CTLA4 (cytotoxic T-lymphocyte-associated protein 4), also known as CD152, competitively binds to B7-1 (CD80) and B7-2 (CD86) on Antigen-Presenting Cells (APCs) to block the T cell activating signal by B7 and CD28 (on T cells) interaction. The inhibition of CTLA4 by its inhibitory antibodies enhances T cell activity. The CTLA4 antibody is the first FDA-approved antibody to treat advanced melanoma. LAG3 (Lymphocyte activation gene 3, CD223) is a lymphocyte activation gene and a member of the Ig family. It is mainly expressed in activated T cells, NK cells, B cells and plasmacytoid DCs. LAG3 is a negative regulator of immunity and mainly binds to MHC class Ⅱ molecules to regulate the function of dendritic cells. The expression of LAG3 is associated with the negative immunoregulatory function of specific T cells. Inhibition of LAG3 function enhances the antitumor effect of specific CD8+ T cells, therefore LAG3 is a potential target for tumor immunotherapy.
Gene targeting strategy for B-hCTLA4/hLAG3 mice. The exon 2 of mouse Ctla4 gene that encodes the extracellular domain was replaced by human CTLA4 exon 2 in B-hCTLA4/hLAG3 mice. The exons 2-3 of mouse Lag3 gene that encode the extracellular domain were replaced by human LAG3 exons 2-3 in B-hCTLA4/hLAG3 mice.
Protein Expression Analysis
Strain specific CTLA4 and LAG3 expression analysis in homozygous B-hCTLA4/hLAG3 mice by flow cytometry. Splenocytes were collected from WT and homozygous B-hCTLA4/hLAG3 (H/H) mice stimulated with anti-CD3ε in vivo, and analyzed by flow cytometry with species-specific anti-CTLA4 and anti-LAG3 antibody. Mouse CTLA4 was detectable in WT mice, mouse LAG3 was detectable in homozygous B-hCTLA4/hLAG3 and WT mice due to the anti-mouse LAG3 antibody cross-reacts with human LAG3. Human CTLA4 and LAG3 were exclusively detectable in homozygous B-hCTLA4/hLAG3 but not WT mice.
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