Establishment and characterization of a human non‐secretory plasmacytoid cell line and its hybridization with human B cells

Ritts, Roy E.; Ruı́z-Argüelles, Alejandro; Weyl, Karin G.; Bradley, Amber; Weihmeir, Bernadette; Jacobsen, Deborah J.; Strehlo, Bonnie L.

doi:10.1002/ijc.2910310202

Cited by 42 publications

(6 citation statements)

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“…We investigated these issues in a myeloma model consisting of eight different myeloma-and plasma-cell leukaemia lines, which have been extensively characterized Nishiuchi et al, 1996;Ritts et al, 1983). Our results indicated a constitutive expression of Bcl-2 and a high variability in the relative expression level of the Apo-1/Fas antigen in all of the eight myeloma cell lines investigated.…”

mentioning

confidence: 94%

“…Two additional cell lines were used in this study (i.e. the ARH-77 (Nishiuchi et al, 1996) and the MC/CAR (Ritts et al, 1983)). The IM-9, OPM-2, LP-1 and L363 cell lines were obtained from DSM (German Collection of Microorganisms and Cell Cultures, Braunschweig, Germany), the RPMI 8226, ARH-77, MC/CAR and HS-Sultan cell lines from the American Type Culture Collection (Rockville, Md., U.S.A.).…”

Section: Methodsmentioning

confidence: 99%

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Expression of Apo‐1/Fas (CD95), Bcl‐2, Bax and Bcl‐x in myeloma cell lines: relationship between responsiveness to anti‐Fas mab and p53 functional status

et al. 1997

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Summary. The down-regulation of apoptosis may be an essential mechanism for tumour cell expansion in slowly proliferating tumours such as multiple myeloma. We studied eight myeloma cell lines for the presence of Bcl-2, which inhibits apoptosis, of Bax, which counteracts Bcl-2, of Bcl-x L and Bcl-x S , which act in an anti-and pro-apoptotic fashion, respectively, and of Apo-1/Fas, which induces programmed cell death, when activated by the Apo-1/Fas ligand or the relevant monoclonal antibody (mab). All cell lines constitutively expressed homogenous amounts of Bcl-2, but displayed different amounts of Bax and Bcl-x proteins. The Apo-1/Fas antigen could be detected in seven out of eight myeloma lines, but expression levels varied considerably. The relative expression levels of Apo-1/Fas correlated with that of Bax, but not with that of Bcl-2 or Bcl-x subtypes. Furthermore, the effectiveness of the Apo-1/Fas mab was associated with the relative expression levels of the Apo-1/Fas and with that of the Bax antigen, but not with that of the Bcl-2 and Bcl-x antigens. We further showed that wild-type p53 function is not required for Apo-1/Fas-induced apoptosis, nor is it necessary for the expression of Bax or Apo-1/Fas antigens in myeloma.In conclusion, our results suggest a p53-independent coregulation of Apo-1/Fas and Bax, as well as a role for Bax in Apo-1/Fas-induced apoptosis in myeloma.

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confidence: 94%

Section: Methodsmentioning

confidence: 99%

Expression of Apo‐1/Fas (CD95), Bcl‐2, Bax and Bcl‐x in myeloma cell lines: relationship between responsiveness to anti‐Fas mab and p53 functional status

et al. 1997

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“…The examination was performed in a myeloma cell line model, consisting of the Apo-1IFas-sensitive IM-9, LP-1, RPMI-8226, ARH-77 and MUCAR cell lines [17][18][19]. IFN-a, exerted a marked growth inhibition on myeloma cells when incubated for 48 h, but this effect was not caused by the induction of apoptosis.…”

Section: Introductionmentioning

confidence: 99%

Modulation of Apo‐1/Fas (CD95)‐induced programmed cell death in myeloma cells by interferon‐α₂

et al. 1996

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The Apo-1/Fas (CD95) antigen is known to be involved in the process of T cell-mediated target cell killing and has recently been shown to be expressed on myeloma cell lines and native malignant plasma cells. Several cytokines have been reported to interfere with spontaneous and even Apo-1/Fas-induced apoptosis, but no attempt has been made yet to investigate these interactions and the possible underlying mechanisms in myeloma cells. Since in myeloma patients Interferon (IFN)-alpha2 displays a profound therapeutic effect in vivo, which is usually attributed to its growth inhibitory and/or immunomodulatory capacity, we set out to study the potential interference of IFN-alpha2 with Apo-1/Fas-induced apoptosis. Contrary to expectations, IFN-alpha2 reduced the degree of apoptosis caused by the treatment of five Apo-1/Fas-sensitive myeloma cell lines with a Fas monoclonal antibody (mAb). Simultaneous application of IFN-alpha2 and Fas mAb was superior to the prolonged (i.e. >8 h) preincubation with the cytokine as far as inhibition of Apo-1/Fas-induced apoptosis was concerned. This effect of IFN-alpha2 was neither explained by a down-regulation of the Apo-1/Fas receptor nor caused by modulation of the expression levels of c-myc, bcl-2-, bcl-xL, bax- or p53 genes. IFN-alpha2 did not alter the Apo-1/Fas-induced activity of Mitogen-activated protein kinase (MAPK) 1 and did not inhibit the Apo-1/Fas-mediated proteolytic cleavage of ADP-ribosyltransferase, a substrate of Interleukin-beta1 converting enzyme (ICE) and homologues. However, activation of protein kinase C (PKC) by phorbol 12-myristate 13-acetate (PMA) mimicked the effects of IFN-alpha2. Furthermore, the bis-indolylmaleimide GF 109203X, a specific inhibitor of PKC, inhibited the effect of PMA as well as that of IFN-alpha2 on Apo-1/Fas-induced apoptosis. These results point to a PKC-dependent mechanism of transient interaction between the intracellular signaling along the IFN-alpha2 and the Apo-1/Fas pathway (downstream of MAPK signaling as well as of ICE homologues), which becomes exhausted by prolonged stimulation with the cytokine. According to our data IFN-alpha2, applied continuously and in high doses resembling the therapeutic situation in vivo, inhibits myeloma growth. However, based on the observed inhibitory effect of IFN-alpha2 on Apo-1/Fas-induced apoptosis, a partial inhibition of the natural immune surveillance on myeloma cells by endogenous IFN-alpha2 present in the bone marrow microenvironment of this malignancy should be investigated.

