• IL-23 is a heterodimeric cytokine composed of p40 and p19 subunits, primarily produced by macrophages and dendritic cells. IL-23 binds to its receptor IL-23R, which regulates the release of downstream pro-inflammatory cytokines.
• Integrin α4β7 is composed of a 150 kD (α4 or CD49d) and a 130 kD (β7) heterodimer, also known as CD49d/β7 or LPAM-1. Belonging to the Ig superfamily, it is found on the majority of peripheral lymphocytes and subsets of thymocytes and bone marrow cells (including mast cell progenitors). Integrin α4β7 binds its ligands, VCAM-1 (CD106), MAdCAM-1 and fibronectin, and plays an important role in lymphocytes adhesion and the direction of migration of blood lymphocytes to the intestine and associated lymphoid tissues.
• The genome of the mouse Il23a gene encoding the full-length protein was replaced with human IL23A counterpart in B-hIL23A/hIL12B/hα4β7 mice. The genome of the mouse Il12b gene encoding the full-length protein was replaced with human IL12B counterpart in B-hIL23A/hIL12B/hα4β7 mice. The genome of the mouse Itga4 gene encoding the extracellular domain was replaced with human ITGA4 counterpart in B-hIL23A/hIL12B/hα4β7 mice. The genome of the mouse Itgb7 gene encoding the extracellular domain was replaced with human ITGB7 counterpart in B-hIL23A/hIL12B/hα4β7 mice.
• Human IL23, ITGA4, and ITGB7 protein were only detectable in homozygous B-hIL23A/hIL12B/hα4β7 mice.
• This product is used for the evaluation of the pharmacodynamics and safety of anti-human IL23/α4β7 antibodies in autoimmune diseases such as inflammatory bowel disease.

Gene targeting strategy for B-hIL23A/hIL12B/hα4β7 mice

The exons 1-4 of mouse Il23a gene that encode the whole molecule (ATG to STOP codon) were replaced by human counterparts in B-hIL23A/hIL12B/hα4β7 mice. The promoter, 5’UTR and 3’UTR region of the mouse gene were retained. The human IL23A expression was driven by endogenous mouse Il23a promoter, while mouse Il23a gene transcription and translation will be disrupted.

The exons 2-8 of mouse Il12b gene that encode the whole molecule (ATG to STOP codon), including 3’UTR were replaced by human counterparts in B-hIL23A/hIL12B/hα4β7 mice. The promoter and 5’UTR region of the mouse gene were retained. The human IL12B expression was driven by endogenous mouse Il12b promoter, while mouse Il12b gene transcription and translation will be disrupted.

The exons 2-27 of mouse Itga4 gene that encode the extracellular domain were replaced by human counterparts in B-hIL23A/hIL12B/hα4β7 mice. The promoter, 5’UTR and 3’UTR region of the mouse gene were retained. The human ITGA4 expression was driven by endogenous mouse Itga4 promoter, while mouse Itga4 gene transcription and translation will be disrupted.

The exons 2-14 of mouse Itgb7 gene that encode the extracellular domain were replaced by human counterparts in B-hIL23A/hIL12B/hα4β7 mice. The promoter, 5’UTR and 3’UTR region of the mouse gene were retained. The human ITGB7 expression was driven by endogenous mouse Itgb7 promoter, while mouse Itgb7 gene transcription and translation will be disrupted.

Mouse IL-23 and human IL-23 expression analysis in B-hIL23A/hIL12B/hα4β7 mice by ELISA. Bone marrow derived dendritic cells were produced by culturing the bone marrow from wild-type C57BL/6JNifdc mice (+/+) and homozygous B-hIL23A/hIL12B/hα4β7 mice (H/H;H/H;H/H;H/H) (female, 7-week-old, n=3), which were stimulated with 1 μg/mL LPS in vitro for 24h. After stimulation, the supernatants were collected and the levels of mouse and human IL23 were analyzed by ELISA. Mouse IL23 was only detectable in wild-type C57BL/6JNifdc mice. Human IL23 was exclusively detectable in homozygous B-hIL23A/hIL12B/hα4β7 mice. Values are expressed as mean ± SEM. ND: not detectable.

