This invention relates to a novel tumor suppressor gene, ING1, to methods for making and using this and related tumor suppressor genes and proteins and peptides, and to nucleic acids encoding this and related tumor suppressor proteins and peptides.
The following references are cited in the application as numbers in brackets ([ ]) at the relevant portion of the application.
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The disclosure of the above publications, patents and patent applications are herein incorporated by reference in their entirety to the same extent as if the language of each individual publication, patent and patent application were specifically and individually included herein.
Many cancers originate and progress by accumulating mutations in one or more genes. Such mutations which result in cancer formation can be in proto-oncogenes or in tumor suppressor genes. Mutations in tumor suppressor genes result in loss of function, and therefore act in a recessive fashion to native genes. Oncogenes, in contrast, act in dominant fashion to native alleles and, therefore, are not usually inherited through the germ lines. The tumor suppressor genes, however, are found in inherited predispositions to cancer and are inherited as a dominant predisposition because of the high frequency of a second genetic event such as reduction in homozygosity[1].
Several tumor suppressor genes have been identified. Examples include the Rb gene, which is involved in retinoblastoma and osteosarcoma; p53, which is involved in osteosarcoma and adrenocortical, breast and brain cancers; WT-1, which is involved in Wilmsxe2x80x2 tumor, nephroblastoma and neurofibromatosis; adenomatous polyposis coli (APC), which is involved in adenomatous polyposis; and deleted colorectal cancer (DCC), which is involved with a somatic mutation in the colon.
The negative regulation of cell growth is effected by tumor suppressor proteins that regulate the cell S cycle by different mechanisms[2]. The gene cloned and sequenced as described herein, ING1 (formerly called p33IG1), represents a new tumor suppressor gene which is expressed in normal mammary epithelial cells, but expressed only at lower levels in several cancerous mammary epithelial cell lines and is not expressed in many primary brain tumors.
Known applications of sequenced genes include use of the DNA sequence (or analogs thereof) or of RNA or amino acid sequences derived from these DNA sequences for diagnosis or treatment of the corresponding disease. Accordingly, the gene ING1 (previously designated p33IG1) is useful for the diagnosis and treatment of breast and brain cancers among others.
The present invention is directed to a novel DNA sequence for an isolated gene (designated ING1). The DNA sequence, an RNA sequence identical to or complementary to the DNA sequence; the protein the DNA sequence encodes, p33ING1, and/or fragments or analogs thereof and antibodies which bind to p33ING1 which are useful for diagnosing and/or treating cancer.
One aspect provides a DNA sequence selected from the group consisting of a DNA isolate substantially identical to the p33ING1 DNA sequence shown in FIG. 2 and a DNA sequence greater than about 10 base pair (bp) in length capable of hybridizing under stringent conditions to the complement of the p33ING1 DNA sequence shown in FIG. 2. Recombinant expression vectors comprising such DNA isolates and cells transformed with such recombinant expression vectors are also provided.
Another aspect of the invention provides peptide or protein encoded by the DNA sequence substantially identical to the DNA sequence of FIG. 2 or a DNA sequence greater than about 10 base pair (bp) in length capable of hybridizing under stringent conditions to the complement of the p33ING1 DNA sequence shown in FIG. 2.
Another aspect of the invention provides a method for decreasing proliferation of mammalian cells comprising selecting said mammalian cells whose proliferation is to be decreased and increasing the expression of p33ING1 in said mammalian cells. It is comtemplated that said mammalian cells may be selected from the group consisting of normal cells and cancerous cells. It is further contemplated that the method of decreasing the expression of p33ING1 will comprise introducing into said mammalian cells at least one composition selected from the group consisting of p33ING1 and nucleotides which code for p33ING1.
Another aspect of the invention provides a method of diagnosing breast cancer comprising obtaining a biological sample comprising mammary cells suspected of being neoplastic and determining whether or not the biological sample contains p33ING1 or the DNA which encodes p33ING1, wherein the presence of p33ING1 or its DNA denotes non-cancerous cells.
Another aspect of the invention provides a method of diagnosing breast cancer comprising obtaining a biological sample comprising mammary cells suspected of being neoplastic, contacting said biological sample with at least one antibody to p33ING1 under conditions wherein antibody binding to p33ING1 occurs; and detecting whether or not said antibody binds to said cells, wherein binding to said cells indicates that said cells are non-cancerous.
One aspect of the invention provides nucleic acid isolates greater than 10 nucleotides in length which are substantially identical to the DNA sequence of FIG. 3 or its complement. Recombinant expression vectors comprising such sequences and cells transformed with such recombinant expression vectors are also provided.
One other aspect of the invention provides for a nucleic acid sequence which encodes the amino acid sequence of FIGS. 2 or 3 and recombinant expression vectors comprising such sequences and cells transformed with such recombinant expression vectors.
One other aspect of the invention provides for peptides and proteins having p33ING1 biological activity. It is contemplated that such peptides will have an amino acid sequence substantially identical to the amino acid sequence set forth in FIGS. 2 or 3.
Still a further aspect of the invention provides for antibodies to the p33ING1 protein.
A further aspect of the invention provides methods for decreasing proliferation of cancer cells in a patient comprising administering an effective amount of the above-described nucleic acid isolates, nucleic acid sequences, proteins or peptides under conditions wherein p33ING1 is expressed in the cancer cells. It is contemplated that the cancer is selected from the group consisting of breast and brain cancers.
Another further aspect of the invention provides methods of increasing cell proliferation of mammalian cells by decreasing expression of p33ING1 in the cells. It is contemplated that such methods could include the administration of either a single-stranded oligonucleotide comprising a sequence substantially identical to the complement of the cDNA sequence of FIG. 3 or the administration of a single or double-stranded oligonucleotide under conditions that a single-stranded oligonucleotide comprising a sequence substantially identical to the complement of the cDNA sequence of FIG. 3 is expressed in the cells. It is further contemplated that chemical inhibitors of p33ING1 activity could also be administered.
A yet further aspect of the invention provides a method for diagnosing cancer comprising obtaining a biological sample comprising cells suspected of being neoplastic and detecting whether or not the biological sample contains only the native ING1 gene, or expresses native ING1 mRNA or p33ING1, wherein the presence of only the native ING1 gene or expression of native ING1 mRNA or p33ING1 denotes non-cancerous cells. Preferably the cancer is breast cancer or brain cancer.
A still further aspect of the invention provides a kit for the detection of neoplastic cells in a biological sample comprising cells suspected of being neoplastic comprising a solid support for attaching the mRNA from the cell or tissue to be tested and a labelled polynucleotide of at least 10 nucleotides which polynucleotide is substantially identical to the sequence of FIG. 3 or its complement.
A still further aspect of the invention provides a kit for the detection of neoplastic cells in a biological sample comprising cells suspected of being neoplastic comprising a solid support for attaching the cells, an anti-p33ING1 antibody and a detectable label.