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criteria are not being met (34). It is unre- alistic to think that the specialist genetics services will expand to cope with this, so the burden will fall on primary care and on hospital surgical clinics. It is here that in- formation and education must be targeted. Explicit guidelines have been published for the tbllow-up care of individuals found to have predisposing mutations for breast, ovarian, and colorectal cancer (20, 35). A widely available consensus statement with similarly explicit guidelines for family his- tory criteria that may merit specialist refer- ral for genetic testing might also be helpful (at present, it seems the best-publicized cri- teria are those put forward by commercial laboratories). Such guidelines would pro- vide reassurance to clinicians beset by de- mand and uncertain how to respond; and they will also encourage providers of health care that they will not be asked to meet an open-ended commitment. REFERENCES ANDNOTES 1. C.J.M. Ups etal., IV. Engl. J. M~. 331,828 (1994); H. Z. Nourani, H. N. Khan, B. L. Gallic, A. S. Detsky, Am. J. Hum. Genet. 59, 301 (1996); S. M. Powetl et aL, N. Engl, J. Med. 3~J, 1982 (199~. 2. B. Healey, N. Engl. J. M~d. ~.'.36, 1448 (1997}. 3. R. Hubbard and R. C. Lewontin, timid. 3,34, 1"192 (1996). 4. P. Kahn, Sc/ence 274, 496 (1996). 5. N. J. Nelson, J. Nat/. Cancer/nst. 88, 70 (1996). 6. A. Marshall, Nature Biotechno/. 14, 1642 (1996). 7. D. Smith, Nature Med. 3, 709 (1997). 8. D. F. Easton eta/., Am. J. Hum. Genet. 56, 265 9. M. P. M. Richardseta/.,J. Genet. Counse#ng4, 219 (1995); C. Lerman et aL, J, Am. Meal. Assoc. 275, 1885 (1996). 10. P. Williamson et al., J. R. Coll. Physicians London 30, 443 (1996). 11. H. J. Jarvinen, J. P. Mecklin, P. Sistonen, Gastroen- terology 108, 1405 (1995). 12. M. Dunlop and H. Campbell, Br. M~I. J. 314, 1779 (1997). 13. J. Streuwing eta/., N. Engl. J. Med. ~, 1401 (19973. 14. S, Lakenetal.,Natore Ge~et. 17', 79(19973. 15. Oncormed Inc., BRCA1/BRCA2 Fact Sheet (1997). 16. M. KraJner et al., N. EngI. J. Med. ~, 1416, 1997; F. J. Couch et aL, ibid., p. 1409. 17. J. Green, M. P. M. Richard& H. Statham, J. Genet. Counse/ing 6, 45 (1997). 18. F. S. Collins, N. Eng/. J. Meal. 334, 186 (1996). 19. S. Mazoye~, A. M. Dunning, O. Serova, Nature Gen- et. 14, 253 (1996). 20. W, Burke et a/., J. Am. Meal. Assoc. 277,997 ( 19973. 21. D. Easton, ~ture C-.-.-.-.-.-.-.-.-~net, 15, 210(19973. 22. N. Hallowell eta/., Psycho/. Hea/th Meal. 2, 149 (~9973. 23. M, P. M. Richard& Pub/ic Understanding ScL 5, 217 (1996). 24. D. Schrag, K. M. Kuntz, J. E. Garber, J. C. Weeks, N. EngL J. Meal. 336, 1465(1~973. 25. K. Rothenberg eta/., Sdence 275, 1755 (1997). 26. K. L. Hudson, K. H. Rofhenberg, L. B. A~drews, M. J. Pllis Kahn, F. S. Collins, ibid. 270, 391 (1995). 27. ~n of British Insurers, Information Sheet: Ufe Insurance and Gen~cs (ABI, I_~ndon, 1997). 28. NAPBC and NIH-DOE FISI Working Group, "Rec- ommandations on genetk~ informat/on and the work- place" (NAPBC, U.S. Public Health Service Office on Women's Health, Washington, DC, 19973. For fur- ther information see wv~t.napbc.org. 29. M. L. Srown and L G. Kessler, J. NatI. Cancer lnst. 1054 SCIENCE 87, 1131 (1995). 30. Task Fome on Genetic Test=ng of the NIH-DOE Working Group on Ethicah Legal and Social Impli- cations of Human Genome Research: Promottng Safe and Effective Genetic Testing tn the Un=ted States. 31. Nuflteld Council on Bioethics, Genetic Screening: Ethical I~sues (Nuffield Foundatioe, London, 1993). 32. Amedcen Society of Clinical Oncology, J. Clin. On- col. 14, 1730 (1996). &3. American Society of Human Genetics, Am. J. Hum. Genet. 55, i (1994), 34, F. M. Giardiello et al., N. Engl. J. Med. 336, 823 (1997). 35. W. BurkeetaI.J. Am. Med. Assoc. 