bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 3534: Proposed pathway of disposition of ON 01910. Na, a novel clinical trial stage anti-cancer agent: Implicaton of Mrp2 in biliary excretion in the isolated perfused rat liver system
Authors: Taft David , Chun Amy , Ren Chen , Maniar Manoj
Published: February 2011

Introduction: ON 01910. Na is a novel targeted anti-cancer agent under clinical investigation in Phase I and Phase II trials. Preclinical studies indicate that the compound preferentially distributes to the liver but is not extensively metabolized in vivo The purpose of this study was to evaluate the disposition of ON 01910. Na employing the isolated perfused rat liver (IPRL) model. The specific goals were to 1) Assess the dose-linearity of ON 01910. Na disposition; 2) Probe the role of the Mrp2 transporter on ON 01910. Na biliary excretion; and 3) Establish the effect of concurrent dosing with oxaliplatin and doxorubicin on ON 01910. Na hepatobiliary disposition. Methods: Perfusion experiments (n=3/group) were performed at 4 doses (0.8, 4, 8, 20 mg), targeting a range of perfusate levels between 10 and 250 ug/ml. ON 01910. Na (10 ug/ml) disposition was then studied in the presence of doxorubicin (2.5 ug/ml) and oxaliplatin (2 ug/ml). IPRL experiments were also conducted using livers from Mrp2 deficient rats. ON 01910. Na was assayed in perfusate and bile samples by HPLC. Results: ON 01910. Na parameter estimates are provided in the table below. The compound displayed nonlinear excretion in the IPRL, and ON 01910. Na disposition was altered in mrp2-deficient rats. The effect of co-administered chemotherapeutic agents is being investigated. Conclusions: ON 01910. Na showed extensive biliary excretion in the IPRL, and IPRL findings correlated with in vivo data in rats. ON 01910. Na biliary transport appears to be mediated in part by Mrp2.ON 01910. Na Disposition in the IPRLaPARAMETERON 01910. Na (µg/ml) 10.050.0100250t1/2 (hr)0.460 (0.0340)0.327 (0.0674)0.585 (0.0810)1.26 (0.318)Cmax (µg/ml)11.3 (1.75)56.1 (7.12)106 (11.2)257 (658)AUC (0-∞) (µg*min/ml)290 (87.1)1480 (73.0)4010 (963)21100 (2580)Cl (ml/min)2.91 (0.753)2.71 (0.134)2.08 (0.557)0.959 (0.125)% Drug Excreted in Bile57.2 (5.04)59.2 (7.40)68.4 (0.870)45.6 (2.66)aData presented as mean (standard deviation)

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 3534.
Abstract 3537: P-glycoprotein expression level is responsible for alterations in plasma membrane microfluidity as probed by fluorescence correlation spectroscopy
Authors: Winckler Pascale , Jaffiol Rodolphe , Morjani Hamid , Deturche Regis , Royer Pascal , Jeannesson Pierre
Published: February 2011

Expression of P-glycoprotein (P-gp), the multidrug resistance (MDR) 1 gene product, leads to MDR in tumors. By using the fluorescent membrane probe DiA and Fluoresence Correlation Spectroscopy (FCS) to explore the plasma membrane microfluidity, we have previousely shown a showed a decrease in plasma membrane microfluidity in LR73R cells (the Chinese hamster ovarian LR73R cells transfected with MDR1 cDNA) (1).

In the present study, and in order to study the relationship between membrane microfluidity and the level of P-gp expression, a strategy using flow cytometry and calcein-AM functional fluorescence-activated cell sorting permitted us to select two cell populations from LR73R cells expressing distinct levels of P-gp (P4 “lower calcein-AM uptake” and P5 “higher calcein-AM uptake”). Data obtained on the cytotoxic effects of doxorubicin (DOX) showed clearly that P4 cells were 10-fold resistant to DOX than P5 cells. A direct correlation between the sensitivity of the cells to DOX and the calcein-AM uptake was observed. In addition, intranuclear DOX measurements using microspectrofluorometry showed a decrease in nuclear accumulation of DOX in P4 cells when compared to P5 cells. This was in agreement with the functional calcein-AM uptake assay.

By using the fluorescent membrane probe DiA FCS analysis, we explored the membrane microfluidity in P4 and P5 cells. We show that P5 cells presented an increase in membrane fluidity when compared to P4 cells. LR73 cells presented the highest microfluidity when compared to P4 and P5 cells. This was in agreement with the previous published data (1) and demonstrate also that the microfluidity of the plasma membrane is strongly related to the level of Pgp expression.

(1) Boutin C, Roche Y, Millot C, Deturche R, Royer P, Manfait M, Plain JM, Jeannesson P, Millot JM, Jaffiol R. High heterogeneity of plasma membrane microfluidity in multidrug-resistant cancer cells. J Biomed Opt. 2009 14(3): 034030.

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 3537.
Abstract 3544: Assessment of chemo-response in cells derived from patients with malignant ascites
Authors: Peedicayil Abraham , Rattan Ramandeep , He Xiaoping , Kalli Kimberly R. , Cliby William A. , Chien Jeremy , Shridhar Vijayalakshmi
Published: February 2011

Objectives. To assess chemo-response of patient derived ovarian cancer cells to first line chemotherapeutic agents as well as targeted therapies.

