Publications by authors named "Mohammad Reza Akbari Eidgahi"

19 Publications

  • Page 1 of 1

Optimization of post-insertion method to conjugate Doxil with anti-CD133 monoclonal antibodies: Investigating the specific binding and cytotoxicity to colorectal cancer cells .

Saudi Pharm J 2020 Nov 12;28(11):1392-1401. Epub 2020 Sep 12.

Department of Biochemistry, Faculty of Medicine, Semnan University of Medical Sciences, Semnan, Iran.

In this paper, Doxil coupled with anti-CD133 monoclonal antibodies made by either routine or optimized post-insertion technique, were compared with respect to their size, drug leakage, release pattern and the number of antibodies conjugated per single liposome. The results demonstrated that the number of antibodies conjugated per liposome in the optimized post-insertion technique was almost two times more than those in the routine post-insertion method. However, the drug release and leakage pattern was almost similar between the two methods. Furthermore, anti-tumor activity and therapeutic efficacy of the preferred CD133-targeted Doxil with Doxil was compared in terms of their in vitro binding, uptake, internalization and cytotoxicity against HT-29 (CD133+) and CHO (CD133-) cells. Flow cytometry analyses and confocal laser scanning microscopy results exhibited a significantly higher cellular uptake, binding and internalization of CD133-targeted Doxil in CD133 cells relative to Doxil. Cytotoxicity results revealed a lower in vitro inhibitory concentration for CD133-targeted Doxil compared to Doxil. However, CHO (CD133) cells displayed a similar uptake and in vitro cytotoxicity for both CD133-Doxil and non-targeted Doxil. Therefore, the results of this study can exhibit that specific recognition and binding of antibodies with CD133 receptors on HT-29 cells can result in enhanced cellular uptake, internalization and cytotoxicity. The research suggests further investigation for in vivo studies and may offer proof-of-principle for an active targeting concept.
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http://dx.doi.org/10.1016/j.jsps.2020.09.003DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7679470PMC
November 2020

Novel Small Molecules against Two Binding Sites of Wnt2 Protein as potential Drug Candidates for Colorectal Cancer: A Structure Based Virtual Screening Approach.

Iran J Pharm Res 2020 ;19(2):160-174

Department of Molecular Medicine, Pasteur Institute of Iran, Tehran, Iran.

Wnts are the major ligands responsible for activating Wnt signaling pathway through binding to Frizzled proteins (Fzd) as the receptors. Among these ligands, Wnt2 plays the main role in the tumorigenesis of several human cancers especially colorectal cancer (). Therefore, it can be considered as a potential drug target. The aim of this study was to identify potential drug candidates against two binding sites of Wnt2. Structure-based virtual screening approaches were applied to identify compounds against binding sites of Wnt2 for inhibiting the interaction Wnt2 and Fzd receptors. The best hit compounds from molecular docking of National Cancer Institute diversity set II database were used for structural similarity search on ZINC database, obtaining large hit compounds query to perform a virtual screening and retrieving potential lead compounds. Eight lead compounds were selected while their binding affinity, binding modes interactions, and molecular dynamics simulations studies were assessed. Molecular docking studies showed that eight selected lead compounds can bind to the desired binding sites of Wnt2 in a high affinity manner. Bioavailability analysis of the selected lead compounds indicated that they possessed significant drug like properties. Thus, these lead compounds were considered as potential drug candidates for inhibiting Wnt signaling pathway through combining with the binding sites of Wnt2 and hindering the interaction of Wnt2 and Fzd receptors. Our findings suggest that Wnt2 binding sites may be a useful target for treatment for CRC fueling the future efforts for developing new compounds against Wnt signaling pathway.
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http://dx.doi.org/10.22037/ijpr.2019.15297.13037DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7667561PMC
January 2020

Optimization and High Level Production of Recombinant Synthetic Streptokinase in Using Response Surface Methodology.

