Sarcoma-180 Cells Research Articles

Synthesis of (R- and (S)-1-[[2-Hydroxy-1-(aminomethyl)ethoxy]methyl]-5- benzyluracil, Potent Inhibitors of Uridine Phosphorylase

Abstract Optically pure (R)- and (S)-1-[[2-hydroxy-1-(aminomethyl) ethoxy]methyl]-5-benzyluracil [(R)-AHPBU and (S)-AHPBU, respectively], two potent uridine phosphorylase inhibitors, have been synthesized via multi-step syntheses starting from independent chiral compounds. The activity of (R)-AHPBU, (S)-AHPBU, and (R,S-AHPBU which have been previously synthesized, on the inhibition of uridine phosphorylase from Sarcoma-180 cells has been studied and compared. The K. values for (R,S)-, (R)- and (S)-AHPBU were determined to be 15·2.3, 17·2.7 and 16·2.0 nM, respectively. This indicates that (R) and (S) optical enantiomers have the same affinity for binding to uridine phosphorylase. These acyclic pyrimidine amino nucleoside analogues represent a new class of potent uridine phosphorylase inhibitors, which has a bulky hydrophobic substituent at the 5-position in the uracil base, yet has remarkably high water solubility.

Natural Killer(NK) cell activity of mice treated with herpes simplex virus type-2, sarcoma-180 cells, cyclosporine and cimetidine.

NK cell activity in beige mice was compared with that in control BALB/c mice and BALB/c mice treated with HSV-2, sarcoma-180 cells, cimetidine and cyclosporine. The NK cell activity in five week-old beige mice was lower than in three week-old BALB/c mice. The NK cell activity in eight week-old BALB/c mice was four to eleven times the activity in BALB/c mice that were three weeks of age. BALB/c mice, injected with 2 mg cyclosporine or 10(7) plaque forming units (PFU) of HSV-2, had decreased NK cell activity. BALB/c mice, injected with 2 mg cimetidine, 10(6) sarcoma-180 cells and 6.0 x 10(5) PFU of HSV-2, had high NK cell activities compared with the control mice. Cimetidine further increased the effect of HSV-2 or sarcoma-180 on NK activity. BALB/c mice that received a transfer of less than 10(7) NK cells developed a high resistance against viral infections, but mice that received more than 10(7) transferred NK cells had a lower resistance to viral infections than the control mice or mice receiving less than 10(7) NK cells.

Antitumor property of the active principle of Jawaharene

The active principle of Jawaharene (JF 1), a mixture of fatty acids, was studied for antitumor properties against Ehrlich Ascites Carcinoma (EAC) and Sarcoma-180 (S-180). Using a Warburg respirometer, respiratory O 2-uptake by EAC and S-180 cells was completely inhibited by JF 1 at the dose of 100 μg ml −1. The in vitro cytotoxicity assay revealed that 100% of JF 1-treated tumor cells were killed within 60 min incubation. In vivo, EAC and S-180 ascites tumor development was completely inhibited by JF 1 at the dose of 5 mg mouse −1 day −1 × 8.

Effects of platinum-thymine on mitosis of sarcoma-180 ascites cells in vitro: ultrastructural study.

Electron microscopic studies of sarcoma-180 ascites cells, which were treated with platinum-thymine at a concentration of 60 micrograms/ml, showed mitotic inhibition. The drug clumped the chromosomes into a compact mass at the center of the cell preventing them from separating during mitosis. Prolonged treatment depolymerized spindle fibers and cytoplasmic microfilament. Degraded cytoplasmic organelles crowded around the clumped chromosomes. Disintegrating and casting off of cortical cytoplasm was apparent. Platinum-thymine-treated cells did not resume mitosis irrespective of the amount of time they were allowed to remain in fresh medium. Perturbation of mitotic machinery by platinum-thymine inhibited the mitotic process and caused eventual demise of the cancer cells.

Characterization of DNA binding proteins released from sarcoma-180 chromatin during brief digestion with DNase-I.

