D1.205 - Immunological characterization of recombinant Sal k 1 produced in Nicotiana benthamiana and potential immunomodulatory role of HSP90 in Salsola kali allergy

In some regions of Spain, Salsola kali is becoming the major cause of pollinosis, due to the climate changes and the soil desertification. Current allergen-specific immunotherapy (AIT) faces challenges due to the high polymorphism of natural extracts. Recombinant allergens produced in genetically modified plants known as "molecular farming" offer a scalable, cost-effective and standardized alternative to produce high quality allergens. Furthermore, the use of immunomodulators like Heat Shock Protein 90 (HSP90) as adjuvants presents a promising strategy to enhance the efficacy of future vaccines. This study aims to characterize rSal k 1 produced in Nicotiana benthamiana and evaluate its potential for epitope mapping and vaccine design.

Eighteen patients with Salsola kali pollen allergy, with positive skin prick tests and specific IgE levels, were recruited in Spain. The recombinant Sal k 1 (rSal k 1) was expressed in N. benthamiana and its identity was confirmed by MALDI-TOF/TOF mass spectrometry (MS). Its ability to bind to IgE was compared with that of its counterpart in Salsola kali pollen (nSal k 1), evaluated by Western blot and inhibition assays. The study is investigating the possible immunomodulatory contribution of HSP90 proteins as part of the approach based on the production of recombinant allergens.

Immunoblotting assays with IgE serum of individual patients revealed comparable results between rSal k 1 and nSal k 1. In inhibition assays, nSal k 1 inhibited the 100% IgE-binding to rSal k 1, while rSal k 1 achieved 74% inhibition of nSal k 1. This partial inhibition highlights the isoform complexity of the pollen but confirms that rSal k 1 displays the major clinical epitopes. These identified epitopes are ideal candidates for vaccine design.

rSal k 1 produced in N. benthamiana is immunologically equivalent to its natural homologue from S. kali pollen and it would be a useful tool for standardizing diagnostic and immunotherapeutic protocols. Combining specific epitopes of this allergen with the adjuvant and immunomodulatory properties of HSP90 could help guide the development of safer and more sustainable vaccination strategies for Salsola kali and other allergies.