bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 5492: The effects of whole body hyperthermia on tumor growth by a novel far infrared emitter
Authors: Uemura Hirotsugu , Ushida Hiroshi , Asahi Chiori , Miyazaki Yuka , Yoshikawa Kazuhiro , De Velasco Marco A.
Published: February 2011

Whole body hyperthermia (WBH) has been used alone or as an adjunct to radio- and/or chemotherapy for the treatment of various malignant diseases. The therapeutic effect of WBH in these conditions has been hypothesized to involve direct or indirect activation of the immune system and heat shock proteins (HSPs). In this study, we investigate the effects of hyperthermia in vitro and in vivo using BioRubber (BR, Yamamoto Bio-Technology Inc., Anaheim, CA), a revolutionary far infrared emitter on tumor growth. BR is a pliable state-of-the-art biomaterial that emits FIR in the 4-25μm spectral ranges.

For the in vitro experiments, cell growth assays and cell cycle analysis was performed in a total of twelve different cancer cell lines (HeLa, bladder, prostate and renal) sandwiched between BR. BR had no inhibitory effects on any of the tumor cells grown in vitro.

To determine the effects of BR therapy on tumor and normal tissue in vivo, KU7/GFP-bearing nude mice were housed in cages lined with BR for 28 days. Tumor size and core body temperature were recorded weekly. The core body temperatures of mice exposed to BR increased significantly by 1oC (P<0.05) after 2 weeks of exposure to BR. Although not significant, tumors of mice exposed to BR showed a clear tendency in tumor size reduction. Tumors were collected and anlyzed for histopathology, TUNEL and PCNA immunohistochemical analysis. Proliferation and apoptotic rates in tumors from mice exposed to BR were similar to control mice. Further histopathological analysis showed that BR-exposed tumors had remarkable granulocyte infiltration accompanied by increased necrosis. This suggest that BR exposure was indeed able to induce and sustain mild hyperthermia in vivo with reduced tumor growth. However, no direct inhibitory on the tumor cells resulted form BR-induced hyperthermia but rather appeared to have been an indirect effect of immuno-stimulation achieved from sustained WBH.

To further investigate the immunological component of hyperthermia-induced immune response, a syngeneic tumor model was carried out using mouse RENCA cells in immunocompetent and immunodeficient mice. Similar tumor growth inhibitory effects were seen in both immunocompetent and immunodeficient mice thus suggesting an innate immune response as a result of sustained WBH.

Together, these data suggest that BR may be a promising biomaterial in for use hyperthermia therapy: however, more research has to be conducted to elucidate the specific mechanisms involved in immuno-stimulation and whether the effects from BR-induced hyperthermia can be maximized with combination therapies.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 5492.
Abstract 5493: A new strategy to assess cytotoxicity using Raman spectroscopy of single isolated nuclei
Authors: Lin Hsin-Hung , Li Yen-Chang , Liu Chun , Chen Chung-Hsuan , Yu Alice L.
Published: February 2011

Single-cell biodetection systems have potential applications in biomedicine and pharmacotherapy, and Raman spectroscopy is an ideal tool for single-cell analysis. Raman spectroscopy biosensors can provide an information-rich “fingerprint” of the overall biochemical composition of single cell undergoing time-dependent changes of cellular biochemistry without reacting with many biologically active compounds. To minimize the interfering signals form cytoplasm, nuclei were isolated from cells for analysis by Raman spectrum to compare the cellar responses and biochemical changes that occur in the nuclei of the HL60 promyelocytic leukemia cell line and drug-resistant cells HL60/ADR after treatment with the antineoplastic drug, doxorubicin. With increasing concentrations of doxorubicin up to 80 nM, the intensities of individual peaks representing nucleic acids and proteins displayed a concentration dependent decrease in the nuclei of HL60 but not HL60/ADR. At 80 nM doxorubicin, there was a 55.6±2.2% (p<0.001) of control in the intensities of Raman peaks representing nuclei acid, but no change of Raman peak intensities of the nuclei of HL60/ADR (96.4±3.1% of control). A significant drop in the intensities of Raman peaks representing nucleic acid was evident around 8 to 10 hours after doxorubicin exposure. Upon incubation with 80nM doxorubicin for 24 hours, thymidine incorporation was 74.2±1.9% and 94±3.8% of control in HL60 and HL60/ADR, respectively and Alamar Blue assay was even less sensitive with the value of 102.0±1.8% and 101.7±2.5% of control in HL60 and HL60/ADR, respectively. To verify the sensitivity and reproducibility of single nuclei Raman spectroscopy, treatment of CCRF-CEM, a T lymphoblastoid cell line, and drug-resistant subline CEM/VBL with anti-microtubule agent, vinblastine was also examined. At 0.4 nM, the intensities of Raman spectrum for nucleic acid dropped to 77.2±1.5% (p<0.001) of control while thymidine incorporation was not significantly inhibited (94.8±4.1% of control). In comparison, CEM/VBL was resistant to vinblastine up to 50 nM with negligible changes in Raman peak intensities of CEM/VBL nuclei (96.8±2.7% of control), and thymidine incorporation was 102.5±1.2% of control. These findings demonstrate that this single nuclei Raman spectroscopy has the ability to detect and identify nuclear changes related to cytotoxicity at lower concentrations and in shorter times than conventional cell based assays. Thus, this new cytotoxicity detection strategy using Raman spectroscopy of single, isolated nuclei may be useful for rapid and sensitive detection of cellular changes in response to chemotherapeutic agents.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 5493.
Abstract 5496: Determination of drug-kinase residence time in a homogenous, low-volume format
Authors: Marks Bryan , Lebakken Connie , Riddle Steve , Somberg Richard , Hereley Sara
Published: February 2011

