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Biomedical subjects

V Alakhov

Publications and source records attributed to V Alakhov.

10 recordsLinked to original sources
PubMed2002

Inducing neutrophil recruitment in the liver of ICAM-1-deficient mice using polyethyleneimine grafted with Pluronic P123 as an organ-specific carrier for transgenic ICAM-1.

Coordinated expression of cell adhesion molecules and chemokines on the surface of vascular endothelium is responsible for the homing of immune effector cells to targeted sites. One way to attract non-activated immune cells to targeted organs is to use transgenically expressed adhesion molecules responsible for leukocyte recruitment. We have previously shown that polyethyleneimine (PEI) grafted with non-ionic amphiphilic Pluronic P123 block copolymer (P123PEI) modifies biodistribution of plasmid DNA toward the liver. In the present study, a P123PEI-formulated plasmid carrying the gene encoding for the murine ICAM-1 molecule was injected i.v. into transgenic ICAM-1-deficient mice. The RT-PCR analysis of ICAM-1 mRNA expression showed that P123PEI induced a dose-dependent expression of ICAM-1 in the liver. Furthermore, this expression of ICAM-1 induced neutrophil invasion in the liver, while no such invasion was observed in mice injected with formulated control plasmid or naked DNA. These results suggest that P123PEI allows functional transgene expression in the liver following i.v. injection and that ICAM-1 could be used to enhance immune response locally by attracting immune effector cells.

Animals↗
PubMed2001

Block copolymeric biotransport carriers as versatile vehicles for drug delivery.

This review describes block copolymer-based systems that are used in drug formulation development. The use of amphiphilic block copolymers to modify pharmacological performance of various classes of drugs attracts more and more attention. This is largely attributable to the high tendency of block copolymer-based drug formulations to self-assemble, as well as flexibility of block copolymer chemistry, which allows precise tailoring of the carrier to virtually any chemical entity. Combination of these features allows adjustment of block copolymer-based drug formulations to achieve the most beneficial balance in drug biological interactions with the systems that control its circulation in and removal from the body and its therapeutic activity. The following major aspects are considered: 1) physical properties of formulations and the methods used to adjust these properties towards the highest pharmacological performance of the product; 2) combinatorial methods for optimisation of block copolymer-based formulations; 3) biological response modifying properties of block copolymer-based formulations.

Animals↗
PubMed2001

Combinatorial approaches to formulation development.

This review describes the use of combinatorial methods for the development of drug formulations. Combinatorial methods are applied to find solutions to various formulation problems, including drug solubilization, controlled release, oral drug administration, and others. Various methods are described, including the synthesis of carrier libraries, high-throughput screening and computational analysis, which are used during the formulation development process, starting from initial assays through the optimization of formulation composition, to the optimization of the manufacturing process. This review also describes an integrated approach to drug formulation development using libraries of block copolymers as the drug carriers.

Animals↗
PubMed2000

A combination of poloxamers increases gene expression of plasmid DNA in skeletal muscle.

Intramuscular administration of plasmid DNA is a promising strategy to express therapeutic genes, however, it is limited by a relatively low level of gene expression. We report here that a non-ionic carrier, SP1017, composed of two amphiphilic block copolymers, pluronics L61 and F127, also known as poloxamers, significantly increases intramuscular expression of plasmid DNA. Two reporter genes, luciferase and beta-galactosidase, and one therapeutic gene, erythropoietin, were injected intramuscularly with and without SP1017 into C57Bl/6 and Balb/C mice and Sprague-Dawley rats. SP1017 increased gene expression by about 10-fold and maintained higher gene expression compared with naked DNA. Comparison of SP1017 with polyvinyl pyrrolidone (PVP) showed that SP1017 exhibited a significantly higher efficacy and its optimal dose was 500-fold lower. Experiments with beta-galactosidase using X-gal staining suggested that SP1017 considerably increased plasmid DNA diffusion through the tissue. SP1017 also improved expression of the erythropoietin gene leading to an increase in its systemic level and hematocrits. Previous toxicity studies have suggested that SP1017 has over a 1000-fold safety margin. Poloxamers used in SP1017 are listed in the US Pharmacopeia as inactive excipients and are widely used in a variety of clinical applications. We believe that the described system constitutes a simple and efficient gene transfer method to achieve local or systemic production of therapeutic proteins.

Animals↗
PubMed2000

Epitope-specific antibody response to HT-1080 fibrosarcoma cells by mimotope immunization.

Mouse monoclonal antibody (mAb) BCD-F9, which recognizes an unknown antigen found on the surface of many tumor cells, was used to screen a phage display library expressing random peptide decamers. The phage that was selected encoded the unique sequence GRRPGGWWMR, representing the peptide capable of binding to the BCD-F9 mAb. The peptide was synthesized and found to specifically inhibit the binding of mAb to HT-1080 fibrosarcoma cells. Alanine mutagenesis of the sequence encoding this peptide indicated that three residues, PXXWW, were critical for its binding to the BCD-F9 mAb. Polyclonal antibodies generated by immunization of rabbits with the synthetic peptide GRRPGGWWMR (anti-mimotope antiserum or AM-F9) bound specifically to HT-1080 cells and inhibited the binding of the BCD-F9 mAb to these cells. Using an experimental animal model in which CD-1 nude mice are inoculated i.v. with HT-1080 cells, develop lung metastasis, and die within 30 days, we have shown that AM-F9 could significantly prolong the life span of these animals. Our results suggest that a peptide mimotope can potentially be used as a novel immunotherapy to induce a beneficial antitumor response.

