Combination Of Anticancer Drugs Research Articles

Fucoxanthin Enhances Cisplatin-Induced Cytotoxicity via NFκB-Mediated Pathway and Downregulates DNA Repair Gene Expression in Human Hepatoma HepG2 Cells

Cisplain, a platinum-containing anticancer drug, has been shown to enhance DNA repair and to inhibit cell apoptosis, leading to drug resistance. Thus, the combination of anticancer drugs with nutritional factors is a potential strategy for improving the efficacy of cisplatin chemotherapy. In this study, we investigated the anti-proliferative effects of a combination of fucoxanthin, the major non-provitamin A carotenoid found in Undaria Pinnatifida, and cisplatin in human hepatoma HepG2 cells. We found that fucoxanthin (1–10 μΜ) pretreatment for 24 h followed by cisplatin (10 μΜ) for 24 h significantly decreased cell proliferation, as compared with cisplatin treatment alone. Mechanistically, we showed that fucoxanthin attenuated cisplatin-induced NFκB expression and enhanced the NFκB-regulated Bax/Bcl-2 mRNA ratio. Cisplatin alone induced mRNA expression of excision repair cross complementation 1 (ERCC1) and thymidine phosphorylase (TP) through phosphorylation of ERK, p38 and PI3K/AKT pathways. However, fucoxanthin pretreatment significantly attenuated cisplatin-induced ERCC1 and TP mRNA expression, leading to improvement of chemotherapeutic efficacy of cisplatin. The results suggest that a combined treatment with fucoxanthin and cisplatin could lead to a potentially important new therapeutic strategy against human hepatoma cells.

Characteristics of potential drug-related problems among oncology patients

Oncology patients are more at risk for drug related problems because of treatment with (combinations of) anticancer drugs, as they have a higher risk for organ failure or altered metabolism with progression of their disease. The aim of this study was to characterize and to evaluate the frequency of potential drug related problems (pDRPs) among oncology patients. Outpatient- and day-care centres for Internal and Pulmonary Medicine at the Deventer Hospital, Deventer, The Netherlands. A prospective, descriptive, observational study was carried out from March 2010 to March 2011 at the Deventer Hospital, Deventer, The Netherlands. All patients older than 18 years receiving anticancer drugs prescribed by an internal medicineoncologist or pulmonologist-oncologist were included. The primary outcome was the number and type of pDRPs according to Dutch guidelines. Among 546 patients with cancer, 952 pDRPs were identified, of which 474 were oncology-related. These were mainly drug interactions (IA) (246 IA in 157 patients) and potential contraindications (pCI) (201 pCI in 143 patients). Most identified pDRPs in cancer patients were IAs and pCIs and involved corticosteroids. The most frequently occurring oncology-related IAs were classified as minor or moderate levels of severity.

Apoptosis induction and inhibition of drug resistant tumor growth in vivo involving daunorubicin-loaded graphene–gold composites

Daunorubicin (DNR) loaded graphene-gold nanocomposites offer a novel strategy for inducing apoptosis in drug resistant leukemia cells (K562/A02; KA). In vitro and in vivo investigations on xenografted tumors in KA nude mice demonstrate that the combination of monoclonal P-glycoprotein (P-gp) antibodies and DNR anticancer drug loaded on graphene-gold nanocomposites (GGN) is an efficient drug delivery vector, with remarkable targeting and binding properties towards drug resistant KA cell lines, and induces apoptosis of KA cells and inhibits tumor growth in KA nude mice. Cellular treatment with DNR-loaded GGN remarkably reduced drug resistant-related P-gp expression and activated apoptosis-related caspase protein expression in KA cells. Cell apoptosis provoked in vitro by such nanocomposites corresponds to a rapid induction of active caspase 8,3 activities and stimulation of poly-(ADP-ribose) polymerase (PARP) proteolytic cleavage. In vivo studies indicate that DNR-loaded GGN nanocomposites effectively overcome the inhibition of drug resistant leukemia cell-induced tumor growth in KA nude mice. This nanocomposite raises the possibility of modulating apoptosis in cancer cells, and of inhibiting tumor growth, showing that nanocomposites of this kind have promising applications in efficient multifunctional therapy.

