Catechol (CAS: 120-80-9) is present in the environment, being a major industrial chemical as well as a major phenolic component of cigarette smoke. Continuous oral treatment of rats with 0.8% catechol for 51 weeks after a single intragastric dose of 150mg/kg of N-methyl-N'-nitro-N-nitrosoguanidine strongly enhanced both forestomach and glandular stomach carcinogenesis. In addition, and more importantly, catechol alone induced adenocarcinoma and adenomatous hyperplasia in the pyloric region of the glandular stomach. These results clearly indicate that this environmental contaminant merits classification as an enhancer of forestomach and glandular stomach carcinogenesis with complete carcinogenic potential for the glandular stomach. Its significance for gastric tumor development in man requires elucidation.

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Two HTLV-I-carrying T cell lines were prepared from peripheral lymphocytes of a virus-infected (B/J×Chbb:HM) F1 rabbit, and these cells were inoculated intraperitoneally into 5 newborn F1 rabbits. These animals were killed 3-5 weeks later. Their leukocyte counts were higher than those in normal control animals, with abnormal lymphocytes amounting to 3-5% of total leukocytes. Histological examination showed leukemic infiltration in liver, spleen, lung and kidneys of all these animals. The peripheral lymphocytes at first lacked HTLV-I antigens, but became antigen-positive after in vitro culture. Southern blot analysis of these cells revealed HTLV-I integration patterns different from those of the inoculated cells.

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High-titer antibody against HTLV-I in carrier mothers is proposed as a secondary parameter for risk of viral transmission to the children via milk. For titration of antibodies, a less expensive modified gelatin particle agglutination assay (approximately 40% of the standard cost) was developed. None of 11 carrier mothers with antibody titers of less than 4000 had carrier children, whereas 11 out of 17 mothers with titers of 256, 000 or higher had carrier children. The antibody titer was correlated with antigen-bearing cells detectable in cultures of peripheral blood T-lymphocytes, which was previously described as a marker of risk for transferring HTLV-I to children.

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Liposomes containing fragment A of diphtheria toxin killed human immunodeficiency virus (HIV)-producing MOLT-4 or TALL-1 cells, and HTLV-I-carrying MT-4 cells infected with HIV, but did not kill these cells when they were not infected with HIV. This killing was not affected by the presence of HIV antibodies. These results show that liposomes containing fragment A of diphtheria toxin selectively killed cells expressing HIV antigens or producing the virus in vitro.

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The inhibitory effects of several polysaccharides, dextran, xylofuranan, and ribofuranan, and their sulfated counterparts on the infectivity and replication of human immunodeficiency virus (HIV) were examined by using an HTLV-I-carrying cell line, MT-4, in vitro. Dextran sulfate [34] (Mw 34×10³), xylofuranan sulfate, and ribofuranan sulfate completely prevented HIV-induced cytopathic effects (CPE) at concentrations >10μg/ml and dextran sulfate [7] (Mw 7×10³) at concentrations >100 μg/ml. However, the non-sulfated compounds did not prevent them at any concentration tested. The anti-HIV effect of these polysaccharides was confirmed by measuring HIV-specific antigen expression in infected MT-4 cells. In cocultures with MOLT-4 and MOLT-4/HIV_HTLV-HIB cells, formation of multinucleated cells was completely inhibited in the presence of 100μg/ml of these sulfated compounds. Dextran sulfate [34] showed 20-30% growth inhibition of uninfected MT-4 cells at 1000μg/ml but dextran sulfate [7], xylofuranan sulfate, and ribofuranan sulfate showed no effect on MT-4 cell growth at this concentration. All of these sulfated polysaccharides efficiently inhibited the reverse transcriptase activity of avian myeloblastosis virus and HIV.

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c-myc protein p62 in human gastric carcinomas was studied by immunohistochemistry using an antiserum raised against the C-terminal portion of the human c-myc gene protein. No immunoreactivity of c-myc p62 was detected in tumor cells of 27 early carcinomas, while c-myc p62-positive tumor cells were observed in 51 (36.4%) of the 140 advanced carcinomas. Many stromal cells including macrophages and fibroblasts around the tumors showed various degrees of c-myc p62 immunoreactivity. Localization of c-myc p62 was predominant in the cytoplasm of tumor cells and stromal cells. The prognosis of patients with p62-positive stromal cells was significantly better than that of patients with p62-negative stromal cells.

