Title:  Modulation of allergic response

United States Patent:  6,773,695

Issued:  August 10, 2004

Inventors:  Kundig; Thomas M. (Zurich, CH); McCormack; Stephen J. (Claremont, CA)

Assignee:  MannKind Corporation (Valencia, CA)

Appl. No.:  804464

Filed:  March 13, 2001

Abstract

The modulation or elimination of an allergic condition according to the invention can be achieved by injecting small amounts of allergen directly into a lymph node, which greatly reduces the potential for side effects.

SUMMARY OF THE INVENTION

It is one object of the present invention to provide an effective method of delivery of allergens.

It is a further object of the invention to provide a method of delivery of allergens which effectively employs a low dosage of allergens.

It is a still further object of the invention to provide a regimen of treatment for allergies which is effective to modulate an allergic condition after only a few injections.

It is an additional object of the invention to provide a mode of treatment with allergens which is characterized by reduced adverse side effects.

This invention contemplates a method of modulating an allergic response of an individual comprising delivery by direct injection of an allergen to a lymph node of said individual whereby the allergic response is modulated. For individuals who lack lymph nodes or who possess defective lymph nodes, the allergen may be delivered to the lymphatic tissue or to an immune cell. In one specific aspect of the invention the allergen is delivered in combination with an adjuvant, or is precipitated on, or bound to a delivery or formulation substance. Still other aspects of the invention are described in the Description of the Preferred Embodiments of the Invention.

This invention contemplates using intranodal delivery of allergens to re-equilibrate the immune system more effectively than can be accomplished utilizing conventional immunotherapy. The invention provides efficient and effective modulation of a wide variety of allergic responses and represents a major improvement over the current approach. For example, using the preferred embodiments, modulation or elimination of an allergic condition can be achieved with as few as 1 to 3 injections. The targeted delivery also allows the use of smaller amounts of allergen than are used in conventional allergy shots, greatly reducing the potential for side effects such as urticaria, dyspnea, syncope, hypotension, myocardial events and even death.

DETAILED DESCRIPTION OF THE INVENTION

The present invention is useful in any animal having an immune system that includes a lymphatic system. Such field of use generally includes mammals and humans. While the primary use of the invention is likely to be in the treatment of humans, it will also be suitable for treatment of animals, including household pets such as dogs and cats.

The present invention involves the delivery of an allergen by injection directly into a lymph node in order to modulate an allergic response of an individual (for example, to elicit an IgG response) more efficiently than is possible by subcutaneous injection. According to the invention, modulation of an allergic response includes, but is not limited to, diminution or elimination of responses such as alterations in specific IgG levels, alterations in IgG ratios, alterations in specific IgE levels, lowered sensitivity to the allergen or to a cross-reactive allergenic agent, alterations in activated basophils (such as the reduction of the amount of surface IgE), alterations in cytokine profiles (such as increase in type 1 cytokines e.g., IL-2 and IFN .gamma., vs. type 2 cytokines e.g., IL-4, IL-5), alterations in Radio-Allergosorbent Test (RAST) results, or skin tests, as well as diminution or elimination of symptoms of an allergic response, such as urticaria, itching, malaise, anxiety, angioedema, constriction of the chest, nausea, vomiting, diarrhea, abdominal pain, dizziness, dyspnea, wheezing, stridor, dysphagia, dysarthria, hoarseness, weakness, confusion, fall in blood pressure, collapse, loss of consciousness, incontinence, cyanosis, mucus production, coughing, shock, stomach cramps, rhinitis, hay fever, asthma, inflammation, and the like.

Intranodal administration of allergens has a number of advantages. Because lower doses of allergen can induce an IgG response more potently when injected directly into a lymph node, there are fewer side effects than observed using the conventional allergy shot regime. Moreover, delivery of the allergen to the lymph node by injection is no more painful to the patient than regular subcutaneous injections. An additional advantage of this method is that only two or three treatments typically are necessary to desensitize an individual against an allergen. This lowers the risk of side effects or reaction to the administration, and results in a significant cost savings compared with traditional allergy treatments.

An "allergen" according to the invention can be any substance or portion thereof that elicits an allergic response. For example, common allergens include bee venom, wasp venom, fire ant venom, pollens, including grass, tree and herb pollens, penicillin and other drugs, anesthetics, serum, animals, animal dander, cockroaches, dust mites, food allergens such as those found in peanuts, tree nuts, milk, fish, shellfish, eggs, soy, wheat, honey, fruits, viruses, bacteria, mold, protozoa, or latex. Allergens also can be any component of the allergen that elicits an allergic response, such as PLA₂ in bee venom or urushiol in poison ivy. Likewise, an allergen can be a mixture of substances or a crude or purified extract of a generally allergenic composition. These allergens can be recovered from a natural source or can be a synthetic or non-naturally occurring substance, such as a recombinant protein, a synthesized peptide, or a mimetic chemical (including a peptide) that elicits an allergic response similar to a naturally occurring allergen.