Everolimus combined with cisplatin has a potential role in treatment of urothelial bladder cancer

Cisplatin (CDDP)-based chemotherapy is a commonly treatment for advanced urothelial carcinoma. However, episodes of cisplatin resistance have been referenced. Recently it has been reported that everolimus (RAD001) could have an important role to play in bladder-cancer treatment and that mTOR inhibitors may restore chemosensitivity in resistant tumours. The aim of this study was to assess RAD001 in vitro ability to enhance CDDP cytotoxicity in three human bladder-cancer cell lines. Over the course of 72h, the cells were exposed to different concentrations of CDDP and RAD001, isolated or combined. Treatment with CDDP statistically (P<0.05) decreased cell proliferation in cell lines in a dose-dependent manner. The anti-proliferative activity of CDDP used in combination with RAD001 was statistically significant (P<0.05) in the cell lines at all concentrations tested. RAD001 had a therapeutic effect when used in combination with CDDP and could therefore be a useful anti-cancer drug combination for patients with bladder cancer.

Comparison of Minimal Residual Disease Detection Using High-Throughput Sequencing and Allele-Specific Oligonucleotide PCR Methods in Pediatric B-Lineage Acute Lymphoblastic Leukemia.

AbstractAbstract 2532▪▪This icon denotes a clinically relevant abstract Background:The clinical management of patients with pediatric B-lineage acute lymphoblastic leukemia (B-ALL) relies on combinations of multiagent anticancer drugs and risk-stratified treatment. The prognostic significance of minimal residual detection (MRD) in pediatric B-ALL has been demonstrated in multiple cohorts. Allele-specific oligonucleotide PCR (ASO-PCR) amplification of immunoglobulin or T-cell receptor rearrangements, a method for MRD detection, requires the development of patient-specific reagents and cannot detect clonal evolution. ASO-PCR also has limited coverage, with clonal rearrangements being detected in only 90% of patients. We developed the sequencing-based LymphoSIGHT platform to address these limitations. Here we report the results of a pilot study of MRD detection using both the sequencing assay and ASO-PCR in paired diagnostic and end-of-induction samples from 7 B-ALL patients. Analysis of 82 additional patients is ongoing. Methods:Using universal primer sets, we amplified immunoglobulin heavy chain (IgH@) variable (V), diversity, and joining gene segments from genomic DNA in diagnostic and follow-up bone marrow samples. Amplified products were sequenced to obtain >1 million reads (20× coverage per B-cell), and were analyzed using standardized algorithms for clonotype determination. Tumor-specific clonotypes were identified in the diagnostic sample of each patient based on high-frequency within the B-cell repertoire. The presence of the tumor-specific clonotype was then assessed in the end-of-induction sample. A quantitative and standardized measure of MRD level among all leukocytes in the sample was determined using internal reference DNA. Following identification of IgH clonal rearrangements and MRD assessment using the sequencing assay, we examined the MRD results obtained at Boston Children’s Hospital using ASO-PCR. Among the 7 patients analyzed to date, 6 patients were in complete remission at the time of the second sample; 1 patient had persistent evidence of disease. Sequencing was performed blinded to all clinical and ASO-PCR information on these patients. Results:With the sequencing platform, we detected a high-frequency IgH clonal rearrangement in all 7 diagnostic ALL samples. The leukemic clonotype that was identified at diagnosis was detected in the end-of-induction sample in each of the 7 patients. The quantitative range of the leukemic sequence in MRD samples ranged over 5 orders of magnitude. MRD results were concordant between sequencing and ASO-PCR in 5 of 7 patients. The detected MRD level differed by > 10 fold in 2 patients. In patient 1, sequencing detected high MRD, while low MRD was detected by ASO-PCR; this patient relapsed 1 year later while still on therapy. In patient 6, sequencing detected low MRD, while ASO-PCR detected high MRD. This patient remains in complete remission after 7 years. In patient 7, sequencing and ASO-PCR concordantly detected low MRD; this patient relapsed after completion of therapy. Conclusions:Results from the application of a high-throughput sequencing method for MRD detection in childhood B-ALL are shown. The sequencing assay does not require development of patient-specific reagents, which will reduce cost and laboratory turnaround time. This data, along with the laboratory workflow improvements, support the use of the sequencing assay as a next-generation MRD test for B-ALL. Analysis of samples from 82 patients is ongoing.Table 1Comparison of MRD results using sequencing and ASO-PCR methodsPatientNumber of input cellsNumber of input leukemic clone moleculesSequencing MRD (%)ASO-PCR MRD (%) (High MRD ≥ 0.1%)Clinical outcome1163,33317,1705.30.008Relapse22,043,8609,958,326∼10018.2Induction failure32,052,6512,4640.066.5Alive and well41,298,291670.0010.001Alive and well5984,91810,0360.50.2Alive and well6863,757474,93827.528.1Relapse731,536730.10.03Relapse Disclosures:Faham:Sequenta, Inc.: Employment, Equity Ownership, Research Funding. Fang:Sequenta, Inc.: Employment, Equity Ownership, Research Funding. Moorhead:Sequenta, Inc.: Employment, Equity Ownership, Research Funding. Zheng:Sequenta, Inc.: Employment, Equity Ownership, Research Funding.

