COVID-19 and Medical Biotechnology 2 2 Novel coronavirus disease (COVID-19) pandemic, caused by Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2), became a global challenge 1. The disease which emerged in Wuhan, China, late 2019, has affected more than 6 million individuals in almost all countries and regions, leading to death in more than 350,000 only in a 6-month period (Date: June 1st, 2020). It should be mentioned that SARS-CoV-2 is responsible for the 3rd respiratory syndrome, caused by coronaviruses during last two decades, while SARS-CoV and Middle East Respiratory Syndrome coronavirus (MERS-CoV) were both connected to the emergence of severe respiratory syndromes in 2003 and 2012, respectively 2. Meanwhile there is no effective treatment or vaccine for the disease. Although the pathogenesis of SARS-CoV-2 has not been clearly understood yet, it is a large enveloped virus, similar to other coronaviruses, which contains several proteins including M (membrane), S (spike), E (envelope), and N (nucleocapsid), which are good candidates for targeting 3. Among them, S glycoprotein, with two domains of S1 and S2, has been as of interest of recent studies, while it is responsible for invasion and entry into the host cells; the Receptor Binding Domain (RBD) of S1 interacts with Angiotensin-Converting Enzyme 2 (ACE2) on the cell surface, while the S2 domain is responsible for virus-cell membrane fusion and viral entry with higher affinity 4. Considering the fact that the immune system is affected by the SARS-CoV-2, immune-based treatment, including corticosteroids, monoclonal antibodies against pro-inflammatory cytokines, plasma therapy, and intravenous immunoglobulin was practiced in some patients in a few studies 5. However, the efforts should not be limited to such treatments, while novel therapeutic approaches could be considered, using medical biotechnology. Such pandemic is complex problem, which needs transdisciplinary studies. The development of medical biotechnology to produce pharmaceutical and diagnostic products is a need, which needs close collaboration with other disciplines 6. It should be emphasized that it has been clear that coronaviruses know no borders; therefore borderless solution is needed to fight COVID-19 7,8. It is to be hoped that the lessons we learned from SARS-CoV-2, help us to prevent possible pandemic in the near future. 139 139 Nima Rezaei Editorial 30422.pdf Hanaei S, Rezaei N. COVID-19: Developing from an outbreak to a pandemic. Arch Med Res 2020. Online ahead of print.##Saghazadeh A, Rezaei N. Immune-epidemiological parameters of the novel coronavirus – a perspective. Expert Rev Clin Immunol 2020;1-6. Online ahead of print.##Lotfi M, Hamblin MR, Rezaei N. COVID-19: transmission, prevention, and potential therapeutic opportunities. Clinica Chimica Acta 2020;508:254-266.##Yazdanpanah F, Hamblin MR, Rezaei N. The immune system and COVID-19: friend or foe? Life Sciences 2020. In Press.##Saghazadeh A, Rezaei N. Towards treatment planning of COVID-19: Rationale and hypothesis for the use of multiple immunosuppressive agents: Anti-antibodies, immunoglobulins, and corticosteroids. Int Immunopharmacol 2020;84(106560):1-6.##Moradian N, Ochs HD, Sedikies C, Hamblin MR, Camargo Jr CA, Alfredo Martinez J, et al. The urgent need for integrated science to fight COVID-19 pandemic and beyond. J Transl Med 2020;18:205.##Mohamed K, Rodríguez-Román E, Rahmani F, Zhang H, Ivanovska M, Makka SA, et al. Borderless collaboration is needed for COVID-19; a disease that knows no borders. Infect Control Hosp Epidemiol 2020;1-2. ##Momtazmanesh S, Ochs HD, Uddin LQ, Perc M, Routes JM, Nuno Vieira D, et al. All together to fight COVID-19. Am J Trop Med Hyg 2020;102(6):1181-1183.##

Pasteurella multocida Vaccine Candidates: A Systematic Review 2 2 Pasteurella multocida (P. multocida) is the highly contagious causative agent of a broad range of diseases in animals as well as an occasional human pathogen. Economically significant infections caused by P. multocida include avian fowl cholera, rabbit snuffles, and hemorrhagic septicemia in cattle, goats and pigs. Chemotherapy of pasteurellosis infections has some limitations, such as high cost of treatment, low efficacy, and the possibility of therapy failure due to antibiotic resistance. Prophylactic immunization offers a safe and effective preventive measure in case of zoonotic diseases. Bacterins, live attenuated and some old traditional vaccines against pasteurellosis remain in use today, beside their limitations. However, the past few years have seen significant progress in research to identify modern, effective vaccine candidates, but there is no new vaccine produced by new strategies. While scientists should struggle with a lot of aspects to design vaccine producing strategies, this review shows how pasteurellosis vaccine evolved and the limitations in its application which need to be overcome.  140 147 Saied Mostaan Department of Molecular Biology, Pasteur Institute of Iran Iran Abbas Ghasemzadeh Department of Molecular Biology, Pasteur Institute of Iran Iran Soroush Sardari Mohammad Ali Shokrgozar National Cell Bank of Iran, Pasteur Institute of Iran Iran Gholamreza Nikbakht Brujeni Department of Microbiology and Immunology, Faculty of Veterinary Medicine, University of Tehran Department of Microbiology and Immunology, Faculty of Veterinary Medicine, University of Tehran Iran Mohsen Abolhassani Parastoo Ehsani Department of Molecular Biology, Pasteur Institute of Iran Iran Mohammad Reza Asadi Karam Department of Molecular Biology, Pasteur Institute of Iran Iran Pasteurella multocidaPasteurellosisVaccines 30423.pdf Samina I, Khinich Y, Peleg BA. Vaccination of turkeys in the wattles (dewlap) with turkey meningo-encephalitis live vaccine and P. multocida killed-in-oil vaccine. Avian Pathology 1999;28(5):495-499.##Arif J, Rahman SU, Arshad M, Akhtar P. Immunopotentiation of outer membrane protein through anti-idiotype P. multocida vaccine in rabbits. Biologicals 2013;41(6):339-344.##Nimtrakul P, Atthi R, Limpeanchob N, Tiyaboonchai W. Development of P. multocida-loaded microparticles for hemorrhagic septicemia vaccine. Drug Dev Ind Pharm 2015;41(3):423-429.##Homayoon M, Tahamtan Y, Kargar M, Hosseini SMH, Ahavan Sepahy A. Pasteurella multocida inactivated with ferric chloride and adjuvanted with bacterial DNA is a potent and efficacious vaccine in Balb/c mice. J Med Microbio 2018;67(9):1383-1390.##Bierer BW, Scott WF. Comparison of attenuated live P. multocida vaccine given in the drinking water every two weeks to an injected oil-base bacterin administered to turkeys. Poult Sci 1969;48(2):520-523.##Bierer BW, Derieux WT. Immunologic response of turkeys to an avirulent P. multocida vaccine in the drinking water. Poult Sci 1972 Jul;51(4):1402-1408.##Rice JT, Dick JW, Bierer BW. Subcutaneous vaccination of chickens with a live, avirulent P. multocida, Vaccine. Poult Sci 1978;57(6):1514-1518.##Coates SR, Jensen MM, Brown ED. The response of turkeys to varying doses of live oral P. multocida vaccine. Poult Sci 1977;56(1):273-276.##Dua SK, Maheswaran SK. Studies on P. multocida. VII. dynamics and temporal development of local humoral immunity induced by a live avirulent fowl cholera vaccine protection against challenge. Avian Dis 2006;22(4):748-764. ##Singer N, Malkinson M. An avirulent live P. multocida vaccine for drinking water and aerosol administration against Turkey cholera. Avian Pathol 1979;8(4):391-399. ##Catt DM, Chengappa MM, Kadel WL, Herren CE. Preliminary studies with a live streptomycin-dependent P. multocida and Pasteurella haemolytica vaccine for the prevention of bovine pneumonic pasteurellosis. Can J Comp Med 1985;49(4):366-371. ##Prantner MM, Harmon BG, Glisson JR, Mahaffey EA. The pathogenesis of P. multocida serotype A:3,4 infection in Turkeys: A comparison of two vaccine strains and a field isolate. Avian Dis 2006;34(2):260-266.