The Importance of Clinical Trials in Drug Development 2 2 While preclinical research answers basic questions about a drug’s safety, it is not a substitute for studies of ways the drug will interact with the human body. “Clinical research” refers to studies, or trials, that are done in people 1,2. As the developers design the clinical study, they will consider what they want to accomplish for each of the different Clinical Research Phases and begin the Investigational New Drug Process, a process they must go through before clinical research begins. The ultimate goal of drug development is to bring a new compound with proven therapeutic effect to the market. In this context, the transition from preclinical research to clinical stages marks a critical turning point, as it nears the new medicinal product to the market 3,4. With the promise of marketing authorization, though far ahead in the road, hanging on the horizon, the approval of a clinical trial usually attracts investors and leads to a respectable rise of the company shares. However, everything comes at a price. Clinical trials are not without risks, and while the perspective of success is encouraging, the crude reality is that most compounds fail before reaching the market. As explained in previous entries, despite higher R&D expenditures, attrition rates are high and, what is worse, on the rise. Data collected between 1990 and 2004 show that the number of unsuccessful clinical trials has been steadily increasing during the last years: from 30% to 50% at Phase 1, from 40% to 70% at Phase 2 and from 20% to nearly 50% at Phase 3 3,4. As a result, less than 10% of the drugs that enter clinical trials end up being approved by regulatory agencies. Clinical trials are only a small part of the research that goes into developing a new treatment. Drugs of the future, for example, first have to be discovered or created, purified, described, and tested in labs (in cell and animal studies) before ever reaching human clinical trials. Of all the substances that are tested in these early stages, very few are promising enough to be tested in humans. Drug development is the process of bringing a new pharmaceutical drug to the market once a lead compound has been identified through the process of drug discovery. It includes pre-clinical research on microorganisms and animals, filing for regulatory status, such as via the United States Food and Drug Administration for an investigational new drug to initiate clinical trials on humans, and may include the step of obtaining regulatory approval with a new drug application to market the drug. Indeed, of every 5,000 cancer molecules identified in the laboratory, about 250 will enter pre-clinical testing. Of this 250, fewer than 10 are tested in clinical trials and on average only one will be approved by regulatory authorities. The process of bringing a new treatment from the research stage (laboratory) to clinic is estimated to take between 10-13 years 5. 151 151 Shahin Akhondzadeh Psychiatric Research Center, Roozbeh Hospital, South Kargar Street Psychiatric Research Center, Roozbeh Hospital, South Kargar Street Iran Editorial 249.pdf Sepanjnia K, Modabbernia A, Ashrafi M, Modabbernia MJ, Akhondzadeh S. Pioglitazone adjunctive therapy for moderate-to-severe major depressive disorder: randomized double blind placebo-controlled trial. Neuropsychopharmacology 2012;37(9):2093-2100.##Akhondzadeh S. Hippocampal synaptic plasticity and cognition. J Clin Pharm Ther 1999;24(4):241-248.##Akhondzadeh S, Malek-Hosseini M, Ghoreishi A, Raznahan M, Rezazadeh SA. Effect of ritanserin, a 5HT2A/2C antagonist, on negative symptoms of schizophrenia: a double-blind randomized placebo-controlled study. Prog Neuropsychopharmacol Biol Psychiatry 2008;32(8):1879-1883.##Amrollahi Z, Rezaei F, Salehi B, Modabbernia AH, Maroufi A, Esfandiari GR, et al. Double-blind, randomized, placebo-controlled 6-week study on the efficacy and safety of the tamoxifen adjunctive to lithium in acute bipolar mania. J Affect Disord 2011;129(1-3):327-331.##Akhondzadeh S, Ahmadi-Abhari SA, Assadi SM, Shabestari OL, Kashani AR, Farzanehgan ZM. Double-blind randomized controlled trial of baclofen vs. clonidine in the treatment of opiates withdrawal. J Clin Pharm Ther 2000;25(5):347-353.##

