Phosphocofilin: cofilin co-localization intensity as a predictor of metastatic recurrence

Abstract

Methods and products are provided for determining if a subject having a tumor is at risk of metastasis of the tumor. Specifically, the methods comprise detecting phosphorylated cofilin, and both phosphorylated and non-phosphorylated cofilin; quantifying the phosphorylated cofilin, and the total of phosphorylated and nonphosphorylated cofilin; and determining if a subject having the tumor is likely to experience metastasis of the tumor, based on the ratio of the amount of detected phosphorylated cofilin: total amount of phosphorylated and non-phosphorylated cofilin detected. Further disclosed are the types of tumor metastases that can be determined using the methods provided.

1 . A method of determining if a subject having a tumor is likely to experience metastasis of the tumor comprising contacting a sample of the tumor with (a) a detectable agent that binds phosphorylated cofilin and (b) a detectable agent which binds both phosphorylated cofilin and non-phosphorylated cofilin and quantifying (i) the phosphorylated cofilin bound by (a) and (ii) the total of phosphorylated cofilin and non-phosphorylated cofilin bound by (b), wherein a ratio of the amount of phosphorylated cofilin bound:total amount of phosphorylated cofilin and non-phosphorylated cofilin bound in excess of a predetermined ratio value, or a colocalization intensity value (IF) within the highly colocalized areas marked by (a) and (b) in excess of a predetermined IF value, indicates that the subject having the tumor is likely to experience metastasis of the tumor. 
     
     
         2 . The method of  claim 1 , wherein the tumor is a breast cancer tumor, or a tumor of the nasopharynx, pharynx, lung, bone, brain, sialaden, stomach, esophagus, testes, ovary, uterus, liver, small intestine, appendix, colon, rectum, gall bladder, pancreas, kidney, urinary bladder, breast, cervix, vagina, vulva, prostate, thyroid or skin. 
     
     
         3 . The method of  claim 1 , wherein IF is intensity of (a) in pixels of an image of the sample x intensity of (b) in pixels of the image of the sample, optionally first masking out pixels with low Pearson distribution. 
     
     
         4 . The method of  claim 1 , wherein the tumor is a solid tumor. 
     
     
         5 . The method of  claim 1 , wherein the sample is a biopsy sample. 
     
     
         6 . The method of  claim 1 , wherein the sample is surgically removed from the subject. 
     
     
         7 . The method of  claim 1 , wherein the detectable agent that binds phosphorylated cofilin comprises an antibody, or antigen-binding fragment thereof, that specifically binds phosphorylated cofilin. 
     
     
         8 . The method of  claim 1 , wherein the detectable agent which binds to phosphorylated cofilin and to non-phosphorylated cofilin comprises an antibody, or antigen-binding fragment thereof, which binds phosphorylated cofilin and which also binds non-phosphorylated cofilin. 
     
     
         9 . The method of  claim 1 , wherein the detectable agent which binds to phosphorylated cofilin and to non-phosphorylated cofilin and/or the detectable agent which binds to phosphorylated cofilin comprises a fluorescent moiety. 
     
     
         10 . The method of  claim 9 , wherein both the detectable agents each comprise a fluorescent moiety and wherein the detectable agent that binds to phosphocofilin comprises a different fluorescent moiety from the fluorescent moiety of the detectable agent that binds to phosphorylated cofilin and to non-phosphorylated cofilin. 
     
     
         11 . (canceled) 
     
     
         12 . The method of  claim 1 , wherein the phosphorylated cofilin and non-phosphorylated cofilin are human cofilin. 
     
     
         13 . The method of  claim 1 , wherein the cofilin comprises SEQ ID NO:1. 
     
     
         14 . The method of  claim 1 , wherein the sample is a paraffin-embedded biopsy sample. 
     
     
         15 . The method of  claim 1 , wherein the detectable agent which binds to phosphorylated cofilin and which binds to non-phosphorylated cofilin comprises an anti-pan-cofilin antibody. 
     
     
         16 . The method of  claim 1 , wherein the detectable agent which specifically binds to phosphorylated cofilin comprises an anti-phospho-Ser-3-cofilin antibody. 
     
     
         17 . The method of  claim 1 , wherein the predetermined ratio value is determined from one or more subjects with malignant tumors which have not metastasized. 
     
     
         18 . (canceled) 
     
     
         19 . The method of  claim 1 , further comprising administering to the subject found likely to experience metastasis of the tumor an anti-metastatic therapy. 
     
     
         20 . A method of mitigating the risk of metastasis of a tumor in a subject comprising administering to a subject identified by the method of  claim 1  as likely to experience metastasis of the tumor an anti-metastatic therapy. 
     
     
         21 . A kit for determining if a subject having a tumor is likely to experience metastasis of the tumor, the kit comprising (a) a detectable agent that specifically binds phosphorylated cofilin and (b) a detectable agent which binds to phosphorylated cofilin and which binds to non-phosphorylated cofilin, and instructions for use. 
     
     
         22 . (canceled) 
     
     
         23 . A computer system comprising a program for determining if a subject having a tumor is likely to experience metastasis of the tumor, the computer system comprising a processor and a memory encoding one or more programs coupled to the processor, wherein the processor effects the one or more programs to quantify phosphorylated cofilin stained with a detectable agent in a sample from the tumor in the subject and quantify total phosphorylated cofilin and un-phosphorylated cofilin stained with a different detectable agent in the sample and determine the ratio thereof. 
     
     
         24 - 30 . (canceled)