Compositions and Methods for Detection of Colorectal Cancer

Abstract

We have identified a new variant of ileal bile acid binding protein (IBABP), designated IBABP-L, which is a biomarker for colorectal cancer. The transcript for IBABP-L arises from an alternative start site and includes three exons that are absent in IBABP. IBABP-L also shares part of a fourth exon with IBABP. The protein encoded by IBABP-L contains a deduced 49 residue N-terminal sequence that is not found in the IBABP protein. The present invention provides methods for diagnosing colorectal cancer and other compositions and methods based on this discovery.

1 . An isolated polynucleotide comprising a sequence that has at least 90% nucleic acid sequence identity to a native IBABP-L polynucleotide and that hybridizes selectively to the native IBABP-L polypeptide. 
     
     
         2 . The isolated polynucleotide of  claim 1  wherein the sequence that has at least 95% identity to a native IBABP-L polynucleotide. 
     
     
         3 . The isolated polynucleotide of  claim 1  wherein the sequence has at least 99% nucleic acid sequence identity to a native IBABP-L polynucleotide. 
     
     
         4 . The isolated polynucleotide of  claim 1  wherein the sequence has 100% nucleotide sequence identity to a native IBABP-L polynucleotide. 
     
     
         5 . An isolated polynucleotide comprising a sequence at least 100 nucleotides in length that has at least 90% nucleic acid sequence identity to a native IBABP-L exon 1-3 polynucleotide. 
     
     
         6 . The isolated polynucleotide of  claim 5  wherein the sequence has at least 95% nucleic acid sequence identity to the native IBABP-L exon 1-3 polynucleotide. 
     
     
         7 . The isolated polynucleotide of  claim 5  wherein the sequence has at least 99% nucleic acid sequence identity to the native IBABP-L exon 1-3 polynucleotide. 
     
     
         8 . An isolated polynucleotide comprising at least 15 contiguous nucleotides of a native IBABP-L exon 1-3 polynucleotide, wherein the isolated polynucleotide hybridizes selectively to a native IBABP-L polynucleotide. 
     
     
         9 . The isolated polynucleotide of  claim 8  comprising at least 20 contiguous nucleotides from the native IBABP-L exon 1-3 polynucleotide. 
     
     
         10 . The isolated polynucleotide of  claim 8  comprising at least 30 contiguous nucleotides from the native IBABP-L exon 1-3 polynucleotide. 
     
     
         11 . The isolated polynucleotide of  claim 8  comprising the native IBABP-L exon 1-3 polynucleotide. 
     
     
         12 . The isolated polynucleotide of  claim 8  comprising a full-length protein-coding sequence of a native IBABP-L mRNA or cDNA. 
     
     
         13 . An isolated polynucleotide that (a) encodes a polypeptide of at least 11 amino acids, wherein the polypeptide comprises at least 4 contiguous amino acids of a native IBABP-L N-terminal polypeptide, and, (b) when introduced into a mammal, elicits an antibody that binds selectively to a native IBABP-L polypeptide. 
     
     
         14 . The isolated polynucleotide of  claim 13  wherein the polypeptide comprises at least 5 contiguous amino acids of the IBABP-L N-terminal polypeptide. 
     
     
         15 . The isolated polynucleotide of  claim 13  wherein the polypeptide comprises at least 6 contiguous amino acids of the IBABP-L N-terminal polypeptide. 
     
     
         16 . The isolated polynucleotide of  claim 13  wherein the polypeptide comprises at least 10 contiguous amino acids of the IBABP-L N-terminal polypeptide. 
     
     
         17 . The isolated polynucleotide of any of  claims 1 - 16  that encodes a polypeptide that binds bile acid. 
     
     
         18 . A cell comprising the isolated polynucleotide of any of  claims 1 - 16 . 
     
     
         19 . A vector comprising the isolated polynucleotide of any of  claims 1 - 16 . 
     
     
         20 . A cell comprising the vector of  claim 19 . 
     
     
         21 . An expression vector of  claim 19 . 
     
     
         22 . A cell comprising the expression vector of  claim 21 . 
     
     
         23 . A probe comprising the isolated polynucleotide of any of  claims 1 - 16 . 
     
     
         24 . A primer comprising the isolated polynucleotide of any of  claims 1 - 16 . 
     