Brief digestion of sarcoma-180 chromatin (less than or equal to 5%) by pancreatic DNase-I releases six non-histone proteins with Mol. wt. 21.5K, 26K, 72.5K, 77K, 90K and 120K dalton and pI values ranging from 4.7 to 12.4. The protein of Mol. wt. 77K dalton was obtained at high alkaline range of pH = 12.4. The antibodies against these proteins induce dose dependent inhibition in transcription of native chromatin. Results suggest a role of these proteins in positive control of gene transcription in sarcoma-180 cells.

Effect of several d-morphinans on ascites tumors in mice.

Dextromethorphan and its analogues (DM 16, DM 34, DM 72, DM 75 and DM 96) were examined for their effect on Ehrlich ascites carcinoma or ascites sarcoma-180 in female mice of the ddY strain. The suspension of Ehrlich carcinoma cells or sarcoma-180 cells was prepared from mice at 10 days after i.p. inoculation of the cells, using Hanks' balanced salt solution, and the cell suspension was inoculated i.p. into mice (2 X 10(6) viable cells/mouse). The chemicals dissolved in physiological saline containing 5% HCO-60 were then injected i.p. into the mice once daily for 5 successive days (5-40 mg/kg/day). In addition, mice given the tumor cells were treated with the saline containing 5% HCO-60 alone for 5 days (untreated mice). In groups of mice bearing Ehrlich ascites carcinoma or ascites sarcoma-180, the mean survival time of mice treated with 20-40 mg/kg/day of DM 96 was more than twice that of the corresponding untreated mice. The mean survival time of mice treated with 20 mg/kg/day of DM 96 was also longer than that of mice treated with 40 mg/kg/day of the other chemicals, irrespective of the ascites tumors. Concerning these survival times, the LD50 (i.p.) of DM 96 in mice differed slightly from that of other chemicals (88 mg/kg and 77-106 mg/kg). These results indicate that DM 96 is more active than the other chemicals against the ascites tumors in mice.

Fluoropyrimidine-mediated changes in small nuclear RNA.

Studies were completed to examine the effects of the antineoplastic agent 5-Fluorouridine (FUrd) on the metabolism of the small molecular weight nuclear RNA (snRNA). Cultured Sarcoma-180 murine tumor cells were exposed to FUrd concurrent with [3H]cytidine for 6 h, the drug was removed, and the RNA was isolated at 0, 24, or 48 h following the drug treatment. The results of these studies demonstrated that FUrd produced three dose-dependent changes in snRNA metabolism. The electrophoretic migration of the U4 and U6 snRNA was altered in nondenaturing 10% polyacrylamide slab gels. These results were not observed in denaturing gels or when RNA was extracted at temperatures exceeding 25 degrees C, suggesting that the incorporation of 5-fluorouracil induced secondary structural changes in these RNA. A dose- and time-dependent selective reduction in the turnover of the U1 snRNA synthesized in the presence of FUrd was observed as well, with levels over 100% higher than control cells at 48 h after exposure to 10 microM FUrd. These changes in snRNA metabolism may contribute to the reported alterations in large molecular weight RNA metabolism that also result in fluoropyrimidine-treated cells, or to cytotoxicity.

Biological activities of mouse haptoglobin elicited by administration of an antitumor polysaccharide, PSK

The concentration of mouse haptoglobin in serum was increased by administration of an antitumor polysaccharide, PSK. The administration of the purified mouse haptoglobin inhibited the growth of Sarcoma-180 cells implanted in ICR mice. Furthermore, this glycoprotein enhanced macrophage activitiesin vitro, as judged from the cytostatic and cytolytic activities, glycose consumption, O2-production, and interleukin-1 production of macrophages. In addition, mouse haptoglobin enhanced the cytolytic effect of cytotoxic T-lymphocytes. These results suggested that haptoglobin has a role in restoring or enhancing the resistance of the host against tumors.

