Ehrlich II –2nd World Conference on Magic Bullets

Conclusions: These results provide a compelling evidence for the role of isoproterenol and insulin in regulating Mg2+ content within mammalian tissues. Together with results provided by other laboratories they highligth a key role of Mg2+ in regulating various cellular funcitons under physiological conditions.

Computed Tomographic Study Of The Paranasal Sinuses And Nasal Washings In Atopic Children Without Sinusitis Symptoms ROSARIO NA; KOVALHUK LC; CARVALHO A; CALLEFE LG Federal University of Parana, Curitiba – Brazil Background: the aim of this study is to correlate the extent of sinus disease with eosinophilia and inflammatory cells in nasal fluid (NF). Methods: We studied 48 atopic children with allergic rhinitis (33 asthmatics); 13 nonatopic children served as a control group. Neither patients nor controls had sinusitis symptoms. Coronal computed tomographies (CT) were graded following a standard protocol. A sinus CT score  12 indicated extensive disease. Nasal wash cytology (NW) expressed as a number of cells/mL of volume recovered, was followed by differential counts. Results: 9/48 patients (19%) had extensive disease, 7 were asthmatics and 2 had only rhinitis; 39/48 and 13 controls had a CT score < 12. Atopics had significant eosinophilia in NF and peripheral blood (Table). Total cell counts in NL were higher in atopics than in the control group. The differential cell counts in NW were similar in atopics regardless the CT score. There was correlation between CT score, peripheral eosinophil and NL eosinophil counts in atopics. Group CT score n Blood eosinophil Cells/mL‡ Eosinophils Neutrophils Epithelial Atopic 12 9 665 1305 757 137 137 <12 39 558 578 208 86 109 Control <12 13 148 390¥ 6¥ 167 113 *Mean §Nasal Fluid ‡Cell counts x 10^3/mL ¥ Mann-Whitney U Test p<0.05 in atopics (I,II) X Control (III) group Conclusion: Extensive sinusal disease is frequent in atopics without sinusitis symptoms (19%). Cytology of NF did not identify these patients among atopics. Eosinophils may be involved in extensive sinus mucosa inflammatory changes.
New Chemical Entities For The Treatment Of Estrogen Receptor Negative Breast Cancer: In Vitro Mechanisms Of Action SOMERS-EDGAR, TJ^1, SCANDLYN MJ^1, LARSEN L^2, ROSENGREN RJ^1 ^1University of Otago, Dunedin, New Zealand. ^2New Zealand Institute for Crop and Food Research Ltd., Dunedin, New Zealand Background: Previously, we showed that combination therapy with natural compounds such as curcumin and epigallocatechin gallate significantly suppressed estrogen receptor (ER)-negative tumor growth in vivo. Therefore, we synthesized 8 novel compounds with the aim to increase efficacy and bioavailability. These compounds were screened for their ability to elicit cytotoxicity, apoptosis and alter cell cycle progression in ER-negative breast cancer cells. The most potent compounds were also examined for their oral bioavailability. Methods: MDA-MB-231 and SkBr3 human breast cancer cells were incubated with new chemical entities (0 – 25 µM) for 5 days and cell number was determined using the sulforhodamine B assay. Cell cycle distribution was assessed using propidium iodide staining. The presence of apoptosis was assessed using Annexin-V-FLUOS/propidium iodide staining. The proportion of cells in each of G0/G1-, S- and G2/M-phases as well as the proportion of apoptotic cells (as a percent of total cells) was determined using a flow cytometer. All data are expressed as the mean ± SEM from 4 independent determinations performed in triplicate. Statistical significance was determined using an ANOVA and a Student Newman-Keuls post hoc test, where p<0.05 was required for a statistically significant difference. Results: Three compounds (RL75, RL84 and RL86) were less potent than curcumin. RL90, RL91 and RL92 had the highest cytotoxic potential in both MDA-MB-231 and SkBr3 cells with IC50s of ~1 µM and 0.5 µM, respectively. RL91 (2 µM) induced apoptosis in 40% of cells from both cell lines. RL92 (4 µM) elicited a 2.5-fold increase in the number of cells in G2 and a 60% decrease in G0/G1 phase cells. RL92 also produced a peak plasma concentration of 551 ng/ml 30 min after an oral dose of 8.5 mg/kg in female mice, while RL90 and RL91 were at the limit of detection. Conclusions: 1) RL90, RL91 and RL92 elicited the greatest cytotoxicity in both ER-negative cell lines. 