CIRCULATING miRNAs AS MARKERS FOR BREAST CANCER

Abstract

The present invention is concerned with a method for diagnosing breast cancer in a subject comprising the steps of determining in a sample of a subject suspected to be afflicted with said breast cancer the amount of at least one miRNA or the amounts of at least the miRNAs of a combination of miRNAs selected from the group consisting of: (i) miR-801, (ii) miR-801 and miR-148b, (iii) miR-801 and miR-376c, (iv) miR-801 and miR-409-3p, (v) miR-801, miR-376c and miR-148b, (vi) miR-801, miR-409-3p and miR-376c, (vii) miR-801, miR-409-3p and miR-148b, miR-801, miR-376c, miR-409-3p and miR-148b, (ix) miR-148b, (x) miR-409-3p, (xi) miR-376c, (xii) miR-376c and miR-409-3p, (xiii) miR-148b and miR-376c, (xiv) miR-148b and miR-409-3p, (xv) miR-148b, miR-376c and miR-409-3p, (xvi) miR-127-3p, (xvii) miR-148b, (xvii) miR-376a, (xix) miR-376c, (xx) miR-409-3p, (xxi) miR-652, (xxii) miR-127-3p, miR-148b, miR-376a, miR-376c, miR-409-3p, miR-652, and miR-801, (xxiii) miR-127-3p, miR-148b, miR-652, and miR-801, (xxiv) miR-376a, miR-148b, miR-652, and miR-801, (xxv) miR-376c, miR-148b, miR-652, and miR-801, and (xxvi) miR-409-3p, miR-148b, miR-652, and miR-801 and comparing said amount with a reference or comparing said amounts with references, whereby breast cancer is to be diagnosed. The present invention is also concerned with methods for diagnosing metastasizing breast cancer in a subject and for determining the circulating tumor cell (CTC) status in a subject comprising the steps of (a) determining in a sample of a subject suspected to be afflicted with said metastasizing breast cancer the amount of at least one miRNA selected from the group consisting of: miR-801, miR-141, miR-200a, miR-200b, miR-200c, miR-203, miR-210, miR-375, miR-142-3p, and miR-768-3p, and (b) comparing said amount with a reference or comparing said amounts with references. Furthermore the present invention is concerned with the use of the miRNAs of the invention for diagnosing breast cancer, metastasizing breast cancer, or for determining the CTC status in a subject. Moreover, the present invention is concerned with devices and kits for carrying methods of the invention.

1 . A method for diagnosing breast cancer in a subject comprising the steps of:
 (a) determining in a sample of a subject suspected to be afflicted with said breast cancer the amount of at least one miRNA or the amounts of at least the miRNAs of a combination of miRNAs selected from the group consisting of: (i) miR-801, (ii) miR-801 and miR-148b, (iii) miR-801 and miR-376c, (iv) miR-801 and miR-409-3p, (v) miR-801, miR-376c and miR-148b, (vi) miR-801, miR-409-3p and miR-376c, (vii) miR-801, miR-409-3p and miR-148b, (viii) miR-801, miR-376c, miR-409-3p and miR-148b, (ix) miR-148b, (x) miR-409-3p, (xi) miR-376c, (xii) miR-376c and miR-409-3p, (xiii) miR-148b and miR-376c, (xiv) miR-148b and miR-409-3p, (xv) miR-148b, miR-376c and miR-409-3p, (xvi) miR-127-3p, (xvii) miR-148b, (xvii) miR-376a, (xix) miR-376c, (xx) miR-409-3p, (xxi) miR-652, (xxii) miR-127-3p, miR-148b, miR-376a, miR-376c, miR-409-3p, miR-652, and miR-801, (xxiii) miR-127-3p, miR-148b, miR-652, and miR-801, (xxiv) miR-376a, miR-148b, miR-652, and miR-801, (xxv) miR-376c, miR-148b, miR-652, and miR-801, and (xxvi) miR-409-3p, miR-148b, miR-652, and miR-801; and   (b) comparing said amount with a reference or comparing said amounts with references, whereby breast cancer is to be diagnosed.   
     
     
         2 . The method of  claim 1 , wherein said sample is a body fluid sample and wherein an amount determined in step (a) of said at least one miRNA or amounts of miRNAs of a combination of miRNAs higher than said reference value is/are indicative of a subject being afflicted with breast cancer. 
     
     
         3 . The method of  claim 2 , wherein said body fluid is selected from the group consisting of: blood, serum, plasma, saliva, urine, and fluids obtainable from the breast glands. 
     
     
         4 . The method of  claim 1 , wherein the sample is a tumor sample and wherein an amount determined in step (a) of said at least one miRNA or amounts of miRNAs of a combination of miRNAs lower than said reference value is/are indicative of a subject being afflicted with breast cancer. 
     
     
         5 . The method of  claim 1 , wherein said breast cancer is early breast cancer. 
     
     
         6 - 10 . (canceled) 
     
     
         11 . A method for determining the circulating tumor cell (CTC) status in a subject comprising the steps of:
 (a) determining in a sample of a subject suspected to be afflicted with breast cancer the amount of at least one miRNA selected from the group consisting of: miR-801, miR-141, miR-142-3p, miR-200a, miR-200b, miR-200c, miR-203, miR-210, miR-375, and miR-768-3p, and   (b) comparing said amount with a reference or comparing said amounts with references, whereby the CTC status is to be determined.   
     
