Fusion Of Tumor Cells Research Articles

Treatment of hepatocellular carcinoma with the cellular tumor vaccines generated by in vitro modification of tumor cells with non gene transfer approaches.

Anti-tumor immune responses are mediated primarily by T cells. Down regulation of major histocompatibility complex (MHC) and the molecules that costimulate the immune responses is associated with defective signaling of tumor cells for T cell activation. In vitro fusion of autologous tumor cells with antigen presenting cells (APCs) or treatment of tumor cells with a combination of cytokines significantly increased the expression of MHC class I and adhesion molecules on tumor cell surfaces that costimulate host immune responses. The hybrid cells generated by fusion of tumor cells with APCs and the tumor cells treated in vitro with a combination of cytokines and pre-incubated with a bispecific monoclonal antibody (bi-Mab) cross-linking antigen on tumor cells to CD28 on T cells, become immunogenic and able to stimulate naive T cells with generation of tumor specific cytotoxic T cells both in vitro and in vivo. Immunization with the modified tumor cells elicits an immune response mediated by both CD4+ and CD8+ T cells. This response protected against a parental tumor cell challenge and cured established tumors. The approach was effective in both low immunogenic and non-immunogenic tumor systems. Modification of tumor cells with tumor:APC fusion or the two-step procedure may provide a strategy for development of tumor vaccines that is effective for cancer immunotherapy.

Pre-B acute lymphoblastic leukemia cells may induce T-cell anergy to alloantigen

Abstract Even if neoplastic cells express tumor associated antigens they still may fail to function as antigen presenting cells (APC) if they lack expression of one or more molecules critical for the induction of productive immunity. These cellular defects can be repaired by physiologic activation, transfection, or fusion of tumor cells with professional APC. Although such defects can be repaired, antitumor specific T cells may still fail to respond in vivo if they may have been tolerized. Here, human pre-B cell acute lymphoblastic leukemia (pre-B ALL) was used as a model to determine if primary human tumor cells can function as alloantigen presenting cells (alloAPC) or alternatively whether they induce anergy. In the present report, we show that pre-B cell ALL express alloantigen and adhesion molecules but uniformly lack B7–1 (CD80) and only a subset express B7–2 (CD86). Pre-B ALL cells are inefficient or ineffective alloAPC and those cases that lack expression of B7–1 and B7–2 also induce alloantigen specific T- cell unresponsiveness. Under these circumstances, T-cell unresponsiveness could be prevented by physiologic activation of tumor cells via CD40, cross-linking CD28, or signaling through the common gamma chain of the interleukin-2 receptor on T cells. Taken together, these results suggest that pre-B ALL may be incapable of inducing clinically significant T-cell-mediated antileukemia responses. This defect may be not only due to their inability to function as APC, but also due to their potential to induce tolerance. Attempts to induce clinically significant antitumor immune responses may then require not only mechanisms to repair the antigen presenting capacity of the tumor cells, but also reversal of tolerance.

Pre-B acute lymphoblastic leukemia cells may induce T-cell anergy to alloantigen

Even if neoplastic cells express tumor associated antigens they still may fail to function as antigen presenting cells (APC) if they lack expression of one or more molecules critical for the induction of productive immunity. These cellular defects can be repaired by physiologic activation, transfection, or fusion of tumor cells with professional APC. Although such defects can be repaired, antitumor specific T cells may still fail to respond in vivo if they may have been tolerized. Here, human pre-B cell acute lymphoblastic leukemia (pre-B ALL) was used as a model to determine if primary human tumor cells can function as alloantigen presenting cells (alloAPC) or alternatively whether they induce anergy. In the present report, we show that pre-B cell ALL express alloantigen and adhesion molecules but uniformly lack B7-1 (CD80) and only a subset express B7-2 (CD86). Pre-B ALL cells are inefficient or ineffective alloAPC and those cases that lack expression of B7-1 and B7-2 also induce alloantigen specific T-cell unresponsiveness. Under these circumstances, T-cell unresponsiveness could be prevented by physiologic activation of tumor cells via CD40, cross-linking CD28, or signaling through the common gamma chain of the interleukin-2 receptor on T cells. Taken together, these results suggest that pre-B ALL may be incapable of inducing clinically significant T-cell-mediated antileukemia responses. This defect may be not only due to their inability to function as APC, but also due to their potential to induce tolerance. Attempts to induce clinically significant antitumor immune responses may then require not only mechanisms to repair the antigen presenting capacity of the tumor cells, but also reversal of tolerance.

Impaired interleukin-3 mRNA decay in autocrine mast cell tumors after transient calcium ionophore stimulation.

