Gene targeting strategy for B-hCD47 mice.
The IgV domain on exon 2 of mouse CD47 gene was replaced by the IgV domain of human CD47 on exon 2 in B-hCD47 mice.

Strain specific analysis of CD47 gene expression in WT and B-hCD47 mice by RT-PCR. Mouse Cd47 mRNA was only detectable in splenocytes of wild-type (+/+) mice. Human CD47 mRNA was only detectable in H/H but not in +/+ mice.

Strain specific CD47 expression analysis in homozygous B-hCD47 mice by flow cytometry.
Splenocytes were collected from WT and homozygous B-hCD47 (H/H) mice stimulated with anti-CD3ε in vivo and analyzed by flow cytometry with species-specific anti-CD47 antibody. Mouse CD47 was only detectable in WT mice. Human CD47 was exclusively detectable in homozygous B-hCD47 mice but not in WT mice.

Antitumor activity of anti-human CD47 antibody in B-hCD47 mice and C57BL/6 mice. Anti-human CD47 antibody inhibited MC38-hCD47 tumor growth in B-hCD47 mice. (A) Murine colon cancer MC38-hCD47 cells (5×10⁵) were subcutaneously implanted into homozygous B-hCD47 mice (female, 6 week-old, n=6). Mice were grouped when tumor volume reached approximately 150 mm³, at which time they were treated with two anti-human CD47 antibodies and schedules indicated in panel A, as shown in panel A, 3 mg/kg of anti-human CD47 Ab2 was efficacious in controlling tumor growth in the homozygous B-hCD47 mice.(B)Body weight. The results demonstrating that B-hCD47 mice provide a powerful preclinical model for in vivo evaluation of anti-human CD47 antibodies. Values are expressed as mean ± SEM.

Homozygous B-hCD47 mice were injected with various anti-hCD47 antibodies via i.p. injection. Body weight was measured daily. The results indicated toxicity of different antibodies. hCD47 Ab4 and hCD47 Ab6 showed high toxicity levels, as mice died. Y-axis showed average body weight ± SEM.

Homozygous B-hCD47 mice were i.p. injected with anti-hCD47 antibodies. Blood from each mice were collected and Blood Routine Test was conducted. Blood from Ab3 and Ab5 groups were not analyzed because of death of the treated mice.

Blood biochemical analysis of toxicity in mice injected with hCD47 antibody

Homozygous B-hCD47 mice were I.P. injected with anti-hCD47 antibodies. Blood from each mice were collected and Blood Biochemical Test was conducted. Blood from Ab3 and Ab5 groups were not analyzed because of death of the treated mice.