Methods of Detecting Cervical Cancer

Abstract

Methods of detecting cervical dysplasia, such as cervical dysplasia likely to progress to carcinoma in a sample of human cervical cells, are provided. Methods of detecting changes in expression of one or more microRNAs or mRNAs associated with cervical dysplasia or cervical cancer are also provided. Compositions and kits are also provided.

1 . A method for detecting the presence of cervical dysplasia in a subject, the method comprising detecting a level of at least one target RNA in a cervical sample from the subject, wherein the at least one target RNA:
 (i) is capable of specifically hybridizing to a nucleic acid having a sequence selected from SEQ ID NOs: 1 to 41 and 133 to 211; or   (ii) comprises a sequence that is complementary to at least 15 contiguous nucleotides of a sequence selected from SEQ ID NOs: 1 to 41 and 133 to 211; or   (iii) comprises at least 15 contiguous nucleotides of a sequence selected from SEQ ID NOs: 345 to 388;   
       wherein a level of at least one target RNA in the sample that is greater than a normal level of the at least one target RNA indicates the presence of cervical dysplasia in the sample. 
     
     
         2 . The method of  claim 1 , wherein the method further comprises comparing the level of the at least one target RNA in the sample to a normal level of the at least one target RNA. 
     
     
         3 . A method for facilitating the detection of cervical dysplasia in a subject, comprising:
 (a) detecting a level of at least one target RNA in a cervical sample from the subject, wherein the at least one target RNA:
 (i) is capable of specifically hybridizing to a nucleic acid having a sequence selected from SEQ ID NOs: 1 to 41 and 133 to 211; or 
 (ii) comprises a sequence that is complementary to at least 15 contiguous nucleotides of a sequence selected from SEQ ID NOs: 1 to 41 and 133 to 211; or 
 (iii) comprises at least 15 contiguous nucleotides of a sequence selected from SEQ ID NOs: 345 to 388; and 
   (b) communicating the results of the detection to a medical practitioner for the purpose of determining whether the subject has cervical dysplasia.   
     
     
         4 . The method of  claim 1 , wherein detecting a level of at least one target RNA in a sample comprises:
 (a) hybridizing nucleic acids of the sample with at least one polynucleotide that is complementary to a target RNA in the sample or to a complement thereof; and   (b) detecting at least one complex comprising a polynucleotide hybridized to at least one nucleic acid selected from the target RNA, a DNA amplicon of the target RNA, and a complement of the target RNA.   
     
     
         5 . A method for detecting the presence of cervical dysplasia in a subject, comprising:
 (a) obtaining a cervical sample from the subject,   (b) providing the cervical sample to a laboratory for detection of the level of at least one target RNA in the sample, wherein the at least one target RNA:
 (i) is capable of specifically hybridizing to a nucleic acid having a sequence selected from SEQ ID NOs: 1 to 41 and 133 to 211; or 
 (ii) comprises a sequence that is complementary to at least 15 contiguous nucleotides of a sequence selected from SEQ ID NOs: 1 to 41 and 133 to 211; or 
 (iii) comprises at least 15 contiguous nucleotides of a sequence selected from SEQ ID NOs: 345 to 388; and 
   (c) receiving from the laboratory a communication indicating the level of at least one target RNA in the cervical sample;   
       wherein a level of at least one target RNA that is greater than a normal level of the at least one target RNA indicates the presence of cervical dysplasia in the sample. 
     
     
         6 . The method of  claim 1 , wherein the method further comprises isolating nucleic acids from the cervical sample. 
     
     
         7 . The method of  claim 6 , wherein the nucleic acids comprise RNA that has been separated from DNA. 
     
     
         8 . The method of  claim 1 , wherein at least one target RNA in its mature form comprises fewer than 30 nucleotides. 
     
     
         9 . The method of  claim 1 , wherein at least one target RNA is a microRNA. 
     