##Aubry P, Warnick LD, Guard CL, Hill BW, Witt MF. Health and performance of young dairy calves vaccinated with a modified-live Mannheimia haemolytica and P. multocida vaccine. J Am Vet Med Assoc 2001;219(12):1739-1742.##Zhang L, Tian X, Zhou F. CpG oligodeoxynucleotides augment the immune responses of piglets to swine P. multocida living vaccine in vivo. Res Vet Sci 2007;83(2):171-181.##Xie Z, Li H, Chen J, Zhang HB, Wang YY, Chen Q, et al. Shuffling of pig interleukin-2 gene and its enhancing of immunity in mice to P. multocida vaccine. Vaccine 2007;25(48):8163-8171.##Crouch CF, LaFleur R, Ramage C, Reddick D, Murray J, Donachie W, et al. Cross protection of a Mannheimia haemolytica A1 Lkt-/P. multocida ΔhyaE bovine respiratory disease vaccine against experimental challenge with Mannheimia haemolytica A6 in calves. Vaccine 2012;30(13):2320-2328.##Kim T, Son C, Lee J, Kim K. Vaccine potential of an attenuated P. multocida that expresses only the N-terminal truncated fragment of P. multocida toxin in pigs. Can J Vet Res 2012;76(1):69-71.##Harper M, Boyce JD. The myriad properties of P. multocida lipopolysaccharide. Toxins 2017;9(8):254.##Oslan SNH, Halim M, Ramle NA, Saad MZ, Tan JS, Kapri MR, et al. Improved stability of live attenuated vaccine gdhA derivative P. multocida B:2 by freeze drying method for use as animal vaccine. Cryobiology 2017;79:1-8.##Liu Q, Hu Y, Li P, Kong Q. Identification of Fur in P. multocida and the potential of its mutant as an attenuated live vaccine. Front Vet Sci 2019;6(February):1-12.##Suckow MA. Immunization of rabbits against P. multocida using a commercial swine vaccine. Lab Anim 2000;34(4):403-408.##Riising HJ, Van Empel P, Witvliet M. Protection of piglets against atrophic rhinitis by vaccinating the sow with a vaccine against P. multocida and Bordetella bronchiseptica. Vet Rec 2002;150(18):569-571.##Suckow MA, Haab RW, Miloscio LJ, Guilloud NB. Field trial of a P. multocida extract vaccine in rabbits. J Am Assoc Lab Anim Sci 2008;47(1):18-21.##Liao CM, Huang C, Hsuan SL, Chen ZW, Lee WC, Liu CI, et al. Immunogenicity and efficacy of three recombinant subunit P. multocida toxin vaccines against progressive atrophic rhinitis in pigs. Vaccine 2005;24(1):27-35. ##Hsuan SL, Liao CM, Huang C, Winton JR, Chen ZW, Lee WC, et al. Efficacy of a novel P. multocida vaccine against progressive atrophic rhinitis of swine. Vaccine 2009;27(22):2923-2929.##Al-Hasani K, Boyce J, McCarl VP, Bottomley S, Wilkie I, Adler B. Identification of novel immunogens in P. multocida. Microbial Cell Factories 2007;6:1-5.##Zhao ZZ, Zhang HB, Chen Q, Su D, Xie Z, Wang YY, et al. Promotion of immunity of mice to P. multocida and hog cholera vaccine by pig interleukin-6 gene and CpG motifs. Comp Immunol Microbiol Infect Dis 2009;32(3):191-205. ##Wu JR, Shien JH, Shieh HK, Chen CF, Chang PC. Protective immunity conferred by recombinant P. multocida lipoprotein E (PlpE). Vaccine 2007;25(21):4140-4148.##Tatum FM, Tabatabai LB, Briggs RE. Cross-protection against fowl cholera disease with the use of recombinant P. multocida FHAB2 peptides vaccine. Avian Dis 2012;56(3):589-591.##Mohd Yasin IS, Mohd Yusoff S, Mohd ZS, Abd Wahid Mohd E. Efficacy of an inactivated recombinant vaccine encoding a fimbrial protein of P. multocida B:2 against hemorrhagic septicemia in goats. Trop Anim Health Prod 2011;43(1):179-187.##Johnson TJ, Abrahante JE, Hunter SS, Hauglund M, Tatum FM, Maheswaran SK, et al. Comparative genome analysis of an avirulent and two virulent strains of avian P. multocida reveals candidate genes involved in fitness and pathogenicity. BMC Microbiol 2013;13(1):1.##Singh S, Singh VP, Cheema PS, Sandey M, Ranjan R, Kumargupta S, et al. Immune response to dna vaccine expressing transferrin binding protein a gene of P. multocida. Braz J Microbiol 2011;42(2):750-760.##Gong Q, Kong LY, Niu MF, Qin CL, Yang Y, Li X, et al. Construction of a ptfA chitosan nanoparticle DNA vaccine against P. multocida and the immune response in chickens. Veterinary J 2018;231:1-7.##Marchart J, Dropmann G, Lechleitner S, Schlapp T, Wanner G, Szostak MP, et al. P. multocida- and Pasteurella haemolytica-ghosts: new vaccine candidates. Vaccine 2003;21(25-26):3988-3997. ##Herath C, Kumar P, Singh M, Kumar D, Ramakrishnan S, Goswami TK, et al. Experimental iron-inactivated P. multocida A: 1 vaccine adjuvanted with bacterial DNA is safe and protects chickens from fowl cholera. Vaccine 2010;28(11):2284-2289. ##Ren W, Zou L, Ruan Z, Li N, Wang Y, Peng Y, et al. Dietary l-proline supplementation confers immunostimulatory effects on inactivated P. multocida vaccine immunized mice. Amino Acids 2013;45(3):555-561.##

Circulating Tumor Cells Detection in Patients with Early Breast Cancer Using MACS Immunomagnetic Flow Cytometry 2 2 Background: Circulating Tumor Cells (CTCs) detection in peripheral blood of epithelial cancer patients is an indicator of the presence of primary tumors and metastasis. The CTC phenotype detection uses epithelial markers in defining, detecting, and isolating CTCs. Circulating cell-separation technologies, with the epithelial origin, can be identified by epithelial biomarkers, with different techniques such as flow cytometry. The purpose of this study was to evaluate the expression of molecular Cytokeratins (CKs), CK7, CK8, CK18, CK19 (Pan-CK) and Epithelial Cell Adhesion Molecule (EpCAM) markers for CTC detection. Methods: The Magnetic Activated Cell Sorting (MACS) was used to identify CTCs in the blood of patients. Specific antibodies to EpCAM and Pan-CK were used and analyzed by flow cytometry. In this study, 35 blood samples of patients with breast cancer were assessed before any treatment and 35 healthy blood samples as the control were evaluated. Results: Expression of CK markers in the peripheral blood of breast cancer patients was statistically significant with p≤0.05, specifically at stages II-IV, but it was not significant in patients at stage I and healthy controls. Biomarkers expression in the blood of patients and healthy controls was assessed along with the pathologic characteristics of patients. Conclusion: CTC assessment by flow cytometry in patients with breast cancer could not only be used for detection but also can be considered as a source of specific and subjective evaluation for monitoring the therapy. Besides, the sensitivity and specificity of CTC detection were shown that could be enhanced by specific CK markers. 148 156 Nasrin Karimi Department of Molecular Biology, Pasteur Institute of Iran Iran Mana Oloomi Zahra Orafa Department of Molecular Biology, Pasteur Institute of Iran Iran Circulating tumor cellsCytokeratinsEpCAMEpithelial cell adhesion moleculeFlow cytometry 30424.pdf Bazzichi A, Guidi FV, Rindi L, Incaprera M, Garzelli C. PCR ELISA for the quantitative detection of Epstein-Barr virus genome. J Virol Methods 1998;74(1):15-20. ##Hamid N, Ahmadian A, Akbari Shaye Y. Effectiveness of cognitive behavior therapy based on religious believes on hope and quality of life in the patients suffering breast cancer. Med J Hormozgan University 2012;16(3):213-221.##Gohari MR, Moghadami fard Z, Abolghasemi J, Mohammadi M, Mokhtari P. Prognostic factors of metastases in breast cancer patients using the recurrent Andersen-Gill model. Koomesh 2013;14(4):483-489.##Oloomi M, Bouzari S, Rasaeian A, Mohagheghi MA. Evaluation of CK-19 biomarker in the peripheral blood of breast cancer patients. Iranian J Surgery 2011;19(1):9-15.##Khazan N, Ghavamzadeh A, Boyajyan A, Mkrtchyan G, Alimoghaddam K, Ghaffari SH. The detection of micrometastases in peripheral blood and bone marrow of breast cancer patients using marker (MUC2) real time PCR. Payavard 2012;6(2):89-100.##Kanwar N, Done S. Circulating Tumour Cells: Implications and Methods of Detection. IN: Dons S. Breast Cancer; Recent Advances in Biology, Imaging and Therapeutics. Shanghai; InTech.; 2011. p. 2-23.##Markou A, Strati A, Malamos NV, Georgoulias E, Lianidou S. Molecular characterization of circulating tumor cells in breast cancer by a liquid bead array hybridization assay. Clin Chem 2011;57(3):421-430.