bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 5262: The role of Bcl-2 in colon cancer metastatic progression
Authors: Wang Wang , Are Chandrakanth , Brattain Michael G.
Published: June 2012

Colon cancer is the second most lethal malignancy in the United States. The majority of deaths are related to metastasis, for which no effective therapies are currently available. While much is known about oncogenic initiation of colon cancer, the mechanisms by which cancer cells survive and metastasize in spite of the cellular stresses associated with this process require further investigation. Recent studies indicated that deregulation of apoptosis contributes to metastasis, and deregulation of Bcl-2 family members, which control the mitochondrial-dependent apoptosis pathway, is correlated with cancer progression.

We have initiated these studies using a unique othotopic implant model based on closely related colon cancer cell lines. HCT116 and HCT116b cells were isolated from the same primary colon tumor and both bear the H1047R PIK3CA mutation and form progressively growing invasive orthotopic primary tumors. However, only HCT116 cells exhibit metastatic potential. Interestingly, we found that Bcl-2 is highly expressed in HCT116 cells, but not in HCT116b cells. Overexpression of exogenous Bcl-2 in HCT116b cells, and complementary studies by shRNA mediated stable knockdown and overexpression of dominant negative forms of Bcl-2 molecule in metastatic HCT116 cells indicated that Bcl-2 is essential for their ability to withstand external stress. Current studies in our orthotopic model demonstrated that HCT116b tumors bearing exogenous Bcl-2 associated with increased cell survival and reduced center necrosis. Future study will determine whether this significant alteration in the expression of Bcl-2 associates with the distal metastatic capability of colon cancer.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 5262. doi:10.1158/1538-7445.AM2011-5262
Abstract 5268: CXCR4 and P-glycoprotein N-glycan patterning in developing cancer cells
Authors: Ivey Stan , Shan Yufei , Owens Tori A.
Published: June 2012

Human CXCR4 and P-glycoprotein (Pgp) are N-glycosylated membrane proteins overexpressed in cancer cells. CXCR4 is thought to be an initiator of neuroblastoma metastasis to bone tissue and is overexpressed in neuroblastomas. Pgp is a heavily glycosylated, ATP-dependent drug efflux pump overexpressed in uterine sarcomas. Pgp is responsible for multidrug resistance in some tumors. Our lab is studying the N-glycan profiles of CXCR4 and Pgp in neuroblastoma (IMR-32) and uterine sarcoma cells (MES-SA and MES-SA/Dx5). We believe CXCR4 and Pgp are closely involved in metastasis. To better understand the molecular mechanisms of the metastatic roles of CXCR4 and Pgp, we are characterizing their respective complements of N-glycans. CXCR4 contains a N-linked oligosaccharide on two separate extracellular loops; moreover, Pgp has 3 N-glycans present on a 9 amino acid stretch near its N-terminus. We believe both protein N-glycan moieties are differentially glycosylated enough to act as markers for identifying and staging cancer cells. Our previous studies have shown that Pgp overexpressed in MES-SA/Dx5 cells contains at least two different complex N-glycans; a high mannose tree and a hybrid, sialic acid-capped oligosaccharide. To build on our previous studies, we have developed a novel approach to further characterize CXCR4 and Pgp N-glycans using polyclonal antibodies (pAbs) raised to the specific amino acid sequences containing the sugars in both cancer proteins. We are using these inimitable pAbs to aid in the isolation, sequencing, and profiling of specific N-glycan changes that are associated with the stage of each of the cancer cell lines mentioned above. Our pAb panels bind the native, expressed glycoforms of CXCR4 and Pgp in cells and on Western blots. We are using pAb affinity columns to capture protease-released glycopeptides. Once captured, we intend to sequence each of the released glycopeptides’ oligosaccharides to verify their saccharide sequences in the different cancer cell stages. Furthermore, we predict that the resultant glycopeptide gel and blot patterns will ‘bar code’ the specific cancer stage of the cells lines studied. We envision that these glycopeptide patterns can be used to identify and catalog the various stages of developing cancer cells.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 5268. doi:10.1158/1538-7445.AM2011-5268
Abstract 5269: Mechanistic insights into regulation of metastasis by Sipa1
Authors: Geiger Thomas R. , Mattaini Katie , Williams Mia , Walker Renard , Alsarraj Jude , Borges Rosan Nieves , Hunter Kent
Published: June 2012

Metastasis is the major cause for morbidity and mortality of cancer patients. Still, the molecular mechanisms underlying metastasis are incompletely understood. Conceivably, a better knowledge of the metastatic process will enable the design of better treatments for cancer patients in the future.

Previously, we have shown that genetic background has a strong influence on metastasis susceptibility in breast cancer mouse models. Single nucleotide polymorphisms (SNP) in the Sipa1 gene are associated with metastasis in mice and human breast cancer patients. Consistent with this observation, we have shown that Sipa1 regulates metastasis in breast cancer cells; however, the molecular mechanisms remain largely unknown.

