Ehrlich II –2nd World Conference on Magic Bullets

Results: Increasing knowledge about genes implicated in pharmacokinetics, mode of action, efficacy, and toxicity of drugs has already provided relevant results for clinical practice, for example:

The strong association of the abacavir hypersensitivity reaction with HLA-B*5701 permits testing patients for the allele, and if present avoiding the drug and therefore preventing the reaction.

Persons with the allele CYP2B6*6 present higher efavirenz “area under the curve” and have increased risk of neuropsychological toxicity.

Additional gene variants are being discovered that influence the action of antiretroviral drugs. And, moreover, it is expected that larger-scale comprehensive genome approaches will profoundly improve the landscape of knowledge of HIV therapy in the future.

Conclusions: Pharmacogenetics is called to play a major role in the treatment of HIV infections in the future.

Predicting The Active Doses Of New Anticancer Agents In Humans Directly From Preclinical Data ROCCHETTI M^1, PESENTI E^2, DE NICOLAO G^3 ^1Pharmacokinetics&Modeling, Accelera, Nerviano Medical Sciences, Nerviano (MI), Italy. ^2Pharmacology Department, Oncology, Nerviano Medical Sciences, Nerviano (MI), Italy. ^3Dipartimento di Informatica e Sistemistica, University of Pavia, Pavia, Italy The success rate of clinical drug development is significantly lower in oncology (less than 5%) than in other therapeutic areas, with cost and development time of over 1 billion of dollars and more than 10 years for a drug to reach marketing approval and successful use in clinical practice. Of these drugs, few cases can really be considered Magic Bullets and, very often, their contribution proves crucial only in sub-populations of patients. This situation clearly reflects the specific difficulties encountered in the development of oncology drugs. For example, a large number of candidates fail in the last phase of development, due to insufficient efficacy observed in the clinics. In most cases, this is not due to errors in the prediction of either the mechanism of action or pharmacological activity in humans, but to the onset of toxicity findings that prevent the achievement of the required drug exposure in patients for obtaining clinical activity. These failures in the final stage of development may have disruptive financial impact even on big multinational companies. In view of this, there is an absolute need for more appropriate ranking and selection methods of compounds in the preclinical phase. In this talk, the improvements that can follow from a model-based pharmacokinetic-pharmacodynamic (PK/PD) approach will be illustrated. In addition, a methodology able to quantitatively predict, right from the first animal studies, the dose levels of new compounds expected to have clinical activity in patients will be presented. This estimate, combined with the toxicological, pharmacokinetic and pharmacodynamic properties of the compounds, provides a valuable input to the decision making process in drug development. On this basis, the whole development process of R&D projects may be optimised so that inappropriate compounds are discarded earlier and the real Magic Bullets are more easily recognized.
Pharmacokinetics And Effects On Adenosine Concentration Of A New 4-Anilinoquinazoline Derivative In Rats ROCCO SA^1, MARIN RM^1, THEIZEN TH^1, LIMA J^1, GUIDO, MC^1, FRANCHINI KG^1 ^1State University of Campinas, Campinas, Brazil Background: The purinic nucleoside adenosine (ADO) regulates multiple cell functions. Its effects are mediated by at least four kinds of P1 purinergic receptors (A1, A2a, A2b and A3). Adenosine kinase (AK) is a key intracellular enzyme regulating intra- and extracellular ADO concentrations. Pharmacological targeting of AK can enhance ADO tissue levels. In this study, we examined the pharmacokinetics of a new adenosine kinase inhibitor, the 4-N-(3’-N,N-dimethylphenyl)amino-6,7-dimethoxyquinazoline (DMAQ) and its effects on ADO concentrations in rat tissues. Methods: Male Wistar rats (n= 6 of each experimental group, body weight (BWT): 200-250g) were dosed orally with 30 and 90 mg/Kg BWT of DMAQ or vehicle (Control group – n=6). Tissue samples were collected 1h after treatment and immediately ground to a powder under liquid nitrogen. To ADO analysis it was extracted from tissues, derivatized with 2-chloroacetaldehyde and analysed by high performance liquid chromatography method (HPLC). To pharmacokinetic studies, blood was sampled in tubes containing heparin at control, 15, 30, 60, 120, 180, 240, 300, 360, 420 and 480 minutes. Samples were collected from a catheter previously implanted into the femoral artery. Results: Oral administration of DMAQ (30 mg/Kg) was demonstrated to increase ADO concentration in rat tissues such as heart (43.05.5%), liver (33.41.8%) and brain (24.04.9%). Pharmacokinetic parameters were determined for orally administered DMAQ (90 mg/Kg). The DMAQ maximal concentration in plasma of 22.90 mol L^-1 was reached at 3h after administration and after 7h, DMAQ concentration approached the limit of HPLC quantification. DMAQ showed a rapid absorption profile with an estimated absorption half-life of 13 min. and a moderately fast rate of elimination (63 min). Values for AUC0-7h and volume of distribution were 970.06 mol L^-1 min.