Ovarian tumor targeting of docetaxel-loaded liposomes mediated by luteinizing hormone-releasing hormone analogues. In vive distribution in nude mice.

Recent studies indicate that luteinizing hormone-releasing hormone (LHRH) analogues (LHRHa), like LHRH, are able to specifically bind to LHRH receptors which are highly expressed on the extracellular membrane of ovarian tumor cells. As a targeting moiety, LHRHa can mediate the ovarian tumor targeting of docetaxel-loaded liposomes. In our study, synthesized negatively charged cholesterol succinimide (CHS) was employed for the preparation of negatively charged docetaxel-loaded liposomes, with which the positively charged LHRHa is linked via electrostatic absorption. An HPLC-based assay for determination of docetaxel (CAS 114977-28-5, Doc) in vivo and the model of ovarian cancer xenograft were established to investigat the biodistribution of docetaxel, docetaxel liposomes (Doc-Lipo), and LHRHa mediated docetaxel-loaded liposomes (LHRHa-Doc-Lipo) in nude mice. Sixty minutes after administration of LHRHa-Doc-Lipo, the concentration of docetaxel in the ovarian tumor was 2.86 times that of Doc-Lipo and 9.02 times that of Doc in the nude mice bearing ovarian tumor. LHRHa-Doc-Lipo decreased the concentration of docetaxel in the liver and spleen by 57% and 34%, respectively, as compared with Doc-Lipo. Therefore, LHRHa-Doc-Lipo exhibits potentiality as an active targeting drug delivery system for chemotherapy of ovarian tumor.

[Research on thymopentin-loaded N-trimethyl chitosan nanoparticles administered through mouth].

OBJECTIVE: To prepare a peroral thymopentin-loaded N-trimethyl chitosan chloride-nanoparticle (Tp5-TMC-NP) ,and observe the pharmacodynamic action when the Tp5-TMC-NP is taken by way of the mouth. METHODS: N-trimethyl chitosan chloride was first synthesized, and then Tp5-TMC-NP was prepared with the formulation technology optimized by the Central Composite Design. The influence of Tp5-TMC-NP on the ratio of CD4+/CD8+ of T-lymphocytes were determined by flow cytometer. RESULTS: The regular global Tp5-TMC-NP prepared with the optimized formulation craft had the mean diameter of 110.6 nm and got the entrapment efficiency of 78.8%. The ratio of lymphocyte CD4+/CD8+ of Wistar rat administered with Tp5-TMC-NP perfusing stomach had 2.59 times higher than that with Tp5. CONCLUSION: Taken orally the Tp5-TMC-NP has much higher efficiency than Tp5.

Preparation of a TK/GCV administration system mediated by transferrin modified pro-cationic liposomes.

Transferrin modified pro-cationic liposomes were prepared and used to investigate the effect of targeting therapeutic genes to human hepatoma carcinoma cells in vitro. The main lipid CHETA, cholest-5-en-3beta-yl[2-[[4-[(carboxymethyl)dithio]-1-iminobutyl]amino]ethyl] carbamate (C36H61N3O4S2), was synthesized and used to prepare pro-cationic liposomes. The thymidine kinase (TK) gene loaded pro-cationic liposomes were prepared by first mixing the plasmid DNA and protamine together, and then incubating the resulted polyplexes with blank pro-cationic liposomes preformed by the thin film dispersion-sonication method. Transferrin (Tf) was adsorbed on the surface of pro-cationic liposomes via electrostatic interactions to form transferrin modified pro-cationic liposomes. Cellular association was measured by fluorimetry at excitation and emission wavelengths of 490 and 520 nm, respectively. The viability of TK gene infected cells following administration of ganciclovir (GCV) was investigated by MTT assay. The transferrin modified TK gene pro-cationic liposomes had a mean diameter of 240 +/- 12 nm and zeta potential of -24.10 +/- 2.5 mV (n = 3). The transmission electron microscopy image indicated that most of the liposomes were relatively regular and spherical with a condensed core inside. Cell-associated fluorescence of Tf-liposomes and unmodified liposomes (without transferrin) was 7.8 x 10(6), and 3.2 x 10(6) per milligram protein, respectively. Compared to Lipofectamine 2000 (Invitrogen, USA) the pro-cationic liposomes and transferrin modified pro-cationic liposomes had less cytotoxicity to cells. The transduced TK gene HepG2 cells were more sensitive to GCV than the un-transduced TK gene ones and the human normal Chang liver cells were not affected by the TK/GCV system mediated by procationic liposomes.

