsive PEG-HPAH micelles for drug delivery and their potential in cancer therapy. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 4477. doi:10.1158/1538-7445.AM2014-4477
Abstract 4485: Synthesis of targeted docetaxel-polymer conjugate and its anti-tumor efficacy
Authors: Lee Young Bok , Kim Deog Joong , Yang Miyoung , Ahn Chang-Ho , Nan Anjan
Published: May 2015

Over the last decade, it has been shown that nanopolymer therapeutics can be engineered to target cancer cells specifically and can deliver the cancer agent in a time-release fashion, thereby leaving normal healthy cells unaffected by toxic drugs such as docetaxel. Docetaxel has been used to treat several types of cancers, but it has provided pharmaceutical challenges due to its poor solubility and toxicities associated with the co-solvents. Given that nanotechnology can enhance the specificity, efficacy and safety of cancer agents and with the need of a new alternative formulation for docetaxel delivery, we developed a poly N-(2-hydroxypropyl) methacrylamide (HPMA)-Docetaxel (Doc) conjugate with Folate (FA) targeting ligand and evaluated the effect of the product in both in vitro human cancer cell lines and in vivo mice xenograft tumor models. Analysis of the product displayed the incorporation of 7.9% of docetaxel and 2.1% of folate with 38.6kDa MW and 1.92 of polydispersity, and an improvement in water solubility. The product inhibited the proliferation of a variety of human cancer cells in nM ranges in in vitro study. The maximal tolerated dose (MTD) of the product in mice was more than 150 mg/kg (as a docetaxel amount), which is much higher than that of free docetaxel (∼25 mg/kg). In mice bearing tumor xenografts, treatment with the polymer-conjugate strongly inhibited the growth of various human tumors, extended survival, and enhanced tumor regression significantly without effects on body weight compared to control animals. The results clearly demonstrate that our new polyHPMA-docetaxel-folate conjugate compound is a promising candidate for anti-tumor chemotherapeutics with reduced toxicity and prolonged survival.

Citation Format: Young Bok Lee, Deog Joong Kim, Miyoung Yang, Chang-Ho Ahn, Anjan Nan. Synthesis of targeted docetaxel-polymer conjugate and its anti-tumor efficacy. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 4485. doi:10.1158/1538-7445.AM2014-4485
Abstract 4493: Medi-573 alone or in combination with mammalian target of rapamycin inhibitors, targets the insulin-like growth factor pathway in sarcomas
Authors: Zhong Haihong , Fazenbaker Christine , Breen Shannon , Chen Cui , Huang Jiaqi , Chris Morehouse , Yao Yihong , Hollingsworth Robert
Published: May 2015

MEDI-573 is a human antibody that neutralizes the insulin-like growth factors, IGF-1 and IGF-2. IGFs are over-expressed in multiple types of cancer; their over-expression is a potential mechanism for resistance to IGF-1 receptor (IGF-1R)-targeting therapy. Effects of IGFs on cell proliferation, differentiation, and survival are mediated through their binding to and activation of IGF-1R or insulin receptor A (IR-A). In this study, we studied the anti-tumor activity and mechanism of action of MEDI-573 in models of sarcoma. MEDI-573 potently inhibited in vitro proliferation of several sarcoma cell lines, with Ewing's sarcoma cell lines being the most sensitive. This inhibition also occurred after growth stimulation with added IGF-1- and IGF-2. The effect of MEDI-573 on IGF signaling was also examined. Treatment with MEDI-573 markedly reduced levels of pIGF-1R, pIR, and pAKT, and significantly blocked IGF-1- and IGF-2-induced activation of the IGF-1R and AKT pathways. MEDI-573 inhibited the growth of sarcoma xenografts in vivo, and inhibition correlated with neutralization of IGF-1 and IGF-2. Combination of MEDI-573 with either rapamycin or another mTOR inhibitor, AZD2014, significantly enhanced the anti-tumor activity of MEDI-573, and this response correlated with modulation of AKT and mTOR signaling. In summary, sarcoma cells respond to autocrine or paracrine growth stimulation by secreted IGF-1 and IGF-2, and inhibition of IGF-1 and IGF-2 by MEDI-573 results in potent anti-tumor activity in several sarcoma models. Our data provide evidence for evaluation of MEDI-573 and mTORi combinations in clinical studies of sarcoma patients.

Note: This abstract was not presented at the meeting.

