Ovarian cancer is the eighth most common type of cancer among women. The American Cancer Society estimates that about 22,220 new cases of ovarian cancer will be diagnosed in the United States during 2005. Ovarian cancer accounts for about 3% of all cancers in women. Because many ovarian cancers cannot be detected early in their development, they account for a disproportionate number of fatal cancers, being responsible for almost half the deaths from cancer of the female genital tract; more deaths than any other reproductive organ cancer. Older women are at higher risk. More than half of the deaths from ovarian cancer occur amongst women between 55 and 74 years of age. About 25% of ovarian cancer deaths occur amongst women between 35 and 54 years of age.
The main treatments for ovarian cancer are surgery, chemotherapy, and radiation therapy. Combinations of these treatments are used to treat ovarian cancer.
Surgery is the usual initial treatment for women diagnosed with ovarian cancer. The ovaries, the fallopian tubes, the uterus, and the cervix are usually removed. Staging during surgery (to find out whether the cancer has spread) generally involves removing lymph nodes, samples of tissue from the diaphragm and other organs in the abdomen, and fluid from the abdomen. If the cancer has spread, the surgeon usually removes as much of the cancer as possible. This reduces the amount of cancer that will have to be treated later with chemotherapy or radiation therapy
Chemotherapy is the use of drugs to kill cancer cells. Chemotherapy may be given to destroy any cancerous cells that may remain in the body after surgery, to control tumour growth, or to relieve symptoms of the disease. Most drugs used to treat ovarian cancer are given intravenously or directly into the abdomen through a catheter.
Radiation therapy, also called radiotherapy, involves the use of high-energy rays to kill cancer cells. Radiation therapy affects the cancer cells only in the treated area. The radiation may come from a machine or women receive a treatment called intraperitoneal therapy in which radioactive is put directly into the abdomen through a catheter.
Deciding on a particular course of treatment is typically based on a variety of prognostic parameters and markers [Fitzgibbons, et al. (2000) Arch. Pathol. Lab. Med. 124:966-978; Hamilton and Piccart (2000) Ann. Oncol. 11:647-663)], including genetic predisposition markers BRCA-1 and BRCA-2 [Robson (2000) J. Clin. Oncol. 18:113sup-118sup].
Although many ovarian cancer patients are effectively treated, the current therapies can all induce serious side effects, which diminish quality of life. Moreover, approximately 85% of the patients that have been effectively treated with platinum- and paclitaxel-based chemotherapy, including complete responses, relapse within two years after treatment.
The identification of novel therapeutic targets is essential for improving the current treatment of ovarian cancer patients. Recent advances in molecular medicine have increased the interest in tumour-specific cell surface antigens that could serve as targets for various immunotherapeutic or small molecule strategies.
Among the various elements of the immune system, T lymphocytes are probably the most adept to recognize and eliminate cells expressing foreign or tumour-associated antigens. Cytotoxic T Lymphocytes (CTLs) express the CD8 cell surface marker and are specialized at inducing lysis of the target cells with which they react via the perforin/granzyme and/or the Fas/Fas-L pathways. The T-cell receptor (TCR) for antigen of CTLs binds to a molecular complex on the surface of the target cell formed by small peptides (8-11) residues derived from processed foreign or tumour associated antigens, which associate with major histocompatibility complex (MHC) class I molecules.
The other major T-cell subset, helper T lymphocytes (HTLs or T helper cells), is characterized by the expression of CD4 surface marker. The T helper cells recognize slightly larger peptides (11-20 residues), also derived from foreign or tumour associated antigens, but in the context of MHC class II molecules, which are only expressed by specialized antigen presenting cells (APCs) such as B lymphocytes, macrophages and dendritic cells (DCs).
As a consequence of TCR stimulation of naive CTLs and HTLs by peptide/MHC complexes on APCs, the CTLs mature into effector killer cells capable of lysing (tumour) cells that express the corresponding peptide/MHC class I complex. Activated HTLs amplify CTL responses by making the APCs more effective at stimulating the naive CTLs and by producing lymphokines that stimulate the maturation and proliferation of CTLs. The potentiating effect of T helper cells occurs both in secondary lymphoid organs where the immune response is initiated and at the tumor site where CTL responses need to be sustained until the tumour cells are eliminated. Thus, one would predict that vaccines should stimulate both tumour-reactive CTLs and HTLs to generate effective antitumour immunity.
