JP2002511421A - Vaccine composition comprising CD-1 antigen and T cell stimulating compound and method of using the same - Google Patents

Abstract

  (57) [Summary] The present invention relates to immunogenic or vaccine compositions comprising a CD1 or lipid antigen and a T cell stimulating compound (eg, an adjuvant). The invention also relates to a method for administering an immunogenic or vaccine composition.

Description

DETAILED DESCRIPTION OF THE INVENTION

[0001] This application claims the benefit of US Provisional Application No. 60 / 081,638, filed April 13, 1998, the entire teachings of which are incorporated herein by reference. .

(Government Assistance) The present invention provides, in whole or in part, National Institutes of Technology.
Supported by a grant from f Health. The government has certain rights in the invention.

BACKGROUND OF THE INVENTION Vaccination induces a specific immune response in a host against foreign viruses, bacteria, or parasites. Many types of pathogens and their products have been used as vaccines. However, not all vaccine compositions can elicit an immune response sufficient to protect the host against the challenge of infection.

[0004] Similarly, there are many diseases, disorders, or conditions in which the immune system is compromised or suppressed. Many treatments for such diseases or disorders are ineffective or ineffective at boosting or stimulating the immune system. An example of such a disease or condition is an AIDS disease, or a cancer patient continuing chemotherapy.

[0005] These illustrate the examples needed to develop compositions or agents that stimulate or enhance the immune response. There is a need for an enhanced immune response, especially for vaccines that have a sufficiently protective immune response in the host. In addition, there is a need for compositions that stimulate the immune system of an compromised individual.

SUMMARY OF THE INVENTION [0006] The present invention relates to a method of enhancing an immune response to at least one CD1 antigen in a vaccination or mammal. The method comprises at least one CD
Comprising administering (eg, co-administering) an effective amount of one antigen (eg, a non-peptide or lipid antigen) and at least one T cell stimulating compound (eg, an adjuvant) to a mammal (eg, an individual). Where the CD1 antigen is C
Initiate a D1 restricted response. Types of lipid antigens are described herein. Examples of adjuvants are: mineral salt adjuvant, incomplete Freund's adjuvant or complete Freund's adjuvant, Bacillus Calmette-Guerin adjuvant,
Block polymer adjuvant, cholera toxin, cytokine, adjuvant containing CPG motif, oil / water emulsion adjuvant, MF-59 adjuvant, L
eIF adjuvant, liposome adjuvant, ISCOM adjuvant, monophosphoryl A adjuvant, biodegradable microsphere adjuvant, muramyl dipeptide adjuvant, polyphosphodine adjuvant or saponin adjuvant (for example, QS-7, QS-17, Q
S-18 and QS-21). The method of the invention elicits at least one immunological parameter, such as an antibody response to an antigen, a cytotoxic T lymphocyte response, a T cell proliferative response, a helper T cell response or a T cell regulatory cytokine response. This method can be used to enhance or boost the immune response of an individual (eg, AIDS or chemotherapeutic recipient) with an immuno-compromise disease, disorder or condition. Using the present invention, at least one bacterial infection (eg, Mycobacteria, Haemophilus, Streptococcus, Staphylococcus or Ch)
elicits or boosts an immune response against Lamydia species and / or at least one parasitic infection (eg, Plasmodium or Trypanosoma species). The CD1 antigens of the present invention may also include tumor-associated or tumor-derived antigens involved in diseases such as cancer (eg, melanoma, breast, prostate and colorectal), or autoimmune diseases (eg, diabetes, lupus, It may be an autoantigen involved in rheumatoid arthritis). Embodiments of the method may further comprise administering a peptide-based antigen from the same organism or cancer type, such that the individual is protected or enhanced immunologically against non-peptide and peptide antigens. You can get the effect.

[0007] The invention also provides at least one T cell stimulating compound (eg, an adjuvant)
And an immunological or vaccine composition comprising at least one type of CD1 antigen (eg, a non-peptide or lipid antigen). Here, the CD1 antigen elicits a CD1 restricted response. CD1 molecules include CD1a, CD1b, CD1
c, a family of molecules including CD1d and CD1e. Examples of adjuvants are described herein. For this method, the antigen of the composition comprises at least one bacterium (eg, for example, the genus Mycobacteria, Haemop).
genus hilu, Streptococcus, Staphylococcus
Or a species of the genus Chlamydia) and / or at least one parasitic infection (eg, a species of the genus Plasmodium or Trypanosoma).
Derived from Examples of non-peptide antigens are glycolipids, phospholipids, triglycerides, glycosylphosphatidylinositol (GPI), mycolic acid, glucose monomycolate (GMM), lipoarabinomannan (LAM), lipooligosaccharides, or phosphatidylinositol mannoside. (PIM). The composition can optionally include a carrier.

[0008] The present invention also provides at least one T cell stimulating compound (eg, an adjuvant)
And a kit comprising at least one type of CD1 antigen (eg, a lipid antigen). Here, the CD1 antigen elicits a CD1 restricted response. The kit comprises a CD1 derived from at least one bacterium and / or at least one parasite.
(Eg, a lipid antigen) and, optionally, a peptide antigen (eg, a peptide vaccine composition). Examples of types of adjuvants and lipid antigens are described further herein.

[0009] The present invention relates to the administration of an adjuvant / CD1 antigen composition to an individual to gain protection from severe bacterial and / or parasitic infections, or to cancer or autoimmune diseases. It provides more effective prevention and treatment options or provides immunotherapy against cancer or autoimmune disease.

DETAILED DESCRIPTION OF THE INVENTION The present invention provides a vaccine composition or immunological composition comprising at least one type of non-peptide antigen (eg, a lipid antigen) and at least one T cell stimulating compound (eg, an adjuvant). Composition. According to the present invention, an antigen-presenting cell (APC)
Take advantage of the discovery of presenting non-peptide antigens to T cells associated with the D1 molecule.

[0011] CD1 antigens, such as lipid antigens or non-peptide antigens (eg, from bacteria or parasites) are taken up by APCs (eg, macrophages, B cells or dendritic cells) and then combined with the CD1 molecule. And thereby induce T cell proliferation (Porcelli, SA et al., "T-cell recog".
Nation of Non-Peptide Antigen "Curren
t Opinion, Immunology 8: 510-516 (1996).
). This pathway is referred to herein as the “CD1 antigen presentation pathway”. An immune response resulting from the CD1 antigen presentation pathway is called a “CD1 restricted response”. These non-peptide antigens are processed independently of the MHC peptide antigen presentation pathway. 1992 1
US patent application Ser. No. 07 / 989,790 filed on Feb. 10, US Ser. No. 08 / 080,072 filed on Jun. 21, 1993; Jun. 2, 1994
PCT Application No. PCT / US94 / 06911, filed on Oct. 1, US Pat. No. 5,679,347, issued Oct. 21, 1997; US Pat. U.S. Pat. No. 5,853,737, issued Dec. 29, 1998; U.S. Patent Application No. 08 / 501,491 filed Jul. 12, 1995; Oct. 1995 PCT Patent Application PCT / US95 / 13274 filed on the 13th; US Patent Application No. 08 / 817,231 filed on April 11, 1997; US Provisional Application filed on September 12, 1997 U.S. Provisional Patent Application No. 60 / 081,638, filed April 13, 1998.

The present invention utilizes the concept that when the CD1 antigen is combined with a T cell stimulating compound (eg, an adjuvant), it elicits or boosts an immune response against the non-peptide antigen. A composition comprising a CD1 antigen and a T cell stimulating compound produces a significantly more effective immunogenic response than the antigen alone. Therefore, the present invention
An immunological or vaccine composition comprising a CD1 antigen and a T cell stimulating compound, and methods of using them.

