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Prognostic Value of p53 Genetic Changes in Colorectal Cancer

From the Departments of Surgery and Oncology, University Hospital, Uppsala; Pharmacia Biotech AB, Uppsala; and Department of Surgery, Umeå University Hospital, Umeå, Sweden.

Address reprint requests to Ulf Kressner, MD, PhD, Department of Surgery, Uddevalla sjukhus, 451 80 Uddevalla, Sweden; email ulf.kressner{at}nu.alvsborg.se

	ABSTRACT

TOP ABSTRACT INTRODUCTION MATERIALS AND METHODS RESULTS DISCUSSION REFERENCES

 
PURPOSE: To explore whether there is a linkage between different mutations in the p53 gene in primary colorectal cancer and the risk of death from colorectal cancer in a large group of patients with long follow-up. We also compared a complementary DNA-based sequencing method and an immunohistochemical (IHC) method for detecting p53 protein overexpression in colorectal cancer.

MATERIALS AND METHODS: The entire coding region of the p53 gene was sequenced in 191 frozen tumor samples collected from January 1988 to November 1992. RNA was extracted and synthesized to cDNA. p53 was amplified by the polymerase chain reaction, and the DO-7 monoclonal antibody was used in the IHC assessments.

RESULTS: Mutations were detected in 99 samples (52%) from 189 patients. There was a significant relationship between the p53 mutational status and the cancer-specific survival time, with shorter survival time for patients who had p53 mutations than for those who did not (P = .01, log-rank test). Mutations outside the evolutionarily conserved regions were associated with the worst prognosis. Multivariate analysis showed that the presence of p53 mutations was an independent prognostic factor (relative hazard, 1.7, P = .03). There was no significant relationship between overexpression of p53 protein, as determined by IHC analysis, and cancer-specific survival.

CONCLUSION: Mutational analyses of the p53 gene, using cDNA sequencing in colorectal cancer, provide useful prognostic information. In addition, cDNA sequencing gives better prognostic information than IHC assessment of p53 protein overexpression.

	INTRODUCTION

TOP ABSTRACT INTRODUCTION MATERIALS AND METHODS RESULTS DISCUSSION REFERENCES

 
THE DUKES CLASSIFICATION SYSTEM and similar systems based on the extent of tumor spread (eg, Astler-Coller and the tumor-node-metastasis classification) represent the most commonly used staging procedures in colorectal cancer.^1,2 The tumor stage provides prognostic information but not to the extent that it can be used to meet current requirements for differentiated therapy. The toxicity and expense of additional therapy can only be justified in the subset of patients at high risk for relapse. A large number of potentially clinically useful prognostic factors have been identified,³ among them, mutational status of the p53 gene.

The p53 suppressor gene, located on the short arm of chromosome 17,⁴ encodes a 53-kd nuclear phosphoprotein that regulates the cell cycle.^5,6 Mutations in this gene constitute some of the most frequently occuring genetic changes found in human malignancies.^4,5 They are thought to be a late development in the adenoma-carcinoma sequence in colorectal cancer.⁷ It has been reported that p53 mutations seem to be associated with poor prognosis in colorectal cancer.^8-11 However, conflicting results have also been claimed.¹² Overexpression of the p53 protein is detectable in 30% to 70% of the tumors, using immunohistochemical (IHC) methods. In a great majority of studies, p53 protein overexpression has been used as a surrogate marker for p53 mutations, an assumption that is not entirely correct,¹³ although it may sometimes be justified for practical and economic reasons. In some studies,^14-17 p53 protein overexpression has been shown to correlate with patient survival, a finding that has not been observed in other studies.^10,18-22

The purpose of this study was to explore and to determine whether there is a valid association between the mutations in the p53 gene, as determined by cDNA sequencing, and the risk of death from colorectal cancer, as studied in a large group of patients with long follow-up. In addition, we investigated the relationship between observed mutations in the p53 gene and the overexpression of the p53 protein, as detected by IHC analysis. We also evaluated whether there is any association between cDNA sequencing and serum p53 antibody levels in preoperative serum samples.

