Affiliations of authors: A. F. Gazdar, The Hamon Center for Therapeutic Oncology Research and Department of Pathology, The University of Texas Southwestern Medical Center, Dallas; B. Czerniak, Department of Pathology, The University of Texas M. D. Anderson Cancer Center, Houston.
Correspondence to: Adi F. Gazdar, M.D., The Hamon Center for Therapeutic Oncology Research, 6000 Harry Hines Blvd., Dallas, TX 753908593 (e-mail: gazdar{at}simmons.swmed.edu).
Because most advanced invasive or metastatic cancers have low cure rates, risk assessment and early detection of the clinically occult premalignant phases of neoplasia are of particular importance. The National Cancer Institute (Bethesda, MD) has recently implemented an innovative new program, the Early Detection Research Network (its website is http://cancer.gov.edrn), where clinicians and scientists collaborate closely to develop and validate new markers and technologies for achieving these goals.
For studies of risk assessment and early detection, carcinomas arising in the urinary bladder are as close to ideal as any of the cancers arising in the internal organs. Urinary bladders are highly accessible and can be monitored by a variety of noninvasive or minimally invasive techniques. The entire mucosal surface at risk, the bladder urothelium, can be examined and biopsy specimens can be taken via a simple endoscopic procedure; moreover, exfoliated cells or secreted products can be examined repeatedly in voided urine or bladder washes with minimal or no risk to the patient. By contrast with voided urine, bladder washes are collected by cystoscopy. However, bladder washes provide the advantage of collecting larger numbers of well-preserved, recently shed cells in an isotonic fluid. Consequently, bladder washes may be useful for studies of some of the less stable cellular constituents, such as messenger RNA. In this issue of the Journal, Hemstreet et al. (1) describe the usefulness of measuring three biomarkers in voided urine for risk assessment and cancer detection in a large cohort at increased risk of bladder cancer.
Urinary bladder cancer is the fifth most common cancer in the Western world and is responsible for about 3% of all cancer-related deaths. Approximately 53 000 cases of bladder cancer were diagnosed in the United States in the year 2000, and approximately 23% of these patients will eventually die of their disease (2). Approximately 90% of bladder tumors are carcinomas derived from the bladder epithelial surface (the urothelium). In most regions of the world, transitional cell carcinoma is the most common histologic type of bladder cancer. Current concepts postulate that the common transitional cell cancers, papillary and nonpapillary, arise via two distinct but overlapping pathogenetic pathways (3,4). Approximately 80% of urothelial bladder cancers are superficial, growing as exophytic papillary lesions, which may recur but usually do not invade and metastasize. These cancers originate from hyperplastic urothelium. The remaining 20% of urothelial bladder cancers are highly aggressive, solid, nonpapillary carcinomas, which have a strong propensity to invade and metastasize. The vast majority of invasive bladder cancers occur in patients without a history of papillary tumors and originate from clinically occult, mild to moderate dysplasia (low-grade intraurothelial neoplasia), progressing to severe dysplasia and carcinoma in situ (high-grade intraurothelial neoplasia) and then to invasive cancer. However, in some patients who present with low-grade superficial papillary lesions, intraurothelial neoplasia may eventually develop and progress to invasive cancer. In such instances, urothelial dysplasia and/or carcinoma in situ may develop in the adjacent urinary bladder epithelium or within the superficially growing papillary lesions.
Habitual exposures to chemical carcinogens, either environmental or industrial, are well-established major risk factors for the development of bladder cancer (5). Cigarette smoking is the most important risk factor, contributing to approximately 50% of all bladder cancers in men. Occupational exposures to aromatic amines, including benzidine, contribute to approximately 25% of bladder cancers in some Western countries and possibly to a higher percentage in some developing countries (6). Inherited polymorphisms of enzymes involved in the metabolism of aromatic amines may modulate the risk of bladder cancer in exposed individuals (7).
