Design of a dual-function agent by fusing a designed anti-VEGF-A binder and CPG-2 enzyme.

Anti-VEGF therapies are common for the treatment of cancer. Carboxypeptidase G (CPG-2) enzyme is a zinc-dependent metalloenzyme that metabolizes non-toxic synthetic 'benzoic mustard prodrugs' to cytotoxic moieties in tumor cells. In this study, we designed a dual-activity agent by combining a designed anti-VEGF- and CPG-2 enzyme to convert methotrexate (MTX). VEGF-A was docked against a set of scaffolds, and suitable inverse rotamers were made. Rosetta design was used for the interface design. The top 1200 binders were chosen by flow cytometry and displayed in yeast. The activity of CPG-2 enzyme was analyzed at different temperature conditions and in the presence of the substrate, MTX. Optimal binders were selected and protein was eluted using immobilized metal affinity chromatography and size-exclusion chromatography. Both, native PAGE and on-yeast flow cytometry confirmed the binding of the binder to VEGF-A. The activity of truncated enzymes was slightly lower than that of full-length enzymes linked to VEGF-A. The method should be generally useful as a dual-activity agent for targeting VEGF-A and combination therapy with the enzyme CPG-2 for metabolizing non-toxic prodrugs to cytotoxic moieties.Communicated by Ramaswamy H. Sarma.

Binder design for targeting SARS-CoV-2 spike protein: An in silico perspective.

INTRODUCTION: The COVID-19 pandemic is now affecting all people around the world and getting worse. New antiviral medications are desperately needed other than the few approved medications that have shown no promising efficacy so far. METHODS: Here we report three blocking binders for targeting SARS-CoV-2 spike protein to block the interaction between the spike protein on the SARS-CoV-2 and the angiotensin-converting enzyme 2 (ACE2) receptors, responsible for viral homing into the alveolar epithelium type II cells (AECII). RESULTS: The design process is based on the collected natural scaffolds and using Rosetta interface for designing the binders. CONCLUSION: Based on the structural analysis, three binders were selected, and the results showed that they might be promising as new therapeutic targets for blocking COVID-19.

Isolation and characterization of mesenchymal stem cells and its antitumor application on ovarian cancer cell line.

The molecular interaction network of Oct-4 (POU5F1) and NANOG connected to regulation and growth of mesenchymal stem cells (MSCs) were supplemented with information of miRNA to find an important micro-RNAs and supplemented molecular interaction network. Following co-culturing of Bone marrow mesenchymal stem cells (BMMSCs) with SKOV3 ovarian cancer cell lines and undifferentiated BMMSCs, MTT was analyzed for cell cytotoxicity. The analyses of the expression of miRNA were performed either after oesteogenesis (hsa-miR-34 and hsa-miR-335) or chondrogenic (hsa-miR-145 and hsa-miR-455) differentiation. This molecular interaction network was imaged in using software. The results from these findings gave an understanding of the main molecular mechanisms regulating MSCs therapeutic activity and their undifferentiated state maintenance. We recommend that the downregulation of miR-335 is crucial role for tissue homeostasis.

Chrysin Alters microRNAs Expression Levels in Gastric Cancer Cells: Possible Molecular Mechanism.

Background Gastric carcinoma still remains the second most common cause of cancer mortality in the world. Chrysin, as a flavone, has showed cancer chemopreventive activity. The cellular and molecular mechanisms of chrysin in cancer cells have not been fully understood. Objective In this study, we investigate expression levels of let-7a, miR-9, mir-18a, miR-21, miR-22, miR-34a, miR-126 and mir-221 to describe the anti-cancer effects of chrysin. Materials and Methods The cytotoxic effects of chrysin were assessed using MTT assay. The effect of chrysin on the microRNAs expression was determined by qRT-PCR. Results The MTT results for different concentrations of chrysin at different times on the Gastric carcinoma cells showed that IC50 for chrysin was 68.24 µM after 24 h of treatment. Expression analysis identified that miR-18, miR-21 and miR-221 were down regulated whereas let-7a, miR-9, miR-22, miR-34a and miR-126 were up regulated in Gastric carcinoma cell line (p<0.05). Conclusion Treatment with chrysin can alter the miRNAs expression and these findings might be an explanation for molecular mechanism of chrysin effect on gastric cancer.

