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Prior research has shown the effectiveness of dalbergin (DL), dalbergin nanoformulation (DLF), and dalbergin-loaded PLGA-galactose-modified nanoparticles (DLMF) in treating hepatocellular carcinoma (HCC) cells. The present investigation constructs upon our previous research and delves into the molecular mechanisms contributing to the anticancer effects of DLF and DLMF. This study examined the anti-cancer effects of DL, DLF, and DLMF by diethyl nitrosamine (DEN)-induced HCC model in albino Wistar rats. In addition, we performed biochemical, antioxidant, lipid profile tests, and histological studies of liver tissue. The anticancer efficacy of DLMF is equivalent to that of 5-fluorouracil, a commercially available therapy for HCC. Immunoblotting studies revealed a reduction in the expression of many apoptotic markers, such as p53, BAX, and Cyt-C, in HCC. Conversely, the expression of Bcl-2, TNF-α, NFκB, p-AKT, and STAT-3 was elevated. Nevertheless, the administration of DL, DLF, and DLMF effectively controlled the levels of these apoptotic markers, resulting in a considerable decrease in the expression of Bcl-2, TNF-α, NFκB, p-AKT, and STAT-3. Specifically, the activation of TNF-alpha and STAT-3 triggers the signalling pathways that include the Bcl-2 family of proteins, Cyt-C, caspase 3, and 9. This ultimately leads to apoptosis and the suppression of cell growth. Furthermore, metabolomic analysis using 1H NMR indicated that the metabolites of animals reverted to normal levels after the treatment.
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We designed a tris-catecholate-based siderophore mimic, H6-T-CATL, to selectively chelate iron(III) from mitochondrial cytochromes and other iron-containing proteins within cellular matrices. This strategic sequestration aims to trigger apoptosis or ferroptosis in cancer cells through the glutathione (GSH)-dependent release of reduced iron and subsequent ROS-mediated cytotoxicity. Synthesis of H6-T-CATL involved precise peptide coupling reactions. Using the Fe(III)-porphyrin model (Fe-TPP-Cl), akin to cytochrome c, we studied H6-T-CATL's ability to extract iron(III), yielding a binding constant (Krel) of 1014 for the resulting iron(III) complex (FeIII-T-CATL)3-. This complex readily underwent GSH-mediated reduction to release bioavailable iron(II), which catalyzed Fenton-like reactions generating hydroxyl radicals (ËOH), confirmed by spectroscopic analyses. Our research underscores the potential of H6-T-CATL to induce cancer cell death by depleting iron(III) from cellular metalloproteins, releasing pro-apoptotic iron(II). Evaluation across various cancer types, including normal cells, demonstrated H6-T-CATL's cytotoxicity through ROS production, mitochondrial dysfunction, and activation of ferroptosis and DNA damage pathways. These findings propose a novel mechanism for cancer therapy, leveraging endogenous iron stores within cells. H6-T-CATL emerges as a promising next-generation anticancer agent, exploiting iron metabolism vulnerabilities to induce selective cancer cell death through ferroptosis induction.
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Antineoplásicos , Ferroptosis , Especies Reactivas de Oxígeno , Sideróforos , Ferroptosis/efectos de los fármacos , Humanos , Especies Reactivas de Oxígeno/metabolismo , Sideróforos/química , Sideróforos/farmacología , Antineoplásicos/farmacología , Antineoplásicos/química , Antineoplásicos/síntesis química , Diseño de Fármacos , Hierro/química , Hierro/metabolismo , Línea Celular Tumoral , Ensayos de Selección de Medicamentos Antitumorales , Supervivencia Celular/efectos de los fármacosRESUMEN
Joint diseases, such as osteoarthritis, often require delivery of drugs to chondrocytes residing within the cartilage. However, intra-articular delivery of drugs to cartilage remains a challenge due to their rapid clearance within the joint. This problem is further exacerbated by the dense and negatively charged cartilage extracellular matrix (ECM). Cationic nanocarriers that form reversible electrostatic interactions with the anionic ECM can be an effective approach to overcome the electrostatic barrier presented by cartilage tissue. For an effective therapeutic outcome, the nanocarriers need to penetrate, accumulate, and be retained within the cartilage tissue. Nanocarriers that adhere quickly to cartilage tissue after intra-articular administration, transport through cartilage, and remain within its full thickness are crucial to the therapeutic outcome. To this end, we used ring-opening polymerization to synthesize branched poly(l-lysine) (BPL) cationic nanocarriers with varying numbers of poly(lysine) branches, surface charge, and functional groups, while maintaining similar hydrodynamic diameters. Our results show that the multivalent BPL molecules, including those that are highly branched (i.e., generation two), can readily adhere and transport through the full thickness of cartilage, healthy and degenerated, with prolonged intra-cartilage retention. Intra-articular injection of the BPL molecules in mouse knee joint explants and rat knee joints showed their localization and retention. In summary, this study describes an approach to design nanocarriers with varying charge and abundant functional groups while maintaining similar hydrodynamic diameters to aid the delivery of macromolecules to negatively charged tissues.