277, 915 (1997), 36, I thank M. Richards for helpful discussions and M. Bobrow, L. Brody, F. Collins, D. Easton, M. Ponder, and M, Richards for cdtical comments on the manu- script, B.P. is a Gibb Fellow of the Cancer Research Campaign (CRC). Nucleic Acid-Based Methods for the Detection of Cancer David Sidransky 2063633082 Continued elucidation of the genetic changes that drive cancer.progression is yielding new and potentially powerful nucleic acid-based markers of neoplastic disease. Pilot studies indicate that these markers can be used to detect cancer cells in a variety of clinical settings with unprecedented precision. Nucleic acid-based markers may prove to be valuable tools for early detection of cancer in asymptomatic individuals, for con- firmation or exclusion of a cancer diagnosis that is based on suspicious but nondiag- nestle clinical material, for assessment of tumor burden in cancer patients, and for assessment of response to preventive approaches applied to healthy individuals who are at high dsk of developing cancer. Examples of these markers, their potential applications, and the current practical limitations on their clinical use are reviewed here. Recent discoveries in genetics and molec- ular biology have revolutionized our un- derstanding of cancer initiation and pro- gression. We now know that cancer is a heterogeneous group of diseases, each composed of a complex array of genetic changes driving uncontrolled growth and metastatic spread. Although this under- standing has stimulated the development of innovative molecular therapies for can- cer, successful introduction of these ther- apies into the clinical setting has been rare. Thus, a simple molecular cure for the most common cancers must still be viewed as a long-term goal. However, the war on cancer has many fronts. Identification of the genetic changes that drive cancer pro- gression is also providing us with a variety of molecular markers and tests that may ultimately redefine the criteria for cancer diagnosis and provide new avenues for early detection. Long before molecular cures for cancer arrive, accurate molecular diagnosis may change our clinical ap- proach to and management of cancer pa- tients. Here I will review the status of promising molecular tests for cancer, fo- cusing primarily on nucleic acid-based agnosis of epithelial cell malignancies, The author is at The Johns Hopkins University School of Medicine, Department of Otola~tngology--Head and which account for the overwhelming number of cancer deaths worldwide. Types of Molecular Markers Strong evidence supports the concept that cancer is a genetic disease that involves clonal evolution of transformed cells (1). Cancer cells arise through the accumula- tion of mutations, either inherited (germ- line) or acquired (somatic), in critical proto-oncogenes and tumor suppressor genes. Each mutation may provide an addi- tional growth advantage to the transformed cells as they dominate their normal coun- terparts (2, 3). The genetic alterations that arise during tumorigenesis can be used as targets for detection of cancer cells in clin- ical samples. DNA is an ideal substmte for molecular diagnosis became it readily sur- vives the adverse conditions experienced by many clinical specimens and it can be rap- idly amplified by polymerase chain reaction (PCR)-based techniques, thus diminishing the amount of starting material needed. In addition to specific mutations in on- cogenes and tumor suppressor genes, chang- es in DNA repeat sequences, called micro- satellites (4), can also be used as markers to detect the clonal evolution of neoplastic cells. Became they are highly polymorphic, microsatellite markers allow distinction of Neck Surgery, Division of Head and Neck Cancer maternal and paternal alleles. Typically, Research, 818 Ross Re~.~arch Building, 720 RutJand • ~ • c I ,-,,. T-- ; . 8A. E ma~l patrea samptes or normat umz~ [muatty Avenue, Baltimore, MD 21205-2195, U - ": dsidrans@we/chlink.welch.j.hu.edu " from blood lymphocytes) and DNA from a • VOL. 278 • 7 NOVEMBER [997 • www.sciencemag.org