Iran J Pharm Res 2019 ;18(2):961-973

Department and Biotechnology Research Center, Semnan University of Medical Sciences, Semnan, Iran.

Streptokinase (SK) is an extracellular protein comprising 414 amino acids with considerable clinical importance as a commonly used thrombolytic agent. Due to its wide spread application and clinical importance designing more efficient SK production platforms worth investigation. In this regard, a synthetic SK gene was optimized and cloned in to pET21b plasmid for periplasmic expression. Response surface methodology was used to design a total of 20 experiments for optimization of IPTG concentration, post-induction period, and cell density of induction (OD). The optimum levels of the selected parameters were successfully determined to be 0.28 mM for IPTG concentration, 9.889 H for post induction period, and 3.40768 for cell density (OD). These settings result in 4.14fold increase in SK production rate of optimum expression conditions (7663 IU/mL) in comparison to the primary expression conditions (1853 IU/mL). Achieving higher yields of SK production in shake flask could lead to more cost effective industrial production of this drug which is the ultimate aim of SK production studies.
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http://dx.doi.org/10.22037/ijpr.2019.1100636DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6706719PMC
January 2019

Dietary approaches to stop hypertension, mediterranean dietary pattern, and diabetic nephropathy in women with type 2 diabetes: A case-control study.

Clin Nutr ESPEN 2019 10 14;33:164-170. Epub 2019 Jun 14.

Department and Biotechnology Research Center, Semnan University of Medical Sciences, Semnan, Iran. Electronic address:

Background And Aims: The association between dietary habits and kidney function in patients with type 2 diabetes (T2D) has been poorly investigated. We aimed to test the relationship between adherences to the Dietary Approaches to Stop Hypertension (DASH) diet and the Mediterranean dietary pattern (Med diet) and likelihood of diabetic nephropathy (DN) in women with T2D.

Methods: In a case-control study, 105 women with T2D and DN (albumin-creatinine ratio ≥ 30 mg/g, mean age: 55.3 ± 7.0 years; diabetes duration: 7.6 ± 2.2 years), and 105 controls with T2D and without DN (mean age: 55.4 ± 7.1 years; diabetes duration: 7.6 ± 2.1 years) who attended at Kowsar diabetes clinic in Semnan, Iran were matched for age and diabetes duration. Dietary intakes were assessed using a validated 147-item semiquantitative food frequency questionnaire. The DASH and Med diet scores were calculated using the methods developed by Fung and Trichopoulou, respectively. A generalized estimating equation model was used to examine the relationship between dietary scores and odds of DN across tertiles of dietary patterns scores.

Results: Type 2 diabetic women with moderate and high Med diet scores had 62% and 86% lower odds of DN in comparison with low adherent (ORs: 0.38, 95%CI: 0.20, 0.73; and 0.14, 95%CI: 0.06, 0.33; respectively). A moderate adherence to the DASH diet was not associated with risk of DN, but a significant inverse relationship was found in those with high adherence (OR: 0.71, 95%CI: 0.57, 0.90).

Conclusions: Adherence to the DASH and Med diets was inversely and dose-dependently associated with risk of DN. Further observational studies are needed to confirm the present results.
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http://dx.doi.org/10.1016/j.clnesp.2019.05.021DOI Listing
October 2019

Epinecidin-1, a highly potent marine antimicrobial peptide with anticancer and immunomodulatory activities.

BMC Pharmacol Toxicol 2019 05 28;20(1):33. Epub 2019 May 28.

Antimicrobial Resistance Research Center, Mashhad University of Medical Sciences, Mashhad, Iran.

Background: Antibiotic-resistant pathogens are an emerging threat in this century. Epinecidin-1 is a multi-functional Antimicrobial Peptide (AMP) produced by Orange-spotted grouper (Epinephelus coioides) has been shown to have extensive potentials as an alternative for current antibiotics. Due to the huge costs for the study and the production of a new drug, if an antimicrobial peptide has other beneficial functions in addition to antimicrobial activities, it would be preferred.

