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We studied the effect of single and multiple injections of Corynebacterium granulosum on weight and histology of lymph nodes and spleen, on peripheral white blood cell count, response of peripheral blood lymphocytes, lymph node, and spleen cells to phytohemagglutinin and pokeweed mitogen, survival of skin allografts, and lung metastases of a syngeneic fibrosarcoma in C3Hf/Bu mice. Corynebacterium parvum was used in some studies on antitumor activity. The weight of lymph nodes and spleen was markedly increased by single and multiple i.p. injections of C. granulosum, the peak enlargement occurring at Day 7 in lymph nodes and at Day 16 in spleen. Histologically, there was an extensive proliferation of nucleated cells in the enlarged organs. C. granulosum did not change the total white blood cell count but caused a temporary lymphopenia. In general, in vitro response to phytohemagglutinin and pokeweed mitogen of blood lymphocytes and spleen cells was decreased. Lymph node cell response to phytohemagglutinin was increased by small doses (0.025 mg) of C. granulosum, was not altered by a single large dose (0.5 mg), and was decreased by multiple doses. The response of lymph node cells to pokeweed mitogen was increased by all treatments. These changes in response to mitogens were demonstrable for about 2 months after treatment. Treatment i.v. with 0.1 or 0.25 mg of C. granulosum given before but not after grafting significantly prolonged the survival of grafted BALB/c skin. Smaller doses of this bacterium were not effective. Splenectomy of skin graft recipients did not prevent the effect of C. granulosum. Treatment i.p. or i.v. with this bacterium significantly decreased the number of lung metastases from i.v.-injected fibrosarcoma cells, even if the cells were injected 3 to 4 months later. The magnitude of this effect varied with the dose and frequency of injection of C. granulosum and C. parvum.

Experiments were designed to investigate whether the tumor-associated macrophage (TAM) content of murine solid tumors correlates with the clonogenic ability of tumor cells to establish s.c. tumors, tumor growth rate, extent of tumor necrosis, tumor metastatic propensity, and tumor radioresponse. Of 13 tumors studied, 6 were sarcomas and 7 were carcinomas; all tumors were of spontaneous origin in C3Hf/Kam mice, with the exception of one sarcoma that was induced by 3-methylcholanthrene. Tumors were growing in the hind thighs of syngeneic mice, and their TAM content was determined when they were 8 mm in diameter. The TAM content varied greatly among tumors, ranging from 9 to 83%. Tumor bearing mice experienced a reduction of 50% or more in the number of peritoneal macrophages, but the degree of reduction was independent of TAM content. A significant negative correlation was noted between TAM content and TD50 values (i.e., the number of tumor cells needed to produce tumors in 50% of injected sites) and between TAM content and the amount of tumor necrosis. Also, an obvious trend toward positive correlation between TAM content and reduced local tumor radiocurability was apparent. No correlation was found between TAM content and tumor growth rate or metastatic spread. TAM from the NFSA sarcoma (a tumor with a low TD50 value, almost without necrosis, and poorly responsive to radiation) stimulated the in vitro growth of NFSA tumor cells. These observations suggest that high TAM content could be conductive to tumor cell proliferation and could be a factor in poor tumor radioresponse.

The ability of the in vitro sister chromatid exchange (SCE) assay to predict in vivo tumor drug sensitivity was investigated using a spontaneous hepatocarcinoma in C3Hf/Kam mice and 3 chemotherapeutic agents: melphalan; cis-platinum; and 1,3-bis(2-chloroethyl)-1-nitrosourea (BCNU). For hepatocarcinoma cells grown in monolayer culture, melphalan was the most efficient at inducing SCEs, and BCNU, the least. cis-Platinum induced a range in SCEs that overlapped those of BCNU and melphalan, suggesting that hepatocarcinoma is not a homogeneous population with intermediate sensitivity, but is a mixture of cis-platinum-sensitive and -resistant cells. According to in vitro cell survival curves, hepatocarcinoma was most sensitive to melphalan, less sensitive to cis-platinum, and essentially resistant to BCNU. The relative antineoplastic effects of melphalan, cis-platinum, and BCNU in vivo were compared by the response of artificial and spontaneous pulmonary metastases and solid tumors to these agents. For artificial metastases, there was a dose-dependent decrease in the number of lung nodules in mice treated with melphalan or cis-platinum, with melphalan being the more effective. BCNU had no effect. Spontaneous pulmonary metastases generated from hepatocarcinoma leg tumors were reduced in those mice treated with melphalan, unaffected by cis-platinum, and increased by BCNU. In hepatocarcinoma leg tumors (5 to 6 mm in diameter), melphalan induced the longest growth delay, and BCNU the least. Therefore, the relative effects produced by these three drugs in vivo were the same as predicted by SCE induction in vitro. The SCE assay may thus have potential clinical application.