Abstract:
Biomarkers (and suites of biomarkers) relating to prostate cancer are provided, as well as methods for using such biomarkers (ans suites thereof), including early prediction of prostate cancer, disease grading, target identification/validation, and monitoring of drug efficacy.

Description:
[0001]    This application claims the benefit of U.S. Provisional Patent Application No. 61/368,434, filed Jul. 28, 2010, the entire contents of which are hereby incorporated herein by reference. 
     
    
     FIELD 
       [0002]    The invention generally relates to biomarkers for prostate cancer and methods based on the same biomarkers. 
       BACKGROUND 
       [0003]    Prostate cancer is the leading cause of male cancer-related deaths and afflicts one out of nine men over the age of 65. The American Cancer Society estimates that over 200,000 American men will be diagnosed with prostate cancer and over 30,000 will die this year. While effective surgical and radiation treatments exist for localized prostate cancer, metastatic prostate cancer remains essentially incurable and most men diagnosed with metastatic disease will succumb over a period of months to years. 
         [0004]    Prostate cancer is detected by either a digital rectal exam (DRE), or by the measurement of levels of prostate specific antigen (PSA), which has an unacceptably high rate of false-positives. The diagnosis of prostate cancer can be confirmed only by a biopsy. Radical prostatectomy, radiation and watchful waiting are generally effective for localized prostate cancer, but it is often difficult to determine which approach to use. Since it is not possible to distinguish between the indolent and more aggressive tumors current therapy takes a very conservative approach. 
         [0005]    While imaging, X-rays, computerized tomography scans and further biopsies can help determine if prostate cancer has metastasized, they are not able to differentiate early stages. Understanding the progression of prostate cancer from a localized, early, indolent state, to an aggressive state, and, ultimately, to a metastatic state would allow the proper clinical management of this disease. Furthermore, early-indolent prostate cancer may be progressive or non-progressive toward aggressive forms. 
       SUMMARY 
       [0006]    In one aspect, the present invention provides a method of diagnosing whether a subject has prostate cancer, comprising analyzing a biological sample from a subject to determine the level(s) of one or more biomarkers for prostate cancer in the sample, where the one or more biomarkers are selected from Tables 1A, 1B, 3A, 3B, 5A, 5B, 7A, 7B, 8, and/or 10 and comparing the level(s) of the one or more biomarkers in the sample to prostate cancer-positive and/or prostate cancer-negative reference levels of the one or more biomarkers in order to diagnose whether the subject has prostate cancer. The one or more biomarkers may be selected from Tables 1A, 1B, 3A, 3B, and 8. When the biological sample is prostate tissue the one or more biomarkers may be selected from Tables 1A, 1B, 3A, 3B, 5A, 5B, 7A, 7B, 8, and/or 10, or may be selected from Tables 1A, 1B, 3A, 3B, 5A, 5B, 7A, 7B, and/or 10. When the biological sample is urine the one or more biomarkers may be selected from Tables 1A, 1B, 3A, 3B, 5A, 5B, 7A, 7B, 8, and/or 10, or may be selected from Table 8. The biological sample may be a DRE urine sample. 
         [0007]    In another aspect, the present invention also provides a method of determining whether a subject is predisposed to developing prostate cancer, comprising analyzing a biological sample from a subject to determine the level(s) of one or more biomarkers for prostate cancer in the sample, where the one or more biomarkers are selected from Tables 1A, 1B, 3A, 3B, 5A, 5B, 7A, 7B, 8, and/or 10; and comparing the level(s) of the one or more biomarkers in the sample to prostate cancer-positive and/or prostate cancer-negative reference levels of the one or more biomarkers in order to determine whether the subject is predisposed to developing prostate cancer. 
         [0008]    In yet another aspect, the invention provides a method of monitoring progression/regression of prostate cancer in a subject comprising analyzing a first biological sample from a subject to determine the level(s) of one or more biomarkers for prostate cancer in the sample, where the one or more biomarkers are selected from Tables 1A, 1B, 3A, 3B, 5A, 5B, 7A, 7B, 8, and/or 10 and the first sample is obtained from the subject at a first time point; analyzing a second biological sample from a subject to determine the level(s) of the one or more biomarkers, where the second sample is obtained from the subject at a second time point; and comparing the level(s) of one or more biomarkers in the first sample to the level(s) of the one or more biomarkers in the second sample in order to monitor the progression/regression of prostate cancer in the subject. 
         [0009]    In another aspect, the present invention provides a method of assessing the efficacy of a composition for treating prostate cancer comprising analyzing, from a subject having prostate cancer and currently or previously being treated with a composition, a biological sample to determine the level(s) of one or more biomarkers for prostate cancer selected from Tables 1A, 1B, 3A, 3B, 5A, 5B, 7A, 7B, 8, and/or 10; and comparing the level(s) of the one or more biomarkers in the sample to (a) levels of the one or more biomarkers in a previously-taken biological sample from the subject, where the previously-taken biological sample was obtained from the subject before being treated with the composition, (b) prostate cancer-positive reference levels of the one or more biomarkers, and/or (c) prostate cancer-negative reference levels of the one or more biomarkers. 
         [0010]    In another aspect, the present invention provides a method for assessing the efficacy of a composition in treating prostate cancer, comprising analyzing a first biological sample from a subject to determine the level(s) of one or more biomarkers for prostate cancer selected from Tables 1A, 1B, 3A, 3B, 5A, 5B, 7A, 7B, 8, and/or 10, the first sample obtained from the subject at a first time point; administering the composition to the subject; analyzing a second biological sample from the subject to determine the level(s) of the one or more biomarkers, the second sample obtained from the subject at a second time point after administration of the composition; comparing the level(s) of one or more biomarkers in the first sample to the level(s) of the one or more biomarkers in the second sample in order to assess the efficacy of the composition for treating prostate cancer. 
         [0011]    In yet another aspect, the invention provides a method of assessing the relative efficacy of two or more compositions for treating prostate cancer comprising analyzing, from a first subject having prostate cancer and currently or previously being treated with a first composition, a first biological sample to determine the level(s) of one or more biomarkers selected from Tables 1A, 1B, 3A, 3B, 5A, 5B, 7A, 7B, 8, and/or 10; analyzing, from a second subject having prostate cancer and currently or previously being treated with a second composition, a second biological sample to determine the level(s) of the one or more biomarkers; and comparing the level(s) of one or more biomarkers in the first sample to the level(s) of the one or more biomarkers in the second sample in order to assess the relative efficacy of the first and second compositions for treating prostate cancer. 
         [0012]    In another aspect, the present invention provides a method for screening a composition for activity in modulating one or more biomarkers of prostate cancer, comprising contacting one or more cells with a composition; analyzing at least a portion of the one or more cells or a biological sample associated with the cells to determine the level(s) of one or more biomarkers of prostate cancer selected from Tables 1A, 1B, 3A, 3B, 5A, 5B, 7A, 7B, 8, and/or 10; and comparing the level(s) of the one or more biomarkers with predetermined standard levels for the biomarkers to determine whether the composition modulated the level(s) of the one or more biomarkers. 
         [0013]    In a further aspect, the present invention provides a method for identifying a potential drug target for prostate cancer comprising identifying one or more biochemical pathways associated with one or more biomarkers for prostate cancer selected from Tables 1A, 1B, 3A, 3B, 5A, 5B, 7A, 7B, 8, and/or 10; and identifying a protein affecting at least one of the one or more identified biochemical pathways, the protein being a potential drug target for prostate cancer. 
         [0014]    In yet another aspect, the invention provides a method for treating a subject having prostate cancer comprising administering to the subject an effective amount of one or more biomarkers selected from Tables 1A, 1B, 3A, 3B, 5A, 5B, 7A, 7B, 8, and/or 10 that are decreased in prostate cancer. In another aspect, the invention also provides a method of distinguishing low grade (less aggressive) prostate cancer from high grade (high aggressive) prostate cancer in a subject having prostate cancer, comprising analyzing a biological sample from a subject to determine the level(s) of one or more biomarkers for low grade prostate cancer and/or high grade prostate cancer in the sample, where the one or more biomarkers are selected from Tables 1A, 1B, 3A, 3B, 5A, 5B, 7A, 7B, 8, and/or 10 and comparing the level(s) of the one or more biomarkers in the sample to low grade prostate cancer-positive reference levels that distinguish over high grade prostate cancer and/or to high grade prostate cancer-positive reference levels that distinguish over low grade prostate cancer in order to determine whether the subject has low grade or high grade prostate cancer. The one or more biomarkers may be selected from Tables 1A, 1B, 5A, 5B, 7A, 7B, 8 and/or 10. When the biological sample is prostate tissue, the one or more biomarkers may be selected from Tables 1A, 1B, 3A, 3B, 5A, 5B, 7A, 7B, 8, and/or 10; may be selected from Tables 1A, 1B, 3A, 3B, 5A, 5B, 7A, 7B, and/or 10; or may be selected from Table 10. When selected from Table 10, the biomarkers may be selected from putrescine, lactate, 5,6-dihydrouracil, 10-nonadecenoate, NAD+, spermine, N-acetylputrescine, succinylcarnitine, 3-(4-hydroxyphenyl)lactate, 2-palmitoylglycerophosphoethanolamine, spermidine, glycerol-2-phosphate, glycylvaline, and/or phosphoethanolamine; may be selected from putrescine, lactate, 5,6-dihydrouracil, 10-nonadecenoate, NAD+, spermine, and/or N-acetylputrescine; may be selected from putrescine, glycerol-2-phosphate, and/or glycylvaline; may be selected from phosphoethanolamine, putrescine, and/or spermidine; may be selected from succinylcarnitine, 3-(4-hydroxyphenyl)lactate, 2-palmitoylglycerophosphoethanolamine, lactate, and/or spermidine; and/or may be selected from putrescine, lactate, 5,6-dihydrouracil, 10-nonadecenoate, NAD+, spermine, N-acetylputrescine, succinylcarnitine, 3-(4-hydroxyphenyl)lactate, 2-palmitoylglycerophosphoethanolamine, spermidine, glycerol-2-phosphate, glycylvaline, and/or phosphoethanolamine. 
     
    
     
       BRIEF DESCRIPTION OF THE DRAWINGS 
         [0015]      FIG. 1  provides a recursive partitioning plot based on one example metabolite (adrenate) to distinguish between subjects with high aggressive prostate cancer and low aggressive prostate cancer (Left) and the corresponding receiver operating characteristic (ROC), or ROC curve, graphical plot of the sensitivity, or true positives, vs. (1—specificity), or false positives (Right). 
           [0016]      FIG. 2  provides boxplots of representative biomarker metabolites that are correlated in abundance with cancer. The AUCs for the individual biomarker metabolites range from 0.73 to 0.84. The level of the biomarker in the benign (non-cancer) DRE urine sediment samples is presented on the left and the cancer samples is on the right. 
           [0017]      FIG. 3  provides a Receiver Operator Characteristics (ROC) curve for the current state of the art tests for prostate cancer detection, the “Post-DRE PCA 3” (PCA3) test and the “Serum PSA” (PSA) test. The Area Under the Curve (AUC) for the PCA3 test was approximately 0.68 and the AUC for the PSA test was approximately 0.61. 
           [0018]      FIG. 4  is a heat map that illustrates the biomarker signatures from DRE urine sediment samples that are associated with prostate cancer. Groups 1 and 2 are biomarker signatures of prostate cancer while Group 3 is a biomarker signature of non-cancer. The cancer biomarker signatures (Group 1 and Group 2) further distinguish subtypes of prostate cancer. 
           [0019]      FIG. 5  shows an ROC curve for the Han nomogram described in Example 7. 
       
    
    
     DETAILED DESCRIPTION 
       [0020]    The present invention relates to biomarkers of prostate cancer, methods for diagnosis of prostate cancer, methods of distinguishing between less aggressive and high aggressive prostate cancer, methods of determining predisposition to prostate cancer, methods of monitoring progression/regression of prostate cancer, methods of assessing efficacy of compositions for treating prostate cancer, methods of screening compositions for activity in modulating biomarkers of prostate cancer, methods of treating prostate cancer, as well as other methods based on biomarkers of prostate cancer. Prior to describing this invention in further detail, however, the following terms will first be defined. 
       DEFINITIONS 
       [0021]    “Biomarker” means a compound, preferably a metabolite, that is differentially present (i.e., increased or decreased) in a biological sample from a subject or a group of subjects having a first phenotype (e.g., having a disease) as compared to a biological sample from a subject or group of subjects having a second phenotype (e.g., not having the disease). A biomarker may be differentially present at any level, but is generally present at a level that is increased by at least 5%, by at least 10%, by at least 15%, by at least 20%, by at least 25%, by at least 30%, by at least 35%, by at least 40%, by at least 45%, by at least 50%, by at least 55%, by at least 60%, by at least 65%, by at least 70%, by at least 75%, by at least 80%, by at least 85%, by at least 90%, by at least 95%, by at least 100%, by at least 110%, by at least 120%, by at least 130%, by at least 140%, by at least 150%, or more; or is generally present at a level that is decreased by at least 5%, by at least 10%, by at least 15%, by at least 20%, by at least 25%, by at least 30%, by at least 35%, by at least 40%, by at least 45%, by at least 50%, by at least 55%, by at least 60%, by at least 65%, by at least 70%, by at least 75%, by at least 80%, by at least 85%, by at least 90%, by at least 95%, or by 100% (i.e., absent). A biomarker is preferably differentially present at a level that is statistically significant (i.e., a p-value less than 0.05 and/or a q-value of less than 0.10 as determined using either Welch&#39;s T-test or Wilcoxon&#39;s rank-sum Test). 
         [0022]    The “level” of one or more biomarkers means the absolute or relative amount or concentration of the biomarker in the sample. 
         [0023]    “Sample” or “biological sample” means biological material isolated from a subject. The biological sample may contain any biological material suitable for detecting the desired biomarkers, and may comprise cellular and/or non-cellular material from the subject. The sample can be isolated from any suitable biological tissue or fluid such as, for example, prostate tissue, blood, blood plasma, urine, or cerebral spinal fluid (CSF). 
         [0024]    “Subject” means any animal, but is preferably a mammal, such as, for example, a human, monkey, mouse, or rabbit. 
         [0025]    A “reference level” of a biomarker means a level of the biomarker that is indicative of a particular disease state, phenotype, or lack thereof, as well as combinations of disease states, phenotypes, or lack thereof A “positive” reference level of a biomarker means a level that is indicative of a particular disease state or phenotype. A “negative” reference level of a biomarker means a level that is indicative of a lack of a particular disease state or phenotype. For example, a “prostate cancer-positive reference level” of a biomarker means a level of a biomarker that is indicative of a positive diagnosis of prostate cancer in a subject, and a “prostate cancer-negative reference level” of a biomarker means a level of a biomarker that is indicative of a negative diagnosis of prostate cancer in a subject. A “reference level” of a biomarker may be an absolute or relative amount or concentration of the biomarker, a presence or absence of the biomarker, a range of amount or concentration of the biomarker, a minimum and/or maximum amount or concentration of the biomarker, a mean amount or concentration of the biomarker, and/or a median amount or concentration of the biomarker; and, in addition, “reference levels” of combinations of biomarkers may also be ratios of absolute or relative amounts or concentrations of two or more biomarkers with respect to each other. Appropriate positive and negative reference levels of biomarkers for a particular disease state, phenotype, or lack thereof may be determined by measuring levels of desired biomarkers in one or more appropriate subjects, and such reference levels may be tailored to specific populations of subjects (e.g., a reference level may be age-matched so that comparisons may be made between biomarker levels in samples from subjects of a certain age and reference levels for a particular disease state, phenotype, or lack thereof in a certain age group). Such reference levels may also be tailored to specific techniques that are used to measure levels of biomarkers in biological samples (e.g., LC-MS, GC-MS, etc.), where the levels of biomarkers may differ based on the specific technique that is used. 
         [0026]    “Non-biomarker compound” means a compound that is not differentially present in a biological sample from a subject or a group of subjects having a first phenotype (e.g., having a first disease) as compared to a biological sample from a subject or group of subjects having a second phenotype (e.g., not having the first disease). Such non-biomarker compounds may, however, be biomarkers in a biological sample from a subject or a group of subjects having a third phenotype (e.g., having a second disease) as compared to the first phenotype (e.g., having the first disease) or the second phenotype (e.g., not having the first disease). 
         [0027]    “Metabolite”, or “small molecule”, means organic and inorganic molecules which are present in a cell. The term does not include large macromolecules, such as large proteins (e.g., proteins with molecular weights over 2,000, 3,000, 4,000, 5,000, 6,000, 7,000, 8,000, 9,000, or 10,000), large nucleic acids (e.g., nucleic acids with molecular weights of over 2,000, 3,000, 4,000, 5,000, 6,000, 7,000, 8,000, 9,000, or 10,000), or large polysaccharides (e.g., polysaccharides with a molecular weights of over 2,000, 3,000, 4,000, 5,000, 6,000, 7,000, 8,000, 9,000, or 10,000). The small molecules of the cell are generally found free in solution in the cytoplasm or in other organelles, such as the mitochondria, where they form a pool of intermediates which can be metabolized further or used to generate large molecules, called macromolecules. The term “small molecules” includes signaling molecules and intermediates in the chemical reactions that transform energy derived from food into usable forms. Examples of small molecules include sugars, fatty acids, amino acids, nucleotides, intermediates formed during cellular processes, and other small molecules found within the cell. 
         [0028]    “Metabolic profile”, or “small molecule profile”, means a complete or partial inventory of small molecules within a targeted cell, tissue, organ, organism, or fraction thereof (e.g., cellular compartment). The inventory may include the quantity and/or type of small molecules present. The “small molecule profile” may be determined using a single technique or multiple different techniques. 
         [0029]    “Metabolome” means all of the small molecules present in a given organism. 
         [0030]    “Prostate cancer” refers to a disease in which cancer develops in the prostate, a gland in the male reproductive system. “Low grade” or “lower grade” prostate cancer refers to non-metastatic prostate cancer, including malignant tumors with low potential for metastisis (i.e. prostate cancer that is considered to be “less aggressive”). Cancer tumors that are confined to the prostate (i.e. organ-confined, OC) are considered to be less aggressive prostate cancer. “High grade” or “higher grade” prostate cancer refers to prostate cancer that has metastasized in a subject, including malignant tumors with high potential for metastasis (prostate cancer that is considered to be “aggressive”). Cancer tumors that are not confined to the prostate (i.e. non-organ-confined, NOC) are considered to be aggressive prostate cancer. Tumors that are confined to the prostate (i.e., organ confined tumors) are considered to be less aggressive than tumors which are not confined to the prostate (i.e., non-organ confined tumors). “Aggressive” prostate cancer progresses, recurs and/or is the cause of death. Aggressive cancer may be characterized by one or more of the following: non-organ confined (NOC), association with extra capsular extensions (ECE), association with seminal vesicle invasion (SVI), association with lymph node invasion (LN), association with a Gleason Score major or Gleason Score minor of 4, and/or association with a Gleason Score Sum of 8 or higher. In contrast “less aggressive” cancer is confined to the prostate (organ confined, OC) and is not associated with extra capsular extensions (ECE), seminal vesicle invasion (SVI), lymph node invasion (LN), a Gleason Score major or Gleason Score minor of 4, or a Gleason Score Sum of 8 or higher. 
       I. Biomarkers 
       [0031]    The prostate cancer biomarkers described herein were discovered using metabolomic profiling techniques. Such metabolomic profiling techniques are described in more detail in the Examples set forth below as well as in U.S. Pat. Nos. 7,005,255, 7,329,489; 7,550,258; 7,550,260; 7,553,616; 7,635,556; 7,682,783; 7,682,784; 7,910,301; 6,947,453; 7,433,787; 7,561,975; 7,884,318, the entire contents of which are hereby incorporated herein by reference. 
         [0032]    Generally, metabolic profiles were determined for biological samples from human subjects diagnosed with prostate cancer, the human subjects were diagnosed with lower grade prostate cancer (e.g., organ-confined tumor) or were diagnosed with metastatic/high grade prostate cancer (e.g., non-organ confined tumor). The metabolic profile for biological samples from a subject having prostate cancer was compared to the metabolic profile for biological samples from the one or more other groups of subjects. Those molecules differentially present, including those molecules differentially present at a level that is statistically significant, in the metabolic profile of tumor samples from subjects with aggressive prostate cancer as compared to another group (e.g., subjects diagnosed with less aggressive prostate cancer) were identified as biomarkers to distinguish those groups. In addition, those molecules differentially present, including those molecules differentially present at a level that is statistically significant, in the metabolic profile of non-tumor samples (i.e., non-cancerous tissue adjacent to a cancer tumor) from subjects with low grade prostate cancer as compared to high grade prostate cancer were also identified as biomarkers to distinguish those groups. 
         [0033]    The biomarkers are discussed in more detail herein. The biomarkers that were discovered correspond with the following group(s):
       Biomarkers for distinguishing subjects having prostate cancer vs. control subjects not diagnosed with prostate cancer (see Tables 1A, 1B, 3A, 3B, and 8); and   Biomarkers for distinguishing subjects having aggressive prostate cancer from subjects with less aggressive prostate cancer (see Tables 1A, 1B, 3A, 3B, 5A, 5B, 7A, 7B, and 10);
 
although the biomarkers in Tables 5A, 5B, 7A, 7B, and 10 may also be used to distinguish subjects having prostate cancer vs. control subjects not diagnosed with prostate cancer, and the biomarkers in Table 8 may also be used to distinguish subjects having aggressive prostate cancer from subjects with less aggressive prostate cancer.
       
