Microvesicles derived from nucleated, mammalian cells and use thereof

Abstract

The present invention relates to a microvesicle that is derived from nucleated mammalian cells, which are smaller than the nucleated cells. The microvesicles of the present invention can be used in the delivery of a therapeutic or diagnostic substance to specific tissues or cells, and more particularly, relates to microvesicles derived from monocytes, macrophages, dendritic cells, stem cells or the like, which can be used to deliver specific therapeutic or diagnostic substances for treating and/or diagnosing tissue associated with cancer, diseased blood vessels, inflammation, or the like.

1 . A composition, comprising sub-cell sized microvesicles derived from nucleated mammalian cells. 
     
     
         2 . The composition of  claim 1 , wherein the nucleated mammalian cells are selected from the group consisting of monocytes, macrophages, dendritic cells and stem cells. 
     
     
         3 . The composition of  claim 1 , wherein the nucleated mammalian cells are guided to specific cells or tissues. 
     
     
         4 . The composition of  claim 1 , wherein the nucleated mammalian cells express therapeutic or diagnostic substances. 
     
     
         5 . The composition of  claim 1 , wherein the nucleated mammalian cells are transformed cells. 
     
     
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         13 . The composition of  claim 1 , wherein the microvesicles further comprise components other than the plasma membrane of the nucleated mammalian cells. 
     
     
         14 . The composition of  claim 13 , wherein the component other than the plasma membrane is cyclodextrin or polyethylene glycol. 
     
     
         15 . The composition of  claim 1 , wherein the microvesicles have membrane components that have been chemically modified. 
     
     
         16 . A pharmaceutical composition, comprising sub-cell sized microvesicles or shedding microvesicles derived from nucleated mammalian cells, loaded with therapeutic or diagnostic substances. 
     
     
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         18 . The pharmaceutical composition of  claim 16 , wherein the therapeutic or diagnostic substances are derived from the nucleated mammalian cells. 
     
     
         19 . The pharmaceutical composition of  claim 16 , wherein the nucleated mammalian cells are transformed cells to express the therapeutic or diagnostic substances. 
     
     
         20 . The pharmaceutical composition of  claim 16 , wherein the therapeutic or diagnostic substances are foreign to the nucleated mammalian cells. 
     
     
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         32 . A method for preparing microvesicles loaded with therapeutic or diagnostic substances from nucleated mammalian cells, comprising:
 (a) applying a process to a suspension of the nucleated mammalian cells to construct microvesicles, said process being selected from the group consisting of extrusion, sonication, cell lysis, homogenization, freezing-thawing, electroporation, mechanical degradation and chemical substance treatment;   (b) isolating the microvesicles from the suspension; and   (c) incubating a suspension of the microvesicles isolates with therapeutic or diagnostic substances.   
     
     
         33 . A method for preparing microvesicles loaded with therapeutic or diagnostic substances from nucleated mammalian cells, comprising:
 (a) incubating a suspension of the nucleated mammalian cells with the therapeutic or diagnostic substances to load the therapeutic or diagnostic substances to the nucleated mammalian cells; and   (b) applying a process to the suspension to construct the microvesicles, said process being selected from the group consisting of extrusion, sonication, cell lysis, homogenization, freezing-thawing, electroporation, mechanical degradation and chemical substance treatment.   
     
     
         34 . A method for preparing microvesicles loaded with therapeutic or diagnostic substances from nucleated mammalian cells, comprising:
 (a) adding therapeutic or diagnostic substances to a suspension of the nucleated mammalian cells to yield a mixed suspension; and   (b) applying a process to the mixed suspension to construct microvesicles, said process being selected from the group consisting of extrusion, sonication, cell lysis, homogenization, freezing-thawing, electroporation, mechanical degradation and chemical substance treatment.   
     
     
         35 . A method for preparing shedding microvesicles loaded with therapeutic or diagnostic substances, from nucleated mammalian cells:
 (a) isolating shedding microvesicles from a suspension of the nucleated mammalian cells; and   (b) incubating a suspension of the shedding microvesicles isolates with the therapeutic or diagnostic substances.   
     
     
         36 . A method for preparing shedding microvesicles loaded with therapeutic or diagnostic substances from nucleated mammalian cells, comprising:
 (a) culturing a suspension of the nucleated, mammalian cells in presence of the therapeutic or diagnostic substances to load the therapeutic or diagnostic substances to the nucleated mammalian cells; and   (b) isolating the shedding microvesicles loaded with the therapeutic or diagnostic substances from the culture.   
     
     
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         48 . A method for delivering a therapeutic or diagnostic substance to specific cells or tissues, comprising using sub-cell sized microvesicles or shedding microvesicles, derived from nucleated mammalian cells, loaded with the therapeutic or diagnostic substances. 
     
     
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         50 . The method of  claim 47 , wherein two or more different therapeutic or diagnostic substances are delivered specifically to cells or tissues. 
     
     
         51 . The method of  claim 50 , wherein the two or more different therapeutic or diagnostic substances are loaded together to single microvesicles or shedding microvesicles. 
     
     
         52 . The method of  claim 47 , wherein two or more different therapeutic or diagnostic substances are loaded to respective microvesicles or shedding microvesicles and used together. 
     
     
         53 . The method of  claim 52 , wherein the microvesicles or shedding microvesicles respectively loaded with the two or more different substances are used together. 
     
     
         54 . The method of  claim 47 , wherein two or more different microvesicles or shedding microvesicles selected from the group consisting of microvesicles or shedding microvesicles loaded with one therapeutic or diagnostic substance, microvesicles or shedding microvesicles loaded with two or more therapeutic or diagnostic substances, and a combination thereof are administered sequentially. 
     
     
         55 . A method for treating or diagnosing a disease, comprising using sub-cell sized, microvesicles, derived from nucleated mammalian cells, with therapeutic or diagnostic substances loaded thereto, to deliver the therapeutic or diagnostic substances to specific cells or tissues. 
     
     
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         66 . A kit for diagnosing a disease, comprising nucleated mammalian cell-derived, sub-cell size microvesicles or shedding microvesicles loaded with primers, probes, antisense nucleic acids or antibodies as active ingredients.