Search Results (254)

Streptococcus canis is an opportunistic pathogen that colonises the mucosal surfaces and skin of its host. Though predominantly a veterinary pathogen affecting cats and dogs, S. canis has also been identified as the causative agent in severe human disease. IdeC is a secreted cysteine protease of S. canis that has a high specificity for IgG, cleaving at the hinge region. We show here that the protein binds back to the surface of the bacteria. Additionally, the protein contains a conserved Arg-Gly-Asp (RGD) motif, the minimal peptide sequence required for integrin binding. Several bacterial proteins containing RGD motifs have been implicated in adhesion and invasion of host cells. This RGD motif along with the ability of IdeC to bind back to the bacterial surface after secretion is the basis for this study into a potential secondary function of IdeC in adhesion and/or invasion. We used protein-coated latex beads to investigate the interaction of IdeC with epithelial and endothelial cells and, further, the extent to which the RGD motif is involved in this interaction by utilising an RGD->RGE recombinant protein. We also report here that the deletion of IdeC in S. canis results in a significant reduction in invasion into epithelial cells. Full article

We utilized molecular dynamics (MD) simulations to explore the interaction of the RGD peptide with the αvβ3 integrin receptor, a key process for targeted drug delivery to tumors. The goal of these simulations was to model the transport of boron atoms by the RGD peptide and to characterize the binding event between this vector and its receptor. The study focused on the interaction processes and spatial arrangements of the solvated RGD–integrin system. Simulations were run for 100 ns to achieve relaxed-state configurations. Our model featured two RGD peptides: one pre-localized within the integrin’s binding site and another initially positioned externally. The external peptide was observed to diffuse freely and subsequently bind to the αvβ3 integrin. This spontaneous binding event provides valuable insights into the pharmacological specificity and mechanisms of the RGD–integrin interaction, informing the design of effective drug delivery systems. Full article

Although the parental recombinant protein rLj-RGD3 exhibits antitumor activity, it carries immunogenicity risks owing to its large molecular size (13.5 kDa). We generated a genetically truncated mutant, rLj-RGD4 (6.27 kDa, four RGD motifs), which inhibited B16 melanoma cell proliferation, migration, and invasion in vitro. However, the in vivo efficacy and mechanisms of action remain unclear. Here, B16 xenograft mice were treated with rLj-RGD4 (5, 10, and 20 μg/kg i.p. daily for 14 days). Tumor growth was measured, and histopathology/apoptosis was evaluated using hematoxylin and eosin (HE), Masson’s dye, Hoechst, and TUNEL staining. Apoptotic pathways (mitochondrial, death receptor, and MAPK) were analyzed via Western blotting, whereas endocytosis mechanisms were explored using inhibitors (filipin III, NaN₃, cytochalasin D), and EGFR (epidermal growth factor receptor) interactions via fluorescence co-localization and phosphoprotein assays. The results demonstrated dose-dependent tumor growth inhibition (21.60–89.26% volume reduction, 41.03–86.51% weight reduction), with histological evidence of tissue loosening, fibrosis, and apoptosis. rLj-RGD4 induced apoptosis by activating the mitochondrial (Bax/Bcl-2 upregulation), death receptor (caspase-8 activation), and MAPK (JNK/p38 phosphorylation) pathways. Internalization was blocked by NaN₃ and cytochalasin D, indicating actin-dependent macropinocytosis. Direct EGFR binding was confirmed, accompanied by reduced EGFR expression and the inhibition of FAK/AKT/Src signaling. In conclusion, rLj-RGD4 exerts potent in vivo antitumor activity via two mechanisms: induction of multi-pathway apoptosis and EGFR-targeted suppression of pro-survival signaling. RGD4 exerts its antitumor function in vivo by targeting and co-internalizing with EGFR. Full article

Monitoring the concentration of doxorubicin (DOX) was critical for tumor treatment, but existing methods failed to cross cell membrane. Here, an electrochemical platform for intracellular DOX detection in MCF-7 cells based on membrane-permeation strategy was developed. A modified gold electrode was prepared via electrodepositing AuNPs and assembling SH-DNA. Concurrently, the silica nanosphere/gold nanocluster-circular transmembrane peptide (SiO₂/AuNCs-iRGD) composite nanoparticles with membrane permeability, tumor targeting, and imaging capability were synthesized. After co-incubation of SiO₂/AuNCs-iRGD with MCF-7 cells and DOX, followed by co-incubation with the DNA-modified electrode, intracellular DOX intercalated into the DNA backbone, and redox-generated electrons were transferred to the electrode to produce a concentration-correlated electrochemical signal. The modification of the electrode, the morphology of the composite nanoparticles and the detection process were characterized by means of SEM, TEM, CV, EIS, DPV, fluorescence spectroscopy and laser confocal imaging. Under the optimized conditions, the proposed method exhibited a wide detection range of 0.05–300 μmol/L, with a detection limit of 0.01 μmol/L. Moreover, the modified electrode demonstrated satisfactory regenerability, and the proposed method showed excellent reproducibility and stability. The development platform could offer a new strategy for real-time assessment of drug concentration within cultured breast cancer cells in vitro. Full article

