Chemistry: natural resins or derivatives; peptides or proteins; – Proteins – i.e. – more than 100 amino acid residues – Blood proteins or globulins – e.g. – proteoglycans – platelet...
Reexamination Certificate
1998-03-09
2001-11-13
Caputa, Anthony C. (Department: 1642)
Chemistry: natural resins or derivatives; peptides or proteins;
Proteins, i.e., more than 100 amino acid residues
Blood proteins or globulins, e.g., proteoglycans, platelet...
C530S387100, C530S388100, C530S388800, C530S387900, C530S389100, C530S389700
Reexamination Certificate
active
06316599
ABSTRACT:
FUNDING
Work described herein was funded by the National Institutes of Health, the Medical Research Council of Canada and the National Cancer Institute.
BACKGROUND
Wilms' tumor (WT) is an embryonal malignancy of the kidney which affects approximately 1 in 10,000 infants and young children. Matsunaga,
Human Genetics,
57:231-246 (1981). The molecular basis of Wilms' tumor is not well understood.
The majority of WTs are sporadic tumors (>95%). A small set of WTs occur in a hereditary setting as familial cases or as part of cogential disorders such as the WAGR syndrome. Wilms' tumor (W) cases (approximately 2%) can occur in association with aniridia (A), a defect in the development of the iris, as well as genitourinary (G) abnormalities and mental retardation (R). Miller et al.,
New Engl. J. Med.,
270:922-927 (1964). These disorders form the WAGR syndrome, and can be attributed to a constitutional deletion of DNA in band 11p13 on human chromosome 11 in a group of genes known as the WAGR complex. Riccardi, et al.,
Pediatrics,
61:604-610 (1978); Francke, et al.,
Cytogenet. Cell Genet.,
24: 185-192 (1979). In these cases, bilateral Wilms' tumors are frequently observed, as are dysplastic changes in surrounding renal tissue (nephroblastomatosis) which are though to precede malignant transformation (i.e., precancerous conditions). Bove and McAdams,
Perspectives on Pediatric Pathol.,
3:185-223 (1976). As a recessive oncogene or anti-oncogene, the Wilms' tumor locus is thought to curtail the growth of undifferentiated nephrotic cells. The genetics of WT generally conform to the two-hit mutational model of carcinogenesis used to describe the retinoblastoma locus on chromosome 13q. According to this concept, the rate-limiting events in tumor formation are two mutations. Sporadic tumors develop following two independent events, while familial cases involve transmission of one altered gene in the germline and a somatic mutation in the second gene. Hereditary cases display both a high penetrance and an increased incidence of bilateral cancers, attesting to the frequency of the somatic second event.
Observations have lead to the conclusion that at least in a subset of Wilms' tumors, the inactivation of a gene in 11p13, analogous to the retinoblastoma (RB) gene, is a key event in tumor formation. Considerable effort has been expended in attempting to localize the gene responsible for WT, as is evidence by the numerous reports describing such efforts. The localization of a Wilms' tumor gene at 11p13 is supported by the fact that 40-60% of individuals with the WAGR syndrome develop Wilms' tumor.
However, it appears that the genetics of Wilms' tumor are more complex than the two-hit mutational model. Based on additional research, it appears that Wilms' tumor may be caused by loss of function at alternative loci. In studies of two families showing hereditary predisposition to Wilms' tumor, linkage to 11p genetic markers was excluded, indicating the presence of at least one additional Wilms' tumor locus. Grundy, et al.,
Nature,
336:374-376 (1988); Huff, et al.,
Nature,
336:377-378 (1988). Further studies showed loss of heterozygosity in Wilms' tumors at 11p15 rather than 11p13. Reeve, et al.,
Mol. Cell Biol.,
9:1799-1803 (1989); Koufos, et al.,
Am. J. Hum. Gen.,
44:711-719 (1989); Koufos, et al.,
Nature,
309:170-172 (1984); Orkin, et al.,
Nature,
309:172-174 (1984); Reeve, et al.,
Nature,
309:174-176 (1984); Fearon, et al.,
Nature,
309:176-178 (1984). Although these data suggest the possibility of additional loci, the 11p13 Wilms' tumor locus is clearly associated with constitutional WAGR deletions and somatic chromosome rearrangements in a subset of sporadic tumors. Lewis, et al.,
Genomics,
3:25-31 (1988).