[0013] The present invention relates to at least one T cell stimulating compound and at least one CD1
An immunological composition comprising an antigen (eg, a CD1 antigen / adjuvant composition or a non-peptide / adjuvant composition), wherein the CD1 antigen elicits a CD1 restricted response. An immunological composition refers to a composition that includes these components, and can generate or boost an immune response via the CD1 antigen presentation pathway, as described herein. A CD1 restricted response is a response that utilizes the CD1 antigen presentation pathway and can be independent of the MHC antigen presentation pathway.

“Boosting” an immune response refers to enhancing the immune response of an individual or mammal when compared to an immune response prior to administering a composition of the present invention. An increase or enhancement of various immunological parameters occurs when compared to baseline (eg, prior to administering the composition). To “elicit” an immune response is to create an immune response to an antigen that is sufficient to confer a protective or immune enhancing effect on the CD1 antigen. In particular, T cell proliferation occurs and / or antibodies are generated in response to the antigen. Various immunological parameters are measured to determine the immune response (eg, antibody response (eg, level or titer) to the antigen, cytotoxic T lymphocyte response, helper T cell response, T cell proliferative response, or T cell Enhancement or induction of a regulatory cytokine response (eg, a cytokine regulated by a T cell response) can be assessed. See Example 1. One or more of these immunological parameters is increased in response to the CD1 / adjuvant composition when compared to the response before administration of the composition. The basal levels of various immunological parameters can be measured prior to administering the compound, and those levels can then be reevaluated after administration of the composition.
An increase in one or more immunological parameters occurs when compared to baseline. Immunological parameters can also be measured relative to levels from a control, ie, a control population that has already been administered the composition (eg, a positive control) or a population that has not been exposed to the composition (eg, a negative control).
These immunological parameters can be measured using methods known in the art. Accordingly, the present invention includes methods and vaccine compositions for eliciting a protective effect against the CD1 antigen, as well as methods and immunological compositions for boosting or eliciting an immune response against the CD1 antigen.

[0015] In addition to methods and compositions that utilize the CD1 antigen / adjuvant composition, the present invention further includes the step of inducing an immunopotentiating effect on the peptide antigen. Some non-peptide antigens can be used to combat one or more diseases and can be combined with peptide-based vaccines to produce more effective and complete vaccine compositions. The non-peptide antigen and adjuvant may be combined with one or more peptide antigens from the same organism or tumor cells to provide a protective or immunopotentiating effect on multiple antigens, including peptide and non-peptide antigens. Can be provided. For example, tuberculosis vaccines are available from tuberculo
sis coding peptide, M. It may include non-peptide antigens and adjuvants from the tuberculosis cell wall. Such a vaccine is described in M. tuberc
The individual may be protected from tuberculosis or disease caused by ulosis. Thus, the present invention includes vaccines or immunogenic compositions that include non-peptide or lipid antigens and adjuvants as well as peptide-based antigens.

[0016] The administration of the CD1 antigen and the T cell stimulatory compound can be performed simultaneously or continuously without delay. The T cell stimulating compound may be administered before, after or simultaneously with the CD1 antigen. Thus, the term "co-administration" is used herein to mean that the CD1 antigen and the T cell stimulating compound are administered simultaneously to achieve a protective or immunopotentiating effect on the CD1 antigen. The method of the present invention provides for the administration of CD1 as long as the adjuvant is administered in a short period of time sufficient to produce the desired effect (eg, boosting or eliciting a CD1 restricted response to the CD1 antigen).
There is no limitation on the order in which the antigen and the adjuvant are administered. In a preferred embodiment, the administration of the non-peptide antigen / adjuvant composition is simultaneous. The method may also involve co-administration with other drugs (eg, cytokines such as IL-2, GM-CSF and IL-12, and / or antibodies) that help elicit or boost an immunogenic response. Including.

The CD1 antigen is an antigen (eg, an antigen that can undergo the CD1 antigen presentation pathway after its administration)
An antigen presented to the immune system associated with the CD1 molecule). The CD1 antigen also contains C
An antigen capable of eliciting or producing a D-1 restricted response, referred to herein as a "CD1-presenting antigen." A non-peptide antigen refers to an antigen that is not wholly or partially non-peptide (eg, one carboxyl group is bonded to the other amino group to eliminate water molecules and not contain more than one amino acid). The CD1 antigen has a hydrophobic moiety that binds non-specifically to the CD1 molecule, which in turn is located on the hydrophilic component of the antigen that interacts highly specifically with the T cell antigen receptor. The form of the CD1 antigen is a lipid antigen.

[0018] Lipid antigens can be obtained from a variety of biological sources or can be synthesized.
Such antigens can be isolated or synthesized by standard methods known to those of skill in the art, such that the characteristics that non-peptide antigens can present in the context of the CD1 antigen presentation pathway remain. These non-peptide or lipid antigens can be isolated from a variety of biological sources, including bacteria (eg, Gram-negative or Gram-positive), parasites, plants, fungi, or other mammalian sources. . The non-peptidic antigens of the invention may exist in whole form, but may be inactivated bacteria or parasites; and / or portions thereof that retain the antigenicity of the bacteria or parasites. These lipid antigens
Found in the cell walls of bacteria, parasites and / or fungi. For example, Myco
The lipid antigen component of Bacteria can be isolated by standard methods described below: Goren, M .; B. And Brennan, P .; J. , Mycob
actial Lipids: Chemistry and Biolog
ic Activities in Tubeculosis, 1979, T
Uberculosis; P. Youmans ed., Pp. 63-193, (Ph
iladelphia, WB Saunders 1979), and Hun
ter S.T. W. J. et al. Biol. Chem. 261: 12345-1253
1 (1986).

As shown in FIG. 1, the lipid fraction contains bacteria (eg, Mycobacteri).
um tuberculosis). The lipid fraction may be obtained using an alcohol mixture such as acetone or methanol, or chloroform. The organic extract (O / E) is separated from the insoluble crude cell wall fraction by extraction with chloroform or methanol. The O / E is placed on a silicon column and eluted with chloroform, acetone and then methanol to give three purified fractions. The chloroform fraction contains triglycerides and free fatty acids. The acetone fraction contains glycolipids and the methanol fraction contains various phospholipids (
For example, phosphatidylinositol, phosphatidylethanolamine and phosphatidylglycerol). Thus, lipid fractions (eg, chloroform, acetone or methanol fractions) can be used to isolate lipid antigens of the invention.
An “isolated” lipid-antigenic fraction is a fraction that contains a substantially purified amount of lipid molecules or antigen (eg, greater than 80%, 85%, 90%, or 95%). Say.

Examples of the CD1 antigen include lipoglycan, glycolipid, phospholipid, triglyceride, glycosylphosphatidylinositol (GPI), mycolic acid, glucose monomycolate (GMM), lipoarabinomannan (LAM), and lipo-oligosaccharide. Or a lipid antigen such as phosphatidylinositol mannoside (PIM). The lipid antigen may include lipid-based molecules other than those described herein.

Accordingly, the present invention provides protection without causing infection (eg, causing disease) of a living organism that contains a portion of the organism (eg, including CD1 or a lipid antigen) or that occurs naturally. Immunogenic or vaccine compositions, including whole organisms that have been modified to provide a response or an immune enhancing response. Examples of these vaccines are live attenuated vaccines, live killed vaccines, subunit vaccines or toxoid vaccines. Examples of bacteria or parasites containing non-peptide or lipid antigens used in the present invention include Plasmodium (malaria), Mycobacteria (tuberculosis and leprosy), Strepto
genus coccus (streptococcal infection), Staphylococcus genus (staphylococcal infection and pneumonia), Haemophilus (influenza), Ch
Bacteria or parasites, such as species of the genus lamydia (chlamydiasis) or Tyrpanosoma (trypanosomiasis).