A composition useful in the practice of the invention comprises one or more allergens or one or more nucleic acid constructs encoding the allergen(s). The nucleic acid construct can be, for example, RNA or DNA or can simply be a naked nucleic acid construct, such as a plasmid or a virus, encoding the allergen. The allergen or nucleic acid construct can, if desired, be delivered to a specific cell type within a lymph node, such as a dendritic cell. A specific cell type can, if desired, be transfected with the nucleic acid construct so that it expresses the allergen. Optionally, the nucleic acid construct can be targeted via a vector to the desired cell type.

Embodiments of the invention can be adapted to address any disease that includes an allergic-type response. For example, a number of newly discovered and/or little understood viral, bacterial, protozoal, and other diseases exist in the human population. To the extent that such diseases have a pathogenic component that involves an allergic response or an IgE response, embodiments of the invention can be used to reduce the virulence of these diseases and/or to protect people against these diseases. The novel delivery method contemplated by this invention can also be used to alleviate the aspects of asthma and anaphylactic shock associated with hypersensitivity to allergens.

The allergen may be encapsulated in a polymeric material to achieve sustained or pulsatile release. The form of the encapsulation can be, for example, nano- or microspheres, injectable gels, or implants. Suitable polymeric materials include, without limitation, biodegradable PLGA (polylactide-co-glycolide) polyesters, polyanhydrides, polysaccharides (e.g., chitosan, starch, alginate), cellulose derivatives (e.g., ethyl cellulose, hydroxypropylmethyl cellulose), proteins (e.g., collagen, albumin), or polyacrylates.

The allergen is preferably delivered in a physiologically acceptable carrier suitable for injection. In general, any physiologically acceptable carrier known for use with vaccines or allergy shots can be used in the practice of this invention. The choice of such carriers includes, without limitation, water, standard saline solutions, dextrose solutions and albumin water, and is easily within the skill of the art.

In one preferred embodiment, the allergen is delivered in combination with an adjuvant. The adjuvant may be, but is not limited to, one or more of the following: alum, BCG, aluminum hydroxide, aluminum phosphate, calcium phosphate, lipid emulsions, lipid or polymeric nano- or microspheres, micelles, liposomes, saponin, lipid A, or muramyl dipeptide, bacterial products, chemokines, cytokines and hormones, chitosan, starch, alginate, cellulose derivatives (e.g., ethyl cellulose, hydroxypropylmethyl cellulose), nucleic acids, or a nucleic acid construct. One or more of these components may be added to enhance the immune response, increase adsorption of the allergen, provide increased comfort to the patient, and/or slow the release of the allergen to prolong exposure. Alternatively, the allergen may be delivered without an adjuvant or in an aqueous form.

The allergen may be delivered in a dose of about 0.01 .mu.g to about 10 .mu.g or about 0.1 .mu.g to 50 .mu.g and more preferably in a dose from about 0.1 .mu.g to about 10 .mu.g, although the optimal dose may vary depending on the allergen being injected, the weight of the patient, the immune system of the patient, and the like. Effective treatment in many cases may be accomplished with one delivery. In some embodiments, treatment includes from 1 to 15 injections. In preferred embodiments, treatment includes from 1 to 5 injections and more preferably 1 to 3 injections. For example, the standard escalation after a test dose of 0.1 .mu.g involves administration of 1 .mu.g followed by 5 .mu.g and 10 .mu.g. Escalation depends on the patient's tolerance of the previous dose. Multiple injections may be delivered periodically, e.g., over a course of days, once or twice per month, or several times per year.

The dose employed during the initial (desensitization) phase can be from about 0.01 .mu.g to about 10 .mu.g or 0.1 .mu.g to 10 .mu.g delivered in from 1 to 5, preferably from 1 to 3, injections of 1 .mu.g, 5 .mu.g and 10 .mu.g over the course of from several days up to 3 months. In preferred embodiments, the allergen is delivered 2 to 3 times, 1 to 2 weeks apart. During desensitization treatment, 50 .mu.l to 200 .mu.l of an allergen-contaiping composition is administered directly into the lymph node starting with very small doses of allergen, from 0.1 .mu.g up to 10 .mu.g. This dose is one-tenth the normal dose for subcutaneous immunotherapy, and therefore the possibility of side effects is minimized.