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We used three highly and two weakly metastatic clones obtained from a rat mammary carcinoma cell line (c-SST-2) to examine the relationship between the metastatic capacity of tumor cells and their intercellular communication with normal fibroblasts. By employing the dye-transfer method, we found that the frequency of intercellular communication between weakly metastatic clone cells and fibroblasts was significantly higher than that between highly metastatic clone cells and fibroblasts. These results suggest that normal fibroblasts may regulate the metastatic capacity of tumor cells by intercellular communication.

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Human granulocyte colony-stimulating factor (hG-CSF) cDNA was cloned, by using a synthetic oligonucleotide probe, from an Okayama-Berg cDNA library of lipopolysaccharide-stimulated human peripheral blood macrophages. The cDNA encodes a polypeptide with an amino acid sequence which completely matches that of the known polypeptide with hG-CSF activity derived from human tumor cell lines. Expression in E. coli of high levels of the protein (about 10% of total cellular proteins) was accomplished under control of the trp promoter, and the purified protein was proved to have hG-CSF activity. Our data provide evidence that human peripheral blood macrophages do produce hG-CSF mRNA when stimulated exogenously, suggesting they are the producer of naturally occurring hG-CSF.

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A new human cell line, SP-49, was established from the peripheral blood of a patient with leukemic conversion of diffuse lymphoma, medium-sized cell type. SP-49 cells were shown to have intracytoplasmic and surface immunoglobulin and Leu-12 antigen, but were negative for Epstein-Barr virus nuclear antigen. Chromosome analysis demonstrated that SP-49 had t(11;14) chromosome translocation involving 11q13 and 14q32, where oncogene bcl-1 and immunoglobulin heavy chain gene, respectively, were reportedly located.

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To evaluate the effects of continuous low-level ionizing radiation on humans, the follow-up data (1980-85) on Japanese thorotrast-exposed patients were analyzed. The patients were 241 war-wounded military personnel registered with and cared for by the Ministry of Health and Welfare since 1979. During this period, a total of 1144 person-years, 94 patients died. Compared with the expected number of deaths calculated from age-and cause-specific death rates in Japan during the same period, the thorotrast-exposed patients were at three times greater risk of death from all causes (P<0.001), had 47 times the risk of liver cancer (P<0.001), 12 times the risk of leukemia (P<0.05), and 20 times the risk of liver cirrhosis (P<0.001). Age at time of thorotrast injection, drinking and smoking habits had little effect on these statistics. Analyses of 30 autopsied patients with liver cancer showed statistically significantly increases in hemangiosarcoma and cholangiocarcinoma. The thorotrast-exposed patients' estimated risk of liver cancer by histological type was 21 times that of the general population for hepatocellular carcinoma, 303 times that for cholangiocarcinoma and 3129 times that for hemangiosarcoma.

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The phalloidin sensitivity of hepatocytes resulting in the formation of cytoplasmic blebs was examined with cells isolated from 73-to 74-week-old and 99-to 100-week-old F344/DuCrj rats of both sexes by a collagenase perfusion method. The cells isolated from aged rats were less sensitive to the toxin than those obtained from 10-to 14-week-old rats. The decrease in the sensitivity was more marked in males than in females, and it appeared at an earlier age in the former than in the latter. Phalloidin consumption experiments showed decreases in the cellular uptake of the toxin in aged rats, and these were more marked in males than in females. The low cellular uptake of the toxin seemed to play an important role in the low sensitivity of the cells in aged rats. Although histological and histochemical examinations showed the development of foci of altered hepatocytes in the aged rat liver, the foci were estimated to account for less than 1.5% of liver tissues. Thus, the decrease in the sensitivity of the cells isolated from whole liver tissues might mainly be attributed to the decrease in the sensitivity of otherwise normal-looking hepatocytes.

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The effects of dietary oils and fats used in Japan on the fecal contents of bile acids in rats were studied. F344/Du Crj female rats (8 weeks old) were fed on diet containing 20% corn oil, rape seed oil, sesame oil, soybean oil, lard, or tallow as high oil or fat diets or on 0.2% linoleic acid diet as a low fatty acid diet for 4 weeks, and then their feces were collected. Bile acids in the feces were partially purified and analyzed by high-performance liquid chromatography. Analyses showed that lard or tallow in the diet resulted in significant increases in the contents of bile acids in the feces, whereas sesame oil in the diet resulted in significant decreases in their contents.

