Abnormalities of cellular oncogenes in 22 cases of human primary gastric cancer were screened by Southern blot analysis using 16 onc probes. Human cellular sequences related to the v-yes oncogene of Y73 avian sarcoma virus (human c-yes-1 gene) were found to be amplified in a primary gastric cancer. The degree of amplification was about 4- to 5-fold. Another v-yes-related cellular gene (human c-yes-2, a pseudogene) was not amplified in this tumor. The normal stomach tissue adjacent to the tumor tissue in the same patient showed no amplification of c-yes-1 gene, suggesting that the amplified c-yes-1 is involved in the onset or the progress of this carcinoma.

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Spontaneous T-cell leukemia similar to human adult T-cell leukemia (ATL) was found in an African green monkey naturally infected with simian retrovirus closely related to human T-cell leukemia virus type I (HTLV-I). Monoclonal integration of the simian retrovirus was detected in the primary leukemic cells, suggesting an association of the retrovirus with ATL-like leukemia in the monkey.

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A monoclonal antibody 9A3 was raised to human gastric carcinoma cell line TMK-1, a poorly differentiated adenocarcinoma of the stomach. 9A3 antibody (IgG1, κ) strongly reacted with various carcinomas immunohistochemically, but did not react with any normal tissues of the whole body tested except for neutrophiles and macrophages. 9A3 antibody also reacted with the luminal surface of the fetal colon and esophagus at six months of gestation. The antigen recognized by 9A3 antibody might be an oncodevelopmental antigen.

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Alterations in γ-glutamyl transpeptidase (γ-GT) and alkaline phosphatase (ALP) activities and binding of specific antibodies to glucose-6-phosphate dehydrogenase (G6PDH), γ-GT and the A, B, C, D, and P forms of glutathione S-transferase (GST-A, -B, -C, -D, and -P, respectively) in preneoplastic and neoplastic lesions induced by N-ethyl-N-hydroxyethylnitrosamine (EHEN) in the rat kidney were investigated. Morphologically the lesions were of basophilic type and were classified either as altered tubules or adenomas, the latter being further subdivided into microadenomas and adenomas depending on size and the presence of compression. The fact that all lesions demonstrated only weak (or negative) γ-GT, ALP and GST-B stainings, all of which are normally evident in proximal tubules, and also the occasional finding of cells bearing a periodic acid-Schiff-positive apical brush border within altered tubules strongly positive for GST-A, indicated their histogenesis from the proximal segment of the nephron. Thus a directed shift to a phenotype opposite to that observed in the cells of origin was demonstrated. Comparison of phenotype within the range of EHEN-induced lesions strongly suggested putative preneoplastic character for the altered tubules. Transition to adenomas appeared to be correlated with progressive loss of GST-A and moderate to slight increase of G6PDH.

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The promoting effects of various chemicals and dietary constituents on bladder carcinogenesis were examined by means of a short-term assay, in which maintenance of concanavalin A agglutination of isolated rat bladder cells caused by a subcarcinogenic dose of N-butyl-N-(4-hydroxybutyl)nitrosamine was used as an indicator. Twenty-seven chemicals were examined as possible promoters. Positive results in this assay were consistent with established promoting effects in the cases of sodium saccharin, saccharin, sodium L-ascorbate, sodium cyclamate, DL-tryptophan, butylated hydroxy-anisole, butylated hydroxytoluene, L-thioproline and phenacetin. Allopurinol was the only established promoter that gave negative results in the agglutination assay. Thus, this method is useful for rapid evaluation of the specific promoting effect of a chemical on bladder carcinogenesis.

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The dose-response curves of benzidine derivatives calculated from Ames test data were analyzed by combining the least-squares method ith cluster analysis. The calculations of the electronic structures of various benzidine derivatives have at the same time been performed by using the CNDO/2 method. The cluster analysis was carried out on various parameters concerning the electronic structure, such as total electron density, frontier electron density, the energy level of HOMO (highest occupied molecular orbital), and so on. It was found that the total electron density on the nitrogen atom gives a similar pattern of classification to that for mutagenic compounds in cluster analysis. In other words, it may be inferred that the nitrogen atoms of the benzidine derivatives play an important role in the metabolic activation of the compounds. Consequently, cluster analysis with a parameter of electronic structure should be a useful initial screening procedure for various analogous mutagenic compounds as well as carcinogenic compounds.

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BglII fragment C mapping between 0.416 and 0.580 map units (mu) on the herpes simplex virus type 2 (HSV-2) genome was used for in vitro translation to identify proteins encoded on this fragment. RNA homologous to the BglII C fragment directs the synthesis of three proteins with approximate molecular weights of 144, 000, 52, 000 and 27, 000. The 27, 000 dalton protein is encoded by sequences within the EcoRI/HindIII AE fragment (0.419-0.525mu) that overlap the immortalizing se quences within BglII C. The 144, 000 (144K) and 52, 000 dalton proteins are encoded by sequences within the BamHI “e” fragment of HSV-2 DNA (0.535-0.585mu). The 144K protein is the only species translated in vitro from mRNA hybrid-selected from cells arrested in the “early” (β) phase of viral protein synthesis. It is precipitated by anti-ICP-10 serum and by monoclonal antibody 48S (previously shown to precipitate the HSV-induced ribonucleotide reductase). The 48S antibody competes with the anti-ICP-10 serum for the 144K protein. Furthermore the in vitro translated 144K protein is structurally similar to ICP-10, an HSV-2-infected cell protein that is antigenically ndentical to AG-4, the cervical tumor-associated antigen.

