Cytotoxicity of sera from cervical cancer patients mediated by peripheral blood lymphocytes against cervical cancer cell lines was studied. Percentages of reduction of target QG-K by sera from healthy controls and cervical cancer patients were 4.7% and 32.0%, respectively. Sera from patients with pulmonary tuberculosis also exhibited antibody-dependent cell-mediated cytotoxicity (ADCC) against the cells. Especially, high degree of activity was observed in the sera from patients with stage IV and the activity decreased to the control level in recurrent cases. The titer of the sera varied against different target cell lines, and the activity was found even at 1:10⁴ dilution of the sera. These activities remained after absorption of the serum with other tumor cells. Complement-dependent cytotoxic activity of the sera was not observed. The meaning of the cytotoxic activity of the sera from cervical cancer patients was discussed.

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Cytotoxicity of peripheral blood lymphocytes against allogeneic target cells of bronchogenic carcinoma was examined by a microcytotoxicity test before, during, and after radiotherapy in primary lung cancer patients. Before the treatment, cytotoxicity was depressed only slightly in patients in stage III and strikingly in those in stage IV, as compared to the values in patients at earlier stages of lung cancer such as stages I and II. Local irradiation scarcely affected cytotoxicity at stages II and III, but augmented remarkably at stage IV. The number of peripheral blood lymphocytes decreased profoundly during and after radiotherapy in all cases of stages II, III, and IV. Although radiotherapy exhibited various effects on the cytotoxic activity of lymphocytes and the number of peripheral blood lymphocytes, only the cytotoxic activity at the end of radiotherapy correlated well with the reduction in tumor size.

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Alkaline phosphatases, which had a unique electrophoretic mobility on polyacrylamide gel electrophoresis, were found in hepatic tissue of a patient with liver cirrhosis. Enzymic and immunological properties of the enzymes examined on electropherogram were similar to those of a fetal intestinal-type alkaline phosphatase in hepatoma with respect to sensitivity to amino acids, heat stability, sensitivity to sodium dodecyl sulfate, and reactivity to antiintestinal alkaline phosphatase antiserum. The enzymes seem to be a variant of a fetal intestinal alkaline phosphatase.
The significance of occurrence of the enzymes in cirrhotic liver is discussed.

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An attempt was made to enhance the plating efficiency of normal human diploid skin fibroblasts by testing various culture media. Eagle's minimum essential medium (MEM), Dulbecco's MEM, Williams' medium E, DM-160, McCoy's 5a, RPMI-1640, Ham's F12, MCDB 102, Waymouth's MB 752/1, and L-15 synthetic media were examined. L-15 was the best among them. Plating efficiency with L-15 supplemented with 20% calf serum ranged from 20 to 60%. Plating efficiency decreased in the order of Eagle's MEM, Dulbecco's MEM, DM-160, and Ham's F12. Waymouth's MB 752/1, MCDB 102, McCoy's 5a, RPMI-1640, and Williams' medium E were not favorable for high plating efficiency. Although L-15 gave the best plating efficiency and Waymouth's MB 752/1 the worst, the Waymouth medium supported better cell proliferation than L-15, when cell proliferation was compared in mass culture with these two media. It was concluded that L-15 was better suited for culture of small number of cells, and especially for work involving colony formation.

