Investigation Of Vitamin B12-Modified Amphiphilic Sodium Alginate Derivatives For Enhancing The Oral Delivery Efficacy Of Peptide Drugs.

PURPOSE: Peptide drugs have been used in therapy various diseases. However, the poor bioavailability of peptide drugs for oral administration has limited their clinical applications, on account of the acidic environment and digestive enzymes inside the human gastrointestinal tract. To enhance stability in the human gastrointestinal tract, bioavailability, and targeted drug delivery of peptide drugs through oral administration, a vitamin B12-modified amphiphilic sodium alginate derivative (CSAD-VB12) was synthesized. MATERIALS AND METHODS: A vitamin B12-modified amphiphilic sodium alginate derivative (CSAD-VB12) was synthesized via the N,N'-dicyclohexylcarbodiimide active method at room temperature, and then characterized using FTIR and 1H NMR spectroscopy. Insulin was used as a model peptide drug and the insulin-loaded CSAD-VB12 (CSAD-VB12/insulin) nanoparticles with negative zeta potentials were prepared in PBS (pH=7.4). Scanning electron microscopy was used to observe CSAD-VB12/insulin as spherical nanoparticles. The CSAD-VB12 derivatives and CSAD-VB12/insulin nanoparticles displayed nontoxicity towards the human colon adenocarcinoma (Caco-2) cells by CCK-8 test. Caco-2 cell model was used to measure the apparent permeability (Papp) of insulin, CSAD/insulin and CSAD-VB12/insulin. Furthermore, confocal was used to confirm the endocytosis of intestinal enterocytes. Type 1 diabetes mice were used to evaluate the intestinal absorption and retention effect of test nanoparticles. RESULTS: They were observed as spherical nanoparticles in the size of 30-50 nm. The CSAD-VB12 derivatives and CSAD-VB12/insulin nanoparticles displayed nontoxicity towards the human colon adenocarcinoma (Caco-2) cells. Comparing with insulin and the CSAD/insulin nanoparticles, the CSAD-VB12/insulin nanoparticles exhibited higher permeation ability through intestinal enterocytes in the Caco-2 cell model. Oral administration of the CSAD-VB12/insulin nanoparticles to Type 1 diabetic mice yields higher intestinal retention effect, targeted absorption, and outstanding efficacy. CONCLUSION: CSAD-VB12 derivatives enhance the small intestinal absorption efficacy and retention of peptide by oral administration, which indicated that it could be a promising candidate for oral peptide delivery in the prospective clinical application.

Tivantinib induces G2/M arrest and apoptosis by disrupting tubulin polymerization in hepatocellular carcinoma.

BACKGROUND: Tivantinib has been described as a highly selective inhibitor of MET and is currently in a phase III clinical trial for the treatment of hepatocellular carcinoma (HCC). However, the mechanism of tivantinib anti-tumor effect has been questioned by recent studies. RESULTS: We show that tivantinib indiscriminately inhibited MET dependent and independent HCC cells proliferation. In contrast, other MET inhibitors, JNJ-38877605 and PHA-665752, just specifically inhibited the growth of MET dependent HCC cells. Tivantinib neither inhibit constitutive MET phosphorylation nor HGF-induced MET phosphorylation in HCC cells. In the microtubule polymerization analysis, tivantinib affected microtubule dynamics by a mechanism as a microtubule depolymerizer. Interesting, unlike other microtubule-targeting agents, paclitaxel and vincristine, tivantinib showed similar anti-proliferative activity in parental and multidrug-resistant cells. Further studies demonstrated that tivantinib induced a G2/M arrest and promoted apoptosis by both intrinsic and extrinsic pathway. The in vivo efficacy evaluation showed that tivantinib exhibited a good anti-tumor growth activity with anti-proliferative and pro-apoptotic effects. CONCLUSIONS: The potent anti-tumor activity of tivantinib in HCC was achieved by targeting microtubule. Tivantinib treatment for patients with HCC should not be selected based on MET status.

Nonadherent culture method downregulates stem cell antigen-1 expression in mouse bone marrow mesenchymal stem cells.

Mesenchymal stem cells (MSCs) are primarily isolated by their adherence to plastic and their in vitro growth characteristics. Expansion of these cells from an adherent culture is the only method to obtain a sufficient number of cells for use in clinical practice and research. However, little is known with regard to the effect of adherence to plastic on the phenotype of the cells. In the present study, bone marrow CD45-CD31-CD44- stem cell antigen (Sca)-1+ MSCs were sorted by flow cytometry and expanded in adherent cultures. The expression levels of the adhesion molecule, Sca-1, in the adherent cultures were compared with those from nonadherent cultures at different time points. The flow cytometry results indicated that the expression levels of Sca-1 decreased in the MSCs in the nonadherent cultures grown in ultra-low-adherent plates. Furthermore, the result was confirmed by quantitative polymerase chain reaction at the same time points. Therefore, the results demonstrated that the loss of plastic adherence downregulated the expression of Sca-1. The observations may provide novel insights into the molecular mechanisms underlying plastic adherent culture.

An efficient nonviral gene-delivery vector based on hyperbranched cationic glycogen derivatives.

BACKGROUND: The purpose of this study was to synthesize and evaluate hyperbranched cationic glycogen derivatives as an efficient nonviral gene-delivery vector. METHODS: A series of hyperbranched cationic glycogen derivatives conjugated with 3-(dimethylamino)-1-propylamine (DMAPA-Glyp) and 1-(2-aminoethyl) piperazine (AEPZ-Glyp) residues were synthesized and characterized by Fourier-transform infrared and hydrogen-1 nuclear magnetic resonance spectroscopy. Their buffer capacity was assessed by acid-base titration in aqueous NaCl solution. Plasmid deoxyribonucleic acid (pDNA) condensation ability and protection against DNase I degradation of the glycogen derivatives were assessed using agarose gel electrophoresis. The zeta potentials and particle sizes of the glycogen derivative/pDNA complexes were measured, and the images of the complexes were observed using atomic force microscopy. Blood compatibility and cytotoxicity were evaluated by hemolysis assay and MTT (3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyltetrazolium bromide) assay, respectively. pDNA transfection efficiency mediated by the cationic glycogen derivatives was evaluated by flow cytometry and fluorescence microscopy in the 293T (human embryonic kidney) and the CNE2 (human nasopharyngeal carcinoma) cell lines. In vivo delivery of pDNA in model animals (Sprague Dawley rats) was evaluated to identify the safety and transfection efficiency. RESULTS: The hyperbranched cationic glycogen derivatives conjugated with DMAPA and AEPZ residues were synthesized. They exhibited better blood compatibility and lower cytotoxicity when compared to branched polyethyleneimine (bPEI). They were able to bind and condense pDNA to form the complexes of 100-250 nm in size. The transfection efficiency of the DMAPA-Glyp/pDNA complexes was higher than those of the AEPZ-Glyp/pDNA complexes in both the 293T and CNE2 cells, and almost equal to those of bPEI. Furthermore, pDNA could be more safely delivered to the blood vessels in brain tissue of Sprague Dawley rats by the DMAPA-Glyp derivatives, and then expressed as green fluorescence protein, compared with the control group. CONCLUSION: The hyperbranched cationic glycogen derivatives, especially the DMAPA-Glyp derivatives, showed high gene-transfection efficiency, good blood compatibility, and low cyto toxicity when transfected in vitro and in vivo, which are novel potential nonviral gene vectors.