##Noori Daloii MR, Fazilaty H, Tabrizi M. Cancer metastasis, genetic and microenvironmental factors of distant tissue. Tehran Univ Med J (TUMJ) 2013; 70(11):671-683.##Graves H, Czerniecki BJ. Circulating tumor cells in breast cancer patients: an evolving role in patient prognosis and disease progression. Patholog Res Int 2011;3:621090.##Huebner H, Fasching PA, Gumbrecht W, Jud S, Rauh C, Matzas M, Paulicka P, et al. Filtration based assessment of CTCs and CellSearch® based assessment are both powerful predictors of prognosis for metastatic breast cancer patients. BMC Cancer 2018;18(1):204.##Larsson AM, Jansson S, Bendahl PO, Levin Tykjaer Jörgensen C, Loman N, Graffman C, et al. Longitudinal enumeration and cluster evaluation of circulating tumor cells improve prognostication for patients with newly diagnosed metastatic breast cancer in a prospective observational trial. Breast Cancer Res 2018;20(1):48.##Andreopoulou E, Yang LY, Rangel KM, Reuben JM, Hsu L, Krishnamurthy S, et al. Comparison of assay methods for detection of circulating tumor cells in metastatic breast cancer: AdnaGen AdnaTest BreastCancer Select/Detect™ versus Veridex CellSearch™ system. Int J Cancer 2012;130(7):1590-1597.##Kirfel J, Magin TM, Reichelt J. Keratins: a structural scaffold with emerging functions. Cell Mol Life Sci 2003;60(1):56-71.##Olszewski-Hamilton U, Buxhofer-Ausch VC, Hamilton G. Cytokeratin 18 (CK18) and CK18 Fragments for Detection of Minimal Residual Disease in Colon Cancer Patients. IN: Hamilton G. Cytokeratins;Tools in Oncology. Shanghai; InTech. 2012. p. 99-118.##Weng YR, Cui Y, Fang JY. Biological functions of cytokeratin 18 in cancer. Mol Cancer Res 2012;10(4):485-493.##Mavroudis D. Circulating cancer cells. Ann Oncol 2010;21(Suppl 7):vii95-vii100. ##Kummalue T, Suntiparpluacha M, Tongkao K, Chuangsuwanich T, Pongpruttipan T, O-charoenrat P. CK19 detection by multiplex real-time RT-PCR correlates with the presence of angiolymphatic and perineural invasion in sentinel lymph nodes of Thai breast cancer patients. J Clin Exp Pathol 2012;2(5):1-4.##Raffel A, Eisenberger CF, Cupisti K, Schott M, Baldus SE, Hoffmann I, et al. Increased EpCAM expression in malignant insulinoma: potential clinical implications. Eur J Endocrinol 2010;162(2):391-398.##Kim Y, Kim HS, Cui ZY, Lee HS, Ahn JS, Park CK, et al. Clinicopathological implications of EpCAM expression in adenocarcinoma of the lung. Anticancer Res 2009; 29(5):1817-1822.##Weissenstein U, Schumann A, Reif M, Link S, Toffol-Schmidt UD, Heusser P. Detection of circulating tumor cells in blood of metastatic breast cancer patients using a combination of cytokeratin and EpCAM antibodies. BMC Cancer 2012;12(206):1-8.##Alunni-Fabbroni M, Sandri MT. Circulating tumour cells in clinical practice: Methods of detection and possible characterization. Methods 2010;50(4):289-297. ##Mirmalek SA, Elham Kani F. Clinical application of breast cancer biology. Iran J Surgery 2009;17(1):1-17.##Chen CC, Hou MF, Wang JY, Chang TW, Lai DY, Chen YF, et al. Simultaneous detection of multiple mRNA markers CK19, CEA, c-Met, Her2/neu and hMAM with membrane array, an innovative technique with a great potential for breast cancer diagnosis. Cancer Lett 2006;240(2):279-288.##Nadal R, Lorente JA, Rosell R, Serrano MJ. Relevance of molecular characterization of circulating tumor cells in breast cancer in the era of targeted therapies. Expert Rev Mol Diagn 2013;13(3):295-307.##Wang R, Li X, Zhang H, Wang K, He J. Cell-free circulating tumor DNA analysis for breast cancer and its clinical utilization as a biomarker. Oncotarget 2017;8(43):75742-75755.##Han X, Wang J, Sun Y. Circulating tumor DNA as biomarkers for cancer detection. Genomics Proteomics Bioinformatics 2017;15(2):59-72. ##Campos-Carrillo A, Weitzel JN, Sahoo P, Rockne R, Mokhnatkin JV, Murtaza M, et al. Circulating tumor DNA as an early cancer detection tool. Pharmacol Ther 2020:107458. ##Kowalik A, Kowalewska M, Góźdź S. Current approaches for avoiding the limitations of circulating tumor cells detection methods-implications for diagnosis and treatment of patients with solid tumors. Transl Res 2017;185:58-84. ##Joosse SA, Hannemann J, Spötter J, Bauche A, Andreas A, Müller V, et al. Changes in keratin expression during metastatic progression of breast cancer: impact on the detection of circulating tumor cells. Clin Cancer Res 2012;18(4):993-1003.##Tanaka F, Yoneda K, Hasegawa S. Circulating tumor cells (CTCs) in lung cancer: current status and future perspectives. Lung Cancer (Auckl) 2010;1:77-84.##Wang L, Wang Y, Liu Y, Cheng M, Wu X, Wei H. Flow cytometric analysis of CK19 expression in the peripheral blood of breast carcinoma patients: relevance for circulating tumor cell detection. J Exp Clin Cancer Res 2009;28:57. ##Allan AL, Keeney M. Circulating tumor cell analysis: technical and statistical considerations for application to the clinic. J Oncol 2010;426218. ##Ross JS, Slodkowska EA. Circulating and disseminated tumor cells in the management of breast cancer. Am J Clin Pathol 2009;132(2):237-245. ##Hu Y, Fan L, Zheng J, Cui R, Liu W, He Y, et al. Detection of circulating tumor cells in breast cancer patients utilizing multi-parameter flow cytometry and assessment of the prognosis of patients in different CTCs levels. Cytometry A 2010;77(3):213-219.##Strati A, Kasimir-Bauer S, Markou A, Parisi C, Lianidou ES. Comparison of three molecular assays for the detection and molecular characterization of circulating tumor cells in breast cancer. Breast Cancer Res 2013;15(2):R20. ##van Bussel MTJ, Pluim D, Bol M, Beijnen JH, Schellens JHM, Brandsma D. EpCAM-based assays for epithelial tumor cell detection in cerebrospinal fluid. J Neurooncol 2018;137(1):1-10.##Oloomi M, Bouzari S, Mohagheghi MA, Khodayaran-Tehrani H. Molecular markers in peripheral blood of Iranian women with breast cancer. Cancer Microenviron 2013; 6(1):109-116.##Keyvani S, Karimi N, Orafa Z, Bouzari S, Oloomi M. Assessment of cytokeratin-19 gene expression in peripheral blood of breast cancer patients and breast cancer cell lines. Biomark Cancer 2016;8:57-63.##

The Role of Dihydropyrimidine Dehydrogenase and Thymidylate Synthase Polymorphisms in Fluoropyrimidine-Based Cancer Chemotherapy in an Iranian Population 2 2 Background: The fluoropyrimidine drug 5-Fluorouracil (5-FU) and the prodrug capecitabine have been extensively used for treatment of many types of cancer including colorectal, gastric, head and neck. Approximately, 10 to 25% of patients suffer from severe fluoropyrimidine-induced toxicity. This may lead to dose reduction and treatment discontinuation. Pharmacogenetics research could be useful for the identification of predictive markers in chemotherapy treatment. The aim of the study was to investigate the role of five genetic polymorphisms within two genes (DPYD, TYMS) in toxicity and efficacy of fluoropyrimidine-based chemotherapy. Methods: Total genomic DNA was extracted from 83 cancer patients treated with fluoropyrimidine-based chemotherapy. In this study, three polymorphisms were genotyped in dihydropyrimidine dehydrogenase gene c.1905+1 G>A (DPYD*2A;  rs3918290), c.1679 T>G (I560S; DPYD*13; rs55886062), and c.2846A>T (D949V; rs67376798) and two polymorphisms, besides the Variable Number of Tandem Repeat (VNTR) polymorphism and 6-bp insertion/deletion polymorphism in thymidylate synthase gene. The analysis of polymorphisms for rs3918290, rs55886062, rs67376798 and 6-bp insertion/deletion in TYMS was done by Polymerase Chain Reaction-restriction Fragment Length Polymorphism (PCR-RFLP) TYMS VNTR analysis. 