Methods: In this study, properties and applications of Epinecidin-1 were investigated and addressed comprehensively. To achieve this, the Google Scholar search engine and three databases of PubMed, Scopus, and Web of Science were used.

Results: Epinecidin-1 is a cationic AMP with an alpha-helical structure. Seven functional usages of this peptide have been reported in the literature including antibacterial, antifungal, antiviral, antiprotozoal, anticancer, immunomodulatory, and wound healing properties. Moreover, this peptide has high potential to be used as an active ingredient in cleaning solutions as well as application in vaccine production.

Conclusion: Due to significant antimicrobial activities tested on bacteria such as Staphylococcus aureus and Helicobacter pylori and also wound healing properties, Epi-1 has high potential to be considered as an important candidate for the production of new drugs and treatment of various infections including diabetic foot ulcer and peptic ulcer. Moreover, adjuvant-like properties of Epi-1 make it a suitable candidate for the studies related to an adjuvant. Other attractive properties such as anticancer effects have also been reported for this peptide which encourages further studies on this peptide.
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http://dx.doi.org/10.1186/s40360-019-0309-7DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6537373PMC
May 2019

Identification of new DNA gyrase inhibitors based on bioactive compounds from streptomyces: structure-based virtual screening and molecular dynamics simulations approaches.

J Biomol Struct Dyn 2020 02 27;38(3):791-806. Epub 2019 Mar 27.

Molecular Medicine Department, Biotechnology Research Center, Pasteur Institute of Iran, Tehran, Iran.

DNA gyrase enzyme has vital role in bacterial survival and can be considered as a potential drug target. Owing to the appearance of resistance to gyrase-targeted drugs, especially fluoroquinolone, screening new compounds which bind more efficiently to the mutant binding pocket is essential. Hence, in this work, using Smina Autodock and through structure-based virtual screening of StreptomeDB, several natural products were discovered based on the SimocyclinoneD8 (SD8) binding pocket of GyrA subunit of DNA gyrase. After evaluation of binding affinity, binding modes, critical interactions and physicochemical and pharmaceutical properties, three lead compounds were selected for further analysis. Afterward 60 ns molecular dynamics simulations were performed and binding free energies were calculated by the molecular mechanics/Poisson-Boltzmann surface area method. Also, interaction of the selected lead compounds with the mutated GyrA protein was evaluated. Results indicated that all of the selected compounds could bind to the both wild-type and mutated GyrA with the binding affinities remarkably higher than SimocyclinoneD8. Interestingly, we noticed that the selected compounds comprised angucycline moiety in their structure which could sufficiently interact with GyrA and block the DNA binding pocket of DNA gyrase, . In conclusion, three DNA gyrase inhibitors were identified successfully which were highly capable of impeding DNA gyrase and can be considered as potential drug candidates for treatment of fluoroquinolone-resistant strains.Communicated by Ramaswamy H. Sarma.
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http://dx.doi.org/10.1080/07391102.2019.1588784DOI Listing
February 2020

Optimizing denaturing HPLC as a robust technique for identification of Short Tandem Repeats (STR) in forensic medicine.

J Forensic Leg Med 2019 Feb 10;61:108-114. Epub 2018 Dec 10.

Department and Biotechnology Research Center, Semnan University of Medical Sciences, Semnan, Iran. Electronic address:

Introduction: Short Tandem Repeats (STRs) are defined as short lengths of 2-7 base pairs spreading through human genome which due to their highly diverse individually distribution are widely applied for identity detection and other forensic medicine purposes. Burdening considerable costs by the conventional methods such as capillary electrophoresis, we aimed to compare concomitant usage of multiplex PCR and denaturing high-performance liquid chromatography (DHPLC) as cheap, fast, highly accurate, and more accessible methods, with capillary electrophoresis (CE) to evaluate their potential for early screening of STRs.