	MATERIALS AND METHODS

TOP ABSTRACT INTRODUCTION MATERIALS AND METHODS RESULTS DISCUSSION REFERENCES

 
Patients
Tumor samples were collected from 194 nonselected patients resected for colorectal cancer in the Uppsala and Falun counties in Sweden between January 1988 and November 1992. One hundred ninety-one meticulously dissected, fresh-frozen samples were available for analyses. Two patients contributed samples from two different tumors. Subsequently, one tumor sample from each patient was included in the survival analyses. Serum samples were collected before surgery from 90 patients (47%). The age and sex distribution, Dukes stage, and tumor differentiation are listed in Table 1. One hundred sixty-seven patients (87%) were resected for cure. In the remaining 24 patients, distant metastases were detected during surgery; consequently, these patients underwent a palliative resection. This group was classified as Dukes stage D. Twenty-eight of the patients (43%) with rectal cancer received preoperative radiotherapy. No patient received any postoperative adjuvant radiotherapy or chemotherapy. At follow-up in January 1997, 74 patients (39%) had died from cancer or from other causes with known tumor burden. Twenty-eight patients (15%) died from other causes without any indication of tumor relapse. No patient was lost to follow-up. The median survival time of the 89 patients alive at follow-up was 87 months (range, 51 to 106 months). Routine biopsy samples were taken from each tumor for histopathologic classification. The tumors were graded according to the World Health Organization classification²³ and staged according to the Dukes classification system.¹

RNA was prepared from the frozen tumor samples, under stringent conditions, to avoid degradation and contamination. This procedure was followed by an enzymatic synthesis of cDNA, using RNA as template. p53 was amplified from the tumor cDNA by PCR, using four overlapping primer pairs covering the entire coding region of the p53 gene. With one of the primers (in each primer pair) modified with a biotin molecule, biotin-labeled PCR products were generated, thus facilitating solid sequencing.²⁴ Manifold (solid-phase) sequencing was performed essentially as described by Lagerkvist et al.²⁵ The sequencing products generated were analyzed using an automated laser fluorescence sequencer (ALFexpress; Pharmacia Biotech, Uppsala, Sweden). The sequence was finally compared with the wild-type p53 sequence, using prototype software program p53 SB Decipher, version 1.00 (Pharmacia Biotech). Nucleotide changes that had an impact on the protein were considered to be mutations. Each identified mutation was verified by sequencing an entirely new PCR product from the corresponding cDNA. Further confirmation was obtained by analysis of the neighboring PCR fragment when the mutation was located in an overlapping segment.

IHC Analysis of Overexpression of p53 Protein
p53 protein overexpression (mutated and wild-type) was evaluated by use of an IHC method in 190 tumor samples.²³ In a previous study, we investigated four different antibodies for overexpression of the p53 protein and compared the homogeneity in multiple biopsy samples collected meticulously and randomly from each tumor.²⁶ A tumor section with positive staining by the the anti-p53 antibody was clearly homogenously stained, ie, virtually all tumor nuclei were positively stained. We concluded that DO-7 did not show intratumor heterogeneity; therefore, we selected DO-7 for further analyses. We lack information about p53 IHC analysis from one of the tumors. Briefly, DO-7 monoclonal antibody was used, in combination with biotinylated horse–anti-mouse IgG antibody as secondary antibody, followed by incubation with Vectastain ABC ELITE reagent (Vector Laboratories, Burlingame, CA) and development with 3-amino-9-ethylcarbazole.

Analysis of Serum p53 Antibody Levels
Serum samples were collected before surgery from 90 patients. The serum samples were stored at -70°C until analyzed. Anti-p53 was measured using an enzyme-linked immunoadsorbent assay (Dianova, Hamburg, Germany), described in detail elsewhere.²⁷

Statistical Analyses
The Cox proportional hazards model was used in both the univariate and the multivariate survival analyses.^28,29 Cancer-specific survival curves were constructed using the Kaplan-Meier method, and differences between curves were tested using the log-rank test. Censored cases in the analyses were patients who died from other causes with no known residual cancer disease. The ² test was used to test for differences in distribution between groups. The statistical software Statistica, version 5.0 (StatSoft Inc, Tulsa, OK), was used for the analyses. P values of less than .05 were considered statistically significant.

