In studying "hemorrhagic necrosis" of tumors produced by endotoxin, it was found that the serum of bacillus Calmette-Guerin (BCG)infected mice treated with endotoxin contains a substance (tumor necrosis factor; TNF) which mimics the tumor necrotic action of endotoxin itself. TNF-positive serum is as effective as endotoxin itself in causing necrosis of the sarcoma Meth A and other transplanted tumors. A variety of tests indicate that TNF is not residual endotoxin, but a factor released from host cells, probably macrophages, by endotoxin. Corynebacteria and Zymosan, which like BCG induce hyperplasia of the reticulo-endothelial system, can substitute for BCG in priming mice for release of TNF by endotoxin. TNF is toxic in vitro for two neoplastic cell lines; it is not toxic for mouse embryo cultures. We propose that TNF mediates endotoxin-induced tumor necrosis, and that it may be responsible for the suppression of transformed cells by activated macrophages.One of the best-known enigmas of cancer biology is the "hemorrhagic necrosis" of tumors induced by endotoxin (1-5). We report here that endotoxin acts indirectly by causing the host to release a substance, which we name tumor necrosis factor (TNF), that is selectively toxic for malignant cells.

(4) and normal and malignant tissues (6, 7) showed that R24 reacts with melanocytes, astrocytes, melanomas, astrocytomas, and a subset of sarcomas. R24 also mediates a variety of biological effector functions, including tumor cell aggregation, human complement-mediated cytotoxicity, and antibody-dependent cell-mediated cytotoxicity with human effector cells (refs. 8-10; unpublished work).In the present study, we have investigated the response of melanoma patients to R24 with regard to different dose levels, toxicity, serological parameters, and tumor response. MATERIALS AND METHODSPatients. Patients' tumors were shown to express GD3 by indirect immunofluorescence tests on frozen sections prior to treatment. All patients had objectively measurable disease, a performance status (Karnofsky scale) of at least 60, and had been off anticancer therapy for at least 4 weeks. No concurrent anticancer therapy was given during evaluation. Patients were considered evaluable 6 weeks after initiation of therapy; 10 patients are evaluable and 2 have not yet reached the 6-week mark.Preparation and Administration of R24. R24 was prepared from ascites of (BALB/c x C57BL/6)F1 mice and purified by ammonium sulfate precipitation, chromatography over protein A-Sepharose and Sephadex G-25 columns, and filtration. Each R24 batch was tested for antibody reactivity and assayed for nucleic acids, 16 mouse viruses, bacteria, fungi, and mycoplasma. Preparations underwent standard safety testing in mice and guinea pigs and pyrogenicity testing in rabbits.R24 was administered by i.v. infusion in 100-200 ml of 0.9% saline/5% human serum albumin. Skin tests with 0.1 ,ug of R24 were done before the first treatment. The schedule of treatment was 1 or 10 mg/M2 every other day for eight treatments or 30 mg/M2 per day by continuous infusion on days 1-5 and 8-12.Serological Tests. R24 antibody titers were determined by testing serum samples in protein A mixed hemadsorption assays (11) against the melanoma target cell line SK-MEL-28. R24 concentrations were measured by an enzyme-linked immunoassay. Falcon 3034 plates were precoated with purified R24 at 125 ,g/ml. Rabbit anti-mouse IgG3 (Bionetics, Kensington, MD) diluted 1:100 was mixed (1:1, vol/vol) with patients' serum samples diluted 1:4 and incubated for 120 min. The mixture was transferred to the precoated wells and incubated for 60 min, and wells were washed with phosphatebuffered saline. Wells were incubated with goat anti-rabbit IgG linked to alkaline phosphatase (Sigma) for 60 min. Alkaline phosphatase activity was determined using p-nitrophenyl disodium phosphate substrate (12). R24 concentrations were determined by comparison with standards using different concentrations of purified R24 diluted in a pretreatment serum sample from the patient.Human IgG antibody against mouse Ig was detected by enzyme-linked immunoassays. Falcon 3034 plates precoated with R24 at 50 ug/ml were incubated with patients' serum samples diluted 1:50 for 60 min and washed with'phosphatebuffered saline. Anti-human I...

The FAP tumor fibroblast antigen is highly expressed in primary and metastatic colorectal carcinomas and shows limited expression in normal adult tissues. This highly selective expression pattern allows imaging of colorectal carcinoma lesions as small as 1 cm in diameter on 131I-mAbF19 scans. Because of the consistent presence of FAP in the stroma of epithelial cancers and the accessibility of FAP-positive tumor stromal fibroblasts to circulating monoclonal antibodies (mAbs), this study suggests possible diagnostic and therapeutic applications of humanized mAbF19 and mAbF19 constructs with novel immune and nonimmune effector functions.

Human cell lines of hematopoietic origin were tested for production of tumor necrosis factor (TNF) hTNF activity was not found in supernatants of cell lines of T-cell, monocytic, or promyelocytic origin. Partially purified hTNF has a molecular weight of approximately 70,000, has no interferon activity, is acid labile, is destroyed by heating at 70°C for 1 hr, induces cross-resistance to mouse TNF in vitro, and causes hemorrhagic necrosis of Meth A mouse sarcoma in the standard in vivo mouse TNF assay. Tests with a panel of 23 human cancer cell lines showed that hTNF is cytotoxic for 7 cell lines, cytostatic for 5, and has no effect on 11. Comparative studies with human a, fi, and y interferons indicated that sensitivity to hTNF and interferon can be distinguished. Combined treatment with hTNF and a or y interferon resulted in a synergistic cytotoxic effect.

Tumor necrosis factor, lymphocyte-activating factor, and enhanced levels of type I interferon were found in serum samples taken 2 h after mice infected with Plasmodium vinckei subsp. petteri received a small intravenous injection of endotoxin. These three mediators are among those released when mice receive an endotoxin injection 2 weeks after Mycobacterium bovis BCG or Corynebacterium parvum have been administered. There is indirect evidence that this wider range of mediators is also released in P. vinckei subsp. petteri-infected mice given parenteral endotoxin. A recent report that endotoxin is detectable in the plasma of malaria-infected mice and children implies that these mediators may also be released in the acute phase of the natural infection. We propose that these macrophage-derived mediators may be important in the glucocorticoid antagonism, bone marrow depression, fever, hypergammaglobulinemia, splenomegaly, elevation of serum amyloid A, consumptive coagulopathy, and shock syndrome with associated organ damage which can accompany malaria. The intraerythrocytic parasite death seen at crisis in some malarias, as well as the subsequent development of specific protective immunity, may also depend on these mediators.