In addition to drug-target affinity, drug-target residence time is becoming recognized as a critical parameter influencing drug efficacy. Desired compound action can only take place when a drug is interacting with its target protein, which is governed both by the affinity of the compound for the target, as well as the dissociative off-rate of the compound from the drug-target complex. A compound with a slower dissociative off-rate may require reduced dosing schedule relative to a compound with a rapid off-rate, and effective target specificity may be increased if the off-rate for the desired drug-target complex is greater than that for off-target complexes. With respect to kinase inhibitors, a slow off-rate for Tykerb relative to the related EGFR inhibitors Tarceva and Iressa has been proposed to explained longer lasting effects in cell culture. Currently, off rates for kinases can be measured by surface plasmon resonance (SPR)-based instruments or by detailed enzyme kinetics experiments. While SPR yields highly quantitative data, it is low-throughput and requires costly instrumentation. Furthermore, it often involves covalent conjugation of protein targets to surfaces, which can lead to loss of protein function. Off-rate measurements by classical enzyme kinetics methods are established, but fairly labor-intensive. We have developed and will present data on a fully homogenous (in-solution) method for determining kinase-compound off-rates that is simple to perform in a standard fluorescence-based plate reader. In this method, a drug-kinase complex is first formed at supra-Kd concentrations of drug, and then rapidly diluted into a solution containing a fluorescent, ATP-site directed small molecule probe. As the compound dissociates from the kinase the fluorescent probe associates with the kinase, causing an increase in the fluorescence signal that can be monitored in real time.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 5496.
Abstract 5502: Therapeutic efficacy prediction by quantitative protein analysis in response to anti-cancer agents
Authors: Nishizuka Satoshi S. , Ishida Kazushige , Matsuo Teppei , Noda Hironobu , Iwaya Takeshi , Wakabayashi Go
Published: February 2011

Background: The conventional chemosensitivity test employs a growth suppression assay. However, surgically removed specimens often have low viability which makes the assay unreliable, since the poor cell growth may be associated with the low viability instead of the chemosensitivity of the cell. On the other hand, we do know that the level of proteins involved in signal transduction is changed prior to cellular responses to the anti-cancer agents. In the present study, we investigated if protein signaling within a 24-hour exposure of the anti-cancer agents can predict the cellular fate at 72-hour time points.

Materials and Methods: The human colon cancer cell line, HCT116, was exposed to three different DNA-damaging anti-cancer agents including gamma irradiation, ultraviolet, and doxorubicin. The anti-cancer agent-exposed cell pellets from different conditions (e.g., doses, and time-points) were collected to produce a lysate array, which finally accommodated more than 500 samples on a single glass slide. Array image and quantitative protein analyses were performed with the P-SCAN and ProteinScan programs.

Results: A total of 19 protein species were tested on a lysate array, providing kinetic information of the protein. In response to all three agents, p53 protein was stabilized and the level increased over time in a dose-dependent manner. Upon detailed time-course analysis, however, an oscillation was seen in response to gamma irradiation, indicating the presence of a feedback loop. Comparing the rate of increasing levels of p21 and CyclinD3 within 24 hours in response to doxorubicin administration, the rates are well associated with the status of cell fate. In other words, when the rate of p21 is higher than CyclinD3, it is associated with a G2 arrest at 72 hours, whereas a higher CyclinD3 rate is associated with apoptosis at 72 hours after drug administration.

Conclusion: Some events occurring at 72 hours can be predicted by seeing the quantitative level of protein kinetics within 24 hours. Most of the primary culture cells can survive for only a short period of time. The protein kinetics may be a useful clinical chemosensitivity predictor even when using low viability specimens.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 5502.
Abstract 5508: Screening Of Novel Thymidylate Synthase Inhibitors
Authors: Repalli Jayanthi , Robinson Matthew R. , Gumina Giuseppe , Berger Sondra H.
Published: February 2011

