Bacteriophages and Green Synthesized Zinc Oxide Nanoparticles in Combination Are Efficient against Biofilm Formation of Pseudomonas aeruginosa.

Bacteriophages (phages) are viruses that infect the bacteria within which their reproduction cycle takes place, a process that ends in the lysis and death of the bacterial cell. Some phages are also able to destroy bacterial biofilms. Due to increased antibiotics resistance, Pseudomonas aeruginosa, another biofilm-forming pathogen, is a problem in many parts of the world. Zinc oxide (ZnO) and other metal nanoparticles (NPs) are biologically active and also possess anti-biofilm properties. ZnO-NPs were prepared by the green synthesis method using orange peels. The vibrational peaks of the ZnO-NPs were analyzed using FTIR analysis, and their size and morphological properties were determined using scanning electron microscopy (SEM). The ability of the ZnO-NPs to reduce or eliminate P. aeruginosa biofilm alone or in combination with phages PB10 and PA19 was investigated. The P. aeruginosa cells were effectively killed in the preformed 48 h biofilms during a 24 h incubation with the ZnO-NP-phage combination, in comparison with the control or ZnO-NPs alone. The treatments on growing biofilms were most efficient in the final stages of biofilm development. All five treatment groups showed a significant biofilm reduction compared to the control group (p < 0.0001) at 48 h of incubation. The influence of the ZnO-NPs and phages on the quorum sensing system of P. aeruginosa was monitored by quantitative real-time PCR (qRT-PCR) of the autoinducer biosynthesis gene lasI. While the ZnO-NPs repressed the lasI gene transcription, the phages slightly activated it at 24 and 48 h of incubation. Also, the effect of the ZnO-NPs and phage PA19 on the viability of HFF2 cells was investigated and the results showed that the combination of NPs with PA19 reduced the toxic effect of ZnO-NPs and also stimulated the growth in normal cells.

Comparative evaluation of hesperetin-loaded graphene oxide nanosheets (Hsp-GO) as a drug delivery system for colon cancer: synthesis and anticancer efficiency assessment.

BACKGROUND: Graphene oxide nanosheets (GONS) are recognized for their role in enhancing drug delivery and effectiveness in cancer treatment. With colon cancer being a prevalent global issue and the significant side effects associated with chemotherapy, the primary treatment for colon cancer alongside surgery, there is a critical need for novel therapeutic strategies to support patients in combating this disease. Hesperetin (HSP), a natural compound found in specific fruits, exhibits anti-cancer properties. The aim of this study is to investigate the effect of GONS on the LS174t colon cancer cell line. METHODS: In this study, an anti-cancer nano-drug was synthesized by creating a hesperetin-graphene oxide nanocomposite (Hsp-GO), which was subsequently evaluated for its efficacy through in vitro cell toxicity assays. Three systems were investigated: HSP, GONS, and HSP-loaded GONS, to determine their cytotoxic and pro-apoptotic impacts on the LS174t colon cancer cell line, along with assessing the expression of BAX and BCL2. The morphology and properties of both GO and Hsp-GO were examined using scanning electron microscopy (SEM), X-ray diffraction, and Fourier transform infrared spectroscopy (FTIR). RESULTS: The Hsp-GO nanocomposite displayed potent cytotoxic and pro-apoptotic effects on LS174t colon cancer cells, outperforming individual treatments with HSP or GONS. Cell viability assays showed a significant decrease in cell viability with Hsp-GO treatment. Analysis of BAX and BCL2 expression revealed elevated BAX and reduced BCL2 levels in Hsp-GO treated cells, indicating enhanced apoptotic activity. Morphological analysis confirmed successful Hsp-GO synthesis, while structural integrity was supported by X-ray diffraction and FTIR analyses. CONCLUSIONS: These study highlight the potential of Hsp-GO as a promising anti-cancer nano-drug for colon cancer therapy.

The effect of chitosan hydrogel containing gold nanoparticle complex with paclitaxel on colon cancer cell line.

