Phage Display as a Medium for Target Therapy Based Drug Discovery, Review and Update.

Phage libraries are now amongst the most prominent approaches for the identification of high-affinity antibodies/peptides from billions of displayed phages in a specific library through the biopanning process. Due to its ability to discover potential therapeutic candidates that bind specifically to targets, phage display has gained considerable attention in targeted therapy. Using this approach, peptides with high-affinity and specificity can be identified for potential therapeutic or diagnostic use. Furthermore, phage libraries can be used to rapidly screen and identify novel antibodies to develop immunotherapeutics. The Food and Drug Administration (FDA) has approved several phage display-derived peptides and antibodies for the treatment of different diseases. In the current review, we provided a comprehensive insight into the role of phage display-derived peptides and antibodies in the treatment of different diseases including cancers, infectious diseases and neurological disorders. We also explored the applications of phage display in targeted drug delivery, gene therapy, and CAR T-cell.

Development of a novel multi­epitope vaccine against the pathogenic human polyomavirus V6/7 using reverse vaccinology.

BACKGROUND: Human polyomaviruses contribute to human oncogenesis through persistent infections, but currently there is no effective preventive measure against the malignancies caused by this virus. Therefore, the development of a safe and effective vaccine against HPyV is of high priority. METHODS: First, the proteomes of 2 polyomavirus species (HPyV6 and HPyV7) were downloaded from the NCBI database for the selection of the target proteins. The epitope identification process focused on selecting proteins that were crucial, associated with virulence, present on the surface, antigenic, non-toxic, and non-homologous with the human proteome. Then, the immunoinformatic methods were used to identify cytotoxic T-lymphocyte (CTL), helper T-lymphocyte (HTL), and B-cell epitopes from the target antigens, which could be used to create epitope-based vaccine. The physicochemical features of the designed vaccine were predicted through various online servers. The binding pattern and stability between the vaccine candidate and Toll-like receptors were analyzed through molecular docking and molecular dynamics (MD) simulation, while the immunogenicity of the designed vaccines was assessed using immune simulation. RESULTS: Online tools were utilized to forecast the most optimal epitope from the immunogenic targets, including LTAg, VP1, and VP1 antigens of HPyV6 and HPyV7. A multi-epitope vaccine was developed by combining 10 CTL, 7 HTL, and 6 LBL epitopes with suitable linkers and adjuvant. The vaccine displayed 98.35% of the world's population coverage. The 3D model of the vaccine structure revealed that the majority of residues (87.7%) were located in favored regions of the Ramachandran plot. The evaluation of molecular docking and MD simulation revealed that the constructed vaccine exhibits a strong binding (-1414.0 kcal/mol) towards the host's TLR4. Moreover, the vaccine-TLR complexes remained stable throughout the dynamic conditions present in the natural environment. The immune simulation results demonstrated that the vaccine design had the capacity to elicit robust immune responses in the host. CONCLUSION: The multi-parametric analysis revealed that the designed vaccine is capable of inducing sustained immunity against the selected polyomaviruses, although further in-vivo investigations are needed to verify its effectiveness.

Strategies to Overcome Antimicrobial Resistance in Nosocomial Infections, A Review and Update.

Nosocomial infections, also known as healthcare-associated infections, are a signif-icant global concern due to their strong association with high mortality and morbidity in both developed and developing countries. These infections are caused by a variety of pathogens, particularly the ESKAPE group of bacteria, which includes the six pathogens Enterococcus faecium, Staphylococcus aureus, Klebsiella pneumoniae, Acinetobacter baumannii, Pseudo-monas aeruginosa, and Enterobacter spp. These bacteria have demonstrated noteworthy re-sistance to different antibiotics. Antimicrobial resistance mechanisms can manifest in various forms, including restricting drug uptake, modifying drug targets, inactivating drugs, active drug efflux, and biofilm formation. Accordingly, various strategies have been developed to combat antibiotic-resistant bacteria. These strategies encompass the development of new antibiotics, the utilization of bacterio-phages that specifically target these bacteria, antimicrobial combination therapy and the use of peptides or enzymes that target the genomes or essential proteins of resistant bacteria. Among promising approaches to overcome antibiotic resistance, the CRISPR/Cas system stands out and offers many advantages. This system enables precise and efficient editing of genetic material at specific locations in the genome. Functioning as a bacterial "adaptive im-mune system," the CRISPR/Cas system recognizes, degrades, and remembers foreign DNA sequences through the use of spacer DNA segments that are transcribed into CRISPR RNAs (crRNA). This paper has focused on nosocomial infections, specifically the pathogens involved in hospi-tal infections, the mechanisms underlying bacterial resistance, and the strategies currently em-ployed to address this issue. Special emphasis has been placed on the application of CRISPR/Cas technology for overcoming antimicrobial resistance.