 
       IIA. Diagnosis of Prostate Cancer 
       [0036]    The identification of biomarkers for prostate cancer allows for the diagnosis of (or for aiding in the diagnosis of) prostate cancer in subjects presenting one or more symptoms of prostate cancer. A method of diagnosing (or aiding in diagnosing) whether a subject has prostate cancer comprises (1) analyzing a biological sample from a subject to determine the level(s) of one or more biomarkers of prostate cancer in the sample and (2) comparing the level(s) of the one or more biomarkers in the sample to prostate cancer-positive and/or prostate cancer-negative reference levels of the one or more biomarkers in order to diagnose (or aid in the diagnosis of) whether the subject has prostate cancer. The one or more biomarkers that are used are selected from Tables 1A, 1B, 3A, 3B, 5A, 5B, 7A, and/or 7B and combinations thereof. In one aspect, the one or more biomarkers may be selected from Tables 1A, 1B, 3A, 3B, and 8. When such a method is used to aid in the diagnosis of prostate cancer, the results of the method may be used along with other methods (or the results thereof) useful in the clinical determination of whether a subject has prostate cancer. 
         [0037]    Any suitable method may be used to analyze the biological sample in order to determine the level(s) of the one or more biomarkers in the sample. Suitable methods include chromatography (e.g., HPLC, gas chromatography, liquid chromatography), mass spectrometry (e.g., MS, MS-MS), enzyme-linked immunosorbent assay (ELISA), antibody linkage, other immunochemical techniques, and combinations thereof. Further, the level(s) of the one or more biomarkers may be measured indirectly, for example, by using an assay that measures the level of a compound (or compounds) that correlates with the level of the biomarker(s) that are desired to be measured. 
         [0038]    The levels of one or more of the biomarkers of Tables 1A, 1B, 3A, 3B, 5A, 5B, 7A, 7B, 8 and/or 10 may be determined in the methods of diagnosing and methods of aiding in diagnosing whether a subject has prostate cancer. For example, the level(s) of one biomarker, two or more biomarkers, three or more biomarkers, four or more biomarkers, five or more biomarkers, six or more biomarkers, seven or more biomarkers, eight or more biomarkers, nine or more biomarkers, ten or more biomarkers, etc., including a combination of all of the biomarkers in Tables 1A, 1B, 3A, 3B, 5A, 5B, 7A, 7, 8, and/or 10 and combinations thereof or any fraction thereof, may be determined and used in such methods. Determining levels of combinations of the biomarkers may allow greater sensitivity and specificity in diagnosing prostate cancer and aiding in the diagnosis of prostate cancer, and may allow better differentiation of prostate cancer from other prostate disorders (e.g. benign prostatic hypertrophy (BPH), prostatitis, etc.) or other cancers that may have similar or overlapping biomarkers to prostate cancer (as compared to a subject not having prostate cancer). For example, ratios of the levels of certain biomarkers (and non-biomarker compounds) in biological samples may allow greater sensitivity and specificity in diagnosing prostate cancer and aiding in the diagnosis of prostate cancer and may allow better differentiation of prostate cancer from other cancers or other disorders of the prostate that may have similar or overlapping biomarkers to prostate cancer (as compared to a subject not having prostate cancer). 
         [0039]    One or more biomarkers that are specific for diagnosing prostate cancer (or aiding in diagnosing prostate cancer) in a certain type of sample (e.g., prostate tissue sample, urine sample, or blood plasma sample) may also be used. For example, when the biological sample is prostate tissue, one or more biomarkers listed in Tables 1A, 1B, 3A, 3B, 5A, 5B, 7A, 7B, and/or 10, may be used to diagnose (or aid in diagnosing) whether a subject has prostate cancer. As another example, when the biological sample is urine (or DRE urine), one or more biomarkers listed in Table 8 may be used to diagnose (or aid in diagnosing) whether a subject has prostate cancer. 
         [0040]    After the level(s) of the one or more biomarkers in the sample are determined, the level(s) are compared to prostate cancer-positive and/or prostate cancer-negative reference levels to aid in diagnosing or to diagnose whether the subject has prostate cancer. Levels of the one or more biomarkers in a sample matching the prostate cancer-positive reference levels (e.g., levels that are the same as the reference levels, substantially the same as the reference levels, above and/or below the minimum and/or maximum of the reference levels, and/or within the range of the reference levels) are indicative of a diagnosis of prostate cancer in the subject. Levels of the one or more biomarkers in a sample matching the prostate cancer-negative reference levels (e.g., levels that are the same as the reference levels, substantially the same as the reference levels, above and/or below the minimum and/or maximum of the reference levels, and/or within the range of the reference levels) are indicative of a diagnosis of no prostate cancer in the subject. In addition, levels of the one or more biomarkers that are differentially present (especially at a level that is statistically significant) in the sample as compared to prostate cancer-negative reference levels are indicative of a diagnosis of prostate cancer in the subject. Levels of the one or more biomarkers that are differentially present (especially at a level that is statistically significant) in the sample as compared to prostate cancer-positive reference levels are indicative of a diagnosis of no prostate cancer in the subject. 
         [0041]    The level(s) of the one or more biomarkers may be compared to prostate cancer-positive and/or prostate cancer-negative reference levels using various techniques, including a simple comparison (e.g., a manual comparison) of the level(s) of the one or more biomarkers in the biological sample to prostate cancer-positive and/or prostate cancer-negative reference levels. The level(s) of the one or more biomarkers in the biological sample may also be compared to prostate cancer-positive and/or prostate cancer-negative reference levels using one or more statistical analyses (e.g., t-test, Welch&#39;s T-test, Wilcoxon&#39;s rank sum test, random forest). 
         [0042]    In addition, the biological samples may be analyzed to determine the level(s) of one or more non-biomarker compounds. The level(s) of such non-biomarker compounds may also allow differentiation of prostate cancer from other prostate disorders that may have similar or overlapping biomarkers to prostate cancer (as compared to a subject not having a prostate disorder). For example, a known non-biomarker compound present in biological samples of subjects having prostate cancer and subjects not having prostate cancer could be monitored to verify a diagnosis of prostate cancer as compared to a diagnosis of another prostate disorder when biological samples from subjects having the prostate disorder do not have the non-biomarker compound. 
         [0043]    The methods of diagnosing (or aiding in diagnosing) whether a subject has prostate cancer may also be conducted specifically to diagnose (or aid in diagnosing) whether a subject has less aggressive prostate cancer and/or high aggressive prostate cancer. Such methods comprise (1) analyzing a biological sample from a subject to determine the level(s) of one or more biomarkers of less aggressive prostate cancer (and/or high aggressove prostate cancer) in the sample and (2) comparing the level(s) of the one or more biomarkers in the sample to less aggressive prostate cancer-positive and/or less aggressive prostate cancer-negative reference levels (or high aggressive prostate cancer-positive and/or high aggressive prostate cancer-negative reference levels) in order to diagnose (or aid in the diagnosis of) whether the subject has less aggressive prostate cancer (or high aggressive prostate cancer). Biomarker specific for low grade prostate cancer are listed in Tables 1, 3, 7 and biomarkers specific for high grade prostate cancer are listed in Tables 1, 3, 7. 
         [0000]    IIB. Methods of Distinguishing Less Aggressive Prostate Cancer (Low Grade) from More Aggressive Prostate Cancer (High Grade) 
         [0044]    The identification of biomarkers for distinguishing less aggressive prostate cancer versus more aggressive prostate cancer allows less aggressive prostate cancer and aggressive prostate cancer to be distinguished in patients. The subjects can then be treated appropriately, with those subjects having more aggressive prostate cancer undergoing more aggressive treatment than those subjects with less aggressive prostate cancer. A method of distinguishing less aggressive prostate cancer from more aggressive prostate cancer in a subject having prostate cancer comprises (1) analyzing a biological sample from a subject to determine the level(s) in the sample of one or more biomarkers of less aggressive prostate cancer that distinguish over high aggressive prostate cancer and/or one or more biomarkers of high aggressive prostate cancer that distinguish over less aggressive prostate cancer, and (2) comparing the level(s) of the one or more biomarkers in the sample to less aggressive prostate cancer-positive reference levels that distinguish over high aggressive prostate cancer and/or high aggressive prostate cancer-positive reference levels that distinguish over less aggressive prostate cancer of the one or more biomarkers in order to determine whether the subject has less aggressive or high aggressive prostate cancer. The one or more biomarkers that are used are selected from Tables 1A, 1B, 3A, 3B, 5A, 5B, 7A, 7B, 8, and/or 10 and combinations thereof. 
         [0045]    In one aspect of the invention, the biomarkers that are used are selected from Tables 1A, 1B, 3A, 3B, 5A, 5B, 7A, 7B, and/or 10 and combinations thereof. 
         [0046]    In another aspect of the invention the one or more biomarkers that are used are selected from putrescine, lactate, 5,6-dihydrouracil, 10-nonadecenoate, NAD+, spermine, N-acetylputrescine, succinylcarnitine, 3-(4-hydroxyphenyl)lactate, 2-palmitoylglycerophosphoethanolamine, spermidine, glycerol-2-phosphate, glycylvaline, and/or phosphoethanolamine. 
         [0047]    In an aspect of the invention, the more aggressive cancer is associated with extracapsular extensions (ECE) and the biomarker metabolites are selected from putrescine, lactate, 5,6-dihydrouracil, 10-nonadecenoate, NAD+, spermine, and/or N-acetylputrescine. 
         [0048]    In an aspect of the invention, the more aggressive cancer is associated with seminal vesicle invasion (SVI) and the biomarkers are selected from putrescine, glycerol-2-phosphate, and/or glycylvaline. 
         [0049]    In an aspect of the invention, the more aggressive cancer is associated with lymph node invasion and the biomarkers are selected from phosphoethanolamine, putrescine, and/or spermidine. 
         [0050]    In an aspect of the invention, the more aggressive cancer is associated with a Gleason Score (GS) greater than 8 and the biomarkers are selected from succinylcarnitine, 3-(4-hydroxyphenyl)lactate, 2-palmitoylglycerophosphoethanolamine, lactate, and/or spermidine. 
         [0051]    Any suitable method may be used to analyze the biological sample in order to determine the level(s) of the one or more biomarkers in the sample. Suitable methods include chromatography (e.g., HPLC, gas chromatography, liquid chromatography), mass spectrometry (e.g., MS, MS-MS), enzyme-linked immunosorbent assay (ELISA), antibody linkage, other immunochemical techniques, and combinations thereof. Further, the level(s) of the one or more biomarkers may be measured indirectly, for example, by using an assay that measures the level of a compound (or compounds) that correlates with the level of the biomarker(s) that are desired to be measured. 
         [0052]    The levels of one or more of the biomarkers of Tables 1A, 1B, 3A, 3B, 5A, 5B, 7A, 7B, 8, and/or 10 may be determined in the methods of diagnosing and methods of aiding in diagnosing whether a subject has prostate cancer. For example, the level(s) of one biomarker, two or more biomarkers, three or more biomarkers, four or more biomarkers, five or more biomarkers, six or more biomarkers, seven or more biomarkers, eight or more biomarkers, nine or more biomarkers, ten or more biomarkers, etc., including a combination of all of the biomarkers in Tables 1A, 1B, 3A, 3B, 5A, 5B, 7A, 7B, 8, and/or 10 or any fraction thereof, may be determined and used in such methods. Determining levels of combinations of the biomarkers may allow greater sensitivity and specificity in distinguishing between low aggressive and high aggressive prostate cancer. 
         [0053]    One or more biomarkers that are specific for distinguishing between less aggressive and high aggressive prostate cancer in a certain type of sample (e.g., prostate tissue sample, urine sample, or blood plasma sample) may also be used. For example, when the biological sample is prostate tissue, one or more biomarkers listed in Tables 1A, 1B, 3A, 3B, 5A, 5B, 7A, 7B, and/or 10 may be used. As another example, when the biological sample is urine (or DRE urine), one or more biomarkers listed in Table 8 may be used. 
         [0054]    After the level(s) of the one or more biomarkers in the sample are determined, the level(s) are compared to less aggressive prostate cancer-positive reference levels that distinguish over high aggressive prostate cancer-negative and/or high aggressive prostate cancer-positive reference levels that distinguish over less aggressive prostate cancer of the one or more biomarkers in order to determine whether the subject has less aggressive or high aggressive prostate cancer. Levels of the one or more biomarkers in a sample matching the less aggressive prostate cancer-positive reference levels that distinguish over high aggressive prostate cancer (e.g., levels that are the same as the reference levels, substantially the same as the reference levels, above and/or below the minimum and/or maximum of the reference levels, and/or within the range of the reference levels) are indicative of less aggressive prostate cancer in the subject. Levels of the one or more biomarkers in a sample matching the high aggressive prostate cancer-positive reference levels that distinguish over low aggressive prostate cancer (e.g., levels that are the same as the reference levels, substantially the same as the reference levels, above and/or below the minimum and/or maximum of the reference levels, and/or within the range of the reference levels) are indicative of high-aggressive prostate cancer in the subject. If the level(s) of the one or more biomarkers are more similar to the less aggressive prostate cancer-positive reference levels that distinguish over high aggressive prostate cancer (or less similar to the high aggressive prostate cancer-positive reference levels), then the results are indicative of less aggressive prostate cancer in the subject. If the level(s) of the one or more biomarkers are more similar to the high aggressive prostate cancer-positive reference levels that distinguish over less aggressive prostate cancer (or less similar to the less aggressive prostate cancer-positive reference levels), then the results are indicative of high aggressive prostate cancer in the subject. 
         [0055]    The level(s) of the one or more biomarkers may be compared to less aggressive prostate cancer-positive reference levels that distinguish over high aggressive prostate cancer and/or high aggressive prostate cancer-positive reference levels that distinguish over less aggressive prostate cancer using various techniques, including a simple comparison (e.g., a manual comparison) of the level(s) of the one or more biomarkers in the biological sample to less aggressive prostate cancer-positive and/or high aggressive prostate cancer-positive reference levels. The level(s) of the one or more biomarkers in the biological sample may also be compared to less aggressive prostate cancer-positive reference levels that distinguish over high aggressive prostate cancer and/or high aggressive prostate cancer-positive reference levels that distinguish over less aggressive prostate cancer using one or more statistical analyses (e.g., t-test, Welch&#39;s T-test, Wilcoxon&#39;s rank sum test, random forest). 
         [0056]    In addition, the biological samples may be analyzed to determine the level(s) of one or more non-biomarker compounds. The level(s) of such non-biomarker compounds may also allow differentiation of less aggressive prostate cancer from high aggressive prostate cancer. 
       III. Methods of Determining Predisposition to Prostate Cancer 
       [0057]    The identification of biomarkers for prostate cancer also allows for the determination of whether a subject having no symptoms of prostate cancer is predisposed to developing prostate cancer. A method of determining whether a subject having no symptoms of prostate cancer is predisposed to developing prostate cancer comprises (1) analyzing a biological sample from a subject to determine the level(s) of one or more biomarkers listed in Tables 1A, 1B, 3A, 3B, 5A, 5B, 7A, 7B, 8, and/or 10 in the sample and (2) comparing the level(s) of the one or more biomarkers in the sample to prostate cancer-positive and/or prostate cancer-negative reference levels of the one or more biomarkers in order to determine whether the subject is predisposed to developing prostate cancer. The results of the method may be used along with other methods (or the results thereof) useful in the clinical determination of whether a subject is predisposed to developing prostate cancer. 
         [0058]    As described above in connection with methods of diagnosing (or aiding in the diagnosis of) prostate cancer, any suitable method may be used to analyze the biological sample in order to determine the level(s) of the one or more biomarkers in the sample. 
         [0059]    As with the methods of diagnosing (or aiding in the diagnosis of) prostate cancer described above, the level(s) of one biomarker, two or more biomarkers, three or more biomarkers, four or more biomarkers, five or more biomarkers, six or more biomarkers, seven or more biomarkers, eight or more biomarkers, nine or more biomarkers, ten or more biomarkers, etc., including a combination of all of the biomarkers in Tables 1A, 1B, 3A, 3B, 5A, 5B, 7A, 7B, 8, and/or 10 or any fraction thereof, may be determined and used in methods of determining whether a subject having no symptoms of prostate cancer is predisposed to developing prostate cancer. 
         [0060]    After the level(s) of the one or more biomarkers in the sample are determined, the level(s) are compared to prostate cancer-positive and/or prostate cancer-negative reference levels in order to predict whether the subject is predisposed to developing prostate cancer. Levels of the one or more biomarkers in a sample matching the prostate cancer-positive reference levels (e.g., levels that are the same as the reference levels, substantially the same as the reference levels, above and/or below the minimum and/or maximum of the reference levels, and/or within the range of the reference levels) are indicative of the subject being predisposed to developing prostate cancer. Levels of the one or more biomarkers in a sample matching the prostate cancer-negative reference levels (e.g., levels that are the same as the reference levels, substantially the same as the reference levels, above and/or below the minimum and/or maximum of the reference levels, and/or within the range of the reference levels) are indicative of the subject not being predisposed to developing prostate cancer. In addition, levels of the one or more biomarkers that are differentially present (especially at a level that is statistically significant) in the sample as compared to prostate cancer-negative reference levels are indicative of the subject being predisposed to developing prostate cancer. Levels of the one or more biomarkers that are differentially present (especially at a level that is statistically significant) in the sample as compared to prostate cancer-positive reference levels are indicative of the subject not being predisposed to developing prostate cancer. 
         [0061]    Furthermore, it may also be possible to determine reference levels specific to assessing whether or not a subject that does not have prostate cancer is predisposed to developing prostate cancer. For example, it may be possible to determine reference levels of the biomarkers for assessing different degrees of risk (e.g., low, medium, high) in a subject for developing prostate cancer. Such reference levels could be used for comparison to the levels of the one or more biomarkers in a biological sample from a subject. 
         [0062]    As with the methods described above, the level(s) of the one or more biomarkers may be compared to prostate cancer-positive and/or prostate cancer-negative reference levels using various techniques, including a simple comparison, one or more statistical analyses, and combinations thereof. 
         [0063]    As with the methods of diagnosing (or aiding in diagnosing) whether a subject has prostate cancer, the methods of determining whether a subject having no symptoms of prostate cancer is predisposed to developing prostate cancer may further comprise analyzing the biological sample to determine the level(s) of one or more non-biomarker compounds. 
         [0064]    The methods of determining whether a subject having no symptoms of prostate cancer is predisposed to developing prostate cancer may also be conducted specifically to determine whether a subject having no symptoms of prostate cancer is predisposed to developing less aggressive prostate cancer and/or high aggressive prostate cancer. Biomarker specific for less aggressive prostate cancer are listed in Tables 1, 3, 5, 7, and 10 and biomarkers specific for high aggressive prostate cancer are listed in Tables 1, 3, 5, 7, and 10. 
         [0065]    In addition, methods of determining whether a subject having less aggressive prostate cancer is predisposed to developing high aggressive prostate cancer may be conducted using one or more biomarkers selected from Tables 1A, 1B, 3A, 3B, 5A, 5B, 7A, 7B, 8, and/or 10. 
       IV. Methods of Monitoring Progression/Regression of Prostate Cancer 
       [0066]    The identification of biomarkers for prostate cancer also allows for monitoring progression/regression of prostate cancer in a subject. A method of monitoring the progression/regression of prostate cancer in a subject comprises (1) analyzing a first biological sample from a subject to determine the level(s) of one or more biomarkers for prostate cancer selected from Tables 1A, 1B, 3A, 3B, 5A, 5B, 7A, 7B, 8, and/or 10, the first sample obtained from the subject at a first time point, (2) analyzing a second biological sample from a subject to determine the level(s) of the one or more biomarkers, the second sample obtained from the subject at a second time point, and (3) comparing the level(s) of one or more biomarkers in the first sample to the level(s) of the one or more biomarkers in the second sample in order to monitor the progression/regression of prostate cancer in the subject. The results of the method are indicative of the course of prostate cancer (i.e., progression or regression, if any change) in the subject. 
         [0067]    The change (if any) in the level(s) of the one or more biomarkers over time may be indicative of progression or regression of prostate cancer in the subject. In order to characterize the course of prostate cancer in the subject, the level(s) of the one or more biomarkers in the first sample, the level(s) of the one or more biomarkers in the second sample, and/or the results of the comparison of the levels of the biomarkers in the first and second samples may be compared to prostate cancer-positive, prostate cancer-negative, less aggressive prostate cancer-positive, less aggressive prostate cancer-negative, high-aggressive prostate cancer-positive, and/or high aggressive prostate cancer-negative reference levels as well as less aggressive prostate cancer-positive reference levels that distinguish over high aggressive prostate cancer and/or high aggressive prostate cancer-positive reference levels that distinguish over low aggressive prostate cancer. If the comparisons indicate that the level(s) of the one or more biomarkers are increasing or decreasing over time (e.g., in the second sample as compared to the first sample) to become more similar to the prostate cancer-positive reference levels (or less similar to the prostate cancer-negative reference levels), to the high aggressive prostate cancer reference levels, or, when the subject initially has less aggressive prostate cancer, to the high aggressive prostate cancer-positive reference levels that distinguish over less aggressive prostate cancer, then the results are indicative of prostate cancer progression. If the comparisons indicate that the level(s) of the one or more biomarkers are increasing or decreasing over time to become more similar to the prostate cancer-negative reference levels (or less similar to the prostate cancer-positive reference levels), or, when the subject initially has high aggressive prostate cancer, to less aggressive prostate cancer reference levels and/or to less aggressive prostate cancer-positive reference levels that distinguish over high aggressive prostate cancer, then the results are indicative of prostate cancer regression. 
         [0068]    As with the other methods described herein, the comparisons made in the methods of monitoring progression/regression of prostate cancer in a subject may be carried out using various techniques, including simple comparisons, one or more statistical analyses, and combinations thereof. 
         [0069]    The results of the method may be used along with other methods (or the results thereof) useful in the clinical monitoring of progression/regression of prostate cancer in a subject. 
         [0070]    As described above in connection with methods of diagnosing (or aiding in the diagnosis of) prostate cancer, any suitable method may be used to analyze the biological samples in order to determine the level(s) of the one or more biomarkers in the samples. In addition, the level(s) one or more biomarkers, including a combination of all of the biomarkers in Tables 1A, 1B, 3A, 3B, 5A, 5B, 7A, 7B, 8, and/or 10 or any fraction thereof, may be determined and used in methods of monitoring progression/regression of prostate cancer in a subject. 
         [0071]    Such methods could be conducted to monitor the course of prostate cancer in subjects having prostate cancer or could be used in subjects not having prostate cancer (e.g., subjects suspected of being predisposed to developing prostate cancer) in order to monitor levels of predisposition to prostate cancer. 
       V. Methods of Assessing Efficacy of Compositions for Treating Prostate Cancer 
       [0072]    The identification of biomarkers for prostate cancer also allows for assessment of the efficacy of a composition for treating prostate cancer as well as the assessment of the relative efficacy of two or more compositions for treating prostate cancer. Such assessments may be used, for example, in efficacy studies as well as in lead selection of compositions for treating prostate cancer. 
         [0073]    A method of assessing the efficacy of a composition for treating prostate cancer comprises (1) analyzing, from a subject having prostate cancer and currently or previously being treated with a composition, a biological sample to determine the level(s) of one or more biomarkers selected from Tables 1A, 1B, 3A, 3B, 5A, 5B, 7A, 7B, 8, and/or 10, and (2) comparing the level(s) of the one or more biomarkers in the sample to (a) level(s) of the one or more biomarkers in a previously-taken biological sample from the subject, wherein the previously-taken biological sample was obtained from the subject before being treated with the composition, (b) prostate cancer-positive reference levels (including less aggressive prostate cancer-positive and/or high aggressive prostate cancer-positive reference levels) of the one or more biomarkers, (c) prostate cancer-negative reference levels (including less aggressive prostate cancer-negative and/or high aggressive prostate cancer-negative reference levels) of the one or more biomarkers, (d) less aggressive prostate cancer-positive reference levels that distinguish over high aggressive prostate cancer, and/or (e) high aggressive prostate cancer-positive reference levels that distinguish over less aggressive prostate cancer. The results of the comparison are indicative of the efficacy of the composition for treating prostate cancer. 
         [0074]    Thus, in order to characterize the efficacy of the composition for treating prostate cancer, the level(s) of the one or more biomarkers in the biological sample are compared to (1) prostate cancer-positive reference levels, (2) prostate cancer-negative reference levels, (3) previous levels of the one or more biomarkers in the subject before treatment with the composition, (4) less aggressive prostate cancer-positive reference levels that distinguish over high aggressive prostate cancer, and/or (5) high aggressive prostate cancer-positive reference levels that distinguish over less aggressive prostate cancer. 
         [0075]    When comparing the level(s) of the one or more biomarkers in the biological sample (from a subject having prostate cancer and currently or previously being treated with a composition) to prostate cancer-positive reference levels and/or prostate cancer-negative reference levels, level(s) in the sample matching the prostate cancer-negative reference levels (e.g., levels that are the same as the reference levels, substantially the same as the reference levels, above and/or below the minimum and/or maximum of the reference levels, and/or within the range of the reference levels) are indicative of the composition having efficacy for treating prostate cancer. Levels of the one or more biomarkers in the sample matching the prostate cancer-positive reference levels (e.g., levels that are the same as the reference levels, substantially the same as the reference levels, above and/or below the minimum and/or maximum of the reference levels, and/or within the range of the reference levels) are indicative of the composition not having efficacy for treating prostate cancer. The comparisons may also indicate degrees of efficacy for treating prostate cancer based on the level(s) of the one or more biomarkers. 