clinical sample (a tumor or bodily fluid such as urine) are compared. The loss of one allele in the clinical sample [loss of het- erozygosity (LOH)] results from chromo- somal deletion or mitotic combination and is commonly thought to represent the sec- ond genetic inactivation step in the com- plete loss of a tumor suppressor gene locus (5). • Microsatellite analysis can also detect the presence of a new allele (detected as a mobility shift on ehctrophoretic gels) in- dicative of microsatellite instability. Wide- spread microsatellite instability, manifested as expansion or deletion of many repeat elements in tumor DNA, is particularly common in colorectal tumors; and in pa- tients with hereditary nonpolyposis colorec- tal carcinoma (HNPCC), it is caused by mutations Of DNA mismatch repair genes (6)~ Microsatellite instability can occur in many tumor types and can inactivate tumor suppressor genes, but more often it occurs in anonymous stretches of noncoding DNA. The detection of either of these genetic changes in a clinical sample (LOH or insta- bility, or both) demonstrates the presence of a clonal population of ceils that share altered genetic information, which is a characteristic of cancer cells. Mutations in proto-oncogenes and tu- mor suppressor genes can produce vast changes in the expression of many other genes. These changes can be assessed at the RNA level, although RNA is a less suitable substrate for clinical diagnosis than DNA because it is readily degraded. However, careful isolation of RNA from clinical sam- ples with subsequent conversion to cDNA and amplification [called reverse transcrip- tase (RT)-PCR] may be a more viable ap- proach to evaluating gene expression in the blood, lymph nodes, and bone marrow of cancer patients. Moreover, normal and neo- plastic cells are distinguished by the differ- ential expression of hundreds of cellular genes. New RNA-based methods for gene discovery that can track these changes in expression, including cDNA chip arrays (7) and serial analysis of gene expression (SAGE).. (8), will produce an ever-growing number of potential tumor markers. Another new marker is telomerase (dis- cussed below), a ribonucleoprotein enzyme that extends the sequences at chromosomal ends (telomeres) and is active in >90% of primary human tumors and cell lines (9) but inactive in most normal cells (I0). Sensitivity and Specificity Ltx~king for cancer cells in a clinical sample that contains a predominance of normal cells can be like looking for the proverbial needle in a haystack. This is especially true for molecular tests, which, unlike cytologi- cal tests breed on morphological assessment of individual or clustered cells, usually begin with the preparation of a specific substrate such as DNA from the admixture. The ratio of tumor cells to normal cells varies consid- erably from one organ s~rem to another and from one individual to another. Thus, molecular tests must he developed with a clear understanding of the clinical problem and the limits of the technology. A simple molecular test to identify bladder cancer cells in urine, where 50% or more of the DNA may be derived firom sloughed-off tu- mor cells, may he wholly inadequate for identification of lung cancer cells in spu- rum, where -<0.2% o~ the DNA is likely to be isolated from tumor cells. Conversely, exquisitely sensitive PCR-based approaches that can detect an abnormal transcript from one cancer cell among KYs normal cells may identify changes in single cells or cell clus- ters that. are not yet clonal or will not definitively progress to cancer (1 I). Identi- fication of molecular changes with this sen- sitivity may serve to identify patients at risk of developing cancer hut may he unsuitable for early detection. The precise cutoff for accurate detection of clinical disease has not been determined. Only prospective testing in patients at risk of cancer will empirically identify the critical threshold for accurate detection of the smallest tumors. Once the reliability of a technique is established through feasibility studies, its sensitivity and specificiW must then be as- sessed in formal clinical trials. Sensitivity refers to how often the test identifies cancer when it is present, and specificity refers to how often the test correctly identifies can- cer. If the prevalence of a specif'tc cancer type is low in the general population, the test must be exquisitely specific; otherwise more patients "without cancer may test positive. Applications Early detection. Because successful treat- ment. of most cancers depends on early de- tection, there is a critical need for new early detection approaches. Based on our emerg- ing knowledge of the underlying genetic events that lead to cancer initiation and progression, pilot studies have shovm that oncogenes and tumor suppressor gene mu- tations