We identified several potential binding partners of SIPA1 in a yeast-two-hybrid screen, and confirmed interactions of SIPA1 with BRD4, RRP1B and SUN2 in subsequent experiments. Our analysis suggests that several complexes of SIPA1 exist in different compartments of the cell. A structure-function analysis that we have begun to carry out indicates that Sipa1 regulates metastasis in several ways, depending on its interaction partners and subcellular localization. Furthermore, the enzymatic GTPase-activating function of Sipa1 appears to play a critical role in regulating metastasis. These results could facilitate developing novel therapeutic approaches for the treatment of metastatic breast cancer.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 5269. doi:10.1158/1538-7445.AM2011-5269
Abstract 5278: Intravital dual-colored visualization of colorectal liver metastasis formation in the living mice using two photon laser scanning microscopy
Authors: Tanaka Koji , Morimoto Yuhki , Matsushita Kohei , Kawamura Mikio , Toiyama Yuji , Saigusa Susumu , Okugawa Yoshinaga , Koike Yuhki , Inoue Yasuhiro , Miki Chikao , Mizoguchi Akira , Kusunoki Masato
Published: June 2012

Background and aim: Metastatic spread is the major cause of mortality in patients with colorectal cancer (CRC). Liver metastasis is the most frequent and thus the main therapeutic target in metastatic CRC patients. The mechanism of colorectal liver metastasis (CLM) is still poorly understood. In this study, we presented high-resolution images of dynamics of tumor-host interaction in liver metastasis model of the living mice using two photon laser scanning microscopy (TPLSM). We also visualized the formation of CLM from single cancer cells to micrometastatic colonies in the same mice by intravital TPLSM at several times.

Materials and methods: Red fluorescent protein (RFP) expressing SL4 murine colorectal cancer cell line (RFP-SL4) and green fluorescent protein (GFP)-transgenic mice (GFP mouse) were used as syngeneic tumor model. RFP expressing HT29 human colorectal cancer cell line (RFP-HT29) and GFP nude mice (C57BL/6-BALB/c-nu/nu-EGFP) were used as xenogeneic tumor model. 1×106 RFP expressing cancer cells were injected into the spleen of GFP (nude) mice for an experimental liver metastasis model. Intravital TPLSM was performed by exteriorizing the liver lobe of living mice and fixing it with a solder lug terminal and an adhesive agent (organ stabilizing system, patent number: 2007-129723). Interval TPLSM was also performed for intravital TPLSM at several times in the same mice.

Results: Viable cancer cells in the vital liver of living mice were successfully imaged at high-resolution in vivo real-time by TPLSM. Single cancer cells were arrested within hepatic sinusoids 2 hours after injection at a magnification of X600∼X1200. Platelet aggregation surrounding a single cancer cell was observed 2h after injection, indicating a phenomenon of tumor-cell induced platelet aggregation (TCIPA). Cancer cells were extravasated from hepatic sinusoid to Disse space 24h after injection. Protrusions of a Kupffer cell surrounding a cancer cell were observed, indicating that a Kupffer cell appears to be phagocytosing a cancer cell. RFP-SL4 were formed micrometastatic colonies with extensive stromal reaction. Liver metastasis of RFP-HT29 was observed as a clustering of micrometastatic nodules.

Conclusions: Our advanced technique using both intravital and interval TLPSM enables a dynamic high-resolution imaging of not only tumor-host interaction in cancer metastasis, but also metastatic process from a single cancer cell to micrometastasis formation in the living animals.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 5278. doi:10.1158/1538-7445.AM2011-5278
Abstract 5286: Evaluation of 64Cu-labeled peptide for in vivo imaging of Thomsen-Friedenreich antigen expressing breast carcinomas
Authors: Kumar Senthil R. , Quinn Thomas , Deutscher Susan
Published: June 2012

Introduction: Thomsen-Friedenreich carbohydrate antigen (TF; Galβ1-3GalNAc) is displayed on cell surface proteins and lipids in ∼90% of adenocarcinomas including those of the breast. We investigated the ability of a peptide, DRD-IVWHRWYAWSPASRI-DRD-NH2 (TF-pep), selected through bacteriophage display to bind breast and prostate carcinoma cells expressing TF. It is hypothesized that TF-pep once radiolabeled with 64Cu, a positron radiation emitter, has the potential to be used as a positron emission tomography (PET) imaging agent for TF expressing breast tumors for diagnostic purposes.

Experimental Procedure: A bifunctional-chelator (1,4,7-triazacyclononane-1,4,7-triacetic acid (NOTA)) was attached to the TF-pep with Gly-Ser-Gly (GSG) spacer. The peptide was radiolabeled with 64Cu in ammonium acetate buffer pH 6.8 at 80oC for 60 minutes. In vitro cell binding studies with both human breast (MDA-MB-435) carcinoma cells, as well as normal breast epithelial (184A1) cells were performed with the radiolabeled peptide. In vivo biodistribution and PET/CT imaging of the tumor with radiolabeled peptide was analyzed in SCID mice bearing MDA-MB-435 tumor xenografts. Results: 64Cu-NOTA-Gly-Ser-Gly-TF-pep peptide bound to human MDA-MB-435 carcinoma cells in a time dependant manner while insignificant binding was observed with normal breast 184A1 cells. In vivo biodistribution studies demonstrated radiolabeled peptide accumulation in MDA-MB-435 tumor-bearing SCID mice (∼1.09 ± 0.34 %, and 0.85 ± 0.06% ID/g in tumor) at 0.5 h and 2 h, respectively. Little accumulation occurred in other organs with the exception of liver (1.12 ± 0.12% ID/g, and kidneys (15.4 ± 1.2% ID/g) at 1 h. Live imaging studies with 64Cu-NOTA-Gly-Ser-Gly-TF-pep (300 μιχρoCi) demonstrated good tumor uptake after 1 h post-injection.