     
         25 . A kit comprising: (a) a first primer comprising at least 15 contiguous nucleotides of a native IBABP-L exon 1-3 polynucleotide, wherein the first primer hybridizes selectively to a native IBABP-L polynucleotide; (b) a second primer comprising at least 15 contiguous nucleotides from the native IBABP-L polynucleotide; and (c) suitable-packaging enclosing the first primer and the second primer, wherein an amplification reaction performed using the first primer, the second primer, and a sample comprising an IBABP-L mRNA produces an amplification product that indicates the presence of the IBABP-L mRNA in the sample. 
     
     
         26 . The kit of  claim 25  wherein the second primer hybridizes selectively to the IBABP-L exon 1-3 polynucleotide. 
     
     
         27 . The kit of  claim 25  further comprising a third primer comprising at least 15 contiguous nucleotides from a native IBABP polynucleotide, wherein the sample further comprises an IBABP mRNA and an amplification reaction performed using the first primer, the second primer, the third primer, and the sample produces a first amplification product that indicates the presence of the IBABP-L mRNA in the sample and a second amplification product that indicates the presence of the IBABP mRNA in the sample. 
     
     
         28 . The kit of  claim 27  wherein the third primer comprises at least 15 contiguous nucleotides from a native IBABP exon 4a polynucleotide. 
     
     
         29 . The kit of  claim 25  further comprising a third primer comprising at least 15 contiguous nucleotides from a native IBABP polynucleotide, and a fourth primer comprising at least 15 contiguous nucleotides from the native IBABP polynucleotide, wherein the sample further comprises an IBABP mRNA, and a polymerase chain reaction performed using the first primer, the second primer, the third primer, the fourth primer and the sample produces a first amplification product that indicates the presence of the IBABP-L mRNA in the sample and a second amplification product that indicates the presence of the IBABP mRNA in the sample. 
     
     
         30 . The kit of  claim 29  wherein the third primer or the fourth primer comprises at least 15 contiguous nucleotides from a native IBABP exon 4a polynucleotide. 
     
     
         31 . An isolated polypeptide comprising a sequence that has at least 90% amino acid sequence identity to a native IBABP N-terminal polypeptide, wherein introduction of the isolated polypeptide into a mammal elicits production of an antibody that selectively binds to IBABP-L. 
     
     
         32 . The isolated polypeptide of  claim 31  wherein the sequence has at least 95% identity to the native IBABP-L N-terminal polypeptide. 
     
     
         33 . The isolated polypeptide of  claim 31  wherein the sequence has at least 99% identity to the native IBABP-L N-terminal polypeptide. 
     
     
         34 . The isolated polypeptide of  claim 31  wherein the sequence is at least 15 amino acids in length. 
     
     
         35 . The isolated polypeptide of  claim 31  wherein the sequence is at least 20 amino acids in length. 
     
     
         36 . The isolated polypeptide of  claim 31  wherein the sequence is at least 30 amino acids in length. 
     
     
         37 . The isolated polypeptide of  claim 31  wherein the sequence is at least 40 amino acids in length. 
     
     
         38 . An isolated polypeptide comprising a sequence of at least 11 contiguous amino acids from an IBABP-L N-terminal polypeptide, wherein introduction of the isolated polypeptide into a mammal elicits production of an antibody that selectively binds to IBABP-L. 
     
     
         39 . The isolated polypeptide of  claim 38  comprising a sequence of at least 12 contiguous amino acids from the IBABP-L N-terminal polypeptide. 
     
     
         40 . The isolated polypeptide of  claim 38  comprising a sequence of at least 13 contiguous amino acids from the IBABP-L N-terminal polypeptide. 
     
     
         41 . The isolated polypeptide of  claim 38  comprising a sequence of at least 15 contiguous amino acids from the IBABP-L N-terminal polypeptide. 
     
     
         42 . The isolated polypeptide of  claim 38  comprising a sequence of at least 20 contiguous amino acid residues from the IBABP-L N-terminal polypeptide. 
     
     
         43 . The isolated polypeptide of  claim 38  comprising a sequence of at least 30 contiguous amino acid residues from the IBABP-L N-terminal polypeptide. 
     
     
         44 . The isolated polypeptide of  claim 38  comprising the IBABP-L N-terminal polypeptide. 
     
     
         45 . The isolated polypeptide of  claim 38  comprising the native IBABP-L polypeptide. 
     