Colon cancer is a significant global health issue, and its primary treatment, chemotherapy, is limited by toxicity and drug resistance. This has led researchers to explore alternative therapeutic approaches. One such approach is the use of chitosan, a natural biopolymer with anti-cancer properties, and paclitaxel, a potent chemotherapeutic agent with promising activity against many types of cancer. In this study, the effectiveness of a chitosan hydrogel that contains a complex of gold nanoparticles with paclitaxel in treating LS174T colon cancer cell line was investigated. The synthesized chitosan hydrogel was characterized and used to treat the colon cancer cells in cell culture. MTT assay and apoptotic gene expression analysis were conducted to evaluate the complex's effectiveness. The results showed that the chitosan hydrogel-loaded gold nanoparticle-paclitaxel complex exhibited a potent cytotoxic effect against the cancer cells. Moreover, the treatment resulted in a significant increase in pro-apoptotic BAX and BAD expression and a decrease in anti-apoptotic BCL2 expression, indicating a pro-apoptotic effect. These findings suggest that utilizing a chitosan hydrogel that contains a complex of gold nanoparticles with paclitaxel shows promise as a viable treatment option for colon cancer. Further research is needed to determine the potential efficacy and safety of this treatment approach in clinical settings.

VEGFR2 Mimicking Peptide Inhibits the Proliferation of Human Umbilical Vein Endothelial Cells (Huvecs) by Blocking VEGF.

INTRODUCTION: A variety of key human physiological processes rely on angiogenesis, ranging from reproduction and fetal growth to wound healing and tissue repair. Furthermore, this process significantly contributes to tumor progression, invasion, and metastasis. As the strongest inducer of angiogenesis, Vascular Endothelial Growth Factor (VEGF) and its receptor (VEGFR) are targets of therapeutic research for blocking pathological angiogenesis. OBJECTIVE: Preventing the interaction between VEGF and VEGFR2 by a peptide is a promising strategy for developing antiangiogenic drug candidates. This study was aimed at designing and evaluating VEGF-targeting peptides using in silico and in vitro techniques. METHODS: The VEGF binding site of VEGFR2 was considered a basis for peptide design. The interaction of VEGF and all three peptides derived from VEGFR2 were analyzed using ClusPro tools. In a complex with VEGF, the peptide with a higher docking score was evaluated to confirm its stability using molecular dynamics (MD) simulation. The gene coding for the selected peptide was cloned and expressed in E. coli BL21. The bacterial cells were cultured on a large scale, and the expressed recombinant peptide was purified using Ni-NTA chromatography. Refolding of the denatured peptide was carried out by the stepwise removal of the denaturant. The reactivity of peptides was confirmed using western blotting and enzyme-linked immunosorbent assay (ELISA) assays. Finally, the inhibition potency of the peptide on human umbilical vein endothelial cells was assessed using the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl- 2H-tetrazolium bromide (MTT) assay. RESULTS: Among three peptides, the peptide with the best docking pose and the highest affinity for VEGF was selected for further studies. Then the stability of the peptide was confirmed over the 100 ns MD simulation. After in silico analyses, the selected peptide was presented for in vitro analysis. Expression of the selected peptide in E. coli BL21 resulted in a pure peptide with a yield of approximately 200 µg/ml. Analysis by ELISA revealed the high reactivity of the peptide with VEGF. Western blot analysis confirmed the specific reactivity of selected peptides with VEGF. The MTT assay revealed the growth inhibitory effect of the peptide on human umbilical vein endothelial cells with an IC50 value of 247.8 µM. CONCLUSION: In summary, the selected peptide demonstrated a promising inhibitory effect on human umbilical vein endothelial cells that could be a valuable anti-angiogenic candidate for further assessment. Additionally, these in silico and in vitro data provide new insights into peptide design and engineering.

An efficient method for cell sheet bioengineering from rBMSCs on thermo-responsive PCL-PEG-PCL copolymer.

