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Control of Human B Cell Tumor Growth in Severe Combined Immunodeficiency Mice by Monoclonal Anti-B Cell Antibodies

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Control of Human B Cell Tumor Growth in Severe Combined Immunodeficiency Mice by Monoclonal Anti-B Cell Antibodies Control of human B cell tumor growth in severe combined immunodeficiency mice by monoclonal anti-B cell antibodies. A Durandy, … , A M Fischer, A Fischer J Clin Invest. 1992;90(3):945-952. https://doi.org/10.1172/JCI115971. Research Article Severe combined immunodeficiency (scid) mice develop EBV (+)B cell tumors after infusion of EBV(+)B cells or of B cells and EBV. In this study, scid mice were infused with B cell lines derived from three patients who developed a B lymphocyte proliferative disorder after bone marrow or organ transplantation. Intraperitoneal injection of 5 x 10(6) B cells induced tumor growth in all mice, leading to death within 60 d. Human B cells were identified in spleen and bone marrow by means of immunofluorescence or EBV genome amplification, and human IgM was detected in serum. Infusion of murine monoclonal antibodies specific for human B cell membrane antigens CD21, CD24, and CD23 was effective in 80% of animals, against two of the three cell lines preventing tumor development or inducing remission according to the time of treatment. The effect was antibody dose dependent and was optimal with four intravenous infusions of at least 0.1 mg 4 d apart. Human IgM in serum and human B cells in spleen and bone marrow became undetectable when peritoneal tumors regressed completely. Infusions of IgG1 isotype-matched anti-CD4 antibody or anti-CD3 antibody had no effect. Tumors developed or recurred in 50% of these animals injected with one of the B cell line 3 mo after treatment was stopped. The same anti-CD21 and anti-CD24 antibodies had been used […] Find the latest version: https://jci.me/115971/pdf Control of Human B Cell Tumor Growth in Severe Combined Immunodeficiency Mice by Monoclonal Anti-B Cell Antibodies A. Durandy,* N. Brousse,t F. Rozenberg,' G. De Saint Basile,* A. M. Fischer,11 and A. Fischer* *Institut National de la Sante et de la Recherche Medicale, U 132, H6pital des Enfants-Malades, 75015 Paris, France; tService d'Anatomopathologie, H6pital des Enfants-Malades, 75015 Paris, France; §Laboratoire de Virologie, H6pital Saint- Vincent-de-Paul, 75014 Paris, France; and I1Laboratoire d'Hematologie, H6pital des Enfants-Malades, 75015 Paris, France Abstract are a significant complication of the immunosuppression re- Severe combined immunodeficiency (scid) mice develop EBV quired for HLA nonidentical bone marrow and organ trans- ( + ) B cell tumors after infusion of EBV( + ) B cells or of B cells plantation (1-4). BLPD also occur in HIV-infected subjects and EBV. In this study, scid mice were infused with B cell lines and patients with profound, primary T cell disorders (5, 6). derived from three patients who developed a B lymphocyte pro- The outcome of BLPD is frequently poor, although some liferative disorder after bone marrow or organ transplantation. forms resolve following reduction of immunosuppression ther- Intraperitoneal injection of 5 x 10' B cells induced tumor apy (7), and treatment with the antiviral drug DHPG (8) or a growth in all mice, leading to death within 60 d. Human B cells 2-interferon plus immunoglobulin- (9). We have recently were identified in spleen and bone marrow by means of immuno- found that nonmonoclonal BLPD can be controlled by the fluorescence or EBV genome amplification, and human IgM administration of CD2 1- and CD24-specific monoclonal anti- was detected in serum. Infusion of murine monoclonal antibod- bodies ( 10). ies specific for human B cell membrane antigens CD21, CD24, Severe combined immunodeficiency (scid) mouse T and B and CD23 was effective in 80% lymphocytes are unable to differentiate because of a defective of animals, against two of the exhibit a three cell lines preventing tumor development or inducing re- recombinase complex and thus profound immunode- mission according to the time of ficiency ( 1 1, 12); this has enabled several groups to transplant treatment. EBV-infected B cells The effect was antibody dose dependent and was optimal human lymphoid cells ( 13, 14). grow well with four intravenous infusions of at least 0.1 mg 4 d apart. in scid mice and give rise to tumors ( 15-18), and their morpho- characteristics are identical to those Human IgM in serum and human B cells in spleen and bone logical and membrane of marrow became undetectable when peritoneal tumors re- EBV-induced B cell lines in vitro and some EBV-induced gressed completely. Infusions of IgGl isotype-matched anti- BLPD in immunosuppressed patients. Lymphocytes contain- CD4 antibody or anti-CD3 antibody had no effect. Tumors de- ing the EBV genome (15, 16), lymphocytes injected before veloped or recurred in 50% of these animals