     
         46 . The isolated polypeptide of  claim 38  that binds bile acid. 
     
     
         47 . An isolated polypeptide at least 11 amino acids in length comprising at least 4 contiguous amino acids of a native IBABP-L N-terminal polypeptide, wherein introduction of the isolated polypeptide into a mammal elicits production of an antibody that binds selectively to the native IBABP-L polypeptide. 
     
     
         48 . The isolated polypeptide of  claim 47  comprising at least 5 contiguous amino acids of the native IBABP-L N-terminal polypeptide. 
     
     
         49 . The isolated polypeptide of  claim 47  comprising at least 6 contiguous amino acids of the native IBABP-L N-terminal polypeptide. 
     
     
         50 . The isolated polypeptide of  claim 47  comprising at least 10 contiguous amino acids of the native IBABP-L N-terminal polypeptide. 
     
     
         51 . The isolated polypeptide of  claim 47  that is at least 12 amino acids in length. 
     
     
         52 . The isolated polypeptide of  claim 47  that is at least 15 amino acids in length. 
     
     
         53 . A pharmaceutical composition comprising an amount of an IBABP-L polypeptide that is effective to treat or prevent colorectal cancer and a pharmaceutically acceptable carrier. 
     
     
         54 . A method of making a medicament for treating a patient with colorectal cancer or at risk for developing colorectal cancer, the method comprising formulating the medicament with a pharmaceutically effective amount of an IBABP-L polypeptide. 
     
     
         55 . A method of treating or preventing colorectal cancer comprising administering to a patient in need thereof a composition comprising an effective amount of an IBABP-L polypeptide. 
     
     
         56 . An antibody that binds selectively to a native IBABP-L polypeptide. 
     
     
         57 . A monoclonal antibody of  claim 56 . 
     
     
         58 . A polyclonal antibody of  claim 56 . 
     
     
         59 . A chimeric antibody of  claim 56 . 
     
     
         60 . A humanized antibody of  claim 56 . 
     
     
         61 . A single-chain antibody of  claim 56 . 
     
     
         62 . A fragment antibody of  claim 56 . 
     
     
         63 . A method of making an antibody that binds selectively to a native IBABP-L polypeptide comprising introducing into a mammal (a) an expression vector comprising a polynucleotide of any of  claims 1 - 16 , or (b) an isolated polypeptide of any of  claims 31 - 52 , thereby eliciting production of an the antibody. 
     
     
         64 . A method of detecting the presence of an IBABP-L polypeptide in a sample comprising the IBABP-L polypeptide, the method comprising contacting the sample with an antibody that binds selectively to the IBABP-L polypeptide, and detecting binding of the antibody to the IBABP-L polypeptide. 
     
     
         65 . The method of  claim 64  wherein the antibody is a monoclonal antibody. 
     
     
         66 . The method of  claim 64  comprising performing an ELISA assay. 
     
     
         67 . The method of  claim 64  comprising performing a bio-barcode assay. 
     
     
         68 . The method of  claim 64  comprising measuring IBABP-L polypeptide in the sample by measuring binding of the antibody to the IBABP-L polypeptide. 
     
     
         69 . A method of determining a ratio of IBABP-L polypeptide to IBABP polypeptide in a sample comprising the IBABP-L polypeptide and the IBABP polypeptide, the method comprising: (a) contacting the sample with a first antibody that binds selectively to the IBABP-L polypeptide, and measuring binding of the first antibody to the IBABP-L polypeptide in the sample; (b) contacting the sample with a second antibody that binds selectively to IBABP polypeptide and to IBABP-L polypeptide, and measuring binding of the second antibody to the IBABP polypeptide and the IBABP-L polypeptide in the sample; and (c) calculating the ratio of IBABP-L polypeptide to IBABP polypeptide in the sample. 
     
     
         70 . The method of  claim 69  wherein steps (a) and (b) are performed in a single reaction. 
     
     
         71 . The method of  claim 69  wherein the first antibody and second antibody are monoclonal antibodies. 
     
     
         72 . The method of  claim 69  comprising performing an ELISA assay. 
     
     
         73 . The method of  claim 69  comprising performing a bio-barcode assay. 
     
     
         74 . A method of detecting the presence of an IBABP-L polynucleotide in a sample comprising the IBABP-L polynucleotide, the method comprising contacting the sample with a probe or primer comprising a polynucleotide sequence that binds selectively to the IBABP-L polynucleotide and detecting binding of the probe or primer to the IBABP-L polynucleotide. 
     