     
         12 . The method of  claim 11 , wherein the amount of least one miRNA selected from the group consisting of: miR-801, miR-203, and miR-768-3p, or the amounts of at least two miRNAs selected from at least two different groups of miRNAs are determined, said groups being selected from:
 (i) a group consisting of miR-142-3p and miR-768-3p,   (ii) a group consisting of miR-203,   (iii) a group consisting of miR-375,   (iv) a group consisting of miR-210 and miR-801,   (v) a group consisting of miR-141, miR-200a, miR-200b, miR-200c.   
     
     
         13 . The method of  claim 11 , wherein said at least two miRNAs comprise (i) miR-141, miR-200b, miR-375, and miR-801, (ii) miR-141, miR-200b, miR-375, miR-801, miR-203, and miR-768-3p, (iii) miR-141, miR-200c, miR-210, miR-801, and miR-768-3p, (iv) miR-141, miR-200b, miR-142-3p, and miR-768-3p, (v) miR-141, miR-200b, miR-210, miR-375, miR-801, miR-142-3p, and miR-768-3p, (vi) miR-141, miR-200b, miR-200c, miR-210, miR-375, miR-801, miR-142-3p, and miR-768-3p, or (vi) miR-141, miR-200b, miR-200c, miR-210, miR-375, miR-203, miR-801, miR-142-3p, and miR-768-3p. 
     
     
         14 . The method of  claim 11 , wherein said combination comprising at least two miRNAs is selected from the group consisting of (i) miR-141 and miR-200b, (ii) miR-141, miR-200b, and miR-200c, (iii) miR-141, miR-200b, miR-210, and miR-200c, (iv) miR-141, miR-200b, miR-210, miR-768-3p, and miR-200c, (v) miR-141, miR-200b, and miR-375, (vi) miR-141, miR-200b, miR-210, miR-375, and miR-203, and (vi) miR200c and miR-210. 
     
     
         15 . The method of  claim 11 , wherein said reference is an amount of miRNA or amounts of miRNAs of a combination of miRNAs which is/are derived from a subject or group of subjects with a known CTC status. 
     
     
         16 . A method for recommending a breast cancer therapy to a subject comprising
 the method of  claim 1 , and   the further step of recommending a breast cancer therapy to the subject if breast cancer has been diagnosed.   
     
     
         17 . The method of  claim 16 , wherein said breast cancer therapy is selected from the group consisting of: chemotherapy, anti-hormone therapy, targeted therapy, immunotherapy, surgery, radiation therapy, and cell based immunotherapy. 
     
     
         18 - 21 . (canceled) 
     
     
         22 . A kit for carrying out the method of  claim 1 , wherein said kit comprises instructions for carrying out said method, a detection agent for determining the amount of at least one miRNA selected from the group consisting of: miR-801, miR-376c, miR-409-3p, miR-148b, miR-127-3p, miR-376a, and miR-652 in a sample of a subject suspected to be afflicted with breast cancer, and standards for a reference. 
     
     
         23 - 24 . (canceled) 
     
     
         25 . A kit for carrying out the method of  claim 11 , wherein said kit comprises instructions for carrying out said method, a detection agent for determining the amount of at least one miRNA selected from the group consisting of: miR-141, miR-200a, miR-200b, miR-200c, miR-203, miR-210, miR-375, miR-142-3p, miR-768-3p, and miR-801 in a sample of a subject suspected to be afflicted with breast cancer, and standards for a reference. 
     
     
         26 . The kit of  claim 25 , wherein the miRNA is selected from the group consisting of: miR-801, miR-203, miR-142-3p, and miR-768-3p. 
     
     
         27 . A method for diagnosing a benign or malignant breast tumor in a subject comprising the steps of:
 (a) determining in a sample of a subject suspected to be afflicted with breast tumor the amount of at least one miRNA or the amounts of at least the miRNAs of a combination of miRNAs selected from the group consisting of: (i) miR-148b, (ii) miR-652, (iii) miR-801, (iv) miR-148b and miR 652, (v) miR-148b and miR-801, (vi) miR-652 and miR-801, and (vii) miR-148b, miR-652, and miR-801; and   (b) comparing said amount with a reference or comparing said amounts with references, whereby a benign or malignant breast tumor is to be diagnosed.   
     
     
         28 . The method of  claim 27 , wherein said sample is a body fluid sample and wherein an amount determined in step (a) of said at least one miRNA or amounts of miRNAs of a combination of miRNAs higher than said reference value is/are indicative of a subject being afflicted with a breast tumor. 
     
     
         29 . The method of  claim 27 , wherein said body fluid is selected from the group consisting of: blood, serum, plasma, saliva, urine, and fluids obtainable from the breast glands. 
     
     
         30 - 35 . (canceled) 
     
     
         36 . A method for recommending a breast cancer therapy to a subject comprising the method of  claim 11 , and the further step of recommending a breast cancer therapy to the subject if breast cancer has been diagnosed. 
     
     
         37 . The method of  claim 36 , wherein said breast cancer therapy is selected from the group consisting of: chemotherapy, anti-hormone therapy, targeted therapy, immunotherapy, surgery, radiation therapy, and cell based immunotherapy.