Paroxetine Can Enhance Neurogenesis During Neurogenic Differentiation of Human Adipose-derived Stem Cells 2 2 Background: Some antidepressant drugs can promote neuronal cell proliferation in vitro as well as hippocampal neurogenesis in human and animal models. Furthermore, adipose tissue is an available source of adult stem cells with the ability to differentiate in to multiple lineages. Therefore, human Adipose-Derived Stem Cells (hADSCs) may be a suitable source for regenerative medical applications. Since there is no evidence for the effect of Paroxetine as the most commonly prescribed antidepressant drug for neurogenic potential of hADSCs, an attempt was made to determine the effect of Paroxetine on proliferation and neural differentiation of hADSCs. Methods: ADSCs were isolated from human abdominal fat. These cells differentiated to neuron-like cells and were treated with Paroxetine. 3-[4, 5-dimethylthiazol-2-yl]-2, 5-diphenyl tetrazolium bromide (MTT) assay and immunofluorescence technique were used for assessment of cell proliferation and neurogenic differentiation potential of induced cells, respectively. Results: MTT assay analysis showed that Paroxetine significantly increased the proliferation rate of induced hADSCs (p<0.05), while immunofluorescent staining indicated that Paroxetine treatment during neurogenic differentiation could enhance the mean percentage of Nestin and MAP2 (Microtubule-associated protein-2) positive cells but the mean percentage of GFAP (Glial acidic fibrillary protein) positive cells significantly decreased relative to control group (p<0.05). Conclusion: Our results provide evidence that Paroxetine can promote proliferation and differentiation rate during neurogenic differentiation of hADSCs. Moreover, Paroxetine can reduce gliogenesis of induced hADSCs during neurogenic differentiation. 152 158 Maliheh Jahromi Department of Anatomical Sciences, Faculty of Medicine, Isfahan University of Medical Sciences Department of Anatomical Sciences, Faculty of Medicine, Isfahan University of Medical Sciences Iran Shahnaz Razavi Department of Anatomical Sciences, Faculty of Medicine, Isfahan University of Medical Sciences Department of Anatomical Sciences, Faculty of Medicine, Isfahan University of Medical Sciences Iran Nushin Amirpour Department of Anatomical Sciences, Faculty of Medicine, Isfahan University of Medical Sciences Department of Anatomical Sciences, Faculty of Medicine, Isfahan University of Medical Sciences Iran Zahra Khosravizadeh Department of Anatomical Sciences and Molecular Biology, Faculty of Medicine, Tehran University of Medical Sciences Department of Anatomical Sciences and Molecular Biology, Faculty of Medicine, Tehran University of Medical Sciences Iran Antidepressant drugsNeurogenic differentiationParoxetineProliferationStem cells 251.pdf Magni LR, Purgato M, Gastaldon C, Papola D, Furukawa TA, Cipriani A, et al. 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Exogenous Secreted Frizzled-Related Protein-4 Modulates Steroidogenesis of Rat ‎Granulosa Cells Through Wnt/Bcatenin and PI3K/AKT Signaling Pathways‎ 2 2 Background: It has been reported that secreted frizzled-related protein-4 known as an antagonist of Wnt signaling pathway plays a role in luteinization process of rodent granulosa cells. The purpose of this study was twofold: 1) to determine whether recombinant human secreted frizzled-related protein-4 (rhSFRP-4) could directly induce terminal differentiation of rat Granulosa Cells (GCs) and 2) to understand how the modulation of β-catenin and Protein Kinase B (PKB)/AKT activity by exogenous SFRP-4 could be involved in steroidogenesis. Methods: GCs were firstly stimulated with Follicle-Stimulating Hormone (FSH) named as FSH-primed cells then were treated with luteinizing hormone (LH). Then estradiol (E2) and progesterone (P4) production levels were assessed in the absence or presence of rhSFRP-4 treatment. The expression levels of activated β-catenin, pAKTser473, pGSK3βser9 were assessed by western blot or immuno-fluoresence. Results: In the presence of rhSFRP-4, there was 38% decreased E2 levels compared to untreated FSH-primed cells (p<0.05), and P4 production subsequently decreased. However, in GCs pre-treated with rhSFRP-4 prior to addition of FSH, P4 levels increased 2-fold compared with untreated cells (p<0.05). Unexpectedly, treatment with rhSFRP-4 prior to LH stimulation inhibited LH-induced P4 secretion. Treatment with low (0.5 ng/ml) but not high (50 ng/ml) concentrations of rhSFRP-4 led to significantly increased levels of pGSK3βser9 (1.6-fold) and nuclear active β-catenin (2.8-fold) in GCs compared with untreated cells. Interestingly, pre-treating GCs with rhsFPR4 prior to LH stimulation resulted in a 38% decrease in pAKTser473 levels compared with those in LH-treated cells (p<0.05). Conclusion: Taken together, our results showed that rhSFRP-4 could directly induce terminal differentiation in GCs via the modulation of β-catenin and PKB/AKT pathways and that it does so in a dose-dependent manner. 159 168 Ghamartaj Hossein Department of Animal Physiology, Developmental Biology Laboratory, Faculty of Biology, University College of Science, University of Tehran Department of Animal Physiology, Developmental Biology Laboratory, Faculty of Biology, University College of Science, University of Tehran Iran Manijeh Khanmohammadi Department of Animal Physiology, Developmental Biology Laboratory, Faculty of Biology, University College of Science, University of TehranReproductive Biotechnology Research Center, Avicenna Research Institute, ACECR Department of Animal Physiology, Developmental Biology Laboratory, Faculty of Biology, University College of Science, University of TehranReproductive Biotechnology Research Center, Avicenna Research Institute, ACECR IranIran Parissa Sahranavard Fard Department of Animal Physiology, Developmental Biology Laboratory, Faculty of Biology, University College of Science, University of TehranDepartment of Stem Cells & Developmental Biology at Cell Science Research Center, Royan Institute for Stem Cell Biology & Technology, ACECR Department of Animal Physiology, Developmental Biology Laboratory, Faculty of Biology, University College of Science, University of TehranDepartment of Stem Cells & Developmental Biology at Cell Science Research Center, Royan Institute for Stem Cell Biology & Technology, ACECR IranIran Yasaman Heidarian Department of Animal Physiology, Developmental Biology Laboratory, Faculty of Biology, University College of Science, University of Tehran Department of Animal Physiology, Developmental Biology Laboratory, Faculty of Biology, University College of Science, University of Tehran Iran Somaieh Kazemnejad Reproductive Biotechnology Research Center, Avicenna Research Institute, ACECR Reproductive Biotechnology Research Center, Avicenna Research Institute, ACECR Iran Mohammad Mehdi Akhondi Reproductive Biotechnology Research Center, Avicenna Research Institute, ACECR Reproductive Biotechnology Research Center, Avicenna Research Institute, ACECR Iran Active β-cateninGSK3βPKB/AKTRat granulosa cellSecreted frizzled-related protein-4 (SFRP-4) 252.pdf Vainio S, Heikkilä M, KispertA, Chin N, McMahon AP. 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Novel Combinations of Synthesized ZnO NPs and Ceftazidime: Evaluation of their Activity Against Standards and New Clinically Isolated Pseudomonas aeruginosa 2 2 Background: Antibiotic resistant bacteria can be considered as a main problem in infection management. Zinc oxide nanoparticles (ZnO NPs), individually or in combination with antibiotics, can be considered as good candidates for struggling against drug resistant bacteria. Methods: In this study, Zinc oxide nanoparticles were synthesized using sol-gel method in low temperature as a cost effective procedure and characterized by X-ray diffraction and Scanning Electron Microscopy. Antibacterial activity of 9 new combinations of Zinc oxide nanoparticles and ceftazidime was assessed against standards and new clinically isolated multi drug resistant Pseudomonas aeruginosa (P. aeruginosa), in order to evaluate enhancement effect of synthesized Zinc oxide nanoparticles on antibacterial activity of ceftazidime. Results: The results indicated that desirable effects can be seen at 6 and 7 mM of Zinc oxide nanoparticles (60 to 100% inhibition). Moreover, after evaluation of 9 new combinations with various concentrations of both components, it was demonstrated that Zinc oxide nanoparticles can enhance the antibacterial activity of ceftazidime, against some bacterial strains of P. aeruginosa. The highest activity was observed with the concentration of 20 μg/ml ceftazidime in the presence of 5, 6 or 7 mM of Zinc oxide nanoparticles. Conclusion: Zinc oxide nanoparticles in appropriate concentrations can be proposed as new and promising candidates for overcoming bacterial resistance. 