Analbuminemic rats were found to be highly susceptible to induction of renal tumors. Thirty- nine weeks after N-dimethylnitrosamine administration, the incidence of renal tumors and average weight of kidneys (including tumors) were 76.0% and 10.8±4.1g in male analbuminemic rats, respectively, whereas they were 37.1% and 3.5± 0.1g, respectively, in normal male SD rats.

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Two new human myeloid cell lines, PL-21 and KCL-22, were established from acute promyelocytic leukemia and chronic myelocytic leukemia, respectively. PL-21 was positive to peroxidase staining and differentiated into mature myeloid cells in vitro. KCL-22 had Ph¹ chromosomes and differentiated into granulocytes in vivo.

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Co-cultivation of lymphocytes from two Japanese monkeys, one of which was seropositive to adult T-cell leukemia (ATL)-associated antigens (ATLA), gave rise to a lymphoid cell line derived from the anti-ATLA negative monkey. This cell line harbors ATLA and type C virus particles identical in morphology to ATL virus.

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Mouse monoclonal antibodies, GIN-2, -7 and -14, against adult T-cell leukemia virus (ATLV) were prepared by a hybridoma procedure. These antibodies belonged to different subclasses of IgG, but they reacted similarly with both ATLV-specific polypeptides p19 and p28. In the reaction with monoclonal antibodies and various ATLV-producer cell lines, it was found that MT-2 and MT-2-related T-cell lines produced both p19 and p28, whereas MT-2-unrelated cell lines produced p19, but not p28.

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Unknown N-nitroso compounds were found in human urine of healthy volunteers by gas chromatography-thermal energy analysis. These compounds were identified as N-nitroso-L-thioproline and cis- and trans-N-nitroso-L-methylthioprolines by gas chromatography-mass spectrometry. The precursors of these new N-nitroso compounds may be formed by the reactions of L-cysteine with formaldehyde and acetaldehyde in the human body.

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A unique fibroblastoid cell line, BMA1, has been established from bone marrow cells of a DDY mouse transfected with adenovirus 5 DNA. BMA1 cells expressed adenoviral early antigens and BMA1-conditioned medium contained mouse granulocyte/macrophage colony-stimulating factor(s). The DNA transfection technique may be useful in establishing cell lines from bone marrow and analyzing the regulation of hemopoiesis.

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The mutagenic effects of 1-nitropyrene, and 1, 3- and, 8-dinitropyrenes on Chinese hamster V79 cells in the presence or absence of X-irradiated Syrian hamster embryo cells were examined. Without Syrian hamster embryo cells, 1, 3-dinitropyrene had weak mutagenic activity and 1, 8-dinitropyrene had strong, dose-related mutagenic activity. With Syrian hamster embryo cells, the mutagenicities of both 1, 3- and 1, 8-dinitropyrenes were appreciably increased. 1-Nitropyrene did not induce ouabain-resistant mutants at concentrations of up to 10μg/ml either with or without Syrian hamster embryo cells.

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Four groups of female Donryu rats were continuously given 400, 200, 100 or 0ppm N-isobutyl-N-nitrosourea (iso-BNU) in their drinking water, and were examined for the development of tumors. The incidence of digestive tract tumors was 25/28 (89%), 14/24 (58%), 6/25 (24%) and 0/17 (0%), in the 400, 200, 100 and 0ppm groups, respectively. The predominant type of digestive tract tumor wasangiogenic, and a few were of the epithelial type. A dose-effect relationship was clearly demonstrated not only in the incidence of digestive tract tumors but also in the average survival period of rats with these tumors.

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The effect of a protein-bound polysaccharide preparation, PS-K, on the induction of intestinal tumors by dimethyl hydrazine (DMH) was assessed in Wistar rats. One hundred and fifty-one rats were randomly divided into two groups. Seventy-two rats were treated with DMH alone and 79 rats were treated with DMH and PS-K. All animals were subjected to a sequential autopsy and all lesions within the gastro-intestinal tract were examined macroscopically and histologically. Tumor incidence in the DMH plus PS-K-treated group was significantly lower than that in the group treated with DMH alone. The most interesting histological finding was marked lymphoid infiltration in and around the tumors of the rats in the PS-K-treated group. The number of circulating lymphocytes dropped below the control range in the 25th and 35th weeks for the group treated with DMH alone, but the drop in the PS-K-treated group was smaller. Delayed-type hypersensitivity reaction to purified protein derivative was well maintained in the PS-K-treated rats. The most interesting findings in these experiments were differences in the serum blocking activities and serum immunosuppressive substance in these two groups; they were markedly reduced in the PS-K-treated rats. The present results may be explained in terms of competitive action of PS-K against the immunosuppressive factor produced by a tumor-bearing host.