Conclusions: The TF-specific peptide once radiolabeled with 64Cu, can function as a non-invasive in vivo tumor imaging agent of human breast and other carcinomas expressing this antigen.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 5286. doi:10.1158/1538-7445.AM2011-5286
Abstract 5311: The targeting effect of radio-labeled LXY1 and LXW7 against glioblastoma on xenograft mouse model
Authors: Xiao Wenwu , Liu Ruiwu , Yao Nianhuan , Lam Kit S.
Published: June 2012

Glioblastoma, the most common type of primary brain tumors in adults, remains largely incurable, and surgical resection followed by radiation and chemotherapy has been shown to only slightly increase patient survival. New approaches to treat this deadly cancer are needed. Small peptide-based compounds have attracted wide interest as cancer targeting agents for the delivery of radionuclides or cytotoxic drugs to tumors. Using the “one-bead-one-compound” (OBOC) combinatorial library method, we have identified a specific ligand, LXY1, which targets the α3β1 integrin on glioblastoma cell line U87MG. We have also discovered another novel ligand containing a RGD motif, LXW7, which targets the αvβ3 integrin of U87MG cells and tumor endothelial cells. LXY1 and LXW7 demonstrated excellent tumor targeting efficacy in in vivo optical imaging studies. To further investigate the targeting property of the peptides and precisely clarify their in vivo biodistribution profile, we have conjugated LXY1 and LXW7 to the metal chelator CB-TE2A complexed with 64Cu to perform PET imaging and biodistribution studies. The results indicate 64Cu-labeled LXY1-CB-TE2A conjugate efficiently targets both orthotopic and subcutaneous glioblastoma. The LXW7-CB-TE2A conjugate also demonstrates good tumor accumulation. Both conjugates show relatively rapid clearance from tumor site and high uptake in the liver and kidney beside to tumor. The undesired high uptake of the peptides in the liver and kidney indicate both peptides in current version may be more promising in radioimaging than radiotherapy for glioblastoma.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 5311. doi:10.1158/1538-7445.AM2011-5311
Abstract 5359: Cytotoxicity of MLN8237 and SAHA in pediatric cancer cell lines
Authors: Muscal Jodi , Scorsone Kathy , Ecsedy Jeffrey , Berg Stacey
Published: June 2012

Purpose: Children with recurrent or refractory tumors often have a poor prognosis. Innovative drug combinations are necessary to increase their disease-free and overall survival. Background: MLN8237 is a reversible inhibitor of the Aurora A serine/threonine kinase that showed excellent activity against several tumor types in the Pediatric Preclinical Testing Program. SAHA is a pan-histone deacetylase (HDAC) inhibitor that has shown pre-clinical activity in leukemia, lymphoma, neuroblastoma, and medulloblastoma. HDAC inhibitors decrease Aurora kinases by a variety of mechanisms, and HDAC inhibitors and Aurora kinase inhibitors disrupt the spindle assembly checkpoint at different points, suggesting that the combination could have increased antitumor activity. Methods: We investigated the combination of MLN8237 and SAHA in pediatric cancer cell lines, Molt-4 (pediatric T-cell ALL), IMR32 (neuroblastoma), and Daoy (medulloblastoma). Cell viability was measured using a modified methyl tetrazolium assay after 72 hours of drug treatment, and the cytotoxicity of the drug combination was analyzed using the universal response surface approach as implemented in ADAPT II modeling. For each single drug experiment, two replicate plates were tested, and for the combination drug experiments, four replicate plates were tested. Results: Our results showed that the single agent IC50 for MLN8237 is 0.02 – 0.04 μM in all cell lines. For SAHA, the single agent IC50 ranged from 0.5 μM for Molt-4 to 2.7 μM for Daoy. Both SAHA and MLN8237 inhibited cell survival in a concentration-dependent fashion. Treatment with SAHA resulted in nearly complete inhibition of cell growth in Molt-4 and IMR32 cells at concentrations > 1 μM and 80% inhibition of Daoy cell growth at the maximum concentration of 10 μM, which is well above the clinically relevant concentration range. Maximum inhibition of cell growth in all three lines occurred with 0.05-0.5 μM MLN8237. At concentrations of MLN8237 exceeding approximately 1 μM, there was a paradoxical increase in viability signal in all three lines, most pronounced for Daoy. This may be explained by inhibition of Aurora B kinase which occurs at higher MLN8237 concentrations. Aurora B inhibition leads to metabolically active multinucleated cells. The combination of MLN8237 plus SAHA showed additive cytotoxicity in all three cell lines and nearly abrogated the survival noted at high single-agent MLN8237 concentrations. Conclusion: These results provide important preclinical support for the development of future clinical studies of MLN8237and SAHA.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 5359. doi:10.1158/1538-7445.AM2011-5359
Abstract 5371: A pooled apoptome shRNA screen to identify pathways that enhance NVP-BEZ235-induced apoptosis
Authors: Buchdunger Elisabeth , Theuer Andreas , Kazic Malika , Hattenberger Marc , Guthy Daniel , Roesel Johannes , Brachmann Saskia , Maira Sauveur-Michel
Published: June 2012