     
         75 . The method of  claim 74  wherein the IBABP-L polynucleotide is an mRNA. 
     
     
         76 . The method of  claim 74  comprising contacting the sample with a first primer that comprises the polynucleotide sequence that hybridizes selectively to the IBABP-L polynucleotide and a second primer comprising a polynucleotide sequence that hybridizes to the IBABP-L polynucleotide, performing an amplification reaction, and detecting an amplification product that indicates the presence of the IBABP-L polynucleotide in the sample. 
     
     
         77 . The method of  claim 76  wherein the amplification reaction is a PCR reaction. 
     
     
         78 . The method of  claim 77  wherein the amplification reaction is a quantitative PCR reaction. 
     
     
         79 . The method of  claim 77  wherein the amplification reaction is an RT-PCR reaction. 
     
     
         80 . The method of  claim 74  comprising performing a bio-barcode assay. 
     
     
         81 . The method of  claim 74  comprising measuring the IBABP-L polynucleotide in the sample by measuring the binding of the probe or primer to the IBABP-L mRNA. 
     
     
         82 . A method of determining a ratio of IBABP-L polynucleotide to IBABP polynucleotide in a sample that comprises IBABP-L polynucleotide and IBABP polynucleotide, the method comprising (a) contacting the sample with a first probe that hybridizes selectively to IBABP-L polynucleotide; (b) measuring hybridization of the first probe to the IBABP-L polynucleotide in the sample, (c) contacting the sample with a second probe that hybridizes selectively to IBABP polynucleotide and IBABP-L polynucleotide; (d) measuring hybridization of the second probe to the IBABP polynucleotide and the IBABP-L polynucleotide in the sample; and (e) calculating the ratio of IBABP-L polynucleotide to IBABP polynucleotide in the sample. 
     
     
         83 . The method of  claim 82  wherein the IBABP-L polynucleotide and the IBABP polynucleotide are mRNAs. 
     
     
         84 . The method of  claim 82  comprising (1) contacting the sample with at least one primer that hybridizes selectively to the IBABP-L polynucleotide and performing a first amplification reaction to produce a first amplification product that indicates the presence of the IBABP-L polynucleotide in the sample; (b) contacting the sample with at least one primer that hybridizes selectively to IBABP polynucleotide and the IBABP-L polynucleotide and performing a second amplification reaction to produce a second amplification product that indicates the presence of IBABP polynucleotide and the IBABP-L polynucleotide in the sample; (c) measuring the first amplification product and the second amplification product; and (d) calculating the ratio of IBABP-L polynucleotide to IBABP polynucleotide in the sample. 
     
     
         85 . The method of  claim 83  wherein the steps of contacting and the steps of performing amplification reactions are performed in a single reaction. 
     
     
         86 . The method of  claim 85  comprising performing PCR. 
     
     
         87 . The method of  claim 86  comprising performing RT-PCR. 
     
     
         88 . The method of  claim 82  comprising performing a bio-barcode assay. 
     
     
         89 . A method of detecting a colorectal cancer in an individual comprising measuring IBABP-L polypeptide in a sample from the individual that comprises the IBABP-L polypeptide. 
     
     
         90 . A method of detecting a colorectal cancer in an individual comprising measuring a ratio of IBABP-L polypeptide to IBABP polypeptide in a sample from the individual that comprises the IBABP-L polypeptide and the IBABP polypeptide. 
     
     
         91 . A method of detecting a colorectal cancer in an individual comprising measuring IBABP-L mRNA in a sample from the individual that comprises the IBABP-L mRNA. 
     
     
         92 . A method of detecting a colorectal cancer in an individual comprising measuring a ratio of IBABP-L mRNA to IBABP mRNA in a sample from the individual that comprises the IBABP-L mRNA and the IBABP mRNA. 
     
     
         93 . A method of identifying an individual who is at increased risk to develop colorectal cancer comprising measuring IBABP-L polypeptide in a sample from the individual that comprises the IBABP-L polypeptide. 
     
     
         94 . A method of identifying an individual who is at increased risk to develop colorectal cancer comprising measuring a ratio of IBABP-L polypeptide to IBABP polypeptide in a sample from the individual that comprises the IBABP-L polypeptide and the IBABP polypeptide. 
     