Antigens suitable for immunotherapeutic cancer strategies should be highly expressed in cancer tissues and ideally not in normal adult tissues. Expression in tissues that are dispensable for life, however, may be acceptable.
A number of antigens suitable for immunotherapeutic strategies in the treatment of ovarian tumors have been described so far, including MUC1, CTs, SP17 and Her2/neu.
Polymorphic epithelial mucin (MUC1) is a transmembrane protein, present at the apical surface of glandular epithelial cells. It is often overexpressed in ovarian cancer (in more than 90% of all ovarian cancers), and typically exhibits an altered glycosylation pattern, resulting in an antigenically distinct molecule. MUC1 is in early clinical trials as a vaccine target [Gilewski, et al. (2000) Clin. Cancer Res. 6:1693-1701; Scholl, et al. (2000) J. Immunother. 23:570-580]. The tumour-expressed protein is often detectable as tumor marker in the circulation [cf. Bon, et al. (1997) Clin. Chem. 43:585-593].
A unique class of differentiation antigens, the cancer/testis (CT) antigens, are not expressed in normal tissues except for testis and, in some cases, placenta. This fact makes CT antigens attractive targets for specific immunotherapy of cancer. The function of the majority of the CT antigens is currently unknown. Tammela et al. [Tamella, et al. (2004) Cancer Immunity 4:10-21] demonstrated that SCP-1, a CT antigen with a known role in gamete development, is expressed in 15% of ovarian cancer cases. It was suggested that because of its restricted expression in normal tissues and its aberrant expression in tumour tissues SCP-1 might serve as a potential target for vaccine therapy in ovarian cancer.
Another potential target for immunotherapy in patients with ovarian carcinoma is the sperm protein 17 (SP17). Sp17 was found to be expressed in the primary tumor cells from 70% of the patients with ovarian carcinoma. The restricted expression of Sp17 in normal tissue makes it an ideal target for tumour vaccine. A recombinant Sp17 protein was used with monocyte-derived dendritic cells and autologous peripheral blood mononuclear cells to generate Sp17 specific cytotoxic T-lymphocytes (CTLs). Human leukocyte antigen (HLA) class II restricted Sp17 specific CTLs were generated successfully from the peripheral blood of three patients with ovarian carcinoma at the time of disease presentation. These CTLs were able to lyse autologous Epstein-Barr virus-transformed lymphoblastoid cells in an Sp17-dependent manner. The CTLs also lysed Sp17-positive autologous tumour cells, suggesting that Sp17 is processed and presented in association with the HLA class I molecules in Sp17-positive tumour cells. [Chiriva et al. (2002) Cancer 94(9):2447-2453]
Human epidermal growth factor receptor 2 (Her2/neu) is an oncogene that is activated by gene amplification with the increased expression of another (normal) gene product. Her2/neu is overexpressed in 20 to 30% of patients with breast and ovarian cancer. Initial studies to develop a peptide based HER-2/neu vaccine were performed in a rat model [Disis et al. (1999) Clinical Cancer Research 5:1289-1297]. No T-cell responses or anti-body responses were observed in animals immunized with intact rat neu protein. By marked contrast, tolerance to rat neu protein in rats, could be circumvented by immunization with a peptide based vaccine. Rats immunized with neu peptides designed for eliciting CD4+ T-cell responses, generated T-cell and antibody responses specific for both the immunizing peptides and the whole protein.
Brossart et al. demonstrated that patients with advanced breast and ovarian cancer could be efficiently vaccinated with autologues dendritic cells (DCs) pulsed with Her2/neu- or MUC1-derived peptides. In 5 out of 10 patients peptide specific CD8+ cytotoxic T lymphocytes could be detected in the peripheral blood. It was reported that MAGE-3- and CEA-peptide-specific CD8+ T cells were observed in one patient treated with MUC-1 peptide-pulsed DCs, and MUC-1 specific T-cells were observed in another patient after vaccination with HER2/neu derived peptides. It was suggested by Brossart et al. [Brossart et al. (2002) Transfus Apher Sci. 27(2):183-186] that this indicated that epitope spreading occurred in these patients upon treatment.