Cytotoxicity of Cortisone-Resistant Lymphocytes from Mice Treated with a Group A Streptococcus or Freund’s Complete Adjuvant against Tumor Cells

The cortisone-resistant lymphocytes (CR lymphocytes) of mice treated with a group A streptococcus, Su strain, or Freund's complete adjuvant (FCA) were examined for their cytotoxicity on Ehrlich carcinoma cells and sarcoma-180 cells. Female mice of the ddY strain, 7-8 weeks of age, were injected subcutaneously with streptococci or FCA in emulsion, and they were killed 14 days later. To obtain CR lymphocytes, mice treated with and without agents were injected intraperitoneally with hydrocortisone acetate (125 mg/kg) 2 days before killing. Tumor cells and CR lymphocytes from thymus, spleen or mesenteric lymph node were suspended in Hanks balanced salt solution supplemented with 2% bovine albumin. The cytotoxicity of CR lymphocytes on tumor cells was examined by the Winn test: Tumor growth was observed in mice inoculated s.c. with the mixture of tumor cells (T) and CR lymphocytes (L) at a T/L ratio of 1/10 (10(6) tumor cells/mouse). The mesenteric and thymic CR lymphocytes of mice treated with streptococci or FCA were more effective than the corresponding lymphocytes of untreated mice in suppressing the tumor growth in animals given the cell mixture. This suggests that the treatment of mice with streptococci or FCA results in an enhancement in the cytotoxicity of mesenteric and thymic CR lymphocytes against the tumor cells.

Cytoflowmetric analysis of binding pattern and a new binding kinetic constant for an enterovirus and receptor.

A bovine enterovirus (MZ-468) was adsorbed to Sarcoma-180 cells at 4°C. The cells were reacted with FITC-labeled IgG antibody for the virus and were studied with a laser fluorescence cytoflowmeter. From the histograms, binding-saturation of receptors was observed and the dissociation constant (Kd) was estimated. A new binding rate constant was deduced from the change of the ratio of virus-adsorbed cells against time and virus concentration. The constant has a dimensions of (cm3/min·receptor-site) and probably represents the receptor class on a cell with the highest rate constant.

Rifampicin-induced chromatid exchanges in Sarcoma-180 mouse ascites cells

Sarcoma-180 ascites tumour-bearing male mice were injected i.p. with single dose of 0.5, 2.5, 5 mg/kg body wt. of rifampicin. Cells were sampled for mitotic chromosome analysis 4, 16 or 24 h after treatment. The maximal yield of chromatid-type aberrations induced was found 24 h after treatment with 5 mg/kg of rifampicin. More than 60% of the cells carried at least one chromatid exchange. The majority of these were exchanges derived from breaks in the centromeric heterochromatin.

Acriflavine-induced surface changes in three tumor cell types and differential sensitivity to lectins.

Phytohemagglutinin (PHA) and concanavalin A (Con A) were used as probes to detect changes in the cell surface of Dalton's lymphoma, sarcoma-180 and Ehrlich's carcinoma after short in vitro exposure to acriflavine. Dye-treated cells showed enhancement of agglutination both by PHA and Con A, and such enhancement was found to be dependent on the time of exposure and concentration of acriflavine. However, PHA-induced percent agglutination seemed to be much higher than that of Con A among the 3 cell types. There were also marked differences among the 3 cell types in order of their sensitivity to lectin-mediated agglutination. The strength of the response was greater in lymphoma to both PHA and Con A than that of sarcoma-180 and carcinoma cells, which appeared to be most resistant. Acriflavine, which is known as an intercalative agent with DNA, induces cell surface changes by promoting lectin-mediated cellular agglutination.

Effect of cisplatin on the plasma membrane phosphatase activities in ascites sarcoma-180 cells: a cytochemical study.