Methods. Ascites was obtained at laparotomy from 14 patients with epithelial ovarian cancer. Short term cultures were established in 11 of these and the cells characterized by morphology and immuno-staining to cytokeratin AE1/AE3 and E-cadherin. MTT assays were done on the 11 samples after 48 hours of exposure to Cisplatin, Paclitaxel, Vandetanib and Flavopiridol.

Results. Three samples did not grow well and ultimately showed senescence or were over-run by fibroblasts. In the remaining 11 samples, morphology showed the cells to be arranged in a cobble-stone or rosette pattern. Most of the cells stained positive for cytokeratin AE1/AE3 and E-cadherin. The majority of samples (8/11, 73%) were sensitive to either Cisplatin or Paclitaxel. Of the three resistant samples, one was sensitive to Vandetanib, one to Flavopiridol and one was resistant to all four drugs. Two patients did not receive chemotherapy due to morbidity or mortality. Among those who were treated, 4 (50%) had complete response and are currently free of disease.

Conclusions. Short term cultures may be a good method of studying chemo-response of tumor cells ex-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 3544.
Abstract 3552: Mirk kinase mediates cisplatin resistance in ovarian cancers
Authors: Hu Jing , Friedman Eileen A.
Published: February 2011

Epithelial carcinoma of the ovary is the fifth most frequent cause of cancer death in women, and the second most frequent gynecological malignancy. Cisplatin based regimens are the standard of care, but efficacy is limited by drug resistance. Cisplatin treatment increased intracellular levels of toxic reactive oxygen species (ROS) in each of four ovarian cancer cell lines in a dose-dependent manner, while concomitantly reducing cell numbers, which suggested that further increasing ROS levels might increase cisplatin toxicity. The serine/threonine kinase Mirk/dyrk1B is known to increase the expression of a cohort of antioxidant genes in various cells. Targeting Mirk expression by RNA interference also increased ROS levels in these ovarian cancer cell lines while decreasing levels of ferroxidase (ceruloplamin) which prevents the generation of hydroxyl radicals. A combination of Mirk depletion and cisplatin treatment enabled low levels of drug to increase ROS to toxic levels in both SKOV3 cells and TOV21G cells. Increased tumor cell death by the combination treatment was confirmed by loss of viable cells capable of dye exclusion, a decrease in cell number, and an increase in pro-apoptotic proteins. Mirk has a second function in ovarian cancer cells, to maintain cells in quiescent state where they are less vulnerable to chemotherapeutic drugs and radiation. The majority of ovarian cancer cells in human tumors are not cycling as shown by lack of Ki67 expression. Some of these cells may be in a reversible quiescent state where they can contribute to cancer spread under favorable growth conditions. Depletion of Mirk in SKOV3 cells enabled them to escape quiescence. Studies using cisplatin revealed a Mirk-dependent G0/G1 checkpoint in SKOV3 and other p53 mutant ovarian cancer cells. The Mirk gene is on the 19q13 amplicon which is found in about one-third of all ovarian cancers. The Mirk gene was amplified in the OVCAR3 cell line where it mediated both quiescence and increased expression of antioxidant genes which decreased cellular ROS levels. Mirk protein was expressed in 21 of 28 resected human ovarian cancers as shown by immunohistochemical analysis, and was detected in each of 7 ovarian cancer cell lines, where it was an active kinase. Mirk is not an essential gene for either murine embryogenesis or the viability of human diploid fibroblasts. Targeting Mirk in ovarian cancers may sensitize these tumors to lower levels of cisplatin, both by increasing ROS levels and by releasing tumor cells from quiescence.

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 3552.
Abstract 3563: Novel anticancer agents modulate multiple signal transduction pathways for cancer therapy
Authors: Jong Ling , Chao Wan-ru , Shi Yihui , Green Carol , Sambucetti Lidia , Collins Nathan
Published: February 2011

Cancer is remarkably heterogeneous and is associated with wide variations in biological behavior. Except for some rare cancers whose growth may depend on a single factor, no single signaling pathway drives the oncogenic behavior of all tumors. In fact, most single-target agents that have undergone development in recent years have lacked the exquisite level of effectiveness. Multitarget drugs and the network approach may thus constitute a useful means for discovering novel drugs; such drugs emerge from the sum of the interactions that, together, are the principal determinants of the system-scale behavior of the cell. A multitarget agent is not necessarily more toxic than single-target agents. These multitarget drugs have presumed low-affinity interactions with several of their targets, and do not necessarily shut down a pathway completely or activate it excessively; thus they may have lower side effects.

We has developed a novel class of dietary indole analogs with oral anticancer activity against a variety of cancer in animal models, including prostate, breast, lung and ovarian tumors. Western blot analysis shows that SR13654, SRI advanced lead, is able to induce apoptosis by caspase activation and poly(ADP-ribose) polymerase (PARP) cleavages. Using the Boyden chamber assay, we showed that SR13654 significantly inhibits PC-3 cells’ invasion at low concentration, and its anti-invasive activity is not due to cell killing or simple cell growth inhibition. In vitro and in vivo mechanistic studies show that SR13654 inhibits Src activation, down-regulates survivn expression, enhances EGFR degradation. SR13654 also produced a decrease in cyclin D1 and B1, and an increase in p21 levels in cancer cells. Screening a full panel of 285 kinases shows that SR13654 is not a kinase inhibitor. More in-depth mechanistic studies are undergoing. Preclinical safety studies indicate that this class of compounds holds great promise for extended therapeutic use to maximize disease control without compromising quality of life.

