WO2010128364A1

WO2010128364A1 - Method for the prevention or treatment of taxodiaceae pollen allergy

Info

Publication number: WO2010128364A1
Application number: PCT/IB2009/053465
Authority: WO
Original assignee: Stallergenes S.A.
Priority date: 2009-05-04
Filing date: 2009-05-04
Publication date: 2010-11-11

Abstract

The present invention relates to the use of at least one extract or of at least one recombinant allergen from a plant of the Cupressaceae family, for the manufacture of a medicament intended for preventing or treating Taxodiaceae pollen allergy in an individual.

Description

Method for the prevention or treatment of taxodiaceae pollen allergy

Field of the invention

The present invention relates to a method for treating Taxodiaceae pollen allergy, in particular Cryptomeria japonica pollen allergy.

Technical background of the invention

Allergies to pollen are continuously on the rise worldwide. Among them, allergies to pollens from grasses, birch, or conifers, such as cedar, are among the most prevalent. Thus, from 2002 to 2008, an increase from about 15% to about 26% of the prevalence of Japanese cedar pollen allergy among the Japanese population has been reported.

The most promising treatments for these allergies are focused on inducing an allergen-specific immune tolerance towards pollen allergen antigens, in particular through sublingual immunotherapy (SLIT), which has been recently explored for antigen administration in a variety of conditions, especially in the field of immunotherapy of allergy.

Allergen-specific sublingual immunotherapy (SLIT) indeed represents a safe and efficient non invasive alternative to subcutaneous immunotherapy (Bahceciler et a/. (2005) Int. Arch. Allergy Immunol. 136:287-294) in particular for the treatment of type I respiratory allergies (Canonica & Passalacqua (2006) Allergy 61 :20-23, Wilson et al. (2005) Allergy 60:4-12). During SLIT, the allergen is captured within the oral mucosa by Langerhans-like dendritic cells expressing high affinity IgE receptors, producing IL10 (interleukin 10) and TGFβ and upregulating the indoleamine dioxygenase (IDO), suggesting that such cells are prone to induce tolerance.

It has thus been shown that the treatment of allergy to Japanese cedar pollen (Cryptomeria japonica, of the Taxodiaceae family) could be carried out by performing SLIT using Japanese cedar pollen extracts (Okubo et al. (2008) Allergol. Int. 57:265-75).

However, production of active extracts from Japanese cedar in quantity sufficient for large scale therapeutic use proves to be an arduous task. Besides, the sticky character of Japanese cedar extracts obtained up to now is an obstacle to the production of tablets, which is the preferred administration form for SLIT.

Accordingly, it is an object of the present invention to provide an alternative treatment, in particular by SLIT, for allergies to Taxodiaceae pollens.

Description of the invention

The present invention arises from the unexpected finding, by the present inventors, that extracts from Mountain cedar pollen (Juniperus ashei, belonging to the Cupressaceae family, mainly growing in Texas, USAJ are at least as potent as extracts from Cryptomeήa japonica pollen for completely inhibiting IgE binding to

Cryptomeria japonica pollen allergens. It has been previously shown that complete inhibition by birch pollen extracts of IgE binding to pollen allergens of other

Betulaceae was predictive of the efficacy of a birch pollen extract for treating pollen allergy to pollen of other plants of the Betulaceae family. This indicates that Juniperus ashei pollen extracts can be used for inducing immune tolerance towards Cryptomeria japonica pollen allergens.

Accordingly, the present invention relates to the use of at least one extract or of at least one recombinant pollen allergen from a plant of the Cupressaceae family, for the manufacture of a medicament intended for preventing or treating Taxodiaceae pollen allergy in an individual, preferably by inducing allergen- specific immune tolerance towards Taxodiaceae pollen allergens.

As intended herein, "Cupressaceae" a n d "Taxodiaceae" relate to two different well-known Linnean taxonomic families.

Preferably, the plant of the Cupressaceae family as defined above is of the Juniperus genus and more preferably of the Juniperus ashei species.