         [0076]    When comparing the level(s) of the one or more biomarkers in the biological sample (from a subject having high aggressive prostate cancer and currently or previously being treated with a composition) less aggressive prostate cancer-positive reference levels that distinguish over high aggressive prostate cancer and/or high aggressive prostate cancer-positive reference levels that distinguish over less aggressive prostate cancer, level(s) in the sample matching the less aggressive prostate cancer-positive reference levels that distinguish over high aggressive prostate cancer (e.g., levels that are the same as the reference levels, substantially the same as the reference levels, above and/or below the minimum and/or maximum of the reference levels, and/or within the range of the reference levels) are indicative of the composition having efficacy for treating prostate cancer. Levels of the one or more biomarkers in the sample matching the high aggressive prostate cancer-positive reference levels that distinguish over less aggressive prostate cancer (e.g., levels that are the same as the reference levels, substantially the same as the reference levels, above and/or below the minimum and/or maximum of the reference levels, and/or within the range of the reference levels) are indicative of the composition not having efficacy for treating prostate cancer. 
         [0077]    When the level(s) of the one or more biomarkers in the biological sample (from a subject having prostate cancer and currently or previously being treated with a composition) are compared to level(s) of the one or more biomarkers in a previously-taken biological sample from the subject before treatment with the composition, any changes in the level(s) of the one or more biomarkers are indicative of the efficacy of the composition for treating prostate cancer. That is, if the comparisons indicate that the level(s) of the one or more biomarkers have increased or decreased after treatment with the composition to become more similar to the prostate cancer-negative reference levels (or less similar to the prostate cancer-positive reference levels) or, when the subject initially has high aggressive prostate cancer, the level(s) have increased or decreased to become more similar to less aggressive prostate cancer-positive reference levels that distinguish over high aggressive prostate cancer (or less similar to the high aggressive prostate cancer-positive reference levels that distinguish over low aggressive prostate cancer), then the results are indicative of the composition having efficacy for treating prostate cancer. If the comparisons indicate that the level(s) of the one or more biomarkers have not increased or decreased after treatment with the composition to become more similar to the prostate cancer-negative reference levels (or less similar to the prostate cancer-positive reference levels) or, when the subject initially has high aggressive prostate cancer, the level(s) have not increased or decreased to become more similar to less aggressive prostate cancer-positive reference levels that distinguish over high aggressive prostate cancer (or less similar to the high aggressive prostate cancer-positive reference levels that distinguish over less aggressive prostate cancer), then the results are indicative of the composition not having efficacy for treating prostate cancer. The comparisons may also indicate degrees of efficacy for treating prostate cancer based on the amount of changes observed in the level(s) of the one or more biomarkers after treatment. In order to help characterize such a comparison, the changes in the level(s) of the one or more biomarkers, the level(s) of the one or more biomarkers before treatment, and/or the level(s) of the one or more biomarkers in the subject currently or previously being treated with the composition may be compared to prostate cancer-positive reference levels (including less aggressive and high aggressive prostate cancer-positive reference levels), prostate cancer-negative reference levels (including less aggressive and high aggressive prostate cancer-negative reference levels), less aggressive prostate cancer-positive reference levels that distinguish over high aggressive prostate cancer, and/or high aggressive prostate cancer-positive reference levels that distinguish over less aggressive prostate cancer. 
         [0078]    Another method for assessing the efficacy of a composition in treating prostate cancer comprises (1) analyzing a first biological sample from a subject to determine the level(s) of one or more biomarkers selected from Tables 1A, 1B, 3A, 3B, 5A, 5B, 7A, 7B, 8, and/or 10, the first sample obtained from the subject at a first time point, (2) administering the composition to the subject, (3) analyzing a second biological sample from a subject to determine the level(s) of the one or more biomarkers, the second sample obtained from the subject at a second time point after administration of the composition, and (4) comparing the level(s) of one or more biomarkers in the first sample to the level(s) of the one or more biomarkers in the second sample in order to assess the efficacy of the composition for treating prostate cancer. As indicated above, if the comparison of the samples indicates that the level(s) of the one or more biomarkers have increased or decreased after administration of the composition to become more similar to the prostate cancer-negative reference levels (or less similar to the prostate cancer-positive reference levels) or, when the subject initially has high aggressive prostate cancer, if the level(s) have increased or decreased to become more similar to less aggressive prostate cancer-positive reference levels that distinguish over high aggressive prostate cancer (or less similar to the high aggressive prostate cancer-positive reference levels that distinguish over less aggressive prostate cancer), then the results are indicative of the composition having efficacy for treating prostate cancer. If the comparisons indicate that the level(s) of the one or more biomarkers have not increased or decreased after treatment with the composition to become more similar to the prostate cancer-negative reference levels (or less similar to the prostate cancer-positive reference levels) or, when the subject initially has high aggressive prostate cancer, the level(s) have not increased or decreased to become more similar to less aggressive prostate cancer-positive reference levels that distinguish over high aggressive prostate cancer (or less similar to the high aggressive prostate cancer-positive reference levels that distinguish over less aggressive prostate cancer), then the results are indicative of the composition not having efficacy for treating prostate cancer. The comparison may also indicate a degree of efficacy for treating prostate cancer based on the amount of changes observed in the level(s) of the one or more biomarkers after administration of the composition as discussed above. 
         [0079]    A method of assessing the relative efficacy of two or more compositions for treating prostate cancer comprises (1) analyzing, from a first subject having prostate cancer and currently or previously being treated with a first composition, a first biological sample to determine the level(s) of one or more biomarkers selected from Tables 1A, 1B, 3A, 3B, 5A, 5B, 7A, 7B, 8 and/or 10 (2) analyzing, from a second subject having prostate cancer and currently or previously being treated with a second composition, a second biological sample to determine the level(s) of the one or more biomarkers, and (3) comparing the level(s) of one or more biomarkers in the first sample to the level(s) of the one or more biomarkers in the second sample in order to assess the relative efficacy of the first and second compositions for treating prostate cancer. The results are indicative of the relative efficacy of the two compositions, and the results (or the levels of the one or more biomarkers in the first sample and/or the level(s) of the one or more biomarkers in the second sample) may be compared to prostate cancer-positive reference levels (including less aggressive and high aggressive prostate cancer-positive reference levels), prostate cancer-negative reference levels (including less aggressive and high aggressive prostate cancer-negative reference levels), less aggressive prostate cancer-positive reference levels that distinguish over high aggressive prostate cancer, and/or high aggressive prostate cancer-positive reference levels that distinguish over less aggressive prostate cancer to aid in characterizing the relative efficacy. 
         [0080]    Each of the methods of assessing efficacy may be conducted on one or more subjects or one or more groups of subjects (e.g., a first group being treated with a first composition and a second group being treated with a second composition). 
         [0081]    As with the other methods described herein, the comparisons made in the methods of assessing efficacy (or relative efficacy) of compositions for treating prostate cancer may be carried out using various techniques, including simple comparisons, one or more statistical analyses, and combinations thereof. Any suitable method may be used to analyze the biological samples in order to determine the level(s) of the one or more biomarkers in the samples. In addition, the level(s) of one or more biomarkers, including a combination of all of the biomarkers in Tables 1A, 1B, 3A, 3B, 5A, 5B, 7A, 7B, 8, and/or 10 or any fraction thereof, may be determined and used in methods of assessing efficacy (or relative efficacy) of compositions for treating prostate cancer. 
         [0082]    Finally, the methods of assessing efficacy (or relative efficacy) of one or more compositions for treating prostate cancer may further comprise analyzing the biological sample to determine the level(s) of one or more non-biomarker compounds. The non-biomarker compounds may then be compared to reference levels of non-biomarker compounds for subjects having (or not having) prostate cancer. 
         [0000]    VI. Methods of Screening a Composition for Activity in Modulating Biomarkers Associated with Prostate Cancer 
         [0083]    The identification of biomarkers for prostate cancer also allows for the screening of compositions for activity in modulating biomarkers associated with prostate cancer, which may be useful in treating prostate cancer. Methods of screening compositions useful for treatment of prostate cancer comprise assaying test compositions for activity in modulating the levels of one or more biomarkers in Tables 1A, 1B, 3A, 3B, 5A, 5B, 7A, 7B, 8, and/or 10. Such screening assays may be conducted in vitro and/or in vivo, and may be in any form known in the art useful for assaying modulation of such biomarkers in the presence of a test composition such as, for example, cell culture assays, organ culture assays, and in vivo assays (e.g., assays involving animal models). 
         [0084]    In one embodiment, a method for screening a composition for activity in modulating one or more biomarkers of prostate cancer comprises (1) contacting one or more cells with a composition, (2) analyzing at least a portion of the one or more cells or a biological sample associated with the cells to determine the level(s) of one or more biomarkers of prostate cancer selected from Tables 1A, 1B, 3A, 3B, 5A, 5B, 7A, 7B, 8, and/or 10; and (3) comparing the level(s) of the one or more biomarkers with predetermined standard levels for the one or more biomarkers to determine whether the composition modulated the level(s) of the one or more biomarkers. As discussed above, the cells may be contacted with the composition in vitro and/or in vivo. The predetermined standard levels for the one or more biomarkers may be the levels of the one or more biomarkers in the one or more cells in the absence of the composition. The predetermined standard levels for the one or more biomarkers may also be the level(s) of the one or more biomarkers in control cells not contacted with the composition. 
         [0085]    In addition, the methods may further comprise analyzing at least a portion of the one or more cells or a biological sample associated with the cells to determine the level(s) of one or more non-biomarker compounds of prostate cancer. The levels of the non-biomarker compounds may then be compared to predetermined standard levels of the one or more non-biomarker compounds. 
         [0086]    Any suitable method may be used to analyze at least a portion of the one or more cells or a biological sample associated with the cells in order to determine the level(s) of the one or more biomarkers (or levels of non-biomarker compounds). Suitable methods include chromatography (e.g., HPLC, gas chromatograph, liquid chromatography), mass spectrometry (e.g., MS, MS-MS), ELISA, antibody linkage, other immunochemical techniques, and combinations thereof. Further, the level(s) of the one or more biomarkers (or levels of non-biomarker compounds) may be measured indirectly, for example, by using an assay that measures the level of a compound (or compounds) that correlates with the level of the biomarker(s) (or non-biomarker compounds) that are desired to be measured. 
       VII. Method of Identifying Potential Drug Targets 
       [0087]    The identification of biomarkers for prostate cancer also allows for the identification of potential drug targets for prostate cancer. A method for identifying a potential drug target for prostate cancer comprises (1) identifying one or more biochemical pathways associated with one or more biomarkers for prostate cancer selected from Tables 1A, 1B, 3A, 3B, 5A, 5B, 7A, 7B, 8, and/or 10 and (2) identifying a protein (e.g., an enzyme) affecting at least one of the one or more identified biochemical pathways, the protein being a potential drug target for prostate cancer. 
         [0088]    Another method for identifying a potential drug target for prostate cancer comprises (1) identifying one or more biochemical pathways associated with one or more biomarkers for prostate cancer selected from Tables 1A, 1B, 3A, 3B, 5A, 5B, 7A, 7B, 8, and/or 10 and one or more non-biomarker compounds of prostate cancer and (2) identifying a protein affecting at least one of the one or more identified biochemical pathways, the protein being a potential drug target for prostate cancer. 
         [0089]    One or more biochemical pathways (e.g., biosynthetic and/or metabolic (catabolic) pathway) are identified that are associated with one or more biomarkers (or non-biomarker compounds). After the biochemical pathways are identified, one or more proteins affecting at least one of the pathways are identified. Preferably, those proteins affecting more than one of the pathways are identified. 
         [0090]    A build-up of one metabolite (e.g., a pathway intermediate) may indicate the presence of a ‘block’ downstream of the metabolite and the block may result in a low/absent level of a downstream metabolite (e.g. product of a biosynthetic pathway). In a similar manner, the absence of a metabolite could indicate the presence of a ‘block’ in the pathway upstream of the metabolite resulting from inactive or non-functional enzyme(s) or from unavailability of biochemical intermediates that are required substrates to produce the product. Alternatively, an increase in the level of a metabolite could indicate a genetic mutation that produces an aberrant protein which results in the over-production and/or accumulation of a metabolite which then leads to an alteration of other related biochemical pathways and result in dysregulation of the normal flux through the pathway; further, the build-up of the biochemical intermediate metabolite may be toxic or may compromise the production of a necessary intermediate for a related pathway. It is possible that the relationship between pathways is currently unknown and this data could reveal such a relationship. 
         [0091]    For example, the data indicates that metabolites in the biochemical pathways involving nitrogen excretion, amino acid metabolism, energy metabolism, oxidative stress, purine metabolism and bile acid metabolism are enriched in prostate cancer subjects. Further, polyamine levels are higher in cancer subjects, which indicates that the level and/or activity of the enzyme ornithine decarboxylase is increased. It is known that polyamines can act as mitotic agents and have been associated with free radical damage. These observations indicate that the pathways leading to the production of polyamines (or to any of the aberrant biomarkers) would provide a number of potential targets useful for drug discovery. 
         [0092]    The proteins identified as potential drug targets may then be used to identify compositions that may be potential candidates for treating prostate cancer, including compositions for gene therapy. 
       VIII. Methods of Treating Prostate Cancer 
       [0093]    The identification of biomarkers for prostate cancer also allows for the treatment of prostate cancer. For example, in order to treat a subject having prostate cancer, an effective amount of one or more prostate cancer biomarkers that are lowered in prostate cancer as compared to a healthy subject not having prostate cancer may be administered to the subject. The biomarkers that may be administered may comprise one or more of the biomarkers in Tables 1A, 1B, 3A, 3B, 5A, 5B, 7A, 7B, 8, and/or 10 that are decreased in prostate cancer. In some embodiments, the biomarkers that are administered are one or more biomarkers listed in Tables 1A, 1B, 3A, 3B, 5A, 5B, 7A, 7B, 8, and/or 10 that are decreased in prostate cancer and that have a p-value less than 0.10. In other embodiments, the biomarkers that are administered are one or biomarkers listed in Tables 1A, 1B, 3A, 3B, 5A, 5B, 7A, 7B, 8, and/or 10 that are decreased in prostate cancer by at least 5%, by at least 10%, by at least 15%, by at least 20%, by at least 25%, by at least 30%, by at least 35%, by at least 40%, by at least 45%, by at least 50%, by at least 55%, by at least 60%, by at least 65%, by at least 70%, by at least 75%, by at least 80%, by at least 85%, by at least 90%, by at least 95%, or by 100% (i.e., absent). 
       IX. Methods of Using the Prostate Cancer Biomarkers for Other Types of Cancer 
       [0094]    It is believed that some of the biomarkers for major prostate cancer described herein may also be biomarkers for other types of cancer, including, for example, lung cancer or kidney cancer. Therefore, it is believed that at least some of the prostate cancer biomarkers may be used in the methods described herein for other types of cancer. That is, the methods described herein with respect to prostate cancer may also be used for diagnosing (or aiding in the diagnosis of) any type of cancer, methods of monitoring progression/regression of any type of cancer, methods of assessing efficacy of compositions for treating any type of cancer, methods of screening a composition for activity in modulating biomarkers associated with any type of cancer, methods of identifying potential drug targets for any type of cancer, and methods of treating any type of cancer. Such methods could be conducted as described herein with respect to prostate cancer. 
       X. Methods of Using the Prostate Cancer Biomarkers for Other Prostate Disorders 
       [0095]    It is believed that some of the biomarkers for prostate cancer described herein may also be biomarkers for prostate disorders (e.g. prostatitis, benign prostate hypertrophy (BHP)) in general. Therefore, it is believed that at least some of the prostate cancer biomarkers may be used in the methods described herein for prostate disorders in general. That is, the methods described herein with respect to prostate cancer may also be used for diagnosing (or aiding in the diagnosis of) a prostate disorder, methods of monitoring progression/regression of a prostate disorder, methods of assessing efficacy of compositions for treating a prostate disorder, methods of screening a composition for activity in modulating biomarkers associated with a prostate disorder, methods of identifying potential drug targets for prostate disorder, and methods of treating a prostate disorder. Such methods could be conducted as described herein with respect to prostate cancer. 
       XI. Other Methods 
       [0096]    Other methods of using the biomarkers discussed herein are also contemplated. For example, the methods described in U.S. Pat. No. 7,005,255, U.S. Pat. No. 7,329,489, U.S. Pat. No. 7,553,616, U.S. Pat. No. 7,550,260, U.S. Pat. No. 7,550,258, U.S. Pat. No. 7,635,556, U.S. patent application Ser. No. 11/728,826, U.S. patent application Ser. No. 12/463,690 and U.S. patent application Ser. No. 12/182,828 may be conducted using a small molecule profile comprising one or more of the biomarkers disclosed herein. 
         [0097]    In any of the methods listed herein, the biomarkers that are used may be selected from those biomarkers in Tables 1A, 1B, 3A, or 3B, 5A, 5B, 7A, 7B, 8, and/or 10 having p-values of less than 0.05 and/or those biomarkers in Tables 1A, 1B, 3A, 3B, 5A, 5B, 7A, 7B, 8, and/or 10 having q-values of less than 0.10. The biomarkers that are used in any of the methods described herein may also be selected from those biomarkers in Tables 1A, 1B, 3A, 3B, 5A, 5B, 7A, 7B, 8, and/or 10 that are decreased in prostate cancer (as compared to the control) or that are decreased in remission (as compared to control or prostate cancer) by at least 5%, by at least 10%, by at least 15%, by at least 20%, by at least 25%, by at least 30%, by at least 35%, by at least 40%, by at least 45%, by at least 50%, by at least 55%, by at least 60%, by at least 65%, by at least 70%, by at least 75%, by at least 80%, by at least 85%, by at least 90%, by at least 95%, or by 100% (i.e., absent); and/or those biomarkers in Tables 1A, 1B, 3A, 3B, 5A, 5B, 7A, 7B, 8, and/or 10 that are increased in prostate cancer (as compared to the control or remission) or that are increased in remission (as compared to the control or prostate cancer) by at least 5%, by at least 10%, by at least 15%, by at least 20%, by at least 25%, by at least 30%, by at least 35%, by at least 40%, by at least 45%, by at least 50%, by at least 55%, by at least 60%, by at least 65%, by at least 70%, by at least 75%, by at least 80%, by at least 85%, by at least 90%, by at least 95%, by at least 100%, by at least 110%, by at least 120%, by at least 130%, by at least 140%, by at least 150%, or more. 
       EXAMPLES 
       [0098]    The invention will be further explained by the following illustrative examples that are intended to be non-limiting. 
       I. General Methods 
       [0099]    A. Identification of Metabolic Profiles for Prostate Cancer 
         [0100]    Each sample was analyzed to determine the concentration of several hundred metabolites. Analytical techniques such as GC-MS (gas chromatography-mass spectrometry) and LC-MS (liquid chromatography-mass spectrometry) were used to analyze the metabolites. Multiple aliquots were simultaneously, and in parallel, analyzed, and, after appropriate quality control (QC), the information derived from each analysis was recombined. Every sample was characterized according to several thousand characteristics, which ultimately amount to several hundred chemical species. The techniques used were able to identify novel and chemically unnamed compounds. 
         [0101]    B. Statistical Analysis 
         [0102]    The data was analyzed using T-tests to identify molecules (either known, named metabolites or unnamed metabolites) present at differential levels in a definable population or subpopulation (e.g., biomarkers for prostate cancer biological samples compared to control biological samples or compared to patients in remission from prostate cancer) useful for distinguishing between the definable populations (e.g., prostate cancer and control, low aggressive prostate cancer and high aggressive prostate cancer). Other molecules (either known, named metabolites or unnamed metabolites) in the definable population or subpopulation were also identified. 
         [0103]    Data was also analyzed using Random Forest Analysis. Random forests give an estimate of how well individuals in a new data set can be classified into existing groups. Random forest analysis creates a set of classification trees based on continual sampling of the experimental units and compounds. Then each observation is classified based on the majority votes from all the classification trees. In statistics, a classification tree classifies the observations into groups based on combinations of the variables (in this instance variables are metabolites or compounds). There are many variations on the algorithms used to create trees. A tree algorithm searches for the metabolite (compound) that provides the largest split between the two groups. This produces nodes. Then at each node, the metabolite that provides the best split is used and so on. If the node cannot be improved on, then it stops at that node and any observation in that node is classified as the majority group. 
         [0104]    Random forests classify based on a large number (e.g. thousands) of trees. A subset of compounds and a subset of observations are used to create each tree. The observations used to create the tree are called the in-bag samples, and the remaining samples are called the out-of-bag samples. The classification tree is created from the in-bag samples, and the out-of-bag samples are predicted from this tree. To get the final classification for an observation, the “votes” for each group are counted based on the times it was an out-of-bag sample. For example, suppose observation 1 was classified as a “Control” by 2,000 trees, but classified as “Disease” by 3,000 trees. Using “majority wins” as the criterion, this sample is classified as “Disease.” 
         [0105]    The results of the random forest are summarized in a confusion matrix. The rows correspond to the true grouping, and the columns correspond to the classification from the random forest. Thus, the diagonal elements indicate the correct classifications. A 50% error would occur by random chance for 2 groups, 66.67% error for three groups by random chance, etc. The “Out-of-Bag” (OOB) Error rate gives an estimate of how accurately new observations can be predicted using the random forest model (e.g., whether a sample is from a diseased subject or a control subject). 
         [0106]    It is also of interest to see which variables are more “important” in the final classifications. The “importance plot” shows the top compounds ranked in terms of their importance. There are different criteria for ranking the importance, but the general idea is that removing an important variable will cause a greater decrease in accuracy than a variable that is less important. The most important identified biomarkers are presented in Tables 3A, 3B, 5A, 5B, 7A, and 7B. 
         [0107]    C. Biomarker Identification 
         [0108]    Various peaks identified in the analyses (e.g. GC-MS, LC-MS, MS-MS), including those identified as statistically significant, were subjected to a mass spectrometry based chemical identification process. 
       Example 1 
       [0109]    Biomarkers were discovered by (1) analyzing tissue samples from different groups of human subjects to determine the levels of metabolites in the samples and then (2) statistically analyzing the results to determine those metabolites that were differentially present in the two groups. 
         [0110]    The tissue samples used for the analysis were 61 control tissues that were cancer free tissues derived from sections of prostate tissue not containing cancer cells (i.e. from cancerous prostate glands and that were determined to be free of cancerous cells), 46 prostate tissue samples from organ confined (T_OC) prostate cancer tumors (i.e. lower aggressive prostate cancer) and 25 prostate tissue samples from non-organ confined (T_NOC) prostate cancer tumors (i.e. high aggressive prostate cancer). After the levels of metabolites were determined, the data was analyzed using univariate T-tests (i.e., Welch&#39;s T-test). 
         [0111]    T-tests were used to determine differences in the mean levels of metabolites between two populations (i.e., Prostate Cancer (T) vs. Control (C), High Aggressive (T_NOC) Prostate Cancer vs. Less Aggressive (T_OC) Prostate Cancer) and Adjacent tissue to High Aggressive Prostate Cancer (N_NOC) vs. Adjacent tissue to Less Aggressive Prostate Cancer Control (N_OC)). 
       Biomarkers: 
       [0112]    As listed below in Tables 1A and 1B, biomarkers were discovered that were differentially present between tissue samples from 1.) prostate cancer tumors and Control prostate tissue that was determined to be free of cancerous cells (i.e. sections of prostate tissue not containing cancerous cells from cancerous prostate glands removed from the patient), 2.) aggressive prostate tumors (i.e. tumors that were non-organ confined, NOC) and less aggressive prostate tumors (i.e. tumors that were organ confined, OC) and 3.) between NOC and OC cancer using non-cancer tissue adjacent to the NOC cancer tumor or the OC cancer tumor. The study was comprised of tissue collected from 25 subjects with non-organ-confined (NOC) prostate tumors and 46 subjects with (OC) organ-confined prostate cancer tumors. 
         [0113]    Tables 1A and 1B include, for each listed biomarker, the p-value and the q-value determined in the statistical analysis of the data concerning the biomarkers and the ratio of the mean level of cancer samples as compared to the control mean level (Tables 1A and 1B, columns 3-5), the p-value and the q-value determined in the statistical analysis of the data concerning the biomarkers and the ratio of the mean level of the non-cancer tissue adjacent to high aggressive prostate cancer (N_NOC) mean level as compared to the non-cancer tissue adjacent to less aggressive (N_OC) mean level (Tables 1A and 1B, columns 6-8), and the p-value and the q-value determined in the statistical analysis of the data concerning the biomarkers and the ratio of the mean level of the cancer tumor from high aggressive prostate cancer (T_NOC) mean level as compared to the cancer tumor from lower aggressive prostate cancer (T_OC) mean level (Tables 1A and 1B, columns 9-11). The term “Isobar” as used in the tables indicates the compounds that could not be distinguished from each other on the analytical platform used in the analysis (i.e., the compounds in an isobar elute at nearly the same time and have similar (and sometimes exactly the same) quant ions, and thus cannot be distinguished). 
         [0114]    Tables 1A and 1B. Prostate Cancer Biomarkers. 
         [0115]    Legend: C, Control non-cancer tissue adjacent to cancer tissue; T, Tumor cancer tissue; N_NOC, Non-cancerous tissue adjacent to cancer tumor that is Non-Organ Confined; N_OC, Non-Cancerous tissue adjacent to cancer tumor that is Organ Confined; T_NOC, Tumor tissue that is Non-Organ Confined; T_OC, Tumor tissue that is Organ Confined 
         [0000]    
       