NVP-BEZ235 is a dual inhibitor of class 1 phosphatidylinositol 3-kinase (PI3K) and mammalian target of rapamycin (mTOR) catalytic activity. It induces cell death in a subset of breast cancer cell lines characterized by amplification of human epidermal growth factor receptor 2 (HER2/ErbB2) and/or activating PIK3CA mutations, but not in cell lines with loss of function of the PTEN tumor suppressor protein or KRAS mutations. In order to better understand the molecular mechanisms leading to cell death, to identify potential biomarkers of activity and to reveal pathways that may even enhance NVP-BEZ235 induced lethality, a pooled short hairpin RNA (shRNA) screen using a lentiviral-based shRNA library targeting the apoptome was performed in NVP-BEZ235-sensitive MDA-MB453 and HCC1954 breast cancer cells. This strategy led to the identification of shRNAs that sensitized cells to NVP-BEZ235-induced apoptosis. Top sensitizers included pro-survival BCL2 family members, BCL2L1 and MCL1. Validation studies were carried out with single shRNAs targeting BCL2L1 and MCL1 as well as drug combination studies of NVP-BEZ235 with ABT-263 or ABT-747, both BCL2 homology domain 3 (BH3)-only protein mimetics. Cell death assays and biochemical readouts including PARP cleavage support the use of BCL2 family inhibitors to enhance NVP-BEZ235-induced cell death in HER2 amplified/PIK3CA mutated breast cancer cells.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 5371. doi:10.1158/1538-7445.AM2011-5371
Abstract 5382: Anti-tumor efficacy of a recombinant human arginase in combination with chemotherapeutic agents in human hepatocellular carcinoma
Authors: Chow Ariel , Pang Roberta , Chu Andrew , Ng Lui , Poon Ronnie
Published: June 2012

Introduction: Systemic chemotherapy of hepatocellular carcinoma (HCC) relies on few drugs and the response rates are low especially in patients with advanced HCC. HCC is considered as auxotrophic for arginine due to the lack of expression of argininosuccinate synthetase (ASS). Several recombinant human arginases were found to be effective in inhibiting liver, pancreatic and leukemic cell proliferation. Recently, a new recombinant human arginase, BCT-100, has been synthesized to deprive arginine and inhibit arginine-dependent tumor growth. In this study, the efficacy of BCT-100 and the combined used of BCT-100 with cisplatin and fluorouracil on the inhibition of in vitro cell proliferation of HCC cell lines and in vivo tumor growth was studied.

Method: The anti-tumor efficacy of BCT-100, cisplatin and fluorouracil, alone or in combination, on cell proliferation, cell cycle distribution and cellular apoptosis were determined in human hepatoma HepG2, Hep3B and PLC/PRF/5 cells. Protein phosphorylation and expression in the Wnt/β-catenin pathways, and expression of cyclin D1, eIF4E, survivin and XIAP, were also analyzed by western blotting. For in vivo animal studies, subcutaneous tumors were established by subcutaneous injections of 1×106 cells into nude mice. BCT-100 in combination with cisplatin or fluorouracil was administrated. Mice were sacrificed week 16 or when tumor sizes exceeded 30% of their body weight.

Results: Treatment with BCT-100 alone was found to inhibit cell proliferation and enhance cellular apoptosis. Additive effect of BCT-100 with cisplatin or fluorouracil was found in inhibiting cell proliferation and increasing apoptosis of HepG2, Hep3B and PLC/PRF/5 cells. Cell cycle arrest at G1/S phase was also observed with BCT-100 treatment. A significant reduction in β-catenin, cyclin D1, phosphorylated eIF4E, survivin and XIAP expression was observed. Furthermore, repeated sequential use of BCT-100 and chemotherapy with either cisplatin or fluorouracil demonstrated synergistic effect of inhibition of tumor growth compared with BCT-100 or chemotherapy alone.

Conclusion: These pre-clinical data suggested additive/synergistic effect of BCT-100 with other chemotherapeutic agents in HCC, which suggested the rationale of combining BCT-100 and chemotherapy in clinical treatment of HCC.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 5382. doi:10.1158/1538-7445.AM2011-5382
Abstract 5393: Prodrug activator gene therapy mediated by prostate-specific replicating retrovirus vectors
Authors: Kamijima Shuichi , Kimura Takahiro , Haga Kazunori , Hiraoka Kei , Inagaki Akihito , Logg Christopher R. , Kao Chinghai , Bochner Bernard H. , Ichikawa Tomohiko , Kasahara Noriyuki
Published: June 2012

Replicating murine leukemia virus (RMLV)-based vectors can propagate specifically within actively dividing cancer cells and achieve highly efficient and tumor-selective gene delivery. However, in view of the known potential for retroviral integration-mediated genotoxicity, additional mechanisms to target the virus specifically to cancer cells are desirable.

