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Abnormal p53 immunohistochemistry is associated with an increased colorectal cancer–related mortality in patients with ulcerative colitis
THE AMERICAN JOURNAL OF GASTROENTEROLOGY © 2003 by Am. Coll. of Gastroenterology Published by Elsevier Inc.
Vol. 98, No. 6, 2003 ISSN 0002-9270/03/$30.00 doi:10.1016/S0002-9270(03)00422-2
Abnormal p53 Immunohistochemistry Is Associated With an Increased Colorectal Cancer–Related Mortality in Patients With Ulcerative Colitis Bret A. Lashner, M.D., William M. Bauer, M.D., Lisa A. Rybicki, M.S., and John R. Goldblum, M.D. Center for Inflammatory Bowel Disease, Departments of Gastroenterology, Biostatistics, and Anatomic Pathology, Cleveland Clinic Foundation, Cleveland, Ohio
OBJECTIVE: Immunohistochemical evaluation of p53 staining patterns has been considered as a complementary test for dysplasia in ulcerative colitis-related colorectal cancer surveillance, but usefulness would be particularly important if it were a marker associated with a poor prognosis. The aim of this study was to determine whether abnormal p53 staining of the tumor correlated with cancer-related mortality in ulcerative colitis patients. METHODS: An historical cohort study was designed to examine all ulcerative colitis patients who developed colorectal cancer between 1978 and 1997 and who had tumor tissue blocks available for staining. Tissue was recut and stained for abnormal p53 immunohistochemistry using the DO-7 antibody. Tumors were considered to be p53 positive if at least 5% of nuclei in a high power field were positive for staining. RESULTS: Among 75 patients entered in the study, 38 (50.7%) had p53 positive tumors. Fourteen patients (36.8%) with p53 positive tumors died from colorectal cancer, compared to five (13.5%) with p53 negative tumors (p ⬍ 0.04, log-rank test). The adjusted relative risk of cancer-related death among patients with p53 positive tumors was 3.03 (95% CI ⫽ 1.05– 8.73). CONCLUSIONS: Abnormal p53 immunohistochemistry of tumors is associated with a poor prognosis among ulcerative colitis patients who develop colorectal cancer. As such, p53 immunohistochemical staining could be a useful histological marker to complement routine histology in cancer surveillance programs in ulcerative colitis patients. (Am J Gastroenterol 2003;98:1423–1427. © 2003 by Am. Coll. of Gastroenterology)
INTRODUCTION Tumor suppressor genes, including p53, act as a G1 checkpoint in the cell replication cycle for cells with abnormal DNA until DNA repair can occur, or until apoptosis ensues in the event of excessive DNA damage. Inactivation of suppressor gene function through mutations perpetuates abnormal DNA through loss of repair and/or apoptotic mech-
anisms, thus increasing the risk of malignant transformation (1). In many cases, a mutated, inactive p53 protein has a much longer half-life than the active wild-type protein, and immunohistochemical staining of p53 often will stain nuclei strongly with accumulated mutated proteins. Therefore, nuclear accumulation of the p53 protein product is a surrogate marker for p53 mutations. Sporadic colorectal cancers with p53 mutations are associated with a poor prognosis (2–5). In ulcerative colitis patients, abnormal p53 staining and loss of heterozygosity have been shown to be present in approximately one half of patients with cancer and an early marker of neoplastic progression, but its relationship to prognosis has not been determined (6 –14). The aim of this study was to determine whether positive p53 immunohistochemical staining of ulcerative colitis–related cancer is associated with a poor prognosis as defined by cancer-related mortality.
