Identification and use of circulating tumor markers

Abstract

Methods for creating a library of recurrently mutated genomic regions and for using the library to analyze cancer-specific and patient-specific genetic alterations in a patient are provided. The methods can be used to measure tumor-derived nucleic acids in patient blood and thus to monitor the progression of disease. The methods can also be used for cancer screening.

What is claimed is: 
     
         1 . A method for creating a library of recurrently mutated genomic regions comprising:
 identifying a plurality of genomic regions from a group of genomic regions that are recurrently mutated in a specific cancer;   wherein the library comprises the plurality of genomic regions;   the plurality of genomic regions comprises at least 10 different genomic regions; and   at least one mutation within the plurality of genomic regions is present in at least 60% of all subjects with the specific cancer.   
     
     
         2 . The method of  claim 1 , wherein the plurality of genomic regions comprises at least 25, at least 50, at least 100, at least 150, at least 200, or at least 500 different genomic regions. 
     
     
         3 . The method of  claim 1 , wherein at least two mutations within the plurality of genomic regions or at least three mutations within the plurality of genomic regions is present in at least 60% of all subjects with the specific cancer. 
     
     
         4 . The method of  claim 1 , wherein at least one mutation within the plurality of genomic regions is present in at least 60%, 70%, 80%, 90%, 95%, 98%, 99%, or 99.9% of all subjects with the specific cancer. 
     
     
         5 . The method of  claim 1 , wherein the identifying step comprises for each genomic region in the plurality of genomic regions, ranking the genomic region to maximize the number of all subjects with the specific cancer having at least one mutation within the genomic region. 
     
     
         6 . The method of  claim 1 , wherein the identifying step comprises for each genomic region in the plurality of genomic regions, ranking the genomic region to maximize the ratio between the number of all subjects with the specific cancer having at least one mutation within the genomic region and the length of the genomic region. 
     
     
         7 . The method of  claim 1 , wherein the library comprises a plurality of genomic regions encoding a plurality of driver sequences. 
     
     
         8 . The method of  claim 7 , wherein the driver sequences are known driver sequences. 
     
     
         9 . The method of  claim 7 , wherein the driver sequences are recurrently mutated in the specific cancer. 
     
     
         10 . The method of  claim 1 , wherein the library comprises a plurality of genomic regions that are recurrently rearranged in the specific cancer. 
     
     
         11 . The method of  claim 1 , wherein the specific cancer is a carcinoma. 
     
     
         12 . The method of  claim 11 , wherein the carcinoma is an adenocarcinoma, a non-small cell lung cancer, or a squamous cell carcinoma. 
     
     
         13 . The method of  claim 1 , wherein the cumulative length of the plurality of genomic regions is at most 30 Mb, 20 Mb, 10 Mb, 5 Mb, 2 Mb, 1 Mb, 500 kb, 200 kb, 100 kb, 50 kb, 20 kb, or 10 kb. 
     
     
         14 . A method for analyzing a cancer-specific genetic alteration in a subject comprising the steps of:
 obtaining a tumor nucleic acid sample and a genomic nucleic acid sample from a subject with a specific cancer;   sequencing a plurality of target regions in the tumor nucleic acid sample and in the genomic nucleic acid sample to obtain a plurality of tumor nucleic acid sequences and a plurality of genomic nucleic acid sequences; and   comparing the plurality of tumor nucleic acid sequences to the plurality of genomic nucleic acid sequences to identify a patient-specific genetic alteration in the tumor nucleic acid sample;   wherein the plurality of target regions are selected from a plurality of genomic regions that are recurrently mutated in the specific cancer;   the plurality of genomic regions comprises at least 10 different genomic regions; and   at least one mutation within the plurality of genomic regions is present in at least 60% of all subjects with the specific cancer.   
     
     
         15 . The method of  claim 14 , wherein the plurality of genomic regions comprises at least 25, at least 50, at least 100, at least 150, at least 200, or at least 500 different genomic regions. 
     
     
         16 . The method of  claim 14 , wherein at least two mutations within the plurality of genomic regions or at least three mutations within the plurality of genomic regions is present in at least 60% of all subjects with the specific cancer. 
     
     
         17 . The method of  claim 14 , wherein at least one mutation within the plurality of genomic regions is present in at least 60%, 70%, 80%, 90%, 95%, 98%, 99%, or 99.9% of all subjects with the specific cancer. 
     
     
         18 . The method of  claim 14 , wherein each genomic region in the plurality of genomic regions is identified by ranking the genomic region to maximize the number of all subjects with the specific cancer having at least one mutation within the genomic region. 
     
     
         19 . The method of  claim 14 , wherein each genomic region in the plurality of genomic regions is identified by ranking the genomic region to maximize the ratio between the number of all subjects with the specific cancer having at least one mutation within the genomic region and the length of the genomic region. 
     
     
         20 . The method of  claim 14 , wherein the plurality of genomic regions comprises genomic regions encoding a plurality of driver sequences. 
     
     
         21 . The method of  claim 20 , wherein the driver sequences are known driver sequences. 
     