	RESULTS

TOP ABSTRACT INTRODUCTION MATERIALS AND METHODS RESULTS DISCUSSION REFERENCES

 
Changes in p53
The p53 mutational status for 189 colorectal cancers out of 191 was determined by cDNA sequencing. From the remaining two samples, no PCR products were generated. Mutations were detected in 99 samples (52%) from 189 patients. One hundred seven genetic changes were found (84 missense mutations, 16 deletions, three insertions, and four stops). Eight samples displayed two different mutations (five tumors had two missense mutations, one tumor had one deletion and one insertion, one tumor had two deletions, and one tumor had an insertion and one missense mutation). p53 mutations were found throughout the entire protein coding region of the gene. Seventy-nine (74%) of the 107 mutations were located in the conserved regions of p53 (Fig 1). Most of the missense mutations (83%) occurred inside conserved regions, whereas other genetic changes more frequently occurred outside. There was no difference in the frequency of mutations within the Dukes stages (Table 1). Tumors in the distal colon and rectum contained more frequent mutations than tumors in the proximal colon.

View larger version (71K): [in this window] [in a new window]   Fig 1. Distribution of missense mutations and other genetic changes along the p53 gene.

Comparison of p53 cDNA Sequencing and p53 IHC Analysis
Overexpression of p53 protein was demonstrated in tumors from 92 (48%) of the 190 patients.²³ In the 188 cases in which a comparison was possible, there was a concordance in the results of the cDNA sequencing and the IHC analysis in 140 tumors (74%; Table 2).

Twenty-eight (29%) of the tumors that were positive by cDNA sequencing were negative on IHC analysis. This discrepancy was observed in three (75%) of four tumors with mutations that created premature stop codons and in nine (64%) of 14 tumors with deletions, but was much more uncommon in tumors with missense mutations (15 of 84, 18%). Twenty (22%) of the tumors that displayed the wild-type gene on cDNA sequencing were positive by IHC analysis (Table 2).

p53 Mutations and Serum p53 Antibody Levels
The association of tumors with p53 genetic changes with increased serum p53 antibody levels was significant: 23 (50%) of 46 patients with tumors showing genetic changes had high levels of serum p53 antibodies; this was the case in only two (5%) of 44 patients who did not have p53 genetic changes (P = .0002) (Table 2). The association between p53 genetic changes and serum p53 antibody levels was observed in 20 (71%) tumors with missense mutations.

p53 Mutations and Prognosis
Patients with p53 mutations, as determined by cDNA sequencing, had a significantly shorter cancer-specific survival time when compared with patients with the wild-type p53 gene. This finding was observed when the total patient material was analyzed (P = .01, log-rank test; Fig. 2A) but not when the analysis was restricted to the potentially cured patients in Dukes stages A to C (P = .12, log-rank test; data not shown).

View larger version (27K): [in this window] [in a new window]   Fig 2. (Left) Life table plots for Dukes stages A to D. The solid line indicates the p53 mutation (n = 97), and the dotted line indicates wild-type p53 (n = 90). (Right) Life table plots for Dukes stages A to D. The solid indicates p53 IHC positivity (n = 90), and the dotted line represents p53 IHC negativity (n = 97). Cancer-specific curves were generated according to the Kaplan-Meier method, and statistical comparisons were made using the log-rank test.

Overexpression of p53 Protein and Prognosis
There was no significant relationship between the overexpression of p53 protein and cancer-specific survival, neither when the total patient material was analyzed (P = .40, log-rank test; Fig. 2B) nor when the analysis was restricted to the potentially cured patients (P = .64, log-rank test; data not shown).

Uni- and Multivariate Analyses
The results of the univariate analyses for p53 mutations overall and for selected subgroups are shown in Table 3. p53 mutations overall, p53 mutations outside conserved regions, and other types of mutations versus wild-type p53 showed significant prognostic information, whereas p53 mutations within conserved regions and missense mutations did not.

Age, sex, p53 mutation, Dukes stage, and tumor differentiation were independent prognostic factors in a multivariate analysis using the Cox proportional hazards model, whereas overexpression of p53 protein and tumor localization were not (Table 4). Furthermore, no prognostic information was obtained from overexpression of p53 protein when tested in a multivariate model (data not shown).

