PD-L1 (Programmed cell death ligand-1), also known as B7-H1 and CD274, is mainly expressed in antigen-presenting cells (APCs) and activated T cells, and is one of the two ligands of PD-1. The interaction between PD1 and PD-L1 plays an important role in the negative regulation of the immune response. PD-L1 is highly expressed in a variety of solid tumors. PD-1 and PD-L1 interactions can reduce T Cell activation and promote tumor immune escape. The PD-1/PD-L1 signaling pathway can be blocked and antitumor immune response can be restored by using by anti-PD-1 or anti-PD-L1 antibodies to block the binding of PD1 to PD-L1.
TNFRSF9 (Tumor necrosis factor receptor super family, member 9), also called CD137 and 4-1BB, is a co-stimulatory molecule and is mainly expressed on the surface of T, NK and mononuclear cells. CD137 is activated by its ligand CD137L or activating anti-CD137 antibodies, resulting in T Cell activation, proliferation and cytokine production. CD137L is expressed on antigen presenting cells such as dendritic cells and macrophages and activated B cells. In vivo studies show that CD137 activation increases anti-tumor immune responses, providing a new target for immunotherapy.
Gene targeting strategy for B-hPD-L1/h4-1BB mice. The exon 3 of mouse Pdl1 gene that encode the IgV domain were replaced by human PD-L1 exon 3 in B-hPD-L1/h4-1BB mice. The exon 2-7 of mouse 4-1BB gene that encode the extracellular domain were replaced by human 4-1BB exon 2-7 in B-hPD-L1/h4-1BB mice.
Protein Expression Analysis
Strain specific PD-L1 and 4-1BB expression analysis in homozygous B-hPD-L1/h4-1BB mice by flow cytometry. Splenocytes were collected from WT and homozygous B-hPD-L1/h4-1BB (H/H) mice stimulated with anti-CD3ε in vivo, and analyzed by flow cytometry with species-specific anti-hPD-L1 and anti-h4-1BB antibody. Mouse Pdl1 and 4-1BB was detectable in WT mice but not in homozygous B-hPD-L1/h4-1BB mice. Human PD-L1 and 4-1BB was exclusively detectable in homozygous B-hPD-L1/h4-1BB mice but not in WT mice.
In vivo efficacy of combination of anti-human PD-L1 antibody and anti-human 4-1BB antibody
Antitumor activity of combination of anti-human PD-L1 antibody (atezolizumab，in house）and anti-human 4-1BB (urelumab, in house) in B-hPD-L1/h4-1BB mice. (A) combination of anti-human PD-L1 antibody and anti-human 4-1BB antibody inhibited MC38 tumor growth in B-hPD-L1/h4-1BB mice. Murine colon cancer MC38 cells were subcutaneously implanted into homozygous B-hPD-L1/h4-1BB mice (female, 6-week-old, n=6). Mice were grouped when tumor volume reached approximately 100 mm3, at which time they were treated with anti-human PD-L1 antibody and anti-human 4-1BB antibody with doses and schedules indicated in panel. (B) Body weight changes during treatment. As shown in panel A, combination of anti-human PD-L1 antibody and anti-human 4-1BB antibody were efficacious in controlling tumor growth in B-hPD-L1/h4-1BB mice, demonstrating that the B-hPD-L1/h4-1BB mice provide a powerful preclinical model for in vivo evaluation of combination of anti-human PD-L1 antibody and anti-human 4-1BB antibody. Values are expressed as mean ± SEM.
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