5-FU-related toxicities such as anemia, febrile neutropenia, neurotoxicity, vomiting, nausea, and mucositis were evaluated according to NCI-CTC criteria version 4.0. T-test and chi-square were used and p-values less than 0.05 were considered statistically significant. Results: DPYD gene polymorphisms were not observed in this study. The frequency of the TYMS +6 bp allele was 40.35% and the -6 bp allele was 59.65% in this study. The frequency of VNTR 2R allele was 48.75% and 3R allele was 51.15%. Toxicity grade II diarrhea, mucositis, nausea, vomiting, and neurotoxicity was 2.2, 24.1, 15.7, 6, and 51.8%, respectively. Thymidylate synthase ins/del polymorphisms were associated with increased grade III neurotoxicity (p=0.02). Furthermore, anemia grade III was significantly associated with 2R/2R genotype (0.009). Conclusion: Thymidylate synthase gene polymorphisms may play a key role in fluoropyrimidne -based chemotherapy. Although rare DPYD polymorphisms were not observed in our study, according to large population studies, DPYD gene polymorphisms could be used as a predictive biomarker for patient treatments. 157 164 Mohammad Hadi Abbasian Department of Medical Biotechnology, National Institute of Genetic Engineering and Biotechnology (NIGEB) Iran Nafiseh Ansarinejad Department of Hematology and Oncology, Hazrat Rasool-e Akram Hospital, Iran University of Medical SciencesCancer Pharmacogenetics Research Group (CPGRG), Iran University of Medical Sciences Department of Hematology and Oncology, Hazrat Rasool-e Akram Hospital, Iran University of Medical SciencesCancer Pharmacogenetics Research Group (CPGRG), Iran University of Medical Sciences IranIran Bahareh Abbasi Cancer Pharmacogenetics Research Group (CPGRG), Iran University of Medical SciencesDepartment of Medical Genetic, Medical Biotechnology Ins., National Institute of Genetic Engineering and Biotechnology (NIGEB) Cancer Pharmacogenetics Research Group (CPGRG), Iran University of Medical Sciences IranIran Masoud Iravani Tehran Gastroenterology and Hepatology Center Iran Tayeb Ramim Department of Medicine, Faculty of Medicine, Tehran University of Medical Sciences Department of Medicine, Faculty of Medicine, Tehran University of Medical Sciences Iran Fahime Hamedi Department of Cell and Molecular Biology, Faculty of Biological Sciences, Kharazmi University Department of Cell and Molecular Biology, Faculty of Biological Sciences, Kharazmi University Iran Ali M. Ardekani 5-fluorouracilDihydropyrimidine dehydrogenaseFluoropyrimidinesPharmacogeneticsThymidylate synthase 30425.pdf de Gramont Ad, Figer A, Seymour M, Homerin M, Hmissi A, Cassidy J, et al. Leucovorin and fluorouracil with or without oxaliplatin as first-line treatment in advanced colorectal cancer. J Clin Oncol 2000;18(16):2938-2947.##Sargent D, Sobrero A, Grothey A, O'Connell MJ, Buyse M, Andre T, et al. Evidence for cure by adjuvant therapy in colon cancer: observations based on individual patient data from 20,898 patients on 18 randomized trials. J Clin Oncol 2009;27(6):872-877.##Bokemeyer C, Bondarenko I, Hartmann J, De Braud F, Schuch G, Zubel A, et al. Efficacy according to biomarker status of cetuximab plus FOLFOX-4 as first-line treatment for metastatic colorectal cancer: the OPUS study. Ann Oncol 2011;22(7):1535-1546.##Bohanes P, Rankin CJ, Blanke CD, Winder T, Ulrich CM, Smalley SR, et al. Pharmacogenetic analysis of INT 0144 trial: association of polymorphisms with survival and toxicity in rectal cancer patients treated with 5-FU and radiation. Clin Cancer Res 2015;21(7):1583-1590.##Rosmarin D, Palles C, Pagnamenta A, Kaur K, Pita G, Martin M, et al. A candidate gene study of capecitabine-related toxicity in colorectal cancer identifies new toxicity variants at DPYD and a putative role for ENOSF1 rather than TYMS. Gut 2015;64(1):111-120.##Jiang H, Shen Y. Methylene tetrahydrofolate reductase (MTHFR) gene rs1801133 C> T polymorphisms and response to 5-FU based chemotherapy in patients with colorectal cancer: a meta-analysis. Pteridines 2019;30(1):126-132.##Chen J, Ying X, Zhang L, Xiang X, Xiong J. Influence of TS and ABCB1 gene polymorphisms on survival outcomes of 5‑FU-based chemotherapy in a Chinese population of advanced gastric cancer patients. Wiener klinische Wochenschrift 2017;129(11-12):420-426.##Liu D, Li X, Li X, Zhang M, Zhang J, Hou D, et al. CDA and MTHFR polymorphisms are associated with clinical outcomes in gastroenteric cancer patients treated with capecitabine-based chemotherapy. Cancer Chemother Pharmacol 2019;83(5):939-949.##Longley DB, Harkin DP, Johnston PG. 5-fluorouracil: mechanisms of action and clinical strategies. Nat Rev Cancer 2003;3(5):330-338.##Lee AM, Shi Q, Pavey E, Alberts SR, Sargent DJ, Sinicrope FA, et al. DPYD variants as predictors of 5-fluorouracil toxicity in adjuvant colon cancer treatment (NCCTG N0147). J Natl Cancer Inst 2014;106(12). pii: dju298.##Boisdron-Celle M, Remaud G, Traore S, Poirier A, Gamelin L, Morel A, et al. 5-Fluorouracil-related severe toxicity: a comparison of different methods for the pretherapeutic detection of dihydropyrimidine dehydrogenase deficiency. Cancer Lett 2007;249(2):271-282.##Morel A, Boisdron-Celle M, Fey L, Soulie P, Craipeau MC, Traore S, et al. Clinical relevance of different dihydropyrimidine dehydrogenase gene single nucleotide polymorphisms on 5-fluorouracil tolerance. Mol Cancer Ther 2006;5(11):2895-2904. ##Schwab M, Zanger UM, Marx C, Schaeffeler E, Klein K, Dippon Jr, et al. Role of genetic and nongenetic factors for fluorouracil treatment-related severe toxicity: a prospective clinical trial by the German 5-FU Toxicity Study Group. J Clin Oncol 2008;26(13):2131-2138.##Offer SM, Lee AM, Mattison LK, Fossum C, Wegner NJ, Diasio RB. A DPYD variant (Y186C) in individuals of African ancestry is associated with reduced DPD enzyme activity. Clin Pharmacol Ther 2013;94(1):158-166. ##Johnson MR, Wang K, Diasio RB. Profound dihydropyrimidine dehydrogenase deficiency resulting from a novel compound heterozygote genotype. Clin Cancer Res 2002;8(3):768-774.##Caudle KE, Thorn CF, Klein TE, Swen JJ, McLeod HL, Diasio RB, et al. 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Clin Cancer Res 2001;7(12):4096-4101. ##Pullarkat S, Stoehlmacher J, Ghaderi V, Xiong Y, Ingles S, Sherrod A, et al. Thymidylate synthase gene polymorphism determines response and toxicity of 5-FU chemotherapy. Pharmacogenomics J 2001;1(1):65-70. ##Mandola MV, Stoehlmacher J, Zhang W, Groshen S, Yu MC, Iqbal S, et al. A 6 bp polymorphism in the thymidylate synthase gene causes message instability and is associated with decreased intratumoral TS mRNA levels. Pharmacogenetics 2004;14(5):319-327. ##Meulendijks D, Jacobs BA, Aliev A, Pluim D, Van Werkhoven E, Deenen MJ, et al. Increased risk of severe fluoropyrimidine‐associated toxicity in patients carrying a G to C substitution in the first 28‐bp tandem repeat of the thymidylate synthase 2R allele. Int J Cancer 2016;138(1):245-253. ##Loganayagam A, Hernandez MA, Corrigan A, Fairbanks L, Lewis C, Harper P, et al. Pharmacogenetic variants in the DPYD, TYMS, CDA and MTHFR genes are clinically significant predictors of fluoropyrimidine toxicity. Br J Cancer 2013;108(12):2505-2515. ##Health UDo, Services H. Common terminology criteria for adverse events (CTCAE) version 4.0. National Cancer Institute. 2009(09-5410).##Deenen MJ, Tol J, Burylo AM, Doodeman VD, De Boer A, Vincent A, et al. Relationship between single nucleotide polymorphisms and haplotypes in DPYD and toxicity and efficacy of capecitabine in advanced colorectal cancer. Clin Can Res 2011;17(10):3455-3468. ##Thomas F, Motsinger-Reif AA, Hoskins JM, Dvorak A, Roy S, Alyasiri A, et al. Methylenetetrahydrofolate reductase genetic polymorphisms and toxicity to 5-FU-based chemoradiation in rectal cancer. Br J Cancer 2011;105(11):1654-1662. ##Rosmarin D, Palles C, Church D, Domingo E, Jones A, Johnstone E, et al. 