Advancements in Melanoma Therapies: From Surgery to Immunotherapy.

OPINION STATEMENT: Melanoma is defined as the most aggressive and deadly form of skin cancer. The treatment of melanoma depends on the disease stage, tumor location, and extent of its spread from its point of origin. Melanoma treatment has made significant advances, notably in the context of targeted and immunotherapies. Surgical resection is the main therapeutic option for earlystage melanoma, and it provides favourable outcomes. With disease metastasis, systemic treatments such as immunotherapy and targeted therapy become increasingly important. The identification of mutations that lead to melanoma has influenced treatment strategies. Targeted therapies focusing on these mutations offer improved response rates and fewer toxicities than conventional chemotherapy. Furthermore, developing immunotherapies, including checkpoint inhibitors and tumor-infiltrating lymphocyte (TIL) therapies, has demonstrated encouraging outcomes in effectively combating cancer cells. These therapeutic agents demonstrate superior effectiveness and a more tolerable side-effect profile, improving the quality of life for patients receiving treatment. The future of melanoma treatment may involve a multimodal approach consisting of a combination of surgery, targeted therapy, and immunotherapy adapted to each patient's profile. This approach may improve survival rates and health outcomes.

Isolation and Characterization of the Vascular Endothelial Growth Factor Receptor Targeting ScFv Antibody Fragments Derived from Phage Display Technology.

Angiogenesis, as a tumor hallmark, plays an important role in the growth and development of the tumor vasculature system. There is a huge amount of evidence suggesting that the vascular endothelial growth factor receptor (VEGFR-2)/VEGF-A axis is one of the main contributors to tumor angiogenesis and metastasis. Thus, inhibition of the VEGFR-2 signaling pathway by anti-VEGFR-2 mAb can retard tumor growth. In this study, we employ phage display technology and solution-phase biopanning (SPB) to isolate specific single-chain variable fragments (scFvs) against VEGFR-2 and report on the receptor binding characteristics of the candidate scFvs A semisynthetic phage antibody library to isolate anti-VEGFR-2 scFvs through an SPB performed with decreasing concentrations of the VEGFR-2-His tag and VEGFR-2-biotin. After successful expression and purification, the specificity of the selected scFv clones was further analyzed by enzyme-linked immunosorbent assay (ELISA), flow cytometry, and immunoblotting. The competition assay was undertaken to identify the VEGFR-2 receptor-blocking properties of the scFvs. Furthermore, the molecular binding characteristics of candidate scFvs were extensively studied by peptide-protein docking. Polyclonal ELISA analysis subsequent to four rounds of biopanning showed a significant enrichment of VEGFR-2-specific phage clones by increasing positive signals from the first round toward the fourth round of selection. The individual VEGFR-2-reactive scFv phage clones were identified by monoclonal phage ELISA. The sequence analysis and complementarity-determining region alignment identified the four unique anti-VEGFR-2-scFv clones. The soluble and purified scFvs displayed binding activity against soluble and cell-associated forms of VEGFR-2 protein in the ELISA and flow cytometry assays. Based on the inference from the molecular docking results, scFvs D3, E1, H1, and E9 recognized domains 2 and 3 on the VEGFR-2 protein and displayed competition with VEGF-A for binding to VEGFR-2. The competition assay confirmed that scFvs H1 and D3 can block the VEGFR-2/VEGF-A interaction. In conclusion, we identified novel VEGFR-2-blocking scFvs that perhaps exhibit the potential for angiogenesis inhibition in VEGFR-2-overexpressed tumor cells.

Sex-specific association of exposure to air pollutants and Nrf2 gene expression and inflammatory biomarkers in exhaled breath of healthy adolescents.

Studies investigating the nuclear factor erythroid 2-related factor 2 (Nrf2) expression levels in the respiratory system of healthy subjects are scarce. Moreover, separate studies on the health-related outcomes of air pollution for each sex are limited. The current panel study investigated sex-specific Nrf2 expression levels and related oxidative stress and inflammatory responses among healthy adolescents exposed to PM2.5, PM10, O3, and PM2.5-bounded metals in a high traffic region. Forty-nine healthy nonsmoking subjects participated in the study for five consecutive months (Nov. 2019 to Feb. 2020). Each subject was asked to provide 1 mL of exhaled breath condensate (EBC). Data were analyzed using linear mixed-effects models. The results showed that PM10, PM2.5, O3, and PM2.5-bounded metals were negatively linked to Nrf2 expression level in EBC of females with -58.3% (95% CI: 79.5, -15.4), -32.1% (95% CI: -50.3, -7.1), -76.2% (95% CI: -92.6, -23.9), and -1.9 (95% CI: -3.4, -0.4), respectively. While our results presented no significant association between the studied pollutants and Nrf2 gene expression in males, significant associations were observed between the pollutants and total nitric oxide (NOx), interleukins 6 (IL-6), and tumor necrosis factor-alpha (TNF-α) in the EBC of females. In the case of males, only EBC cytokines showed a significant association with air pollutants. Overall, this study suggests that exposure to ambient air pollutants may affect the respiratory system with biologically different mechanisms in males and females. PM2.5 concentration had a positive correlation with exhaled TNF-α and IL6 values in females while positive correlation with TNF-α and negative correlation with IL6 values in males. O3 had a negative correlation with TNF-α in males.

