Methods and compositions for the detection of chromosomal aberrations

Abstract

This invention relates generally to methods and compositions for direct detection of specific nucleic acid flanking sequences associated with structural chromosomal aberration breakpoints, by forming hybrids between the sequences and genetic probes, and detecting the probes. In particular aspects, the invention concerns detection of nucleic acid sequences in situ in chromosomes, and more specifically in cells, including interphase cells. Compositions of probes useful for detecting chromosomal translocations, in particular those associated with human leukemias, are also disclosed. An aspect of the invention is labelled probes that, when juxtaposed by formation of an aberration, are distinguishable and provide a pattern different from that of normal cells.

What is claimed is:  
     
         1 . A method of detecting a structural chromosomal aberration comprising: 
 (a) preparing a plurality of nucleic acid probes each capable of hybridizing with a separate nucleic acid flanking sequence brought together by the chromosome aberration;    (b) contacting the probes with chromatin under conditions of appropriate stringency to allow hybridization of the probes to sequences homologous with the probe sequences; and    (c) detecting the presence of the probes.    
     
     
         2 . The method of detecting a chromosomal aberration of  claim 1  wherein the probes are labelled.  
     
     
         3 . The method of detecting a chromosomal aberration of  claim 2  wherein each probe label is distinct from each other.  
     
     
         4 . The method of detecting a chromosomal aberration of  claim 3  wherein the probes are further defined as at least approximately 800 kb apart.  
     
     
         5 . The method of detecting a chromosomal aberration of  claim 4  wherein the labels comprise fluorescent labels.  
     
     
         6 . The method of detecting a chromosomal aberration of  claim 5  wherein the fluorescent labels are microscopically distinct as different colors.  
     
     
         7 . The method of detecting a chromosomal aberration of  claim 6  wherein the fluorescent labels comprise digoxigenin-11-dUTP and biotin-11-dUTP.  
     
     
         8 . The method of detecting a chromosomal aberration of  claim 1  wherein the chromatin-probe contacts occur in situ in cells.  
     
     
         9 . The method of detecting a chromosomal aberration of  claim 8  wherein the cells comprise those in interphase of mitotic division.  
     
     
         10 . The method of detecting a chromosomal aberration of  claim 9  wherein the probes are juxtaposed in interphase as doublets if a chromosomal aberration is present.  
     
     
         11 . The method of detecting a chromosomal aberration of  claim 10  wherein the chromosomal aberration is further defined as comprising a translocation.  
     
     
         12 . The method of detecting a chromosomal aberration of  claim 11  wherein the translocation is formed by breakpoints which occur on the long arms of human chromosomes No. 9 and No. 22.  
     
     
         13 . The method of detecting a chromosomal aberration of  claim 12  wherein the translocation breakpoints are further defined as occurring at the locations designated t(9;22)(q11;q34).  
     
     
         14 . The method of detecting a chromosomal aberration of  claim 13  wherein the translocation breakpoints are further defined to occur in the BCR and ABL genes respectively, and a fusion gene is formed by the translocation, and said fusion gene comprises portions of the BCR and ABL genes.  
     
     
         15 . The method of detecting a chromosomal aberration of  claim 14  wherein the fusion gene is designated as p190.  
     
     
         16 . The method of detecting a chromosomal aberration of  claim 10  wherein the probes consist of those selected from probes designated PEM12, c-H-abl and MSB-1.  
     
     
         17 . The method of detecting a chromosomal aberration of  claim 8  wherein the cells comprise samples of human tissues.  
     
     
         18 . The method of detecting a chromosomal aberration of  claim 17  wherein the human tissue samples comprise peripheral blood.  
     
     
         19 . The method of detecting a chromosomal aberration of  claim 17  wherein the human tissue samples comprise bone marrow.  
     
     
         20 . The method of detecting a chromosomal aberration of  claim 8  wherein the cells comprise a sample of cultured cells.  
     
     
         21 . A genetic probe capable of hybridizing to the 5′region of the major breakpoint cluster region (M-bcr) of chromosome 22 as illustrated in FIG. 2A and FIG. 4.  
     
     
         22 . A genetic probe capable of hybridizing to the first exon region of the BCR gene as illustrated in FIG. 2A.  
     
     
         23 . A genetic probe designated as c-H-abl and capable of hybridizing to the 3′ end of the ABL gene as illustrated in FIG. 5 and FIGS. 2B and 2C.  
     
     
         24 . The genetic probe of  claim 21  wherein the probe comprises the designation PEM12.  
     
     
         25 . The genetic probe of  claim 22  wherein the probe comprises designation MSB-1.  
     
     
         26 . The genetic probe of  claim 23  wherein the probe comprises designation c-H-abl.  
     
     
         27 . The method of detecting a chromosomal aberration of  claim 1  wherein the plurality of probes comprise MSB-1, PEM12 and c-H-abl, and said probes are contacted to chromosomes in pairs.  
     
     
         28 . The method of detecting chromosomal aberrations of  claim 27  wherein a first pair comprises MSB-1 and c-H-abl, and a second pair comprises PEM12 and c-H-abl.  
     
     
         29 . A kit for the detection of chromosomal aberrations comprising at least two genetic probes selected from claims  21 ,  22  and  23 , and appropriate controls, each in separate containers.  
     
     
         30 . A kit for the detection of cancer in human cells, comprising: 
 a) a carrier being compartmentalized to hold multiple containers;    b) a first pair of containers including the pair of genetic probes of claims  21  and  23 ; and    c) a second pair of containers containing the pair of genetic probes of claims  22  and  23 .