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The significance of cancer stem cells (CSCs) in initiation and progression of colon cancer (CC) has been established. In this study, we investigated the utility of measuring mRNA expression levels of CSC markers EpCAM, LGR5 and LGR4 for predicting survival outcome in surgically treated CC patients. Expression levels were determined in 5 CC cell lines, 66 primary CC tumors and 382 regional lymph nodes of 121 CC patients. Prognostic relevance was determined using Kaplan-Meier survival and Cox regression analyses. CC patients with lymph nodes expressing high levels of EpCAM, LGR5 or LGR4 (higher than a clinical cutoff of 0.07, 0.06 and 2.558 mRNA copies/18S rRNA unit, respectively) had a decreased mean survival time of 32 months for EpCAM and 42 months for both LGR5 and LGR4 at a 12-year follow-up (p = 0.022, p = 0.005 and p = 0.011, respectively). Additional patients at risk for recurrence were detected when LGR5 was combined with the biomarkers CXCL17 or CEA plus CXCL16. In conclusion, the study underscores LGR5 as a particularly useful prognostic biomarker and illustrates the strength of combining biomarkers detecting different subpopulations of cancer cells and/or cells in the tumor microenvironment for predicting recurrence.

G protein-coupled receptor 55 (GPR55) probably plays a role in innate immunity and tumor immunosurveillance through its effect on immune cells, such as T cells and NK cells. In this study, the prognostic value of GPR55 in colon cancer (CC) was investigated. mRNA expression levels of GPR55 were determined in 382 regional lymph nodes of 121 CC patients with 12 years observation time after curative surgery. The same clinical material had previously been analyzed for expression levels of CEA, CXCL16, CXCL17, GPR35 V2/3 and LGR5 mRNAs. Clinical cutoffs of 0.1365 copies/18S rRNA unit for GPR55 and 0.1481 for the GPR55/CEA ratio were applied to differentiate between the high- and low-GPR55 expression groups. Kaplan-Meier survival analysis and Cox regression risk analysis were used to determine prognostic value. Improved discrimination between the two groups was achieved by combining GPR55 with CEA, CXCL16 or CXCL17 compared with GPR55 alone. The best result was obtained using the GPR55/CEA ratio, with an increased mean survival time of 14 and 33 months at 5 and 12 years observation time, respectively (

Aim: This study investigates how often bowel continuity was restored after anastomotic leakage in anterior resection for rectal cancer and assesses the clinical factors associated with permanent stoma. Method: The Swedish Colorectal Cancer Registry was used to identify cases of anastomotic leakage registered in southern Sweden between January 2001 and December 2011. Patient characteristics, surgical details and clinical information about the anastomotic leakages were retrieved from medical records. Results: Of the 1442 patients operated on with anterior resection in 11 hospitals, 144 (10%) were diagnosed with anastomotic leakage after anterior resection for rectal cancer. After a median follow-up of 87 months (range 21–165), the overall rate of permanent stoma among patients with anastomotic leakage was 65%. Age ≥ 70 years (P = 0.02) and re-laparotomy (P < 0.001) were independently related to permanent stoma. Compared with nondefunctioned patients with anastomotic leakage, defunctioned patients with anastomotic leakage at the index procedure less often required re-laparotomy at some point during the entire clinical course (P < 0.001), but nondefunctioned and defunctioned patients with anastomotic leakage both had permanent stoma to the same extent (67% and 62%, respectively). Conclusion: Anastomotic leakage is highly associated with permanent stoma after anterior resection, especially in patients aged ≥ 70 years. In this cohort of patients with anastomotic leakage, 65% had permanent stoma at long-term follow-up. A defunctioning stoma ameliorates the clinical course but does not affect the end result of bowel continuity in established anastomotic leakage after anterior resection. (Less)

Supplemental Digital Content is available in the text., BACKGROUND: Lymph node metastasis is the single most important prognostic risk factor for recurrence in patients with colon cancer who have undergone curative surgery. The routine method for detecting disseminated tumor cells in lymph nodes is microscopic examination of one or a few hematoxylin and eosin-stained tissue sections by a trained pathologist. This method, however, is insensitive mainly because less than 1% of the lymph node volume is examined, leading to misclassification. OBJECTIVE: This study aimed to investigate whether analysis of a selected group of biomarker mRNAs improves detection and characterization of lymph node metastases/micrometastases compared with the routine method. DESIGN: This study is a side-by-side comparison of biomarker mRNA analysis and histopathology of 185 lymph nodes from patients with colon cancer representing stages I to IV, and an investigation of the importance of lymph node tissue volume for tumor cell detection. SETTINGS: This is a collaborative study between a high-volume central hospital and a preclinical university institution. PATIENTS: Fifty-seven patients who had undergone tumor resection for colon cancer were included. MAIN OUTCOME MEASURES: The primary outcomes measured were mRNA copies per 18S rRNA copy of CEACAM5, KLK6, SLC35D3, POSTN, and MUC2 by multiplex assay and metastases/micrometastases detected by histopathology. RESULTS: The number of tumor cell-positive lymph nodes was 1.33-fold higher based on CEACAM5 mRNA levels compared with histopathological examination. Increasing the tissue volume analyzed for CEACAM5 levels from an 80-µm section to half a lymph node increased the number of positive nodes from 34 of 107 to 80 of 107 (p < 0.0001). Similarly, the number of positive nodes for the aggressiveness marker KLK6 increased from 9 of 107 to 24 of 107. LIMITATIONS: Only a limited number of individual lymph nodes per patient was available for analysis. CONCLUSIONS: mRNA analysis of CEACAM5, KLK6, and SLC35D3 improves the detection of tumor cells in lymph nodes from patients surgically treated for colon cancer, and, together with POSTN and MUC2, it further allows characterization of the tumor cells with respect to aggressiveness and the tumor cell environment. See Video Abstract at http://links.lww.com/DCR/B650. EL ANÁLISIS DE ARNM DE CEACAM5, KLK6, SLC35D3, POSTN Y MUC2 MEJORA LA DETECCIÓN Y PERMITE LA CARACTERIZACIÓN DE CÉLULAS TUMORALES EN LOS GANGLIOS LINFÁTICOS DE PACIENTES CON CÁNCER DE COLON ANTECEDENTES: Las metástasis en los ganglios linfáticos son el factor de riesgo pronóstico más importante de recurrencia en pacientes con cáncer de colon que se han sometido a cirugía curativa. El método de rutina para detectar células tumorales diseminadas en los ganglios linfáticos es el examen microscópico de una o algunas secciones de tejido teñidas con hematoxilina-eosina por un patólogo capacitado. Sin embargo, este método es insensible principalmente porque se examina menos del 1% del volumen de los ganglios linfáticos, lo que conduce a una clasificación errónea. OBJETIVO: Investigar si el análisis de un grupo seleccionado de ARNm de biomarcadores mejora la detección y caracterización de metástasis / micrometástasis en los ganglios linfáticos en comparación con el método de rutina. DISEÑO: Una comparación en paralelo del análisis de ARNm de biomarcadores y la histopatología de 185 ganglios linfáticos de pacientes con cáncer de colon que representan las etapas I-IV, e investigación de la importancia del volumen de tejido de los ganglios linfáticos para la detección de células tumorales. ENTORNO CLINICO: Estudio colaborativo entre un hospital central de alto volumen y una institución universitaria preclínica. PACIENTES: Cincuenta y siete pacientes que han sido sometidos a resección tumoral por cáncer de colon. PRINCIPALES MEDIDAS DE VALORACION: copias de ARNm / copia de ARNr 18S de CEACAM5, KLK6, SLC35D3, POSTN y MUC2 mediante análisis múltiple y metástasis / micrometástasis detectadas por histopatología. RESULTADOS: El número de ganglios linfáticos con células tumorales positivas fue 1,33 veces mayor según los niveles de ARNm de CEACAM5 en comparación con el examen histopatológico. El aumento del volumen de tejido analizado para los niveles de CEACAM5 de una sección de 80 µm a la mitad de un ganglio linfático aumentó el número de ganglios positivos de 34/107 a 80/107 (p