Citation Format: Haihong Zhong, Christine Fazenbaker, Shannon Breen, Cui Chen, Jiaqi Huang, Morehouse Chris, Yihong Yao, Robert Hollingsworth. Medi-573 alone or in combination with mammalian target of rapamycin inhibitors, targets the insulin-like growth factor pathway in sarcomas. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 4493. doi:10.1158/1538-7445.AM2014-4493
Abstract 4495: Sym013, novel pan-HER monoclonal antibody mixture, augments radiation response in human lung and head and neck tumors
Authors: Francis David , Huang Shyhmin , Werner Lauryn , Lantto Johan , Horak Ivan D. , Kragh Michael , Harari Paul M.
Published: May 2015

Sym013 represents a novel HER family targeting approach consisting of a mixture of six monoclonal antibodies against EGFR, HER2, and HER3. Sym013 effectively induces rapid, simultaneous down-regulation of all HER members. Compared with single receptor or dual receptor targeting of the HER family, Sym013 provides broader inhibition and greater down-regulation efficacy. In this current study, we examined the capacity of Sym013 in combination with ionizing radiation to augment tumor response in lung (NSCLC) and head and neck (HNSCC) cancer model systems.

The anti-proliferative effects of Sym013 were confirmed showing 30-60% growth inhibition across a variety of NSCLC and H&N cancer cell lines that express EGFR, HER2, and/or HER3. Furthermore, Sym013 inhibited the growth of tumor cells that exhibit resistance to cetuximab. Western blot analysis showed a dose-dependent down-regulation of HER family members and downstream MAPK and AKT signaling pathways following 24-hour treatment with Sym013. Clonogenic survival analysis revealed that Sym013 enhanced radiosensitivity in tumor cells following radiation exposure. We characterized the DNA damage profile following radiation via flow cytometric analysis of phosphorylated histone 2AX (γH2AX) foci. This revealed a significant increase of γH2AX following treatment with Sym013 and 2 Gy radiation when compared with Sym013 or radiation alone. Furthermore, combined treatment of Sym013 and radiation induced a significant G1 and G2 arrest. Using Annexin V/PI staining, we also found a significant increase in the apoptotic cell population following treatment of Sym013 and 6 Gy radiation. These results suggest that Sym013 augments radiation response via the induction of cell cycle arrest followed by the induction of apoptosis and cell death, likely reflecting inhibitory effects on DNA damage repair. Additional studies to examine the impact of Sym013 to augment radiation response as well as overcome acquired resistance to cetuximab in xenograft models are in progress. Overall, these data suggest the significant capacity of Sym013 to augment radiation response in lung and head and neck cancers. This unique antibody mixture warrants additional investigation as a promising novel HER family targeting strategy in cancer therapy.

Citation Format: David Francis, Shyhmin Huang, Lauryn Werner, Johan Lantto, Ivan D. Horak, Michael Kragh, Paul M. Harari. Sym013, novel pan-HER monoclonal antibody mixture, augments radiation response in human lung and head and neck tumors. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 4495. doi:10.1158/1538-7445.AM2014-4495
Abstract 4497: Therapeutic benefit of targeting ERK in mutant KRAS pancreatic ductal adenocarcinoma
Authors: Hayes Tikvah K.
Published: May 2015

The limited success of anti-MEK and anti-RAF therapies with KRAS mutant cancers has been attributed, in part, to de novo or acquired resistance mechanisms that arise after prolonged treatment. A majority of resistance mechanisms involve kinome reprograming to reactivate ERK downstream of inhibitor activity. Therefore, we hypothesized that directly targeting ERK would be an effective method for overcoming these resistance mechanisms. To address this possibility, we evaluated the sensitivity of a panel of mutant KRAS pancreatic ductal adenocarcinoma (PDAC) cell lines to the highly selective SCH772984 ERK1/2 protein kinase inhibitor. First, we found that the anchorage-dependent growth of 6 of 12 cell lines were sensitive to SCH772984 treatment (IC50<2 uM). Some sensitive PDAC lines were resistant to the AZD6244 MEK1/2-selective inhibitor, indicating different mechanisms of action of these two inhibitors. Second, we identified phospho-AKT as a potential biomarker for PDAC SCH772984 sensitivity. Third, we assessed the molecular basis for anchorage-dependent growth inhibition. After 72 h treatment, flow cytometry analyses indicated impaired cell cycle progression through G1. Additionally, we found increased apoptosis as measured by formation of cleaved caspase-3. Together these observations suggest that sensitivity is a result of both impaired cell cycle progression and induction of apoptosis. Surprisingly prolonged SCH772984 treatment (2 weeks) resulted in senescence, as sensitive lines stained positive for senescence-associated beta-galactocidase. Furthermore, we found that senescent cells induced p16 and lost phospho-RB two common markers associated with induction of senescence. Onset of senescence was dependent on the loss of total MYC protein expression. Finally, we found that treatment with SCH772984 reduced tumorigenic growth in an in vivo xenograft mouse model. Our ongoing studies include determining the mechanisms of inhibitor resistance. In summary, our findings suggest that targeting ERK will be therapeutically beneficial for the treatment of KRAS mutant PDAC.