Abstract A1: SNAIL inhibitor GN-25 induces MET through Global Reprograming of EMT Network Genes: a Systems and Network Analysis

Abstract Epithelial-to-mesenchymal transition (EMT) is a complex mechanism that arises through a cohesive and intricate biological cross-talk between numerous cancer-related networks. Among them sonic hedgehog directed snail network is one of the major components that drives EMT. This has led to strategies targeting either hedgehog signaling or snail protein using highly specific agents. However, it is needless to say that removal of individual components through single protein-targeted drug (here snail) from robust EMT networks may surprisingly have little functional consequence in cancer. We propose that, to have an impact, interventions within EMT network need to be multiple but highly selective. Further, predicting the functional outcome of interventions, such as those originating from single pathway targeted drug treatment, is not as straightforward as originally thought and computational/integrated tools are needed. The emergence of computational network biology has enhanced our knowledge of multi-pathway interactions in cancer and helped in making sense of drug response signature datasets that collectively decode the complex mechanisms of drug action. These holistic approaches have laid the foundation of the emerging concepts of “network pharmacology” that is solidifying its position in cancer medicine. Unlike earlier reductionist one-drug, one-target approaches, network pharmacology invokes the idea of drug promiscuity often engages a fortuitous combination of appropriate high-value targets in cancer cells to produce treatment success. Network pharmacology is being used in the identification of weak nodes in global cancer networks, predicting drug toxicity, drug repurposing, identifying multiscale mechanisms of drug action and in the rational design of potent anticancer drug combinations. To this end, we had earlier developed a novel snail-p53 interaction inhibitor (GN-25) that was shown to suppress snail function. However, this drug was not investigated for its effects against EMT pathways. Here using network pharmacology drug repurposing principles, we show that our novel SNAIL inhibitor (GN-25), through its promiscuous action induces mesenchymal-to-epithelial transition (MET) in well-recognized HMLE-snail cell line model developed by Dr. Weinberg. Network modeling of microarray gene expression profiling data from GN-25 treated cells showed an orchestrated global reorganization of EMT network genes. Most importantly, the expression signatures were validated both at the mRNA and protein expression levels, and mechanistically studied by RNA interference which provided unequivocal evidence on the role of GN-25 targeting EMT specific genes. This proof-of-concept investigation shows that computational biology and network pharmacology can be very powerful tools for rational design and development of agents that are typically promiscuous like the snail inhibitor as a candidate EMT-targeting drug. Citation Format: Asfar S. Azmi, Ramzi M. Mohammad, Fazlul H. Sarkar. SNAIL inhibitor GN-25 induces MET through Global Reprograming of EMT Network Genes: a Systems and Network Analysis [abstract]. In: Proceedings of the AACR Special Conference on Chemical Systems Biology: Assembling and Interrogating Computational Models of the Cancer Cell by Chemical Perturbations; 2012 Jun 27-30; Boston, MA. Philadelphia (PA): AACR; Cancer Res 2012;72(13 Suppl):Abstract nr A1.