Despite considerable interest in identifying the Wilms' tumor gene and work focusing on doing so, to the present time, a transcript mapping to the region identified as containing the Wilms' tumor gene has not been identified.
SUMMARY OF THE INVENTION
The present invention relates to a method of analyzing cells for the Wilms' tumor gene, as well as to a method of analyzing cells for the Wilms' tumor gene transcript, the encoded polypeptides, and antibodies (polyclonal or monoclonal) which recognize all, or a portion of, the encoded polypeptides. As used herein, the term Wilms' tumor gene or Wilms' tumor locus refers to a specific gene in chromosome 11 band 13 (11p13) which is characteristicly altered in the WAGR syndrome or some sporadic Wilms' tumors (i.e., found in cells affected in these conditions), but which can reasonably be expected to be associated with or causative of other tumor types.
The present invention further relates to DNA sequences (SEQ ID NOS: 1 and 3), both genomic and cDNA clones, which map within the boundaries of constitutional and tumor deletions which physically define the Wilms' tumor locus on human chromosome 11 band p13 (11p13). For the first time, a gene which maps to the region containing the Wilms' tumor locus has been identified. The gene has been characterized and shown to span approximately 50 kb and to encode an mRNA (referred to as WT1 mRNA or transcript) approximately 3.1 kb in length. The WT1 mRNA has been shown to be expressed in a number of cell types. However, it is predominantly expressed in kidney and gonadal cells.
The amino acid sequence of the polypeptides encoded by the sequences have also been derived and features of the polypeptides have been examined (SEQ ID NOS: 2 and 4). Several of these features, such as the presence of four zinc finger domains and of a region rich in proline and glutamine (SEQ ID NO: 5), are indicative of a role in transcription regulation. The localization of the gene to 11p13, its tissue-specific expression and its predicted function, as well as the finding that this gene is specifically mutated in some Wilms' tumors, support the conclusion that it is the 11p13 Wilms' tumor gene. The present invention includes a method of identifying the Wilms' tumor gene; the isolated Wilms' tumor gene, the isolated gene transcript; the isolated encoded polypeptide; and diagnostic methods and reagents based thereon.
The present invention makes available for the first time a method of identifying in a sample, DNA which is clearly the 11p13 Wilms' tumor gene, an mRNA transcript thereof or a Wilms' tumor-encoded polypeptide, as well as materials (e.g., nucleic acid probes, anti-Wilms' tumor polypeptide antibodies) useful in these methods of identification. This is particularly valuable because the treatment of Wilms' tumor represents one of the clearest examples of success in pediatric oncology, as a result of the development of effective therapeutic regimens. However, for treatment to be most effective, the tumor must be diagnosed early. The present invention provides a means by which the risk of developing Wilms' tumor, for example, in diseases such as WAGR and Denys-Drash syndrome, can be assessed prior to its current method of detection. Often a Wilms' tumor mass is identified inadvertently by a parent when bathing the child. A diagnostic test would allow much earlier detection of the disease. The presence of the disease, once it has occurred, can be confirmed, thus making it possible to intervene therapeutically prior to or at an earlier stage in the development of the disease. It also provides a method by which the alteration of the WT1 gene can be detected in other tumor types which are known to express the WT1 transcript (e.g., leukemia cells, testicular tumors, ovarian tumors), using DNA probes or antibodies specific for Wilms' tumor gene-encoded polypeptide.
REFERENCES:
patent: PCT/UA90/06629 (1990-11-01), None
patent: WO 91/07509 (1991-05-01), None
patent: 95/29995 (1995-11-01), None
patent: 98/13494 (1998-04-01), None
Goding, J.W., Monoclonal Antibodies:Principles & Practice, 1986, Academic Press, pp. 281-282.*
Reeck et al., Cell, v
Breuning Wendy
Buckler Alan J.
Call Katherine M.
Darveau Andre
Glaser Thomas M.
Caputa Anthony C.
Hamilton Brook Smith & Reynolds P.C.
Massachusetts Institute of Technology
Nickol Gary B
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