Autocrine interleukin-3 (IL-3) expression of v-H-ras transformed mast cell tumors involves either IL-3 mRNA stabilization (class-I tumors) or enhanced IL-3 transcription (class-II tumors). Since calcium ionophores induce IL-3 expression in untransformed PB-3c cells by transcript stabilization, we asked whether class-I tumor could still respond to calcium ionophores. We found that ionomycin treatment further increased IL-3 mRNA expression of class-I tumor cells. Following ionomycin wash-out, IL-3 mRNA decay was slower in class-I tumor cells than in class-II tumor or precursor cell lines (t1/2 > 50 min versus < 20 min, respectively). Somatic cell fusion of the class-I tumor cells with the non-tumorigenic PB-3c cells resulted in reversion to rapid decay after ionomycin wash-out. The data indicate that a recessive defect of IL-3 mRNA degradation can be revealed in class-I tumor cells by transient calcium ionophore stimulation. However, IL-3 mRNA stabilization operating constitutively in class-I tumor cells appears to be distinct from the ionomycin induced process.

The fusion of tumor cells with host cells; reflections on an ovarian tumor.

The fusion of tumor cells with host cells; reflections on an ovarian tumor.

3] Sendai virus-induced cell fusion

Publisher Summary This chapter discusses the mechanism and details of cell fusion by hemagglutinating virus of japan (HVJ). Sendai virus (HVJ) can fuse cells with viral receptors on their surface, including almost all cultured cells derived from mammals and fowls. In general, cells of established lines and transformed cells are fused more readily than differentiated cells, ∼ and HVJ can fuse both cells in suspension and plated cells in culture dishes. The use of cells in suspension is easier for quantitative estimation of cell fusion efficiency. The chapter describes the mechanism of fusion of Ehdich ascites tumor (EAT) cells induced by HVJ. This chapter discusses that Ehdich ascites tumor cells are rapidly fused by HVJ at 37 °, under conditions supplying calcium ions and allowing energy generation. The initial reaction is disruption of the plasma membranes, induced by the function of the fusogenic N-terminal domain of F 1 .

The T cell repertoire may be biased in favor of MHC recognition

The receptors of two T cell hybridomas that recognize class I and class II major histocompatibility complex (MHC) molecules, respectively, have been compared. In both cases these receptors are hybrid molecules formed as a result of cellular fusion. The receptors contain the same α chain, contributed by the tumor cell fusion partner, and related β chains, contributed by the normal T cell component. Thus, surprisingly, the same α chain can contribute to recognition of class I and class II MHC molecules. Moreover, the finding that in two independent examples hybrid receptor molecules created randomly by in vitro cell fusion recognize MHC supports the theory that the T cell repertoire has an intrinsic affinity for MHC.

Экспрессия генов Т-района в гибридах соматических клеток, полученных путем слияния нормальной и опухолевой клеток

Экспрессия генов Т-района в гибридах соматических клеток, полученных путем слияния нормальной и опухолевой клеток

Somatic cell fusion as a source of genetic rearrangement leading to metastatic variants.

Tumor cell populations displaying metastatic properties often have higher gene dosage than their less malignant progenitor tumors, as shown by increased ploidy levels, chromosome duplication and gene amplification. The acquisition by tumor cells of high chromosome numbers may be due to endoreduplication or somatic hybridization either between tumor cells or between tumor and host cells. All such mechanisms increase genetic variability and instability in tumor cells since they trigger a polyploidization-segregation cycle. Among the wide variety of segregants which may emerge from high-ploidy cells, variants with increased malignancy can be positively selected in vivo. Evidence for in vivo fusion of tumor and normal host cells has been reported in different tumor systems. However the attainment by tumor-host hybrids of a higher degree of malignancy has only been observed following substantial chromosome segregation. The involvement of a cell of bone marrow origin as preferential host partner in the fusion process has been proved both by studies on tumor-host hybrids in bone marrow radiation chimeras and in vitro hybridization experiments between non-metastatic tumors and normal lymphoreticular cells which have led to the establishment of metastatic variants. Several different segregational mechanisms may bring about homozygosity or hemizygosity of recessive alleles in tumor-host hybrids, leading to their expression. The marked chromosome dynamics of tumor-host hybrids are also responsible for extensive chromosome rearrangements. At the molecular level these may represent mechanisms causing altered oncogene activity. The activation of new oncogenes by transposition or amplification as well as the amplification of previously activated oncogenes are the mechanisms most likely to be responsible for transition from low to high malignancy, occurring through ploidy changes, such as those produced by somatic mating.

