Novel method of diagnosing, monitoring, and staging colon cancer

Abstract

The present invention provides a new method for detecting, diagnosing, monitoring, staging, and prognosticating colon cancer.

Description

FIELD OF THE INVENTION

[0001] This invention relates, in part, to newly developed assays for detecting, diagnosing, monitoring, staging, and prognosticating cancers, particularly colon cancer.

BACKGROUND OF THE INVENTION

[0002] Colon cancer is the second most frequently diagnosed malignancy in the United States. Cancer of the gastrointestinal tract, especially colon cancer, is a highly treatable and often a curable disease when localized to the bowel. However, currently colon cancer is the second most common cause of cancer death. Surgery is the primary treatment and results in cure in approximately 50% of patients. Recurrence following surgery is a major problem and often is the ultimate cause of death. The prognosis of colon cancer is clearly related to the degree of penetration of the tumor through the bowel wall and the presence or absence of nodal involvement. These two characteristics form the basis for all staging systems developed for this disease. Bowel obstruction and bowel perforation are indicators of poor prognosis. Elevated pretreatment serum levels of carcinoembryonic antigen (CEA) and carbohydrate antigen 19-9 (CA 19-9) also have negative prognostic significance.

[0003] Because of the frequency of the disease (approximately 160,000 new cases of colon cancer per year), the identification of high-risk groups, the demonstrated slow growth of primary lesions, the better survival of early-stage lesions, and the relative simplicity and accuracy of screening tests, screening for colon cancer should be a part of routine care for all adults starting at age 50, especially those with first-degree relatives with colorectal cancer.

[0004] Procedures used for detecting, diagnosing, monitoring, staging, and prognosticating colon cancer are of critical importance to the outcome of the patient. For example, patients diagnosed with early colon cancer generally have a much greater five-year survival rate as compared to the survival rate for patients diagnosed with distant metastasized colon cancer. Treatment decisions are usually made in reference to the older Dukes or the Modified Astler-Coller (MAC) classification schema for staging. However, new diagnostic methods which are more sensitive and specific for detecting early colon cancer are clearly needed.

[0005] Further, colon cancer patients must be closely monitored following initial therapy and during adjuvant therapy to determine response to therapy and to detect persistent or recurrent disease of metastasis. Thus, there is clearly a need for a colon cancer marker which is more sensitive and specific in detecting colon cancer recurrence.

[0006] Another important step in managing colon cancer is to determine the stage of the patient's disease. Stage determination has potential prognostic value and provides criteria for designing optimal therapy. Currently, pathological staging of colon cancer is preferable over clinical staging as pathological staging provides a more accurate prognosis. However, clinical staging would be preferred were the method of clinical staging at least as accurate as pathological staging because it does not depend on an invasive procedure to obtain tissue for pathological evaluation. Staging of colon cancer would be improved by detecting new markers in cells, tissues, or bodily fluids which could differentiate between different stages of invasion.

[0007] In the present invention, methods are provided for detecting, diagnosing, monitoring, staging, and prognosticating colon cancers, particularly colon, stomach, and small intestine cancer, via nine (9) Colon Specific Genes (CSGs). The nine CSGs refer, among other things, to native proteins expressed by the genes comprising the polynucleotide sequences of any of SEQ ID NO: 1, 2, 3, 4, 5, 6, 7, 8 or 9. In the alternative, what is meant by the nine CSGs as used herein, means the native mRNAs encoded by the genes comprising any of the polynucleotide sequences of SEQ ID NO: 1, 2, 3, 4, 5, 6, 7, 8 or 9 or levels of the genes comprising any of the polynucleotide sequences of SEQ ID NO: 1, 2, 3, 4, 5, 6, 7, 8 or 9.

[0008] Other objects, features, advantages and aspects of the present invention will become apparent to those of skill in the art from the following description. It should be understood, however, that the following description and the specific examples, while indicating preferred embodiments of the invention, are given by way of illustration only. Various changes and modifications within the spirit and scope of the disclosed invention will become readily apparent to those skilled in the art from reading the following description and from reading the other parts of the present disclosure.

SUMMARY OF THE INVENTION

[0009] Toward these ends, and others, it is an object of the present invention to provide a method for diagnosing the presence of colon cancer in a patient which comprises measuring levels of CSG in a sample of cells, tissue or bodily fluid from the patient and comparing the measured levels of CSG with levels of CSG in preferably the same cells, tissue, or bodily fluid type of a control, wherein an increase in the measured CSG levels in the patient versus levels of CSG in the control is associated with colon cancer.

[0010] Another object of the present invention is to provide a method of diagnosing metastatic colon cancer in a patient which comprises measuring CSG levels in a sample of cells, tissue, or bodily fluid from the patient and comparing the measured CSG levels with levels of CSG in preferably the same cells, tissue, or bodily fluid type of a control, wherein an increase in measured CSG levels in the patient versus levels of CSG in the control is associated with a cancer which has metastasized.

[0011] Another object of the present invention is to provide a method of staging colon cancer in a patient which comprises identifying a patient having colon cancer, measuring levels of CSG in a sample of cells, tissues, or bodily fluid obtained from the patient, and comparing the measured CSG levels with levels of CSG in preferably the same cells, tissue or bodily fluid type of a control. An increase in measured CSG levels in the patient versus CSG levels in the control can be associated with a cancer which is progressing while a decrease or equivalent level of CSG measured in the patient versus the control can be associated with a cancer which is regressing or in remission.

[0012] Another object of the present invention is to provide a method of monitoring colon cancer in a patient for the onset of metastasis. The method comprises identifying a patient having colon cancer that is not known to have metastasized, periodically measuring levels of CSG in a sample of cells, tissues, or bodily fluid obtained from the patient, and comparing the measured CSG levels with levels of CSG in preferably the same cells, tissue, or bodily fluid type of a control, wherein an increase in measured CSG levels versus control CSG levels is associated with a cancer which has metastasized.

[0013] Yet another object of the present invention is to provide a method of monitoring the change in stage of colon cancer in a patient which comprises identifying a patient having colon cancer, periodically measuring levels of CSG in a sample of cells, tissue, or bodily fluid obtained from the patient, and comparing the measured CSG levels with levels of CSG in preferably the same cells, tissues, or bodily fluid type of a control wherein an increase in measured CSG levels versus the control CSG levels is associated with a cancer which is progressing and a decrease in the measured CSG levels versus the control CSG levels is associated with a cancer which is regressing or in remission.

[0014] Other objects, features, advantages and aspects of the present invention will become apparent to those of skill in the art from the following description. It should be understood, however, that the following description and the specific examples, while indicating preferred embodiments of the invention, are given by way of illustration only. Various changes and modifications within the spirit and scope of the disclosed invention will become readily apparent to those skilled in the art from reading the following description and from reading the other parts of the present disclosure.

DESCRIPTION OF THE INVENTION

[0015] The present invention relates to diagnostic assays and methods, both quantitative and qualitative for detecting, diagnosing, monitoring, staging, and prognosticating cancers by comparing levels of CSG with those of CSG in a normal human control. What is meant by "levels of CSG" as used herein, means levels of the native protein expressed by the genes comprising the polynucleotide sequence of any of SEQ ID NO: 1, 2, 3, 4, 5, 6, 7, 8 or 9. In the alternative, what is meant by "levels of CSG" as used herein, means levels of the native mRNA encoded by any of the genes comprising any of the polynucleotide sequences of SEQ ID NO: 1, 2, 3, 4, 5, 6, 7, 8 or 9 or levels of the gene comprising any of the polynucleotide sequence of SEQ ID NO: 1, 2, 3, 4, 5, 6, 7, 8 or 9. Such levels are preferably measured in at least one of, cells, tissues and/or bodily fluids, including determination of normal and abnormal levels. Thus, for instance, a diagnostic assay in accordance with the invention for diagnosing over-expression of any one of the CSG proteins compared to normal control bodily fluids, cells, or tissue samples may be used to diagnose the presence of cancers, including colon cancer. Any of the nine CSGs may be measured alone in the methods of the invention, or all together or any combination of the nine.

[0016] By "control" it is meant a human patient without cancer and/or non cancerous samples from the patient, also referred to herein as a normal human control; in the methods for diagnosing or monitoring for metastasis, control may also include samples from a human patient that is determined by reliable methods to have colon cancer which has not metastasized.

[0017] All the methods of the present invention may optionally include measuring the levels of other cancer markers as well as CSG. Other cancer markers, in addition to CSG, useful in the present invention will depend on the cancer being tested and are known to those of skill in the art.

[0018] Diagnostic Assays

[0019] The present invention provides methods for diagnosing the presence of colon cancer by analyzing for changes in levels of CSG in cells, tissues or bodily fluids compared with levels of CSG in cells, tissues or bodily fluids of preferably the same type from a normal human control, wherein an increase in levels of CSG in the patient versus the normal human control is associated with the presence of colon cancer. Without limiting the instant invention, typically, for a quantitative diagnostic assay a positive result indicating the patient being tested has cancer is one in which cells, tissues, or bodily fluid levels of the cancer marker, such as CSG, are at least two times higher, and most preferably are at least five times higher, than in preferably the same cells, tissues, or bodily fluid of a normal human control.

[0020] The present invention also provides a method of diagnosing metastatic colon cancer in a patient having colon cancer which has not yet metastasized for the onset of metastasis. In the method of the present invention, a human cancer patient suspected of having colon cancer which may have metastasized (but which was not previously known to have metastasized) is identified. This is accomplished by a variety of means known to those of skill in the art. For example, in the case of colon cancer, patients are typically diagnosed with colon cancer following traditional detection methods.