[0022] In particular, mycobacteria are a genus of aerobic intracellular bacterial organisms that survive in the endosomal compartment of monocytes and macrophages when invading their hosts. Human mycobacterial diseases include tuberculosis (caused by M. tuberculosis), leprosy (caused by M. leprae), Bairnsda.
le ulcer (caused by M. ulcerans) and M. ulcerans Marim
um, M. kansasii, M .; scrofluaceum, M.P. szulg
ai, M .; xenopi, M .; fortuitum, M.P. chelonei, M
. haemophilum and M.P. Includes various infections caused by intracellulare. Wolinsky, E .; Chapter 37, Microb
ology: Included Immunology and Mole
cultural Genetics, Third Edition, Harper and Row, Phil
adelphia 1980, Daniel. T. M. Miller, R .; A
. And Freedman, S .; D. Chapters 119, 120 and 121, respectively, Harrison's Principles of Inte
rnal Medicine, 11th Edition, Braunwald. E. FIG. Hen, Mc
Graw-Hill, New York, 1987.

With the advent of AIDS, tuberculosis has increased at an approximately logarithmic rate, and multidrug resistant strains have emerged, and now one-third of all cases in New York City. Deserve. Bloom, B .; R. And Murry
, C.I. J. L. Science 257: 1055-1064 (1992);
U. S. Congress, Office of Technology As
sessment, The Continuing Challenge of
Tuberculosis, OTA-H-574. U. S. Governme
nt Printing Office, Washington, D.C. C. , 1
993. Mycobacterial strains previously considered non-pathogenic strains (eg, M.
avium) is currently the main killer of immunosuppressed AID patients.
In addition, currently mycobacterial vaccines are In the case of the BCG vaccine against th. leprae is either unavailable. Kaugmann S.A. Microbiol. Sci. 4: 324-328
(1987); S. Congress, Office of Techno
logic Assessment. The Counting Chal
length of Tuberculosis, pp. 62-67. OTAH-57
4U. S. Government Printing office, Wash
inton, D .; C. 1993. The methods and compositions of the present invention that boost the immune system are important in combating such conditions.

The discovery of structural motifs for antigen presentation by the CD1 protein provides a means by which synthetic antigens can be used to extend the range of antigens presented by the CD1 molecule. The present invention relates to one or more gram-negative or gram-positive bacteria (Streptococ).
cus and / or Staphylococcus species) and vaccine compositions comprising one or more parasites (including malaria) and non-peptide synthetic derived antigens effective against one or more tumor antigens. All Gram-negative bacteria contain lipopolysaccharide (LPS), which is structurally similar to lipomannan. Most Gram-positive bacteria contain structurally related glycolipids (eg, lipoteichoic acid). In addition, protozoan chemical compositions that cause many diseases include glycolipids (eg, Leishm
lipophosphoglycan of C. ania). Orlandi, P .; A. And S.I.
J. Turco, J.M. Biol. Chem. 262: 10384-10391.

The fungal cell wall and other cellular components also contain lipoglycans. Accordingly, the present invention includes a vaccine composition comprising a non-peptide / lipogenic antigen / adjuvant complex that can be used for the prevention or treatment of a fungal infection in a mammal. These antigens can include partial derivatives of LAM, GMM, PIM molecules, or similar glycolipid derivatives from microorganisms, either prokaryotic or eukaryotic in nature.

The present invention uses various parasites or parts thereof as non-peptide antigens. Examples of diseases caused by protozoa include malaria, trichinosis,
Include, but are not limited to, filariasis, trypanosomiasis, schistosomiasis, toxoplasmosis and leishmaniasis. Accordingly, the present invention provides methods and compositions for treating a parasitic or protozoal infection.

The present invention may utilize a synthetic inducible antigen that utilizes the CD1 restriction pathway. According to the present invention, a synthetic antigen is an antigen that is not naturally present in the organism. Synthetic antigens
It may be an antigen that is chemically synthesized, or a portion thereof synthesized by combining elements, molecules or compounds. In addition, a synthetic antigen combines two or more components, one or more of which are antigens isolated or derived from an organism, thus producing a hybrid or chimeric antigen. Can be constructed by Synthetic antigens comprise a lipid component and a chemically synthesized hydrophilic component, an AP that associates with the CD1 molecule through the CD1 antigen presentation pathway.
C can be presented to T cells.

[0028] The CD1 or lipid antigen is combined with a T cell stimulating compound. T cell stimulating compounds are compounds that can increase T cell proliferation or T cell assisting effects when an antigen is introduced into the immune system. T cell stimulating compounds include adjuvants. Adjuvants are substances that enhance the immunogenic response to an antigen. In particular, adjuvants are generally considered to be in one of two categories:
1) a "vehicle" that helps transport and retain antigens in lymphoid tissues, or 2) an "immunomodulator" that stimulates secreted cytokines locally. Adjuvants are
It may consist of bacteria or bacterial products that contribute to the immunogenicity of the antigen. Examples of adjuvants are inorganic salt adjuvants (alum or calcium-based adjuvants)
Incomplete or Complete Freund's Adjuvant, Basille Calmett
e-Guerin (BCG) adjuvant, block polymer adjuvant (T
itermax), cholera toxin (Cholera venom B subunit, enterotoxin (LT) variant), cytokine, adjuvant containing CPG motif, oil / water emulsion adjuvant (oil and water adjuvant; water and oil adjuvant; water, oil and water adjuvant) MF-59 adjuvant, LeIF adjuvant (eg, protein based), liposome adjuvant, ISCOM
Adjuvant, monophosphoryl A adjuvant (MPL), biodegradable microsphere adjuvant, muramyl dipeptide adjuvant, polyphosphodine adjuvant, and saponin adjuvant (QS-7, QS-17, QS-18 or QS-21). Preferably, the composition comprises two or more adjuvants (eg, QS-21 and MPL).

[0029] Other lipid-based vaccines are included in the present invention. Individuals with the disease (eg, cancer and autoimmune disease) can be vaccinated with the lipid / adjuvant composition. Cancers include melanoma, breast, prostate and colorectal cancer. Tumor-derived lipid-based antigens can be used as non-peptide antigens for the present invention. Similarly, autoimmune diseases such as diabetes, lupus and rheumatoid arthritis can be treated with non-peptide / lipid antigens.

The family of CD1 molecules exists to participate in the CD1 antigen presentation pathway. In humans, the CD family is located on a single chromosome and is encoded by five closely linked, non-polymorphic forms of the gene. These show similarities to the intron / exon structure of the MHC class I gene and encode polypeptides with modest, but significant levels of homology to both MHC class I and II proteins. There are five human CD1 proteins, and they are CD1a, CD1b, CD1c, CD1d and CD1e. These CD1 molecules are involved in the CD1 antigen presentation pathway. Porcelli, S.M. Et al.,
Immun. Rev .. , 120: 137-183 (1991).

The present invention can be used in mammals, including humans, cats, dogs, horses, rodents, guinea pigs, and the like. The compositions and methods of the present invention are intended for human as well as veterinary use.

