Release from apoptosis correlates with tumor progression in the AKR lymphoma.

Disturbance of apoptosis is an established factor in tumorigenesis. The role of apoptosis in tumor progression is not yet clear. In the present study we compared the tendency to spontaneous apoptosis (and the proliferative capacity) of tumor cells derived from primary (PT) and metastatic tumor (MT) cells of several AKR lymphoma variants. Apoptosis-related gene expression was also compared. Our results indicate that release from apoptosis has a role in the tumor progression of this T cell lymphoma. At the cellular level, a markedly lower apoptotic tendency was observed in MT than in PT cells. The existence of macrophages only in PT also supports the presence of apoptotic cells in local but not in MTs. By contrast, proliferative capacity does not determine tumor aggressiveness in this system. At the molecular level, we found a higher staining intensity for bcl-2 in MT than in PT cells, suggesting that bcl-2 might be responsible for the reduced apoptosis in MT compared to PT cells. Evidence for p53 overexpression was found in the MT cells of one of the variants but in none of the PT. Comparison of Fas receptor, unexpectedly showed an increased expression in MT versus PT cells, possibly indicating resistance to Fas-induced apoptosis in the MT cells.

Apoptotic cell death and related gene expression in metastatic tumors of AKR lymphomas of varying malignancy.

Resistance to apoptosis may be related to tumor progression, due to the implications it might have on both tumor mass and genetic instability. We compared the tendency to spontaneous apoptosis and the proliferative capacity of metastatic growths of several AKR lymphoma variants (TAU-45, TAU-47, TAU-44, TAU-33, TAU-42 and TAU-46, in the order of increasing metastatic potential). We further compared the expression of several apoptosis-related genes. Cell proliferative capacity did not appear to determine malignant behavior since, on the whole, a decrease in S + G2M fraction was observed with increasing malignancy. Sensitivity to apoptotic cell death decreased with increasing malignancy when comparing the TAU-45, TAU-47, TAU-44 and TAU-33 variants, suggesting a role of reduced apoptosis in this T-cell lymphoma. An increase in Bcl-2 content with increasing aggressiveness among these variants, implicates this protein in this tumor progression-related resistance to apoptosis. However, the two variants of highest malignancy, TAU-42 and TAU-46, did not follow the same trend, since they displayed a relatively high content in apoptotic cells and a low Bcl-2 content. Fas receptor expression did not correlate with tendency to apoptosis, indicating that malignant behavior in the AKR lymphoma does not depend on CD95/Fas/APO1 downregulation. Overexpression of p53 was observed only in one of the variants of lowest malignancy.

Antitumoral activity of an immunomodulatory fraction of Nocardia opaca: mechanism of action.

Immunomodulatory substances have been used as antineoplastic agents in experimental and human systems. Many of these agents were derived from microorganisms. Several biologically active fractions have been isolated from Nocardia. These derivatives were shown to induce interferon production, to activate natural killer cells and macrophages and to exert an antitumoral effect. We attempted to examine the mechanism of the antitumoral activity of the Nocardia water-soluble mitogen (NWSM). The tumor tested was the Lewis lung carcinoma (3LL). Regular histological examination and identification of the cellular immune reaction by monoclonal antibodies against macrophages (Mac 1 antigen), B- (IgG expressing) and T-lymphocytes (anti-Lyt-1), analysed by flow cytometry, were performed on samples of the tumor site and of the spleen. Intratumoral administration of the immunomodulators resulted in a massive accumulation of inflammatory cells around the tumor in mice treated with NWSM. The thick rim of infiltrating cells consisted of macrophages and lymphocytes, while the nontreated tumor was found to provoke only a scanty lymphocyte infiltration. Macrophages were, therefore, present at the tumor site and were directly implicated in the antitumoral effect of the Nocardia immunomodulator. T-lymphocytes were also observed at the site of the tumor. The spleen reaction consisted of marked extramedullary hematopoiesis and enlarged follicles containing prominent germinal centres (assessed also by a FACS-demonstrated increase in B-lymphocytes). In view of the inefficiency of chemotherapy in the treatment of advanced cancer, it is of major importance to explore alternative cancer treatment modalities. Immunotherapy is a particularly interesting alternative since it can potentially affect metastatic disease.