2) RL91 elicited the greatest apoptotic response. 3) RL92 elicited the greatest degree of G0/G1 arrest, mirroring its increase in G2 phase cells and was orally available in female mice. 4) Currently, RL92 is our best drug candidate and is undergoing further testing in mouse models of ER-negative breast cancer.	Personalized Management Of Breast Cancer ROUKOS DH Surgical Oncology Research Unit, Department of Surgery, Ioannina University School of Medicine, Ioannina, Greece Basic science data provide major promise to achieve the goal of personalized medicine in clinical practice. If this goal will be achieved million of people worldwide will benefit. But multiple hurdles and challenges should be overcome and many scientists are skeptical whether this revolution in medicine is realistic or elusive. Breast cancer managements represents one of cancer types with faster advances toward personalized prevention and treatment. Genetic BRCA testing allows effective both prevention and appropriate local control of healthy individual high-risk women with breast cancer family history and patients with inherited BRCA mutations respectively. Current standard targeted agents tamoxifen or aromatase inhibitors and trastuzumab are tailored only to patients with hormone receptor-positive and HER2-positive tumors respectively. Here, we provide for the first time two comprehensive models for personalized prevention of BRCA mutation carriers and a surgery-guided algorithm for patients with familial or sporadic breast cancer.
Cannabis And Cannabinoids: The Forgotten Magic Bullet? ROUSSEAUX CG University of Ottawa, Ottawa, Canada Abstract: Cannabis sp. is a ubiquitous dioecious plant genus that grows wild in most countries. This plant, and its 66 pharmacologically active cannabinoids (CB), has been used for industrial purposes for at least 10,000 years and medically for more than 2,700 years. Historically, wars have focused upon the need for industrial hemp by seafaring nations. For example, Napoleons attempted to block passage of hemp from Russia to England in 1807. Because of the legal and political nature of the psychoactive compound, tetrahydrocannabinol (THC), development of CB based pharmaceutical interventions have provided the medical community with only nabilone (Cesamet^®) and dronabinol (Marinol^®). These synthesized CBs are used for treatment of cancer related symptoms such as nausia and anorexia. Current research has revealed that humans, and other mammals, have an intimate relationship with CBs as there is an endogenous CB system distributed throughout most organs of the body. Two endogenous CBs, anandamide and 2-arachnoylglycerol (2-AG), and exogenous CBs have been implicated in a number of disease processes including Alzheimer’s disease, amyotrophic lateral sclerosis, Tourette’s syndrome, diabetes, dystonia, fibromalgia, gastrointestinal disease, gliomas, hepatitis C, human immunodeficiency virus related symptoms, hypertension, incontinence, multiple sclerosis, osteoporosis, rheumatoid arthritis and sleep apnea, via the CB receptors CB1 and CB2. More recently, CB receptors have been shown to interact with a number of neurotransmitters, such as the opiate and glutamate systems. Generally, CBs dampen the excitatory effects of other excitatory neurotransmitters in the central nervous system; hence, the development of the novel synthetic CB for appetite control, e.g., rimonabant. Conclusions: Once past the political hyperbole regarding the pros and cons of medical marijuana, research may indeed reveal that the CB system is one of Professor Ehrlich’s “magic bullets.”	