Modares Journal of Biotechnology

Abstract Rapid and sensitive diagnosis of breast cancer, especially in the early stages of its formation, is very important. One of the methods of detecting cancer cells is the use of electrochemical sensors. Here, a new nanocomposite including an organic metal framework and silver nanoclusters are used. The resulting nanocomposite can be used as a scaffold to attach antibodies for the detection of HER2-positive cells. In the final nanocomposite structure, silver nanoclusters are placed in the internal cavities of the metal-organic framework, leading to strong electron transport, good biocompatibility, and high electrochemical activity. Our results showed that the designed electrochemical sensor has a high sensitivity in identifying HER2 positive cells, with a detection limit of 3 cells and a linear range of 100 to 5000 cells/ml. Also, the investigations showed that the introduced sensor has stability, good selectivity and acceptable application. The proposed strategy for the development of sensors based on metal-organic frameworks provides a promising approach for early detection of cancer markers and living cancer cells.

Abstract Abstract
The wound healing process is a complex and dynamic process that involves many metabolic pathways. This process consists of three phases inflammation, cell proliferation, and tissue regeneration stages. Successful wound healing depends on careful regulation and coordination between the factors involved. Until recent years, the strategy of treating chronic wounds was limited to wound preparation, removal of necrotic tissue, and control of infection and inflammation, but recently the use of growth factors has been approved to accelerate the healing process and heal the wound. Human recombinant growth factor PDGF-BB is one of the first types of recombinant growth factors approved in treating diabetic wounds. Several studies have reported that PDGF is an important mediator in wound healing that helps to accelerate healing, improve inflammation, cell proliferation, angiogenesis, and tissue regeneration. In this study, the human PDGF-B gene sequence was inserted into the pET 21a (+) expression vector for cloning and then inserted under the T7 promoter for its expression in the E. coli shuffle host. Purification was done using a nickel agarose column and to check the activity of the purified protein, cell proliferation, migration, and interaction were checked. The results of this study showed that the dimer type of PDGF expressed and purified in the bacterial host, probably due to maintaining the correct folded structure, has both main activities, i.e., cell proliferation due to the active binding to the cell receptor, as well as the ability to bind to fibrinogen.

Abstract Immune checkpoints are molecules that regulators the immune system. However, some tumor cells can express the ligands of immune checkpoints to escape from antitumor immune responses. Some agents, such as antibodies, can inhibit these checkpoints that prevent the immune system from targeting and killing cancer cells. The aim of this study was to express a novel bispecific diabody in periplasmic space of E.coli for simultaneous targeting of two immune checkpoints, cytotoxic T‑lymphocyte‑associated protein 4 (CTLA‑4) and programmed death- ligand 1 (PD‑L1).
The bispecific diabody was constructed based on the variable regions gene of anti PD-L1 and anti CTLA‑4 antibodies. The optimum codon for expression in E. coli was chemically synthesized and subcloned in pET21 expression plasmid. After transformation, the effect of cultivation conditions on periplasmic expression of the protein in E. coli BL21(DE3) was evaluated. Then, the bispecific diabody was purified .
Expression of diabody with a molecular weight of 55 kDa was verified by Sodium dodecyl sulfate‑polyacrylamide gel electrophoresis and western blotting analysis. The best condition for soluble periplasmic expression was obtained to be incubation with 0.5 mM isopropyl β‑D‑1‑thiogalactopyranoside at 23°C. The protein was successfully purified using affinity chromatography with a final yield of 0.4 mg/L. The affinity of the purified protein interaction were checked by ELISA.
Recombinant Diabody protein was cloned, expressed, and purified in a bacterial system and Diabody Interaction with PDL-1 receptor conformed by Cell-Elisa.

Abstract
The green synthesis of nanoparticles is performed in a low-cost, environmentally friendly, and efficient manner. Compared to other methods of nanoparticle production, green synthesis has proven its superiority and unique benefits, eliminating the need for expensive, toxic, time-consuming, and undesirable methods. In this study, the green synthesis of silver nanoparticles using the extract of Tribulus terrestris was investigated. Tribulus terrestris is a medicinal plant used in traditional medicine to treat urinary and reproductive tract infections, especially burning, kidney stone elimination, relief of rheumatic pains, reduction of blood pressure, and stimulation of the liver. In this research, the morphology, size, and structural properties of nanoparticles were investigated using XRD, UV-visible, FT-IR, and SEM. Given the antibacterial and anticancer properties of the Tribulus terrestris extract and the importance of silver nanoparticles, the antimicrobial effects of silver nanoparticles were examined against a number of standard strains, as well as gram-negative and gram-positive bacteria. UV-visible spectroscopy revealed a peak in the 429 nm, indicating the presence of synthesized silver nanoparticles. The results of X-ray diffraction (XRD) also confirmed the formation of the crystalline structure of nanoparticles. The results of the non-growth halo diameter for Bacillus subtilis bacterium were more than that of Escherichia coli, in other words, Escherichia coli showed more resistance against synthesized nanoparticles. The results of this research show that the green synthesis of silver nanoparticles using milk thistle seed extract can be used as a suitable antibacterial agent against clinical pathogens.

