Sanne Kjær-Frifeldt

The prognostic importance of miR-21 in stage II colon cancer: a population-based study

Background:Despite several years of research and attempts to develop prognostic models a considerable fraction of stage II colon cancer patients will experience relapse within few years from their operation. The aim of the present study was to investigate the prognostic importance of miRNA-21 (miR-21), quantified by in situ hybridisation, in a unique, large population-based cohort.Patients and methods:The study included 764 patients diagnosed with stage II colon cancer in Denmark in the year 2003. One section from a representative paraffin-embedded tumour tissue specimen from each patient was processed for analysis of miR-21 and quantitatively assessed by image analysis.Results:The miR-21 signal was predominantly observed in fibroblast-like cells located in the stromal compartment of the tumours. We found that patients expressing high levels of miR-21 had significantly inferior recurrence-free cancer-specific survival (RF-CSS): HR=1.26; 95% CI: 1.15–1.60; P<0.001. In Cox regression analysis, a high level of miR-21 retained its prognostic importance and was found to be significantly related to poor RF-CSS: HR=1.41; 95% CI: 1.19–1.67; P<0.001.Conclusion:The present study showed that increasing miR-21 expression levels were significantly correlated to decreasing RF-CSS. Further investigations of the clinical importance of miR-21 in the selection of high-risk stage II colon cancer patients are merited.

Prognostic importance of VEGF-A haplotype combinations in a stage II colon cancer population

AIM To investigate the prognostic effect of three VEGF-A SNPs, -2578, -460 and 405, as well as the corresponding haplotype combinations, in a unique population of stage II colon cancer patients. PATIENTS & METHODS The study included all patients diagnosed with stage II colon cancer in Denmark in 2003 (698 patients). One paraffin-embedded tissue block from each patient was used for DNA extraction and analysis of the three VEGF SNPs. RESULTS The homozygous genotype VEGF -2578 AA had significant effect on time to tumor recurrence (hazard ratio [HR] = 2.01 [95% CI: 1.13-3.56]; p = 0.02) as well as -460TT (HR = 0.50 [95% CI: 0.29-0.89]; p = 0.02). Patients harboring the haplotype combinations ACG,CTC and ACG,ACG displayed a significantly shorter time to tumor recurrence in both univariate (HR = 1.87 [95% CI: 1.21-2.89]; p = 0.008) and multivariate analysis (HR = 1.76 [95% CI: 1.09-2.82]; p = 0.02). CONCLUSION We found that the gene polymorphism in VEGF-A holds prognostic information and should be considered as a potential adjunct in identification of high-risk stage II colon cancer patients.

Redefining high-risk patients with stage II colon cancer by risk index and microRNA-21: results from a population-based cohort

Background:The aim of the present study was to analyse the prognostic value of microRNA-21 (miRNA-21) in patients with stage II colon cancer aiming at a risk index for this group of patients.Methods:A population-based cohort of 554 patients was included. MicroRNA-21 was analysed by qPCR based on tumour tissue. An index was created using the coefficients obtained from a collective multiple Cox regression. The entire procedure was cross-validated (10-fold). The performance of the index was quantified by time-dependent receiver operating characteristics curves.Results:High miRNA-21 expression was associated with an unfavourable recurrence-free cancer-specific survival (RF-CSS), hazard ratio 1.35 (95% confidence interval, 1.03–1.76) (P=0.028). The generated RF-CSS index divided the traditional high-risk patients into subgroups with 5-year RF-CSS rates of 87% and 73%, respectively (P<0.001). The overall survival (OS) index identified three different subgroups (P<0.001). Cross-validated 5-year OS rates were 88%, 68%, and 50%, respectively.Conclusions:This population-based study supports miRNA-21 as an additional prognostic biomarker in patients with stage II colon cancer. Furthermore, the introduction of a risk index may guide the use of postoperative adjuvant treatment in a more appropriate way compared with current practice.

