Iranian Journal of Biotechnology
Volume:20 Issue: 2, Spring 2022

The immune system is directly linked to the tumors, from tumor formation to the tumor’s development and metastasis. So, the interest of scientists over the protective immunological mechanisms has increased and shown gifted strategy in cancer treatment. 

Genetic engineering and cellular immunotherapy are two different advanced molecular mechanisms to modify the immune responses and genome. Gene manipulation is the bioengineering technology that allows vectors to transfer new genetic information into the target cells. Cellular immunotherapy is an excellent strategy that connects the body’s immune system to fight cancer. 

This review described that combination of genetic engineering and cellular immunotherapy has brought the novel antitumor repressive molecules stopping the tumor tissue immune tolerance and significantly expanding  cancer therapy’s effectiveness. Usually, cell immunotherapy and genetic engineering are considered two independent  processes, and, in this review, we believe them in combinations. Here, we review these two novel approaches, and they  are both combinations in terms of technological advances and clinical experience.

One of the challenges in using stem cells to neural repair is to induce their differentiation into neurons and lack of glial formation. Mesenchymal stem cells have revealed great potential for neural reorganization and renewal by taking advantage of differentiation capabilities. Here we explored the potential use of olibanum extract in freeze-dried scaffolds for induction of stem cells differentiation. In this study, gelatin/ collagen/olibanum/ graphene oxide (GEL/COL/OL/GO) freeze-dried scaffolds were synthesized and then adult rat bone marrow mesenchymal stem cells (BMMSCs) were seeded on scaffolds. The viability of cells was evaluated using MTT test on days 1, 3 and 5. The morphology of the cells seeded on scaffolds was studied using SEM and specific protein expression detected by immunohistochemical analysis. Real-time PCR was applied to detect the expression of Chat, Pax6, Hb-9, Nestin, Islet-1, and neurofilament-H (NF-H). The data were analyzed using Tukey test and one-way ANOVA and the means difference was considered significant at P<0.05, P<0.01, and P<0.001. Showed that the pore size is increased in GEL/COL/OL/GO scaffolds compared with GO-free scaffolds and higher attachment and proliferation of BMMSCs on GEL/COL/OL /1.5% GO scaffolds compared to GEL/COL/OL/3% GO scaffolds. The cell viability results after 5 days of incubation showed the significant biocompatibility of GEL/COL/OL /1.5% GO freeze-dried scaffold. The results of immunohistochemical and PCR analysis revealed positive role of GEL/COL/OL/1.5% GO scaffolds in upregulation of neuron-specific markers. These results reveal the great potential of GEL/COL/OL/GO scaffolds for nerve regeneration. Our data suggested that both OL extract and GO can regulate the MSCs differentiation into neurons.

Epithelial-mesenchymal transition (EMT) is a biological process in embryonic development and cancer progression, and different gene families, such as HOX genes, are closely related to this process. 

Our aim in this study was to investigate the correlation between TWIST1 and EVX1 mRNA expression in ESCC patients and also examine the probable regulatory function of TWIST1 on EVX1 expression in human ESCC cell line. 

TWIST1 and EVX1 gene expression patterns were assessed in ESCC patients by relative comparative Real-time PCR then correlated with their clinical characteristics. In silico analysis of the EVX1 gene was conducted. KYSE-30 cells were transduced by a retroviral system to ectopically express TWIST1, followed by qRT-PCR to reveal the correlation between TWIST1 and EVX1 gene expression. 

The expression of TWIST1 and EVX1 was correlated to each other significantly (p=0.005) in ESCC. Of 28 patients with under/normal expression of TWIST1, 22 samples (78.57%) had over/normal expression of EVX1. TWIST1 overexpression was correlated with advanced stages of the tumor (III, IV) (P = 0.019) and lymph node metastasis. However, EVX1 under expression was associated with lymph node metastasis (p = 0.027) and invasiveness of the disease (P = 0.037) in ESCC. Furthermore, retroviral transduction enforced significant overexpression of TWIST1 in GFP-hTWIST-1 approximately 9-fold compared to GFP control cells, causing a – 8.83- fold reduction in EVX1 mRNA expression significantly. 

Our results indicated the repressive role of TWIST1 on EVX1 gene expression in ESCC. Therefore, our findings can help dissect the molecular mechanism of ESCC tumorigenesis and discover novel therapeutic targets for ESCC invasion and metastasis.

