Prognostic factors in colorectal cancer : studies of thymidylate synthase expression, mismatch repair protein expression, tumor budding and t-cell infiltration

Abstract

Colorectal cancer (CRC) is a major health problem in the Western world. CRC is treatable with surgery and often curable when the disease is diagnosed at an early stage. With improved surgery and adjuvant chemotherapy treatment, the disease-free-survival (DFS) rate for colon cancer is approximately 80% for stage II and for stage III close to 65%. There is a continual need for the established risk stratification of CRC based on the tumor, node, metastasis (TNM) staging to evolve. The overall aim of this thesis was to detect new potential prognostic and predictive factors for molecular and clinical responses in primary CRC. One specific aim was to establish whether thymidylate synthase (TS) expression in a large group of patients with stage II and stage III CRC is a prognostic and/or predictive factor alone or in combination with mismatch repair (MMR) status in the colon cancer subgroup. Another aim was to explore in primary CRC the potential association between tumor budding and MMR status and the impact of tumor budding as a factor of tumor recurrence and development of distant metastases. The last aim was to investigate the prognostic and predictive value of tumor budding, tumor border configuration and T-cell infiltration in primary colon cancer with known MMR status. Tumor material for the studies was derived from the adjuvant Nordic trials which randomized from 1991 to 1997, 2224 patients with stage II or stage III CRC to surgery alone or surgery plus adjuvant 5-FU-based chemotherapy. Immunohistochemistry (IHC) was used to detect and evaluate TS expression, MMR-status, tumor budding and infiltration of CD3+ and CD8+ T-cells.

Paper I. TS expression was assessed in tumors from 1,389 patients with stage II and III CRC. TS expression with the classification of TS grade 0-1 versus 2-3 as well as TS 1-2 versus TS 3 was prognostic in the surgery alone group. A high TS expression was associated with a shorter overall survival (OS). Among patients with TS grade 3, the subgroup treated with adjuvant chemotherapy had a significant longer OS compared with the group treated with surgery alone. Paper II. Primary colon cancer with a deficient MMR status (dMMR) is associated with an improved prognosis. Data indicates that colon cancer with a proficient MMR status (pMMR) and with high TS expression has an improved response to adjuvant 5-FU-based chemotherapy. This study evaluated if a combined analysis of MMR status and TS expression in 716 colon cancer patients added prognostic value and better predicted response to adjuvant 5-FU-based chemotherapy. In the group of patients treated with surgery alone, patients with dMMR tumors and low TS grade had a trend to a better OS compared to patients with pMMR tumors and high TS grade. In stage III, patients with pMMR tumors and high TS grade who received adjuvant 5-FU-based chemotherapy had a significantly improved OS compared to patients treated with surgery alone. No relationship was found between MMR status and TS expression. Paper III. A pMMR status compared to a dMMR status and tumor budding are considered adverse prognostic factors in primary CRC. This exploratory study included 134 patients with primary CRC. It assessed tumor budding grade in tumors with pMMR status compared to dMMR status to see if it differed as to whether local recurrence or metastases did or did not develop. The 29 available dMMR cases which developed recurrence or distant metastases (met+) were matched with a dMMR group with no recurrence or metastases (met-), and the pMMR/met+ group with pMMR/met- cases. A significantly higher percentage of high-grade tumor budding was only found in the dMMR/met+ group compared to the pMMR/met+ group. Paper IV. Tumor budding is correlated to the development of local and distant metastases in CRC while high density of tumor infiltrating immune cells is associated with an improved prognosis. In this study, tumors from 478 patients with stage II-III colon cancer and known MMR status were examined to determine the prognostic value of tumor budding, tumor border configuration and CD3+ / CD8+ T-cell infiltration. An association was found between high grade tumor budding and more advanced stage, higher N-stage, pMMR status, an infiltrating tumor border configuration and lower levels of high score CD3+ and CD8+ T-cells. High grade tumor budding was correlated to worse OS in univariate analysis but not in multivariate. In the entire study population, an infiltrative tumor border configuration was an adverse prognostic factor for OS and a dense infiltration of CD8+ T-cells was independently associated with a better OS.

In conclusion, TS is a prognostic factor in CRC patients treated with surgery alone and patients with the highest level of TS expression had an improved clinical outcome following adjuvant 5-FU-based chemotherapy. In stage III colon cancer, a combined rather than a single marker analysis of MMR-status and TS expression, can improve the prediction of response to 5-FU-based chemotherapy. Whether tumor budding can provide prognostic information for patients with primary CRC and a dMMR status should be further explored in larger studies. An independent prognostic impact was found for CD8+ T cell infiltration and tumor border configuration as well as an association between them and tumor budding. Our study supports the inclusion of tumor border configuration, tumor budding, CD8+ T cell infiltration in the risk assessment for stage II-III colon cancer patients.