[0032] The compositions of the present invention can be administered with or without a carrier.
The terms "pharmaceutically acceptable carrier" or "carrier" generally refer to any acceptable excipient or drug delivery composition that is relatively inert and non-toxic.
Exemplary carriers include sterile water, saline (eg, Ringer's solution), alcohol, gelatin, talc, viscous paraffin, fatty acid esters, hydroxymethylcellulose, polyvinylpyrrolidone, calcium carbonate, carbohydrates (eg, lactose, sucrose, dextrose, Mannose), albumin, starch,
Examples include cellulose, silica gel, polyethylene glycol (PEG), dried skim milk, rice flour, magnesium stearate and the like. Suitable formulations and additional carriers are available from Remington's Pharmaceuticals
Sciences (17th edition, Mack Pub. Co., Easton, PA)
), The teachings of which are incorporated herein by reference in their entirety. Such preparations may be sterilized and, if desired, adjuvants such as lubricants, preservatives, stabilizers, wetting agents, emulsifiers, which do not deleteriously react with the active compound,
(Eg, salts, buffers, coloring substances, and / or fragrances that affect osmotic pressure). They can also be combined, if desired, with other active agents to reduce metabolic degradation, such as enzyme inhibitors. Carrier (for example,
A pharmaceutically acceptable carrier) is preferred but not required to administer the compound.

[0033] The present invention provides various pharmaceutical compositions. Such compositions include a therapeutically (or prophylactically) effective amount of an immunogenic or vaccine composition (including CD1 antigen, adjuvant, peptide antigen and / or carrier). If desired, the composition may also contain minor amounts of wetting or emulsifying agents, or pH buffering agents, or preservatives. Representative preservatives may include potassium sorbate, sodium metabisulfite, methyl paraben, propyl paraben, thimerosal, and the like.

[0034] The composition can be a liquid solution, suspension, emulsion, tablet, pill, capsule, sustained release formulation, or powder. The method of administration can dictate how to formulate the composition. For example, the composition can be formulated as a suppository, with classical binders and carriers such as triglycerides. Oral formulations can include standard carriers such as pharmaceutical grades of mannitol, lactose, starch, magnesium stearate, sodium saccharine, cellulose, magnesium carbonate, and the like.

The carrier may be added to the composition at any convenient time. For example, in the case of a lyophilized vaccine, the carrier can be added just prior to administration. Alternatively, the end product can be manufactured with a carrier.

The immunogenic vaccine compositions of the present invention are administered intravenously, parenterally, intramuscularly, subcutaneously, orally, nasally, topically, by inhalation, by implantation, by injection, or by suppository. obtain. The composition may be administered in a single dose or in more than one dose (eg, boost) over a period of time to provide the desired effect.

For enteral or mucosal application, including via the oral and nasal mucosa, tablets, liquids, drops, suppositories or capsules are particularly suitable. syrup,
Elixir and the like may be used, where a sweetened vehicle is used. Topical application can also be used, for example, in intraocular administration. An alternative method of administration is described in US Pat. No. 4,900,549 (or European Patent Application No. 0 604 727 A).
1 (published June 6, 1994)). In addition to liposomes, viral vectors, microspheres, and microcapsules are available and can be used. Rabinovich
See above.

For pulmonary diseases such as tuberculosis, pulmonary administration, such as an inhaler, may be suitable for prophylactic purposes, or for immediate treatment and certain topical treatments. Pulmonary administration, for example,
This can be accomplished using any of a variety of delivery devices known in the art. For example,
S. P. Newman (1984), Aerosols and the Lu
ng, Clarke and Davia, Butterworths, London
on, England, pp. 197-224; PCT Publication No. WO 92/1619.
2; see PCT Publication No. WO 91/08760; NTIS Patent.

For parenteral administration, sterile injectable solutions are particularly suitable, preferably oily or aqueous solutions, and suspensions, emulsions or implants, including suppositories. In particular, carriers for parenteral administration include dextrose aqueous solution, physiological saline, pure water, ethanol, glycerol, propylene glycol, peanut oil, sesame oil, polyoxyethylene-polyoxypropylene block polymer and the like. An ampoule is a convenient unit dosage.

The mode of administration will vary according to the type of disorder, and will require that the microorganism be controlled or extinct. The vaccine dose will depend on the amount of the antigen, its level of antigenicity, and the route of administration. One skilled in the art can easily and immediately titrate the dose for the immunogenic response of each antigen-adjuvant complex and method of administration.

The actual effective amount of the compound or drug may vary, for example, according to the particular composition employed, the mode of administration, and the age, weight and condition of the patient. As used herein, an effective amount of a drug is an amount of the drug that elicits or boosts an immune response to the non-peptide antigen. The dosage for a particular patient can be determined by one of ordinary skill in the art using conventional considerations (eg, by appropriate conventional pharmacological protocols).

The present invention also provides at least one T cell stimulating compound (eg, an adjuvant)
And a kit comprising at least one non-peptide antigen (eg, a lipid antigen), and the non-peptide antigen elicits a CD1 restricted response. As described herein, the antigen can be obtained or isolated from at least one bacterium and / or parasite, or any other organism that contains a lipid-based antigen. The antigen can also be a tumor-associated antigen (eg, cancer), or a self-antigen (eg, an autoimmune disease). The kit may include various adjuvants as described herein. Preferably, the adjuvant is QS-7, QS
-17, QS-18 or QS-21. Further, the kit may include a peptide antigen or vaccine as described herein.

The following examples are illustrative and are not intended to be limiting in any way.

EXAMPLES Example 1 Measurement of Immunogenic Parameters Including Antibody Response, Cytokine Response and T Cell Response Methods for evaluating antibody response or cytokine response to non-peptide antigens are described in the art. Includes several suitable assays that are known. Samples from an individual given an adjuvant / non-peptide antigen composition can be analyzed. T
The level of cytokines resulting from cell growth and the level of antibodies specific for non-peptide antigens produced can be measured. An increase in the level of cytokines and / or antibodies specific for non-peptide antigens is indicated by the adjuvant /
Occurs after administration of a non-peptide antigen composition. The sample can be any biological component that includes a cytokine or antibody to the antigen (eg, a blood sample, urine sample, sputum sample, etc.). Suitable assays include immunological methods, radioimmunoassays, flow cytometry assays, enzyme-linked immunosorbent assays (ELISAs) and chemiluminescent assays.

In determining the amount of an antibody specific for an antigen to which an individual is to be immunized, the assay will evaluate a sample to be tested under conditions appropriate for the formation of a complex between the antibody and the antigen. Combining or contacting with a non-peptidic antigen (eg, a bacterial and / or parasite antigen or a portion thereof). The level of complex formation is then detected or measured (directly or indirectly).

Immunological assays can be used to detect cytokine levels for cytokines affected by T cell proliferation. Instead of contacting the sample with a non-peptide antigen, under appropriate conditions for the formation of a complex between the cytokine and the anti-cytokine antibody, as is done to assess the level of the antibody in the sample, Can be combined with an antibody specific for the cytokine. The level of the cytokine / anti-cytokine complex can then be detected.

The sample can be obtained directly or indirectly (eg, provided by a health care provider) and can be prepared by methods appropriate to the particular sample and assay format selected. For example, a suitable method for collecting whole blood is venipuncture. That is, the method obtains blood from an indwelling arterial line. The container where the healthcare provider stores blood is ACD-A, heparin, or EDTA.
And anticoagulants such as

An appropriate label can be detected directly (eg, a radioactive, fluorescent, or chemiluminescent label). These can also be detected indirectly using labels such as enzyme labels and other antigenic or specific binding partners such as biotin. Examples of such labels include fluorescent labels such as fluorescein, rhodamine, CY5, chemiluminescent labels such as luciferase, radioisotope labels such as ³² P, ¹²⁵ I, ¹³¹ I, and horseradish peroxidase. Enzyme labeling, and alkaline phosphatase, β-galactosidase, biotin, avidin, spin labeling and the like. Detection of the antibody in the complex can also be done immunologically using a secondary antibody, which is then detected (eg, using a label). Conventional or other suitable methods may label the antibody directly or indirectly. Antibody levels or cytokine levels can be assessed using methods known in the art or later developed.