Preferably, the extract as defined above is a pollen extract. Preferably also the extract is a protein extract. The protein extract can notably be obtained by an aqueous extract of pollen, in particular with an ammonium bicarbonate solution, preferably at a concentration of 4 g/l. The extract is preferably such that it contains pollen allergens. Preferably, also the extract can be purified so as to contain essentially only pollen allergens. Procedures for obtaining extracts are well known to one of skill in the art and can be performed essentially as described in Hrabina et a/. (2003) Allergy 58:808-813 for Juniperus ashei extracts. As intended herein an "allergen" is a substance wh ich elicits the production of IgE antibodies in predisposed individuals. Similar definitions are presented in the following references: Clin. Exp. Allergy, No. 26, pp. 494-516

(1996); MoI . Biol, of Allergy and Immunology, ed. R. Bush, Immunology and Allergy Clinics of North American Series (August 1996).

Preferably, the pollen allergens comprised in the extract, or the recombinant pollen allergens, are selected from Juniperus sabinoides or virginiana pollen allergens, such as Jun s I and Jun v I and Juniperus ashei pollen allergens, such as Jun a 1 , Jun a 2, and Jun a 3. The cloning and expression of Jun a 1 , Jun a 2, and Jun a 3 can be easily carried out by one of skill in the art and are notably respectively described in Midoro-Horiuti et al. (1999) J. Allergy CHn. Immunol. 104:613-7; Yokoyama et al. (2000) Biochem. Biophys. Res. Commun. 275:195-202; Moehnke et al. (2008) Biotech nol. Lett. 30:1259-64. Preferably, the Taxodiaceae pollen allergy as defined above is an allergy to pollen from a plant of the Cryptomeria genus, more preferably of the Cryptomeria japonica (i.e. Japanese cedar) species. As will be clear to one of skill in the art, the allergic reaction is directed against pollen allergens of Cryptomeria japonica, such as Cry j 1 , Cry j 2, and Cry j 3. As intended herein "pollen allergy" notably encompasses pollinosis, and pollen-induced asthma. Determining that the pollen allergy is a Taxodiaceae pollen allergy can be easily carried out by one of skill in the art, for instance by carrying skin prick test with pollen extract from a Taxodiaceae, in particular from a Cryptomeria, and more particularly from Cryptomeria japonica. Preferably also, the individual as defined above has stayed in Japan for at least one week at a period of pollen emission by plants of the Taxodiaceae family, such as Cryptomeria japonica, and more preferably the individual is a Japanese resident, i.e. an individual whose main residence is in Japan.

In an embodiment of the invention, the medicament further comprises another extract, in particular a pollen extract, or another recombinant pollen allergen from a plant. The plant can notably be another Cupressaceae, such as Chamaecyparis obtuse (i.e. Japanese cypress) or a Taxodiaceae, such as Cryptomeria japonica. Preferably, the medicament as defined above is suitable for administration by the mucosal route, more preferably by the oromucosal route, and most preferably by the sublingual route. As such, the medicament is preferably formulated in a way suitable for such administration routes. Mucosal administration denotes any administration method wherein the formulation in part or in full comes into contact with a mucosa. Mucosa refers to the epithelial tissue that lines the internal cavities of the body. The mucosal surface may be selected from the group consisting of a nasal, buccal, oral, vaginal, ocular, auditory, pulmonary tract, urethral, digestive tract, and rectal surface.

Oromucosal administration comprises any administration method, wherein the formulation in part or in full comes into contact with the mucosa of the oral cavity and/or the pharynx of the patient.

Oromucosal administration includes in particular sublingual, perlingual (i.e. through the tongue mucosa) and oral administrations.

Where the medicament is administered sublingually to the patient (i.e. under the tongue), the sublingual mucosa, located on the underside of the tongue, facilitates capture of the antigen and adjuvant by Langerhans-like cells that migrate to draining lymph nodes to prime T lymphocytes. A route of administration of particular interest is to keep the composition under the tongue a few minutes, e.g. about 2 minutes, before swallowing or spitting it out.

The medicaments according to the invention can be administered in various forms, such as dispersed forms, e.g. in suspensions or gels, or as dry forms, e.g. in powders, tablets, capsules, lyoc, or forms suitable to be administered in a metered-dosing device.