The nucleotide stress response (NSR) is defined as activation of nucleoside salvage pathways to rescue cells from the detrimental consequences caused by a deficiency of or increased demand for nucleotides. Thymidylate depletion due to loss of function of the enzyme, thymidylate synthase (TS) or exposure to TS inhibitors disrupts the balance in nucleotide pools and damages DNA leading to stress-induced cell death. In order to replete nucleotide pools, cells activate nucleoside (ENT 1) transporters to increase the influx of extracellular nucleosides. TS catalyzes the de-novo synthesis of thymidylate for DNA replication. The enzyme in humans undergoes conformational switching between active and inactive forms. We are testing the hypothesis that stabilization of the inactive conformation will not induce the resistance mechanisms that are observed after exposure to active site-targeted TS inhibitors. To identify lead compounds that bind to the inactive conformation, a NSR assay has been developed, which measures a stress response to TMP depletion. TS inhibitors that bind to either the active or inactive conformations will induce TMP stress which is measured as induction of uptake of a fluorescent probe, N2, N3-etheno-6-thiomethylpurine riboside (ETMPR). In order to differentiate TS inhibitors that are selective for active and inactive conformations, we conducted an experiment on paired isogenic cell lines that express either wild-type hTS (undergoes conformational switching) or a mutant hTS (R163K - stabilized in the active conformation). Cells exposed to the potent active site TS inhibitor, raltitrexed (RTX) exhibited a similar level of uptake of ETMPR. When cells are exposed to glutarate (inactive site inhibitor predicted by structure-based drug design), fluorescent nucleoside uptake was preferentially induced in cells expressing wild-type hTS, consistent with the prediction that glutarate is an inactive site targeted drug. Protein turnover studies revealed that glutarate decreases TS expression, which is opposite to the effect of active TS inhibitors, which stabilize TS to degradation. Collectively, the data indicate that the NSR assay will be useful in identifying lead inactive site-targeted drugs and that these drugs will not stabilize TS, a mechanism thought to cause resistance to active state TS inhibitors.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 5508.
Abstract 551: Protein biomarkers of human prostate cancer stromal cells
Authors: Khamis Zahraa I. , Iczkowski Kenneth A. , Sang Qing-Xiang Amy
Published: February 2011

In situ human prostate tumors are not lethal unless they become invasive and metastatic. For cells to become invasive, the prostate gland must undergo disruption of the basal cell layer and degradation of the basement membrane underneath the luminal epithelial cell layer. Although the roles of proteinases in breaking down the basement membrane have been well-studied, little is known about the factors that induce basal cell layer disruption, degeneration, and its eventual disappearance in invasive cancer. It is hypothesized that microenvironmental factors may affect the degradation of the basal cell layer which if protected may prevent tumor progression and invasion. In this study, we have revealed differential protein expression patterns between cells separated from different prostate pathologies and identified several important stromal proteins that may contribute to inflammation and malignant transformation of benign prostate tissues to cancerous tissues using mass spectrometry and proteomics methods. The identifications of these proteins may generate new strategies for the prevention of basal cell layer disruption leading to the eventual blockade of invasive prostate carcinomas. Furthermore, human matrix metalloproteinase-26/endometase/matrilysin-2 was also investigated and its expression in preinvasive human prostate cancer cells may be associated with the expression of some apoptotic markers. This work may contribute to our understanding of epithelial-stromal interaction in the progression of human prostate cancer. (This work was supported in part by grants DAMD17-02-1-0238 and W81XWH-07-1-0225 from DOD, US Congressionally Directed Medical Research Programs, a grant from the Elsa U. Pardee Foundation, and grants from Florida State University to Dr. Q.-X. Sang)

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 551.
Abstract 5513: Synergistic antitumor activity of NK012, a micelle-forming macromolecular prodrug of SN-38, combined with platinum compounds in human cancer xenograft models
Authors: Masuda Kuniko , Onda Takeshi , Akatsu Yuichi , Ichimura Eiji , Nakanishi Takeshi , Okamoto Kazuya , Morino Tomio
Published: February 2011

Irinotecan (CPT-11) is clinically used to treat various tumors, but low enzymatic conversion to its active metabolite, SN-38, limits its therapeutic effect. NK012, a micelle-forming macromolecular prodrug of SN-38, accumulates in tumor tissue and gradually releases SN-38 in enzyme-independent manner. NK012 showed much stronger growth inhibitory effects than CPT-11 at maximum tolerated dose (MTD) against all tested xenografted human tumors. Furthermore, NK012 showed much stronger antitumor activities than other cytotoxic agents clinically used for cancer treatment (taxanes, doxorubicin, carboplatin etc.).

