1 Medical Oncology Unit, Hospital of Urbino, Urbino; 2 Division of Medical Oncology, Azienda Ospedale di Parma, Parma, Italy
Received 3 September 2002; revised 27 January 2003; accepted 14 March 2003
Abstract
The benefit of postoperative adjuvant chemotherapy in patients with Dukes B colorectal cancer is still uncertain and its routine use is not recommended. Prognostic biomarkers may be useful for identifying high-risk patients with resected, node-negative disease, and this stratification may represent an innovative strategy for designing adjuvant chemotherapy trials. Featured prognostic molecular markers can be divided into the following categories: cell proliferation indeces (Ki-67, Mib-1, proliferating cell nuclear antigen); oncogenes/tumor suppressor genes [p53, K-ras, Deleted in Colorectal Cancer (DCC), Bcl-2, c-erbB2]; DNA repair (microsatellite instability); markers of angiogenesis (vascular count, vascular endothelial growth factor); markers of invasion/metastasis (plasminogen-related molecules, matrix metalloproteinases); and biochemical markers (thymidylate synthase). Studies that have investigated their prognostic role in Dukes B colorectal cancer patients are reviewed here. Current data do not provide sufficient evidence for the incorporation of available prognostic biomarkers into clinical practice. However, a biomarker-based approach could be an effective strategy for improving results of postoperative adjuvant treatments in high-risk Dukes B colorectal cancer patients. Markers of altered DCC function have shown promising prognostic role and sufficient prevalence in retrospective investigations and they deserve further assessment in prospective studies.
Key words: colorectal cancer, Dukes B, prognosis, tumor markers
Introduction
Colorectal cancer remains a significant health care problem worldwide, and the tumornodemetastasis (TNM) system represents the main tool for identifying prognostic differences among patients with early-stage disease [1]. After surgery, fluorouracil-based adjuvant chemotherapy is indicated in patients with node-positive, stage Dukes C disease. Although up to 40% of patients with Dukes B colorectal cancer will develop recurrent disease during their lifetime, the role of adjuvant chemotherapy in this setting is still unclear [1].
The identification of categories of patients with high-risk colorectal cancer would be of great help for improving treatment strategies in the node-negative disease, and probably, patients outcome [2]. A growing burden of data suggests that several prognostic molecular markers might be useful in defining individual patients risk after radical surgery and determining which patients might benefit most from adjuvant chemotherapy [3, 4]. However, since it is unlikely that all featured prognostic biomarkers will be investigated in large, prospective trials, time and effort should be given to address, in hypothesis-generating retrospective studies, those that show sufficient prevalence in colorectal carcinomas and have promising prognostic roles.
In this perspective, our aim was to evaluate current results of molecular prognostic molecular markers in Dukes B colorectal cancer and data in support of their validation in future prospective studies.
Materials and methods
Studies on prognostic molecular markers in early-stage colorectal cancer were searched for in peer-review journals; the search was restricted to English-language publications. Investigations published in abstract form only were excluded. Where results were reported or updated in more than one publication, only the most recent publication was used. The content terms colon, rectal or colorectal with prognosis, prognostic and survival were used for every featured biomarker in colorectal cancer. The CancerLit and Medline databases were used.
In the next step, each study was evaluated and those including patients with Dukes B disease were selected. These studies were assessed for methodological quality according to a six-point checklist [5] (Table 1), and only studies that fulfilled these quality features were selected for the review. The assessment of evidence was limited to and performed in studies that investigated the prognostic role of the biomarker in the subgroup of patients with Dukes B disease (Tables 28). In these studies, adjustment with multivariate analysis for all important clinico-pathological features and post-operative treatments like adjuvant chemotherapy had to be provided.
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Proliferation indeces
Antibodies that recognize nuclear proteins associated with tumor cell proliferation can be determined by immunohistochemistry and have represented an attractive alternative to the analysis of cell proliferation as determined by flow cytometry. Currently, PCNA and Ki-67, and its epitope Mib-1, are the most popular methods that have been investigated as prognostic factors in colorectal cancer. PCNA is a DNA polimerase accessory protein that complexes with cyclin D and cyclin-dependent kinases. The specific antibody recognizes PCNA protein, which is maximally elevated in late G1 and S phase of proliferating cells. The Ki-67 antigen and its epitope Mib-1 are expressed in all phases of the cell cycle except G0.
Among prognostic studies of PCNA and Ki-67 in early-stage colorectal cancer [3, 4, 613], eight studies investigated survival of Dukes B patients, and their data are summarized in Table 2 [613]. In the largest series of 293 carcinomas, which included 101 Dukes B patients, PCNA results did not correlate with survival [6]. In addition, Ki-67 analysis in two large series of 255 [7] and 465 [8] Dukes B and Dukes C patients did not reveal prognostic differences depending on the high or low labeling index. In particular, Allegra et al. [8] failed to demonstrate any significant prognostic role of Ki-67 in 465 colorectal cancer patients, with 220 patients being Dukes B2 disease. The only positive study was reported by Palmqvist et al. [9], who found an independent prognostic role of Ki-67 in 56 stage Dukes B patients when determined at the invasive margin.
Available data do not support routine evaluation of cell proliferation indeces in early stage colorectal cancer. In addition, the lack of positive prognostic indications in Dukes B disease does not support their validation in prospective trials.
In recent years, new molecules involved in the cell cycle regulation, such as cyclines or the putative tumor suppressor gene p27, seem to be more promising prognosticators than PCNA and Ki-67 [14, 15]. However, these promising data are still limited and have not been confirmed in large series of patients with node-negative disease.
Angiogenesis
Angiogenesis plays a key role in tumor growth and metastasis. This phenomenon may have prognostic relevance and it can be assessed by the vascular density in the tumor and/or by the analysis of angiogenesis promoting molecules. VEGF is a glycoprotein similar to platelet-derived growth factor, and it is considered to be the main angiogenic stimulator [16]. Microvessel counts in human tumors are performed by marking endothelial cells with specific antibodies (CD34, CD31, anti-VIII factor). VEGF expression can be determined by the analysis of mRNA levels or by immunohistochemistry with anti-VEGF antibodies, in fresh or paraffin-embedded tumor tissues [3].
The prognostic role of angiogenesis features in early stage colorectal cancer has been investigated in retrospective series of patients [12, 1732]. Seven studies [12, 1721, 23] showed the prognostic analysis in stage Dukes B patients, and their data are summarized in Table 3. In this group, three investigations found an independent prognostic role of vascular count [12, 18, 19], and one investigation with VEGF found worse disease-free survival in Dukes B patients whose tumors showed expression in >10% of tumor cells [23].
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Markers of tumor angiogenesis may discriminate prognostic differences in early stage colorectal cancer. However, lack of standardization of methods and differences in the choice of cut-off levels are relevant problems for the interpretation of results. More retrospective data are necessary before planning prospective investigations with tumor microvessell counts or VEGF expression for identifying high-risk Dukes B patients.