169 174 Elham Isaei Department of Microbiology, Afzalipour Faculty of Medicine, Kerman University of Medical Sciences Department of Microbiology, Afzalipour Faculty of Medicine, Kerman University of Medical Sciences Iran Shahla Mansouri Department of Microbiology, Afzalipour Faculty of Medicine, Kerman University of Medical Sciences Department of Microbiology, Afzalipour Faculty of Medicine, Kerman University of Medical Sciences Iran Fereshteh Mohammadi Nanobiotechnology Research Center, Avicenna Research Institute, ACECR Nanobiotechnology Research Center, Avicenna Research Institute, ACECR Iran Sadegh Taheritarigh Department of Plant Breeding and Biotechnology, Faculty of Plant Production, Gorgan University of Agricultural Sciences and Natural Resources Department of Plant Breeding and Biotechnology, Faculty of Plant Production, Gorgan University of Agricultural Sciences and Natural Resources Iran Zohreh Mohammadi Nanobiotechnology Research Center, Avicenna Research Institute, ACECR Nanobiotechnology Research Center, Avicenna Research Institute, ACECR Iran Antibiotic resistanceCeftazidimePseudomonas aeruginosaZnO NPs 253.pdf Li Q, Mahendra S, Lyon DY, Brunet L, Liga MV, Li D, et al. 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Enhanced bioactivity of ZnO nanoparticles-an antimicrobial study. Sci Technol Adv Mater 2008;9(3):035004.##Tokumoto MS, Briois V, Santilli CV, Pulcinelliet SH. Preparation of ZnO nanoparticles: structural study of the molecular precursor. J Solgel Sci Technol 2003;26(1-3):547-551.##Kim JH, Choi WCh, Kim HY, Kang Y, Park YK. Preparation of mono-dispersed mixed metal oxide micro hollow spheres by homogeneous precipitation in a micro precipitator. Powder Technol 2005;153(3):166-175.##Damonte LC, Mendoza Zélis LA, Marí Soucase B, Hernández Fenollosa MA. Nanoparticles of ZnO obtained by mechanical milling. Powder Technol 2004;148(1):15-19.##Kahn ML, Mong M, Colliere V, SDenocq F, Maisonnat A, Chaudret B. Size-and shape-control of crystalline zinc oxide nanoparticles: a new organometallic synthetic method. Adv Funct Mater 2005;15(3):458-468.##Komarneni S, Bruno M, Mariani E. Synthesis of ZnO with and without microwaves. Mater Res Bull 2000;35(11):1843-1847.##Tani T, Mädler L, Pratsinis SE. 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Cloning and Expression of Soluble Recombinant HIV-1 CRF35 Protease-HP Thioredoxin Fusion Protein 2 2 Background: As a drug target and an antigenic agent, HIV-1 protease (HIV-1 PR) is at the center of attention for designing anti-AIDS inhibitors and diagnostic tests. In previous studies, the production of the recombinant protease has been faced with several difficulties; therefore, the aims of this study were the easy production, purification of the soluble form of protease in E. coli and investigation of its immunoreactivity. Methods: Protease coding region was isolated from the serum of an infected individual, amplified by RT-PCR and cloned into PTZ57R using TA-cloning. Protease coding frame was isolated by PCR and cloned in pET102/D. TOPO expression vector and cloned protease was expressed in Escherichia coli (E. coli) BL21. Produced recombinant protein was purified by affinity Ni-NTA column and protein concentration was checked by BCA protein assay kit. Subsequently, immunoreactivity of recombinant protease (rPR) was assayed by Western blotting and ELISA. Results: Cloning of the HIV protease by TOPO cloning system in pET102/D.TOPO was confirmed with PCR and sequencing. The concentration range of purified recombinant protein was 85 to 100 μg/ml. Immunogenicity of rPR was confirmed by Western blotting and ELISA. Conclusion: Soluble production of recombinant HIV-1 protease (HIV-1 rPR) was performed successfully. This recombinant protein disclosed 86% specificity and 90% sensitivity in immunoassay tests. 