     
         10 . The method of  claim 1 , wherein levels of at least two target RNAs are detected, wherein at least two of the target RNAs:
 (i) are capable of specifically hybridizing to a nucleic acid having a sequence selected from SEQ ID NOs: 1 to 41 and 133 to 211; or   (ii) comprise a sequence that is complementary to at least 15 contiguous nucleotides of a sequence selected from SEQ ID NOs: 1 to 41 and 133 to 211; or   (iii) comprise at least 15 contiguous nucleotides of a sequence selected from SEQ ID NOs: 345 to 388; and   
       wherein the at least two target RNAs are different. 
     
     
         11 . The method of  claim 10 , wherein detection of a level of at least one target RNA that is greater than a normal level of the at least one target RNA indicates the presence of cervical dysplasia. 
     
     
         12 . The method of  claim 10 , wherein detection of levels of at least two target RNAs that are greater than normal levels of the at least two target RNAs indicates the presence of cervical dysplasia. 
     
     
         13 . The method of  claim 10 , wherein levels of at least three target RNAs are detected, wherein at least three of the target RNAs:
 (i) are capable of specifically hybridizing to a nucleic acid having a sequence selected from SEQ ID NOs: 1 to 41 and 133 to 211; or   (ii) comprise a sequence that is complementary to at least 15 contiguous nucleotides of a sequence selected from SEQ ID NOs: 1 to 41 and 133 to 211; or   (iii) comprise at least 15 contiguous nucleotides of a sequence selected from SEQ ID NOs: 345 to 388; and   
       wherein the at least three target RNAs are different. 
     
     
         14 . The method of  claim 13 , wherein detection of a level of at least one target RNA that is greater than a normal level of the at least one target RNA indicates the presence of cervical dysplasia. 
     
     
         15 . The method of  claim 13 , wherein detection of levels of at least two target RNAs that are greater than normal levels of the at least two target RNAs indicates the presence of cervical dysplasia. 
     
     
         16 . The method of  claim 13 , wherein detection of levels of at least three target RNAs that are greater than normal levels of the at least three target RNAs indicates the presence of cervical dysplasia. 
     
     
         17 . The method of  claim 10 , wherein levels of at least five target RNAs are detected. 
     
     
         18 . The method of  claim 1 , wherein a level is detected of at least one target RNA that:
 (i) does not specifically hybridize to a nucleic acid having a sequence selected from SEQ ID NOs: 1 to 41 and 133 to 211; or   (ii) does not comprise a sequence that is complementary to at least 15 contiguous nucleotides of a sequence selected from SEQ ID NOs: 1 to 41 and 133 to 211; and   (iii) does not comprise at least 15 contiguous nucleotides of a sequence selected from SEQ ID NOs: 345 to 388.   
     
     
         19 . The method of  claim 1 , wherein the method further comprises detection of a level of at least one target RNA that is an mRNA. 
     
     
         20 . The method of  claim 19 , wherein the mRNA is selected from CDKN2A, MKI67, TOP2A, MCM5, BIRC5, MMP9, and MCM2. 
     
     
         21 . The method of  claim 1 , wherein at least one target RNA:
 (a) is capable of specifically hybridizing to a sequence selected from SEQ ID NOs: 1, 2, 3, 4, 5, 6, 7 and 8; or   (b) comprises a sequence that is complementary to at least 15 contiguous nucleotides of a sequence selected from SEQ ID NOs: 1, 2, 3, 4, 5, 6, 7 and 8.   
     
     
         22 . The method of  claim 1 , wherein at least one target RNA:
 (a) is capable of specifically hybridizing to a sequence selected from SEQ ID NOs: 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11 and 12; or   (b) comprises a sequence that is complementary to at least 15 contiguous nucleotides of a sequence selected from SEQ ID NOs: 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11 and 12.   
     
     
         23 . The method of  claim 1 , wherein at least one target RNA:
 (a) is capable of specifically hybridizing to a sequence selected from SEQ ID NOs: 1, 5, 7, and 32; or   (b) comprises a sequence that is complementary to at least 15 contiguous nucleotides of a sequence selected from SEQ ID NOs: 1, 5, 7, and 32.   
     