Metastasis and invasion
Plasminogen-related molecules and MMPs are considered crucial enzymes for tumor invasion and metastasis. They have been found to be overexpressed in colorectal carcinomas and their prognostic role has been investigated in retrospective series of patients with early stage disease.
Enzyme-linked immunosorbent essay (ELISA) and immunohistochemistry techniques have been employed to determine plasminogen-related molecules in tumor tissues. In the present review 10 studies are quoted on the prognostic role of urokinase-type plasminogen activator (uPA) and its receptor (uPAR), tissue-type plasminogen activator (tPA), plasminogen and the plasminogen inhibitors 1 and 2 (PAI-1, PAI-2) [3342]. In these studies, one or more of these molecules showed independent prognostic role in early stage disease, but only three investigations were addressed to Dukes B and Dukes C patients [4042] (Table 4). These studies found that uPAR [40], tPA [41] and uPA [42] were independent prognostic indicators in patients with stage Dukes B colorectal cancer. The largest investigation was reported by Stephens et al. [40], who determined preoperative plasma levels of soluble uPAR in 591 patients. In this study, 219 patients were classified as having Dukes B disease and those who had high plasma suPAR levels showed shorter survival than patients who had levels below the median value (P <0.0001).
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Plasminogen-related molecules are promising prognostic parameters in early-stage colorectal cancer, and the ease with which they can be analyzed supports further investigation. However, it is necessary to clarify which of the plasminogen-related molecules has the best prognostic power, and to define cut-off levels before planning their validation in prospective studies.
Thymidylate synthase
TS is a rate-limiting enzyme in the DNA synthesis pathway and is the main intracellular target of 5-fluorouracil and raltitrexed, two of the most important drugs used in the treatment of colorectal cancer. TS quantification can be performed by immunohistochemistry or RNA levels, and high TS levels have been correlated with poor response rates to fluorouracil-based chemotherapy in advanced gastrointestinal carcinomas [53]. In colorectal cancer, fluorouracil-based chemotherapy is widely used as post-surgical adjuvant treatment, and the prognostic role of TS quantitation has been investigated in patients with early stage disease [8, 5462].
Since the first report in 1994 by Johnston et al. [54], who found a significant association between TS levels and survival in 294 patients with rectal cancer, further studies have addressed the prognostic role of TS in early stage colorectal cancer. Ten studies that included stage Dukes B patients have been identified [8, 5462], and in the majority of these TS quantitation was found to be an independent prognosticator of postoperative outcome; in particular, patients with TS-positive tumors showed poorer survival than patients with TS-negative tumors [5462]. Only four studies were performed with prognostic analysis in the subgroup of Dukes B patients (Table 5), and only two of these had a sufficiently large sample to explore the efficacy of fluorouracil-based adjuvant chemotherapy according to TS expression [8, 60]. In 465 patients with Dukes B2 (220 patients) and Dukes C (245 patients) disease, Allegra et al. [8] failed to demonstrate a consistent and significant association between TS quantification and either disease-free survival or overall survival. In this study, the prognostic role of TS was evaluated according to different TS scores that were based on staining intensity and staining patterns. Edler et al. [60] found that TS expression was an independent prognostic factor in the whole group of 862 patients and in the 442 Dukes B patients who underwent surgery alone. On the contrary, in 420 patients treated with fluorouracil-based adjuvant chemotherapy, TS quantitation lost its prognostic value and patients with low TS levels showed even worse outcome than patients treated with surgery alone. In this study, TS expression was dichotomized in high and low categories according to staining intensity.
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p53
p53 is a tumor suppressor gene located on chromosome 17p13.1, encoding a protein that is involved in cell cycle regulation, DNA replication and apoptosis in response to DNA damage [63]. Accordingly, p53 status has been studied as prognostic factor, and more recently as predictor of response to cancer chemotherapy.
Two major techniques are used for p53 analysis: DNA analysis to detect a variety of mutations in the p53 gene [63] and immunohistochemistry to detect abnormal nuclear accumulation of the p53 protein [64]. Immunohistochemically detected p53 overexpression has been generally used as a surrogate marker of p53 mutations, but this assumption may not always be correct. Many genetic changes do not result in p53 overexpression, and positive immunohistochemistry analysis of p53 may occur in the absence of p53 mutations. In colorectal carcinomas, the correlation between p53 gene status and p53 staining has been estimated at 70% of cases or more [63, 64].
It is worth noting that controversies also exist on the prognostic significance of genetic p53 abnormalities. In a recently published report, a specific p53 mutational site was found to be associated with patients prognosis; in particular, patients with p53 mutations affecting a region of the core domain (the L3 zinc-binding domain) had a significantly shorter cancer-related survival [65]. Conversely, a number of p53 mutations in colorectal cancer do not seem to promote disease progression. Some mutations within conserved regions may even counteract negative functional effects of other p53 structural alterations [66].
Despite the above-mentioned difficulties in p53 analysis, this genetic marker has provoked great enthusiasm among researchers, and over 3000 patients have been included in prognostic studies in colorectal cancer between 1990 and 2000 [3, 67].
In a pooled analysis of published studies on the p53 prognostic value in colorectal cancer, neither p53 overexpression nor p53 mutations emerged as a powerful prognostic indicator in patients with colorectal cancer [67]. This analysis was performed after including survival data from 4416 patients from 28 published studies up to June 1999. In recent years, further retrospective studies have attempted to evaluate the prognostic role of p53 abnormalities in early stage colorectal cancer, but again, the results do not suggest that this marker can provide significant prognostic information. Twelve studies that included stage Dukes B patients [8, 12, 17, 18, 66, 6874], and the results of seven studies with prognostic analysis in the node-negative disease [8, 12, 17, 6871], are summarized in Table 6.
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More investigations are needed to evaluate the prognostic role of p53 abnormalities in colorectal carcinomas. Immunohistochemistry may represent an easy and accessible technique for determining p53 overexpression, but available data are inconclusive and the definition of standardized methods of analysis and cut-off levels are mandatory. To date, p53 analyses do not seem to be the best candidate for further prognostic evaluation in prospective studies.
K-ras
K-ras belongs to the RAS family (K-ras, H-ras, N-ras) of cellular proto-oncogenes and encodes a 21 kDa protein (p21) located in the inner surface of the plasma membrane. This protein controls cell growth and differentiation by transduction of extracellular mitogenic signals [75]. The functions of K-ras support its putative prognostic role in colorectal cancer, and several studies have been performed in recent years in this setting. K-ras abnormalities have been studied by molecular genetic assay for determination of point mutations at codons 12, 13, 31 and 61. The point mutations that trigger the oncogenic potential of ras have been detected in more than 80% of cases on codons 12 and 13. In addition, immunohistochemistry has been used to evaluate the ras/p21 protein expression on fixed fresh or paraffin-embedded tissue.