New state of nanofibers in regenerative medicine.

Regenerative medicine has discovered engineered nanofiber scaffolds enhancing regeneration process. These agents have an attractive property to mimic the native environment. They are excellent agents in binding the extracellular matrix of a cell to another cell. They help in the growth and multiplication of the cell and help in the differentiation of the cells which are required before the regeneration process. Regenerative medicine focuses on cellular therapies, origins of stem and progenitor cells, and on explaining how they persevere (or do not) in adult organisms and improvement of biomaterials. The focus of this review is on the application of nanofiber scaffolds.

Drug loading and delivery using nanofibers scaffolds.

In recent times, notable advancement has been made in the field of electrospinning for the fabrication of numerous types of nanofiber scaffolds. Due to the ultrathin fiber diameter, electrospun nanofiber scaffolds are considered to be an operational delivery system for biomolecules, genes, as well as drugs due to the high specific surface area and stereological porous structure. Here, we introduce some of methods for the integration of drugs and biomolecules within electrospun nanofiber scaffolds, such as blending, surface modification, coaxial process, and emulsion methods. Then, we describe some important biomedical applications of nanofibers in drug delivery systems along with their suitable examples in transdermal systems and wound dressings, cancer therapy, growth factor delivery, nucleic acid delivery, and stem cell delivery.

Upregulation of miR-9 and Let-7a by nanoencapsulated chrysin in gastric cancer cells.

Chrysin, as a flavone, has showed cancer chemopreventive activity. The present study utilized the PLGA-PEG-chrysin to evaluate the expression of miR-9 and Let-7a in human gastric cells. The structure of nanoparticles and encapsulated chrysin was evaluated using 1H NMR, FT-IR, and SEM. MTT assay was used for the evaluation of cytotoxicity effect of nanoencapsulated chrysin. Expression levels of miR-9 and Let7-a were studied by real-time PCR. The results demonstrated that chrysin-PLGA-PEG nanoparticles are more effective than pure chrysin in upregulation of miR-9 and Let-7a due to enhanced uptake by cells. Therefore, PLGA-PEG could be a superior carrier for this kind of hydrophobic agent.

Effects of Chrysin-PLGA-PEG Nanoparticles on Proliferation and Gene Expression of miRNAs in Gastric Cancer Cell Line.

BACKGROUND: Recently, Chrysin, as a flavone, has revealed cancer chemo-preventive activity. The present experiment utilized the PLGA-PEG-chrysin complex, and free chrysin, to evaluation of the expression of miR-22, miR-34a and miR-126 in human gastric cell line. OBJECTIVES: The purpose of this study was to examine whether nano encapsulating chrysin improves the anti-cancer effect of free chrysin on AGS human gastric cell line. METHODS: Properties of the chrysin encapsulated in PLGA-PEG nanoparticles were investigated by SEM, H NMR, and FTIR. The assessment of cytotoxicity on the growth of the human gastric cell line was carried out through MTT assay. After treating the cells with a prearranged amount of pure and encapsulated chrysin, RNA was extracted and the expressions of miR-22, miR-34a and miR-126 were measured by using real-time PCR. RESULTS: With regard to the amount of the chrysin loaded in PLGA-PEG nanoparticles, IC50 value was significantly decreased in nanocapsulatedchrysin, in comparison with free chrysin. This finding has been proved through the further increase of miR-22, miR-34a and miR-126 gene expression of nanocapsulatedchrysin, in comparison with free chrysin. CONCLUSIONS: In this study, we revealed that the PLGA-PEG-chrysin is more effective than free chrysin in inhibiting the growth of human gastric cell line.

Down regulation of miR-18a, miR-21 and miR-221 genes in gastric cancer cell line by chrysin-loaded PLGA-PEG nanoparticles.

Chrysin were well-documented as having significant biological roles particularly cancer chemo-preventive activity. However, the poor water solubility of chrysin limited their bioavailability and biomedical applications. In this study, we encapsulate the chrysin into PLGA-PEG nanoparticles for local treatment. In regard to the amount of the drug load, IC50 was significant decreased in nanocapsulated chrysin in comparison with free chrysin. This was confirmed through decrease of miR-18a, miR-21, and miR-221 genes expression by real-time PCR. The results demonstrated that PLGA-PEG-chrysin complexes can be more effective than free chrysin. Therefore, PLGA-PEG can be a better nanocarrier for this kind of hydrophobic flavonoid.