OBJECTIVE: The microenvironment of colon cancer (CC) is heterogeneous including cells of myeloid lineage affecting tumor growth and metastasis. Two functional subtypes of myeloid cells have been identified; one (M1) is tumor-inhibitory and the other one (M2) is tumor-promoting. Whether the three myeloid markers EMR1, CD206 and CD86 are expressed only in the infiltrating myeloid cells or also in the tumor cells was investigated. METHODS: Expression of the myeloid markers was investigated in CC at the mRNA and protein levels in primary tumors and lymph nodes. mRNA expression was also determined in 5 CC cell lines. Protein expression was investigated by two-color immunofluorescence and consecutive-sections-immune-staining combined with morphometry using specific antibodies for the myeloid cell markers and the epithelial cell markers CEACAM5 and EpCAM. RESULTS: EMR1 and CD86, but not CD206, mRNA levels were significantly higher in CC primary tumors compared to apparently normal colon tissue (P < 0.0001). EMR1 mRNA levels were significantly higher in both hematoxylin-eosin positive (H&E(+)) and H&E(–) lymph nodes of CC patients compared to control nodes (P = 0.03 and P = 0.01, respectively). EMR1 and CD206 mRNAs were expressed in 4/5 and 5/5 CC cell lines, respectively, while CD86 mRNA was not expressed. Immuno-morphometry revealed that about 20% of the tumor cells expressed EMR1 and CD206. Positive cells were tumor cells as revealed by anti-CEACAM5 and anti-EpCAM staining. The number of EMR1, CD206 and CD86 positive cells were significantly increased in CC primary tumors compared to normal colon tissue (P < 0.0001). However, CD206 was also expressed in normal colonocytes. Only EMR1 showed significantly increased numbers of positive tumor cells in H&E(+) nodes compared to H&E(-) nodes (P = 0.001). EMR1 expression in CC tumor cells correlated with CXCL17 expressing tumor cells. CONCLUSION: EMR1, like the chemokine CXCL17, is ectopically expressed in colon cancer possibly in the same cancer cells.

Objective: Several studies indicate that macrophage migration inhibitory factor 1 plays a role for tumor progression in colon cancer. We investigated whether determination of migration inhibitory factor 1 mRNA expression levels in lymph nodes of colon cancer patients could be used as a prognostic marker. Methods: Expression levels of migration inhibitory factor 1 and carcinoembryonic antigen mRNAs were assessed in primary tumors and regional lymph nodes of 123 colon cancer patients (stages I–IV), and in colon cancer- and immune cell lines using quantitative reverse transcriptase–polymerase chain reaction. Expression of migration inhibitory factor 1 protein was investigated by two-color immunohistochemistry and immunomorphometry. Results: Migration inhibitory factor 1 mRNA was expressed at 60 times higher levels in primary colon cancer tumors compared to normal colonic tissue (medians 8.2 and 0.2 mRNA copies/18S rRNA unit; p Conclusion: Although migration inhibitory factor 1 mRNA expression levels were related to severity of disease and lymph node analysis revealed that colon cancer patients with high levels had a shorter survival time after surgery than those with low levels, the difference was small and probably not useful in clinical practice.

Chemokines are important in the development and progression of tumors. We investigated the expression of CXCL14 and CXCL16 in colon cancer. Expression of mRNA was assessed in primary tumors and lymph nodes and CXCL16 mRNA levels were correlated to patient&rsquo, s survival. Protein expression was investigated by two-color immunofluorescence and immunomorphometry. CXCL14 and CXCL16 mRNA levels and protein expression were significantly higher in colon cancer primary tumors compared to apparently normal colon tissue. Positive cells were tumor cells, as revealed by anti-CEA and anti-EpCAM staining. CXCL16, but not CXCL14, mRNA levels were significantly higher in hematoxylin and eosin positive (H&amp, E(+)) compared to H&amp, E(&minus, ) colon cancer lymph nodes or control nodes (P &lt, 0.0001). CXCL16 mRNA was expressed in 5/5 colon cancer cell lines while CXCL14 was expressed significantly in only one. Kaplan-Meier analysis revealed that colon cancer patients with lymph nodes expressing high or very high levels (7.2 and 11.4 copies/18S rRNA unit, respectively) of CXCL16 mRNA had a decreased mean survival time of 30 and 46 months at the 12-year follow-up (P = 0.04, P = 0.005, respectively). In conclusion, high expression of CXCL16 mRNA in regional lymph nodes of colon cancer patients is a sign of a poor prognosis.