MATERIALS AND METHODS Between 1978 and 1997, all patients with ulcerative colitis– related colorectal cancer who had surgery at the Cleveland Clinic and who had specimens available in paraffin blocks were included. Patients with Crohn’s colitis or indeterminate colitis were excluded. Sections were cut and stained with a 1:200 dilution of the D0 –7 antibody for p53 (Dako, Carpinteria, CA) (15). Specimens were considered as positive for abnormal p53 if 5% of nuclei in any high power field stained. An example of a p53 positive staining in a tumor is shown in Figure 1. All specimens were read by an experienced gastrointestinal pathologist (J.R.G.) who was blinded regarding the outcome of each patient. Clinical information was gathered from medical records. A historical cohort study was performed. Patients whose specimens were p53 positive were compared to those who were p53 negative from the time of tumor resection. The principal outcome was colorectal cancer–related death. Possible counfounding variables included age at symptom onset, extent and duration of disease, primary sclerosing cholangitis, medication use and folic acid supplementation for at least 6 months before the development of cancer, family history of colorectal cancer, family history of inflam-
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Table 1. Characteristics of 75 Patients With Ulcerative Colitis– related Colorectal Cancer
Characteristic Male Sex Age at diagnosis (yr) Age at colectomy (yr) Duration of disease at colectomy (yr) Extensive colitis Primary sclerosing cholangitis Family history of inflammatory bowel disease Family history of colorectal cancer Current cigarette smoking Positive alcohol intake history
Figure 1. Positive p53 immunohistochemistry staining in an ulcerative colitis patient with colorectal cancer.
matory bowel disease, history of alcohol use, and history of cigarette smoking. Unadjusted survival curves were constructed for p53 positive and p53 negative tumors using the method of Kaplan and Meier and were compared using the log-rank test. Survival was measured from the time of diagnosis of cancer. Death from colorectal cancer was counted as an event; all other outcomes were censored. Cox proportional hazards analysis was used to assess the relationship between p53 positivity and death from colorectal cancer, adjusting for possible confounding variables (16). Proportional hazards results are summarized as the relative risk (hazard rate ratio) and corresponding 95% CI. This study was approved by Institutional Review Board of the Cleveland Clinic.
RESULTS Between 1978 and 1997, a total of 75 patients were identified to have ulcerative colitis–related colorectal cancer and had paraffin-embedded histology specimens available. Of
p53 Positive Tumors (N ⫽ 38)
p53 Negative Tumors (N ⫽ 37)
27 (71.0%) 30.5 ⫾ 18.9 46.2 ⫾ 14.7 15.8 ⫾ 10.9
22 (59.5%) 29.2 ⫾ 16.7 48.8 ⫾ 14.2 20.1 ⫾ 12.3
37 (97.4%) 5 (13.2%) 5 (13.2%)
36 (97.3%) 8 (21.6%) 9 (24.3%)
1 (2.6%)
1 (2.7%)
7 (18.4%) 17 (44.7%)
9 (24.3%) 14 (37.8%)
these tumors, 38 (50.7%) demonstrated p53 immunoreactivity and 37 (49.3%) did not. Chacteristics of these patients are shown in Table 1. There were no significant differences in clinical characteristics between patients with p53 positive tumors and those with p53 negative tumors. Also, there were no differences among tumors that were positive or negative for p53 with regard to location or Dukes’ stage (Table 2). Fourteen patients (36.8%) with p53 positive tumors died of metastatic colorectal cancer compared to only five (13.5%) patients with p53 negative tumors. Death from colorectal cancer was significantly worse among patients with p53 positive tumors (Fig. 2). Simultaneously adjusting for Dukes’ stage and primary sclerosing cholangitis, the relative risk of death was 3.03 (95% CI ⫽ 1.05– 8.73) for patients with p53 positive tumors compared to those with p53 negative tumors (Table 3). There were 13 patients with concomitant primary sclerosing cholangitis. Three of the five patients with tumors that were p53 positive died of metastatic disease, compared to one of eight patients with p53-negative tumors (not significant). Adjusting for p53 positivity and Dukes’ stage, the relative risk of death from metastatic disease was 2.64 (95% CI ⫽ 0.82– 8.56) for PSC patients (Table 3). Table 2. Location and Dukes’ Stage of 75 Patients With Ulcerative Colitis–related Colorectal Cancer
Location and stage
p53 Positive Tumors, n (%) (N ⫽ 38)
p53 Negative Tumors, n (%) (N ⫽ 37)
Rectum Sigmoid colon Descending colon Transverse colon Ascending colon Cecum Dukes’ A Dukes’ B Dukes’ C Dukes’ D
13 (34.2) 7 (18.4) 8 (21.1) 6 (15.8) 6 (15.8) 12 (31.6) 13 (34.2) 9 (23.7) 8 (21.1) 8 (21.1)
11 (29.8) 8 (21.6) 3 (8.1) 8 (21.6) 10 (27.0) 10 (27.0) 16 (43.2) 7 (18.9) 7 (18.9) 7 (18.9)
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Figure 2. Colorectal cancer survival among 75 ulcerative colitis patients: survival curves for p53 positive and p53 negative patients.