Here, we report combination antitumor effects of NK012 with platinum compounds in human breast cancer, non-small cell lung cancer, colorectal cancer and hepatocellular cancer xenografts in nude mice. The dose of NK012 was set at 1.9-3.8 mg/kg/day to be sub-optimum for complete regression depending on the cell line. NK012 and carboplatin were administered by q7dx3. Although both of NK012 and carboplatin were ineffective as single agent (minimum T/C values >50%), their combination was effective (minimum T/C value = 8%) to cause tumor regression in MC-05-JCK breast cancer. In MC-05-JCK breast cancer, the TGDs (Tumor Growth Delay; tumor tripling time over control group delayed by treatment / tumor tripling time of control group × 100) by NK012 1.9 mg/kg/day, carboplatin 50 mg/kg/day (67% of MTD), and their combination were 41%, 29%, and 286%, respectively. In NCI-H460 non-small cell lung cancer, the TGDs by NK012 3.8 mg/kg/day, carboplatin 75 mg/kg/day (MTD), and their combination were 90%, 71%, and 403%, respectively. In Co-3 colorectal cancer, the TGDs by NK012 3.8 mg/kg/day, carboplatin 75 mg/kg/day, and their combination were 358%, 97%, and >489%, respectively. Thus, combination of NK012 with carboplatin showed remarkably synergistic effects, and it didn't accompany toxicity exacerbation. Similarly, combination of NK012 with cisplatin exhibited remarkable synergistic effects in Li-07-JCK hepatocellular cancer. The TGDs by NK012 3.8 mg/kg, one shot, cisplatin 10 mg/kg, one shot, and their combination were 58%, 136%, and 455%, respectively.

These results indicate that NK012 and platinum compounds combination regimen would be effective for treatment of patients with solid tumors including breast, non-small cell lung, colorectal and hepatocellular carcinomas.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 5513.
Abstract 5518: Bladder tumor-targeting peptide-mediated anti-tumor activity of a pro-apoptotic peptide
Authors: Jung Hyun-Kyung , Kwak Mi-Kyung , Lee Eun-Ju , Son Ji Woong , Park Rang-Woon , Kim In-San , Lee Byung-Heon
Published: February 2011

Bladder carcinoma is the most common urothelial malignancy in developed countries and the rate of recurrence is pretty high. Using phage display, we have previously identified a bladder tumor-targeting peptide with the amino acid sequence of CSNRDARRC (namely, Bld-1). Bld-1 was shown to target bladder tumor in carcinogen-induced tumor-bearing rats. A synthetic pro-apoptotic peptide KLAKLAKKLAKLAK, called (KLAKLAK)2, has been demonstrated to induce apoptotic cell death of tumor cells. In this study, we used Bld-1 as a homing domain to help the pro-apoptotic peptide selectively bind to bladder tumor cells and allow its internalization. Bld-1 preferentially bound to HT-1376 bladder tumor cells, while little binding to A549 lung tumor cells was observed. Confocal microscopic analysis demonstrated that Bld-1 was efficiently internalized into HT-1376 cells. Bld-1-conjugated D-enantiomer D(KLAKLAK)2 was synthesized and incubated with HT-1376 cells for 24, 48, and 72 h. Bld-1-D(KLAKLAK)2 exerted its cytotoxic activity on HT-1376 cells in a dose- and time-dependent manner. In addition, D(KLAKLAK)2 alone also showed cytotoxic activity after 48 h treatment but this was in a lesser extent as compared to Bld-1-D(KLAKLAK)2. The cell death induced by Bld-1-D(KLAKLAK)2 was due to apoptosis as shown by annexin V and propidium iodide staining. Bld-1-D(KLAKLAK)2 more efficiently inhibited tumor growth in HT-1376 bladder tumor-bearing mice than D(KLAKLAK)2. These results suggest that Bld-1-D(KLAKLAK)2 might be a useful therapeutic agent for targeted therapy of bladder tumor.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 5518.
Abstract 552: Microglia-stimulation of glioma cell invasion is dependent on EGFR and CSF1R signaling
Authors: Coniglio Salvatore J. , Kedrin Dmitriy , Dobrenis Kostantin , Symons Marc , Segall Jeffrey E.
Published: February 2011

The ability of glioma tumor cells to invade surrounding brain parenchyma is what makes complete removal of tumor nearly impossible resulting in a median survival of only 12 months for patients afflicted by this disease. Targeting the motile properties of this tumor therefore, is an attractive strategy for therapeutic intervention. Many recent studies have demonstrated a key role for the tumor microenvironment in promoting tumor cell invasion and metastasis. Our laboratory and collaborators have demonstrated that tumor associated macrophages (TAMs) play a critical role in breast carcinoma invasion and metastasis. Here I propose a similar mechanism may occur between glioma cells and microglia within the brain. In this study we demonstrate that murine microglia are able to enhance GL261 glioma cell invasion into Matrigel in co-culture assays by up to 3 fold. Furthermore, we have found using pharmacological and/or antibody mediated inhibition of EGFR or CSF1R signaling abrogates this effect. GL261 cells constituitively secrete CSF-1, the levels of which are unaffected by EGF stimulation, EGFR inhibition or coculture with microglia. Disruption of EGF binding to EGFR using an EGF-specific function-blocking antibody also inhibits microglia-stimulated invasion, showing that EGF is responsible for mediating invasion. This data strongly suggests a paracrine loop exists between microglia and glioma similar to that seen in the breast cancer model. It is our hope that the results of this study may offer novel therapeutic approaches against glioblastoma by targeting functions of the tumor associated microglia as well as the glioma cells.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 552.
Abstract 5520: Thermally targeted proapoptotic peptide for cancer therapy
Authors: Moktan Shama , Raucher Drazen
Published: February 2011

Peptide based therapy is a promising field of anti-cancer therapy. The biologically active therapeutic peptides, derived from an endogenous protein or designed synthetically, can inhibit or activate important cellular pathways to promote cancer cell death. The major challenges of peptide based therapy are stability and tumor-specificity of the therapeutic peptides. One of the methods to increase stability and target the therapeutic peptide to the tumor site is to utilize stimulus-responsive macromolecular carriers.

