The role of complement in the therapeutic activity of rituximab in a murine B lymphoma model homing in lymph nodes.
OBJECTIVE
We have set up a murine B lymphoma model stably expressing human CD20 and homing in lymph nodes in immunocompetent mice to study the mechanism of action of rituximab.


METHODS
The B lymphoma line 38C13 was stably transduced with the human CD20 cDNA by retroviral infection and injected into syngeneic mice.


RESULTS
The transduced 38C13-CD20(+) cells stably expressed human CD20 on 100% of cells. Rituximab alone did not inhibit 38C13-CD20+ cell growth but relocalized the human CD20 into lipid rafts and induced complement-mediated lysis in vitro. Inoculation of 4x10(3) 38C13-CD20(+) intravenously into syngeneic mice led to the development of tumor masses in the spleen, bone marrow and lymph nodes, detectable from day 15 by polymerase chain reaction (PCR) analysis, and with a median survival of 21-24 days. Treatment with 250 mg rituximab i.p. given 1-10 days after tumor inoculation cured 100% of animals, with disappearance of tumor documented by immunohistochemistry and PCR analysis. Depletion of both NK cells and neutrophils did not affect the therapeutic activity of rituximab in vivo. Similarly, removal of phagocytic macrophages using clodronate-liposomes did not modify the capacity of rituximab to control tumor growth. In contrast, the protective activity of the antibody was completely abolished after complement depletion with cobra venom factor. Complement was also required when cells were inoculated subcutaneously in nude mice.


CONCLUSIONS
These data demonstrate that complement is required for the therapeutic activity of rituximab in vivo in a murine model of B-cell lymphoma, independently of its localization in lymph nodes or subcutaneously.
Golay
Josee
J
Laboratory of Cellular and Gene Therapy G. Lanzani, Division of Haematology, Ospedali Riuniti di Bergamo, Bergamo, Italy. jgolay@ospedaliriuniti.bergamo.it
Cittera
Elena
E
Di Gaetano
Nicola
N
Manganini
Massimiliano
M
Mosca
Michela
M
Nebuloni
Manuela
M
van Rooijen
Nico
N
Vago
Luca
L
Introna
Martino
M
D000818:Animals; D000911:Antibodies, Monoclonal / Q000627:therapeutic use*; D058846:Antibodies, Monoclonal, Murine-Derived; D018951:Antigens, CD20 / Q000235:genetics; D045744:Cell Line, Tumor; D003165:Complement System Proteins / Q000502:physiology*; D004195:Disease Models, Animal; D006801:Humans; D008198:Lymph Nodes / Q000473:pathology*; D016393:Lymphoma, B-Cell / Q000188:drug therapy / Q000473:pathology*; D051379:Mice; D008819:Mice, Nude; D009368:Neoplasm Transplantation; D000069283:Rituximab; D014161:Transduction, Genetic
D000911:Antibodies, Monoclonal; D058846:Antibodies, Monoclonal, Murine-Derived; D018951:Antigens, CD20; D000069283:Rituximab; D003165:Complement System Proteins