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 in104 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 CD4lympho-cytes (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.

Dendritic cells (DCs) are the most efficient antigen-presenting cells and play a key role in a cellular antitumor immune response. In this study we investigated the exact localization of DCs within colorectal tumors and their relationship to tumor-infiltrating lymphocytes as well as clinical outcome of the patients. Primary tumor specimens of 104 patients with a diagnosis of colorectal cancer were identified retrospectively and analyzed with the dendritic cell markers S-100 protein and human leukocyte antigens (HLA) class II. The markers were individually combined with laminin as a second marker to facilitate the observation of the different tumor localizations. S-100 or HLA class II positive cells were found in the three different compartments of colorectal tumors: tumor epithelium, tumor stroma, and advancing tumor margin, but mainly present in tumor stroma and advancing tumor margin. S-100-positive tumor-infiltrating DCs in direct contact with tumor cells, i.e., in tumor epithelium, significantly correlated to the intraepithelial infiltration of CD4+ (p=0.02) and CD8+ (p=0.01) lymphocytes. High HLA class II+ cell infiltration in the tumor stroma correlated to a lower intraepithelial infiltration of CD8+ (p=0.02) lymphocytes. High intraepithelial infiltration of S-100-positive DCs suggested increased disease-free survival, but was not statistically significant, while high amounts of HLA class II+ cells in the tumor stroma correlated with an adverse survival outcome. Our results show that the infiltration of DCs in colorectal cancer, depending on both location and type of marker, is correlated with local immune interactions and patient prognosis, suggesting a central role for DCs in controlling local tumor immunity.

SUMMARY:Several techniques to determine apoptotic frequencies in tumors have been described. In this study, we report that biochemical detection of enzymatic caspase-3 activity is a simple and quantitative technique to measure apoptosis in colorectal tumor cells. The relevance of the level of apoptosis in colorectal cancer for the clinical course remains unclear. Therefore, we studied the correlation between caspase-3 activity and prognosis of the disease in relation to different factors known to be involved in apoptosis induction. High caspase-3 activity significantly correlated with a higher risk of recurrence and was preferentially found in tumors of the right side of the colon. No correlation was detected between high caspase-3 activity and altered protein expression of p53, ␤-catenin, or proteins of mismatched repair genes. This indicates that high caspase-3 activity has no evident correlation with the genetic Wnt-signaling or the mismatch repair mutational pathways. The caspase-3 activity significantly correlated with CD57 ϩ tumor infiltrating cells. Therefore, high caspase-3 activity in right-sided tumors might be induced by a specific lymphocytic reaction. (Lab Invest 2001, 81:681-688).A poptosis is an essential biologic process. As well as having a role in controlling cell number during early development, apoptosis is important for the removal of infected or genetically altered cells (Duke et al, 1996). Defects in the apoptotic mechanism are often found in neoplastic growth (Duke et al 1996;Green and Martin, 1995;Tompson, 1995), which is also the case in colorectal cancer (Evertsson et al, 1999;Gryfe et al, 1997;Tsujitani et al, 1996). Development of colorectal cancer proceeds through a series of genetic alterations that result in the activation of oncogenes and loss of tumor suppressor genes (Gryfe et al 1997;Rafter and Glinghammar, 1998;Ilyas et al, 1999). Mutations in genes known to be involved in cell cycle regulation, such as APC, p53, ␤-catenin, deleted in colorectal cancer (DCC), and K-ras, have been reported in colorectal cancer (reviewed in Gryfe et al, 1997). Furthermore, mutated genes involved in DNA mismatch repair can contribute to tumor growth. Alterations in these mismatch repair genes will lead to a microsatellite instability phenotype (MSI) (reviewed in Lothe, 1997). During tumorigenesis one of the target genes that is preferentially inactivated due to MSI is transforming growth factor ␤ receptor II (TGF␤RII) (Markowitz et al, 1995;Akiyama et al, 1996). Inactivation of TGF␤RII on cells down-regulates the suppressive function of TGF␤ on cell proliferation and in that way also results in disturbance of growth control (DeVisser and Kast, 1999). Most of the hereditary nonpolyposis colorectal tumors (HNPCC) exhibit MSI, whereas in sporadic colorectal cancers only 3% to 10% show this phenotype. This low rate of MSI in sporadic colorectal cancers seems to be caused by somatic inactivation of hMLH1 (Lothe, 1997).As well as the occurrence of apoptosis in cells caused by intrinsic factors, apopto...

Peripheral blood natural killer (NK) cells are usually defined as a homogeneous cell population. However, NK cells show heterogeneous expression of a diversity of cell surface molecules, which might reflect the diversity of NK-cell functions. Therefore, a more specific phenotypic definition of NK cells is necessary. In this study, we made an inventory of phenotypic subsets that are present within the peripheral blood NK-cell population of healthy donors based on differential expression of nine cell-surface markers. Using threecolour flow cytometric analysis we were able to define at least 48 different CD56 1 NK-cell subsets within the peripheral blood. This phenotypic heterogeneity appeared to be stable among healthy individuals, and was also steady within CD56 dim and CD56 bright NK populations, indicating a possible role for these subsets in NK-cell function or differentiation.

Interleukin‐2‐activated, cultured NK cells (A‐NK) cells were adoptively transferred into a syngeneic rat liver‐tumor model. The kinetics of tumor infiltration by NK cells, originating either from adoptively transferred or from endogenous sources, the localization of these cells in the tumor, and their interactions with extracellular‐matrix proteins were studied by immunohistochemistry and transmission‐electron microscopy. The adoptive transfer of A‐NK cells via the hepatic artery and s.c. injections of IL‐2 into rats bearing sub‐capsularly induced CC531 liver tumors, but also IL‐2 monotherapy, resulted in a significant increase of the number of NK cells both at the tumor border and in the tumor center. The majority of tumor‐infiltrating NK cells was present in the tumor stroma and only occasionally was an NK cell observed in a tumor nodule in direct contact with tumor cells. Observations by electron microscopy suggested that matrix proteins, abundantly present in the tumor stroma but absent in the tumor nodules, provide a substrate for migration of infiltrating cells, whereas tight structures of matrix proteins surrounding tumor nodules provide a barrier for establishment of direct NK‐cell‐to‐tumor‐cellcontact. Our results suggest that direct NK‐cell‐to‐target‐cell‐contact‐mediated lysis is of minor importance for attaining an anti‐tumor effect in this model. We hypothesize that treatment of tumor‐bearing rats with A‐NK cells and/or IL‐2 initiates a cascade of events (e.g., secretion of tumor‐killing cytokines and/or infiltration of other immune cells) ultimately leading to tumor regression. Int. J. Cancer 78:783–789, 1998. © 1998 Wiley‐Liss, Inc.