Materials And Methods: The present study randomly included 20 blood samples from the subjects referred to forensic medicine of Semnan, Iran. According to the size and allele frequency, we selected 8 major STR loci including CSF1PO, VWA, D18S51, TPOX, Amelogenin, FGA, SE33, and Penta D. A quad-STR multiplex PCR was performed for each locus and the PCR products were then analyzed using DHPLC machine and compared with the basic genetic properties obtained by capillary electrophoresis.

Results: By optimizing the PCR and DHPLC conditions, our findings suggest this strategy as an effective method for STR detection. The genotypes were determined using size of loci which led to comparable results with capillary electrophoresis confirming an insignificant variation in the detection of TOPX, Amelogenin, CSF1PO, and D18S5 (p = 0.331), but discrepant results for FGA and VWA loci (p = 0.002).

Conclusion: Our study proposed DHPLC method as an effective screening method to characterize TOPX, Amelogenin, CSF1PO, and D18S51 as frequently used STR loci during identity detection in forensic medicine.
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http://dx.doi.org/10.1016/j.jflm.2018.12.003DOI Listing
February 2019

Review of antimicrobial peptides with anti-Helicobacter pylori activity.

Helicobacter 2019 Feb 15;24(1):e12555. Epub 2018 Nov 15.

Antimicrobial Resistance Research Center, Mashhad University of Medical Sciences, Mashhad, Iran.

Background: The emergence of antibiotic-resistant Helicobacter pylori strains in recent years has increased the need for finding an alternative in the post-antibiotic era. One of the fields being considered for this purpose is antimicrobial peptides. The aim of this review was to provide an obvious scheme from the studied anti-H. pylori peptides and to investigate their common features.

Method: First, all of the antimicrobial peptides with their anti-H. pylori effects have been proved up to September 2018 were selected and their information including structure, mechanism of action, and function was reviewed. To achieve this, three databases of PubMed, Scopus, and Web of science were used.

Results: A total of 9 groups containing 22 antimicrobial peptides were found with demonstrated anti-H. pylori effects. The nine groups included pexiganan, tilapia piscidins, epinecidin-1, cathelicidins, defensins, bicarinalin, odorranain-HP, PGLa-AM1, and bacteriocins. Most of the antimicrobial peptides, not all, had common features such as the ability to kill antibiotic-resistant strains, having α-helical structure, being cationic, with high positive charge and isoelectric point.

Conclusion: Antimicrobial peptides with anti-H. pylori effects have the potential to replace the antibiotics, especially in the post-antibiotic era, if a rapid and low-cost production method would be found.
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http://dx.doi.org/10.1111/hel.12555DOI Listing
February 2019

Structural and dynamic characterization of human Wnt2-Fzd7 complex using computational approaches.

J Mol Model 2018 Sep 6;24(10):274. Epub 2018 Sep 6.

Molecular Medicine Department, Pasteur Institute of Iran, Tehran, Iran.

Wnt and Frizzled (Fzd) family members play crucial roles in the self-renewal of tumor-initiating cells. Until now, only a few studies have addressed the distinct mechanism of Wnt-Fzd interactions. In this study, we suggest a possible interaction mode of Wnt2 with the Fzd7 cysteine-rich domain (CRD)-both of which are up-regulated in some types of cancer. A combination of homology modeling, molecular docking and molecular dynamics (MD) simulations was carried out to study this ligand-receptor complex in great detail. The results demonstrated the unique dynamic behavior of Wnt2 upon binding to Fzd7. Interestingly, the β-strand content of the C-terminal binding site of Wnt2 was obviously reduced when bound to Fzd7 CRD. Moreover, the N-terminal and C-terminal binding sites of Wnt2 appeared to interact with the C-terminal and N-terminal binding sites of Fzd7, respectively. Calculation of the binding energies uncovered the pivotal role of electrostatic and hydrophobic interactions in the binding of Wnt2 to Fzd7 CRD. In conclusion, this study provides valuable insights into the mechanism of the Wnt2-Fzd7 CRD interaction for application in colorectal cancer prevention programs. Graphical abstract Flowchart representation of different steps used in this study.
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http://dx.doi.org/10.1007/s00894-018-3788-3DOI Listing
September 2018

Evaluation of the Effect of Promoter Type on the Immunogenicity of the Live Recombinant Vaccines Expressing Heat-labile Enterotoxins (LTB).