Upregulation of Mir-34a in AGS Gastric Cancer Cells by a PLGA-PEG-PLGA Chrysin Nano Formulation.

BACKGROUND: Nano-therapy has the potential to revolutionize cancer therapy. Chrysin, a natural flavonoid, was recently recognized as having important biological roles in chemical defenses and nitrogen fixation, with anti-inflammatory and anti-oxidant effects but the poor water solubility of flavonoids limitstheir bioavailability and biomedical applications. OBJECTIVE: Chrysin loaded PLGA-PEG-PLGA was assessed for improvement of solubility, drug tolerance and adverse effects and accumulation in a gastric cancer cell line (AGS). MATERIALS AND METHODS: Chrysin loaded PLGA-PEG copolymers were prepared using the double emulsion method (W/O/W). The morphology and size distributions of the prepared PLGA-PEG nanospheres were investigated by 1H NMR, FT-IR and SEM. The in vitro cytotoxicity of pure and nano-chrysin was tested by MTT assay and miR-34a was measured by real-time PCR. RESULTS: 1H NMR, FT-IR and SEM confirmed the PLGA-PEG structure and chrysin loaded on nanoparticles. The MTT results for different concentrations of chrysin at different times for the treatment of AGS cell line showed IC50 values of 68.2, 56.2 and 42.3 µM and 58.2, 44.2, 36.8 µM after 24, 48, and 72 hours of treatment, respectively for chrysin itslef and chrysin-loaded nanoparticles. The results of real time PCR showed that expression of miR-34a was upregulated to a greater extent via nano chrysin rather than free chrysin. CONCLUSIONS: Our study demonstrates chrysin loaded PLGA-PEG promises a natural and efficient system for anticancer drug delivery to fight gastric cancer.

Endovascular treatment of very small and very large ruptured aneurysms of the anterior cerebral circulation: a single-center experience.

INTRODUCTION: Endovascular treatment of aneurysms has been introduced as a less invasive method for decreasing the rate of aneurysm rerupture and subsequent subarachnoid hemorrhage. The outcome and complication rate for endovascular treatment of very small (≤3 mm) and very large (15-25 mm) intracranial aneurysms has been controversial. Here we report our experience with endovascular coiling of very small and very large ruptured aneurysms of the anterior cerebral circulation. METHODS: Patients were included in the study if the maximum dimension of the intracranial ruptured aneurysm was reported to be ≤3 mm or 15-25 mm and if the aneurysm was within the anterior cerebral circulation. The largest dimension was calculated using CT angiography and was confirmed by digital subtraction angiography. Endovascular coiling was performed using Guglielmi detachable coils. All patients underwent follow-up contrast MR angiography every 6 months. RESULTS: A total of 40 cases (18 females and 22 males) were included in this single-center study. Twenty-one very small and 19 very large ruptured aneurysms were analyzed. Preprocedural Hunt and Hess grades were determined. Endovascular coiling was performed successfully in most cases (97.5%), with unsuccessful coiling in 1 patient with a very small ruptured aneurysm. In the very small aneurysm group, the most common location was the anterior communicating artery and, in the large aneurysm group, the most common location was the middle cerebral artery (MCA) bifurcation. The mean follow-up time was 15.08 months (range: 6-30 months). The 6th month modified Rankin scale (mRS) values for very small aneurysm cases were 0 (no symptoms at all) in 16 cases (76.2%) and 1 (no significant disability despite symptoms) in 5 cases (23.80%). For the very large aneurysm cases, the mRS values were 1 in 2 cases (10.5%), 2 in 7 cases (36.8%), 3 in 6 cases (31.6%), 4 in 3 cases (15.8%) and 6 in 1 case (died due to vasospasm 72 h later; 5.2%). The immediate complications that were observed were MCA branch occlusion in 1 very small aneurysm patient and early vasospasms in 3 very large aneurysm patients. The late complication that was observed was recanalization in 1 very small aneurysm case (1/21, 4.76%) and in 5 very large aneurysm cases (5/18, 27.77%). CONCLUSION: Endovascular treatment of very small aneurysms is an effective method of treatment with acceptable immediate and long-term outcomes. Immediate and long-term complications were more prevalent in very large ruptured aneurysms.