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Carcinogenicity of phenacetin was tested using Sprague-Dawley rats. Two groups of animals containing 50 males and 50 females per group were fed respectively with 2.5% and 1.25% phenacetin diet for 18 months and fed thereafter with basal diet for 6 months. Control animals containing 65 males and 65 females were fed with basal diet for 24 months. Animals surviving more than 24 months were regarded as effective animals and killed. Rats that died of tumor development within 24 months were also regarded as effective animals. Every organ from the killed and dead animals was fixed in 10% formaldehyde solution and examined histopathologically. Effective number of rats was 27 males and 27 females in 2.5% phenacetin feeding group, and 22 males and 25 females in 1.25% phenacetin feeding group. In control group, 19 males and 25 females were effective. Neoplasms including spontaneous tumors were detected in 26 out of 27 males (96.3%) and 21 out of 27 females (77.8%) of 2.5% phenacetin feeding group, and in 20 out of 22 males (90.9%) and 19 out of 25 females (76.0%) of 1.25% phenacetin feeding group. In control group, 1 out of 19 males (5.3%) and 6 out of 25 females (24.0%) showed spontaneous tumor development. Histopathologically, carcinomas of the nasal cavity, such as adenocarcinoma, squamous cell carcinoma, and transitional cell carcinoma, and the urinary passage, as renal cell carcinoma of the kidney pelvis, and transitional cell carcinoma of the urinary bladder, were most conspicuous, suggesting the target organs of phenacetin carcinogenesis. Males showed higher tumor incidence compared to females. The higher the concentration of phenacetin given, higher incidence of tumors was observed.

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Nicotinamide, a specific inhibitor of poly (ADP-ribose) synthetase, was found to be a moderate inducer of hemoglobin synthesis in Friend erythroid leukemia cells (FLC). Therefore, the effect of other inducers, such as dimethyl sulfoxide (DMSO), hexamethylene-bisacetamide (HMBA), and butyrate, on poly (ADP-ribose) synthesis was examined. The extent of poly (ADP-ribose) synthesis in nuclei of FLC treated with DMSO or HMBA began to decrease before many phenotypic changes including hemoglobin production and reached 30∼50% of the level of nontreated control when the cells enter the stationary phase. FLC variants unresponsive to HMBA or DMSO did not exhibit as low an activity of poly (ADP-ribose) synthesis as their parent cells did by treatment with these inducers. In contrast, butyrate stimulated poly (ADP-ribose) synthesis transiently but distinctly (about 50%) at an early stage of culture (6∼24hr), but suppressed it at a later stage. Neither the cell growth nor degradation of poly (ADP-ribose) is correlated with the effect of inducers. These results suggest that the level of poly (ADP-ribose) synthesis is correlated with the differentiation of FLC.

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Thymidine kinase, dTMP kinase, and DNA polymerase activities were determined in cell lines of AH hepatoma, L1210 leukemia, and Yoshida sarcoma that were sensitive and resistant to 5-fluorouracil (5-FU). It was found that the levels of these enzymes in tumor cells were not consistently related to the property of sensitivity to 5-FU.
A marked difference was observed between sensitive and resistant cell lines of L1210 leukemia and Yoshida sarcoma in the uptake of labeled 5-FU into the acid-soluble, nucleotide, and RNA fractions, the rate of incorporation of 5-FU into these fractions being 3 to 5 times greater in sensitive tumor cells than in resistant tumor cells.
The radioactivities in the acid-soluble fractions of AH44 (sensitive) and AH109A (resistant) were similar after incubation of these cells with labeled 5-FU in vitro. However, a smaller volume of ascites and lower cell number were observed in AH44 (sensitive)-bearing rats than in AH109A (resistant)-bearing rats.
These in vivo results indicate that the 5-FU injected intraperitoneally was diluted by ascites more in AH109A (resistant)-bearing rats than in AH44 (sensitive)-bearing rats.

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Administration of oil-attached Nocardia rubra cell-wall skeleton (N. rubra-CWS) was evaluated for the effect on the induction of lung cancers in ACI/N rats. Lung cancers were induced by 15 weekly intratracheal instillations of 3mg benzo [a]-pyrene with 3mg ferric oxide. After the 10th instillation of the carcinogen, rats received seven subcutaneous injections of 100μg of N. rubra-CWS at 2-week intervals. In the observation period of 56 weeks, the cumulative incidence of lung cancer was 71.4% in the control group and 48.0% in the N. rubra-CWS treated group. The latent period of tumor induction was prolonged in the group treated with N. rubra-CWS.