The utility of mRNA and protein determinations of G protein-coupled receptor 35, that is, GPR35a (GPR35 V1) and GPR35b (GPR35 V2/3), as indicators of outcome for colon cancer patients after curative surgery was investigated. Expression levels of V1 and V2/3 GPR35, carcinoembryonic antigen and CXCL17 mRNAs were assessed in primary tumours and regional lymph nodes of 121 colon cancer patients (stage I–IV), colon cancer cell lines and control colon epithelial cells using real-time quantitative reverse transcriptase-polymerase chain reaction. Expression of G protein-coupled receptor 35 was investigated by two-colour immunohistochemistry and immunomorphometry. GPR35 V2/3 mRNA, but not V1 mRNA, was expressed in colon cancer cell lines, primary colon tumours and control colon epithelial cells. Haematoxylin and eosin positive (H&E(+)), but not H&E(–), lymph nodes expressed high levels of GPR35 V2/3 mRNA ( P

Lymph node metastasis is the most important prognostic characteristic of colorectal cancer. Carcinoembryonic antigen messenger RNA was shown to detect tumor cells that have disseminated to lymph nodes of colorectal cancer patients and to be at least as good as the hematoxylin and eosin method to predict survival in colorectal cancer patients. CXCL17 was recently shown to be ectopically expressed in colon cancer tumors. Therefore, CXCL17 may serve as prognostic marker alone or in combination with carcinoembryonic antigen. CXCL17 and carcinoembryonic antigen messenger RNA levels were determined using quantitative reverse transcription polymerase chain reaction with RNA copy standard in 389 lymph nodes of 120 colon cancer patients (stages I-IV) and 67 lymph nodes of 12 control patients with inflammatory bowel disease as well as in 68 primary tumors and 30 normal colon tissue samples. Lymph nodes of colon cancer patients were analyzed for CXCL17 and carcinoembryonic antigen protein expression by immunohistochemistry. CXCL17 messenger RNA was expressed in primary tumors at high levels, while it was barely detected in normal colon tissue ( p 0.0001). Similarly, CXCL17 messenger RNA levels were significantly higher in hematoxylin- and eosin-positive (hematoxylin and eosin (+)) lymph nodes compared to hematoxylin- and eosin-negative nodes ( p 0.0001). CXCL17 messenger RNA levels were investigated in lymph nodes grouped according to carcinoembryonic antigen messenger RNA levels: low (-), intermediate (int), and high (+). CXCL17 messenger RNA levels were higher in the carcinoembryonic antigen (int) and carcinoembryonic antigen (+) groups compared to the carcinoembryonic antigen (-) group ( p = 0.03 and p 0.0001, respectively). In lymph nodes of stage III and IV patients, CXCL17 messenger RNA levels correlated with carcinoembryonic antigen messenger RNA levels ( p 0.0001, r = 0.56 and p = 0.0002, r = 0.66, respectively). Staining of consecutive lymph node sections for CXCL17 and carcinoembryonic antigen demonstrated that the same cells expressed both proteins. Altogether, these results indicate that CXCL17 in lymph nodes is expressed by tumor cells. Patients were grouped according to the CXCL17 messenger RNA levels in the highest lymph node with low levels (-) and high levels (+). CXCL17(+) colon cancer patients showed 2.8-3.6 fold increased risk for recurrence ( p = 0.03) and decreased mean disease-free survival time of 8 months compared to CXCL17(-) colon cancer patients ( p = 0.03). CXCL17(+) carcinoembryonic antigen (int) colon cancer patients showed increased risk for recurrence by 8.3 fold ( p = 0.04) and decreased mean disease-free survival time of 46 months compared to CXCL17(-) carcinoembryonic antigen (int) colon cancer patient at follow-up after 12 years ( p = 0.02). The presence of tumor cells expressing CXCL17 in regional lymph nodes is a sign of poor prognosis. Analysis of CXCL17 messenger RNA is particularly useful to detect less differentiated colon cancer tumors expressing relatively low carcinoembryonic antigen messenger RNA levels. Thus, CXCL17 messenger RNA in combination with carcinoembryonic antigen messenger RNA may be used as a complementary tool to the hematoxylin and eosin method for detection of poorly differentiated, aggressive tumors.

Background During rectal cancer surgery the bowel may contain viable, exfoliated cancer cells, a potential source for local recurrence (LR). The amount and viability of these cells can be reduced using intraoperative rectal washout, a procedure that reduces the LR risk after anterior resection. The aim of this study was to analyse the impact of washout on oncological outcome when performed in Hartmann’s procedure (HP) for rectal cancer. Methods A national cohort study on data for patients registered from 1995 to 2007 in the Swedish Colorectal Cancer Registry was carried out. The final analysis included patients belonging to TNM stages I–III who had undergone R0 HP with a registered 5-year follow-up. Multivariate analysis was performed. Results A total of 1188 patients were analysed (686 washout and 502 no washout). No differences were detected between the washout group and the no washout group concerning rates of LR [7% (49/686) vs. 10% (49/502); p = 0.13], distant metastasis (DM) [17% (119/686) vs. 18% (93/502); p = 0.65], and overall recurrence (OAR) [21% (145/686) vs. 24% (120/502); p = 0.29]. For both groups, the 5-year cancer-specific survival was below 50%. In multivariate analysis, washout neither decreased the risk of LR, DM, or OAR nor increased overall or the cancer-specific 5-year survival. Conclusions The oncological outcome did not improve when washout was performed in HP for rectal cancer. Electronic supplementary material The online version of this article (doi:10.1007/s10151-017-1637-5) contains supplementary material, which is available to authorized users.

Improved outcome after rectal cancer surgery requires identification of novel risk factors of tumour recurrence in order to personalise therapy, that is, enhanced selection of high-risk patients to additional radiochemotherapy or intensified follow-up. In several tumour types, including colorectal cancer, high expression of the membrane–cytoskeleton linker ezrin has been suggested to impair prognosis but has not yet reached clinical application. We evaluated the expression of ezrin in rectal cancer with a focus on the identification of a marker for local tumour recurrence. Immunohistochemical expression of ezrin was analysed in 104 primary rectal cancers from patients who developed local recurrences despite being treated with R0 major abdominal surgery. Time to local recurrence and distant metastasis as well as 5-year overall and cancer-specific survival were used as end points. Ezrin expression was weak in 17% of the tumours, moderate in 62%, and intense in 21%. The time to local recurrence was significantly shorter (p = 0.0004) for patients with tumours showing high ezrin expression. No correlation between ezrin expression and time to distant metastasis was identified. Survival data were similar between groups irrespective of ezrin expression in the primary tumours. Our findings suggest that increased expression of ezrin may represent a marker of aggressive biological behaviour in rectal cancer. Although further validation is needed, ezrin may represent a relevant marker for personalised treatment of rectal cancer with respect to risk of local recurrence after R0 surgery.