Folic acid supplementation, which was believed to prevent patients from developing p53 mutations, was documented in seven patients (18.4%) with p53 positive tumors and five patients (13.5%) with p53 negative tumors (not significant).
DISCUSSION Despite major advances in diagnosis, treatment, and cancer surveillance techniques, colorectal cancer still is the principal cause of death in ulcerative colitis patients (17, 18). Furthermore, the young age of cancer development (often in patients’ 30s and 40s) makes colorectal cancer a particularly important concern (19). Colorectal cancer risk is further increased 3-fold in ulcerative colitis patients with primary sclerosing cholangitis (20, 21). Currently, ulcerative colitis patients with high cancer risk (i.e., patients with primary sclerosing cholangitis or with extensive colitis of at least 7 yr) are offered cancer surveillance colonoscopy with extensive biopsy every 1–3 yr (22). Before 1993 or so, patients with high grade dysplasia or asymptomatic cancer were advised to have total colectomy (23). Surveillance programs with these parameters generally have failed to decrease cancer-related mortality (24, 25). One factor cited as being Table 3. Adjusted Relative Risk of Cancer-Related Death Characteristic Abnormal p53 immunohistochemistry Dukes’ A Dukes’ B Dukes’ C Dukes’ D Primary sclerosing cholangitis
Relative-Risk (95% CI)
p Value
3.03 (1.05–8.73)
0.04
1.0 2.59 (0.41–16.34) 5.19 (0.92–29.38) 35.87 (6.86–187) 2.64 (0.82–8.56)
Reference 0.31 0.06 ⬍0.001 0.11
responsible is poor performance of high grade dysplasia as the criterion for a positive test (26 –29). Although dysplasia is known to be a premalignant lesion, it is an imperfect criterion for a positive test because its localization is not necessarily widespread throughout the colon, leading to sampling errors. Also, when dysplasia is present in areas of chronic inflammation, histology often is interpreted as being indefinite for dysplasia (30). More recently, total colectomy has been recommended to patients with any grade of dysplasia, either low grade or high grade, that is found on any biopsy specimen. Although this study was designed to determine prognosis from p53 in colorectal tumors, the results have direct implications for cancer surveillance if p53 can be used as a marker complementary to dysplasia. An ideal complementary marker of malignancy that may be useful in cancer surveillance in ulcerative colitis would be objective, have a high sensitivity and specificity, and be inexpensive. Of the available biomarkers of colonic neoplasia (i.e., DNA aneuploidy, DNA hypomethylation, oncogene mutations, abnormal mucin expression, microsatellite instability, and suppressor gene mutations and loss of heterozygosity), p53 gene mutations with detection of the mutated protein products using immunohistochemistry offer promising possibilities as a test complementary to dysplasia for use in cancer surveillance programs (31). Testing for abnormal p53 staining in ulcerative colitis patients could improve sensitivity of surveillance if: 1) it occurs earlier in the sequence of genetic and histological events leading to cancer than does dysplasia, and 2) p53 is a marker of a poor cancer-related prognosis. Studies that have mapped areas of p53 mutations and dysplasia from resected colonic specimens have found that p53 mutations are distributed across a larger area than is dysplasia, suggesting that p53 mutations are earlier events
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than is dysplasia (6). If a tumor marker is not associated with a poor prognosis, it is not likely to identify patients with a high mortality risk who would benefit from colectomy. The current study documents the poor cancer related prognosis of p53 positive tumors. Previously, we performed a historical cohort study on 95 patients with long-standing ulcerative colitis, examining biopsy specimens for abnormal p53 immunohistochemistry during the course of cancer surveillance (32). In all, 39% of patients were p53 positive, 73% of whom had cancer or dysplasia, compared to only 14% with cancer or dysplasia in patients who were p53-negative. Abnormal p53 staining was significantly associated with cancer or dysplasia (adjusted relative risk ⫽ 4.53, 95% CI ⫽ 2.16 –9.48). From survival curve analysis, abnormal p53 staining preceded dysplasia in the sequence of events leading to cancer. When surveillance biopsy specimens with no dysplasia preceding the cancer were available, two of three p53 positive patients were p53 positive when no dysplasia was seen, compared to none of two patients with p53 negative cancers. Thus, abnormal p53 immunohistochemistry from archival biopsy specimens was significantly associated with dysplasia and cancer, and seemed to be an early marker of malignancy. Of note, all three patients who died of colorectal cancer, including one patient with a Dukes’ A tumor, were p53 positive. The current study of 75 patients with colorectal cancer and ulcerative colitis, 12 of whom were included in the previous study of cancer surveillance (and no additional patient was followed in our cancer surveillance program), demonstrated that abnormal p53 staining of tumors is associated with a significantly increased cancer-related mortality. Taken together, these two studies provide encouraging epidemiological evidence that p53 staining, when used as a test complementary to histological evaluation of dysplasia, could improve outcomes of cancer surveillance programs in ulcerative colitis. Possibly, patients who are found to be p53 positive would be placed in a “high mortality risk” group, regardless of the degree of dysplasia, and treated differently from those who are p53 negative by having frequent colonoscopy or, perhaps, prophylactic colectomy.