[Preparation and gene expression of transferrin modified gene loaded procationic liposomes].

A novel transferrin modified non-viral gene delivery system Tf-PLPD was developed and the related characteristics was investigated. Blank procationic liposomes were prepared by film dispersion-filteration method. PLPD was prepared as follows by first mixing the plasmid DNA and protamine together, then the resulted polyplexes were incubated for 10 min at room temperature, followed by addition of preformed blank procationic liposomes. Transferrin was adsorbed at the surface of PLPD via electrostatic interactions to form Tf-PLPD. Central composite design (CCD) was employed to optimize the formulation. The HepG2 cells were transfected using lacZ as reporter gene and characteristics such as the morphology, the mean particle size, the zeta potential and the transfection efficiency in HepG2 cells were further investigated by different methods. The resulting PLPD had a regular spherical surface with an average size of (228. 9 +/- 8. 0) nm (polydispersity index, PDI = 0. 122 +/- 0. 02, n = 3) , a zeta potential of ( - 25. 08 +/-2. 50) mV (n = 3) and a transfection efficiency of (12. 18 +/- 3. 80) mU x mg(-1) (protein). The Tf-PLPD had an average size of (240 +/- 12) nm (polydispersity index, PDI = 0. 150 +/- 0. 03, n = 3), a zeta potential of ( - 24. 10 +/- 2. 50) mV ( n = 3) and a transfection efficiency of (24. 26 +/- 2. 60) mU x mg(-1) (protein) , 20 times greater than that of the naked plasmid DNA. The presence of serum didn' t affect the tansfection activity of PLPD or Tf-PLPD. Compared to one kind of cationic liposomes (liposome-protamine-DNA, LPD), the PLPD and Tf-PLPD had much less cytotoxicity to three hepatic cell lines (including HepG2, SMMC7721 and Chang' s normal hepatocyte). The results indicated that the Tf-PLPD is a perspective non-viral vector for gene delivery systems.

Preparation and characterization of a novel nonviral gene transfer system: procationic-liposome-protamine-DNA complexes.

Procationic-liposome-protamine-DNA (PLPD) vector, a novel nonviral gene delivery system, that may further adsorb transferrin (Tf) at its surface via electrostatic interactions to form Tf-PLPD, was prepared from soybean phosphatidylcholine (PC), cholesterol (Chol), and a kind of cholesterol derivative, CHETA(cholest-5-en-3-ol(3beta)-[2-[[4-[(carboxymethyl)dithio]-1-iminobutyl] amino] ethyl] carba- mate) containing disulfide bond by film dispersion-filteration method. Central composite design was used to optimize the formulation. The presence of serum did not affect the transfection activity of PLPD or Tf-PLPD and the cell viability was not affected significantly when the cells were incubated with the complexes for 4 hr at 37 degrees C. Compared with one kind of cationic liposomes(liposome-protamine-DNA), the PLPD had much less cytotoxicity to three hepar cell lines(including HepG2, SMMC7721, and Chang's normal heptocyte). The procationic lipoplex described here, combining the condensing effect of protamine and the targeting capability of Tf, was a perspective nonviral vector for gene delivery system.

Characteristics comparison before and after lyophilization of transferrin modified Procationic-Liposome-Protamine-DNA complexes (Tf-PLPD).