Antibacterial Activity Of Glucosamine Sulfate And Chondroitine Sulfate? ROZIN AP^1, GOLDSTEIN M^2, SPRECHER H^2 ^1B. Shine Department of Rheumatology, ^2Department of Microbiology, Rambam Health Care Campus and Technion, Haifa, Israel Background: Glucosamine sulfate (GS) and chondroitine sulfate (CS) were shown to delay X-Ray progression of OA. Glucosamine chloride along with CS was shown to reduce symptoms of moderate-severe painful knee OA. GS and CS supplements declined 5-year operative risk after its discontinuation. The mechanism of action of this therapy is still unknown. Recent data demonstrating the efficacy of co-trimoxazole administered as prophylaxis for urinary tract infections in relieving symptoms of patients with knee OA have raised the possibility of participation of the fecal flora in pathogenesis of OA [1]. Expression of Toll-like receptors 2 and 4 [lipopolysaccharide(LPS)-binding] has recently been found up-regulated in lesion areas of OA cartilage [2]. Innate production of tumor necrosis factor-alpha and interleukin-10 upon LPS-stimulation has been associated with radiological progression of knee osteoarthritis [3]. Objectives: To examine the antibacterial activity of GS, CS separately and both in one solution and as a trademark compound Megagluflex on E. coli growth in vitro. Methods: Working solutions of GS-CS [Megagluflex (MGF), “American Health”, NY)] in concentration ranging from 40mcg/ml to 100mg/ml and GS (Sigma), CS (Sigma) (1mg/ml, 50mg/ml) were prepared in normal saline as dissolvent. Inoculums of up to 104 /ml E. coli (strain ATCC 25922) was prepared in serum supplemented Brain heart growth media. One ml of the inoculum solution was mixed with 0.2 ml of different concentration of GS-CS, GS, CS solutions or with control normal saline and incubated in at 37°C for 16 hours. Number of colonies was counted. pH testing was performed for every mixture. We examined antibacterial properties of solutions with components of MGF: vitamine C and MnSO4 (0.166mg/ml, Riedel-De Haen) and solutions with similar pH (5.0) (HCL diluted by normal saline), osmolality (933mOsm/kg) (glucose solution with normal saline) like that of the tested MGF solution. Experiments with 3 repeatable results were taken for consideration. Results: MGF inhibited E. coli growth significantly (p=0.001) in MIC of 1mg/ml and higher. Close to expired time of the drug antibacterial activity declined and persisted at concentration of 100 mg/ml. Solutions of GS and CS separately and in one solution mildly inhibited E. coli growth in one of 5 experiments only in concentration of 50mg/ml. Solutions of vitamine C and manganese sulfate, the components of Megagluflex, and control media, such as control solutions with pH and osmolality like that of MGF solution did not affect E.coli growth. MGF, GS, CS solutions were negative for bacterial contamination. Conclusion: Our data suggest that MGF trademark compound has certain antibacterial activity against E. coli in vitro. Sustained antibacterial activity of GS, CS of another manufacturer in a separate and one solution was not found. Further trials are needed to clarify the antibacterial activity of GC. References: 1.ROZIN AP, et al: OARSI response criteria in assessment of co-trimoxazole influence on refractory knee osteoarthritis during prophylaxis of recurrent UTI. EULAR-Berlin 2004, Ann Rheum Dis 2004; Sup.1, 63:358. 2.KIM HA, et al.: The catabolic pathway mediated by Toll-like receptors in human osteoarthritic chondrocytes. Arthritis Rheum 2006; 54: 2152-2163. 3.BOTHA-SCHEEPERS S, et al.: Innate production of tumor necrosis factor-alpha and interleukin-10 upon lypopolysaccharide stimulation are associated with radiological progression of knee osteoarthritis. ACR 2006, 266.