SR13654 has relatively poor water solubility, which could limit its use in intravenous dosing. We have developed the second generation of water-soluble analogs, such as SR13677, which has significantly improved water solubility and is at least 5-fold more potent than SR13654 against various cancer cell lines. With support from the 2009 DOD/Prostate Cancer Research Program (PCRP) “Laboratory-Clinical Transition” Award, we will advance the optimized candidate through preclinical development and future IND filing.

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 3563.
Abstract 3569: Photodynamic activity of the glucoconjugated silicon(IV) phthalocyanine SiPcGlu
Authors: FONG Wing-Ping , Cheang Wai-San , Lo Pui-Chi , Ng Dennis K.P.
Published: February 2011

Photodynamic therapy involves the use of a non-toxic photosensitizer which exhibits killing effect upon activation by light. It has been used successfully in treating age-related macular degeneration, actinic keratosis and various cancers like lung, bladder and esophageal cancers. The success of photodynamic therapy depends on the properties of the photosensitizer and there is a continuous search for better photosensitizers with desirable photophysical and biochemical effects. In the last few years, we have synthesized a series of silicon(IV) phthalocyanines with different axial substituents and demonstrated their photocytotoxic effects in the in vitro heptocellular carcinoma HepG2 cells. Among them the glucoconjugated silicon(IV) phthalocyanine SiPcGlu is most promising with IC50 value down to nanomolar range in the cell culture system. In the present investigation, the in vivo effect of SiPcGlu was studied in the HepG2 tumor-bearing nude mice model. The biodistribution of SiPcGlu was followed to evaluate its tumor-selectivity and the tumor volume was measured after photodynamic therapy. The level of intrinsic toxicity induced was also investigated by examining the level of plasma enzyme markers. The distribution and clearance results showed that the concentration of SiPcGlu in the tumor was relatively higher at 24 hr post-injection when compared with all the other tissues examined. One percent of initial dose per gram tissue could be found in the tumor while the corresponding values for the other tissues ranged from 0.8% (lung) to 2.5% (skin). Thus, a drug-light interval of 24 hr was selected in the photodynamic therapy experiment. A laser light dose of 30 J/cm2 (bandwidth 675 ± 3 nm) was spotted onto the tumor 24 hr after intravenous injection of the drug (one μmol per kg body weight). Significant tumor regression was observed for the experimental group of mice. Fifteen days after photodynamic treatment, the volume of the tumor was less than 20% of the initial volume while mice in the two control groups, viz. dark control in which the mice received drug but no irradiation and the light control group in which the mice received irradiation but no drug, did not show any tumor regression. The photodynamic treatment did not cause any apparent toxicity to the animals as the plasma levels of the three enzyme markers, alanine transaminase, aspartate transaminase and creatine kinase, were similar in all the three groups of mice. To conclude, the present results suggest that SiPcGlu is a good candidate for further evaluation on its anti-tumor potential. (This project is supported by a grant from the Research Grants Council of the Hong Kong Special Administrative Region, Project No. CUHK 4569/05M.)

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 3569.
Abstract 357: Induction of galectin-1 decreases radiosensitivity of patient-generated primary cell lines through AKT activation
Authors: Siedow Mike , Chakravarti Arnab , Lautenschlaeger Tim
Published: February 2011

Glioblastoma, grade IV malignant glioma based on the WHO classification, is one of the most common brain tumors found in adults and has an average survival time of twelve to sixteen months. The dismal prognosis associated with these tumors is largely due to their ability to grow and proliferate despite radiotherapy, which along with resection and Temozolomide is the current standard of care. A better understanding of the molecular mechanisms that define this malignant phenotype may result in new therapeutic targets and lead to improvements in how patients are treated. Galectin 1(GAL1), a beta-galactosidase binding protein, is strongly expressed in a variety of human cancers and has been shown to be upregulated by irradiation. In this study, we investigated the role of GAL1 upregulation following irradiation in a variety of patient-generated primary cell lines and its role in cell survival. Patient-generated primary cell lines were characterized using a variety of assays such as MTS, Annexin V, and cell viability. Western Blot analysis was performed probing for a variety of proteins implicated in proliferation, motility, and apoptosis. Radiation sensitivities were established by performing Clonogenic Survival Assays comparing the surviving fraction of patient-derived cell lines with established stable cell lines. 6 Gy irradiation upregulated Galectin 1 protein levels in the majority of primary cells used in our study. Furthermore, cell lines which Gal1was upregulated also displayed induction of Phospho-AKT and were significantly less sensitive to irradiation. The combination of GAL1 upregulation and Phosho-AKT induction following irradiation was associated with a significant decrease in the radiosensitization of the patient-derived primary cell lines used in our study. Interestingly, the patient-derived cell lines which showed no upregulation of GAL1 or Phospho-AKT induction displayed markedly increased sensitivity to radiation elucidating an interesting association between GAL1, Phospho-AKT, and radiation-sensitivity. The underlying mechanism by which GAL1 may medicate radioresistence seems to be through AKT activation, a well-documented protein involved in cell survival. Additional studies are ongoing to validate Galectin 1 as a target for radiation sensitization.