Epitope spreading is a recognized phenomenon of autoimmune responses and is believed to be an exacerbating factor in CD4+ T cell-mediated autoimmune diseases. The phenomenon has been demonstrated in murine relapsing-remitting experimental autoimmune encephaleomyelitis (EAE), Theiler's murine encephalomyelitis virus-induced demyelineating disease and diabetes in the non-obese diabetic (NOD) mouse. A model has been suggested for how epitope spreading in autoimmune diseases mediated by CD4+ T cells occurs. This model is supported by direct evidence that tissue damage, TCR ligation on CD4+ T cells by MHC class II-peptide complexes, CD40-CD40 ligand interactions and CD28-mediated co-stimulation are required for epitope spreading to become manifest. It is thought that an initiating self-antigen or a persistent viral epitope, presented in MHC class II molecules on the surface of professional antigen-presenting cells (APC) residing in the target tissue, causes the activation of CD4+ T cells specific for that antigen. This T cell activation results in chronic inflammation, leading to damage of the target tissue. Tissue debris is subsequently taken up by APC which have up-regulated expression of MHC class II and co-stimulatory molecules in response to inflammatory cytokines. These APC are then capable of activating CD4+ T cells specific for secondary tissue epitopes presented by the APC. The newly activated T cells then aid in destruction of the target tissue.
Due to the requirement for presentation by APC of exogenous antigen, epitope spreading has historically been thought of as a phenomenon unique to CD4+ T cell responses. However, recent data have indicated that cross-priming by APC can participate in the induction of CD8+ cytotoxic T lymphocyte (CTL) responses as well. In particular, bone marrow chimera studies in murine tumour models have shown that tumour-specific CTL are predominately restricted to the MHC of the host rather than that of the tumour, suggesting that indirect presentation by host APC is involved in the generation of tumour-specific CTL. Moreover, there is increasing evidence that a pathway exists whereby exogenous antigen can be presented for eventual peptide loading onto class I MHC molecules. This phenomenon is best described for dendritic cells (DC) and provides a cellular mechanism to explain the process of cross-priming. Collectively, these data suggest that it may be possible for epitope spreading to occur during a class I MHC-restricted CTL response. Because re-presentation of MHC class I-restricted tumour antigens is known to occur, it has been postulated that if tumour-bearing hosts could initiate a CTL response against a single tumour antigen, that following tumor cell damage caused by the CTL, epitope spreading might occur via a mechanism analogous to that described in CD4+ T cell-mediated autoimmune diseases. Unlike during an autoimmune response, however, CTL epitope spreading during an anti-tumour response could be beneficial to the host by possibly allowing for elimination of variant tumor cells that have lost expression of the antigen (antigen negative tumour cells). [Markiewicz et al., (2001) International Immunology 13:625-632].
Markiewicz et al found that immunization with the single tumour peptide P1A followed by tumour rejection led to CTL activity against a P1A− tumour, indicating that the phenomenon of epitope spreading is not limited to CD4+ T cell responses. The population of CTL included cells recognized the unrelated antigen PIE. Since this epitope was not included in the vaccine and is a mutated peptide not presented in normal tissues, the source of PIE antigen must have been the tumor cell challenge.
Because many patients have ovarian tumours that express neither one of the aforementioned antigens there is a need to uncover additional antigenic targets for immunotherapy to manage localized and metastatic disease. Accordingly, provided herein are molecular targets for immunotherapeutic intervention in ovarian cancers.
The zona pellucida (ZP) forms an extracellular glycoprotein matrix surrounding the developing and ovulated oocyte and the preimplantation embryo and is also detectable in atretic follicles. The ZP induces acrosome reaction on sperm, determines the species specificity for fertilization and prevents polyspermy in mammals. The zona pellucida contains four major glycoproteins, ZP1, ZP2, ZP3 and ZP4. In vitro studies in mice indicate that O-linked oligosaccharide side chains of ZP3 are involved in the primary binding of the sperm to the ZP3, while ZP2 contributes to the subsequent and persistent ZP binding and functions as a secondary sperm receptor.