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Fabrication of stimuli-responsive superstructure capable of delivering chemotherapeutics directly to the cancer cell by sparing healthy cells is crucial. Herein, we developed redox-responsive hollow spherical assemblies through self-assembly of disulfide-linked cysteine-diphenylalanine (SN). These fluorescent hollow spheres display intrinsic green fluorescence, are proteolytically stable and biocompatible, and allow for real-time monitoring of their intracellular entry. The disulfide bond facilitates selective degradation in the presence of high glutathione (GSH) concentrations, prevalent in cancer cells. We achieved efficient encapsulation (68.72%) of the anticancer drug doxorubicin (Dox) and demonstrated GSH-dependent, redox-responsive drug release within cancerous cells. SN-Dox exhibited a 20-fold lower effective concentration (2.5 µM) for compromising breast cancer cell viability compared to non-malignant cells (50 µM). The ability of SN-Dox to initiate DNA damage signaling and trigger apoptosis was comparable to that of the unencapsulated drug. Our findings highlight the potential of SN for creating site-specific drug delivery vehicles for sustained therapeutic release.
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In addition to their classical role in ATP generation, mitochondria also contribute to Ca2+ buffering, free radical production, and initiation of programmed cell death. Mitochondrial dysfunction has been linked to several leading causes of morbidity and mortality worldwide including neurodegenerative, metabolic, and cardiovascular diseases as well as several cancer subtypes. Thus, there is growing interest in developing drug-delivery vehicles capable of shuttling therapeutics directly to the mitochondria. Here, we functionalized the conventional 10,12-pentacosadiynoic acid/1,2-dimyristoyl-sn-glycero-3-phosphocholine (PCDA/DMPC)-based liposome with a mitochondria-targeting triphenylphosphonium (TPP) cationic group. A fluorescent dansyl dye (DAN) group was also included for tracking mitochondrial drug uptake. The resultant PCDA-TPP and PCDA-DAN conjugates were incorporated into a 1,2-dimyristoyl-sn-glycero-3-phosphocholine (DMPC)-based lipid bilayer, and these modified liposomes (Lip-DT) were studied for their cellular toxicity, mitochondrial targeting ability, and efficacy in delivering the drug Doxorubicin (Dox) to human colorectal carcinoma (HCT116) and human breast (MCF7) cancer cells in vitro. This Lip-DT-Dox exhibited the ability to shuttle the encapsulated drug to the mitochondria of cancer cells and triggered oxidative stress, mitochondrial dysfunction, and apoptosis. The ability of Lip-DT-Dox to trigger cellular toxicity in both HCT116 and MCF7 cancer cells was comparable to the known cell-killing actions of the unencapsulated drug (Dox). The findings in this study reveal a promising approach where conventional liposome-based drug delivery systems can be rendered mitochondria-specific by incorporating well-known mitochondriotropic moieties onto the surface of the liposome.
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BACKGROUND: Prior evidence demonstrated that Regulator of G protein Signaling 6 (RGS6) translocates to the nucleolus in response to cytotoxic stress though the functional significance of this phenomenon remains unknown. METHODS: Utilizing in vivo gene manipulations in mice, primary murine cardiac cells, human cell lines and human patient samples we dissect the participation of a RGS6-nucleolin complex in chemotherapy-dependent cardiotoxicity. RESULTS: Here we demonstrate that RGS6 binds to a key nucleolar protein, Nucleolin, and controls its expression and activity in cardiomyocytes. In the human myocyte AC-16 cell line, induced pluripotent stem cell derived cardiomyocytes, primary murine cardiomyocytes, and the intact murine myocardium tuning RGS6 levels via overexpression or knockdown resulted in diametrically opposed impacts on Nucleolin mRNA, protein, and phosphorylation.RGS6 depletion provided marked protection against nucleolar stress-mediated cell death in vitro, and, conversely, RGS6 overexpression suppressed ribosomal RNA production, a key output of the nucleolus, and triggered death of myocytes. Importantly, overexpression of either Nucleolin or Nucleolin effector miRNA-21 counteracted the pro-apoptotic effects of RGS6. In both human and murine heart tissue, exposure to the genotoxic stressor doxorubicin was associated with an increase in the ratio of RGS6/Nucleolin. Preventing RGS6 induction via introduction of RGS6-directed shRNA via intracardiac injection proved cardioprotective in mice and was accompanied by restored Nucleolin/miRNA-21 expression, decreased nucleolar stress, and decreased expression of pro-apoptotic, hypertrophy, and oxidative stress markers in heart. CONCLUSION: Together, these data implicate RGS6 as a driver of nucleolar stress-dependent cell death in cardiomyocytes via its ability to modulate Nucleolin. This work represents the first demonstration of a functional role for an RGS protein in the nucleolus and identifies the RGS6/Nucleolin interaction as a possible new therapeutic target in the prevention of cardiotoxicity.
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MicroARNs , Proteínas RGS , Animales , Humanos , Ratones , Cardiotoxicidad , MicroARNs/genética , Miocitos Cardíacos , Nucleolina , Proteínas RGS/genética , Transducción de Señal/fisiologíaRESUMEN
Efficient delivery of bioactive ingredients into cells is a major challenge. Cell-penetrating peptides (CPPs) have emerged as promising vehicles for this purpose. We have developed novel CPPs derived from the flexible and disordered tail extensions of DNA-binding Ku proteins. Ku-P4, the lead CPP identified in this study, is biocompatible and displays high internalization efficacy. Biophysical studies show that the proline residue is crucial for preserving the intrinsically disordered state and biocompatibility. DNA binding studies showed effective DNA condensation to form a positively charged polyplex. The polyplex exhibited effective penetration through the cell membrane and delivered the plasmid DNA inside the cell. These novel CPPs have the potential to enhance the cellular uptake and therapeutic efficacy of peptide-drug or gene conjugates.