The mouse Cd47 gene was replaced by human CD47 coding sequence in B-hCD47 MC38 cells. Human CD47 is highly expressed on the surface of B-hCD47 MC38 cells.

Gene targeting strategy for B-hCD47 MC38 cells. The exogenous promoter and human CD47 coding sequence were inserted to replace part of murine exon 2 and all of exon 3. The insertion disrupts the endogenous murine Cd47 gene, resulting in a non-functional transcript.

CD47 expression analysis in B-hCD47 MC38 cells by flow cytometry. Single cell suspensions from wild-type MC38 and B-hCD47 MC38 cultures were stained with species-specific anti-CD47 antibody. Human CD47 was detected on the surface of B-hCD47 MC38 cells but not wild-type MC38 cells. The 1-B3 clone of B-hCD47 MC38 cells was used for in vivo experiments.

Subcutaneous homograft tumor growth of B-hCD47 MC38 cells. B-hCD47 MC38 cells (1x10⁶) were subcutaneously implanted into B-hSIRPA/hCD47 mice (female, 7-week-old, n=6). Tumor volume and body weight were measured twice a week. (A) Average tumor volume ± SEM. (B) Body weight (Mean± SEM). Volume was expressed in mm³ using the formula: V=0.5 X long diameter X short diameter². As shown in panel A, B-hCD47 MC38 cells were able to establish tumors in vivo and can be used for efficacy studies.

Subcutaneous homograft tumor growth of B-hCD47 MC38 cells. B-hCD47 MC38 cells (5x10⁵, 1x10⁶, 5x10⁶) and wild-type MC38 cells (5x10⁵) were subcutaneously implanted into B-hCD3E mice (female, 7-week-old). Tumor volume and body weight were measured twice a week. (A) Average tumor volume ± SEM. (B) Body weight (Mean± SEM). Volume was expressed in mm³ using the formula: V=0.5 × long diameter × short diameter². As shown in panel A, B-hCD47 MC38 cells were able to establish tumors in vivo and can be used for efficacy studies.

Subcutaneous homograft tumor growth of B-hCD47 MC38 cells. B-hCD47 MC38 cells and wild-type MC38 cells were subcutaneously implanted into B-h4-1BB mice (female, 9-week-old, n=6). Tumor volume and body weight were measured twice a week. (A) Average tumor volume. (B) Body weight. Volume was expressed in mm³ using the formula: V=0.5 X long diameter X short diameter². As shown in panel A, B-hCD47 MC38 cells were able to establish tumors in vivo and can be used for efficacy studies. Values are expressed as mean ± SEM.