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Iran J Immunol 2018;15(4):321-328. ##Terrazzino S, Cargnin S, Del Re M, Danesi R, Canonico PL, Genazzani AA. DPYD IVS14+ 1G> A and 2846A> T genotyping for the prediction of severe fluoropyrimidine-related toxicity: a meta-analysis. Pharmacogenomics 2013;14(11):1255-1272. ##Deenen MJ, Meulendijks D, Cats A, Sechterberger MK, Severens JL, Boot H, et al. Upfront genotyping of DPYD* 2A to individualize fluoropyrimidine therapy: a safety and cost analysis. J Clin Oncol 2015;34(3):227-234. ##Del Re M, Cinieri S, Michelucci A, Salvadori S, Loupakis F, Schirripa M, et al. DPYD* 6 plays an important role in fluoropyrimidine toxicity in addition to DPYD* 2A and c. 2846A> T: a comprehensive analysis in 1254 patients. Pharmacogenomics J 2019;19(6):556-563.##Van Kuilenburg A, Baars J, Meinsma R, Van Gennip A. Lethal 5-fluorouracil toxicity associated with a novel mutation in the dihydropyrimidine dehydrogenase gene. Ann Oncol 2003;14(2):341-342. ##Tsalic M, Bar-Sela G, Beny A, Visel B, Haim N. 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Monoclonal Antibody Against ROR1 Induces Apoptosis in Human Bladder Carcinoma Cells 2 2 Background: Receptor tyrosine kinase-like Orphan Receptor 1 (ROR1) is one of the promising cell surface antigens for targeting cancer cells. The aim of this study was to evaluate ROR1 cell surface expression in bladder cancer cells using a murine anti-ROR1 monoclonal antibody (mAb) called 5F1-B10 as well as investigate its potential in apoptosis induction. Methods: Expression of ROR1 in two human bladder cell lines, 5637 and EJ138, as well as a non-cancerous human cell line, Human Fetal Foreskin Fibroblast (HFFF), was examined by flow cytometry and immunocytochemistry. Immunohistochemical staining of cancer and normal bladder tissues was also performed. Results: The flow cytometry results showed that 5F1-B10 mAb could recognize ROR1 molecules in 86.1% and 45.6% of 5637 and EJ138 cells, respectively. The expression level of ROR1 was 5.49% in HFFF cells. The immunocytochemistry and immunohistochemistry staining results also confirmed the presence of ROR1 on the surface of both bladder cancer cells and tissues, respectively. The obtained data from apoptosis assay demonstrated that 5F1-B10 mAb could induce apoptosis in both 5637 and EJ138 cell lines. Conclusion: Taken together, our finding indicates the role of ROR1 in bladder cancer cell survival and suggests this receptor might be a promising target for developing novel therapeutic agents against bladder carcinoma. 165 171 Ali Ahmad Bayat Monoclonal Antibody Research Center, Avicenna Research Institute, ACECR Iran Niloufar Sadeghi Monoclonal Antibody Research Center, Avicenna Research Institute, ACECR Iran Ramina Fatemi Monoclonal Antibody Research Center, Avicenna Research Institute, ACECR Iran Mohammad Reza Nowroozi Uro-Oncology Research Center, Tehran University of Medical Sciences Uro-Oncology Research Center, Tehran University of Medical Sciences Iran Solmaz Ohadian Moghadam Uro-Oncology Research Center, Tehran University of Medical Sciences Uro-Oncology Research Center, Tehran University of Medical Sciences Iran Mohadeseh Borzuee Uro-Oncology Research Center, Tehran University of Medical Sciences Uro-Oncology Research Center, Tehran University of Medical Sciences Iran Amin Radmanesh Legal Medicine Research Center, Legal Medicine Organization Iran Mahmood Khodadoost Faculty of Traditional Medicine, Traditional Medicine and Materia Medica Research Center, Shahid Beheshti University of Medical Sciences Faculty of Traditional Medicine, Traditional Medicine and Materia Medica Research Center, Shahid Beheshti University of Medical Sciences Iran Ali reza Sarrafzadeh Department of Pathology, Khatam Al Anbia Hospital Iran Omid Zarei Cellular and Molecular Research Center, Research Institute for Health Development, Kurdistan University of Medical Sciences Cellular and Molecular Research Center, Research Institute for Health Development, Kurdistan University of Medical Sciences Iran Hodjattallah Rabbani Bladder cancerFlow cytometryMonoclonal antibodyROR1 protein 30428.pdf Bray F, Ferlay J, Soerjomataram I, Siegel RL, Torre LA, Jemal A. 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Sci Rep 2014;4:5811. ##Zheng YZ, Ma R, Zhou JK, Guo CL, Wang YS, Li ZG, et al. ROR1 is a novel prognostic biomarker in patients with lung adenocarcinoma. Sci Rep 2016;6:36447. ##Zhou JK, Zheng YZ, Liu XS, Gou Q, Ma R, Guo CL, et al. ROR1 expression as a biomarker for predicting prognosis in patients with colorectal cancer. Oncotarget 2017;8(20):32864-32872. ##Masiakowski P, Carroll RD. A novel family of cell surface receptors with tyrosine kinase-like domain. J Biol Chem 1992;267(36):26181-26190. ##Reddy UR, Phatak S, Allen C, Nycum LM, Sulman EP, White PS, et al. Localization of the human Ror1 gene (NTRKR1) to chromosome 1p31-p32 by fluorescence in situ hybridization and somatic cell hybrid analysis. Genomics 1997;41(2):283-285. ##Borcherding N, Kusner D, Liu GH, Zhang W. ROR1, an embryonic protein with an emerging role in cancer biology. Protein Cell 2014;5(7):496-502. ##Daneshmanesh AH, Hojjat-Farsangi M, Khan AS, Jeddi-Tehrani M, Akhondi MM, Bayat AA, et al. Monoclonal antibodies against ROR1 induce apoptosis of chronic lymphocytic leukemia (CLL) cells. Leukemia 2012;26(6):1348-1355. ##Daneshmanesh AH, Hojjat-Farsangi M, Moshfegh A, Khan AS, Mikaelsson E, Österborg A, Mellstedt H. The PI3K/AKT/mTOR pathway is involved in direct apoptosis of CLL cells induced by ROR1 monoclonal antibodies. Br J Haematol 2015;169(3):455-458. ##Ma X, Liu B, Yang J, Hu K. Solution structure, dynamics and function investigation of Kringle domain of human receptor tyrosine kinase-like orphan receptor 1. J Biomol Struct Dyn 2019:1-11. ##Nomi M, Oishi I, Kani S, Suzuki H, Matsuda T, Yoda A, et al. Loss of mRor1 enhances the heart and skeletal abnormalities in mRor2-deficient mice: redundant and pleiotropic functions of mRor1 and mRor2 receptor tyrosine kinases. Mol Cell Biol 2001;21(24):8329-8335. ##Yoda A, Oishi I, Minami Y. Expression and function of the Ror-family receptor tyrosine kinases during development: lessons from genetic analyses of nematodes, mice, and humans. J Recept Signal Transduct Res 2003;23(1):1-15. ##Zhang S, Chen L, Wang-Rodriguez J, Zhang L, Cui B, Frankel W, et al. The onco-embryonic antigen ROR1 is expressed by a variety of human cancers. Am J Pathol 2012;181(6):1903-1910. ##Balakrishnan A, Goodpaster T, Randolph-Habecker J, Hoffstrom BG, Jalikis FG, Koch LK, et al. Analysis of ROR1 protein expression in human cancer and normal tissues. Clin Cancer Res 2017; 23(12):3061-3071. ##Karvonen H, Niininen W, Murumägi A, Ungureanu D. Targeting ROR1 identifies new treatment strategies in hematological cancers. Biochem Soc Trans 2017;45(2):457-464. ##Henry CE, Llamosas E, Djordjevic A, Hacker NF, Ford CE. Migration and invasion is inhibited by silencing ROR1 and ROR2 in chemoresistant ovarian cancer. Oncogenesis 2016;5(5):e226. ##Henry C, Hacker N, Ford C. Silencing ROR1 and ROR2 inhibits invasion and adhesion in an organotypic model of ovarian cancer metastasis. Oncotarget 2017;8(68):112727-112738. ##Xu GL, Shen J, Xu YH, Wang WS, Ni CF. ROR1 is highly expressed in circulating tumor cells and promotes invasion of pancreatic cancer. Mol Med Rep 2018;18(6):5087-5094.##Karvonen H, Barker H, Kaleva L, Niininen W, Ungureanu D. Molecular mechanisms associated with ROR1-mediated drug resistance: Crosstalk with Hippo-YAP/TAZ and BMI-1 pathways. Cells 2019;8(8):812. ##Hojjat-Farsangi M, Ghaemimanesh F, Daneshmanesh AH, Bayat AA, Mahmoudian J, Jeddi-Tehrani M, et al. Inhibition of the receptor tyrosine kinase ROR1 by anti-ROR1 monoclonal antibodies and siRNA induced apoptosis of melanoma cells. PLoS One 2013;8(4):e61167. ##Bayat AA, Ghods R, Shabani M, Mahmoudi AR, Yeganeh O, Hassannia H, et al., Production and characterization of monoclonal antibodies against human prostate specific antigen. 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Effect of Conditioned Medium from IGF1-Induced Human Wharton’s Jelly Mesenchymal Stem Cells (IGF1-hWJMSCs-CM) on Osteoarthritis 2 2 Background: Osteoarthritis (OA) is a chronic disease that attacks joints and bones which can be caused by trauma or other joint diseases. Stem cell and Conditioned Medium (CM) of stem cells are developed for OA therapy, which is minimally invasive. It can decrease inflammation and be a replacement for knee surgery.  