Citation Format: Tikvah K. Hayes. Therapeutic benefit of targeting ERK in mutant KRAS pancreatic ductal adenocarcinoma. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 4497. doi:10.1158/1538-7445.AM2014-4497
Abstract 4543: Identification of downstream targets of polo-like kinase 1 in melanoma by large-scale label-free comparative proteomics profiling
Authors: Cholewa Brian D. , Hahn Molly P. , Scarlett Cameron O. , Ahmad Nihal
Published: May 2015

Polo-like kinase 1 (Plk1) is a serine-threonine kinase that plays a critical role in the cell cycle by regulating mitotic entry, progression and exit. In recent years, Plk1 has gained considerable attention as a promising mechanism-based therapeutic target in a variety of cancers. Pre-clinical studies have suggested that Plk1 inhibition is capable of reducing cancer cell growth both in vitro and in vivo with tremendous efficacy and with minimal detriment to normal cells. This evidence has spurred the development of many Plk1-specific inhibitors that has culminated in over 30 clinical trials, either completed or ongoing. However, the remarkable pre-clinical success of Plk1 inhibitors has yet to be validated at the clinical level, thus stressing on an urgent need for a better understanding of Plk1 signaling. In an effort to further elucidate the molecular function of Plk1 inhibitors, we performed a comparative quantitative proteomics strategy to compare the proteome of human melanoma cells (A375) following treatment with the Plk1-specific ATP competitive inhibitor BI 6727 (25 nM, 24 h). We employed label-free nano-LC/MS/MS technology on a Q-exactive followed by SIEVE processing to identify more than 1800 proteins, ≤ 1% false discover rate, 343 of which had significantly altered expression. Using ingenuity pathway analysis (IPA) to select for proteins with more than two unique peptides and greater than a two-fold change, our proteomic analysis resulted in the identification of more than twenty proteins that were significantly dysregulated following Plk1-inhibition, many of which have not been previously shown to be associated with Plk1 signaling. Interestingly, the metabolic and proteasome pathways harbor the greatest portion of differentially expressed proteins, including five subunits of the 20S proteasome core particle and five members of the glycolysis pathway. Given the importance of these two pathways in cancer cell survival, this study provides two potential mechanisms of Plk1 inhibition that have yet to be explored. Further studies are currently ongoing in our laboratory to validate our findings in additional experiments.

Citation Format: Brian D. Cholewa, Molly P. Hahn, Cameron O. Scarlett, Nihal Ahmad. Identification of downstream targets of polo-like kinase 1 in melanoma by large-scale label-free comparative proteomics profiling. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 4543. doi:10.1158/1538-7445.AM2014-4543
Abstract 4548: The anti-proliferative activity of TAS-108 (a steroid anti-estrogen) is augmented by high ERβ levels in breast cancer cells
Authors: Cheong Ye-Hwang , Choi Young J.
Published: May 2015

Background: TAS-108 is an anti-estrogen that binds to both ERα and ERβ and has potent anti-proliferative activity in vitro and in vivo assay. Recently, the phase II clinical study of TAS-108 demonstrated competitive clinical benefits with decreased adverse effects. However, the molecular mechanism TAS-108 remains uncharacterized.

Methods: Plasmids containing ERα or ERβ fused with N- and C-terminal fragments of firefly luciferase were constructed for split luciferase complementation assays. The resulting plasmids were transfected in HEK293 cells for 24 hr and TAS-108 was added to the media for 6 hr. For RT-qPCR of AREG and TFF1 and proliferation assay, ERβ was transfected in MCF7 cells for 24hr. The proliferation level was determined by measuring the fluorescent level induced by Prestoblue® reagent.