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A simple and sensitive procedure is described for the quantitative determination of N-butyl-N-(3-carboxypropyl)nitrosamine (BCPN), the principal metabolite of the bladder carcinogen N-butyl-N-(4-hydroxybutyl)nitrosamine (BHBN-4), in urine. BCPN was analyzed by gas-liquid chromatography as its methyl ester after extraction with ethyl acetate from urine followed by esterification with diazomethane. Species and strain variations in susceptibility to bladder carcinogenesis induced by BHBN-4 are discussed in relation to the urinary excretions of BCPN; determined by the new procedure, after oral administration of the compound to rats (4 strains), mice, hamsters, and guinea pigs at a dose approximately equal to the daily dose used in chronic carcinogenicity assay.

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Spontaneous tumors in 296 male and 297 female F-344/DuCrj rats used as control groups in six carcinogenicity tests were tabulated and evaluated. In males the most frequent tumors were testicular interstitial cell tumors, followed by mammary fibromas and fibroadenomas, mononuclear cell leukemias, pheochromocytomas, C-cell adenomas of the thyroid, pituitary adenomas, preputial adenomas, neoplastic nodules of the liver, and lung adenomas. In contrast, in females pituitary adenomas, uterine endometrial stromal polyps, mammary fibroadenomas, mononuclear cell leukemias and C-cell adenomas were the most common. Various other tumors of almost all other organs and/or tissues were found, although their incidences were low. Variability in the rates of incidence in some tumors was observed in different control groups.

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A transplantable colorectal adenocarcinoma and the normal colonic mucosa derived from rats of ACI/N strain were digested successively with pronase, deoxyribonuclease, chondroitinase ABC, and heparitinase to obtain the corresponding glycopeptide fractions. The amino acid compositions of these two fractions suggested that the polypeptide backbones were quite similar. However, the electrostatic net charges of these fractions were shown to be different by cellulose acetate membrane electrophoresis, ion exchange chromatography, and measurement of sialic acid contents. The glycopeptide fraction derived from adenocarcinoma contained much greater quantities of less acidic glycopeptides than that derived from the normal colonic mucosa. The former exhibited much stronger blood group A and H activities than the latter. Moreover, the former reacted with Ricinus communis lectin I, whereas the latter did not react with this lectin. These results indicate that the carbohydrate structures of tumor sialoglycoproteins are different from those of the corresponding ones in the normal tissue from which the tumor has originated.

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Glyosphingolipids of neutrophils, lymphocytes and leukocytes from patients with various types of human leukemia [acute lymphoblastic (ALL), acute unclassified type (AUL), acute myeloblastic (AML), acute monocytic (AMoL), chronic myeloblastic (CML)] and the hypereosinophilic syndrome (HES) were analyzed chemically and immunochemically. No distinct difference was found in the molar ratio of lipid-bound sialic acid to lipid-bound phosphorus in these cells, but a low ratio of cholesterol to lipid-bound phosphorus was found in ALL (3 of 4 cases), AML, CML and AMoL (one of 2 cases). The predominant glycosphingolipid was ceramide dihexoside (CDH) in all cells analyzed, but the amount and the molar ratio of lipid-bound phosphorus to CDH were clearly different in different cell types, indicating that the molar ratio is a useful criterion in the classification of types of leukemia. In addition, molecular diversity of minor glycosphingolipid components was observed in various leukemic cells. Two of the neutral glycosphingolipids in AMoL were tentatively identified as asialo GM1 and Forssman glycolipids by comparing their mobilities on thin-layer chromatography with those of standard glycolipids and by observing the formation of precipitin lines on a double diffusion agar plate with anti-asialo GM1 and anti-Forssman antibodies.

