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Fusogenic properties of saposin c and related proteins and peptides for transmembrane drug delivery systems

A lipid and phospholipid technology, applied in animal/human protein, drug combination, drug delivery, etc.

Inactive Publication Date: 2011-11-30
CHILDRENS HOSPITAL MEDICAL CENT CINCINNATI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, the major barriers to the successful use of imaging agents to monitor liposome delivery are ease of detection, availability of relevant technologies, and ease and efficiency of delivery.

Method used

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  • Fusogenic properties of saposin c and related proteins and peptides for transmembrane drug delivery systems
  • Fusogenic properties of saposin c and related proteins and peptides for transmembrane drug delivery systems
  • Fusogenic properties of saposin c and related proteins and peptides for transmembrane drug delivery systems

Examples

Experimental program
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preparation example Construction

[0126] A number of methods are available in connection with the preparation of liposome compositions. Thus, liposomes can be prepared using any of a number of conventional liposome preparation techniques, which will be apparent to those skilled in the art. Such techniques include, for example, solvent dialysis, French press, extrusion (with or without freeze-thaw), reverse phase evaporation, simple freeze-thaw, sonication, chelate dialysis, Homogenization, solvent infusion, microemulsions, spontaneous formation, solvent evaporation, solvent dialysis, French pressure cell techniques, controlled detergent dialysis, etc., each involving in different ways Prepare vesicles. See, eg, Madden et al., Chemistry and Physics of Lipids, 1990 53, 37-46, the disclosure of which is incorporated herein by reference in its entirety. Suitable freeze-thaw techniques are described, for example, in International Application Serial No. PCT / US89 / 05040, filed November 8, 1989, the disclosure of whi...

Embodiment 1

[0529] Example 1: Saposin C and liposome formulations and in vitro and in vivo delivery

[0530] Materials - The following materials were obtained from commercial sources: mouse laminin, P / S, fetal bovine serum, and DMEM (Gibco BRL, Gaithersborg, MD); Neurobasal medium with B27 supplement (Life Technologies, ); Dicer (New England Biolabs, Beverly, MA); pET21a(+) DNA vector, Escherichia coli (E.Coli) host strain [BL21(DE3)] and His Bind resin (Novagen, Medison, WI); conjugated with Monoclonal anti-His antibody to Alexa Fluor488 (QIAGEN, Valencia, CA); goat anti-rabbit antibody conjugated to fluorescein and sheep anti-mouse antibody conjugated to rhodamine (ICN / CAPPEL, Aurora, OH); anti-fading Reagents (Ventana Medical Systems, Tucson, AZ); C 4 Reverse-phase HPLC column (Alltech Association Inc., Deerfield, IL); DOPS and 1,2-dioleoyl-sn-glycero-3-phospho-L-serine-N-(7-nitro-2-1,3 -benzoxadiazol-4-yl) (NBD-DOPS) (as stock solution in chloroform) (Avanti Polar Lipids, Alabaster,...

Embodiment 2

[0541] Example 2: Synthesis of Liposomes Using Acidic Long-Chain Lipids, Neutral Long-Chain Lipids and Neutral Short-Chain Lipids

[0542] Materials and methods

[0543] All phospholipids DOPS, DPPC and DHPC were purchased from Avanti Polar Lipids in powder form and used without further purification. For dynamic light scattering (DLS) measurements, the molar ratio of DOPS to DPPC in the mixture was from about 10 to about 1, for all samples ([DPPC]+[DOPS]) / DHPC=about 4. The lipid mixture was dissolved in filtered ultrapure HO at a total lipid concentration of 10 wt.% using a combination of vortexing and temperature cycling (between 50 and 4 °C). 2 O (Millipore EASYpure UV). then filtered with H 2 O The homogenized 10 wt.% solution was diluted progressively to 5, 2, 1, 0.5 and 0.1 wt.%.

[0544] Lipid sample stocks were diluted 5, 50, and 200-fold with N4 prior to DLS + Particle size analyzer (Coulter, Miami, FL) for analysis. It was determined that the dilution system had...

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Abstract

The present invention includes methods of using fusogenic proteins to deliver pharmaceutical agents and / or imaging agents into and / or through the membranes of the skin, mucous membranes, and other cells, and across the blood-brain barrier. Fusogenic proteins associate with phospholipid membranes such as liposomes. Liposomes may include dioleoylphosphatidylserine, a negatively charged long-chain lipid. Alternatively, the liposomes comprise negatively charged long-chain lipids, neutral long-chain lipids and neutral short-chain lipids. Preferred fusogenic proteins include saposin C and other proteins, polypeptides and peptide analogs derived from saposin C. Active agents contained in liposomes may include biomolecules and / or organic molecules. This technology can be used in cosmetic and medical applications where the aim is to deliver active agents into and / or under biofilms or across the blood-brain barrier and neuronal membranes.

Description

[0001] This invention was made with government support in part under Grant No. RO1 DK57690-01 awarded by the National Institutes of Health. The Government may have certain rights in this invention. [0002] Cross reference to related applications [0003] This application is a continuation-in-part of, and claims the benefit of, PCT Patent Application Serial No. PCT / US2007 / 010357, filed April 27, 2007, and U.S. Provisional Patent Application Serial No. 60 / 745,969, filed April 28, 2006, for Said application is hereby incorporated by reference in its entirety for all purposes. field of invention [0004] The present application relates to compositions and methods for the targeted delivery of imaging agents and / or therapeutic agents to cells or tissues of interest, wherein the agents are contained or otherwise incorporated within protein / lipid nanovesicles. More specifically, the present invention relates to compositions and methods for the targeted delivery of imaging agents an...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): A61K9/127
CPCA61K9/1271C07K14/475C12N15/88C12N2810/85A61K9/1277A61K9/0019A61P35/00
Inventor 祁晓阳
Owner CHILDRENS HOSPITAL MEDICAL CENT CINCINNATI
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