Challenging the effectiveness of green tea in primary and tertiary cancer prevention

Drinking green tea daily is part of Japanese culture, and various studies have revealed that green tea is a cancer preventive. We here review our progress in cancer prevention with green tea on 12 main topics, from basic to clinical level. TOPICS AND METHODS: Biochemical and biological studies of green tea catechins, a prospective cohort study, preclinical safety trials with tablets of green tea extract, double-blind randomized clinical phase II prevention trial for recurrence of colorectal adenomas, and synergistically enhanced inhibition by the combination of green tea catechins and anticancer drugs. All results were significant, including human studies with informed consent. Drinking 10 Japanese-size cups of green tea per day delayed the cancer onset of humans 7 years for females. For tertiary cancer prevention, consuming 10 cups of green tea per day fortified by green tea tablets, 50 %, significantly prevented the recurrence of colorectal adenomas. A minimum effective amount of green tea catechins for cancer prevention was found in humans. In addition, the combination of green tea catechins and anticancer drugs engendered a new cancer therapeutic strategy. The consumption of 10 Japanese-size cups of green tea per day is a significant factor in primary cancer prevention for the general population, and the preventive effect on recurrence of colorectal adenomas in patients is vital evidence in tertiary cancer prevention.

Multifunctional Magnetic Nanoparticles for Synergistic Enhancement of Cancer Treatment by Combinatorial Radio Frequency Thermolysis and Drug Delivery

Few-layer, carbon-coated, iron (C/Fe) magnetic nanoparticles (MNPs) were synthesized with controlled sizes ranging from 7 to 9 nm. The additional loading of two anti-cancer drugs, doxorubicin and erlotinib, was achieved through - stacking onto the carbon shells. Controlled release of the drugs was successfully triggered by radio frequency (RF) heating or pH variation. Based on the experimental results, C/Fe MNPs act as heat-inducing agents and are able to thermally destroy cancer cells when RF is applied. It was found that the combination of anti-cancer drugs (in particular a low dose of doxorubicin) and RF treatment demonstrates a synergistic effect in inducing cell death in pancreatic cancer cells. Our findings demonstrate that MNPs can be used as highly efficient multimodal nanocarrier agents for an integrated approach to cancer treatment involving triggered delivery of antineoplastic drugs and RF-induced thermal therapy.

Green tea: An effective synergist with anticancer drugs for tertiary cancer prevention

Green tea is now an acknowledged cancer preventive in Japan. Based on evidence that colorectal adenomas and prostate cancer in humans have been prevented, we review here the concept that the combination of anticancer drugs with green tea catechin synergistically induces apoptosis of human cancer cells, inhibits tumor formation in mice, and enhances inhibition of tumor growth in xenograft mouse models. As a molecular mechanism by the combination, the induction of growth arrest and DNA damage-inducible 153 (GADD153, CHOP) gene expression is discussed in relation to death receptor 5 and TRAIL-apoptotic pathway. The combination of anticancer drugs with green tea could be a new cancer therapeutic strategy in humans.

Reversion of Multidrug Resistance in a Chemoresistant Human Breast Cancer Cell Line by β-Elemene

Background: Multidrug resistance (MDR) presents a problem in cancer chemotherapy, and developing new agents to overcome MDR is important. This study intends to investigate the reversal effect of β-elemene on MDR in human breast carcinoma MCF-7 and doxorubicin-resistant MCF-7 cells. Methods: MTT cytotoxicity assays, flow cytometry, and Western blot analysis were performed to investigate the antiproliferative effects of the combination of anticancer drugs with β-elemene, to study the reversal of drug resistance, and to examine the inhibitory effects on protein expression. Results: The results showed that β-elemene (30 µmol/l) had a strong potency to increase the cytotoxicity of doxorubicin to MCF-7/DOX cells, with a reversal fold of 6.38. In addition, the mechanisms of β-elemene in reversing P-glycoprotein (P-gp)-mediated MDR demonstrated that β-elemene significantly increases the intracellular accumulations of doxorubicin and Rh123 via inhibition of the P-gp transport function in MCF-7/DOX cells. Flow cytometry and Western blot analyses revealed that β-elemene could inhibit the expression of P-gp, while it had little effect on the expression of MRP1 protein. In addition, β-elemene had little inhibitory effect on the intracellular GSH levels and GST activities in MCF-7/DOX cells. Conclusions: β-Elemene might represent a promising agent for overcoming MDR in cancer therapy.