Preferably, the medicament comprises a mucoadhesive carrier. As intended herein, a mucoadhesive carrier enables close and prolonged contact with a mucosa, in particular a mucosa of the oral cavity, and more particularly the sublingual mucosa, thereby enhancing-antigen specific tolerance induction. Preferred mucoadhesive carriers as defined herein notably comprise chitosan, polymers of maltodexthn or carboxymethylcellulose.

The medicament as defined above may further comprise at least one adjuvant and/or at least one pharmaceutically acceptable excipient. Preferably, the medicament as defined above comprises an adjuvant and/or a mucoadhesive carrier.

As intended herein an "adjuvant" enhances antigen-specific tolerance induction. Adjuvants have been used for many years to improve the host immune responses to, for example, vaccines. The adjuvant as defined herein may include any conventional or exploratory, synthetic or biological adjuvant for vaccination, including heat-labile enterotoxin (LT), cholera-toxin (CT), cholera toxin B subunit (CTB), polymerised liposomes, mutant toxins, probiotic bacteria, saponins complexed to membrane protein antigens (immune stimulating complexes), pluronic polymers with mineral oil, killed mycobacteria in mineral oil, Freund's complete adjuvant, bacterial products, such as muramyl dipeptide (MDP) and lipopolysaccharide (LPS), as well as lipid A, or biological or synthetic ligands of Toll like receptors (eg TLR2, 4, 5, 7 or 9).

As used herein, the term "pharmaceutically acceptable excipient" includes solvents, dispersion media, coatings, isotonic and absorption delaying agents, mucoadhesive excipients, and the like, that do not produce an adverse or other untoward reaction when administered to an animal, or a human, as appropriate. Excipients may further include, but are not limited to disintegrants, binders, lubricants, flavoring, colorants, preservatives. Suitable disintegrants include dry starch, calcium carbonate, polyoxyethylene sorbitan fatty acid esters, sodium lauryl sulfate, stearic monoglyceride, lactose, as well as cross-linked polyvinylpyrrolidones, such as crospovidone (e.g., Polyplasdone™. XL, which may be obtained from GAF), cross-l inked carboxylic methylcelluloses, such as croscarmelose (e.g., Ac-di-sol™, which may be obtained from FMC), alginic acid, and sodium carboxymethyl starches (e.g., Explotab™, which may be obtained from Edward Medell Co., Inc.), methylcellulose, agar bentonite and alginic acid. Binders, if used, are those that enhance adhesion. Examples of such binders include, but are not limited to, starch, gelatin and sugars such as sucrose, dextrose, molasses, and lactose. Preferred lubricants are stearates and stearic acid. Lactose, mannitol, and croscarmelose are preferred excipients.

Additional components that may be incorporated into the dosage forms are known, or will be apparent, to those skilled in this art; for example, see Remington: The Science and Practice of Pharmacy, 19th edition (Mack Publishing, 1995). The invention will be further illustrated in view of the following figures and examples.

Description of the figures

Figure 1 represents inhibition patterns obtained with the sera from a French patient allergic to cypress pollen. Cryptomeria japonica proteins were resolved by SDS Page. Proteins were transferred onto a nitrocellulose membrane, cut in 3 different strips (lanes 1 , 2 and 3) and incubated with either the serum from a

French patient allergic to cypress pollen properly diluted (lane 1 ), the serum previously incubated with Cryptomeria japonica pollen extract in excess (self inhibition, lane 2), the serum previously incubated with Juniperus ashei pollen extract in excess (lane 3).

Examples

Example 1 : Capacity of Juniperus ashei pollen allergens to inhibit the IgE binding of sera from a French cypress allergic patient on Cryptomeria japonica pollen allergens

The objective of the experiment was to study the capacity of Juniperus ashei pollen allergens to inhibit the IgE binding, on Cryptomeria japonica pollen allergens, of sera from a French patient allergic to cypress, by Western-blot. Cryptomeria japonica proteins from a pollen extract obtained with sodium bicarbonate (by following the general teachings of Hrabina et al. (2003) Allergy 58:808-813) were resolved by SDS-Page. Proteins were transferred onto a nitrocellulose membrane, cut in 3 different strips (lanes 1 , 2 and 3) and incubated with either a serum from a French patient allergic to cypress pollen properly diluted (lane 1 ), the serum previously incubated with Cryptomeria japonica pollen extract in excess (self inhibition, lane 2), and the serum previously incubated with Juniperus ashei pollen extract obtained with sodium bicarbonate (for example as described by Hrabina et al. (2003) Allergy 58:808-813) in excess (lane 3). For the revealing step, luminescence signals were detected and quantified using a highly sensitive Chemismart 2000 camera.