Utilizing both medium enrichment and a thermos-responsive substrate to maintain the cell-to-cell junctions and extracellular matrix (ECM) intact, cell sheet technology has emerged as a ground-breaking approach. Investigating the possibility of using sodium selenite (as medium supplementation) and PCL-PEG-PCL (as vessel coating substrate) in the formation of the sheets from rat bone marrow-derived mesenchymal stem cells (rBMSCs) was the main goal of the present study. To this end, first, Polycaprolactone-co-Poly (ethylene glycol)-co-Polycaprolactone triblock copolymer (PCEC) was prepared by ring-opening copolymerization method and characterized by FTIR, 1 H NMR, and GPC. The sol-gel-sol phase transition temperature of the PCEC aqueous solutions with various concentrations was either measured. Next, rBMSCs were cultured on the PCEC, and let be expanded in five different media containing vitamin C (50 µg/ml), sodium selenite (0.1 µM), vitamin C and sodium selenite (50 µg/ml + 0.1 µM), Trolox, and routine medium. The proliferation of the cells exposed to each material was evaluated. Produced cell sheets were harvested from the polymer surface by temperature reduction and phenotypically analyzed via an inverted microscope, hematoxylin and eosin (H&E) staining, and field emission scanning electron microscopy (FESEM). Through the molecular level, the expression of the stemness-related genes (Sox2, Oct-4, Nanog), selenium-dependent enzymes (TRX, GPX-1), and aging regulator gene (Sirt1) were measured by q RT-PCR. Senescence in cell sheets was checked by beta-galactosidase assay. The results declared the improved ability of the rBMSCs for osteogenesis and adipogenesis in the presence of antioxidants vitamin C, sodium selenite, and Trolox in growth media. The data indicated that in the presence of vitamin C and sodium selenite, the quality of the cell sheet was risen by reducing the number of senescent cells and high transcription of the stemness genes. Monolayers produced by sodium selenite was in higher-quality than the ones produced by vitamin C.

Network-Based and Machine-Learning Approaches Identify Diagnostic and Prognostic Models for EMT-Type Gastric Tumors.

The microsatellite stable/epithelial-mesenchymal transition (MSS/EMT) subtype of gastric cancer represents a highly aggressive class of tumors associated with low rates of survival and considerably high probabilities of recurrence. In the era of precision medicine, the accurate and prompt diagnosis of tumors of this subtype is of vital importance. In this study, we used Weighted Gene Co-expression Network Analysis (WGCNA) to identify a differentially expressed co-expression module of mRNAs in EMT-type gastric tumors. Using network analysis and linear discriminant analysis, we identified mRNA motifs and microRNA-based models with strong prognostic and diagnostic relevance: three models comprised of (i) the microRNAs miR-199a-5p and miR-141-3p, (ii) EVC/EVC2/GLI3, and (iii) PDE2A/GUCY1A1/GUCY1B1 gene expression profiles distinguish EMT-type tumors from other gastric tumors with high accuracy (Area Under the Receiver Operating Characteristic Curve (AUC) = 0.995, AUC = 0.9742, and AUC = 0.9717; respectively). Additionally, the DMD/ITGA1/CAV1 motif was identified as the top motif with consistent relevance to prognosis (hazard ratio > 3). Molecular functions of the members of the identified models highlight the central roles of MAPK, Hh, and cGMP/cAMP signaling in the pathology of the EMT subtype of gastric cancer and underscore their potential utility in precision therapeutic approaches.

Preparation and evaluation of chitosan skin patches containing mesoporous silica nanoparticles loaded by doxycycline on skin wound healing.

This study aims to prepare and evaluate a skin patch containing mesoporous silica nanoparticles with doxycycline for skin wound healing in a rat model. For this purpose, 84 female rats were randomly placed in four equal groups: (A) Control group with only skin defects and no therapeutic intervention; (B) Chitosan group in which a chitosan skin patch without loading any drug was placed on their skin defect; (C); The ChMesN group had a chitosan skin patch containing drug-free mesoporous silica nanoparticles; (D) ChMesND group had a skin patch loaded with doxycycline loaded with mesoporous silica nanoparticles on their skin defect. The histological results showed that on the 3rd day of the study, collagen fiber orientation was significantly higher in the ChMesND group than in the other groups. On the 7th day of the study, neovascularization, and inflammation in the ChMesND group were significantly higher and lower than in the other groups, respectively. On day 21, the most re-epithelialization was observed in the ChMesND group. It was found that on day 7, the wound area in the ChMesND group was significantly less than in other groups. On the 21st day of the study, the minimal experimental wound area was related to chitosan and ChMesND groups. Although chitosan has anti-inflammatory effects, its combination with doxycycline with several beneficial biological effects can have significant therapeutic effects with chitosan. Hence, it can be concluded that chitosan skin patch containing doxycycline can be suitable dressings for managing and accelerating the healing of skin wounds.