         
               
               
               
               
               
               
               
               
               
               
               
             
               
               
               
               
               
               
               
               
               
               
               
             
           
               
                 TABLE 1A 
               
               
                   
               
               
                   
                   
                   
                   
                 Ratio 
                   
                 N_NOC 
                 Ratio 
                 T_NOC 
                   
                   
               
               
                   
                   
                   
                   
                 Cancer 
                 N_NOC 
                 VS 
                 NOC/ 
                 VS 
                 T_NOC 
                 Ratio 
               
               
                   
                   
                   
                   
                 Tumor/ 
                 VS 
                 N_OC 
                 OC 
                 T_OC 
                 VS 
                 T_NOC/ 
               
               
                 Comp 
                   
                 C VS T P- 
                 C VS T 
                 Control 
                 N_OC 
                 Q- 
                 Adja- 
                 P- 
                 T_OC Q- 
                 T_OC 
               
               
                 ID 
                 Name 
                 VALUE 
                 Q-VALUE 
                 (T/C) 
                 P-VALUE 
                 VALUE 
                 cent 
                 VALUE 
                 VALUE 
                 Tumor 
               
               
                   
               
             
             
               
                   
               
             
          
           
               
                 35439 
                 glutaroyl carnitine 
                 2.2E−13 
                 1.076E−11 
                 2.5428 
                 0.7797 
                 0.5723 
                 0.9867 
                 0.8926 
                 0.3790 
                 0.9852 
               
               
                 1356 
                 nonadecanoate (19:0) 
                 2.9E−13 
                 1.076E−11 
                 1.8780 
                 0.2707 
                 0.3190 
                 1.1401 
                 0.0357 
                 0.0412 
                 1.3227 
               
               
                 33972 
                 10-nonadecenoate (19:1n9) 
                   6E−13 
                 1.365E−11 
                 1.9738 
                 0.0022 
                 0.0281 
                 1.3829 
                 0.0040 
                 0.0129 
                 1.4216 
               
               
                 19324 
                 1-stearoylglycerophosphoinositol 
                 1.5E−11 
                 1.705E−10 
                 1.7487 
                 0.0362 
                 0.1082 
                 1.4025 
                 0.0040 
                 0.0129 
                 1.4886 
               
               
                 27728 
                 glycerol 2-phosphate 
                 2.1E−11 
                 1.987E−10 
                 2.0245 
                 0.9731 
                 0.6302 
                 0.9925 
                 0.0872 
                 0.0760 
                 1.2966 
               
               
                 37459 
                 ergothioneine 
                 4.5E−11 
                 3.407E−10 
                 1.7200 
                 0.2226 
                 0.2852 
                 1.1414 
                 0.1806 
                 0.1228 
                 1.2472 
               
               
                 36747 
                 deoxycarnitine 
                 6.7E−11 
                  4.46E−10 
                 1.3905 
                 0.0464 
                 0.1223 
                 1.1204 
                 0.0963 
                 0.0801 
                 1.1071 
               
               
                 37097 
                 tryptophan betaine 
                 7.7E−11 
                 4.879E−10 
                 1.3584 
                 0.1732 
                 0.2401 
                 1.0891 
                 0.0997 
                 0.0823 
                 1.3327 
               
               
                 37455 
                 glycerophosphoethanolamine 
                 3.7E−10 
                 1.607E−09 
                 2.1207 
                 0.4521 
                 0.4293 
                 0.9035 
                 0.2214 
                 0.1401 
                 1.0885 
               
               
                 32452 
                 propionylcarnitine 
                 1.4E−09 
                 4.967E−09 
                 1.4653 
                 0.0446 
                 0.1201 
                 1.2477 
                 0.1466 
                 0.1064 
                 1.2842 
               
               
                 18467 
                 eicosapentaenoate (EPA; 20:5n3) 
                 1.7E−09 
                 5.666E−09 
                 1.6414 
                 0.1555 
                 0.2282 
                 1.2593 
                 0.3180 
                 0.1803 
                 1.0792 
               
               
                 32654 
                 3-dehydrocarnitine 
                 1.9E−09 
                 6.024E−09 
                 1.2935 
                 0.2482 
                 0.3025 
                 1.0944 
                 0.1089 
                 0.0883 
                 1.1905 
               
               
                 32412 
                 butyrylcarnitine 
                 3.2E−09 
                 8.956E−09 
                 1.4534 
                 0.0771 
                 0.1586 
                 1.1538 
                 0.0172 
                 0.0280 
                 1.2936 
               
               
                 33587 
                 eicosenoate (20:1n9 or 11) 
                 3.4E−09 
                 9.39E−09 
                 1.7489 
                 0.0105 
                 0.0602 
                 1.3890 
                 0.0001 
                 0.0019 
                 1.6222 
               
               
                 1638 
                 arginine 
                 3.8E−09 
                 9.953E−09 
                 1.6913 
                 0.2783 
                 0.3269 
                 1.2801 
                 0.0337 
                 0.0399 
                 1.4488 
               
               
                 17805 
                 dihomo-linoleate (20:2n6) 
                 6.8E−09 
                 1.646E−08 
                 1.7543 
                 0.0053 
                 0.0447 
                 1.4025 
                 0.0006 
                 0.0044 
                 1.5861 
               
               
                 15772 
                 ribitol 
                 7.7E−09 
                  1.83E−08 
                 1.6384 
                 0.0002 
                 0.0149 
                 1.5060 
                 0.0000 
                 0.0002 
                 1.7684 
               
               
                 15720 
                 N-acetylglutamate 
                 8.5E−09 
                 1.962E−08 
                 0.6162 
                 0.1223 
                 0.2005 
                 1.1085 
                 0.8721 
                 0.3726 
                 0.9202 
               
               
                 35305 
                 1-palmitoylglycerophosphoinositol 
                 1.3E−08 
                 2.796E−08 
                 1.6574 
                 0.1761 
                 0.2420 
                 1.2487 
                 0.0095 
                 0.0195 
                 1.3145 
               
               
                 19260 
                 1-oleoylglycerophosphoserine 
                   2E−08 
                 4.021E−08 
                 1.4208 
                 0.1119 
                 0.1909 
                 1.1938 
                 0.0754 
                 0.0698 
                 1.2107 
               
               
                 36593 
                 2-linoleoylglycerophospho- 
                 2.1E−08 
                 4.021E−08 
                 1.6367 
                 0.0140 
                 0.0666 
                 1.5003 
                 0.0017 
                 0.0077 
                 1.5788 
               
               
                   
                 ethanolamine 
               
               
                 1577 
                 2-aminobutyrate 
                 2.6E−08 
                 5.039E−08 
                 1.2443 
                 0.0346 
                 0.1064 
                 1.2226 
                 0.0049 
                 0.0141 
                 1.3036 
               
               
                 35433 
                 hydroxyisovaleroyl carnitine 
                 2.7E−08 
                 5.146E−08 
                 1.8954 
                 0.1565 
                 0.2285 
                 1.2063 
                 0.0456 
                 0.0476 
                 1.3227 
               
               
                 33080 
                 N-ethylglycinexylidide 
                 3.3E−08 
                 6.118E−08 
                 1.4492 
                 0.5567 
                 0.4712 
                 1.1998 
                 0.0435 
                 0.0459 
                 1.5639 
               
               
                 37948 
                 2-oleoylglycerophosphoserine 
                 5.2E−08 
                 9.064E−08 
                 1.4802 
                 0.0204 
                 0.0832 
                 1.3687 
                 0.0134 
                 0.0243 
                 1.5106 
               
               
                 32198 
                 acetylcarnitine 
                 9.6E−08 
                 1.49E−07 
                 1.2642 
                 0.9134 
                 0.6222 
                 0.9900 
                 0.9119 
                 0.3806 
                 0.9668 
               
               
                 32635 
                 1-linoleoylglycerophospho 
                 1.2E−07 
                 1.762E−07 
                 1.7959 
                 0.3015 
                 0.3392 
                 1.2398 
                 0.0321 
                 0.0391 
                 1.4287 
               
               
                   
                 ethanolamine 
               
               
                 32415 
                 docosadienoate (22:2n6) 
                 2.3E−07 
                 3.131E−07 
                 1.5734 
                 0.0006 
                 0.0204 
                 1.4680 
                 0.0001 
                 0.0020 
                 1.6508 
               
               
                 3141 
                 betaine 
                 2.8E−07 
                 3.737E−07 
                 1.2893 
                 0.9353 
                 0.6302 
                 0.9951 
                 0.4788 
                 0.2438 
                 0.9541 
               
               
                 34437 
                 1-stearoylglycerophosphoglycerol 
                 2.8E−07 
                 3.737E−07 
                 2.0348 
                 0.0828 
                 0.1640 
                 1.5698 
                 0.1718 
                 0.1184 
                 1.4281 
               
               
                 35162 
                 UDP-N-acetylglucosamine 
                 3.5E−07 
                 4.525E−07 
                 1.9109 
                 0.4994 
                 0.4470 
                 0.9214 
                 0.9489 
                 0.3900 
                 1.0176 
               
               
                 32504 
                 docosapentaenoate 
                 3.9E−07 
                 5.061E−07 
                 1.4968 
                 0.0125 
                 0.0646 
                 1.4746 
                 0.0082 
                 0.0175 
                 1.4506 
               
               
                   
                 (n3 DPA; 22:5n3) 
               
               
                 34565 
                 1-palmitoleoylglycerophospho- 
                 4.2E−07 
                 5.327E−07 
                 2.2261 
                 0.6581 
                 0.5234 
                 1.0482 
                 0.2032 
                 0.1341 
                 1.1898 
               
               
                   
                 ethanolamine 
               
               
                 32417 
                 docosatrienoate (22:3n3) 
                 7.9E−07 
                 9.345E−07 
                 1.7688 
                 0.0012 
                 0.0251 
                 1.5248 
                 0.0011 
                 0.0060 
                 2.1304 
               
               
                 33971 
                 10-heptadecenoate (17:1n7) 
                 8.3E−07 
                 9.742E−07 
                 1.2217 
                 0.0147 
                 0.0667 
                 1.1435 
                 0.0826 
                 0.0745 
                 1.1477 
               
               
                 37419 
                 1-heptadecanoylglycerophospho- 
                 9.8E−07 
                 1.118E−06 
                 1.8968 
                 0.1528 
                 0.2263 
                 1.2312 
                 0.0327 
                 0.0391 
                 1.3785 
               
               
                   
                 ethanolamine 
               
               
                 21127 
                 1-palmitoylglycerol 
                 3.1E−06 
                 3.298E−06 
                 1.5124 
                 0.9054 
                 0.6194 
                 1.0302 
                 0.0246 
                 0.0329 
                 1.3123 
               
               
                   
                 (1-monopalmitin) 
               
               
                 19323 
                 docosahexaenoate (DHA; 22:6n3) 
                 3.3E−06 
                 3.369E−06 
                 1.6100 
                 0.0753 
                 0.1578 
                 1.3489 
                 0.0434 
                 0.0459 
                 1.5001 
               
               
                 15506 
                 choline 
                 3.7E−06 
                 3.725E−06 
                 1.1487 
                 0.0560 
                 0.1310 
                 1.0781 
                 0.0027 
                 0.0105 
                 1.1544 
               
               
                 35718 
                 dihomo-linolenate (20:3n3 or n6) 
                 4.2E−06 
                 4.158E−06 
                 1.6088 
                 0.0443 
                 0.1201 
                 1.3770 
                 0.0067 
                 0.0162 
                 1.5242 
               
               
                 2134 
                 flavin adenine dinucleotide (FAD) 
                 4.8E−06 
                 4.665E−06 
                 1.2276 
                 0.6157 
                 0.5021 
                 1.0230 
                 0.4335 
                 0.2269 
                 1.0488 
               
               
                 34035 
                 linolenate [alpha or gamma; 
                 9.8E−06 
                 8.992E−06 
                 1.3425 
                 0.0305 
                 0.0983 
                 1.3680 
                 0.0180 
                 0.0286 
                 1.4184 
               
               
                   
                 (18:3n3 or 6)] 
               
               
                 33487 
                 glutamate, gamma-methyl ester 
                 1.1E−05 
                 9.867E−06 
                 1.7460 
                 0.1457 
                 0.2198 
                 0.7586 
                 0.2105 
                 0.1368 
                 0.7030 
               
               
                 3108 
                 adenosine 5′-diphosphate (ADP) 
                 1.3E−05 
                 1.164E−05 
                 0.7466 
                 0.1627 
                 0.2313 
                 0.8626 
                 0.0064 
                 0.0157 
                 0.7410 
               
               
                 37058 
                 succinylcarnitine 
                 1.4E−05 
                 1.198E−05 
                 1.5749 
                 0.7291 
                 0.5540 
                 0.9290 
                 0.0152 
                 0.0255 
                 1.3840 
               
               
                 37202 
                 4-androsten-3beta,17beta-diol 
                 1.5E−05 
                 1.242E−05 
                 0.7759 
                 0.2829 
                 0.3309 
                 1.3527 
                 0.3546 
                 0.1930 
                 1.2828 
               
               
                   
                 disulfate 1 
               
               
                 1361 
                 pentadecanoate (15:0) 
                 1.6E−05 
                 1.375E−05 
                 0.8034 
                 0.8579 
                 0.6007 
                 1.0474 
                 0.5080 
                 0.2524 
                 0.9049 
               
               
                 1301 
                 lysine 
                 2.2E−05 
                 1.858E−05 
                 1.5717 
                 0.6977 
                 0.5404 
                 1.3324 
                 0.2270 
                 0.1416 
                 1.4210 
               
               
                 22171 
                 glycylproline 
                   3E−05 
                 2.422E−05 
                 1.4058 
                 0.0120 
                 0.0646 
                 3.0403 
                 0.0103 
                 0.0205 
                 2.8213 
               
               
                 22175 
                 aspartylphenylalanine 
                   3E−05 
                 2.426E−05 
                 1.6947 
                 0.0530 
                 0.1278 
                 2.6327 
                 0.0072 
                 0.0168 
                 2.9412 
               
               
                 32197 
                 3-(4-hydroxyphenyl)lactate 
                 3.1E−05 
                 2.481E−05 
                 1.2467 
                 0.0049 
                 0.0441 
                 1.8140 
                 0.0241 
                 0.0324 
                 1.3510 
               
               
                 35626 
                 1-myristoylglycerophosphocholine 
                 3.2E−05 
                 2.523E−05 
                 2.3929 
                 0.6642 
                 0.5264 
                 1.0678 
                 0.2114 
                 0.1369 
                 1.2595 
               
               
                 35627 
                 1-myristoylglycerophospho- 
                 4.1E−05 
                 3.197E−05 
                 1.7902 
                 0.9700 
                 0.6302 
                 1.0040 
                 0.2896 
                 0.1694 
                 0.9431 
               
               
                   
                 ethanolamine 
               
               
                 35428 
                 tiglyl carnitine 
                 4.7E−05 
                  3.64E−05 
                 1.5202 
                 0.0149 
                 0.0667 
                 1.5850 
                 0.3087 
                 0.1770 
                 1.5909 
               
               
                 3155 
                 3-ureidopropionate 
                 4.8E−05 
                 3.694E−05 
                 0.6847 
                 0.6433 
                 0.5167 
                 1.1618 
                 0.0858 
                 0.0752 
                 1.3666 
               
               
                 32380 
                 nicotinamide adenine dinucleotide 
                 4.9E−05 
                 3.694E−05 
                 2.0890 
                 0.0385 
                 0.1120 
                 0.6852 
                 0.2570 
                 0.1563 
                 0.7039 
               
               
                   
                 phosphate (NADP+) 
               