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 357.
Abstract 3578: New Angiochem-modified doxorubicin with increased brain penetration and efficacy against brain tumors
Authors: Demeule Michel , Régina Anthony , Yang Goagiang , Ché Christian , Béliveau Richard , Castaigne Jean-Paul
Published: February 2011

The blood-brain barrier (BBB) is mainly formed by brain capillary endothelial cells which are closely sealed by tight junctions and express high levels of active efflux transport proteins, including P-glycoprotein (Pgp). As a result, the overwhelming majority of small molecules, proteins and peptides do not cross the BBB. Angiochem's engineered peptide compounds (EPiC) provides a non-invasive and flexible platform for small and large molecules to treat brain diseases. Based on these properties, we have created a portfolio of new drug entities composed of siRNA, peptides and mAbs, the most advanced of which is ANG1005 formed by chemical conjugation of our peptide to three molecules of paclitaxel. ANG1005 demonstrated safety and efficacy in two phase ½ clinical trials for the treatment of primary and secondary brain tumors in humans. In the present study, we have investigated the brain uptake of a new chemical entity formed by conjugation of the peptide Angiopep-2 (An2) with 3 molecules of the anti-cancer drug doxorubicin. Despite the clinical effectiveness of doxorubicin in the treatment of many malignant tumors, clinical trials involving systemic administration have demonstrated very limited efficacy in the treatment of gliomas. This limited efficacy can be explained by the poor penetration of the drug thru the BBB and by the effect of the multidrug resistance pump, P-gp. Here, we show by mice in-situ brain perfusion that the ANG-doxorubicin (ANG1007) is transported very efficiently across the BBB. The transport rate is higher than that of unconjugated doxorubicin by at least 10-fold. In vitro, ANG1007 inhibits cancer cell proliferation, with highly cytotoxic activities against various tumor cell lines. In addition, brain perfusion studies performed with P-gp knock-out mice showed that ANG1007 bypasses the drug efflux pump P-gp at the BBB. Furthermore, ANG1007 inhibits growth of glioblastoma s.c. tumors and increases survival of mice implanted with brain tumors. In conclusion, these data confirm that conjugation of chemotherapeutic drugs such as doxorubicin to the peptide Angiopep-2 significantly enhances their entry into the brain and further validate the use of Angiochem's technology for new brain tumor 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 3578.
Abstract 3582: In vitro pharmacological studies of water soluble garlic extract (WSGE) in the management of acute promyelocytic leukemia (APL)
Authors: Yedjou Clement G. , Tchounwou Paul B.
Published: February 2011

Garlic (allium sativum) is one of the oldest cultivated plants, used in spices, food, and herbal medicine for over 4000 years. The therapeutic efficacy of garlic has a wide variety of ailments, including cardiovascular, cancer, hepatic and microbial infections. Although the therapeutic efficacy of garlic in the prevention of cancer and other diseases has received increasing attention in recent years, little is known about its mechanism for therapeutic action. In this research, we hypothesize that the pharmacological effect of garlic may be mediated through oxidative stress, DNA damage, modulation of annexin-5 and caspase-3 in human leukemia (HL-60) cells. To test this hypothesis, HL-60 cells were treated with various doses of water soluble garlic extract (WSGE) for 12 h. Cell survival was determined by MTT-assay. The extent of oxidative cell/tissue damage was determined by measuring the level of malonaldiahyde (MDA) concentrations. Cell apoptosis was measured by flow cytometry using annexin V and caspase-3 kits, respectively. MTT assay indicated a strong dose-response relationship with regard to the cytotoxic efficacy of WSGE in HL-60 cells. There was a slight increase in MDA concentrations in WSGE-treated cells compared to the control, but this was not statistically significant (P > 0.05) probably due to the antioxidant property of garlic. WSGE-induced apoptosis was characterized by a significant (P < 0.05) increase in the percentages of annexin-5 and caspase-3 positive cells. In summary, the pharmacotherapy of WSGE may be associated through induction of moderate toxicity, induction of DNA damage, phosphatidylserine externalization and caspase-3 activation characteristic of apoptosis.

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 3582.
Abstract 3584: Comparison of the severity of imatinib-induced cardiotoxicity in normotensive Sprague-Dawley (SD) and spontaneously hypertensive (SHR) rats
Authors: Herman Eugene H. , Knapton Alan , Rosen Elliot , Zhang Jun , Lu Quynh A. , Estis Joel , Todd John , Lipshultz Steven
Published: February 2011