[0021] In the present invention, determining the presence of CSG level in cells, tissues, or bodily fluid, is particularly useful for discriminating between colon cancer which has not metastasized and colon cancer which has metastasized. Existing techniques have difficulty discriminating between colon cancer which has metastasized and colon cancer which has not metastasized and proper treatment selection is often dependent upon such knowledge.

[0022] In the present invention, the cancer marker levels measured in such cells, tissues, or bodily fluid is CSG, and are compared with levels of CSG in preferably the same cells, tissue, or bodily fluid type of a normal human control. That is, if the cancer marker being observed is just CSG in serum, this level is preferably compared with the level of CSG in serum of a normal human patient. An increase in the CSG in the patient versus the normal human control is associated with colon cancer which has metastasized.

[0023] Without limiting the instant invention, typically, for a quantitative diagnostic assay a positive result indicating the cancer in the patient being tested or monitored has metastasized is one in which cells, tissues, or bodily fluid levels of the cancer marker, such as CSG, are at least two times higher, and most preferable are at least five times higher, than in preferably the same cells, tissues, or bodily fluid of a normal patient.

[0024] Normal human control as used herein includes a human patient without cancer and/or non cancerous samples from the patient; in the methods for diagnosing or monitoring for metastasis, normal human control may also include samples from a human patient that is determined by reliable methods to have colon cancer which has not metastasized.

[0025] Staging

[0026] The invention also provides a method of staging colon cancer in a human patient.

[0027] The method comprises identifying a human patient having such cancer; analyzing a sample of cells, tissues, or bodily fluid from such patient for CSG. Then, the method compares CSG levels in such cells, tissues, or bodily fluid with levels of CSG in preferably the same cells, tissues, or bodily fluid type of a normal human control sample, wherein an increase in CSG levels in the patient versus the normal human control is associated with a cancer which is progressing and a decrease in the levels of CSG is associated with a cancer which is regressing or in remission.

[0028] Monitoring

[0029] Further provided is a method of monitoring colon cancer in a human having such cancer for the onset of metastasis. The method comprises identifying a human patient having such cancer that is not known to have metastasized; periodically analyzing a sample of cells, tissues, or bodily fluid from such patient for CSG; comparing the CSG levels in such cells, tissue, or bodily fluid with levels of CSG in preferably the same cells, tissues, or bodily fluid type of a normal human control sample, wherein an increase in CSG levels in the patient versus the normal human control is associated with a cancer which has metastasized.

[0030] Further provided by this inventions is a method of monitoring the change in stage of colon cancer in a human having such cancer. The method comprises identifying a human patient having such cancer; periodically analyzing a sample of cells, tissues, or bodily fluid from such patient for CSG; comparing the CSG levels in such cells, tissue, or bodily fluid with levels of CSG in preferably the same cells, tissues, or bodily fluid type of a normal human control sample, wherein an increase in CSG levels in the patient versus the normal human control is associated with a cancer which is progressing in stage and a decrease in the levels of CSG is associated with a cancer which is regressing in stage or in remission.

[0031] Monitoring such patient for onset of metastasis is periodic and preferably done on a quarterly basis. However, this may be more or less frequent depending on the cancer, the particular patient, and the stage of the cancer.

[0032] Assay Techniques

[0033] Assay techniques that can be used to determine levels of gene expression, such as CSG of the present invention, in a sample derived from a host are well-known to those of skill in the art. Such assay methods include radioimmunoassays, reverse transcriptase PCR (RT-PCR) assays, immunohistochemistry assays, in situ hybridization assays, competitive-binding assays, Western Blot analyses and ELISA assays. Among these, ELISAs are frequently preferred to diagnose a gene's expressed protein in biological fluids. An ELISA assay initially comprises preparing an antibody, if not readily available from a commercial source, specific to CSG, preferably a monoclonal antibody. In addition a reporter antibody generally is prepared which binds specifically to CSG. The reporter antibody is attached to a detectable reagent such as radioactive, fluorescent or enzymatic reagent, for example horseradish peroxidase enzyme or alkaline phosphatase.

[0034] To carry out the ELISA, antibody specific to CSG is incubated on a solid support, e.g., a polystyrene dish, that binds the antibody. Any free protein binding sites on the dish are then covered by incubating with a non-specific protein such as bovine serum albumin. Next, the sample to be analyzed is incubated in the dish, during which time CSG binds to the specific antibody attached to the polystyrene dish. Unbound sample is washed out with buffer. A reporter antibody specifically directed to CSG and linked to horseradish peroxidase is placed in the dish resulting in binding of the reporter antibody to any monoclonal antibody bound to CSG. Unattached reporter antibody is then washed out. Reagents for peroxidase activity, including a calorimetric substrate are then added to the dish. Immobilized peroxidase, linked to CSG antibodies, produces a colored reaction product. The amount of color developed in a given time period is proportional to the amount of CSG protein present in the sample. Quantitative results typically are obtained by reference to a standard curve.

[0035] A competition assay may be employed wherein antibodies specific to CSG attached to a solid support and labeled CSG and a sample derived from the host are passed over the solid support and the amount of label detected attached to the solid support can be correlated to a quantity of CSG in the sample. Nucleic acid methods may be used to detect CSG mRNA as a marker for colon cancer. Polymerase chain reaction (PCR) and other nucleic acid methods, such as ligase chain reaction (LCR) and nucleic acid sequence based amplification (NASABA), can be used to detect malignant cells for diagnosis and monitoring of various malignancies. For example, reverse-transcriptase PCR (RT-PCR) is a powerful technique which can be used to detect the presence of a specific mRNA population in a complex mixture of thousands of other mRNA species. In RT-PCR, an mRNA species is first reverse transcribed to complementary DNA (cDNA) with use of the enzyme reverse transcriptase; the cDNA is then amplified as in a standard PCR reaction. RT-PCR can thus reveal by amplification the presence of a single species of mRNA. Accordingly, if the mRNA is highly specific for the cell that produces it, RT-PCR can be used to identify the presence of a specific type of cell.

[0036] Hybridization to clones or oligonucleotides arrayed on a solid support (i.e., gridding) can be used to both detect the expression of and quantitate the level of expression of that gene. In this approach, a cDNA encoding the CSG gene is fixed to a substrate. The substrate may be of any suitable type including but not limited to glass, nitrocellulose, nylon or plastic. At least a portion of the DNA encoding the CSG gene is attached to the substrate and then incubated with the analyte, which may be RNA or a complementary DNA (cDNA) copy of the RNA, isolated from the tissue of interest. Hybridization between the substrate bound DNA and the analyte can be detected and quantitated by several means including but not limited to radioactive labeling or fluorescence labeling of the analyte or a secondary molecule designed to detect the hybrid. Quantitation of the level of gene expression can be done by comparison of the intensity of the signal from the analyte compared with that determined from known standards. The standards can be obtained by in vitro transcription of the target gene, quantitating the yield, and then using that material to generate a standard curve. The above tests can be carried out on samples derived from a variety of patients' cells, bodily fluids and/or tissue extracts (homogenates or solubilized tissue) such as from tissue biopsy and autopsy material. Bodily fluids useful in the present invention include blood, urine, saliva, or any other bodily secretion or derivative thereof. Blood can include whole blood, plasma, serum, or any derivative of blood.

EXAMPLES

[0037] The present invention is further described by the following examples. These examples are provided solely to illustrate the invention by reference to specific embodiments. These exemplifications, while illustrating certain specific aspects of the invention, do not portray the limitations or circumscribe the scope of the disclosed invention.

Example 1

CSGs

[0038] Searches were carried out and CSGs identified using the following Search Tools as part of the LIFESEQ.RTM. database available from Incyte Pharmaceuticals, Palo Alto, Calif.:

[0039] 1. Library Comparison (compares one library to one other library) allows the identification of clones expressed in tumor and absent or expressed at a lower level in normal tissue.

[0040] 2. Subsetting is similar to library comparison but allows the identification of clones expressed in a pool of libraries and absent or expressed at a lower level in a second pool of libraries.

[0041] 3. Transcript Imaging lists all of the clones in a single library or a pool of libraries based on abundance. Individual clones can then be examined using Electronic Northerns to determine the tissue sources of their component ESTs.

[0042] 4. Protein Function: Incyte has identified subsets of ESTs with a potential protein function based on homologies to known proteins. Some examples in this database include Transcription Factors and Proteases. We identified some leads by searching in this database for clones whose component ESTs showed disease specificity.

[0043] Electronic subtractions, transcript imaging and protein function searches were used to identify clones, whose component ESTs were exclusively or more frequently found in libraries from specific tumors. Individual candidate clones were examined in detail by checking where each EST originated.

1TABLE 1 CSGs SEQ ID NO: Clone ID # Gene ID # 1 238330 242807 Transcript Imaging 2 1285234 239588 Subsetting 3 1341701 29634 Transcript Imaging 4 816257 233421 Subsetting 5 775133 245080 Subsetting 6 1335450 245811 Subsetting 7 2348122 233711 Transcript Imaging 8 3228674 230273 Subsetting 9 1632174 229022 Transcript Imaging

[0044] The following example was carried out using standard techniques, which are well known and routine to those of skill in the art, except where otherwise described in detail. Routine molecular biology techniques of the following example can be carried out as described in standard laboratory manuals, such as Sambrook et al., MOLECULAR CLONING: A LABORATORY MANUAL, 2nd Ed.; Cold Spring Harbor Laboratory Press, Cold Spring Harbor, N.Y. (1989).