can be successfully identified in bodily fluids that drain from the organ af- fected by the tumor (Table 1). Using sen- sitive assays, investigators have found ras or p53 mutations in many bodily fluids of pa- tients, including blood (12-18). In all of these studies, the identical mutation present in the primary tumor was identified in the bodily fluid tested from affected patients. ras and p53 mutations have been used as molecular markers in these studies because they commonly occur in the tumor types tested (19) and because they may provide information about the staging of the tumor; for example, in colon cancer, ras mutations are an early event in tumorigenesis, whereas p53 mutations usually occur in invasive tu- mors (2). It would follow that APC muta- tions, which occur in more than 70% of colon adenomas (precursors to cancer), might also be valid markers (20). However, identification of all the possible mutations in the coding region of a gene, especially when it is the size of APC (8.5 kb) is daunting. Such "mutation scanning" to de- tect these alterations in an admixture of normal and neoplastic cells is not presently feasible. With the exception of K-ras muta- tions (clustered at codons 12 and 13) or a few mutation "hotspots" in p53 (1.2 kb), this technological barrier is preventing the development of the necessary assays for ini- tiation of clinical trials to validate this approach. Because of these technical limitations, there is a great need to identify other clonal DNA-based markers. Microsatellite analysis is emerging as an important and relatively easy alternative for cancer detection. In contrast to the use of specific probes to identify oncogene mutations, these molec- ular alterations are easily identified with one set of primers for all samples. In a retrospective analysis of urine samples from 25 patients, microsatellite markers success- fully detected over 90% of bladder tumors (21). In a follow-up study, 10 of 11 recur- rences were detected prospectively and two patients had a positive test several months before the clinical cancer was visualized by bladder inspection (cystoscopy) (22). A multi-institutional trial to test for bladder cancer recurrence using a panel of 20 mi- crosatellite markers is already under way. As noted above, microsatellite alter- ations (low-level instability) can be also be found in tumors without mismatch repair deficits. PCR with subsequent electro- phoretic separation of the PCR products can identify these shifts at a sensitivity of - 1 neoplastic cell among 500 normal cells, which appears to be sufficient for clinical detection in many situations. Certain mi- crosatellite markers are particularly unsta- ble in human tumors; these markers often contain larger repeats, particularly tet- ranucleotides (23). Interestingly, although some microsatellite markers are unstable in virtually all tumors, others are unstable only in specific tumor types. The mechanism underlying this phenomenon is unknown but may involve flanking DNA sequences, oo o3 o o~ w,vw,q~-iencema~.or~ • SCIENCE • VOL 27g • ? NOVEMBER 1007 1055

tissue-specific expression of genes in the surrounding chromosomal region,.or an un- derlying DNA repair deficit. As mentioned above, the ribonucleopro- rein enzyme telomemse is expressed selec- tively in virtually all primary tumors and therefore has emerged as a promising mo- lecular marker fbr cancer detection. Cur- rently, telomemse activity in clinical sam- ples is measured by the TRAP (telomerase repeat amplification protocol) assay, which requires protein extraction and subsequent primer-directed PCR amplification of telo- mere extensions (24). The specificity of this approach has been lower thab that reported in studies using detection of DNA alter- ations (25-27) (Table 1). The recent clon- ing of the human telomerase catalytic com- ponent (28) may allow development of im- proved assays. Tumor burden. Molecular markers can also be used to assess the migration of tumor cells locally or into the bloodstream. Sur- gery remains the most effective treatment for most localized primary tumors, but tu- mor cells often spread beyond the surgical margins and may evade detection by stan- dard light microscopy. Because the