Imatinib (IMNB) is a promising type of chemotherapeutic agent that specifically targets tyrosine kinase pathway-dependent tumors. However, cardiotoxicity has been reported with clinical use of IMNB. The characteristics of IMNB-induced cardiac alterations are not completely defined. The present study sought to examine the influence of a concomitant disease such as hypertension on the cardiotoxic effects of IMNB. Groups of adult male SD or SHR were dosed with 50 (5/group) or 100 mg/kg (10/group) IMNB or water (10/group)(p.o.) daily for 14 days (dose of 50 mg/kg is approximately 2x recommended human dose of 600 mg/m2). Tissues and blood samples were collected 24 hours after the last dosing. The 100 mg/kg dose caused a slight reduction in the rate of body weight gain. Serum levels of glucose were decreased and alanine transaminase (ALT) were increased in both SD and SHR dosed with IMNB. Changes from control were most pronounced in SD given 100mg/kg IMNB (ALT=175% vs 118% and glucose=81% vs 91% compared to SHR). White blood cell counts were depressed in SHR but not in SD rats. Dose-dependent cardiac lesions were noted in the groups of SD and SHR given either dose of IMNB (blinded evaluation). Cardiac lesions were characterized by cytoplasmic vacuolization, myofibrillar loss, interstitial infiltration with chronic inflammatory cells and fibrosis (proliferation of myofibroblasts). Mean lesion scores (based on a scale of 0 to 3) were higher in SHR than in SD (100mg/kg-1.9 vs 1.25 and 50 mg/kg-1.5 vs 1.1, p<0.05)(Tukey-Kramer test). Increased serum levels of cardiac troponin I were detected in all IMNB-treated groups. The overall mean cTnI levels were higher in SHR (31.5, 41.3 and 53.9 pg/ml) compared to SD (6.8, 25 and 30 pg/ml) at the vehicle control, 50 and 100 mg/kg doses, respectively. These results indicate that hypertension as expressed in SHR appears to be a factor that can intensify the cardiotoxic effects of IMNB and that monitoring for cardiac troponin I may provide a sensitive means of detecting IMNB toxicity.

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 3584.
Abstract 3594: Development of a TRPV1 inducible expression model in MCF-7 breast cancer cells and its potential role in the evaluation of calcium channels as viable therapeutic targets in cancer
Authors: Wu Tina , Robinson Jodie , Monteith Gregory R. , Roberts-Thomson Sarah J.
Published: February 2011

Calcium is an important intracellular messenger that regulates many physiological functions such as cellular differentiation, proliferation and apoptosis. Abnormal functioning of calcium signaling pathways may occur during tumorigenesis. Calcium signals are tightly controlled by calcium influx channels and many calcium channels, including those of the transient receptor potential (TRP) ion channel family have altered expression levels in specific cancer types, such as the upregulation of TRPM8 in some prostate cancers. Plasma membrane calcium channels belong to a protein class that can be effectively pharmacologically modulated. Pharmacological agents are available that modify specific TRP calcium channels and some voltage gated calcium channels, such as the inhibition of L-type calcium channels by the anti-hypertensive nifedipine. Both activators and inhibitors of calcium channels overexpressed in cancer cells are potential anti-cancer drugs. This is due to the duality of the calcium signal, whereby calcium can be both a promoter of proliferation but also an inducer of apoptosis. Therefore, calcium channels with increased expression and/or activity in cancer could potentially serve as anti-cancer therapeutic targets. To test this and to explore some of the likely issues related to calcium channels as drug targets in cancer, we have generated a Tet-Off inducible model system where the transient receptor potential vanilloid 1 (TRPV1) calcium channel is over-expressed in the MCF-7 Tet-Off breast cancer cell line. TRPV1 has been chosen in this model, due to the wide availability of selective inhibitors and activators to this channel. TRPV1 functional activity in the model system was measured by calcium influx using the Fluorescent Imaging Plate Reader (FLIPR) and the calcium sensitive probe Fluo-4. Capsaicin, a TRPV1 agonist had an EC50 of 2 × 10-7 M for increased cytosolic free calcium in the absence of doxycycline and an EC50 of 1 × 10-6 M in the presence of doxycycline (1µg/ml). In this same stable cell line, peak cytosolic responses were significantly higher in the absence of doxycycline compared to the presence of doxycycline. This suggests a tight regulation of TRPV1 expression in the Tet-Off system. Future studies will characterize the consequences of TRPV1 over-expression and TRPV1 pharmacological modulators on cellular proliferation and apoptosis pathways. This model system could be an important tool to address the feasibility of calcium channels as drug targets in cancer.

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 3594.
Abstract 3596: Comparison of intravenous microdialysis sampling to standard blood sampling to determine free etoposide exposure in non-human primates
Authors: Pastakia Devang , McCully Cynthia , Cole Diane , Bacher John , Cruz Rafael , Blakeley Jaishri , Widemann Brigitte , Balis Frank , Fox Elizabeth
Published: February 2011