To study the effects of cisplatin [cis-dichlorodiammine-platinum (II)] on tumor cells in the presence or absence of the immune system, animals with ascites sarcoma-180 tumor burden were treated with therapeutic dose levels (9 mg/kg). Similarly, ascites sarcoma-180 cells were maintained in tissue culture media containing the same levels of the drug. Cell samples were taken from the animals at 12-hr intervals for 3 days, whereas samples were drawn from the tissue cultures at 15-, 30-, 45-, and 60-min and at 2-, 3-, 4-, and 5-hr intervals. Treated and untreated cells from in vitro and in vivo experiments, when checked for alkaline phosphatase, 5'-nucleotidase, Ca2+-ATPase, and Na+-K+-ATPase, show a gradual decrease in activity on the plasma membrane. It takes about 60 min for inactivation of any enzyme in vitro, whereas it takes 2 days in in vivo experiments. Quantitative analysis show alkaline phosphatase activity drops from 9.7 to 4.9 nmol in just 15 min, and drops further to 0.79 nmol after 2 hr. Inactivation of various plasma membrane enzymes, resulting in permeability changes, is probably responsible for cell death.

Effect of the centrally acting antitussives on ascites tumor cells.

Dextromethorphan, dimemorfan, dihydrocodeine and oxymethebanol, centrally acting antitussives, were examined for their effect on Ehrlich carcinoma cells and sarcoma-180 cells in vitro or in vivo. The tumor cells were suspended in Hanks balanced salt solution (pH 7.4) supplemented with 2% bovine albumin, and they were incubated with and without 1 mM drugs at 37 degrees C for 120 min. The incubation of the tumor cells with dextromethorphan or dimemorfan resulted in a decrease in the proportion of the viable cells (less than 25% after 120 min). However, no significant change was observed in the proportion of the viable tumor cells during the incubation with and without the other drugs (80-83% after 120 min). In addition, mice given the tumor cells i.p. were injected intraperitoneally with drugs (20-80 mg/kg/day) once daily for 5 successive days, and their survival time was observed. There was a slight difference in the survival time between mice treated with and without dextromethorphan or dimemorfan. However, a significant difference was found in the survival time between mice treated with and without dextromethorphan when mice given Ehrlich carcinoma cells were injected with the drug (40 mg/kg/time) twice a day for 5 days (about 18 days and 29 days). These results indicate that dextromethorphan and dimemorfan are cytotoxic to the tumor cells in vitro and in vivo.

EFFECT OF THE CENTRALLY ACTING ANTITUSSIVES ON ASCITES TUMOR CELLS

Dextromethorphan, dimemorfan, dlhydrocodeine and oxymethebanol, centrally acting antitussives, were examined for their effect on Ehrlich carcinoma cells and sarcoma-180 cells in vitro or in vivo. The tumor cells were suspended in Hanks balanced salt solution (pH 7.4) supplemented with 2% bovine albumin, and they were incubated with and without 1 mM drugs at 37°C for 120 min. The incubation of the tumor cells with dextromethorphan or dimemorfan resulted in a decrease in the proportion of the viable cells (less than 25% after 120 min). However, no significant change was observed in the proportion of the viable tumor cells during the incubation with and without the other drugs (80-83% after 120 min). In addition, mice given the tumor cells i.p. were injected intraperitoneally with drugs (20-80 mg/kg/day) once daily for 5 successive days, and their survival time was observed. There was a slight difference in the survival time between mice treated with and without dextromethorphan or dimemorfan. However, a significant difference was found in the survival time between mice treated with and without dextromethorphan when mice given Ehrlich carcinoma cells were injected with the drug (40 mg/kg/time) twice a day for 5 days (about 18 days and 29 days). These results indicate that dextromethorphan and dimemorfan are cytotoxic to the tumor cells in vitro and in vivo.

Cytotoxicity of cortisone-resistant lymphoid cells from mice treated with adjuvants.