A review on targeting tumor microenvironment: The main paradigm shift in the mAb-based immunotherapy of solid tumors.

Monoclonal antibodies (mAbs) as biological macromolecules have been remarked the large and growing pipline of the pharmaceutical market and also the most promising tool in modern medicine for cancer therapy. These therapeutic entities, which consist of whole mAbs, armed mAbs (i.e., antibody-toxin conjugates, antibody-drug conjugates, and antibody-radionuclide conjugates), and antibody fragments, mostly target tumor cells. However, due to intrinsic heterogeneity of cancer diseases, tumor cells targeting mAb have been encountered with difficulties in their unpredictable efficacy as well as variability in remission and durable clinical benefits among cancer patients. To address these pitfalls, the area has undergone two major evolutions with the intent of minimizing anti-drug responses and addressing limitations experienced with tumor cell-targeted therapies. As a novel hallmark of cancer, the tumor microenvironment (TME) is becoming the great importance of attention to develop innovative strategies based on therapeutic mAbs. Here, we underscore innovative strategies targeting TME by mAbs which destroy tumor cells indirectly through targeting vasculature system (e.g., anti-angiogenesis), immune system modulation (i.e., stimulation, suppression, and depletion), the targeting and blocking of stroma-based growth signals (e.g., cancer-associated fibroblasts), and targeting cancer stem cells, as well as, their effector mechanisms, clinical uses, and relevant mechanisms of resistance.

Acute responses of airway oxidative stress, inflammation, and hemodynamic markers to ambient PM2.5 and their trace metal contents among healthy adolescences: A panel study in highly polluted versus low polluted regions.

Particulate air pollutants are known contributors to global cardiorespiratory mortality through several pathways. We examined the effects of varied exposure to PM2.5 and trace metals on biological markers of airway inflammation, oxidative stress, and hemodynamic function of young individuals living in two different exposure settings. We enrolled and followed a panel of 97 healthy nonsmoking participants aged 15-18 years living in a highly polluted metropolitan city of Tabriz (TBZ) and a much less polluted semi-urban town of Hadishahr (HDS). For five consecutive months, the subjects were examined by a physician, and fractional exhaled nitric oxide levels (FENO) were measured. Samples of exhaled breath condensation (EBC) were obtained for measuring interleukin 6 (IL-6), tumor necrosis factor α (TNF-α), and total nitric oxide (NOx). We measured daily outdoor PM2.5 mass concentration in a fixed station in each location for all this period. The PM-metal content was analyzed by ICP-MS. The linear mixed-effects regression models were applied for data analysis. The averages of PM2.5 mass and total metals in TBZ were nearly two and four times higher than in HDS, respectively. In TBZ, an increased IQR of PM2.5 mass during 0-5 days was -correlated with a significant rise in diastolic blood pressure, heart rate, TNF-α, FENO, and NOx and reduction of IL-6. Moreover, exposure to low PM2.5 concentration is significantly -correlated with an elevation in diastolic blood pressure in HDS. We also observed that exposure to metal constituents in the highly polluted region is correlated with increased TNF-α and IL-6 with 131.80% (95% CI: 56.01, 244.39) and 47.51% (95% CI: 33.01, 62.05) per IQR of Hg, respectively. This study suggests that exposure to ambient PM2.5 and their metal contents in highly polluted areas may incite significant changes in airway inflammation, oxidative stress, and hemodynamic parameters in healthy subjects.

Improved Soluble ScFv ELISA Screening Approach for Antibody Discovery Using Phage Display Technology.

Phage display technology (PDT) is a powerful tool for the isolation of recombinant antibody (Ab) fragments. Using PDT, target molecule-specific phage-Ab clones are enriched through the "biopanning" process. The individual specific binders are screened by the monoclonal scFv enzyme-linked immunosorbent assay (ELISA) that may associate with inevitable false-negative results. Thus, in this study, three strategies were investigated for optimization of the scFvs screening using Tomlinson I and J libraries, including (1) optimizing the expression of functional scFvs, (2) improving the sensitivity of ELISA, and (3) preparing different samples containing scFvs. The expression of all scFv Abs was significantly enhanced when scFv clones were cultivated in the terrific broth (TB) medium at the optimum temperature of 30 °C. The protein A-conjugated with horseradish peroxidase (HRP) was found to be a well-suited reagent for the detection of Ag-bound scFvs in comparison with either anti-c-myc Ab or the mixing procedure. Based on our findings, it seems there is no universal media supplement for an improved expression of all scFvs derived from both Tomlinson I and J libraries. We thus propose that expression of scFv fragments in a microplate scale is largely dependent on a variety of parameters, in particular the scFv clones and relevant sequences.