WO1998014207A1

WO1998014207A1 - Immunological therapy for cancer

Info

Publication number: WO1998014207A1
Application number: PCT/US1997/017260
Authority: WO
Inventors: Baruj Benacerraf
Original assignee: Dana-Farber Cancer Institute
Priority date: 1996-10-01
Filing date: 1997-09-26
Publication date: 1998-04-09
Also published as: AU4502797A

Abstract

The invention relates to a method for treating a solid tumor in somatic tissue, or treating or preventing metastasis of such a tumor, in a patient comprising administering to the patient an amount of one or more antigens specific to the somatic tissue effective to induce an immune response against the somatic tissue. In distinction to the above-described immunotherapies, the antigen or antigens administered to the patient are not specific to the tumor, but are specific to the somatic tissue from which the tumor originated. Tissue specific antigens (TSAs) are expected to be more immunogenic than TSTAs and are more numerous. As such, the therapy of the invention has the advantage of initiating and sustaining a strong immune response against the somatic tissue and tumor until the tumor has been rejected. The therapy also has the advantage of eliciting an immune response against metastatic cancer. In another embodiment, the invention relates to vaccines comprising the tissue specific antigen in a pharmaceutically acceptable carrier and in a size and manner to elicit a class I MHC immune response. In another embodiment, the invention relates to a method for the identification and isolation of a tissue specific antigen which can elicit or induce a class I MHS response.

Description

IMMU O OGICAL THERAPY FOR CANCER

BACKGROUND OF THE INVENTION

Ever since tumor specific associated antigens (TSTA) have been demonstrated in experimental murine tumors (Hellstrom, K.E. and Hellstrom I., Advances in Iiimunology , 18 : 209 (1974); Germain, R.N. et al . , J. Exp . Med . , 142 : 1023 (1975); Doherty, P.C. et al., Adv. Cancer Res . , 42 : 1 (1984)), attempts to generate immune responses against tumors have lacked success. The lack of effective result is believed to be attributed to a number of factors. One factor is that the dynamic growth of the tumors overwhelms the immune responses. Also, the TSTA' s identified to date are only weakly immunogenic with the development of a tolerance against them by the animal . Specific epitopes have been identified which are capable, in association with Class I MHC antigens, of initiating cytotoxic T cell (CTL) responses (Yewdell, J. . and Bennink, J.R., Adv. Immunol . 52 : 1 (1992); Townsend, A. and Trowsdale, A., Sem. Cell Biol . 4:53 (1993)), required for the rejection of solid tissue, and of the necessary secondary signals which must be delivered during antigen presentation to initiate effective T cell responses (Reiser, H. and Benacerraf, B., Proc. Na t . Acad . Sci . USA, 85:10069 (1989); Freeman, G.J. et al . , J. Immunol . 143:2714 (1989); Razi- olf, Z. et al., Proc. Nat . Acad . Sci . USA 89 : 4210 (1992); Galvin, F. et al . , J. Immunol . 149 : 3802 (1992)) . It has become increasingly recognized that only T cell mediated immune responses specific for sequential determinants of internally synthesized tumor antigens associated with Class I MHC antigens have the capacity to invade and destroy established growing solid tumors. Although, several such epitopes, such as prostate specific antigen (PSA) , have been shown to be produced by cancer cells (De Plaen, E. et al . , Proc . Na t . Acad . Sci . USA 85:2274 (1988); Lurquin, C. et al . , Cell 58:293 (1989); Boon, T. Adv. Cancer Res . 58:177 (1992); van der Bruggen, P. et al . , Eur. J. Immunol . 24:3038 (1993); Peoples, G.E. et al . , Proc . Na t . Acad . Sci . USA, 52:432 (1995)), these antigens still have the disadvantage of being weak antigens, necessitating an improved initial and sustained immune response until the tumors have been definitively rejected. Secondly, prostate specific antigen, like many tumor antigens, has the significant disadvantage of being a soluble protein. As such, the immune response is not focused at the site of the tumor. Accordingly, there is a great need for the development of improved immunotherapies for the treatment of cancers .

SUMMARY OF THE INVENTION

The invention relates to a method for treating a solid tumor in somatic tissue, or treating or preventing metastasis of such a tumor, in a patient comprising administering to the patient an amount of one or more antigens specific to the somatic tissue effective to induce an immune response against the somatic tissue. In distinction to the above-described immunotherapies, the antigen or antigens administered to the patient are not specific to the tumor, but are specific to the somatic tissue from which the tumor originated. Tissue specific antigens (TSAs) are more immunogenic than TSTAs and are more numerous. As such, the therapy of the invention has the advantage of initiating and sustaining a strong immune response against the somatic tissue and tumor until the tumor has been rejected. The therapy also has the advantage of eliciting an immune response against metastatic cancer. Furthermore, since the TSA' s are tissue specific, the immune response is expected to include essentially any tumor which is originating from the tissue. In contrast, TSTA's, in being specific to the tumor, are generally not as "universal" as TSA' s between patients or tumors .