We have designed a therapeutic peptide, SynB1-ELP-(KLAKLAK)2 based on the thermally responsive biopolymer, elastin-like polypeptide (ELP) and the mitochondrial membrane disrupting proapoptotic peptide, (KLAKLAK)2. Additionally, a cell penetrating peptide, SynB1 has been included in the N-terminus of this polypeptide in order to increase the polypeptide's membrane permeability. SynB1 is an 18-mer peptide derived from protegrins, which are antimicrobial peptides found in porcine leukocytes. The utilities of ELP are two-fold - 1) it is a macromolecule and therefore can provide stability to the overall polypeptide system, and 2) it is thermally responsive and therefore can be targeted to the tumor site by external application of local hyperthermia. The unique phase transition property of ELP allows it to remain soluble at physiological temperature but forces it to aggregate in response to mild hyperthermia (40 - 42 °C). Therefore, SynB1-ELP-(KLAKLAK)2 can be targeted to solid tumors by the application of local hyperthermia without being degraded before reaching the tumor.

The anti-proliferative effect of SynB1-ELP-(KLAKLAK)2 was examined in MCF-7 breast cancer cells. While the polypeptide was not cytotoxic to cells at 37 °C, it killed about 40% of cells at 42 °C after 6 days with two one hour treatments. Similar treatment schedule resulted in approximately 4-fold increase in apoptosis induction at 42 °C compared to 37 °C. Confocal microscopy showed that SynB1-ELP-(KLAKLAK)2 localizes in the mitochondria which supports the mitochondrial membrane disrupting feature of the (KLAKLAK)2 peptide. In conclusion, SynB1-ELP-(KLAKLAK)2 localizes in the mitochondria and inhibits the proliferation of MCF-7 cells in combination with hyperthermia by inducing apoptosis. Further studies include determining the proapoptotic effects of SynB1-ELP-(KLAKLAK)2 in vivo.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 5520.
Abstract 5521: IL13 linked lipsomal doxorubicin for malignant peripheral nerve sheath tumors
Authors: Madhankumar Achuthamangalam B. , Slagle-Webb Becky , Park Annie , Klinger Marianne , Pang Min , Harbaugh Kimberly , Sheehan Jonas M. , Connor James R.
Published: February 2011

We have previously demonstrated the therapeutic effect of interleukin-13 (IL13) linked liposomal doxorubicin (IL13LDXR) on glioma tumors. These tumors selectively overexpress IL13 receptor α2, a receptor for IL13. In our subsequent study, we found that malignant peripheral nerve sheath tumors (MPNST) also express the receptor for IL13. This observation prompted us to investigate the therapeutic value of IL13LDXR in a MPNST model. IL13LDXR was formulated and characterized in our laboratory using our previously established method. In vitro binding and internalization studies and subsequent fluorescence microscopy demonstrated the effective nuclear delivery of the doxorubicin in the MPNST cells expressing IL13Rα2 receptor. The cytotoxic potential of IL13LDXR was demonstrated in MPNST cell lines sNF96.2, ST88-14 and #215. Immunohistochemistry was performed on malignant and benign peripheral nerve sheath tumor (PNST) sections ex vivo to determine the expression of IL13Rα2 receptor. In addition we also demonstrated the expression of P-glycoprotein, a multidrug resistant protein in MPNST cell lines by immunocytochemistry. We also studied the effect of IL-13 protein in altering the expression of Pgp expression in established MPNST cell lines. We confirmed by an immunoblot that IL13 protein at 5µM concentration was able to decrease the Pgp expression in ST88-14 and sNF96.2 cells. This may explain the mechanism by which the IL-13 conjugated liposomes can deliver the encapsulated doxorubicin in a much more effective manner to drug resistant MPNST cell lines. We investigated the correlation of IL13Rα2 receptor expression with P-glycoprotein a drug resistance associated protein by immunocytochemistry. We confirmed the expression of IL13Rα2 in PNST tumors and established MPNST cell lines. We attempted to develop a MPNST tumor model in athymic nude mice by matrigel implantation method. We administered a single injection of 15 mg/kg of IL13 linked liposomal doxorubicin in a mouse tumor model by intraperitoneal route and observed the selective accumulation of doxorubicin in the tumor by endogenous fluorescence of doxorubicin, whereas no fluorescence was observed in the normal skin region as observed under fluorescence microscope. In conclusion consistent expression of IL13Ra2 receptors in PNST and MPNST cells and tissues indicate a viable approach to use this receptor as a target for MPNST therapy. The doxorubicin accumulation property and its cytotoxic potential on MPNST cells when delivered though the targeted liposomes envisage IL13LDXR as a potential drug for MPNST.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 5521.
Abstract 5523: Gold nanoshells in the treatment of high-grade gliomas in a mouse model
Authors: Thompson Patrick A. , Day Emily S. , Zhang Linna , Wong Kelvin , Gaber Mostafa W. , Blaney Susan M. , West Jennifer L.
Published: February 2011