The large collaborative RASCAL studies [76, 77] have collected and reanalyzed survival data of patients worldwide whose tumors have been investigated for ras mutations. The first RASCAL analysis in 2721 patients was published in 1998 [76] and the second, in 4268 patients, was published in 2001 [77]. In the first assessment [76], mutations of K-ras codon 12 or 13 were detected in 37.7% of tumors, and overall survival was reduced by any mutation [hazard ration (HR) = 1.22; 95% confidence interval (CI) 1.071.40; P = 0.004]. When the authors repeated the multivariate analysis taking into account individual mutations, only glycine to valine on codon 12 (10% of the study population) was found to be an independent factor for increased risk of death (P = 0.004). The second RASCAL analysis [77] confirmed the significant association of the valine mutation on codon 12 with aggressive biological behavior in colorectal cancer (HR = 1.29; 95% CI 1.081.55; P = 0.008). The large study sample allowed subgroup analysis in patients with stage Dukes B and Dukes C disease and this genetic change (8.6% of all patients) lost its prognostic role in the node-negative disease.
After the publication of the latest RASCAL analysis, three additional studies investigated the prognostic role of K-ras mutations in early stage colorectal cancer [7880] and two found an independent prognostic role of K-ras mutations [78, 79]. Font et al. [79] found poor survival of Dukes B patients whose tumors had specific aspartic and serine mutations on codon 12 (P = 0.03). Bazan et al. [78] showed that codon 13 K-ras mutations, but not any mutation, were independently related to risk of relapse or death.
Data on the prognostic role of ras/p21 oncoprotein expression in Dukes B patients are still limited and unconfirmed [8183].
According to current data, K-ras may act as prognostic factor in colorectal cancer, but this effect seems related to a limited number of defined mutations, probably in the node-positive disease. K-ras mutations occur in 30% of colorectal carcinomas, but the prevalence of the potentially more aggressive genotypes seems lower [76, 77]. For these reasons, K-ras mutational status does not seem to be the best candidate for prognostic validation in Dukes B patients. In addition, further investigations are needed to clarify the influence of specific mutations on the biological behavior of colorectal carcinomas.
Deleted in Colorectal Cancer (DCC)
The DCC gene is localized on chromosome 18q and encodes a transmembrane protein with high homology to cell adhesion molecules [84]. Lack of the DCC protein may lead to impaired contacts between cells and contribute to tumor growth and invasion. For these reasons, the DCC gene and its protein product may have a role as natural history prognostic factor. In human colorectal carcinomas, DCC status has been investigated by molecular genetic assays or immunohistochemistry [84, 85].
Fifteen studies including stage Dukes B patients investigated the prognostic role of loss of heterozygosity (LOH) at chromosome 18q or absence of DCC expression in early-stage colorectal cancer [2, 79, 8698]. A specific subgroup prognostic analysis in stage Dukes B patients was performed in 10 studies, which are listed in Table 7. Overall, two studies were based on the DCC expression analysis [86, 93] and the remaining eight studies investigated LOH at chromosome 18q with microsatellite markers. All these investigations were retrospective and a consistent and significant prognostic role of the DCC status was found in eight of these studies [79, 8690, 92, 93].
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On the whole, data on the DCC functional assessment for identifying high-risk patients with early-stage colorectal cancer are encouraging. The prognostic role seems to be confirmed by the subgroup analyses of stage Dukes B patients, and the prevalence of DCC abnormalities in colorectal carcinomas is 4050% of all cases. These data are in favor of further testing of the DCC status in prospective clinical trials.
Microsatellite instability
The presence of a defective DNA mismatch repair mechanism results in somatic alterations in the size of simple repeat nucleotide sequences (microsatellites). This phenomenon is known as microsatellite instability (MSI) and is due to silencing of the mismatch repair genes, primarily MLH1 and MSH2 [99]. Analysis of MSI can be performed by molecular assays with microsatellite markers in tumor and corresponding normal DNA [99]. In 1999, the National Cancer Institute sponsored an International Workshop and a validated panel of five microsatellites was proposed for the identification and the assessment of MSI [100]. Tumors may be characterized as high-frequency MSI (MSI-H), if two or more of the five markers show instability, and low-frequency MSI (MSI-L), if only one of the five markers shows instability. MSI-H colorectal tumors seem to share a less aggressive clinical course than stage-matched MSI-L or microsatellite stable (MSS) tumors [100]. In addition, analysis of the specific non-coding mononucleotide BAT-26, a component of the consensus panel, was shown to be highly correlated with generalized dinucleotide repeat instability [101]. Immunohistochemistry for hMLH1 and hMSH2 expression may represent a practical test for identifying DNA mismatch repair-deficient tumors, and it has been used in retrospective prognostic analyses in colorectal carcinomas [102].
MSI was detected in 90% of tumor samples with the hereditary non-polyposis colorectal cancer syndrome (HNPCC), and patients prognosis in this group appeared to be better compared with patients with sporadic colorectal carcinomas [100]. In perspective of future adjuvant chemotherapy trials for high-risk stage Dukes B patients, this review focused on the prognostic role of MSI in early-stage, sporadic colorectal carcinomas. In this setting, among 26 selected studies [97, 98, 103126], 17 retrospective investigations [109125] showed a significant association between MSI-H or abrogated hMLH1 and hMSH2 expressions and improved prognosis. The majority of studies showed a 520% frequency of MSI-H in sporadic tumors, and only a minority of these investigations were performed with a large sample of patients that allowed the prognostic analysis in the subgroup of stage Dukes B patients [98, 107, 108, 120, 121, 124, 125] (Table 8). Only one of these studies confirmed a consistent and independent association between MSI-H and improved survival in stage Dukes B patients [125].
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Available data are insufficient to support a prognostic role of MSI in stage Dukes B colorectal cancer patients. In addition, the low frequency of this phenomenon is a major limitation for planning large prospective trials with MSI-based identification of high-risk patients.
Other prognostic biomarkers
Newly discovered molecular markers of prognosis are under investigation in early stage colorectal cancer. Some of them showed potential prognostic significance, but specific survival data in stage Dukes B patients are almost lacking, and further studies are needed [127, 128]. An overview of these new biomarkers is reported in this section.
Transforming growth factors (TGF)- and -ß1 are two components of a superfamily of molecules with mitogenic properties and promoting the growth of colon cancer cell lines in vitro. Their expression has been associated with poor survival in early-stage colorectal cancer patients, and in recent investigations TGF-ß1 emerged as independent prognostic factor [98, 129].
The c-erbB2 proto-oncogene, also known as HER-2, is a glycoprotein with tyrosine kinase activity that has been largely investigated in breast cancer. Its overexpression in colorectal carcinomas seems to be detectable in 2040% of tumors, but its prognostic role remains unclear [130, 131].
Epidermal growth factor receptor (EGFR) is another transmembrane protein with tyrosine kinase activity. The frequency of EGFR overexpression in colorectal carcinomas seems to be higher than that reported for the HER-2 glycoprotein [131], and EGFR inhibitors are under investigation in clinical trials for the treatment of colorectal cancer. However, the role of EGFR as an independent prognostic marker has not been clearly defined yet [132].