175 181 Asaad Azarnezhad Cellular and Molecular Research Center, Kurdistan University of Medical SciencesDepartment of Medical Genetics, Faculty of Medicine, Tehran University of Medical Science Cellular and Molecular Research Center, Kurdistan University of Medical SciencesDepartment of Medical Genetics, Faculty of Medicine, Tehran University of Medical Science IranIran Zohreh Sharifi Blood Transfusion Research Center, Institute for Research and Education in Transfusion Medicine Blood Transfusion Research Center, Institute for Research and Education in Transfusion Medicine Iran Rahmatollah Seyedabadi Department of Molecular Medicine and Genetics, Faculty of Medicine, Hamedan University of Medical Sciences Department of Molecular Medicine and Genetics, Faculty of Medicine, Hamedan University of Medical Sciences Iran Arshad Hosseini Department of Medical Biotechnology, Faculty of Allied Medicine, Iran University of Medical Sciences Department of Medical Biotechnology, Faculty of Allied Medicine, Iran University of Medical Sciences Iran Behrooz Johari Department of Medical Biotechnology, Faculty of Allied Medicine, Iran University of Medical Sciences Department of Medical Biotechnology, Faculty of Allied Medicine, Iran University of Medical Sciences Iran Mahsa Sobhani Fard Department of Immunology, Faculty of Medicine, Hamedan University of Medical SciencesStudent Research Center, Hamedan University of Medical Sciences Department of Immunology, Faculty of Medicine, Hamedan University of Medical SciencesStudent Research Center, Hamedan University of Medical Sciences IranIran Human immunodeficiency virusMolecular cloningProteaseRecombinant proteins 254.pdf Robertson DL, Sharp PM, McCutchan FE, Hahn BH. 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Evaluation of Recombinant Human Growth Hormone Secretion in E. coli using the L-asparaginase II signal Peptide 2 2 Background: In the recent years, there has been an increasing interest in secretory production of recombinant proteins, due to its various advantages compared with intracellular expression. Signal peptides play a critical role in prosperous secretion of recombinant proteins. Accordingly, different signal peptides have been assessed for their ability to produce secretory proteins by trial-and-error experiments. The aim of this study was to evaluate the effect of L-asparaginase II signal peptide on the recombinant human Growth Hormone (hGH) protein secretion in the Escherichia coli (E. coli) host. Methods: Cloning and expression of a synthetic hGH gene, containing L-asparaginase II signal sequence was performed in E. coli BL21 (DE3) using 0.1 mM IPTG as an inducer at 23oC overnight. Periplasmic protein extraction was performed using three methods, including osmotic shock, osmotic shock in the presence of glycine and combined Lysozyme/EDTA osmotic shock. Afterwards, the hGH expression was determined by SDS-PAGE. Results: Based on experimentally obtained results, hGH protein is expressed as inclusion body even in the presence of L-asparaginase II signal peptide. Conclusion: Therefore, this signal peptide is not effective for secretory production of the recombinant hGH. 182 187 Mozhdeh Zamani Pharmaceutical Sciences Research Center, Shiraz University of Medical SciencesDepartment of Pharmaceutical Biotechnology, Faculty of Pharmacy, Shiraz University of Medical Sciences Pharmaceutical Sciences Research Center, Shiraz University of Medical SciencesDepartment of Pharmaceutical Biotechnology, Faculty of Pharmacy, Shiraz University of Medical Sciences IranIran Navid Nezafat Pharmaceutical Sciences Research Center, Shiraz University of Medical Sciences Pharmaceutical Sciences Research Center, Shiraz University of Medical Sciences Iran Younes Ghasemi Pharmaceutical Sciences Research Center, Shiraz University of Medical SciencesDepartment of Pharmaceutical Biotechnology, Faculty of Pharmacy, Shiraz University of Medical Sciences Pharmaceutical Sciences Research Center, Shiraz University of Medical SciencesDepartment of Pharmaceutical Biotechnology, Faculty of Pharmacy, Shiraz University of Medical Sciences IranIran Escherichia coli (E. coli)Human growth hormoneL-asparaginase IIRecombinant proteinsSignal peptide 255.pdf Tritos NA, Mantzoros CS. 