Purpose: To evaluate the anti-tumor activity of near-infrared (NIR) absorbing gold-silica nanoshells against high grade gliomas in an in vivo subcutaneous mouse model. Background: Nanoshells with a silica core and a gold outer layer, which are optically tunable with peak absorption between 790 and 810 nm, have been shown to preferentially accumulate in colon cancer models after i.v. administration as a result of the enhanced vascular permeability and retention effect. When the tumor is illuminated with NIR light, the gold nanoshells are thermally activated. This results in tumor ablation without damage to normal tissues, since NIR light is not appreciably absorbed by tissue components at these wavelengths. High-grade gliomas are an attractive target for this type of therapy because the vast majority of tumors remain incurable with multi-modality therapeutic approaches, with local control being the primary manifestation of disease progression or recurrence. Methods: Gold silica nanoshells were synthesized according to previously published procedures. Survival studies were performed in NOD SCID mice with subcutaneous U373 glioma tumors in the right hind flank. When the tumor diameter was 3-5 mm, either 100 µl of PEG-coated nanoshells (treatment group, n =7) or 100 µl of saline (control group, n=8) were injected via tail vein. Twenty-four hours later tumors were exposed to an 808 nm diode laser at 4 W/cm2 for 3 minutes. Mice were monitored daily following treatment and euthanized when tumors reached 10 mm in largest diameter. Results: In the subcutaneous xenografts the overall survival at 90 days was 57% in the treatment group and 0% in the control group. None of the control group mice survived past 24 days. In addition, all of the mice in the treatment group that were still alive at 90 days were completely tumor-free without any sign of recurrence. Conclusions: These studies demonstrate that gold nanoshells can accumulate in subcutaneous glial tumors at levels sufficient to improve survival following thermal activation with NIR light.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 5523.
Abstract 5527: Short-chain glucosylceramide improves doxorubicin delivery in spontaneous mouse breast carcinomas
Authors: van Hell Albert J. , van Lummel Menno , Koning Gerben A. , van Blitterswijk Wim J. , Verheij Marcel
Published: February 2011

Background: Insufficient delivery of chemotherapeutic agents to their intended intracellular targets within the tumor remains a major obstacle in clinical oncology. Most of these drugs are amphiphilic and their diffusion through the plasma membrane is, therefore, slow and a major limitation for sufficient uptake within the tumor cell. We have found that certain short-chain sphingolipids enhance the intratumoral drug accumulation by modulation of inward translocation of cytostatics over the plasma membrane. Here, we examine this novel approach in an in vivo mouse model of aggressive invasive lobular breast carcinoma (ILC), designed to closely mimic its human counterpart.

Results: In a panel of cell lines, short-chain sphingolipids, such as C8-glucosylceramide (GC) increase the intracellular accumulation of doxorubicin up to 4-fold. A clinically approved drug carrier (100 nm liposomes) was employed for co-delivery of both lipid and drug to the tumor tissue. The addition of 10 mol% GC markedly enhanced the cellular doxorubicin content in vivo, as was demonstrated with intravital fluorescence microscopy. In the metastatic and aggressive mouse ILC tumors, the efficacy of GC doxorubicin liposomes was significantly improved compared to the currently applied conventional doxorubicin liposomes. The enrichment with the GC membrane modulator resulted in a significant tumor growth delay (p = 0.0119) and an overall survival benefit (p = 0.0143) for mice treated with the GC enriched liposomal doxorubicin as compared to conventional liposomes.

Conclusion: Short-chain sphingolipids improve the translocation of various cytostatics, including doxorubicin, over the plasma membrane of tumor cells. This membrane modulating strategy can be successfully applied in vivo, e.g. as a co-formulation with doxorubicin liposomes, thereby enhancing their anti-tumor efficacy. Furthermore, our data suggest that GC-enriched doxorubicin liposomes hold promise as anti-neoplastic therapy in human breast cancer.

