Basic Information
Description
B-NDG B2m KO mice plus combines the severely immunodeficient B-NDG mouse background with the loss of the MHC class I molecule β2m, which does not show a difference in the metabolism of IgG drugs compared to wild-type mice. B-NDG B2m KO mice plus can be useful for studying the in vivo mechanism of xenograft versus host disease and for therapeutic drug efficacy evaluation.
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Details
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Flow cytometric analysis of MHC class l expression
Blood cells, splenocyte, Bone marrow cells were collected from B-NDG B2m KO plus, B-NDG, NOD-scid mice (n=5) and analyzed by flow cytometry with MHC class I(H-2 Kd). MHC class I was detected in B-NDG and NOD-scid mice, but not B-NDG B2m KO plus.
Protein expression analysis(MHC class lI expression in DC)
Flow cytometric analysis of MHC class lI expression. Blood cells, splenocyte, Peritoneal lavage fluids were collected from B-NDG B2m KO plus, B-NDG, NOD-scid mice (n=3) and analyzed by flow cytometry with MHC class II(mIA-IE). Single live cells were gated for DC (CD45+CD11b+CD11c+) population and used for further analysis as indicated here. MHC class II was detectable in B-NDG, NOD-scid mice, and B-NDG B2m KO plus DC subpopulations.
Protein expression analysis(MHC class lI expression in Macrophages)
Flow cytometric analysis of MHC class lI expression. Blood cells, splenocyte, Peritoneal lavage fluids were collected from B-NDG B2m KO plus, B-NDG, NOD-scid mice (n=3) and analyzed by flow cytometry with MHC class II(mIA-IE). Single live cells were gated for macrophages (CD45+CD11b+F4/80+) population and used for further analysis as indicated here. MHC class II was detectable in B-NDG, NOD-scid mice, and B-NDG B2m KO plus macrophage subpopulations.
Flow cytometric analysis of T , B and NK cells
Blood cells, splenocyte, cells were collected from B-NDG B2m KO plus, B-NDG, NOD-scid and C57BL/6 mice (n=5) and analyzed by flow cytometry. T, B and NK cells were not detectable in B-NDG B2m KO plus and B-NDG mice. T, B, cells were undetectable and NK cells were detectable in NOD scid mice. T, B and NK cells were detectable in C57BL/6 mice.
Flow cytometric analysis of Macrophages
Flow cytometric analysis of DC cells and monocytes
Histopatholoical sections of tissues from B-NDG B2M plus and control mice
(A) Spleen from C57BL/6 has normal structure with well-defined follicles. (B) Spleen from NOD-scid show hypoplasia of white pulp. (C, D) Spleen from both B-NDG and B-NDG B2M plus show complete loss of follicular structure. (E) Thymic lobes from C57BL/6 have normal structure with a well-defined cortex. (G) Thymic lobes from NOD scid mouse are hypoplastic and lack a defined cortex.(H) Thymic lobes from both B-NDG 2 mouse are severely hypoplastic,lack a defined cortex. (F) B-NDG B2M plus show no thymic lobes on the normal anatomy location.
Determination of plasma concentrations of human IgG in different mice
Homozygous B-NDG, B-NDG B2m KO mice,B-NDG B2m KO mice plus and C57BL/6 mice were treated with human IgG (n=5). Blood samples were collected at different time point for the PK assay.The results showed that the PK results of B-NDG and B-NDG B2m KO mice plus groups were basically in line with the pharmacokinetic characteristics, with no difference compared with wild-type mice, and the drug concentration of B-NDG B2m KO mice group could not be measured at the time point 2 days later.
B-NDG B2m KO mice plus vs B-NDG mice in human PBMC induced GvHD model
Conclusions: B2M/FcRn mice significantly extend survival compared with B-NDG mice in the human PBMC induced GvHD model. B2M/FcRn mice delay the onset and reduce the severity of GvHD.
Methods: B2M/FcRn (11-week-old female, n=5) and B-NDG (10-week-old female, n=6) mice were engrafted i.v. with 20M human PBMC from three healthy donors (D1-3) on day 0. Clinical signs of GvHD were scored approximately twice a week according to the table below. Data was analyzed in Prism. Values were expressed as mean ± SEM. **: P<0.01.
B-NDG B2m KO mice plus are well suited for human immune system engraftment
Human PBMC (5E6) were intravenous implanted into homozygote B-NDG B2m KO mice plus and B-NDG mice (female, 7-week-old, n=6). Representative flow cytometric analysis of PBMCs from mice after engraftment with human PBMC. B-NDG B2m KO mice plus show a little change in body weight and exhibit longer survival compared with B-NDG. The results showed that the human PBMC engrafted humanized mice model was successfully constructed.
Anti-tumor efficacy in PBMC RKO model in B-NDG B2m KO mice plus (preliminary)
Conclusions: Check point inhibitors inhibited RKO tumor growth in PBMC engrafted B-NDG B2m KO mice plus. B-NDG B2m KO mice plus appeared to show less GvHD symptoms as indicated by maintenance of body weight.
Method: B-NDG B2m KO mice plus (B-NDG background) mice (n=7/8), engrafted with human PBMC and implanted with human RKO tumor, were treated with pembro/ipi (200 ug) BIW, ip.
CAR-T in vivo efficacy study using NCI-H226 CDX Tumor model in B-NDG B2m KO mice plus
Antitumor activity of CAR-T therapy in B-NDG B2m KO mice plus. Human NCI-H226(A) (1E7) were implanted into B-NDG B2m KO mice plus. (A) Mice were grouped when tumor size was approximately 150±50 mm3. at which time they were treated with CAR-T cells (5E5) with schedules indicated in panel. (B) Body weight changes during treatment. The results showed that CAR-T cells differently inhibited tumor growth in B-NDG B2m KO mice plus. B-NDG is a powerful model for human CAR-T cells efficacy study. Values are expressed as mean ± SEM.
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Summary
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1. Flow cytometric analysis of MHC class l and Il expression indicates that MHC class I is not detectable in B-NDG B2m KO mice plus.
2. Flow cytometric analysis using specific markers for T, B and NK cells indicates that T, B and NK cells were not detectable in B-NDG B2m KO plus.
3. Determination of plasma concentrations of human IgG indicates that that the PK results of B-NDG and B-NDG B2m KO mice plus groups were similar to wild-type mice.
4. B-NDG B2m KO mice plus show a little change in body weight and exhibit longer survival after human PBMC engraftment compared with B-NDG.
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Poster
