Ehrlich II –2nd World Conference on Magic Bullets

Dual mechanistic anorexigenic and anti-adipogenic therapeutic for the treatment of obesity CHIN KV^1,4,5, QASEM A^1, SAUNDERS RA^1, SZLUDLAREK, M^1, CHIN A^1, BOSIO RM^2, WU Q^1, SHAPIRO J^1, NAJJAR S^3,4 Department of Medicine^1, Department of Surgery^2, Department of Physiology, Pharmacology, Metabolism, and Cardiovascular Sciences^3, Center for Diabetes and Endocrine Research^4, and Department of Biochemistry and Cancer Biology^5, The University of Toledo, College of Medicine, 3000 Arlington Avenue, Toledo, Ohio 43614, USA. Background: The sharp rise in obesity in the last decade is one of the most serious public health risks worldwide. Currently approved therapies for obesity exhibit modest efficacy and limiting side effects. We show here the identification of a novel pharmacological agent, ECP00068, for the treatment of obesity. Methods: The morbidly obese leptin-deficient, ob/ob, and the leptin receptor-deficient db/db mice, as well as the glucose intolerant/type II diabetes proned Ceacam^-/-, and the diet-induced obese (DIO) mice, were treated with ECP00068. Effects of ECP00068 on the differentiation of preadipocytes into adipocytes and on differentiated adipocytes were examined. Mechanisms of action of ECP00068 was investigated by whole genome DNA microarray during the differentiation of preadipocytes into adipocytes. Results: We showed that ECP00068 causes appetite suppression that results in up to 50% reduction in food intake, decrease in visceral and subcutaneous adipose tissues, and weight loss in ob/ob and db/db, Ceacam^-/-, and DIO mice. ECP00068 inhibits the proliferation and differentiation of preadipocytes, and causes either dedifferentiation or delipidation of adipocytes. Gene expression profiling showed that inhibition of differentiation by ECP00068 was accompanied by the transcriptional inhibition of a large cluster of fat regulatory genes with functional equivalence in C. elegans. Additionally, ECP00068 increases the expression of a transcriptional repressor, zinc finger protein 68 (Zfp68), which inhibits the CCAAT/enhancer binding proteins (C/EBPs) and the peroxisome proliferator-activated receptor  (PPAR). Conclusions: 1) The unique dual anorexigenic and anti-adipogenic pharmacological profile of ECP00068 suggests a first-in-class anti-obesity drug. 2) Drug-induced tolerance resulting from chronic pharmacotherapy for obesity can be overcome by a cyclical drug holiday treatment strategy. 3) Zfp68 is a repressor of C/EBP and , and PPAR, master regulators of adipogenesis.

A potential molecular link for metabolic stress and carcinogenesis: AMP-activated protein kinase CHING YP The University of Hong Kong, Pokfulam, Department of Anatomy, LKS medical school Hong Kong Background: AMP-activated protein kinase (AMPK), which is a serine/theronine protein kinase, is found to be a key regulator in glucose and lipid metabolism in response to cellular stress and hormones such as leptin and adiponectin. Metabolic stresses, like heat shock, hypoxia or ischemia, and glucose deprivation, have been demonstrated to activate AMPK activity. Activation of AMPK aims to switch off pathways that consume ATP, like inhibition of carbohydrates and lipid synthesis; while switch on pathways that generate ATP, like fatty acid oxidation, glucose transport and glycolysis. Recently, AMPK has been shown to be the molecular target of a wildly used anti-diabetic drug, metformin. Recent publications suggest that AMPK activation results in suppressing cell proliferation. AMPK lies upstream and downstream of two tumor suppressors, TSC2 and LKB1, respectively, indicating that AMPK may also involve in carcinogenesis. Liver cancer (hepatocellular carcinoma, HCC) is one of the most common cancers worldwide. However, the molecular mechanism underlying the development of HCC is still unclear. Here, we examined if AMPK is related to HCC formation. Methods and Results: Using real-time quantitative PCR, we observed that AMPK2 was significantly underexpressed in HCCs (47.6%) as compared to its corresponding nontumorous liver samples. To further confirm the effect of AMPK on hepatocarcinogenesis, we established stable HCC clones expressing short-hairpin RNA (shRNA) that can specifically knockdown the endogenous AMPK2 and assay for the proliferation rate as well as anchorage-independent growth in soft agar. The results showed that the stable AMPK2 knockdown clones proliferated much faster and formed more colonies than the vector control. Since p53, an important metabolic checkpoint protein, was recently reported to be regulated by AMPK, we query whether AMPK-regulated HCC cell growth is p53-dependent. Using in vitro kinase assay, we showed that AMPK2 catalytic domain can directly phosphorylate p53. Interestingly, we found that AMPK phosphorylated p53 at a novel site T150, which affect the stability of p53 protein. Conclusions: our results suggest that AMPK may mediate its tumor suppression function through regulation of p53 in HCC.