The dynamicity of mutant KRAS ß2 strand modulates its downstream activation and predicts anticancer KRAS inhibition.

AIM: Oncogenic mutations involving KRAS are human cancer's most common driving force. We aimed to determine specific conformational features of the active KRAS regarding downstream signaling activation, especially in mutant forms of KRAS. MAIN METHODS: We applied Molecular Dynamics (MD) simulations in triplicate and post-MD analytical methods on the KRAS and its G12 mutant structures. In addition, clustering, umbrella sampling, and principal component analysis were conducted to improve the significant conformations related to the activity of the KRAS variants. The results were generally represented as the probability of the conformations regarding different structural aspects, including ß2-strand length, main residual distances, and critical residue interactions. KEY FINDINGS: Our results showed that the KRAS ß2-strand length was a convenient structural criterion to show the KRAS activity. Accordingly, the active conformations of KRAS were the most probable to have 9-10 residue numbers of ß2-strand. Based on this observation, it was also shown that the GDP forms of KRAS G12 mutants could be in the active mode because of increased ß2-strand length. Moreover, the distance between the E37 and A59 residues differed in relation to ß2-ß3 sheet length and can be considered another KRAS activity indicator. Interestingly, ß2-strand length could also predict the KRAS activity in the presence of a direct mutant KRAS inhibitor. SIGNIFICANCE: As a result, our observations provide a new mechanism regarding the high efficacy of direct inhibition of KRAS-GDP in cancer therapy. In addition, designing and screening the mutant KRAS inhibitors can be more achievable using the ß2-strand length probability.

Evaluation of Inflammatory Response Due to Use of Controlled Release Drug Delivery System of Chitosan Hydrogel Loaded with Buprenorphine and Ketorolac in Rat with Experimental Proximal Tibial Epiphysis Defect.

Aims:A controlled release drug delivery system loaded with buprenorphine and ketorolac was synthesized and used in the experimental model of bone defect and while evaluating the inflammatory response, the repair process in the defects was investigated.Materials and methods:To determine the effectiveness of the synthesized the mentioned systems, 5 groups were defined; the control group, the chitosan hydrogel receiving group (chitosan group), the ketorolac-loaded chitosan hydrogel group (ketorolac group), the buprenorphine-loaded chitosan hydrogel receiving group (buprenorphine group), and the chitosan hydrogel-loading group loaded with a combination of ketorolac and buprenorphine (ketorolac-buprenorphine group). Results:The results showed that the population of leukocytes (tWBC) and neutrophils on different days of the study in the control group compared to other groups had a significant increase (P < 0.05) while on day 7 of the study in the ketorolac group these parameters decreased significantly compared to other groups (P < 0.05). While examining the histological changes in the experimental defect created in the proximal tibia of rats at different times, some inflammatory indices such as total and differential leukocyte population, plasma concentrations of TNF-α and IL-6 were compared in different groups (P < 0.05). The various evaluated data showed that among the different groups, in the control and ketorolac-buprenorphine groups, there was the lowest and highest control of inflammatory response and bone repair, respectively.Conclusion:In the ketorolac group due to the impact of ketorolac on leukocyte populations the best bone healing can be expected among the different treatment groups.

Absorption of daunorubicin and etoposide drugs by hydroxylated and carboxylated carbon nanotube for drug delivery: theoretical and experimental studies.

Anti-cancer daunorubicin and etoposide drugs are mostly used in chemotherapy medicine to treat a wide variety of cancers. Many of the side effects and specific delivery to a target tissue are the main challenges of using chemotherapeutic agents. To avoid serious toxic side effects and improve treatment outcomes, functionalized carbon nanotubes (f-CNTs) are considered promising nano-carriers for the delivery of chemotherapeutic drugs to cancerous cells. We examined the effects of -OH and -COO- groups on CNTs surface for absorption of two anticancer drugs including daunorubicin and etoposide using molecular dynamics simulation and experimental assays. To evaluate the absorption of each drug in each CNT, the complexes of drugs/CNTs in water were simulated separately. Theoretical investigation demonstrated that CNT-OH and CNT-COO- are more suitable for absorption of daunorubicin and etoposide, respectively. Experimental findings also confirmed molecular dynamics simulation results. Communicated by Ramaswamy H. Sarma.