Tamoxifen in Trimodal Therapy with Cytotoxic Drugs and Hyperthermia in Vivo Significantly Enhance Therapeutic Efficacy against B16-F10 Melanoma

The aim of the study was to investigate whether use of the antiestrogen tamoxifen and heat treatment in combined therapy with the well-known anticancer drugs cisplatin, dacarbazine and cyclophosphamide enhances their therapeutic efficacy on mouse B16-F10 melanoma in vivo. The results of systemic melanoma therapy have been mostly disappointing. Therefore, there is still a great need for strategies that can improve existing chemotherapy options. The tumor model for the investigation of antitumor activity was a mouse B16-F10 melanoma transplanted into the footpad of C57BL/6 Zgr/Hr mice. Drugs were given intraperitoneally 15 min before the application of local hyperthermia, and tumor growth and mouse survival were followed. Hyperthermia alone determined a significant delay of tumor growth, but mouse survival was not affected. In bimodal combinations with hyperthermia, all the tested antitumor drugs significantly increased both tumor growth delay and mouse survival. Tamoxifen alone did not show any inhibitory effect on B16-F10 melanoma in vivo. However, in the trimodal therapy with a particular drug and hyperthermia, it potentiated the inhibitory effects of the respective bimodal treatments, especially that of cyclophosphamide and hyperthermia. Our results obtained on the mouse B16-F10 melanoma in vivo confirmed the enhanced therapeutic efficacy of the trimodal therapy tamoxifen, hyperthermia and anticancer drug combinations in melanoma treatment. Further studies should optimize the heat-drug time scheduling and drug doses that will result in the best possible therapeutic achievement for these trimodal therapy options.

Mechanisms of Apoptosis Induced by Anticancer Compounds in Melanoma Cells

The sensitivity of cancer cells to apoptosis induced by anticancer drugs in vitro may be a predictor of their sensitivity to these drugs in vivo. In this review I summarize recent data describing anticancer drug-induced apoptosis in human melanoma cells. Proteasome inhibitors alone, or in combination with other drugs, efficiently induce apoptosis in melanoma cells. It has been shown that apoptosis induced by proteasome inhibitors is linked to suppression of transcription factor FoxM1 and upregulation of the proapoptotic Noxa protein. In addition, proteasome inhibitors stabilize the antiapoptotic Mcl-1 protein, and its suppression leads to more robust apoptosis in melanoma cells. Drugs targeting B-Raf (BAY 54-9085) or IKKb (BMS-345541) have been tested in melanoma cell lines, and it has been shown that the proapoptotic activity of both drugs depends on the inhibition of NF-kB in melanoma cells. A synthetic analog of dsRNA in complex with a polycation stimulated autophagy via induction of dsRNA helicase MDA-5 followed by apoptosis that was partially modulated by Noxa. These data may provide important information needed for designing more efficient combinations of anticancer drugs against melanoma.

Abstract A36: An anticancer drug combination and regimen database for research and clinical optimization.

Abstract Physiomics and pharmacometrics have collaborated to design a new database of anti-cancer drugs and therapeutic treatment information aimed at researchers in oncology and clinicians. This database, accessible through the web, offers data on more than 130 anti-cancer drugs (small molecules and biologics) used in research and in the clinic. It contains information on drug combination as well as a several hundreds of cancer chemotherapy regimens routinely used in the clinic. These data are classified according to tumor type, species, or source (in vitro, in vivo or simulated). It will be constantly expanded and curated with the most recent information. Furthermore, it provides ways to standardize the expression and nomenclature of chemotherapy regimens unambiguously and uniformly is of paramount importance to improve efficacy, as well as to reduce medication errors. Individual drug information covers pharmacokinetic profiles, mechanism of action and of resistance, dose-response effect, dosing limits, therapeutic index, and immunosuppression data. Drug combinations are also referenced. The database covers synergy or antagonism, as well as a combination therapeutic index and cross-resistance information. Some drug combination having level of synergy depending on the drug schedule, drug sequence and administration timings are also referenced and thoroughly discussed. The user can also browse and compare chemotherapeutic regimens, analyze the overall drug dose over a course of treatment, by tumor type, in animal and clinical models. Advanced functions include the ability to do statistical analysis on drug usage and dosing in various contexts. It can also help determine which drug candidates are likely to be used in combination with a new chemical or biological entity, given the mechanism of action and other PK/PD data. Also, it will allow users to design new combinations and regimens, which obey dosing constraints, such as MLD and MTD. Finally, data can be exported and used in spread sheets, modeling software or simulation packages. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2011 Nov 12-16; San Francisco, CA. Philadelphia (PA): AACR; Mol Cancer Ther 2011;10(11 Suppl):Abstract nr A36.