This study aimed to utilize human Wharton’s Jelly-Mesenchymal Stem Cells (hWJMSCs) as an alternative OA therapy. Methods: CM from hWJMSCs induced by IGF1 was collected. The OA cells model (IL1β-CHON002) culture was treated as follows: 1) with hWJMSCs-CM 15% (v/v); 2) with hWJMSCs-CM  30% (v/v); 3) with IGF1-hWJMSCs (IGF1-hWJMSCs-CM) 15% (v/v); 4) with IGF1-hWJMSCs-CM 30% (v/v). Parameters including inflammatory cytokines (IL10 and TNFα), extracellular matrix degradation (MMP3 expression), and chondrogenic marker (COL2 expression) were determined. Results: The most significant increase in COL2 chondrogenic markers was found in IL1β-CHON002 treatment using 15% CM of hWJMSCs induced with IGF1. CM of hWJMSCs can reduce inflammatory cytokines (TNFα and IL10) and matrix degradation mediator MMP3. Better result was gained from IGF1-induced hWJMSCs-CM. Conclusion: CM of IGF1-hWJMSCs reduce inflammation while repairing injured joint in the human chondrocyte OA model. 172 178 Hanna Sari Widya Kusuma Biomolecular and Biomedical Research Center, Aretha Medika, Utama, Bandung, Biomolecular and Biomedical Research Center, Aretha Medika, Utama, Bandung, Indonesia Wahyu Widowati Rimonta Febby Gunanegara Faculty of Medicine, Maranatha Christian University, Bandung Faculty of Medicine, Maranatha Christian University, Bandung Indonesia Berry Juliandi Department of Biology, Faculty of Mathematics and Natural Sciences, Bogor Agricultural University, IPB Darmaga Campus, Bogor Department of Biology, Faculty of Mathematics and Natural Sciences, Bogor Agricultural University, IPB Darmaga Campus, Bogor Indonesia Nyoman Ehrich Lister Universitas Prima Indonesia Universitas Prima Indonesia Indonesia Seila Arumwardana Biomolecular and Biomedical Research Center, Aretha Medika, Utama, Bandung Biomolecular and Biomedical Research Center, Aretha Medika, Utama, Bandung Indonesia Dewani Tediana Yusepany Biomolecular and Biomedical Research Center, Aretha Medika, Utama, Bandung Biomolecular and Biomedical Research Center, Aretha Medika, Utama, Bandung Indonesia Dwi Surya Artie Biomolecular and Biomedical Research Center, Aretha Medika, Utama, Bandung Biomolecular and Biomedical Research Center, Aretha Medika, Utama, Bandung Indonesia Enden Dea Nataya Biomolecular and Biomedical Research Center, Aretha Medika, Utama, Bandung Biomolecular and Biomedical Research Center, Aretha Medika, Utama, Bandung Indonesia Kamila Yashfa Gunawan Biomolecular and Biomedical Research Center, Aretha Medika, Utama, Bandung Biomolecular and Biomedical Research Center, Aretha Medika, Utama, Bandung Indonesia Ika Adhani Sholihah Biomolecular and Biomedical Research Center, Aretha Medika, Utama, Bandung Biomolecular and Biomedical Research Center, Aretha Medika, Utama, Bandung Indonesia Ermi Girsang Universitas Prima Indonesia Universitas Prima Indonesia Indonesia Chrismis Novalinda Ginting Universitas Prima Indonesia Universitas Prima Indonesia Indonesia Indra Bachtiar Stem Cell and Cancer Institute Stem Cell and Cancer Institute Indonesia Harry Murti Stem Cell and Cancer Institute Stem Cell and Cancer Institute Indonesia ChondrocyteIGF1OsteoarthritisProinflammatoryWharton’s jelly 30429.pdf Ahmed U, Anwar A, Savage RS, Costa ML, Mackay N, Filer A, et al. 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Conus coronatus and Conus frigidus Venom: A New Source of Conopeptides with Analgesic Activity 2 2 Background: Cone snails are a natural source of complex peptides with analgesic properties called conotoxins. These peptides are secreted in a complex venomic mixture and are predominantly smaller than 5 kDa. The present study aimed to document the analgesic activity of two species of Conus coronatus (C. coronatus) and Conus frigidus (C. frigidus) venom collected off the Iranian coast in a mouse behavioral test. Methods: Conotoxin containing fractions was extracted from the venom ducts and initially purified by column chromatography. The analgesic effect of the fractions was determined on formalin pain model and hot-plate test. Results: The results led to the identification of four fractions with analgesic activity in C. coronatus and two in C. frigidus. Only one fraction was able to reduce the flinching and licking in both acute pain and chronic pain phases of the formalin test. Moreover, the activity of this fraction remained 30 minutes on the hot-plate test. Purification of the fractions was carried out by RP-HPLC. LC-ESI-MS analysis of the fractions showed that the conotoxins of the analgesic fraction had molecular weights not previously reported. Conclusion: The findings give insight into the venom of two previously under-investigated Conus species and reveal the therapeutic potential of the containing conopeptides. 179 185 Halimeh Rajabi Khorramshahr University of Marine Science and Technology Khorramshahr University of Marine Science and Technology Iran Hossein Zolgharnein Khorramshahr University of Marine Science and Technology Khorramshahr University of Marine Science and Technology Iran Mohammad Taghi Ronagh Khorramshahr University of Marine Science and Technology Khorramshahr University of Marine Science and Technology Iran Jamshid Amiri Moghaddam Leibniz Institute for Natural Product Research and Infection Biology- Hans Knöll Institute Germany Max Crüsemann Institute for Pharmaceutical Biology, University of Bonn Institute for Pharmaceutical Biology, University of Bonn Germany AnalgesicsConotoxinConus frigidusPainVenoms 30430.pdf Pillandre N, Bouchet P, Duda TF, Kauferstein S, Kohn A, Olivera BM, et al. Molecular phylogeny and evolution of the cone snails. Mol Phylogenet Evol 2014;78:290-303. ##Pillandre N, Duda TF, Meyer C, Olivera BM, Bouchet P. One, four or 100 genera? A new classification of the cone snails. J Molluscan Stud 2015;81(1):1-23. ##Dobson R, Collodor M, Gilles N, Turtoi A, De Pauw E, Quinton L. Secretion and maturation of conotoxins in the venom ducts of Conus textile. Toxicon 2012;60(8):1370-1379. ##Favreau P, Benoit E, Hocking HG, Carlier L, Hoedt DD, Leipold E, et al. A novel µ-conotoxin, CnШC, exerts potent and preferential inhibition of NaV 1.2/1.4 channels and blocks neuronal nicotinic acetylcholine receptors. British J Pharm 2012;166(5):1654-1668.##Neves J, Campos A, Osorio H, Antunes A, Vitor V. Conotoxin from Cape Verde Conus crotchii. Mar Drug 2013;11(6):2203-2215. ##Rodriguez AM, Dutertre S, Lewis RJ, Mari F. Intraspecific variation in Conus purpurascens injected venom using LC/MALDI-TOF-MS and LC-ESI-Triple TOF-MS. Anal Bioanal Chem 2015;407(20):6105-6116. ##Marshall J, Kelley WP, Rubakhin SS, Bingham JP, Sweedler JV, Gilly WF. Anatomical correlates of venom production in Conus californicus. Biol Bull 2002;203(1):27-41.##Page LR. Metamorphic remodeling of a planktotrophic larva to produce the predatory feeding system of a cone snail (Mollusca, Neogastropoda). Biol Bull 2011,221(2):176-188. ##Tayo LL, Lu B, Cruz LJ, Yates JR 3rd. Proteomic analysis provides insights on venom processing in Conus textile. J Proteome Res 2010;9(5):2292-2301. ##Biass D, Violette A, Hulo N, Lisacek F, Favreau P, Stocklin R. Uncovering intense protein diversification in a cone snail venom gland using an integrative venomics approach. J Proteome Res 2015;14(2):628-638. ##Dutertre S, Jin AH, Vetter I, Hamilton B, Sunagar K, Lavergne V, et al. Evolution of separate predation and defence evoked venoms in carnivorous cone snails. Nat Commun 2014;5:1-9.##Heghinian M, Mari F. Discovery and biological characterization of conotoxin from the venom of Conus brunneus in Drosophila melanogaster. Presented for the Ph.D., Florida Atlantic University. 2014.