Chemistry: molecular biology and microbiology – Measuring or testing process involving enzymes or... – Involving nucleic acid
Reexamination Certificate
2001-06-28
2003-12-16
Benzion, Gary (Department: 1637)
Chemistry: molecular biology and microbiology
Measuring or testing process involving enzymes or...
Involving nucleic acid
C536S023100, C536S024320, C536S024330, C435S091100, C435S091200
Reexamination Certificate
active
06664057
ABSTRACT:
FIELD OF THE INVENTION
This invention pertains to the field of cancer genetics and cytogenetics. In particular, this invention pertains to the identification of nucleic acid sequences associated with a novel amplicon on human chromosome 20 which is associated with cancer. More particularly this invention pertains to the identification of a novel “amplicon” in a region of genomic nucleic acid amplification at about 20q13. These nucleic acid sequences can be used as probes in the diagnosis and prognosis of various cancers.
BACKGROUND OF THE INVENTION
Chromosome abnormalities are often associated with genetic disorders, degenerative diseases, and cancer. The deletion or multiplication of copies of whole chromosomes and the deletion or amplifications of chromosomal segments or specific regions are common occurrences in cancer (Smith (1991)
Breast Cancer Res. Treat.
18: Suppl. 1:5-14; van de Vijer (1991)
Biochim. Biophys. Acta.
1072:33-50). In fact, amplifications and deletions of DNA sequences can be the cause of a cancer. For example, proto-oncogenes and tumor-suppressor genes, respectively, are frequently characteristic of tumorigenesis (Dutrillaux (1990)
Cancer Genet. Cytogenet.
49: 203-217). Clearly, the identification and cloning of specific genomic regions associated with cancer is crucial both to the study of tumorigenesis and in developing better means of diagnosis and prognosis.
Studies using comparative genomic hybridization (CGH) have revealed approximately twenty amplified genomic regions in human breast tumors (Muleris (1994)
Genes Chromosomes Cancer
10:160-170; Kalliioniemi (1994)
Proc. Natl. Acad. Sci. USA
91:2156-2160; Isola (1995)
Am. J. Pathol.
147:905-911). These regions are predicted to encode dominantly acting genes that may play a role in tumor progression or response to therapy. Three of these amplified regions have been associated with established oncogenes: ERBB2 at 17q12, MYC at 8q24 and CCND1 and EMS1 at 11q13. In breast cancer, ERBB2 and CCND1/EMS1 amplification and overexpression are associated with decreased life expectancy (Gaudray (1992)
Mutat. Res.
276:317-328; Borg (1991)
Oncogene
6:137-143). MYC amplification has been associated with lymph node involvement, advanced stage cancer and an increased rate of relapse (Borg (1992)
Intern. J. Cancer
51:687-691; Berns (1995) gene 159:11-18). Clearly, the identification of additional amplified genomic regions associated with breast cancer or other tumor cells is critical to the study of tumorigenesis and in the development of cancer diagnostics.
One of the amplified regions found in the CGH studies was on chromosome 20, specifically, 20q13. Amplification of 20q13 was subsequently found to occur in a variety of tumor types and to be associated with aggressive tumor behavior. Increased 20q13 copy number was found in 40% of breast cancer cell lines and 18% of primary breast tumors (Kalliioniemi (1994) supra). Copy number gains at 20ql3 have also been reported in greater than 25% of cancers of the ovary (Iwabuchi (1995)
Cancer Res.
55:6172-6180), colon (Schlegel (1995)
Cancer Res.
55:6002-6005), head-and-neck (Bockmuhl (1996)
Laryngor.
75:408-414), brain (Mohapatra (1995)
Genes Chromosomes Cancer
13:86-93), and pancreas (Solinas-Toldo (1996)
Genes Chromosomes Cancer
20:399-407). The 20q13 region was analyzed at higher resolution in breast tumors and cell lines using fluorescent in situ hybridization (FISH). A 1.5 megabase (Mb) wide amplified region within 20q13 was identified (Stokke (1995)
Genomics
26:134-137); Tanner (1994)
Cancer Res.
54:4257-4260). Interphase FISH revealed low-level (>1.5×) and high level (>3×) 20q13 sequence amplification in 29% and 7% of breast cancers, respectively (Tanner (1995)
Clin. Cancer Res.