The dose employed dinjng the maintenance phase can be from about 0.01 .mu.g to about 10 .mu.g or 0.1 .mu.g to 50 .mu.g, preferably 0.1 .mu.g to 20 .mu.g, delivered periodically over the course of from several months to several years. During maintenance treatment, the patient's lymph node is injected with from 0.1 .mu.g to 50 .mu.g of allergen in injections of typically 50 .mu.l to 200 .mu.l each. One skilled in the art will recognize that even smaller quantities of carrier are feasible.

Optionally, a lymph node may be visualized during the injection procedure. Ultrasound, radiological, or other visualization means such as computerized axial tomography (CAT scan), can be used to visualize the lymph node and monitor location of the needle and changes in the lymph node, such as swelling. Injection into the axillary and inguinal lymph nodes is preferred due to ease of ultrasound guided location and injection.

During administration of the allergen, the patient's vital signs typically are closely monitored, and the lymph node reaction is monitored, for example, by ultrasound or other visualization methods.

The technique used for injection is within the skill of the art. One method is to use a dual-chamber syringe in which the allergen is included in one chamber and a liquid carrier in the other, to be mixed prior to injection.

In preferred embodiments of the invention, the allergen is delivered directly to the lymph node during both desensitization treatment and maintenance treatment. Alternatively, the allergen may be delivered directly to the lymph node during the desensitization phase and subcutaneously during the maintenance phase. Although less preferred, some of the benefits of the invention may be conferred if intranodal therapy is employed during desensitization and subcutaneous therapy is employed during maintenance.

To determine the efficacy of the allergen administration, the patient can be tested for baseline reactions before administration begins, using assays such as those for the measurement of IgG and IgE levels, T-cell stimulation, basophil activation, and/or controlled allergen exposures, such as skin tests and bee sting challenge. To determine whether a patient has been desensitized, one or more of these measurements can be compared to measurements taken after administration.  It is well within the skill of the art to identify and employ alternative assays.

Efficacy of intranodal administration has been demonstrated in mouse models. In mouse models, intranodal therapy induces high IgG2a levels and no detectable IgE response, whereas conventional immunization induces high T_H 2-dependent IgE levels and only low T_H 1-dependent IgG2a levels. Stated otherwise, delivery of the intranodal formulations in minute and otherwise non-immunogenic concentrations induces strong T_H 1 responses, leading to higher and longer lasting IgG2a responses than conventional immunotherapy.

The fact that intranodal administration of PLA₂ induces high IgG2a responses was shown when different amounts of PLA₂ were delivered in mice by direct injection into an inguinal lymph node and compared to conventional subcutaneous PLA₂ injections. Blood was sampled for IgG2a antibody responses at 2, 4, 6, 8 and 10 weeks. In contrast to conventional results on IgG2a induction, low dose intranodal therapy with PLA₂ is sufficient to induce a strong IgG2a response, but only a weak IgE response. With these low dose injections, no significant IgE titers were induced by either delivery.

The intranodal therapy, using approximately 1/1000 of the antigens required in the conventional formulation, induced IgG2a titers in five out of five mice with an average titer of 1:1000, whereas conventional therapy with the same dose induced no measurable IgG2a titers. A strong IgG2a response was induced in two out of three mice by intranodal injection of only 0.1 .mu.g of PLA₂. In contrast, conventional treatment with the same dose induced no significant IgG2a response. To induce even a very low IgG2a response against PLA₂ using the conventional therapy, 10 .mu.g of the allergen must be used for immunization.

Allergens or compositions comprising allergens can be provided in a kit. The kit can contain a composition comprising an allergen and a physiologically acceptable carrier, as well as instructions for the methods described herein. The kit also can contain a syringe, such as a dual-chambered syringe. Optionally, the syringe can be prefilled with the allergen-containing composition. If prefilled, the syringe contains an appropriate dosage of the composition, typically not exceeding a concentration of 100 .mu.g/ml.

Claim 1 of 16 Claims

What is claimed is:

1. A method of desensitizing a human to an allergen against which the individual mounts an allergic response, wherein the allergen is selected from the group consisting of venom, animal dander, pollen, and dust mite comprising the step of delivering the allergen directly into a lymph node of the human, wherein no more than three injections of 1 ug once per month of the allergen are sufficient to desensitize the human to the allergen as assessed by sensitivity of the human to the allergen.

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