Rob A. E. M. Tollenaar

Micrometastases and Survival in Stage II Colorectal Cancer

BACKGROUND Standard treatment of colorectal cancer includes adjuvant chemotherapy for patients with stage III disease (defined by the presence of lymph-node metastases), but not for patients with stage II tumors (who have no lymph-node metastases). However, 20 percent of patients with stage II tumors die of recurrent disease. We investigated whether the detection of micrometastases can be used to identify patients with stage II disease who are at high risk for recurrence. METHODS We analyzed 192 lymph nodes from 26 consecutive patients with stage II colorectal cancer, using a carcinoembryonic antigen-specific nested reverse-transcriptase polymerase chain reaction. Five-year follow-up information was obtained on all patients. Observed and adjusted survival rates were assessed in the patients with and the patients without micrometastases. RESULTS Micrometastases were detected in one or more lymph nodes from 14 of 26 patients (54 percent). The adjusted five-year survival rate (for which only cancer-related deaths were considered) was 50 percent in this group, whereas in the 12 patients without micrometastases, the survival rate was 91 percent (P=0.02 by the log-rank test). The observed five-year survival rates were 36 percent and 75 percent, respectively (P=0.03). The groups were similar with respect to age, sex, tumor side (location in relation to the flexura lienalis), degree of tumor differentiation (grade), and diameter of the primary tumor. CONCLUSIONS Molecular detection of micrometastases is a prognostic tool in stage II colorectal cancer.

Risk factors for anastomotic failure after total mesorectal excision of rectal cancer

Anastomotic leakage is a major complication of rectal cancer surgery. The aim of this study was to investigate risk factors associated with symptomatic anastomotic leakage after total mesorectal excision (TME).

Regions of focal DNA hypermethylation and long-range hypomethylation in colorectal cancer coincide with nuclear lamina-associated domains

Extensive changes in DNA methylation are common in cancer and may contribute to oncogenesis through transcriptional silencing of tumor-suppressor genes. Genome-scale studies have yielded important insights into these changes but have focused on CpG islands or gene promoters. We used whole-genome bisulfite sequencing (bisulfite-seq) to comprehensively profile a primary human colorectal tumor and adjacent normal colon tissue at single-basepair resolution. Regions of focal hypermethylation in the tumor were located primarily at CpG islands and were concentrated within regions of long-range (>100 kb) hypomethylation. These hypomethylated domains covered nearly half of the genome and coincided with late replication and attachment to the nuclear lamina in human cell lines. We confirmed the confluence of hypermethylation and hypomethylation within these domains in 25 diverse colorectal tumors and matched adjacent tissue. We propose that widespread DNA methylation changes in cancer are linked to silencing programs orchestrated by the three-dimensional organization of chromatin within the nucleus.

Gene Expression Signature to Improve Prognosis Prediction of Stage II and III Colorectal Cancer

PURPOSE This study aims to develop a robust gene expression classifier that can predict disease relapse in patients with early-stage colorectal cancer (CRC). PATIENTS AND METHODS Fresh frozen tumor tissue from 188 patients with stage I to IV CRC undergoing surgery was analyzed using Agilent 44K oligonucleotide arrays. Median follow-up time was 65.1 months, and the majority of patients (83.6%) did not receive adjuvant chemotherapy. A nearest mean classifier was developed using a cross-validation procedure to score all genes for their association with 5-year distant metastasis-free survival. RESULTS An optimal set of 18 genes was identified and used to construct a prognostic classifier (ColoPrint). The signature was validated on an independent set of 206 samples from patients with stage I, II, and III CRC. The signature classified 60% of patients as low risk and 40% as high risk. Five-year relapse-free survival rates were 87.6% (95% CI, 81.5% to 93.7%) and 67.2% (95% CI, 55.4% to 79.0%) for low- and high-risk patients, respectively, with a hazard ratio (HR) of 2.5 (95% CI, 1.33 to 4.73; P = .005). In multivariate analysis, the signature remained one of the most significant prognostic factors, with an HR of 2.69 (95% CI, 1.41 to 5.14; P = .003). In patients with stage II CRC, the signature had an HR of 3.34 (P = .017) and was superior to American Society of Clinical Oncology criteria in assessing the risk of cancer recurrence without prescreening for microsatellite instability (MSI). CONCLUSION ColoPrint significantly improves the prognostic accuracy of pathologic factors and MSI in patients with stage II and III CRC and facilitates the identification of patients with stage II disease who may be safely managed without chemotherapy.

