Using statistical analyses of Fourier transform-IR spectra, we show that DNA of the histologically normal prostates of men 16 -80 years old undergoes structural changes in the bases and backbone with increasing age. Of the older men (ages 55-80), 42% exhibited a DNA phenotype mimicking that of primary prostate tumors from a comparable age group. This cancer-like phenotype, which was not found in the younger men (ages 16 -36), appears to arise from progressive age-related damage to DNA. The mean concentrations of 8-hydroxypurine lesions (e.g., 8-hydroxyguanine) were substantially higher for the older men than for the younger men. This finding suggests that the hydroxyl radical contributed to the structural changes that characterize the cancer-like phenotype. Strikingly, we were additionally able to discriminate between the DNA of primary prostate tumors and the DNA of primary prostate tumors from which distant metastases had been identified. Moreover, logistic regression analysis was able to predict the probability that a tumor had metastasized with Ϸ90% sensitivity and specificity. Collectively, these findings are particularly promising for identifying men at risk for developing prostate cancer, as well as for the early determination of whether a primary tumor has progressed to the metastatic state. This is highly important because the prognosis of histologically similar prostate carcinomas varies, thus creating a need to predict which cancers are most likely metastatic.I n the United States, prostate cancer is the most prevalent of all cancers and the second-leading cause of cancer mortality in men (1). The incidence rates for this disease rise markedly as a function of increasing age (1-3). Notably, 70% of all newly diagnosed prostate cancers occur in men over age 65 (2). The greatest threat from prostate cancer lies in its potential to metastasize (3). Clinical diagnosis of metastatic prostate cancer presently depends on histological identification of distant metastases. Yet, by the time the tumor has metastasized, the success of intervention is severely limited. Clearly, there is a need to identify early subcellular changes in the prostate, such as in DNA, associated with aging, primary tumor development, and the progression to metastasis.We have reported (4) that the ratio of mutagenic 8-hydroxy to putatively nonmutagenic ring-opened purine lesions in prostate DNA increased Ϸ3-fold in men between the ages of 16 and 83. In addition, a related study (5) showed a substantial accumulation of 8-hydroxyguanine (8-OH-Gua) in the liver, kidney, and intestine as rats aged. It was later reported (6) that levels of this base lesion increased 186% in the livers of rats between 5 and 30 months. Several factors have been suggested to explain agerelated increases in DNA base lesion concentrations in various tissues, among them reduced enzyme repair (7-10), a slow loss of DNA nuclease activity (5), and an increase in base oxidation (11). These findings focus attention on the importance of age-related free radical damage to...

Using the carcinogen 3-methylcholanthrene (MCA), we demonstrate with Fourier transform-infrared spectroscopy that a cancer DNA phenotype is produced well in advance of palpable tumors. We further demonstrate that the administration of cyclophosphamide markedly inhibits the development of the cancer phenotype and concomitantly delays tumor formation. MCA, injected into the hind legs of mice, produced a variety of significant structural changes in the nucleotide bases and phosphodiester-deoxyribose backbone, as reflected in a substantial (34%) difference between the mean DNA spectra of the control and the MCA-injected mice. Strikingly, 57 days before the mean appearance of tumors, we could not distinguish the DNA structure of the histologically normal tissues of the MCA-injected mice from the DNA structure of the tumor tissues. This finding indicates the early development of a cancer phenotype. Confirmatory evidence was obtained when tissues from a group of mice injected with both MCA and cyclophosphamide did not manifest the cancer phenotype, and their mean DNA structure closely resembled that of the control mice. Accordingly, we propose that the cancer DNA phenotype, as evinced by Fourier transform-infrared spectroscopy, is a promising early indicator of tumor formation, and we postulate that agents capable of inhibiting this phenotype may delay or prevent carcinogenesis.3-methylcholanthrene ͉ antineoplastic agents ͉ cyclophosphamide ͉ cancer inhibition ͉ cancer prediction