tumor cells are vastly outnumbered by normal cells in this situation, very sensitive detection techniques are required. In one study of patients, specific p53 mutations were used to identify infiltrating tumor cells in surgi- cal margins beyond the resection border (29). In -50% of the patients, tumor cells harboring the same mutations identified in the primary tumor were detected in appar- ently "clean" margins or lymph nodes. De- spite radiation treatment, "about one-third of the patients with these mutations went on to recur, often developing new tumors adjacent to or within the area identified as positive by molecular analysis. A similar analysis of p53 and ras mutations has iden- tified tumor cells in apparently disease-free lymph nodes of colorectal and lung cancer patients, but the clinical outcome of posi- tive patients was not provided for critical appraisal (30). The determination of node status is critical for precise staging of tumors and for treatment decisions. In addition to local spread, malignant cells can metastasize; that is, enter the bloodstream, disseminate, and grow in oth- er organs. In light of earlier studies indicat- ing daat cancer patients have large amounts of circulating DNA in serum or plasma (31), blood samples are now being analyzed for nucleic acid markers such as K-ras mu- tations and microsatellite alterations (32, 33). In 29% of 21 patients with head and neck squamous cell carcinoma (HNSCC) cancer and in 71% of 21 patients with small-cell lung cancer (SCLC), LOH or microsatellite alterations were detected in serum or plasma (33). In the HNSCC study, the positive patients had larger tu- mors and a poorer prognosis. The higher incidence of plasma DNA alterations in SCLC patients may reflect the tendency of these tumors to metastasize early (33). Al- though analysis of serum nucleic acid mark- ers does not currently allow early detection of tumors, it may provide useful information on tumor burden and response to therapy. Analysis of whole blood and bone mar- row (BM) for abnormal transcripts derived from neoplastic cells is routinely used to monitor patients with chronic myelogenous leukemia. More recently, transcripts ex- pressed exclusively or preferentially in can- cer cells have been targets of RT-PCR- based detection strategies in patients with solid tumors. For example, tyrosine hydrox- ylase transcripts were found to correlate with micrometastatic 8M disease in neuro- blastoma, and tyrosine transcript levels in melanoma may predict a poor prognosis (34). RT-PCR approaches targeting cyto- keratins, adhesion molecules, tyrosine ki- nases, and prostate-specific markers [includ- ing prostate-specific antigen (PSA) and prostate-specific membrane antigen PSM] to detect micrometastatic disease have been tested in various tumor types, including pri- mary bremt, gastric, colorectal, lung, and prostate cancer (35). However, issues of specificity remain because of illegitimate expression of these markers in normal cells and down-regulation of the markers in tu- mor cells. One recent study suggests that RT-PCR of PSA in bone marrow shows high specificity (no false positives in 53 control patients) for micrometastatic dis- ease in prostate cancer (36). Testing BM may be more relevant in some cases, be- cause animal studies suggest that only a small port/on of metastatic cells actually settle and develop into metastatic deposits in various organs. Adjuncts to cytolog~ and histop~tholo~. Needle aspirates from various organs are often used to establish the cancer diagnosis when there is a suspicious mass. In some cases, it is difficult to distinguish bet~'een benign or preneoplastic lesions and frank cancer. Recently, telomerase was detected in all 1 ! follicuhr carcinomas of the thyroid but in only 8 of 33 benign follicular tumors and never in normal thyroid tissue (37). Table 1. Selected feasibility tdals employing molecular diagnosis of clinical samples. This table lists selected pilot or feasibility trials using accessible clinical bodily samples in molecular detection approaches. These studies have employed both retrospective and prospective collection of samples, but only the study by Steiner et a/. (22) reports prospective follow-up of the patient