The mouse Tpbg gene was replaced by human TPBG coding sequence in B-hTPBG CT26.WT cells. Human TPBG is highly expressed on the surface of B-hTPBG CT26.WT cells.

Gene targeting strategy for B-hTPBG CT26.WT cells. The exogenous promoter and human TPBG coding sequence was inserted to replace part of murine Tpbg exon 2. The insertion disrupts the endogenous murine Tpbg gene, resulting in a non-functional transcript.

TPBG expression analysis in B-hTPBG CT26.WT cells by flow cytometry. Single cell suspensions from wild-type CT26.WT and B-hTPBG CT26.WT cultures were stained with species-specific anti-TPBG antibody. Human TPBG was detected on the surface of B-hTPBG CT26.WT cells but not wild-type CT26.WT cells. The 4-H05 clone of B-hTPBG CT26.WT cells was used for in vivo experiments.

B-hTPBG CT26.WT cells were subcutaneously transplanted into BALB/c mice. At the end of the experiment, tumor cells were harvested and assessed for human TPBG expression by flow cytometry. As shown, human TPBG was highly expressed on the surface of tumor cells. Therefore, B-hTPBG CT26.WT cells can be used for in vivo efficacy studies of novel TPBG therapeutics.

Subcutaneous homograft tumor growth of B-hTPBG CT26.WT cells. B-hTPBG CT26.WT cells (1x10⁵) and wild-type CT26.WT cells (1x10⁵) were subcutaneously implanted into BALB/c mice (female, 6-week-old, n=8). 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-hTPBG CT26.WT cells were able to establish tumors in vivo and can be used for efficacy studies.

B-hTPBG CT26.WT tumor cells growth of individual mice. B-hTPBG CT26.WT cells (1x10⁵) and wild-type CT26.WT cells (1x10⁵) were subcutaneously implanted into BALB/c mice (female, 6-week-old, n=8). As shown in panel, B-hTPBG CT26.WT cells were able to establish tumors in vivo and can be used for efficacy studies.