N-acetylcysteine (NAC), a strong antioxidant, has antigenotoxic and anticarcinogenic properties currently being investigated in clinical trials. NAC detoxifies free radicals (e.g., the hydroxyl radical, ⅐OH) that cause DNA changes implicated in disease (e.g., cancer). The ⅐OH reacts with purines to form mutagenic 8-hydroxypurine (8-OH) and putatively nonmutagenic formamidopyrimidine (Fapy) lesions. Fapy lesions inhibit DNA synthesis likely modulating the mutagenic potential of the 8-OH lesions, which would suggest that the ratio of these oxidized bases is biologically important. However, little is known about how NAC modifies oxidized DNA structure or how such modifications may affect cellular processes, such as replication and transcription. By using gas chromatography-mass spectrometry and Fourier transform-infrared spectroscopy, we found that dietary NAC (5% in the diet for 14 days) affected ⅐OH-induced structural changes in DNA of the hind leg of the BALB͞c mouse. For example, mutagenic 8-hydroxyguanine (8-OH-Gua) was reduced Ϸ50% (P ‫؍‬ 0.02) in mice fed NAC compared with controls. NAC reduced the log10 (8-OH-Gua͞FapyGua) ratio from 0.58 ؎ 0.15 to essentially zero, a virtually neutral redox status. DNA from control mice had a remarkably high variance compared with mice fed NAC. Moreover, the DNA from treated and control mice was distinct with respect to base structure and vertical base-stacking interactions. The findings showing that NAC lowered the concentration of 8-OH-Gua, the log ratio, and the variance (previously associated with neoplastic changes) suggest that NAC reduces the mutagenic potential of oxidized DNA. These benefits could be offset by the other structural changes found after NAC exposure, which may affect the fidelity of DNA synthesis.DNA damage ͉ free radicals ͉ N-acetylcysteine ͉ oxidation ͉ redox

A cancer DNA phenotype, identical to the DNA structure of tumors, has been identified in the prostate glands of certain healthy men over 55 years of age. We now show that the same DNA signature exists in normal tissues adjacent to tumors. This finding implies that the phenotype is maintained in normal prostate cells from its inception through tumor development. The presence of the phenotype in tumors, adjacent normal cells, and in the normal prostate cells of certain older men suggests that it is a potentially critical factor in tumor development and may serve as an early biomarker for cancer risk assessment. Intervention to inhibit the development of the phenotype in healthy men, or to eliminate it once formed, may suppress or even prevent tumor formation.cancer phenotype ͉ DNA structure ͉ cancer biomarkers ͉ cancer prediction and intervention ͉ Fourier transform-infrared spectroscopy U sing established Fourier transform-infrared (FT-IR) statistical models (1-5), we have demonstrated previously a cancer DNA phenotype in the limbs of mice 10 weeks after injecting the carcinogen 3-methylcholanthrene (4). This phenotype was identified in normal tissues 8 weeks before tumor formation. Strikingly, administration of the prodrug cyclophosphamide every 10 days after 3-methylcholanthrene injection delayed tumor formation by about 30% and almost completely suppressed development of the phenotype, thus suggesting that it may also be inhibited by other intervention strategies with less toxic side effects (4).In a previously reported study (2), a similar cancer DNA phenotype was identified in the prostate glands of certain healthy older men (55-80 years old). Both studies showed differences between the DNA of normal tissues and those tissues with the phenotype characterized by significant alterations in the base and backbone structures. In the prostate, for example, the development of the phenotype appeared to be age-related in that no evidence was found for it in men younger than 36 years of age. A statistical comparison of the FT-IR spectrum of the cancer phenotype with the DNA spectrum of normal prostate tissues of the healthy younger men revealed a broad array of differences in base structures (e.g., N-H and C-O), as well as in vertical base-stacking interactions. Various differences were also found in the phosphodiester-deoxyribose backbone (2). These differences likely arise from constituents in the microenvironment (e.g., reactive oxygen species) capable of inducing conformational changes in the backbone via disruptions in the planar base structure (6).The previous evidence (2) for a cancer DNA phenotype in certain older men suggests that this structure may respond differently in transcription and replication from the DNA of younger men, thus potentially increasing cancer risk. We now report that the cancer DNA phenotype in the prostate glands of older men is indistinguishable from the phenotype found in histologically normal tissues surrounding tumors. This finding implies that the phenotype is structurally stable, ...