Bispecific humanized anti-IL-2 receptor alpha beta antibodies inhibitory for both IL-2- and IL-15-mediated proliferation.
Humanized anti-Tac (HAT) and Mik beta1 (HuMik beta 1) Abs directed at IL-2R alpha and IL-2R beta, respectively, inhibit IL-2 binding and biological activity and together act synergistically in vitro. The Abs have been used successfully in primate models of allograft rejection, graft-vs-host disease, and autoimmunity. We produced bifunctional humanized anti-IL-2R alpha beta Abs (BF-IgG) to combine the specificity of the two Abs into one entity by fusing HAT-producing NSO cells and HuMik beta 1-producing Sp2/0 cells. BF-IgG was purified using protein G-Sepharose affinity chromatography, followed by IL-2R alpha and IL-2R beta affinity chromatography and hydrophobic interaction chromatography. BF-IgG exhibited both anti-IL-2R alpha and anti-IL-2R beta specificities in binding assays. While the Ab binds the IL-2R with intermediate affinity (Kd = 2.82 nM), it does not inhibit IL-15 binding to its high affinity IL-15R. In Kit225/K6 (IL-2R alpha beta gamma+) cells, BF-IgG was 10-fold more potent than a HAT/HuMik beta 1 equimolar mixture in blocking IL-2-induced proliferation and, unexpectedly, was at least 65-fold more active than the mixture in blocking IL-15-induced proliferation. This dual inhibitory activity may be due to cross-linking of the IL-2R alpha and IL-2R beta, thus blocking IL-2 binding and possibly impeding the association of IL-2R beta with IL-15R. BF-IgG has potent immunosuppressant activities against both IL-2- and IL-15-mediated responses, and this antagonist could be more efficacious than HAT and/or HuMik beta 1 for the treatment of autoimmunity and the prevention of allograft rejection.
Pilson
R S
RS
Roche Research Center, Inflammation and Autoimmune Diseases, Hoffmann-La Roche, Inc., Nutley, NJ 07110, USA.
Levin
W
W
Desai
B
B
Reik
L M
LM
Lin
P
P
Korkmaz-Duffy
E
E
Campbell
E
E
Tso
J Y
JY
Kerwin
J A
JA
Hakimi
J
J
D018033:Antibodies, Bispecific / Q000096:biosynthesis / Q000302:isolation & purification / Q000494:pharmacology*; D019138:Antibodies, Blocking / Q000494:pharmacology; D000915:Antibody Affinity; D000918:Antibody Specificity; D001666:Binding Sites, Antibody; D002455:Cell Division / Q000276:immunology; D002999:Clone Cells; D004591:Electrophoresis, Polyacrylamide Gel; D004797:Enzyme-Linked Immunosorbent Assay; D006131:Growth Inhibitors / Q000276:immunology* / Q000494:pharmacology*; D006801:Humans; D006822:Hybrid Cells; D019409:Interleukin-15 / Q000502:physiology*; D015458:Leukemia, T-Cell / Q000276:immunology / Q000378:metabolism; D015375:Receptors, Interleukin-2 / Q000276:immunology* / Q000502:physiology*; D012967:Sodium Dodecyl Sulfate; D014407:Tumor Cells, Cultured
D018033:Antibodies, Bispecific; D019138:Antibodies, Blocking; D006131:Growth Inhibitors; D019409:Interleukin-15; D015375:Receptors, Interleukin-2; D012967:Sodium Dodecyl Sulfate