Example 2

Relative Quantitation of CSG Gene Expression

[0045] Real-Time quantitative PCR with fluorescent Taqman probes is a quantitation detection system utilizing the 5'-3' nuclease activity of Taq DNA polymerase. The method uses an internal fluorescent oligonucleotide probe (Taqman) labeled with a 5' reporter dye and a downstream, 3' quencher dye. During PCR, the 5'-3' nuclease activity of Taq DNA polymerase releases the reporter, whose fluorescence can then be detected by the laser detector of the Model 7700 Sequence Detection System (PE Applied Biosystems, Foster City, Calif., USA).

[0046] Amplification of an endogenous control is used to standardize the amount of sample RNA added to the reaction and normalize for Reverse Transcriptase (RT) efficiency. Either cyclophilin, glyceraldehyde-3-phosphate dehydrogenase (GAPDH) or 18S ribosomal RNA (rRNA) is used as this endogenous control. To calculate relative quantitation between all the samples studied, the target RNA levels for one sample are used as the basis for comparative results (calibrator). Quantitation relative to the "calibrator" can be obtained using the standard curve method or the comparative method (User Bulletin #2: ABI PRISM 7700 Sequence Detection System).

[0047] To evaluate the tissue distribution, and the level of CSGs in normal and tumor tissue, total RNA was extracted from normal tissues, tumor tissues, and from tumors and the corresponding matched normal tissues. Subsequently, first strand cDNA was prepared with reverse transcriptase and the polymerase chain reaction was done using primers and Taqman probe specific to the CSG. The results were analyzed using the ABI PRISM 7700 Sequence Detector. The absolute numbers are relative levels of expression of the CSG compared to the calibrator.

Comparative Examples

[0048] Similar mRNA expression analysis for genes coding for the diagnostic markers PSA (Prostate Specific Antigen) and PLA2 (Phospholipase A2) was performed for comparison. PSA is currently the only cancer screening marker available in clinical laboratories. When the panel of normal pooled tissues was analyzed, PSA was expressed at very high levels in prostate, with a very low expression in breast and testis. After analysis of more than 55 matching samples from 14 different tissues, the data corroborated the tissue specificity seen with normal tissue samples. PSA expression was compared in cancer and normal adjacent tissue for 12 matching samples of prostate tissue. The relative levels of PSA were higher in 10 cancer samples (83%). Clinical data recently obtained support the utilization of PLA2 as a staging marker for late stages of prostate cancer. mRNA expression data described herein showed overexpression of the mRNA in 8 out of the 12 prostate matching samples analyzed (66%). PLA2 had high levels of mRNA expression in small intestine, prostate, liver, and pancreas.

[0049] Measurement of SEQ ID NO:3; Clone ID 1341701; Gene ID 29634 (Cln106)

[0050] Absolute numbers are depicted in Table 2 as relative levels of expression of Cln106 (SEQ ID NO:3) in 12 normal different tissues. All the values are compared to normal testis (calibrator). These RNA samples are commercially available pools, originated by pooling samples of a particular tissue from different individuals.

2TABLE 2 Relative levels of Cln106 Expression in Pooled Samples Tissue NORMAL Colon-Ascending 110 Endometrium 0 Kidney 0 Liver 0 Ovary 0 Pancreas 0 Prostate 16 Small Intestine 0 Spleen 0 Stomach 0 Testis 1 Uterus 0

[0051] The relative levels of expression in Table 2 show for the CSG Cln106 (SEQ ID NO:3), mRNA expression is more than 6 fold higher in the pool of normal ascending colon (110) compared with prostate (16). Testis, the calibrator, with a relative expression level of 1, is the only other tissue expressing the mRNA for Cln106 (SEQ ID NO:3). These results demonstrate that mRNA expression of this CSG is highly specific for colon.

[0052] The absolute numbers in Table 2 were obtained analyzing pools of samples of a particular tissue from different individuals. They can not be compared to the absolute numbers originated from RNA obtained from tissue samples of a single individual in Table 3.

[0053] The absolute numbers in Table 3 are relative levels of expression of Cln106 (SEQ ID NO:3) in 57 pairs of matching samples. All the values are compared to normal testis (calibrator). A matching pair is formed by mRNA from the cancer sample for a particular tissue and mRNA from the normal adjacent sample for that same tissue from the same individual.

3TABLE 3 Relative levels of Cln106 Expression in Individual Samples Matching Normal Sample ID Tissue Cancer Adjacent Sto AC93 Stomach 1 4 96 Sto AC99 Stomach 2 0.4 0.5 Sml 21XA Small Intestine 1 0 0 Sml H89 Small Intestine 2 0.93 1.28 Cln B56 Colon-Cecum(A)1 317 101 Cln AS45 Colon-Ascending(A)2 316.3 146.5 Cln CM67 Colon-Cecum(B)3 481.0 217.5 Cln AS67 Colon-Ascending(B)4 858.1 220.6 Cln AS43 Colon-Ascending(C)5 1370 98 Cln AS46 Colon-Ascending(C)6 3051 375 Cln AS98 Colon-Ascending(C)7 26 42 Cln AS89 Colon-Ascending(D)8 524.6 11.0 Cln TX01 Colon-Transverse(B)9 2886.3 1992.0 Cln TX89 Colon-Transverse(B)10 146.0 35.9 Cln TX67 Colon-Transverse(C)11 2.9 421.7 Cln MT38 Colon-Splenic 1681 187 Flexture (M) 12 Cln SG89 Colon-Sigmoid(B)13 1063.8 31.1 Cln SG67 Colon-Sigmoid(C)14 8.5 9.4 Cln SG33 Colon-Sigmoid(C)15 264 549 Cln SG45 Colon-Sigmoid(D)16 580.0 114.6 Cln B34 Colon-Rectosigmoid(A)17 97 244 Cln CXGA Colon-Rectum(A)18 45.1 273.4 Cln RC67 Colon-Rectum(B)19 2.7 20.0 Cln C9XR Colon-Rectosigmoid(C)20 609 460 Cln RS45 Colon-Rectosigmoid(C)21 472.8 144.0 Cln RC01 Colon-Rectum(C)22 568 129 Cln RC89 Colon-Rectum(D)23 4.6 322.91 Bld 46XK Bladder 1 0.2 0 Bld 66X Bladder 2 1 1 Bld 32XK Bladder 3 0.0 0.0 Kid 126XD Kidney 1 0 0 Kid 12XD Kidney 2 0 0 Kid 5XD Kidney 3 0.0 1.0 Kid 6XD Kidney 4 0.0 0.0 Kid 106XD Kidney 5 0.4 0.0 Liv 42X Liver 1 0.0 0.0 Liv 15XA Liver 2 0.0 0.0 Liv 94XA Liver 3 0.0 0.0 Lng AC69 Lung 1 2 0 Lng BR94 Lung 2 0 0 Lng 47XQ Lung 3 0 0 Mam 59X Mammary Gland 1 0 0 Mam B011X Mammary Gland 2 0 0 Mam A06X Mammary Gland 3 0 0 Ovr 103X Ovary 1 0.04 2.08 Ovr 130X Ovary 2 0.1 2.76 Pan 71XL Pancreas 1 4.08 0.1 Pan 82XP Pancreas 2 0 0 Pro 12B Prostate 1 0.3 0 Pro 23B Prostate 2 3 4 Pro 13XB Prostate 3 2 7 Pro 34B Prostate 4 0.54 4.01 Pro 20XB Prostate 5 4.8 4.3 Pro 65XB Prostate 6 0.7 1.3 Tst 39X Testis 1 2.78 0 End 8XA Endometrium 1 0 0.2 Utr 85XU Uterus 1 1.26 0 0 = Negative

[0054] When matching samples were analyzed, the higher levels of expression were in the colon, showing a high degree of tissue-specificity for this tissue. These results confirm the tissue specificity results obtained with the panel of normal pooled samples (Table 2). Furthermore, the level of mRNA expression in cancer samples and the isogenic normal adjacent tissue from the same individual were compared. This comparison provides an indication of specificity for the cancer stage (e.g. higher levels of mRNA expression in the cancer sample compared to the normal adjacent). Table 3 shows overexpression of Cln106 (SEQ ID NO:3) in 15 colon cancer tissues compared with their respective normal adjacent (colon samples #1, 2, 3, 4, 5, 6, 8, 9, 10, 12, 13, 16, 20, 21, and

[0055] 22). There is overexpression in the cancer tissue for 65% of the colon matching samples tested (total of 23 colon matching samples). The matching sample Pan 71XL is a secondary cancer in pancreas, the primary cancer in that individual was a duodenal cancer.

[0056] Altogether, the high level of tissue specificity, plus the mRNA overexpression in 65% of the colon matching samples tested are demonstrative of CSG Cln106 (SEQ ID NO:3) being a diagnostic marker for colon cancer.

[0057] Measurement of SEQ ID NO:4; Clone ID 816257; Gene ID 406452 (Cln107)

[0058] Absolute numbers as depicted in Table 4 are relative levels of expression of CSG Cln107 (SEQ ID NO:4) in 12 normal different tissues. All the values are compared to normal small intestine (calibrator). These RNA samples are commercially available pools, originated by pooling samples of a particular tissue from different individuals.

