Hybrid PEGylated chitosan/PLGA nanoparticles designed as pH-responsive vehicles to promote intracellular drug delivery and cancer chemotherapy.

To achieve effective intracellular anticancer drug release for boosted antitumor efficacy, the acidity-responsive nanovehicles for doxorubicin (DOX) delivery were fabricated by tailor-made co-assembly of amphiphilic PEGylated chitosan20k and hydrophobic poly(lactic-co-glycolic acid) (PLGA) segments at pH 8.5. The attained DOX-loaded PEGylated chitosan20k/PLGA nanoparticles (DOX-PC20kPNs) were characterized to have a spherical shape composed of drug-encapsulated chitosan20k/PLGA-constituted solid core surrounded by hydrophilic PEG shells. Compared to non-pH-sensitive DOX-loaded PLGA nanoparticles (DOX-PNs), the DOX-PC20kPNs displayed outstanding colloidal stability under serum-containing condition and tended to swell in weak acidic milieu upon increased protonation of chitosan20k within hybrid cores, thus accelerating drug release. The in vitro cellular uptake and cytotoxicity studies revealed that the DOX-PC20kPNs after being endocytosed by prostate TRAMP-C1 cancer cells rapidly liberated drug, thus promoting drug accumulation in nuclei to enhance anticancer potency. Moreover, the hydrated PEG shells of DOX-PC20kPNs remarkably reduced their uptake by macrophage-like RAW264.7 cells. Importantly, in vivo animal findings showed that the DOX-PC20kPNs exhibited the capability of inhibiting TRAMP-C1 tumor growth superior to free hydrophobic DOX molecules and DOX-PNs, demonstrating the great potential in cancer chemotherapy.

Tumor acidity-responsive polymeric nanoparticles to promote intracellular delivery of zoledronic acid by PEG detachment and positive charge exposure for enhanced antitumor potency.

Zoledronic acid (ZA), a third-generation bisphosphonate, has been extensively used to treat osteoporosis and cancer bone metastasis and demonstrated to suppress proliferation of varied cancer cells and selectively kill tumor-associated microphages (TAMs). However, the clinical applications of ZA in extraskeletal tumor treatment are largely restricted due to its rapid renal clearance and binding to bones. In this study, to promote intracellular delivery of ZA for amplified antitumor efficacy, tumor acidity-responsive polymeric nanoparticles with high ZA payload (ca. 12.3 wt%) and low premature ZA leakage were designed. As a pivotal material for surface coating, the acidity-sensitive and amphiphilic methoxy poly(ethylene glycol) (mPEG)-benzoic imine-octadecane (C18) (mPEG-b-C18) was synthesized by conjugation of mPEG-CHO with 1-octadecylamine upon Schiff base reaction. Through tailor-made co-assembly of the hydrophobic poly(lactic-co-glycolic acid) (PLGA), amphiphilic tocopheryl polyethylene glycol succinate (TPGS) and mPEG-b-C18 to encapsulate ionic complexes composed of ZA molecules and branched poly(ethylenimine) (PEI) segments, the attained therapeutic polymeric nanoparticles, characterized to have a hydrophobic PLGA/ZA/PEI-constituted core covered with mPEG-b-C18 and TPGS, were able to not only detach mPEG shielding upon acidity-triggered hydrolysis of benzoic imine bonds but also expose surface positive charges of protonated PEI segments. The in vitro cellular uptake and cytotoxicity studies demonstrated that the internalization of acidity-sensitive ZA-encapsulated nanoparticles by TRAMP-C1 mouse prostate cancer cells and murine macrophages RAW 264.7 was considerably promoted upon acidity-elicited PEG detachment and surface charge conversion, thus remarkably boosting intracellular ZA delivery and anticancer potency. Compared to PEG non-detachable ZA-loaded nanoparticles with poor tumor deposition and antitumor effect, the PEG-detachable ZA-carrying nanoparticles markedly accumulated in TRAMP-C1 solid tumors in vivo and inhibited tumor growth, thereby increasing the survival rate of the treated mice. The collective data suggest the great promise of tumor acidity-sensitive ZA-carrying hybrid nanoparticles in the treatment of extraskeletal solid tumors.

Potential of antiviral drug oseltamivir for the treatment of liver cancer.