In another embodiment, the invention relates to vaccines comprising the tissue specific antigen in a pharmaceutically acceptable carrier and in a size and manner to elicit an immune response such as a Class I or II MHC immune response. In another embodiment, the invention relates to a method for the identification and isolation of a tissue specific antigen which can elicit or induce an MHC immune response.

DETAILED DESCRIPTION OF THE INVENTION

As set forth above, the invention relates to a method for treating a solid tumor in somatic tissue, or treating or preventing metastasis of such a tumor, in a patient comprising administering to the patient a protein or antigen specific to the somatic tissue in an amount effective to induce an immune response against the somatic tissue. Since the antigen is specific to the somatic tissue, the immune response generated by the animal will target not only tumor cells, but also normal somatic tissue cells. In light of this effect, the method of the claimed invention is particularly suitable for the treatment of tumors in non-essential tissues or organs. A "non- essential" tissue or organ is defined herein as tissue or an organ that is not required by the animal for survival or to sustain life. Examples of preferred non-essential tissues and organs include breast tissue, prostate, testes, cervix, ovaries, uterus, thyroid and pancreas, for example. It is particularly preferred to administer the antigen after surgical removal of the tumor and, optionally, the somatic tissue or organ associated with the tumor. In this manner, the immune response generated against the somatic tissue and tumor cells need only destroy the cells remaining after surgery and tumor cells which are circulating or may have metastasized.

A "patient" is intended to include an animal in need of the described therapy. Particularly preferred patients include mammals, such as humans or other primates, rodents, etc .

The tissue specific antigens (TSAs) are administered such that an immune response, such as a class I and/or II MHC immune response, is elicited or induced. As such, the invention further relates to vaccines comprising the tissue specific antigen in a pharmaceutically acceptable carrier and in a size and manner to elicit an immune response. This can be accomplished by administering the antigen or protein in association with particles of appropriate size, as described, for example, in Kovacsovics-Bankowski , M. and Rock, K.L., Eur. J. Immunol . , 24:2421 (1994), Rock, K.L. et al . , J. Immunol . , 150:438 (1993), and Rock, 091/16924 (published November 14, 1991) which are incorporated herein by reference in their entireties. In these antigen presenting cells particle associated proteins are interiorized in phagolysosomes, gain access to the cytosol, and then share a common pathway with endogenously synthesized proteins for class I MHC presentation. In one embodiment, the antigen is administered as membrane fragments of the somatic cells.

One preferred method of administering the antigen is in a vector encoding the antigen which, upon administration, results in the expression of the antigen. The vector can be, for example, a recombinant host cell or virus which comprises a recombinant nucleic acid molecule which encodes the antigen operably linked to a promoter. In this embodiment, the recombinant host cell expresses and, optionally, secretes the antigen. The recombinant host cell can be any suitable cell, such as a bacteria, yeast, fungus, plant or animal cell. Where the host cell is an animal cell containing, in its genome, a nucleic acid which encodes the antigen (or an allelic variant or homolog of the antigen) , the host cell can be transformed or transfected with a nucleic acid which increases expression of the nucleic acid encoding the antigen, thereby causing over-expression of the antigen upon administration. Where the vector is a viral vector containing a nucleic acid encoding the antigen, the vector, upon infection, results in the expression of the antigen. The delivery of proteins by this method are generally described in, for example, Anderson et al . , United States Patent No. 5,399,346, which is incorporated herein by reference.

Examples of suitable viral vectors include retroviruses , polioviruses, vaccinia adenovirus and herpesviruses , particularly attenuated viruses or other viruses characterized by altered genomes to decrease infectivity, as is known in the art.

In a particularly preferred embodiment, the vector is a recombinant bacterium encoding the antigen. Particularly preferred bacteria are bacterial vaccines, such as is described in Cirillo et al . , Clinical Infect . Diseases , 20:1801-9 (1995), which is incorporated herein by reference. An example of a pharmaceutically acceptable bacterial vaccine is MycoJacteriuiτi bovis Bacillus Calmette- Guerin (BCG) . Transfection of foreign genes, including human genes, into BCG has been reported. See, for example, Aldovini A. and Young, R.A., Na ture, 351:479-482 (1991), Stover, C.K. et al . , J^". Exp . Med . , 178:197-209 (1993), Lagranderie M. , et al . , Vaccine, 11:1283-1290 (1993), Connell, N.D., et al . , PNAS , 90:11473-11477 (1993) and

Stover, C.K., et al . , Nature, 351:456-460 (1991), which are incorporated herein by reference. BCG has also been suggested as a treatment for bladder carcinoma by, for example, Cheng et al . , Journal of Urology 152 (4 ) -.1275-1280 (1994) .