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The tumor-initiating and -promoting effects of ionizing radiation were tested, using mouse skin as the target organ and ⁹⁰Sr-⁹⁰Y beta-rays as the source of radiation. Neither single 2400 rad beta-irradiation followed by repetitive treatment with 12-O-tetradecanoylphorbol-13-acetate (TPA) nor single pretreatment with 7, 12-dimethylbenz[a]anthracene (DMBA) followed by repetitive 470 rad beta irradiation produced tumors above the level of significance within a period of 210 days, while a positive control, DMBA+TPA, yielded a high incidence of papilloma in a shorter period. DMBA exerted an action antagonistic to beta-rays in the induction of malignant tumors. It is concluded that the tumor-enhancing activity of repetitive radiation is qualitatively different from the promoting activity of TPA.

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Using 40 tumor cell strains derived from various organs of Japanese tumor patients and also 12 normal cell strains, we have measured the activity of O⁶-methylguanine-DNA methyltransferase (MT), which can repair O⁶-methylguanine produced in DNA by alkylating agents. Then, the lethal sensitivities of the strains to the anti-tumor drug 1-(4-amino-2-methyl-5-pyrimidinyl)methyl-3-(2-chloroethyl)-3-nitrosourea hydrochloride (ACNU) were measured. The MT activity was assayed by measuring the ³H radioactivity transferred from the substrate DNA containing [methyl-³H]-O⁶-methylguanine to acceptor molecules in the cell extracts. Extracts from the majority of tumor strains and all normal cell strains contained substantial MT activity of varying degree, while the extracts of 6 tumor strains showed virtually undetectable MT activity. Hence these 6 strains were assigned as Mer^- the phenotype which is characterized by the inability to repair O⁶-methyl-guanine in DNA due to the lack of MT. The Mer^- tumor strains were much more sensitive to ACNU than the rest of Mer⁺ strains, as measured by colony-forming ability. Furthermore, with all the tumor and normal strains tested, a good correlation was observed between MT activity and cellular resistance to ACNU. These results indicate that the frequency of Mer^- strain is about 15% among Japanese tumor cell strains so far analyzed, and further suggest that MT may be the only system able to repair lethal DNA damage induced by ACNU.

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Human amnion membrane epithelial (FL) and Madin-Darby canine kidney (MDCK) cells formed colonies which consisted of tightly packed cells. A bundle of actin filaments in the cells at the edge of the colonies ran parallel to the outline of these cells. The cells were attached to the substratum by regions which were observed as black solid lines by interference-reflection microscopy. Treatment with 12-O-tetradecanoylphorbol-13-acetate (TPA) for 24hr induced dissociation of and morphological changes in the cells in the colonies. TPA also disrupted the organization of the actin filaments and the attachment of cells to the substratum. Addition of herbimycin A for 24hr inhibited the TPA-induced dissociation of the cells in colonies. Treatment with herbimycin A also inhibited TPA-induced modulation of the organization of actin filaments and of the adhesion of cells to the substratum. However, herbimycin A did not inhibit the rapid spreading of cells induced by TPA in the absence of serum. These results suggest that herbimycin A inhibits the dissociation of cells and the morphological changes induced by TPA through suppression of the disruption of a bundle of actin filaments and modulation of adhesion of cells to the substratum.

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Media conditioned by the human bladder carcinoma cell line HTB9 contained high leukemic blast growth factor activity and also showed granulocyte-macrophage colony-stimulating factor (GM-CSF) activity. Northern blotting analysis of total RNA from HTB9 cells using GM-CSF cDNA as a probe demonstrated abundant expression of the GM-CSF gene. Thus, this cell line secretes GM-CSF, and this factor contributes to the proliferation of clonogenic leukemic blast cells.