Female-biased association of NOS2-c.1823C>T (rs2297518) with co-susceptibility to metabolic syndrome and asthma.

The nitric oxide (NO) pathway contributes to the pathogeneses of metabolic syndrome (MetS) and asthma. NOS2 encodes inducible-NO synthase, which is an important enzyme of the pathway, and its variations could affect the risk of asthma and MetS and thereby co-susceptibility to them. This study aims to estimate the association of NOS2-c.1823C>T with risk of asthma, MetS, and asthma with MetS condition (ASMetS), and with asthma stages: intermittent, mild, moderate, and severe asthma. The study included asthmatics (n = 555), MetS (n = 334), and ASMetS cases (n = 232) and 351 controls, which were genotyped by the PCR-RFLP method. The T allele was significantly associated with an increased risk of asthma and MetS in the sample population and females. CT genotype and CT+TT model were significantly associated with increased risk of ASMetS in females. A significant association between CT genotype and increased risk of ASMetS in the sample population and females was found in ASMetS versus MetS. In the sample population and among females, the T allele was significantly associated with severe asthma. The rs2297518 single nucleotide polymorphism of NOS2 contributes to the risk of MetS, asthma, and co-susceptibility to them, and this contribution may be stronger in females compared to males.

Regulatory Effect of let-7f Transfection in Non-Small Cell Lung Cancer on its Candidate Target Genes.

Background: Let-7f has essential impacts on biological processes; however, its biological and molecular functions in lung cancer pathogenesis have yet been remained unclear. We aimed to investigate the expression level of let-7f and its candidate target genes both in lung cancer tissues and A549 cell line. Methods: Bioinformatics databases were first used to select candidate target genes of let-7f. Then the relative gene and protein expressions of let-7f and its target genes, including HMGA2, ARID3B, SMARCAD1, and FZD3, were measured in lung tissues of NSCLC patients and A549 cell line using qRT-PCR and Western blotting. The electroporation method was used to transfect A549 cells with let-7f mimic and microRNA inhibitor. The impact of let-7f transfection on the viability of A549 cells was assessed using MTT assay. The expression data of studied genes were analyzed statistically. Results: Results indicated significant downregulated expression level of let-7f-5p (p = 0.0013) and upregulated level of the HMGA2 and FZD3 in NSCLC cases (p < 0.05). In A549 cells, after transfection with let-7f mimic, the expression of both mRNA and protein levels of HMGA2, ARID3B, SMARCAD1, and FZD3 decreased. Also, the overexpression of let-7f significantly inhibited the A549 cell proliferation and viability (p = 0.017). Conclusion: Our findings exhibited the high value of let-7f and HMGA2 as biomarkers for NSCLC. The let-7f, as a major tumor suppressor regulatory factor via direct targeting genes (e.g. HMGA2), inhibits lung cancer cell viability and proliferation and could serve as a marker for the early diagnostic of NSCLC.

Association Between NOX4 And Nrf2 Genes in Non-Small-Cell Lung Carcinoma: A Case-Control Study.

BACKGROUND: Epithelial malignancy in lung cancer, which is initiated with myofibroblast differentiation and remodeling, promotes hypoxia and intracellular ROS generation most affected by the prototypical enzyme, NADPH oxidase 4 (NOX4). In addition, nuclear factor erythroid 2-related factor 2 (Nrf2) acts as a critical transcription factor by stimulating antioxidant proteins as redox homeostasis regulators. The aim of this study was to investigate a possible correlation between lung tissue NOX4 and Nrf2 genes (NOX4 and Nrf2) mRNA expression and bronchoalveolar lavage fluid (BALF) protein expression in non-small-cell lung carcinoma (NSCLC) patients. METHODS: Samples from 25 patients with various NSCLC types and stages and 20 healthy controls were collected. NOX4 and Nrf2 mRNA were measured by qRT-PCR, and protein by western blot analysis. RESULTS: NOX4 mRNA and protein expression was significantly up-regulated in NSCLC patients' lung tissues and BALFs (p= 0.03 and 0.01, respectively). In addition, by adjusting for age, sex, and NSCLC types and stages, a significant and positive correlation was observed between NOX4 and Nrf2 mRNA expression (r= 0.927, p= 0.001). This was also true when not adjusted as above (r= 0.944, p< 0.001). CONCLUSION: NOX4 mRNA and protein expression is significantly up-regulated in NSCLC patients' lung tissues and BALFs, and NOX4 and Nrf2 mRNA expression is positively correlated in NSCLC tissues.