Results: The split luciferase complementation assay demonstrates that TAS-108 increases ERα/β heterodimerization by ≈100-fold than ERα/α homodimerization. The potency of TAS-108 in MCF7 cells was increased significantly by the enforced expression of ERβ via transfection (MCF7-ERβ); 30nM TAS-108 was suitable to block the E2 mediated increases of AREG and TFF1 expression and significantly decreased proliferation of MCF7-ERβ.

Conclusions: TAS-108 has a higher affinity for ERα/β heterodimerization than ERα/α homodimerization, and TAS-108 anti-proliferative activity is augmented by blocking the expression of E2-induced proliferative genes when coupled with increased levels of ERβ. Our findings underscore the molecular role of ERβ in the biology and suggest that promoting ERα/β heterodimerization may be an effective strategy to increase anti-tumor activity in breast cancer.

Note: This abstract was not presented at the meeting.

Citation Format: Ye-Hwang Cheong, Young J. Choi. The anti-proliferative activity of TAS-108 (a steroid anti-estrogen) is augmented by high ERβ levels in breast cancer cells. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 4548. doi:10.1158/1538-7445.AM2014-4548
Abstract 456: ERG regulation of intracrine androgen production and castration-resistant prostate cancer progression
Authors: Powell Katelyn A. , Semaan Louie , Maddipati Krishna R. , Conley-Lacomb Katie (Mary) , Li Yanfeng , Cher Michael , Chinni Sreenivasa R.
Published: May 2015

Introduction and Objective: Intratumoral androgen synthesis in prostate cancer (PC) contributes to the development of castration-resistant prostate cancer (CRPC). Several enzymes responsible for androgen biosynthesis have been shown to be overexpressed in CRPC, thus, contributing to CRPC in a castrated environment. However, little is known regarding their mode of regulation and expression in this context. The TMPRSS2-ERG transcription factor has been shown to be present in primary PC tumors as well as CRPC tumors; however, its biological function contributing to the development CRPC in the context of intratumoral androgen synthesis is not known. We hypothesize that TMPRSS2-ERG fusions regulate androgen biosynthetic enzyme (ABE) gene expression and the production of androgens, which contributes to the development of CRPC.

Methods: We used a variety of methods including lentiviral knockdown and overexpression of ERG, qPCR, Western blot, ChIP-PCR, and Mass Spectrometry.

Results: We found that (a) shRNA knockdown of ERG in VCaP PC cells significantly reduces the expression of ABE's AKR1C3, HSD17B6, and HSD17B4; (b) overexpression of truncated ERG or wild-type ERG in LNCaP PC cells significantly increases the expression of AKR1C3 and HSD17B6; (c) ERG binds to the AKR1C3 gene in VCaP PC cells; and (d) DHT synthesis is reduced in VCaP shERG cells treated with 5α-Androstanedione, a DHT precursor metabolite, compared to VCaP shScrambled controls.

Conclusions: These data suggest that AKR1C3 catalyzes the biochemical reduction of 5α-Androstanedione to DHT in PC cells, and that ERG may regulate this step through upregulation of AKR1C3 expression. Elucidation of ERG regulation of ABEs in CRPC may help to stratify TMPRSS2-ERG fusion-positive PC patients in the clinic for anti-hormone driven therapies. Additionally, not all PC tumors express CYP17A1 enzyme, therefore this study emphasizes the importance of investigating ABEs other than CYP17A1, such as AKR1C3 and HSD17B6. In summary, this study addresses alternative ABE drug targets as well as possible drug resistance modes in PC treatment.

Citation Format: Katelyn A. Powell, Louie Semaan, Krishna R. Maddipati, Katie (Mary) Conley-Lacomb, Yanfeng Li, Michael Cher, Sreenivasa R. Chinni. ERG regulation of intracrine androgen production and castration-resistant prostate cancer progression. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 456. doi:10.1158/1538-7445.AM2014-456
Abstract 4566: Fusarochromanone inhibits cell proliferation and induces cell death in COS7 cells
Authors: Gu Ying , Jiang Shanxiang , Mahdavian Elahe , Huang Shile
Published: May 2015