Rational Design of Anti-Angiogenic Peptides to Inhibit VEGF/VEGFR2 Interactions for Cancer Therapeutics.

BACKGROUND: Angiogenesis is a critical physiological process that plays a key role in tumor progression, metastatic dissemination, and invasion. In the last two decades, the vascular endothelial growth factor (VEGF) signaling pathway has been the area of extensive researches. VEGF executes its special effects by binding to vascular endothelial growth factor receptors (VEGFRs), particularly VEGFR-2. OBJECTIVE: The inhibition of VEGF/VEGFR2 interaction is known as an effective cancer therapy strategy. The current study pointed to design and model an anti-VEGF peptide based on VEGFR2 binding regions. METHOD: The large-scale peptide mutation screening was used to achieve a potent peptide with high binding affinity to VEGF for possible application in inhibition of VEGF/VEGFR2 interaction. The AntiCP and Peptide Ranker servers were used to generate the possible peptides library with anticancer activities and prediction of peptides bioactivity. Then, the interaction of VEGF and all library peptides were analyzed using Hex 8.0.0 and ClusPro tools. A number of six peptides with favorable docking scores were achieved. All of the best docking scores of peptides in complexes with VEGF were evaluated to confirm their stability, using molecular dynamics simulation (MD) with the help of the GROMACS software package. RESULTS: As a result, two antiangiogenic peptides with 13 residues of PepA (NGIDFNRDFFLGL) and PepC (NGIDFNRDKFLFL) were achieved and introduced to inhibit VEGF/VEGFR2 interactions. CONCLUSIONS: In summary, this study provided new insights into peptide-based therapeutics development for targeting VEGF signaling pathway in tumor cells. PepA and PepC are recommended as potentially promising anticancer agents for further experimental evaluations.

Synthesis, Characterization and Fluorescence Properties of Novel Porous Fe/ZnO Nano-Hybrid Assemblies by Using Berberis thunbergii Extract.

In this work, novel Fe/ZnO nanocomposites (NCs) and Fe nanoparticles loaded onto porous ZnO nanostructures have been synthesized via a simple biotechnological route by using Berberis thunbergii extract. In this direction, the as-synthesized bio-based porous ZnO derivatives and human serum albumin (HSA), as a biopolymeric model, form nano-hybrid assemblies. The effect of loading Fe on properties of porous ZnO nanostructures as well as the behavior of the nano-hybrid assemblies were evaluated by using XRD, SEM, EDX, DLS, PL, CD, FTIR and UV/Visible-diffuse reflectance spectra (UV/Vis-DRS) techniques. The fluorescence results revealed that the interaction of Fe/ZnO NCs with HSA biopolymer led to the formation of a ground state complexes as nano-hybrid assemblies. The calculated thermodynamic parameters indicated that the binding process occurred spontaneously. The CD and FTIR spectra confirmed the changes in helicity of HSA as well as the random coil and ß-turn in the secondary structure of HSA upon interaction with Fe/ZnO NCs.

Classification of seed members of five riboswitch families as short sequences based on the features extracted by Block Location-Based Feature Extraction (BLBFE) method.

Introduction: Riboswitches are short regulatory elements generally found in the untranslated regions of prokaryotes' mRNAs and classified into several families. Due to the binding possibility between riboswitches and antibiotics, their usage as engineered regulatory elements and also their evolutionary contribution, the need for bioinformatics tools of riboswitch detection is increasing. We have previously introduced an alignment independent algorithm for the identification of frequent sequential blocks in the families of riboswitches. Herein, we report the application of block location-based feature extraction strategy (BLBFE), which uses the locations of detected blocks on riboswitch sequences as features for classification of seed sequences. Besides, mono- and dinucleotide frequencies, k-mer, DAC, DCC, DACC, PC-PseDNC-General and SC-PseDNC-General methods as some feature extraction strategies were investigated. Methods: The classifiers of the Decision tree, KNN, LDA, and Naïve Bayes, as well as k-fold cross-validation, were employed for all methods of feature extraction to compare their performances based on the criteria of accuracy, sensitivity, specificity, and f-score performance measures. Results: The outcome of the study showed that the BLBFE strategy classified the riboswitches indicating 87.65% average correct classification rate (CCR). Moreover, the performance of the proposed feature extraction method was confirmed with average values of 94.31%, 85.01%, 95.45% and 85.38% for accuracy, sensitivity, specificity, and f-score, respectively. Conclusion: Our result approved the performance of the BLBFE strategy in the classification and discrimination of the riboswitch groups showing remarkable higher values of CCR, accuracy, sensitivity, specificity and f-score relative to previously studied feature extraction methods.

