Monique Martin

Tumor-infiltrating dendritic cells are defective in their antigen-presenting function and inducible B7 expression in rats

Tumors are tolerated by the immune system notwithstanding the expression of tumor‐associated antigens. PROb tumor cells, derived from a rat colon carcinoma, are rejected by tumor‐immune hosts but give rise to progressive tumors in naive hosts. Paradoxically, these tumors are heavily infiltrated by dendritic cells that express MHC class II and ICAM‐1. These tumor‐infiltrating dendritic cells (TiDCs) could be expected to process and present to T cells the antigens released by the adjacent tumor cells. Indeed, we report here that TiDCs, compared with splenic dendritic cells, are poor stimulators of primary allogeneic T‐cell proliferation and cytokine [interleukin‐2 (IL‐2) and interferon‐γ] production. Most of them (89–97%) do not express B7, an essential co‐stimulatory signal for T cells, even after a culture period allowing B7 up‐regulation on epidermal Langerhans cells. GM‐CSF in association with tumor necrosis factor‐α or IL‐4, or cell‐associated CD40‐ligand, all known to be potent stimulators of B7 expression on other dendritic cells, did not restore B7 expression by TiDCs. After a first exposure to TiDCs, allogeneic T‐cell response to a second challenge to splenic dendritic cells was decreased. The failure of most dendritic cells infiltrating PROb tumors to express B7, even after stimulation, may contribute to their poor capacity to stimulate T cells and could play a role in the immune tolerance allowing tumor growth. Int. J. Cancer 72:619–624, 1997.

Role of Stromal Myofibroblasts Infiltrating Colon Cancer in Tumor Invasion

In the normal colon, myofibroblasts are closely apposed to colonocytes where they deposit type IV collagen, the main basement membrane component. In colon carcinomas, this epithelial-mesenchymal association is physically disrupted, leading to the production of an abnormal, type IV collagen defective, basement membrane. Tumor-infiltrating myofibroblasts are migratory cells that accumulate at the invasive front of the colorectal carcinomas. They produce lytic enzymes able to degrade the basement membrane surrounding tumor glands. They also participate in the synthesis of the extracellular matrix components of the tumor stroma, which could subsequently alter the adhesive and migratory properties of the epithelial colon cancer cells. These results suggest that tumor-infiltrating myofibroblasts play a role in the invasion and metastasis of colorectal tumor cells.

Identification of Tumor-Infiltrating Macrophages as the Killers of Tumor Cells After Immunization in a Rat Model System

Immunization can prevent tumor growth, but the effector cells directly responsible for tumor cell killing in immunized hosts remain undetermined. The present study compares tumor grafts that progress in naive syngeneic rats with the same grafts that completely regress in hosts preimmunized with an immunogenic cell variant. The progressive tumors contain only a few macrophages that remain at the periphery of the tumor without direct contact with the cancer cells. These macrophages do not kill tumor cells in vitro. In contrast, tumors grafted in immunized hosts and examined at the beginning of tumor regression show a dramatic infiltration with mature macrophages, many of them in direct contact with the cancer cells. These macrophages are strongly cytotoxic for the tumor cells in vitro. In contrast to macrophages, tumor-associated lymphocytes are not directly cytotoxic to the tumor cells, even when obtained from tumor-immune rats. However, CD4+ and CD8+ T cells prepared from the regressing tumors induce tumoricidal activity in splenic macrophages from normal or tumor-bearing rats and in macrophages that infiltrate progressive tumors. These results strongly suggest that the main tumoricidal effector cells in preimmunized rats are macrophages that have been activated by adjacent tumor-immune lymphocytes.