Fusarochromanone (FC101) is a mycotoxin produced by Fusarium equiseti. Contamination of FC101 occurs frequently in grains and feedstuffs, which has been found to cause tibial dyschondroplasia in chickens and Kashin-Beck disease in children. However, little is known about the toxicity of FC101 in the kidney. In this study, we found that FC101 inhibited cell proliferation and induced cell death in monkey kidney fibroblasts (COS7) in a concentration-dependent manner. Furthermore, we found that FC101 inhibited cell proliferation by arresting cells in G1 phase of the cell cycle, which was associated with reduced expression of CDK4, cyclin D1, and Cdc25A, as well as decreased phosphorylation of Rb. FC101 induced cell death by downregulating expression of Bcl-2, Bcl-xl and survivin, and upregulating expression of Bad, leading to increased cleavages of caspase 3 and PARP. Also, FC101 induced translocation of apoptosis inducing factor (AIF) from the cytosol to the nucleus. Furthermore, FC101 activation of JNK cascade also contributed to cell death, as inhibition of JNK with SP600125 or expression of dominant negative c-Jun attenuated FC101-induced cell death. In addition, we observed that FC101-induced cell death linked to induction of reactive oxygen species (ROS) in the cells, since addition of antioxidants (co-enzyme Q10 and N-acetyl-cysteine) attenuated the toxic effect of FC101 on the cells. Our findings suggest that FC101 may exert its toxicity in COS7 cells through oxidative stress.

Citation Format: Ying Gu, Shanxiang Jiang, Elahe Mahdavian, Shile Huang. Fusarochromanone inhibits cell proliferation and induces cell death in COS7 cells. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 4566. doi:10.1158/1538-7445.AM2014-4566
Abstract 4569: Digitizing single-cell expression patterns in urine for prostate cancer detection
Authors: Lin Chun-Lin , Chen Chun-Liang , Wang Chiou-Miin , Liu Joseph , Huang Susan , Huang Tim
Published: May 2015

Gene expression profiles of microdissected tissue are frequently used to stratify cancer subtypes and to predict patients' clinical outcome. This type of analysis usually operates under the assumption that uniform gene expression is present in a cancer cell population. However, high fluctuation of gene expression in single cells stemming from intra-clonal heterogeneity within a cancer cell population can lead to misinterpretation of the data. An important challenge then lies in developing approaches to decipher this expression chaos for cancer diagnostic and/or prognostic strategies.

We have developed a novel digitizing approach to analyze single-cell gene expression in exfoliated prostate cells isolated from urine sediment. In analyzing 1220 single cells, we observed a panel of genes with dichotomous expression patterns (DEP) and further digitized their expression levels to either 1 for high or 0 for low expression. Of 35 androgen-responsive genes examined, 6 genes exhibited DEP characteristics. When aligning the dichotomous numbers of these genes in the order of CXCL6-TGFBR2-GSK3B-CDKN1C-GATA3-EIF4EBP1, we identified 64 (26) types of code; e.g., 1-0-1-0-1-0. With the conversion from original expression levels to a series of dichotomous numbers, these new codes not only reduced the complexity associated with single cell expression, they also enabled us to decipher expression heterogeneity in the samples by considering multiple gene expression codes simultaneously.

Importantly, these digitized codes identified distinct clonal prostate cell populations in the urine samples. At the single-cell level, high volumes of cells with the same code (common codes) in a collection of cells suggested the presence of a large clonal cell population. By examining the types of code and the size of clonal populations, we found that increasing clonal size of cells carrying common codes was associated with high-grade cancers. Certain common codes were also found in cells from patients diagnosed with benign prostate hyperplasia and prostatic intraepithelial neoplasia. On the other hand, digitized code types appeared to be highly diverse in normal cell populations.

Our data suggest that digitizing single-cell expression can stratify benign, low grade and high grade prostate cancer. This analysis pipeline has a wider application for disease diagnosis/prognosis using cancer cells isolated by non-invasive means from biological fluids or washes from biopsy needles and surgical blades.

Citation Format: Chun-Lin Lin, Chun-Liang Chen, Chiou-Miin Wang, Joseph Liu, Susan Huang, Tim Huang. Digitizing single-cell expression patterns in urine for prostate cancer detection. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 4569. doi:10.1158/1538-7445.AM2014-4569
Abstract 4570: Role of ERRalpha in ovarian cancer
Authors: Stevens Ellen V. , Whitaker Regina , Guinet Audrey , Chang Ching-Yi , Grenier Carole , Marks Jeffrey , McDonnell Donald P. , Murphy Susan K. , Berchuck Andrew , Gaillard Stephanie
Published: May 2015

Objective. The estrogen receptor-related receptor-alpha (ERRα, encoded by ESRRA) has emerged as a key transcriptional regulator of mitochondrial function and has been implicated as a prognostic marker in many cancers, including epithelial ovarian cancer. ERRα has shown promise as a therapeutic target in breast cancer where ERRα transcriptional activity regulates growth of breast cancer cells. This study aims to evaluate whether ERRα expression in ovarian cancer correlates with patient outcomes, determine the effect of inhibiting ERRα transcriptional activity on ovarian cancer cell growth, and identify molecular signaling pathways regulated by ERRα in ovarian cancer cells.