Background: Microdialysis (MD) is a method for continuously sampling small molecules in tissue extracellular fluid in vivo. To determine if free (non-protein bound) drug exposure (AUC) in plasma could be accurately measured by MD under non-steady state conditions, we compared intravenous MD to standard serial plasma sampling during and after a short infusion of etoposide. Methods: CMA IView 64 catheters (10 mm PAES membrane with 20 kD MWT cutoff) were surgically implanted into the external jugular vein of Rhesus monkeys (Macaca mulatta). Catheter inflow and outflow tubing was tunneled subcutaneously from neck to the area on back between shoulder blades, brought through skin and attached to a MD pump (CMA 106 pump with fixed flow rate of 0.3 μL/min). The MD pump was secured in a fitted mesh jacket on the dorsum of the awake animal. Retrodialysis using 10 µM etoposide was used to measure in vivo relative recovery (RR) in each catheter. Following retrodialysis and catheter re-equilibration with normal saline, etoposide (5 mg/kg) was administered IV over 1 h. Serial plasma samples were collected during the infusion and for 8 h post-infusion. Free etoposide was extracted by ultrafiltration. MD samples were collected continuously over a single collection interval (0-9 h) or split into two collection intervals (0-3 h and 3-9 h). Etoposide was quantified using HPLC-electrochemical detection (linear range 0.05 - 10 μM). The etoposide concentration in MD samples was corrected for in vivo RR. The AUC was calculated by the linear trapezoidal method for serial plasma sampling and by summation of the concentration x time intervals for MD. Results: Simultaneous MD and serial plasma sampling was performed in 5 animals. The in vivo RR of etoposide ranged from 86.9 - 99.4%. The mean AUC0-9h for plasma free etoposide was 3.27 (median 3.81, range 2.11 - 5.15) μM·h. The mean AUC0-9h for MD sampling was 2.68 (median 3.02, range 1.75 - 3.65) μM·h. The mean ratio of AUCMD:AUCplasma was 0.81. The mean (95% CI) for the difference in the paired MD and plasma AUC was − 0.7 (−1.94, 0.54) µM·h. Conclusions: Quantitative MD sampling for pharmacokinetic studies is feasible and the in vivo RR of etoposide is high. Free etoposide exposure by MD slightly underestimates the AUC compared to serial plasma sampling. Intravenous MD sampling may be a promising alternative to standard plasma sampling for calculating free etoposide exposure.

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 3596.
Abstract 3599: The plasma and cerebrospinal fluid pharmacokinetics of satraplatin after intravenous administration in non-human primates
Authors: Marcus Leigh J. , Murphy Robert F. , Fox Elizabeth , McCully Cynthia L. , Cruz Rafeal , Warren Kathy , Balis Frank M. , Widemann Brigitte C. , Meyer Thorsten
Published: February 2011

Background: Satraplatin is an orally bioavailable platinum analog with preclinical activity (IC50 range in tumor cell lines 0.04 - 16 µM) in cisplatin sensitive and resistant models and clinical activity in adults with refractory cancers including prostate cancer. The cerebrospinal fluid (CSF) penetration of cisplatin and carboplatin in non-human primates (NHP) is limited (3.6% and 3.8%, respectively). We evaluated the plasma and CSF pharmacokinetics (PK) of satraplatin, which is more lipophilic than other platinum analogs, after an intravenous (IV) dose in a NHP model. Methods: Satraplatin 120 mg/m2 was administered as 1 h IV infusion in DMSO (5%) and normal saline to 5 NHP. Serial blood and CSF samples were obtained over 48 hours. Plasma was separated immediately and ultrafiltrate (UF) was prepared by centrifugation through a Microcon 10K MWCO filter. CSF was obtained from an indwelling Ommaya reservoir (n=4) or from a temporary lumbar catheter (n=1). Platinum was quantified in the plasma UF and CSF using a validated atomic absorption spectroscopy assay with lower limit of quantification of 0.03 µM in UF and CSF. Pharmacokinetic parameters were estimated using non-compartmental analyses. CSF penetration was calculated from the AUCCSF:AUCPlasma. Results: Satraplatin was well tolerated by all NHP. Pharmacokinetic parameters (median and range) for satraplatin plasma UF were Cmax 8.3 (5.7-10.6) µM, AUC0-48h 28.3 (22.6-33.2) µM·h, AUC0-inf 34.2 (24.7-38.2) µM·h, clearance 3.6 (2.4-4.3) L/h, and t1/2 18.8 (13.4-25) h. Satraplatin was detected in the CSF of all NHP. For the 4 NHP with Ommaya reservoirs PK parameters were: tmax 1.5 (1.3-2.1) h from the start of infusion, Cmax 0.07 (0.02-0.12) µM, AUC0-48h 1.2 (0.49-2.43) µM·h, ratio AUC0-48h CSF:AUC0-48h Plasma was 4.3 (2.2-7.4) %. For the NHP with lumbar catheter and limited CSF samples the tmax was 1.1 h, and Cmax was 0.62 µM. Conclusions: Satraplatin penetration into CSF is similar to that of carboplatin and cisplatin, despite its greater lipophilicity. The development of a phase I trial of satraplatin for refractory childhood solid tumors including brain tumors is in progress.

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 3599.
Abstract 360: Tamoxifen treatment induces mitogen-activated protein kinase phosphatase-1 expression in T47D breast cancer cells
Authors: Haagenson Kelly K. , Peng Dongjun , Wu Gen Sheng
Published: February 2011

