Dna methylation biomarkers for lung cancer

Abstract

The present invention relates to the identification of novel DNA biomarkers and the use of the aberrant methylation patterns of the biomarkers to diagnose a disease or a condition (e.g., a cancer) associated therewith. In particular, the present invention relates to the use of the novel DNA biomarkers to diagnose lung cancers, e.g., squamous cell carcinomas and adenocarcinomas.

What is claimed is: 
     
         1 . A method of diagnosing lung cancer comprising:
 obtaining a lung tissue test sample from a subject;   measuring a methylation level of one or a combination of DNA biomarkers selected from the group consisting of SEQ ID NOS. 1-21 and 23-111 in the lung tissue test sample;   comparing the methylation level of the one or a combination of DNA biomarkers with the methylation level of a corresponding one or combination of DNA biomarkers in a normal lung tissue sample or lung standard sample; and   predicting that an increase in the methylation level of the lung tissue test sample in relation to that of the normal lung tissue sample or lung standard sample indicates that the subject is likely to have lung cancer.   
     
     
         2 . The method of  claim 1  wherein the aberrant methylation is hypermethylation or hypomethylation. 
     
     
         3 . The method of  claim 2  wherein the condition is a lung cancer. 
     
     
         4 . The method of  claim 1  wherein the lung cancer is squamous cell carcinoma. 
     
     
         5 . The method of  claim 4  wherein the one or a combination of DNA biomarkers are selected from one or more genes listed in Table 2 (SEQ ID NOS. 1-59). 
     
     
         6 . The method of  claim 4  wherein the one or a combination of DNA biomarkers are selected from the group consisting of BARHL2 (SEQ ID NO. 3), EVX2 (SEQ ID NO. 14), IRX2 (SEQ ID NO. 24), MEIS1 (SEQ ID NO. 11), MSX1 (SEQ ID NO. 22), NR2E1 (SEQ ID NO. 33), OC2 (SEQ ID NO. 55), OSR1 (SEQ ID NO. 7), OTX1 (SEQ ID NO. 10), PAX6 (SEQ ID NO. 44), TFAP2A (SEQ ID NO. 30), and ZNF577 (SEQ ID NO. 56). 
     
     
         7 . The method of  claim 4  wherein the lung cancer is adenocarcinoma. 
     
     
         8 . The method  claim 7  wherein the one or combination of DNA biomarkers are selected from one or more genes listed in Table 4 (SEQ ID NOS. 60-111). 
     
     
         9 . The method of  claim 7  wherein the one or combination of DNA biomarkers are selected from the group consisting of CHAD (SEQ ID NO. 63), DLX4 (SEQ ID NO. 64), GRIK2 (SEQ ID NO. 68), KNCG3 (SEQ ID NO. 74), NR2E1 (SEQ ID NO. 78), OSR1 (SEQ ID NO. 79), OTX1 (SEQ ID NO. 80), OTX2 (SEQ ID NO. 83), PROX1 (SEQ ID NO. 88), RUNX1 (SEQ ID NO. 90), and VAX1 (SEQ ID NO. 98). 
     
     
         10 . The method of  claim 1  wherein the one or a combination of DNA biomarkers are selected from the group consisting of BARHL2 (SEQ ID NO. 3), EVX2 (SEQ ID NO. 14), IRX2 (SEQ ID NO. 24), MEIS1 (SEQ ID NO. 11), MSX1 (SEQ ID NO. 22), NR2E1 (SEQ ID NO. 33), OC2 (SEQ ID NO. 55), OSR1 (SEQ ID NO. 7), OTX1 (SEQ ID NO. 10), PAX6 (SEQ ID NO. 44), TFAP2A (SEQ ID NO. 30), ZNF577 (SEQ ID NO. 56), CHAD (SEQ ID NO. 63), DLX4 (SEQ ID NO. 64), GRIK2 (SEQ ID NO. 68), KNCG3 (SEQ ID NO. 74), NR2E1 (SEQ ID NO. 78), OSR1 (SEQ ID NO. 79), OTX1 (SEQ ID NO. 80), OTX2 (SEQ ID NO. 83), PROX1 (SEQ ID NO. 88), RUNX1 (SEQ ID NO. 90), and VAX1 (SEQ ID NO. 98). 
     
     
         11 . The method of  claim 1  wherein the methylation level is measured by a methylated-CpG island recovery assay (MIRA), a combined bisulfite-restriction analysis (COBRA), or a methylation-specific PCR (MSP). 
     