^-1 and 20.00 L/kg. Conclusions: 1) DMAQ rapidly and consistently increased ADO levels in rat tissues. 2) DMAQ was shown to have favorable pharmacokinetics, indicated by a rapid absorbtion after orally dosed, wide distribution and moderate rate of excretion. Therefore, DMAQ may be considered as a good alternative for systemic inhibition of AK that can potentially be used for the treatment of cardiovascular and neurological diseases. Authors’ disclosure statement: DMAQ concentration in plasma and ADO tissue concentrations were measured by developed and validated reversed–phase high performance liquid chromatography methods.	Antimicrobial Prophylaxis In Surgery And Emergence Of Super Bugs: Dilemma In Developing Countries RODRIGUES G^1, KHAN S^2 ^1Kasturba Medical College and ^2Manipal College of Pharmaceutical Sciences, Manipal University, Manipal, India Background: Wound infections are the commonest hospital-acquired infections in surgical patients. There is a significant difference between developed and developing countries with respect to surgical site infections (SSIs). Approximately 30-50% of antibiotic use in hospitals is now for surgical prophylaxis. However, between 30-90% of this prophylaxis is inappropriate, that increases the selective pressure favoring the emergence of antimicrobial resistance. Judicious use of antibiotics in the hospital through effective antibiotic policy and guideline development is thus essential. The objective is to evaluate the reason for noncompliance in adherence to guideline for prophylactic antibiotic usage in surgery especially with developing countries in context and to recommend the practical approach for safe and efficacious use of magic bullet in this area. Methods: We searched electronic databases including MEDLINE,EMBASE, International Pharmaceutical Abstracts,Surgical abstracts, Surgical Infection Prevention (SIP) database and transcripts on SSIs. Papers were retrieved andcategorized according to content and recommendation status by an independent reviewer. Articles highlighting the incidence of SSIs and reason for non-adherence to guideline were extracted. Results: Incidence of SSIs range from 7-15% in developing countries as compared to 2-5% in western set-up. Patients who develop SSIs are upto 60% more likely to spend time in an intensive care unit (ICU), 5 times more likely to be readmitted to the hospital and 2 times more likely to die than are patients without SSIs. There were numerous reasons for non-adherence to the clinical guideline. Irrational use of prophylactic antibiotic is rampant; especially the widespread use of the same to cover-up the aseptic techniques during surgery. Use of prophylactic antibiotic even for clean procedures (without recommendation) was very common. Choice of antibiotic too differs with the recommendation. Antibiotics were administered for prolonged duration (around two weeks) without supportive laboratory evidence. Emergence of Methicillin Resistant Staphylococcus Aureus (MRSA) and higher resistant organisms is very common is such setting. Major barriers involved in non-adherence to guidelines include lack of awareness about the guidelines and a general perception of guideline as a bureaucratic rather than educational tool. Local guidelines seem more likely to be accepted and followed than those developed nationally. Some practitioners perceive guidelines as "cookbook medicine" that does not permit them to make their own medical decisions. Other barriers are complex, multi-step systems that create confusion and decrease accountability. Methods for guideline adherence include surveillance and data analysis, new systems to facilitate documentation and improving workflow, education regarding current evidence-based guidelines and promoting the development of local guidelines or protocol, development and implementation of reminders to facilitate adherence to the local guidelines. Conclusions: 1) A multifaceted educational intervention involving a team effort of healthcare professionals can have a significant effect on effective antibiotic utilization and reducing the incidence of surgical site infections 2) Local guidelines seem more likely to be accepted and followed than those developed nationally 3) Both pros and cons should be taken into consideration while administering the prophylactic antibiotic in surgery 4) Development and adherence to antibiotic guidelines is essential to prevent emergence of resistant pathogens and to rationalize the use of antibiotics in the most cost effective manner and preventing the incidence of hospital acquired infections 5) The gap between evidence-based guidelines and practice is populated by individual values, professional conflicts, and organizational conflicts that must be addressed in order to achieve optimal practice in this domain 6) Using group interviews to reveal these factors to team members and managers may be a first step to resolving the gap and reducing surgical site infections.