To further improve their tumor specificity and safety profile, we have developed RMLV vectors regulated by two different tissue-specific promoters.

We generated GFP-expressing RMLV vectors ACE-emd (wild type), ACE-At-emd (regulated by ARR2PB promoter, a synthetic variant of the probasin promoter which exhibits high specificity for androgen receptor (AR)-positive prostate cancer cells) and ACE-PSES-emd (regulated by PSES promoter, an androgen-independent promoter which is highly active in PSA/PSMA-positive prostate cancer cells, both in the presence and absence of androgen). We also developed ACE-yCD, ACE-At-yCD and ACE-PSES-yCD carrying the yeast cytosine deaminase (yCD) prodrug-activator gene, which converts the nontoxic prodrug 5-fluorocytosine (5FC) into the chemotoxin 5-fluorouracil.

These vectors were first in vitro tested in both prostate (LNCaP, MDA PCa 2b and PC-3) and non-prostate (293T) cell lines in the presence and absence of androgen. RMLV vector ACE-At-emd exhibited high specificity and robust replication in AR-positive prostate cancer cell lines, but only in the presence of androgen. RMLV vector ACE-PSES-emd exhibited high specificity for PSA/PSMA-positive prostate cancer cell line both in the presence and absence of androgen.

For in vivo studies, male nude mice were inoculated subcutaneously with LNCaP cells (AR-positive, PSA/PSMA-positive) in Matrigel. RMLV vectors expressing GFP were injected intratumorally, and tumors were analyzed by flow cytometry 2 and 4 weeks after vector injection. The results showed that both ARR2PB- and PSES-regulated RMLV vectors achieve efficient gene delivery to LNCaP s.c. tumors in vivo.

Cytotoxic activity of RMLV-yCD vectors was examined in vitro in LNCaP cells in the presence and absence of 5FC after virus infection. Cell-specific expression of each vector correlated with reduced cell viability compared to uninfected cells after 5FC treatment. We are now conducting studies to evaluate therapeutic efficacy vs. potential genotoxicity of these vectors for prodrug-activator gene therapy in vivo.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 5393. doi:10.1158/1538-7445.AM2011-5393
Abstract 5408: PEG surface modification of Poly(D/L Asp-DET)/DNA polyplexes greatly increases tumor accumulation and improves biodistribution
Authors: Mirosevich Janni , Cardoen Gregoire , Burke Brian , Costich Tara , Sill Kevin
Published: June 2012

Cationic polymers have received much attention as promising non-viral vectors for gene transfer. However, generating polymers that are stable and can systemically deliver large amounts of nucleic acids to specific disease sites continues to be a significant hurdle to non-viral vector success. Therefore, the purpose of this study was to develop a new biocompatible and biodegradable cationic polymer system for non-viral gene delivery of plasmid DNA. Poly-d/l Aspartate-Diethylenetriamine [Poly(d/l Asp-DET)] polymers were synthesized and used to complex plasmid DNA. Polymer/DNA polyplexes were further modified by covalent coupling with various polyethylene glycol (PEG) polymers. Polyplexes were characterized and evaluated for particle size, zeta potential and morphology by TEM. Plasmid DNA stability was assessed by challenging polyplexes with serum nucleases, and the efficacy of DNA delivery and gene expression was examined by performing transfection experiments on cancer cells. In vivo DNA tumor delivery and biodistribution was also investigated using tumor bearing nude mice. Poly(d/l Asp-DET) polymers were able to bind to plasmid DNA and formed polyplex particles the were amenable to surface coating with PEG. PEG-Polyplexes were approximately ∼70nm in diameter with neutral zeta potentials. TEM confirmed that the polyplexes were uniform in size. PEG-Polyplexes were also extremely stable and maintained the structural integrity of DNA following incubation in nucleases, and were also capable of transfecting HCT-116 and PC-3 culture cells with no cell toxicity. Tail vein administration of PEG-Polyplexes to tumor bearing nude mice did not reveal any observable toxicities. Moreover, improvements in PEG polymer chemistry increased nucleic acid accumulation in tumors while reducing DNA accumulation in other tissues including liver, kidney and spleen. Poly(d/l Asp-DET) cationic polymers show enormous potential as non-viral drug delivery agents. Further, the surface modification of polyplexes with PEG allowed for sustained systemic delivery of nucleic acids to tumors. The characterization of PEG-Poly(Asp-DET)/DNA polyplexes provides useful insights that can be used to the design vectors for targeted nucleic acid delivery.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 5408. doi:10.1158/1538-7445.AM2011-5408
Abstract 5410: Down regulating heavy chain ferritin gene increasesthe radiation therapeutic efficacy for treatment of glioma
Authors: Liu Xiaoli , Madhankumar A.B. , Sheehan Jonas , Slagle-Webb Becky , Connor James R.
Published: June 2012