Primary hepatocyte cultures serve as an ex vivo model of liver physiology. This study aims to employ poly(vinyl alcohol) (PVA) nanofiber membranes (NMs) to establish a three-dimensional (3D) culture system that supports the long-term functionality of primary hepatocytes. Primary hepatocytes were monocultured on a PVA NM or indirectly cocultured with NIH3T3 fibroblasts on a distinct polycaprolactone (PCL) NM layer. Monocultured and cocultured hepatocytes maintained prolonged survival without supplemental growth factors. Cocultured hepatocytes formed larger aggregates composed of cell clusters attached to untreated nanofibers than monocultured cells. However, most primary hepatocytes cultured on NaOH-treated PVA NM and Arg–Gly–Asp (RGD) peptide-blended PVA (RGD-PVA) NM, under monoculture and coculture conditions, formed non-aggregated cells in a single-cell layer. In a bioinert assay, unstimulated dendritic cells were activated on untreated but not NaOH-treated PVA NM. CYP3A4 activity was higher in cocultured cells on RGD-PVA NM with fibroblasts than in monocultured cells on PVA and RGD-PVA NM. Functional hepatocyte cultures were successfully maintained in a 3D single-cell layer on RGD-PVA NM, along with fibroblasts in a layer-by-layer coculture, for a prolonged period. The prolonged culture of hepatocytes in a 3D single-cell layer may facilitate further drug discovery, toxicity studies, and translational liver research. Full article

Bioactive peptide coatings modulate cell–implant interactions on TiO₂ surfaces; however, most molecular-level studies of peptide adsorption are performed under low or fixed ionic conditions. Physiological environments exhibit non-negligible and variable electrolyte concentrations, so understanding ionic strength effects is crucial for designing effective peptide-functionalized titanium implants. An amorphous TiO₂ surface was generated from a crystalline rutile precursor and simulated in explicit water using classical molecular dynamics at nine NaCl concentrations. For each condition, seven independent simulations with different initial peptide placements/orientations were performed. Peptide backbone RMSD, minimum peptide–surface distance, and adsorption time ratio were analysed as functions of NaCl concentration. For both peptides, backbone RMSD remained stable and showed no statistically significant correlation with NaCl concentration. KRSR exhibited a significant increase in minimum distance with increasing NaCl concentration and a significant decrease in adsorption time ratio, indicating reduced persistence of close surface contact at higher salt levels. In contrast, RGD showed no significant dependence of either minimum distance or adsorption time ratio within the tested range. Within the limits of the applied force-field MD framework and the investigated NaCl range, KRSR adsorption on TiO₂ is more sensitive to ionic strength than RGD, consistent with the stronger electrostatic contribution for the net-positively charged KRSR motif. Full article

Due to their therapeutic potential and effects on cells, exosomes derived from bovine colostrum (BCE) and milk (BME) are molecules that have been at the center of recent studies. Their properties include the ability to cross biological barriers, their natural biocompatibility, and their structure, which enable them to act as stable nanocarriers. Exosomes derived from milk and colostrum stand out in cancer prevention and treatment due to these properties. BMEs can be enriched with bioactive peptides, lipids, and nucleic acids. The targeted drug delivery capacity of BMEs can be made more efficient through these enrichment processes. For example, BME enriched with an iRGD peptide and developed using hypoxia-sensitive lipids selectively transported drugs and reduced the survival rate of triple-negative breast cancer (TNBC) cells. ARV-825-CME formulations increased antitumor activity in some cancer types. The anticancer effects of exosomes are supported by these examples. In addition to their anticancer activities, exosomes also exhibit effects that maintain immune balance. BME and BCE can regulate inflammatory responses with their miRNA and protein loads. These effects of BMEs have been demonstrated in studies on colon, breast, liver, and lung cancers. The findings support the safety and scalability of these effects. However, significant challenges remain in terms of their large-scale isolation, load heterogeneity, and regulatory standardization. Consequently, BMEs represent a new generation of biogenic nanoplatforms at the intersection of nutrition, immunology, and oncology, paving the way for innovative therapeutic approaches. Full article