Iran J Pharm Res 2018 ;17(Suppl2):98-110

Department of Pharmaceutical Biotechnology, School of Pharmacy, Shahid Beheshti University of Medical Sciences, Tehran, Iran.

Enterotoxigenic (ETEC)-induced diarrhoea is the second most common cause of death in children in the developing countries. Heat labile toxin (LT) is responsible for ETEC-induced diarrhoea. In the present study, a novel live ETEC vaccine based on subunit B of LT (LTB) expression in attenuated PhoP strain was developed. Herein, we aimed to compare the activity of promoters including constitutive , IPTG inducible and -inducible (B and B78-23) in PhoP. Additionally, the ability of these recombinant PhoP/pLTBs to induce LTB-specific antibody responses in BALB/c mice after nasal immunization was evaluated. studies demonstrated that PhoP has the ability to produce rLTB. Furthermore, B promoter directed significantly more LTB expression in PhoP/pnirBLTB under anaerobic condition without induction compared to the amount of rLTB secreted by PhoP/ptrcLTB in bacterial soup under uninduced condition (6.06 ± 0.05 1.4 ± 0.46 μg/10 cfu, < 0.01). In addition, the constitutive rLTB expression from promoter was more than its expression from uninduced promoter in bacterial soup (4.2 ± 0.92 1.4 ± 0.46 (μg/10 cfu)) and pellet (27.4 ± 0.89 13.4 ± 1.42 (μg/10 cfu), < 0.0001). However, the mice immunized with PhoP/ptrcLTB elicited the superior anti-LTB responses among the PhoP containing the examined prompters, which were significantly higher than those induced by PhoP/pnirB78-23LTB and PhoP/pnirB, 6 weeks after the first immunization. Totally, it could be concluded that analysis of promoters for LTB expression in PhoP may not necessarily predict the recombinant PhoP immunogenicity.
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http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6447867PMC
January 2018

PhiC31 integrase can improve the efficiency of different construct designs for monoclonal antibody expression in CHO cells.

Protein Expr Purif 2017 Jun 9;134:89-95. Epub 2017 Apr 9.

Biotechnology Research Center, Pasteur Institute of Iran, Tehran, Iran. Electronic address:

Objectives: Several types of expression vectors have been used for recombinant protein expression in Chinese hamster ovary cells (CHO) which usually result in variable and unstable levels of expression.

Methods And Results: In this study, we have compared the mAb0014 expression level of single ORF/IRES vector and dual ORF vector in the presence and absence of phiC31 integrase targeting system. Both expression vectors contain an elongation factor 1α (EF1α) promoter upstream of LC and harboring an attB site. CHO-S cells were co-transfected with single ORF/IRES or dual ORF vectors along with a phiC31 integrase expression vector which can catalyze recombination between attB site and pseudo-attP sites presented in the mammalian genome. Our results demonstrated that dual ORF vector in the presence of phiC31 integrase expression vectors (+FC31 2P) generated more recombinant antibody in comparison to its negative control (-FC31 2P). Moreover, both of +FC31 2P and -FC31 2P cell pools yield higher recombinant protein in comparison to single ORF/IRES vector (FC31 IRES) cell pools. Stability of expression in phiC31 co-transfected cell pools (+FC31 2P and +FC31 IRES) had no considerable changes.