Methods. Affymetrix U133A gene expression microarray analysis from frozen tumor specimens of serous epithelial ovarian cancer was used to compare ESRRA transcripts levels in patients with short-term (<3 years) verses long-term survival (>7 years). ESRRA transcript levels were also measured in normal ovarian surface and fallopian tube epithelial tissues. Protein expression levels were assessed in ovarian cancer cell lines and patient tumor samples. Overall survival was estimated using the Kaplan-Meier Method. The effect of ERRα inhibition on proliferation and anchorage-independent growth was determined after treatment with the ERRα antagonist, XCT790. The activity of 49 different phospho-receptor tyrosine kinases (RTKs) was analyzed after down-regulation of ERRα with shRNA.

Results. ESRRA expression was significantly higher in cancer tissues verses normal ovarian surface and fallopian tube epithelial tissues. Expression was also higher in patients with short-term versus long-term survival. Inhibition of ERRα with XCT790 led to a >50% reduction in proliferation and anchorage-independent growth. The induced knockdown of ERRα decreased the phosphorylation of RTKs, including HER2, suggesting that important growth promoting pathways are downstream of ERRα.

Conclusions. In this study we determined that higher expression of ESRRA correlates with worse survival in ovarian cancer patients. However, inhibition of ERRα results in decreased growth potential. This effect may be manifest through the regulation of receptor tyrosine kinase activity. Thus, ERRα may be an important therapeutic target for the treatment of ovarian cancer and warrants future studies to determine the role of ERRα regulated signaling networks in ovarian oncogenesis.

Citation Format: Ellen V. Stevens, Regina Whitaker, Audrey Guinet, Ching-Yi Chang, Carole Grenier, Jeffrey Marks, Donald P. McDonnell, Susan K. Murphy, Andrew Berchuck, Stephanie Gaillard. Role of ERRalpha in ovarian cancer. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 4570. doi:10.1158/1538-7445.AM2014-4570
Abstract 4573: A novel bispecific Fap-Dr5 antibody inducing potent and tumor-specific death receptor 5 (Dr5) activation by fibroblast activation protein (Fap)-dependent crosslinking
Authors: Wartha Katharina , Weiser Barbara , Friess Thomas , Majety Meher , Runza Valeria , Herting Frank , Weber Thomas , Scheuer Werner , Harring Suzana Vega , Sade Hadassah , Niu Huifeng , Bruenker Peter
Published: May 2015

Background:

Activation of the extrinsic apoptosis pathway in tumor cells through agonistic death receptor 5 (DR5) antibodies has been evaluated in the clinic with limited success so far. In this context, several reports show that DR5 activation is strongly dependent on receptor hyperclustering on the cell surface. Therefore a therapeutic principle that induces DR5 hyperclustering specifically at the tumor site may provide superior efficacy, potency and safety compared to conventional DR5 agonistic antibodies. Fibroblast activation protein (FAP) is a marker for activated fibroblasts and abundantly expressed in cancer associated fibroblasts of various epithelial tumor indications and as a tumor antigen on tumors of mesenchymal origin. Due to its relative absence from normal tissues, FAP can be used as a tumor targeting antigen. Here, we are using the broad expression of FAP in tumor stroma for crosslinking of DR5 by a bispecific antibody.

Aim:

In order to achieve superior tumor targeting and tumor located DR5 hyperclustering we have generated a bispecific antibody, RG7386, comprised of an agonistic DR5 binder and a FAP targeting moiety.

Results:

RG7386 shows potent and selective binding to FAP and DR5 and can simultaneously bind to both targets. In in vitro co-culture assays, using human DLD-1 colon cancer cells and FAP expressing fibroblasts, RG7386 induces potent, FAP dependent DR5 hyperclustering and apoptosis induction in DR5 positive tumor cells (IC50: 0.05 nM). In preclinical in vivo models with co-injection of DLD-1 tumor cells and fibroblasts as well as patient-derived colorectal cancer models, RG7386 shows FAP dependent efficacy and apoptosis induction superior to conventional DR5 antibodies. Furthermore the superior induction of apoptosis could be confirmed by in vivo and ex vivo analysis of cleaved Caspase-3 with imaging, Luminex and histopathology.