Background: Lactate Dehydrogenase C (LDHC) is a promising therapeutic target due to its highly tumor-specific expression, immunogenicity, and oncogenic functions. We previously showed that LDHC silencing in triple-negative breast cancer (TNBC) cells enhances treatment response to DNA-damage response-related drugs, supporting its therapeutic potential. However, no selective LDHC inhibitors exist, highlighting the need for innovative targeting strategies. Methods: We assessed the physicochemical properties and evaluated the delivery efficiency, anti-tumor activity, and safety of four cell-penetrating peptides (CPPs)—R10, 10R-RGD, cRGD-10R, and iRGD-10R—for siRNA-mediated LDHC silencing in TNBC. Clonogenic assays were used to evaluate effects on olaparib sensitivity, and TNBC zebrafish xenografts were utilized to study in vivo anti-tumor activity. Results: All CPP:siRNA complexes formed uniform nanocomplexes (129–168 nm) with low polydispersity indices (<0.25) and positive zeta potentials (+6.47 to +29.6 mV). Complexes remained stable in human serum for 24 h and showed no significant cytotoxicity in TNBC and non-cancerous cell lines. The 10R-RGD and cRGD-10R:siLDHC complexes achieved 40% LDHC protein knockdown, reduced TNBC clonogenicity by 30–36%, and enhanced olaparib sensitivity. Treatment of TNBC zebrafish xenografts with 10R-RGD or cRGD-10R:siLDHC complexes significantly reduced tumor growth by approximately 50% without major toxicity. Conclusions: These results demonstrate that CPP-mediated siRNA delivery enables selective LDHC silencing with tumor growth inhibition in triple-negative breast cancer models. This approach represents a novel, effective, and safe proof-of-concept therapeutic strategy to target LDHC, with potential translational relevance as a standalone therapy or in combination with common anti-cancer drugs. Full article

Cutaneous melanoma is an extremely dangerous tumor disease with poor prognosis at advanced stages. Accounting for a small percentage of all skin tumors, malignant melanoma leads the mortality rate in this group of cancers. Clearly, the search for new drugs and therapeutic approaches for the treatment of cutaneous melanoma is a highly pressing issue in modern medicine. In this study, novel recombinant proteins with anti-melanoma activity, called RGD-apoptins, were produced in an E. coli expression system, and their properties were evaluated in human cell models. These chimeric proteins consist of two parts, each tumor-specific. One part of the chimeric molecule is the RGD peptide, which binds to αVβ3 integrins widely expressed on the surface of malignant melanocytes. The other part is the viral protein apoptin, known to induce programmed cell death in tumor cells but not in normal cells. This molecular design aims to enhance the specificity of potential therapeutic agent toward malignant melanoma cells while reducing cytolytic effects on healthy tissue. In a resazurin assay, RGD-apoptins decreased the viability of MeWo human melanoma cells and did not affect the viability of HaCaT human keratinocyte cell line and primary skin fibroblasts. Using an annexin V assay, we confirmed that malignant melanocytes death occurs via apoptosis. Transcriptomic analysis allowed us to dynamically evaluate the spectrum of differentially expressed genes 24 and 48 h after treating melanoma cells with recombinant RGD-apoptin. Full article

Selenium nanoparticles (SeNPs) have emerged as promising metal-based nanoparticles for drug delivery due to their unique physicochemical properties, intrinsic bioactivity, and biocompatibility. SeNPs offer a lower toxicity, higher bioavailability, and flexibility to be customized for surface chemistry compared to traditional selenium compounds. Advances in synthetic strategies, including chemical reduction, green biosynthesis, and surface functionalization with polymers, peptides, or ligands, have improved their stability, targeting capability, and circulation time. SeNP-based systems have demonstrated unique anticancer, antimicrobial, and anti-inflammatory activities, as they can function as drug carriers and active therapeutic agents. The surface of SeNPs has been functionalized with ligands such as Arginylglycylaspartic acid (RGD) peptides, hyaluronic acid, or chitosan to enhance their receptor-mediated targeting abilities in tumor tissues. In addition, SeNPs have shown a synergistic effect in the presence of drugs such as doxorubicin and paclitaxel. Even though SeNPs have demonstrated significant potential in pre-clinical investigations, their use in clinical studies has not been expanded due to several limiting challenges, including large-scale production, long-term safety, pharmacokinetic properties, and regulations required for FDA approval. Continued research into optimizing formulation strategies and expanding in vivo validation will be critical to translating SeNP-based drug delivery systems into clinical applications. In this review, we focus on the methods for synthesizing SeNPs, their physicochemical properties, the structure of ligands attached to SeNPs for drug delivery applications, and the specific biological targets of functionalized SeNPs. Full article