     
         22 . The method of  claim 20 , wherein the driver sequences are recurrently mutated in the specific cancer. 
     
     
         23 . The method of  claim 14 , wherein the plurality of genomic regions comprises genomic regions that are recurrently rearranged in the specific cancer. 
     
     
         24 . The method of  claim 14 , wherein the specific cancer is a carcinoma. 
     
     
         25 . The method of  claim 24 , wherein the carcinoma is an adenocarcinoma, a non-small cell lung cancer, or a squamous cell carcinoma. 
     
     
         26 . The method of  claim 14 , wherein the cumulative length of the plurality of genomic regions is at most 30 Mb, 20 Mb, 10 Mb, 5 Mb, 2 Mb, 1 Mb, 500 kb, 200 kb, 100 kb, 50 kb, 20 kb, or 10 kb. 
     
     
         27 . The method of any one of  claims 14 - 26 , further comprising the steps of:
 obtaining a cell-free nucleic acid sample from the subject; and   identifying the patient-specific genetic alteration in the cell-free nucleic acid sample.   
     
     
         28 . The method of  claim 27 , wherein the step of identifying the patient-specific genetic alteration in the cell-free nucleic acid sample comprises sequencing a genomic region comprising the patient-specific genetic alteration in the cell-free sample. 
     
     
         29 . The method of  claim 27 , wherein the step of obtaining a tumor nucleic acid sample and a genomic nucleic acid sample comprises the step of enriching the plurality of target regions in the tumor nucleic acid sample and the genomic nucleic acid sample. 
     
     
         30 . The method of  claim 29 , wherein the enriching step comprises use of a custom library of biotinylated DNA. 
     
     
         31 . The method of  claim 27 , wherein the step of obtaining a cell-free nucleic acid sample comprises the step of enriching the plurality of target regions in the cell-free nucleic acid sample. 
     
     
         32 . The method of  claim 27 , further comprising the step of quantifying the cancer-specific genetic alteration in the cell-free sample. 
     
     
         33 . A method for screening a cancer-specific genetic alteration in a subject comprising the steps of:
 obtaining a cell-free nucleic acid sample from a subject;   sequencing a plurality of target regions in the cell-free sample to obtain a plurality of cell-free nucleic acid sequences; and   identifying a cancer-specific genetic alteration in the cell-free sample;   wherein the plurality of target regions are selected from a plurality of genomic regions that are recurrently mutated in the specific cancer;   the plurality of genomic regions comprises at least 10 different genomic regions; and   at least one mutation within the plurality of genomic regions is present in at least 60% of all subjects with the specific cancer.   
     
     
         34 . The method of  claim 33 , wherein the plurality of genomic regions comprises at least 25, at least 50, at least 100, at least 150, at least 200, or at least 500 different genomic regions. 
     
     
         35 . The method of  claim 33 , wherein at least two mutations within the plurality of genomic regions or at least three mutations within the plurality of genomic regions is present in at least 60% of all subjects with the specific cancer. 
     
     
         36 . The method of  claim 33 , wherein at least one mutation within the plurality of genomic regions is present in at least 60%, 70%, 80%, 90%, 95%, 98%, 99%, or 99.9% of all subjects with the specific cancer. 
     
     
         37 . The method of  claim 33 , wherein each genomic region in the plurality of genomic regions is identified by ranking the genomic region to maximize the number of all subjects with the specific cancer having at least one mutation within the genomic region. 
     
     
         38 . The method of  claim 33 , wherein each genomic region in the plurality of genomic regions is identified by ranking the genomic region to maximize the ratio between the number of all subjects with the specific cancer having at least one mutation within the genomic region and the length of the genomic region. 
     
     
         39 . The method of  claim 33 , wherein the plurality of genomic regions comprises genomic regions encoding a plurality of driver sequences. 
     
     
         40 . The method of  claim 39 , wherein the driver sequences are known driver sequences. 
     
     
         41 . The method of  claim 39 , wherein the driver sequences are recurrently mutated in the specific cancer. 
     
     
         42 . The method of  claim 33 , wherein the plurality of genomic regions comprises genomic regions that are recurrently rearranged in the specific cancer. 
     
     
         43 . The method of  claim 33 , wherein the specific cancer is a carcinoma. 
     
     
         44 . The method of  claim 43 , wherein the carcinoma is an adenocarcinoma, a non-small cell lung cancer, or a squamous cell carcinoma. 
     
     
         45 . The method of  claim 33 , wherein the cumulative length of the plurality of genomic regions is at most 30 Mb, 20 Mb, 10 Mb, 5 Mb, 2 Mb, 1 Mb, 500 kb, 200 kb, 100 kb, 50 kb, 20 kb, or 10 kb. 
     
     
         46 . The method of  claim 33 , wherein the step of obtaining a cell-free nucleic acid sample comprises the step of enriching the plurality of target regions in the cell-free nucleic acid sample. 
     
     
         47 . The method of  claim 46 , wherein the enriching step comprises use of a custom library of biotinylated DNA.