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The frequency of intramucosal cysts underneath focal (elevated) neoplastic lesions (which include epithelial dysplasia and early gastric cancer) was recorded in eighty-eight resected stomachs. The mean cyst index (i.e. the amount of intramucosal cysts divided by the amount-in cm-of gastric mucosa analyzed) underneath such lesions was 30.9 (SD±16.9), while in adjacent normal-looking gastric mucosa it was 9.4 (SD±8). The difference was significant (P<0.001). The cyst index was highest in female patients, underneath focal (elevated) dysplasias. Previous work indicated that the normal-looking gastric mucosa in Japanese patients with peptic ulcer disease was 3.4 (SD±6.6). The present findings appear to suggest that cystic formation in gastric mucosa may herald an early neoplastic growth at the luminal aspect in some of the cysts.

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The antitumor activity of lectins were examined by studying the effect of lectins on the in vivo growth of murine tumors. The lectin wheat germ agglutinin (WGA) inhibited the growth of MM46 ascites tumors in vivo as effectively as syngeneic antitumor antiserum, but other lectins were not effective in vivo. WGA also had an inhibitory effect on other syngeneic and allogeneic ascites tumors, a fact which is consistent with its wide range of target tumor cells demonstrated in the in vitro cytolysis system. These results suggest that WGA is a better antitumor agent than other plant lectins. The therapeutic effect of WGA might be, in part, due to its ligand activity between tumor cells and macrophages that induces lectin-dependent macrophage-mediated cytolysis, in addition to its direct cytostatic activity.

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Several lectins from marine animals, such as Balanus roseus hemagglutinin, B. balanoides hemagglutinin, Tetraclita squamosa japonica hemagglutinin and Aplysia kurodai agglutinin, were tested for induction of tumor lysis mediated by macrophages. Among them, B. roseus and B. balanoides lectins agglutinated several murine tumor cells and induced binding of macrophages to tumor cells. Binding of these cells was inhibited by galacturonic acid, suggesting that carbohydrate moieties on the cell membrane of the two types of cells are recognized by these lectins. These lectins did not induce tumor lysis in co-operation with various macrophages, but after inactivation of tumor cells with glutaraldehyde, they induced extensive lysis of target cells in the presence of marophages. B. roseus lectin was also effective in vivo. These results suggest that tumor cells can be recognized via carbohydrate moieties on the cell membrane as well as tumor-associated antigen and that some animal lectins participate in macrophage-mediated cytolysis and tumor rejection.

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The immunogenicity of concanavalin A (Con A)-treated tumor cells was examined in 4 histologically distinct tumors originated from 3 different strains of mice. In all of these tumors, Con A-treated tumor cells induced stronger tumor-specific immunity than Con A-free tumor cells as determined from the survival of sensitized mice after live tumor cell inoculation. However, Con A-induced potentiation of tumor cell immunogenicity was dependent on 2 experimental factors associated with the tumor cell vaccine preparation. One was the agent used for inhibiting in vivo transplantability of tumor cells and the other was the dose of tumor cell vaccine. In Meth A, Meth 1, and Lewis lung tumors, glutaraldehyde and Con A-treated cells, but not mitomycin C and Con A-treated cells, induced an enhanced immune resistance, whereas in L1210 leukemia, mitomycin C was more beneficial than glutaraldehyde. Higher doses of Con A-treated tumor cells (up to 10⁷) induced stronger immune resistance in Meth A and Meth 1 tumors, as was the case with L1210 tumor, whereas in Lewis lung tumor there existed an optimal vaccine dose. These results indicate a tumor species dependence of pretreatment agents and cell vaccine doses in inducing immune resistance, although Con A potentiated the immunogenicity of all the tumors tested.

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The natural killer (NK) activity of peripheral blood lymphocytes of gastric cancer patients in all pathological stages from I to IV was less than that of healthy persons or patients with benign diseases. However, there was no significant difference in the antibody-dependent cellular cytotoxicity (ADCC) of cancer patients and healthy individuals, and the correlation between NK and ADCC activities was somewhat lower in cancer patients than in healthy persons. The peripheral blood lymphocytes from digesitive tract cancer patients were divided into two fractions on the basis of low affinity erythrocyte (E)-rosette formation. The NK activity of the low affinity E-receptor negative fraction from cancer patients was significantly lower than that of the same fraction from healthy persons. The binding of lymphocytes with target cells is considered to be a first step of natural killing. In cancer patients, there was a decrease in the target-binding capacity of lymphocytes in both fractions. Therefore, the decreased NK activity of cancer patients appeared to be related to a reduced target-binding capacity.