Immune system and prognosis in colorectal cancer: a detailed immunohistochemical analysis

The efficacy of tumor cell–immune cell interactions depends on a number of factors, for example, the expression of HLA-I on tumor cells, the type of immune cell, the accessibility of tumor cells for immune cells and the expression of immunogenic epitopes. We assessed infiltration of CD4+, CD8+, CD56+ and CD57+ cells in the tumor epithelium, tumor stroma and advancing tumor margin of 93 colorectal carcinomas and correlated this to clinicopathological parameters, the expression of HLA-A and HLA-B/C on tumor cells, the presence of a basal membrane (BM)-like structure surrounding tumor nodules and the presence of microsatellite instability/mutator phenotype (absent MLH-1 expression). The median intraepithelial CD4+, CD8+, CD56+ and CD57+ cell infiltrations were 3, 23, 0 and 0 cells/mm2 tumor, respectively. HLA-A/BC expression by tumor cells was normal in 28/43%, heterogeneous in 59/48% and absent in 13/9% of the cases. A BM-like structure surrounding the tumor nodules was absent, present and thick in 47, 38 and 15% of the cases. Six cases lost MLH1 expression. There was a positive correlation between leukocyte infiltration in the three compartments for CD4+, CD8+, CD56+ (partly) and CD57+ (all P<0.05) cell infiltration. Intraepithelial CD8+ cell infiltration inversely correlated with HLA-A (P=0.04) and HLA-B/C expression (P=0.04). Intraepithelial CD57+ cell infiltration inversely correlated with HLA-B/C expression (P=0.04). Moreover, intraepithelial infiltration of CD8+ and CD57+ cells was inversely correlated to the presence of a BM-like structure (P=0.003 and 0.04, respectively). Uni- and multivariate analyses showed that a lower tumor stage (P=0.004) and marked infiltration of CD8+ (P=0.04) and CD57+ cells (P=0.05) at the advancing tumor margin were independent prognostic factors for a longer disease-free survival. Loss of MLH1 expression was correlated with a significantly higher intraepithelial CD8+ and CD57+ cell infiltration. We conclude that infiltration of CD8+ and CD57+ cells are important prognostic factors in colorectal cancer. However, their interaction with tumor cells is inversely correlated to the presence of HLA-I on tumor cells and a thick BM-like structure around tumor islets. Our data indicate that NK cells might play an important role in the immune surveillance in colorectal cancer patients.

Systematic review and meta‐analysis of the volume–outcome relationship in pancreatic surgery

Many studies have shown lower mortality and higher survival rates after pancreatic surgery with high‐volume providers, suggesting that centralization of pancreatic surgery can improve outcomes. The methodological quality of these studies is open to question. This study involves a systematic review of the volume–outcome relationship for pancreatic surgery with a meta‐analysis of studies considered to be of good quality.

Mechanisms of oncogenesis in colon versus rectal cancer.

Observations support the theory that development of left‐ and right‐sided colorectal cancers may involve different mechanisms. This study investigated different genes involved in oncogenesis of colon and rectal cancers and analysed their prognostic value. The study group comprised 35 colon and 42 rectal cancers. Rectal cancer patients had been treated with standardized surgery performed by an experienced rectal cancer surgeon. Mutation analysis was performed for p53 in eight colon cancers and for APC and p53 in 22 rectal cancers. MLH1, MSH2, Bcl‐2, p53, E‐cadherin and β‐catenin were investigated by immunohistochemistry in all colorectal tumours. APC mutation analysis of the MCR showed truncating mutations in 18 of 22 rectal tumours (82%), but the presence of an APC mutation was not related to nuclear β‐catenin expression (p=0.75). Rectal cancers showed significantly more nuclear β‐catenin than colon cancers (65% versus 40%, p=0.04). p53 mutation analysis corresponded well with p53 immunohistochemistry (p<0.001). Rectal cancers showed significantly more immunohistochemical expression of p53 than colon cancers (64% versus 29%, p=0.003). In rectal cancers, a significant correlation was found between positive p53 expression and worse disease‐free survival (p=0.008), but not in colon cancers. Cox regression showed that p53‐expression (p=0.03) was an independent predictor for disease‐free survival in rectal cancers. This study concluded that rectal cancer may involve more nuclear β‐catenin in the APC/β‐catenin pathway than colon cancer and/or nuclear β‐catenin may have another role in rectal cancer independently of APC. The p53‐pathway seems to be more important in rectal cancer, in which it also has independent prognostic value. When prognostic markers are investigated in larger series, differences in biological behaviour between colon and rectal cancer should be considered. Copyright