4TABLE 4 Relative levels of Cln107 Expression in Pooled Samples Tissue NORMAL Colon-Ascending 3.2 Endometrium 0 Kidney 0.2 Liver 0 Ovary 0 Pancreas 0 Prostate 0.1 Small Intestine 1 Spleen 0 Stomach 0.3 Testis 0 Uterus 0

[0059] The relative levels of expression in Table 4 show that mRNA expression of the CSG Cln107 (SEQ ID NO:4) is more than 10 fold higher in the pool of normal ascending colon (3.2), five fold higher in small intestine (1), and 1.5 fold higher in stomach (0.3), compared with the next higher expressor (0.2 for kidney). Seven of the pooled tissues samples analyzed were negative and prostate showed a relative expression of 0.1 for Cln107 (SEQ ID NO:4). These results demonstrate that Cln107 mRNA expression is highly specific for colon, small intestine, and in a lower degree for stomach.

[0060] The absolute numbers in Table 4 were obtained analyzing pools of samples of a particular tissue from different individuals. They can not be compared to the absolute numbers originated from RNA obtained from tissue samples of a single individual in Table 5.

[0061] The absolute numbers in Table 5 are relative levels of expression of Cln107 (SEQ ID NO:4) in 57 pairs of matching samples. All the values are compared to normal small intestine (calibrator). A matching pair is formed by mRNA from the cancer sample for a particular tissue and mRNA from the normal adjacent sample for that same tissue from the same individual.

5TABLE 5 Relative levels of Cln107 Expression in Individual Samples Matching Normal Sample ID Tissue Cancer Adjacent Sto AC93 Stomach 1 8.9 13.4 Sto AC99 Stomach 2 6.0 0.9 Sml 21XA Small Intestine 1 1.07 1.42 Sml H89 Small Intestine 2 0.97 4.13 Cln B56 Colon-Cecum(A)1 2 16 Cln AS45 Colon-Ascending(A)2 0.7 2.1 Cln CM67 Colon-Cecum(B)3 1.6 2.1 Cln AS67 Colon-Ascending(B)4 1.2 6.2 Cln AS43 Colon-Ascending(C)5 13.5 0.5 Cln AS46 Colon-Ascending(C)6 9.7 23.6 Cln AS98 Colon-Ascending(C)7 28.1 1.4 Cln AS89 Colon-Ascending(D)8 0.9 3.1 Cln TX01 Colon-Transverse(B)9 3.0 10.6 Cln TX89 Colon-Transverse(B)10 4.5 0.6 Cln TX67 Colon-Transverse(C)11 3.6 3.4 Cln MT38 Colon-Splenic 4.0 2.6 Flexture(M)12 Cln SG89 Colon-Sigmoid(B)13 4.7 0.9 Cln SG67 Colon-Sigmoid(C)14 1.0 1.3 Cln SG33 Colon-Sigmoid(C)15 14.2 7.6 Cln SG45 Colon-Sigmoid(D)16 4.8 6.0 Cln B34 Colon-Rectosigmoid(A)17 3 2 Cln CXGA Colon-Rectum(A)18 4.4 1.9 Cln RC67 Colon-Rectum(B)19 0.1 0.4 Cln C9XR Colon-Rectosigmoid(C)20 5 3 Cln RS45 Colon-Rectosigmoid(C)21 11.4 4.6 Cln RC01 Colon-Rectum(C)22 1.8 2.3 Cln RC89 Colon-Rectum(D)23 0.1 5.35 Bld 46XK Bladder 1 0.2 0 Bld 66X Bladder 2 1 1 Bld 32XK Bladder 3 0.1 0.1 Kid 126XD Kidney 1 0 0.02 Kid 12XD Kidney 2 0.1 0.2 Kid 5XD Kidney 3 0.3 0.0 Kid 6XD Kidney 4 0.1 0.1 Kid 106XD Kidney 5 0.0 0.1 Liv 42X Liver 1 7.9 0.002 Liv 15XA Liver 2 0.0 0.0 Liv 94XA Liver 3 0.0 0.0 Lng AC69 Lung 1 1.6 0.2 Lng BR94 Lung 2 0.4 0 Lng 47XQ Lung 3 0.78 0.2 Mam 59X Mammary Gland 1 0.05 0.3 Mam B011X Mammary Gland 2 0.01 0.004 Mam A06X Mammary Gland 3 0.22 0 Ovr 103X Ovary 1 0.01 0.01 Ovr 130X Ovary 2 0.09 0.1 Pan 71XL Pancreas 1 2.51 2.81 Pan 82XP Pancreas 2 0 0.62 Pro 12B Prostate 1 0.3 0.1 Pro 23B Prostate 2 0.3 0.2 Pro 13XB Prostate 3 0 0 Pro 34B Prostate 4 0.04 0.22 Pro 20XB Prostate 5 0.4 0.1 Pro 65XB Prostate 6 0.0 0.1 Tst 39X Testis 1 0.02 0.01 End 8XA Endometrium 1 0.01 0.5 Utr 85XU Uterus 1 0.03 0 0 = Negative

[0062] When matching samples were analyzed, the higher levels of expression were in colon, stomach, and small intestine, showing a high degree of tissue specificity for colon tissues. These results confirm the tissue specificity results obtained with normal pooled samples (Table 4). Furthermore, the level of mRNA expression in cancer samples and the isogenic normal adjacent tissue from the same individual were compared. This comparison provides an indication of specificity for the cancer stage (e.g. higher levels of mRNA expression in the cancer sample compared to the normal adjacent). Table 5 shows overexpression of Cln107 (SEQ ID NO:4) in 11 colon cancer tissues compared with their respective normal adjacent (colon samples #5, 7, 10, 11, 12, 13, 15, 17, 18, 20, and 21). There is overexpression in the cancer tissue for 48% of the colon matching samples tested (total of 23 colon matching samples). The matching sample Pan 71XL is a secondary cancer in pancreas, the primary cancer in that individual was a duodenal cancer.

[0063] Altogether, the high level of tissue specificity, plus the mRNA overexpression in almost half of the colon, stomach, and small intestine matching samples tested are demonstrative of CSG Cln107 (SEQ ID NO:4) being a diagnostic marker for colon cancer.

[0064] Measurement of SEQ ID NO: 5; Clone ID 775133; Gene ID 24508 (Cln108)

[0065] The absolute numbers shown in Table 6 are relative levels of expression of CSG Cln108 (SEQ ID NO:5) in 12 normal different tissues. All the values are compared to normal small intestine (calibrator). These RNA samples are commercially available pools, originated by pooling samples of a particular tissue from different individuals.

6TABLE 6 Relative levels of Cln108 Expression in Pooled Samples Tissue NORMAL Colon-Ascending 2846.5 Endometrium 1 Kidney 5.5 Liver 18.7 Ovary 3.4 Pancreas 198.1 Prostate 1024 Small Intestine 810.8 Spleen 32.2 Stomach 9981.2 Testis 0 Uterus 294.1

[0066] The relative levels of expression in Table 6 show that mRNA expression of CSG Cln108 (SEQ ID NO:5) is more than 10 fold higher in the pool of normal ascending colon (2846.5) and almost ten fold higher in stomach (9981.2), compared to the expression level in any other tissue analyzed. These results demonstrate that mRNA expression of this CSG is also highly specific for colon and stomach.

[0067] The absolute numbers in Table 6 were obtained analyzing pools of samples of a particular tissue from different individuals. They can not be compared to the absolute numbers originated from RNA obtained from tissue samples of a single individual in Table 7.

[0068] The absolute numbers depicted in Table 7 are relative levels of expression of Cln108 (SEQ ID NO:5) in 57 pairs of matching samples. All the values are compared to normal small intestine (calibrator). A matching pair is formed by mRNA from the cancer sample for a particular tissue and mRNA from the normal adjacent sample for that same tissue from the same individual.

