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 classifying a tissue of origin of a biological sample, the method comprising:
 (a) obtaining a biological sample from a subject;   (b) determining an expression profile in said sample of nucleic acid sequences selected from the group consisting of SEQ ID NOS: 1-49, or a sequence having at least about 80% identity thereto; and   (c) comparing said expression profile to a reference expression profile by using a classifier algorithm; whereby the expression of any of said nucleic acid sequences allows the classification of the tissue of origin of said sample.   
     
     
         2 . The method of  claim 1 , wherein said tissue is selected from the group consisting of liver, lung, bladder, prostate, breast, colon, ovary, testis, stomach, thyroid, pancreas, brain, head and neck, kidney, melanocytes, thymus, biliary tract and esophagus. 
     
     
         3 . A method of classifying a cancer, said method comprising:
 (a) obtaining a biological sample from a subject;   (b) measuring the relative abundance in said sample of nucleic acid sequences selected from the group consisting of SEQ ID NOS: 1-49 or a sequence having at least about 80% identity thereto; and   (c) comparing said obtained measurement to a reference abundance of said nucleic acid by using a classifier algorithm;   whereby the relative abundance of said nucleic acid sequences allows the classification of said cancer.   
     
     
         4 . The method of  claim 3 , wherein said classifier algorithm is selected from the group consisting of a decision tree classifier, logistic regression classifier, nearest neighbor classifier, neural network classifier, Gaussian mixture model (GMM), Support Vector Machine (SVM) classifier, nearest centroid classifier, linear regression classifier and random forest classifier. 
     
     
         5 . The method of  claim 3 , wherein said sample is obtained from a subject with cancer of unknown primary (CUP), with a primary cancer or with a metastatic cancer. 
     
     
         6 . The method of  claim 3 , wherein said cancer is selected from the group consisting of liver cancer, biliary tract cancer, lung cancer, bladder cancer, prostate cancer, breast cancer, colon cancer, ovarian cancer, testicular cancer, stomach cancer, thyroid cancer, pancreas cancer, brain cancer, head and neck cancer, kidney cancer, melanoma, thymus cancer and esophagus cancer. 
     
     
         7 . The method of  claim 6 , wherein said testicular cancer is selected from the group consisting of testicular non-seminoma and testicular seminoma. 
     
     
         8 . The method of  claim 6 , wherein said lung cancer is selected from the group consisting of lung carcinoid, lung small cell carcinoma, lung adenocarcinoma and lung squamous cell carcinoma. 
     
     
         9 . The method of  claim 6 , wherein said ovarian cancer is selected from the group consisting of ovarian serous and ovarian endometrioid cancer. 
     
     
         10 . The method of  claim 6 , wherein said brain cancer is selected from the group consisting of brain astrocytoma and brain oligodendroglioma. 
     
     
         11 . The method of  claim 6 , wherein said thyroid cancer is selected from the group consisting of thyroid papillary, thyroid follicular and thyroid medullary cancer. 
     
     
         12 . The method of  claim 3 , wherein said biological sample is selected from the group consisting of bodily fluid, a cell line, a tissue sample, a biopsy sample, a needle biopsy sample, a fine needle biopsy (FNA) sample, a surgically removed sample, and a sample obtained by tissue-sampling procedures such as endoscopy, bronchoscopy, or laparoscopic methods. 
     
     
         13 . The method of  claim 12 , wherein said tissue is a fresh, frozen, fixed, wax-embedded or formalin-fixed paraffin-embedded (FFPE) tissue. 
     
     
         14 . The method of  claim 3  for classifying a cancer of liver origin, the method comprising measuring the relative abundance of a nucleic acid sequence selected from the group consisting of SEQ ID NOS: 6, 9, 25, 26, or a sequence having at least about 80% identity thereto in said sample; wherein the abundance of said nucleic acid sequence is indicative of a cancer of liver origin. 
     
     
         15 . The method of  claim 3  for classifying a cancer of testicular origin, the method comprising measuring the relative abundance of a nucleic acid sequence selected from the group consisting of SEQ ID NOS: 6, 26, 41, or a sequence having at least about 80% identity thereto in said sample; wherein the abundance of said nucleic acid sequence is indicative of a cancer of testicular origin. 
     