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Glycolipids from human leukemia cells were analyzed by gas-liquid chromatography, enzymatic hydrolysis and high-performance thin-layer chromatography with immunostaining by the use of mouse monoclonal antibody. Glucosylceramide, lactosylceramide, and ganglioside GM3 were found in various leukemia cases. Acute lymphoblastic leukemia cells contained little or none of the glycolipids with lacto-series structures such as neolactotetraosylceramide (nLc4Cer), NeuAcα2-3Galβ1-4GlcNAcβ1-3Galβ1-4Glcβ1-Cer, or NeuAcα2-6Galβ1-4GlcNAcβ1-3Galβ1-4Glcβ1-Cer (IV⁶NeuAc-nLc4Cer), which were found in every case of various myeloid leukemias. GM3 was a major ganglioside (45-100%) in acute leukemia cells, whereas IV⁶NeuAc-nLc4Cer was a major one (37%) in chronic myelogenous leukemia cells.

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We studied the relationship between the metastatic potential of murine B16 melanoma cells and their surface expression of heparan sulfate glycosaminoglycan (HS-GAG) by using HepSS-1, a monoclonal antibody specific to HS-GAG. Firstly, among five B16 sublines, those capable of developing lung colonies with high efficiencies, such as B16-F10, had relatively low levels of surface HS-GAG. Secondly, a subline freshly prepared from metastatic lung colonies (F1) displayed a level of surface HS-GAG lower than that of injected B16 cells. Thirdly, in vitro selection of B16 cells with low surface HS-GAG by repeated HepSS-1 staining and cell-sorting resulted in cells with a higher metastatic efficiency than that of the original B16 cells. Collectively, B16 melanoma cells with high metastatic activities seem to have low surface HS-GAG. We also found that there was a positive correlation between the surface level of HS-GAG and the susceptibility to natural killer cells in eight B16 sublines.

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Tumor antigen TA-4 is divided into two subgroups; acidic and neutral TA-4. The tissue localizations of these TA-4 subgroups were examined by using monoclonal antibodies, i.e., Mab-21 which reacts with both acidic and neutral TA-4, and Mab-317 which is specific to acidic TA-4. Immunohistochemical staining with Mab-21 showed positive cells in most parts of the cancer nest and in the intermediate layer of the non-cancerous squamous epithelium of the uterine cervix, whereas positive staining with Mab-317 was observed only in the cells at the peripheral parts of the cancer nest adjacent to the surrounding stromal tissue. Thus, examination of the subgroups of TA-4 may be a useful aid for investigating the biologic behavior of squamous cells.

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The in vivo administration of a recently described recombinant human-mouse chimeric antibody specific for a human common acute lymphocytic leukemia antigen (cALLA) caused significant inhibition of the tumorigenic growth of human leukemic cells (Manca cells, expressing cALLA on the surface) which were implanted into nude mice. Intratumor as well as intraperitoneal administration of the human-mouse chimeric antibody repressed the tumor growth of Manca cells in nude mice. In order to investigate the in vivo localization of the antibody molecules, the chimeric antibody was labeled with radioiodine (¹³¹I) and injected into nude mice transplanted with Manca cells. The labeled antibody was significantly localized in the tumor and the location of the tumor was successfully visualized by scintiphotoscanning. These results indicated that the recombinant human-mouse chimeric antibody can exert a significant antitumor effect in vivo and can be utilized for radio-immunoimaging. Since the chimeric human-mouse monoclonal antibody would be expected to have a much lower antigenicity to humans and much higher efficiency in the interaction with human effector cells, such recombinant chimeric antibodies may be beneficial for immunotherapy and immunoimaging of cancer patients.

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The antiproliferative activity of recombinant human interferon (rIFN)-β and rIFN-γ, alone or in combination, against a human colon cancer cell line (RPMI 4788) was examined in vitro and in nude mice. rIFN-β and rIFN-γ interacted synergistically in vitro. In experimental pulmonary metastasis and intraabdominal carcinomatosis in nude mice, rIFN-β and rIFN-γ were administered iv and ip, respectively, alone or in combination, for 10 consecutive days beginning 2 days after RPMI 4788 cells were inoculated. In both models, rIFN-β and rIFN-γ alone had significant antitumor effects compared with the saline-control group. Combined administration of rIFN-β and rIFN-γ resulted in marked antitumor effects. In the pulmonary metastasis model, there were no pulmonary metastatic nodules in the group treated with the combination of rIFN-β and rIFN-γ (P<0.001), while there were 324.6±83.1 (mean±SD) nodules in the control group. In the intraabdominal carcinomatosis model, the mean survival time was 114.0±8.2 days (P< 0.01) for the combination therapy group, but only 41.8±5.6 days for the control group. These results suggest that combined treatment with rIFN-β and rIFN-γ might be a promising therapy for pulmonary metastasis and intraabdominal carcinomatosis of human cancers.