The titer is determined by titration of the antiserum by serial dilution to determine the point at which binding decreases to 50% of maximum. This titer is expressed as the reciprocal of the serum dilution defining the endpoint.

[0050] T cell responses can also be measured using methods known in the art. Sample from an individual (eg, blood, bone marrow, lymphoid organ)
Epithelium and tissues or cells) can be obtained from the site of inflammation. The function of T cells can be determined by various methods (eg, target cell killing).
) (CLT assay), macrophage activation, T cell proliferation, B cell activation,
Or lymphokine production).

T cells are detected by their effect on the secretion of target cell presenting antigens, or of certain cytokines acting on such target cells. Measuring these effector functions underlies T cell bioassays used to assess both T cell specificity for antigen function and T cell effector function.

Activated T cells (eg, CD8 T cells) generally kill any cell that presents a particular peptide or lipid. Thus, the function of CD8 T cells can be determined using the simplest and fastest T cell bioassay (killing of target cells by cytotoxic T cells). This may be detected in a ⁵¹ Cr release assay.
Living cells take up, but do not spontaneously release, radioactively labeled sodium chromate (Na ₂ ⁵¹ CrO ₄ ). When these labeled cells are killed, radioactive chromate is released, and the presence of radioactive chromate in the supernatant of the mixture of target cells and cytotoxic T cells can be measured.

In a similar assay, proliferating lymphocytes can be labeled with ³ H-thymidine, which is incorporated into replicating DNA. The cells are then harvested and the incorporated CPM is determined. These assays provide a quick, sensitive, and specific measure of T cell activity.

The function of CD4 T cells is involved in activation, rather than killing, of cells with specific antigens. The activating effect of CD4 T cells on B cells or macrophages is mediated primarily by non-specific mediator proteins called cytokines.
This cytokine is released by T cells when recognizing the antigen. Therefore, the function of CD4 T cells is usually studied by measuring the type and amount of these released proteins. Since different effector T cells release different amounts and types of cytokines, by measuring the proteins they produce, one can know about the potential effector of that T cell. Where cytokines serve as either growth factors or growth inhibitors, the cytokines may be detected by their activity in biological assays of cell proliferation.
That is, more specifically, cytokines may be detected by a modified version of the ELISA, as described herein, known as a capture or sandwich ELISA. In this assay, cytokines are characterized by the ability of the cytokine to crosslink between two monoclonal antibodies that react with different epitopes on the cytokine molecule. Sandwich ELISA can also be performed by placing the cells themselves on a surface coated with antibodies to the cytokine. After a short incubation, the cytokine released by each cell is captured on the antibody coat and the presence of cytokine-secreting cells may become apparent when the cells are washed away and a labeled secondary anti-cytokine antibody is added. The cytokine released by each cell creates a distinct spot in this assay. Therefore, this is known as the ELISPOT assay.
ELISPOT can also be used to detect specific antibody secretion by B cells.
In this case, an antigen-coated surface for capturing the specific antibody and a labeled anti-immunoglobulin for detecting the bound antibody are used. Sandwich ELIS
A avoids the main problem of cytokine bioassays (the ability of different cytokines to stimulate a similar response in the bioassay). Bioassay,
It must always be confirmed by inhibition of the response using monoclonal antibodies to the cytokine.

An alternative method is to identify cytokine mRNA either in a cell population by reverse transcriptase-polymerase chain reaction (RT-PCR) or by single-cell in situ hybridization. Reverse transcriptase
It is an enzyme used by RNA viruses. This RNA virus is similar to the human immunodeficiency virus that causes acquired immunodeficiency syndrome (AIDS),
Convert A genome to DNA copy or cDNA. By collecting mRNA from T cells stimulated with the antigen, a cDNA copy can be made which is then
Selectively amplify by polymerase chain reaction using cytokine specific primers. The amount of product is proportional to the representation of the amount of product in the RNA in the responding cells. In situ hybridization is performed using labeled antisense RN.
The A probe is used to hybridize to sense RNA in single cells isolated either from culture or directly in tissue sections. This allows to determine the number of cells that make RNA encoding a particular cytokine. Janeway, Charles A. , And Travers, Paul,
"T-cell effectors functions can be me
assured in four ways-target-cell kill
ing, macrophage activation, B-cell act
Ivation, or lymphokine production ", In
Immuno Biology, The Immuno System in
Health and Diesease, (New York and L
ondon: Garland Publishing Inc. ), 2: 31-
2:33, these teachings are incorporated herein by reference in their entirety.

Example 2: Methods for preparing lipid fractions, methods for immunizing mammals, and other procedures for evaluating immunological parameters Lipids disclosed herein Fractions (acetone Fx, methanol Fx, and chloroform Fx) were collected by standard extraction methods for Mycobacterium t
Uberculosis cells were prepared from cultures. These extracts are illustrated in FIG. After extraction with chloroform and methanol, the organic extract (O /
E) was separated from the insoluble, delipidated cell wall fraction. The O / E was placed on a silica column and eluted with chloroform, acetone and then methanol to give three purified fractions. The chloroform fraction contained triglycerides and free fatty acids and represented 34% by weight of the total extract. Various glycolipids eluted with the acetone fraction, which contained 12% of the extract. The methanol fraction contained 29% of the extract and contained various phospholipids such as phosphatidylinositol, phosphatidylethanolamine and phosphatidylglycerol. The delipidated cell wall was saponified and precipitated to obtain isolated mycolic acid corresponding to 24% of the total cell extract.

All fractions were tested for the presence of protein by gel electrophoresis and standard protein assays (eg, Bradford assay) and found to have no detectable protein. By acid hydrolysis and quantitative amino acid analysis by HPLC, it was determined that the four fractions were relatively free amino acids and all had less than 5% amino acid content. In fact, the acetone and chloroform fractions each had less than 1% amino acid content.

[0058] Glucose monomycolate (GMM) is converted to a rapidly growing Mycobactate.
Purified from Rium phlei culture to homogeneity. M. GMM was obtained by hydrolyzing phlei cells, drying on glass and treating with 2M TFA at 121 ° C. for 2 hours (GS Besra et al. (1994) Proc.
. Nat. Acad. Sci. , USA 91: 12737). The resulting glycolipid product was characterized by TLC as compared to a standard GMM standard.
ESI-MS analysis showed the expected m / z ion for GMM.

For immunization of Hartley strain guinea pigs, total heat-killed M. tubercu
Fractions eluted with losis cells or a mixture of acetone, chloroform, and methanol were mixed with various adjuvants and, in some samples, incorporated into liposomes. Ovalbumin (OVA) was also added to the mixture. Ovalbumin has a MHC class II restricted response (restricte
d response, which stimulated the immune response of CD4 ⁺ T cells and was a positive control showing T cell activity against protein antigens.

Some adjuvants were mixed with the lipid mixture to enhance the immunogenicity of the antigen mixture. Incomplete Freund's adjuvant (IFA) is an oil-in-water emulsion. Titermax ^™ is a combination of squalene and a particulate stabilizer (micropar)
ticulate stabilizer) (Vaxcel, Inc., Nor)
cross, Georgia, USA) and a block copolymer (CR)
L89-41 or CRL-8300). QS-21 (Aquila B
iopharmaceuticals, Inc. Supra) and bone marrow derived dendritic cells (BMDC) were also used as adjuvants. Monophosphoryl lipid A (M
PL) from RIBI ImmunoChem Research, Inc. , H
amilton, MT 59840-3131.