	DISCUSSION

TOP ABSTRACT INTRODUCTION MATERIALS AND METHODS RESULTS DISCUSSION REFERENCES

 
Our results show that determination of p53 mutations using cDNA sequencing in colorectal cancer provides independent prognostic information about colorectal cancer–related deaths. This finding is in accordance with the findings of Hamelin et al,⁸ who also evaluated p53 mutational status in a multivariate model. In two other studies,^9,11 a correlation between p53 mutations and patient survival, using Kaplan-Meier life table analyses, was also observed. The majority of published studies focusing on p53 gene status and prognosis in colorectal cancer have been based solely on IHC analyses. Some studies have then shown a correlation with survival^14-17; this finding could not, however, be confirmed by us²³ or by others.^10,18-20,22 Our results suggest that the choice of methodology could have influenced the outcome of the studies and thus the overall clinical significance of p53 mutations. Direct comparisons among different IHC studies are difficult. Different mono- and polyclonal antibodies, plus both fresh tissue samples and archival material, have been used. Moreover, there is a lack of consensus as to how to interpret nuclear staining, ie, the number of cells that need to be stained for a tumor to be considered positive for p53 overexpression. In addition, our present results show that cDNA sequencing gives better prognostic information than IHC assessment of the p53 protein (relative hazard [RH], 1.63, P = 0.04; v RH, 1.02, P = .72), in agreement with results previously reported for breast cancer.¹³

We are the first to report that mutations that occur outside evolutionary conserved regions seem to be related to a significantly poorer prognosis (RH, 2.23, P = .01). This finding is in contrast to that of Goh et al,⁹ who stated that missense mutations within conserved regions were associated with the worst prognosis. In that study, however, only exons 4 to 9 in the p53 gene were studied and not the entire p53 gene, as was the case in our study. Mutations in the p53 gene occur predominantly in the segment of the genome that is responsible for DNA binding.³⁰ This genomic region, coded mostly by exons 5 to 8, is referred to as the evolutionary conserved domain,³¹ in which evolutionary conserved regions are located. We found that the frequency of missense mutations was higher inside the conserved regions (82%), which was in agreement with other reports,^9,11 whereas other types of mutations seemed to be more widely distributed.

The concordance between the cDNA sequencing and IHC analysis in our material was only 74%. Although the correlation is highly statistically significant (P < .001), it indicates that the two methods have partly different sensitivity for detecting aberrant p53. The samples used for IHC analysis and cDNA sequencing were selected from each tumor and located adjacent to each other. Normal intestine was always excluded from the samples, but microdissection was not performed. The possible impact of intratumoral connective tissue was considered negligible, as the tumor tissue always dominates intratumoral stroma demonstrated by IHC analyses. Similar results have been reported in studies comparing p53 mutations and overexpression of p53 protein with polyclonal antibody 1801^11,32 or with monoclonal antibody DO-7.³³ As most genetic changes other than missense mutations do not result in the expression of any p53 protein, it is therefore natural that these genetic changes more frequently fail detection by the IHC method than by cDNA sequencing. This failure indicates a weakness of the IHC method to find these types of mutations, which represent 20% of all mutations in colorectal cancer.⁸ In contrast, the IHC method was relatively effective in identifying missense mutations (81%), which may be due to the fact that these mutations result in excessive p53 protein expression not sufficiently counterbalanced by Mdm2.^34,35 The cDNA sequencing method used in this study has previously been used for the detection of p53 changes in breast cancer.¹³ We have had no indication that the method generated false-positive results. It could theoretically be the case as a consequence of contamination of the samples. Stringent procedures during the pre-PCR phase of the analysis, in combination with appropriate negative controls, reduced to a minimum the risk of obtaining false-positive results.

Baas et al³⁶ showed that strong rather than weak IHC staining is associated to a great extent with p53 genetic changes detected by gene analyses. Twenty tumors with no detectable p53 changes were positive according to IHC analysis, thus possibly indicating an overexpression of the p53 protein without genetic changes. In theory, this may be caused by an inability to detect certain mutations. However, cDNA sequencing is a very sensitive method, and the frequency of missed mutations is extremely low.¹³ The theory of stabilization of the p53 protein is more likely, possibly involving a nonmutational pathway responsible for the overexpression of wild-type p53 protein.³⁷

False-negative results can occur putatively, if tumor samples used for the analyses contain a low proportion of tumor cells in relation to normal cells. As a result, false-negative results dilute the signal corresponding to mutation below the threshold of detection of cDNA sequencing. The biopsy specimens selected for this study were carefully chosen to contain abundant tumor cells, which can explain the strong signal seen in virtually all biopsy specimens.