A promising strategy for pancreatic cancer therapy involves developing nanocarriers capable of simultaneously delivering various antitumor substances with diverse physicochemical properties, often resulting in synergistic effects. In the present work, novel biocomposites were developed using amphiphilic N-vinylpyrrolidone polymer incorporating bortezomib (BTZ) and modified with either the DR5-selective TRAIL cytokine (DR5-B) or its fusion with the iRGD effector peptide (DR5-B-iRGD), resulting in AmphPVP-BTZ-DR5-B and AmphPVP-BTZ-DR5-B-iRGD formulations. The release of BTZ was most extensive at acidic pH 5.6, mimicking endolysosomal compartments, while at near-neutral pH 7.4 and alkaline pH 8.2 the release was slower and less complete, indicating a smart pH-responsive behavior suitable for triggered release in the tumor microenvironment. Both AmphPVP-BTZ-DR5-B and AmphPVP-BTZ-DR5-B-iRGD significantly inhibited the growth of pancreatic adenocarcinoma cell lines PANC-1, BxPC-3, and MIA PaCa-2 and induced more rapid internalization of the DR5 receptor in MIA PaCa-2 cells than unmodified particles and free DR5-B or DR5-B-iRGD. Importantly, AmphPVP-BTZ-DR5-B-iRGD exhibited a more pronounced DR5 internalization rate and cytotoxic effect than AmphPVP-BTZ-DR5-B owing to the presence of fusion protein with internalizing iRGD peptide. Both biocomposites induced cell death via the apoptotic pathway while exhibiting minimal cytotoxic effects on healthy cells. Therefore, biocomposites incorporating BTZ and functionalized with DR5-selective TRAIL variants DR5-B or DR5-B-iRGD represent a promising avenue for future studies in pancreatic cancer animal models. Full article

Liposomes modified with slightly acidic pH-sensitive peptides (SAPSp-lipo) are effectively delivered to tumor tissues, followed by cellular uptake in the tumor microenvironment. Although SAPSp-lipo can penetrate tumor tissues via the interspace route between cancer cells and the extracellular matrix (ECM), penetration needs to be enhanced to deliver liposomes into tumor cores comprising malignant cancer cells. To enhance the intratumoral penetration of SAPSp-lipo, we focused on the internalizing RGD peptide (iRGD), which can penetrate tumor tissue, differing from the penetration mechanism of SAPSp. In this study, we developed liposomes modified with iRGD-conjugated SAPSp (SAPSp-iRGD-lipo). Compared with SAPSp-lipo, SAPSp-iRGD-lipo was delivered to deeper regions within both spheroids and tumor tissues. The enhanced penetration was suppressed by a co-treatment with a Neuropilin-1 inhibitor, and the fluorescence signals from intratumorally injected SAPSp-iRGD-lipo were localized in Neuropilin-1-expressing regions, indicating a Neuropilin-1-mediated tumor penetration. Moreover, SAPSp-iRGD-lipo reduced F-actin formation in monolayered cells and was not localized in F-actin-rich regions in tumors, suggesting that SAPSp-iRGD-lipo facilitates tumor penetration through actin depolymerization. In addition, anticancer siRNA delivered by SAPSp-iRGD-lipid nanoparticles effectively induced apoptosis in cells under slightly acidic conditions. Taken together, SAPSp-iRGD-modified nanoparticles represent a novel class of tumor-penetrable and microenvironment-responsive drug carriers capable of efficient intratumoral delivery and therapeutic activity. Full article

Objectives: This paper reports the preclinical evaluation of stable tumor-specific gold nanoparticles (AuNPs) activated by neutron irradiation as a therapeutic option for the treatment of cancers characterized by high tumor angiogenesis. Methods: A selection of promising AuNPs with high avidity to α_vβ₃-expressing glioma (U-87 MG) cells (IC₅₀ = 82–104 nM) were chosen with different surface loading of Arg-Gly-Asp (RGD) peptides as tumor targeting vectors for integrin α_vβ₃, a target which is overexpressed in tissues displaying high tumor angiogenesis. Three different [¹⁹⁸Au]AuNPs were evaluated applying three injection methods, intravenous (i.v.), intraperitoneal (i.p.), and intratumoral (i.t.), each in a group of six U-87 MG xenograft–bearing mice (54 female athymic nude mice in total). Their biodistribution and tumor accumulation was assessed by in vivo imaging within 1–7 days after injection and 7 days after injection by ex vivo measurement. Results: The developed [¹⁹⁸Au]AuNPs exhibited suboptimal biodistribution by i.v. application (accumulation pattern tail > liver > spleen, no significant tumor accumulation) and by i.p. application (accumulation pattern spleen >> liver > pancreas, slight tumor accumulation of <0.3 %ID/g). However, an acceptable biodistribution by i.t. application was observed (5.5 %ID/g in liver, 4.9 %ID/g in spleen, and 3.0 %ID/g in tumor). Conclusions: Despite the very promising in vitro results, the in vivo evaluation suggests that the [¹⁹⁸Au]AuNPs represent a platform for the development of restricted therapeutic strategies. Full article

