The exogenous promoter and mouse Ccl2 coding sequence were inserted into the mouse genome randomly. Mouse Ccl2 can be secreted by B-Tg(mCcl2) MC38 cells.

Gene targeting strategy for B-Tg(mCcl2) MC38 cells. The exogenous promoter and mouse Ccl2 coding sequence were inserted into the mouse genome randomly.

Ccl2 expression analysis in B-Tg(mCcl2) MC38 cells by ELISA. Mouse Ccl2 was detected in the supernatant of B-Tg(mCcl2) MC38 cells. The 1-C06 clone of B-Tg(mCcl2) MC38 cells was used for in vivo experiments

Subcutaneous homograft tumor growth of B-Tg(mCcl2) MC38 cells. B-Tg(mCcl2) MC38 cells (5x10⁵) and wild-type MC38 cells (5x10⁵) were subcutaneously implanted into C57BL/6 mice (female, 6-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-Tg(mCcl2) MC38 cells were able to establish tumors in vivo and can be used for efficacy studies.

B-Tg(mCcl2) MC38 tumor growth of individual mice. B-Tg(mCcl2) MC38 cells (5x10⁵) and wild-type MC38 cells (5x10⁵) were subcutaneously implanted into C57BL/6 mice (female, 6-week-old, n=5). As shown in panel, B-Tg(mCcl2) MC38 cells were able to establish tumors in vivo and can be used for efficacy studies.

Tumor cells were harvested at the end of experiment and assessed for mouse Ccl2 expression by ELISA. As shown, mouse Ccl2 was highly expressed in the tumor homogenate. Data was shown as Mean±SEM, and analyzed using T tests compared with G1. (*p<0.05, **p<0.01, ***p<0.001, ****p<0.0001)