New Developments in Anti-Angiogenic Therapy of Cancer, Review and Update.

BACKGROUND: Angiogenesis is one of the critical physiological processes, by which the new blood vessels are generated from the pre-existing vessels in the early stage of vasculogenesis. While normal angiogenesis is critical for development and tissue growth, pathologic angiogenesis is important for the growth and spread of cancers by supplying nutrients and oxygen as well as providing a conduit for distant metastasis. In the last two decades, angiogenesis has been the area of extensive researches, indicating antiangiogenic target therapy as an effective strategy for cancer therapy. At present, this field has become a major avenue for research and development of novel therapeutics. OBJECTIVE: This review is dedicated to an updated review of the most prominent antiangiogenic agents, emphasizing the novel advancements and their applications, in particular, in the fields of antibodies, peptides, vaccines, endogenous inhibitors, Nanoparticles (NPs), antiangiogenic oligonucleotides and small molecules. Also, the potential role of 3D microfluidic models as an affordable and time-saving tool for angiogenesis investigations are discussed. METHODS: Firstly, we collected and summarized new developments that have occurred in all review and research articles in databases. Then, we used important keywords related to antiangiogenic target therapy and their applications for retrieval of most relevant data. RESULTS: This review is based on recent research and review articles and intended to cover all newly discovered agents inhibiting tumor angiogenesis and in particular, VEGF. CONCLUSION: New research studies have shown that anti-angiogenesis agents especially, in the form of combination therapy are effective in various cancers treatment.

Simulations on the dual effects of flavonoids as suppressors of Aß42 fibrillogenesis and destabilizers of mature fibrils.

Structural studies of the aggregation inhibition of the amyloid-ß peptide (Aß) by different natural compounds are of the utmost importance due to their great potential as neuroprotective and therapeutic agents for Alzheimer's disease. We provided the simulation of molecular dynamics for two different states of Aß42, including "monomeric aggregation-prone state (APS)" and "U-shaped pentamers of amyloidogenic protofilament intermediates" in the absence and presence of polyphenolic flavonoids (Flvs, myricetin and morin) in order to verify the possible mechanism of Flvs fibrillogenesis suppression. Data showed that Flvs directly bind into Aß42 species in both states of "monomeric APS ß-sheets" and "pentameric amyloidogenic intermediates". Binding of Flvs with amyloidogenic protofilament intermediates caused the attenuation of some inter-chains H-bonds, salt bridges, van der Waals and interpeptide interaction energies without interfering with their secondary ß-sheets. Therefore, Flvs redirect oligomeric amyloidogenic intermediates into unstructured aggregates by significant disruption of the "steric zipper" motif of fibrils-pairs of self-complementary ß-sheets-without changing the amount of ß-sheets. It is while Flvs completely destruct the disadvantageous secondary ß-sheets of monomeric APS conformers by converting them into coil/helix structures. It means that Flvs suppress the fibrillogenesis process of the monomeric APS structures by converting their ß-sheets into proper soluble coil/helices structures. The different actions of Flvs in contact with two different states of Aß conformers are related to high interaction tendency of Flvs with additional H-bonds for monomeric APS ß-sheet, rather than oligomeric protofilaments. Linear interaction energy (LIE) analysis confirmed the strong binding of monomeric Aß-Flvs with more negative ∆Gbinding, rather than oligomeric Aß-Flvs system. Therefore, atomic scale computational evaluation of Flvs actions demonstrated different dual functions of Flvs, concluded from the application of two different monomeric and pentameric Aß42 systems. The distinct dual functions of Flvs are proposed as suppressing the aggregation by converting ß-sheets of monomeric APS to proper soluble structures and disrupting the "steric zipper" fibril motifs of oligomeric intermediate by converting on-pathway into off-pathway. Taken together, our data propose that Flvs exert dual and more effective functions against monomeric APS (fibrillogenesis suppression) and remodel the Aß aggregation pathway (fibril destabilization).

