The mouse Tpbg gene was replaced by human TPBG coding sequence in B-hTPBG MC38 cells. Human TPBG is highly expressed on the surface of B-hTPBG MC38 cells.

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

TPBG expression analysis in B-hTPBG MC38 cells by flow cytometry. Single cell suspensions from wild-type MC38 and B-hTPBG MC38 cultures were stained with species-specific anti-TPBG antibody. Human TPBG was detected on the surface of B-hTPBG MC38 cells, but not on the surface of wild-type MC38 cells. The 2-C06 clone of B-hTPBG MC38 cells was used for in vivo experiments.

B-hTPBG MC38 cells were subcutaneously transplanted into C57BL/6N mice (n=5). 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 MC38 cells can be used for in vivo efficacy studies of novel TPBG therapeutics.

Subcutaneous homograft tumor growth of B-hTPBG MC38 cells. B-hTPBG MC38 cells (5x10⁵) and wild-type MC38 cells (5x10⁵) were subcutaneously implanted into C57BL/6N mice (female, 7-week-old, n=5). 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 MC38 cells were able to establish tumors in vivo and can be used for efficacy studies.

B-hTPBG MC38 tumor growth of individual mice. B-hTPBG MC38 cells (5x10⁵) and wild-type MC38 cells (5x10⁵) were subcutaneously implanted into C57BL/6N mice (female, 7-week-old, n=5). As shown in panel, B-hTPBG MC38 cells were able to establish tumors in vivo and can be used for efficacy studies.

Subcutaneous homograft tumor growth of B-hTPBG MC38 cells. B-hTPBG MC38 cells (5x10⁵, 1x10⁶, 5x10⁶) and wild-type MC38 cells (5x10⁵) were subcutaneously implanted into B-hCD3E 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-hTPBG MC38 cells were able to establish tumors in vivo and can be used for efficacy studies.

B-hTPBG MC38 tumor growth of individual mice. B-hTPBG MC38 cells (5x10⁵, 1x10⁶, 5x10⁶) and wild-type MC38 cells (5x10⁵) were subcutaneously implanted into B-hCD3E mice (female, 8-week-old, n=6). As shown in panel, B-hTPBG MC38 cells were able to establish tumors in vivo and can be used for efficacy studies.

Subcutaneous homograft tumor growth of B-hTPBG MC38 cells. B-hTPBG MC38 cells and wild-type MC38 cells were subcutaneously implanted into B-h4-1BB mice (female, 7-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-hTPBG MC38 cells were able to establish tumors in vivo and can be used for efficacy studies. Values are expressed as mean ± SEM.

Subcutaneous homograft tumor growth of B-hTPBG MC38 cells. B-hTPBG MC38 cells and wild-type MC38 cells were subcutaneously implanted into B-h4-1BB mice (female, 7-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-hTPBG MC38 cells were able to establish tumors in vivo and can be used for efficacy studies. Values are expressed as mean ± SEM.

B-hTPBG MC38 tumor growth of individual mice. B-hTPBG MC38 cells and wild-type MC38 cells were subcutaneously implanted into B-h4-1BB mice (female, 7-week-old, n=6). As shown in panel, B-hTPBG MC38 cells were able to establish tumors in vivo and can be used for efficacy studies.