Genetic Polymorphisms Of Cytochromes P450 Influence The Leflunomide Treatment And Toxicity In Rheumatoid Arthritis Patients ROZMAN B^1, LOGAR D^1, TOMŠIČ M^1,PRAPROTNIK S^1, ŠUPUT D^1, GRABAR I^2, MRHAR A^2, BOHANEC GRABAR P^3, DOLŽAN V^3 ^1Department of Rheumatology, University Medical Centre Ljubljana, Vodnikova 62, Ljubljana, Slovenia, ^2Faculty of Pharmacy, University of Ljubljana, Aškerčeva 7, Ljubljana, Slovenia, ^3 Institute of Biochemistry, Faculty of Medicine, University of Ljubljana, Vrazov trg 2, Ljubljana, Slovenia Background: Leflunomide is a disease-modifying antirheumatic drug that is used for the treatment of rheumatoid arthritis (RA). Upon oral absorption, it is converted to the active metabolite (A77 1726) that inhibits de novo synthesis of pyrimidine ribonucleotides. In vitro studies have demonstrated that cytochromes P450 (CYPs), mainly CYP1A2 and CYP2C19 may be involved in leflunomide activation. The hypothesis of our study was that genetic polymorphisms of CYP1A2 and CYP2C19 influence the A77 1726 serum concentrations and the response of leflunomide treatment in RA patients. Methods: A genotyping approach was used to determine CYP1A2 -163C>A, CYP2C19*2 and CYP2C19*17 genotypes in 112 RA patients. Trough steady-state A77 1726 serum concentrations were determined by validated HPLC with UV detection in all patients on leflunomide treatment. Results: The leflunomide treatment was well tolerated by 62 patients, while 50 patients discontinued the treatment within the first year due to inefficacy (N= 7), toxicity (N= 37) or both (N= 6). CYP2C19 genotypes did not inluence the treatment response (P= 0.258), while an association between CYP1A2 C-163A polymorphism and leflunomide toxicity was observed. Patients with the CYP1A2 -163CC genotype had a 11.9-fold higher risk for leflunomide-induced toxicity as compared to carriers of the CYP1A2 -163A allele (P= 0.001, OR= 11.933, 95% CI= 2.793-50.980). On the other hand, mean values of A77 1726 serum concentrations were not significantly different between CYP1A2 -163CC and CYP1A2 -163CA+AA genotypes (43.9± 39.5mg/L and 33.7± 28.9 mg/L, respectively). Nevertheless, carriers of the CYP2C19*2 allele had significantly lower mean values of A77 1726 serum concentrations as compared to patients with CYP2C19*1/*1 genotype (18.4± 12.8 mg/L vs. 43.7± 33.7 mg/L, P= 0.005). Conclusion: Our results suggest that the CYP1A2 -163C>A polymorphism influenced the leflunomide toxicity, while genetic polymorphisms of CYP2C19 had an impact on the A77 1726 serum concentrations in RA patients.	Dissemination And Communication: "Selling" Vaccines To Peers And The General Public RUNDBLAD G King’s College London, London, United Kingdom Background: In a society that relies on herd immunity as a health protection measure, informing peers and the general public about immunisation can be precarious without access and knowledge of effective communication strategies. Aims: 1) To compare immunisation discourses on two vaccines: MMR and DTP. 2) To compare peer versus general public directed discourse. Focus include both descriptions of vaccines and the diseases they target. Special attention will be given to controversies. Methods: Discourses analysed include: medical research articles in the British Medical Journal and the Lancet, British newspaper articles, and British websites maintained by the National Health Service or by private organisations. Discourses were coded manually. Cognitive Discourse Analysis was used and both qualitative and quantitative measures were obtained. Results: Vaccine discourses were found to be very similar. The main differences lie in the target audience and in the purpose of the discourse. Medical research articles instigating controversies differ significantly in several respects (e.g. vague language, “name dropping” and high variability of titles for professionals) from typical ones. Patterns of “over-compensation” were found in both lay and professional discouses. The concept of herd immunity is very strikingly present even in professionals’ discourse aimed at the general public, whereas peer-to-peer discourse empowers the reader in the discourse. Conclusions: 1) Proponents, opponents and reporters on vaccines employ discourse as a key tool. 2) Different sources and different target audiences yield different discourses, with both intended and unintended drastic effects as a result.