Carboxymethyl chitosan (CMC)-based hydrogels have emerged as promising candidates for wound dressing applications due to their excellent biocompatibility and tunable physicochemical properties. In this study, a novel hydrogel functionalized with hyaluronic acid (HA) and RGD peptides (RGD) was fabricated and evaluated for its structural characteristics and wound-healing potential. Using CMC as the base matrix and EDC/NHS as crosslinking agents, four hydrogel variants were fabricated: CMC gel, CMC-HA gel, CMC-RGD gel, and CMC-HA-RGD gel. The preliminary cell compatibility experiment identified the optimal formulation as 1% CMC, 0.9% HA, and 0.02 mg/mL RGD, crosslinked with 1 vol% EDC and 0.05 wt% NHS. Scanning electron microscopy showed a porous architecture (100–400 μm), conducive to fibroblast viability and proliferation. Zeta potential measurements (|ζ| > 30 mV) indicated colloidal stability of the hydrogel system. Fourier-transform infrared spectroscopy confirmed successful crosslinking and integration of HA and RGD via hydrogen bonding and electrostatic interactions, forming a stable three-dimensional network. Thermogravimetric analysis revealed enhanced thermal stability upon HA/RGD incorporation. CCK-8 assays demonstrated significantly improved cell viability with HA/RGD loading (p < 0.05), while Ki-67 immunofluorescence confirmed enhanced fibroblast proliferation, with the CMC-HA-RGD gel showing the most pronounced effect. In vitro scratch assay results demonstrated that the CMC-HA-RGD hydrogel dressing significantly enhanced cellular migration compared to other carboxymethyl chitosan-based hydrogel groups (p < 0.05). The observed statistically significant improvement in cell migration rate versus controls underscores the distinctive enhancement of synergistic HA and RGD modification in accelerating cellular migration and facilitating wound repair. Collectively, these findings suggest that the CMC-HA-RGD hydrogel possesses favorable physicochemical and biological properties and holds strong potential as an advanced wound dressing for the treatment of chronic and refractory wounds. Full article

Background/Objectives: Conventional therapeutic strategies exhibit limited efficacy against pancreatic cancer, primarily due to its profoundly hypoxic tumor microenvironment and dense fibrotic stroma. Chemodynamic therapy (CDT) holds promise; however, its application in pancreatic cancer is restricted by insufficient endogenous hydrogen peroxide (H₂O₂) levels and the activation of protective autophagy in response to oxidative stress. Methods: To overcome these obstacles, we developed a tumor microenvironment-responsive, pancreatic cancer-targeted CDT nanoamplifier—H-MnO₂/GOX&CQ-iRGD—comprising a hollow mesoporous MnO₂ shell co-loaded with glucose oxidase (GOX) and chloroquine (CQ), and surface-functionalized with the tumor-penetrating peptide iRGD. GOX catalyzes glucose oxidation to generate H₂O₂, enhancing Fenton-like reactions. CQ suppresses autophagy induced by oxidative stress, thereby alleviating therapy resistance. The iRGD peptide targets integrin α_vβ₃, which is overexpressed on pancreatic cancer cells and tumor vasculature, promoting deep tumor penetration and enhanced delivery efficiency. Results: We comprehensively characterized the nanoplatform’s physicochemical properties, tumor microenvironment triggered degradation, controlled drug release, glucose-driven H₂O₂ generation, and hydroxyl radical production in vitro. Cellular studies assessed nanoparticle uptake, intracellular H₂O₂ production, autophagy inhibition, and cytotoxicity. In vivo experiments further demonstrated effective tumor targeting and significant therapeutic outcomes in pancreatic cancer models. Conclusions: This nanoplatform addresses major barriers of CDT—namely, insufficient H₂O₂ levels, autophagy-mediated resistance, and limited intratumoral penetration—offering a promising strategy for pancreatic cancer treatment. Full article