The aim was to explore the utility for staging and prognostic impact of carcinoembryonic antigen (CEA), cytokeratin 20 (CK20), guanylyl cyclase C (GCC), CUB (complement protein subcomponents C1r/C1s, urchin embryonic growth factor, and bone morphogenic protein 1) containing domain protein 1 (CDCP1) and mucin 2 (MUC2) mRNA levels in mesenteric lymph nodes of colorectal cancer (CRC) patients. Lymph nodes were collected at surgery and bisected; one half was subjected to biomarker mRNA analysis using real-time quantitative RT-PCR and the other half to routine histopathology. Lymph nodes from 174 CRC patients and 24 controls were analyzed. The median follow-up time was 59 (range 17-131) months. Cut-off levels were defined by analyzing quintiles by Cox regression model. CEA mRNA showed the best discriminating power between patients with recurrence in CRC after surgery and patients who were apparently disease-free (p = 0.015). The risk of recurrence for the CEA(+) patients was 4.6 times greater than for the CEA(-) patients (p < 0.0001). The other biomarkers gave lower hazard ratios. Cumulative survival analysis demonstrated that the average survival time was 99 months for CEA(-) patients compared to 39 months for CEA(+) patients, a difference of 60 months (p < 0.0001). Six to nine percent of the Stage I and Stage II patients [H&E(-)] had CEA(+), CK20(+), GCC(+) and/or MUC2(+) lymph nodes. Two of these patients died from recurrent CRC. Low lymph node MUC2/CEA mRNA ratio identified patients with high risk for recurrence (p = 0.011). Thus, quantitative reverse transcriptase-polymerase chain reaction of CEA mRNA is a sensitive method to identify tumor cells in lymph nodes of CRC patients and, in combination with MUC2 mRNA, allows improved prediction of clinical outcome.

Aim The impact of anastomic leakage (AL) on the oncological outcome after anterior resection (AR) for rectal cancer is still controversial. We explored the impact of AL regarding local recurrence ( ...

Background Adenocarcinomas of the rectum shed viable cells, which have the ability to implant. Intraoperative rectal washout decreases the amount and viability of these cells, but there is no conclusive evidence of the effect of rectal washout on local recurrence after rectal cancer surgery. Methods Data were analysed from a population-based registry of patients who had anterior resection from 1995 to 2002 and were followed for 5 years. Rectal washout was performed at the discretion of the surgeon. National inclusion of patients with rectal cancer and follow-up was near complete (approximately 97 and 98 per cent respectively). Results A total of 4677 patients were analysed (3749 who had washout, 851 no washout and 77 with information missing); 52·0 per cent of patients in the washout group and 41·4 per cent in the no-washout group had preoperative radiotherapy (P < 0·001). Local recurrence rates were 6·0 and 10·2 per cent respectively (P < 0·001). Univariable and multivariable logistic regression analyses produced odds ratios that favoured washout: 0·56 (95 per cent confidence interval (c.i.) 0·43 to 0·72) and 0·61 (0·46 to 0·80) respectively (both P < 0·001). In multivariable analysis restricted to patients who had curative surgery, the odds ratio was 0·59 (95 per cent c.i. 0·44 to 0·78; P < 0·001). Conclusion There was a more favourable outcome in patients after rectal washout than without.

Identification of risk factors of poor oncological outcome in rectal cancer surgery is of utmost importance. This study examines the impact of incidental perforation on the oncological outcome.Using the Swedish Rectal Cancer Registry, patients were selected who received major abdominal surgery for rectal cancer between 1995 and 1997 with registered incidental perforation. A control group was also selected for analysis of the oncological outcome after 5-year follow-up. Multivariate analysis was performed. Registry data were validated, and additional data were supplemented from medical records.After validation and exclusion of non-radically operated patients, 118 patients with incidental perforation and 155 controls in TNM stages I-III were included in the analysis. The rate of local recurrence (LR) [20% (23/118) vs. 8% (12/155) (p = 0.007)] was significantly higher among patients with perforation, whereas the rates of distant metastasis [27% (32/118) vs. 21% (33/155) (p = 0.33)] and overall recurrence (OAR) [35% (41/118) vs. 25% (38/155) (p = 0.087)] were not significantly different between the groups. Overall as well as cancer-specific 5-year survival rates were significantly reduced for the patients with perforation [44 vs. 64% (p = 0.002) and 66 vs. 80% (p = 0.026), respectively]. In the multivariate analysis, perforation was a significant risk factor of increased rates of LR and OAR as well as reduced 5-year overall and cancer-specific survival.Incidental perforation in rectal cancer surgery is an important risk factor of poor oncological outcome and should be considered in the discussion concerning postoperative adjuvant treatment as well as the follow-up regime.

Aim: Despite advances in rectal cancer treatment, local recurrence (LR) remains a significant problem. To select high-risk patients for different treatment options aimed at reducing LR, it is essen ...

Background An audit of all patients with rectal cancer in Sweden was launched in 1995. This is the first report from the Swedish Rectal Cancer Registry (SRCR). Methods Between 1995 and 2003, 13 434 patients treated for adenocarcinoma of the rectum were registered with the SRCR; there were approximately 1500 new patients annually. Results Approximately half had an anterior resection, a quarter an abdominoperineal resection and 15 per cent a Hartmann's procedure. The median 30-day postoperative mortality rate was 2·4 per cent and the overall postoperative morbidity rate was 35·0 per cent. The 5-year cancer-specific survival rate was 62·3 per cent. The 5-year relative survival rate was 70·1 per cent after anterior resection, 59·8 per cent after abdominoperineal resection and 39·8 per cent after a Hartmann's procedure. The crude 5-year local recurrence rate was 9·5 per cent overall, 6·1 per cent after preoperative radiotherapy and 11·4 per cent after surgery alone. For 3868 patients who had a locally curative procedure the local recurrence rate was 7·4 per cent overall, 5·9 per cent for those who had radiotherapy and 10·2 per cent for those who did not. The local recurrence rate was 2·9 per cent (28 of 968) for stage I disease, 7·9 per cent (112 of 1418) for stage II, 13·9 per cent (188 of 1357) for stage III and 8·5 per cent (45 of 532) for stage IV. Conclusion These good population-based results are due, in part, to the nationwide prospective quality assurance registration.