               
                 33449 
                 adenosine 5′-triphosphate (ATP) 
                 0.0001 
                 4.416E−05 
                 0.6983 
                 0.1912 
                 0.2538 
                 0.7455 
                 0.0000 
                 0.0008 
                 0.4480 
               
               
                 32562 
                 pregnen-diol disulfate 
                 0.0001 
                 0.0001 
                 0.7838 
                 0.3739 
                 0.3876 
                 1.0807 
                 0.5852 
                 0.2779 
                 0.9883 
               
               
                 37538 
                 15-HETE 
                 0.0001 
                 0.0001 
                 1.5407 
                 0.8138 
                 0.5876 
                 1.1126 
                 0.2931 
                 0.1701 
                 1.1631 
               
               
                 37083 
                 alanylproline 
                 0.0001 
                 4.399E−05 
                 1.5274 
                 0.0801 
                 0.1618 
                 2.3264 
                 0.0490 
                 0.0499 
                 2.0471 
               
               
                 37093 
                 alanylleucine 
                 0.0002 
                 0.0001 
                 2.0969 
                 0.4398 
                 0.4227 
                 1.1477 
                 0.0235 
                 0.0324 
                 2.2294 
               
               
                 31591 
                 androsterone sulfate 
                 0.0003 
                 0.0002 
                 0.7987 
                 0.0455 
                 0.1214 
                 1.3709 
                 0.1190 
                 0.0930 
                 1.2336 
               
               
                 32980 
                 adrenate (22:4n6) 
                 0.0003 
                 0.0002 
                 1.2378 
                 0.0236 
                 0.0898 
                 1.2236 
                 0.0000 
                 0.0008 
                 1.4992 
               
               
                 31609 
                 N1-methylguanosine 
                 0.0003 
                 0.0002 
                 1.2092 
                 0.0113 
                 0.0632 
                 1.6745 
                 0.0095 
                 0.0195 
                 1.4342 
               
               
                 35128 
                 ketamine 
                 0.0003 
                 0.0002 
                 1.4679 
                 0.6281 
                 0.5084 
                 1.1403 
                 0.0421 
                 0.0456 
                 1.4483 
               
               
                 35431 
                 2-methylbutyroylcarnitine 
                 0.0003 
                 0.0002 
                 1.3277 
                 0.0011 
                 0.0251 
                 1.5389 
                 0.0686 
                 0.0647 
                 1.3283 
               
               
                 37203 
                 4-androsten-3beta,17beta- 
                 0.0004 
                 0.0003 
                 0.7788 
                 0.4971 
                 0.4470 
                 1.0682 
                 0.8235 
                 0.3584 
                 1.0066 
               
               
                   
                 dioldisulfate 2 
               
               
                 27716 
                 bilirubin (Z,Z) 
                 0.0006 
                 0.0004 
                 0.8097 
                 0.9752 
                 0.6302 
                 0.9625 
                 0.4850 
                 0.2445 
                 0.8558 
               
               
                 34406 
                 valerylcarnitine 
                 0.0007 
                 0.0004 
                 1.2819 
                 0.0542 
                 0.1278 
                 1.2441 
                 0.0194 
                 0.0295 
                 1.4141 
               
               
                 34398 
                 glycylleucine 
                 0.0008 
                 0.0005 
                 1.4343 
                 0.0022 
                 0.0281 
                 2.5922 
                 0.0012 
                 0.0061 
                 2.9299 
               
               
                 37752 
                 13-HODE + 9-HODE 
                 0.0011 
                 0.0006 
                 1.3264 
                 0.2977 
                 0.3378 
                 1.2262 
                 0.0270 
                 0.0351 
                 1.3782 
               
               
                 15821 
                 fucose 
                 0.0011 
                 0.0006 
                 1.4559 
                 0.4180 
                 0.4147 
                 1.1233 
                 0.4588 
                 0.2357 
                 1.2518 
               
               
                 34396 
                 choline phosphate 
                 0.0012 
                 0.0007 
                 1.6764 
                 0.0285 
                 0.0960 
                 0.3963 
                 0.0000 
                 0.0002 
                 0.0430 
               
               
                 34418 
                 cytidine 5′-diphosphocholine 
                 0.0013 
                 0.0007 
                 1.2818 
                 0.3645 
                 0.3834 
                 1.2992 
                 0.3282 
                 0.1846 
                 1.1698 
               
               
                 36602 
                 1-oleoylglycerophosphoinositol 
                 0.0014 
                 0.0008 
                 1.3831 
                 0.3366 
                 0.3660 
                 1.1374 
                 0.0069 
                 0.0164 
                 1.3173 
               
               
                 35628 
                 1-oleoylglycerophospho- 
                 0.0016 
                 0.0009 
                 1.3535 
                 0.5192 
                 0.4561 
                 1.0950 
                 0.0192 
                 0.0295 
                 1.2800 
               
               
                   
                 ethanolamine 
               
               
                 21188 
                 1-stearoylglycerol (1-monostearin) 
                 0.0017 
                 0.0009 
                 1.3353 
                 0.8483 
                 0.5955 
                 1.0815 
                 0.0001 
                 0.0019 
                 1.6631 
               
               
                 1118 
                 arachidate (20:0) 
                 0.0018 
                 0.001 
                 1.3959 
                 0.7790 
                 0.5723 
                 1.0688 
                 0.3320 
                 0.1848 
                 1.2144 
               
               
                 21184 
                 1-oleoylglycerol (1-monoolein) 
                 0.0019 
                 0.001 
                 1.4805 
                 0.7232 
                 0.5530 
                 0.9054 
                 0.2151 
                 0.1377 
                 1.2539 
               
               
                 34656 
                 2-arachidonoylglycerophospho- 
                 0.0024 
                 0.0012 
                 0.7781 
                 0.5602 
                 0.4712 
                 0.9951 
                 0.8264 
                 0.3584 
                 0.9278 
               
               
                   
                 ethanolamine 
               
               
                 1589 
                 N-acetylmethionine 
                 0.0024 
                 0.0012 
                 1.3539 
                 0.0884 
                 0.1700 
                 2.3971 
                 0.0143 
                 0.0251 
                 2.3350 
               
               
                 35687 
                 2-oleoylglycerophospho- 
                 0.0027 
                 0.0013 
                 1.2656 
                 0.5786 
                 0.4819 
                 1.1005 
                 0.2050 
                 0.1341 
                 1.1747 
               
               
                   
                 ethanolamine 
               
               
                 1561 
                 alpha-tocopherol 
                 0.0029 
                 0.0014 
                 1.1977 
                 0.4378 
                 0.4227 
                 0.9839 
                 0.2878 
                 0.1688 
                 1.0442 
               
               
                 32672 
                 pyroglutamine 
                 0.0032 
                 0.0016 
                 0.9551 
                 0.8632 
                 0.6008 
                 1.0450 
                 0.4190 
                 0.2214 
                 0.7556 
               
               
                 20714 
                 methyl-alpha-glucopyranoside 
                 0.0036 
                 0.0017 
                 1.5768 
                 0.3013 
                 0.3392 
                 0.6076 
                 0.3168 
                 0.1801 
                 1.1074 
               
               
                 32379 
                 scyllo-inositol 
                 0.0038 
                 0.0018 
                 0.8927 
                 0.9696 
                 0.6302 
                 0.9992 
                 0.5025 
                 0.2503 
                 1.0929 
               
               
                 32553 
                 phenol sulfate 
                 0.0038 
                 0.0018 
                 0.8015 
                 0.6235 
                 0.5059 
                 1.1328 
                 0.7288 
                 0.3255 
                 0.8787 
               
               
                 31530 
                 threonylphenylalanine 
                 0.0038 
                 0.0018 
                 1.8909 
                 0.5790 
                 0.4819 
                 1.1509 
                 0.0305 
                 0.0376 
                 2.4724 
               
               
                 1497 
                 ethanolamine 
                 0.0042 
                 0.0019 
                 1.2250 
                 0.0055 
                 0.0447 
                 1.2915 
                 0.0000 
                 0.0011 
                 1.5381 
               
               
                 37478 
                 docosapentaenoate 
                 0.0045 
                 0.0021 
                 1.5229 
                 0.0900 
                 0.1711 
                 1.2613 
                 0.0057 
                 0.0148 
                 1.8681 
               
               
                   
                 (n6 DPA; 22:5n6) 
               
               
                 32792 
                 andro steroid monosulfate 2 
                 0.0048 
                 0.0022 
                 0.8343 
                 0.1448 
                 0.2198 
                 1.1349 
                 0.7907 
                 0.3473 
                 0.9667 
               
               
                 18357 
                 glycylvaline 
                 0.0048 
                 0.0022 
                 1.2595 
                 0.0489 
                 0.1235 
                 1.0407 
                 0.0042 
                 0.0133 
                 1.4685 
               
               
                 31260 
                 glucose-6-phosphate (G6P) 
                 0.005 
                 0.0023 
                 0.7327 
                 0.9487 
                 0.6302 
                 0.8770 
                 0.5594 
                 0.2694 
                 0.6543 
               
               
                 18790 
                 acetylcholine 
                 0.0052 
                 0.0024 
                 0.8183 
                 0.2137 
                 0.2783 
                 0.7852 
                 0.0130 
                 0.0243 
                 0.6454 
               
               
                 27447 
                 1-linoleoylglycerol 
                 0.0053 
                 0.0024 
                 1.3016 
                 0.1160 
                 0.1941 
                 1.2719 
                 0.0834 
                 0.0746 
                 1.5063 
               
               
                   
                 (1-monolinolein) 
               
               
                 35159 
                 cysteine-glutathione disulfide 
                 0.0053 
                 0.0024 
                 1.2938 
                 0.0098 
                 0.0586 
                 1.7915 
                 0.0079 
                 0.0175 
                 1.7349 
               
               
                 33970 
                 cis-vaccenate (18:1n7) 
                 0.0054 
                 0.0024 
                 1.2878 
                 0.8072 
                 0.5858 
                 1.0118 
                 0.0607 
                 0.0589 
                 1.3005 
               
               
                 35256 
                 2-arachidonoylglycerophospho- 
                 0.0057 
                 0.0025 
                 0.7951 
                 0.3157 
                 0.3492 
                 1.1561 
                 0.5369 
                 0.2629 
                 0.9577 
               
               
                   
                 choline 
               
               
                 17945 
                 2-hydroxystearate 
                 0.0062 
                 0.0027 
                 1.3275 
                 0.0081 
                 0.0511 
                 1.3578 
                 0.0235 
                 0.0324 
                 1.3367 
               
               
                 32807 
                 taurocholenate sulfate 
                 0.0064 
                 0.0028 
                 0.7875 
                 0.1495 
                 0.2236 
                 1.1135 
                 0.9729 
                 0.3967 
                 0.9418 
               
               
                 36103 
                 p-cresol sulfate 
                 0.0067 
                 0.003 
                 0.7883 
                 0.5740 
                 0.4802 
                 1.2395 
                 0.9884 
                 0.3998 
                 0.9829 
               
               
                 36738 
                 gamma-glutamylglutamate 
                 0.0078 
                 0.0034 
                 1.2069 
                 0.0979 
                 0.1771 
                 1.7623 
                 0.3395 
                 0.1878 
                 1.2287 
               
               
                 27672 
                 3-indoxyl sulfate 
                 0.0086 
                 0.0038 
                 0.4768 
                 0.3137 
                 0.3481 
                 1.2350 
                 0.3332 
                 0.1848 
                 1.2041 
               
               
                 34585 
                 4-hydroxybutyrate (GHB) 
                 0.0107 
                 0.0046 
                 1.4057 
                 0.2391 
                 0.2981 
                 1.2594 
                 0.0134 
                 0.0243 
                 1.9568 
               
               
                 19503 
                 stearoyl sphingomyelin 
                 0.012 
                 0.0051 
                 0.8476 
                 0.7864 
                 0.5758 
                 0.9210 
                 0.3437 
                 0.1894 
                 0.9947 
               
               
                 12102 
                 phosphoethabolamine 
                 0.0124 
                 0.0053 
                 1.4084 
                 0.4304 
                 0.4201 
                 0.7657 
                 0.0056 
                 0.0148 
                 0.1747 
               
               
                 35186 
                 1-arachidonoylglycerophospho- 
                 0.0126 
                 0.0054 
                 0.9210 
                 0.1578 
                 0.2285 
                 1.1177 
                 0.2446 
                 0.1503 
                 1.0836 
               
               
                   
                 ethanolamine 
               
               
                 27727 
                 glutathione, oxidized (GSSG) 
                 0.0132 
                 0.0055 
                 0.9154 
                 0.3395 
                 0.3679 
                 0.9189 
                 0.3655 
                 0.1983 
                 0.8581 
               
               
                 37418 
                 1-pentadecanoylglycero- 
                 0.0144 
                 0.006 
                 1.4082 
                 0.9776 
                 0.6302 
                 1.2763 
                 0.1024 
                 0.0841 
                 1.6725 
               
               
                   
                 phosphocholine 
               
               
                 35320 
                 catechol sulfate 
                 0.0145 
                 0.006 
                 0.5918 
                 0.4747 
                 0.4390 
                 1.4079 
                 0.8987 
                 0.3793 
                 0.8798 
               
               
                 37190 
                 5alpha-androstan-3beta,17beta-diol 
                 0.0152 
                 0.0062 
                 0.8095 
                 0.2861 
                 0.3323 
                 1.3632 
                 0.5289 
                 0.2615 
                 1.3012 
               
               
                   
                 disulfate 
               
               
                 33935 
                 piperine 
                 0.02 
                 0.008 
                 1.1046 
                 0.6552 
                 0.5226 
                 0.9456 
                 0.3512 
                 0.1917 
                 1.1069 
               
               
                 35631 
                 1-palmitoylglycerophospho- 
                 0.0216 
                 0.0085 
                 1.1989 
                 0.3337 
                 0.3640 
                 1.1561 
                 0.0157 
                 0.0261 
                 1.2994 
               
               
                   
                 ethanolamine 
               
               
                 12110 
                 isocitrate 
                 0.0221 
                 0.0087 
                 0.8190 
                 0.7406 
                 0.5588 
                 0.9831 
                 0.5637 
                 0.2705 
                 1.0695 
               
               
                 34407 
                 isovalerylcarnitine 
                 0.0226 
                 0.0089 
                 1.3073 
                 0.0089 
                 0.0555 
                 1.5247 
                 0.1406 
                 0.1031 
                 1.3621 
               
               
                 27738 
                 threonate 
                 0.0252 
                 0.0098 
                 0.5796 
                 0.2981 
                 0.3378 
                 0.8809 
                 0.1986 
                 0.1315 
                 1.1153 
               
               
                 34258 
                 2-docosahexaenoylglycero- 
                 0.0257 
                 0.01 
                 0.8530 
                 0.4578 
                 0.4310 
                 0.9878 
                 0.7711 
                 0.3409 
                 0.8962 
               
               
                   
                 phosphoethanolamine 
               
               
                 32506 
                 2-linoleoylglycerol 
                 0.0269 
                 0.0104 
                 1.2801 
                 0.0293 
                 0.0964 
                 1.3141 
                 0.0252 
                 0.0335 
                 1.4894 
               
               
                   
                 (2-monolinolein) 
               
               
                 36808 
                 dimethylarginine 
                 0.0289 
                 0.0111 
                 1.2149 
                 0.7786 
                 0.5723 
                 0.9359 
                 0.9511 
                 0.3901 
                 0.9653 
               
               
                   
                 (SDMA + ADMA) 
               
               
                 37496 
                 N-acetylputrescine 
                 0.0327 
                 0.0123 
                 0.7432 
                 0.5030 
                 0.4470 
                 0.8853 
                 0.0803 
                 0.0731 
                 1.3542 
               
               
                 18369 
                 gamma-glutamylleucine 
                 0.0336 
                 0.0126 
                 1.2712 
                 0.0649 
                 0.1445 
                 1.4672 
                 0.0959 
                 0.0801 
                 1.2004 
               
               
                 31787 
                 3-carboxy-4-methyl-5-propyl-2- 
                 0.0363 
                 0.0135 
                 0.8852 
                 0.9592 
                 0.6302 
                 0.9358 
                 0.3935 
                 0.2119 
                 0.8851 
               
               
                   
                 furanpropanoate (CMPF) 
               
               
                 37253 
                 2-hydroxyglutarate 
                 0.0371 
                 0.0138 
                 4.3978 
                 0.9989 
                 0.6362 
                 0.9446 
                 0.4595 
                 0.2357 
                 0.5822 
               
               
                 27718 
                 creatine 
                 0.0377 
                 0.014 
                 0.9427 
                 0.9436 
                 0.6302 
                 0.9917 
                 0.1831 
                 0.1237 
                 1.0718 
               
               
                 12035 
                 pelargonate (9:0) 
                 0.0388 
                 0.0143 
                 1.0872 
                 0.9872 
                 0.6322 
                 0.9928 
                 0.0187 
                 0.0292 
                 0.8775 
               
               
                 37070 
                 methylphosphate 
                 0.0411 
                 0.015 
                 1.0885 
                 0.6486 
                 0.5198 
                 0.8520 
                 0.1230 
                 0.0943 
                 1.1018 
               
               
                 2849 
                 guanosine 5′-monophosphate 
                 0.0561 
                 0.0199 
                 1.1068 
                 0.0430 
                 0.1196 
                 0.5421 
                 0.0051 
                 0.0141 
                 0.5192 
               
               
                   
                 (GMP) 
               
               
                 34214 
                 1-arachidonoylglycero 
                 0.0598 
                 0.021 
                 1.0651 
                 0.1326 
                 0.2079 
                 1.1316 
                 0.0033 
                 0.0121 
                 1.2019 
               
               
                   
                 phosphoinositol 
               
               
                 1585 
                 N-acetylalanine 
                 0.0793 
                 0.0272 
                 1.2058 
                 0.0025 
                 0.0289 
                 2.3072 
                 0.1328 
                 0.0992 
                 1.7737 
               
               
                 34534 
                 laurylcarnitine 
                 0.0964 
                 0.0323 
                 1.5214 
                 0.0810 
                 0.1620 
                 1.3196 
                 0.0307 
                 0.0376 
                 1.4644 
               
               
                 33961 
                 1-stearoylglycerophosphocholine 
                 0.1053 
                 0.0348 
                 1.0164 
                 0.0375 
                 0.1110 
                 1.2052 
                 0.0432 
                 0.0459 
                 1.5333 
               
               
                 32492 
                 caprylate (8:0) 
                 0.1139 
                 0.0373 
                 1.1163 
                 0.9880 
                 0.6322 
                 1.0062 
                 0.0125 
                 0.0235 
                 0.6925 
               
               
                 35255 
                 2-stearoylglycerophosphocholine 
                 0.133 
                 0.043 
                 1.0537 
                 0.0539 
                 0.1278 
                 1.3310 
                 0.0200 
                 0.0300 
                 1.6258 
               
               
                 33441 
                 isobutyrylcarnitine 
                 0.1492 
                 0.0475 
                 0.9942 
                 0.0048 
                 0.0441 
                 1.4825 
                 0.1358 
                 0.1007 
                 1.3567 
               
               
                 35855 
                 ribulose 
                 0.1684 
                 0.0529 
                 1.1928 
                 0.0128 
                 0.0646 
                 2.2699 
                 0.0136 
                 0.0244 
                 1.2616 
               
               
                 33952 
                 myristoylcarnitine 
                 0.1965 
                 0.0604 
                 1.7507 
                 0.0490 
                 0.1235 
                 1.2622 
                 0.0201 
                 0.0300 
                 1.7254 
               
               
                 33958 
                 glycyltyrosine 
                 0.2102 
                 0.0639 
                 1.2061 
                 0.0106 
                 0.0602 
                 2.3342 
                 0.1073 
                 0.0875 
                 2.2093 
               
               
                 35688 
                 2-palmitoylglycerophospho- 
                 0.2263 
                 0.0673 
                 1.1190 
                 0.2626 
                 0.3118 
                 1.2662 
                 0.0421 
                 0.0456 
                 1.2722 
               
               
                   
                 ethanolamine 
               
               
                 34416 
                 1-stearoylglycerophospho- 
                 0.2409 
                 0.0711 
                 1.0623 
                 0.0494 
                 0.1235 
                 1.2114 
                 0.0133 
                 0.0243 
                 1.4767 
               
               
                   
                 ethanolamine 
               
               
                 35637 
                 cysteinylglycine 
                 0.266 
                 0.0779 
                 1.1549 
                 0.3845 
                 0.3942 
                 0.9484 
                 0.0360 
                 0.0414 
                 1.5609 
               
               
                 35137 
                 N2,N2-dimethylguanosine 
                 0.2977 
                 0.0854 
                 0.8784 
                 0.4907 
                 0.4459 
                 1.6160 
                 0.0352 
                 0.0412 
                 1.2912 
               
               
                 36761 
                 isoleucylisoleucine 
                 0.3175 
                 0.0898 
                 1.0246 
                 0.1900 
                 0.2535 
                 0.7555 
                 0.0021 
                 0.0090 
                 1.8901 
               
               
                 35114 
                 7-methylguanine 
                 0.3398 
                 0.0946 
                 0.9146 
                 0.0540 
                 0.1278 
                 1.7909 
                 0.0033 
                 0.0122 
                 1.2966 
               
               
                 35675 
                 2-hydroxypalmitate 
                 0.3815 
                 0.1032 
                 1.1376 
                 0.0349 
                 0.1064 
                 1.2941 
                 0.2574 
                 0.1563 
                 1.3723 
               
               
                 33960 
                 1-oleoylglycerophosphocholine 
                 0.4486 
                 0.1183 
                 1.1937 
                 0.1699 
                 0.2365 
                 1.1700 
                 0.0307 
                 0.0376 
                 1.6297 
               
               
                 32342 
                 adenosine 5′-monophosphate 
                 0.6021 
                 0.1507 
                 0.7935 
                 0.0191 
                 0.0810 
                 0.5291 
                 0.0013 
                 0.0067 
                 0.4850 
               
               
                   
                 (AMP) 
               