An Experimental Model for Cancer of the Colon and Rectum

1,2-Dimethylhydrazine(DMH) was administered s.c. to a group of 20 inbred BD-IX rats at the dose of 15 mg/kg body weight, weekly for 7 months. Intestinal adenocarcinoma have been found in all of the tr

Role of Tumor Cell Apoptosis in Tumor Antigen Migration to the Draining Lymph Nodes

Establishment of an immune response against cancer may depend on the capacity of dendritic cells to transfer tumor Ags into T cell-rich areas. To check this possibility, we used a colon cancer cell variant that yields tumors undergoing complete T cell-dependent rejection when injected into syngeneic rats. We previously demonstrated that immunogenicity of these tumors depended on the early apoptosis of a part of these tumor cells. In this paper we show that fluorescent tumor cell proteins are released from FITC-labeled tumor cells and undergo engulfment by tumor-infiltrating monocytes without a phenotype of mature dendritic cells or macrophages. Fluorescence-labeled mononuclear cells with a phenotype of MHC class II+ dendritic cells are also found in the T cell areas of the draining lymph nodes. Interestingly, no fluorescent cell can be found in lymph nodes after a s.c. injection of Bcl2-transfected apoptosis-resistant tumor cells that yielded progressive tumors. Proliferation of tumor-immune T lymphocytes was induced by dendritic cells isolated from the draining lymph nodes recovered after a s.c. injection of apoptosis-sensitive, but not apoptosis-resistant, tumor cells. These results show that tumor cell apoptosis releases proteins that are engulfed by inflammatory cells in the tumor, then transported to lymph node T cell areas where they can induce a specific immune response leading to tumor rejection.

Effects of cyclosporin A on progressive and regressive tumors induced by two cancer lines derived from a single colon carcinoma chemically induced in the rat

The effects of cyclosporin-induced immunosuppression were assessed in a rat model of progressive and regressive colonic tumors. Two cloned cell variants, obtained from the same chemically induced colonic carcinoma, differ in their capacity to grow when injected into the syngeneic rat. PROb cells yield progressive tumors and often metastases; in contrast, REGb cells produce tumors which regress in 3 to 6 weeks. Cyclosporin A (CsA) administered daily, 20 mg/kg subcutaneously (s.c.) for 30 days after tumor cell inoculation, drastically enhanced the local growth of PROb tumors and increased the number of metastases. It increased the local growth and prevented the regression of REGb tumors which persisted even as long as 8 weeks after the termination of CsA administration and occasionally yielded metastases. CsA prevented the accumulation of inflammatory cells with the T lymphocyte phenotype at the periphery of both PROb and REGb tumors but did not alter the tumor infiltration by macrophages and NK cells. CsA did not modify the natural cytotoxicity of peripheral blood mononuclear cells against PROb and REGb target cells. These results suggest that CsA-induced suppression of T lymphocyte activity may enhance tumor progression and suppress tumor regression in this model.

Abnormal basement membrane in tumors induced by rat colon cancer cells

BACKGROUND/AIMS Colonic mucosa basement membrane results from a cooperation between epithelial cells and pericryptal fibroblasts characterized as myofibroblasts. This cooperation may be abnormal in colorectal carcinoma resulting in basement membrane alteration. METHODS Basement membrane composition and myofibroblast distribution were studied in normal rat colon and two colon carcinoma models by immunohistochemistry. Colon cancer cells and tumor-associated myofibroblasts were also studied for their capacity to deposit three basement membrane components (laminin, heparan sulfate proteoglycan, and type IV collagen) in vitro. RESULTS A continuous, type IV collagen-containing basement membrane, such as that observed in normal colon, was found only in the most differentiated tumor model and was restricted to the areas in which myofibroblasts were closely apposed to carcinoma cells. In other areas of this tumor and in the poorly differentiated tumor model, myofibroblasts dissociated from the epithelial cells and the basement membrane was devoid of type IV collagen. In vitro, carcinoma cells deposited laminin and heparan sulfate proteoglycan but not type IV collagen. Tumor-associated myofibroblasts deposited type IV collagen only in the presence of tumor cell extracellular matrix or laminin coating. CONCLUSIONS The colon cancer basement membrane defect in type IV collagen may result from a physical disruption in the association between epithelial cancer cells and myofibroblasts.