cohort, Studies using gene targets (ras and p53) report sensitMty of the molecular assays as a fractfon of patients with tumors containing the sought-after mutation. The other studies report sensitivity as a fraction of all patients with cancer, regardless of the molecular status of their tumor. When microsatellites are used as targets, the number of markers in each study varies and is listed in parentheses. The upper limit of detection denotes the upper limit of sensitivity for the assay as a dilution of cancer cells among normal cells. Nipple aspirates and ejaculates are potential but unproven clinical samples for the detection of breast and prostate cancer, respec'dvely. SensitMty and spec~city for detecting cancer are listed as reported in each study. Upper limit Patients Controls ~ Specificity Cancer type Clinical Genetic marker Reference sample of detection (n) (n) (%) (%) Head and neck Saliva p53 1/10,000 7 0 71 100 Telomerase 1/10,000 44 22 32 95 Lung Sputum ras/p53 1/10,000 10 5 80 100 ms 1,10,000 5 30 100 100 Microsatellites (4) 1/500 5 0 60 100 Colon Stool ras 1/10,000 9 6 88 100 Telomerase 1/10,000 15 9 60 100 Pancreas Stool ms 1/10,000 11 3 66 100 Juice ras 1/100,000 7 3 100 100 Bladder Urine p53 1/10,000 3 3 100 100 Microsatellites (13) 1/500 20 5 95 1 O0 Microsatellites (20) 1/500 21 0 91 1 O0 Telomerase 1/10,000 26 83 62 96 49 .25 17 16 50 13 26 15 14 12 21 22 27 SCIENCE • VOL. 278 • 7 NOVEMBER t997 • www.sciencemag.org

Thus, detection of telomerase in needle biopsies from suspicious thyroid nodules may help establish the diagnosis of follicular carcinoma before proceeding to thyroidec- tomy. Others have used telomerase to cor- rectly establish the diagnosis from three suspicious but not diagnostic needle biop- sies taken from breast cancers (38). The Pap smear of the cervix is the single most successful effort in screening for can- cer that has been made in this century. Virtually all cervical cancers are associated with human papilloma virus (HPV) infec- tion, and investigators have developed a sensitive molecular test to identify HPV sequences in liquid cytology medium. In older women (who have a low prevalence of HPV infection) with borderline abnormal- ities, HPV testing identifies -90% of indi- viduals who have underlying high-grade neoplasia (39). Another perplexing problem in pathol- ogy is the need to identify the primary tumor when a patient presents only with a metastatic lymph node. This is especially common in the neck, where an occult pri- mary HNSCC tumor may be difficult to identify. Microsatellite analysis has proven useful even when random biopsies of the oral cavity and hypopharynx do not reveal the primary malignant focus (40). In pre- liminary studies with HNSCC, it was found that in 6 of 10 patients, the same genetic changes identified in the metastatic depos- its were present in at least one random mucosal biopsy (40). In two cases, the pri- mary tumor subsequently recurred in the predicted anatomical site identified by mo- lecular analysis. Intermediate biomarkers. Clonal genetic changes that can reliably predict the occur- rence of neoplasia well in advance of clin- ical cancer may have a role as intermediate biomarkers in cancer prevention studies. New opportunities may thus exist to test chemopreventive agents in populations without waiting many years for accurate statistical analysis from commonly used endpoints, such as survival or the onset of The notion that oncogene or tumor sup- pressor gene mutations can be used as mark- ers for early preneoplastic disease is support- ed by the recent observation of clonal K-ras mutations in hyperplastic intestinal crypts (41). Until the emergence of an APC mu- tation in these cell populations leads to' dysplasia, progression to cancer is unlikely (41). Other studies have demonstrated the presence of K-ras and APC mutations as well as microsatellite instability in the pre- neoplastic mucosa of patients with ulcer- ative colitis, who are also at high risk for colon cancer (42). K-ras mutations have also been found in the sputum of smokers without lung cancer (43). Thus, the iden- tification of K-ras mutations, especially in low proportion to normal DNA (for exam- ple, 1 in 10,000 to 100,000), may