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Monoclonal antibodies against tumor-associated antigens (TAAs) expressed in the leukemic cells of enzootic bovine leukosis (EBL) were examined for their possible usefulness in detecting EBL tumor cells. The antibodies immunohistochemically reacted with EBL tumor cells and had cytotoxic activity against the cells but not against normal adult bovine peripheral blood lymphocytes (PBL) even after activation with mitogens. The TAA-positive EBL tumor cells also had B cell surface markers. The TAA was also found in several samples of PBL taken from BLV-infected healthy cattle (8 positive out of 81 tested), as well as in PBL from 13 out of 14 cattle with lymphocytosis but no evidence of tumor. Thus, monoclonal antibodies directed against TAAs might be a useful tool not only for diagnosing EBL but also for screening of BLV-infected cattle with the potential to develop tumors in the future.

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Two culture cell lines (C-Lu65, C-Lu99) were established from human giant cell carcinomas of the lung transplanted in athymic nude mice (BALB/c, nu/nu). During early passage in tissue culture, C-Lu65 grew as a loosely adherent monolayer with some piling-up and with floating cells. After 30 successive subcultures, C-Lu65 began to grow in suspended cell clusters, showing a faster growth rate. C-Lu65 was characterized by multinucleated giant cells with large abnormal nuclei and prominent nucleoli. C-Lu99 grew as adherent cells, and fewer multinucleated giant cells were observed. C-Lu65 and C-Lu99 showed some ultrastructural differences in cell surface and cytoplasmic features. Chromosomal analysis revealed numerical and structural abnormalities in both cell lines. Cell-free supernatants from both cell lines stimulated the colony formation of mouse bone marrow cells in vitro. In addition, mice bearing tumors induced by transplanting C-Lu65 and C-Lu99 showed remarkable leukocytosis without evidence of infection. These results suggest that these two cell lines release colony-stimulating factor both in vitro and in vivo.

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Fragile sites appear to be associated with a higher rate of breakage and specific chromosome rearrangement in cancer. A fragile site located on the human X chromosome at band Xq27 is known to be expressed under conditions of thymidylate stress. In order to obtain a model cell system suitable for studying the mechanism of expression of the fragile X site, interspecific somatic cell hybrids were constructed by cell fusion between human skin fibroblasts derived from a male patient with fragile X-linked mental retardation and thymidylate synthase-negative mouse mutant cells. The primary isolated hybrid clones were thymidine-prototrophic and expressed the fragile X site under conditions of thymidylate stress caused by 5-fluoro-2'-deoxyuridine treatment. In a thymidine-auxotrophic hybrid clone segregated from a thymidine-prototrophic hybrid clone, the expression of the fragile X site was induced in thymidylate stress conditions achieved by thymidine deprivation alone. This result provides direct evidence that expression of the fragile X site is dependent upon a lowered supply of thymidylate.

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To establish cell systems appropriate for investigating the mode of action of antiherpetic nucleoside analogs, mutants were constructed from murine FM3A mammary carcinoma cells, which were deficient in both thymidine kinase (EC 2.7.1.21) and thymidylate synthase (EC 2.1.1.45), but were transformed with a recombinant plasmid DNA containing the herpes simplex virus type 1 thymidine kinase gene. The transformed cells expressed viral thymidine kinase activity and showed unusually high sensitivity to the cytostatic action of the antiherpetic agent (E)-5-(2-iodovinyl)-2'-deoxyuridine, which was only weakly inhibitory to the growth of the parent cells.

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The inhibitory effect of tumor necrosis serum (TNS) on the artificial metastasis of B-16 melanoma cells and the spontaneous metastasis of Lewis lung carcinoma cells was investigated. The results obtained were as follows: 1) When tumor necrosis serum (TNS) was administered either 20min or 2 days after injection of B-16 melanoma cells, a very strong inhibitory effect (99%) relative to the control was noted. 2) TNS showed a 60% inhibitory effect on spontaneous metastasis, and the weight of the original tumor regressed by 60%. 3) No histological changes in normal tissues were observed microscopically following TNS injection. The above results confirm that TNS is extremely effective in preventing metastasis.