Mitogen Activated Protein Kinase Phosphatase-1 (MKP-1) is a dual specificity phosphatase known to target the Extracellular Signal Regulated Kinases (ERK), c-Jun N-terminal Kinases (JNK) and the p38 Mitogen Activated Protein Kinases (p38), which make up the Mitogen Activated Protein Kinase (MAP Kinase) family. These pathways play vital roles in determining whether cells continue to survive or undergo apoptosis. ERK mainly contributes to cell survival while JNK and p38 signaling can be involved in both cell survival and apoptosis. The function of MKP-1 is to attenuate the signals perpetuated by ERK, JNK and p38. Previous research in our lab has shown that knockdown of either MKP-1 or ERK-2 leads to increased cisplatin sensitivity in ovarian cancer. ERK activation has also been linked to tamoxifen resistance and poor prognosis in breast cancer, but the effects of MKP-1, JNK and p38 in this area are not well studied. A previous study by Cui et al has shown that overexpression of MKP-3, which mainly targets ERK, increased tamoxifen resistance in breast tumors. We chose to focus this investigation on the contribution of MKP-1 to tamoxifen resistance because of its ability to dephosphorylate all three MAP Kinase family members. In our study, MKP-1 protein expression was shown to be elevated in MDA-MB-231, T47D and MCF7 breast cancer cells as compared to non-malignant MCF10A cells. After four and six hours of treatment with 1 μM tamoxifen, MKP-1 protein expression was increased in T47D cells compared to untreated parental control cells. Very little change in phosphorylated ERK protein levels was observed at these same time points. There were, however, decreases in phosphorylated JNK protein levels after four and six hours of 1μM tamoxifen treatment when compared to untreated parental control cells. Since some chemotherapeutic agents induce apoptosis in a JNK mediated manner, a decrease in JNK expression in tamoxifen treated breast cancer cells could contribute to decreased therapeutic efficacy. In conclusion, by examining changes in protein expression levels in tamoxifen treated breast cancer cells we have identified that increased MKP-1 expression leads to a decrease in phosphorylated JNK expression. This finding could constitute a mechanism contributing to tamoxifen resistance in estrogen receptor positive breast cancer cells and suggests that modulation of MKP-1 expression could increase tamoxifen sensitivity in these 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 360.
Abstract 3603: Endoxifen pharmacokinetics and bioavailability in female mice
Authors: Ames Matthew M. , Reid Joel M. , Buhrow Sarah A. , Walden Chad A. , Safgren Stephanie L. , Goetz Matthew P.
Published: February 2011

In collaboration with NCI, we are developing endoxifen (END), the active metabolite of tamoxifen (TAM), as a drug for the treatment of estrogen receptor positive breast cancer. Tamoxifen treated women with reduced or absent CYP2D6 enzyme activity have significantly lower END plasma concentrations. In a retrospective analysis of two different prospective tamoxifen trials (NCCTG and ABCSG), we have demonstrated that genetic or drug-induced reductions in CYP2D6 metabolism are associated with a higher risk of breast cancer recurrence in Tam treated ER positive breast cancer. To test the hypothesis that clinically relevant concentrations and exposures of END are achieved following oral administration of END, we characterized the pharmacokinetics of i.v. and oral END and oral TAM in female mice. Endoxifen was synthesized and provided by NCI/DTP. Plasma samples were analyzed using a validated reverse-phase HPLC method with fluorescence detection. Following i.v. administration of a 1 mg/kg dose, peak plasma concentration, terminal elimination half-life and plasma clearance values were 0.26 µM, 6.5 h and 11.8 L/h/kg, respectively. Oral pharmacokinetics and bioavailability were determined following doses of 50 and 200 mg/kg. Peak plasma concentrations of 0.76 µM were achieved with a 50 mg/kg dose and plasma concentrations above 0.1 µM were maintained for longer than 8 h. Peak plasma concentrations of 8 µM were achieved with a 200 mg/kg dose and plasma concentrations above 2 µM were maintained for 24 h. Bioavailability was 50% and greater than 100% following the 50 mg/kg and 200 mg/kg doses, respectively. Based on pharmacokinetic data for oral END and TAM normalized to a 50 mg/kg oral dose, END plasma concentrations and total exposure were 20-fold greater following oral END as compared to oral TAM. In conclusion, END has high oral bioavailability in mice and substantial plasma concentrations are achieved and maintained after a single oral END dose. Compared to an equivalent oral dose of Tam, substantially higher END concentrations are achieved following oral END. These data suggest that the primary administration of End may overcome the limitations related to human CYP2D6 metabolism, and support the ongoing development of endoxifen as a primary therapy for ER breast cancer. Supported by the Mayo Comprehensive Cancer Center Grant (CA15083) and the Mayo Clinic Breast Cancer SPORE (CA 116201; MMA, JMR, MPG).

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 3603.
Abstract 3606: Gene and miRNA expression profiles in lung cancer cells cooperated with drug sensitivity to PKC inhibitor enzastaurin
Authors: Masahiro Seike , Okano Tetsuya , Shimokawa Tsuneo , Gemma Akihiko
Published: February 2011

