Anti-CD20 treatment depletes B-cells in blood and lymphatic tissue of cynomolgus monkeys.
BACKGROUND
Macaque species offer a valuable model for translational allo-transplantation and tolerance studies. Cardiac allograft vasculopathy in Macaca fascicularis is associated with elaboration of anti-donor antibodies. Since T-independent pathways of B cell activation have been described, and anti-B cell strategies have proven to be a fruitful tolerogenic adjunct in rodent and xenogenic models, here we investigate whether an anti-CD20 antibody (rituximab) would be useful to deplete B-cells in a pre-clinical allo-transplantation setting in macaques.


METHODS
Three cynomolgus macaques which had previously rejected a cardiac allograft and one with concurrent subacute vascular rejection were treated weekly with rituximab 20 mg/kg i.v. for 4 and 2 weeks, respectively. B-cell levels (CD19+ cells) were measured by flow cytometry in peripheral blood, spleen, lymph node and bone marrow cells at various intervals after initiation of treatment. B-cells and plasma cells were also analyzed by immunohistochemistry at necropsy in spleen, lymph node, tonsil and thymus tissue sections. Anti-donor antibody titers were measured by flow cytometry.


RESULTS
B-cells expressing CD19 were not detectable in the peripheral blood in any animal within 24 h after initial treatment, or over the ensuing month. At necropsy, the germinal centers in spleen and lymph node were completely depleted of CD20+ B-cells in 2 animals, leaving a hypocellular trabecular pattern around preserved plasma cell follicles. Substantial but incomplete depletion of B-cells was demonstrated in the other 2 animals, in each instance immunohistochemical findings in spleen and lymph node exhibiting higher sensitivity for residual B-cells compared to FACS. Anti-donor antibody titers exhibited kinetics similar to untreated animals over this short follow-up.


CONCLUSIONS
Treatment with anti-CD20 very efficiently depletes peripheral and tissue B-cells but not plasma cells in this macaque species. Biopsy of lymph node is necessary and may be sufficient to assess B-cell clearance in secondary lymphoid organs in this model.
Schröder
Carsten
C
Department of Cardiothoracic Surgery, Vanderbilt University, Nashville, TN, USA.
Azimzadeh
Agnes M
AM
Wu
Guosheng
G
Price
James O
JO
Atkinson
James B
JB
Pierson
Richard N
RN
D000818:Animals; D000906:Antibodies / Q000097:blood / Q000494:pharmacology; D000911:Antibodies, Monoclonal / Q000494:pharmacology / Q000627:therapeutic use*; D058846:Antibodies, Monoclonal, Murine-Derived; D018941:Antigens, CD19 / Q000032:analysis / Q000097:blood; D018951:Antigens, CD20 / Q000276:immunology*; D001402:B-Lymphocytes / Q000737:chemistry / Q000276:immunology*; D001854:Bone Marrow Cells / Q000737:chemistry; D023201:CD40 Ligand / Q000276:immunology; D016572:Cyclosporine / Q000494:pharmacology; D005434:Flow Cytometry; D006087:Graft vs Host Reaction; D016027:Heart Transplantation; D007074:Immunoglobulin G / Q000032:analysis / Q000276:immunology; D007075:Immunoglobulin M / Q000032:analysis / Q000276:immunology; D007150:Immunohistochemistry; D008198:Lymph Nodes / Q000166:cytology / Q000187:drug effects; D008221:Lymphoid Tissue / Q000166:cytology / Q000187:drug effects* / Q000276:immunology; D008252:Macaca fascicularis; D008297:Male; D014066:Palatine Tonsil / Q000166:cytology / Q000187:drug effects; D010950:Plasma Cells / Q000166:cytology; D000069283:Rituximab; D013154:Spleen / Q000166:cytology / Q000187:drug effects; D013601:T-Lymphocytes / Q000276:immunology; D013950:Thymus Gland / Q000166:cytology / Q000187:drug effects; D016042:Transplantation, Heterotopic
D000906:Antibodies; D000911:Antibodies, Monoclonal; D058846:Antibodies, Monoclonal, Murine-Derived; D018941:Antigens, CD19; D018951:Antigens, CD20; D007074:Immunoglobulin G; D007075:Immunoglobulin M; D023201:CD40 Ligand; D000069283:Rituximab; D016572:Cyclosporine