Gene targeting strategy for B-CAG-hPD-L1（v2）/hCD47 MC38. The human full-length CD47 CDS region was inserted in the Cd47 exon2 of wild-type MC38. The human full-length CD274 CDS region was inserted in the Cd274 exon3 of wild-type MC38.
Protein expression analysis
The PD-L1 and CD47 expression analysis in B-CAG-hPD-L1(v2)/hCD47 MC38 cells by flow cytometry. Single cells were collected from B-CAG-hPD-L1(v2)/hCD47 MC38 cells, and analyzed by flow cytometry with species-specific anti-PD-L1 and anti-CD47 antibody. Human PD-L1 and CD47 was detectable in B-CAG-hPD-L1(v2)/hCD47 MC38 cells.
Tumor Growth Curve & Body Weight Changes
Subcutaneous xenograft tumor growth of B-CAG-hPD-L1(v2)/hCD47 MC38 cells. B-CAG-hPD-L1(v2)/hCD47 MC38 cells (5×105) were subcutaneously implanted into the B-hPD-L1/hSIRPA/hCD47 mice (n=5).Tumor size and mice body weight were measured twice a week. (A) Tumor average volume ± SEM, (B) Mice body weight (Mean± SEM). Volume was expressed in mm3 using the formula: V=0.5a X b2, where a and b were the long and short diameters of the tumor, respectively. As shown in panel A, B-CAG-hPD-L1(v2)/hCD47 MC38 were able to establish tumor in vivo and can be used for efficacy study.