Macrophage involvement in the antitumoral effect of Nocardia-delipidated cell mitogen (NDCM).

Several immunomodulatory fractions derived from Nocardia have been found to inhibit the growth of several experimental tumors, including Lewis lung carcinoma (3LL). An involvement of both macrophages and lymphocytes in the antitumoral effect of Nocardia fractions has been suggested. The mechanism of the Nocardia delipidated Cell Mitogen (NDCM)-induced tumor-inhibiting effect was investigated further in the present study. Macrophages activated by NDCM exerted a cytotoxic effect on 3LL cells in vitro, indicating a direct influence of macrophages on the tumor cells. These results correlate with previous findings which showed a local accumulation of macrophages (but also lymphocytes) at the tumor site in NDCM-treated mice. In tumor-bearing mice--both treated and non-treated with NDCM--a splenomegaly due to a pronounced extramedullary hematopoiesis was seen. Concomitant with the gradual evolution of the extramedullary hematopoiesis in the red pulp, a depletion in white pulp component was observed, more pronounced in the control 3LL-bearing mice than in the 3LL-inoculated NDCM-treated animals. The disappearance of the lymphatic follicles in 3LL-bearing mice may be responsible for the failure to cope with the tumor. It is therefore possible that by delaying white pulp depletion, NDCM favors a better host defense against the tumor. Examination of lungs in 3LL-bearing mice treated by NDCM showed a rich infiltration of macrophages in the vicinity of isolated tumor cells, probably indicating a defensive role of NDCM-activated macrophages against metastatic spread of the tumor. Although the macrophage appears to be of major importance in the NDCM-induced host response against the tumor, other components of the immune system are probably system are probably also activated by the Nocardia fraction in defense against the neoplasm.

Sensitivity to thermochemotherapy of AKR lymphoma and B16 melanoma variants of malignancy.

Drug resistance, which so often accompanies tumor progression, has been shown to be related to changes in membrane properties which may result in decreased drug accumulation in the tumor cell. A correlation between sensitivity to thermochemotherapy and degree of malignancy was found in the AKR lymphoma system. Hyperthermia increased adriamycin (ADR) uptake and concomitantly its cytotoxicity to AKR lymphoma cells. Moreover, these effects were more pronounced on a variant of high malignancy (HM) than on a low malignancy (LM) one. Fluorescent microscopy, as well as cytofluorometry, indicated that lymphoma cells treated by ADR at 43 degrees C were more permeable to the cytotoxic agent than those exposed to the chemotherapeutic substance at 37 degrees C. Cytofluorometry indicated the presence of a minor cell subpopulation with low ADR uptake in the HM variant, not found in the LM one. Fluorocytometry also showed that the temperature-dependent increased ADR uptake was more marked in the HM than in the LM variant, explaining the differential effect of thermochemotherapy on the two lymphoma variants. However, correlation between degree of malignancy and sensitivity to thermochemotherapy is not a general feature. In contrast to the results obtained in the AKR lymphoma system, in the B16 melanoma the low malignancy variant, F1, was more markedly affected by the combined treatment than the F10 variant. The increased cytotoxic effect of ADR by supranormal temperatures in the F1 variant was shown to be due to an augmented drug uptake. The results suggest that drug resistance in late stages of tumor progression can be overcome by an agent acting on the cell membrane. However, the data also indicate the necessity of assaying cancer treatment modalities, including those designed to circumvent drug resistance, on various tumor system models.

In vitro effect of levan-activated macrophages on Lewis lung carcinoma cells.

The polysaccharide levan (polyfructose) has previously been shown to exert an inhibitory effect on the growth of several murine tumors. This activity is mediated by a host reaction, involving mainly macrophages but also other elements of the immune system. It was not clear, however, whether levan-activated macrophages act by a direct cytocidal effect on the tumor cells or via the activation of a specific immune response to the tumor. In the present study, the possibility of a direct cytotoxicity of levan-activated macrophages against Lewis lung carcinoma cells was tested by coculture in vitro. It was found that levan-induced (as well as paraffin oil induced) macrophages actually exert a direct cytotoxic effect on Lewis lung carcinoma cells. The tumor cell killing is mediated by cell to cell contact. A cytoplasmic bridge was often seen between the macrophage and the tumor cell. The remaining tumor cells in the lysed area appear slender, shrunken and non-dividing.