Grass Pollen Allergen-Induced Surface Expression of CD203c, CD63 and CD107a on CRTH2+ Basophils: Novel Biomarkers for Monitoring Efficacy of Allergen-Specific Immunotherapy

Background: Grass pollen immunotherapy is associated with reduction in symptoms, the need for rescue medication and improvement of quality of life in patients with severe seasonal pollinosis. Although, the suppression of the early allergic response following in vivo cutaneous allergen challenge is associated with inhibition of basophil histamine release, the effects of immunotherapy on basophil reactivity is yet to be fully determined. We hypothesized that basophil reactivity as measured by increased expression of surface activation markers CD203c, CD63 and CD107a on CRTH2+ basophils is increased in grass pollen allergic individuals following in vitro allergen stimulation. We further hypothesized that this hypereactivity is reduced in immunotherapy-treated patients.

Methods: Heparinized blood obtained from grass pollen allergics (n=7), immunotherapy treated patients (n=6) and non-atopic controls (n=9) was incubated with 0, 0.1, 1, 10 and 100ng/mL of P. Pratense extract at 37°C for 15 minutes. Cells were stained with anti-CD3, CRTh2, CD123, CD303, CD203c, CD63 and CD107a. Additionally, whole blood basophil histamine release was measured pre/post immunotherapy by ELISA (n=6).

Results: A dose-dependent increase in the proportion of CD203c+, CD63+ and CD107a+ CRTH2+ basophils was observed following in vitro grass pollen stimulation in allergics but not in non-atopic controls. At 100ng/mL of P. Pratense extract, CD203c+, CD63+ and CD107a+CRTH2+ basophils were significantly elevated in allergics compare to non-topics (p<0.001, p<0.001 and p<0.009). This increase in CD203c+, CD63+CRTH2+ basophils in allergic individuals significantly correlated with timothy-specific IgE (r=0.84, p<0.0001; r=0.85, p<0.0001). Interestingly, 10-100-fold more allergen was required for CRTH2+ basophils to express CD203c, CD63 and CD107a in immunotherapy-treated patients compare to grass pollen allergics. At suboptimal allergen-concentration (10ng/mL), CD203c+, CD63+ and CD107a+CRTH2+ basophils were significantly reduced in immunotherapy treated subjects compare to allergics (p<0.001, p<0.001 and p<0.002). Basophil histamine release measured after treatment was significantly reduced compared to pre-treatment levels (p< 0.03).

Conclusions: Basophil reactivity and histamine release is significantly reduced following grass pollen immunotherapy. The use of surface activation markers CD203c, CD63 and CD107a on basophils for monitoring clinical efficacy requires further investigations.