Gliomas, especially glioblastoma multiforme are resistant to radiation and chemotherapy. Therefore, efforts to develop new course of therapy to enhance efficacy of treatment strategies are obviously critical. We hypothesized that disrupting iron homeostasis in malignant tumor cells could prove effective at negatively affecting tumor growth because cancer cells have a robust iron requirement consistent with their rapid growth and high metabolic rates. The high levels of iron uptake require a mechanism to limit iron induced oxidative damage. This function is the responsibility of ferritin; an iron storage protein with 24 subunits of two types of chains light and heave chains. Heavy chain ferritin (H-ferritin) has been found in the nucleus of tumor cells where it appears to protect DNA from oxidative damage and promote transcription. We hypothesized that silencing the H-ferritin gene could increase the sensitivity of tumors cells to radiation. On the other hand, our previous data already demonstrated that silencing the H-ferritin gene increased chemotherapeutic efficacy for treating gliomas. To test our present hypothesis, the H-ferritin gene will be silenced via a cationic liposome carrying siRNA. We have chosen glioma cell lines (U251, U87 and MNSF) as our in vitro model. Subcutaneous glioma tumor in mouse model (athymic nude mice) was chosen as our in vivo model. Our preliminary in vitro data demonstrated silencing the H-ferritin gene in U251 cells leads to a 50% increase in cell death at 20 Gy for 24, 48 and 72 hours post radiation. Intratumoral injections of cationic liposomes containing H-ferritin siRNA reduced the effective in vivo dose of radiation for tumor suppression by more than 90% at 7 weeks. A supercoil relaxation assay demonstrated that H-ferritin, but not L-ferritin directly protects DNA from radiation by maintaining DNA in a relaxed from. We will further test our hypothesis in glioma-U87, neurofibroma-sNF96.2 and breast cancer-MCF-7 cells. As of now our data shows that silencing the H-ferritin gene appears to be an effective way to increase the sensitivity of the glioma tumors to radiation, which sheds light on increasing the radiation therapeutic efficacy for treatment of malignant tumors.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 5410. doi:10.1158/1538-7445.AM2011-5410
Abstract 5423: New diazonamide analogs demonstrate remarkably larger therapeutic window compared to taxanes and vinca alkaloids in rodents
Authors: Li Lin , Qi Wei , Zhou Ming , Xu Xiaoming , Harran Patrick G. , Wang Xiaodong , Wang Lai
Published: June 2012

Diazonamide A is a natural product originally isolated from a marine organism. It potently induces G2/M cell cycle arrest. Its molecular mode of action is not fully understood but its potential as a cancer therapeutic is high. To streamline our diazonamide synthesis and to define a minimal pharmacophore, we prepared a series of truncated structures. Its simplified structure and shortened synthesis allowed for extensive structure activity relationship analyses. Several modifications markedly improved activity. Among them, JP2295 and JP2362 were studied in details. Both compounds were highly efficacious in a wide spectrum of tumors from different origin both in vitro and in vivo. They were >20 fold more potent than AB5, a closely related analog of the natural product, in a MIA Paca-2 pancreatic cancer xenograft model. At dose 10-fold higher than the efficacious dose, JP2295 or JP2362 did not cause any apparent toxicity. Three out of 7 mice were completely free of tumor even 6 months after the initial treatment. In the same model, taxanes, i.e. paclitaxel and docetaxel, and vinca alkaloids, i.e. vinorelbine, vinblastine, and vincristine, were either less efficacious or causing severe body-weight loss, even animal death. Similar effects were observed in other xenograft models. Safety studies were further carried out in NIH Swiss mice and Sprague Dawley rats with similar results. In summary, JP2295 and JP2362 demonstrated over 20-fold bigger therapeutic window than taxanes and vinca alkaloids in rodents. Such a large therapeutic window is unique for a small molecule anti-mitotic and suggests further development is warranted.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 5423. doi:10.1158/1538-7445.AM2011-5423
Abstract 5458: Identification of functional downstream gene of ABCB1 2677 G>T/A polymorphism, LAMP-1, as a predictor gene to inhibitors of microtubule dynamics using integrative analysis of pharmacogenetics and genomics
Authors: Kwon Woo Sun
Published: June 2012

Objectives Overexpression of ABCB1 in cancers was first reported to encode a protein responsible for drug resistance. ABCB1 expression has been shown to be associated with single nucleotide polymorphisms.

Methods We studied chemosensitivity-related functionally downstream genes of ABCB1 non-synonymous 2677G>T/A polymorphism using integrative genomics studies of ABCB1 (polymorphisms, expression profiling), chemo-sensitivity to various anticancer drugs, and functional association study of the selected gene.

Results The genes were identified based on the results of the significance analysis of microarray, and prediction accuracy was evaluated using the prediction analysis of microarray. Functional assay of the selected gene (LAMP1) was done by siRNA and drug accumulation. Association studies demonstrated correlations between 2677 genotypes and sensitivity to docetaxel, vincristine, and vinblastine. Based on the pharmacogenetic association of 2677 polymorphism, we performed expression profiling to select seven genes. Chemosensitivity was restored with LAMP1 siRNA by increased intracellular drug accumulation.