The main aims were to explore time trends in the management and outcome of patients with rectal cancer in a national cohort and to evaluate the possible impact of national auditing on overall outcomes. A secondary aim was to provide population-based data for appraisal of external validity in selected patient series.Data from the Swedish ColoRectal Cancer Registry with virtually complete national coverage were utilized in this cohort study on 29 925 patients with rectal cancer diagnosed between 1995 and 2012. Of eligible patients, nine were excluded.During the study period, overall, relative and disease-free survival increased. Postoperative mortality after 30 and 90 days decreased to 1.7% and 2.9%. The 5-year local recurrence rate dropped to 5.0%. Resection margins improved, as did peri-operative blood loss despite more multivisceral resections being performed. Fewer patients underwent palliative resection and the proportion of non-operated patients increased. The proportions of temporary and permanent stoma formation increased. Preoperative radiotherapy and chemoradiotherapy became more common as did multidisciplinary team conferences. Variability in rectal cancer management between healthcare regions diminished over time when new aspects of patient care were audited.There have been substantial changes over time in the management of patients with rectal cancer, reflected in improved outcome. Much indirect evidence indicates that auditing matters, but without a control group it is not possible to draw firm conclusions regarding the possible impact of a quality control registry on faster shifts in time trends, decreased variability and improvements. Registry data were made available for reference.

Accurate identification of lymph node involvement is critical for successful treatment of patients with colorectal carcinoma (CRC). Real-time quantitative RT-PCR with a specific probe and RNA copy standard for biomarker mRNA has proven very powerful for detection of disseminated tumour cells. Which properties of biomarker mRNAs are important for identification of disseminated CRC cells? Seven biomarker candidates, CEA, CEACAM1-S/L, CEACAM6, CEACAM7-1/2, MUC2, MMP7 and CK20, were compared in a test-set of lymph nodes from 51 CRC patients (Dukes' A-D) and 10 controls. Normal colon epithelial cells, primary tumours, and different immune cells were also analysed. The biomarkers were ranked according to: (1) detection of haematoxylin/eosin positive nodes, (2) detection of Dukes' A and B patients, who developed metastases during a 54 months follow-up period and (3) identification of patients with Dukes' C and D tumours using the highest value of control nodes as cutoff. The following properties appear to be of importance; (a) no expression in immune cells, (b) relatively high and constant expression in tumour tissue irrespective of Dukes' stage and (c) no or weak downregulation in tumours compared to normal tissue. CEA fulfilled these criteria best, followed by CK20 and MUC2.

Biallelic germline mutations in the base excision repair gene MYH have been shown to predispose to a proportion of multiple colorectal adenomas and cancer. To evaluate the contribution of MYH mutations to non- FAP, non-HNPCC familial colorectal cancer, 84 unrelated Swedish individuals affected with colorectal cancer from such families were screened for germline mutations in the coding sequence of the gene. None of the cases was found to carry any pathogenic sequence change. We then determined the prevalence of the two most common pathogenic MYH mutations found in Caucasians, Y165C and G382D, in 450 Swedish sporadic colorectal cancer cases and 480 Swedish healthy controls. The frequency of both variants in Swedish cases and controls was similar to those previously reported. In addition, we found that previously unknown sequence variations at the position of amino acid 423 (R423Q, R423P, and R423R) appear to occur more frequently in cases than in controls (p = 0.02), a finding that warrants future studies.

The purpose of our study was to develop specific, sensitive, objective assays for early detection of disseminated tumour cells in patients with colorectal cancer (CRC). Carcinoembryonic antigen (CEA) and cytokeratin 20 (CK20) were chosen as markers because they are selectively expressed in epithelial cells with maintained expression in CRC. Real-time quantitative RT-PCR assays with RNA copy standards were constructed. Regional lymph nodes were collected from patients with CRC (n = 51) and benign intestinal disease (n = 10). Results were compared to routine histopathology and anti-CEA immunohistochemistry. Lymph node levels of CEA and CK20 mRNA correlated strongly (p < 0.0001, r = 0.8). Lymph nodes from non-CRC patients had

PURPOSE: Carcinoembryonic antigen is the classical tumor marker for colorectal cancer. The main clinical utility is in monitoring patients with colorectal cancer. Like carcinoembryonic antigen, the plasma level of CA 242 is elevated in patients with colorectal cancer. The purpose of this study was to investigate whether the plasma levels of carcinoembryonic antigen and/or CA 242 were elevated before clinical diagnosis of colorectal cancer. METHODS: The Northern Sweden Health and Disease Cohort was linked to the Swedish National and Regional Cancer registries, and 124 prospective cases with colorectal cancer were identified. Two referents for each case were randomly selected and matched for gender, age, date of sampling, and fasting time. Plasma from the included patients was analyzed for carcinoembryonic antigen and CA 242 using specific immunoassays. RESULTS: An elevated level of carcinoembryonic antigen before diagnosis was associated with an increased risk of developing manifest colorectal cancer (adjusted odds ratio, 7.9; 95 percent confidence interval, 2.1–29.1; P = 0.002). An elevated level of CA 242 was not significantly related to colorectal cancer risk. Elevated carcinoembryonic antigen levels were only seen in samples collected in the two-year time interval immediately before diagnosis. In this group, 30.4 percent of all plasma samples from cases were carcinoembryonic antigen-positive and 71.4 percent were future Dukes A or B cases. The specificity of the carcinoembryonic antigen test for identifying future colorectal cancer patients was 0.99 with a sensitivity of 0.12. For CA 242 the specificity was 0.92 and the sensitivity was 0.1. CONCLUSIONS: Elevated carcinoembryonic antigen levels strongly indicate occult colorectal cancer. Although the specificity of the carcinoembryonic antigen test in its present form is high, the sensitivity is disappointingly low, prohibiting the use of the carcinoembryonic antigen test for mass screening.

One of the most prevalent hereditary syndromes associated with colorectal cancer is hereditary nonpolyposis colorectal cancer (HNPCC). The inherited gene defects in HNPCC have been shown to reside in DNA mismatch repair genes, mostly hMSH2 or hMLH1. Most HNPCC patients are heterozygous with regard to the relevant mismatch repair gene; they have one normal and one mutated allele, and mismatch repair in normal somatic cells is functional. Cancer predisposition in HNPCC is believed to be associated with the loss of the wild-type allele in somatic cells, resulting in defective DNA mismatch repair. This gives rise to DNA microsatellite instability (MSI), an increased somatic mutation rate, and eventually, to the accumulation of mutations in genes involved in colorectal carcinogenesis. In support of this theory, colorectal tumors in HNPCC patients and in mice deficient for hMSH2 or hMLH1 show MSI. Here, we describe two missense mutations in hMLH1 exon 16 associated with colorectal cancer. Interestingly, the tumors do not show MSI. This raises some potentially important issues. First, even microsatellite-negative colorectal tumors can be associated with germline mutations and these will be missed if an MSI test is used to select patients for mutation screening. Second, the lack of MSI in these cases suggests that the mechanism involved in carcinogenesis could be different from that generally hypothesized.