     
         16 . The method of  claim 3  for classifying a cancer of testicular seminoma origin, the method comprising measuring the relative abundance of a nucleic acid sequence selected from the group consisting of SEQ ID NOS: 6, 26, 31, 41, 45, 48, or a sequence having at least about 80% identity thereto in said sample; wherein the abundance of said nucleic acid sequence is indicative of a cancer of testicular seminoma origin. 
     
     
         17 . The method of  claim 3  for classifying a cancer of melanoma, the method comprising measuring the relative abundance of a nucleic acid sequence selected from the group consisting of SEQ ID NOS: 6, 15, 17, 26, 41, 46, or a sequence having at least about 80% identity thereto in said sample; wherein the abundance of said nucleic acid sequence is indicative of a cancer of melanoma origin. 
     
     
         18 . The method of  claim 3  for classifying a cancer of kidney origin, the method comprising measuring the relative abundance of a nucleic acid sequence selected from the group consisting of SEQ ID NOS: 6, 7, 15, 17, 26, 41, 46, 47, or a sequence having at least about 80% identity thereto in said sample; wherein the abundance of said nucleic acid sequence is indicative of a cancer of kidney origin. 
     
     
         19 . The method of  claim 3  for classifying a cancer of brain origin, the method comprising measuring the relative abundance of a nucleic acid sequence selected from the group consisting of SEQ ID NOS: 6, 7, 15, 17, 26, 41, 46, 47, or a sequence having at least about 80% identity thereto in said sample; wherein the abundance of said nucleic acid sequence is indicative of a cancer of brain origin. 
     
     
         20 . The method of  claim 3  for classifying a cancer of brain astrocytoma origin, the method comprising measuring the relative abundance of a nucleic acid sequence selected from the group consisting of SEQ ID NOS: 6, 7, 10, 15, 17, 26, 41, 46, 47, or a sequence having at least about 80% identity thereto in said sample; wherein the abundance of said nucleic acid sequence is indicative of a cancer of brain astrocytoma origin. 
     
     
         21 . The method of  claim 3  for classifying a cancer of thyroid medullary origin, the method comprising measuring the relative abundance of a nucleic acid sequence selected from the group consisting of SEQ ID NOS: 6, 17-19, 24, 26, 32, 41, 42, or a sequence having at least about 80% identity thereto in said sample; wherein the abundance of said nucleic acid sequence is indicative of a cancer of thyroid medullary origin. 
     
     
         22 . The method of  claim 3  for classifying a cancer of lung carcinoid origin, the method comprising measuring the relative abundance of a nucleic acid sequence selected from the group consisting of SEQ ID NOS: 3, 6, 17-19, 24, 26, 32, 36, 41, 42, or a sequence having at least about 80% identity thereto in said sample; wherein the abundance of said nucleic acid sequence is indicative of a cancer of lung carcinoid origin. 
     
     
         23 . The method of  claim 3  for classifying a cancer of lung small cell carcinoma origin, the method comprising measuring the relative abundance of a nucleic acid sequence selected from the group consisting of SEQ ID NOS: 3, 6, 17-19, 24, 26, 32, 36, 41, 42, or a sequence having at least about 80% identity thereto in said sample; wherein the abundance of said nucleic acid sequence is indicative of a cancer of lung small cell carcinoma origin. 
     
     
         24 . The method of  claim 3  for classifying a cancer of colon origin, the method comprising measuring the relative abundance of a nucleic acid sequence selected from the group consisting of SEQ ID NOS: 1, 3, 4, 6, 17-19, 21, 26, 29, 34, 37, 41, 42, 48, or a sequence having at least about 80% identity thereto in said sample; wherein the abundance of said nucleic acid sequence is indicative of a cancer of colon origin. 
     
     
         25 . The method of  claim 3  for classifying a cancer of stomach origin, the method comprising measuring the relative abundance of a nucleic acid sequence selected from the group consisting of SEQ ID NOS: 1, 3, 4, 6, 17-19, 21, 26, 29, 34, 37, 41, 42, 48, or a sequence having at least about 80% identity thereto in said sample; wherein the abundance of said nucleic acid sequence is indicative of a cancer of stomach origin. 
     