Note: This abstract was not presented at the AACR 101st Annual Meeting 2010 because the presenter was unable to attend.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 5527.
Abstract 5528: Toxicology and drug delivery by Cucurbit[n]urils and derivatives
Authors: Hettiarachchi Gaya , Nguyen Duc , Wu Jing , Lucas Derick , Ma Da , Isaacs Lyle , Briken Volker
Published: February 2011

Drug candidates are continuously being discovered using high-through put screens of small compound chemical libraries, however, many of these drugs are not pursued into development because of low bioavailability. Therefore, drug delivery systems that improve bioavailability are of great importance. Cucurbit[n]urils (CB[n]) are a new class of molecular container compounds that are beginning to show great promise as a novel drug delivery system. Here we analyzed the toxicology of two unmodified cucurbit[n]urils, (CB[5] and CB[7]), and three CB[n] derivatives (Pentamer, Me-Hexamer and Ph-Hexamer). All five containers showed high cell tolerance at concentrations of up to 1mM in HEK293, HepG2 and RAW264.7 cell lines using assays for metabolic activity and cytotoxicity. We also studied uptake, and bioactivity using CB[7], the most soluble of the containers presented above. Bioactivity assays conducted with RAW264.7 cells infected with M. Smegmatis showed no inhibition of the activity of the antituberculosis drug ethambutol (EMB) by CB[7] after 3 days of incubation. This study provides a proof of principle towards the use of these containers as a novel drug delivery system.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 5528.
Abstract 5532: Folate receptor expression in human malignancies to select patients who would benefit from folate conjugated therapy
Authors: Covello Kelly L. , Wiebesiek Amy , Alland Leila , She Yijin , Wu Qiuyan , Lee Francis
Published: February 2011

The folate receptor (FR) is expressed on a number of human tumors, including ovarian, renal, lung, and breast. Folic acid is the ligand for the FR and many innovative strategies for targeting therapeutics to tumors make use of this ligand/receptor pair. Novel therapeutic approaches include folate conjugation to cytotoxics or radiopharmaceuticals, folate-coated liposomes containing antisense oligonucleotides, folate conjugates of pro-drugs, and folate-linked nanoparticle carriers for therapeutic drugs and genes. Potential patient enrichment strategies are necessary to allow for better targeting of these agents to those patients who have FR expressing tumors. Preclinical experiments to evaluate the molecular correlation of FR levels by three different methods, qRT-PCR, IHC, and the radiolabeled ligand binding assay, were conducted. A total of seventeen samples (four xenografts and thirteen primary patient ovarian and endometrial tumor biopsies) were evaluated for FR expression in these three independent assays and the FR levels correlated in this sample size. Compared to a preclinical model that was sensitive to epothilone-folate conjugate therapy, most patient samples expressed higher FR levels. These data provide a reliable and useful method to identify patient tumors that express the FR above a baseline determined to be sufficient for response to folate-conjugated therapy in preclinical models.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 5532.
Abstract 5535: Delivery of siRNA via functionalized lipopolyamine systems
Authors: Polach Kevin J. , Sparks Jeff , Congo Richard , Slobodkin Gregory , Rea-Ramsey Angela , Pence Casey , Matar Majed , McClure Diane , Rice Jennifer , Brunhoeber Elaine , Wilkinson Leslie , Anwer Khursheed , Fewell Jason G.
Published: February 2011

The efficient delivery of siRNA molecules to target cells is critical for the application of RNAi to the treatment of human cancers. To this end, we have developed novel cationic lipopolyamime structures that can be used to complex or encapsulate siRNA into nanoparticles for efficient in vitro or in vivo delivery. Unlike conventional cationic delivery systems, these lipopolyamines are chemically flexible which allow for covalent attachment of functional moieties which can modulate their biological activity. For example, modification of these systems with serum stabilizing agents allows for modulation of physical parameters important for biological properties such as toxicity and biodistribution. Intravenous (iv) injection of these functionalized nanoparticles showed preferential distribution to the lungs and corresponding depletion of targeted mRNA transcripts from lung tissue. We have investigated these systems for use in delivering siRNA targeting VEGF in mice with advanced disseminated ovarian cancer through intraperitoneal (ip) injection of siRNA-lipopolyamine nanoparticles. This model was established by injecting animals (ip) with 2.5×106 ID8 cells (malignant transformed ovarian surface epithelial cells that have been further modified to overexpress VEGF protein). Delivery of serum stabilized complexes to the peritoneal cavity of mice with disseminated ovarian cancer resulted in significant target gene knockdown in cells obtained from the malignant ascites fluid of these animals with modest knockdown in the peritoneal tumors themselves. A further modification of the core lipopolyamine structure incorporated attachment of a targeting peptide ligand through a linker molecule. Physico-chemical characterization of this targeted system and in vitro assays with ID8 cancer cells demonstrated specific delivery of siRNAs dependent on the presence of the targeting ligand on the liposome and the targeted receptor on the cell surface. These results have prompted in vivo testing using the targeted delivery systems to assess siRNA distribution as a function of delivered dose. Additional studies are underway that will establish the optimized delivery conditions for use in tumor bearing animals.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 5535.
Abstract 554: Crosstalk between TNF-α and IGF signalling increases IL-6 production and protects tumor cells from TNF-α-induced apoptosis in the liver
Authors: Li Shun , Yang Long , Lin Rongtuan , Hiscott John , Brodt Pnina
Published: February 2011