Systemic Immunosuppresive Therapy With Oral Sirolimus After Bare Metal Stent Implantation: The Missing Alternative To Prevent Restenosis Following Percutaneous Coronary Interventions RODRIGUEZ AE On behalf of the ORAR III Investigators. Cardiovascular Research Ccenter (CECI),Otamendi Hospital,Buenos Aires School of Medicine,Buenos Aires,Argentina Background: In previous randomized study, ORAR II (Oral Rapamycin in Argentina), we demonstrated a significant reduction of restenosis with oral rapamycin (OR) in patients undergoing bare metal stent (BMS) implantation. Its role in comparison with drug eluting stent (DES) was unknown. Methods: Since January 2006 to September 2007 at the Cardiac facilities of three Hospitals in Buenos Aires, Argentina, 200 patients with de novo coronary lesions were randomized to treated with OR plus BMS (100 pts) or with DES (100 pts). OR was given as a bolus of 10 mg per day before PCI followed by daily doses of 3 mg plus diltiazem during 13 days after. Primary End points were to compare Hospital, Follow Up and Overall cost per patient at one, two, three and five years of follow up. Safety end point such death from any cause, Q and non Q myocardial infarction (MI) and stroke was analyzed as Major Adverse Cardiovascular Events . Target Vessel (TVR)and Target Lesion Revascularization (TLR) were independently analyzed as Efficacy end point . Direct cost in U$ dollars included procedural resources, hospitalization, medications, repeat revascularization procedures and profesional fees. Results: Baseline demographic, clinical and angiographic characteristics were similar in both groups. At 12.1 + 6.1 months of follow up, The rates of clinically driven TLR and TVR were similar with both strategies: TVR per vessel was 8.4% with OR versus 7.7% with DES, TLR per lesion was 5.7% with OR versus 5.8% with DES (p=ns).Cumulative incidence of MACCE was not significant different, although during follow up patients treated with DES had significant high incidence of MACCE compared to OR group( 13% vs 3% p=0.03 ) Initial and cumulative cost were significant high in DES treated patients (p=0.0001). Conclusions: At 12 months of follow up and initial strategy with OR plus BMS is cost saving and effective compared to DES in patients with de novo lesions undergoing PCI.	Cytochrome P450 Dependent Drug Response In Oncology RODRÍGUEZ-ANTONA C Hereditary Endocrine Cancer Group, Spanish National Cancer Research Center (CNIO), C/ Melchor Fernández Almagro, 3, 28029 Madrid, Spain Cytochromes P450 (CYP) in families 1-3 are responsible for most of the phase I hepatic biotransformations of drugs. The polymorphic nature of these enzymes causes large inter-individual differences in drug disposition that can result in differences in drug efficacy and toxicity. In this way, specific genotypes within patients could be used to individualize drug therapy. In addition to hepatic metabolism, drug efficacy can be modified by the target cells. For example, an altered drug metabolizing capacity in cancer cells could result in resistance to anticancer drugs. Here we illustrate these different situations with two recently performed studies and present our novel findings. Concerning the relevance of CYP variations for drug toxicities, we investigated the association between paclitaxel neurotoxicity and the polymorphisms of the CYPs mediating paclitaxel metabolism. Paclitaxel is an effective anti-cancer drug widely used, and neurotoxicity is one of its most important and often dose-limiting toxicities. The neurotoxicity has a large inter-individual variability of unknown molecular basis, and it represents a major challenge for the improvement of paclitaxel therapy. Since paclitaxel pharmacokinetic parameters have been shown to be related to neurotoxicity, we selected polymorphisms in the CYPs involved in paclitaxel metabolism, CYP2C8, CYP3A4 and CYP3A5, and investigated their association with paclitaxel neurotoxicity. In addition, we also selected polymorphisms potentially affecting paclitaxel hepatic uptake and biliar excretion, through the organic anion transporting polypeptides 1B1 and 1B3 and P-glycoprotein, respectively. In total 13 polymorphisms were