Abstract Matrix metalloproteinases (MMPs) are a zinc endopeptidase family that increases the metastatic behavior of human malignant tumors. Epigallocatechin gallate (EGCG) is a major component of green tea polyphenols and is used as an MMP inhibitor in cancer treatment. This study aims to develop and optimize the loading of EGCG in the liposomal delivery system in an experimental/ computational way. In this study, nanoliposomes were prepared by passive loading and thin-film hydration method. Size, zeta potential, stability, encapsulation efficiency, and nanoliposome drug release profile were investigated. Cytotoxicity of nanoliposomes was evaluated on three breast cancer cell lines using an MTT viability assay. To investigate the EGCG-Liposome interaction, coarse-grained Molecular Dynamic simulations were carried out. The mean diameter of liposome was 73.6±6.9 nm, the surface charge was -14.6 mV and the encapsulation efficiency was 78.5±7.3%. The encapsulation of EGCG into the liposome caused a continuous release of the drug after 72 h, which also increased the potency of the drug. Due to the EGCG hydrophobic properties, the major distribution is located at the hydrophobic part of the membrane. The energy and radial distribution function results indicate the stability of liposomes. Simulation results demonstrate that the majority of the drug is surrounded by liposomes, which indicates high encapsulation efficiency and confirms the developed synthesis method. Due to the low solubility of the drug, it seems that the use of liposomal carriers to deliver and release EGCG is a suitable solution to increase the efficiency of the drug.

Abstract The COVID-19 pandemic has created a global health crisis, and developing effective treatments is essential to prevent the spread of the disease and save millions of lives. One of the key proteins involved in the replication cycle of SARS-CoV-2, the virus that causes COVID-19, is the main protease enzyme, 3CL^pro. Due to its high importance, this enzyme is the subject of molecular, structural, and clinical investigations, and efforts have been made to develop drugs that can inhibit its activity. One such drug is the chemical compound N3, which has been found to have a high inhibitory effect against 3CL^pro. However, traditional medicine perspectives on this issue have been less explored. In this research, molecular docking interaction simulation and all-atom molecular dynamics (MD) simulation were conducted to study the potential inhibitory capability of generally available 21 plant-extracted compounds against the 3CL^pro enzyme. Three compounds with the highest inhibition probability were selected from the molecular docking results and subjected to 100 ns of MD simulation to investigate their stability and structural-dynamic-energetic features. Beside the complexes stability, the results from the simulation demonstrated that, all our selected three compounds induce N3 comparable structural-dynamics characteristics to 3CL^pro and, therefore, are expected to have a similar inhibitory ability against this enzyme. Compound number 5 was found to have the most favorable binding energy and was proposed as the best plant substitute for N3. The results from this research can be directly used to design experimental research for 3CLpro enzyme inhibition, saving the time-financial cost.

Abstract Phenolic compoundes are present in nature and have gained extensive research attention because of their unique physiochemical properties and widespread industrial use. These phenolic compoundes are one of the most numerous and ubiquitous groups of plant metabolites and have many benefits for human health. However, low bioavailability of polyphenols is a big challenge in their therapeutic and nutritional effectiveness. Nanotechnology is an emerging field of science, and nanotechnological concepts have been studied for potential applications in the food and biomedical industry. Nanoparticles have specific characteristics and better functionality, thanks to their size and other physicochemical properties. Nanotechnology can overcome challenges of Phenolic compoundes and lead to improved bioavailability and targeted drug delivery and sustained release of them, while also reducing the required drug dose. Based on plant phenolic compoundes, this study reviews the chemical classification, metabolism and bioavailability of these compounds and also provides a brief description for nano-delivery systems of them to improve their potential in food and biomedical applications.

Abstract Hepatitis C virus (HCV) NS3/4A Serine protease is an important drug target for treating patients with hepatitis C virus. However, its amino acid mutations, particularly A156G, commonly lead to the rapid emergence of drug resistance. Bosiprevir, simiprevir, and viniprevir drugs approved by the FDA show distinct resistance profiles against the HCV NS3/4A protease. In order to show the behavior of each of these drugs in the interaction with the protease in the wild type and A156G mutant, molecular dynamics simulation and binding free energy calculations were performed. MMPBSA-based binding free energy calculations showed that the binding affinity of each of the drugs in the interaction with NS3/4A protease in the wild type is significantly more than the interaction with the protease in the A156G mutant state. Free energy landscape (FEL) calculations revealed that in the presence of each of the drugs, more basins of conformations are formed. We hope that our data can provide useful insights for the design of a new effective inhibitory drug for the treatment of patients with the hepatitis C virus.