EGF receptor inhibitors increase ErbB3 mRNA and protein levels in breast cancer cells

The potential benefits of drugs directly targeting the ErbB receptors for cancer therapy have led to an extensive development within this field. However, the clinical effects of ErbB receptor-targeting drugs in cancer treatment are limited due to a high frequency of resistance. It has been reported that, when inhibiting the epidermal growth factor receptor (EGFR) with the tyrosine kinase inhibitor gefitinib, increased activation of ErbB3 via MET, or by re-localization of ErbB3 mediates cell survival. Here we show further evidence that members of the ErbB receptor family facilitate resistance to EGFR inhibitor treatment in ErbB2 overexpressing breast cancer cells. We found that gefitinib treatment increased ErbB3 expression, both at protein and mRNA levels. ErbB3 expression was upregulated not only by gefitinib but also by a panel of different EGFR inhibitors, suggesting that inhibition of EGFR in general affects ErbB3 expression. In addition, we found that gefitinib treatment increased ErbB2 expression levels while EGFR inhibitors decreased the activity of ErbB2. Concentrations of gefitinib that decreased phospho-ErbB2 reversely increased ErbB3 levels. We further examined changes induced by gefitinib treatment on mRNA levels of the most common genes known to be involved in breast cancer. As expected, we found that gefitinib downregulated genes whose functions were linked to cellular proliferation, such as Ki-67, topoisomerase II alpha and cyclins, and surprisingly downregulated gene expression of FAS which is involved in apoptotic signaling. Together, our data strongly suggest that resistance to EGFR inhibitors may result from the compensation of other family members and that combinations of anti-cancer drugs are required to increase the sensitivity of these treatments.

6140 POSTER Cetuximab-mediated Immune-enhancing Effects in Vitro and in Metastatic Colorectal Cancer (mCRC) Patients

6140 POSTER Cetuximab-mediated Immune-enhancing Effects in Vitro and in Metastatic Colorectal Cancer (mCRC) Patients P. Correale1, C. Botta1, M.G. Cusi2, E. Bestoso1, S. Apollinari1, M. Caraglia3, M.T. Del Vecchio4, A. Abbruzzese3, A. Aquino5, P. Tagliaferri6. 1“S. Maria alle Scotte” Siena University Hospital, Department of Oncology, Siena, Italy; 2“S. Maria alle Scotte” Siena University Hospital, Department of Molecular Biology Microbiology Section, Siena, Italy; 3II University of Naples, Department of Biochemistry and Biophysics, Naples, Italy; 4“S. Maria alle Scotte” University Hospital of Siena, Section of Pathological Anatomy Department of Human Pathology and Oncology, Siena, Italy; 5“Tor Vergata” University, Department of Neurosciences Pharmacology Section, Rome, Italy; 6Magna Graecia University and Tommaso Campanella Cancer Center Catanzaro, Medical Oncology, Catanzaro, Italy Introduction: Cetuximab is a chimeric human-murine monoclonal IgG1 to the Epidermal-growth-factor-Receptor, approved for colorectal cancer treatment with chemotherapy. Its human constant fragment (Fc), throughout receptor binding, trigger additional immune-mediated effects, which may offer significant contribute to the final therapeutic effect. We investigated cetuximab ability in vitro to promote colon cancer cell phagocytosis and antigen-cross-priming by dendritic cells (DCs) to tumour-specific cytotoxic- T-cell (CTL) precursors. We also carried-out an immunological study in 26 mCRC patients enrolled in an on-going phase 2 trial, receiving an experimental biochemotherapy (GILFICet) regimen combining gemcitabine, irinotecan, fluoruracil, levofolinate, cetuximab, and metronomic sc. aldesleukine. Material and Methods: Transmission-electron-microscopy (TEM) and Flow cytometry were used to evaluate the susceptibility of multiple colon cancer cell lines to DC-mediated phagocytosis. Human DCs loaded with cetuximab-coated colon cancer cells, exposed or not to a combination of anti-cancer drugs, were used to in vitro sensitize human PBMCs from normal donors and cancer patients collected at baseline and after 3 GILFICet courses. T-cell cultures were characterized for immunephenotype and tumour-antigen specific CTL activity by Flow cytometry, LDH release and IFNg-ELISPOT. Results: ILF (irinotecan+ folinate+5-flurouracil) and GILF (gemcitabine+ ILF) poly-chemotherapies confirmed their ability to induce antigen remodelling and danger signals in colon cancer cells. After exposure to poly-chemotherapy and cetuximab these cells became highly susceptible to Fc-receptor-mediated phagocytosis/trogocytosis by human DCs, promoted their activation, and increased their ability to elicit a highly efficient CTL response on human PBMCs in vitro. Our study on the PBMCs of colon cancer patients enrolled in the GILFICet trial, revealed a significant treatment-related increase in na¨ive and central-memory-Tcells, activated-CTLs, NK/NKTs, mature-activated DCs and IFNg-releasing cells. Substantial differences were observed in T-cell lines generated from patients’ PBMCs taken before and after biochemotherapy. In the latter group there was in fact, a remarkable increase in proliferating CD8+Ki67+CTLs and tumour-antigen specific CTLs’ precursors. Discussion: These results suggest that cetuximab may exert immuneenhancing effects with potential antitumour activity