The mean time from the initial carcinogen exposure to the development of urothelial bladder cancers is 18 years (5). This time period provides ample opportunities for finding the genotypic and phenotypic fingerprints of clinically occult premalignant conditions that are the antecedents to clinically aggressive invasive disease. Cystoscopy (a minimally invasive procedure) and histopathologic examination are the gold standards for bladder cancer diagnosis but are unsuitable for mass screening. However, examination of urine, which is voided as a natural physiologic function, is suitable for mass screening studies. Although routine screening by urine cytology is not helpful for risk assessment or identification of early preinvasive phases of bladder neoplasia, the technique is effective in diagnosing high-grade invasive cancer and carcinoma in situ (8). Because many genotypic and phenotypic changes precede progression to invasive cancer and may begin in cytologically normal cells, urine testing for changes antecedent to the development of invasive disease may detect occult intraurothelial neoplasia. Thus, the identification of genotypic and phenotypic changes that predict the tendency of in situ preneoplastic conditions to progress to invasive carcinoma is of particular importance. The U.S. Food and Drug Administration has approved urine tests for the detection of recurrent bladder tumors (9). However, none of these tests is approved or has been used for widespread screening, initial diagnosis, or risk assessment.
What important lessons can we learn from the study by Hemstreet et al. (1)? The authors studied a large cohort of 1788 Chinese men occupationally exposed to benzidine and 373 nonexposed workers. All workers provided urine samples that were tested by cytologic examination for hematuria and for measurement of three biomarkers initially and after 3 years. On the basis of the findings of the initial examination, workers were assigned to low-, moderate-, or high-risk groups, and each group was subject to a predefined intervention protocol. Twenty-eight cancers developed in the exposed group, and two developed in the nonexposed group. Surprisingly, the difference between the incidence rates of these groups was only threefold but may have been biased by the smaller size of the control group. Although the high- and moderate-risk groups constituted about 21% of the total subjects, 87% of the cancers arose in these groups. Subjects were predicted to have bladder cancer a mean time of 15 months (for those in the high-risk group with a positive biomarker profile) or 33 months (for those in the moderate-risk group with a positive biomarker profile) before clinical detection of their tumors. By contrast, a positive hematuria test preceded cancer detection by only 3 months. These findings validate the classification schema for cancer prediction. Unfortunately, we are not informed about the relationship between the smoking status of the workers and their risk classification or tumor development.
Because of the ease of obtaining voided urine, a large number of prospective biomarkers for bladder cancer risk assessment, early diagnosis, or recurrence has been investigated. These biomarkers include, but are not limited to, DNA ploidy and quantitative fluorescence image analysis (10,11), molecular cytogenetics (12), telomerase expression (13), various tumor-associated intracellular or secreted products (1417), oncogene mutations (18), and microsatellite alterations (19). Of the various markers, aneuploidy (i.e., altered DNA content and numeric chromosomal aberrations) is the most prevalent somatic alteration identified in human solid tumors (20,21). It has been well documented that aneuploidy is an early event in carcinogenesis and may drive tumor progression from a preneoplastic condition to invasive disease. Therefore, measurements of the total nuclear DNA content by image analysis or flow cytometry are used not only to evaluate the clinical aggressiveness of advanced tumors but also to assess risk in the clinically occult phases of neoplasia, as convincingly documented by Hemstreet et al. (1). These investigators examined cells in voided urine sediments of workers at increased risk of bladder cancer for three biomarkers: DNA ploidy, a bladder tumor antigen (p300), and a cytoskeletal protein (G-actin). These biomarkers were selected because of the investigators' extensive experience with them over the past several years (8,10). In this study (1), the risk of developing bladder cancer was nearly 20 times higher in workers positive for either aneuploidy analysis or p300 expression than in workers negative for both markers. G-actin was a poor marker for risk assessment.
The findings by Hemstreet et al. (1) do not exclude the numerous other markers awaiting large-scale, multi-institutional validation studies, although the cost, time, and enormous resources needed for such studies mandate that only the most promising can be fully tested. Most human cancers are considered to be genomic diseases in which multiple transforming and tumor suppressor genes additionally modified by epigenetic phenomenon contribute to the cancer phenotype. To address such global alterations, investigators are currently testing methods for the detection of literally thousands of expressed genes at the RNA or protein levels in clinical samples. These methods remain to be applied and tested in large-scale, population-based studies, and high throughput, cost-effective methodologies need to be developed. In the meantime, the study by Hemstreet et al. (1) will remain a landmark investigation for risk assessment of a large cohort of subjects at increased risk of bladder cancer.
NOTES
Supported by Public Health Service grants U01CA84971 (to A. F. Gazdar) and U01CA85078 (to B. Czerniak) from the National Cancer Institute, National Institutes of Health, Department of Health and Human Services.
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