Hereditary non-polyposis colorectal cancer (HNPCC) is linked to an inherited defect in the DNA mismatch repair system. DNA from HNPCC tumours shows microsatellite instability (MSI). It has been reported that HNPCC patients have a better prognosis than patients with sporadic colorectal cancer. We examined whether the presence of MSI in a series of unselected colorectal tumours carries prognostic information. In a series of 181 unselected colorectal tumours, 22 tumours (12%) showed MSI. Survival analysis at 5–10 years follow-up showed no statistically significant difference in prognosis between MSI-positive and -negative tumours. Our results suggest that the MSI phenotype as such is not an independent prognostic factor. © 1999 Cancer Research Campaign

PURPOSE: The aim was to investigate the significance of lymph node micrometastases in Dukes Stages A and B colorectal cancer. METHODS: Archival specimens were examined from 147 patients (96 colon, 51 rectum; 44 Stage A, 103 Stage B) who had surgery between 1987 and 1994. One lymph node section from each node (colon, 1–11; median, 4; rectum, 1–15; median, 3) was examined with use of an anticytokeratin antibody. RESULTS: Forty-seven (32 percent) patients had micrometastases. At follow-up in June 1996, 23 patients had died of cancer or with known tumor relapse, after a median time of 28 (range, 5–67) months; 8 of 47 (17 percent) patients had micrometastases, 15 of 100 (15 percent) did not. No statistically significant differences were observed according to micrometastases when the results were analyzed with respect to Dukes stage or survival time. The median survival time of living patients with micrometastases was 48 (range, 18–97) months, and for patients without micrometastases, 48 (range, 19–111) months. Six of 96 living patients had a tumor relapse; three of these displayed micrometastases. CONCLUSION: Lymph node micrometastases are not a useful prognostic marker in Dukes Stages A and B and do not imply different strategies for additional therapy or follow-up.

Mutations in the KRAS gene is a key event in the carcinogenesis of many human cancers and may serve as a diagnostic marker and a target for therapeutic intervention. In this study we have applied three different techniques for mutation detection of KRAS exon 1 mutations: Allele specific polymerase chain reaction (AS-PCR), temporal temperature gradient electrophoresis (TTGE) and constant denaturant capillary electrophoresis (CDCE). Samples from 191 sporadic colon carcinomas were analyzed. AS-PCR were performed with oligonucleotides specific for know mutations in codon 12 and 13 of the KRAS gene. In TTGE analyses, linear ramping of the temperature were performed during electrophoresis in a constant denaturant gel. CDCE analyses were performed using fluorescin labeled PCR-products. Separation was achieved under constant denaturing conditions using high temperature in a gel-filled capillary followed by laser detection. A mutated KRAS gene was found in 42/191 (22.0%) of the samples using AS-PCR, in 62/191 (32.5%) using TTGE and in 66/191 (34.6%) of the samples using CDCE. In the TTGE and CDCE analyses the sequence of the mutant were determined by comparing the electrophoretic pattern to that of known mutations or by mixing the sample with known mutations prior to reanalysis. In a titration experiment mixing mutant and wild-type alleles prior to PCR, the sensitivity for mutation detection was shown to be 10(-2) for TTGE and under optimized conditions 10(-3) for CDCE.

Background Biofragmentable anastomosis ring (BAR) has been proven to be a safe anastomotic device in elective surgery. The use of this anastomotic ring in high-risk patients has not been established. Methods During a 5-year period (1990-1995), 100 high-risk patients undergoing colonic resection and suitable for a primary anastomosis were allocated randomly to a standard suture technique or to anastomosis performed with a BAR. High risk was defined as large bowel obstruction, complicated diverticular disease, Crohn's disease, local cancer recurrence, previously irradiated colon, and trauma to the colon or rectum. The patients were equally distributed to the two groups regarding sex, age, emergency surgery and concomitant diseases. Results In three patients allotted to the BAR group, the device could not be used. There were three (6 per cent) postoperative deaths in each group; none was related to anastomotic problems. Three anastomotic dehiscences were diagnosed, two (4 per cent) in the BAR group and one (2 per cent) in the suture group. Postoperative complications and postoperative recovery were similar. Conclusion This study shows that the BAR anastomosis probably is as safe as the standard hand-sewn anastomosis in high-risk colorectal surgery. As the cost of a BAR anastomosis is substantially higher than that for a hand-sewn anastomosis, the latter technique is still the preferred method in the authors' unit.

To investigate whether endosonography is reliable in making radiation therapy decisions in rectal cancer, with possible downstaging taken into consideration.Ninety patients (52 men, 38 women; median age, 69 years) with rectal adenocarcinoma underwent endosonography within 2 weeks before surgery and radiation therapy (performed in 54 patients). The tumor invasive edge was used for radiation therapy decision making.The local stage was accurately assessed in 65 patients (39 with and 26 without irradiation). The tumor invasive edge was accurately assessed in 63 patients. Overstaging was present in 19 patients; the tumor had grown almost through the muscularis propria in six. The invasive edge (P = .1) and lymph node status were overstaged more often in the patients with than in the patients without irradiation. Tumor was understaged in eight patients: The invasive edge did not penetrate but there was budding beyond the muscularis propria in five; the invasive edge penetrated the muscularis propria in two. In seven of the eight patients, growth beyond the muscularis propria was smaller than the endosonographic resolution. Three patients with understaged, nonirradiated tumors developed pelvic recurrence. None of the patients with irradiation and none of the 16 patients without irradiation but with correct assessment developed pelvic recurrence.Preoperative irradiation decision making on the basis of endosonographic findings is uncertain. Downstaging after preoperative irradiation must be considered.