In one aspect of the invention, the claimed method results in the initiation of destructive autoimmunity of the somatic tissue and tumor cell. The existence of autoimmune disease specific for several tissues, such as thyroid and pancreas (Wick, G. et al . , Immunol . Rev. 94 : 113 (1986); Smilek, D.E. et al . , Immunol . Rev. , 118 : 31 (1990); Klareskog, L. and Olsson, T. Immunol . Rev. 118:285 (1990)), and the demonstration that destructive autoimmunity could be initiated in experimental animals by immunization with unfractionated tissue extracts in Freund's adjuvant indicate that self-tolerance to tissue specific antigens can be circumvented. It is expected that the immunization with two, or even more, TSAs can further increase the immune response against the tissue and tumor. The vaccines of the present invention can be administered orally (e.g., capsule, tablet or liquid formulation), parenterally (e.g., intra-muscularly, intravenously, subcutaneously) , topically, nasally, intravaginally or rectally (e.g., contraceptive formulation or suppository) , or via slow releasing microcarriers in dosage formulations containing a physiologically acceptable vehicle and optional adjuvants and preservatives. Suitable physiologically acceptable vehicles include saline sterile water, Ringer's solutions, and isotonic sodium chloride solutions. Specifically, Sodium Chloride Injection USP (0.9%), Ringer's Injection USP, Lactated Ringer's Injection USP, Sodium Lactate Injection USP, Dextrose Injection USP (5% or 10%) , Bacteriostatic Water for Injection USP and Sterile Water for Injection USP can be used, for example. The specific dosage level of active ingredient will depend upon a number of factors, including biological activity of the particular preparation, age, body weight, sex, general health and the clinical stage of the disease.

"Tissue specific protein" is defined as a protein which is unique or specific to the somatic tissue or organ where the solid tumor resides or was originated. The protein is present in both normal cells or tissue and tumor cells . The protein is preferably not secreted from the cells, or a "soluble protein".

A "tissue specific antigen" is defined as an immunogenic tissue specific protein or an immunogenic fragment or derivative thereof. An immunogenic fragment is defined as a protein or polypeptide which contains one or more epitopes of the protein. In general, a fragment has a contiguous string of amino acids which are the same as the protein and possess the same or similar three dimensional structure or folding. An immunogenic derivative is defined as a protein, polypeptide or other molecule which contains one or more epitopes of the protein. Examples of immunogenic derivatives are proteins or polypeptides wherein one or more amino acids have been deleted, added or substituted. It is particularly preferred that the amino acids which comprise the epitope are not substituted, deleted or interrupted. It may be particularly desirable that an amino acid substitution, addition or deletion occur at a site in the protein structurally distinct or remote from the epitope. It may also be particularly desirable that an amino acid substitution be conservative, e.g., substituting structurally similar amino acids.

Tissue specific proteins can be identified by a number of known methods. For example, the protein or nucleic acid encoding the protein can be identified through a substractive hybridization method. Subtractive or differential hybridization is described, for example, in Sambrook et al . , Molecular Cloning, A Laboratory Manual, Second Edition, pages 10.40-10.43 and Maniatis et al .

Molecular Cloning, A Laboratory Manual, pp.227-228 (1982), which is incorporated herein by reference. In this embodiment, cDNA and/or RNA libraries are prepared from the somatic cell or tissue of interest and one or more different tissues or cells. The two libraries are then contacted under conditions which permit annealing or hybridization of complementing strands found in the two libraries. Nucleic acids which are unique to one library do not anneal or hybridize. These nucleic acids can then be cloned and identified. To more clearly identify nucleic acids which originate from the library of interest, it may be desirable to label the nucleic acid molecules of one library with, for example, ³²P.

The cells which can be used to generate a library which will subtract nucleic acids encoding non-specific proteins can be, for example, cells which are derived or obtained from organs or tissues which are essential for survival to the animal. Examples include hematopoietic, muscle, neural, epithelial or dermal cells. In another embodiment, the TSA can be identified through differential display, as is described by Pardee and Liang (US Patent 5,262,311 and Science, 251 : 967-971 (1992)) and Liang et al . (Methods in Molecular Genetics, Vol.5, pp 3-16 (1994)), which are incorporated herein by reference. Generally, a set of oligonucleotide primers, one which anchors the polyadenylate tail of a subset of mRNAs, the other being short and arbitrary in sequence so that it anneals different positions relative to the first primer. The mRNA subpopulations defined by these primer pairs can then be amplified after reverse transcription and resolved on a DNA sequencing gel . This method has the advantage of amplifying polynucleotides which are present in even low abundance. A comparison of the nucleotide libraries obtained by this method from cells, as described above, can establish those nucleotides that are constitutively or differentially expressed.