7TABLE 7 Relative levels of Cln108 Expression in Individual Samples Matching Normal Sample ID Tissue Cancer Adjacent Sto AC93 Stomach 1 28696 34842 Sto AC99 Stomach 2 21523 30862 Sml 21XA Small Intestine 1 2944 964.4 Sml H89 Small Intestine 2 244.5 3513.2 Cln B56 Colon-Cecum(A)1 27242 24637 Cln AS45 Colon-Ascending(A)2 5827.0 8771.0 Cln CM67 Colon-Cecum(B)3 4251.0 4684.0 Cln AS67 Colon-Ascending(B)4 564.0 1949.0 Cln AS43 Colon-Ascending(C)5 50310 10949 Cln AS46 Colon-Ascending(C)6 246044 120073 Cln AS98 Colon-Ascending(C)7 40442 17482 Cln AS89 Colon-Ascending(D)8 5730.0 1581.0 Cln TX01 Colon-Transverse(B)9 22281.0 114784.0 Cln TX89 Colon-Transverse(B)10 11026.0 1639.0 Cln TX67 Colon-Transverse(C)11 17004.0 11654.0 Cln MT38 Colon-Splenic 77589 31620 Flexture(M)12 Cln SG89 Colon-Sigmoid(B)13 140339.0 49617.0 Cln SG67 Colon-Sigmoid(C)14 4951.0 7905.0 Cln SG33 Colon-Sigmoid(C)15 60875 120490 Cln SG45 Colon-Sigmoid(D)16 30437.0 47267.0 Cln B34 Colon-Rectosigmoid(A)17 5848 5861 Cln CXGA Colon-Rectum(A)18 13877.0 9787.0 Cln RC67 Colon-Rectum(B)19 1703.0 26589.0 Cln C9XR Colon-Rectosigmoid(C)20 2458 19071 Cln RS45 Colon-Rectosigmoid(C)21 95523 61939 Cln RC01 Colon-Rectum(C)22 98891.0 80047.0 Cln RC89 Colon-Rectum(D)23 17.0 1775 Bld 46XK Bladder 1 0 8 Bld 66X Bladder 2 397 44 Bld 32XK Bladder 3 0.0 16.0 Kid 126XD Kidney 1 32 22 Kid 12XD Kidney 2 6 0 Kid 106XD Kidney 3 4.0 33.0 Liv 42X Liver 1 4783 0 Liv 15XA Liver 2 4.0 10.0 Liv 94XA Liver 3 159.0 21.0 Lng AC69 Lung 1 222 295 Lng BR94 Lung 2 112 0 Lng 47XQ Lung 3 30 69 Lng AC66 Lung 4 29 137 Mam 59X Mammary Gland 1 56 0 Mam B011X Mammary Gland 2 54 31 Mam A06X Mammary Gland 3 12 0 Ovr 103X Ovary 1 37 0 Pan 71XL Pancreas 1 13203 4163 Pan 82XP Pancreas 2 39.1 0 Pro 12B Prostate 1 386 88 Pro 23B Prostate 2 250 23 Pro 13XB Prostate 3 92 731 Pro 34B Prostate 4 33.3 265.7 Pro 20XB Prostate 5 454.6 1908.9 Pro 65XB Prostate 6 733.5 922.0 End 8XA Endometrium 1 5 92 Utr 85XU Uterus 1 98.9 21.8 Utr 23XU Uterus 2 35.3 0 Utr 135XO Uterus 3 39.2 43.8 Utr 141XO Uterus 4 212.1 55.9 0 = Negative

[0069] When matching samples were analyzed, the higher levels of expression were in colon and stomach, showing a high degree of tissue specificity for these two tissues. These results confirm the tissue specificity results obtained with normal pooled samples (Table 6). Furthermore, the level of mRNA expression in cancer samples and the isogenic normal adjacent tissue from the same individual were compared. This comparison provides an indication of specificity for the cancer stage (e.g. higher levels of mRNA expression in the cancer sample compared to the normal adjacent). Table 7 shows overexpression of CSG Cln108 (SEQ ID NO:5) in 13 colon cancer tissues compared with their respective normal adjacent (colon samples #1, 5, 6, 7, 8, 9, 10, 11, 12, 13, 18, 21, and 22). There is overexpression in the cancer tissue for 56% of the colon matching samples tested (total of 23 colon matching samples). The matching sample Pan 71XL is a secondary cancer in pancreas, the primary cancer in that individual was a duodenal cancer.

[0070] Altogether, the high level of tissue specificity, plus the mRNA overexpression in more than half of the colon, stomach, and small intestine matching samples tested are demonstrative of this CSG, Cln108 (SEQ ID NO:5), also being a diagnostic marker for colon cancer.

[0071] Measurement of SEQ ID NO:7; Clone ID 2348122; Gene ID 23371 (Cln109)

[0072] The absolute numbers depicted in Table 8 are relative levels of expression of CSG Cln109 (SEQ ID NO:7) in 12 normal different tissues. All the values are compared to normal ovary (calibrator). These RNA samples are commercially available pools, originated by pooling samples of a particular tissue from different individuals.

8TABLE 8 Relative levels of Cln109 Expression in Pooled Samples Tissue NORMAL Colon-Ascending 28.8 Endometrium 0.45 Kidney 0.41 Liver 0.72 Ovary 0.07 Pancreas 82.8 Prostate 124.3 Small Intestine 626.4 Spleen 1.2 Stomach 12.05 Testis 1.51 Uterus 52.99

[0073] The relative levels of expression in Table 8 show that mRNA expression of CSG Cln109 (SEQ ID NO:7), is more than 5 fold higher in the pool of normal small intestine (626.4) compared to the expression level in any other tissue analyzed. These results demonstrate that Cln109 (SEQ ID NO:7) mRNA expression is highly specific for small intestine.

[0074] The absolute numbers in Table 8 were obtained analyzing pools of samples of a particular tissue from different individuals. They can not be compared to the absolute numbers originated from RNA obtained from tissue samples of a single individual in Table 9.

[0075] The absolute numbers depicted in Table 9 are relative levels of expression of Cln109 (SEQ ID NO:7) in 53 pairs of matching samples. All the values are compared to normal ovary (calibrator). A matching pair is formed by mRNA from the cancer sample for a particular tissue and mRNA from the normal adjacent sample for that same tissue from the same individual.

9TABLE 9 Relative levels of Cln109 Expression in Individual Samples Matching Normal Sample ID Tissue Cancer Adjacent Sto AC93 Stomach 1 2574 1310 Sto AC99 Stomach 2 4153 5 Sml 21XA Small Intestine 1 2667 13663.8 Sml H89 Small Intestine 2 57.8 904.29 Cln B56 Colon-Cecum(A)1 6794 299 Cln AS45 Colon-Ascending(A)2 814.6 105.8 Cln CM67 Colon-Cecum(B)3 294.6 36.1 Cln AS67 Colon-Ascending(B)4 2.2 26.3 Cln AS43 Colon-Ascending(C)5 111 377 Cln AS46 Colon-Ascending(C)6 1180 352 Cln AS98 Colon-Ascending(C)7 1075 92 Cln AS89 Colon-Ascending(D)8 14022.7 87.5 Cln TX01 Colon-Transverse(B)9 1027.6 282.1 Cln TX89 Colon-Transverse(B)10 2.5 23.7 Cln TX67 Colon-Transverse(C)11 0.1 72.3 Cln MT38 Colon-Splenic 372 88 Flexture(M)12 Cln SG89 Colon-Sigmoid(B)13 179.2 33.4 Cln SG67 Colon-Sigmoid(C)14 85.0 94.7 Cln SG33 Colon-Sigmoid(C)15 5461 377 Cln SG45 Colon-Sigmoid(D)16 762.7 15.9 Cln B34 Colon-Rectosigmoid(A)17 460 1 Cln RC67 Colon-Rectum(B)18 64.5 136.2 Cln C9XR Colon-Rectosigmoid(C)19 441 34 Cln RS45 Colon-Rectosigmoid(C)20 1931 195 Cln RC01 Colon-Rectum(C)21 72.8 19.1 Cln RC89 Colon-Rectum(D)22 4.8 90.2 Bld 46XK Bladder 1 4 3 Bld 66X Bladder 2 1 0 Bld 32XK Bladder 3 0.1 307.6 Kid 126XD Kidney 1 0 2 Kid 12XD Kidney 2 3 16 Kid 5XD Kidney 3 0.0 0.3 Kid 6XD Kidney 4 18.5 1.2 Liv 42X Liver 1 21 0.03 Liv 15XA Liver 2 0.5 0.4 Liv 94XA Liver 3 0.4 0.0 Lng AC69 Lung 1 0.1 0 Lng BR94 Lung 2 3 0 Lng 60XL Lung 3 0.1 0 Mam 59X Mammary Gland 1 0 4 Mam B011X Mammary Gland 2 8 13 Mam A06X Mammary Gland 3 4.7 9.6 Pan 71XL Pancreas 1 8902.5 1428.2 Pan 82XP Pancreas 2 0.2 9.3 Pro 12B Prostate 1 9 20 Pro 23B Prostate 2 191 88 Pro 13XB Prostate 3 12 460 Pro 34B Prostate 4 3.2 80.4 Tst 39X Testis 1 29.9 0 End 8XA Endometrium 1 0.3 21 Utr 85XU Uterus 1 244.7 592.2 Ovr 63A Ovary 1 11.4 0 Ovr A1C Ovary 2 68.4 0 0 = Negative

[0076] When matching samples were analyzed, the higher levels of expression were in small intestine, colon and stomach, showing a high degree of tissue specificity for these three colon tissues. These results confirm the tissue specificity results obtained with normal pooled samples for small intestine (Table 8). Furthermore, the level of mRNA expression in cancer samples and the isogenic normal adjacent tissue from the same individual were compared. This comparison provides an indication of specificity for the cancer stage (e.g. higher levels of mRNA expression in the cancer sample compared to the normal adjacent). Table 9 shows overexpression of CSG, Cln109 (SEQ ID NO:7) in 15 colon cancer tissues compared with their respective normal adjacent (colon samples #1, 2, 3, 6, 7, 8, 9, 12, 13, 15, 16, 17, 19, 20, and 21). There is overexpression in the cancer tissue for 68% of the colon matching samples tested (total of 22 colon matching samples). The matching sample Pan 71XL is a secondary cancer in pancreas, the primary cancer in that individual was a duodenal cancer.

[0077] Altogether, the high level of tissue specificity, plus the mRNA overexpression in more than half of the colon, stomach, and small intestine matching samples tested are demonstrative of CSG Cln109 (SEQ ID NO:7) being a diagnostic marker for colon cancer. The amino acid sequence encoded by the open reading frame of Cln109 is depicted in SEQ ID NO:10.