Liposomes were made from phospholipids known to be non-irritating to the mammalian immune system. Ovalbumin and MPL contained the minor components of the liposomes used to deliver the lipid mixture.

[0062] Guinea pigs were inoculated with one of the immunogenic complexes described above. This administration was parenteral and subcutaneous. A second dose was given 2-4 weeks later. Cell samples
Obtained for ex vivo measurement of T cell responses 3 weeks after the last immunization. The results are shown in the table below.

Example 3 Results Demonstrating that Lipid Antigen and Adjuvant Compositions Increase Immune Response in Mammals Glycolipids mixed with QS-21, MPL or bone marrow derived dendritic cells (BMDC) , Produced the greatest response from the T cells. QS-21 has been shown to boost the immune response when combined with an antigen specific for a wide variety of diseases. The improved immunogenicity with adjuvants such as QS-21 compared to using liposomes alone is significant.

[0064] Lymphocytes isolated from guinea pigs immunized with liposomes containing QS-21 and MPL or immunized with purified antigen responded to lipid antigens in vitro. Peripheral blood mononuclear cells responded to all antigens compared to exposure to cell culture medium alone. Medium lacking the antigen was used as a negative control. See FIG. 2A. The strongest response was to OVA, GMM, and the combined organic extracts of all hydrophobic fractions. Peripheral blood mononuclear cells were prepared by passing blood over a gradient to extract erythrocytes and granular monoocytes. The eluate was T cells, B cells,
And consisted of mononuclear cells. However, this procedure results in an enriched T cell population with less than 10% of immune responder cells, including B cells. Thus, the response shown in FIG. 2A is primarily a T cell response.

Spleen lymphocytes were purified by passing cells over a gradient and then through nylon wool by standard means known to those skilled in the art to enrich the T cell population. These procedures resulted in a T cell to B cell ratio of greater than 10: 1. Irradiated spleen cells are added at a 1: 1 ratio to contain CD1 ⁺ antigen presenting cells. These spleen fractions showed growth in the presence of OVA, GMM, organic extract, acetone fraction and mycolic acid fraction. See FIG. 2B.

When the cell suspension was treated with an anti-CD4 antibody to remove the CD4 ⁺ T cell fraction, the response to OVA was reduced as shown in FIGS. 3A-3B and FIG. The T cell fraction in these suspensions that responds to the presence of antigen is CD4 ^- C
D8 ^- consisting of T-cells and CD8 ⁺ T cells. A comparison of these responses showing significant responses to GMM, acetone fraction, and mycolic acid is shown in FIG. This antigen-specific response was dose-dependent and comparable to the highest values.

This data indicates the type of immunogenic complex that can stimulate an immune response that protects animals from disease. Any CD1 presenting cell can be used for the immunogenic conjugate of the present invention. CD
Examples of structural requirements for 1-presenting antigens are described in Moody, D .; , B. , Et al., Science
, 278: 283-286 (1997). Synthetic CD-1 presenting antigens may also be found therein. For the adjuvant used, the best response was observed by QS-21 and MPL or BMDC. See Table 1.

[0068]

[Table 1] (Example 4: Results of FIGS. 5-7) FIGS. 5A-C show strain 2 guinea pigs immunized with purified mycobacterial lipid antigens reconstituted as immunogenic compositions by incorporation into liposomes using adjuvants. 1 shows a major ex vivo cytotoxic response of isolated T lymphocytes. M. was inserted into liposomes composed of inert phospholipids and cholesterol and mixed twice with the adjuvants QS-21 and MPL in the form of liposomes. Strain 2 guinea pigs were immunized with a purified lipid-antigen mixture containing tuberculosis glycolipids (ie, the lipid fraction eluted from a silica column using acetone as a solvent; see FIG. 1 for details). After immunization, the guinea pigs were sacrificed and spleen T cells were obtained. Spleen T cells were
. Tuberculosis was incubated with the glycolipid fraction for 6 days in the presence of irradiated spleen adherent cells (ie, the source of CD1 + antigen presenting cells) and 0.87 nM human recombinant interleukin-2. Live cells were isolated and used as effector cells in a standard chromium release assay for cytotoxic T cell activity. These T cells are transformed into two different guinea pig CD1 molecules (gpCD1b1 or gpC1) by transfection of a cDNA encoding these molecules into a guinea pig cell line (104C1) that normally lacks expression of the CD1 molecule.
Target cells engineered to express any of D1c2) were lysed. Target cells were initially treated at a concentration of 50 μg / ml (ie, designated as “acetone 50” in FIG. Lysis of target cells was observed when incubated with the tuberculosis glycolipid antigen, and when the target cells were incubated with the glycolipid antigen at a concentration of 25 μg / ml (denoted “Acetone 25” in FIG. 5),
Lysing of the target cells to a lesser extent was observed. When this target was cultured in a medium containing no mycobacterial lipid antigen (represented as "medium" in FIG. 5), no cytotoxic activity of T cells against any target cells was observed. This demonstrated that the cytotoxic T cells present in the culture were specific for lipid antigens and antigen recognition was dose dependent. Furthermore, if the target cells do not express the CD1 molecule (this is the case for 104C1 mimicking the transfected target), no target cell lysis was observed (FIG. 5C). This demonstrated that lipid antigen-specific T cells present in the culture were restricted by either CD1b1 or CD1c2 molecules. Therefore, these data are
Both D1b1 and CD1c2 molecules, in this pathway, produce specific cytotoxic T from animals immunized with the appropriate immunogenic lipid antigen / adjuvant composition.
It is utilized and effective for producing cells.

FIG. 6 shows CD1 evoked from animals immunized with an antigen displaying CD-1.
-Restricted M. T. tuberculosis lipid antigen-specific T cell line (S2-CD8
. 1 shows the proliferative response of 1). Line 2 guinea pigs were immunized twice by the subcutaneous route with an immunogenic lipid antigen / adjuvant formulation. The proliferative response of the T cell line (count / min (CPM) of tritiated thymidine incorporation uptake) was measured in the presence of: T cell line alone; bone marrow-derived dendritic cells (BM-DC; CD1 + antigen presentation) T cell lines cultured with T. cell lines; monoclonal antibodies specific for all known forms of T cell lines and BM-DC and guinea pig CD1 (anti-C
D1 mAb); T cell lines and BM-DC; tuberculosi
s lipid antigen (total lipid chloroform / methanol extract of M. tuberculosis, expressed as "M.Tbc O / E"; see FIG. 1 for details); and T cell lines and BM-DC, and M. tuberculo
sis lipid antigen and anti-CD1 mAb. This result indicates that T cells proliferated strongly when exposed to lipid antigens in the presence of CD1 + antigen presenting cells (BM-DC),
This indicates that the cells did not proliferate in the absence of CD1 + antigen-presenting cells. This growth is
Dependent on the presence of lipid antigen and was strongly inhibited by anti-CD1 mAb. These data indicate that T cell lines that are specific for lipid antigens and are restricted by the CD1 molecule can be derived from guinea pigs immunized with an appropriate immunogenic lipid antigen / adjuvant composition.