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Sera of Japanese women with breast cancer and without disorders of the mammary gland gave positive immunofluorescence reaction with mouse mammary tumor cells (MMT cells) producing type-A and type-B virus particles. Among these, 55.1% (49/89) of the sera from patients with breast cancer reacted with MMT cells, whereas 60% (3/5) of the sera from patients with benign mammary hyperplasia (2 of 3 mastopathies, 1 of 2 cystosarcoma phyllodes), 30% (12/40) of the patients with other than breast cancer (4 of 14 stomach cancers, 6 of 20 uterine cancers, 1 of 3 lung cancers, 1 of 3 rectum cancers), and 26.5% (18/68) of the sera from apparently healthy women showed positive reaction with MMT cells. Distribution pattern of the specific fluorescence obtained with positive human sera was similar to that resulting from the reaction with rabbit antiserum to type-A particles. Results of absorption studies and blocking tests also suggested that the reaction was due to the antigenic components common to type-A and -B particles. Furthermore, the results of membrane immunofluorescence tests suggested that some human sera also react with envelope antigen (s) of B particles. Antinuclear antibody was found in sera from 15.7% (14/89) of breast cancer patients by the indirect immunofluorescence tests, and antibody cross-reacting with mouse mammary tumor virus was not found in most of the sera from breast cancer patients who have antinuclear antibody.

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The pretreatment of B6D2F₁ mice with OK-432 alone was not so effective to retard the growth of L1210 leukemia, and the leukemic cell was poorly immunogenic in the mice. However, when OK-432 was given by ip injection in combination with mitomycin-C-treated L1210 cells, the growth of L1210 leukemia was significantly retarded and some mice did not "take" the leukemia.
Meanwhile, BCG was also effective in this respect. Differences in properties as an immunopotentiator between OK-432 and BCG were suggested by the experiments of in vitro cytotoxicity test of spleen cells.

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Effect of 1, 1, 1-trichloropropene 2, 3-oxide and 7, 8-benzoflavone on benzo[a]-pyrene and 3-methylcholanthrene carcinogenesis in the subcutaneous tissues of C3H/He and DBA/2 mice was examined. By a single application of the carcinogen, the incidence of fibrosarcoma was higher and the latency of tumorigenesis was shorter in C3H/He mice than in DBA/2 mice. Treatment with 1, 1, 1-trichloropropene 2, 3-oxide increased the incidence of fibrosarcoma in DBA/2 mice, but decreased the rate in C3H/He mice, when benzo [a] pyrene was used as a carcinogen. On the other hand, in 3-methylcholanthrene carcinogenesis, no effect of the oxide on the tumor incidence was observed. By the simultaneous application of 7, 8-benzoflavone with benzo [a] pyrene, the tumor incidence increased, but not so significantly in DBA/2 mice, compared to that treated with benzo [a] pyrene alone, and no appreciable effect was observed in C3H/He mice treated with benzo [a] pyrene, and in both strains of mice with 3-methyl-cholanthrene. The relationship between the activity of arylhydrocarbon hydroxylase and the change in polycyclic hydrocarbon carcinogenesis is briefly discussed.

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To define the relationship between cell-mediated immunity and responses to chemotherapy or prognosis, delayed cutaneous hypersensitivity, E- and active E-rosette tests, and mitogenic responses of lymphocytes were examined in 15 patients with acute nonlymphocytic leukemia. Its results indicated that cellmediated immunity before remission induction chemotherapy did not correlate with the outcome of treatment. In contrast, delayed cutaneous hypersensitivity and mitogenic response tested after remission induction correlated with therepeutic effect. Further a "rebound" or overshoot of phytohemagglutinin responsiveness was observed in patients achieving remission. Serial studies on cell-mediated immunity may be useful for predicting therapeutic efficacy and prognosis in patients with acute leukemia.

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While lymphocytes from normal persons exhibited cytotoxicity against T24 cells, prednisolone at a concentration as low as 10^-7M significantly inhibited the cytotoxic reactions and almost complete inhibition was observed at the concentration of 10^-4M. The inhibitory effect was also shown to be progressively decreased when prednisolone was added during the cytotoxicity assay of normal human lymphocytes. These findings suggest that the cytotoxicity of lymphocytes from normal persons can be suppressed by immunosuppressive effect of prednisolone and that the early phase in the cytotoxic reaction is considerably sensitive to prednisolone.