The Bcl-2/BAX complex plays a key role in the regulation of apoptosis. The bcl-2 proto-oncogene encodes a protein with anti-apoptotic properties, whereas the BAX protein is a promoter of apoptosis [128]. According to this biological background, bcl-2 overexpression and/or BAX reduced expression may have negative prognostic role in colorectal cancer. Early investigations seem to confirm this hypothesis [17], but data are still limited.
Gene damage and allelic loss in diverse chromosomes seemed to correlate with poorer survival of colorectal cancer patients [132136]. These data have not been extensively evaluated and additional studies are required.
Conclusions
Several prognostic biomarkers have been discovered over the last decades, and as we move into the 21st century, their introduction in the planning of adjuvant chemotherapy trials may help to improve and prolong lives of patients with surgically resected colorectal cancer. Unfortunately, puzzling data from current retrospective studies, poor homogeneity of inclusion criteria, variability in the techniques of analysis, and differences in the choice of cut-off levels and statistical methods all represent major barriers for interpreting results and widening applications of biomarkers in the overall management of early-stage colorectal cancer. Lack of standardization across studies is a relevant problem, especially for evaluating the results of immunohistochemically detected biomarkers. Whereas molecular markers may be assessed as a relatively simple yes/no signal for the presence of gene damage, several parameters may be included in the analysis of immunohistochemistry; number of positive tumor cells, staining intensity and staining patterns. One or more of these features can be considered in the choice of a cut-off level.
There is no clear indication of the level of evidence to be met for validating prognostic biomarkers in prospective studies. Repeated positive results in retrospective studies may suggest the prognostic utility of a molecular marker. Among these promising molecular markers, those, or the one, with sufficient frequency in colorectal carcinomas and with reliable and repeatable methods of assessment could be tested for validation in prospective trials. In this perspective, angiogenesis features, plasminogen-related molecules and loss of DCC function may warrant further investigation. Their frequency in about half of colorectal carcinomas allows the possibility of performing well-designed prospective studies with reasonable sample sizes [137]. In addition, many of these markers can be determined by immunohistochemistry, thereby enabling their analysis in the majority of laboratories worldwide. The analysis of DCC function could be preferable. In fact, it is still not clear which of the several angiogenic features and palsminogen-related molecules do have the best prognostic power. Also, immunohistochemistry for DCC showed unambiguous dichotomization between negative (05% stained cells) and positive cases, and consequently has been described an all or nothing phenomenon [86, 93].
The simultaneous testing of multiple molecular predictors of survival may supply major information [137], but the necessity of applying a multivariate model of analysis mandate for a large sample size. In a more realistic way, confirmatory studies of prognostic factors should test a single biomarker in a prospective fashion. These investigations can be embedded in trials evaluating new therapies, where the prognostic biomarker is used to identify categories of high- and low-risk patients. The population of patients with Dukes B colorectal cancer represents the target population for biomarker-based trials. In these patients, the role of adjuvant chemotherapy is unclear, and the optimal treatment strategy is still matter of debate [138, 139]. In a prospective adjuvant trial, Dukes B patients with preserved DCC could not be treated following surgical resection, whereas patients with loss of DCC function could be randomized to observation or chemotherapy.
We believe that hopes for improving the efficacy of postoperative adjuvant chemotherapy in early-stage colorectal cancer should not rely only on new drugs. Biomarker-based clinical trials do not receive the same commercial hype as new chemotherapeutic compounds, but clinicians should consider the opportunity of individualized cancer treatments based on the molecular characteristics of the tumor.
Footnotes
+ Correspondence to: Dr Francesco Graziano, Unità Operativa di Oncologia Medica, Ospedale di Urbino, via Bonconte da Montefeltro, 61029 Urbino, Italy. Tel: +39-722-301251; Fax: +39-722-301289; E-mail: frada{at}tin.it
References
1. Mcdonald JS. Adjuvant therapy of colorectal cancer. CA Cancer J Clin 1999; 49: 202219.
2. OConnell MJ, Schaid DJ, Ganju V et al. Current status of adjuvant therapy of colorectal cancer: can molecular markers play a role in predicting prognosis? Cancer 1992; 70 (Suppl): 17321739.[ISI][Medline]
3. McLeod HL, Murray GI. Tumor markers of prognosis in colorectal cancer. Br J Cancer 1999; 79: 191203.[ISI][Medline]
4. American Society of Clinical Oncology. Clinical practice guidelines for the use of tumor markers in breast and colorectal cancer. J Clin Oncol 1996; 14: 28432877.[Abstract]
5. Altman DG. Systematic reviews in heatlh care. Systematic reviews of evaluations of prognostic variables. Br Med J 2001; 323: 224228.
6. Sun XF, Carstensen JM, Stal O et al. Proliferating cell nuclear antigen (PCNA) in relation to ras, c-erbB-2, p53, clinico-pathological variables and prognosis in colorectal adenocarcinoma. Int J Cancer 1996; 69: 58.[CrossRef][ISI][Medline]
7. Jansson A, Sun XF. Ki-67 expression in relation to clinicopathological variables and prognosis in colorectal adenocarcinomas. APMIS 1997; 105: 730734.[ISI][Medline]
8. Allegra CJ, Parr AL, Wold LE et al. Investigation of the prognostic and predictive value of thymidylate synthase, p53, and Ki-67 in patients with locally advanced colon cancer. J Clin Oncol 2002; 20: 17351743.