Gastric cancer (GC) is a malignancy cause associated with a high death rate in the world. Cancer stem cells (CSCs) are a rare immortal subpopulation of cells within tumors with characteristics of the ability to self-renew, initiate tumor, and differentiate into defined progenies as well as and high resistance to conventional therapies.  Despite the use of surgery and chemotherapy for GC therapy, there are no efficient therapeutic protocols for it to date. Therefore, rapid isolation of CSCs in order to therapeutic targets, especially immunotherapy is very important. Cancerous cell suspension isolated from patients with GC was cultured in the serum-free medium containing EGF, bFGF, LIF, and heparin under non-adherent culture conditions to generate spheres. Expression of mRNA level stemness transcription factors (OCT4, SOX2, SALL4, and Cripto-1), CD44 variable isoforms (CD44s, CD44v3, CD44v6, CD44V8-10) of spheroid-forming single cells compared with gastric normal tissue cells using real time PCR and molecules of CD44, CD54, and EpCAM as gastric CSC markers, and stemness factor Oct4 using flow cytometry, as well as tumorgenicity using subcutaneous injection of sphere-forming cells to nude mice were investigated. Few cancerous cells isolated from patients with GC were able to generate three-dimensional spheroid colonies in the serum-free medium containing EGF, bFGF, LIF, and heparin under non-adherent culture conditions, and form xenograft tumors in immunodeficient nude mice after subcutaneous injection. Spheroid-forming single cells upregulated stemness transcription factors OCT4, SOX2, SALL4, and Cripto-1 that are associated with pluripotency and self-renewal and CD44 isoforms (CD44s, CD44v3, CD44v6, CD44V8-10) compared with gastric normal tissue cells. Finally, molecules of CD44, CD54, and EpCAM as gastric CSC markers and stemness factor Oct4 were expressed in sphere-forming cells. We suggested that the sphere formation and tumorigenicity assays are two procedures, leading to the rapid isolation of cancer cells with certain stem-like properties in order to target CSCs using autologous dendritic cell therapy, especially in patients with advanced disease.

Cancer is a group of diseases that have received much attention in biological research because of its high mortality rate and the lack of accurate identification of its root causes. In such studies, researchers usually try to identify cancer driver genes (CDGs) that start cancer in a cell. The majority of the methods that have ever been proposed for the identification of CDGs are based on gene expression data and the concept of mutation in genomic data. Recently, using networking techniques and the concept of influence maximization, some models have been proposed to identify these genes. We aimed to construct the cancer transcriptional regulatory network and identify cancer driver genes using a network science approach without the use of mutation and genomic data. In this study, we will employ the social influence network theory to identify CDGs in the human gene regulatory network (GRN) that is based on the concept of influence and power of webpages. First, we will create GRN Networks using gene expression data and Existing nodes and edges. Next, we will implement the modified algorithm on GRN networks being studied by weighting the regulatory interaction edges using the influence spread concept. Nodes with the highest ratings will be selected as the CDGs. The results show our proposed method outperforms most of the other computational and network-based methods and show its superiority in identifying CDGs compared to many other methods. In addition, the proposed method can identify many CDGs that are overlooked by all previously published methods.  Our study demonstrated that the Google’s PageRank algorithm can be utilized and modified as a network based method for identifying cancer driver gene in transcriptional regulatory network. Furthermore, the proposed method can be considered as a complementary method to the computational-based cancer driver gene identification tools.

Newcastle disease, is one of the most important illnesses in the aviculture industry which shows a constant threat. In this case, the vaccine could be considered an important solution to prevent and control this disease. So, the development of a new and more effective vaccine against Newcastle disease is an urgent need. Immune informatics is an important field that provides insight into the experimental procedure and could facilitate the analysis of large amounts of immunological data generated by experimental research and help to design a new vaccine candidate.  This study is aimed at bioinformatics to investigate and select the most immunogenic and conserved epitopes derived from F and HN glycoproteins, which play a key role in pathogenesis and immunity. This strategy could cover a wide range of Newcastle disease viruses. For expression in both E. coli (as an injectable recombinant vaccine candidate) and maize plant (as an edible vaccine candidate) host, two constructs were designed and analyzed separately. Furthermore, the role of LTB as an effective bio-adjuvant for general eliciting of the immune system and simultaneous expressions with those two antigens was evaluated. Hence, here a multimeric recombinant protein with the abbreviation LHN2F from the highly immunogenic part of HN, F and LTB proteins were designed. The synthetic construct was analyzed based on different bioinformatics tools.  The proper immunogenicity and stability of this multimeric fusion protein have been shown by immunoinformatic methods from various servers. To confirm the function of the designed protein, the final molecule was docked to chicken MHC class I using the Pyrex-python 0.8 program. the results of Immune Epitope analysis were confirmed by the docking results between protein and receptor. ‎The results of structural and immunological computational studies proposed that the protein deduced from this novel construct could act as a vaccine candidate for Newcastle disease virus‎ control and prophylactic.

Polycyclic Aromatic Hydrocarbons (PAHs) as resistant compounds in the environment has received much attention in recent years due to their adverse effects on ecological health. Among the various methods studying the removal of PAHs, enzyme biotechnology is one of the most effective and appropriate method.  In the present study, a halophilic laccase was used for bioremoval of anthracene in the presence of 1-Hydroxybenzotriazole (HBT). Halophilic laccase from Alkalibacillus salilacus was tested for anthracene degradation. Residual concentration of anthracene at various concentrations of NaCl (0‒4 M), incubation time, pH, solvent, and surfactants in the enzymatic reaction mixtures was determined by HPLC. The maximum removal of substrate was achieved after 72 h at 40 °C, pH 8, and NaCl concentration 1.5 M. Besides, the addition of 1% (v/v) ionic and non-ionic surfactants and 25% (v/v) of various organic solvents increased removal efficiency. The kinetic parameters Km and Vmax of the laccase for removing of anthracene were 0.114 μM and 0.546 μmoL. h.−1 mg−1, respectively.  Laccase showed the maximum removal efficiency of anthracene in the presence of 1-Hydroxybenzotriazole (HBT).