     
         26 . The method of  claim 3  for classifying a cancer of pancreas origin, the method comprising measuring the relative abundance of a nucleic acid sequence selected from the group consisting of SEQ ID NOS: 1, 3, 6, 17-19, 21, 26, 28, 29, 33, 37, 41, 42, or a sequence having at least about 80% identity thereto in said sample; wherein the abundance of said nucleic acid sequence is indicative of a cancer of pancreas origin. 
     
     
         27 . The method of  claim 3  for classifying a cancer of biliary tract origin, the method comprising measuring the relative abundance of a nucleic acid sequence selected from the group consisting of SEQ ID NOS: 1, 3, 6, 9, 17-19, 21, 25, 26, 28, 29, 33, 37, 41, 42, or a sequence having at least about 80% identity thereto in said sample; wherein the abundance of said nucleic acid sequence is indicative of a cancer of biliary tract origin. 
     
     
         28 . The method of  claim 3  for classifying a cancer of prostate origin, the method comprising measuring the relative abundance of a nucleic acid sequence selected from the group consisting of SEQ ID NOS: 3, 6, 17-21, 26, 41, 42, or a sequence having at least about 80% identity thereto in said sample; wherein the abundance of said nucleic acid sequence is indicative of a cancer of prostate origin. 
     
     
         29 . The method of  claim 3  for classifying a cancer of ovarian origin, the method comprising measuring the relative abundance of a nucleic acid sequence selected from the group consisting of SEQ ID NOS: 3, 5, 6, 11, 17-21, 26, 30, 41, 42, or a sequence having at least about 80% identity thereto in said sample; wherein the abundance of said nucleic acid sequence is indicative of a cancer of ovarian origin. 
     
     
         30 . The method of  claim 3  for classifying a cancer of ovarian endometrioid origin, the method comprising measuring the relative abundance of a nucleic acid sequence selected from the group consisting of SEQ ID NOS: 2, 3, 5, 6, 11, 17-22, 26, 30, 41, 42, or a sequence having at least about 80% identity thereto in said sample; wherein the abundance of said nucleic acid sequence is indicative of a cancer of ovarian endometrioid origin. 
     
     
         31 . The method of  claim 3  for classifying a cancer of breast origin, the method comprising measuring the relative abundance of a nucleic acid sequence selected from the group consisting of SEQ ID NOS: 3, 5, 6, 11, 17-22, 26, 30, 39, 41, 42, or a sequence having at least about 80% identity thereto in said sample; wherein the abundance of said nucleic acid sequence is indicative of a cancer of breast origin. 
     
     
         32 . The method of  claim 3  for classifying a cancer of lung adenocarcinoma origin, the method comprising measuring the relative abundance of a nucleic acid sequence selected from the group consisting of SEQ ID NOS: 3, 5, 6, 8, 11, 16-22, 26, 27, 30, 37, 39, 41, 42, or a sequence having at least about 80% identity thereto in said sample; wherein the abundance of said nucleic acid sequence is indicative of a cancer of lung adenocarcinoma origin. 
     
     
         33 . The method of  claim 3  for classifying a cancer of papillary thyroid origin, the method comprising measuring the relative abundance of a nucleic acid sequence selected from the group consisting of SEQ ID NOS: 3, 5, 6, 8, 11, 16-22, 26, 27, 29, 30, 37-39, 41, 42, or a sequence having at least about 80% identity thereto in said sample; wherein the abundance of said nucleic acid sequence is indicative of a cancer of papillary thyroid origin. 
     
     
         34 . The method of  claim 3  for classifying a cancer of follicular thyroid origin, the method comprising measuring the relative abundance of a nucleic acid sequence selected from the group consisting of SEQ ID NOS: 3, 5, 6, 8, 11, 16-22, 26, 27, 29, 30, 37-39, 41, 42, or a sequence having at least about 80% identity thereto in said sample; wherein the abundance of said nucleic acid sequence is indicative of a cancer of follicular thyroid origin. 
     
     
         35 . The method of  claim 3  for classifying a cancer of thymus origin, the method comprising measuring the relative abundance of a nucleic acid sequence selected from the group consisting of SEQ ID NOS: 3, 5, 6, 11, 16-22, 26, 27, 29, 30, 35, 39, 41, 42, or a sequence having at least about 80% identity thereto in said sample; wherein the abundance of said nucleic acid sequence is indicative of a cancer of thymus origin. 
     