Background: Liver metastasis is a common and often fatal occurrence in cancer patients. A better understanding of the biology of liver metastasis is essential to improve survival statistics for these patients. We have previously shown that metastatic lung and colon carcinoma cells that invade the liver initiate a rapid host inflammatory response that entails increased production of cytokines TNF-α and IL-1 and results in increased tumor adhesion, transmigration and metastasis. TNF-α is a pleiotropic cytokine that can also induce tumor cell apoptosis. We hypothesized that the ability of tumor cells to form liver metastases in the face of increased local TNF-α levels is due to an acquired resistance to the tumoricidal effects of TNF-α. The objective of the present study was therefore to identify molecular mechanism(s) that regulate the tumor cell response and sensitivity to TNF-α.

Methodology: We used a Lewis lung carcinoma-based tumor model consisting of poorly metastatic subline M-27 cells in which ectopic expression of the type 1 insulin-like growth factor receptor (IGF-IR) markedly increased the liver-metastasizing potential of the cells. TNF-α-mediated signalling in these (M-27IGFIR) and wild-type (M-27) cells were analyzed by a combination of Western blotting, electrophoretic mobility shift assays, cytokine profiling, immunofluorescence labelling and flow cytometry.

Results: We found that IGF-I receptor overexpression in the tumor cells altered TNF-α-mediated signalling by accelerating TNF-α-induced IκBα phosphorylation and degradation and enhancing nuclear translocation of NFκB. This resulted in increased IL-6 production in M-27IGFIR, but not M-27 cells in response to TNF-α. IL-6 expression could be further augmented by co-stimulation with IGF-I, an effect that was mediated via PI3-K signalling. Furthermore, the level of activated STAT-3 in M-27IGFIR cells was significantly increased, suggesting that IL-6 could activate STAT-3 in these cells through an autocrine mechanism. Finally, when apoptosis induction by TNF-α in these cells was compared using antibodies to cleaved caspase 3, a significant reduction in the proportion of apoptotic cells was seen in TNF-α-treated M-27IGFIR as compared to M-27 cells.

Conclusions: Taken together, the results suggest that overexpression of IGF-IR altered TNF-α signalling in the tumor cells, shifting the response towards increased IL-6 production and autocrine STAT-3 activation, thereby providing the cells with a survival advantage in the presence of TNF-α.

Supported by Canadian Institute for Health Research grant MOP- 81201 (PB) and a McGill University Health Center Research Institute fellowship (SL).

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 554.
Abstract 5543: Combining antibody-targeted radiation (radioimmunotherapy) and antibody-SN-38 conjugates (ADC) improves pancreatic cancer therapy
Authors: Karacay Habibe , Govindan Serengulam V. , Sharkey Robert M. , Goldenberg David M.
Published: February 2011

We previously reported effective anti-tumor activity in nude mice bearing human pancreatic tumors with 90Y-humanized PAM4 IgG (hPAM4; 90Y-clivatuzumab tetrexetan) that was enhanced when combined with gemcitabine (GEM). These studies led to clinical testing of fractionated 90Y-hPAM4 IgG combined with GEM that is showing encouraging objective responses. While GEM is known for its radiosensitizing ability, alone it is not a very effective therapeutic agent for pancreatic cancer and its dose is limited by hematologic toxicity, which is also limiting for 90Y-hPAM4 IgG. We recently reported promising anti-tumor activity with an ADC composed of hRS7 IgG (humanized anti-epithelial glycoprotein-1; EGP-1) and SN-38, the active component of irinotecan. This ADC is very well tolerated in mice (eg, ≥ 60 mg), yet just 4.0 mg (0.5 mg, twice-weekly x 4) is significantly therapeutic. Since EGP-1 is also expressed in most pancreatic cancers, we have examined combinations of 90Y-hPAM4 IgG with RS7-SN-38 in nude mice bearing 0.35 cm3 subcutaneous xenografts of the human pancreatic cancer cell line, Capan-1. Mice (n=10) were treated with a single dose of 90Y-hPAM4 IgG alone (130 µCi, ie, the maximum tolerated dose or 75 µCi), with RS7-SN-38 alone (as above), or combinations of the 2 agents at the two 90Y-hPAM4 dose levels, with the first ADC injection given the same day as the 90Y-hPAM4. All treatments were tolerated, with ≤ 15% loss in body weight. Objective responses occurred in most animals, but they were more robust in both of the combination groups as compared to each agent given alone. All animals in the 0.13 mCi 90Y-hPAM4 IgG + hRS7-SN-38 achieved a tumor-free state within 4 weeks, while other animals continued to have evidence of persistent disease. These studies provide the first evidence that combined radioimmunotherapy and ADC can enhance efficacy at safe doses. (Supported in part by SBIR grant 2R44CA1148 from the NCI.)

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 5543.