Conclusion:

RG7386 is a promising novel therapeutic entity for the treatment of solid tumors with FAP positive tumor stroma inducing DR5 activation by FAP dependent DR5 hypercrosslinking which results in potent anti-tumor activity.

Citation Format: Katharina Wartha, Barbara Weiser, Thomas Friess, Meher Majety, Valeria Runza, Frank Herting, Thomas Weber, Werner Scheuer, Suzana Vega Harring, Hadassah Sade, Huifeng Niu, Peter Bruenker. A novel bispecific Fap-Dr5 antibody inducing potent and tumor-specific death receptor 5 (Dr5) activation by fibroblast activation protein (Fap)-dependent crosslinking. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 4573. doi:10.1158/1538-7445.AM2014-4573
Abstract 4581: A novel modular polymer platform for the treatment of oral and head and neck carcinoma
Authors: Lin Yuan , Sharma Sherven , Dubinett Steven M. , Wu Ben , John Maie St.
Published: May 2015

Objective: Head and neck Squamous cell carcinoma (HNSCC) is difficult to resect completely by surgery due to complicated context and therefore exhibits high recurrence rate in the patient. To reduce tumor recurrence after surgery, we developed a biodegradable polymer to deliver CCL21 cytokine and cisplatin, and evaluate its efficacy in a partially resected xenograft animal model of HNSCC.

Study Design: in vivo study.

Setting: Academic research laboratory.

Subjects and Methods: After xenograft HNSCC tumors were established by subcutaneous injection, C3H/HeJ mice underwent surgery to debulk their tumors to a size of 0.5x0.5x0.5 cm3 and received different polymer implantation. For cytokine studies, mice were grouped into: (1) no polymer; (2) plain polymer; (3) plain polymer with intratumoral injection of recombinant CCL21 twice a week; (4) polymer containing parental dendritic cells; (5) polymer containing dendritic cells secreting CCL21 (DC-CCL21). For cisplatin studies, mice were randomized to receive implantation of (1) no polymer; (2) plain polymer; (3) plain polymer with local cisplatin injection; (4) cisplatin polymer. The two groups of mice implanted with cisplatin polymer or no polymer were further randomized to receive (1) 4 Grays external beam radiation for 4 days; (2) no radiation. Tumor size was measured until the mice were euthanized. At necropsy, the tumors were excised and weighed.

Results: Our results using this novel polymer platform demonstrate a significant reduction in tumor growth. The DC-CCL21 polymer reduced SCCVII/SF tumors in the C3H/HeJ mice by over 41% compared to the control groups (p<0.01). The cisplatin secreting polymer effectively reduced SCCVII/SF tumors in the C3H/HeJ mice by over 16-fold (P < 0.01) as compared to control, plain polymer group. We also observed a statistically significant lower tumor weight among mice treated with cisplatin polymer and concomitant CCL21 compared to the control group.

Conclusion: Herein we demonstrate the efficacy of a novel polymer platform in delivering cisplatin and cytokines to a partially resected SCC in a murine model. We also demonstrate that we can effectively grow dendritic cells in the polymer that can actively secrete CCL21. Our results indicate that this polymer may represent a new therapeutic modality for patients with HNSCC. Once this polymer platform is optimized we will plan for validation in the context of a prospective trial in patients with unresectable advanced or recurrent HNSCC.

Citation Format: Yuan Lin, Sherven Sharma, Steven M. Dubinett, Ben Wu, Maie St. John. A novel modular polymer platform for the treatment of oral and head and neck carcinoma. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 4581. doi:10.1158/1538-7445.AM2014-4581
Abstract 4591: The folate receptor-targeted agents, vintafolide and etarfolatide, are not substrates for the reduced folate carrier or the proton-coupled folate transporter
Authors: Parker Nikki , Cherian Christina , Matherly Larry , Leamon Christopher P.
Published: May 2015