Conclusions We identified seven genes that predict the chemosensitivity to microtubule dynamics modulators using integrative pharmacogenetic genotyping and pharmacogenomic profiling assays. Biological validation of the role of the seven downstream genes of non-synonymous ABCB1 2677G>T/A polymorphism, including the membrane function of LAMP1, and clinical validation of their predictivity are required for clinical application of these results.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 5458. doi:10.1158/1538-7445.AM2011-5458
Abstract 5480: Effect of the Smac mimetic TL32711 in combination with TRAIL or TNF alpha on a panel of melanoma cell lines
Authors: Krepler Clemens , Chunduru Srinivas K. , He Xu , Gimotty Phyllis A. , Vultur Adina , Villanueva Jessie , Herlyn Meenhard
Published: June 2012

Although major advances have been made recently in the treatment of malignant melanoma, it remains an incurable disease when not surgically excised at a very early stage. This is due to the development of resistance to targeted compounds currently being tested in patients. In addition, the genetic heterogeneity of melanoma often limits the effects of those therapies to specific subsets of patients. New targets that are exploitable in multiple genetic subgroups of melanoma and that can overcome resistance are therefore still urgently needed. IAP's (inhibitor of apoptosis proteins) have been described as major players in conferring resistance to therapy by blocking the apoptotic cascade in a high percentage of melanomas. Thus, the sensitivity to apoptotic stimuli, such as TNF alpha or TRAIL, could be restored in melanomas using the novel Smac (second mitochondria-derived activator of caspases) mimetic, TL32711. A panel of patient-derived human melanoma cell lines was clustered according to mutational status, representing all major genetic subgroups of cutaneous melanoma. Cells were treated with TL32711 as a single agent or in combination with TRAIL or TNF-α for 72h. Cell viability was assessed by MTS assay. Target inhibition and initiation of apoptosis were also evaluated. To more accurately predict in vivo efficacy, cells were grown as three dimensional spheroids in a collagen matrix and treated as described above. Treatment effects on spheroids were assessed through confocal microscopy using live/ dead immune fluorescent staining, as well as alamar blue cell viability assay for objective quantification.We observed that seventeen out of eighteen cell lines tested were resistant to TL32711 as a single agent in vitro, even at high doses of the compound. Similarly, treatment with TRAIL or TNF-α alone did not show any significant increase in apoptosis. However, when TL32711 was combined with either TRAIL or TNF-α, a strong synergistic activity was observed in twelve out of eighteen cell lines at low doses, resulting in a dramatic increase in cell death of adherent and three dimensional cultures (spheroids). In all treated cell lines, degradation of the target IAP proteins was observed. However, only in sensitive cell lines, activation of apoptotic cascade was observed.In conclusion, the Smac mimetic TL32711 exhibits strong synergistic activity in combination with TRAIL or TNF-α leading to apoptosis in the majority of the genetically diverse cell lines tested. Since TNF-α and TRAIL are present in a high percentage of melanoma tumors in vivo, single agent activity of TL32711 may be observed in vivo.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 5480. doi:10.1158/1538-7445.AM2011-5480
Abstract 5487: Anti-proliferative, apoptotic and immunological effects of ganoderic acid DM on prostate cancer cells
Authors: Haque Azizul , Johnson Benjamin M. , Doonan Bently P. , Hossain Azim , Radwan Faisal F.
Published: June 2012

Natural extracts have been the focus of recent investigation in the design of novel chemotherapeutics. One potential natural therapeutic agent is ganoderic acid DM (GA-DM), an extract from the Ganoderma lucidum mushroom previously shown to inhibit 5-α-reductase activity and prevent binding of dihydrotestosterone to the androgen receptor in prostate cancer cells. Studies in our laboratory used nonradioactive cell proliferation and cytotoxicity assays to investigate GA-DM's effect on both androgen dependent and independent prostate cancer cell lines, revealing dose dependent killing and inhibition of proliferation by this natural extract. Microscopic analysis using Hoechst staining showed DNA breakdown consisted with apoptosis in GA-DM-treated prostate cancer cells. Annexin V staining confirmed low levels of apoptotic cell death when treated with GA-DM. Western blotting and biochemical analysis revealed that GA-DM induced apoptotic cell death that was independent of caspase activation, cytochrome c release, and Bax induction. Further investigation showed that GA-DM treatment activated HLA protein expression, implying some immunological involvement contributing to GA-DM-induced cell death. This study outlines a novel mechanism of GA-DM-induced inhibition of proliferation, induction of apoptosis and immune components in both androgen dependent and independent prostate cancer cells, the implications of which support GA-DM's potential use in multimodal chemoimmunotherapeutics.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 5487. doi:10.1158/1538-7445.AM2011-5487
Abstract 5489: An amiloride derivative capable of inducing ER stress and cell death in highly invasive breast cancer
Authors: Leon Leonardo J. , Harley William , Carraway Kermit , Gorin Fredric
Published: June 2012