Conclusions: Our results indicated that the dual ORF vector using integrase can support the generation of cell lines with stable transgene expression at an elevated mAb relative to single ORF/IRES vector.
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http://dx.doi.org/10.1016/j.pep.2017.04.005DOI Listing
June 2017

Frequency of VanA, VanB and VanH variants amongst vancomycin-resistant enterococci isolated from patients in central region of Iran.

Gastroenterol Hepatol Bed Bench 2016 ;9(4):308-315

Semnan Biotechnology Research Center, Semnan University of Medical Sciences, Semnan, Iran.

Aim: The aim of this study was to investigate the VRE frequency and the rate of each gene in isolated enterococci from patients with intestinal infection in the central region of Iran.

Background: infections are a public health growing concern due to the glycopeptide antibiotics resistance especially vancomycin. Genes, , , and contribute to the influence of vancomycin-resistant enterococci (VRE).

Patients And Methods: This study was conducted from January to July 2014 in Shahrood university hospital. Enterococci isolation and its antibacterial susceptibility were performed by culturing in Aesculin Azide agar and Kirby-Bauer method, respectively. Vancomycin-resistant genes were screened through conventional PCR, and subsequently sequenced.

Results: Among 265 specimens, 100 isolates revealed enterococci, in which (91%) and (9%). The isolated enterococci were resistant to vancomycin (6%) and chloramphenicol (21%), whereas their large proportions (94% to 100%) were multi-drug resistant. All VRE isolates belonged to conversely, the were susceptible to the same antibiotic. Both and genes were identified in all VRE isolates, although, no gene was indicated. Homology analysis of sequenced amplicons verified the full length compatibility to the worldwide reported genes.

Conclusion: The present study revealed VR in gastroenteritis patients and resistance factor for and genes are coordinated. Since enterococci isolates were all multidrug resistance, increase in VR vanA / vanH in this area could be expected.
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http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5118856PMC
January 2016

Survivin, a Promising Gene for Targeted Cancer Treatment.

Asian Pac J Cancer Prev 2016 ;17(8):3711-9

Faculty of Medicine, Semnan University of Medical Sciences, Semnan, Iran E-mail : shahbazimajid@ yahoo.co.uk,

Drawbacks of conventional cancer treatments, with lack of specificity and cytotoxicity using current approaches, underlies the necessity for development of a novel approach, gene-directed cancer therapy. This has provided novel technological opportunities in vitro and in vivo. This review focuses on a member of an apoptosis inhibitor family, survivin, as a valuable target. Not only the gene but also its promoter are applicable in this context. This article is based on a literature survey, with especial attention to RNA interference as well as tumor- specific promoter action. The search engine and databases utilized were Science direct, PubMed, MEDLINE and Google. In addition to cell-cycle modulation, apoptosis inhibition, interaction in cell-signaling pathways, cancer-selective expression, survivin also may be considered as specific target through its promoter as a novel treatment for cancer. Our purpose in writing this article was to create awareness in researchers, emphasizing relation of survivin gene expression to potential cancer treatment. The principal result and major conclusion of this manuscript are that survivin structure, biological functions and applications of RNA interference systems as well as tumor-specific promoter activity are of major interest for cancer gene therapy.
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January 2017

Evaluating the efficiency of phiC31 integrase-mediated monoclonal antibody expression in CHO cells.

Biotechnol Prog 2016 11 30;32(6):1570-1576. Epub 2016 Sep 30.

Biotechnology Research Center, Pasteur Institute of Iran, Tehran, Iran.

