CYP3A4 NFSE variant and methods of use therefore

Details CYP3A4 NFSE variant and methods of use therefore CYP3A4 NFSE variant and methods of use therefore

1999-08-11

2001-01-16

Fredman, Jeffrey (Department: 1655)

Chemistry: molecular biology and microbiology

Measuring or testing process involving enzymes or...

Involving nucleic acid

C435S091100, C435S091200, C536S023100, C536S024300

Reexamination Certificate

active

06174684

ABSTRACT:

FIELD OF THE INVENTION
The present invention relates to a nucleic acid sequence encoding a novel CYP3A4 variant, referred to herein as CYP3A4-V. CYP3A4 is a metabolizer of multiple drugs. Thus, identification of this polymorphism is believed to be useful in predicting susceptibility of individuals to a broad spectrum of diseases and/or treatments. For example, detection of this variant can be used to predict risk for development of prostate cancer in a patient. Detection of this variant can also be used to predict risk for development of treatment-related leukemia in a patient upon administration of an epipodophyllotoxin. Kits for detection of this variant are thus provided. Also provided in the present invention are methods of identifying more effective treatment regimes for individuals with prostate cancer and safer anticancer drugs which will not lead to treatment-related leukemia through the identification of individuals carrying CYP3A4 variants.
BACKGROUND OF THE INVENTION
Prostate cancer is the most commonly diagnosed nondermatologic cancer in the United States among men. It has been estimated that over 325,000 new cases of prostate cancer are diagnosed in the United States each year, with over 40,000 fatalities annually.
The etiology of prostate cancer involves the effects of androgens as well as inherited genotypes that regulate androgen metabolism. Candidate prostate cancer genes include those involved in androgen metabolism, such as the androgen receptor (Giovannucci et al. Proc. Natl Acad. Sci. USA 1997 94(7):3320-3; Ingles et al. J. Nat. Cancer Inst. 1997 89(2):166-170) or 5-&agr;-reductase type II (Reichardt et al. Cancer Res. 1995 55(18):3973-5) genes. Additional candidates include members of the cytochrome P450 supergene family involved in androgen metabolism.
One member of this multigene family is CYP3A4, a gene involved in the oxidation of testosterone to 2&bgr;-, 6&bgr;-, or 15&bgr;-hydroxytestosterone (Waxman et al. Arch. Biochemical Pharmacology 1988 263:242-436). Substantial interindividual variability in metabolism of specific compounds by CYP3A4 has been reported (Kleinbloesem et al. Biochemical Pharmacology 1984 33:3721-3724), yet no genetic basis for this variability has been found. CYP3A4 protein has been reported to be expressed in only 61% of prostate tumors (Murray et al. J. Pathology 1995 177:147-152).
Second cancers are uncommon events occurring at a frequency of about 7% in survivors of primary malignant neoplasms. Leukemias are the major type of second cancers resulting from chemotherapy. There are two main forms of treatment-related leukemia, those with chromosome 5 and 7 monosomies induced by alkylating agents, and those with MLL gene translocations and other translocations related to DNA topoisomerase II inhibitors. Since only a minority of patients develop leukemia following chemotherapy, it has been suggested that differences in drug interactions with the host may be the predisposing factors (Boice et al. Proc. AACR 1997 38:645).
Genetic polymorphisms can account for large differences in the pharmacokinetics of chemotherapeutic agents, but metabolism of the majority is polygenetically determined and unimodally distributed. There is a 5- to 20-fold interindividual variability in drug clearance, which is a consequence of genetic and non-genetic factors. CYP3A-mediated first pass metabolism occurs after oral drug administration and has been suggested to contribute to the variability. CYP3A activity can also be modulated by inducers such as rifampin and anticonvulsants, inhibitors such as azole antifungal agents and macrolide antibiotics, by liver disease and by aging (Wilkinson, G. R. J. Pharmacokinet. Biopharm. 1996 24:475-490).