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Spontaneous radiation-induced or radiation leukemia virus (RadLV)-induced leukemias were treated in vivo in an early-to-advanced lymphomatous state by using 1, 3-bis(2-chloroethyl)-1-nitrosouea (BCNU). BCNU was given at various times after the tumor induction procedure. Treatment was based upon the findings that the RadLV-derived transplant lymphosarcoma (LSA) tumor was cured with a frequency of>90% and a tumor-resistant state was produced by tumor cure. Death from RadLV lymphomas which had been initiated in newborn C57BL mice by ip injection of RadLV was scored in untreated or BCNU-treated mice. At 150 days after initiation, control mice had developed extensive thymic lymphomas, whereas BCNU-treated mice had developed less extensive tumors, had a reduced incidence by -50% and had an increased survival time. Similar findings were noted for the radiation (190 rad of cobalt-60 gamma rays weekly×4)-induced thymic lymphoma-bearing C57BL mice. Injection of 10⁶ living LSA cells before BCNU treatment greatly decreased the tumor load for all groups treated but also shortened the mean survival time. Challenge of long-term survivors with small numbers of viable LSA tumor cells did not identify any host resistance of the survivors to the transplant LSA tumor and indicated that the induction procedure, whether RadLV or radiation, was strongly immunosuppressive.

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The effects of cyclophosphamide (CPA) and its active form, 4-hydroperoxy-CPA, against human breast carcinoma transplanted into nude mice (BALB/c nu/nu) were evaluated in terms of the decreases of hepatic drug-metabolizing enzymes in nude mice. A human breast carcinoma, MX-1, was implanted into the subcutaneous tissue of nude mice and a drug was administered intravenously once at a dose of 0.05, 0.1 or 0.15mmol/kg, 1 or 3 weeks after tumor inoculation. 4-Hydroperoxy-CPA was more effective than CPA as regards inhibition of tumor growth, and the difference in effect was greater when the drugs were administered 3 weeks after tumor inoculation. The activity of CPA was depressed by the decrease of the hepatic drug-metabolizing enzymes in proportion to the tumor-bearing period. Therefore, the effects of masked derivatives of CPA may correlate with the changes in drugmetabolizing activities of tumor-bearing mice. The human tumor xenografts-nude mice system is considered to be suitable for chemosensitivity tests with masked compounds.

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In order to clarify differences in mode of action between N⁴-behenoyl-1-β-D-arabinofuranosylcytosine (behenoyl-ara-C) and 1-β-D-arabinofuranosylcytosine (ara-C), comparative studies on both agents were undertaken. When human erythrocytes incubated with behenoyl-ara-C[acyl-1-¹⁴C] were fractionated into stroma and stroma-free lysate, a marked accumulation of radioactivity in stroma was observed. In contrast, ara-C[cytosine-2-¹⁴C] was rapidly incorporated into the stroma-free lysate. Thin-layer chromatography of the extracts of L1210 cells incubated with behenoyl-ara-C[acyl-1-¹⁴C] or behenoyl-ara-C[cytosine-2-¹⁴C] at 37° for 60min revealed that most of the incorporated drug remained as unmetabolized behenoyl-ara-C. After incubation of 20μM behenoyl-ara-C or ara-C with L1210 cells at 37°for 60min, subcellular fractionation of the cell suspension was performed; behenoyl-ara-C was accumulated markedly in the membrane, mitochondria and microsome fractions. In contrast, most of the ara-C was found in the 105, 000g supernatant fraction. The accumulation of behenoyl-ara-C in membrane structures may result from the lipophilic nature of the agent, which may have a prolonged inhibitory action on leukemic cell proliferation.

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The mode of action of the direct antiproliferative effect of human fibroblast interferon (HuIFN-β) on tumor cells was analyzed in vitro with a human malignant melanoma cell line, HMV-1. HuIFN-β inhibited the growth of HMV-1 cells in a time-dependent fashion (50%-inhibition concentration: <50IU/ml). Its action was cytostatic. DNA synthesis was inhibited before the antitumor effect appeared, and apparent dose-dependent inhibitions of RNA and protein synthesis were induced. An increase of cell protein content was seen with the occurrence of growth inhibition. Increase of cell volume and prolongation of doubling time were seen from the second day after the addition, which corresponded well with the effects of HuIFN-β on the cell cycle (the accumulation of cells in the S phase). These results indicate that the direct antitumor effect of HuIFN-β on HMV-1 cells may result from cell cycle retardation mediated through obstruction of DNA synthesis.

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