ACKNOWLEDGMENTS This study was supported by a Clinical Research Award from the American College of Gastroenterology and the Ann and E. Bradley Jones Research Fund for the Study of Inflammatory Bowel Disease. Reprint requests and correspondence: Bret A. Lashner, M.D., Cleveland Clinic Foundation/A30, 9500 Euclid Avenue, Cleveland, OH 44195. Received Apr. 29, 2002; accepted Nov. 20, 2002.
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REFERENCES 1. Carson DA, Lois A. Cancer progression and p53. Lancet 1995;346:1009 –11. 2. Auvinen A, Isola J, Visakorpi T, et al. Over-expression of p53 and long-term survival in colon carcinoma. Br J Cancer 1994; 70:293–6. 3. Hamelin R, Laurent-Puig P, Olschwang S, et al. Association of p53 mutations with short survival in colorectal cancer. Gastroenterology 1994;106:42–8. 4. Pricolo VE, Finkelstein SD, Wu T-T, et al. Prognostic value of p53 andK-ras-2 mutational analysis in stage III carcinoma of the colon. Am J Surg 1996;171:41–6. 5. Remvikos Y, Tominaga O, Hammel P, et al. Increased p53 protein content of colorectal tumors correlates with poor survival. Br J Cancer 1992;66:758 –64. 6. Brentnall TA, Crispin DA, Rabinovitch PS, et al. Mutations in the p53 gene: an early marker of neoplastic progression in ulcerative colitis. Gastroenterology 1994;107:369 –78. 7. Ajioka Y, Watanabe H, Matsuda K. Over-expression of p53 protein in neoplastic changes in ulcerative colitis: Immunohistochemical study. J Gastroenterol 1995;30(suppl VIII): 33–5. 8. Burmer GC, Crispin DA, Kolli VR, et al. Frequent loss of a p53 allele in carcinomas and their precursors in ulcerative colitis. Cancer Commun 1991;3:167–72. 9. Burmer GC, Rabinovitch PS, Haggitt RC, et al. Neoplastic progression in ulcerative colitis: histology, DNA content, and loss of a p53 allele. Gastroenterology 1992;103:1602–10. 10. Greenwald BD, Harpaz N, Yin J, et al. Loss of heterozygosity affecting the p53, Rb, and mcc/apc tumor suppressor gene loci in dysplastic and cancerous ulcerative colitis. Cancer Res 1992;52:741–5. 11. Harpaz N, Peck AL, Yin J, et al. p53 Protein expression in ulcerative colitis-associated colorectal dysplasia and carcinoma. Hum Pathol 1994;25:1069 –74. 12. Taylor HW, Boyle M, Smith SC, et al. Expression of p53 in colorectal cancer and dysplasia complicating ulcerative colitis. Br J Surg 1993;80:442–4. 13. Yin J, Harpaz N, Tong Y, et al. p53 Point mutations in dysplastic and cancerous ulcerative colitis lesions. Gastroenterology 1993;104:1633–9. 14. Ilyas M, Talbot IC. p53 Expression in ulcerative colitis: A longitudinal study. Gut 1995;37:802–4. 15. Baas IO, Mulder JR, Offerhaus GJ, et al. An evaluation of 6 antibodies for immunohistochemistry of mutant p53 gene product in archival colorectal neoplasms. J Pathol 1994;172: 5–12. 16. Breslow NE, Day NE. Statistical methods in cancer research, vol 2: The design and analysis of cohort studies. Lyon: International Agency for Research in Cancer, 1987. 17. Ekbom A, Helmick C, Zack M, Adami H-O. Ulcerative colitis and colorectal cancer: A population-based study. N Engl J Med 1990;323:1228 –33. 18. Karlen P, Lofberg R, Brostron O, et al. Increased risk of cancer in ulcerative colitis: A population-based cohort study. Am J Gastroenterol 1999;94:1047–52. 19. Eaden JA, Abrams KR, Mayberry JF. The risk of colorectal cancer in ulcerative colitis: A meta-analysis. Gut 2001;48: 526 –35. 20. Shetty K, Rybicki L, Brzezinski A, et al. The risk for cancer or dysplasia in ulcerative colitis patients with primary sclerosing cholangitis. Am J Gastroenterol 1999;94:1643–9. 21. Loftus EV, Jr, Aguilar HI, Sandborn WJ, et al. Risk of colorectal neoplasia in patients with primary sclerosing cholangitis following orthotopic liver transplantation. Hepatology 1998; 27:685–90.