Epipodophyllotoxins are associated with leukemias characterized by translocation of the MLL gene at chromosome band 11q23 and other translocations. The epipodophyllotoxins, etoposide and teniposide and cyclophosphamide, ifosphamide, vinblastine and vindesine are substrates for metabolism by CYP3A.
A variant in the 5′ promoter region of the CYP3A4 gene has now been identified. This polymorphism comprises an A→G transition in the nifedipine-specific response element (NFSE) of the gene. Detection of this polymorphism is useful as a biomarker in predicting prostate cancer and epipodophyllotoxin-induced leukemogenesis.
SUMMARY OF THE INVENTION
An object of the present invention is to provide a nucleic acid sequence encoding a variant of CYP3A4.
Another object of the present invention is to provide a method of identifying patients with heightened risk of developing or having prostate cancer which comprises obtaining a biological sample from the patient and testing for the presence of a nucleic acid sequence encoding a variant of CYP3A4 in the sample, wherein the presence of this variant is indicative of a heightened risk of the patient developing or having prostate cancer.
Another object of the present invention is to provide a method of identifying patients at risk for developing treatment-related leukemia upon administration of an epipodophyllotoxin which comprises obtaining a biological sample from the patient and testing the sample for the presence of wildtype CYP3A4 or a variant CYP3A4, wherein the presence of wildtype CYP3A4 is indicative of an increased risk of the patient developing treatment-related leukemia upon administration of an epipodophyllotoxin while the presence of variant CYP3A4 is indicative of a decreased risk of the patient developing treatment-related leukemia upon administration of an epipodophyllotoxin.
Another object of the present invention is to provide a kit for identifying patients with heightened risk of developing or having prostate cancer or decreased risk for developing treatment-related leukemia upon administration of an epipodophyllotoxin, wherein said kit comprises a means for detecting wildtype CYP3A4 or a variant CYP3A4 in a biological sample.
Another object of the present invention is to provide methods for identifying more effective treatment regimes for prostate cancer which comprises identifying individuals suffering from prostate cancer who carry a variant CYP3A4 gene and selecting a treatment regime which is more effective in the presence of the variant CYP3A4 gene.
Yet another object of the present invention is to provide a method of identifying safer anticancer treatments for a patient suffering from cancer which comprises identifying whether a patient carries wildtype CYP3A4 or a variant CYP3A4 and selecting a safer treatment regime for patients carrying wildtype CYP3A4 or individualizing treatment doses based on whether the CYP3A4 genotype is variant or wildtype.
DETAILED DESCRIPTION OF THE INVENTION
CYP3A4 (OMIM*124010, GenBank D11131, SEQ ID NO:1) is a member of the cytochrome P450 supergene family involved in the metabolism of numerous compounds including aflatoxin B1, steroid hormones such as testosterone and estrogen, and numerous drugs (Li et al. Toxicol. 1995 104:1-8). A variant in the 5′ promoter region of the CYP3A4 gene has now been identified. This variant is referred to herein as CYP3A4-V (SEQ ID NO:2). This genetic variant has been found to disrupt a regulatory element upstream of CYP3A4. Specifically, CYP3A4-V comprises an A to G transition that alters the 10 base pair nifedipine-specific element located −287 to −296 base pairs from the transcription start site of CYP3A4. As a major metabolizer of hundreds of drugs, the applicability of this polymorphism to susceptibility of individuals to various treatments and/or diseases is believed to be quite broad.
For example, it has also now been found that prostate cancer patients who carry this variant allele have a higher clinical stage than patients who do not carry this variant. It is believed that CYP3A4-V genotype carriers may have decreased CYP3A4 activity, and thus decreased 2&bgr;-, 6&bgr;-, and 15&bgr;-testosterone oxidation. This decreased oxidation is believed to increase the bioavailability of testosterone for conversion to its intracellular mediator, dihydrotestosterone (DHT)

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