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Prophylactic effect of repeated intravenous administrations of oil-attached BCG cell-wall skeleton (BCG-CWS) on the induction of tumor by 7, 12-dimethylbenz[a]anthracene (DMBA) was investigated in various strains of mice.
The subcutaneous injection of DMBA emulsified in oil induced squamous cell carcinoma in almost all of the strains of mice. Treatment of C57BL/6, BALB/c, and ddO strains with BCG-CWS with appropriate route and timing resulted in the retardation of DMBA-induced tumor development manifested by a prolonged latent period of tumor outgrowth. In contrast, the same BCG-CWS treatment of C3H/He and BTK mice was incapable in preventing such DMBA-induced carcinogenesis. Thus, the treatment with BCG-CWS was effective for preventing the DMBA-induced carcinogenesis in certain strains of mice, but the effectiveness varied depending on the strain.
The implication of such a strain variation of the BCG-CWS effect on the prophylaxis of chemical carcinogenesis was discussed in the context of the differences in the magnitude of immunopotentiation of the host by BCG-CWS.

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Thirty-five primary tumors and 14 metastatic lymph nodes from stomach cancer patients were transplanted in nude mice. Successful transplantation was obtained in 16 cases of primary tumors and 5 cases of metastatic lymph nodes. The tumors growing in the nude mouse displayed histological features similar to those of the original human tumors. Eight cases of the successful takes have been maintained by consecutive transplantation. Nine cases have been stored in liquid nitrogen.
The transplanted tumors were observed to spread to the distant lymph nodes in 3 cases out of the 17 successive transplants.

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Canine transmissible sarcoma has been maintained since 1967 as an allogeneic tumor cell line. The tumor originated from a naturally occurring venereal sarcoma in a 7-year-old female dog, Hokkaido-ken. The animals used for the serial transfer from 1st to 50th passages were 1- to 8-month-old puppies. Histological features and karyological characteristics of the tumor were not modified by the passive transfer. In most of the dogs, the palpable growths in the subcutaneous tissues were present at approximately the same time. These findings suggest that the sarcoma is an only established tumor cell line in dogs.

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Addition of a potent tumor promoter, 12-O-tetradecanoylphorbol 13-acetate (TPA), to mouse myeloid leukemia line cells (M1) in suspension cultures inhibited both functional and morphological differentiation of the cells induced by dexamethasone or protein inducer. A positive correlation was found between the tumor-promoting activities of several plant diterpenes and their inhibition of cell differentiation. The inhibition of cell differentiation by TPA was reversible and was unrelated to its cytotoxic action.

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Sequential quantitative analyses were made of hyperplastic liver nodules induced by treating male Fischer rats first with diethylnitrosamine (DEN) and then with N-(2-fluorenyl)acetamide (2-FAA), α-isomer of 1, 2, 3, 4, 5, 6-hexachlorocyclohexane (α-BHC), or Phenobarbital. The test rats were injected ip with 200mg/kg body weight of DEN, given basal diet containing 2-FAA, α-BHC, or phenobarbital from week 2 to 12, and subjected to partial hepatectomy at the end of week 3. From week 6, significantly higher percentage areas of hyperplastic nodules per unit area of liver were found in all experimental groups than in the control group. From week 8, significantly more hyperplastic nodules were found in all experimental groups than in the control group. Continuous administration of α-BHC or phenobarbital for 12 weeks with partial hepatectomy did not induce hyperplastic nodules. Continuous administration of 2-FAA did induce hyperplastic nodules, but the percentage areas and number of these nodules were significantly lower than in rats injected with DEN and then given 2-FAA orally, with partial hepatectomy.

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Using the chromium release technique, the cytolysis of murine leukemia virus-infected (xenogenized) tumor cells was studied with spleen cells from tumor-bearing rats. The cytolysis was found to be higher in xenogenized tumor cells than in non-xenogenized tumor cells. Tumor specificity of increased sensitivity was found.

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