Magic Bullets With Multiple Warheads: Multi-Targeted Antineoplastic Agents RUSSU WA, SHALLAL HM School of Pharmacy, University of the Pacific, Stockton, CA, USA Background: While extensive work in the development of selective multi-targeted kinase inhibitors has been ongoing the design of multi-targeted antineoplastic agents that interact with target proteins of different functional families is much rarer. Multi-targeted antineoplastic agents are anticipated to have the efficacious advantages of drug combinations embodied in one molecule while avoiding the time, cost, pharmacokinetic and other disadvantages of developing and using drug combinations. Here we describe employment of a cross-docking approach to the design of molecules with antiproliferative activity that can potentially interact with both Bcl-XL and HDACs. Methods: We employed structure based design taking advantage of available protein/ligand complex structural data (RCSB), and docking software (Autodock 3.0.5 and Fred). Target molecules were synthesized and characterized by NMR, MS and HPLC. Cell-line antiproliferation assay was performed for a 48 or 72 hour duration and assessed using sulforhodamine B. Results: The initial set of 36 molecules was designed to bind and inhibit the anti-apoptosis Bcl-2 family member Bcl-XL. Molecular docking and synthetic tractability guided the design process. Molecules were successfully synthesized and characterized. Two molecules in this initial set exhibited near or greater than 50% growth inhibition in leukemia, breast, non small cell lung cancer (NSCLC) and renal cell lines at 10 μM concentration. Structure activity relationship (SAR) revealed a key hydrogen bond donor group necessary for significant activity. The active molecules were then cross-docked against other protein targets relevant to cancer. Histone deacetylase (HDAC) was identified as a potential secondary target. A second set of 6 molecules were design that replaced the key hydrogen bond donor group of the initially identified active molecules with an hydroxamic acid group. These newly designed molecules were successfully synthesized and characterized. These displayed 2 fold greater potency. Conclusions: 1) Multi-targeted antineoplastic agents can be designed by utilizing a cross-docking approach of known active compounds against other potential protein targets. 2) Cross-docking results may be exploited by employing traditional medicinal chemistry techniques of lead optimization.	Some New Derivatives Of Vindoline, Monoindole Catharanthus Alkaloid RUSZKOWSKA J, CHROBAK R, ŻERO P, STUDZIAN M, CZARNOCKI Z Laboratory of Natural Products, Faculty of Chemistry, Warsaw University, Warszawa, PL Vindoline and catharanthine, monoindole alkaloids from Catharanthus roseus (L.) G. Don, are valuable precursors of oncolytic drugs, vinblastine and vincristine, used in treatment for some types of cancer. Due to the availability of Polonovsky-Potier and some kinds of oxidation reactions resulting in coupling of monomeric alkaloids into bisindole ones, the search for semi-synthetic analogues of natural precursors is still worth of interest. In this communication five new vindoline derivatives obtained by means of Suzuki-Miyaura reaction, some new (hetero)dimers and other vindoline oxidation products as well as results of their preliminary cytotoxicity tests are reported.
Cationic Liposomes:How A Magic Bullet Turns Into An Anti-Inflammatory Agent LONEZ C, VANDENBRANDEN M, RUYSSCHAERT JM Laboratoire de Structure et Fonctions des Membranes Biologiques, Université Libre de Bruxelles – Brussels, Belgium Background: Cationic lipids are mainly used as efficient DNA, RNA or protein carriers for gene therapy or immunization trials.Significant progress has been made in the understanding of the cellular pathways and mechanisms involved in lipoplex-mediated gene transfection but the interaction of cationic lipids with cell components and the consequences of such an interaction on cell physiology remains poorly described. Methods: RAW 264.7 macrophage cell line was cultured in DMEM (Invitrogen) supplemented by 5%FBS, 1 mM sodium pyruvate, 1 mMglutamine and antibiotics (Invitrogen). Peripheral blood mononuclear cells (PBMC) were isolated from buffy coats (obtained from local routine blood donations) by density centrifugation. Results: Interestingly, diC14-amidine liposomes stimulate, like LPS, myeloid dendritic cells through Toll-like receptor 4 activation . These findings suggest that cationic liposomes alone are potent immunostimulatory adjuvants that could stimulate efficiently the immune system and open the way to the use of cationic lipids as unique adjuvant components of vaccine.On the other hand, diC14-amidine liposomes inhibit CpG sequences or lipopolysaccharides- induced cytokine secretion by macrophages. This inhibitory effect was also mediated by the phospholipidic fraction of lipoproteins and synthetic phospholipids suggesting that cationic liposomes confer to these phospholipids new anti-inflammatory properties.