     
         95 . A method of identifying an individual who is at increased risk to develop colorectal cancer comprising measuring IBABP-L mRNA in a sample from the individual that comprises the IBABP-L mRNA. 
     
     
         96 . A method of identifying an individual who is at increased risk to develop colorectal cancer comprising measuring a ratio of IBABP-L mRNA to IBABP mRNA in a sample from the individual that comprises the IBABP-L mRNA and the IBABP mRNA. 
     
     
         97 . A method of identifying an individual who is likely to respond to a particular therapy for colorectal cancer comprising measuring IBABP-L polypeptide in a sample from the individual that comprises the IBABP-L polypeptide. 
     
     
         98 . A method of identifying an individual who is likely to respond to a particular therapy for colorectal cancer comprising measuring a ratio of IBABP-L polypeptide to IBABP polypeptide in a sample from the individual that comprises the IBABP-L polypeptide and the IBABP polypeptide. 
     
     
         99 . A method of identifying an individual who is likely to respond to a particular therapy for colorectal cancer comprising measuring IBABP-L mRNA in a sample from the individual that comprises the IBABP-L mRNA. 
     
     
         100 . A method of identifying an individual who is likely to respond to a particular therapy for colorectal cancer comprising measuring a ratio of IBABP-L mRNA to IBABP mRNA in a sample from the individual that comprises the IBABP-L mRNA and the IBABP mRNA. 
     
     
         101 . A method of assessing the progress of a course of treatment for a patient suffering from colorectal cancer comprising (a) measuring IBABP-L polypeptide in a first sample from the patient taken at a first time point during the course of treatment, wherein the first sample comprises the IBABP-L polypeptide; (b) measuring IBABP-L polypeptide in a second sample from the patient taken at a second time point during the course of treatment, wherein the second sample comprises the IBABP-L polypeptide; and (c) comparing the measurements of IBABP-L polypeptide in the first sample and the second sample. 
     
     
         102 . A method of assessing the progress of a course of treatment for a patient suffering from colorectal cancer comprising (a) measuring a ratio of IBABP-L polypeptide to IBABP polypeptide in a first sample from the patient taken at a first time point during the course of treatment, wherein the first sample comprises the IBABP-L polypeptide and the IBABP polypeptide; (b) measuring a ratio of IBABP-L polypeptide to IBABP polypeptide in a second sample from the patient taken at a second time point during the course of treatment, wherein the second sample comprises the IBABP-L polypeptide and the IBABP polypeptide; and (c) comparing the measurements of the ratio of IBABP-L polypeptide to IBABP polypeptide in the first sample and the second sample. 
     
     
         103 . A method of assessing the progress of a course of treatment for a patient suffering from colorectal cancer comprising (a) measuring IBABP-L mRNA in a first sample from the patient taken at a first time point during the course of treatment, wherein the first sample comprises the IBABP-L mRNA; (b) measuring IBABP-L mRNA in a second sample from the patient taken at a second time point during the course of treatment, wherein the second sample comprises the IBABP-L mRNA; and (c) comparing the measurements of IBABP-L mRNA in the first sample and the second sample. 
     
     
         104 . A method of assessing the progress of a course of treatment for a patient suffering from colorectal cancer comprising (a) measuring a ratio of IBABP-L mRNA to IBABP mRNA in a first sample from the patient taken at a first time point during the course of treatment, wherein the first sample comprises the IBABP-L mRNA and the IBABP mRNA; (b) measuring a ratio of IBABP-L mRNA to IBABP mRNA in a second sample from the patient taken at a second time point during the course of treatment, wherein the second sample comprises the IBABP-L mRNA and the IBABP mRNA; and (c) comparing the measurements of the ratio of IBABP-L mRNA to IBABP mRNA in the first sample and the second sample. 
     
     
         105 . The method of any of  claims 64 - 104  wherein the sample is a cell. 
     
     
         106 . The method of any of  claims 64 - 104  wherein the sample is a tissue sample. 
     
     
         107 . The method of any of  claims 64 - 104  wherein the sample is a gastrointestinal tissue sample. 
     
     
         108 . The method of any of  claims 64 - 104  wherein the sample is a fecal sample. 
     
     
         109 . The method of any of  claims 64 - 104  wherein the sample is a blood sample. 
     
     
         110 . An automated method of any of  claims 64 - 104 .