9. Palmqvist R, Sellberg P, Oberg A et al. Low tumour cell proliferation at the invasive margin is associated with a poor prognosis in Dukes stage B colorectal cancers. Br J Cancer 1999; 79: 577581.[CrossRef][ISI][Medline]
10. Guerra A, Borda F, Javier Jimenez F et al. Multivariate analysis of prognostic factors in resected colorectal cancer: a new prognostic index. Eur J Gastroenterol Hepatol 1998; 10: 5158.[ISI][Medline]
11. Chen YT, Henk MJ, Carney KJ et al. Prognostic significance of tumor markers in colorectal cancer patients: DNA index, S-phase fraction, p53 expression, and Ki-67 index. J Gastrointest Surg 1997; 1: 266273.[CrossRef][Medline]
12. Bhatavdekar JM, Patel DD, Chikhlikar PR et al. Molecular markers are predictors of recurrence and survival in patients with Dukes B and Dukes C colorectal adenocarcinoma. Dis Colon Rectum 2001; 44: 523533.[ISI][Medline]
13. Buglioni S, DAgnano I, Cosimelli M et al. Evaluation of multiple bio-pathological factors in colorectal adenocarcinomas: independent prognostic role pf p53 and bcl-2. Int J Cancer 1999; 84: 545552.[CrossRef][ISI][Medline]
14. Handa K, Yamakawa M, Takeda H et al. Expression of cell cycle markers in colorectal carcinoma: superiority of cyclin A as an indicator of poor prognosis. Int J Cancer 1999; 84: 225233.[CrossRef][ISI][Medline]
15. Tenjo T, Toyoda M, Okuda J et al. Prognostic significance of p27(kip1) protein expression and spontaneous apoptosis in patients with colorectal adenocarcinomas. Oncology 2000; 58: 4551.[CrossRef][ISI][Medline]
16. Folkman J. Tumor angiogenesis and tissue factors. Nat Med 1996; 2: 167168.[ISI][Medline]
17. Giatromanolaki A, Stathopoulos GP, Tsiobanou E et al. Combined role of tumor angiogenesis, bcl-2, and p53 expression in the prognosis of patients with colorectal carcinoma. Cancer 1999; 86: 14211425.[CrossRef][ISI][Medline]
18. Vermeulen PB, Van den Eynden GG, Huget P et al. Prospective study of intratumoral microvessel density, p53 expression and survival in colorectal cancer. Br J Cancer 1999; 79: 316322.[CrossRef][ISI][Medline]
19. Frank RE, Saclarides TJ, Leurgans S et al. Tumor angiogenesis as a predictor of recurrence and survival in patients with node-negative colon cancer. Ann Surg 1995; 222: 695699.[ISI][Medline]
20. Banner BF, Whitehouse R, Baker SP, Swanson RS. Tumor angiogenesis in stage II colorectal carcinoma: association with survival. Am J Clin Pathol 1998; 109: 733737.[ISI][Medline]
21. Takahashi Y, Tucker SL, Kitadai Y et al. Vessel counts and expression of vascular endothelial growth factor as prognostic factors in node-negative colon cancer. Arch Surg 1997; 132: 541546.[Abstract]
22. Lindmark G, Gerdin B, Sundberg C et al. Prognostic significance of the microvascular count in colorectal cancer. J Clin Oncol 1996; 14: 461466.[Abstract]
23. Cascinu S, Staccioli MP, Gasparini G et al. Expression of vascular endothelial growth factor can predict event-free survival in stage II colon cancer. Clin Cancer Res 2000; 6: 28032807.
24. Tanigawa N, Amaya H, Matsumura M et al. Tumor angiogenesis and mode of metastasis in patients with colorectal cancer. Cancer Res 1997; 57: 10431046.[Abstract]
25. Takebayashi Y, Aklyama S, Yamada K et al. Angiogenesis as an unfavorable prognostic factor in human colorectal carcinoma. Cancer 1996; 78: 226231.[CrossRef][ISI][Medline]
26. Galindo Gallego M, Fernandez Acenero MJ, Sanz Ortega J, Aljama A. Vascular enumeration as a prognosticator for colorectal carcinoma. Eur J Cancer 2000; 36: 5560.[CrossRef][ISI][Medline]
27. Maeda K, Nishiguchi Y, Yashiro M et al. Expression of vascular endothelial growth factor and thrombospondin-1 in colorectal carcinoma. Int J Mol Med 2000; 5: 373378.[ISI][Medline]
28. Kang SM, Maeda K, Chung YS. Vascular endothelial growth factor expression correlates with hematogenous metastasis and prognosis in colorectal carcinoma. Oncol Rep 1997; 4: 381384.[ISI]
29. Amaya H, Tanigawa N, Lu C et al. Association of vascular endothelial growth factor expression with tumor angiogenesis, survival and thymidine phosphorylase/platelet-derived endothelial growth factor expression in human colorectal cancer. Cancer Lett 1997; 119: 227235.[CrossRef][ISI][Medline]
30. Lee JC, Chow NH, Wang ST, Huang SM. Prognostic value of vascular endothelial growth factor expression in colorectal cancer patients. Eur J Cancer 2000; 36: 748753.[CrossRef][ISI][Medline]
31. Ishigami SI, Arii S, Furutani M et al. Predictive value of vascular endothelial growth factor (VEGF) in metastasis and prognosis of human colorectal cancer. Br J Cancer 1998; 78: 13791384.[ISI][Medline]
32. Tokunaga T, Oshika Y, Abe Y et al. Vascular endothelial growth factor (VEGF) mRNA isoform expression pattern is correlated with liver metastasis and poor prognosis in colon cancer. Br J Cancer 1998; 77: 9981002.[ISI][Medline]
33. Yang JL, Seetoo D, Wang Y et al. Urokinase-type plasminogen activator and its receptor in colorectal cancer: independent prognostic factors of metastasis and cancer-specific survival and potential therapeutic targets. Int J Cancer 2000; 5: 431439.[CrossRef]
34. Abe J, Urano T, Konno H et al. Larger and more invasive colorectal carcinoma contains larger amounts of plasminogen activator inhibitor type 1 and its relative ratio over urokinase receptor correlates well with tumor size. Cancer 1999; 12: 26022611.[CrossRef]
35. Fujii T, Obara T, Tanno S et al. Urokinase-type plasminogen activator and plasminogen activator inhibitor-1 as a prognostic factor in human colorectal carcinomas. Hepatogastroenterology 1999; 28: 22992308.
36. Skelly MM, Troy A, Duffy MJ et al. Urokinase-type plasminogen activator in colorectal cancer: relationship with clinicopathological features and patient outcome. Clin Cancer Res 1997; 10: 18371840.
37. Sato T, Nishimura G, Yonemura Y et al. Association of immunohistochemical detection of urokinase-type plasminogen activator with metastasis and prognosis in colorectal cancer. Oncology 1995; 4: 347352.
38. Ganesh S, Sier CF, Griffioen G et al. Prognostic relevance of plasminogen activators and their inhibitors in colorectal cancer. Cancer Res 1994; 15: 40654071.
39. Raigoso P, Junco A, Andicoechea A et al. Tissue-type plasminogen activator (tPA) content in colorectal cancer and in surrounding mucosa: relationship with clinicopathologic parameters and prognostic significance. Int J Biol Markers 2000; 15: 4450.[ISI][Medline]
40. Stephens RW, Nielsen HJ, Christensen IJ et al. Plasma urokinase receptor levels in patients with colorectal cancer: relationship to prognosis. J Natl Cancer Inst 1999; 10: 869874.[CrossRef]
41. Ganesh S, Sier CF, Heerding MM et al. Contribution of plasminogen activators and their inhibitors to the survival prognosis of patients with Dukes stage B and C colorectal cancer. Br J Cancer 1997; 12: 17931801.
42. Mulcahy HE, Duffy MJ, Gibbons D et al. Urokinase-type plasminogen activator and outcome in Dukes B colorectal cancer. Lancet 1994; 8922: 583584.[CrossRef]
43. Masaki T, Matsuoka H, Sugiyama M et al. Matrilysin (MMP-7) as a significant determinant of malignant potential of early invasive colorectal carcinomas. Br J Cancer 2001; 84: 13171321.[CrossRef][ISI][Medline]
44. Bodey B, Bodey B Jr, Siegel SE, Kaiser HE. Prognostic significance of matrix metalloproteinase expression in colorectal carcinomas. In Vivo 2000; 14: 659666.[ISI][Medline]
45. Oberg A, Hoyhtya M, Tavelin B et al. Limited value of preoperative serum analyses of matrix metalloproteinases (MMP-2, MMP-9) and tissue inhibitors of matrix metalloproteinases (TIMP-1, TIMP-2) in colorectal cancer. Anticancer Res 2000; 20: 10851091.[ISI][Medline]
46. Zeng ZS, Cohen AM, Zhang ZF et al. Elevated tissue inhibitor of metalloproteinase 1 RNA in colorectal cancer stroma correlates with lymph node and distant metastases. Clin Cancer Res 1995; 8: 899906.