signal the presence of preneoplastic clones in addition to overt clinical lesions and supports the role of K.ras as an intermediate marker for monitoring and chemopreventive studies. p53 mutations are induced by a variety of endogenous and exogenous compounds. In patients with skin cancer, pyrimidine dimer mutations occur at specific sites in I)53 as a result of ultraviolet exposure. These muta- tions can also be found in normal skin of sun-exposed individuals, and their frequen- detection Sputum Tumor burden Blood [~ Establish diagnosis Needle biopsy Surgmy Removal of primary tumor Treatment Chemotherapy Radiation Extent of disease Lymph nodes, margins, bone marrow Prognosis Response to therapy Diagnosis Management Monltodng I~B/ood Rg. 1. Molecular diagnostic applications in' the overall management of cancer patients. The schematic depicts the opportunities for molecular analysis of clinical samples in a lung cancer patient as the patient moves from diagnosis, to surgery (for a localized tumor), to additional treatment, and eventually to long-term monitoring, These applications (with the exception of prognosis) are discussed in the text. cy correlates with overall sun exposure (44). Recently, a sensitive assay to detect these changes was used to compare the efficacy of various sunscreens in a mouse model (45). p53 mutation hotspots can be found in oth- er tumor types (for example, codon 249 mutations in liver cancer) and are much easier to test for than a whole army of mutations. From the Bench to the Bedside Figure i summarizes some of the clinical applications of the nucleic acid markers. Continued development of these markers will require the establishment of large biorepositories containing paired clinical samples of blood, tumors, and bodily fluids. To bring these new molecular approaches to the clinic, it is essential to carry out large well-controlled trials. Newly defined high- risk populations, such as carriers of germline mutations in cancer genes, will be crucial for successful implementation of these tri- als. These new molecular approaches can be tested quickly in populations at high risk for disease. The information garnered from these triaLs can then be incorporated into routine monitoring and perhaps screening 'Time (rain) Fig. ~. Molecular detection of bladder cancer by fluorescence-based PCR, microcapillary electro- phoresis, and laser detection. The figure shows electropherograms of PCR products dedved from PCR amplif~.,ations of the inte¢feron (IFNA) micro- satellite locus (chromosoma~ arm 9p21) in vadous samples from the same patient (provided by R. Mathie, University of California, Berkeley). Prod- ucts from the normal sample (h/rnphocytes) were generated with energy transfer (ET) pdmers and detected in tt~e green channel (537 to 567 rim), and those from tumor and udne samples were gener- ated ~ E-I" pdmers detected in the red channel (>590 nm). The reduced intensity of one allelic band in the udne and tumor samples as compared to ~e corresponding norrna~ DNA reflects LOH. The recent development of 96-well capillary array devices supports this assay as a high-throughput approach fo~ cancer detection (47). www.gciencema~.or~ • SCIENCE • \'OL. 278 • 7 NOVEMBER 1997 1057

for these patients and the population at large. As these patients are recruited for such trials, imlx~rtant ethical issues, includ- ing informed consent and insurance cover- age, must be appropriately addressed. Despite the great promise of these new molecular approaches for cancer detection, much of the current technology limits their implementation into routine clinical use even for high-risk populations. High- throughput technologies have to be devel- oped and integrated to make these assays a reality (Fig. 2). Genosensor arrays and mi- crocapillaty systems may make these tests accessible in the near future (46, 47). A positive molecular test is only useful if the tumor can be localized and eradicated. Current imaging approaches cannot reliably detect small tumor masses. For many pa- tients, identification of the primary tumor will result in cure but for others, a positive molecular test may be followed by negative imaging studies. 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