The prognostic value of microRNA-126 and microvessel density in patients with stage II colon cancer: results from a population cohort

BackgroundAngiogenesis plays a pivotal role in malignant tumour growth and the metastatic process. We analysed the prognostic value of two angiogenesis parameters, microRNA-126 (miRNA-126) and microvessel density (MVD), in a population based cohort of patients operated for stage II colon cancer.MethodsA total of 560 patients were included. Analyses were performed on formalin fixed paraffin embedded tissue from the primary tumours. The analysis of miRNA-126 expression was performed by qPCR. Microvessels were visualised by CD105 and quantified in hot spots using a light microscope. The analyses were correlated with recurrence-free cancer specific survival (RF-CSS) and overall survival (OS).ResultsLow miRNA-126 expression was significantly correlated to T4, high malignancy grade, tumour perforation, fixation, and the presence of microsatellite instability. A prognostic impact on OS was detected in the simple analysis favouring patients with high miRNA-126 expression p = 0.03, and borderline significance as to RF-CSS, p = 0.08. The impact on OS demonstrated borderline significance in a following multiple Cox regression analysis, hazard ratio 0.76 (95% confidence interval, 0.58-1.00), p = 0.051. The MVD estimate was not associated with either RF-CSS, p = 0.49, or OS, p = 0.94.ConclusionThe current population based study of patients operated for stage II colon cancer demonstrated correlations between several prognostic unfavourable characteristics and miRNA-126 and argues for a possible prognostic impact on overall survival. An influence on survival by the MVD estimate was not detected.

Stroma-High Lymph Node Involvement Predicts Poor Survival More Accurately for Patients with Stage III Colon Cancer

Objective: The tumor microenvironment has ample impact on the behavior of the malignant process in colon cancer (CC). Patients with a high percentage of stroma within the primary tumor, determined by the tumor-stroma ratio (TSR), have a poor prognosis. In metastatic lymph nodes from patients with stage III CC, the TSR is heterogeneous, but the impact on patients’ prognosis is unknown. Methods: Haematoxylin and eosin stained tissue slides of primary tumor (PT) and associated lymph nodes (LNs) metastases from 102 patients with stage III CC were analyzed for the TSR. Stroma-high (>50% stroma) and stromalow (≤ 50% stroma) groups were evaluated with respect to disease free survival (DFS). Results: Of 102 analyzed primary tumors, 47 (46.1%) scored as stroma-high and 55 (53.9%) as stroma-low. In total, 33 patients had at least one stroma-high LN and 69 patients had one or more stroma-low LNs. Interestingly, 28 patients (27.5%) had both stroma-high and stroma-low LNs, but in another 44 cases the TSR between PT and LNs differed: 29 patients had a stroma-high PT with stroma-low LNs, while 15 patients displayed the opposite. As a result of the combination of the TSR analysis of the PT and the involved metastatic LNs, 62 patients (60.8%) were classified as stroma-high and 40 (39.2%) as stroma-low, restaging 14.7% of the patients to stroma-high with a significantly worse 5-year DFS compared to stroma-low patients (59% vs. 82%, HR=2.83 (95%CI 1.34–5.97), P=0.006). In multivariate analysis, the TSR retained its independent prognostic impact (HR=2.85 (95%CI 1.33-6.10), P=0.007). Conclusion: The presence of abundant stroma in metastatic LNs from patients with stage III CC adds to the prognostic information learned from the primary tumor independently, and supports selective patient treatment.

Tumor–stroma ratio predicts recurrence in patients with colon cancer treated with neoadjuvant chemotherapy

Abstract Background: Neoadjuvant chemotherapy represents a new treatment approach to locally advanced colon cancer. The aim of this study was to analyze the ability of tumor–stroma ratio (TSR) to predict disease recurrence in patients with locally advanced colon cancer treated with neoadjuvant chemotherapy. Material and methods: This study included 65 patients with colon cancer treated with neoadjuvant chemotherapy in a phase II trial. All patients were planned for three cycles of capecitabine and oxaliplatin before surgery. Hematoxylin and eosin stained tissue sections from surgically resected primary tumors were sampled and analyzed by conventional microscopy. Patients were divided into stroma-high (>50%, i.e. TSR low) and stroma-low (≤50%, i.e. TSR high) for the comparison with clinical data. Results: A low TSR was found in 47% of the surgically resected primary tumors and correlated to a significantly higher T- and N-category compared, to tumors with a high TSR (p < .01). A low TSR was also significantly associated with disease recurrence (p = .008), translating into significant differences in disease free survival (DFS) and overall survival, p < .002. The 5-year DFS rate for patients with a low TSR was 55%, compared to 94% in the group of patients with a high TSR. Conclusions: TSR assessed in the surgically resected primary tumor from patients with locally advanced colon cancer treated with neoadjuvant chemotherapy provides prognostic value and may serve as a relevant parameter in selecting patients for post-operative treatment.