Morphological changes in Ehrlich ascites tumor cells during the cell fusion reaction with HVJ (sendai virus): III. Morphological characterization of HVJ glycoproteins integrated into the plasma membrane and their internalization by coated vesicles

On cell-cell fusion of Ehrlich ascites tumor (EAT) cells with HVJ, HVJ envelopes also fuse with the cell membrane, resulting in integration of the viral envelope glycoproteins into the fused cell membranes. Morphological characterization of the glycoproteins in the plasma membrane and the mode of their internalization were investigated in detail. In the fusion reaction, the glycoproteins were rapidly integrated into the cell membrane within 2 or 3 min on incubation at 37 °C and they remained at the fusion sites, not dispersing widely, during further incubation. Thus they were still present in clusters in the plasma membrane at the end of the fusion reaction. On culture of fused cells in culture medium, internalization of the viral glycoproteins was initiated by formation of coated vesicles and most of the integrated glycoproteins were endocytosed into the cytoplasm within 30 min. Soon after internalization, the coated vesicles fused with each other, losing their coat materials. The intact virions that remained unfused on the cell surface were also internalized, but coat materials did not appear on the inside surface of the cell membrane, unlike in the case of integrated glycoproteins.

Tumor-specific immunity induced by somatic hybrids: III. Persistence of adoptively transferred immunity specific for TEPC-15 plasmacytoma in normal BALB/c mice

Immunity against TEPC-15 tumor cells was induced in BALB/c mice by injecting semi-allogeneic hybrid cells derived from fusion of TEPC-15 tumor cells with LM(TK −) cells of the C3H origin. Adoptive transfer of spleen cells from the immune mice into normal BALB/ c recipients rendered them free from tumors following tumor challenge; the recipients were most significantly protected from the tumor when tumor cells were injected 7–14 days after the adoptive transfer of immune cells. Such immunity following adoptive transfer appeared to persist in the recipients for at least 60 days. Moreover, the tumor-specific immunity was consecutively transferable (more than nine passages) into normal BALB/c recipients by serially passing spleen cells from the recipients every 14 days, without further stimulation with the hybrid cells or inactivated TEPC-15 tumor cells. Such consecutive transfer of the immune spleen cells induced splenomegaly in the recipients: a two- to five-fold increase over normal spleen cell recipients. The ability of spleen cells to transfer immunity, but not splenomegaly, was abrogated by treatment with mitomycin C. These results suggest that proliferation of donor cells is necessary to transfer immunity, and that splenomegaly alone does not manifest such immunity in the recipients.

A better cell line for making hybridomas secreting specific antibodies.

FUSION of myeloma cells which grow in tissue culture with spleen cells from an immunised mouse provides a general method for obtaining cell lines (hybridomas) which make antibody of the desired specificity1–3. Hybrids derived from these myelomas make the immunoglobulin (Ig) heavy and light chains of the myeloma parent as well as the antigen-specific heavy and light chains of the spleen cell parent. In conditions in which the two heavy and two light chains associate randomly, a hybridoma would make 10 distinct Ig molecules, and the specific antibody would comprise only 1/16 of the total Ig4,5. To obtain hybridomas making only the specific antibodies requires a tumour cell fusion partner that itself makes no Ig but which can nevertheless be fused with spleen cells to obtain hybrids secreting only the specific antibody. We report here the identification of such a cell line, Sp2/0-Ag14.

Proof of in vivo fusion of murine tumor cells with host cells by universal fusers.

Two mouse cell lines (A9HT CI.3C and 501-1), each carrying both a recessive and a dominant mutation (and therefore designated "universal fusers"), were utilized for selection of in vivo hybrids from tumors produced intraperitoneally and subcutaneously in appropriated hosts. The double selective medium eliminated both parental (tumor and host) cells but allowed the survival and proliferation of the fused product. This proved that hybridization between tumor and host cells occurred in vivo.

Effects of cyclic AMP and calcium ions on virus-induced fusion of Ehrlich ascites tumor cells