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The growth-inhibitory activity of recombinant human tumor necrosis factor (rH-TNF: PAC-4D) against 32 human cultured cell lines derived from leukemias and lymphomas (7 T cell, 11 B cell, 5 non-T, non-B cell and 9 myeloid cell lines) was measured quantitatively by in vitro regrowth assay. The growth of two non-T, non-B-cell lines (Reh, P30/OHK) and six myeloid cell lines (ML-1, U937, THP-1-0, P31/FUJ, P39/TSU, HEL) was found to be significantly inhibited by a 72hr treatment with PAC-4D. Although the levels of sensitivity of these cell lines to PAC-4D were different, it was common to all these cell lines that increasing the dose of PAC-4D did not augment the growth-inhibitory action above a certain level. Neither dose-dependent nor time-dependent growth-inhibitory action was observed, namely, exposure to 100U/ml of PAC-4D for 48hr was sufficient to exhibit maximum growth-inhibitory action. Furthermore U937 cells were found to become completely resistant to PAC-4D during a continuous 12-day exposure to it. This resistance, however, was lost on culture of the cells with PAC-4D-free growth medium for 15 days. These results suggested that some non-T, non-B acute lymphoblastic leukemias and acute myelogenous leukemias might show an initial response to PAC-4D therapy, but prolonged administration might induce resistance to PAC-4D rather than increase the anti-tumor effect.

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The colony-inhibitory effects of recombinant human tumor necrosis factor (rH-TNF) and recombinant human interferon-γ (rH-IFN-γ) were evaluated in four human lung cancer cell lines and their cisplatin-resistant sublines. The cell lines tested were PC-7 and PC-9 (adenocarcinoma), H69 and N231 (small cell lung cancer) and four cisplatin-resistant sublines, PC-7/1.0, PC-9/0.5, H69/0.2 and N231/0.2, which were 20.0, 7.1, 4.8 and 8.4 fold resistant to cisplatin, respectively, compared to the respective parental cell line in terms of IC₅₀ in a soft agar colony assay. All parental cell lines were resistant to rH-TNF and rH-IFN-γ, alone or in combination. However, two resistant sublines showed sensitivity to rH-TNF and rH-IFN-γ. Colony formation by PC-9/0.5 was significantly inhibited, in the absence or presence of cisplatin, by 10²U/ml of rH-TNF (<50% of control) and the inhibition was synergistic with that produced by 10³ or 10⁴U/ml of rH-IFN-γ. RH-IFN-γ inhibited the colony formation of H69/0.2 only at the highest concentration tested (10⁴U/ml) (<50% of control) and the combined effect with rH-TNF was additive. These results suggest that rH-TNF and rH-IFN-γ may have some potential in overcoming cisplatin resistance by virtue of collateral sensitivity.

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The tissue concentrations of doxorubicin (DOX, 15mg/kg, ip) and daunorubicin (DAU, 15mg /kg, ip) in mice were investigated to clarify their relationship to the cardiotoxicity, which is considered to be closely related to the lipid peroxide level in the heart. The largest Cmax of DOX after intraperitoneal injection was obtained in the liver and was equivalent to 1.8 times that in the heart. Elimination of DOX from the heart was not delayed. Nevertheless, the increases of lipid peroxide in the heart found in in vivo and in vitro experiments were considerably higher than those in the liver. Therefore, the cardiotoxicity of DOX can not be explained simply in terms of the relative concentrations of DOX in various tissues. On the other hand, experiments on the tissue concentration of DAU, which shows weaker cardiotoxicity than DOX, and on lipid peroxidation in vitro, suggested that the relative cardiotoxicities of DOX and DAU are directly related to their. relative concentrations in the mouse heart. We observed a metabolite of DOX, doxorubicinone, and an unknown metabolite of DAU in various tissues of mouse after drug injection, but these metabolites did not seem to be involved in the cardiotoxicities of DOX and DAU.

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