47. Ring P, Johansson K, Hoyhtya M et al. Expression of tissue inhibitor of metalloproteinases TIMP-2 in human colorectal cancer: a predictor of tumour stage. Br J Cancer 1997; 6: 805811.
48. Zeng ZS, Huang Y, Cohen AM, Guillem JG. Prediction of colorectal cancer relapse and survival via tissue RNA levels of matrix metalloproteinase-9. J Clin Oncol 1996; 12: 31333140.
49. Murray GI, Duncan ME, ONeil P et al. Matrix metalloproteinase-1 is associated with poor prognosis in colorectal cancer. Nat Med 1996; 4: 461462.
50. Liabakk NB, Talbot I, Smith RA et al. Matrix metalloprotease 2 (MMP-2) and matrix metalloprotease 9 (MMP-9) type IV collagenases in colorectal cancer. Cancer Res 1996; 56: 190196.[Abstract]
51. Holten-Andersen MN, Stephens RW, Nielsen HJ et al. High preoperative plasma tissue inhibitor of metalloproteinase-1 levels are associated with short survival of patients with colorectal cancer. Clin Cancer Res 2000; 11: 42924299.
52. Mysliwiec AG, Ornstein DL. Matrix metalloproteinases in colorectal cancer. Clin Colorectal Cancer 2002; 1: 208219.[Medline]
53. Johnston PG, Lenz HJ, Leichman CG et al. Thymidylate synthase gene and protein expression correlate and are associated with response to 5-fluorouracil in human colorectal and gastric tumors. Cancer Res 1995; 55: 14041412.
54. Johnston PG, Fisher ER, Rockette HE et al. The role of thymidylate synthase expression in prognosis and outcome of adjuvant chemotherapy in patients with rectal cancer. J Clin Oncol 1994; 12: 26402647.[Abstract]
55. Takenoue T, Nagawa H, Matsuda K et al. Relation between thymidylate synthase expression and survival in colon carcinoma, and determination of appropriate application of 5-fluorouracil by immunohistochemical method. Ann Surg Oncol 2000; 7: 193198.
56. Edler D, Hallstrom M, Johnston PG et al. Thymidylate synthase expression: an independent prognostic factor for local recurrence, distant metastasis, disease-free and overall survival in rectal cancer. Clin Cancer Res 2000; 6: 13781384.
57. Lenz HJ, Danenberg KD, Leichman CG et al. p53 and thymidylate synthase expression in untreated stage II colon cancer: associations with recurrence, survival, and site. Clin Cancer Res 1998; 4: 12271234.[Abstract]
58. Edler D, Kresser U, Ragnhammar P et al. Immunohistochemically detected thymidylate synthase in colorectal cancer: an independent prognostic factor of survival. Clin Cancer Res 2000; 6: 488489.
59. Yamachika T, Nakanishi H, Inada K et al. A new prognostic factor for colorectal carcinoma, thymidylate synthase, and its therapeutic significance. Cancer 1998; 82: 7077.[CrossRef][ISI][Medline]
60. Edler D, Glimelius B, Hallstrom M et al. Thymidylate synthase expression in colorectal cancer: a prognostic and predictive marker of benefit from adjuvant fluorouracil-based chemotherapy. J Clin Oncol 2002; 20: 17211728.
61. Kralovanszky J, Koves I, Orosz Z et al. Prognostic significance of the thymidylate biosynthetic enzymes in human colorectal tumors. Oncology 2002; 2: 167174.[CrossRef]
62. Kornmann M, Link KH, Galuba I et al. Association of time to recurrence with thymidylate synthase and dihydropyrimidine dehydrogenase mRNA expression in stage II and III colorectal cancer. J Gastrointest Surg 2002; 6: 331337.[CrossRef][ISI][Medline]
63. Harris CC. Structure and function of the p53 tumor suppressor gene: clues for rational cancer therapeutic strategies. J Natl Cancer Inst 1996; 88: 14421455.
64. Baas IO, Mulder JW, Offerhaus GJ et al. An evaluation of six antibodies for immunohistochemistry of mutant p53 gene product in archival colorectal neoplasms. J Pathol 1994; 172: 512.[ISI][Medline]
65. Børresen-Dale AL, Lothe RA, Meling GI et al. TP53 and long-term prognosis in colorectal cancer: mutations in the L3Zinc-binding domain predict poor survival. Clin Cancer Res 1998; 4: 203210.[Abstract]
66. Forslund A, Lonnroth C, Andersson M et al. Mutations and allelic loss of p53 in primary tumor DNA from potentially cured patients with colorectal carcinoma. J Clin Oncol 2001; 19: 28292836.
67. Petersen S, Thames HD, Nieder C et al. The results of colorectal cancer treatment by p53 status: treatment-specific overview. Dis Colon Rectum 2001; 44: 322333.[ISI][Medline]
68. Gervaz P, Bouzourene H, Cerottini JP et al. Dukes B colorectal cancer: distinct genetic categories and clinical outcome based on proximal or distal tumor location. Dis Colon Rectum 2001; 44: 364372.[ISI][Medline]
69. Buglioni S, DAgnano I, Vaselli S et al. p53 nuclear accumulation and multiploidy are adverse prognostic factors in surgically resected stage II colorectal cancers independent of fluorouracil-based adjuvant chemotherapy. Am J Clin Pathol 2001; 116: 360368.[CrossRef][ISI][Medline]
70. Soong R, Powell B, Elsaleh H et al. Prognostic significance of TP53 gene mutation in 995 cases of colorectal carcinoma. Influence on tumor site, stage, adjuvant chemotherapy and type of mutation. Eur J Cancer 2000; 36: 20532060.[CrossRef][ISI][Medline]
71. Bouzourene H, Gervaz P, Cerottini JP et al. p53 and K-ras as prognostic factors for Dukesstage B colorectal cancer. Eur J Cancer 2000; 36: 10081015.[CrossRef][ISI][Medline]
72. Gallego MG, Acenero MJ, Ortega S et al. Prognostic influence of p53 nuclear overexpression in colorectal carcinoma. Dis Colon Rectum 2000; 43: 971975.[ISI][Medline]
73. Kahlenberg MS, Stoler DL, Rodriguez-Bigas MA et al. p53 tumor suppressor gene mutations predict decreased survival of patients with sporadic colorectal carcinoma. Cancer 2000; 88: 18141819.[CrossRef][ISI][Medline]
74. Schwandner O, Schiedeck THK, Bruch H et al. p53 and Bcl-2 as significant predictors of recurrence and survival in rectal cancer. Eur J Cancer 2000; 36: 348356.[CrossRef][ISI][Medline]
75. Bos JL. Ras oncogenes in human cancer: a review. Cancer Res 1989; 49: 46824689.[Abstract]
76. Andreyev HJ, Norman AR, Cunningham D et al. Kirsten ras mutations in patients with colorectal cancer: the multicenter RASCAL study. J Natl Cancer Inst 1998; 90: 675684.