Strain specific ITGA4 and ITGB7 expression analysis in wild-type C57BL/6JNifdc mice and homozygous humanized B-hIL23A/hIL12B/hα4β7 mice by flow cytometry. Splenocytes were collected from wild-type C57BL/6JNifdc mice (+/+) and homozygous B-hIL23A/hIL12B/hα4β7 mice (H/H;H/H;H/H;H/H). Protein expression was analyzed with anti-mouse ITGA4 antibody (Biolegend, 103705), anti-mouse LPAM-1/α4β7 antibody (Biolegend, 120607), anti-human ITGA4 antibody (Biolegend, 304307) anti-human ITGB7 antibody (Invitrogen, MA5-23541) by flow cytometry. Mouse ITGA4 and ITGB7 were only detectable in wild-type C57BL/6JNifdc mice. Human ITGA4 and ITGB7 were exclusively detectable in homozygous B-hIL23A/hIL12B/hα4β7 mice, but not in wild-type C57BL/6JNifdc mice.

Strain specific ITGA4 and ITGB7 expression analysis in wild-type C57BL/6JNifdc mice and homozygous humanized B-hIL23A/hIL12B/hα4β7 mice by flow cytometry. Splenocytes were collected from wild-type C57BL/6JNifdc mice (+/+) and homozygous B-hIL23A/hIL12B/hα4β7 mice (H/H;H/H;H/H;H/H). Protein expression was analyzed with anti-mouse ITGA4 antibody (Biolegend, 103705), anti-mouse LPAM-1/α4β7 antibody (Biolegend, 120607), anti-human ITGA4 antibody (Biolegend, 304307) anti-human ITGB7 antibody (Invitrogen, MA5-23541) by flow cytometry. Mouse ITGA4 and ITGB7 were only detectable in wild-type C57BL/6JNifdc mice. Human ITGA4 and ITGB7 were exclusively detectable in homozygous B-hIL23A/hIL12B/hα4β7 mice, but not in wild-type C57BL/6JNifdc mice.

Strain specific ITGA4 and ITGB7 expression analysis in wild-type C57BL/6JNifdc mice and homozygous humanized B-hIL23A/hIL12B/hα4β7 mice by flow cytometry. Splenocytes were collected from wild-type C57BL/6JNifdc mice (+/+) and homozygous B-hIL23A/hIL12B/hα4β7 mice (H/H;H/H;H/H;H/H). Protein expression was analyzed with anti-mouse ITGA4 antibody (Biolegend, 103705), anti-mouse LPAM-1/α4β7 antibody (Biolegend, 120607), anti-human ITGA4 antibody (Biolegend, 304307) anti-human ITGB7 antibody (Invitrogen, MA5-23541) by flow cytometry. Mouse ITGA4 and ITGB7 were only detectable in wild-type C57BL/6JNifdc mice. Human ITGA4 and ITGB7 were exclusively detectable in homozygous B-hIL23A/hIL12B/hα4β7 mice, but not in wild-type C57BL/6JNifdc mice.

Strain specific ITGA4 and ITGB7 expression analysis in wild-type C57BL/6JNifdc mice and homozygous humanized B-hIL23A/hIL12B/hα4β7 mice by flow cytometry. Splenocytes were collected from wild-type C57BL/6JNifdc mice (+/+) and homozygous B-hIL23A/hIL12B/hα4β7 mice (H/H;H/H;H/H;H/H). Protein expression was analyzed with anti-mouse ITGA4 antibody (Biolegend, 103705), anti-mouse LPAM-1/α4β7 antibody (Biolegend, 120607), anti-human ITGA4 antibody (Biolegend, 304307) anti-human ITGB7 antibody (Invitrogen, MA5-23541) by flow cytometry. Mouse ITGA4 and ITGB7 were only detectable in wild-type C57BL/6JNifdc mice. Human ITGA4 and ITGB7 were exclusively detectable in homozygous B-hIL23A/hIL12B/hα4β7 mice, but not in wild-type C57BL/6JNifdc mice.

Strain specific ITGA4 and ITGB7 expression analysis in wild-type C57BL/6JNifdc mice and homozygous humanized B-hIL23A/hIL12B/hα4β7 mice by flow cytometry. Splenocytes were collected from wild-type C57BL/6JNifdc mice (+/+) and homozygous B-hIL23A/hIL12B/hα4β7 mice (H/H;H/H;H/H;H/H). Protein expression was analyzed with anti-mouse ITGA4 antibody (Biolegend, 103705), anti-mouse LPAM-1/α4β7 antibody (Biolegend, 120607), anti-human ITGA4 antibody (Biolegend, 304307) anti-human ITGB7 antibody (Invitrogen, MA5-23541) by flow cytometry. Mouse ITGA4 and ITGB7 were only detectable in wild-type C57BL/6JNifdc mice. Human ITGA4 and ITGB7 were exclusively detectable in homozygous B-hIL23A/hIL12B/hα4β7 mice, but not in wild-type C57BL/6JNifdc mice.