In this study, we were also able to demonstrate a significant association between p53 genetic changes in tumor sections, as determined by cDNA sequencing, and elevated levels of serum p53 antibodies (P = .0002), although increased levels were seen in only 50% of the patients with p53 mutations. Missense mutations were more commonly associated with high serum p53 antibody levels (71%). A missense mutation results in an accumulation of mutant p53 protein, and this seems to be necessary for the development of a humoral response with detectable levels of anti-p53 antibodies in the peripheral circulation.

Fluorouracil is a cell cycle–specific cytotoxic agent used in colorectal cancer that induces DNA damage, both by inhibition of thymidylate synthase and by direct fragmentation of DNA. The tumor cell may respond to DNA damage by undergoing G₁ arrest or by inducing cell death by apoptosis. This response may rely on a normal p53 gene and a functional p53 protein.³⁸ Thus, it seems that normal p53 protein is essential for optimal cytotoxic drug action. Lowe et al³⁹ have shown that the status of the p53 gene may play an important role in the responsiveness to treatment with cytotoxic drugs. This also seems to be the case for radiotherapy.⁴⁰ No patients in our series received adjuvant chemotherapy treatment, which has now become routine in clinical practice for Dukes stage C patients. Whereas normal p53 protein facilitates apoptosis induced by chemotherapy, p53 mutation instead leads to chemotherapy resistance³⁸ and thus to overtreatment in a large number of patients.

Detecting genetic changes in the p53 gene with cDNA sequencing is technically demanding. It is likely that methods as accurate as cDNA sequencing but requiring fewer technical skills will be used for routine screening of p53 gene mutations. Such a method has recently been described.⁴¹ Using this method, identification of wild-type samples would only require cDNA sequencing of aberrant samples. This kind of strategy would significantly reduce the amount of work needed to establish the relationship between genotype and phenotype in a clinical context.

In essence, p53 cDNA sequencing in colorectal cancer provides an accurate and sensitive method for localization of genetic changes. Our observations demonstrate that cDNA sequencing is superior to IHC analysis in this respect. However, if p53 cDNA sequencing or any other method is to have an impact on clinical practice in the determination of treatment, it must also be possible to analyze preoperative tumor biopsy specimens more efficiently. The question of tumor heterogeneity in relation to biopsy sampling²⁶ will need further attention before cDNA sequencing can be used routinely.

	ACKNOWLEDGMENTS

 
Supported by grants from the Swedish Cancer Society (grant no. 1921-B97-15XCC), the University Hospital Cancer Foundation, and Pharmacia Biotech, Uppsala

The skillful technical assistance of Fariba Sabounchi is gratefully acknowledged.

	REFERENCES

TOP ABSTRACT INTRODUCTION MATERIALS AND METHODS RESULTS DISCUSSION REFERENCES

 
1. Dukes CE, Bussey HJR. The spread of rectal cancer and its effect on prognosis. Br J Cancer 12:309-320, 1958

2. UICC: TNM Classification of Malignant Tumors (rev ed 4). Berlin, Germany, Springer-Verlag, 1992

3. Bosman FT: Prognostic value of pathological characteristics of colorectal cancer. Eur J Cancer 31A:1216-1221, 1995

4. Levine AJ, Momand J, Finlay CA: The p53 tumor suppressor gene. Nature 351:453-456, 1991[Medline]

5. Harris CC, Hollstein M: Clinical implications of the p53 tumor-suppressor gene. N Engl J Med 329:1318-1327, 1993[Free Full Text]

6. Chang F, Syrjanen S, Syrjanen K: Implications of the p53 tumor-suppressor gene in clinical oncology. J Clin Oncol 4:1009-1022, 1995

7. Fearon E, Vogelstein B: A genetic model for colorectal tumorigenesis. Cell 61:759-767, 1990[Medline]

8. Hamelin R, Laurent-Puig GP, Olschwang S, et al: Association of p53 mutations with short survival in colorectal cancer. Gastroenterol 106:42-48, 1994[Medline]

9. Goh HS, Yao J, Smith DR: p53 point mutation and survival in colorectal cancer patients. Cancer Res 55:5217-5221, 1995[Abstract/Free Full Text]

