B-hIL31/hIL31RA/hOSMR/hIL4/hIL4RA mice_Biocytogen Pharmaceuticals (Beijing) Co., Ltd._Biocytogen,drug development,antibody discovery

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動物・細胞モデル

Humanized Mouse Models

Cytokine Humanized Mice

Text

B-hIL31/hIL31RA/hOSMR/hIL4/hIL4RA mice

Strain Name

C57BL/6-Il31^{tm1(IL31)Bcgen} Il31ra^{tm1(IL31RA)Bcgen} Osmr^{tm1(OSMR)Bcgen}

Il4^{tm2(IL4)Bcgen} Il4ra^{tm1(IL4RA)Bcgen}/Bcgen

Common Name

B-hIL31/hIL31RA/hOSMR/hIL4/hIL4RA mice

Background

C57BL/6

Catalog number

112908

Related Genes

CD124, IL-4RAA, IL4R;

CRL, CRL3, GLM-R, GLMR, GPL, IL-31RA, PLCA2, PRO21384, hGLM-R;

BCGF-1, BCGF1, BSF-1, BSF1, IL-4

mRNA expression analysis-IL31

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Strain specific analysis of IL31 mRNA expression in wild-type C57BL/6 mice and B-hIL31/hIL31RA/hOSMR/hIL4/hIL4RA mice by RT-PCR. Testis RNA were isolated from wild-type C57BL/6 mice (+/+) and homozygous B-hIL31/hIL31RA/hOSMR/hIL4/hIL4RA mice (H/H), then cDNA libraries were synthesized by reverse transcription, followed by PCR with mouse or human IL31 primers. Mouse IL31 mRNA was only detectable in wild-type mice. Human IL31 mRNA was exclusively detectable in homozygous B-hIL31/hIL31RA/hOSMR/hIL4/hIL4RA mice but not in wild-type mice.

mRNA expression analysis-IL31RA

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Strain specific analysis of IL31RA mRNA expression in wild-type C57BL/6 mice and B-hIL31/hIL31RA/hOSMR/hIL4/hIL4RA mice by RT-PCR. Testis RNA were isolated from wild-type C57BL/6 mice (+/+) and homozygous B-hIL31/hIL31RA/hOSMR/hIL4/hIL4RA mice (H/H), then cDNA libraries were synthesized by reverse transcription, followed by PCR with mouse or human IL31RA primers. Human IL31RA mRNA was exclusively detectable in homozygous B-hIL31/hIL31RA/hOSMR/hIL4/hIL4RA mice but not in wild-type mice.

mRNA expression analysis-OSMR

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Strain specific analysis of OSMR mRNA expression in wild-type C57BL/6 mice and B-hIL31/hIL31RA/hOSMR/hIL4/hIL4RA mice by RT-PCR. Kidney RNA were isolated from wild-type C57BL/6 mice (+/+) and homozygous B-hIL31/hIL31RA/hOSMR/hIL4/hIL4RA mice (H/H), then cDNA libraries were synthesized by reverse transcription, followed by PCR with mouse or human OSMR primers. Human OSMR mRNA was exclusively detectable in homozygous B-hIL31/hIL31RA/hOSMR/hIL4/hIL4RA mice but not in wild-type mice.

Protein expression analysis

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Strain specific IL4 expression analysis in homozygous B-hIL31/hIL31RA/hOSMR/hIL4/hIL4RA mice by ELISA. Serum were collected from wild-type mice C57BL/6 mice (n=3, 6-week-old, female) and homozygous B-hIL31/hIL31RA/hOSMR/hIL4/hIL4RA mice (n=3, 6-week-old, female) that stimulated with anti-mCD3e (7.5 μg/mice, i.p.) and anti-mCD28 (5 μg/mice, i.p.) in vivo for 3 h, and analyzed by ELISA with species-specific IL4 ELISA kit (mIL4 kit: BioLegend 431104; hIL4 kit: BioLegend 430304). Mouse IL4 was only detectable in wild-type mice (A). Human IL4 was exclusively detectable in homozygous mice but not in wild-type mice (B).

Protein expression analysis in B cells

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Strain specific IL4RA expression analysis in homozygous B-hIL31/hIL31RA/hOSMR/hIL4/hIL4RA mice by flow cytometry. Splenocytes were collected from wild-type C57BL/6 mice and homozygous B-hIL31/hIL31RA/hOSMR/hIL4/hIL4RA mice stimulated with LPS (200 μg/mice, i.p.) for 2 h, and analyzed by flow cytometry with anti-mouse IL4RA antibody (Biolegend, 144803) or anti-human IL4RA antibody (Biolegend, 355005). Mouse IL4RA was only detectable in wild-type mice, while hIL4RA was exclusively detectable in homozygous mice.

Efficacy evaluation of bispecific antibody in AD model of B-hIL31/hIL31RA/hOSMR/hIL4/hIL4RA

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Efficacy evaluation of bispecific antibody in AD model of B-hIL31/hIL31RA/hOSMR/hIL4/hIL4RA mice. A. OXA was applied to dorsal and ear skin of B-hIL31/hIL31RA/hOSMR/hIL4/hIL4RA mice (female, 8-week-old, n=6) on day 0, and then challenge to the same site of skin nine times from days 7 to 25. Dupilumab, bispecific antibody (BsAb) (Dupilumab, or BsAb provided by client) were intraperitoneally injected in G3-G7. Ear tissues and serum were collected at the endpoint on day 26. B. Ear thickness. C. Ear thickness change. Scratching behavior was recorded on day 11 (D), day 23 (E). F. Day 16 Serum-Total IgE. G. Day 26 Serum-Total IgE. The results showed that ear thicknesses and serum IgE levels were decreased in TA1,TA2,TA3, and Dupilumab with or without TA4 treated group. The combination of Dupilumab and TA4 significantly attenuates scratching behaviors on day 11. The data is expressed as mean ± SEM (* p< 0.05, ** p< 0.01, *** p< 0.001, **** p< 0.0001). AD: atopic dermatitis; OXA: oxazolone.

Note: This experiment is a collaboration with the client.

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Efficacy evaluation of bispecific antibody in AD model of B-hIL31/hIL31RA/hOSMR/hIL4/hIL4RA mice. A. Hematoxylin and eosin (H&E) staining. B. Total pathological score. C. Epidermal thickness. D. Eosinophil score. Ear thickness and infiltration scores of eosinophils in ear skin of the groups treated with TA1,TA2 and Dupilumab analog with or without TA4 were decreased significantly compared to that in the isotype control, demonstrating that the B-hIL31/hIL31RA/hOSMR/hIL4/hIL4RA mice provide a powerful preclinical model for in vivo evaluation of bispecific antibody. Infiltration score of eosinophils: 1=slight; 2=mild; 3=moderate; 4=severe. The content of the pathology total score evaluation includes the following aspects: epidermal hyperplasia in skin, erosion/crusting, hyperkeratosis and parakeratosis; inflammatory cell infiltration in dermis and subcutaneous. The data is expressed as mean ± SEM (* p< 0.05, ** p< 0.01, *** p< 0.001, **** p< 0.0001, n.s. no significance).

Note: This experiment is a collaboration with the client.