Sequence CWU 1

1

10 1 487 DNA Homo sapien 1 tctgcatctg gccctcccag tgcacctgtt caatcccagc ycctccctga cctgtacaaa 60 tacacctgag gaccggctcg agcccagact tcctgcccct gctctgcact ctcaggtatt 120 ccctgctctt actccaaaaa gatggaccca ggtccgaagg ggcactgcca ctgtgggggg 180 catggccatc ctccaggtca ctgcgggcga acccctggcc atggcccagg gccctgcggg 240 ccaccccctg gccatggccc agggccctgc gggcaacccc ctggccatgg cccagggccc 300 tgcgggcctc cccctggcca tggcccaggt cacccacccc ctggtccaca tcactgagga 360 agtagaagaa aacaggacac aagatggcaa gcctgagaga attgcccagc tgacctggaa 420 tgaggcctaa accacaatct tctcttccta ataaacagcc tcctagaggc cacattctat 480 tctttaa 487 2 739 DNA Homo sapiens misc_feature (693)..(693) n=a, c, g or t 2 tctgaaactg tcagttccac cagcactgct tggatactgg taagtttcca gggggctgct 60 ttgcatctga aactgtcagc cccagaatgt tgacagtcgc tctcctagcc cttctctgtg 120 cctcagcctc tggcaatgcc attcaggcca ggtcttcctc ctatagtgga gagtatggaa 180 gtggtggtgg aaagcgattc tctcattctg gcaaccagtt ggacggcccc atcaccgccc 240 tccgggtccg agtcaacaca tactacatcg taggtcttca ggtgcgctat ggcaaggtgt 300 ggagcgacta tgtgggtggt cgcaacggag acctggagga gatctttctg caccctgggg 360 aatcagtgat ccaggtttct gggaagtaca agtggtacct gaagaagctg gtatttgtga 420 cagacaaggg ccgctatctg tcttttggga aagacagtgg cacaagtttc aatgccgtcc 480 ccttgcaccc caacaccgtg ctccgcttca tcagtggccg gtctggttct ctcatcgatg 540 ccattggcct gcactgggat gtttacccca ctagctgcag cagatgctga gcctcctctc 600 cttggcaggg gcactgtgat gaggagtaag aactccctta tcactaaccc ccatccaaat 660 ggctcaataa aaaaatatgg ttaaggctaa aanaaaanng gannnaanan nnnnnntnca 720 aannnnantt cncctgnta 739 3 428 DNA Homo sapiens misc_feature (392)..(392) n=a, c, g or t 3 aattgtccgg ggtcaaacag aggagagcat gaatgagagt catcctcgca agtgtgcaga 60 gtcttttgag atgtgggatg atcgtgactc ccactgtagg cgccctaagt ttgaagggca 120 tccccctgag tcttggaagt ggatccttgc accggtcatt ctttatatct gtgaaaggat 180 cctccggttt taccgctccc agcagaaggt tgtgattacc aaggttgtta tgcacccatc 240 caaagttttg gaattgcaga tgaacaagcg tggcttcagc atggaagtgg ggcagtatat 300 ctttgttaat tgcccctcaa tctctctcct gggaatggca tccttttact ttgacctctg 360 ctccagagga agatttcttc ttcattcata tncgagcagc aggggacttg acagaaaatc 420 tataaggg 428 4 1347 DNA Homo sapiens 4 ggaaaacccc tgagcacaaa gcaagaggca tcgaagcccc ctcggggatg cccgcaagcc 60 aacaggggtg tcgtgcggtg ggagtacttc cgcctgcgtc ctctgcggtt cagggcccca 120 gacgagcccc agcaggccca agtcccccat gtctggggct gggaggtggc tggggcccct 180 gcactgaggc tgcagaagtc ccagtcatct gatctgctgg aaagggagag ggagagtgtc 240 ctgcgccggg agcaagaggt ggcagaggag cggagaaatg ctctcttccc agaggtcttc 300 tccccaacgc cagatgagaa ctctgaccag aactccagga gctcctccca ggcatccggc 360 atcacgggca gttactcggt gtctgagtct cccttcttca gccccatcca cctacactca 420 aacgtggcgt ggacagtgga agatccagtg gacagtgctc ctcccgggca gagaaagaag 480 gagcaatggt acgctggcat caacccctcg gacggtatca actcagaggt cctggaagcc 540 atacgggtga cccgtcacaa gaacgccatg gcagagcgct gggaatcccg catctacgcc 600 agtgaggagg atgactgagc ctcgggatgg ggcgcccacc ccctgccctg ccctgaccct 660 cgtgggaact gccaagacca tcgccaagcc cccaccctag gaaatgggtc ctaggtccag 720 gatccaagaa ccacagctca tctgccaaca atcccaccat gggcacattt gggactgttg 780 ggtttttcgt ttccgtttct atcttccttt agaaatgttt ctgcctttgg ggtctaaagc 840 ttttggggat gaaatgggga cccctgctga ttctttctgc ttctaagact ttgccaaatg 900 ccctgggtct aagaaagaaa gagacccgct cctccacttt caggtgtaat ttgcttccgc 960 tagtctgagg gcagagggac cggtcaaaga gggtggcaca gatcgcagca ccttgagggg 1020 ctgcgggtct gagggaggag acactcagct cctccctctg agaagtccca agctgagagg 1080 ggagacctgc ccctttccaa ccctgggaaa ccatccagtc tgagggagga ggccaaactc 1140 ccagtgctgg gggtccctgt gcagccctca aacccttcac cttggtgcac ccagccacac 1200 ctggtggaca caaagctctc acatcgatag gatcccatga ggatggtccc cttcacctgg 1260 gagaaaagtg acccagttta ggagctggag gggggtcttt gtcccccacc cccaaactgc 1320 cctgaaataa acctggagtg agctgcc 1347 5 1249 DNA Homo sapiens misc_feature (1034)..(1056) n=a, c, g or t 5 ggcagagcct gcgcagggca ggagcagctg gcccactggc ggcccgcaac actccgtctc 60 accctctggg cccactgcat ctagaggagg gccgtctgtg aggccactac ccctccagca 120 actgggaggt gggactgtca gaagctggcc cagggtggtg gtcagctggg tcagggacct 180 aacggcacct ggctgggacc acctcgcctt ctccatcgaa gcaggggaag tgggagcctc 240 gagccctcgg gtggaagctg accccaagcc acccttcacc tggacaggat gagagtgtca 300 ggtgtgcttc gcctcctggc cctcatcttt gccatagtca cgacatggat gtttattcga 360 agctacatga gcttcagcat gaaaaccatc cgtctgccac gctggctggc ctcgcccacc 420 aaggagatcc aggttaaaaa gtacaagtgt ggcctcatca agccctgccc agccaactac 480 tttgcgttta aaatctgcag tggggccgcc aacgtcgtgg gccctactat gtgctttgaa 540 gaccgcatga tcatgagtcc tgtgaaaaac aatgtgggca gaggcctaaa catcgccctg 600 gtgaatggaa ccacgggagc tgtgctggga cagaagtcat ttgacatgta ctctggagat 660 gttatgcacc tagtgaaatt ccttaaagaa attccggggg gtgcactggt gctggtggcc 720 tcctacgacg atccagggac caaaatgaac gatgaaagca ggaaactctt ctctgacttg 780 gggagttcct acgcaaaaca actgggcttc cgggacagct gggtcttcat aggagccaaa 840 gacctcaggg gtaaaagccc ctttgagcag ttcttaaaga acagcccaga cacaaacaaa 900 tacgagggat ggccagagct gctggagatg gagggctgca tgcccccgaa gccattttag 960 ggtggctgtg gctcttcctc agccaggggc ctgaagaagc tcctgcctga cttaggagtc 1020 agagcccggc aggnnnnnnn nnnnnnnnnn nnnnnntgct gcgtggaagg tgctgcaggt 1080 ccttgcacgc tgtgtcgcgc ctctcctcct cggaaacaga accctcccac agcacatcct 1140 acccggaaga ccagcctcag agggtccttc tggaaccagc tgtctgtgga gagaatgggg 1200 tgctttcgtc agggactgct gacggctggt cctgaggaag gacaaactg 1249 6 1220 DNA Homo sapiens 6 gctttctgca cctcattcca catcaggagc gtttttggag aaagctgcac tctgttgagc 60 tccagggcgc agtggaggga gggagtgaag gagctctctg tacccaagga aagtgcagct 120 gagactcaga caagattaca atgaaccaac tcagcttcct gctgtttctc atagcgacca 180 ccagaggatg gagtacagat gaggctaata cttacttcaa ggaatggacc tgttcttcgt 240 ctccatctct gcccagaagc tgcaaggaaa tcaaagacga atgtcctagt gcatttgatg 300 gcctgtattt tctccgcact gagaatggtg ttatctacca gaccttctgt gacatgacct 360 ctgggggtgg cggctggacc ctggtggcca gcgtgcacga gaatgacatg cgtgggaagt 420 gcacggtggg cgatcgctgg tccagtcagc agggcagcaa agcagtctac ccagaggggg 480 acggcaactg ggccaactac aacacctttg gatctgcaga ggcggccacg agcgatgact 540 acaagaaccc tggctactac gacatccagg ccaaggacct gggcatctgg cacgtgccca 600 ataagtcccc catgcagcac tggagaaaca gctccctgct gaggtaccgc acggacactg 660 gcttcctcca gacactggga cataatctgt ttggcatcta ccagaaatat ccagtgaaat 720 atggagaagg aaagtgttgg actgacaacg gcccggtgat ccctgtggtc tatgattttg 780 gcgacgccca gaaaacagca tcttattact caccctatgg ccagcgggaa ttcactgcgg 840 gatttgttca gttcagggta tttaataacg agagagcagc caacgccttg tgtgctggaa 900 tgagggtcac cggatgtaac actgagcacc actgcattgg tggaggagga tactttccag 960 aggccagtcc ccagcagtgt ggagattttt ctggttttga ttggagtgga tatggaactc 1020 atgttggtta cagcagcagc cgtgagataa ctgaggcagc tgtgcttcta ttctatcgtt 1080 gagagttttg tgggagggaa cccagacctc tcctcccaac catgagatcc caaggatgga 1140 gaacaactta cccagtagct agaatgttaa tggcagaaga gaaaacaata aatcatattg 1200 actcaaaaaa aaaaaaaaag 1220 7 2796 DNA Homo sapiens 7 cggctcgagg gacaggatga ggcccggcct ctcatttctc ctagcccttc tgttcttcct 60 tggccaagct gcaggggatt tgggggatgt gggacctcca attcccagcc ccggcttcag 120 ccctttccca ggtgttgact ccagctccag cttcagctcc agctccaggt cgggctccag 180 ctccagccgc agcttaggca gcggaggttc tgtgtcccag ttgttttcca atttcaccgg 240 ctccgtggat gaccgtggga cctgccagtg ctctgtttcc ctgccagaca ccacctttcc 300 cgtggacaga gtggaacgct tggaattcac agctcatgtt ctttctcaga agtttgagaa 360 agaactttcc aaagtgaggg aatatgtcca attaattagt gtgtatgaaa agaaactgtt 420 aaacctaact gtccgaattg acatcatgga gaaggatacc atttcttaca ctgaactgga 480 cttcgagctg atcaaggtag aagtgaagga gatggaaaaa ctggtcatac agctgaagga 540 gagttttggt ggaagctcag aaattgttga ccagctggag gtggagataa gaaatatgac 600 tctcttggta gagaagcttg agacactaga caaaaacaat gtccttgcca ttcgccgaga 660 aatcgtggct ctgaagacca agctgaaaga gtgtgaggcc tctaaagatc aaaacacccc 720 tgtcgtccac cctcctccca ctccagggag ctgtggtcat ggtggtgtgg tgaacatcag 780 caaaccgtct gtggttcagc tcaactggag agggttttct tatctatatg gtgcttgggg 840 tagggattac tctccccagc atccaaacaa aggactgtat tgggtggcgc cattgaatac 900 agatgggaga ctgttggagt attatagact gtacaacaca ctggatgatt tgctattgta 960 tataaatgct cgagagttgc ggatcaccta tggccaaggt agtggtacag cagtttacaa 1020 caacaacatg tacgtcaaca tgtacaacac cgggaatatt gccagagtta acctgaccac 1080 caacacgatt gctgtgactc aaactctccc taatgctgcc tataataacc gcttttcata 1140 tgctaatgtt