asialo-gm1-ganglioside and Leukemia-L5178

We have investigated the effect of recombinant polypeptides with cell-binding domain (C-274) or with heparin-binding domain (H-271) and their fusion polypeptide (CH-271) on liver metastasis of murine lymphoid tumor. The polypeptides containing heparin-binding domain, H-271 and CH-271, were able to inhibit liver metastasis when co-injected i.v. with L5178Y-ML25 T-lymphoma cells, while C-274 with cell-binding domain showed much weaker antimetastatic activity. Treatment with H-271 or CH-271 substantially prolonged the survival time of mice injected i.v. with L5178Y-ML25 cells. CH-271, containing cell- and heparin-binding domains, was more antimetastatic than H-271. The reason why CH-271 was more effective in inhibiting liver metastasis than H-271 can not be explained in terms of a difference in the stability in the circulation or in the molecular size of the polypeptide. The polypeptides used in this study did not affect the tumor cell growth or viability in vitro. CH-271 was found to be still active in inhibiting liver metastasis even when natural killer cells or macrophages were removed from this system. Furthermore, multiple administrations of CH-271 after tumor implantation effectively inhibited liver metastasis and enhanced the survival rate as compared with H-271, C-274 and untreated control. Thus, the fusion of H-271 with C-274 (i.e. CH-271) augments the antimetastatic property of H-271, possibly through the interaction between tumor cells and the heparin-binding domain of fibronectin.

Topics: 2-Chloroadenosine; Animals; Binding Sites; Cell Adhesion; Fibronectins; G(M1) Ganglioside; Glycosphingolipids; Heparin; Leukemia L5178; Liver Neoplasms; Mice; Neoplasm Metastasis; Recombinant Fusion Proteins; Structure-Activity Relationship; Survival Analysis

DBA/2 mice implanted i.p. with an aclarubicin (ACR)-resistant subline of L5178Y cells survived 4- to 5-fold longer than those with the parental cells; and animals with the Adriamycin- or bleomycin-resistant subline displayed an intermediate survival period. The i.p. treatment of mice with cyclophosphamide markedly enhanced i.p. growth of the ACR-resistant cells, suggesting that a certain host defense mechanism participates in the lower transplantability. In vitro, the ACR-resistant subline showed much higher sensitivity to natural killer cells. The i.p. pretreatment with anti-asialo-GM1 antibody markedly reduced the mean survival period of mice implanted i.p. with the ACR-resistant cells, suggesting that natural killer cells play an important role in the defense against transplantation of the ACR-resistant cells.

Topics: Aclarubicin; Animals; Antigen-Antibody Reactions; Cyclophosphamide; Cytotoxicity, Immunologic; Drug Resistance; G(M1) Ganglioside; Glycosphingolipids; Immunity, Cellular; Killer Cells, Natural; Leukemia L5178; Leukemia, Experimental; Macrophages; Mice; Mice, Inbred DBA; Naphthacenes; Neoplasm Transplantation

