The mouse Muc1 gene was replaced by human MUC1 coding sequence in B-hMUC1 MC38 cells. Human MUC1 is highly expressed on the surface of B-hMUC1 MC38 cells.

Gene targeting strategy for B-hMUC1 MC38 cells. The exogenous promoter and human MUC1 coding sequence were inserted to replace murine exon 1~7. The insertion disrupts the endogenous murine Muc1 gene, resulting in a non-functional transcript.

MUC1 expression analysis in B-hMUC1 MC38 cells by flow cytometry. Single cell suspensions from wild-type MC38 and B-hMUC1 MC38 cultures were stained with species-specific anti-MUC1 antibody. Human MUC1 was detected on the surface of B-hMUC1 MC38 cells, but not on the surface of wild-type MC38 cells. The 2-C07 clone of B-hMUC1 MC38 cells was used for in vivo experiments.

Subcutaneous homograft tumor growth of B-hMUC1 MC38 cells. B-hMUC1 MC38 cells (5x10⁵) and wild-type MC38 cells (5x10⁵) were subcutaneously implanted into B-hMUC1 mice (female, 8-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-hMUC1 MC38 cells were able to establish tumors in vivo and can be used for efficacy studies.

B-hMUC1 MC38 cells were subcutaneously transplanted into B-hMUC1 mice (n=6), and on 31 days post inoculation, tumor cells were harvested and assessed for human MUC1 expression by flow cytometry. As shown, human MUC1 was highly expressed on the surface of tumor cells. Therefore, B-hMUC1 MC38 cells can be used for in vivo efficacy studies of novel MUC1 therapeutics.