##Bernaldez J, López, O, Licea A, Salceda E, Arellano R. O, Vega R, et al. Electrophysiological characterization of a novel small peptide from the venom of Conus californicus that targets voltage-gated neuronal Ca2+ channels. Toxicon 2013;57(1):60-6. ##Lewis RJ, Dutertre S, Vetter I, Christie MJ. Conus venom peptide pharmacology. Pharm Rev 2012;64(2):259-298. ##Tabaraki N, Shahbazzadeh D, Moradi AM, Vosughi G, Mostafavi PG. Analgesic effect of Persian Gulf Conus textile venom. Iran J Basic Med Sci 2014;17(10):793-797. ##Kumar P, Venkateshvaran K, Srivastava PP, Nayak SK, Shivaprakash SM, Chakraborty, SK. Pharmacological studies on the venom of the marine snail Conus lentiginosus Reeve 1844. Int J Fish Aqua Stud 2014;1(3):79-85. ##Kaas Q, Yu R, Jin AH, Dutertre S, Craik DJ. Conoserver: updated content, knowledge, and discovery tools in the conotoxin database. Nucleic Acids Res 2012;40(Database issue):D325-D330. ##Ellison J, McIntosh M, Olivera BM. Alpha-conotoxin ІmЏ: similar α7 nicotinic receptor antagonists act at different sites. J Biol Chem 2003;278(2):757-764. ##Jimenez EC, Olivera BM. Divergent M-and O-superfamily peptides from venom of fish-hunting Conus parius. Peptides 2010;31(9):1678-1683. ##Sarasa A, Mohammadi SA, Christie MJ. Conotoxin interactions with α9α10-nAChRs: Is the α9α10-nicotinic acetylcholine receptor an important therapeutic target for pain management? Toxins 2015;7(10):3916-3932. ##Moller C, Vanderweit N, Mari F. Comparative analysis of proteases in the injected and dissected venom of cone snail species. Toxicon 2013;65:59-67.##Violette A, Biass D, Dutertre S, Koua D, Piquemal D, Pierrat F, et al. Large-scale discovery of conotoxins and conoproteins in the injectable venom of a fish-hunting cone snail using a combined proteomic and transcriptomic approach. J Proteom 2012;75(17):5215-5225. ##Khoobdel M, Dehghani H, Tavana AM, Ghasemi M, Dakhteh SM, Hesni MA, Rezaie-Atagholipour M. Faunal data and envenomation emergency first aid of cone snails (Conus spp.) in Qeshm Island, the Persian Gulf. Asian Pac J Trop Med 2017;10(12): 1167-1171. ##Kruger NJ. The Bradford method for protein quantitation. Methods Mol Biol 1994;32:9-15. ##Green BR, Catlin P, Zhang M, Fiedler B, Bayudan W, Morrison A, et al. Conotoxins containing nonnatural backbone spacers: cladistic-based design, Chemical synthesis, and improved analgesic activity. J Biol Chem 2007;14(4):399-407. ##Han TS, Teichert RW, Olivera BM, Bulaj G. Conus Venoms-A rich source of peptide-based therapeutics. Curr Pharm Des 2008;14(24):2462-2479.##Lee S, Kim Y, Back SK, Choi HW, Lee JY, Jung HH, et al. 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Bioactivity of Bac70 Produced by Bacillus atrophaeus Strain DDBCC70 2 2 Background: Recently, using antibacterial peptides has been considered as a strategy to manage the worldwide antibiotic-resistance crisis. Screening of Dasht-Desert Bacterial Culture Collection (DDBCC) for bacteriocin or bacteriocin-like producer was aimed in this study to introduce native antibacterial agent(s). Methods: In this study, 170 isolates were examined by the cross-streak method against G+ and G- indicators. Isolates with antimicrobial activity were compared using turbidity and well diffusion tests. The candidate isolate, DDBCC70, was molecularly and biochemically characterized. Then, the production of an antibacterial agent was physicochemically optimized. The supernatant was saturated ammonium sulfate. SDS-PAGE and Thin-Layer Chromatography (TLC) analyses, cytotoxicity, and hemagglutination tests were performed. Results: First, 23 isolates were detected with antimicrobial activity against at least three of the indicator strains. DDBCC70 was distinguished with the broad-spectrum of antibacterial effects of the Cell-Free Supernatants (CFSs). The black pigments on BHI and a 98% similarity in 16S rDNA and similarity in biochemical tests confirmed the strain of DDBCC70 as Bacillus atrophaeus (B. atrophaeus). The highest amount of the antibacterial agent, Bac70, was obtained from the modified brain heart infusion medium. It was revealed that 70% ammonium sulfate-saturated Bac70 was 3.8 and 1.6 times more effective on Pseudomonas aeuroginosa (P. aeuroginosa) and Klebsiella pneumoniae (K. pneumoniae). Bac70, a >25 kDa protein and a safe compound for blood cells, neither agglutinated human erythrocyte nor lysed sheep blood. The purified bacteriocin-like molecule destroyed biofilms from P. aeruginosa and Staphylococcus aureus (S. aureus). Moreover, the fraction of Bac70 from the TLC plate showed higher inhibitory effects against K. pneumoniae. Conclusion: Based on the above-mentioned features, Bac70 is a potential alternative therapeutic agent in pharmaceutical, food preservative and biotech-related industries. 186 193 Mohammad Reza Sarjoughian Department of Biotechnology, Faculty of Biotechnology, Semnan University Department of Biotechnology, Faculty of Biotechnology, Semnan University Iran Shamsozoha Abolmaali Shakiba Darvish Alipour Astaneh Department of Biotechnology, Faculty of Biotechnology, Semnan University Department of Biotechnology, Faculty of Biotechnology, Semnan University Iran AntibiofilmBacillus atrophaeusBacteriocinKlebsiella pneumoniaePseudomonas aeruginosa 30431.pdf Silver LL. Challenges of antibacterial discovery. Clin Microbiol Rev 2011;24(1):71-109. ##Dias DA, Urban S, Roessner U. A historical overview of natural products in drug discovery. Metabolites 2012;2(2):303-336. ##Yang H, Yu J, Wei H. 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Immunogenic Potency of Formalin and Heat Inactivated E. coli O157:H7 in Mouse Model Administered by Different Routes 2 2 Background: Enterohemorrhagic Escherichia coli (E. coli) (EHEC) O157:H7 is a major foodborne pathogen causing severe disease in humans worldwide. Cattle are important reservoirs of E. coli O157:H7 and developing a specific immunity in animals would be invaluable. The administration of Whole Cell Vaccines (WCV) is a well-established method of vaccination against bacterial infections. Route of administration, inactivation and using suitable adjuvant have significant effects on the characteristics and efficacy of WCV. Methods: In the present study, an attempt was made to evaluate the immunogenic potency of heat and formalin inactivated cells administered orally and subcutaneously in mouse model by ELISA. Mice pretreated with streptomycin were used as a model to evaluate the efficacy of subcutaneous versus oral administration of the vaccine. Following immunization, mice were infected with E. coli O157:H7 and feces were monitored for shedding. Results: Both forms of inactivated cells induced immune response and hence protection against infectious diseases caused by E. coli O157:H7. However, formalin inactivated cells of E. coli O157:H7 showed superior antigenicity compared to heat inactivated cells. Subcutaneous immunization of mice with both heat and formalin inactivated E. coli O157:H7 induced significant specific levels of IgG antibodies but did not lead to significant antigen-specific IgA rise in feces, whereas oral immunization elicited significant levels of IgG antibodies with some animals developing antigen-specific IgA in feces. Conclusion: Inactivated E. coli O157:H7 is highly immunogenic and can induce protective immune responses via oral immunization. 194 200 Nasim Arshadi Department of Biology, Faculty of Basic Sciences, Shahed University Department of Biology, Faculty of Basic Sciences, Shahed University Iran Seyed Latif Mousavi Jafar Amani Applied Microbiology Research Center, Systems Biology and Poisonings Institute, Baqiyatallah University of Medical Sciences Applied Microbiology Research Center, Systems Biology and Poisonings Institute, Baqiyatallah University of Medical Sciences Iran Shahram Nazarian Department of Biology, Imam Hossein University Department of Biology, Imam Hossein University Iran Escherichia coli O157:H7FormaldehydeHot temperatureImmunizationMiceVaccines 30432.pdf Cai K, Tu W, Liu Y, Li T, Wang H. Novel fusion antigen displayed-bacterial ghosts vaccine candidate against infection of Escherichia coli O157: H7. Sci Rep 2015;5:17479. ##Mohawk KL, Melton-Celsa AR, Zangari T, Carroll EE, O'Brien AD. Pathogenesis of Escherichia coli O157: H7 strain 86-24 following oral infection of BALB/c mice with an intact commensal flora. Microb Pathog 2010;48(3-4):131-142.