     
         36 . The method of  claim 3  for classifying a cancer of bladder origin, the method comprising measuring the relative abundance of a nucleic acid sequence selected from the group consisting of SEQ ID NOS: 3-6, 11, 16-22, 26, 27, 29, 30, 35, 39, 41, 42, 44, or a sequence having at least about 80% identity thereto in said sample; wherein the abundance of said nucleic acid sequence is indicative of a cancer of bladder origin. 
     
     
         37 . The method of  claim 3  for classifying a cancer of lung squamous origin, the method comprising measuring the relative abundance of a nucleic acid sequence selected from the group consisting of SEQ ID NOS: 3-6, 11, 16-23, 26, 27, 29, 30, 32, 35, 39, 41, 42, 44, or a sequence having at least about 80% identity thereto in said sample; wherein the abundance of said nucleic acid sequence is indicative of a cancer of lung squamous origin. 
     
     
         38 . The method of  claim 3  for classifying a cancer of head and neck origin, the method comprising measuring the relative abundance of a nucleic acid sequence selected from the group consisting of SEQ ID NOS: 3-6, 11, 14, 16-23, 26, 27, 29, 30, 32, 35, 37, 39, 41, 42, 44, 45, or a sequence having at least about 80% identity thereto in said sample; wherein the abundance of said nucleic acid sequence is indicative of a cancer of head and neck origin. 
     
     
         39 . The method of  claim 3  for classifying a cancer of esophagus origin, the method comprising measuring the relative abundance of a nucleic acid sequence selected from the group consisting of SEQ ID NOS: 3-6, 11, 14, 16-23, 26, 27, 29, 30, 32, 35, 37, 39, 41, 42, 44, 45, or a sequence having at least about 80% identity thereto in said sample; wherein the abundance of said nucleic acid sequence is indicative of a cancer of esophagus origin. 
     
     
         40 . The method of  claim 3 , wherein the nucleic acid sequence expression profile or relative abundance is determined by a method selected from the group consisting of nucleic acid hybridization and nucleic acid amplification. 
     
     
         41 . The method of  claim 40 , wherein the nucleic acid hybridization is performed using a solid-phase nucleic acid biochip array or in situ hybridization. 
     
     
         42 . The method of  claim 40 , wherein said nucleic acid amplification method is real-time PCR. 
     
     
         43 . The method of  claim 42 , wherein said real-time PCR comprises forward and reverse primers. 
     
     
         44 . The method of  claim 43 , wherein said forward primer comprises a sequence selected from the group consisting of SEQ ID NOS: 50-98 and 150. 
     
     
         45 . The method of  claim 43 , wherein said reverse primer comprises SEQ ID NO: 288. 
     
     
         46 . The method of  claim 43 , wherein said real-time PCR method further comprises a probe. 
     
     
         47 . The method of  claim 46 , wherein said probe comprises a sequence selected from the group consisting of a sequence that is complementary to a sequence selected from SEQ ID NOS: 1-49; a fragment thereof and a sequence having at least about 80% identity thereto. 
     
     
         48 . The method of  claim 47 , wherein said probe comprises a sequence selected from the group consisting of SEQ ID NOS: 99-149 and 151. 
     
     
         49 . A kit for cancer classification, said kit comprising a probe comprising a sequence selected from the group consisting of a sequence that is complementary to a sequence selected from SEQ ID NOS: 1-49; a fragment thereof and a sequence having at least about 80% identity thereto. 
     
     
         50 . The kit of  claim 49 , wherein the probe comprises a sequence selected from the group consisting of SEQ ID NOS: 99-149 and 151. 
     
     
         51 . The kit of  claim 50 , wherein said cancer is selected from the group consisting of liver cancer, biliary tract cancer, lung cancer, bladder cancer, prostate cancer, breast cancer, colon cancer, ovarian cancer, testicular cancer, stomach cancer, thyroid cancer, pancreas cancer, brain cancer, head and neck cancer, kidney cancer, melanoma, thymus cancer and esophagus cancer.