CD47 is an integrin-associated protein (IAP) and is an immunoglobulin superfamily member. CD47 is widely expressed on cell surface and interacts with the signal regulatory protein α (SIRPα), thrombospondin-1 (TSP1) and integrins to mediate a series of responses including apoptosis, proliferation, and immunity. CD47 is an important “self” mark on the cell surface and inhibits macrophagocytosis by interacting with SIRPα on macrophage surface. Animal studies have shown that CD47 antibody is an effective treatment for multiple types of tumors. Therefore, CD47 is a promising new target for tumor immunotherapy following PD- 1/PD-L1 axis.
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.
mRNA Expression Analysis
RT-PCR analysis of CD47 mRNA expression. The CD47, but not mCD47, mRNA was detectable in splenocytes of the homozygous B-hCD47 mice.
Protein Expression Analysis
Splenocytes from both wild type (WT) C57BL/6 and homozygous B-hCD47 mice were analyzed by flow cytometry. Splenocytes from WT mice express murine CD47 while human CD47 was detectable on spleen cells from B-hCD47 homozygous mice.
Human CD47 mAb toxicity evaluation
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.
Human CD47 mAb toxicity evaluation
Homozygous B-hCD47 mice were i.p. injected with anti-hCD47 antibodies. Blood from each mouse were collected and Blood Routine Test was conducted. Blood from Ab3 and Ab5 groups were not analyzed because of death of the treated mice.
CD47 Abs toxicity evaluation
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 mouse were collected and Blood Biochemical Test was conducted. Blood from Ab3 and Ab5 groups were not analyzed because of death of the treated mice.
Human CD47 antibody efficacy evaluation
Modified murine colon cancer MC38 cells (mCD47 is replaced with the hCD47 extracellular domain, named as MC38-hCD47) were subcutaneously implanted into homozygous B-hCD47 mouse. Mice were divided into control and experimental groups (n=6) when the tumor size was about 150±50 mm3. Anti-hCD47 antibodies differently inhibited tumor growth in the homozygous B-hCD47 mice at 3 different doses, suggesting that B-hCD47 mice is an effective model for in vivo CD47 antibody efficacy study. The average ±SEM of tumor sizes are shown in the figure.
- Wang Y, Ni H, Zhou S, He K, Gao Y, Wu W, Wu M, Wu Z, Qiu X, Zhou Y, Chen B, Pan D, Huang C, Li M, Bian Y, Yang M, Miao L, Liu J. Tumor-selective blockade of CD47 signaling with a CD47/PD-L1 bispecific antibody for enhanced anti-tumor activity and limited toxicity. Cancer Immunol Immunother. 2020 Aug 6. doi: 10.1007/s00262-020-02679-5. Epub ahead of print. PMID: 32761423.（B-hCD47 mice，Department of Clinical Pharmacology, First Affiliated Hospital of Soochow University）
- Wang Y, Pan D, Huang C, Chen B, Li M, Zhou S, Wang L, Wu M, Wang X, Bian Y, Yan J, Liu J, Yang M, Miao L. Dose escalation PET imaging for safety and effective therapy dose optimization of a bispecific antibody. MAbs. 2020 Jan-Dec;12(1):1748322. doi: 10.1080/19420862.2020.1748322. PMID: 32275842; PMCID: PMC7153848.（B-hCD47 mice，Department of Clinical Pharmacology, The First Affiliated Hospital of Soochow University）
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