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Preliminary Study on Cost-effective L-Tryptophan Production from Indole and L-Serine by E. coli Cells 2 2 Background: L-tryptophan is used widespread in the pharmaceutical industry. The majority of L-Trp production depends on microbial processes that produce L-tryptophan from indole and L-serine. These processes are very costly due to the costs of precursors, especially L-serine. Use of inexpensive substitutions as the L-serine source of L-tryptophan production enables us to reach a cost-effective process. In this paper, effect of Triton X-100 on L-Trp production and the ability to use Iranian cane molasses as inexpensive L-serine source was investigated. Methods: Escherichia coli (E. coli) ATCC 11303 cells were grown in 10-L fermenter containing minimal medium supplemented with beet molasses as an inexpensive carbon source and indole as tryptophan synthase inducer. Whole cells of stationary phase were used as biocatalyst for L-Trp production. Triton X-100 addition to the production medium as indole reservoir was investigated. Then, cane molasses was used as L-Ser source in L-Trp production medium. Amount of L-Tryptophan and theoretical yield of L-Trp production was determined by HPLC and by a colorimetrically method on the basis of remaining indole assay, respectively. Results: As a result, triton X-100 increased L-Trp production three times.  Also, the result showed that 0.68 mM L-Tryptophan was produced in the presence of cane molasses at 37oC for 8 hr. Conclusion: This result showed that cane molasses of Qazvin sugar factory includes significant amounts of L-Ser that makes it a suitable substitution for L-Ser in L-Trp production. Therefore, it has the potential to be used for cost-effective L-Trp production in industrial scale. 188 192 Tahereh Sadeghiyan-Rizi Department of Pharmaceutical Biotechnology, Faculty of Pharmacy, Isfahan University of Medical Sciences Department of Pharmaceutical Biotechnology, Faculty of Pharmacy, Isfahan University of Medical Sciences Iran Jamshid Fooladi National Laboratory of Industrial Microbiology, Department of Biology, Faculty of Science, Alzahra University National Laboratory of Industrial Microbiology, Department of Biology, Faculty of Science, Alzahra University Iran Sima Sadrai Department of Pharmaceutics, Faculty of Pharmacy, Tehran University of Medical Sciences Department of Pharmaceutics, Faculty of Pharmacy, Tehran University of Medical Sciences Iran MolassesPLPTryptophan synthaseTryptophanase 256.pdf Azuma S, Tsunekawa H, Okabe M, Okamoto R, Aiba S. Hyper-production of L-tryptophan via fermentation with crystallization. Appl Microbiol Biotechnol 1993;39(4):471-476.##Bang WG, Lang S, Sahm H, Wagner F. Production of L-tryptophan by Escherichia coli cells. Biotechnol Bioeng 1983;25(4):999-1011.##Ren J, Zhao M, Wang J, Cui Ch, Yang B. Spectrophotometric method for determination of Tryptophan in protein hydrolysates. Food Technol Biotechnol 2007;45(4):360-366.##Bongaerts J, Krämer M, Müller U, Raeven L, Wubbolts M. Metabolic engineering for microbial production of aromatic amino acids and derived compounds. Metab Eng 2001;3(4):289-300.##Muller CP, Jacobs BL. Handbook of the Behavioral Neurobiology of Serotonin. 1st ed. London: Elsevier press; 2010. p. 9-11.##Kino K, Nakanishi T, Sugimoto M. Process for preparation of L-tryptophan. United States patent US 4742007. 1988.##Nyeste L, Pécs M, Sevella B, Hollo J. Production of L-Tryptophan by Microbial Processes. Adv Biochem Eng Biotechnol 1983;26:176-202.##Nakazawa H, Enei H, Okumura S, Yoshida H, Yamada H. Enzymatic preparation of L-Tryptophan and 5-hydroxy-L-tryptophan. FEBS Lett 1972;25(1):43-45.##Ebihara Y, Niitsu N, Terui G. Fermentative production of tryptophan from indole by Hansenula anomala. J Ferment Technol 1969;47:733-738.##Chan EC, Tsai HL, Shen SL, Mou DG. Amplification of tryptophan operon gene in Escherichia coli chromosome to increase L-tryptophan biosynthesis. Appl Microbiol Biotechnol 1993;40(2):301-305.##Sadeghiyan-Rizi T, Fooladi J, Momhed Heravi M, Sadrai S. Optimization of L-Tryptophan Biosynthesis from L-Serine of processed Iranian Beet and Cane Molasses and Indole by induced Escherichia coli ATCC 11303 Cells. Jundishapur J Microbiol 2014;7(6):e10589.##Eggeling L, Pfefferle W, Sahm H. Amino acids. In: Ratledge C, Kristiansen B, editors. Biotechnology. England: Cambridge University Press; 2006. p. 335-359.##Sumpter WC, Miller FM. The chemistry of heterocyclic compounds, indole and carbazole systems. 