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22. Lashner BA, Hanauer SB, Silverstein MD. Optimal timing of colonoscopy to screen for cancer in ulcerative colitis. Ann Intern Med 1988;108:274 –8. 23. Riddell RH, Goldman H, Ransohoff DF, et al. Dysplasia in inflammatory bowel disease: Standardized classification with provisional clinical applications. Hum Pathol 1983;14:931– 68. 24. Lashner BA. Cancer surveillance in ulcerative colitis: Much ado about little?. In: Barkin JS, Rogers AI, eds. Difficult decisions in digestive diseases. St. Louis: Mosby, 1994:357– 61. 25. Provenzale D, Onken J. Surveillance issues in inflammatory bowel disease. J Clin Gastroenterol 2001;32:99 –105. 26. Lashner BA. Recommendations for colorectal cancer screening in ulcerative colitis: A review of research from a single university-based surveillance program. Am J Gastroenterol 1992;87:168 –75. 27. Collins RH, Feldman M, Fordtran JS. Colon cancer, dysplasia,
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and surveillance in patients with ulcerative colitis: A critical review. N Engl J Med 1987;316:1654 –8. Connell WR, Lennard-Jones JE, Williams CB, et al. Factors affecting the outcome of endoscopic surveillance for cancer in ulcerative colitis. Gastroenterology 1994;107:934 –44. Mayer R, Wong WD, Rothenberger DA, et al. Colorectal cancer in inflammatory bowel disease: A continuing problem. Dis Colon Rectum 1999;42:343–7. Bernstein CN, Shanahan F, Weinstein WM. Are we telling patients the truth about surveillance colonoscopy in ulcerative colitis? Lancet 1994;343:71–4. Shapiro BD, Lashner BA. Cancer biology in ulcerative colitis and potential use in endoscopic surveillance. Gastrointest Endosc Clin North Am 1997;7:453–68. Lashner BA, Shapiro BD, Husain A, Goldblum JR. Evaluation of the usefulness of testing for p53 mutations in colorectal cancer surveillance for ulcerative colitis. Am J Gastroenterol 1999;94:456 –62.
