Gene expression signature for classification of tissue of origin of tumor samples

Abstract

The present invention provides a process for classification of cancers and tissues of origin through the analysis of the expression patterns of specific microRNAs and nucleic acid molecules relating thereto. Classification according to a microRNA tree-based expression framework allows optimization of treatment, and determination of specific therapy.

1 . A method of producing thyroid medullary cancer cDNA sequences, the method comprising:
 isolating RNA from a sample from a human subject, wherein the sample comprises a tumor cell;   contacting the RNA with a polyadenylation agent under conditions that are sufficient to form a polyadenylated RNA;   reverse transcribing the polyadenylated RNA in the presence of a universal poly(T) adapter to produce cDNA sequences comprising a poly(T) tail; and   amplifying the cDNA sequences using a forward primer that is specific for SEQ ID NO:42;   thereby producing thyroid medullary cancer cDNA sequences.   
     
     
         2 . The method of  claim 1 , wherein the amplifying further comprises amplifying the cDNA sequences with a reverse primer that is complementary to the poly(T) tail. 
     
     
         3 . The method of  claim 1 , wherein the sample is a biopsy. 
     
     
         4 . The method of  claim 1 , wherein the sample is a fine-needle aspiration. 
     
     
         5 . The method of  claim 1 , wherein the amplifying comprises quantitative PCR. 
     
     
         6 . A reaction mixture for generating cDNA sequences derived from a thyroid medullary cancer, the reaction mixture comprising:
 a nucleic acid sample obtained from a biological sample from a human subject, wherein the biological sample comprises a tumor cell;   a primer for generating cDNA sequences, wherein the primer is specific for SEQ ID NO:42; and   a detectable probe.   
     
     
         7 . The reaction mixture of  claim 6 , wherein the nucleic acid sample comprises cDNA sequences comprising a poly(T) tail, wherein the cDNA sequences are produced from an RNA sample that has been polyadenylated and reverse transcribed in the presence of a universal poly(T) adapter. 
     
     
         8 . The reaction mixture of  claim 7 , wherein the reaction mixture further comprises a reverse primer that is complementary to the poly(T) tail. 
     
     
         9 . The reaction mixture of  claim 6 , wherein the biological sample is a biopsy or a fine-needle aspiration. 
     
     
         10 . A method of detecting gene expression in a sample from a human subject, the method comprising:
 detecting the presence of SEQ ID NO:42 in a nucleic acid sample from the subject, wherein the detecting step comprises:   contacting the nucleic acid sample with a primer specific for SEQ ID NO:42;   amplifying at least one nucleic acid sequence in the sample; and   detecting the presence of an amplified nucleic acid sequence comprising SEQ ID NO:42;   thereby detecting the gene expression in the sample.   
     
     
         11 . The method of  claim 10 , wherein the nucleic acid sample is an RNA sample isolated from a biological sample from the subject, wherein the biological sample comprises a tumor cell. 
     
     
         12 . The method of  claim 11 , wherein the biological sample is a biopsy. 
     
     
         13 . The method of  claim 11 , wherein the biological sample is a fine-needle aspiration. 
     
     
         14 . The method of  claim 11 , wherein prior to the contacting step, the method comprises contacting the RNA sample with a polyadenylation agent under conditions that are sufficient to form a polyadenylated RNA and reverse transcribing the polyadenylated RNA in the presence of a universal poly(T) adapter to produce a nucleic acid sample comprising cDNA sequences. 
     
     
         15 . The method of  claim 10 , wherein the amplifying step comprises quantitative PCR. 
     
     
         16 . A method of identifying a subject as having a cancer of thyroid medullary origin, the method comprising:
 obtaining a nucleic acid sample from a human subject, wherein the nucleic acid sample is from a biological sample comprising a tumor cell;   contacting the nucleic acid sample with a primer specific for SEQ ID NO:42;   amplifying the nucleic acid sequences in the biological sample; and   detecting the presence of an amplified nucleic acid sequence comprising SEQ ID NO:42;   thereby identifying the subject as having a cancer of thyroid medullary origin.   
     
     
         17 . The method of  claim 16 , wherein the nucleic acid sample is an RNA sample, and wherein prior to the contacting step, the method comprises contacting the RNA sample with a polyadenylation agent under conditions that are sufficient to form a polyadenylated RNA and reverse transcribing the polyadenylated RNA in the presence of a universal poly(T) adapter to produce a nucleic acid sample comprising cDNA sequences. 
     
     
         18 . The method of  claim 16 , wherein the biological sample is a biopsy. 
     
     
         19 . The method of  claim 16 , wherein the biological sample is a fine-needle aspiration. 
     
     
         20 . The method of  claim 16 , wherein the amplifying step comprises quantitative PCR. 
     
     
         21 . The method of  claim 16 , wherein the detecting step comprises measuring the relative abundance of an amplified nucleic acid sequence comprising SEQ ID NO:42, relative to a reference value, and identifying the subject as having a cancer of thyroid medullary origin based on the relative abundance of the amplified nucleic acid sequence comprising SEQ ID NO:42.