Synergistic effects of the combination of β-ionone and sorafenib on metastasis of human hepatoma SK-Hep-1 cells

The combination of anti-cancer drugs with nutritional factors is a potential strategy for improving the efficacy of chemotherapy, particularly for hepatocellular carcinoma because its conventional therapies are mostly ineffective. Using a highly invasive hepatoma SK-Hep-1 cell line, we investigated the possible synergistic anti-metastatic efficacy of a combination of sorafenib (SF), a multi-kinase inhibitor, and β-ionone (BI), a precursor of carotenoids. We found that SF (1 μM) in combination with BI (1 μM) synergistically inhibited cell invasion and additively inhibited cell migration, especially at 48 h of incubation. Mechanistically, the combination of SF and BI was found to decrease the protein expression of focal adhesion kinase (FAK) and Rho, and to enhance the protein expression of tissue inhibitor matrix metalloproteinase (TIMP)-1 and TIMP-2. In addition, the combination of SF and BI inhibited the activity of matrix metalloproteinase (MMP)-2 and MMP-9 and decreased the phosphorylation of FAK and of Rac1 proteins. Importantly, SF enhanced the suppressing effect of BI (1-50 μM) on the viability of SK-Hep-1 cells, but not on murine hepatic BNL CL.2 cells, indicating the selective cytotoxicity of this combination on tumor cells. The combination of SF and BI could be a potential therapeutic strategy against human hepatoma cells.

Hair Cell Toxicity in Anti-cancer Drugs: Evaluating an Anti-cancer Drug Library for Independent and Synergistic Toxic Effects on Hair Cells Using the Zebrafish Lateral Line

Inner ear hair cell loss is the most common pathology seen after ototoxic drug injury. While certain drugs such as aminoglycosides and cisplatin are well-known to have dramatic ototoxic effects, it is probable that there are other drugs that cause occult degrees of hair cell loss and lesser degrees of hearing loss. Anti-cancer drugs are particularly strong candidates due to their general cytotoxicity. We have screened a library of 88 anti-cancer drugs (National Cancer Institute Approved Oncology Drugs Set) for drugs that damage hair cells of the zebrafish lateral line. The screen identified four out of five known ototoxic drugs. The screen also identified four out of seven suspected ototoxic drugs (drugs that have isolated case reports of patients developing hearing loss after administration). Five additional drugs with no known ototoxicity were identified as potentially novel ototoxins. Additional dose-response curves were performed to evaluate relative toxicity. Since anti-cancer drugs are often used clinically in combination, we also performed dose-response curves for a variety of anti-cancer drug combinations and demonstrated synergistic toxicity in five out of ten drug combinations that we tested. These findings support the use of the zebrafish lateral line as a screening tool to detect ototoxic effects in drugs and also suggest that ototoxicity should be considered in terms of drug regimens rather than individual drugs.

Abstract 4918: Predicting the effect of drug combination schedules on xenograft growth using the Virtual Tumor

Abstract Since the early 1960s, drug combination therapy has been used to treat cancer, because of the limited number of malignancies that could respond to single-agent chemotherapy. Combination chemotherapy regimens have been designed on the basis of mechanism of action of the drugs, tumor cell specificity, balance between effectiveness and toxicity, and synergy between drugs. But when multiple drugs, combination schedules, sequences and doses are considered, the number of possibilities increases combinatorically, and can not be realistically tested either clinically or in animal models. We have developed a “Virtual Tumor” model to aid with the design of optimal drug schedules. The model combines disparate data, at the cell and tumor level, into a consistent picture, and leverages them to make testable predictions about tumor response. Thousands of simulations can be performed if necessary to find the best treatment regime. We present here a validation study of our Virtual Tumor. We predicted xenograft growth of two anti-cancer drug combinations using experimental data collected from single drug exposure uniquely. We accurately predicted the xenograft course for two different regimens – one simultaneous and one sequential – of the two drugs, which were compared with experimental results in a single-blind test. We show how a computational approach helps explain how multiple drug exposure and correct sequence leads to synergy, and how it can be used to subsequently design optimal schedule and combination treatments. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 4918. doi:10.1158/1538-7445.AM2011-4918