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Spleen cells of mice inoculated intravenously (iv) with concanavalin A (Con A) suppressed the in vitro normal spleen cell blastogenesis. The suppressor cells, as determined by measuring the inhibition of in vitro T cell blastogenesis by Con A, were resistant to X-irradiation. They lacked surface Ig and were abrogated by treatment with αThy 1.2 antibody and complement, but not with complement alone, indicating that these suppressor cells are not B cells but T cells. Inoculation of 5-fluorouracil (5-FU) inhibited the induction and/or activity of these suppressor cells, whereas that of cyclophosphamide (CY), adriamycin, vincristine, or 3-[(4-amino-2-methyl-5-pyrimidinyl)methyl]-1-(2-chloroethyl)-1-nitrosourea (ACNU) did not at any interval tested. Since two other antimetabolic agents, 6-mercaptopurine and 2'-deoxyguanosine, were also effective in inhibiting these suppressor T cells, it was supposed that Con A-induced suppressor T cells are metabolically more active than the other T subpopulations. The findings that CY and ACNU decreased the number of T cells as much as or more than 5-FU did, but that they did not inhibit these suppressor T cells, suggests that 5-FU more or less selectively inhibited this suppressor T-cell subpopulation.

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The effects of interleukin-2 (IL-2) on natural killer (NK) cell activity were examined in both healthy persons and cancer patients. The NK activity of peripheral blood lymphocytes from 29 healthy donors was augmented by IL-2 treatment in all cases. However, in 14 out of 50 cancer patients NK activity was not augmented by IL-2. The rate of NK augmentation in a group of cancer patients was significantly lower than that in a healthy control group. When the cancer patients were divided into each clinical stage, NK augmentation was as follows: in stage I, 3 out of 3; in stage II, 11 out of 12; in stage III, 10 out of 18; and in stage IV, 12 out of 17. The rate of NK augmentation was significantly lower in cancer patients of stages III and IV. The rate of NK augmentation was also dependent on the performance status (PS) of cancer patients. The NK augmentation of cancer patients of PS 0 to 2 was not significantly different from that of normal controls, whereas the rate of NK augmentation was significantly lower in cancer patients of PS 3 and 4.

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(2"R)-4'-O-Tetrahydropyranyladriamycin (THP), a new anthracycline antibiotic, showed stronger inhibition of the growth of L1210, CCMT (mouse mammary adenocarcinoma) and Yoshida sarcoma in rats than did adriamycin (ADM). The antitumor activity of THP against P388 and Meth-A (mouse fibrosarcoma) was equal or slightly superior to that of ADM. Moreover, THP was active against an ADM-resistant subline of P388. THP at the optimal effective dose against experimental tumors had no toxic effect on the cellular immune system in normal and tumor-bearing mice.

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The effect of squalene (SQ) on the antitumor activity of 3-[(4-amino-2-methyl-5-pyrimidinyl) methyl]-1-(2-chloroethyl)-1-nitrosourea (ACNU) was studied in a murine tumor system. SQ at 4.2g/kg exhibited a significant potentiating effect on the activity of 10mg/kg of ACNU against lymphocytic leukemia P388 and resulted in some long-term survivors without toxicity to the host. Simultaneous administration of SQ and ACNU was most effective.

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Sarcophaga lectin, a lectin purified from the hemolymph of Sarcophaga peregrina (flesh fly) larvae, was found to be therapeutically effective against both ascitic and solid tumors. It was especially effective when injected directly into or around nodules of the syngeneic tumor Meth A transplanted intradermally into BALB/c mice. It also induced cytotoxic activity against L-929 cells when it was added to the culture medium of macrophages in vitro. Similar cytotoxic activity was detected transiently in the serum when it was injected into the abdominal cavity of mice, in which sarcoma 180 cells had been inoculated. Probably, this cytotoxic activity induced by Sarcophaga lectin partly contributes to the elimination of tumors. The possible role of this cytotoxic activity is discussed from the viewpoint of ontogeny.

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The antitumor effects of three 5-fluorouracil-related compounds, 5'-deoxy-5-fluorouridine (5'-DFUR), 1-(2-tetrahydrofuryl)-5-fluorouracil (FT-207) and 5-fluorouracil (5-FU) itself, on several experimental murine tumors were compared after oral administration. 5'-DFUR showed strong antitumor activity against the solid type of four kinds of tumors tested with a wide range of effective doses, and also showed moderate antitumor activity against three kinds of solid tumors with a narrow range of effective doses. 5'-DFUR was effective against a few kinds of ascites tumors. In general, the antitumor activity of FT-207 was not very strong, with narrow ranges of effective doses under the present conditions. 5-FU showed strong toxicity but at lower doses its antitumor effectiveness was almost the same as that of FT-207. When the doses were divided into three and the divided dose was given orally three times a day for five consecutive days to mice bearing L1210 leukemia, this modality (with any of the three drugs) enhanced the ILS of the mice by two to three times in the case of the ascites type but not the solid type of L1210. The chemotherapeutic index in oral treatment of the solid type of tumors was higher for 5'-DFUR than for FT-207 or 5-FU. The minimum lethal doses in oral administration for five consecutive days were about 3, 1.5 and 0.5mmol/kg/day for 5'-DFUR, FT-207 and 5-FU, respectively. In conclusion, 5'-DFUR appeared to have stronger antitumor activity and less toxicity than FT-207 and 5-FU, and it is therefore expected to be clinically useful.

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