The main goal of this project is to investigate whether over-expression of cytochrome P450 1A1 (CYP1A1) and cytochrome P4501 B1 (CYP1B1) genes can be risk factors for race-related prostate cancer. Rationale is that African-Americans have twice the risk of Whites for presenting with advanced-stage prostate cancer. The mortality of African-American men 40-60 years of age due to prostate cancer is almost 2-3 folds greater than that of White men of the same age. The genetic basis for racial / ethnic differences in prostate cancer incidence is not known. To address this problem, we have targeted the CYP1A1 and CYP1 B1 genes because their products (2-OH-E2 and 4-OH-E2) are highly carcinogenic. This is a first step towards investigating the genetic basis for racial / ethnic differences in prostate cancer incidence. Based on our prior publications and preliminary data, we hypothesize that the CYP1A1 and CYP1 B1 genes are risk factors for race-related prostate cancer.
Specific Aim # 1: To test the hypothesis that CYP1A1 and CYP1 B1 genes are hyper-activated during malignant transformation of race-related prostate cancer. Under this specific aim, we will analyze mRNA expression of the CYP1A1 and CYP1 B1 genes by real-time PCR in different stages and grades of race-related prostate cancer. In situ hybridization will be used to localize gene expression in prostate tissues. Enzyme activities of CYP1 A1 and CYP1 B1 will be analyzed in prostate tissues by radio-isotopic methods.
Specific Aim # 2: To test the hypothesis that single nucleotide polymorphisms of CYP1A1 and CYP1 B1 genes are risk factors for prostate cancer. Under this specific aim, we will analyze single nucleotide polymorphisms of four polymorphic sites of the CYPIA1 gene and six polymorphic sites of the CYP1 B1 gene in different stages and grades of race-related prostate cancer. SNPs will be analyzed by PCR-RFLP (restriction fragment length polymorphism) and direct genomic sequencing.
Specific Aim # 3: To test the hypothesis that transfection of polymorphic variants of CYP1A1 and CYP1 B1 in E.Coli can hyperactivate estrogen hydoxylase activities using site-directed mutagenesis assays. We will investigate whether polymorphic variants of CYP1 A1 and CYP1 B1 have higher estrogen hydroxylase activities as compared to wild-type. We will also investigate whether estrogen hydroxylase activities and estrogen metabolites (2-OHE2 and 4-OH-E2) are associated with prostate cancer recurrence.
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