10. Pricolo VE, Finkelstein SD, Wu TT,et al: Prognostic value of TP53 and K-ras-2 mutational analysis in stage III carcinoma of the colon. Am J Surg 171:41-46, 1996[Medline]

11. Smith DR, Ji CY, Goh HS: Prognostic significance of p53 overexpression and mutation in colorectal adenocarcinomas. Br J Cancer 74:216-223, 1996[Medline]

12. Dix B, Robbins P, Soong R, et al: The common molecular genetic alterations in Dukes B and C colorectal carcinomas are not short-term prognostic indicators of survival. Int J Cancer 59:747-751, 1994[Medline]

13. Sjögren S, Inganäs M, Norberg T, et al: The 53 gene in breast cancer: Prognostic value of complementary DNA sequencing versus immunohistochemistry. J Natl Cancer Inst 88:173-182, 1996

14. Remvikos Y, Tominaga O, Hammel P, et al: Increased p53 protein content of colorectal tumors correlates with poor survival. Br J Cancer 66:758-764, 1992[Medline]

15. Sun XF, Carstensen JM, Zhang H, et al: Prognostic significance of cytoplasmic p53 oncoprotein in colorectal adenocarcinoma. Lancet 340:1369-1373, 1992[Medline]

16. Auvinen A, Isola J, Visakorpi T, et al: Overexpression of p53 and long-term survival in colon carcinoma. Br J Cancer 70:293-296, 1994[Medline]

17. Bosari S, Giuseppe V, Bossi P, et al: Cytoplasmic accumulation of p53 protein: An independent prognostic indicator in colorectal adenocarcinoma. J Natl Cancer Inst 86:681-687, 1994[Abstract/Free Full Text]

18. Bell SM, Scott N, Cross D, et al: Prognostic value of p53 overexpression and c-Ki-ras gene mutations in colorectal cancer. Gastroenterol 104:57-64, 1993[Medline]

19. Morrin M, Kelly M, Barret N, Delaney P: Mutations of Ki-ras and p53 genes in colorectal cancer and their prognostic significance. Gut 35:1627-1631, 1994[Abstract/Free Full Text]

20. Mulder JWR, Baas IO, Polak MM, et al: Evaluation of p53 protein expression as a marker for long term prognosis in colorectal carcinoma. Br J Cancer 71:1257-1262, 1995[Medline]

21. Kressner U, Lindmark G, Gerdin B, et al: Immunohistochemical p53 staining is of limited value in the staging and prognostic prediction of colorectal cancer. Anticancer Res 16:951-958, 1996[Medline]

22. Poller DN, Baxter KJ, Shepherd NA: p53 and Rb1 protein expression: Are they prognostically useful in colorectal cancer? Br J Cancer 75:87-93, 1997[Medline]

23. Morson B, Sobin L: Histological typing of Intestinal Tumors. International Histological Classification of Tumors, No. 15, Geneva, Switzerland, World Health Organization, 1976

24. Ståhl S, Hultman T, Olsson A, et al: Solid phase DNA sequencing using the biotin-avidin system. Nucleic Acids Res 16:3025-3038, 1988[Abstract/Free Full Text]

25. Lagerkvist A, Stewart J, Lagerström-Fermér M, et al: Manifold sequencing: Efficient processing of large sets of sequencing reactions. Proc Natl Acad Sci USA 91:2245-2249, 1994[Abstract/Free Full Text]

26. Kressner U, Lindmark G, Gerdin B, et al: Heterogeneity in proliferation markers in colorectal cancer. Anticancer Res 15:2755-2762, 1995[Medline]

27. Kressner U, Glimelius B, Bergström R, et al: Increased serum p53 antibody levels indicate poor prognosis in patients with colorectal cancer. Br J Cancer 77:1848-1851, 1998[Medline]

28. Cox DR: Regression models and life-tables (with discussion). J R Stat Soc B34:187-220, 1972

29. Peto R, Pike M, Armitage P, et al: Design and analysis of randomized clinical trials requiring prolonged observations of each patient: II. Analysis and examples. Br J Surg 35:1-39, 1977