The ZP glycoproteins have been studied extensively for the development of vaccines for the fertility control of animals and humans. The proposed vaccine action is the induction in female subjects of effective sustained, but reversible levels of ZP-specific antibodies that inhibit sperm-egg binding and/or prevent sperm penetration of the ZP. Passive immunization of female mice with rat monoclonal antibodies against mouse ZP2 or ZP3 resulted in localization of the antibodies to intra-ovarian oocytes and long-lasting but reversible contraception. Active immunization of female mice with ZP3-derived peptides ZP3328-342, comprising a B-cell epitope recognized by the ZP3-specific contraceptive antibody, also led to reversible albeit incomplete contraception. These ZP3 peptides also induced a T cell response to the ZP3 peptide. These CD4+ ZP3 specific T cells adoptively transfer autoimmune ovarian disease (AOD) to syngeneic recipients. Since the desired contraceptive effect of ZP3 immunization is known to be mediated by antibodies, an acceptable contraceptive ZP vaccine should induce an adequate antibody response without activation of ZP3-specific T cells. Indeed a chimeric peptide consisting of a foreign T-cell epitope from bovine ribonuclease (RNase) and a minimal and modified murine ZP3335-342 B cell epitope has been designed that elicits antibodies to ZP and has a significant contraceptive effect without causing significant oophoritis/AOD. The bovine RNase T-cell epitope stimulates helper T cell (helper T lymphocytes, HTL) responses in mice, thus potentiating the contraceptive effectiveness without inducing ZP(3)-specific T cell action and T-cell mediated ovarian damage.
Immunisation with (self)ZP antigen has also been used to study autoimmune ovarian disease (AOD). More in particular, animal models suitable for studying AOD have been reported wherein autoimmune disease was induced using ZP antigen vaccination. For example, it was demonstrated by Rhim et al. [Rhim et al. (1992) J. Clin. Invest. 89:28-35] that in B6AF1 mice T-cell and antibody response were induced by vaccination with mouse ZP3328-342 peptide. Further studies on truncated ZP3328-342 peptides substantiated that a T cell response is sufficient for induction of oophoritis; seven of such peptides lacking antibody binding sites, elicited severe oophoritis without concomitant antibody response. These peptides include a minimal oophoritogenic peptide of eight amino acids, ZP3330-337, which overlaps the seven amino acid antibody binding site, ZP3336-342, by two residues.
It was reported by Bagavant et al. [Bagavant et al. (1999) Biology of Reproduction 61:635-642] that transfer of ZP3 peptide-specific T-cells into naïve recipient mice resulted in granulomatous oophoritis and enhanced ovarian expression of IL-1, TNF-α and IFN-γ. However the ovarian function of cell recipients was normal and the mice remained fertile. Antibody to ZP3 alone does not cause any ovarian pathology. Co-transfer of pathogenic T cells and ZP antibody together targets the inflammation into developing follicles leading to their destruction and the development of ovarian atrophy. In another study Bagavant et al [Bagavant et al. (2002) American Journal of Pathology 160:141-149] demonstrated that ZP3 peptide (human ZP3328-341, macaque ZP3328-341 and mouse ZP3330-342) immunization in primates can elicit a T-cell response and cause ovarian immunopathology that is similar to murine AOD.
International patent application no. WO 2005/026735 (Buschmann et al.) relates to differentially expressed tumour-specific immunogenic membrane proteins and to their uses, in particular for finding at least one therapeutic molecule or compound which specifically regulates the expression of at least one of said membrane proteins, or for finding a therapeutic molecule that specifically binds to and/or interacts with any of said membrane proteins. The membrane protein can be SYPL, STOML2, RAGA, CLNS1A, PRNP, GNB2L1, GNG4, ITM2B, ITM1, TM9SF2, TM4SF6, OPRL1, LRP4, GLEPP1, TLR3 and/or ZP3. WO 2005/026735 teaches to administer the aforementioned therapeutic molecule or compound to neoplastic target cells for modulating proliferation, differentiation and/or cell migration of said neoplastic target cells. It is stated that the non-steroid dependent cancer to be treated results from the aberrant expression and/or biological activity of at least one of said immunogenic membrane proteins. It is also briefly mentioned in WO 2005/026735 that the development of a specific lesion, such as a pro-neoplastic lesion that can be found in epithelial tissues, into a neoplastic lesion can be inhibited by inoculating a subject with one of said membrane proteins adequate to produce antibody and/or T cell immune response. It is further specified that according to another embodiment the method comprises delivering one of the immunogenic membrane proteins via a vector directing expression of the said protein in vivo in order to induce such an immunological response to produce antibody to protect the subject from disease. Thus, WO 2005/026735, teaches several methods of suppressing the expression and/or biological activity of, amongst others, ZP3 membrane proteins in neoplastic target cells in order to modulate proliferation, differentiation and/or migration of said target cells, using either siRNA, receptor antagonists or antibody. WO 2005/026735 only discloses the expression of ZP3 membrane protein in certain colon cancer cells. No other reports of tumour associated expression of any ZP glycoprotein are known.