Vol. 98, No. 6, 2003 ISSN 0002-9270/03/$30.00 doi:10.1016/S0002-9270(03)00422-2
Abnormal p53 Immunohistochemistry Is Associated With an Increased Colorectal Cancer–Related Mortality in Patients With Ulcerative Colitis Bret A. Lashner, M.D., William M. Bauer, M.D., Lisa A. Rybicki, M.S., and John R. Goldblum, M.D. Center for Inflammatory Bowel Disease, Departments of Gastroenterology, Biostatistics, and Anatomic Pathology, Cleveland Clinic Foundation, Cleveland, Ohio
OBJECTIVE: Immunohistochemical evaluation of p53 staining patterns has been considered as a complementary test for dysplasia in ulcerative colitis-related colorectal cancer surveillance, but usefulness would be particularly important if it were a marker associated with a poor prognosis. The aim of this study was to determine whether abnormal p53 staining of the tumor correlated with cancer-related mortality in ulcerative colitis patients. METHODS: An historical cohort study was designed to examine all ulcerative colitis patients who developed colorectal cancer between 1978 and 1997 and who had tumor tissue blocks available for staining. Tissue was recut and stained for abnormal p53 immunohistochemistry using the DO-7 antibody. Tumors were considered to be p53 positive if at least 5% of nuclei in a high power field were positive for staining. RESULTS: Among 75 patients entered in the study, 38 (50.7%) had p53 positive tumors. Fourteen patients (36.8%) with p53 positive tumors died from colorectal cancer, compared to five (13.5%) with p53 negative tumors (p ⬍ 0.04, log-rank test). The adjusted relative risk of cancer-related death among patients with p53 positive tumors was 3.03 (95% CI ⫽ 1.05– 8.73). CONCLUSIONS: Abnormal p53 immunohistochemistry of tumors is associated with a poor prognosis among ulcerative colitis patients who develop colorectal cancer. As such, p53 immunohistochemical staining could be a useful histological marker to complement routine histology in cancer surveillance programs in ulcerative colitis patients. (Am J Gastroenterol 2003;98:1423–1427. © 2003 by Am. Coll. of Gastroenterology)
INTRODUCTION Tumor suppressor genes, including p53, act as a G1 checkpoint in the cell replication cycle for cells with abnormal DNA until DNA repair can occur, or until apoptosis ensues in the event of excessive DNA damage. Inactivation of suppressor gene function through mutations perpetuates abnormal DNA through loss of repair and/or apoptotic mech-
anisms, thus increasing the risk of malignant transformation (1). In many cases, a mutated, inactive p53 protein has a much longer half-life than the active wild-type protein, and immunohistochemical staining of p53 often will stain nuclei strongly with accumulated mutated proteins. Therefore, nuclear accumulation of the p53 protein product is a surrogate marker for p53 mutations. Sporadic colorectal cancers with p53 mutations are associated with a poor prognosis (2–5). In ulcerative colitis patients, abnormal p53 staining and loss of heterozygosity have been shown to be present in approximately one half of patients with cancer and an early marker of neoplastic progression, but its relationship to prognosis has not been determined (6 –14). The aim of this study was to determine whether positive p53 immunohistochemical staining of ulcerative colitis–related cancer is associated with a poor prognosis as defined by cancer-related mortality.
MATERIALS AND METHODS Between 1978 and 1997, all patients with ulcerative colitis– related colorectal cancer who had surgery at the Cleveland Clinic and who had specimens available in paraffin blocks were included. Patients with Crohn’s colitis or indeterminate colitis were excluded. Sections were cut and stained with a 1:200 dilution of the D0 –7 antibody for p53 (Dako, Carpinteria, CA) (15). Specimens were considered as positive for abnormal p53 if 5% of nuclei in any high power field stained. An example of a p53 positive staining in a tumor is shown in Figure 1. All specimens were read by an experienced gastrointestinal pathologist (J.R.G.) who was blinded regarding the outcome of each patient. Clinical information was gathered from medical records. A historical cohort study was performed. Patients whose specimens were p53 positive were compared to those who were p53 negative from the time of tumor resection. The principal outcome was colorectal cancer–related death. Possible counfounding variables included age at symptom onset, extent and duration of disease, primary sclerosing cholangitis, medication use and folic acid supplementation for at least 6 months before the development of cancer, family history of colorectal cancer, family history of inflam-
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Table 1. Characteristics of 75 Patients With Ulcerative Colitis– related Colorectal Cancer
Characteristic Male Sex Age at diagnosis (yr) Age at colectomy (yr) Duration of disease at colectomy (yr) Extensive colitis Primary sclerosing cholangitis Family history of inflammatory bowel disease Family history of colorectal cancer Current cigarette smoking Positive alcohol intake history
Figure 1. Positive p53 immunohistochemistry staining in an ulcerative colitis patient with colorectal cancer.
matory bowel disease, history of alcohol use, and history of cigarette smoking. Unadjusted survival curves were constructed for p53 positive and p53 negative tumors using the method of Kaplan and Meier and were compared using the log-rank test. Survival was measured from the time of diagnosis of cancer. Death from colorectal cancer was counted as an event; all other outcomes were censored. Cox proportional hazards analysis was used to assess the relationship between p53 positivity and death from colorectal cancer, adjusting for possible confounding variables (16). Proportional hazards results are summarized as the relative risk (hazard rate ratio) and corresponding 95% CI. This study was approved by Institutional Review Board of the Cleveland Clinic.