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A colorimetric method for the determination of indole, and its application to assay of tryptophanase. Anal Biochem 1978;86:457-462.##Mohammad A, Zehra A. Specific separation of thiamine from hydrophilic vitamins with aqueous dioxane on precoated silica TLC plates. Acta Chromatogr 2008;20(4):637-642.##Chapman HL, Kidder RW, Koger M, Crockett JR, Merherson WK. Blackstrap molasses for beef cows. 1st ed. USA: Gainesville, Fla: Agricultural Experiment Stations, Institute of Food and Agricultural Sciences, University of Florida; 1965. P. 701-711.##Ribeiro MHL, Prazeres DMF, Cabral JMS, Fonseca MMR. Adsorption studies for the separation of L-tryptophan from L-serine and indole in a bioconversion medium. Bioprocess Eng 1995;12(1):95-102.##Hagino H, Nakayama K. L-Tryptophan production by analog-resistant mutants derived from a phenylalanine and tyrosine double auxotroph of Corynebacterium glutamicum. Agric Biol Chem 1975;39(2):343-349.##Mateus DMR, Alves SS, Fonseca MMR da. 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Development of a High-Resolution Melting Analysis Method for CYP2C19*17 Genotyping in Healthy Volunteers 2 2 Background: Genetic polymorphisms of drug metabolisms by cytochrome P450 (P450s) could affect drug response, attracting particular interest in the pharm-acogenetics. Due to the importance of CYP2C19* 17 allele and its capability of super- fast metabolism and also lack of information about distribution of the alleles in Iranian population, this research aimed to use High Resolution Melting (HRM) method compared to PCR-RFLP for genotyping healthy Iranian population. Methods: Blood samples were collected from 100 healthy Iranian volunteers. DNA was extracted by salting out method. Real-time PCR was used for amplification of the CYP2C19 gene and the alleles were identified by HRM. Sequencing was used to confirm the amplified DNA fragments and data were analyzed using SPSS software ver.18. Results: The frequency of alleles CYP2C19*1/*1, CYP2C19*1/*17 and CYP2C19*17/*17 were estimated as 58.33, 29.1 and 11.1%, respectively. Specificity and sensitivity of HRM method were 90% and 100%, with respect to PCR-RFLP. Also, HRM analysis has been evaluated as a faster and more effective approach. Conclusion: Comparison of our results based on HRM analysis with PCR-RFLP showed that our developed method is rapid, accurate, fast and economic to study the CYP2C19*17 allele and it is appropriate for other similar population genetic studies. 193 199 Zahra Ghasemi Department of Genetics, Tehran Medical Sciences Branch, Islamic Azad University Department of Genetics, Tehran Medical Sciences Branch, Islamic Azad University Iran Mehrdad Hashemi Department of Genetics, Tehran Medical Sciences Branch, Islamic Azad University Department of Genetics, Tehran Medical Sciences Branch, Islamic Azad University Iran Mahsa Ejabati Department of Biotechnology, Faculty of Pharmacy, Zanjan University of Medical Sciences Department of Biotechnology, Faculty of Pharmacy, Zanjan University of Medical Sciences Iran Seyyed Meisam Ebrahimi Department of Medical Surgical Nursing, Abhar Faculty of Nursing, Zanjan University of Medical Sciences Department of Medical Surgical Nursing, Abhar Faculty of Nursing, Zanjan University of Medical Sciences Iran Hamidreza Kheiri Manjili Department of Biotechnology, Faculty of Pharmacy, Zanjan University of Medical Sciences Department of Biotechnology, Faculty of Pharmacy, Zanjan University of Medical Sciences Iran Ali Sharafi Department of Biotechnology, Faculty of Pharmacy, Zanjan University of Medical Sciences Department of Biotechnology, Faculty of Pharmacy, Zanjan University of Medical Sciences Iran Ali Ramazani Department of Biotechnology, Faculty of Pharmacy, Zanjan University of Medical Sciences Department of Biotechnology, Faculty of Pharmacy, Zanjan University of Medical Sciences Iran Cytochrome P-450 CYP2C19PharmacogeneticsReal time polymerase chain reaction 257.pdf Meyer UA. 