               
                 15335 
                 mannitol 
                 0.6702 
                 0.1631 
                 0.8962 
                 0.1857 
                 0.2488 
                 1.4046 
                 0.0207 
                 0.0305 
                 1.2965 
               
               
                 33957 
                 1-heptadecanoylglycero 
                 0.6734 
                 0.1635 
                 1.4504 
                 0.0611 
                 0.1370 
                 1.3593 
                 0.0120 
                 0.0229 
                 1.8571 
               
               
                   
                 phosphocholine 
               
               
                 35160 
                 oleoylcarnitine 
                 0.6903 
                 0.1672 
                 1.3997 
                 0.0145 
                 0.0667 
                 1.4870 
                 0.0037 
                 0.0127 
                 2.0050 
               
               
                 33477 
                 erythronate 
                 0.704 
                 0.1694 
                 0.9496 
                 0.0587 
                 0.1354 
                 1.4643 
                 0.0002 
                 0.0021 
                 1.3726 
               
               
                 35127 
                 pro-hydroxy-pro 
                 0.7314 
                 0.1745 
                 0.9133 
                 0.0877 
                 0.1700 
                 1.5127 
                 0.0403 
                 0.0451 
                 1.1761 
               
               
                 33871 
                 1-eicosadienoylglycero- 
                 0.7961 
                 0.1865 
                 1.2787 
                 0.0901 
                 0.1711 
                 1.2593 
                 0.0110 
                 0.0217 
                 1.8045 
               
               
                   
                 phosphocholine 
               
               
                 34409 
                 stearoylcarnitine 
                 0.9017 
                 0.2057 
                 1.5564 
                 0.0258 
                 0.0942 
                 1.5893 
                 0.0037 
                 0.0127 
                 2.1241 
               
               
                 22189 
                 palmitoylcarnitine 
                 0.9084 
                 0.2064 
                 1.6256 
                 0.0134 
                 0.0657 
                 1.4246 
                 0.0089 
                 0.0185 
                 2.0203 
               
               
                   
               
             
          
         
       
     
         [0000]    
       
         
               
               
               
               
               
               
               
               
               
               
               
             
               
               
               
               
               
               
               
               
               
               
               
             
           
               
                 TABLE 1B 
               
               
                   
               
               
                   
                   
                   
                   
                 Ratio 
                   
                   
                 Ratio 
                   
                   
                   
               
               
                   
                   
                   
                   
                 Cancer 
                 N_NOC 
                 N_NOC 
                 NOC/ 
                 T_NOC 
                 T_NOC 
                 Ratio 
               
               
                   
                   
                   
                   
                 Tumor/ 
                 VS 
                 VS 
                 OC 
                 VS 
                 VS 
                 T_NOC/ 
               
               
                 Comp 
                   
                 C VS T P- 
                 C VS T 
                 Control 
                 N_OC P- 
                 N_OC Q- 
                 Adja- 
                 T_OC P- 
                 T_OC Q- 
                 T_OC 
               
               
                 ID 
                 Name 
                 VALUE 
                 Q-VALUE 
                 (T/C) 
                 VALUE 
                 VALUE 
                 cent 
                 VALUE 
                 VALUE 
                 Tumor 
               
               
                   
               
             
             
               
                   
               
             
          
           
               
                 15500 
                 carnitine 
                 1.4E−12 
                 2.728E−11 
                 1.2543 
                 0.5028 
                 0.4470 
                 1.0387 
                 0.0907 
                 0.0780 
                 1.0933 
               
               
                 1898 
                 proline 
                 5.3E−12 
                 8.588E−11 
                 1.3923 
                 0.0044 
                 0.0441 
                 1.2297 
                 0.0020 
                 0.0086 
                 1.2368 
               
               
                 54 
                 tryptophan 
                 2.9E−11 
                 2.349E−10 
                 1.2512 
                 0.0047 
                 0.0441 
                 1.2270 
                 0.0001 
                 0.0018 
                 1.2947 
               
               
                 32975 
                 taurine 
                 1.4E−10 
                 7.779E−10 
                 0.6409 
                 0.9504 
                 0.6302 
                 1.0364 
                 0.1102 
                 0.0883 
                 0.8222 
               
               
                 1284 
                 threonine 
                 1.9E−10 
                 1.035E−09 
                 1.3993 
                 0.0597 
                 0.1357 
                 1.1837 
                 0.0058 
                 0.0149 
                 1.2350 
               
               
                 606 
                 uridine 
                   3E−10 
                 1.421E−09 
                 1.3379 
                 0.1055 
                 0.1852 
                 1.1128 
                 0.0010 
                 0.0059 
                 1.2784 
               
               
                 60 
                 leucine 
                 4.7E−10 
                 1.897E−09 
                 1.2454 
                 0.0003 
                 0.0155 
                 1.3605 
                 0.0002 
                 0.0024 
                 1.3898 
               
               
                 6146 
                 2-aminoadipate 
                 5.3E−10 
                 2.085E−09 
                 1.6525 
                 0.3913 
                 0.3961 
                 0.8972 
                 0.3144 
                 0.1794 
                 1.0246 
               
               
                 1359 
                 oleate (18:1n9) 
                 8.1E−10 
                 2.959E−09 
                 1.4134 
                 0.7704 
                 0.5721 
                 1.0609 
                 0.0049 
                 0.0141 
                 1.3210 
               
               
                 1419 
                 5-methylthioadenosine (MTA) 
                 2.1E−09 
                 6.647E−09 
                 1.5658 
                 0.9395 
                 0.6302 
                 1.0373 
                 0.2711 
                 0.1613 
                 1.1081 
               
               
                 64 
                 phenylalanine 
                 2.9E−09 
                  8.43E−09 
                 1.2459 
                 0.0029 
                 0.0318 
                 1.4145 
                 0.0016 
                 0.0076 
                 1.4104 
               
               
                 1299 
                 tyrosine 
                 4.3E−09 
                  1.11E−08 
                 1.2343 
                 0.0038 
                 0.0392 
                 1.5168 
                 0.0028 
                 0.0108 
                 1.4687 
               
               
                 11777 
                 glycine 
                 4.7E−09 
                 1.162E−08 
                 1.3676 
                 0.0340 
                 0.1064 
                 1.1579 
                 0.0299 
                 0.0376 
                 1.1764 
               
               
                 1105 
                 linoleate (18:2n6) 
                   1E−08 
                 2.266E−08 
                 1.4084 
                 0.0008 
                 0.0204 
                 1.4241 
                 0.0010 
                 0.0057 
                 1.4434 
               
               
                 513 
                 creatinine 
                 1.2E−08 
                 2.573E−08 
                 0.7005 
                 0.5418 
                 0.4666 
                 1.2272 
                 0.6213 
                 0.2896 
                 0.9873 
               
               
                 2766 
                 N-acetylgalactosamine 
                 1.3E−08 
                 2.723E−08 
                 2.0376 
                 0.4920 
                 0.4459 
                 1.3785 
                 0.2991 
                 0.1719 
                 1.3636 
               
               
                 1494 
                 5-oxoproline 
                 3.2E−08 
                  5.87E−08 
                 1.3941 
                 0.0151 
                 0.0670 
                 1.6118 
                 0.0572 
                 0.0560 
                 1.3254 
               
               
                 605 
                 uracil 
                 3.9E−08 
                 6.966E−08 
                 1.8625 
                 0.0006 
                 0.0204 
                 1.8463 
                 0.0003 
                 0.0027 
                 2.0160 
               
               
                 15365 
                 glycerol 3-phosphate (G3P) 
                 6.5E−08 
                 1.093E−07 
                 1.4659 
                 0.1355 
                 0.2103 
                 0.8200 
                 0.8962 
                 0.3790 
                 0.9890 
               
               
                 35661 
                 lidocaine 
                 6.6E−08 
                 1.102E−07 
                 1.6411 
                 0.2454 
                 0.3014 
                 1.4764 
                 0.0148 
                 0.0254 
                 1.9789 
               
               
                 3127 
                 hypoxanthine 
                 8.4E−08 
                 1.378E−07 
                 1.3214 
                 0.0000 
                 0.0028 
                 1.5438 
                 0.0003 
                 0.0028 
                 1.3975 
               
               
                 15990 
                 glycerophosphorylcholine (GPC) 
                 8.5E−08 
                 1.378E−07 
                 1.5443 
                 0.3578 
                 0.3800 
                 0.9399 
                 0.5657 
                 0.2705 
                 1.0659 
               
               
                 15136 
                 xanthosine 
                 9.1E−08 
                 1.437E−07 
                 1.9673 
                 0.0324 
                 0.1027 
                 1.5805 
                 0.0415 
                 0.0456 
                 1.3590 
               
               
                 15948 
                 S-adenosylhomocysteine (SAH) 
                 9.9E−08 
                 1.517E−07 
                 1.2312 
                 0.0007 
                 0.0204 
                 1.3262 
                 0.0082 
                 0.0175 
                 1.2211 
               
               
                 31453 
                 cysteine 
                 1.3E−07 
                 1.851E−07 
                 1.9429 
                 0.0066 
                 0.0499 
                 1.3016 
                 0.0025 
                 0.0102 
                 1.7826 
               
               
                 15096 
                 N-acetylglucosamine 
                 1.4E−07 
                 2.019E−07 
                 2.4319 
                 0.1096 
                 0.1908 
                 1.8005 
                 0.0239 
                 0.0324 
                 1.7337 
               
               
                 33447 
                 palmitoleate (16:1n7) 
                   4E−07 
                 5.071E−07 
                 1.2929 
                 0.0466 
                 0.1223 
                 1.2014 
                 0.1757 
                 0.1203 
                 1.1595 
               
               
                 1649 
                 valine 
                 4.3E−07 
                 5.377E−07 
                 1.1475 
                 0.0014 
                 0.0262 
                 1.2464 
                 0.0007 
                 0.0044 
                 1.2735 
               
               
                 554 
                 adenine 
                 4.7E−07 
                  5.79E−07 
                 1.4385 
                 0.3697 
                 0.3864 
                 1.0740 
                 0.0587 
                 0.0572 
                 1.1661 
               
               
                 1508 
                 pantothenate 
                 4.8E−07 
                 5.834E−07 
                 1.1803 
                 0.0303 
                 0.0983 
                 1.3433 
                 0.0062 
                 0.0156 
                 1.3840 
               
               
                 1302 
                 methionine 
                 5.8E−07 
                 6.937E−07 
                 1.2140 
                 0.0211 
                 0.0850 
                 1.6127 
                 0.0569 
                 0.0560 
                 1.5274 
               
               
                 1648 
                 serine 
                 1.4E−06 
                 1.517E−06 
                 1.3310 
                 0.0919 
                 0.1716 
                 1.2496 
                 0.0144 
                 0.0251 
                 1.3279 
               
               
                 1493 
                 ornithine 
                 2.4E−06 
                 2.597E−06 
                 1.5806 
                 0.8271 
                 0.5903 
                 1.2284 
                 0.5654 
                 0.2705 
                 1.1409 
               
               
                 1125 
                 isoleucine 
                 2.6E−06 
                 2.795E−06 
                 1.1602 
                 0.0002 
                 0.0149 
                 1.4599 
                 0.0006 
                 0.0044 
                 1.3873 
               
               
                 59 
                 histidine 
                 3.1E−06 
                 3.233E−06 
                 1.1863 
                 0.0098 
                 0.0586 
                 1.1381 
                 0.0075 
                 0.0171 
                 1.1625 
               
               
                 1303 
                 malate 
                 3.2E−06 
                 3.314E−06 
                 1.4488 
                 0.0692 
                 0.1490 
                 1.1460 
                 0.0050 
                 0.0141 
                 1.3388 
               
               
                 1126 
                 alanine 
                 3.4E−06 
                 3.473E−06 
                 1.3058 
                 0.1843 
                 0.2479 
                 1.1000 
                 0.0112 
                 0.0218 
                 1.1653 
               
               
                 1604 
                 urate 
                   5E−06 
                 4.861E−06 
                 0.8080 
                 0.1322 
                 0.2079 
                 1.1180 
                 0.9965 
                 0.4023 
                 0.9742 
               
               
                 1336 
                 palmitate (16:0) 
                 6.2E−06 
                 5.928E−06 
                 1.1014 
                 0.0023 
                 0.0281 
                 1.1677 
                 0.0051 
                 0.0141 
                 1.1558 
               
               
                 514 
                 cytidine 
                 7.2E−06 
                 6.756E−06 
                 1.5003 
                 0.0248 
                 0.0920 
                 0.6040 
                 0.8239 
                 0.3584 
                 1.1129 
               
               
                 1444 
                 pipecolate 
                   1E−05 
                 9.154E−06 
                 1.2978 
                 0.2277 
                 0.2884 
                 1.2135 
                 0.0307 
                 0.0376 
                 1.2782 
               
               
                 1110 
                 arachidonate (20:4n6) 
                 1.1E−05 
                 9.775E−06 
                 1.2443 
                 0.0669 
                 0.1459 
                 1.1853 
                 0.0022 
                 0.0091 
                 1.2709 
               
               
                 15996 
                 aspartate 
                   3E−05 
                 2.422E−05 
                 1.2468 
                 0.1315 
                 0.2079 
                 1.1669 
                 0.2381 
                 0.1472 
                 1.1181 
               
               
                 1558 
                 4-acetamidobutanoate 
                 0.0001 
                 4.178E−05 
                 0.7334 
                 0.8365 
                 0.5917 
                 1.2146 
                 0.5996 
                 0.2820 
                 1.0557 
               
               
                 32425 
                 dehydroisoandrosterone 
                 0.0001 
                 0.0001 
                 0.8013 
                 0.3882 
                 0.3955 
                 1.0169 
                 0.6068 
                 0.2842 
                 0.9558 
               
               
                   
                 sulfate (DHEA-S) 
               
               
                 1366 
                 trans-4-hydroxyproline 
                 0.0001 
                 0.0001 
                 1.2889 
                 0.3749 
                 0.3876 
                 0.8460 
                 0.8738 
                 0.3726 
                 0.9048 
               
               
                 12083 
                 ribose 
                 0.0001 
                 0.0001 
                 1.3406 
                 0.0015 
                 0.0262 
                 1.8538 
                 0.0002 
                 0.0021 
                 1.8022 
               
               
                 15915 
                 S-adenosylmethionine (SAM) 
                 0.0001 
                 0.0001 
                 1.5259 
                 0.9543 
                 0.6302 
                 1.0203 
                 0.0765 
                 0.0702 
                 1.1843 
               
               
                 11398 
                 asparagine 
                 0.0001 
                 0.0001 
                 1.4370 
                 0.0125 
                 0.0646 
                 1.3092 
                 0.0958 
                 0.0801 
                 1.2640 
               
               
                 22185 
                 N-acetylaspartate (NAA) 
                 0.0001 
                 0.0001 
                 1.5287 
                 0.0495 
                 0.1235 
                 1.2414 
                 0.0650 
                 0.0622 
                 1.1846 
               
               
                 1592 
                 N-acetylneuraminate 
                 0.0001 
                 0.0001 
                 1.8207 
                 0.8843 
                 0.6115 
                 1.0967 
                 0.3229 
                 0.1825 
                 1.0969 
               
               
                 53 
                 glutamine 
                 0.0002 
                 0.0002 
                 1.1291 
                 0.7241 
                 0.5530 
                 1.0155 
                 0.0183 
                 0.0287 
                 1.1182 
               
               
                 19934 
                 myo-inositol 
                 0.0003 
                 0.0002 
                 0.9093 
                 0.9886 
                 0.6322 
                 0.9898 
                 0.1048 
                 0.0858 
                 1.0898 
               
               
                 36984 
                 Isobar: fructose 1,6-diphosphate, 
                 0.0004 
                 0.0002 
                 0.6610 
                 0.5238 
                 0.4572 
                 0.7740 
                 0.0008 
                 0.0051 
                 0.3565 
               
               
                   
                 glucose 1,6-diphosphate 
               
               
                 4966 
                 xylitol 
                 0.0004 
                 0.0002 
                 1.2413 
                 0.0290 
                 0.0964 
                 1.5918 
                 0.0001 
                 0.0019 
                 1.8129 
               
               
                 1559 
                 5,6-dihydrouracil 
                 0.0005 
                 0.0003 
                 1.4286 
                 0.0051 
                 0.0441 
                 1.5557 
                 0.0073 
                 0.0171 
                 1.4564 
               
               
                 35133 
                 N2-methylguanosine 
                 0.0007 
                 0.0004 
                 1.2336 
                 0.2199 
                 0.2840 
                 1.2910 
                 0.0758 
                 0.0698 
                 1.2182 
               
               
                 1827 
                 riboflavin (Vitamin B2) 
                 0.0007 
                 0.0004 
                 1.2503 
                 0.0482 
                 0.1235 
                 1.5993 
                 0.0259 
                 0.0342 
                 1.5053 
               
               
                 2132 
                 citrulline 
                 0.0008 
                 0.0004 
                 1.3540 
                 0.4104 
                 0.4103 
                 0.9233 
                 0.3324 
                 0.1848 
                 0.8118 
               
               
                 57 
                 glutamate 
                 0.0011 
                 0.0006 
                 1.0944 
                 0.1128 
                 0.1909 
                 1.0538 
                 0.0209 
                 0.0305 
                 1.1244 
               
               
                 1365 
                 myristate (14:0) 
                 0.0011 
                 0.0006 
                 1.0944 
                 0.4987 
                 0.4470 
                 1.0207 
                 0.2218 
                 0.1401 
                 0.9406 
               
               
                 2856 
                 uridine 5′-monophosphate (UMP) 
                 0.0014 
                 0.0008 
                 0.7520 
                 0.0071 
                 0.0510 
                 0.4653 
                 0.0003 
                 0.0027 
                 0.2778 
               
               
                 37059 
                 malonylcarnitine 
                 0.0016 
                 0.0009 
                 1.3228 
                 0.1931 
                 0.2546 
                 1.1975 
                 0.2659 
                 0.1587 
                 1.1999 
               
               
                 1516 
                 sarcosine (N-Methylglycine) 
                 0.0018 
                 0.001 
                 1.6614 
                 0.1669 
                 0.2344 
                 1.0950 
                 0.2567 
                 0.1563 
                 1.0535 
               
               
                 1643 
                 fumarate 
                 0.0019 
                 0.001 
                 1.3148 
                 0.0120 
                 0.0646 
                 1.2963 
                 0.6971 
                 0.3190 
                 0.9504 
               
               
                 2372 
                 cytidine 5′-monophosphate 
                 0.0021 
                 0.0011 
                 1.1698 
                 0.4186 
                 0.4147 
                 0.8668 
                 0.0939 
                 0.0801 
                 0.8575 
               
               
                   
                 (5′-CMP) 
               
               
                 527 
                 lactate 
                 0.0022 
                 0.0011 
                 1.0960 
                 0.2045 
                 0.2673 
                 1.1001 
                 0.1096 
                 0.0883 
                 1.1083 
               
               
                 1437 
                 succinate 
                 0.0025 
                 0.0013 
                 1.2840 
                 0.8469 
                 0.5955 
                 1.0776 
                 0.0282 
                 0.0364 
                 1.3244 
               
               
                 1566 
                 3-aminoisobutyrate 
                 0.0026 
                 0.0013 
                 1.2252 
                 0.5469 
                 0.4677 
                 1.9943 
                 0.7731 
                 0.3410 
                 0.8360 
               
               
                 15122 
                 glycerol 
                 0.0029 
                 0.0014 
                 1.1375 
                 0.0012 
                 0.0251 
                 1.2338 
                 0.0001 
                 0.0016 
                 1.3959 
               
               
                 1121 
                 margarate (17:0) 
                 0.009 
                 0.0039 
                 1.1160 
                 0.0025 
                 0.0289 
                 1.1720 
                 0.0469 
                 0.0483 
                 1.1391 
               
               
                 12055 
                 galactose 
                 0.0096 
                 0.0042 
                 1.2630 
                 0.0187 
                 0.0805 
                 1.2559 
                 0.0006 
                 0.0044 
                 1.4373 
               
               
                 5278 
                 nicotinamide adenine 
                 0.0131 
                 0.0055 
                 1.7379 
                 0.0203 
                 0.0832 
                 0.5728 
                 0.1203 
                 0.0933 
                 0.7146 
               
               
                   
                 dinucleotide (NAD+) 
               
               
                 15140 
                 kynurenine 
                 0.0134 
                 0.0056 
                 1.3670 
                 0.0264 
                 0.0942 
                 1.6187 
                 0.2937 
                 0.1701 
                 1.0544 
               
               
                 32328 
                 hexanoylcarnitine 
                 0.0188 
                 0.0076 
                 1.2092 
                 0.2526 
                 0.3053 
                 1.1399 
                 0.0425 
                 0.0456 
                 1.3009 
               
               
                 1574 
                 histamine 
                 0.0191 
                 0.0077 
                 1.1915 
                 0.2213 
                 0.2847 
                 0.9140 
                 0.8185 
                 0.3580 
                 0.9509 
               
               
                 1572 
                 glycerate 
                 0.0288 
                 0.0111 
                 1.0776 
                 0.0073 
                 0.0510 
                 2.0098 
                 0.0067 
                 0.0162 
                 1.9263 
               
               
                 11438 
                 phosphate 
                 0.0323 
                 0.0122 
                 1.1204 
                 0.3605 
                 0.3816 
                 1.1112 
                 0.4569 
                 0.2356 
                 1.0013 
               
               
                 63 
                 cholesterol 
                 0.0401 
                 0.0147 
                 1.0525 
                 0.6863 
                 0.5367 
                 1.0132 
                 0.6003 
                 0.2820 
                 0.9845 
               
               
                 15753 
                 hippurate 
                 0.043 
                 0.0156 
                 0.3444 
                 0.3429 
                 0.3691 
                 2.7235 
                 0.4425 
                 0.2305 
                 2.1170 
               
               
                 15053 
                 sorbitol 
                 0.0513 
                 0.0185 
                 1.3776 
                 0.3475 
                 0.3703 
                 1.0848 
                 0.0053 
                 0.0144 
                 1.4424 
               
