Background:

• S100A4 is secreted by immune cells and promotes cell motility, invasion, cell survival, and angiogenesis by binding to receptors. Its functions include promoting tumor metastasis and the progression of fibrotic and chronic inflammatory diseases.
• S100A4 is expressed at low levels in certain tissues, such as the lungs and lymph nodes, rather than being expressed systemically. It is also secreted by some tumor cells, and neutralizing antibodies against S100A4 have been demonstrated to inhibit tumor metastasis and growth in mouse models.The S100A4 protein is expressed in both lymphoid and myeloid lineages across multiple cell types, including macrophages, neutrophils, mast cells, and memory T cells, as well as in various fibroblasts and tumor cells.

Targeting strategy:

• The exons 1-3 of mouse S100a4 gene that encode the whole molecule (ATG to STOP codon) are replaced by the exons 1-4 of human S100A4 gene in B-hS100A4 mice. The promoter, 5’UTR and 3’UTR region are also replaced by human counterparts in B-hS100A4 mice. The human S100A4 expression is driven by endogenous human S100A4 promoter, while mouse S100a4 gene transcription and translation will be disrupted.

Validation:

• Human S100A4 was detectable in plasma of B-hS100A4 mice.
• Mouse S100a4 mRNA was detectable only in wild-type mice. Human S100A4 mRNA was detectable only homozygous B-hS100A4 mice but not in wild-type mice.

Application:

• B-hS100A4 mice can be used as a preclinical disease model to evaluate the in vivo efficacy and safety of drugs targeting human S100A4.

Gene targeting strategy for B-hS100A4 mice. The exons 1-3 of mouse S100a4 gene that encode the whole molecule (ATG to STOP codon) are replaced by the exons 1-4 of human S100A4 gene in B-hS100A4 mice. The promoter, 5’UTR and 3’UTR region are also replaced by human counterparts in B-hS100A4 mice. The human S100A4 expression is driven by endogenous human S100A4 promoter, while mouse S100a4 gene transcription and translation will be disrupted.

Strain specific S100A4 expression analysis in wild-type C57BL/6JNifdc mice and homozygous humanized B-hS100A4 mice by ELISA. Plasma was collected from wild-type C57BL/6JNifdc mice (+/+) (male, n=3, 7-week-old) and homozygous B-hS100A4 mice (H/H) (male, n=3, 7-week-old). Expression level of mouse and human S100A4 were analyzed by ELISA (anti-mouse S100A4 ELISA kit: abcam, ab309320; anti-human S100A4 ELISA kit: abcam, ab283547). Mouse S100A4 was only detectable in wild-type C57BL/6JNifdc mice. Human S100A4 was detectable in plasma of B-hS100A4 mice and wild-type C57BL/6JNifdc mice, the antibody was cross-reactive between human and mouse. Values are expressed as mean ± SEM.

Strain specific analysis of S100A4 mRNA expression in wild-type C57BL/6JNifdc mice and B-hS100A4 mice by RT-PCR. Kidney and lung RNA were isolated from wild-type C57BL/6JNifdc mice (+/+) and homozygous B-hS100A4 mice (H/H), then cDNA libraries were synthesized by reverse transcription, followed by PCR with mouse or human S100A4 primers. Mouse S100a4 mRNA was detectable only in wild-type C57BL/6JNifdc mice. Human S100A4 mRNA was detectable only in homozygous B-hS100A4 mice but not in wild-type mice.