Biomarkers for Predicting the Recurrence of Colorectal Cancer Metastasis

A method for classifying microsatellite instability (MSI) in a primary colorectal cancer (CRC) comprising:

• providing a panel comprising of mono-, di-, and tetranucleotide repeat loci markers to be used in a MSI assay, wherein the markers are selected from the group consisting of BAT25;
• BAT26;
• D2S123;
• D5S346;
• D175250;
• D18564;
• D18569;
• MYCL1;
• D20582;
• D20585;
• L17835;
• D8S321;
• D9S242;
• and D195394;
• providing a genomic DNA isolated from one or more biological samples from a human subject suffering from or the CRC;
• determining a presence or an absence of the MSI in the primary CRC from the isolated genomic DNA obtained from the human subject;
• and
• classifying the MSI or determining a tumor phenotype based on a scheme, wherein the scheme comprises:
• a MSI-H phenotype indicative of a presence of MSI at three or more of the mono- or dinucleotide markers;
• a MSI-L phenotype indicative of a presence of MSI at least one but no more than two of the mono- or dinucleotide markers;
• a MSS phenotype indicative no MSI at any of the mono- or dinucleotide markers;
• a EMAST+ phenotype indicative of a non MSI-H phenotype with MSI at least one of the tetranucleotide markers;
• a EMAST− phenotype indicative of a non MSI-H phenotype with no MSI at any of the tetranucleotide markers;
• a MSI-M phenotype indicative of a MSI-L, EMAST, or both MSI-L and EMAST phenotype;
• and
• a H-MSS phenotype indicative of non MSI at any of the mono-, di-, and tetranucleotide markers.

A method for selecting a cancer therapy in a patient diagnosed with primary colorectal cancer (CRC), the method comprising:

• identifying the patient diagnosed with the primary CRC;
• determining a level of microsatellite instability (MSI) at one or more mononucleotide, dinucleotide, tetranucleotide repeats (EMAST), or any combinations thereof in cells obtained from one or more biological samples from the patient, wherein a presence of a MSI-M phenotype in a majority of cells in a sample from the stage II or III CRC subject is indicative of a high risk for recurrence, a high risk for liver metastasis (LM), a lowered possibility of success with the cancer therapy or any combinations thereof and presence of a H-MSS phenotype is indicative of a high probability for recurrence-free survival in the human subject;
• and
• selecting the cancer therapy based on identifying agents to lower or suppress the MSI-M, MSS phenotype.

A method of performing a clinical trial to evaluate a candidate drug believed to be useful in treating colorectal liver metastasis, promoting recurrence-free survival, or both, the method comprising:

• a) determining a level of microsatellite instability at least one of one or more tetranucleotide repeats (EMAST), a mono- and dinucleotide repeat loci (MSI-L), or a SMARCA2R-LOH, in cells obtained from a patient, wherein a MSI-M phenotype in a majority of cells in a sample from the patient is indicative of a highest risk for recurrence, a high risk for liver metastasis (LM), or any combinations thereof and presence of MSI-H phenotype is indicative of lowest risk and H-MSS phenotype is indicative of an intermediate risk for recurrent distant metastasis;
• b) administering a candidate drug to a first subset of the patients, and
• a placebo to a second subset of the patients;
• a comparator drug to a second subset of the patients;
• or
• a drug combination of the candidate drug and another active agent to a second subset of patients;
• c) repeating step a) after the administration of the candidate drug or the placebo, the comparator drug or the drug combination;
• and
• d) monitoring a recurrent-free survival rate exhibited by stage II and III primary CRC patients with an MSI-H, an MSI-M, or an H-MSS phenotype that is statistically significant as compared to the rate exhibited by the patients with the MSI-H, the MSI-M, the H-MSS and the SMARCA2R-LOH, phenotypes occurring in the second subset of patients, wherein a statistically significant increase indicates that the candidate drug is useful in treating said disease state.

A method for determining the risk for development of colorectal liver metastasis in a human subject suffering from colorectal cancer (CRC) comprising the steps of:

• identifying the human subject suffering from the primary CRC;
• obtaining one or more biological samples from the subject, wherein the biological samples are selected from the group consisting of a frozen or fresh tissue sample, a FFPE tissue sample, a fecal sample, one or more biological fluids, or any combinations thereof;
• measuring or determining a level of a microsatellite instability (MSI) using a microsatellite assay comprising a panel of a mononucleotide repeat loci, a dinucleotide repeat loci, and a tetranucleotide (EMAST) repeat loci selected from the group consisting of BAT25;
• BAT26;
• D2S123;
• D5S346;
• D17S250;
• D18S64;
• D18S69;
• MYCL1;
• D20S82;
• D20S85;
• L17835;
• D8S321;
• D9S242;
• and D19S394 and a SMARCA2R-LOH;
• determining a presence or an absence of the MSI in the primary CRC from the isolated genomic DNA obtained from the human subject;
• classifying the MSI in the primary CRC by using a classification scheme, wherein the classification scheme comprises:
• a MSI-H phenotype indicative of a presence of MSI at three or more of the mono- or dinucleotide markers;
• a MSI-L phenotype indicative of a presence of MSI at at least one but no more than two of the mono- or dinucleotide markers;
• a MSS phenotype indicative no MSI at any of the mono- or dinucleotide markers;
• a EMAST+ phenotype indicative of a non MSI-H phenotype with MSI at at least one of the tetranucleotide markers;
• a EMAST− phenotype indicative of a non MSI-H phenotype with no MSI at any of the tetranucleotide markers;
• a MSI-M phenotype indicative of a MSI-L or EMAST or both MSI-L and EMAST phenotype;
• and
• a H-MSS phenotype indicative of non MSI at any of the mono-, di-, and tetranucleotide markers;
• and
• determining the risk for colorectal cancer liver metastasis in the human subject based on a presence or an increase in the MSI-M phenotype in the sample.