Liposomes, as a biological membrane, is successfully used for drug delivery, reduces toxicity in normal cells and improves bio-accessibility of the drug to the target cells. Curcumin, as a bioactive substance with pleiotropic biological activities, is an anti-inflammatory compound and has several anticancer effects in different cancers such as pancreatic and breast cancer.  This study was conducted to determine the bio-distribution of arginine-glycine-aspartic acid (RGD)-modified nanoliposomes containing curcumin in different tissues of rats.  The amount of curcumin in each tissue was examined by HPLC analysis. The distribution of liposomal Hoechst in the rats was evaluated by using fluorescence spectrophotometry, live animal imaging analyses and histological methods.  HPLC analysis showed the mean of curcumin in the blood significantly increased in the liposomal curcumin modified with RGD compared to free curcumin. These results were confirmed by fluorescence measurement for RGD modified liposome containing Hoechst dye. There was negligible fluorescent intensity in the blood rats, which received Hoechst alone. Live animal imaging analysis showed the presence of fluorescent color in heart tissue for all groups. It was also detected in kidney tissue for liposomal Hoechst modified with RGD group.  The present study demonstrated that RGD-modified nano-liposomes can significantly improve drug retention time in the blood of rats.

Glycinebetaine (GB) accumulation in many halophytic plants, animals, and microorganisms confers abiotic stress tolerance to salinity, drought, and extreme temperatures. Although there are a few genetic and biochemical pathways to synthesize GB, but isolation of a single gene choline oxidase (codA) from Arthrobacter spp. have opened a new hatch to engineer the susceptible plants.  The effects of overexpressed codA gene, through multiple copy insertion and GB accumulation on salinity tolerance in rice were studied.  Seed-derived embryogenic calli of ‘Tarom Molaie’ cultivar were targeted with two plasmids pChlCOD and pCytCOD both harboring the codA gene using the biolistic mediated transformation. The regenerated T0plants were screened by PCR analysis. A line containing three copies of codA gene and harboring pChlCOD and pCytCOD was identified by Southern blot analysis. The expression of codA gene in this transgenic line was then confirmed by RTPCR. The Mendelian segregation pattern of the inserted sequences was accomplished by the progeny test using PCR. The effects of overexpression of codA on salinity tolerance were evaluated at germination and seedling stage using T2-pChl transgenic line and control seeds in the presence of 0, 100, 200, and 300 mM NaCl. Finally, leaf growth dynamics of T2-pChlCOD transgenic line and control line under hydroponic conditions in the presence of 0, 40, 80, and 120 mM NaCl were assessed. The seed germination experiment results showed that the transformed seeds had a higher germination rate than the controls under all salinity treatments. But also, the leaf growth dynamics showed that the control plants had a more favorable leaf growth dynamic in all of the treatments. Although, the transgenic lines (T0, T1 and T2) exhibited lower performance than the wild type, the transgenic line varied for GB and choline contents and increasing codA gene copy number led to increased GB content.  In a salinity sensitive crop such as rice, GB may not significantly contribute to the plant protection against salt stress. Also, insufficiency of choline resources as GB precursor might have affected the overall growth ability of the transgenic line and resulted in decreased leaf growth dynamics.

Intestinal metaplasia (IM) is a benign lesion with no serious concern for patients’ health. On the other hand, gastric cancer (GC) is a malignant lesion that has to be differentially diagnosed from benign intestinal metaplasia. Epigenetic modifications have been suggested to play an important role in cancer initiation and development, and they have been investigated as a reliable biomarker tool even for early cancer diagnosis. Whole blood leucocytes (WBC) are potentially the most accessible tissue for cancer early diagnosis, especially for GC, which is hard to diagnose in the early stage.

This study aims to investigate the methylation status of DOK7 gene CpG island in blood leukocytes of patients with IM and GC compared to normal control groups. 

DNA was extracted from the whole blood of 30 IM patients, 30 GC patients, and 34 normal controls samples, and MSRE-PCR was utilized to evaluate the loci methylation status.

Significant hypermethylation of DOK7 gene CpG has been observed in GC 88.1 % (p < 0.001) and IM 66.0 % (p = 0.03) in comparison to the normal control group 56.8%. A cut‑off upper than 84.5 % of hypermethylation is considered as a presence of gastric cancer malignant lesions.

This is the first reported on hypermethylation in DOK7 CPG in blood leukocytes of patients with GC and IM and establishing a laboratory blood based test that may be useful as a novel biomarker test in the early diagnosis and screaning of GC and IM.