The intra-tumor stroma percentage in colon cancer (CC) patients has previously been reported by our group as a strong independent prognostic parameter. Patients with a high stroma percentage within the primary tumor have a poor prognosis. Tissue samples from the most invasive part of the primary tumor of 710 patients (52% Stage II, 48% Stage III) participating in the VICTOR trial were analyzed for their tumor-stroma percentage. Stroma-high (>50%) and stroma-low (≤50%) groups were evaluated with respect to survival times. Overall and disease-free survival times (OS and DFS) were significantly lower in the stroma-high group (OS P < 0.0001, hazard ratio (HR) = 1.96; DFS P < 0.0001, HR = 2.15). The 5-year OS was 69.0% versus 83.4% and DFS 58.6% versus 77.3% for stroma-high versus stroma-low patients. This study confirms the intra-tumor stroma ratio as a prognostic factor. This parameter could be a valuable and low cost addition to the TNM status and next to current high-risk parameters such as microsatellite instability status used in routine pathology reporting. When adding the stroma-parameter to the ASCO criteria, the rate of ‘undertreated’ patients dropped from 5.9% to 4.3%, the ‘overtreated’ increased with 6.8% but the correctly classified increased with an additional 14%.

Background: Tumor staging insufficiently discriminates between colon cancer patients with poor and better prognosis. We have evaluated, for the primary tumor, if the carcinoma‐percentage (CP), as a derivative from the carcinoma‐stromal ratio, can be applied as a candidate marker to identify patients for adjuvant therapy. Methods: In a retrospective study of 63 patients with colon cancer (stage I–III, 1990–2001) the carcinoma‐percentage of the primary tumor was estimated on routine H&E stained histological sections. Additionally these findings were validated in a second independent study of 59 patients (stage I–III, 1980–1992). (None of the patients had received preoperative chemo‐ or radiation therapy nor adjuvant chemotherapy.) Results: Of 122 analyzed patients 33 (27.0%) had a low CP and 89 (73.0%) a high CP. The analysis of mean survival revealed: overall‐survival (OS) 2.13 years, disease‐free‐ survival (DFS) 1.51 years for CP‐low and OS 7.36 years, DFS 6.89 years for CP‐high. Five‐year survival rates for CP‐low versus CP‐high were respectively for OS: 15.2% and 73.0% and for DFS: 12.1% and 67.4%. High levels of significance were found (OS p < 0.0001, DFS p < 0.0001) with hazard ratio’s of 3.73 and 4.18. In a multivariate Cox regression analysis, CP remained an independent variable when adjusted for either stage or for tumor status and lymph‐node status (OSp < 0.001, OSp < 0.001). Conclusions: The carcinoma‐percentage in primary colon cancer is a factor to discriminate between patients with a poor and a better outcome of disease. This parameter is already available upon routine histological investigation and can, in addition to the TNM classification, be a candidate marker to further stratify into more individual risk groups.

The tumor-stroma ratio (TSR) has been reported as a strong, independent prognostic parameter in colon cancer as well as in other epithelial cancer types, and may be implemented to routine pathology diagnostics. The TSR is an easy technique, based on routine hematoxylin and eosin stained histological sections, estimating the amount of stroma present in the primary tumor. It links tumors with high stromal content to poor prognosis. The analysis time is less than 2 min with a low inter-observer variation. Scoring of the TSR has been validated in a number of independent international studies. In this manuscript, we provide a detailed technical description of estimating the TSR in colon cancer, including examples, pitfalls, and recommendations.

The tumor microenvironment has a critical role in regulating cancer cell behavior. Tumors with high stromal content are associated with poor patient outcome. The tumor-stroma ratio (TSR) identifies colorectal cancers (CRC) with poor patient prognosis based on hematoxylin & eosin stained sections. The desmoplastic reaction consists to a great extent of cancer-associated fibroblasts (CAFs) of which different subtypes are known. The aim of this study is to investigate and quantify CAFs present in the tumor stroma of CRC stratified by the TSR to possibly add prognostic significance to the TSR. The expression of established CAF markers was compared between stroma-low and stroma-high tumors using transcriptomic data of 71 stage I – III CRC. Based on literature, fibroblast and stromal markers were selected to perform multiplex immunofluorescent staining on formalin fixed, paraffin-embedded tumor sections of patients diagnosed with stage III colon cancer. Antibodies against the following markers were used: αSMA, PDGFR -β, FAP, FSP1 and the stromal markers CD45 and CD31 as reference. The markers were subsequently quantified in the stroma using the Vectra imaging microscope. The transcriptomic data showed that all CAF markers except one were higher expressed in stroma-high compared to stroma-low tumors. Histologically, stroma-high tumors showed a decreased number of FSP1+/CD45+ cells and a trend of an increased expression of FAP compared to stroma-low tumors. FAP was higher expressed at the invasive part compared to the tumor center in both stroma-high and stroma-low tumors. The increased expression of FAP at the invasive part and in stroma-high tumors might contribute to the invasive behavior of cancer cells. Future functional experiments should investigate the contribution of FAP to cancer cell invasion. Combining the quantity of the stroma as defined by the TSR with the activity level of CAFs using the expression of FAP may result in an expanded stroma-based tool for patient stratification.