Extracellular vesicles for agent delivery

Abstract

The present invention relates to the field of extracellular vesicles. More specifically, the present invention provides methods and compositions for using extracellular vesicles as a vector for nucleic acid treatment in vivo of various diseases. In a specific embodiment, the present invention provides an extracellular vesicle isolated from a cell comprising one or more microRNAs (miRNAs) that have been loaded ex vivo into the vesicle so that the miRNAs are present in a higher concentration than when measured in the same extracellular vesicle isolated directly from the cell. In another embodiment, the present invention provides a method for treating cholangiocarcinoma in a subject comprising the step of administering to the subject a plurality of exosomes comprising miR- 195.

What is claimed is: 
     
         1 - 4 . (canceled) 
     
     
         5 . An extracellular vesicle isolated from a cancer associated fibroblast (CAF), wherein the vesicle comprises a small molecule, and wherein the extracellular vesicle selectively targets a cancer cell. 
     
     
         6 . (canceled) 
     
     
         7 . The extracellular vesicle of  claim 5 , wherein the small molecule is loaded into the cell or extracellular vesicle ex vivo. 
     
     
         8 . The extracellular vesicle of  claim 5 , further comprising a recombinant polypeptide or polynucleotide that is heterologously expressed in the CAF or is loaded into the cell or extracellular vesicle ex vivo. 
     
     
         9 . The extracellular vesicle of  claim 8 , wherein the recombinant polynucleotide is a microRNA. 
     
     
         10 . The extracellular vesicle of  claim 9 , wherein the microRNA is miR-195, miR-126, or miR-192. 
     
     
         11 . The extracellular vesicle of  claim 5 , wherein the small molecule is a lipid or other hydrophobic small molecule. 
     
     
         12 . The extracellular vesicle of  claim 5 , wherein the small molecule is doxorubicin, cisplatin, or phosphatidyl ethanolamine. 
     
     
         13 . The extracellular vesicle of  claim 12 , wherein the phosphatidyl ethanolamine is derivatized with an agent selected from the group consisting of rhodamine, fluorescein, biotin, streptavidin, a small molecule, a polynucleotide, and a polypeptide. 
     
     
         14 . The extracellular vesicle of  claim 5 , wherein the small molecule can be used for imaging purposes. 
     
     
         15 - 25 . (canceled) 
     
     
         26 . The extracellular vesicle of  claim 5 , wherein the vesicle expresses increased levels of one or more markers selected from the group consisting of alpha-SMA, Collagen, Vimentin (FSP-1), S100, Metalloproteinases, NG2, PDGFR-B, SDF1/CXCL12, CD34, Fibroblast activation protein (FAP), FSP-1, CD31, Thy-1, and Gremlin, and/or expresses reduced levels of laminin. 
     
     
         27 . (canceled) 
     
     
         28 . The extracellular vesicle of  claim 5 , wherein the CAF is derived from a fibroblast cultured for at least 1-14 days in the presence of a cancer cell or in the presence of conditioned media derived from a cancer cell culture. 
     
     
         29 - 32 . (canceled) 
     
     
         33 . A method for obtaining the extracellular vesicle of  claim 5 , the method comprising culturing a fibroblast or stromal cell in conditioned media obtained from a cancer cell culture, and isolating extracellular vesicles from the media. 
     
     
         34 - 29 . (canceled) 
     
     
         40 . An extracellular vesicle produced according to the method of  claim 33 . 
     
     
         41 . A pharmaceutical composition comprising the extracellular vesicle of  claim 5 . 
     
     
         42 . A method of delivering a small molecule to a cell, the method comprising contacting the cell with the extracellular vesicle of  claim 5 , thereby delivering the small molecule agent to the cell. 
     
     
         43 . (canceled) 
     
     
         44 . The method of  claim 46 , wherein the cancer is breast cancer, pancreatic cancer, glioblastoma, melanoma, lung cancer, ovarian cancer, or liver cancer. 
     
     
         45 . A method of altering gene expression in a cell, the method comprising contacting the cell with the extracellular vesicle of  claim 5 . 
     
     
         46 . A method for treating cancer in a subject comprising administering to the subject a pharmaceutical composition comprising an effective amount of the extracellular vesicle of  claim 5 . 
     
     
         47 . The method of  claim 46 , wherein the cancer is cholangiocarcinoma, hepatocellular carcinoma, or hepatoma. 
     
     
         48 . An extracellular vesicle isolated from a cancer associated fibroblast (CAF), wherein the extracellular vesicle comprises a small molecule and a heterologous polynucleotide comprising miR-195, miR-126, or miR-192, and wherein the extracellular vesicle selectively targets a cancer cell. 
     
     
         49 . A method for treating cancer in a subject comprising administering to the subject a pharmaceutical composition comprising an effective amount of the extracellular vesicle of  claim 48 . 
     
     
         50 . A pharmaceutical composition comprising an effective amount of a first and a second extracellular vesicle, wherein the first extracellular vesicle comprises a small molecule and the second extracellular vesicle comprises a heterologous polynucleotide comprising miR-195, miR-126, or miR-192. 
     
     
         51 . A method for treating cancer in a subject comprising administering to the subject the pharmaceutical composition of  claim 50 . 
     
     
         52 . (canceled) 
     
     
         53 . A composition for imaging studies, the composition comprising the extracellular vesicle of  claim 5 , wherein the vesicle comprises a detectable or imaging agent. 
     
     
         54 . (canceled) 
     
     
         55 . The composition of  claim 54 , wherein the imaging agent is a nanoparticle, magnetite, nanoparticle, paramagnetic particle, microsphere, nanosphere, and is selectively targeted to cancer cells. 
     
     
         56 - 57 . (canceled) 
     
     
         58 . A kit for delivering an agent to a cell the kit, wherein the agent comprises the extracellular vesicle of  claim 5 .