PKC-beta inhibitor, Enzastaurin, is being evaluated in a Phase III trial as a maintenance therapy for the treatment of diffuse large B-cell lymphoma (DLBCL), as well as being evaluated in several Phase II studies across a variety of more common tumor types including: breast, colon, lung, ovarian and prostate cancers. Sequence-dependent, synergistic antiproliferative and proapoptotic effects of the combination of cytotoxic drugs and enzastaurin in non small cell lung cancer cells (NSCLC). To ascertain the potential for PKC-beta inhibitor-based treatment in NSCLC, we analyzed the antitumor effects of enzastaurin in 22 NSCLC cell lines via MTT assay. To identify a molecular model of sensitivity to PKC-beta inhibitor treatment in NSCLC, we conducted a gene and miRNA expression profiling study using cDNA arrays and Taqman MicroRNA Arrays on the same set of cell lines and related the cytotoxic activity of enzastaurin to corresponding gene and miRNA expression pattern. In addition, pathway analysis was done with Pathway Architect software. We used eight genes, which were identified by gene-drug sensitivity correlation and pathway analysis, to build a support vector machine algorithm model by which sensitive cell lines were distinguished from resistant cell lines. We also identified unique miRNA expression patterns correlated with enzastaurin sensitivity. Our results suggested that PKCβ inhibitor may be promising anticancer drugs to NSCLC and the eight-gene classifier is useful in predicting drug sensitivity to PKC-beta inhibitors and may contribute to achieving individualized therapy for NSCLC patients.

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 3606.
Abstract 3608: PTEN mutation associated with EGFR TKI de novo resistance
Authors: Kim Youngwook , Ko Jeonghun , Cui Zheng Yun , Ahn Myung Ju , Park Keunchil
Published: February 2011

Epidermal growth factor receptor (EGFR) is overexpressed in 60-80% of non-small cell lung cancer patients. Recent identification that EGFR-tyrosine kinase inhibitors (EGFR-TKI), such as erlotinib and gefitinib, has marked therapeutic effect in a subset of lung cancer patients has prompted extensive clinical trials, which revealed strong association between EGFR-TKI responder groups and the presence of activating mutations in the tyrosine kinase domain of EGFR. Molecular biomarkers used in predicting EGFR TKI responsiveness, however, can vary depending on individual circumstances. Albeit small number of patients, a subset of lung cancer patients does not respond to gefitinib or erlotinb de novo, despite the presence of EGFR activating mutations. The molecular basis of the de novo resistance to EGFR-TKI in EGFR mutant remains largely unidentified.

Here we report that EGFR mutated H1650 cells harbor PTEN deletion mutation, which confers EGFR mutant lung cancer cells with EGFR-TKI resistance. We show that erlotinib in combination with dual PI3K/mTOR inhibitor BEZ235 inhibits the cell viability of H1650 cells in vitro, consistent with a recent report by Sos et al. that showed PTEN etiology in EGFR-TKI de novo resistance in H1650 cells. Combination of erlotinib and PI3K/mTOR inhibitors in xenograft animal model studies. Unexpectedly, we also show that tumor growth of H1650 cells can be proprely controlled in xenograft in vivo animal model with erlotinib alone, raising the possibility that patients with PTEN deletion in the context of EGFR dependent lung cancer can still benefit from EGFR-TKI therapy.

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 3608.
Abstract 3612: Potent preclinical anti-tumor activity of MGCD265, an orally active Met/VEGFR multitargeted kinase inhibitor in Phase II clinical development, in combination with an EGFR inhibitor
Authors: Fournel Marielle , Dupont Isabelle , Bonfils Claire , Dubay Marja , Ste-Croix Hélène , Beaulieu Carole , Beaulieu Normand , Lemoyne Christian , Wang James , Isakovic Lubo , Claridge Stephen , Saavedra Oscar , Raeppel Franck , Raeppel Stephane , Mannion Michael , Vaisburg Arkadii , Martell Robert E. , Besterman Jeffrey M. , Maroun Christiane R.
Published: February 2011

MGCD265 is an oral multitargeted receptor tyrosine kinase inhibitor in Phase II clinical development. MGCD265 targets the Met receptor tyrosine kinase and blocks Met activities which contribute to cancer development and progression such as cell proliferation, motility/invasion, angiogenesis and tumor cell survival. Although there is a large body of literature supporting Met activities as key to epithelial-mesenchymal transition and tumorigenesis, it is now accepted that the coordination of Met signaling with other regulators is central to oncogenesis. Met engages in cross-talks with several membrane proteins including the EGFR. Met and EGFR are coexpressed on tumor cells and functionally cooperate to amplify activating signals. Moreover, in NSCLC, Met gene amplification or overexpression of HGF, has been identified as a major molecular mechanism through which tumors evade EGFR inhibition by specific inhibitors such as gefitinib and erlotinib. Taken together, these studies provide a compelling rationale for concomitantly inhibiting Met and EGFR. In the present study, we have analyzed the in vivo anti-tumor activity of MGCD265 in combination with an EGFR-specific inhibitor, erlotinib. We demonstrate that this combination achieves greater antitumor responses than treatment with either agent alone, in multiple xenografts including NSCLC models. This is observed in the absence of overt toxicity. Furthermore, the PK of MGCD265 and erlotinib was analyzed when the two agents were co-administered, and indicated that there are no drug-drug interactions. In addition, in a NSCLC xenograft model that expresses an EGFR mutant resistant to erlotinib (T790M), MGCD265 significantly improved the anti-tumor activity when combined with erlotinib. The mechanisms underlying efficient tumor growth inhibition by this combination are presently under investigation. Preliminary results suggest that in addition to downregulating Met activation, MGCD265 may modulate EGFR activation by inhibiting the expression of the EGFR ligands TGFα and EREG. These preclinical studies provide support for the clinical development of MGCD265 in combination with erlotinib.

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 3612.