Once a nucleic acid molecule has been identified as being specific or unique to a somatic cell or tissue, PCR can be used to obtain a full length nucleic acid, if required. The full length nucleic acid can then be cloned under control of a suitable promoter, thereby obtaining expressed protein. Optionally, the protein and/or the nucleic acid molecule can be sequenced for further characterization. The protein which is produced can then be evaluated for immunogenic properties, non-secretion and/or for its presence on the surface of the cell. These tests can be readily established by raising antibodies to the protein, labeling the antibody, subjecting a cell expressing the protein to the antibody and detecting the location of the label. Where the protein is a surface or membrane protein with an extra-cellular epitope, the labeled antibody will be detected on the surface of the cell.

In one embodiment, the epitopes of the TSA(s) can be mapped and/or sequenced by known methods. For example, fragments of the antigen can be recombinantly expressed (including expression via phage display) , produced by proteolysis or chemically synthesized. The peptide fragments can be screened for T cell or antibody recognition. Examples of epitope mapping can be found in Petersen, et al . , Mol ecular & General Genetics , 249 (4) :425- 31 (1995), Morris, et al . , Molecular Biotechnology, 4(l):45-54 (1995), Stephen, et al . , Journal of Molecular Biology, 248 { ! ) -. 58 - 18 (1995), Ellgaard, et al . , Journal of Immunological Methods, 180(1) .53-61 (1995), Carter, Methods in Molecular Biology, 36: 201- 23 (1994), Zhao and Chalt,

Analytical Chemistry, 66 (21) : 3723-6 (1994), Mole, Mol ecular Biotechnology, 1(3) -. 211 - 81 (1994) and Beck-Sickinger and Jung, Pharmaceutica Acta Helvetiae, 68 { 1 ) . - 3 - 20 (1993). CTL assays which can be used in mapping the epitope of an antigen or protein are also known and have been described by, for example, Papdopoulos, et al . , Journal of Immunological Methods, 177 (1-2) : 101-111 (1994) and Burrows, et al . , Immunology, 76 (1) :174-5 (1992). The assays can be used to identify epitope (s) or antigen (s) which possess superior activity or immunogenicity .

The peptide fragments comprising one or more epitopes can then be sequenced and employed to manufacture additional tissue specific antigens which can be employed in the methods described above . In another embodiment, tissue specific proteins can be identified by raising antibodies, according to methods known in the art, against membrane or membrane-bound proteins of the somatic cell. These antibodies can then be tested for cross-reactivity with other cell types to ensure uniqueness or specificity of the protein to the cell.

Thus, in another embodiment, the invention relates to a method for the identification and isolation of a tissue specific antigen which can elicit or induce a class I MHC response .

EQUIVALENTS

While this invention has been particularly shown and described with references to preferred embodiments thereof, it will be understood by those skilled in the art that various changes in form and details may be made therein without departing from the spirit and scope of the invention as defined by the appended claims. Those skilled in the art will recognize or be able to ascertain using no more than routine experimentation, many equivalents to the specific embodiments of the invention described specifically herein. Such equivalents are intended to be encompassed in the scope of the claims .

Claims

CLAIMSWhat is claimed is:

1. A method for treating a solid tumor in somatic tissue of a mammal in need thereof comprising administering to said mammal an amount of an antigen specific to the somatic tissue effective to induce an immune response against the somatic tissue.

2. The method of Claim 1 wherein the somatic tissue is in an organ which is not essential to the survival of the mammal.

3. The method of Claim 2 wherein the organ is selected from the group consisting of the breast, testes, uterus, pancreas, prostate, ovary, thyroid or cervix.

4. The method of Claim 1 wherein the antigen is administered as a vector comparising a nucleic acid encoding the antigen.

5. The method of Claim 4 wherein the vector is a recombinant bacterium comprising a nucleic acid coding for the antigen operably linked to a promoter.

6. The method of Claim 5 wherein the host cell is a recombinant MycoJbacteriuπ. bovis Bacillus Calmette- Guerin comprising a nucleic acid coding for the antigen operably linked to a promoter.

7. A method for treating a solid tumor in somatic tissue of a mammal in need thereof comprising inducing a selective immune response against said somatic tissue.

8. The method of Claim 7 wherein the somatic tissue is in an organ which is not essential to the survival of the mammal .

9. The method of Claim 8 wherein the organ is selected from the group consisting of the breast, testes, uterus, pancreas, prostate, ovary and cervix.

10. The method of Claim 9 wherein the selective immune response is against a protein specific to said organ.

11. A method for treating or preventing metastasis of a solid tumor in a somatic tissue of a mammal in need thereof comprising administering to said mammal an amount of an antigen specific to the somatic tissue effective to induce an immune response against the somatic tissue.