T‐cell co‐stimulation by the CD28 ligand B7 is involved in the immune response leading to rejection of a spontaneously regressive tumor

Cell variants from experimental tumors may lose their tumorigenicity or give rise to tumors that regress after a short period of progression in immunocompetent syngeneic animals. Rejection of these tumor cells is often T‐cell‐dependent. It has recently been reported that, besides the specific signal delivered through the clonogenic receptor, T‐cell activation requires a co‐stimulatory signal, delivered through its CD28 receptor by B7‐1 and/or B7‐2 molecules expressed at the surface of the antigen‐presenting cells. CTLA4Ig, a fusion molecule that specifically inhibits B7‐1 and B7‐2 binding to their receptor on T cells, was used to investigate the role of B7 in the spontaneous regression of the tumors induced in syngeneic rats by REGb cells, a regressor cell line established from a chemically induced colon carcinoma. When rats received either 1 or 3 CTLA4Ig injections, REGb tumors grew 3 or 7 times larger than in control animals, respectively. However, in most animals, single or repeated CTLA4Ig injections delayed rather than suppressed REGb tumor rejection. Antibodies to CTLA4Ig appeared in treated rats and could explain this transient effect. Neither REGb cells nor freshly isolated MHC class‐11+ antigen‐presenting cells infiltrating REGb tumors expressed B7, establishing that the target of CTLA4Ig was not located inside the tumor. In contrast, MHC class‐II+ B7+ accessory cells were found in the REGb tumor‐draining lymph nodes, suggesting that lymphoid tissue, rather than the tumor itself, was the site of tumor‐antigen presentation to tumor‐specific T cells. These results establish the role of the B7/CD28 co‐stimulation pathway in the control of a spontaneously regressive tumor.

Effect of indomethacin on the growth of colon cancer cells in syngeneic rats

A cell culture line has been established from a colonic tumor induced in a rat by dimethylhydrazine. Indomethacin had no effect on the growth of tumors induced in syngeneic rats by s.c. injection of this cell line. Neither indomethacin, nor prostaglandin E2 modify the growth of this established cell line in vitro or alter its destruction by endotoxin-activated macrophages. Indomethacin has been reported to suppress the growth of colonic tumors induced in rats by various carcinogenic drugs. The difference between these results obtained by others on chemically-induced colon cancer and the data we have obtained with a cell line derived from a similar cancer suggests that cancer cells in established culture may loose their sensitivity to prostaglandins, prostaglandin inhibitors or prostaglandin-dependent effector host cells.

New insights into the kinetic resistance to anticancer agents

Kinetic resistance plays a major role in the failure of chemotherapy towards many solid tumors. Kinetic resistance to cytotoxic drugs can be reproduced in vitro by growing the cells as multicellular spheroids (Multicellular Resistance) or as hyperconfluent cultures (Confluence-Dependent Resistance). Recent findings on the cell cycle regulation have permitted a better understanding why cancer cells which arrest in long quiescent phases are poorly sensitive to cell-cycle specific anticancer drugs. Two cyclin-dependent kinase inhibitors (CDKI) seem particularly involved in the cell cycle arrest at the G1 to S transition checkpoint: the p53-dependent p21cip1 protein which is activated by DNA damage and the p27kip1 which is a mediator of the contact inhibition signal. Cell quiescence could alter drug-induced apoptosis which is partly dependent on an active progression in the cell cycle and which is facilitated by overexpression of oncogenes such as c-Myc or cyclins. Investigations are yet necessary to determine the influence of the cell cycle on the balance between antagonizing (bcl-2, bcl-XL...) or stimulating (Bax, Bcl-XS, Fas...) factors in chemotherapy-induced apoptosis. Quiescent cells could also be protected from toxic agents by an enhanced expression of stress proteins, such as HSP27 which is induced by confluence. New strategies are required to circumvent kinetic resistance of solid tumors: adequate choice of anticancer agents whose activity is not altered by quiescence (radiation, cisplatin), recruitment from G1 to S/G2 phases by cell pretreatment with alkylating drugs or attenuation of CDKI activity by specific inhibitors.