Structure-based designing efficient peptides based on p53 binding site residues to disrupt p53-MDM2/X interaction.

MDM2 and MDMX are known as overexpressed oncoproteins in several wild-type p53 cancer cells. The development of potent and dual antagonist peptides for p53-MDM2/X is a continuous challenge. In this study, we intended to investigate the pivotal structural points respecting the development of potent and dual inhibitors of MDM2/X. Correspondingly, MD simulation was performed on the experimentally confirmed peptides, comprising p53, pDI, pDIQ, PMI, and computationally screened mutant pDI and pDIQ. A follow-up secondary structure analysis showed the last three C-terminal residues provide the helicity reservation of peptides bound to MDM2/X. Furthermore, a delicate residue-residue examination displayed Met 11 and Ser12 in the modified peptides contribute significantly to dual inhibition of MDM2/X. Additionally, the peptides_MDM2/X complexes' ΔGbinding extracted by the umbrella sampling method were in agreement with the pattern of their experimental affinity values. It was concluded the screened pDI mutants were considered as suitable anti-MDM2/X peptides, and the data obtained could be exploited as the theoretical structure-based guide for rational peptide design. Taking account of results, the suitable C-terminal residues of p53-based peptides especially Met11, and Ser12, as well as higher umbrella sampling, generated ΔGbinding to MDM2/X would be considered as the positive structural markers of a promising anti-cancer agent.

Design and development of high affinity dual anticancer peptide-inhibitors against p53-MDM2/X interaction.

AIMS: Inhibition of P53-MDM2/X interaction is known as an effective cancer therapy strategy. In this regard, pDI peptide was introduced previously with the potential of targeting MDM2. In this research, the large-scale peptide mutation screening was used to achieve the best sequence of pDI with the highest affinity for inhibition activity against MDM2/X. MAIN METHODS: Three mutant peptides of pDI as dual inhibitor peptides including single mutations of pDIm/4W, pDIm/11M and double mutations of pDIdm/4W11M were presented with the high affinities to inhibit both MDM2/X. The selected mutants were then evaluated comprehensively to confirm their ability as potent MDM2/X inhibitors, using a theoretical simulation approach. KEY FINDINGS: MD simulations analyses confirmed their dual inhibition potential against both MDM2/X interactions with p53 protein. The developed pDIm and mainly pDIdm peptides showed stable conformations over the simulation time with conserved secondary structure and effective interaction with MDM2/X by physical binding such as hydrogen bonding. Besides, umbrella sampling free energy calculation indicated higher binding energy, ΔGbinding, of pDIm-MDM2/X and pDIdm-MDM2/X compared to pDI-MDM2/X. SIGNIFICANCE: The optimized and improved mutant pDI, pDIdm, with more effective ΔGbinding values of -30 and -25 kcal/mol to MDMX and MDM2, respectively, is recommended as a promising anticancer agent and suitable candidate for experimental evaluations.

Classification of Riboswitch Families Using Block Location-Based Feature Extraction (BLBFE) Method.

Purpose: Riboswitches are special non-coding sequences usually located in mRNAs' un-translated regions and regulate gene expression and consequently cellular function. Furthermore, their interaction with antibiotics has been recently implicated. This raises more interest in development of bioinformatics tools for riboswitch studies. Herein, we describe the development and employment of novel block location-based feature extraction (BLBFE) method for classification of riboswitches. Methods: We have already developed and reported a sequential block finding (SBF) algorithm which, without operating alignment methods, identifies family specific sequential blocks for riboswitch families. Herein, we employed this algorithm for 7 riboswitch families including lysine, cobalamin, glycine, SAM-alpha, SAM-IV, cyclic-di-GMP-I and SAH. Then the study was extended toward implementation of BLBFE method for feature extraction. The outcome features were applied in various classifiers including linear discriminant analysis (LDA), probabilistic neural network (PNN), decision tree and k-nearest neighbors (KNN) classifiers for classification of the riboswitch families. The performance of the classifiers was investigated according to performance measures such as correct classification rate (CCR), accuracy, sensitivity, specificity and f-score. Results: As a result, average CCR for classification of riboswitches was 87.87%. Furthermore, application of BLBFE method in 4 classifiers displayed average accuracies of 93.98% to 96.1%, average sensitivities of 76.76% to 83.61%, average specificities of 96.53% to 97.69% and average f-scores of 74.9% to 81.91%. Conclusion: Our results approved that the proposed method of feature extraction; i.e. BLBFE method; can be successfully used for classification and discrimination of the riboswitch families with high CCR, accuracy, sensitivity, specificity and f-score values.