1:1455-1461). High level amplification was associated with an aggressive tumor phenotype (Tanner (1995) supra; Courjal (1996)
Br. J. Cancer
74:1984). Another study, using FISH to analyze 14 loci along chromosome 20q in 146 uncultured breast carcinomas, identified three independently amplified regions, including RMC20C001 region at 20q13.2 (highly amplified in 9.6% of the cases), PTPN1 region 3 Mb proximal (6.2%), and AIB3 region at 20q11 (6.2%) (Tanner (1996)
Cancer Res.
56:3441-3445). Clearly, definitive characterization of amplified regions within 20q13 would be an important step in the diagnosis and prognosis of these cancers.
Increased copy number of chromosome 20q in cultured cells also has been associated with phenotypes characteristic of progressing tumors, including immortalization and genomic instability. For example, increased copy number at 20q11-qter has been observed frequently in human uro-epithelial cells (HUC) (Reznikoff (1994)
Genes Dev.
8:2227-2240) and keratinocytes (Solinas-Toldo (1997)
Proc. Natl. Acad. Sci. USA
94:3854-3859) after transfection with human papilloma virus (HPV)16 E7 or HPV16, respectively. In addition, increased copy number at 20q13.2 has been associated with p53 independent genomic instability in some HPV16 E7 transfected HUC lines (Savelieva (1997)
Oncogene
14:551-560). These studies suggest that increased expression of one or more genes on 20q and especially at 20q13.2 contribute to the evolution of breast cancer and other solid tumors. Several candidate oncogenes have been identified as amplified on 20q, including
AIB
1 (Anzick (1997)
Science
277:965-968),
BTAK
(Sen (1997)
Oncogene
14:2195-200),
CAS
(Brinkmann (1996)
Genome Res.
6:187-194) and
TFAP
2
C
(Williamson (1996)
Genomics
35:262-264). Clearly, definitive characterization of nucleic acid sequences in 20q13 associated with tumor phenotypes would be an important step in the diagnosis and prognosis of these cancers. The present invention fulfills these and other needs.
SUMMARY OF THE INVENTION
The present invention relates to the identification and genomic mapping of new regions of nucleic acid associated with cancer and tumorigenesis.
The invention provides a novel method for screening for the presence of an amplicon in a sample of human nucleic acid. The first step of this method provides a sample of nucleic acid derived from a human cell and a probe, where the probe comprises nucleic acid which hybridizes specifically to a nucleic acid sequence including from D20S211 through D20S120. The second step involves contacting the human nucleic acid with the probe, where the probe is contacted with the human genomic nucleic acid under conditions in which the probe binds selectively under stringent conditions to the human genomic nucleic acid to form a hybridization complex. The last step is detecting the formation of the hybridization complex. In one embodiment, the human nucleic acid can be genomic DNA, which can be isolated from a breast tumor cell. The detection step can further comprise determining the copy number of the amplicon.
In this method, the probe can also comprise a nucleic acid which hybridizes specifically to a nucleic acid sequence spanning the distance between D20S 120 and D20S211. In alternative embodiments, the probe can comprise a nucleic acid which hybridizes specifically to a STS marker selected from the group consisting of AFMa233wg1, AFM080ya1, AFM069ya1, WI-16748, WI-9939, AFMa072zb9, WI-6578, AFM224zd12, WI-9227, and AFM276xh1. The probe can also comprise a nucleic acid which hybridizes specifically to a GDB locus nucleic acid sequence selected from the group consisting of D20S211, D20S854, D20S876, D20S1044, D20S913, D20S720, and D20S 120. Alternatively, the probe can comprise a nucleic acid which hybridizes specifically to a cloned genomic nucleic acid sequence selected from the group consisting of RMC20B4097, RMC20B4103, RMC20P4016, RMC20B4130, RMC20P4185, RMC20B4188, RMC20B4109, RMC20P4010, RMC20P4028, RMC20P4003, RMC20B4099, RMC20P4018, RMC20P4069, RMC20B4121, RMC20B4087, and RMC20P4070.
In another embodiment, the probe can comprise a polymerase chain reaction primer pair capable of amplifying some or all of the nucleic acid sequence including from D20S211 through D20S120. The detection step can
Albertson Donna G.
Collins Colin Conrad
Gray Joe W.
Pinkel Daniel
Benzion Gary
The Regents of the University of California
Townsend and Townsend / and Crew LLP
Wilder Cynthia
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