A total of 169 colorectal adenocarcinomas, obtained from patients with a median follow-up of 6.5 years, were studied with immunohistochemical staining on cryosections using a monoclonal anti-tenascin antibody to evaluate the possible association between the staining patterns and tumour stage, tumour differentiation and survival. We found two different staining patterns in the tumour stroma--a diffuse stromal fibrillar staining in 92 out of 169 (54%) tumours and a subglandular staining in the remaining 77 tumours. When the entire group of patients (P < 0.01) and the group of potentially cured patients (P < 0.03) were analysed univariately, it was found that diffuse stromal fibrillar staining was associated with a shorter survival time than subglandular staining. In a multivariate analysis, the Dukes' stage and age were independent prognostic factors, whereas the tenascin expression did not retain a clear independent relationship to survival (P = 0.06). Hence, it appears that the tumour expression of tenascin may be a potential prognostic marker in colorectal cancer, in so far as a diffuse stromal fibrillar staining pattern seems to indicate an increased risk of poor outcome. However, after adjustment for age and Dukes' stage, the additional prognostic value of tenascin remains to be established in further analyses. Images Figure 1

Founded in 1995, the Swedish Rectal Cancer Registry (SRCR) is frequently used for rectal cancer research. However, the validity of the registry has not been extensively studied. This study aims to validate a large amount of registry data to assess SRCR quality.The study comprises 906 patients treated with major abdominal surgery registered in the SRCR between 1995 and 1997. SRCR data for 14 variables were scrutinized for validity against the medical records. Kappa's and Kendall's correlation coefficients for agreement between SRCR data and medical records data were calculated for 13 variables.For 11 variables, concerning the tumor, neoadjuvant therapy, the surgical procedure, local radicality and TNM stage, data were missing in 5% or less of the registrations; for the remaining three variables, anastomotic leakage, local and distant recurrence, data were missing in 13-38%. For the variables surgery performed or not and type of surgical procedure, no data were missing. Erroneous registrations were found in less than 10% of all variables; for the variables preoperative chemotherapy and surgery performed or not, all registrations were correct. For the variables concerning neoadjuvant therapy, local radicality according to the surgeon as well as the pathologist and distant metastasis, the false-positive or -negative registrations were equally distributed, and for the variables rectal washout, rectal perforation, anastomotic leakage and local recurrence there was a discrepancy in distribution. The correlation coefficient for 12 variables ranged from 0.82 to 1.00, and was 0.78 for the remaining variable.The validity of the SRCR was good for the initial three registry years. Thus, research based on SRCR data is reliable from the beginning of the registry's use.

This study aimed to investigate whether immunohistochemical staining for nm23-H1 protein in the primary tumour is correlated with tumour stage, tumour differentiation, DNA ploidy, cell proliferative index, p53 status and patient survival time in colorectal cancer. Full-cross colorectal cancer biopsies were collected from 202 consecutive surgical specimens between 1987 and 1990. Immunohistochemical expression of nm23-H1 protein was investigated in cryosections, using a monoclonal anti-nm23-H1 antibody (clone NM 301). The staining pattern was classified as follows: strong homogeneous intensity, moderate homogeneous intensity, moderate focal intensity, or as negative. Immunohistochemical expression of p53 was investigated using a monoclonal anti-p53 antibody (DO-7). The DNA ploidy and cell proliferative index were determined by flow cytometry. Possible correlation between nm23-H1 staining patterns and the other studied tumour characteristics was explored at the end of 1994. Median survival time of living patients was 66 months, range 50-93 months. No correlation was found between various nm23-H1 staining patterns and tumour stage, cell proliferative index or p53 status. Nm23-H1-negative tumours and tumours with moderate focal staining intensity were less differentiated than tumours with strong homogeneous or moderate homogeneous staining intensity (P < 0.05). Of the nm23-H1-negative tumours, a significantly higher number was near-diploid rather than aneuploid, as compared with those expressing positive nm23-H1 (P < 0.05). The number of dead patients in Dukes' stages B and C did not correlate significantly with the nm23-H1 staining pattern. The nm23-H1 staining pattern alone, or combined with either of the other explored tumour characteristics, did not correlate with patient survival time. Immunohistochemical studies of the nm23-H1 protein expression are of minor value in the staging and prognostic prediction of colorectal cancer. Images Figure 1

The tumour markers carcinoembryonic antigen (CEA), tissue polypeptide antigen (TPA), TPS, CA 19-9, CA 50 and CA 242 were analysed in serum from 203 potentially curable colorectal cancer patients. The levels of all markers increased with increasing tumour stage, and all markers correlated with survival. Multivariate analyses indicated that the Dukes stage had the best prognostic explanatory power, followed by TPA. In the subset of 166 potentially cured patients, the prognostic information by the markers was substantially reduced. We conclude that preoperative serum tumour marker measurements have the potential to aid therapy selection, but also that their clinical usefulness is not immediately apparent.

A defunctioning loop ileostomy in low anterior resection reduces the incidence and morbidity of an anastomotic leakage, but complications related to the stoma may occur. We explored stoma-associated complications during the stoma period and after stoma reversal. A retrospective analysis of rectal cancer patients operated with low anterior resection and a defunctioning loop ileostomy at Helsingborg Hospital and Malmo University Hospital from January 2007 to June 2009 was undertaken. Ninety-two patients were included, of whom 82 (89 %) underwent stoma reversal. The median stoma period was 6.2 ± 3.2 months. Sixty-six percent of the patients suffered from minor or major stoma-associated morbidity. The complication rate was significantly related to the stoma time (p

Background:A most important characteristic feature for poor prognosis in colorectal cancer (CRC) is the presence of lymph node metastasis. Determination of carcinoembryonic antigen (CEA) mRNA levels in lymph nodes has proven powerful for quantification of disseminated tumour cells. Here, we investigate the utility of human tissue kallikrein-related peptidase 6 (KLK6) mRNA as a progression biomarker to complement CEA mRNA, for improved selection of patients in need of adjuvant therapy and intensified follow-up after surgery. Methods:Lymph nodes of pTNM stage I-IV CRC- (166 patients/503 lymph nodes) and control (23/108) patients were collected at surgery and analysed by quantitative RT–PCR. Results:Lymph node KLK6 positivity was an indicator of poor outcome (hazard ratio 3.7). Risk of recurrence and cancer death increased with KLK6 lymph node levels. Patients with KLK6 lymph node levels above the 90th percentile had a hazard ratio of 6.5 and 76 months shorter average survival time compared to patients with KLK6 negative nodes. The KLK6 positivity in lymph nodes with few tumour cells, that is, low CEA mRNA levels, also indicated poor prognosis (hazard ratio 2.8). Conclusion:In CRC patients, lymph node KLK6 positivity indicated presence of aggressive tumour cells associated with poor prognosis and high risk of tumour recurrence. Originally included in thesis in manuscript form with title: "Lymph node kallicrein 6 mRNA: a new progression marker for colorectal cancer".