Liver cancer is a leading cause of cancer­related mortality globally. Since hepatitis virus infections have been strongly associated with the incidence of liver cancer, studies concerning the effects of antiviral drugs on liver cancer have attracted great attention in recent years. The present study investigated the effects of two anti­hepatitis virus drugs, lamivudine and ribavirin, and one anti­influenza virus drug, oseltamivir, on liver cancer cells to assess alternative methods for treating liver cancer. MTT assays, wound healing assays, Τranswell assays, flow cytometry, immunoblotting, ELISA, immunofluorescence staining and a xenograft animal model were adopted to verify the effects of lamivudine, ribavirin and oseltamivir on liver cancer cells. Treatment with ribavirin and oseltamivir for 24 and 48 h significantly decreased the viability of both Huh-7 and HepG2 cells compared with that of THLE­3 cells in a dose­dependent manner. The subsequent investigations focused on oseltamivir, considering the more serious clinical adverse effects of ribavirin than those of oseltamivir. Significantly decreased migration and invasion were observed in both Huh-7 and HepG2 cells that were treated with oseltamivir for 24 and 48 h. In addition, oseltamivir significantly increased autophagy in Huh­7 cells, as revealed by the significantly higher ratios of LC3­II/LC3­I, increased expression of Beclin­1, and decreased expression of p62, whereas no significant increases in the expression of apoptosis­related proteins, including Apaf­1, cleaved caspase­3, and cleaved PARP­1, were detected. Notably, apoptosis and autophagy were significantly increased in HepG2 cells in the presence of oseltamivir, as revealed by the significant increases in the expression of Apaf­1, cleaved caspase­3, and cleaved PARP­1, the higher ratios of LC3­II/LC3­I, the increased expression of Beclin­1, and the decreased expression of p62. Additionally, significant inhibitory effects of oseltamivir on xenografted Huh­7 cells in athymic nude mice were observed. The present study, for the first time to the best of our knowledge, reported the differential effects of oseltamivir on inducing liver cancer cell death both in vitro and in vivo and may provide an alternative approach for treating liver cancer.

Effect of N­terminal region of human parvovirus B19­VP1 unique region on cardiac injury in naïve mice.

A unique region of human parvovirus B19 virus­VP1 (B19V­VP1u) has been linked to a variety of cardiac disorders. However, the precise role of B19V­VP1u in inducing cardiac injury remains unknown. The present study investigated the effects of B19V­VP1u and different regions of B19V­VP1u, including B19V­VP1uA (residues 1­60), B19V­VP1uB (residues 61­129), B19V­VP1uC (residues 130­195) and B19V­VP1uD (residues 196­227), on inducing cardiac injury in naïve mice by zymography, immunoblotting, H&E staining and cytokine immunoassay. A significantly higher MMP­9/MMP­2 ratio and increased levels of inflammatory cytokines, including IL­6 and IL­1ß, were detected in the left ventricles of the mice injected with B19V­non­structural protein 1 (B19V­NS1) and B19V­VP1u, accompanied by increased expression levels of phosphorylated (p­)ERK and p­P38. Significantly upregulated expression levels of atrial natriuretic peptide (ANP), heart­type fatty acid­binding protein (H­FABP) and creatine kinase isoenzyme­MB (CK­MB), which are well­known cardiac injury markers, as well as increased infiltration of lymphocytes, were detected in the left ventricles of the mice injected with B19V­VP1, B19V­NS1 and B19V­VP1u. Moreover, a significantly higher MMP­9/MMP­2 ratio and increased levels of IL­6 and IL­1ß were observed in the left ventricles of the mice injected with B19V­VP1u, B19V­VP1u­A, B19V­VP1u­B and B19V­VP1u­C, accompanied by upregulated p­ERK and p­P38 expression. Notably, significantly lower levels of IL­6 and IL­1ß were observed in the left ventricles of the mice injected with B19V­VP1uD. Furthermore, significantly increased ANP, H­FABP and CK­MB expression levels were detected in the left ventricles of the mice injected with B19V­VP1u, B19V­VP1u­A and B19V­VP1u­B, along with enhanced infiltration of lymphocytes. Significantly higher serum IL­1ß, IL­6, TNF­α and IFN­Î³ levels were also detected in the mice injected with B19V­VP1u, B19V­VP1u­A and B19V­VP1u­B. To the best of our knowledge, the findings of the present study were the first to demonstrate that the N­terminal region (residues 1­129) of B19V­VP1u induces an increase in the levels of cardiac injury markers, thus providing evidence for understanding the possible functional regions within B19V­VP1u.

Differential Effects of Wedelia chinensis on Human Glioblastoma Multiforme Cells.

INTRODUCTION: Glioblastoma multiforme (GBM) is the most aggressive glioma, and its diffuse nature makes resection of it difficult. Moreover, even with the administration of postoperative radiotherapy and chemotherapy, prolonged remission is often not achieved. Hence, innovative or alternative treatments for GBM are urgently required. Traditional Chinese herbs and their functional components have long been used in the treatment of various cancers, including GBM. The current study investigated the antitumor activity of Wedelia chinensis and its major functional components, luteolin and apigenin, on GBM. MATERIALS AND METHODS: MTT assay, Transwell migration assay, and flow cytometry analysis were adopted to assess the cell viability, invasive capability, and cell cycle. Immunofluorescence staining and Western blotting were used to detect the expressions of apoptotic and autophagy-related signaling molecules. RESULTS: The W. chinensis extract (WCE) significantly inhibited the proliferation and invasive ability of both GBM8401 and U-87MG cells in a dose-dependent manner. Moreover, differential effects of WCE on GBM8401 and U-87MG cells were observed: WCE induced apoptosis in GBM8401 cells and autophagy in U-87MG cells. Notably, WCE had significant effects in reducing the cell survival and invasive capability of both GBM8401 and U-87MG cells than the combination of luteolin and apigenin. CONCLUSIONS: Taken together, these findings indicate the potential of using WCE and the combination of luteolin and apigenin for GBM treatment. However, further investigations are warranted before considering recommending the clinical use of WCE or the combination of luteolin and apigenin as the standard for GBM treatment.