Unprotected carotid artery stenting: complications in 6 months follow-up.

INTRODUCTION: The most serious complications of carotid artery stenting (CAS) are related to the release of particles into the cerebral circulation. Although embolic protection devices may reduce the incidence of cerebral embolization, their use may be associated with additional complications including spasm or dissection. In this report, we present our experience with patients who underwent unprotected CAS. METHODS: We prospectively enrolled patients with symptomatic internal carotid artery stenosis from October 2008 to June 2010.Stenosis diagnosed based on carotid duplex and/or CT angiography findings and confirmed by digital subtraction angiography. All stenting procedures were done without using a protection device. Neurologist evaluated neurological situation of the patients, 24 h, 30 days, 3 and 6 months after stenting. Patients had carotid duplex scan scheduled on the third and sixth month after the procedure. RESULTS: Overall, 116patients underwent stenting; technical success rate was 100% .Mean age of patients was 68.48 ± 9.84 years, and 69% of the patients were male. One patient (1/116, 0.86%) developed transient ischemic attack during the procedure. One non-neurologic death (1/116, 0.86%) occurred 18 h after the procedure because of myocardial infarction. In 1-month follow-up, only one patient (1/115, 0.87%) presented with neurological complication as a consequence of thalamic hemorrhagic. Six months follow-up revealed two cases (2/115, 1.74%) with in-stent restenosis which one of them was severe and symptomatic and underwent angioplasty. CONCLUSION: Unprotected carotid stenting appears to accompany with low early and late complications. Cerebral embolization during CAS is not the only cause of these complications and the use of cerebral protection devices may therefore not prevent all major complications including myocardial infarction, hyperperfusion syndrome, and re-stenosis.

A Comparison of Stent Implant versus Medical Treatment for Severe Symptomatic Intracranial Stenosis: A Controlled Clinical Trial.

BACKGROUND: Atherosclerotic stenosis of the major intracranial arteries is the most common cause of ischemic stroke. There are limited treatments for severe intracranial stenosis, and stent placement versus medical treatment remains controversial. The aim of this study was to compare functional outcomes of these two modalities in patients with severe symptomatic intracranial stenosis. METHODS: At a single center, between 2008 and 2011, patients with angiographically demonstrated severe (70-90%) symptomatic intracranial atherosclerosis were divided into two groups: group A, which received only medical treatment, and group B, which underwent endovascular stent implant treatment. The severity and location of the stenosis was determined by digital subtraction angiography and the Warfarin-Aspirin Symptomatic Intracranial Disease (WASID) trial criteria in all patients. The exclusion criteria were: specific causes other than atherosclerosis, such as artery dissection, fibromuscular dysplasia, vasculitis, radiation and intracranial hemorrhage, focal neurological deficit that did not correlate to internal carotid artery or middle cerebral artery stenosis. All procedures were done under light anesthesia. Technical success was defined as the reduction of stenosis to <30% with complete enveloping of the lesion after the procedure. Early and late adverse events and functional outcomes were compared between the groups using the modified Rankin Scale (mRS). RESULTS: Overall, 63 patients (29 in group A and 34 in group B) were evaluated and followed for a mean period of 15.22 months (range 6-25). The technical success rate was 97% in a total of 34 stents in 34 patients. There was no difference between the early (within 30 days) adverse event rates of the two groups. The median follow-up duration for the stent implant patients was 15 months (range 6-25), and for the medically treated cohort it was 14 months (range 8-25). The re-stenosis rate was 5.8% and the total number of late (>30 days) adverse events, including stroke, myocardial infarction and death, was 1 (2.9%) and 6 (20.7%) in the stent implant and medical groups, respectively (p = 0.042). The stent implant group had significantly better favorable functional outcomes according to the mRS than the medical group (93.9 vs. 63.0%). The cumulative secondary adverse event-free survival was significantly lower in the stent implant group. CONCLUSION: Stent implants can be considered more durable and safe for patients with symptomatic severe stenosis of the internal carotid artery or middle cerebral artery, despite optimal medical therapy. Randomized, multicenter trials are required to confirm these results.