The Prognostic Value of Tumor-Infiltrating lymphocytes in Stage II Colon Cancer. A Nationwide Population-Based Study

BACKGROUND: Additional prognostic markers are needed for better treatment stratification of stage II colon cancer (CC). We investigated the prognostic value of tumor-infiltrating lymphocytes (TILs) in a true population-based cohort of patients with stage II CC. MATERIAL AND METHODS: A total of 573 patients were included. Tumor blocks representing the deepest invasive part of the primary tumor were used for analysis. CD3+ and CD8+ TILs at the invasive front were evaluated by immunohistochemistry on whole tumor sections. The invasive area was manually outlined, and Visiopharm Integrator System software was used for quantification. Data were dichotomized for comparison with clinical data. The prognostic value was investigated in Cox proportional-hazard models for recurrence-free survival (RFS) and overall survival (OS). RESULTS: Low CD3+ or CD8+ TILs were significantly associated with poor RFS and OS (P = .0021 and P ≤ .0009, respectively, log-rank test). In multiple Cox regression analysis, low CD3+ and CD8+ TILs were associated with reduced RFS with hazard ratio (HR) = 1.386 (95% CI 1.039-1.850), P = .026, and HR = 1.394 (95% CI 1.029-1.890), P = .032, respectively, independent of age, T-stage, localization, perforation, and microsatellite instability (MSI). In the subgroups of patients with low CD3+ or CD8+ TILs, there was no difference in survival between patients with MSI and microsatellite-stable tumors, (P = .821 and P = .907, respectively). CONCLUSION: Low CD3+ and CD8+ TILs in the invasive area are both related to inferior prognosis of stage II CC, and we recommend either of these parameters to be considered as additional high-risk factor.

Limitations of tissue micro array in Duke's B colon cancer

Tissue micro array (TMA) is widely used in cancer research in search of new predictive and prognostic markers. Colon cancer is known to be heterogeneous and the present study addresses some methodological aspects using cores of different size and analysing markers with different cellular distribution. We selected 61 paraffin‐embedded tissue blocks representing patients diagnosed with Dukes B colon cancer. Two 1 mm and two 2 mm cores were taken from both the centre and the invasive front of the tumour respectively. The immunostaining included MLH1, MSH2, PMS2, p53, COX‐2, TIMP and Betacatenin. Twenty‐five percent of the cores taken from paraffin blocks less than 0.5 cm was lost and the total loss was 8%. The homogeneous stains (MLH1, MSH2 and PMS2) all showed high agreement between TMA and whole tissue stains (kappa = 0.96,1 and 1 respectively). The COX‐2, p53 and Betacatenin illustrated moderate to high agreement (kappa = 0.54–0.9) whereas TIMP‐1 had the lowest score (kappa 0.19–0.25). The application of TMA in Dukes B colon cancer has several pitfalls and depends substantially on the immunohistochemical marker in question. Therefore a validation study seems justified before applying large scale TMA in this setting.

The prognostic value of tumour stroma ratio and tumour budding in stage II colon cancer. A nationwide population-based study

PurposeHigh-risk patients with stage II colon cancer (CC) may benefit from adjuvant chemotherapy, but additional prognostic markers are needed for better stratification. We investigated the prognostic value of tumour stroma ratio (TSR) and tumour budding (TB).MethodsA nationwide population-based cohort of 573 patients with stage II CC was included. TSR was scored on hematoxylin and eosin sections as low TSR (> 50% stroma) and high TSR (≤ 50% stroma). TB was evaluated in hotspots on pan-cytokeratin stained sections in 10 high power fields (HPF) at the invasive front and classified by the mean number of buds per HPF as high grade budding (≥ 10 buds) or low-grade budding (< 10 buds). The prognostic value was investigated in Cox proportional hazard models for recurrence-free survival (RFS) and overall survival (OS).ResultsLow TSR was associated with worse RFS (HR = 1.342 (95% CI 1.006–1.791), p = 0.045) and OS (HR = 1.376 (95% CI 1.016–1.862), p = 0.039). Furthermore, an association was found between low TSR and microsatellite stabile tumours (p < 0.001). The mean number of buds per HPF was associated to TSR with increasing number of buds related to a lower TSR (p = 0.026). No statistically significant prognostic impact of TB regarding OS or RFS was detected.ConclusionsTSR provided valuable prognostic information, and adding TSR to the current risk stratification may contribute to better patient selection. The estimates of TSR and TB were found to be associated, but no prognostic value of TB was documented.