gcttggcaag atattgactt tgctgtggat gagaatggat tgtgggttat 1200 ttattcaact gaagccagca ctggtaacat ggtgattagt aaactcaatg acaccacact 1260 tcaggtgcta aacacttggt ataccaagca gtataaacca tctgcttcta acgccttcat 1320 ggtatgtggg gttctgtatg ccacccgtac tatgaacacc agaacagaag agatttttta 1380 ctattatgac acaaacacag ggaaagaggg caaactagac attgtaatgc ataagatgca 1440 ggaaaaagtg cagagcatta actataaccc ttttgaccag aaactttatg tctataacga 1500 tggttacctt ctgaattatg atctttctgt cttgcagaag ccccagtaag ctgtttagga 1560 gttagggtga aagagaaaat gtttgttgaa aaaatagtct tctccactta cttagatatc 1620 tgcaggggtg tctaaaagtg tgttcatttt gcagcaatgt ttaggtgcat agttctacca 1680 cactagagat ctaggacatt tgtcttgatt tggtgagttc tcttgggaat catctgcctc 1740 ttcaggcgca ttttgcaata aagtctgtct agggtgggat tgtcagaggt ctaggggcac 1800 tgtgggccta gtgaagccta ctgtgaggag gcttcactag aagccttaaa ttaggaatta 1860 aggaacttaa aactcagtat ggcgtctagg gattctttgt acaggaaata ttgcccaatg 1920 actagtcctc atccatgtag caccactaat tcttccatgc ctggaagaaa cctggggact 1980 tagttaggta gattaatatc tggagctcct cgagggacca aatctccaac ttttttttcc 2040 cctcactaca cctggaatga tgctttgtat gtggcagata agtaaatttg gcatgcttat 2100 atattctaca tctgtaaagt gctgagtttt atggagagag gcctttttat gcattaaatt 2160 gtacatggca aataaatccc agaaggatct gtagatgagg cacctgcttt ttcttttctc 2220 tcattgtcca ccttactaaa agtcagtaga atcttctacc tcataacttc cttccaaagg 2280 cagctcagaa gattagaacc agacttacta accaattcca ccccccacca acccccttct 2340 actgcctact ttaaaaaaat taatagtttt ctatggaact gatctaagat tagaaaaatt 2400 aattttcttt aatttcatta tggactttta tttacatgac tctaagacta taagaaaatc 2460 tgatggcagt gacaaagtgc tagcatttat tgttatctaa taaagacctt ggagcatatg 2520 tgcaacttat gagtgtatca gttgttgcat gtaatttttg cctttgttta agcctggaac 2580 ttgtaagaaa atgaaaattt aatttttttt tctaggacga gctatagaaa agctattgag 2640 agtatctagt taatcagtgc agtagttgga aaccttgctg gtgtatgtga tgtgcttctg 2700 tgcttttgaa tgactttatc atctagtctt tgtctgtttt tcctttgatg ttcaagtcct 2760 agtctatagg attggcagtt taaatgcttt actccc 2796 8 2331 DNA Homo sapiens misc_feature (675)..(675) n=a, c, g or t 8 tttatcacgg gctcaactgc aacaaaacac ttccttgaca gctccacaaa ctcaggccac 60 agtgaggaat caacaatatt ccacagcagc ccagatgcaa gtggaacaac accctcatct 120 gcccactcca caacctcagg tcgtggagaa tctacaacct cacgcatcag tccaggctca 180 actgaaataa caacgttacc tggcagtacc acaacaccag gcctcagtga ggcatctacc 240 accttctaca gtagccccag atcaccagac caaacactct cacctgccag catgagaagc 300 tccagcatca gtggagaacc caccagcttg tatagccaag cagagtcaac acacacaaca 360 gcgttccctg ccagcaccac cacctcaggc ctcagtcagg aatcaacaac tttccacagt 420 aagccaggct caactgagac aacactgtcc cctggcagca tcacaacttc atcttttgct 480 caagaattta ccacccctca tagccaacca ggctcagctc tgtcaacagt gtcacctgcc 540 agcaccacag tgccaggcct tagtgaggaa tctaccacct tctacagcag cccaggctca 600 actgaaacca cagcgttttc tcacagcaac acaatgtcca ttcatagtca acaatctaca 660 cccttccctg acagnccagg cttcactcac acagtgttac ctgccaccct cacaaccaca 720 gacattggtc aggaatcaac agccttccac agcagctcag acgcaactgg aacaacaccc 780 ttacctgccc gctccacagc ctcagacctt gttggagaac ctacaacttt ctacatcagc 840 ccatccccta cttacacaac actctttcct gcgagttcca gcacatcagg cctcactgag 900 gaatctacca ccttccacac cagtccaagc ttcacttcta caattgtgtc tactgaaagc 960 ctggaaacct tagcaccagg gttgtgccag gaaggacaaa tttggaatgg aaaacaatgc 1020 gtctgtcccc aaggctacgt tggttaccag tgcttgtccc ctctggaatc cttccctgta 1080 gaaaccccgg aaaaactcaa cgccacttta ggtatgacag tgaaagtgac ttacagaaat 1140 ttcacagaaa agatgaatga cgcatcctcc caggaatacc agaacttcag taccctcttc 1200 aagaatcgga tggatgtcgt tttgaagggc gacaatcttc ctcagtatag aggggtgaac 1260 attcggagat tgctcaacgg tagcatcgtg gtcaagaacg atgtcatcct ggaggcagac 1320 tacactttag agtatgagga actgtttgaa aacctggcag agattgtaaa ggccaagatt 1380 atgaatgaaa ctagaacaac tcttcttgat cctgattcct gcagaaaggc catactgtgc 1440 tatagtgaag aggacacttt cgtggattca tcggtgactc cgggctttga cttccaggag 1500 caatgcaccc agaaggctgc cgaaggatat acccagttct actatgtgga tgtcttggat 1560 gggaagctgg cctgtgtgaa caagtgcacc aaaggaacga agtcgcaaat gaactgtaac 1620 ctgggcacat gtcagctgca acgcagtgga cccccgctgc ctgtgcccaa atacgaacac 1680 acactggtac tggggagaga cctgtgaatt caacatcgcc aagagcctcg tgtatgggat 1740 cgtgggggct gtgatggcgg tgctgctgct cgcattgatc atcctaatca tcttattcag 1800 cctatcccag agaaaacggc acagggaaca gtatgatgtg cctcaagagt ggcgaaagga 1860 aggcacccct ggcatcttcc agaagacggc catctgggaa gaccagaatc tgagggagag 1920 cagattcggc cttgagaacg cctacaacaa cttccggccc accctggaga ctgttgactc 1980 tggcacagag ctccacatcc agaggccgga gatggtagca tccactgtgt gagccaacgg 2040 gggcctccca ccctcatcta gctctgttca ggagagctgc aaacacagag cccaccacaa 2100 gcctccgggg cgggtcaaga ggagaccgaa gtcaggccct gaagccggtc ctgctctgag 2160 ctgacagact tggccagtcc cctgcctgtg ctcctgctgg ggaaggctgg gggctgtaag 2220 cctctccatc cgggagcttc cagactccca gaagcctcgg cacccctgtc tcctcctggg 2280 tggctcccca ctctggaatt tccctaccaa taaaagcaaa tctgaaagct c 2331 9 909 DNA Homo sapiens 9 gaggaggtgg gcgccaacag acaggcgatt aatgcggctc ttacccaggc aaccaggact 60 acagtataca ttgtggacat tcaggacata gattctgcag ctcgggcccg acctcactcc 120 tacctcgatg cctactttgt cttccccaat gggtcagccc tgacccttga tgagctgagt 180 gtgatgatcc ggaatgatca ggactcgctg acgcagctgc tgcagctggg gctggtggtg 240 ctgggctccc aggagagcca ggagtcagac ctgtcgaaac agctcatcag tgtcatcata 300 ggattgggag tggctttgct gctggtcctt gtgatcatga ccatggcctt cgtgtgtgtg 360 cggaagagct acaaccggaa gcttcaagct atgaaggctg ccaaggaggc caggaagaca 420 gcagcagggg tgatgccctc agcccctgcc atcccaggga ctaacatgta caacactgag 480 cgagccaacc ccatgctgaa cctccccaac aaagacctgg gcttggagta cctctctccc 540 tccaatgacc tggactctgt cagcgtcaac tccctggacg acaactctgt ggatgtggac 600 aagaacagtc aggaaatcaa ggagcacagg ccaccacaca caccaccaga gccagatcca 660 gagcccctga gcgtggtcct gttaggacgg caggcaggcg caagtggaca gctggagggg 720 ccatcctaca ccaacgctgg cctggacacc acggacctgt gacaggggcc cccactcttc 780 tggacccctt gaagaggccc taccacaccc taactgcacc tgtctccctg gagatgaaaa 840 tatatgacgc tgccctgcct cctgcttttg gccaatcacg gcagacaggg gttggggaaa 900 tattttatt 909 10 510 PRT Homo sapiens 10 Met Arg Pro Gly Leu Ser Phe Leu Leu Ala Leu Leu Phe Phe Leu Gly 1 5 10 15 Gln Ala Ala Gly Asp Leu Gly Asp Val Gly Pro Pro Ile Pro Ser Pro 20 25 30 Gly Phe Ser Pro Phe Pro Gly Val Asp Ser Ser Ser Ser Phe Ser Ser 35 40 45 Ser Ser Arg Ser Gly Ser Ser Ser Ser Arg Ser Leu Gly Ser Gly Gly 50 55 60 Ser Val Ser Gln Leu Phe Ser Asn Phe Thr Gly Ser Val Asp Asp Arg 65 70 75 80 Gly Thr Cys Gln Cys Ser Val Ser Leu Pro Asp Thr Thr Phe Pro Val 85 90 95 Asp Arg Val Glu Arg Leu Glu Phe Thr Ala His Val Leu Ser Gln Lys 100 105 110 Phe Glu Lys Glu Leu Ser Lys Val Arg Glu Tyr Val Gln Leu Ile Ser 115 120 125 Val Tyr Glu Lys Lys Leu Leu Asn Leu Thr Val Arg Ile Asp Ile Met 130 135 140 Glu Lys Asp Thr Ile Ser Tyr Thr Glu Leu Asp Phe Glu Leu Ile Lys 145 150 155 160 Val Glu Val Lys Glu Met Glu Lys Leu Val Ile Gln Leu Lys Glu Ser 165 170 175 Phe Gly Gly Ser Ser Glu Ile Val Asp Gln Leu Glu Val Glu Ile Arg 180 185 190 Asn Met Thr Leu Leu Val Glu Lys Leu Glu Thr Leu Asp Lys Asn Asn 195 200 205 Val Leu Ala Ile Arg Arg Glu Ile Val Ala Leu Lys Thr Lys Leu Lys 210 215 220 Glu Cys Glu Ala Ser Lys Asp Gln Asn Thr Pro Val Val His Pro Pro 225 230 235 240 Pro Thr Pro Gly Ser Cys Gly His Gly Gly Val Val Asn Ile Ser Lys 245 250 255 Pro Ser Val Val Gln Leu Asn Trp Arg Gly Phe Ser Tyr Leu Tyr Gly 260 265 270 Ala Trp Gly Arg Asp Tyr Ser Pro Gln His Pro Asn Lys Gly Leu Tyr 275 280 285 Trp Val Ala Pro Leu Asn Thr Asp Gly Arg Leu Leu Glu Tyr Tyr Arg 290 295 300 Leu Tyr Asn Thr Leu Asp Asp Leu Leu Leu Tyr Ile Asn Ala Arg Glu 305 310 315 320 Leu Arg Ile Thr Tyr Gly Gln Gly Ser Gly Thr Ala Val Tyr Asn Asn 325 330 335 Asn Met Tyr Val Asn Met Tyr Asn Thr Gly Asn Ile Ala Arg Val Asn 340 345 350 Leu Thr Thr Asn Thr Ile Ala Val Thr Gln Thr Leu Pro Asn Ala Ala 355 360 365 Tyr Asn Asn Arg Phe Ser Tyr Ala Asn Val Ala Trp Gln Asp Ile Asp 370 375 380 Phe Ala Val Asp Glu Asn Gly Leu Trp Val Ile Tyr Ser Thr Glu Ala 385 390 395 400 Ser Thr Gly Asn Met Val Ile Ser Lys Leu Asn Asp Thr Thr Leu Gln 405 410 415 Val Leu Asn Thr Trp Tyr Thr Lys Gln Tyr Lys Pro Ser Ala Ser Asn 420 425 430 Ala Phe Met Val Cys Gly Val Leu Tyr Ala Thr Arg Thr Met Asn Thr 435 440 445 Arg Thr Glu Glu Ile Phe Tyr Tyr Tyr Asp Thr Asn Thr Gly Lys Glu 450 455