               
                 590 
                 hypotaurine 
                 0.0541 
                 0.0193 
                 1.1282 
                 0.8236 
                 0.5903 
                 0.9378 
                 0.9161 
                 0.3806 
                 1.0150 
               
               
                 37506 
                 palmitoyl sphingomyelin 
                 0.0544 
                 0.0194 
                 1.0672 
                 0.1479 
                 0.2221 
                 0.9104 
                 0.6280 
                 0.2921 
                 1.0152 
               
               
                 35153 
                 1-docosahexaenoylglycerol (1- 
                 0.0572 
                 0.0203 
                 1.3518 
                 0.7668 
                 0.5719 
                 0.9644 
                 0.1369 
                 0.1012 
                 1.2366 
               
               
                   
                 monodocosahexaenoin) 
               
               
                 594 
                 nicotinamide 
                 0.0625 
                 0.0219 
                 1.0741 
                 0.0791 
                 0.1606 
                 1.0855 
                 0.0074 
                 0.0171 
                 1.1850 
               
               
                 27743 
                 triethyleneglycol 
                 0.0642 
                 0.0225 
                 0.9022 
                 0.7707 
                 0.5721 
                 0.9372 
                 0.4665 
                 0.2387 
                 0.9358 
               
               
                 32418 
                 myristoleate (14:1n5) 
                 0.0967 
                 0.0323 
                 1.1350 
                 0.0513 
                 0.1264 
                 1.1489 
                 0.5749 
                 0.2743 
                 0.9037 
               
               
                 1414 
                 3-phosphoglycerate 
                 0.0982 
                 0.0327 
                 0.7285 
                 0.9125 
                 0.6222 
                 1.0783 
                 0.1583 
                 0.1127 
                 0.5864 
               
               
                 33936 
                 octanoylcarnitine 
                 0.0987 
                 0.0328 
                 0.9296 
                 0.7781 
                 0.5723 
                 1.0674 
                 0.5247 
                 0.2600 
                 1.1034 
               
               
                 35665 
                 N-acetyl-aspartyl-glutamate 
                 0.11 
                 0.0363 
                 1.1213 
                 0.1111 
                 0.1909 
                 1.1907 
                 0.2628 
                 0.1574 
                 1.1697 
               
               
                   
                 (NAAG) 
               
               
                 34592 
                 ophthalmate 
                 0.1109 
                 0.0364 
                 0.9763 
                 0.6946 
                 0.5404 
                 0.9727 
                 0.9168 
                 0.3806 
                 1.0580 
               
               
                 36776 
                 7-alpha-hydroxy-3-oxo-4- 
                 0.1255 
                 0.0408 
                 1.1101 
                 0.3455 
                 0.3696 
                 1.1728 
                 0.5488 
                 0.2674 
                 0.8501 
               
               
                   
                 cholestenoate (7-Hoca) 
               
               
                 35253 
                 2-palmitoylglycerophosphocholine 
                 0.1263 
                 0.0409 
                 1.0446 
                 0.0884 
                 0.1700 
                 1.2786 
                 0.1172 
                 0.0928 
                 1.7129 
               
               
                 33230 
                 1-palmitoleoylglycero- 
                 0.1358 
                 0.0437 
                 1.5581 
                 0.4553 
                 0.4299 
                 1.1682 
                 0.0742 
                 0.0693 
                 1.4246 
               
               
                   
                 phosphocholine 
               
               
                 32675 
                 C-glycosyltryptophan 
                 0.1373 
                 0.0441 
                 1.1181 
                 0.3456 
                 0.3696 
                 1.0446 
                 0.2867 
                 0.1686 
                 1.0904 
               
               
                 35638 
                 xylonate 
                 0.1576 
                 0.0498 
                 0.8350 
                 0.5519 
                 0.4690 
                 1.0363 
                 0.1676 
                 0.1171 
                 1.2666 
               
               
                 34875 
                 2-docosapentaenoylglycero- 
                 0.1576 
                 0.0498 
                 0.8239 
                 0.4444 
                 0.4246 
                 0.9173 
                 0.7033 
                 0.3199 
                 0.7928 
               
               
                   
                 phosphoethanolamine 
               
               
                 15496 
                 agmatine 
                 0.1632 
                 0.0514 
                 1.0578 
                 0.5715 
                 0.4793 
                 2.2947 
                 0.1774 
                 0.1211 
                 1.3248 
               
               
                 1358 
                 stearate (18:0) 
                 0.1712 
                 0.0536 
                 1.0409 
                 0.0045 
                 0.0441 
                 1.1534 
                 0.0008 
                 0.0053 
                 1.2131 
               
               
                 18371 
                 GDP-mannose 
                 0.1776 
                 0.0553 
                 1.1528 
                 0.8376 
                 0.5917 
                 0.9405 
                 0.4462 
                 0.2312 
                 1.0278 
               
               
                 35884 
                 2-eicosatrienoylglycero- 
                 0.1794 
                 0.0556 
                 1.0390 
                 0.3172 
                 0.3496 
                 1.5740 
                 0.2154 
                 0.1377 
                 1.4313 
               
               
                   
                 phosphocholine 
               
               
                 2342 
                 serotonin (5HT) 
                 0.2002 
                 0.0612 
                 0.8725 
                 0.7476 
                 0.5607 
                 0.8934 
                 0.5892 
                 0.2792 
                 1.0478 
               
               
                 33955 
                 1-palmitoylglycerophosphocholine 
                 0.2237 
                 0.0672 
                 1.0422 
                 0.0665 
                 0.1459 
                 1.1209 
                 0.0636 
                 0.0611 
                 1.3890 
               
               
                 35257 
                 2-linoleoylglycerophosphocholine 
                 0.2247 
                 0.0672 
                 1.0362 
                 0.1207 
                 0.1999 
                 1.2879 
                 0.1919 
                 0.1286 
                 1.2142 
               
               
                 20488 
                 glucose 
                 0.2257 
                 0.0673 
                 0.8800 
                 0.2625 
                 0.3118 
                 0.8116 
                 0.3295 
                 0.1847 
                 1.1816 
               
               
                 2730 
                 gamma-glutamylglutamine 
                 0.2538 
                 0.0745 
                 1.1874 
                 0.6071 
                 0.4989 
                 0.9534 
                 0.8343 
                 0.3610 
                 0.7011 
               
               
                 485 
                 spermidine 
                 0.2714 
                 0.0788 
                 1.6646 
                 0.1527 
                 0.2263 
                 0.7318 
                 0.2606 
                 0.1573 
                 1.3201 
               
               
                 32394 
                 pyroglutamylvaline 
                 0.2724 
                 0.0788 
                 0.9394 
                 0.5280 
                 0.4582 
                 0.6961 
                 0.0855 
                 0.0752 
                 1.4176 
               
               
                 1573 
                 guanosine 
                 0.2856 
                 0.0824 
                 0.9741 
                 0.9621 
                 0.6302 
                 0.9953 
                 0.1314 
                 0.0990 
                 1.0567 
               
               
                 15488 
                 acetylphosphate 
                 0.2907 
                 0.0836 
                 1.0627 
                 0.2663 
                 0.3150 
                 0.9202 
                 0.2037 
                 0.1341 
                 0.8871 
               
               
                 35126 
                 phenylacetylglutamine 
                 0.3003 
                 0.086 
                 0.3934 
                 0.2904 
                 0.3349 
                 2.2048 
                 0.2243 
                 0.1408 
                 1.8656 
               
               
                 34410 
                 cytidine-5′-diphosphoethanolamine 
                 0.3077 
                 0.0879 
                 1.0042 
                 0.4929 
                 0.4459 
                 0.9996 
                 0.2148 
                 0.1377 
                 0.8323 
               
               
                 34419 
                 1-linoleoylglycerophosphocholine 
                 0.3093 
                 0.0881 
                 1.0467 
                 0.3624 
                 0.3825 
                 1.2830 
                 0.1375 
                 0.1012 
                 1.5680 
               
               
                 15705 
                 cystathionine 
                 0.3259 
                 0.0917 
                 1.1588 
                 0.1648 
                 0.2324 
                 0.8484 
                 0.1815 
                 0.1230 
                 1.2588 
               
               
                 542 
                 3-hydroxybutyrate (BHBA) 
                 0.335 
                 0.0938 
                 1.0394 
                 0.6978 
                 0.5404 
                 1.3721 
                 0.8846 
                 0.3764 
                 1.3159 
               
               
                 55 
                 beta-alanine 
                 0.3465 
                 0.0961 
                 0.9366 
                 0.2596 
                 0.3105 
                 1.2564 
                 0.9808 
                 0.3983 
                 0.8974 
               
               
                 569 
                 caffeine 
                 0.3492 
                 0.0963 
                 0.9603 
                 0.9852 
                 0.6322 
                 1.3570 
                 0.7264 
                 0.3255 
                 1.3668 
               
               
                 37475 
                 4-acetaminophen sulfate 
                 0.3594 
                 0.0982 
                 0.8691 
                 0.4377 
                 0.4227 
                 1.1959 
                 0.7216 
                 0.3253 
                 1.3768 
               
               
                 33420 
                 gamma-tocopherol 
                 0.3751 
                 0.102 
                 1.0016 
                 0.1585 
                 0.2285 
                 1.4161 
                 0.2057 
                 0.1341 
                 0.8640 
               
               
                 17747 
                 sphingosine 
                 0.3771 
                 0.1023 
                 1.3711 
                 0.2878 
                 0.3331 
                 1.0895 
                 0.1104 
                 0.0883 
                 1.3760 
               
               
                 15650 
                 N1-methyladenosine 
                 0.3855 
                 0.1038 
                 1.0138 
                 0.1452 
                 0.2198 
                 1.1624 
                 0.0049 
                 0.0141 
                 1.1902 
               
               
                 599 
                 pyruvate 
                 0.3873 
                 0.1039 
                 1.1099 
                 0.5588 
                 0.4712 
                 1.1298 
                 0.2401 
                 0.1480 
                 0.8845 
               
               
                 35819 
                 2-palmitoleoylglycero- 
                 0.407 
                 0.1086 
                 1.1112 
                 0.6094 
                 0.4992 
                 1.0559 
                 0.5325 
                 0.2620 
                 0.9220 
               
               
                   
                 phosphocholine 
               
               
                 587 
                 gluconate 
                 0.4457 
                 0.1178 
                 0.8638 
                 0.1217 
                 0.2004 
                 0.7099 
                 0.2239 
                 0.1408 
                 0.8142 
               
               
                 35174 
                 mead acid (20:3n9) 
                 0.4507 
                 0.1186 
                 1.6095 
                 0.4894 
                 0.4459 
                 0.7850 
                 0.3147 
                 0.1794 
                 0.7945 
               
               
                 577 
                 fructose 
                 0.4691 
                 0.1228 
                 1.0157 
                 0.4277 
                 0.4198 
                 1.0615 
                 0.0045 
                 0.0140 
                 1.2793 
               
               
                 584 
                 mannose 
                 0.4831 
                 0.1256 
                 1.0744 
                 0.8389 
                 0.5917 
                 0.9744 
                 0.0037 
                 0.0127 
                 1.2834 
               
               
                 15806 
                 maltose 
                 0.5027 
                 0.1301 
                 1.0284 
                 0.1129 
                 0.1909 
                 1.2530 
                 0.4100 
                 0.2174 
                 1.4206 
               
               
                 18392 
                 theobromine 
                 0.5097 
                 0.1316 
                 0.9932 
                 0.8970 
                 0.6150 
                 1.2033 
                 0.9859 
                 0.3996 
                 1.2210 
               
               
                 1416 
                 gamma-aminobutyrate (GABA) 
                 0.5183 
                 0.1332 
                 0.9446 
                 0.1783 
                 0.2430 
                 1.3029 
                 0.1174 
                 0.0928 
                 1.4597 
               
               
                 32352 
                 guanine 
                 0.548 
                 0.1384 
                 1.0322 
                 0.0005 
                 0.0204 
                 1.3769 
                 0.2783 
                 0.1651 
                 1.2774 
               
               
                 35623 
                 1-arachidoylglycerophosphocholine 
                 0.5483 
                 0.1384 
                 1.1119 
                 0.9773 
                 0.6302 
                 0.9718 
                 0.2587 
                 0.1566 
                 1.4464 
               
               
                 1564 
                 citrate 
                 0.5553 
                 0.1399 
                 1.0084 
                 0.3903 
                 0.3961 
                 0.8374 
                 0.0949 
                 0.0801 
                 1.0972 
               
               
                 33442 
                 pseudouridine 
                 0.5749 
                 0.1445 
                 0.8560 
                 0.4023 
                 0.4047 
                 1.2958 
                 0.0629 
                 0.0608 
                 1.1218 
               
               
                 37063 
                 gamma-glutamylalanine 
                 0.5844 
                 0.1466 
                 1.2530 
                 0.3037 
                 0.3394 
                 1.0456 
                 0.1923 
                 0.1286 
                 0.6337 
               
               
                 555 
                 adenosine 
                 0.6033 
                 0.1507 
                 0.9109 
                 0.0020 
                 0.0281 
                 0.2716 
                 0.0014 
                 0.0069 
                 0.3267 
               
               
                 1642 
                 caprate (10:0) 
                 0.6071 
                 0.1513 
                 1.0349 
                 0.6324 
                 0.5105 
                 0.9736 
                 0.1426 
                 0.1042 
                 0.9005 
               
               
                 2127 
                 glutathione, reduced (GSH) 
                 0.6168 
                 0.1531 
                 1.0221 
                 0.2153 
                 0.2792 
                 0.9604 
                 0.9483 
                 0.3900 
                 1.1482 
               
               
                 20675 
                 1,5-anhydroglucitol (1,5-AG) 
                 0.6212 
                 0.1538 
                 0.9881 
                 0.4121 
                 0.4108 
                 0.9482 
                 0.9027 
                 0.3802 
                 1.0704 
               
               
                 3147 
                 xanthine 
                 0.628 
                 0.1551 
                 1.2651 
                 0.0283 
                 0.0960 
                 1.2086 
                 0.4887 
                 0.2455 
                 1.2618 
               
               
                 35254 
                 2-oleoylglycerophosphocholine 
                 0.6345 
                 0.1564 
                 1.2696 
                 0.0781 
                 0.1596 
                 1.3242 
                 0.1315 
                 0.0990 
                 1.3567 
               
               
                 603 
                 spermine 
                 0.6612 
                 0.1622 
                 1.0424 
                 0.0993 
                 0.1771 
                 0.6612 
                 0.2365 
                 0.1467 
                 0.8200 
               
               
                 15877 
                 maltotriose 
                 0.6697 
                 0.1631 
                 1.2089 
                 0.2341 
                 0.2930 
                 1.1637 
                 0.9571 
                 0.3918 
                 1.1865 
               
               
                 1123 
                 inosine 
                 0.6703 
                 0.1631 
                 1.0061 
                 0.1627 
                 0.2313 
                 1.0762 
                 0.0021 
                 0.0090 
                 1.1462 
               
               
                 33937 
                 alpha-hydroxyisovalerate 
                 0.6941 
                 0.1674 
                 1.0000 
                 0.0748 
                 0.1578 
                 1.2299 
                 0.9100 
                 0.3806 
                 1.1201 
               
               
                 1670 
                 urea 
                 0.7166 
                 0.1721 
                 1.0283 
                 0.0127 
                 0.0646 
                 1.1853 
                 0.0235 
                 0.0324 
                 1.2021 
               
               
                 1481 
                 inositol 1-phosphate (I1P) 
                 0.7226 
                 0.1732 
                 1.0016 
                 0.2534 
                 0.3053 
                 0.8317 
                 0.1555 
                 0.1117 
                 0.8076 
               
               
                 19266 
                 2-arachidonoyl glycerol 
                 0.756 
                 0.1797 
                 1.0469 
                 0.9683 
                 0.6302 
                 0.9004 
                 0.1203 
                 0.0933 
                 1.2179 
               
               
                 1645 
                 laurate (12:0) 
                 0.7578 
                 0.1797 
                 1.0118 
                 0.0874 
                 0.1700 
                 0.9217 
                 0.0001 
                 0.0019 
                 0.8124 
               
               
                 34397 
                 1-arachidonylglycerol 
                 0.7603 
                 0.1799 
                 1.0623 
                 0.2549 
                 0.3060 
                 0.8914 
                 0.9292 
                 0.3850 
                 0.9237 
               
               
                 15910 
                 maltotetraose 
                 0.7886 
                 0.1854 
                 1.0561 
                 0.8152 
                 0.5876 
                 0.9876 
                 0.9485 
                 0.3900 
                 1.1374 
               
               
                 37060 
                 methylglutaroylcarnitine 
                 0.7984 
                 0.1866 
                 0.6899 
                 0.0972 
                 0.1771 
                 2.7956 
                 0.0936 
                 0.0801 
                 1.6467 
               
               
                 12025 
                 cis-aconitate 
                 0.8028 
                 0.1873 
                 0.9883 
                 0.6763 
                 0.5329 
                 0.9026 
                 0.2627 
                 0.1574 
                 1.0276 
               
               
                 1640 
                 ascorbate (Vitamin C) 
                 0.821 
                 0.1911 
                 1.0018 
                 0.8119 
                 0.5876 
                 1.0169 
                 0.2942 
                 0.1701 
                 1.1529 
               
               
                 558 
                 adenosine 5′diphosphoribose 
                 0.8463 
                 0.1962 
                 0.9337 
                 0.1841 
                 0.2479 
                 0.7385 
                 0.7111 
                 0.3220 
                 0.9482 
               
               
                 33173 
                 2-hydroxyacetaminophen sulfate 
                 0.8555 
                 0.1979 
                 0.6505 
                 0.4525 
                 0.4293 
                 1.4420 
                 0.6008 
                 0.2820 
                 1.1797 
               
               
                 1408 
                 putrescine 
                 0.884 
                 0.2025 
                 1.0554 
                 0.3823 
                 0.3932 
                 0.8668 
                 0.4838 
                 0.2445 
                 1.0544 
               
               
                 33821 
                 1-eicosatrienoylglycero- 
                 0.904 
                 0.2058 
                 0.9828 
                 0.2406 
                 0.2987 
                 1.5043 
                 0.0562 
                 0.0558 
                 1.5172 
               
               
                   
                 phosphocholine 
               
               
                 27665 
                 1-methylnicotinamide 
                 0.9469 
                 0.2134 
                 0.9365 
                 0.8594 
                 0.6007 
                 1.0928 
                 0.0951 
                 0.0801 
                 1.1174 
               
               
                 21044 
                 2-hydroxybutyrate (AHB) 
                 0.9665 
                 0.2174 
                 1.0117 
                 0.0058 
                 0.0454 
                 1.2906 
                 0.0686 
                 0.0647 
                 1.2024 
               
               
                 20699 
                 erythritol 
                 0.9684 
                 0.2174 
                 0.9460 
                 0.0982 
                 0.1771 
                 1.2939 
                 0.0180 
                 0.0286 
                 1.2313 
               
               
                   
               
             
          
         
       
     
         [0116]    To summarize the results in Tables 1A and 1B, 315 biomarkers were identified. Of these, 206 biomarkers were statistically significantly different between tumors (T) and non-cancer tissue adjacent to tumors (C), 131 biomarkers were identified as significantly different between high aggressive tumors (T_NOC) and less aggressive tumors (T_OC), and 86 biomarkers were identified as significantly different between non-cancer tissue adjacent to high aggressive cancer tumors (N_NOC) and non-cancer tissue adjacent to less aggressive cancer tumors (N_OC). Of the biomarkers that are statistically significantly changed in tumors that are high aggressive cancer (T_NOC) compared to tumors that are less aggressive cancer (T_OC) 34 biomarkers increase or decrease 10%-30%, 49 biomarkers increase or decrease 30%-50%, 37 biomarkers increase or decrease 50%-100% and 12 biomarkers increase or decrease &gt;100%. The range of percent change is 10%-239%. The False Discovery Rate was less than or equal to 5% (i.e., q≦0.05). 
       Example 2 
     Random Forest Analysis for the Classification of Tissue Samples 
       [0117]    The data obtained in Example 1 concerning the tissue samples was used to create a Random Forest model. Random Forest Analysis was carried out on the data obtained from tissue samples in Example 1 to classify them as Control, non-cancer tissue (C), Organ Confined Tumor (T_OC) (i.e. lower aggressive) or Non-Organ Confined Tumor (T_NOC) (i.e. high aggressive cancer). 
         [0118]    It was found that 83% (Table 2) accuracy was achieved by Random Forest Classification of Non-cancer, control tissue compared to organ confined tumor tissue. A list of identified biomarker compounds that effectively separate the groups are presented in Tables 3A and 3B. 
         [0000]    
       
         
               
             
               
               
               
               
             
               
               
               
               
             
               
               
               
               
               
               
             
           
               
                 TABLE 2 
               
             
             
               
                   
               
               
                 Random Forest Classification of Cancer (Tumor) 
               
               
                 vs. Non-cancer (Control) Tissue. 
               
             
          
           
               
                   
                   
                 Predicted 
                   
               
             
          
           
               
                   
                 Control 
                 Tumor 
                 class. error 
               
               
                   
                   
               
             
          
           
               
                   
                 Actual 
                 Control 
                 59 
                 12 
                 0.17 
               
               
                   
                   
                 Tumor 
                 13 
                 60 
                 0.18 
               
               
                   
                   
               
               
                   
                 OOB error = 17% 
               
             
          
         
       
     
         [0119]    The diagnostic parameters based on the Random Forest Analysis are that the Accuracy=83%; the Sensitivity=82, the Specificity=83, the Positive Predictive Value (PPV)=83, the Negative Predictive Value (NPV)=82 and the Area Under the Curve (AUC)=0.87. 
         [0000]    
       
         
               
               
             
           
               
                 TABLE 3A 
               
               
                   
               
             
             
               
                 Glutaroyl-carnitine 
                 Glycerophosphoethanolamine 
               
               
                 Glycerol 2-phosphate 
                 N-acetylglutamate 
               
               
                 Nonadecanoate (19:0) 
                 1-stearoylglycerophosphoinositol 
               
               
                 1-myristoylglycerolphosphocholine 
                 Creatine 
               
               
                 UDP-N-acetylglucosamine 
               
               
                   
               
             
          
         
       
     
         [0000]    
       
         
               
               
               
             
           
               
                   
                 TABLE 3B 
               
               
                   
                   
               
             
             
               
                   
                 Carnitine 
                 5-methylthioadenosine (MTA) 
               
               
                   
                 2-aminoadipate 
                 Proline 
               
               
                   
                   
               
             
          
         
       
     
         [0120]    Random Forest analysis of tissue from less aggressive, organ confined tumors (T_OC) and high aggressive, non-organ confined tumors (T_NOC) resulted in 66% accuracy. The results are presented in Table 4. A list of named biomarkers that effectively separate the genotypes are presented in Table 5. 
         [0000]    
       
         
               
             
               
               
               
               
             
               
               
               
               
             
               
               
               
               
               
               
             
           
               
                 TABLE 4 
               
             
             
               
                   
               
               
                 Random Forest Classification of the organ confined 
               
               
                 tumor vs. non-organ confined cancer. 
               