Traditional methods to generate CHO cell lines rely on random integration(s) of the gene of interest and result in unpredictable and unstable protein expression. In comparison, site-specific recombination methods increase the recombinant protein expression by inserting transgene at a locus with specific expression features. PhiC31 serine integrase, catalyze unidirectional integration that occurs at higher frequency in comparison with the reversible integration carried out by recombinases such as Cre. In this study, using different ratios of phiC31 serine integrase, we evaluated the phiC31 mediated gene integration for expression of a humanized IgG1 antibody (mAb0014) in CHO-S cells. Light chain (LC) and heavy chain (HC) genes were expressed in one operon under EF1α promoter and linked by internal ribosome entry site (IRES) element. The clonal selection was carried out by limiting dilution. Targeted integration approach increased recombinant protein yield and stability in cell pools. The productivity of targeted cell pools was about 4 mg/L and about 40 µg/L in the control cell pool. The number of integrated transgenes was about 19 fold higher than the control cells pools. Our results confirmed that the phiC31 integrase leads to mAb expression in more than 90% of colonies. The productivity of the PhiC31 integrated cell pools was stable for three months in the absence of selection as compared with conventional transfection methods. Hence, utilizing PhiC31 integrase can increase protein titer and decrease the required time for protein expression. © 2016 American Institute of Chemical Engineers Biotechnol. Prog., 32:1570-1576, 2016.
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http://dx.doi.org/10.1002/btpr.2362DOI Listing
November 2016

Construction and immunogenicity of a new Fc-based subunit vaccine candidate against Mycobacterium tuberculosis.

Mol Biol Rep 2016 Sep 1;43(9):911-22. Epub 2016 Jun 1.

Antimicrobial Resistance Research Center, Bu-Ali Research Institute, Mashhad University of Medical Sciences, Mashhad, Iran.

As an ancient disease, tuberculosis (TB) is a major global health threat. Therefore, there is an urgent need for an effective and safe anti-TB vaccine. In the current study, a delivery system of Fc domain of mouse IgG2a and early secreted antigenic target protein 6 (ESAT-6) was evaluated for the selective uptake of antigens by antigen-presenting cells (APCs). Thus, it was based on the immunogenicity of a fusion protein. The study was initiated by the transfer of recombinant expression vectors of pPICZαA-ESAT-6:Fcγ2a and pPICZαA-ESAT-6: His into Pichia pastoris (P. pastoris). Recombinant proteins were assessed for immunogenicity following the immunoblotting analysis. High levels of IFN-γ and IL-12 were produced to induce Th1-type cellular responses through vaccination with both recombinant proteins [ESAT-6:Fcγ2a (EF) and ESAT-6:His (EH)]. The Fc-tagged recombinant protein induced more effective Th1-type cellular responses with a low increment in IL-4 compared to PBS, BCG, and EH groups. Although in all the immunized groups, the ratio of IFN-γ/IL-4 was in favor of Th1 responses, the highest Th1/Th2 balance was observed in EF immunized group. Fc fragment of mouse IgG2a may induce a selective uptake of APCs towards the cross-presentation and formation of Th1 responses in favor of an appropriate protective anti-tuberculosis reaction. Thus, further research on Fc-fusion proteins is required to develop Fc-based TB vaccines.
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http://dx.doi.org/10.1007/s11033-016-4024-9DOI Listing
September 2016

Utilization of Site-Specific Recombination in Biopharmaceutical Production.

Iran Biomed J 2016 25;20(2):68-76. Epub 2015 Nov 25.

Biotechnology Research Center, Pasteur Institute of Iran, Tehran, Iran.

Mammalian expression systems, due to their capacity in post-translational modification, are preferred systems for biopharmaceutical protein production. Several recombinant protein systems have been introduced to the market, most of which are under clinical development. In spite of significant improvements such as cell line engineering, introducing novel expression methods, gene silencing and process development, expression level is unpredictable and unstable because of the random location of integration in the genome. Site-specific recombination techniques are capable of producing stable and high producer clonal cells; therefore, they are gaining more importance in the biopharmaceutical production. Site-specific recombination methods increase the recombinant protein production by specifically inserting a vector at a locus with specific expression trait. The present review focused on the latest developments in site-specific recombination techniques, their specific features and comparisons.
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http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4726886PMC
http://dx.doi.org/10.7508/ibj.2016.02.001DOI Listing
October 2016

Construction of a Synthetically Engineered nirB Promoter for Expression of Recombinant Protein in Escherichia coli.

