Gene targeting strategy for B-hIL33 mice(C). The exons 2-8 of mouse Il33 gene that encode the full-length protein were replaced by human IL33 exons 2-8 in B-hIL33 mice(C).

Measurement of enhanced pause (Penh) by whole-body plethysmography. Airway responses following the exposure to increasing doses of methacholine (MCh) were measured for each mouse 24 hr after the final allergen or PBS exposure using the whole-body plethysmography. The y-axis represents the Penh absolute value. Increasing doses of methacholine were administered by aerosols. B-hIL33 mice(C) (n = 5) exposed to OVA showed a significant increase in airway hyperreactivity to MCh when compared to exposed PBS inhalation (n = 5) (*P<0.05, *** P<0.01). Penh values were significantly decreased treated with anti-IL33 antibodies (in house) when compared to B-hIL33 mice(C) treated with hIgG4 (n = 5) after allergen exposure [MCh doses of 6.25, 12.5, 25 and 50mg/mL (***P<0.001)].

Analysis of immune cells in BALF. B-hIL33 mice(C) (female, 10-week-old, n=8) were immunized with OVA etc to induce asthma. Anti-human  IL33 antibody (Itepekimab analog, synthesized in-house) and anti-human IL33 antibody (etokimab analog, synthesized in-house) were intraperitoneally injected to B-hIL33 mice(C). (A&B) The number of CD45+ cells and eosinophils of BALF in the Itepekimab and the etokimab treated groups decreased significantly compared with the OVA etc-induced hIgG4 treated group. Values are expressed as mean ± SEM. Significance was determined by two-way ANOVA test.  *P < 0.05, **P < 0.01, ***P < 0.001. 

Analysis of mouse total IgE in serum. B-hIL33 mice(C) (female, 10-week-old, n=8) were immunized with OVA etc to induce asthma. Anti-human  IL33 antibody (Itepekimab analog, synthesized in-house) and anti-human IL33 antibody (etokimab analog, synthesized in-house) were intraperitoneally injected to B-hIL33 mice(C). Serum was collected at the study endpoint. IgE level was analyzed by ELISA. The results showed that the levels of total IgE in mice treated with Itepekimab and etokimab groups showed a significant reduction compared with untreated mice. Values are expressed as mean ± SEM. Significance was determined by two-way ANOVA test.  *P < 0.05, **P < 0.01, ***P < 0.001. 

H&E staining of asthma-like model in B-hIL33 mice(C). Lung tissues were collected at the study endpoint and analyzed with H&E staining. The results showed that the group of mice treated with Itepekimab and etokimab in inflammatory infiltration and mucus secretion in lung tissue was lower than that in untreated mice, indicating that B-hIL33 mice(C) provide a powerful preclinical model for in vivo evaluation of anti-human IL33 antibodies. Values are expressed as mean ± SEM. Significance was determined by unpaired t-test.  *P < 0.05, **P < 0.01, ***P < 0.001.