Vintafolide (EC145), a folate-targeted small molecule drug conjugate (SMDC) of desacetylvinblastine hydrazide (DAVLBH), and etarfolatide (EC20), a folate-targeted companion imaging agent, are currently being evaluated in global phase 2 and 3 studies for the selection and treatment of patients with folate receptor (FR)-positive tumors. Both of these compounds have been shown to bind with high affinity and specificity to the FR, which is known to be internalized via endocytosis. Though the FR is the major route for delivery of folic acid (an oxidized folate congener) into a cell at physiological pH, two other transporters, the reduced folate carrier (RFC) and the proton-coupled folate transporter (PCFT), play key roles in transporting folates and antifolates across the cell membrane at neutral and acidic pH, respectively. It is not known, however, if these two transporters play any role in the uptake of folic acid-based SMDCs. Therefore, this study was designed to determine whether vintafolide and/or etarfolatide are substrates for active transport by the RFC or PCFT, or whether their cellular uptake is specific only to the FR. To achieve this, we utilized a set of isogenic Chinese hamster ovary cell lines which have been engineered from transporter-null R2 cells to express the RFC, PCFT, or FR-α, to evaluate cellular uptake of 3H-vintafolide and 3H-etarfolatide:Re. The results of the uptake studies showed that no RFC or PCFT-specific transport of either SMDC occurred under the optimal conditions for transporter activity (pH 7.2 for RFC, and pH 5.5 for PCFT). Conversely, 3H-MTX, a known RFC and PCFT substrate, showed significant uptake by both the RFC and PCFT-expressing cells, and uptake was blocked by PT523 (specific inhibitor for RFC) and AG94 (specific inhibitor for PCFT). Though levels of 3H-vintafolide uptake by the RFC-expressing PC43 cells were similar to that of 3H-MTX, no inhibition of 3H-vintafolide uptake was seen with PT523 suggesting that this uptake is not mediated by the RFC. As expected, very high 3H-vintafolide and 3H-etarfolatide:Re binding and uptake was seen in the FRα-expressing RT-16 cells, and this uptake was competable with excess FA indicating that the interaction was specific to the FR. Collectively, these results show that vintafolide and etarfolatide are specifically transported via the FR and are not substrates for the RFC or PCFT.

Citation Format: Nikki Parker, Christina Cherian, Larry Matherly, Christopher P. Leamon. The folate receptor-targeted agents, vintafolide and etarfolatide, are not substrates for the reduced folate carrier or the proton-coupled folate transporter. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 4591. doi:10.1158/1538-7445.AM2014-4591
Abstract 4602: Blocking of p14/ARF and DX2 binding by novel small chemical can improve the chemo-sensitivity of small cell lung cancer and non-small cell lung carcinoma
Authors: Oh Ah Young , Jung Youn-Sang , Lee Su-Jin , Jo Jung Hyun , Chun Ho Young , Park Bum-Joon
Published: May 2015

In recent, it has been reported that alternative splicing variant of AIMP2, AIMP2/DX2 (DX2), is frequently expressed in human lung cancer and is related with p53-mediated tumor suppression pathway. Here we show that DX2 can promote tumor progression and increase incidence, cooperatively with oncogenic K-Ras in transgenic mouse model. Moreover, it can induce small cell lung carcinoma (SCLC) as well as contribute to progression of non-small cell lung carcinoma (NSCLC). In fact, DX2 expression is elevated in human small cell lung cancer cell lines. Based on the cellular localization and responsibility of si-DX2, we revealed that DX2 is an inhibitor of p14/ARF. Elimination of DX2 can induce p14/ARF expression and DX2 Transgenic (Tg) mouse cells show the low expression of p19/ARF. Since DX2, but not its original product AIMP2, is selectively interacted with p14/ARF, we screened the specific binding inhibitor and obtained the single compound (SLC36) from about 9000 chemicals. This chemical can block the interaction of p14/ARF-DX2 and also AIMP2-DX2, but not the binding of p14/ARF-p53 or AIMP2-p53. In addition, it can induce p14/ARF expression and cell death in human lung cancer cell lines including SCLC cell lines. In our in vivo study, treatment of SLC36, combined with low dosage of adriamycin (1 mg/kg) can suppress the cancer incidence as well as progression in K-RasLA2 and K-Ras/DX2 double Tg model. These results indicate that DX2 can contribute to lung cancer progression and development, including SCLC, through inhibition of p14/ARF, and blocking of DX2-p14/ARF binding would be useful therapeutic strategy of human lung cancers including SCLC as well as NSCLC.

Citation Format: Ah Young Oh, Youn-Sang Jung, Su-Jin Lee, Jung Hyun Jo, Ho Young Chun, Bum-Joon Park. Blocking of p14/ARF and DX2 binding by novel small chemical can improve the chemo-sensitivity of small cell lung cancer and non-small cell lung carcinoma. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 4602. doi:10.1158/1538-7445.AM2014-4602