genotyped in 124 cancer patients treated with paclitaxel. Three polymorphisms from the selected P450s were significantly associated with grade 2 neurotoxicity: the two alleles associated with an increased risk to develop neurotoxicity conferred a high metabolizing capacity, while the allele associated with protection was a low activity allele. On the other hand, for MDR1 the results were inconclusive and no association was found for the other genes. Altogether, the genetic variants identified in this work could be potential targets for individualized pharmacotherapy with paclitaxel, reducing neurotoxicity risks by improving paclitaxel dosing. Concerning the efficacy of anti-cancer drugs and the genetic background of the cancer cells, we showed that the expression of CYP3A4 in peripheral T-cell lymphoma (PTCL) tumors was inversely associated to the survival of the patients. PTCLs are clinically aggressive tumors typically showing less than 30% 5-years overall survival. To date, the most effective therapy is a combination chemotherapy regimen, in many cases CHOP (cyclophosphamide, doxorubicin, vincristine and prednisone), but the poor clinical outcome of most patients clearly reveals the need to improve the therapy by identifying factors affecting the response. Since the CYP3A subfamily enzymes are involved in the inactivation of chemotherapy drugs, we hypothesized that CYP3A expression in these lymphomas could result in a poor clinical response. We measured tumoral CYP3A and MDR1 mRNA content in 44 T-cell lymphomas finding a large variation in CYP3A expression. A high tumoral CYP3A4 expression was significantly associated with a poor drug response and survival of the patients. In conclusion, drugs efficacy and toxicity can be altered by genetic variations in CYPs, in the case of toxicity by altering drug hepatic disposition while, in the case of efficacy, alterations in the target cells can also be critical.
Design Of Aquasomes Loaded With Indomethacin And Their Released Profiles ROJAS-OVIEDO I^1, LÓPEZ-TORRES A^1, REYES-GASCAJ^2, QUIRINO-BARREDA C T^1, GARCÍA-GARCÍA R^2 ^1Universidad Autónoma Metropolitana-Xochimilco. México D.F., ^2 Universidad Nacional Autónoma de México, México, D.F. Background: Diverse technological strategies have been proposed in order to obtain nanoparticles, of a distinct nature. Kossovsky proposed a system to prepare nanoparticles transporting the so called aquasomes, whose particle size (lower than 1000 nm) is appropiate to parenteral administration. Aquasomes have a water absorbent nature which not only makes their aqueous transport permissible but also confers the possibility of establishing non-covalent links with distinct molecules and macromolecules like proteins promoting a mayor stability compared than liposomes. The aim of this study was to prepare nanoparticles in form of aquasomes with Indomethacin and determined their released profiles Methods: The inorganic cores obtained from calcium phosphate were prepared from the precipitation of a 0.25 M NaH2PO4 solution and a 0.75 M CaCl2 solution. The process variables were the level of ultrasound frequency and the influence of the sonication on the particle size of the inorganic cores. An experimental design 2^2 has involved sonication at 30 and 90 pulses of frequency after and during the precipitation of the inorganic cores. Lactose coating. 1.0 mg of inorganic cores was resuspended into bi-distilled water and was added to a 100 ml solution of Lactose with mechanical agitation. Drug loading. A solution 0.06 M of Indomethacin in acetone was added at a dispersion of 1mg/ml of the polyhydroxylated cores. After each step was carried out a filtration and a lyophilization. Structural analyses, particle size and morphology were evaluated by X-ray powder diffractometry, transmission electron microscopy (TEM), and scanning electron microscopy (SEM). Released profiles were determined by dialysis. Results: The X-ray analysis of the samples and their observation through TEM and SEM allowed us to identify the inorganic calcium phosphate nucleus formation, as well as the layers of Lactose and Indomethacin. The final particle size of the aquasomes was in the range of 60 to 120 nm, with a media of 90 nm. The results of the dialysis showed that 1.5 times more of Indomethacin was released from the aquasomes than from the physical mixture at the end of 24 hours Conclusions: The method to produce aquasomes was reproducible under the used conditions and they were able to release the drug.	