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Lack of Association between Interleukin-12 Gene Polymorphisms and Recurrent Aphthous Stomatitis 2 2 No Abstract 200 201 Isaac Firouze Moqadam Molecular Medicine Research Center, Hormozgan University of Medical Sciences Molecular Medicine Research Center, Hormozgan University of Medical Sciences Iran Shamsolmoulouk Najafi Dental Research Center, Faculty of Dentistry, Tehran University of Medical Sciences Dental Research Center, Faculty of Dentistry, Tehran University of Medical Sciences Iran Mahsa Mohammadzadeh Orthodontic Department, Dental Branch, Islamic Azad University Orthodontic Department, Dental Branch, Islamic Azad University Iran Alireza Zare Bidoki Molecular Immunology Research Center, Tehran University of Medical Sciences Molecular Immunology Research Center, Tehran University of Medical Sciences Iran Hila Yousefi Nature of Gene Mutation Group (NGMG), Universal Scientific Education and Research Network (USERN)Department of Endodontics, Dental Branch, Islamic Azad UniversityResearch Center for Immunodeficiencies, Children's Medical Center, Tehran University of Medical Sciences Nature of Gene Mutation Group (NGMG), Universal Scientific Education and Research Network (USERN)Department of Endodontics, Dental Branch, Islamic Azad UniversityResearch Center for Immunodeficiencies, Children's Medical Center, Tehran University of Medical Sciences IranIranIran Elham Farhadi Department of Hematology, Faculty of Allied Medical Sciences, Iran University of Medical Sciences Department of Hematology, Faculty of Allied Medical Sciences, Iran University of Medical Sciences Iran Arghavan Tonekaboni Dental Research Center, Faculty of Dentistry, Tehran University of Medical Sciences Dental Research Center, Faculty of Dentistry, Tehran University of Medical Sciences Iran Ghasem Meighani Department of Pediatrics, Faculty of Dentistry, Tehran University of Medical Sciences Department of Pediatrics, Faculty of Dentistry, Tehran University of Medical Sciences Iran Mohsen Mohammadzadeh Department of Ophthalmology, Khatam Hospital, Mashhad University of Medical Sciences Department of Ophthalmology, Khatam Hospital, Mashhad University of Medical Sciences Iran Ali Akbar Amirzargar Molecular Immunology Research Center, Tehran University of Medical Sciences Molecular Immunology Research Center, Tehran University of Medical Sciences Iran Nima Rezaei Research Center for Immunodeficiencies, Children's Medical Center, Tehran University of Medical SciencesNetwork of Immunity in Infection, Malignancy and Autoimmunity (NIIMA), Universal Scientific Education and Research Network (USERN) Research Center for Immunodeficiencies, Children's Medical Center, Tehran University of Medical SciencesNetwork of Immunity in Infection, Malignancy and Autoimmunity (NIIMA), Universal Scientific Education and Research Network (USERN) IranUSA Interleukin 12Recurrent aphthous stomatitisSingle nucleotide polymorphisms 258.pdf Shulman JD. An exploration of point, annual, and lifetime prevalence in characterizing recurrent aphthous stomatitis in USA children and youths. J Oral Pathol Med 2004;33(9):558-566.##Jurge S, Kuffer R, Scully C, Porter SR. Mucosal disease series. Number VI. Recurrent aphthous stomatitis. Oral Dis 2006;12(1):1-21.##Chavan M, Jain H, Diwan N, Khedkar S, Shete A, Durkar S. Recurrent aphthous stomatitis: a review. J Oral Pathol Med 2012;41(8):577-583.##D'Elios MM, Benagiano M, Della Bella C, Amedei A. T-cell response to bacterial agents. J Infect Dev Ctries 2011;5(9):640-645.##Ship JA, Chavez EM, Doerr PA, Henson BS, Sarmadi M. Recurrent aphthous stomatitis. Quintessence Int 2000;31(2):95-112.##Amirzargar AA, Naroueynejad M, Khosravi F, Dianat SS, Rezaei N, Mytilineos J, et al. Cytokine single nucleotide polymorphisms in Iranian populations. Eur Cytokine Netw 2008;19(2):104-112.##Najafi S, Firooze Moqadam I, Mohammadzadeh M, Bidoki AZ, Yousefi H, Farhadi E, et al. Interleukin-10 gene polymorphisms in recurrent aphthous stomatitis. Immunol Invest 2014;43(4):405-409.##Najafi S, Yousefi H, Mohammadzadeh M, Bidoki AZ, Firouze Moqadam I, Farhadi E, et al. Association study of interleukin-1 family and interleukin-6 gene single nucleotide polymorphisms in recurrent aphthous stomatitis. Int J Immunogenet 2015;42(6):428-431##Bazrafshani MR, Hajeer AH, Ollier WE, Thornhill MH. Polymorphisms in the IL-10 and IL-12 gene cluster and risk of developing recurrent aphthous stomatitis. Oral Dis 2003;9(6):287-291.##Huang D, Cancilla MR, Morahan G. Complete primary structure, chromosomal localisation, and definition of polymorphisms of the gene encoding the human interleukin-12 p40 subunit. Genes Immun 2000;1(8):515-520.##