     
         24 . The method of  claim 1 , wherein at least one target RNA:
 (a) is capable of specifically hybridizing to a sequence selected from SEQ ID NOs: 142, 151, 153, 193, 194, 205, 172, 208, 210, and 211; or   (b) comprises a sequence that is complementary to at least 15 contiguous nucleotides of a sequence selected from SEQ ID NOs: 142, 151, 153, 193, 194, 205, 172, 208, 210, and 211.   
     
     
         25 . A synthetic polynucleotide comprising a first region, wherein the first region comprises a sequence of at least 8 contiguous nucleotides that is identical or complementary to a sequence of at least 8 contiguous nucleotides of one of SEQ ID NOs: 1 to 7, 9 to 37, 133 to 201, and 345 to 388. 
     
     
         26 . The synthetic polynucleotide of  claim 25 , wherein the first region comprises a sequence of at least 9 contiguous nucleotides that is identical or complementary to a sequence of at least 9 contiguous nucleotides of one of SEQ ID NOs: 1 to 7, 9 to 37, 133 to 201, and 345 to 388. 
     
     
         27 . The synthetic polynucleotide of  claim 25 , wherein the first region comprises a sequence of at least 10 contiguous nucleotides that is identical or complementary to a sequence of at least 10 contiguous nucleotides of one of SEQ ID NOs: 1 to 7, 9 to 37, 133 to 201, and 345 to 388. 
     
     
         28 . The synthetic polynucleotide of  claim 25 , wherein the first region comprises a sequence of at least 12 contiguous nucleotides that is identical or complementary to a sequence of at least 12 contiguous nucleotides of one of SEQ ID NOs: 1 to 7, 9 to 37, 133 to 201, and 345 to 388. 
     
     
         29 . The synthetic polynucleotide of  claim 25 , wherein the polynucleotide comprises a detectable label. 
     
     
         30 . The synthetic polynucleotide of  claim 29 , wherein the detectable label is a FRET label. 
     
     
         31 . The synthetic polynucleotide of  claim 25 , wherein the first region is identical or complementary to a region of a target RNA. 
     
     
         32 . The synthetic polynucleotide of  claim 31 , wherein the polynucleotide further comprises a second region that is not identical or complementary to a region of the target RNA. 
     
     
         33 . A composition comprising a plurality of synthetic polynucleotides, wherein at least one polynucleotide comprises a first region comprising a sequence of at least 8 contiguous nucleotides that is identical or complementary to a sequence of at least 8 contiguous nucleotides of one or more of SEQ ID NOs: 1 to 7, 9 to 37, 133 to 201, and 345 to 388. 
     
     
         34 . The composition of  claim 33 , wherein at least two polynucleotides of the plurality of synthetic polynucleotides comprise a first region comprising a sequence of at least 9 contiguous nucleotides that is identical or complementary to a sequence of at least 9 contiguous nucleotides of one or more of SEQ ID NOs: 1 to 7, 9 to 37, 133 to 201, and 345 to 388, and wherein the first regions of the at least two polynucleotides are different. 
     
     
         35 . The composition of  claim 33 , wherein at least three polynucleotides of the plurality of synthetic polynucleotides comprise a first region comprising a sequence of at least 10 contiguous nucleotides that is identical or complementary to a sequence of at least 10 contiguous nucleotides of one or more of SEQ ID NOs: 1 to 7, 9 to 37, 133 to 201, and 345 to 388, and wherein the first regions of the at least three polynucleotides are different. 
     
     
         36 . The composition of  claim 33 , wherein at least five polynucleotides of the plurality of synthetic polynucleotides comprise a first region comprising a sequence of at least 12 contiguous nucleotides that is identical or complementary to a sequence of at least 12 contiguous nucleotides of one or more of SEQ ID NOs: 1 to 7, 9 to 37, 133 to 201, and 345 to 388, and wherein the first regions of the at least five polynucleotides are different. 
     
     
         37 . A kit comprising a synthetic polynucleotide of  claim 25 . 
     
     
         38 . A kit comprising a composition of  claim 33 . 
     
     
         39 . The kit of  claim 37 , wherein the kit further comprises at least one polymerase. 
     
     
         40 . The kit of  claim 37 , wherein the kit further comprises dNTPs.