##McNeilly TN, Mitchell MC, Rosser T, McAteer S, Low JC, Smith DG, et al. Immunization of cattle with a combination of purified intimin-531, EspA and Tir significantly reduces shedding of Escherichia coli O157: H7 following oral challenge. Vaccine 2010;28(5):1422-1428. ##Garcia-Angulo VA, Kalita A, Torres AG. Advances in the development of enterohemorrhagic Escherichia coli vaccines using murine models of infection. Vaccine 2013;31(32):3229-3235. ##Yousif A, Al-Taai N, Mahmood N. Humoral and cellular immune response induced by E. coli [O157: H7 and O157: H7: K99] vaccines in mice. Int J Immunol Res 2013;3(1):17.##Saeedi P, Yazdanparast M, Behzadi E, Salmanian AH, Mousavi SL, Nazarian S, et al. 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CRISPR/Cas as a Potential Diagnosis Technique for COVID-19 2 2 Coronaviruses are positive single stranded RNA viruses, and are members of Coronaviridae family. Coronaviruses localize in respiratory tract and are usually known as common cold viruses 1,2. Seven strains of coronavirus family can infect humans and can cause different signs ranging from cold with major symptoms such as fever and sore throat to upper and lower respiratory tract infections resulting in  pneumonia, severe respiratory tract infection and even death.  These seven strains include HCoV-229E, HCoV-OC43, SARS-Co- V, human coronavirus NL63, human coronavirus HK-U1, MERS-CoV and SARS-CoV-2, known as 2019-nCoV or "novel corona virus 2019" 3. At present, "severe acute respiratory syndrome coronavirus 2"  or "coronavirus disease 2019" (COVI D-19) which is closely related to SARS has become a global health problem. The first-ever COVID-19 case was identified in December 2019 in Wuhan, China; however, since then the virus has spread rapidly across the world and has become a worldwide pandemic and an international concern 4. To the best of our knowledge until March 2020, COVID-19 has been reported in 161 countries. COVID-19 is typically transmitted by respiratory droplets during sneezing and coughing 5. There is no evidence of vertical transmission or transmission during pregnancy 6,7. Incubation period of COVID-19 is estimated between 2-14 days and during this time, infected peoples are considered as asymptomatic carriers. Although infection may be asymptomatic, patients typically have fever, cough and shortness of breath. Occasionally disease progresses acutely and causes severe pneumonia, multiple organ failure and death 8. Patients with underlying medical conditions such as heart and respiratory diseases, asthma, diabetes and immunodeficiency diseases, in addition to elderly age group are high risk and more susceptible to COVID-19 infection 9. At present, there is no certain treatment or vaccination for prevention of COVID-19 and infected people are either isolated or, in critical conditions, take nonspecific or supportive care 9. Diagnosis is made based on the symptoms of the disease, chest CT (Computed tomography scan) scan and qRT-PCR (Quantitative reverse transcription polymerase chain reaction). qRT-PCR technique is the current COVID-19 (SARS-CoV-2) gold standard molecular detection method approved by CDC and World Health Organization (WHO) 10,11.  Recently, researchers have proposed a coronavirus rapid detection method based on CRISPR/Cas system 12. CRISPR/Cas (Clustered Regularly Interspaced Short Palindromic Repeats) is an adaptive immune system in archaea and bacteria against foreign genetic elements such as phages. Recently, CRISPR/Cas has become a powerful gene editing tool  and a promising treatment for genetic diseases and cancers 13,14. In this technique, a programmable protein attaches to the target site by a guide RNA for cleavage of the target sequence. There are several types of  Cas proteins that have different properties. Among them, Cas9 protein has received more attention for gene editing whereas, Cas12a and Cas13a have been more efficient in diagnosis of diseases 15,16. Cas12a is DNA-specific but Cas13a works with RNA which makes it convenient in detection of SARS-CoV-2. Recently, Zhang et al reported specific high-sensitivity enzymatic reporter unlocking (SHERLOCK) technology which is a CRISPR/Cas13 based nucleic acid detection technique for rapid detection of SARS-CoV-2 17,18. They targeted S and ORF1ab protein genes in coronavirus genome. Cas13 identifies and binds to previously determined target sequence which leads to fairly random cleavage of surrounding ssRNA molecules. SHERLOCK technology utilizes a quenched fluorescent ssRNA reporter. The presence of ssRNA coronavirus genome in samples activates Cas13 resulting in the production of quantifiable signals. Amplification of targeted DNA or RNA by Recombinase Polymerase Amplification (RPA) or reverse transcriptase-RPA (RT-RPA) prior to the start of reaction improves the sensitivity of the assay. Subsequently, amplified DNA is converted to RNA by combination of  RPA and T7 transcription. Ultimately, detection is made by simultaneous incorporation of the ssRNA reporter (Biotin-RNA-FITC). Viral genome is detected at attomolar concentration using SHERLOCK technology 19. The test can be carried out starting with RNA purified from patient samples, as used for qRT-PCR assays, and can be read out using a dipstick in less than an hour, without requiring elaborate instrumentation 18. As a result, application of CRISPR/Cas13 based diagnosis or SHERLOCK for SARS-CoV-2 detection is much faster than detection by qRT-PCR and has high sensitivity. Consequently, SHERLOCK technology could swiftly replace qRT-PCR technique considering the high demand for rapid diagnosis tests in current global pandemic state of COVID-19. 201 202 Mahintaj Dara Mahdieh Talebzadeh Department of Molecular Medicine, Faculty of Advanced Medical Science and Technology, Shiraz University of Medical Sciences Department of Molecular Medicine, Faculty of Advanced Medical Science and Technology, Shiraz University of Medical Sciences Iran CoronavirusCOVID-19CRISPR/Cas9SARS-CoV-2 30433.pdf Sexton NR, Smith EC, Blanc H, Vignuzzi M, Peersen OB, Denison MR. 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Mil Med Res 2020;7(1):4. ##Shan F, Gao Y, Wang J, Shi W, Shi N, Han M, et al. Lung Infection Quantification of COVID-19 in CT Images with Deep Learning. arXiv preprint arXiv:200304655. 2020.##Broughton JP, Deng W, Fasching CL, Singh J, Chiu CY, Chen JS. A protocol for rapid detection of the 2019 novel coronavirus SARS-CoV-2 using CRISPR diagnostics: SARS-CoV-2 DETECTR. Medrxiv : the Preprint Server for Health Sciences. 2020 Mar.##Hryhorowicz M, Lipiński D, Zeyland J, Słomski R. CRISPR/Cas9 immune system as a tool for genome engineering. Arch Immunol Ther Exp (Warsz) 2017;65(3):233-240. ##Li Y, Glass Z, Huang M, Chen ZY, Xu Q. Ex vivo cell-based CRISPR/Cas9 genome editing for therapeutic applications. Biomaterials 2020;234:119711. ##Myhrvold C, Freije CA, Gootenberg JS, Abudayyeh OO, Metsky HC, Durbin AF, et al. Field-deployable viral diagnostics using CRISPR-Cas13. Science 2018;360(6387):444-448.##Li SY, Cheng QX, Wang JM, Li XY, Zhang ZL, Gao S, et al. CRISPR-Cas12a-assisted nucleic acid detection. Cell Discov 2018;4(1):20.##Metsky HC, Freije CA, Kosoko-Thoroddsen T-SF, Sabeti PC, Myhrvold C. CRISPR-based COVID-19 surveillance using a genomically-comprehensive machine learning approach. BioRxiv 2020.##Zhang F, Abudayyeh OO, Jonathan SG. A protocol for detection of COVID-19 using CRISPR diagnostics. https://www.broadinstitute.org/files/publications/special/COVID-19%20detection%20(updated).pdf.##Kellner MJ, Koob JG, Gootenberg JS, Abudayyeh OO, Zhang F. Author Correction: SHERLOCK: nucleic acid detection with CRISPR nucleases. Nat Protoc 2020;15(3):1311. ##