Jundishapur J Microbiol 2014 Jul 1;7(7):e15942. Epub 2014 Jul 1.

Semnan Biotechnology Research Center, Semnan University of Medical Sciences, Semnan, IR Iran.

Background: Anaerobic-inducible promoters are alternatives of chemical-inducible promoters for expression of recombinant proteins especially in conditions where chemical induction is not possible or anaerobic conditions are preferable. The nirB promoter is the promoter of the first gene of nir operon in Escherichia coli, which encodes NADH-dependent nitrite reductase. This promoter is naturally induced under anaerobic conditions and upregulated by nitrite and nitrate.

Objectives: The current study was carried out to construct a synthetic nirB promoter that does not respond to chemical inducers (nitrite or nitrate), but instead responds to anaerobic induction. For this purpose, a new plasmid was constructed (pFSnirB78-23LTB), which contains a synthetic nirB promoter. The activity of this plasmid was evaluated in E. coli under both aerobic and anaerobic conditions and in response to chemical inducers, nitrite and nitrate.

Materials And Methods: A synthetic nirB promoter was firstly cloned into a pKK223 derivative plasmid and then the heat labile toxin B subunit gene (LTB) of entrotoxigenic E. coli was cloned under the control of this promoter. The inducibility of this plasmid in E. coli was measured under anaerobic conditions in the presence or absence of nitrite or nitrate by ganglioside GM1 ELISA.

Results: Our data showed that this promoter is strongly induced under anaerobic conditions while it showed much lower activity (11%) under aerobic conditions. In contrast to the native promoter, this promoter was not induced by chemical inducers, nitrite or nitrate.

Conclusions: This study showed that the recombinant protein produced under the control of synthetic nirB promoter has critical characteristics such as pentamer formation, receptor recognition ability and conservation of antigenic epitopes. In addition, the data showed anaerobiosis and chemical inducers had no adverse effects on recombinant proteins. Based on the results, this synthetic promoter is suitable for use in live delivery vaccines or drug systems and for production of recombinant proteins especially oxygen sensitive proteins.
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http://dx.doi.org/10.5812/jjm.15942DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4216584PMC
July 2014

SstI Polymorphism of the Apolipoprotein CIII Gene in Iranian Hyperlipidemic Patients: A Study in Semnan Province.

Iran J Basic Med Sci 2011 Nov;14(6):506-13

Department of Biochemistry, Faculty of Medicine, Semnan University of Medical Sciences, Semnan, Iran.

Objectives: The Sst-I polymorphic site on the 3' untranslated region of the apo CIII gene, has been previously reported to be associated with hypertriglyceridemia. The aim of the present study was to explore the association between Sst-I polymorphism with plasma lipid and lipoprotein levels in hyperlipidemic (HLP) patients from Semnan province, Iran.

Materials And Methods: Genomic DNA was prepared from 76 patients with HLP and 75 matched healthy subjects. DNA samples were amplified by polymerase chain reaction. The samples were analyzed by restriction fragment length polymorphism (RFLP) method using SstI enzyme.

Results: The genotype and allelic frequencies for this polymorphism were significantly different between HLP and normolipidemic groups (P< 0.002). Plasma triglyceride (TG) level was higher in both groups, in S2S2 genotype was more than in the S1S1and S1S2 genotypes, however, there was no significant difference in comparison with the control group. Subjects with S1S2 + S2S2 genotypes in compare to S1S1 genotype had odd ratio of 2.8 (95% CI: 1.41-5.56, P< 0.003) for developing hypertriglyceridemia.

Conclusion: The results showed that the presence of rare S2 allele was associated with change in TG level in the selected population.
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http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3586853PMC
November 2011