77. Andreyev HJ, Norman AR, Cunningham D et al. Kirsten ras mutations in patients with colorectal cancer: the RASCAL II study. Br J Cancer 2001; 85: 692696.[CrossRef][ISI][Medline]
78. Bazan V, Migliavacca M, Zanna I et al. Specific codon 13 K-ras mutations are predictive of clinical outcome in colorectal cancer patients, whereas codon 12 K-ras mutations are associated with mucinous histotype. Ann Oncol 2002; 13: 14381446.
79. Font A, Abad A, Monzo M et al. Prognostic value of K-ras mutations and allelic imbalance on chromosome 18q in patients with resected colorectal cancer. Dis Colon Rectum 2001; 44: 549557.[ISI][Medline]
80. Esteller M, Gonzalez S, Risques RA et al. K-ras and p16 aberrations confer poor prognosis in human colorectal cancer. J Clin Oncol 2001; 19: 299304.
81. Karelia NH, Patel DD, Desai NS et al. Prognostic significance of DNA aneuploidy and p21 ras oncoprotein expression in colorectal cancer and their role in the determination of treatment modalities. Int J Biol Markers 2001; 16: 97104.[ISI][Medline]
82. Zirbes TK, Baldus SE, Moenig SP et al. Prognostic impact of p21/waf1/cip1 in colorectal cancer. Int J Cancer 2000; 89: 1418.[CrossRef][ISI][Medline]
83. Sun XF, Ekberg H, Zhang H et al. Overexpression of ras is an independent prognostic factor in colorectal adenocarcinoma. APMIS 1998; 106: 657664.[ISI][Medline]
84. Fearon ER, Cho KR, Nigro JM et al. Identification of a chromosome 18q gene that is altered in colorectal cancers. Science 1990; 247: 4958.[ISI][Medline]
85. Turley H, Pezzella F, Kocialkowski S et al. The distribution of the deleted in colon cancer (DCC) protein in human tissues. Cancer Res 1995; 55: 56285631.[Abstract]
86. Shibata D, Reale MA, Lavin P et al. The DCC protein and prognosis in colorectal cancer. N Engl J Med 1996; 335: 17271732.
87. Jernvall P, Makinen MJ, Karttunen TJ et al. Loss of heterozygosity at 18q21 is indicative of recurrence and therefore poor prognosis in a subset of colorectal cancers. Br J Cancer 1999; 79: 903908.[CrossRef][ISI][Medline]
88. Jen J, Kim H, Piantadosi S et al. Allelic loss of chromosome 18q and prognosis in colorectal cancer. N Engl J Med 1994; 331: 213221.
89. Ogunbiyi OA, Goodfellow PJ, Herfarth K et al. Confirmation that chromosome 18q allelic loss in colon cancer is a prognostic indicator. J Clin Oncol 1998; 16: 427433.[Abstract]
90. Lanza G, Matteuzzi M, Gafa R et al. Chromosome 18q allelic loss and prognosis in stage II and III colon cancer. Int J Cancer 1998; 79: 390395.[CrossRef][ISI][Medline]
91. Carethers JM, Hawn MT, Greenson JK et al. Prognostic significance of allelic loss at chromosome 18q21 for stage II colorectal cancer. Gastroenterology 1998; 114: 11881195.[ISI][Medline]
92. Martinez-Lopez E, Abad A, Font A et al. Allelic loss on chromosome 18q as a prognostic marker in stage II colorectal cancer. Gastroenterology 1998; 114: 11801187.[ISI][Medline]
93. Sun XF, Rutten S, Zhang H, Nordenskjold B. Expression of the deleted in colorectal gene is related to prognosis in DNA diploid and low proliferative colorectal adenocarcinoma. J Clin Oncol 1999; 17: 17451750.
94. Reymond MA, Dworak O, Remke S et al. DCC protein as a predictor of distant metastases after curative surgery for rectal cancer. Dis Colon Rectum 1998; 41: 755760.[ISI][Medline]
95. Laurent-Puig P, Olschwang S, Delattre O et al. Survival and acquired genetic alterations in colorectal cancer. Gastroenterology 1992; 102: 11361141.[ISI][Medline]
96. Dix BR, 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 1994; 59: 747751.[ISI][Medline]
97. Ko JM, Cheung MH, Kwan MW et al. Genomic instability and alterations in Apc, Mcc and Dcc in Hong Kong patients with colorectal carcinoma. Int J Cancer 1999; 84: 404409.[CrossRef][ISI][Medline]
98. Watanabe N, Wu TT, Catalano PJ et al. Molecular predictors of survival after adjuvant chemotherapy for colon cancer. N Engl J Med 2001; 344: 11961206.
99. Ionov Y, Peinado MA, Malkhosyan S et al. Ubiquitous somatic mutations in simple repeated sequences reveal a new mechanism for colonic tumorigenesis. Nature 1993; 363: 558561.[CrossRef][ISI][Medline]
100. Boland CR, Thibodeau SN, Hamilton SR et al. A National Cancer Institute workshop on microsatellite instability for cancer detection and familial predisposition: development of international criteria for the determination of microsatellite instability in colorectal cancer. Cancer Res 1998; 58: 52485257.[Abstract]
101. Hoang JM, Cottu P, Thuille B et al. BAT-26 and indicator of the replication error phenotype in colorectal cancer and cell lines. Cancer Res 1997; 300303.
102. Marcus VA, Madlensky L, Gryfe R et al. Immunohistochemistry for hMLH1 and hMSH2: a practical test for DNA mismatch repair-deficient tumors. Am J Surg Pathol 1999; 23: 12481255.[CrossRef][ISI][Medline]
103. Johannsdottir JT, Bergthorsson JT, Gretarsdottir S et al. Replication error in colorectal carcinoma: association with loss of heterozygosity at mismatch repair loci and clinicopathological variables. Anticancer Res 1999; 19: 18211826.[ISI][Medline]
104. Farrington SM, McKinley AJ, Cunningham C et al. Evidence for an age-related influence of microsatellite instability on colorectal cancer survival. Int J Cancer 2002; 98: 844850.[CrossRef][ISI][Medline]
105. Salahshor S, Kressner U, Fisher H et al. Microsatellite instability in sporadic colorectal cancer is not an independent prognostic factor. Br J Cancer 1999; 81: 190193.[CrossRef][ISI][Medline]
106. Feeley KM, Fullard JF, Heneghan MA et al. Microsatellite instability in sporadic colorectal carcinoma is not an indicator of prognosis. J Pathol 1999; 188: 1417.[CrossRef][ISI][Medline]
107. Curran B, Lenehan K, Mulcahy H et al. Replication error phenotype, clinicopathological variables and patient outcome in Dukes B stage II (T3N0M0) colorectal cancer. Gut 2000; 46: 200204.
