Oral Immunotherapy for Pollen Allergy Using T-Cell Epitope-Containing Egg White Derived from Genetically Manipulated Chickens

Abstract

Peptide immunotherapy using T-cell epitopes is expected to be an effective treatment for allergic diseases such as Japanese cedar (Cryptomeria japonica; Cj) pollinosis. To develop a treatment for pollen allergy by inducing oral tolerance, we generated genetically manipulated (GM) chickens by retroviral gene transduction, to produce a fusion protein of chicken egg white lysozyme and a peptide derived from seven dominant human T-cell epitopes of Japanese cedar pollen allergens (cLys-7crp). The transgene sequence was detected in all chickens transduced with the retroviral vector. Transduction efficiency in blood cells correlated to transgene expression. Western blot analysis revealed that cLys-7crp was expressed in the egg white of GM hens. Mice induced to develop allergic rhinitis by Cj pollinosis were fed with cLys-7crp-containing egg white produced by GM chickens. Total and Cj allergen (Cry j 1)-specific IgE levels were significantly decreased in allergic mice fed with cLys-7crp-containing egg white compared with allergic mice fed with normal egg white. These results suggest that oral administration of T-cell epitope-containing egg white derived from GM chickens is effective for the induction of immune tolerance as an allergy therapy.

Transcutaneous Peptide Immunotherapy of Japanese Cedar Pollinosis Using Solid-in-Oil Nanodispersion Technology

“…Auricle sections were prepared from mouse auricles subjected to the S/O nanodispersion for 24 h and analyzed by fluorescence microscopy with an objective lens of ×20 determinants and, thus, was hydrophobic and intrinsically had a low solubility in water. Therefore, 7crp was not produced in large quantity in E. coli unless it was fused to a solubilityenhancing glutathione S-transferase (GST)-tag as reported previously (16). Since we assumed that a rather higher solubility in body fluid (more than 1 mg/mL) was desirable for administration by injection or by S/O nanodispersion system, we introduced small linkers (triarginine) between the epitopes in 7crp and designed a new peptide 7crpR (Fig.…”

“…4a) (16). Sensitization was confirmed by induction of serum total IgE which was not detected in the serum of an unprimed mouse.…”

“…Mice (9 weeks old) were sensitized as previously described (16,22). Briefly, a mixture of 10 μg of Cj pollen extract and 4 mg of Imject Alum was subcutaneously injected to mice three times with 1-week intervals.…”

“…Sensitization was confirmed by induction of serum total IgE which was not detected in the serum of an unprimed mouse. The mouse model was reported to be suitable as a mouse model developing Japanese cedar pollinosis with nasal symptoms (24), and its symptoms to be reduced by inoculation of peptide 7crp (16,17). We examined the therapeutic effect of transcutaneous administration of 7crpR to a rhinitis mouse model.…”

“…For the Cj pollinosis, two enzymes (Cry j 1 and Cry j 2) present in Cj pollen are considered the major allergens (10), and several immunodominant human T cell epitopes were determined and evaluated using blood samples from Cj pollinosis patients (11)(12)(13). Peptides comprised of multiple T cell epitopes administered orally effectively reduced allergic responses represented by allergen-specific antibody responses, T cell proliferation, and number of sneezing events (14)(15)(16)(17). Oral administration of protein/peptide drugs including edible vaccines is needle free and convenient and utilizes the whole length of the gastrointestinal tract; however, dosage and vaccine-food interactions are the concerns (18).…”