The results are shown in Figure 1. The magnitude of signal in lane 1 is 100 %. The signal in lane 2 (self inhibition control) is equal to 0 %. The signal in lane 3 is equal to 0 % in presence of Juniperus ashei pollen extract as inhibitor which indicates that Juniperus ashei pollen allergens totally inhibit the binding of allergen-specific IgEs to Cryptomeria japonica pollen allergens. Accordingly, Juniperus ashei pollen extract is useful for treating allergy to Cryptomeria japonica pollen.

Example 2: Capacity of Juniperus ashei pollen allergens to inhibit the IgE bind ing of sera from 22 Cryptomeria japonica allergic patients on Cryptomeria japonica pollen allergens In a first step, a Western-blot study similar to that of Example 1 is performed, Cryptomeria japonica proteins from a pollen extract are resolved by SDS-Page. Proteins are transferred onto a nitrocellulose membrane, cut in 3 different strips (lanes 1 , 2 and 3) and incubated with either a serum from a Japanese patient allergic to Japanese cedar pollen properly diluted (lane 1 ), the serum previously incubated with Cryptomeria japonica pollen extract in excess (self inhibition, lane 2), and the serum previously incubated with Juniperus ashei pollen extract in excess (lane 3). For the revealing step, luminescence signals are detected and quantified using a highly sensitive Chemismart 2000 camera. The results show that Juniperus ashei pollen allergens totally inhibit the binding of Cryptomeria yapon/ca-reactive IgE to Cryptomeria japonica pollen allergens.

In a second step, both western blot experiments an Elisa-inhibition assay were performed using sera from Japanese patients. Briefly, the wells of an ELISA microtitration plate are coated with Cryptomeria japonica pollen allergens, and filled with mixtures of the serum from a Japanese patient previously incubated with

Cryptomeria japonica pollen extract at varying concentrations and the serum previously incubated with Juniperus ashei pollen extract at varying concentration. The percentage of inhibition of IgE binding to the coated allergens is then determined for each mixture, and two curves plotting the percentage of inhibition vs the respective concentrations of Cryptomeria japonica or Juniperus ashei pollen extracts in the mixtures are established.

It is confirmed that Juniperus ashei pollen extracts can be used for treating Cryptomeria japonica pollen allergy when essentially 100% inhibition, parallel curves, and, optionally, concentrations of Juniperus ashei pollen extracts lower than that of Cryptomeria japonica, to achieve the same percentage of inhibition, are obtained.

Claims

1 . The use of at least one extract or of at least one recombinant pollen allergen from a plant of the Cupressaceae family, for the manufacture of a medicament intended for preventing or treating Taxodiaceae pollen allergy in an individual.

2. The use according to claim 1 , wherein the plant of the Cupressaceae family is of the Juniperus genus.

3. The use according to claim 1 or 2, wherein the plant of the Cupressaceae family is of the Juniperus ashei species.

4. The use according to any of claims 1 to 3, wherein the extract is a pollen extract.

5. The use according to any of claims 1 to 4, wherein the extract is a protein extract.

6. The use according to any of claims 1 to 5, wherein the Taxodiaceae pollen allergy is an allergy to pollen from a plant of the Cryptomeria genus.

7. The use according to any of claims 1 to 6, wherein the Taxodiaceae pollen allergy is an allergy to pollen from a plant of the Cryptomeria japonica species.

8. The use according to any of claims 1 to 7, wherein the individual has stayed in Japan for at least one week at a period of pollen emission by plants of the Taxodiaceae family.

9. The use according to any of claims 1 to 8, wherein the medicament is suitable for administration by the sublingual route.

10. The use according to any of claims 1 to 9, wherein the medicament comprises an adjuvant and/or a mucoadhesive carrier.