     
         12 . The method of  claim 11  wherein the methylation levels of the one or a combination of DNA biomarkers are measured by an MIRA-assisted microarray analysis. 
     
     
         13 . The method of  claim 1  wherein the one or combination of DNA biomarkers are selected from the group consisting of OTX1 (SEQ ID NO. 10), IRX2 (SEQ ID NO. 24), OC2 (SEQ ID NO. 55), and EVX2 (SEQ ID NO. 14). 
     
     
         14 . The method of  claim 1  wherein the increase is more than 2 fold. 
     
     
         15 . The method of  claim 14  wherein the increase is more than 3 fold. 
     
     
         16 . A method of diagnosing lung cancer comprising:
 1) obtaining a lung tissue test sample from a subject;   2) obtaining a genome DNA from the lung tissue test sample from the subject;   3) obtaining methylated regions from the genome DNA;   4) hybridizing the methylated regions to a DNA microarray comprising one or a combination of DNA biomarkers selected from the group consisting of SEQ ID NOS. 1-21 and 23-111;   5) comparing the hybridization of the methylated regions from the genome DNA with the hybridization of the corresponding methylated regions of a normal lung tissue sample or lung standard sample genome DNA; and   6) predicting that an increase in the methylated regions of the genome DNA hybridizing to the DNA biomarker relative to the methylated regions of the normal lung tissue sample or lung standard sample genome DNA hybridizing to the one or a combination of DNA biomarkers indicates that the subject is likely to have lung cancer.   
     
     
         17 . The method of  claim 16  wherein the lung cancer is squamous cell carcinoma and the one or combination of DNA biomarkers are selected from is one or more genes listed in Table 2 (SEQ ID NOS. 1-59). 
     
     
         18 . The method of  claim 17  wherein the one or a combination of DNA biomarkers are selected from the group consisting of BARHL2 (SEQ ID NO. 3), EVX2 (SEQ ID NO. 14), IRX2 (SEQ ID NO. 24), MEIS1 (SEQ ID NO. 11), MSX1 (SEQ ID NO. 22), NR2E1 (SEQ ID NO. 33), OC2 (SEQ ID NO. 55), OSR1 (SEQ ID NO. 7), OTX1 (SEQ ID NO. 10), PAX6 (SEQ ID NO. 44), TFAP2A (SEQ ID NO. 30), and ZNF577 (SEQ ID NO. 56). 
     
     
         19 . The method of  claim 16  wherein the condition is adenocarcinoma and the one or combination of DNA biomarkers are selected from one or more genes listed in Table 4 (SEQ ID NOS. 60-111). 
     
     
         20 . The method of  claim 19  wherein the one or combination of DNA biomarkers are selected from the group consisting of CHAD (SEQ ID NO. 63), DLX4 (SEQ ID NO. 64), GRIK2 (SEQ ID NO. 68), KNCG3 (SEQ ID NO. 74), NR2E1 (SEQ ID NO. 78), OSR1 (SEQ ID NO. 79), OTX1 (SEQ ID NO. 80), OTX2 (SEQ ID NO. 83), PROX1 (SEQ ID NO. 88), RUNX1 (SEQ ID NO. 90), and VAX1 (SEQ ID NO. 98). 
     
     
         21 . A method of diagnosing a tumor from a test sample in a subject comprising the step of:
 measuring the methylation level of one or a combination of DNA biomarkers from the test sample and a standard sample:   wherein the fold difference between the methylation level of the test sample in relation to the methylation level of the standard sample indicates the likelihood of the test sample having the condition; and   wherein the one or combination of DNA biomarkers are selected from the group consisting of OTX1 (SEQ ID NO. 10), IRX2 (SEQ ID NO. 24), OC2 (SEQ ID NO. 55), EVX2 (SEQ ID NO. 14).   
     
     
         22 . The method of  claim 21  wherein the methylation level is measured by a methylated-CpG island recovery assay (MIRA), a combined bisulfite-restriction analysis (COBRA), or a methylation-specific PCR (MSP). 
     
     
         23 . A method of diagnosing a tumor from a test sample in a subject comprising the step of:
 measuring the methylation level of one or a combination of DNA biomarkers from the test sample and a standard sample:   wherein the fold difference between the methylation level of the test sample in relation to that of the standard sample indicates the likelihood of the test sample having the condition; and   wherein the DNA biomarker is the 3′ end of C8orf72 gene.