EMT is the dominant program in human colon cancer

BackgroundColon cancer has been classically described by clinicopathologic features that permit the prediction of outcome only after surgical resection and staging.MethodsWe performed an unsupervised analysis of microarray data from 326 colon cancers to identify the first principal component (PC1) of the most variable set of genes. PC1 deciphered two primary, intrinsic molecular subtypes of colon cancer that predicted disease progression and recurrence.ResultsHere we report that the most dominant pattern of intrinsic gene expression in colon cancer (PC1) was tightly correlated (Pearson R = 0.92, P < 10-135) with the EMT signature-- both in gene identity and directionality. In a global micro-RNA screen, we further identified the most anti-correlated microRNA with PC1 as MiR200, known to regulate EMT.ConclusionsThese data demonstrate that the biology underpinning the native, molecular classification of human colon cancer--previously thought to be highly heterogeneous-- was clarified through the lens of comprehensive transcriptome analysis.

BRCA1-Associated Breast Cancers Present Differently From BRCA2-Associated and Familial Cases: Long-Term Follow-Up of the Dutch MRISC Screening Study

PURPOSE The Dutch MRI Screening Study on early detection of hereditary breast cancer started in 1999. We evaluated the long-term results including separate analyses of BRCA1 and BRCA2 mutation carriers and first results on survival. PATIENTS AND METHODS Women with higher than 15% cumulative lifetime risk (CLTR) of breast cancer were screened with biannual clinical breast examination and annual mammography and magnetic resonance imaging (MRI). Participants were divided into subgroups: carriers of a gene mutation (50% to 85% CLTR) and two familial groups with high (30% to 50% CLTR) or moderate risk (15% to 30% CLTR). RESULTS Our update contains 2,157 eligible women including 599 mutation carriers (median follow-up of 4.9 years from entry) with 97 primary breast cancers detected (median follow-up of 5.0 years from diagnosis). MRI sensitivity was superior to that of mammography for invasive cancer (77.4% v 35.5%; P<.00005), but not for ductal carcinoma in situ. Results in the BRCA1 group were worse compared to the BRCA2, the high-, and the moderate-risk groups, respectively, for mammography sensitivity (25.0% v 61.5%, 45.5%, 46.7%), tumor size at diagnosis≤1 cm (21.4% v 61.5%, 40.9%, 63.6%), proportion of DCIS (6.5% v 18.8%, 14.8%, 31.3%) and interval cancers (32.3% v 6.3%, 3.7%, 6.3%), and age at diagnosis younger than 30 years (9.7% v 0%). Cumulative distant metastasis-free and overall survival at 6 years in all 42 BRCA1/2 mutation carriers with invasive breast cancer were 83.9% (95% CI, 64.1% to 93.3%) and 92.7% (95% CI, 79.0% to 97.6%), respectively, and 100% in the familial groups (n=43). CONCLUSION Screening results were somewhat worse in BRCA1 mutation carriers, but 6-year survival was high in all risk groups.

Prognostic Value of Tumor-Infiltrating Dendritic Cells in Colorectal Cancer: Role of Maturation Status and Intratumoral Localization

The clinical significance of tumor-infiltrating dendritic cells has been reported in a variety of human solid tumors as shown by the correlations found between the presence of tumor-infiltrating dendritic cells and clinical prognosis. In this study, we evaluated whether there is an association between the presence and maturation status of tumor-infiltrating dendritic cells, T lymphocytes, and clinical course in 104 primary tumor samples of patients with colorectal cancer. Dendritic cells were identified with four different markers (S-100, HLA class II, CD208, and CD1a) in double immunohistochemistry, with laminin as second marker to support the exact localization. Tumor-infiltrating dendritic cells showed a distinct infiltration pattern based on their maturation status. CD1a-positive dendritic cells resided in the advancing tumor margins in relatively high numbers, whereas mature CD208-positive dendritic cells were sparsely present in the tumor epithelium but mainly distributed in the tumor stroma and advancing tumor margin. Furthermore, high infiltration of CD1a-positive dendritic cells in the tumor epithelium was significantly correlated to the infiltration of CD4 lymphocytes (P = 0.006). Patients with relatively high numbers of mature CD208-positive infiltrating dendritic cells in the tumor epithelium had a shorter overall survival (P = 0.004). In addition, patients with relatively high numbers of CD1a-positive dendritic cells in the advancing margin of the tumor had a shorter disease-free survival (P = 0.03). We found that tumor-infiltrating dendritic cells had preferential infiltration sites within a tumor, affected local tumor cell-immune cell interactions, and correlated to the clinical prognosis of colorectal cancer patients.