A novel non-viral gene delivery system, Procationic-Liposome-Protamine-DNA complexes (PLPD) which could further adsorb transferrin on the surface as a targeting ligand to form Tf-PLPD, was prepared and characterized before and after lyophilization. The size distribution of Tf-PLPD was in the range of 240 +/- 12 nm and the zeta potential was -24.10 +/- 2.5 mV. The transfection efficiencies of PLPD and Tf-PLPD were 12.18 +/- 3.8 and 24.26 +/- 2.6 mU beta-galactosidase/mg protein respectively. The lyophilization and the presence of serum didn't affect the tansfectivities of PLPD or Tf-PLPD. Compared to Lipofectamine 2000 (Invitrogen, U.S.A.), the procationic liposomes had less cytotoxicity to cells. In summary the procationic lipoplex described here, combining the condensing effect of protamine and the targeting capability of Tf, was a perspective non-viral vector for gene delivery system.

Characterization of transferrin-modified procationic-liposome protamine-DNA complexes.

We developed a novel transferrin modified non-viral gene delivery system, transferrin-modified procationic-liposome-protamine-DNA complexes (Tf-PLPD) and investigated its characteristics. Blank procationic liposomes were prepared using the film dispersion filter method. Protamine was used to condense plasmid DNA to form protamine-DNA complexes and the complexes were further incubated with blank procationic liposomes to form PLPD. Transferrin was adsorbed onto the surface of PLPD via an electrostatic interaction, and thus Tf-PLPD was produced. Characteristics such as stability in rat serum, morphology, average particle size, zeta potential, and transfection efficiency in HepG2 cells were further investigated. The results indicated that the procationic liposomes remained stable in rat serum for 24 h. Tf-PLPD protected plasmid DNA from enzymatic degradation even after lyophilization. The size distribution of Tf-PLPD was in the range of 240+/-12 nm and the zeta potential was -24.10+/-2.5 mV (n=3), respectively. The transfection efficiencies of Tf-PLPD were 24.26+/-2.6 mU beta-galactosidase/mg protein. Lyophilization and the presence of serum did not affect the transfectivity of Tf-PLPD and the procationic liposomes also had low cytotoxicity to cells.

Research on thymopentin loaded oral N-trimethyl chitosan nanoparticles.

Peptides, although high efficacy and specificity in their physiological function, usually have low therapeutical activities due to their poor bioavailability when administrated orally. Nanoparticles have been regarded as a useful vector for targeted drug delivery system because they can protect drug from being degraded quickly and pass the gastrointestinal barriers. Here we described a novel oral N-trimethyl chitosan nanoparticles formulation containing thymopentin (Tp5-TMC-NP). N-trimethyl chitosan (TMC) was synthesized and then used to prepare Tp5-TMC-NP by ionotropic gelation. A three-factor, five-level CCD (Central Composite Design) design was used in the optimization procedure, with HPLC as the analyzing method. The resulting Tp5-TMC-NP had a regular spherical surface and a narrow particle size range with a mean diameter of 110.6 nm. The average entrapment efficiency was 78.8%. The lyophilized Tp5-TMC-NP formulation was stable in 4 degrees C or -20 degrees C after storage of 3 months without obvious changes in morphology, particle size, pH and entrapment ratio. The results of the flow cytometer determination showed that the ratio of CD4+/CD8+ of Wistar female rat givenTp5-TMC-NP (ig) was 2.59 time that of the group given Tp5 (ig).

[Determination of trace bismuth in traditional Chinese medicines by hydride generation atomic fluorescence spectrometry].

A method is reported for the determination of trace bismuth in traditional chinese medicine by hydride generation atomic fluorescence spectrometry. The effect of different means of digestion, the medium amounts of acid and reducing agent on the determination of Bi is investigated. In the given conditions, the linear range of determination is 0.1-200 microg x L(-1), and the detection limit is 0.0946 microg x L(-1). The instrumental relative standard deviation is about 0.55% and the recovery is about 94%-107%. The method is accurate, rapid and convenient with satisfactory results.