460 Gly Lys Leu Asp Ile Val Met His Lys Met Gln Glu Lys Val Gln Ser 465 470 475 480 Ile Asn Tyr Asn Pro Phe Asp Gln Lys Leu Tyr Val Tyr Asn Asp Gly 485 490 495 Tyr Leu Leu Asn Tyr Asp Leu Ser Val Leu Gln Lys Pro Gln 500 505 510

Claims

What is claimed is:

1. A method for diagnosing the presence of colon cancer in a patient comprising: (a) measuring levels of CSG in a sample of cells, tissue or bodily fluid obtained from the patient; and (b) comparing the measured levels of CSG with levels of CSG in a sample of cells, tissue or bodily fluid obtained from a control, wherein an increase in measured levels of CSG in the patient versus the CSG levels in the control is associated with the presence of colon cancer.

2. A method of diagnosing metastatic colon cancer in a patient comprising: (a) measuring levels of CSG in a sample of cells, tissue, or bodily fluid obtained from the patient; and (b) comparing the measured levels of CSG with levels of CSG in a sample of cells, tissue, or bodily fluid obtained from a control, wherein an increase in measured CSG levels in the patient versus the CSG levels in the control is associated with a cancer which has metastasized.

3. A method of staging colon cancer in a patient comprising: (a) identifying a patient suffering from colon cancer; (b) measuring levels of CSG in a sample of cells, tissue, or bodily fluid obtained from the patient; and (c) comparing the measured levels of CSG with levels of CSG in a sample of cells, tissue, or bodily fluid obtained from a control, wherein an increase in the measured levels of CSG versus the levels of CSG in the control is associated with a cancer which is progressing and a decrease in the measured levels of CSG versus the levels of CSG in the control is associated with a cancer which is regressing or in remission.

4. A method of monitoring colon cancer in a patient for the onset of metastasis comprising: (a) identifying a patient having colon cancer that is not known to have metastasized; (b) periodically measuring CSG levels in samples of cells, tissue, or bodily fluid obtained from the patient; and (c) comparing the periodically measured levels of CSG with levels of CSG in cells, tissue, or bodily fluid obtained from a control, wherein an increase in any one of the periodically measured levels of CSG in the patient versus the levels of CSG in the control is associated with a cancer which has metastasized.

5. A method of monitoring changes in a stage of colon cancer in a patient comprising: (a) identifying a patient having colon cancer; (b) periodically measuring levels of CSG in samples of cells, tissue, or bodily fluid obtained from the patient; and (c) comparing the measured levels of CSG with levels of CSG in a sample of the same cells, tissue, or bodily fluid of a control, wherein an increase in any one of the periodically measured levels of CSG versus levels of CSG in the control is associated with a cancer which is progressing in stage and a decrease in any one of the periodically measured levels of CSG versus the levels of CSG in the control is associated with a cancer which is regressing in stage or in remission.

6. The method of claim 1, 2, 3, 4 or 5 wherein the CSG comprises SEQ ID NO:3, 4, 5 or 7.