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“…These problems have also been encountered in most, though not all, of the fusions of antigen-primed human B lymphocytes with EBV-transformed B lymphoblastoid cell. lines (LCL) (8,(21)(22)(23)(24)(25)(26)(27)(28). Moreover, such hybridomas usually secrete permuted immunoglobulin molecules derived from both fusionpartners.…”

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confidence: 99%

Construction and testing of mouse--human heteromyelomas for human monoclonal antibody production.

Teng¹,

Lam²,

Riera³

et al. 1983

Proc. Natl. Acad. Sci. U.S.A.

157

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FU-266, a mutant human myeloma cell line sensitive to hypoxanthine/aminopterin/thymidine (HAT), was transfected by protoplast fusion with DNA of the recombinant plasmid vector pSV2-neo , thus acquiring a dominant marker conferring resistance to the antibiotic G418. One of the resultant neoR clones, E-1, was fused to irradiated (500 rads) or unirradiated cells of the HAT-sensitive, G-418-sensitive, nonproducer mouse myeloma line X63-Ag8.653. Hybrid clones were selected in G-418 plus ouabain, thus preserving their HAT sensitivity. Small numbers of human chromosomes were retained in all such hybrids, but most of them ceased secreting human myeloma IgE(A). Selected hybrid clones were then tested as malignant fusion partners in a series of fusions with polyclonally activated human B lymphocytes and with antigen-primed human B lymphocytes, in some instances after transformation of the latter with Epstein-Barr virus. The yield of viable chimeric hybridomas has been consistently high, as has the proportion of hybridomas secreting human immunoglobulin molecules unpermuted with mouse or human myeloma heavy or light chains. Secretion by many subcloned hybridomas has been stable for over 6 months, and several antigen-specific human monoclonal antibodies have been generated. Thus these heteromyeloma cell lines appear to have significant advantages for human monoclonal antibody production.The development by Kohler and Milstein (1, 2) of the hybridoma procedure for mouse monoclonal antibody production has opened a new era in immunology. The fact that the cell lines thus derived are cloned and immortal assures the monoclonality and permanent availability of their antibody products, and antibody yield is limited only by cell culture volume. However, clinical use of xenoantibodies in human patients is likely to be severely limited by the fact that they will be treated as foreign proteins by the human immune system. Thus, for diagnostic and therapeutic applications in man, the production of human rather than mouse or other rodent monoclonal antibodies would clearly be desirable.Early attempts to generate immortalized human immunoglobulin-producing cells involved the fushion of human lymphoid cells with mouse myeloma cells to create chimeric hybridomas (3-9). However, with rare exceptions (6-8), such mouse-human hybridomas have tended to cease human immunoglobulin production due to the selective loss of human chromosomes (10, 11), or to disturbances of gene expression (12). Transformation of antigen-primed human B lymphocytes with the Epstein-Barr virus (EBV) (13-20) has had some success, but such cultures usually secrete low levels of antibody and have tended to cease antibody production entirely after a variable period (13). These problems have also been encountered in most, though not all, of the fusions of antigen-primed human B lymphocytes with EBV-transformed B lymphoblastoid cell. lines (LCL) (8,(21)(22)(23)(24)(25)(26)(27)(28). Moreover, such hybridomas usually secrete permuted immunoglobulin molecules derived from...

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Establishment and characterization of a human non‐secretory plasmacytoid cell line and its hybridization with human B cells

Cited by 42 publications

References 37 publications

Expression of Apo‐1/Fas (CD95), Bcl‐2, Bax and Bcl‐x in myeloma cell lines: relationship between responsiveness to anti‐Fas mab and p53 functional status

Expression of Apo‐1/Fas (CD95), Bcl‐2, Bax and Bcl‐x in myeloma cell lines: relationship between responsiveness to anti‐Fas mab and p53 functional status

Modulation of Apo‐1/Fas (CD95)‐induced programmed cell death in myeloma cells by interferon‐α₂

Construction and testing of mouse--human heteromyelomas for human monoclonal antibody production.

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Establishment and characterization of a human non‐secretory plasmacytoid cell line and its hybridization with human B cells

Cited by 42 publications

References 37 publications

Expression of Apo‐1/Fas (CD95), Bcl‐2, Bax and Bcl‐x in myeloma cell lines: relationship between responsiveness to anti‐Fas mab and p53 functional status

Expression of Apo‐1/Fas (CD95), Bcl‐2, Bax and Bcl‐x in myeloma cell lines: relationship between responsiveness to anti‐Fas mab and p53 functional status

Modulation of Apo‐1/Fas (CD95)‐induced programmed cell death in myeloma cells by interferon‐α2

Construction and testing of mouse--human heteromyelomas for human monoclonal antibody production.

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Modulation of Apo‐1/Fas (CD95)‐induced programmed cell death in myeloma cells by interferon‐α₂