Method and Apparatus for Correlating Levels of Biomarker Products with Disease

Abstract

In one aspect the invention is a method of testing for one or more colorectal pathologies or one or more subtypes of colorectal pathology (in one embodiment colorectal cancer) in a test individual by providing data corresponding to a level of products of selected biomarkers and applying the data to a formula to provide an indication of whether the test individual has one or more colorectal pathologies or one or more subtypes of colorectal pathology. In some aspects the method is computer based and a computer applies the data to the formula. In other aspects a computer system is configured with instructions that cause the processor to provide a user with the indication of whether the test individual has colorectal pathology. Also encompassed are kits for measuring data corresponding to the products of selected biomarkers which in some embodiments include a computer readable medium. Also encompassed are kits and methods of monitoring therapeutic efficacy of treatments for one or more colorectal pathologies.

1 - 34 . (canceled) 
     
     
         35 . A method of determining a probability of whether a human test subject is more likely to have colorectal cancer than to not have colorectal cancer comprising,
 (a) amplifying a test cDNA from blood of the human test subject complementary to a test mRNA to obtain values for levels of test mRNA expressed for each gene of a set of genes consisting of MS4A1 and one or more genes selected from the group consisting of MGC20553, CD163, CDA, BANK1, and BCNP1, comprising the steps of:   (i) generating test cDNA from test mRNA expressed specifically by each of the genes of the set of genes from blood of the human test subject; and   (ii) reacting the test cDNA under conditions to amplify DNA with an appropriate primer pair comprising a first and a second primer, wherein the first and the second primer of each primer pair represents a pair of forward and reverse primers corresponding to MS4A1, MGC20553, CD163, CDA, BANK1 and BCNP1 respectively, to obtain the values of the levels of the test mRNAs expressed specifically by the genes of the set of genes: and   (b) applying to the values of the levels of the test mRNAs a mathematical model formulated using logistic regression analysis of levels of control cDNA generated from RNA encoded by the set of genes in blood of human control subjects having colorectal cancer and levels of control cDNA generated from control RNA encoded by the set of genes in blood of human control subjects not having colorectal cancer, wherein the mathematical model is formulated for determining the probability that a test subject has colorectal cancer as opposed to not having colorectal cancer.   
     
     
         36 . The method of  claim 35 , wherein the set of genes consists of MS4A1 and MGC20553. 
     
     
         37 . The method of  claim 35 , wherein the appropriate primer pairs are selected from the primers listed in Table 14. 
     
     
         38 . The method of  claim 35 , wherein the value of the levels of control cDNA generated from RNA encoded by the set of genes in blood of human subject having colorectal cancer, and not having colorectal cancer, was obtained by the method comprising:
 (i) generating control cDNA from the RNA expressed by the genes of the set of genes from the blood of each of the human control subjects; and   (ii) reacting the control cDNA under conditions to amplify DNA with appropriate primers, wherein the first and the second primer of each primer pair represents a pair of forward and reverse primers corresponding to MS4A1, MGC20553, CD163, CDA, BANK1, and BCNP1 respectively, to obtain the values of the levels of the control mRNAs expressed by the set of genes.   
     
     
         39 . The method of  claim 36 , wherein the value of the levels of control cDNA generated from RNA encoded by the set of genes in blood of human subject having colorectal cancer, and not having colorectal cancer, was obtained by the method comprising:
 (i) generating control cDNA from the RNA expressed by the genes of the set of genes from the blood of each of the human control subjects; and   (ii) reacting the control cDNA under conditions to amplify DNA with appropriate primers, wherein the first and the second primer of each primer pair represents a pair of forward and reverse primers corresponding to MS4A1 and MGC20553 respectively, to obtain the values of the levels of the control mRNAs expressed by the set of genes.   
     
     
         40 . The method of  claim 37 , wherein the value of the levels of control cDNA generated from RNA encoded by the set of genes in blood of human subject having colorectal cancer, and not having colorectal cancer, was obtained by the method comprising:
 (i) generating control cDNA from the RNA expressed by the genes of the set of genes from the blood of each of the human control subjects; and   (ii) reacting the control cDNA under conditions to amplify DNA with appropriate primer pairs, wherein the first and the second primer of each primer pair represents a pair of forward and reverse primers corresponding to MS4A1, MGC20553, CD163, CDA, BANK1, and BCNP1 respectively, to obtain the values of the levels of the control mRNAs expressed by the set of genes.   
     
     
         41 . The method of  claim 35 , wherein the conditions to amplify DNA comprise:
 (a) combining the test cDNA, the set of primer pairs, a thermostable DNA polymerase, and a plurality of free nucleotides comprising adenine, thymine, cytosine, and guanine in a reaction mixture;   (b) heating the reaction mixture to a first predetermined temperature for a first predetermined time to separate the strands of the test cDNA from each other;   (c) cooling the reaction mixture to a second predetermined temperature for a second predetermined time under conditions to allow the first and second primers to hybridize with their complementary sequences on the first and second strands of the test cDNA, and to allow the polymerase to extend the primers; and   (d) repeating steps (b) and (c).