Ultra-small apob-containing particles and methods of use thereof

Abstract

The present disclosure provides an isolated particle comprising very high density, ultra small, lipid depleted apo B containing particles, and may also contain cytokeratin 8. The isolated particle is useful in diagnostic assays, which are also provided.

What is claimed is: 
     
         1 . An isolated, very high density, ultra small, lipid-depleted particle, the particle comprising:
 a) apolipoprotein B (apoB);   b) a cytokeratin 8 polypeptide,   wherein the isolated particle has an average particle diameter in a range of from about 7.1 nm to about 22 nm,   wherein the isolated particle has a density >1.21 g/mL, and   wherein the isolated particle has substantially no cholesterol and substantially no triglyceride.   
     
     
         2 . The isolated particle of  claim 1 , wherein the apoB comprises an amino acid sequence having at least about 95% amino acid sequence identity with the amino acid sequence set forth in SEQ ID NO:1. 
     
     
         3 . The isolated particle of  claim 1 , wherein the cytokeratin 8 comprises an amino acid sequence having at least about 95% amino acid sequence identity with the amino acid sequence set forth in SEQ ID NO:2. 
     
     
         4 . The isolated particle of  claim 1 , wherein the particle is at least 85% pure. 
     
     
         5 . The isolated particle of  claim 1 , wherein the particle is at least 95% pure. 
     
     
         6 . A method for assessing risk of cardiovascular disease (CVD) in an individual, the method comprising:
 detecting a level of the particle of  claim 1  in a biological sample from the individual; and   assessing the risk of CVD based on the results of said detecting, wherein a level of the particle that is higher than a normal control level indicates that the individual has an increased risk of CVD.   
     
     
         7 . The method of  claim 6 , wherein the biological sample is blood or a blood fraction. 
     
     
         8 . The method of  claim 6 , further comprising generating a report that provides an indication of the risk that the individual will develop CVD. 
     
     
         9 . The method of  claim 6 , wherein the individual is a human. 
     
     
         10 . The method of  claim 6 , wherein the individual exhibits at least one clinical symptom or sign of cardiovascular disease. 
     
     
         11 . The method of  claim 6 , further comprising communicating to the individual various treatment options based on the results of the detecting step. 
     
     
         12 . The method of  claim 6 , further comprising treating the individual for CVD. 
     
     
         13 . A method of assessing the efficacy of a treatment for a cardiovascular disease in an individual, the method comprising:
 a) analyzing the level of the particle of  claim 1  in a biological sample obtained from the subject following the treatment; and   b) comparing the post-treatment level to a pre-treatment level,   wherein a post-treatment level that is lower than the pre-treatment level indicates that the treatment was efficacious.   
     
     
         14 . The method of  claim 13 , wherein the individual is a human. 
     
     
         15 . A method of determining the risk of mortality due to a cardiovascular disease (CVD) in an individual, the method comprising detecting a level of the particle of  claim 1  in a biological sample from the individual, wherein a level of the particle that is higher than a normal control level indicates that the individual has an increased risk of mortality due to a CVD. 
     
     
         16 . A kit for assessing risk of cardiovascular disease, the kit comprising:
 a) a reagent that specifically binds apolipoprotein B-100; and   b) a reagent that specifically binds cytokeratin 8.   
     
     
         17 . The kit of  claim 16 , further comprising an isolated particle of  claim 1 . 
     
     
         18 . The kit of  claim 16 , further comprising instructions for use. 
     
     
         19 . The kit of  claim 16 , wherein each of (a) and (b) is in a separate container. 
     
     
         20 . The kit of  claim 16 , wherein each of (a) and (b) is an antibody. 
     
     
         21 . The kit of  claim 20 , wherein each of the antibodies is immobilized on an insoluble support or is detectably labeled. 
     
     
         22 . A method of detecting the particle of  claim 1  in a biological sample, the method comprising:
 a) contacting the biological sample with an antibody that binds apolipoprotein B-100 and/or an antibody that binds cytokeratin 8; and 
 b) detecting binding of the antibody to molecules in the biological sample. 
 
     
     
         23 . The method of  claim 22 , wherein the biological sample is obtained from an individual who is being evaluated for possible cardiovascular disease (CVD) or CVD risk. 
     
     
         24 . A method of isolating the particle of  claim 1 , wherein the method comprises subjecting a sample comprising the particle to an immunoaffinity method, a density gradient method, or a gradient gel electrophoresis method. 
     
     
         25 . The method of  claim 24 , wherein the method comprises contacting a sample comprising the particle with an immobilized antibody specific for apoB; and eluting particles bound to the apoB.