30. Hollstein M, Sidransky D, Vogelstein B, et al: p53 mutations in human cancers. Science 253:49-53, 1992

31. Lane D: p53 and human cancers. Br Med Bull 50:582-599, 1994[Abstract/Free Full Text]

32. Costa A, Marasca R, Valentinis B, et al: p53 gene missense mutations in relation to p53 nuclear protein accumulation in colorectal cancers. J Pathol 176:45-53, 1995[Medline]

33. Dix B, Robbins P, Carello A, et al: Comparison of p53 gene mutation and protein overexpression in colorectal carcinoma. Br J Cancer 70:585-590, 1994[Medline]

34. Haupt Y, Maya R, Oren M: Mdm2 promotes the rapid degradation of p53. Nature387:296-299, 1997[Medline]

35. Kubbutat M, Jones S, Vousden K: Regulation of p53 stability by Mdm2. Nature 387:299-303, 1997[Medline]

36. Baas IO, Mulder JWR, Offerhaus GJ, et al: An evaluation of six antibodies for immunohistochemistry of mutant p53 gene product in archival colorectal neoplasms. J Pathol 72:5-12, 1994

37. Cripps K, Purdie CA, Carder PJ, et al: A study of p53 protein versus point mutation in colorectal cancer. Oncogene 9:2739-2743, 1994[Medline]

38. Brett MC, Pickard M, Green B, et al: p53 protein overexpression and response to biomodulated 5-fluorouracil chemotherapy in patients with advanced colorectal cancer. Eur J Surg Oncol 22:182-185, 1996[Medline]

39. Lowe S, Bodis S, McClatchey A, et al: p53 status and the efficacy of cancer therapy in vivo. Science 266:807-810, 1994[Abstract/Free Full Text]

40. McIlwrath AJ, Vasey PA, Ross GM, et al: Cell cycle arrest and radiosensitivity of human tumor cell lines: Dependence on wild-type p53 for radiosensitivity. Cancer Res 54:3718-3722, 1994[Abstract/Free Full Text]

41. Mashal RD, Koontz J, Sklar J: Detection of mutations by cleavage of DNA heteroduplexes with bacteriophage resolvases. Nat Gen 9:177-183, 1995[Medline]

Submitted May 18, 1998; accepted October 8, 1998.

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				A. Forslund, C. Lonnroth, M. Andersson, H. Brevinge, and K. Lundholm Mutations and Allelic Loss of p53 in Primary Tumor DNA From Potentially Cured Patients With Colorectal Carcinoma J. Clin. Oncol., June 1, 2001; 19(11): 2829 - 2836. [Abstract] [Full Text] [PDF]

				H. Elsaleh, B. Powell, K. McCaul, F. Grieu, R. Grant, D. Joseph, and B. Iacopetta P53 Alteration and Microsatellite Instability Have Predictive Value for Survival Benefit from Chemotherapy in Stage III Colorectal Carcinoma Clin. Cancer Res., May 1, 2001; 7(5): 1343 - 1349. [Abstract] [Full Text]

				M. Inganas, S. Byding, A. Eckersten, S. Eriksson, T. Hultman, A. Jorsback, E. Lofman, F. Sabounchi, U. Kressner, G. Lindmark, et al. Enzymatic Mutation Detection in the P53 Gene Clin. Chem., October 1, 2000; 46(10): 1562 - 1573. [Abstract] [Full Text] [PDF]

				E. M. J. J. Berns, J. A. Foekens, R. Vossen, M. P. Look, P. Devilee, S. C. Henzen-Logmans, I. L. van Staveren, W. L. J. van Putten, M. Inganäs, M. E. M.-v. Gelder, et al. Complete Sequencing of TP53 Predicts Poor Response to Systemic Therapy of Advanced Breast Cancer Cancer Res., April 1, 2000; 60(8): 2155 - 2162. [Abstract] [Full Text]

				L. You, C.-T. Yang, and D. M. Jablons ONYX-015 Works Synergistically with Chemotherapy in Lung Cancer Cell Lines and Primary Cultures Freshly Made from Lung Cancer Patients Cancer Res., February 1, 2000; 60(4): 1009 - 1013. [Abstract] [Full Text]

				H. Zhang Evaluation of Four Antibodies in Detecting p53 Protein for Predicting Clinicopathological and Prognostic Significance in Colorectal Adenocarcinoma Clin. Cancer Res., December 1, 1999; 5(12): 4126 - 4132. [Abstract] [Full Text] [PDF]

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