Altered Morphine Metabolism In Asphyxiated Term Neonates During Prolonged, Moderate Whole Body Hypothermia Treatment ROKA A^1, KELEN D^1, VASARHELYI B^2, AZZOPARDI D^3, SZABO M^1 ^1First Department of Paediatrics, Semmelweis University, Budapest, Hungary, ^2Research Group of Paediatrics and Nephrology, Hungarian Academy of Sciences, Budapest, Hungary; ^3Division of Clinical Sciences, Hammersmith Campus, Imperial College, London, UK Background: Hypothermia is a new treatment of cerebral injury caused by perinatal asphyxia. Both asphyxia and hypothermia may modify drug pharmacokinetics. We investigated whether analgesia with morphine in neonates with hypoxic ischaemic encephalopathy treated with prolonged moderate systemic hypothermia resulted in elevated serum morphine concentrations compared to normothermic infants. Methods: Infants from one centre participating in a multicentre randomized study of moderate whole body hypothermia following perinatal asphyxia (the TOBY study), were randomly selected for treatment with hypothermia (n=10) or for standard care on normothermia (n=6). Hypothermia (33-34 0C) was started before six hours of age, and was maintained for 72 hours. All infants were treated with a continuous infusion of morphine-HCl, with the rate adjusted according to clinical status. Serum morphine concentrations were determined at 6, 12, 24, 48 and 72 hours after birth. Results: Serum morphine concentrations at 24-72 hours after birth were, median (range), 292 (137-767) ng.ml^-1 in the infants treated with hypothermia and 206 (88-327) ng.ml^-1 in the infants on normothermia, p=0.014, despite similar morphine infusion rates, and cumulative doses. Morphine concentrations correlated with morphine infusion rate (r=0.663, p<0.0001), with cumulative dose (r=0.646, p<0.0001), and with treatment with hypothermia (r=0.441, p=0.004). Serum morphine concentrations reached a steady state after 24 hours in the normothermic infants but continued to increase throughout the assessment period in the hypothermia group. Serum morphine concentrations greater than 300 ng.ml-1 occurred more often in the hypothermia group (p<0.025), and when morphine infusion rate was >10 μg.kg^-1.hr^-1 (p<0.01). Conclusions: Infants with hypoxic ischaemic encephalopathy have reduced morphine clearance and elevated, potentially toxic serum morphine concentrations when morphine infusion rates are based on clinical state.
Isoproterenol And Insulin: The Ying-Yang Of Regulation Of Mg2+ Homeostasis In Mammalian Cells ROMANI A Department of Physiology and Biophysics – Case Western Reserve University, Cleveland, USA Background:The investigation of the mechanisms regulating cellular Mg2+ homeostasis has highlighted a novel effect of isoproterenol and insulin on mammalian cells. Methods: Over the years, studies were conducted in whole anesthetized animals, perfused organs, isolated cells as well as purified subcellular organelles from normal or diabetic animals. Results: Isoproterenol administration results in the rapid extrusion of a sizable amount of cellular Mg2+ via a putative Na+/Mg2+ exchanger. This extrusion primarily mobilizes Mg2+ from cytoplasm and mitochondria. In contrast, treatment with insulin abolishes isoproterenol-induced Mg2+ extrusion and elicits per se an accumulation of Mg2+ from the extra-cellular compartment into the cell, which in certain cell types (e.g. cardiac myocytes) is associated with glucose accumulation. Further evidence of this opposite regulation is provided by diabetic conditions. The reduced or absent responsiveness to insulin, in fact, results in a marked decrease of Mg2+ within several tissues including skeletal muscles, heart, and liver, and in an enhanced responsiveness to isoproterenol or catecholamine stimulation, which further decreases tissue Mg2+ level and impairs the operation of several Mg2+-regulated enzymes.