Abstract 2547: NCI-60 combination screening matrix of approved anticancer drugs

Abstract Anticancer drugs are rarely curative as single agents, thus most treatment regimens utilize a combination of agents. Selection of rational combinations based upon presumed mechanisms of action is an active area of research. Nonetheless, treatment regimens may have built up over time by addition of new agents to existing standards of care. In order to establish a framework for extending our understanding combination therapy, we have initiated an additional approach: the systematic testing of all pairwise combinations of FDA-approved anticancer drugs in the NCI-60 panel of human tumor cell lines. About 100 small molecule drugs are approved for cancer treatment worldwide – testing all 5000 pairwise combinations in each of 60 cell lines will require 300,000 experiments. Combinations are tested at 3 or more concentrations of each agent, and single agents are tested on the same plates, requiring ∼ 5 million wells. To date, about 100 pairs of drugs have been tested in the NCI-60 in a pilot phase – the high-throughput phase has just begun. Some combinations of drugs, such as bortezomib combined with cladribine, show better than single agent activity in nearly all cell lines tested. Other combinations, such as doxorubicin plus lapatinib, show more restricted benefit, with some cell lines showing enhanced growth inhibition or cell kill, while in other cell lines the combination is antagonistic or identical to the more active single agent. The NCI-60 panel of cell lines has been extensively molecularly characterized, with publicly available data including gene mutation, DNA copy number, DNA methylation, and expression of mRNA, protein and microRNA. The patterns of cell line sensitivity for a particular drug combination can be used to probe these molecular characterization data, generating hypotheses about potential predictive markers and/or mechanisms of action. Probing the doxorubicin plus lapatinib patterns reveals a strong correlation with MDR activity, with high MDR expressing lines showing enhanced combination activity. The most promising combinations identified from NCI-60 testing are being tested in vivo to determine the therapeutic index. Already 2 combinations have demonstrated promising xenograft activity. The combination screening data, along with tools to analyze the data, will be made available to the public through the NCI web site. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 2547. doi:10.1158/1538-7445.AM2011-2547

Reversal of Multidrug Resistance by Guggulsterone in Drug-Resistant MCF-7 Cell Lines

Background: Multidrug resistance (MDR) presents a serious problem in cancer chemotherapy. Our previous studies have shown that the MDR of K562/DOX cells could be reversed by guggulsterone through inhibiting the function and expression of P-glycoprotein. The purpose of this study was to investigate the reversal effect of guggulsterone on MDR in drug-resistant MCF-7 cells and the parental MCF-7 cells. Methods: MTT cytotoxicity assays, flow cytometry, and Western blot analysis were performed to investigate the antiproliferative effects of the combination of anticancer drugs with guggulsterone, to study the reversal of drug resistance and to examine the inhibitory effects on MRP1 expression. Results: The results showed that co-administration of guggulsterone (10 µM) resulted in a significant increase in chemosensitivity of MCF-7/DOX cells to doxorubicin, compared with doxorubicin treatment alone (p < 0.01). The fold reversal of 10 µM guggulsterone (11.48) was comparable to that of 10 µM verapamil (13.23). Rhodamine123 and doxorubicin accumulation in MCF-7/DOX cells was significantly enhanced after the incubation with guggulsterone (10 µM), compared with untreated MCF-7/DOX cells (p < 0.01). When doxorubicin (10 µM) was combined with guggulsterone (10 µM), the mean apoptotic population of MCF-7/DOX cells was 24.91%. It was increased by 6.15 times, compared with doxorubicin (10 µM) treatment alone. However, guggulsterone had little inhibitory effect on the expression of MRP1 proteins. Conclusion: Guggulsterone is a novel and potent MDR reversal agent with the potential to be an adjunctive agent for tumor chemotherapy.