Use of circulating cell-free rna for diagnosis and/or monitoring cancer

Abstract

The invention provides compositions, methods, and systems for using cell-free RNA for improved detection of rare cells and/or species that are useful for diagnosing and/or monitoring cancer. The invention also provides for compositions, methods, and systems for early detection of cells that are resistant and/or developing resistance to cancer therapies.

What is claimed is: 
     
         1 . A method for detecting mRNA of a biomarker in a biological sample from an individual, the method comprising:
 a. isolating cfRNA from the biological sample on a solid support, wherein the blood sample has been interacted with RNA stabilizer;   b. digesting DNA from the biological sample while the cfRNA is on the solid support;   c. eluting cfRNA at least once from the solid support;   d. reverse transcribing the cfRNA to cDNA;   e. reacting the cDNA with at least one primer that is specific for
 i. a gene mutation specific to the biomarker, or 
 ii. a fusion transcript of the biomarker, or 
 iii. gene expression of the biomarker; and 
   wherein the biomarker is selected from the group consisting of ERCC1, EGFR, TS, AREG, EREG, VEGFR2, EML4ALK, ROS1, RET, c-Met, FGFR1, KRAS, BRAF, NRAS, Her-2, PIK3CA, KIT, GNAQ, and GNA11.   
     
     
         2 . The method of  claim 1  wherein the eluate of step (c) is passed over the same column for a dual elution prior to the reverse transcribing the cfRNA to cDNA. 
     
     
         3 . The method of  claim 1  wherein the individual has or is suspected of having cancer. 
     
     
         4 . The method of  claim 1 , wherein the biological sample is a blood sample or a plasma sample. 
     
     
         5 . The method of  claim 4 , wherein the plasma sample is processed within 7 days of interacting with the RNA stabilizer. 
     
     
         6 . The method of  claim 1  wherein random hexamers are used in step (d) to reverse transcribe the RNA to cDNA. 
     
     
         7 . The method of  claim 1 , further comprising sequencing the cDNA. 
     
     
         8 . The method of  claim 7 , wherein the sequencing comprises polymerase chain reaction (PCR)-based sequencing. 
     
     
         9 . The method of  claim 8 , wherein the PCR-based sequencing is next generation sequencing.