An Entry to Clavams from Chiral Vinyl Ethers and Chlorosulfonyl Isocyanate CHMIELEWSKI M Institute of Organic Chemistry of the Polish Academy of Sciences, 01-224 Warsaw, Kasprzaka 44/52, Poland, e-mail:[e-mail] [2+2]Cycloaddition between chiral vinyl ethers 1-4 and chlorosulfonyl isocyanate leading to corresponding β-lactams 5-8 was investigated. Reactions proceeded with excellent diastereoselectivity. The special attention was focused on the problem of stereocontrol in the formation of a desired configuration of the C-4 carbon atom of the azetidinone ring. Adducts 5-8 were transformed into clavams 9-12. The adduct 5 was subjected to the sequence of reactions, in which glycolic cleavage and intramolecular alkylation of the nitrogen played crucial roles. Adducts 6-8 were transformed into clavams 10-12 via intramolecular alkylation of the nitrogen atom. The antibacterial and antifungal properties of all clavams obtained were investigated to show, however, only in few cases interesting activity.

Bulletprooves to Abacavir-related Hypersensitivity Reaction in HIV-1-infected Population CHOLEWINSKA G Hospital for Infectious Diseases. Warsaw, Poland   Background: Current antiretroviral (ARV) therapy are effective, allowing HIV-positive individuals to live longer, but at the same time they have resulted in many drug-related complications. One of it, is hypersensitivity reaction (HSR) to abacavir (ABC)treatment. Pharmacogenetics associations can influence on HIV-1 treatment. Few recent studies have shown that patients who were positive for HLA B*5701 alleles have a high risk allergic reaction to abacavir. The individual loci of alleles haplotype has been combined to susceptibility to ABC-HSR. Prevalence of HLA B*5701 among Caucasian are 5-7%. Methods: This is retrospective analysis of clinically diagnosed ABC-HSR among 217 treated with abacavir-containing ARV regimen HIV-1 infected individuals, hospitalized between 2003-2008 in the Hospital for Infection Diseases in Warsaw. The aim of analysis was attempt to determine of *B57 alleles carriage in 12 patients (M-9, F-3, av. age 39), who developed ABC-HSR incidence. Multiple symptoms were observed in this cases, defined as abacavir-related allergic reaction, according to HSR characteristic signs and time onset between 1-6 weeks after abacavir initiation. The HLA test was performed in all 12 patients defined as ABC-HSR, using of method Sequens Based Typing [Atria Genetics]. Results: During recent 5 years, abacavir-containing antiretroviral therapy received 217 hospitalized patients. In 12 (5,5%) out of this group have clinically confirmed ABC-HSR and discontinued ABC treatment in consequence. Simply symptoms, such as skin and gastrointestinal reaction, were observed in 4 (33%) individuals. In 2 other (17%) constitutional and severe respiratory symptoms were reported. Among 6 other (50%), 3 or more HSR signs including fever, paresthesis, hypotension, tachycardia, liver parameters elevation, were observed. The onset of HSR occurred between 6 day and 6 week afterwards. No death registered. The HLA B*5701 test performed in subsequence HSR, using the sequence typing method. Only 1 patient was HLA B*5701 positive. It concernd 36y men with AIDS-C3 category, who received 5-th ARV-drug regimen due to resistance and intolerance previous antiretroviral agents. Conclusions: 1. HLA B*5701 screening in HIV-1-infected individuals has the predictive value to ABC-HSR risk. As a result, cases of abacavir-related hypersensitivity reaction incorrectly diagnosed clinically, as well as abacavir unnecessarily discontinuation, can be reduced in clinical practice.