             
          
           
               
                   
                   
                 Predicted 
                   
               
             
          
           
               
                   
                 T_NOC 
                 T_OC 
                 class. error 
               
               
                   
                   
               
             
          
           
               
                   
                 Actual 
                 T_NOC 
                 18 
                 7 
                 0.28 
               
               
                   
                   
                 T_OC 
                 18 
                 30 
                 0.38 
               
               
                   
                   
               
               
                   
                 OOB error = 34% 
               
             
          
         
       
     
         [0121]    The diagnostic parameters based on the Random Forest Analysis are that the Accuracy=66%; the Sensitivity=63%, the Specificity=72%, the Positive Predictive Value (PPV)=81%, the Negative Predictive Value (NPV)=50% and the Area Under the Curve (AUC)=0.73. 
         [0000]    
       
         
               
               
             
           
               
                 TABLE 5A 
               
               
                   
               
             
             
               
                 Adrenate (22:4n6) 
                 Ribitol 
               
               
                 Adenosine-5-triphosphate (ATP) 
                 Isoleucylisoleucine 
               
               
                 1-stearoylglycerol (1-monostearin) 
                 Laurylcarnitine 
               
               
                 Choline phosphate 
                 1-heptadecanoylglycerophospho- 
               
               
                 Ethanolamine 
                 choline 
               
               
                 Caprylate (8:0) 
                 Guanosine 5′-monophosphate (GMP) 
               
               
                 1-stearoylglycerophosphocholine 
                 2-aminobutyrate 
               
               
                 Docosadienoate (22:2n6) 
                 acetylcholine 
               
               
                   
               
             
          
         
       
     
         [0000]    
       
         
               
               
               
             
           
               
                   
                 TABLE 5B 
               
               
                   
                   
               
             
             
               
                   
                 Xylitol 
                 Laurate 
               
               
                   
                 Tryptophan 
                 Valine 
               
               
                   
                 Glycerol 
                 Uracil 
               
               
                   
                   
               
             
          
         
       
     
         [0122]    Random Forest Analysis was also carried out to classify the tissue samples from the non-cancer tissue adjacent the high aggressive cancer tumor (N_NOC) and the non-cancer tissue adjacent the less aggressive cancer tumor (N_OC). This analysis resulted in 62% correct classification of the two tissue types. The results of the Random Forest analysis are presented in Table 6, and a list of named biomarkers that effectively separate the genotypes are presented in Tables 7A and 7B. 
         [0000]    
       
         
               
             
               
               
               
               
             
               
               
               
               
             
               
               
               
               
               
               
             
           
               
                 TABLE 6 
               
             
             
               
                   
               
               
                 Random Forest Classification of non-cancer tissue adjacent 
               
               
                 to high aggressive cancer tumor (N_NOC) vs. non-cancer tissue 
               
               
                 adjacent to less aggressive cancer tumor (N_OC). 
               
             
          
           
               
                   
                   
                 Predicted 
                   
               
             
          
           
               
                   
                 NOC 
                 OC 
                 class. error 
               
               
                   
                   
               
             
          
           
               
                   
                 Actual 
                 NOC 
                 15 
                 10 
                 0.40 
               
               
                   
                   
                 OC 
                 17 
                 29 
                 0.37 
               
               
                   
                   
               
               
                   
                 OOB error = 38% 
               
             
          
         
       
     
         [0123]    The diagnostic parameters based on the Random Forest Analysis are that the Accuracy=62%; the Sensitivity=63, the Specificity=60, the Positive Predictive Value (PPV)=74, the Negative Predictive Value (NPV)=47 and the Area Under the Curve (AUC)=0.71. 
         [0000]    
       
         
               
               
               
             
           
               
                   
                 TABLE 7A 
               
               
                   
                   
               
             
             
               
                   
                 Oleoylcarnitine 
                 Palmitoylcarnitine 
               
               
                   
                 3-(4-hydroxyphenyl)lactate 
                 Taurocholenate sulfate 
               
               
                   
                 Isovalerylcarnitine 
                 Ribitol 
               
               
                   
                 Tiglyl carnitine 
                 Docosadienoate (22:2n6) 
               
               
                   
                   
               
             
          
         
       
     
         [0000]    
       
         
               
               
               
             
           
               
                   
                 TABLE 7B 
               
               
                   
                   
               
             
             
               
                   
                 Hypoxanthine 
                 Tyrosine 
               
               
                   
                 Isoleucine 
                 Phenylalanine 
               
               
                   
                 Valine 
                 Glycerol 
               
               
                   
                 Leucine 
                 5,6-dihydrouracil 
               
               
                   
                 Tryptophan 
                 Palmitate 
               
               
                   
                 Fumarate 
                 Kynurenine 
               
               
                   
                 S-adenosylhomocysteine (SAH) 
                 Pantothenate 
               
               
                   
                   
               
             
          
         
       
     
       Example 3 
     Biomarkers Useful to Rule Out Aggressive Cancer 
       [0124]    We investigated the ability of the biomarkers identified in Example 1 to rule out aggressive cancer. We selected the biomarker adrenate (22:4n6) to test this idea. The level of adrenate was measured in 19 subjects with high aggressive (i.e., NOC) cancer and 47 subjects with less aggressive (i.e., OC) cancer. The recursive partitioning analysis shows that 19 of 19 subjects with NOC cancer were classified correctly and 26 of the 47 OC subjects were classified correctly based on adrenate levels. The Sensitivity is 100% and the Specificity is 55% and the AUC is 0.74. The results are presented in  FIG. 1 . When these biomarkers were used to evaluate cancer aggressivity in subjects having DRE T1 or T2 and a Gleason score of 6-7, ˜40% (26/66) could be ruled out for having the aggressive form of cancer. 
       Example 4 
     Biomarkers Add Value to Clinical Nomograms 
       [0125]    Currently clinicians utilize clinical parameters such as PSA, biopsy Gleason score, and DRE stage to determine PCa tumor aggressiveness. This method is not very accurate for Gleason 6-7 range. We evaluated the effects of adding metabolite biomarkers to help further stratify those with aggressive and non-aggressive disease. According to the published literature the Partin Nomogram for clinical parameters performs with an AUC of 0.68-0.73 for determining non-organ confined cancer (i.e., less aggressive cancer). We evaluated the subjects described in Example 1 using the Partin nomogram. In our dataset the Partin probabilities yielded an AUC of 0.71, consistent with the literature. 
         [0126]    We then tested the effect of adding a pre-Rule Out Test first and then performing the Partin Nomogram on the remaining records (those not ruled out). In the dataset described in Example 1 for the Partin probabilities for subjects having Gleason 6-7 the AUC=0.65. Using the top Random Forest top hit biomarker for Gleason 6-7 subjects the AUC=0.72. For Gleason 6-7 subjects, using adrenate, the top Random Forest top hit biomarker described in Example 3 as a Rule out test first, then using the Partin probability on the remaining records the AUC increased to 0.83. These results indicate that the biomarkers identified in the instant invention can improve the performance of a currently used clinical tool for evaluating prostate cancer. 
       Example 5 
     DRE Urine Biomarkers 
       [0127]    Biomarkers were identified in urine collected from subjects following a digital rectal examination (DRE) that distinguish subjects that have prostate cancer from those subjects that do not have prostate cancer. The urine was collected from the subjects (16 subjects having prostate cancer, 8 subjects not having prostate cancer) following a DRE, transferred into conical centrifuge tubes and spun in a centrifuge to separate the urine sediment from the urine liquid. The metabolites were extracted from the sediment pellet to measure the small molecules present using GC-MS and LC-MS/MS as described in the General Methods. The small molecule profiles measured in urine sediment from subjects with prostate cancer were compared with the small molecule profiles measured in urine sediment from subjects that did not have prostate cancer to identify the small molecules that are biomarkers for prostate cancer. Biomarkers were identified that correlated with the presence of cancer and were useful cancer biomarkers. The biomarkers identified that distinguish subjects having cancer from those subjects that do not have cancer are listed below in Table 8. 
         [0000]    
       
         
               
             
               
               
             
           
               
                 TABLE 8 
               
               
                   
               
               
                 Biomarkers 
               
               
                   
               
             
             
               
                   
               
             
          
           
               
                   
                 1-stearoylglycerol 
               
               
                   
                 3-indoxylsulfate 
               
               
                   
                 5-oxoproline 
               
               
                   
                 catechol sulfate, 
               
               
                   
                 glycerol 3-phosphate (G3P) 
               
               
                   
                 isobutyrylcarnitine 
               
               
                   
                 pro-hydroxy-pro 
               
               
                   
                 propionylcarnitine 
               
               
                   
                 pyruvate 
               
               
                   
                 uridine 
               
               
                   
                 threonine 
               
               
                   
                 3-hydroxyanthranilate 
               
               
                   
                 3-hydroxyhippurate 
               
               
                   
                 4-hydroxyhippurate 
               
               
                   
                 glucose 
               
               
                   
                 mesaconate 
               
               
                   
                 N-tigloylglycine 
               
               
                   
                 tyramine 
               
               
                   
                 cysteine 
               
               
                   
                 glycine 
               
               
                   
                 alanine 
               
               
                   
                 glutamate 
               
               
                   
                 sarcosine (N-methylglycine) 
               
               
                   
                 2-methylbutyroylcarnitine 
               
               
                   
                 4-acetylphenol sulfate 
               
               
                   
                 7-methylxanthine 
               
               
                   
                 arachidonate (20:4n6) 
               
               
                   
                 fucose 
               
               
                   
                 homovanillate (HVA) 
               
               
                   
                 indoleacetate 
               
               
                   
                 isovalerylcarnitine 
               
               
                   
                 kynurenate 
               
               
                   
                 leucine 
               
               
                   
                 N-(2-furoyl)glycine 
               
               
                   
                 N-acetylarginine 
               
               
                   
                 octanoylcarnitine 
               
               
                   
                 phenylacetylglycine 
               
               
                   
                 phenylalanine 
               
               
                   
                   
               
             
          
         
       
     
         [0128]    The diagnostic parameters of these biomarkers to predict prostate cancer were: Sensitivity of 81%; Specificity of 88%; PPV of 93%; NPV of 70%. The individual biomarker metabolites distinguished cancer from non-cancer with an AUC ranging from 0.73 to 0.84. Box plot graphs for representative biomarkers are presented in  FIG. 3 . 
         [0129]    We determined that these biomarkers were useful to distinguish prostate cancer subtypes. We showed that the levels of the prostate cancer biomarkers not only produced distinct signatures that classified the subjects into prostate cancer or non-cancer groups, but also produced biomarker signatures useful to classify the prostate cancer subjects into cancer subgroups. The biomarkers and the biomarker signatures are presented in  FIG. 4 . 
       Example 6 
     Tissue Panel Biomarkers to Determine Cancer Aggressivity 
       [0130]    Biomarkers for prostate cancer were identified in prostate tissue. The study cohort is described in Table 9. The metabolites were extracted from the prostate tissue samples that contained cancer or prostate tissue samples that did not contain cancer and the small molecules present were measured using GC-MS and LC-MS/MS as described in the general methods. To identify the prostate cancer biomarkers, the small molecule profiles measured in prostate cancer tumors were compared with the small molecule profiles measured in non-cancer prostate tissue. 
         [0000]    
       
         
               
             
               
               
               
             
               
               
               
             
           
               
                 TABLE 9 
               
             
             
               
                   
               
               
                 Study Cohort Description 
               
             
          
           
               
                   
                 Number of 
                 5 year 
               
               
                 Classification 
                 subjects 
                 recurrence 
               
               
                   
               
             
          
           
               
                 Organ Confined (OC)** 
                 73 
                  8/45 
               
               
                 Extra Capsular Extension (ECE) 
                 116 
                 19/60 
               
               
                 (SVI negative and LN negative) 
               
               
                 Seminal vesicle invasion positive (SVI+) 
                 54 
                 34/43 
               
               
                 Lymph node negative (LN−) 
               
               
                 SVI − 
                 7 
                 6/7 
               
               
                 Lymph node positive (LN+) 
               
               
                 SVI+ and LN+ 
                 25 
                 19/24 
               
               
                 Total subjects 
                 268 
               
               
                   
               
             
          
         
       
     
         [0131]    The biomarkers identified in prostate tissue that distinguish subjects having cancer from those subjects that do not have cancer are listed below in Table 10. 
         [0000]    
       
         
               
             
               
               
             
           
               
                 TABLE 10 
               
               
                   
               
               
                 Biomarkers 
               
               
                   
               
             
             
               
                   
               
             
          
           
               
                   
                 1-methylhistidine 
               
               
                   
                 1-palmitoylplasmenylethanolamine 
               
               
                   
                 adenosine 5′-diphosphate (ADP) 
               
               
                   
                 arabonate 
               
               
                   
                 N6-acetyllysine 
               
               
                   
                 N-acetylglucosamine-6-phosphate 
               
               
                   
                 N-acetylserine 
               
               
                   
                 N-formylmethionine 
               
               
                   
                 nicotinamide adenine dinucleotide reduced (NADH) 
               
               
                   
                 nicotinamide-ribonucleotide (NMN) 
               
               
                   
                 nicotinamide-riboside 
               
               
                   
                 ribulose 5-phosphate 
               
               
                   
                 xylulose 5-phosphate 
               
               
                   
                 quinate 
               
               
                   
                 trans-aconitate 
               
               
                   
                 ribose 
               
               
                   
                 xylulose 
               
               
                   
                 ethanolamine 
               
               
                   
                 sarcosine (N-methylglycine) 
               
               
                   
                 ascorbate (Vitamin C) 
               
               
                   
                 citrate 
               
               
                   
                 creatinine 
               
               
                   
                 inositol-1-phosphate (I1P) 
               
               
                   
                 kynurenine 
               
               
                   
                 N-acetylaspartate (NAA) 
               
               
                   
                 10-nonadecenoate (19:1n9) 
               
               
                   
                 2-palmitoylglycerophosphoethanolamine 
               
               
                   
                 3-(4-hydroxyphenyl)lactate 
               
               
                   
                 5,6-dihydrouracil 
               
               
                   
                 glycerol 2-phosphate 
               
               
                   
                 glycylvaline 
               
               
                   
                 lactate 
               
               
                   
                 N-acetylputrescine 
               
               
                   
                 nicotinamide-adenine-dinucleotide (NAD+) 
               
               
                   
                 phosphoethanolamine 
               
               
                   
                 putrescine 
               
               
                   
                 spermidine 
               
               
                   
                 spermine 
               
               
                   
                 succinylcarnitine 
               
               
                   
                 10-heptadecenoate (17:1n7) 
               
               
                   
                   
               
             
          
         
       
     
         [0132]    Prostate cancer that is no longer confined to the prostate organ, that is, when it is not organ confined (N_OC) is considered more aggressive than prostate cancer that is confined to the prostate, that is when it is organ confined (OC). Non-organ confined prostate cancer is associated with a higher Gleason Score (GS), with detection of cancer cells in the lymph nodes (LN), with tumors that have extra-capsular extensions (ECE), and with seminal vesicle invasion (SVI). We identified biomarkers that are indicative of each of these types of aggressiveness indicators by measuring the small molecule profiles of cancer tumors with each of these aggressiveness indicators using GC-MS and LC-MS/MS as described in the general methods. The small molecule profiles obtained were compared with the small molecule profiles from non-tumor and non-aggressive cancer tumors to identify the biomarkers. The biomarkers identified in the test cohort were evaluated using a receiver operator characteristic (ROC) curve and the area under the curve (AUC) was determined for each of the aggressiveness indicators using a new cohort of subjects. 
         [0133]    The biomarkers putrescine, lactate, 5,6-dihydrouracil, 10-nonadecenoate, NAD+, spermine, and N-acetylputrescine were useful biomarkers to indicate subjects with prostate cancer tumors that had extracapsular extensions (ECE). The AUC was 0.84. 
         [0134]    The biomarkers putrescine, glycerol-2-phosphate, and glycylvaline were useful biomarkers to indicate subjects with prostate cancer tumors that had invaded the seminal vesicles. The AUC was 0.75. 
         [0135]    The biomarkers phosphoethanolamine, putrescine, spermidine were useful biomarkers to indicate the subjects with prostate cancer tumors that had cancer cells detected in the lymph nodes (LN). The AUC was 0.73. 
         [0136]    The biomarkers succinylcarnitine, 3-(4-hydroxyphenyl)lactate, 2-palmitoylglycerophosphoethanolamine, lactate, and spermidine were useful biomarkers for identifying the cancer tumors associated with a higher Gleason Score. The AUC was 0.73. 
       Example 7 
     Biomarkers of Prostate Cancer Recurrence 
       [0137]    Biomarkers indicative of prostate cancer recurrence were identified that were useful to determine the individuals with prostate cancer that will recur in 5 years. Cancer recurrence is an indicator of cancer tumor aggressiveness. The levels of the biomarkers were initially measured in subjects that had prostate cancer and determined to be biomarkers for cancer aggressivity. The biomarkers were measured in an independent cohort of subjects that had been treated for prostate cancer and underwent a prostatectomy. Of this group of 61 prostate cancer subjects, the prostate cancer did not recur within 5 years in 33 subjects and prostate cancer did recur within 5 years in 28 subjects. Based on the levels of the biomarkers putrescine, lactate, 5,6-dihydrouracil, 10-nonadecenoate, NAD+, spermine, N-acetylputrescine, succinylcarnitine, 3-(4-hydroxyphenyl)lactate, 2-palmitoylglycerophosphoethanolamine, spermidine, glycerol-2-phosphate, glycylvaline, and/or phosphoethanolamine, measured in the cancer tumor tissue, the subjects were predicted to have non-aggressive cancer tumors or aggressive cancer tumors. As presented in Table 11, 25 of 28 cancer tumors that recurred within 5 years were classified as aggressive using the biomarkers while 14 of the 33 non-recurrent tumors were classified as aggressive. 
         [0000]    
       
         
               
             
               
               
               
             
               
               
               
               
             
               
               
               
               
             
           
               
                 TABLE 11 
               
             
             
               
                   
               
               
                 Cancer 5 Year Recurrence Study Cohort Description. 
               
             
          
           
               
                   
                 5 Year Recurrence (Actual) 
                   
               
             
          
           
               
                   
                 Predicted 
                 Non Recurrent 
                 Recurrent 
               
               
                   
                   
               
             
          
           
               
                   
                 Non Aggressive 
                 19 
                 3 
               
               
                   
                 Aggressive 
                 14 
                 25 
               
               
                   
                   
               
             
          
         
       
     
         [0138]    The biomarkers were useful to predict 5 year cancer recurrence. The biomarkers predicted prostate cancer recurrence in 5 years in prostate cancer subjects with a Sensitivity of 89%, Specificity of 58%, PPV of 65%, and an NPV of 86%. 
         [0139]    The same subjects were evaluated using the currently used clinical Han nomogram. Using the Han nomogram, 5 year cancer recurrence 23 of 27 subjects were classified correctly as recurrent. The nomogram correctly predicted non-recurrence for only 7 of 33 subjects. The results of the Han nomogram are presented in Table 12. The ROC curve for the Han nomogram is presented in  FIG. 5 . In contrast to the performance of the biomarkers of the instant invention, the Han nomogram had a Sensitivity of 85%, Specificity of 22%, PPV of 47% and NPV of 64%. The performance of the biomarkers in the instant invention was superior to that of the current clinical standard Han nomogram to predict the subjects with 5 year cancer recurrence. 
         [0000]    
       
         
               
             
               
               
               
             
               
               
               
               
             
               
               
               
               
             
           
               
                 TABLE 12 
               
             
             
               
                   
               
               
                 Cancer 5 Year Recurrence Predicted using Han Nomogram. 
               
             
          
           
               
                   
                 5 Year Recurrence (Actual) 
                   
               
             
          
           
               
                   
                 Han-Predicted: 
                 Recurrent 
                 Non-Recurrent 
               
               
                   
                   
               
             
          
           
               
                   
                 Recurrent 
                 23 
                 26 
               
               
                   
                 Non-recurrent 
                 4 
                 7 
               
               
                   
                   
               
             
          
         
       
     
         [0140]    The performance characteristics of the biomarkers of the instant invention and the Han nomogram are presented in Table 13. 
         [0000]    
       
         
               
             
               
               
               
             
               
               
               
               
             
           
               
                 TABLE 13 
               
             
             
               
                   
               
               
                 Comparison of Biomarkers with Han Nomogram 
               
               
                 to predict cancer 5 year recurrence. 
               
             
          
           
               
                   
                 Han Nomogram 
                 Biomarkers 
               
               
                   
                   
               
             
          
           
               
                   
                 Sensitivity 
                 0.85 
                 0.89 
               
               
                   
                 Specificity 
                 0.22 
                 0.58 
               
               
                   
                 PPV 
                 0.47 
                 0.64 
               
               
                   
                 NPV 
                 0.64 
                 0.86 
               
               
                   
                   
               
             
          
         
       
     
         [0141]    While the invention has been described in detail and with reference to specific embodiments thereof, it will be apparent to one skilled in the art that various changes and modifications can be made without departing from the spirit and scope of the invention.