Cell sheet biofabrication by co-administration of mesenchymal stem cells secretome and vitamin C on thermoresponsive polymer.

Cell sheet technology aims at replacement of artificial extracellular matrix (ECM) or scaffolds, popular in tissue engineering, with natural cell derived ECM. Adipose tissue mesenchymal stem cells (ASCs) have the ability of ECM secretion and presented promising outcomes in clinical trials. As well, different studies found that secretome of ASCs could be suitable for triggering cell free regeneration induction. The aim of this study was to investigate the effect of using two bio-factors: secretome of ASCs (SE) and vitamin C (VC) for cell sheet engineering on a thermosensitive poly N-isopropyl acryl amide-Methacrylic acid (P(NIPAAm-MAA)) hydrogel. The results revealed that using thermosensitive P(NIPAAm-MAA) copolymer as matrix for cell sheet engineering lead to a rapid ON/OFF adhesion/deadhesion system by reducing temperature without enzymatic treatment (complete cell sheet release takes just 6 min). In addition, our study showed the potential of SE for inducing ASCs sheet formation. H&E staining exhibited the properties of a well-formed tissue layer with a dense ECM in sheets prepared by both SE and VC factors, as compared to those of VC or SE alone. Functional synergism of SE and VC exhibited statistically significant enhanced functionality regarding up-regulation of stemness genes expression, reduced ß-galactosidase associated senescence, and facilitated sheet release. Additionally, alkaline phosphatase activity (ALP), mineralized deposits and osteoblast matrix around cells confirmed a better performance of ostogenic differentiation of ASCs induced by VC and SE. It was concluded that SE of ASCs and VC could be outstanding biofactors applicable for cell sheet technology.

17-DMAG-loaded nanofibrous scaffold for effective growth inhibition of lung cancer cells through targeting HSP90 gene expression.

Up-regulation of heat shock protein 90 (HSP90) gene takes place in lung cancer cells. Therefore, targeting HSP90 in lung cancer may be promising step in lung cancer therapy. The present study aimed to evaluate the efficiency of implantable 17-dimethylaminoethylamino-17-demethoxy geldanamycin (17-DMAG)-loaded Poly(caprolactone)-poly(ethylene glycol) (PCL/PEG) nanofibers to increase the anti-cancer effects via inhibition of HSP90 expression and telomerase activity. For this purpose, 17-DMAG-loaded PCL/PEG nanofibers were successfully fabricated via electrospinning and characterized using FE-SEM and FTIR. Colorimetric MTT assay was used to determine the drug cytotoxicity. Also, the expression levels of HSP90 mRNA in the A549 cells treated with the nanofibers were assessed using Quantitative Real-Time PCR. The effect of free 17-DMAG and 17-DMAG-loaded PCL/PEG nanofiber treatment on telomerase activity was monitored by TRAP assay. MTT assay confirmed that loading of 17-DMAG into PCL/PEG nanofiber enhanced dramatically cytotoxicity in the lung cancer cells. This finding was associated with reduction of HSP90 mRNA expression and telomerase activity in the cells seeded on 17-DMAG-loaded PCL/PEG nanofibers in relative to free 17-DMAG. In conclusion, the findings demonstrated that 17-DMAG-loaded PCL/PEG nanofibers are more effectual than free 17-DMAG against A549 lung cancer cells via modulation of Hsp90 expression and inhibition of telomerase activity. Hence, the implantable 17-DMAG-loaded nanofibrous scaffolds might be an excellent tool for efficiently killing of the lung residual cancer cells and avoid the local cancer recurrence.