B-hB7-H4 mice

Basic Information

mRNA Expression Analysis

Species-specific B7-H4 gene expression analysis in wild type and B-hB7-H4 mice by RT-PCR. Murine B7-h4 mRNA was detected in ovaries isolated from wild-type (+/+) mice, while human B7-H4 mRNA was detected in homozygous B-hB7-H4 (H/H) mice.

Protein Expression Analysis

B7-H4 protein expression analysis in homozygous B-hB7-H4 mice by western blot. Testis tissue was collected from wild type (+/+) mice and homozygous B-hB7-H4 (H/H) mice and analyzed by western blot using an anti-B7-H4 antibody. Mouse B7-H4 protein was detected in wild type mice, while human B7-H4 protein was detected in homozygous B-hB7-H4 mice. Anti-B7-H4 antibody is cross-reactive with B7-H4 in human and mice.

Analysis of spleen leukocytes

Analysis of spleen leukocyte subpopulations. Splenocytes were isolated from female C57BL/6 and B-hB7-H4 mice (n=3, 7-week-old). Flow cytometry analysis of the splenocytes was performed to assess leukocyte subpopulations. (A) Representative flow cytometry plots. Single live cells were gated on CD45+ cells and used for further analysis as indicated. (B) Percent of T cells, B cells, NK cells, dendritic cells, granulocytes, monocytes and macrophages in homozygous B-hB7-H4 mice were similar to those in the C57BL/6 mice, demonstrating that introduction of hB7-H4 in place of its mouse counterpart dose not change the overall development, differentiation or distribution of these spleen cell types. Values are expressed as mean ± SEM.

Analysis of spleen T cells

Analysis of spleen T cells. Splenocytes were isolated from female C57BL/6 and B-hB7-H4 mice (n=3, 7-week-old). Flow cytometry analysis of the splenocytes was performed to assess leukocyte subpopulations. (A) Representative flow cytometry plots. Single live CD45+ cells were gated on TCRβ+ T cells and used for further analysis as indicated. (B) Percent of CD4+ T cells, CD8+ T cells, and Tregs in homozygous B-hB7-H4 mice were similar to those in the C57BL/6 mice, demonstrating that B7-H4 humanization does not change the overall development, differentiation or distribution of these spleen T cell subtypes. Values are expressed as mean ± SEM.

In vivo Efficacy of Anti-Human B7-H4 Antibody

Antitumor activity of an anti-human B7-H4 antibody in humanized B-hB7-H4 mice. Humanized murine colon cancer B-hB7-H4 MC38 cells (3ⅹ10⁶) were subcutaneously implanted into homozygous B-hB7-H4 mice (female, 8-week-old, n=6). B-hB7-H4 mice were grouped when tumor volume reached approximately 100 mm³, at which time FPA-150 (anti-human B7-H4 Ab, in house) was administered at doses and schedules indicated in panel A. (B) Body weight changes during treatment. As shown, FPA-150 inhibited tumor growth without negatively impacting body weight changes, demonstrating that B-hB7-H4 mice provide a powerful preclinical model for in vivo evaluation of anti-human B7-H4 antibodies. Values are expressed as mean ± SEM.

Poster

AACR 2022: A Humanized Mouse Model of the Promising Immune Checkpoint Molecule B7-H4