Gene targeting strategy for B-hTIGIT mice. The exon 2 of mouse Tigit gene that encode the extracellular domain was replaced by human TIGIT exon 2 in B-hTIGIT mice.

Strain specific analysis of TIGIT gene expression in WT and hTIGIT mice by RT-PCR. Mouse Tigit mRNA was detected in splenocytes of wild-type (+/+). Human TIGIT mRNA was detected only in homozygous B-hTIGIT mice (H/H), but not in WT mice.

Strain specific TIGIT expression analysis in homozygous B-hTIGIT mice by flow cytometry. Splenocytes were collected from WT and homozygous B-hTIGIT (H/H) mice stimulated with anti-CD3ε in vivo (7.5 µg/mice), and analyzed by flow cytometry with species-specific anti-TIGIT antibody. Mouse TIGIT was exclusively detected in WT mice. Human TIGIT were exclusively detected in homozygous B-hTIGIT mice (H/H) but not WT mice.

TIGIT expression analysis in wild-type C57BL/6 mice and homozygous B-hTIGIT mice by flow cytometry. Spleen NK cells were collected from wild-type C57BL/6 mice (+/+) and homozygous B-hTIGIT mice (H/H) (female, 6-week-old), that stimulated with anti-mCD3ε antibody in vivo (7.5 μg/mice for 24 hours, i.p.; BioXcell, BE0001-2). Protein expression was analyzed with anti-hTIGIT mAb Belrestotug analog (in house) by flow cytometry. Human TIGIT was detected in NK cells of homozygous B-hTIGIT mice.

TIGIT expression analysis in wild-type C57BL/6 mice and homozygous B-hTIGIT mice by flow cytometry. Spleen T cells were collected from wild-type C57BL/6 mice (+/+) and homozygous B-hTIGIT mice (H/H) (female, 6-week-old), that stimulated with anti-mCD3ε antibody in vivo (7.5 μg/mice for 24 hours, i.p.; BioXcell, BE0001-2). Protein expression was analyzed with anti-hTIGIT mAb Belrestotug analog (in house) by flow cytometry. Human TIGIT was detected in T cells of homozygous B-hTIGIT mice.

TIGIT expression analysis in wild-type C57BL/6 mice and homozygous B-hTIGIT mice by flow cytometry. Spleen T cells were collected from wild-type C57BL/6 mice (+/+) and homozygous B-hTIGIT mice (H/H) (female, 6-week-old), that stimulated with anti-mCD3ε antibody in vivo (7.5 μg/mice for 24 hours, i.p.; BioXcell, BE0001-2). Protein expression was analyzed with anti-hTIGIT mAb Belrestotug analog (in house) by flow cytometry. Human TIGIT was detected in CD8+ T cells of homozygous B-hTIGIT mice.

TIGIT expression analysis in wild-type C57BL/6 mice and homozygous B-hTIGIT mice by flow cytometry. Spleen T cells were collected from wild-type C57BL/6 mice (+/+) and homozygous B-hTIGIT mice (H/H) (female, 6-week-old), that stimulated with anti-mCD3ε antibody in vivo (7.5 μg/mice for 24 hours, i.p.; BioXcell, BE0001-2). Protein expression was analyzed with anti-hTIGIT mAb Belrestotug analog (in house) by flow cytometry. Human TIGIT was detected in CD4+ T cells of homozygous B-hTIGIT mice.

TIGIT expression analysis in wild-type C57BL/6 mice and homozygous B-hTIGIT mice by flow cytometry. Spleen T cells were collected from wild-type C57BL/6 mice (+/+) and homozygous B-hTIGIT mice (H/H) (female, 6-week-old), that stimulated with anti-mCD3ε antibody in vivo (7.5 μg/mice for 24 hours, i.p.; BioXcell, BE0001-2). Protein expression was analyzed with anti-hTIGIT mAb Belrestotug analog (in house) by flow cytometry. Human TIGIT was detected in Treg cells of homozygous B-hTIGIT mice.

Antitumor activity of anti-human TIGIT antibody in B-hTIGIT mice. (A) Anti-human TIGIT antibody inhibited MC38 tumor growth in B-hTIGIT mice. Murine colon cancer MC38 cells (5ⅹ105) were subcutaneously implanted into homozygous B-hTIGIT mice (male, 4-6 week-old, n=5). Mice were grouped when tumor volume reached approximately 150±50 mm3, at which time they were treated with anti-human TIGIT antibody and schedules indicated in panel. (B) Body weight changes during treatment. As shown in panel A, anti-human TIGIT antibody was efficacious in controlling tumor growth in B-hTIGIT mice, demonstrating that the B-hTIGIT mice provide a powerful preclinical model for in vivo evaluation of anti-human TIGIT antibodies. Values are expressed as mean ± SEM.

Antitumor activity of anti-human TIGIT antibodies in B-hTIGIT mice. (A) Anti-human TIGIT antibodies inhibited MC38 tumor growth in B-hTIGIT mice. Murine colon cancer MC38 cells (5ⅹ105) were subcutaneously implanted into homozygous B-hTIGIT mice (female, 8-week-old, n=6). Mice were grouped when tumor volume reached approximately 150±50 mm3, at which time they were treated with anti-human TIGIT antibodies with different doses and schedules indicated in panel. (B) Body weight changes during treatment. As shown in panel A, anti-human TIGIT antibodies were efficacious in controlling tumor growth in B-hTIGIT mice, demonstrating that the B-hTIGIT mice provide a powerful preclinical model for in vivo evaluation of anti-human TIGIT antibodies. Values are expressed as mean ± SEM.

Antitumor activity of anti-human TIGIT antibodies in B-hTIGIT mice. (A) Anti-human TIGIT antibodies inhibited MC38 tumor growth in B-hTIGIT mice. Murine colon cancer MC38 cells (1ⅹ106) were subcutaneously implanted into homozygous B-hTIGIT mice (female, 7-week-old, n=7). Mice were grouped when tumor volume reached approximately 150±50 mm3, at which time they were treated with two anti-human TIGIT antibodies and schedules indicated in panel. (B) Body weight changes during treatment. As shown in panel A, anti-human TIGIT antibodies were efficacious in controlling tumor growth in B-hTIGIT mice, demonstrating that the B-hTIGIT mice provide a powerful preclinical model for in vivo evaluation of anti-human TIGIT antibodies. Values are expressed as mean ± SEM.