Amino Acid Sequence↗
PubMed1999

Fundamental relationships between the composition of pluronic block copolymers and their hypersensitization effect in MDR cancer cells.

PURPOSE: Previous studies have demonstrated that Pluronic block copolymers hypersensitize multiple drug resistant (MDR) cancer cells, drastically increasing the cytotoxic effects of anthracyclines and other anticancer cytotoxics in these cells. This work evaluates the dose dependent effects of these polymers on (i) doxorubicin (Dox) cytotoxicity and (ii) cellular accumulation of P-glycoprotein probe, rhodamine 123 (R123) in MDR cancer cells. METHODS: Dox cytotoxicity and R123 accumulation studies are performed on monolayers of drug-sensitive (KB, MCF-7, Aux-B1) and MDR (KBv, MCF-7/ADR, CHrC5) cells. RESULTS: Both tests reveal strong effects of Pluronic copolymers observed at concentrations below the critical micelle concentration (CMC) and suggest that these effects are due to the copolymer single chains ("unimers"). Using block copolymers with various lengths of hydrophobic propylene oxide (PO) and hydrophilic ethylene oxide (EO) segments these studies suggest that the potency of Pluronic unimers in MDR cells increases with elevation of the hydrophobicity of their molecule. Optimization of Pluronic composition in R123 accumulation and Dox cytotoxicity studies reveals that Pluronic copolymers with intermediate lengths of PO chains and relatively short EO segments have the highest net efficacy in MDR cells. CONCLUSIONS: The relationship between the structure of Pluronic block copolymers and their biological response modifying effects in MDR cells is useful for determining formulations with maximal efficacy with respect to MDR tumors.

ATP Binding Cassette Transporter, Subfamily B, Mem↗
PubMed1998

Multidrug-resistance drug-binding peptides generated by using a phage display library.

A phage display library of random decapeptides was used to generate peptide ligands that can bind multidrug-resistance (MDR) drugs mimicking, in this respect, the drug-binding activity of P-glycoprotein. Seven peptide sequences were identified that specifically bound doxorubicin. Five of these sequences expressed the core consensus motif WXXW. The displacement assay showed that the phages expressing these peptides bound MDR type drugs (vinblastine, doxorubicin, verapamil, and genistein) with the same selectivity as P-glycoprotein and did not interact with non-MDR type drugs, such as arabinosylcytosine (Ara-C) and melphalan. One of the selected peptides that showed a highest capacity for the binding (VCDWWGWGIC) was synthesized and displayed competition with the phage for doxorubicin binding. The structure modeling suggested that all the selected sequences contained a hydrophobic envelope in which MDR drugs could be docked with substantial energy minimization. Western blot analysis showed that monospecific antibody obtained against the phage expressing VCDWWGWGIC peptide could specifically recognize P-glycoprotein in the membrane fraction of MDR phenotype MCF-7ADR cells. The MDR drug-binding sequences generated during this work could provide an important tool for design and screening of new chemotherapeutic agents.

ATP Binding Cassette Transporter, Subfamily B, Mem↗
PubMed1996

Hypersensitizing effect of pluronic L61 on cytotoxic activity, transport, and subcellular distribution of doxorubicin in multiple drug-resistant cells.

The present study demonstrated that poly(oxypropylene) and poly(oxyethylene) block copolymer pluronic L61 (L61)-hypersensitized multidrug-resistant CHRC5 Chinese hamster ovary cells and MCF-7/ADR human breast carcinoma cells to the cytotoxic action of doxorubicin (Dox). CHRC5 and MCF-7/ADR cells manifested 290- and 700-fold increases, respectively, in their sensitivity to Dox/L61 formulation compared with free Dox. Their sensitive counterparts Aux-B1 and MCF-7 displayed only marginal or no increase at all in their response to Dox/L61. The study of the drug transport performed by flow cytometry showed that L61 enhanced the drug uptake and reduced the P-glycoprotein-mediated drug efflux. Visualization of Dox subcellular distribution in CHRC5 cells by fluorescent microscopy revealed that Dox was sequestered in cytoplasmic vesicles, whereas incubation of the cells with Dox/L61 altered the drug compartmentalization by releasing the drug from these vesicles and shifting it to the nucleus. These findings suggested that the hypersensitive response of multidrug-resistant cells to the action of Dox/L61 was caused by an increase in the drug accumulation and changes in its subcellular distribution.

ATP Binding Cassette Transporter, Subfamily B, Mem↗
PubMed1996

Enhancement of the polycation-mediated DNA uptake and cell transfection with Pluronic P85 block copolymer.

Polyelectrolyte complexes formed between DNA and poly(N-ethyl4-vinylpyridinium) cations were shown to effectively transfect mammalian cells [7]. This work suggests that the polycation-mediated uptake of the plasmid DNA and cell transfection are significantly enhanced when these complexes are administered simultaneously with a poly(ethylene oxide)-block-poly(propylene oxide)-block-poly(ethylene oxide) copolymer, Pluronic P85. The uptake studies were performed using radioactively labeled pRSV CAT plasmid on NIH 3T3, MDCK, and Jurkat cell lines. The transfection was investigated by chloramphenicol acetyltransferase assay using 3T3 cells as a model. The effects reported may be useful for the enhancement of the polycation-mediated cell transfection.

3T3 Cells↗