FIG. T. tuberculosis lipid antigen-specific T cell line (S2-CD
The CD1 restriction element of 8.1) is exemplified. The S2-CD8.1 T cell line contains 1
04C1 / CD1b1, 104C1 / CD1b2 and 104C1 / CD1c3
M. with transfectants The tuberculosis lipid antigen was lysed but not the mock transfectants expressing CD1b3, CD1b5 or CD1c2. Inserted into liposomes composed of inert phospholipids and cholesterol and mixed with adjuvants QS-21 and MPL.
from a guinea pig of line 2 immunized with an immunogenic lipid antigen / adjuvant composition consisting of a tuberculosis total lipid extract.
1 was induced. After immunization, guinea pigs were sacrificed and T cells were isolated from spleen. These T cells were converted to CD1 + antigen presenting cells (BM-DC) and 20 μg / m
l. The cells were stimulated in vitro four times with cultures containing tuberculosis lipid extract. The resulting T cell line is expanded by the addition of human recombinant interleukin 2, and the cells are harvested and used in a standard chromium release assay to determine the presence of lipid antigen-specific cytotoxic T cells and The restriction of T cells by individual members of the CD1 family was evaluated. This result indicates that the CD1b1 molecule, C
Significant lipid antigen-dependent lysis of 104C1 target cells transfected for expression with either D1b2 or CD1c3 molecules was demonstrated. This recognition is dose dependent, indicating that target cells generally respond more strongly when incubated with 20 μg / ml lipid antigen compared to 10 μg / m lipid antigen. In contrast, C in the presence or absence of lipid antigen
Significant lysis of mock transfected 104C1 cells that do not express the D1 molecule
Not observed. In addition, three other forms of guinea pig CD1 (CD1b3,
CD1b5, and CD1c2) are least active in providing 104C1 cells with the ability to present lipid antigens to the T cell line S2-CD8.1,
Or provided that it is inert. Thus, these data indicate that the CD1 antigen presentation pathway is effective in generating an immune response for mammals immunized with an appropriate immunogenic lipid antigen / adjuvant composition.

The relevant teachings of all references, patents and / or patent applications cited herein are hereby incorporated by reference in their entirety.

Although the present invention has been shown and described in detail with reference to preferred embodiments thereof, various changes in form and detail may occur in accordance with the present invention as defined in the appended claims. It will be understood by those skilled in the art that changes may be made therein without departing from the spirit and scope of the invention.

[Brief description of the drawings]

FIG. 1 illustrates the preparation of mycobacterial lipid antigen from Mycobacterium tuberculosis.

FIG. 2A shows CD1-presenting antigens obtained from blood: ovalbumin (OVA), My.
cobacterium phlei glucose monomycolate (GMM), organic extract (O / E), acetone fraction (acetone Fx), chloroform fraction (C
9 is a bar graph showing the response of lymphocytes to HC13 Fx) and mycolic acid fraction (M. acid) in number of times per minute (CPM).

FIG. 2B shows CD1-presenting antigens: ovalbumin (OVA), M obtained from spleen.
ylobacterium phlei glucose monomycolate (GMM),
Organic extract (O / E), acetone fraction (acetone Fx), chloroform fraction (
CHC13 Fx), lymphocyte response to mycolic acid fraction (M. acid)
It is a bar graph shown by M.

FIG. 3A shows in vitro lymphocyte response to purified lipid antigen (acetone Fx (μg / mL)) in animals (guinea pigs) immunized in combination with monophosphoryl lipid A (MPL) adjuvant and QS-21 adjuvant. To CPM
FIG.

FIG. 3B shows Mycoba in animals (guinea pigs) immunized in combination with monophosphoryl lipid A (MPL) adjuvant and QS-21 adjuvant.
cterium phlei glucose monomycolate (GMM) (μg / mL
5 is a graph illustrating in vitro lymphocyte response to CPM.

FIG. 3C shows in vitro lymphocyte response to ovalbumin (OVA) (μg / mL) in animals (guinea pigs) immunized in combination with monophosphoryl lipid A (MPL) adjuvant and QS-21 adjuvant with CPM. It is a graph shown.

FIG. 4A shows the following CD1-presenting antigen in combination with MPL and QS-21 adjuvants: GMM, acetone fraction, CHC13 Fx, M.P. FIG. 4 is a graph showing the maximal proliferative response of CD4 depleted cells from animals immunized with acid and OVA by CPM.

FIG. 4B shows the following CD1-presenting antigens in combination with MPL and QS-21 adjuvants: GMM, acetone fraction, CHC13 Fx, M.P. Figure 4 is a bar graph showing the maximal proliferative response from animals immunized with acid and OVA by CPM.

FIG. 5A shows MPL adjuvant and QS against CD1b1 antigen presentation pathway.
50 mg / ml (acetone 50) or 2 in combination with -21 adjuvant
E from animals immunized with either acetone fraction at 5 mg / ml (acetone 25)
Major ex vivo cytotoxic T lymphocyte response to T ratio (specific lysis (%))
FIG.

FIG. 5B shows MPL adjuvant and QS against CD1c2 antigen presentation pathway.
50 mg / ml (acetone 50) or 2 in combination with -21 adjuvant
E from animals immunized with either acetone fraction at 5 mg / ml (acetone 25)
Major ex vivo cytotoxic T lymphocyte response to T ratio (specific lysis (%))
FIG.

FIG. 5C shows animals immunized with either 50 mg / ml (acetone 50) or 25 mg / ml (acetone 25) acetone fractions combined with MPL adjuvant and QS-21 adjuvant in mock or control systems. Major ex vivo cytotoxic T lymphocyte response to ET ratio from (specific lysis (%
FIG.

FIG. 6 shows CD1-restricted My induced from animals immunized with these lipid antigens.
C. bacterium turbuculosis lipid antigen-specific T cell line (
FIG. 2 is a bar graph showing the proliferative response of S2-CD8.1) in CPM.

FIG. 7 shows the concentration of Mycobacterium at 1, 10, and 20 μg / ml.
urbuculosis lipid antigen organic extract specific T cell line (S2-CD8.
It is a bar graph which shows the cytotoxic T lymphocyte response (specific lysis (%)) of 1).
This cell line is 104C pulsed with turbuculosis lipid antigen
1-CD1b1, CD1b2 and CD1c3 transfectants were lysed.

──────────────────────────────────────────────────続 き Continued on the front page (51) Int.Cl. ⁷ Identification symbol FI Theme coat ゛ (Reference) A61K 39/09 A61K 39/09 39/102 39/102 39/118 39/118 39/39 39/39 A61P 31/04 A61P 31/04 31/06 31/06 31/08 31/08 31/16 31/16 33/02 33/02 33/06 33/06 37/04 37/04 (81) Designated country EP ( AT, BE, CH, CY, DE, DK, ES, FI, FR, GB, GR, IE, IT, LU, MC, NL, PT, SE), OA (BF, BJ, CF, CG, CI, CM, GA, GN, GW, ML, MR, NE, SN, TD, TG), AP (GH, GM, KE, LS, MW, SD, SL, SZ, UG, ZW), EA (AM, AZ, BY, KG, KZ, MD, RU, TJ, TM), AE, AL, AM, AT AU, AZ, BA, BB, BG, BR, BY, CA, CH, CN, CU, CZ, DE, DK, EE, ES, FI, GB, GD, GE, GH, GM, HR, HU, ID , IL, IN, IS, JP, KE, KG, KP, KR, KZ, LC, LK, LR, LS, LT, LU, LV, MD, MG, MK, MN, MW, MX, NO, NZ, PL, PT, RO, RU, SD, SE, SG, SI, SK, SL, TJ, TM, TR, TT, UA, UG, US, UZ, VN, YU, ZA, ZW (72) Inventor Brenner , Michael B. United States Massachusetts 02159, Newton, The Ledges Road 27 (72) Inventor Dasher, Christopher Sea. United States Massachusetts 02114, Boston, Military Street 81 (72) Inventor Kenji Hiromatsu Massachusetts 02146, Brooklyn, Number 47, Kent Street 197 F term (reference) 4C085 AA03 BA03 BA06 BA09 BA13 BA14 BA18 BA26 BB21 CC07 DD23 FF03 FF01 FF20FF GG02 GG03 GG04 GG08