The cortisone-resistant lymphoid cells (CR cells) from mice treated with and without adjuvants were examined for their cytotoxicity on Ehrlich carcinoma cells (EC cells) or sarcoma-180 cells by the Winn test. Female mice of ddY strain were injected s.c. with Freund’s complete adjuvant (FCA), lipopolysaccharide (LPS) from E.coli or a group A hemolytic streptococcus (Cocci), and they were killed 14 days later. Using Hanks balanced salt solution, CR cells were prepared from thymus, spleen or mesenteric lymph node of mice injected i.p. with hydrocortisone acetate (125 mg/kg) 2 days before killing. CR cells were mixed with tumor cells (tumor cells/CR cells: 1/10), and the cell mixture was inoculated s.c. into mice (106 tumor cells/mouse). When EC cells were mixed with the mesenteric CR cells from untreated mice, 9 out of 20 mice given the cell mixture developed solid tumor. However, no tumor growth was observed in groups of mice receiving the mixture of EC cells and the mesenteric CR cells from animals treated with FCA or Cocci (0/20). In groups of mice given the mixture of EC cells and the thymic CR cells, the tumor incidence was 15/20 for CR cells from untreated mice and 6-7/20 for CR cells from mice treated with FCA or Cocci. On the other hand, the tumor growth was observed in almost all mice given the mixture of sarcoma-180 cells and the thymic or mesenteric CR cells form mice treated with FCA or Cocci.

Inhibition of uridine transport through sarcoma-180 cell membrane by anthracycline antibiotics

Inhibition of uridine transport through sarcoma-180 cell membrane by anthracycline antibiotics

Histopathological studies on antitumor effect of sporamycin. Cell-mediated immunity against allogeneic tumor-bearing mice.

Mice that had received transplants of sarcoma-180 followed by treatment with sporamycin were examined histopathologically at periodic intervals. A marked degeneration of tumor cells was observed at an early stage after the administration of sporamycin, but the degeneration subsequently ceased and regrowth of the tumor was seen. Marked infiltration of lymphoid cells, granulation tissue, and fibrosis was seen in the stroma or surrounding tissue of the tumor at a late stage after the administration of sporamycin, and the regression of tumor cells became marked. With a few exceptions the mice were completely cured by about the 40th day. In the peripheral lymphoid tissues, a transitory decrease in the number of cells was observed after the administration of sporamycin, but this was followed by regeneration of the cells, followed by a marked increase in the B cell system. On the other hand, lymphoid cell depletion of the thymus had persisted. Transplantation of intact sarcoma-180 to mice preliminarily inoculated with sporamycin-treated sarcoma-180 cells resulted in inhibition of tumor growth in most of the mice, and qualitatively the same tissue reactions as those in mice cured of sarcoma-180 by sporamycin were seen. The results suggest that enhancement both of antigenicity of the tumor (cells) and of the subsequent immune response of the host by sporamycin may be involved in the cure of the experimental tumor.

Metabolism of 1, N6-Ethenoadenosine

The metabolic fate of 1, N 6-ethenoadenosine ( I) has been studied in rat. The predominant metabolic pathway is the scission of the glycosidic bond in I to yield the free base, 1, N 6-ethenoadenine ( III). Incubation of I with the extract of sarcoma-180 cells in presence of ATP and Mg 2+ did not result in phosphorylation of the nucleoside. No significant uptake of this compound was observed in L-1210 mouse leukemia cells grown in culture.

Chromosome analysis with special reference to centromeric heterochromatin and ploidy variation in mouse sarcoma-180 cells.

Chromosome analysis of an ascitic form of mouse sarcoma 180 has been made both by conventional Giemsa staining as well as by C-banding. The modal number has been found as 75 with one biarmed marker in the present ascitic tumour. C-banding analysis reveals that this submetacentric marker is actually a dicentric chromosome with two closely situated C-positive heterochromatic zones. In addition to this marker, the cell line also possesses another stable and transmissible dicentric chromosome with two distinctly terminal centromeres, and an another unusual chromosome with an extended centromeric heterochromatin.Spontaneous chromosome elimination during anaphase separation, endoreduplication, multipolar spindle formation, etc. have been recorded in a high frequency which are assumed to be responsible for the characteristic ploidal variation in this cell line.