Association of NOS3-c.894G>T transversion with susceptibility to metabolic syndrome in Azar-cohort population: A case-control study and in silico analysis of the SNP molecular effects.

OBJECTIVES: We investigated whether NOS3-c.894G>T transversion (rs1799983), which causes the substitution of glutamate with aspartate (E298D) in the oxygenase domain of endothelial nitric oxide synthase (eNOS), is associated with susceptibility to metabolic syndrome (MetS) risk in Iranian-Azerbaijanis. MATERIALS AND METHODS: The frequencies of the alleles and genotypes were compared in the 300 cases and 300 controls using PCR-RFLP assay. Also, higher-order MetS interaction with the genotypes, gender, age, and body mass index (BMI) was evaluated by classification and regression tree (CART) analysis. In silico analysis was done to introduce a hypothesis describing the molecular effects of NOS3-c.894G>T. RESULTS: The T allele (OR:1.46; CI:1.054-2.04; P=0.02), GT genotype (OR:1.44; CI:1.02-2.03; P=0.03), and dominant model (TT+GT vs GG, OR:1.48; CI:1.06-2.06; P=0.01) were found to be associated with increased risk of MetS. In the male subpopulation TT genotype (OR:7.19; CI:1.53-33.70; P=0.01) was discovered to be associated with increased odds of MetS. CART analysis showed that NOS3-c.894G>T genotypes and BMI significantly contribute to modulating MetS risk. Furthermore, in silico investigation revealed that c.894G>T may alter eNOS function through affecting interactions of its oxygenase domain with proteins such as B2R, b-actin, CALM1, CAV1, GIT1, HSP90AA1, NOSIP, and NOSTRIN. CONCLUSION: We showed that NOS3-c.894G>T was associated with an increased risk of MetS in Iranian-Azerbaijanis, and BMI modulates the effects of NOS3-c.894G>T genotypes on MetS risk. Also, in silico analysis found that NOS3-c.894G>T may affect the interaction of the eNOS oxygenase domain with its several functional partners.

Long non-coding RNAs as potential biomarkers in the prognosis and diagnosis of lung cancer: A review and target analysis.

Long non-coding RNAs (lncRNA) have been emerged as a novel class of molecular regulators in cancer. They are dysregulated in many types of cancer; however, there is not enough knowledge available on their expression and functional profiles. Lung cancer is the leading cause of the cancer deaths worldwide. Generally, lncRNAs may be associated with lung tumor pathogenesis and they may act as biomarkers for the cancer prognosis and diagnosis. Compared to other invasive prognostic and diagnostic methods, detection of lncRNAs might be a user-friendly and noninvasive method. In this review article, we selected 27 tumor-associated lncRNAs by literature reviewing to further discussing in detail for using as diagnostic and prognostic biomarkers in lung cancer. Also, in an in silico target analysis, the "Experimentally supported functional regulation" approach of the LncTarD web tool was used to identifying the target genes and regulatory mechanisms of the selected lncRNAs. The reports on diagnostic and prognostic potential of all selected lncRNAs were discussed. However, the target genes and regulatory mechanisms of the 22 lncRNAs were identified by in silico analysis and we found the pathways that are controlled by each target group of lncRNAs. They use epigenetic mechanisms, ceRNA mechanisms, protein interaction and sponge mechanism. Also, 10, 23, 5, and 28 target genes for each of these mechanisms were identified, respectively. Finally, each group of target genes controls 50, 12, 7, and 2 molecular pathways, respectively. In conclusion, LncRNAs could be used as biomarkers in lung cancer due to their roles in control of several signaling pathways related to lung tumors. Also, it seems that lncRNAs, which use epigenetic mechanisms for modulating a large number of pathways, could be considered as important subjects for lung cancer-related diagnostic and prognostic biomarkers.