RESULTS Between 1978 and 1997, a total of 75 patients were identified to have ulcerative colitis–related colorectal cancer and had paraffin-embedded histology specimens available. Of
p53 Positive Tumors (N ⫽ 38)
p53 Negative Tumors (N ⫽ 37)
27 (71.0%) 30.5 ⫾ 18.9 46.2 ⫾ 14.7 15.8 ⫾ 10.9
22 (59.5%) 29.2 ⫾ 16.7 48.8 ⫾ 14.2 20.1 ⫾ 12.3
37 (97.4%) 5 (13.2%) 5 (13.2%)
36 (97.3%) 8 (21.6%) 9 (24.3%)
1 (2.6%)
1 (2.7%)
7 (18.4%) 17 (44.7%)
9 (24.3%) 14 (37.8%)
these tumors, 38 (50.7%) demonstrated p53 immunoreactivity and 37 (49.3%) did not. Chacteristics of these patients are shown in Table 1. There were no significant differences in clinical characteristics between patients with p53 positive tumors and those with p53 negative tumors. Also, there were no differences among tumors that were positive or negative for p53 with regard to location or Dukes’ stage (Table 2). Fourteen patients (36.8%) with p53 positive tumors died of metastatic colorectal cancer compared to only five (13.5%) patients with p53 negative tumors. Death from colorectal cancer was significantly worse among patients with p53 positive tumors (Fig. 2). Simultaneously adjusting for Dukes’ stage and primary sclerosing cholangitis, the relative risk of death was 3.03 (95% CI ⫽ 1.05– 8.73) for patients with p53 positive tumors compared to those with p53 negative tumors (Table 3). There were 13 patients with concomitant primary sclerosing cholangitis. Three of the five patients with tumors that were p53 positive died of metastatic disease, compared to one of eight patients with p53-negative tumors (not significant). Adjusting for p53 positivity and Dukes’ stage, the relative risk of death from metastatic disease was 2.64 (95% CI ⫽ 0.82– 8.56) for PSC patients (Table 3). Table 2. Location and Dukes’ Stage of 75 Patients With Ulcerative Colitis–related Colorectal Cancer
Location and stage
p53 Positive Tumors, n (%) (N ⫽ 38)
p53 Negative Tumors, n (%) (N ⫽ 37)
Rectum Sigmoid colon Descending colon Transverse colon Ascending colon Cecum Dukes’ A Dukes’ B Dukes’ C Dukes’ D
13 (34.2) 7 (18.4) 8 (21.1) 6 (15.8) 6 (15.8) 12 (31.6) 13 (34.2) 9 (23.7) 8 (21.1) 8 (21.1)
11 (29.8) 8 (21.6) 3 (8.1) 8 (21.6) 10 (27.0) 10 (27.0) 16 (43.2) 7 (18.9) 7 (18.9) 7 (18.9)
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Figure 2. Colorectal cancer survival among 75 ulcerative colitis patients: survival curves for p53 positive and p53 negative patients.
Folic acid supplementation, which was believed to prevent patients from developing p53 mutations, was documented in seven patients (18.4%) with p53 positive tumors and five patients (13.5%) with p53 negative tumors (not significant).