Metastasis, which is the spread of tumors from their place of origin to other locations in the body via the lymph or blood, is associated with a significant increase in morbidity and mortality. Studies on amiloride, an FDA-approved diuretic, have shown it to suppress metastatic characteristics such as cell invasion and suppression, and impair their proliferation and viability. Here we show that 38B, a cell-permeant amiloride derivative, is more efficacious at causing cell death of cancer cells than amiloride. We also show that the cell death produced by 38B does not seem to be caused by apoptosis, autophagy, or calcium-dependent necrosis. However, bright field and immunofluorescence microscopy reveal vacuole formation and ER stress in cells, characteristics typically associated with a recently discovered type of cell death known as paraptosis. Since cancer cells have shown to be efficient at evading common forms of cell death such as apoptosis, the discovery of alternative pathways of cell death and a good understanding of the drugs that activate them can be can be key in the improvement of cancer therapy.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 5489. doi:10.1158/1538-7445.AM2011-5489
Abstract 5490: Mice deficient in uracil DNA glycosylase (UDG) display increased sensitivity to the antifolate pemetrexed
Authors: Weeks Lachelle D. , Liu Min , Liu Lili , Gerson Stanton L.
Published: June 2012

Pemetrexed, a multi-target antifolate, causes genomic uracil incorporation through inhibition of the enzymes thymidylate synthetase (TS) and dihydrofolate reductase (DHFR). Base excision repair (BER), initiated by uracil DNA glycosylase (UDG), actively recognizes and removes misincorporated uracil from the genome. Mice deficient in UDG have been described and display increased genomic uracil accumulation, elevated spontaneous mutation frequency, and higher incidence of B-cell lymphoma. We have previously reported the in vitro induction of UDG in H460 human lung cancer cells after pemetrexed exposure and have illustrated hypersensitivity to pemetrexed in human colon cancer cell lines lacking UDG expression (DLD1 UDG-/-). Here, we have investigated the role of UDG expression on pemetrexed sensitivity in vivo using gene-targeted UDG-/- mice. Mice received either a single 150mg/kg dose of pemetrexed on day 1 or received 3 consecutive 150 mg/kg doses of pemetrexed on days 1-3 and were sacrificed on day 4 or 5, respectively. Compared to UDG+/+ and UDG+/- littermates, UDG-/- mice were markedly myelosuppressed following pemetrexed treatment, as evidenced by a ∼50% reduction in bone marrow cell numbers and a ∼70% reduction in total colony forming units (CFU). Moreover, marrow from pemetrexed-treated UDG-/- mice expressed higher levels of DNA damage markers γ-H2AX and cleaved-PARP than control samples suggesting the formation of DNA double strand breaks and activation of apoptosis in UDG-/- marrow following pemetrexed exposure. Western blot analysis further evinced Chk-1 phosphorylation (p-Chk1, Ser345) in marrow from UDG-/- mice but not UDG+/+ or UDG+/- samples, suggesting inadequate removal of pemetrexed-induced genomic uracil may stall DNA replication and contribute to the cytotoxicity observed. These data in addition with our novel observations of enhanced pemetrexed sensitivity in UDG-/- mouse embryonic fibroblast (MEFs), UDG mutant human lymphoblastoid cell lines (LCLs), and DLD1 UDG-/- cells, strongly suggest that UDG deficiency potentiates pemetrexed cytotoxicity in in vitro as well as in in vivo models. Additionally, our findings indicate a role for genomic uracil accumulation in the mechanism of cytotoxicity for pemetrexed. Furthermore, our data illustrate a synthetic lethal interaction between TS/DHFR inhibition and UDG deficiency and promote UDG as a target for improving pemetrexed efficacy.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 5490. doi:10.1158/1538-7445.AM2011-5490
Abstract 5526: Epigenetic repolarization of T lymphocytes from chronic lymphocytic leukemia patients using 5-aza-2’-deoxycytidine
Authors: Dubovsky Jason A. , Powers John J. , Gao Yang , Mariusso Luis F. , Sotomayor Eduardo M. , Pinilla-Ibarz Javier A.
Published: June 2012

T cell immune dysfunction has an important role in the profound immunosupression that characterizes chronic lymphocytic leukemia (CLL). Improper polarization of T cells has been proposed as one of the mechanism involved. Mounting data implicates chromatin regulation, namely promoter methylation, in the plasticity of naïve human T cells. Recent in-vitro evidence indicates that this plasticity may be phenotypically altered by using methylation inhibitors which are approved for clinical use in certain types of cancer. These results beg the question: can the ineffective polarization of T lymphocytes in the context of CLL be effectively modulated using methylation inhibitors in a sustainable therapeutic fashion? To answer this question our laboratory has studied the effects of 5-aza-2’-deoxycytidine (5A2) in helper and cytotoxic T lymphocytes from healthy donors and CLL patients in well characterized molecular and epigenetic signaling pathways involved in effective polarization. Moreover, we sought to investigate the consequences of methylation inhibitor treatment on lymphocyte survival, activation intensity, and naïve cell polarization. Our data indicates that 5A2 treatment can repolarize Th2 cells to effectively secrete interferon gamma, signal via T-bet, and achieve demethylation of critical Th1 specific promoters. Moreover, we demonstrate that 5A2 can force Th1 polarization of naïve T cells despite a strong IL-4 stimuli and a lack of IL-12. In conclusion our data seeks to define a modality in which improper or ineffective T cell polarization can be altered by 5AZA and could be incorporated in future therapeutic interventions.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 5526. doi:10.1158/1538-7445.AM2011-5526