injected with one of Epstein-Barr virus infection ( 17) and B lymphoblastoid cell the B cell line 3 mo after treatment was stopped. lines can also grow in scid mice ( 18). To determine whether The same anti-CD21 and anti-CD24 antibodies had been scid mice were suitable as an in vivo model for screening poten- used to treat the three patients, and shown similar degrees of tial anti-BLPD treatments, we inoculated scid mice with EBV- B lines from three immunodeficient who effectiveness as in the scid mouse model. These results indicate induced cell patients had developed BLPD. We then tested the effectiveness of mu- that scid mice may be suitable for assessing therapeutic ap- in B cell proaches to human B cell proliferation. (J. Clin. Invest. 1992. rine anti-human B cell antibodies arresting tumor growth. 90:945-952.) Key words: severe combined immunodeficiency mice * B lymphoproliferative syndrome- B lymphoid cell lines. Epstein Barr virus * anti-B cell monoclonal antibodies Methods Introduction scid mice. 4- to 6-wk-old CB17 scid/scid mice were purchased from Iffa-Credo (L'Arbresle, France) or were a kind gift from Dr. J. L. Transformation of human lymphocytes by Epstein-Barr virus Guenet (Institut Pasteur, Paris, France). The immunological defect leads to B cell activation and indefinite B cell proliferation. In was confirmed in each animal by a blood lymphocyte count < I00/1. immunosuppressed patients, infection by EBV (particularly The mice were kept in a sterile isolator and received three nonabsorb- 1 can to able antibiotics (colimycin, tobramycin, and vancomycin) in sterile type ) lead uncontrolled polyclonal or monoclonal pro- water. liferation. Such B lymphocyte proliferative disorders (BLPD) ' Monoclonal antibodies and immunofluorescence studies. The fol- lowing murine monoclonal antibodies directed against various human Address correspondence to Anne Durandy, M. D., INSERM U 132, antigens were used to treat scid mice: anti-CD2 1 (IOB la-IgG I ), anti- Hopital des Enfants-Malades, 149 rue de Sevres, 75015 Paris, France. CD24 (IOB3-IgG 1), anti-CD23 (IOB8-IgGl ) (all three from Immun- Receivedfor publication 16 January 1992 and in revisedform 7April otech, Marseille-Luminy, France), anti-CD3 (OKT3-IgG2a) (Ortho 1992. Pharmaceutical, Raritan, NJ) and anti-CD4 (BF5-IgG I from Dr. Wid- jenes, Centre Regional de Transfusion Sanguine, Besan9on, France). 1. Abbreviations used in this paper: BLPD, B lymphocyte proliferative The following mAb (Immunotech) against human cell surface anti- disorder; IF, immunofluorescence; LCL, lymphoid cell line; PCR, poly- gens were used in immunofluorescence (IF) studies. Anti-CD2 1 merase chain reaction; scid, severe combined immunodeficiency. (IOBla), anti-CD24 (IOB3), FITC-anti-CD19 (IOB4-IgG1), FITC- anti-CD23 (IOB8-IgG 1), FITC-anti-CD1 8 (IOT8-IgG 1): mAb J. Clin. Invest. against human immunoglobulin heavy chains (iu, 6, y, a) and light © The American Society for Clinical Investigation, Inc. chains (K and X) were purchased from Caltag Laboratories (South San 0021-9738/92/09/0945/08 $2.00 Francisco, CA). A two-step immunofluorescence assay using an FITC- Volume 90, September 1992, 945-952 labeled goat anti-mouse immunoglobulin (GAMIG; Caltag) was used Treatment ofHuman B Cell Tumors in Severe Combined Immunodeficiency Mice 945 for staining with unlabeled antibodies. Analysis was performed using a resis and further characterized using restriction enzyme analysis, as Facscan® (Becton Dickinson and Co., Mountain View, CA). Mem- previously described (20). Each round of PCR included internal, posi- brane immunofluorescence studies were performed on cell suspen- tive, and negative controls. sions. Mouse organs (liver, spleen, and bone marrow) were homoge- Serum immunoglobulin determination. Human IgM immunoglob- nized and filtered, and mononuclear cells were isolated on density gra- ulin was determined in mouse serum by means of nephelometry using dients (Lymphoprep; Nuyagaard, Oslo, Norway). Peritoneal cells were an anti-human-IgM antibody. Monoclonal immunoglobulin was obtained by gentle washing of the peritoneal cavity. identified using the Paragon immunofixation electrophoresis kit Intracytoplasmic immunoglobulin content was assessed by means (Beckman Instruments, Inc., Fullerton, CA) according to the method of direct immunofluorescence using FITC-labeled anti-human heavy described by Ritchie (21 ). or light chain antibodies, on smears of fixed cells that were examined 48 h after the second injection of anti-CD21 mAb (0.1 mg intrave- under a Leitz microscope. nously), anti-CD21 levels in mouse serum were determined using IF Immunohistology. Fragments of abdominal tumors were snap fro- with an FHTC-Gamig on a B lymphoid cell line; serum from untreated zen in isopentane in liquid nitrogen. Cryostat sections 5-sm thick were mice and unlabeled mAb were used as controls. placed on glass sides, air dried for at least
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