The effects of cyclic nucleotides and Ca 2+ on the fusion of Ehrlich ascites tumor cells induced by HVJ (Sendai virus) were studied. 1. 1. Addition of inhibitors of cyclic nucleotide phosphodiesterase increased the frequency of fusion up to 2 fold. None of these drugs affected the fusion and haemolysis of human erythrocytes. 2. 2. Cyclic AMP content reached a maximum (about 2 fold) after 2 min from the start of the fusion reaction and the increase in cyclic AMP was almost proportional to the dose of the virus added. 3. 3. Reagents which might increase intracellular cyclic GMP, such as insulin and carbamylcholine partially inhibited the fusion process (40% inhibition at 6 μM). 4. 4. The stimulatory effect of theophylline was dependent on the extracellular concentration of Ca 2+ with an optimal concentration of 0.5 mM. 5. 5. Theophylline partially antagonized the inhibition of the fusion reaction by cytochalasin D (0.1 μg/ml or 0.5 μg/ml). The inhibition of the fusion reaction by cytochalasin D was influenced by the extracellular concentration of Ca 2+. 6. 6. However, fusion of human erythrocytes which required neither Ca 2+ nor ATP was not affected by cyclic AMP and cytochalasin D. Therefore, fusion of Ehrlich ascites tumor cells has more complex regulatory systems than human erythrocytes. A possible mechanism of the regulation is discussed.

Production of hybrid cells by fusion of human malignant tumour cells and primary mouse embryo cells

Hybrid cells with a subtetraploid mouse chromosome complement were produced by fusion of three types of human tumour cells with primary mouse embryo cells. The most frequently present presumptive human chromosome was 21. Numerous chromosome rearrangements were present. Some hybrid cells produced regressing tumours in mice.

Inhibition of virus-induced fusion of ehrlich ascites tumor cells by cytochalasin B and D

Cytochalasin B and D were found to inhibit HVJ (Sendai virus)-induced fusion of Ehrlich ascites tumor cells. Nearly complete inhibition was attained by 4 uM (2 μg/ml) cytochalasin D, whereas cytochalasin B was a less effective inhibitor. The inhibition was largely reversible. Since the transport of 2-deoxy-glucose into the tumor cells was not affected by cytochalasin D (though inhibited by cytochalasin B), the observed inhibition was not related to the effect of the drugs on sugar transport. Instead, it was suggested that the inhibition was due to the action of the drugs on microfilaments. The requirement of ATP for the cell fusion could be explained at least partly by the involvement of microfilaments in the cell fusion process.

Polyethylene glycol-mediated fusion of human tumor cells with mouse cells

Polyethylene glycol (PEG)-mediated somatic cell fusion is shown to be dependent on temperature, the molecular weight of PEG used, and the fusion technique. Time-sequential formation of multinucleated cells is described. Cultures were established of hybrids obtained by PEG-mediated fusion of mouse cells and human tumor cells.

Degranulating activity in hybrid cells derived from rat peritoneal mast cells and ehrlich ascites.

Fusion of rat mast cells and Ehrlich ascites tumor cells was mediated by HVJ. Compound 48/80-induced degranulation occurred in the fused cells formed from two mast cells and one tumor cell, but not in the fused cells from one mast cell and two or more tumor cells.

Stimulation of virus-induced fusion of ehrlich ascites tumor cells by 3′,5′-cyclic AMP

HVJ(Sendai virus)-induced fusion of Ehrlich ascites tumor cells was found to be stimulated by treatments which increase the intracellular level of cyclic AMP. This stimulation was optimal at an external concentration of Ca ++ of about 0.5 mM. During the process of cell fusion, the intracellular concentration of cyclic AMP was increased with a maximum at 2 min after the initiation of the fusion reaction. Evidence is also presented which suggests that the increase of the cyclic nucleotide is a part of control mechanism of HVJ-induced fusion of eukariotic cells. Thus, this cyclic AMP-stimulated process could be one of the step(s) requiring ATP and Ca ++, both of which are necessary for the overall fusion process of the tumor cells.

Sarcoma in a Hamster Inoculated With BK Virus, a Human Papovavirus&lt;xref ref-type="fn" rid="fn2"&gt;2&lt;/xref&gt;

An undifferentiated sarcoma occurred in 1 of 52 hamsters inoculated when newborn with BK virus (BKV), A SIMIAN VIRUS 40 (SV40)-related human papovavirus. It was transplantable and grew in tissue culture. Sera of tumor-bearing hamsters were without antibodies reactive to BKV virion antigens in hemagglutination-inhibition and neutralizationtexts, but contained antibodies reactive in immunofluorescence (IF) tests to SV40 T antigen in SV40-TRANSFORMED CELLS AND TO ANTIGENS IN CELLS ACUTELY INFECTED WITH BKV or SV40. The IF reaction between tumor cells and sera of tumor-bearing hamsters was minimal in texts of tumor cells of an early passage but satisfactory in tests of cells from a later passage. Fusion of tumor cells with permissive (monkey and human) cells, induced by inactivated Sendai virus, did not lead to recovery of infectious BKV. Immunization of hamster with BKV or SV40 failed to protect against challenge with tumor cells. There was considerable cross-reactivity between the T antigens of BKV and SV40 viruses