DISCUSSION Despite major advances in diagnosis, treatment, and cancer surveillance techniques, colorectal cancer still is the principal cause of death in ulcerative colitis patients (17, 18). Furthermore, the young age of cancer development (often in patients’ 30s and 40s) makes colorectal cancer a particularly important concern (19). Colorectal cancer risk is further increased 3-fold in ulcerative colitis patients with primary sclerosing cholangitis (20, 21). Currently, ulcerative colitis patients with high cancer risk (i.e., patients with primary sclerosing cholangitis or with extensive colitis of at least 7 yr) are offered cancer surveillance colonoscopy with extensive biopsy every 1–3 yr (22). Before 1993 or so, patients with high grade dysplasia or asymptomatic cancer were advised to have total colectomy (23). Surveillance programs with these parameters generally have failed to decrease cancer-related mortality (24, 25). One factor cited as being Table 3. Adjusted Relative Risk of Cancer-Related Death Characteristic Abnormal p53 immunohistochemistry Dukes’ A Dukes’ B Dukes’ C Dukes’ D Primary sclerosing cholangitis
Relative-Risk (95% CI)
p Value
3.03 (1.05–8.73)
0.04
1.0 2.59 (0.41–16.34) 5.19 (0.92–29.38) 35.87 (6.86–187) 2.64 (0.82–8.56)
Reference 0.31 0.06 ⬍0.001 0.11
responsible is poor performance of high grade dysplasia as the criterion for a positive test (26 –29). Although dysplasia is known to be a premalignant lesion, it is an imperfect criterion for a positive test because its localization is not necessarily widespread throughout the colon, leading to sampling errors. Also, when dysplasia is present in areas of chronic inflammation, histology often is interpreted as being indefinite for dysplasia (30). More recently, total colectomy has been recommended to patients with any grade of dysplasia, either low grade or high grade, that is found on any biopsy specimen. Although this study was designed to determine prognosis from p53 in colorectal tumors, the results have direct implications for cancer surveillance if p53 can be used as a marker complementary to dysplasia. An ideal complementary marker of malignancy that may be useful in cancer surveillance in ulcerative colitis would be objective, have a high sensitivity and specificity, and be inexpensive. Of the available biomarkers of colonic neoplasia (i.e., DNA aneuploidy, DNA hypomethylation, oncogene mutations, abnormal mucin expression, microsatellite instability, and suppressor gene mutations and loss of heterozygosity), p53 gene mutations with detection of the mutated protein products using immunohistochemistry offer promising possibilities as a test complementary to dysplasia for use in cancer surveillance programs (31). Testing for abnormal p53 staining in ulcerative colitis patients could improve sensitivity of surveillance if: 1) it occurs earlier in the sequence of genetic and histological events leading to cancer than does dysplasia, and 2) p53 is a marker of a poor cancer-related prognosis. Studies that have mapped areas of p53 mutations and dysplasia from resected colonic specimens have found that p53 mutations are distributed across a larger area than is dysplasia, suggesting that p53 mutations are earlier events
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p53 in Cancer in Ulcerative Colitis
than is dysplasia (6). If a tumor marker is not associated with a poor prognosis, it is not likely to identify patients with a high mortality risk who would benefit from colectomy. The current study documents the poor cancer related prognosis of p53 positive tumors. Previously, we performed a historical cohort study on 95 patients with long-standing ulcerative colitis, examining biopsy specimens for abnormal p53 immunohistochemistry during the course of cancer surveillance (32). In all, 39% of patients were p53 positive, 73% of whom had cancer or dysplasia, compared to only 14% with cancer or dysplasia in patients who were p53-negative. Abnormal p53 staining was significantly associated with cancer or dysplasia (adjusted relative risk ⫽ 4.53, 95% CI ⫽ 2.16 –9.48). From survival curve analysis, abnormal p53 staining preceded dysplasia in the sequence of events leading to cancer. When surveillance biopsy specimens with no dysplasia preceding the cancer were available, two of three p53 positive patients were p53 positive when no dysplasia was seen, compared to none of two patients with p53 negative cancers. Thus, abnormal p53 immunohistochemistry from archival biopsy specimens was significantly associated with dysplasia and cancer, and seemed to be an early marker of malignancy. Of note, all three patients who died of colorectal cancer, including one patient with a Dukes’ A tumor, were p53 positive. The current study of 75 patients with colorectal cancer and ulcerative colitis, 12 of whom were included in the previous study of cancer surveillance (and no additional patient was followed in our cancer surveillance program), demonstrated that abnormal p53 staining of tumors is associated with a significantly increased cancer-related mortality. Taken together, these two studies provide encouraging epidemiological evidence that p53 staining, when used as a test complementary to histological evaluation of dysplasia, could improve outcomes of cancer surveillance programs in ulcerative colitis. Possibly, patients who are found to be p53 positive would be placed in a “high mortality risk” group, regardless of the degree of dysplasia, and treated differently from those who are p53 negative by having frequent colonoscopy or, perhaps, prophylactic colectomy.
ACKNOWLEDGMENTS This study was supported by a Clinical Research Award from the American College of Gastroenterology and the Ann and E. Bradley Jones Research Fund for the Study of Inflammatory Bowel Disease. Reprint requests and correspondence: Bret A. Lashner, M.D., Cleveland Clinic Foundation/A30, 9500 Euclid Avenue, Cleveland, OH 44195. Received Apr. 29, 2002; accepted Nov. 20, 2002.
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