Natural killer (NK) activity in the rat and human has been attributed to cells having the morphology of large granular lymphocytes (LGL). However, this association has been less clear in the mouse, largely because of difficulties in obtaining highly enriched populations of LGL from normal spleen and blood. We have previously observed that the administration of the biological response modifier (BRM) maleic anhydride divinyl ether (MVE-2) strongly augmented NK activity in lung and liver, and the augmented NK activity coincided with increased resistance to the formation of experimental metastases in these organs. The degree of NK augmentation was most striking in the liver, an unexpected and previously unreported observation. In the present study, both MVE-2 or Corynebacterium parvum induced a dramatic augmentation of liver NK activity, which reached maximum levels 3-5 d after treatment. This augmentation of NK activity in the liver coincided with a large increase in the number of lymphoid cells with the morphological characteristics of LGL that could be isolated from enzymatically digested suspensions of perfused liver. The yield of LGL per liver following BRM treatment corresponded to a 10-50-fold increase as compared to normal mice. LGL were purified from these enzymatically digested suspensions of perfused liver by depletion of adherent cells on nylon wool columns and subsequent enrichment for low-density lymphoid cells by fractionation on Percoll density gradients. The enrichment of LGL correlated with greatly increased NK activity against YAC-1. Conversely, the higher-density fractions were depleted of both LGL and NK activity. This increase in NK activity in the liver was suppressed by in vivo treatment with anti-asialo GM1 (asGM1) serum. This treatment also resulted in a corresponding reduction in both the total number and percentage of LGL. By flow cytometry analysis, the phenotype of the majority of these highly cytolytic LGL isolated from the livers of BRM-treated mice were asGM1+, Thy-1+, Ly-5+, Qa-5+, Mac-1+, and Gma-1+, whereas these LGL were Ly-1-, Lyt-2-, L3T4-, and surface Ig-. We conclude that the livers of BRM-treated mice can provide a rich source of highly active mouse LGL that could be used for further characterization of this lymphocyte subset. Further, these studies imply a potential for BRM therapy of neoplastic or viral diseases through augmentation of organ-associated immune responses.

Topics: Adjuvants, Immunologic; Animals; Antigens, Surface; Ascitic Fluid; Cell Line; Cell Separation; Cytotoxicity, Immunologic; Female; G(M1) Ganglioside; Glycosphingolipids; Immune Sera; Killer Cells, Natural; Leukemia L5178; Liver; Lymphoma; Male; Mice; Mice, Inbred C57BL; Organ Specificity; Phenotype; Polymers; Propionibacterium acnes; Pyran Copolymer; Spleen

Intravenous injection of rabbit anti-asialo-GM1 serum, an antiserum previouslY shown to eliminate splenic natural killer (NK) activity in vitro, profoundly depressed NK activity in CBA, DBA/2 and BALB/c nu/nu mice. The effect on NK activity was selective, as treatment of mice with anti-asialo-GM1 serum did not affect the development of other cytotoxic cells including cytotoxic macrophages following injection of poly I:C, or cytotoxic T cells in response to allogeneic cells. The role of NK cells in controlling tumor cell growth was investigated using an NK-sensitive (cl 27v-1C2) and an NK-resistant (cl 27av) subline of the murine lymphoma L5178Y. Initial studies showed that cl 27v-1C2 cells were at least 100 times less tumorigenic than were cl 27av cells in both syngeneic DBA/2 mice and BALB/c nu/nu mice. In addition, treatment of DBA/2 mice with poly I:C, which boosted NK activity, markedly depressed the growth of cl 27v-1C2 cells, but not of cl 27av cells. On the other hand, treatment of DBA/2 mice and BALB/c nu/nu mice with anti-asialo-GM1 serum led to a marked increase in tumorigenicity of cl 27v 1C2 cells, but had no effect on the tumorigenicity of cl 27av cells. In addition, the protection against cl 27v-1C2 growth afforded by poly-I:C treatment was abrogated by injection oif anti-asialo-GM1 serum. The possibility that the effects observed were caused by binding of the injected antibodies to the tumor cells was minimized by: (1) using a clone of tumor cells (cl 27v-1C2) that lacks chemically detectable asialo-GM1, and (2) pretreating animals with anti-asialo-GM1 rather than administering antiserum and tumor cells concurrently. These studies provided compelling evidence that NK cells could play an active role in controlling tumor growth. Selective depletion of NK activity by injection of anti-asialo-GM1 serum is a method which would be generally applicable to studying the role of NK cells in disease processes.

Topics: Animals; Cell Division; Cell Line; Cell Transformation, Neoplastic; G(M1) Ganglioside; Glycosphingolipids; Immune Sera; Immunity, Cellular; Killer Cells, Natural; Leukemia L5178; Male; Mice; Mice, Inbred Strains; Neoplasm Transplantation; Neoplasms, Experimental; Poly I-C