108. Gervaz P, Cerottini JP, Bouzourene H et al. Comparison of microsatellite instability and chromosomal instability in predicting survival of patients with T3N0 colorectal cancer. Surgery 2002; 131: 190197.[CrossRef][ISI][Medline]
109. Bubb YJ, Curtis LJ, Cunningham C et al. Microsatellite instability and the role of hMSH2 in sporadic colorectal cancer. Oncogene 1996; 12: 26412649.[ISI][Medline]
110. Lukish JR, Muro K, DeNobile J et al. Prognostic significance of DNA replication errors in young patients with colorectal cancer Ann Surg 1998; 227: 5156.[CrossRef][ISI][Medline]
111. Lothe RA, Peltomaki P, Meling GI et al. Genomic instability in colorectal cancer: relationships to clinicopathologic variables and family history. Cancer Res 1993; 53: 58495852.[Abstract]
112. Colombino M, Cossu A, Manca A et al. Prevalence and prognostic role of microsatellite instability in patients with rectal carcinoma. Ann Oncol 2002; 13: 14471453.
113. Choi SW, Lee KJ, Bae YA et al. Genetic classification of colorectal cancer based on chromosomal loss and microsatellite instability predicts survival. Clin Cancer Res 2002; 8: 23112312.
114. Michael-Robinson JM, Reid LE, Purdie DM et al. Proliferation, apoptosis, and survival in high-level microsatellite instability sporadic colorectal cancer. Clin Cancer Res 2001; 7: 23472356.
115. Ward R, Meagher A, Tomlinson I et al. Microsatellite instability and the clinicopathological features of sporadic colorectal cancer. Gut 2001; 48: 821829.
116. Hawkins NJ, Tomlinson I, Meagher A, Ward RL. Microsatellite-stable diploid carcinoma: a biologically distinct and aggressive subset of sporadic colorectal cancer. Br J Cancer 2001; 84: 232236.[CrossRef][ISI][Medline]
117. Gafa R, Maestri I, Matteuzzi M et al. Sporadic colorectal adenocarcinomas with high-frequency microsatellite instability. Cancer 2000; 89: 20252037.[CrossRef][ISI][Medline]
118. Massa MJ, Iniesta P, Gonzalez-Quevedo R et al. Differential prognosis of replication error phenotype and loss of heterozygosity in sporadic colorectal cancer. Eur J Cancer 1999; 35: 16761682.[CrossRef][ISI][Medline]
119. Thibodeau SN, Bren G, Schaid D. Microsatellite instability in cancer of the proximal colon. Science 1993; 260: 816819.[ISI][Medline]
120. Guidoboni M, Gafa R, Viel A et al. Microsatellite instability and high content of activated cytotoxic lymphocytes identify colon cancer patients with a favorable prognosis. Am J Pathol 2001; 159: 297304.
121. Samowitz WS, Curtin K, Ma KN et al. Microsatellite instability in sporadic colon cancer is associated with an improved prognosis at the population level. Cancer Epidemiol Biomarkers Prev 2001; 10: 917923.
122. Kruschewski M, Noske A, Haier J et al. Is reduced expression of mismatch repair genes MLH1 and MSH2 in patients with sporadic colorectal cancer related to their prognosis? Clin Exp Metastasis 2002; 19: 7177.[CrossRef][ISI][Medline]
123. Perrin J, Gouvernet J, Parriaux D et al. MSH2 and MLH1 immunodetection and the prognosis of colon cancer. Int J Oncol 2001; 19: 891895.[ISI][Medline]
124. Halling KC, French AJ, McDonnell SK et al. Microsatellite instability and 8p allelic imbalance in stage B2 and C colorectal cancers. J Natl Cancer Inst 1999; 91: 12951303.
125. Gryfe R, Kim H, Hsieh ETK et al. Tumor microsatellite instability and clinical outcome in young patients with colorectal cancer. N Engl J Med 2000; 342: 6977.
126. Gonzalez-Garcia I, Moreno V, Navarro M et al. Standardized approach for microsatellite instability detection in colorectal carcinomas. J Natl Cancer Inst 2000; 92: 544549.
127. Nicholl ID, Dunlop MG. Molecular markers of prognosis in colorectal cancer. J Natl Cancer Inst 1999; 91: 12671269.
128. Watanabe T, Chung DC. Molecular prognostic markers and colorectal cancer: the search goes on. Gastroenterology 1998; 114: 13301332.[ISI][Medline]
129. Barozzi C, Ravaioli M, DErrico A et al. Relevance of biologic markers in colorectal carcinoma: a comparative study of a broad panel. Cancer 2002; 94: 647657.[CrossRef][ISI][Medline]
130. Lee JC, Wang ST, Chow NH, Yang HB. Investigation of the prognostic value of coexpressed c-erbB family members for the survival of colorectal cancer patients after curative surgery. Eur J Cancer 2002; 38: 10651071.[CrossRef][ISI][Medline]
131. Vizoso F, Vilar C, Rodriguez JC et al. C-erbB-2 oncoprotein content in colorectal cancer and in surrounding mucosa: relationship with clinicopathologic parameters and prognostic significance. Int J Surg Investig 2000; 1: 483493.[Medline]
132. Nicholson RI, Gee JM, Harper ME. EGFR and cancer prognosis. Eur J Cancer 2001; 37 (Suppl 4): 915.[ISI][Medline]
133. Bisgard ML, Jager AC, Dalgaard P et al. Allelic loss of chromosome 2p21-16.3 is associated with reduced survival in sporadic colorectal cancer. Scand J Gastroenterol 2001; 36: 405409.[CrossRef][ISI][Medline]
134. Iniesta P, Massa MJ, Gonzalez-Quevedo R et al. Loss of heterozygosity at 3p23 is correlated with poor survival in patients with colorectal carcinoma. Cancer 2000; 89: 12201227.[CrossRef][ISI][Medline]
135. Arribas R, Ribas M, Risques RA et al. Prospective assessment of allelic losses at 4p14-16 in colorectal cancer: two mutational patterns and a locus associated with poorer survival. Clin Cancer Res 1999; 5: 34543459.
136. Gebert J, Sun M, Ridder R et al. Molecular profiling of sporadic colorectal tumors by microsatellite analysis. Int J Oncol 2000; 16: 169179.[ISI][Medline]
137. Sargent D, Allegra C. Issues in clinical trial design for tumor marker studies. Semin Oncol 2002; 29: 222230.[CrossRef][ISI][Medline]
138. Mamounas EP. Adjuvant chemotherapy for stage II colon cancer: the time has come. Eur J Surg Oncol 2000; 8: 725729.[CrossRef]
139. Wein A, Hahn EG, Merkel S, Hohenberger W. Adjuvant chemotherapy for stage II (Dukes B) colon cancer: too early for routine use. Eur J Surg Oncol 2000; 8: 730732.[CrossRef]