We have investigated the expression and cellular source of vitronectin in colorectal adenocarcinoma. Immunofluorescence staining of tissue sections revealed the presence of vitronectin in the stroma of the 11 tumors studied, but not in adjacent normal colon. A method was devised for the isolation from colorectal adenocarcinomas of fibroblast-like cells that stained positive for vimentin but negative for cytokeratin. These tumor-derived stromal cells synthesized and secreted vitronectin, as revealed by metabolic labeling and immunoprecipitation. This was confirmed by Southern blot analysis of polymerase chain reaction amplification products from reverse-transcribed RNA. Normal skin fibroblasts did not synthesize vitronectin. Immunofluorescence staining showed vitronectin deposited at focal contact sites in the tumor-derived cells, where it colocalized with vinculin and the alpha v integrin subunit. The deposition of vitronectin into focal contact sites was not dependent on the presence of serum. The finding that vitronectin can be synthesized and secreted by tumor-derived fibroblast-like cells in culture indicates that vitronectin expression can be promoted by as yet unknown signals provided in disease states, such as cancer.

PURPOSE: This study was designed to investigate, in a population-based setting, the surgical outcome in patients with rectal cancer according to the hospital volume. METHODS: Since 1995 all patients with rectal cancer have been registered in the Swedish Rectal Cancer Registry. Hospitals were classified, according to number treated per year, as low-volume, intermediate-volume, or high-volume hospitals ( 25 procedures per year). Postoperative mortality, reoperation rate within 30 days, local recurrence rate, and overall five-year survival were studied. For postoperative morbidity and mortality the whole cohort from 1995 to 2003 (n = 10,425) was used. For cancer-related outcome only, those with five-year follow-ups, from 1995 to 1998, were used (n = 4,355). RESULTS: In this registry setting the postoperative mortality rate was 3.6% in low-volume hospitals, and 2.2% in intermediate-volume and high-volume hospitals (P = 0.002). The reoperation rate was 10%, with no differences according to volume. The overall local recurrence rates were 9.4%, 9.3%, and 7.5%, respectively (P = 0.06). Significant difference was found among the nonirradiated patients (P = 0.004), but not among the irradiated patients (P = 0.45). No differences were found according to volume in the absolute five-year survival. CONCLUSION: Postoperative mortality and local recurrence in nonirradiated patients were lower in high-volume hospitals. No difference was seen between volumes in reoperation rates, overall local recurrence, or absolute five-year survival.

Colorectal cancer is ranked third in worldwide incidence for women and fourth for men representing 9% of the world cancer or approximately 1 million new cases for 2002 (Parkin et al., 2005). Two ...

No surveillance is recommended after radical excision of low-risk adenomas (pedunculated adenoma irrespective of size, sessile adenomaor = 10 mm, numberor = 2. An endoscopic check-up is recommended 3-6 months after radical excision of high-risk adenomas (sessile adenoma10 mm, numberor = 3), as well as after excision of a pedunculated or a sessile adenoma with an unclear resection margin. All above is irrespective of histopathological adenoma classification. An endoscopic check-up is recommended 3 months after radical excision of a highly or moderately differentiated malignant polyp with no sign of invasion into blood or lymph vessels and with a maximum invasion depth stage T1-sm1. Surgical resection is necessary if the malignant polyp is poorly differentiated, and/or invades into blood or lymph vessels, and/or is stage T1-sm3, or is excised with unclear resection margins. Treatment for stage T1-sm2 polyps may be individualized. Individuals with low-risk adenomas and a first degree relative with colorectal cancer, individuals having high-risk adenomas or malignant polyps removed, as well as individuals operated on for colorectal cancer should be subjected to colonoscopy after three years and then every fifth year whenor = 75 years of age.

Hereditary nonpolyposis colorectal cancer (HNPCC) is commonly associated with at least three currently known DNA mismatch repair genes: (a) hMSH2; (b) hMLH1; and (c) hMSH6. A majority of HNPCC families has identifiable mutations in hMLH1 and hMSH2. When these mutations cause an inherited risk of colorectal cancer, they are also most often associated with microsatellite instability in the tumors. Recently, hMLH3 was suggested to be causative in HNPCC. We screened 70 index patients suggestive of a genetic predisposition for germ-line mutations in hMLH3 with denaturing high-performance liquid chromatography. One frameshift mutation and 11 missense mutations were identified in 16 index patients (23%). Most families presented evidence against hMLH3 as a high risk factor in familial colorectal cancer, and most of the mutations were found in the low risk patients, suggesting hMLH3 to be a low risk gene for colorectal cancer. We demonstrate in one family that a hMLH3 mutation segregated with disease together with a missense mutation in hMSH2, which makes us hypothesize that these mutations work together in an additive manner and result in an elevated risk of colorectal tumors in the family. None of the tumors with hMLH3 mutations showed microsatellite instability, which demonstrates that hMLH3 does not make its contribution to carcinogenesis through an impaired DNA mismatch repair function.

Mutations in p53 is a most common genetic alteration in human cancer and has been related to outcome, also in colorectal cancer. However, published results are not unanimous on this point without clear-cut explanations. Different analytical methods have been applied which could explain some discrepancies. Another factor of importance may be the source of nucleic acids used for identification of sequence alterations. Therefore we compared mutations in exons 7 and 8 of p53 found in genomic DNA, with mutations in cDNA from the same patients and evaluated to what extent LOH and mutations in p53 occurred simultaneously, which are prerequisites for a complete loss of p53 function according to the classic concept. cDNA and genomic DNA from tumor tissue from 123 patients were used. Thirty-four mutations were found in both tumor cDNA and genomic DNA. Twenty missense mutations were the same in both cDNA and genomic DNA. Two missense mutations, one 1 bp deletion and one nonsense mutation were only found in genomic DNA, while nine missense mutations and one 9 bp deletion were found in cDNA only. Statistical analysis showed no significant difference among mutations in cDNA and genomic DNA (p=1.00). Only 2 patients (5%) had both LOH and mutations in exons 2-11 of p53. One patient had LOH without p53 mutation, while 18 patients (44%) showed p53 mutation without LOH and 20 patients (49%) had wt p53 without LOH. Forty-four patients (52%) were non-informative for LOH of the entire cohort consisting of 85 patients. Our findings suggest that different results reported on the role mutated p53 may have in colorectal cancer are probably not explained by the source of nucleic acids (RNA vs DNA) for sequence determinations. Rather, unknown information about simultaneous but different presence of p53 LOH in previously published reports is more likely. However, an unexpectedly infrequent occurrence (5-10%) of complete and isolated p53 ablation (LOH + mutation) would demand considerably larger patient populations (1500-2000 patients) than ever published for relating p53 function to survival in patients with colorectal cancer. Therefore, the role of p53 dysfunction in progression of colorectal cancer remains uncertain.