Claims (45)

[Claims] 1. A method for enhancing an immune response to at least one CD1 antigen in a mammal, comprising co-administering an effective amount of at least one CD1 antigen and at least one saponin adjuvant to the mammal. 2. The saponin adjuvant comprises the following: QS-7, QS-17
, QS-18, and QS-21. 3. The method of claim 1, wherein said CD1 antigen is a lipid antigen. 4. The method according to claim 3, wherein at least one immunological parameter is elicited. 5. The immunological parameter is selected from the group consisting of: an antibody response to an antigen, a cytotoxic T lymphocyte response, a helper T cell response, a proliferative T cell response, and a T cell regulatory cytokine response. The method of claim 4, wherein 6. The mammal has an immune-damage disease, disorder or condition.
The method of claim 5. 7. The method of claim 1 wherein said composition is used to produce at least one bacterial infection and / or
Or eliciting or boosting an immune response to at least one parasitic infection. 8. Use of the composition to treat malaria, leprosy, tuberculosis, streptococcal infection, staphylococci, pneumonia, influenza, chlamydiasis, or trypanosomiasis.
8. The method of claim 7, wherein the method elicits or boosts an immune response to ossis). 9. A method of vaccinating a mammal against at least one CD1 antigen, comprising administering to said mammal an effective amount of at least one saponin adjuvant. 10. The method of claim 10, wherein at least one immunological parameter is elicited.
The method according to claim 9. 11. The immunological parameter is selected from the group consisting of: an antibody response to an antigen, a cytotoxic T lymphocyte response, a helper T cell response, a proliferative T cell response, and a T cell regulatory cytokine response. The method of claim 10, wherein 12. The CD1 antigen comprises the following: glycolipid, phospholipid, triglyceride, glycosylphosphatidylinositol, mycolic acid, glucose monomycolate, lipoarabinomannan, lipooligosaccharide, phosphatidylinositol mannoside, autoantigen and tumor 2. The method according to claim 1, wherein the selected antigen is selected from the group consisting of antigens of interest.
2. The method according to 1. 13. The method according to claim 12, wherein said response is elicited against at least one bacterial infection and / or at least one parasite infection. 14. Use of the composition to treat malaria, leprosy, tuberculosis, streptococcal infection, staphylococci, pneumonia, influenza, chlamydiasis, or trypanosomosis.
14. The method according to claim 13, which elicits an immune response against iosis). 15. The saponin adjuvant comprises the following: QS-7, QS-1
10. The method of claim 9, wherein the method is selected from the group consisting of 7, QS-18, and QS-21. 16. The method of claim 9, further comprising administering at least one other antigen, including a peptide antigen. 17. A method of stimulating a CD1-restricted immune response in a mammal, comprising administering to said mammal an effective amount of a composition comprising at least a saponin adjuvant and at least one lipid antigen. The method wherein the lipid antigen elicits a CD1 restricted response. 18. The lipid antigen is from the group consisting of: glycolipids, phospholipids, triglycerides, glycosylphosphatidylinositol, mycolic acid, glucose monomycolate, lipoarabinomannan, lipooligosaccharides, and phosphatidylinositol mannoside. 18. The method according to claim 17, which is selected. 19. The method according to claim 18, wherein said response is elicited against at least one bacterial infection and / or at least one parasite infection. 20. Use of the composition to treat malaria, leprosy, tuberculosis, streptococcal infection, staphylococci, pneumonia, influenza, chlamydiasis, or trypanosomosis.
20. The method according to claim 19, wherein said method elicits an immune response against iosis). 21. The saponin adjuvant comprises: QS-7, QS-1
18. The method of claim 17, wherein the method is selected from the group consisting of 7, QS-18, and QS-21. 22. The following: a. At least one saponin adjuvant; and b. An immunogenic composition comprising at least one CD1 antigen that elicits a CD1 restricted response. 23. The composition according to claim 22, wherein said CD1 antigen is a lipid antigen. 24. The composition according to claim 23, wherein the antigen is derived from at least one bacterium and / or at least one parasite. 25. The bacterium of the genus Mycobacteria, Hae
genus mophilus, Streptococcus, Staphylococ
25. The composition of claim 24, wherein the composition is from a species selected from the group consisting of the genus cus and the genus Chlamydia. 26. The parasite of the genus Plasmodium or Tryp
26. The composition of claim 25, wherein the composition is from a species of the genus anosoma. 27. The saponin adjuvant comprises: QS-7, QS-1
23. The composition of claim 22, wherein the composition is selected from the group consisting of 7, QS-18, and QS-21. 28. The method according to claim 28, wherein the antigen is: glycolipid, phospholipid, triglyceride,
The composition according to claim 23, wherein the composition is selected from the group consisting of glycosylphosphatidylinositol, mycolic acid, glucose monomycolate, lipoarabinomannan, lipooligosaccharide, phosphatidylinositol mannoside, autoantigen and tumor-derived antigen. object. 29. The antigen comprises: CD1a, CD1b, CD1c, CD
23. The composition of claim 22, which elicits a CD1 restricted response using a CD1 molecule selected from the group consisting of 1d and CD1e. 30. The composition of claim 22, further comprising a carrier. 31. A method for raising an immune response in a mammal, comprising the step of administering the composition of claim 22. 32. The following: a. At least one saponin adjuvant; and b. A vaccine composition comprising at least one lipid antigen that elicits a CD1 restricted response. 33. The vaccine composition according to claim 3, wherein said vaccine composition elicits an immune response against at least one bacterial infection and / or at least one parasitic infection.
3. The vaccine composition according to 2. 34. The vaccine composition according to claim 17, wherein said vaccine composition comprises malaria, leprosy, tuberculosis, streptococcal infection, staphylococci, pneumonia, influenza, chlamydiasis, or trypanosomosis.
34. The vaccine composition according to claim 33, which elicits an immune response against iosis). 35. The saponin adjuvant comprises: QS-7, QS-1
33. The vaccine of claim 32, wherein the vaccine is selected from the group consisting of 7, QS-18, and QS-21. 36. The lipid antigen is from the group consisting of: glycolipids, phospholipids, triglycerides, glycosylphosphatidylinositol, mycolic acid, glucose monomycolate, lipoarabinomannan, lipooligosaccharides, and phosphatidylinositol mannoside. 33. The vaccine composition of claim 32, which is selected. 37. The antigen may be: CD1a, CD1b, CD1c, CD
33. The vaccine composition of claim 32, wherein the vaccine composition elicits a CD1 restricted response using a CD1 molecule selected from the group consisting of 1d and CD1e. 38. The vaccine composition according to claim 32, further comprising a carrier. 39. The vaccine composition of claim 33, further comprising a peptide antigen, wherein said peptide antigen elicits a non-CD1 restricted response. 40. A method of vaccinating a mammal, comprising the step of administering the vaccine composition of claim 32. 41. A kit comprising: a) at least one saponin adjuvant, and b) at least one CD1 antigen that elicits a CD1 restricted response. 42. The kit of claim 41, wherein said CD1 antigen is a lipid antigen. 43. The kit of claim 42, wherein said antigen is from at least one bacterium and / or at least one parasite. 44. The saponin adjuvant comprises: QS-7, QS-1
42. The kit of claim 41, wherein the kit is selected from the group consisting of 7, QS-18, and QS-21. 45. The kit of claim 41, further comprising a carrier.