1. Murdoch Children’s Research Institute,
2. Department of Haematology and Oncology,
Royal Children’s Hospital and University of Melbourne,
Department of Paediatrics,
Cancer immunotherapy aims to exploit and maximise the body’s own immune system in order to target and destroy cancer cells. The immune system is a complex and effective integrated network of specialised cells, organs and factors (cytokines and antibodies) that can quickly and efficiently identify and remove foreign agents such as bacteria and viruses, and “self agents” such as cellular debris and malfunctioning cells that are dangerous to the host. There are several mechanisms in place to alert the immune system to these dangerous self cells in order to safeguard against the development of cancer and other diseases. This process is termed immunosurveillance, and it allows for constant screening by the immune system for malfunctioning cells in order to eliminate them before they can cause harm1. However, when cancer develops, immunosurveillance mechanisms have been averted and the harmful cells are allowed to survive. Furthermore, cancer cells no longer respond to appropriate growth controls and therefore multiply without constraint and become dangerous to the host. Therefore, the coordinate failure of cells to respond to growth control signals and the failure of the effective immunosurveillance mechanisms to alert the immune system to destroy aberrant cells can lead to malignant disease.
Cancer poses a particularly difficult problem to the immune system, as these cells have overcome immunosurveillance mechanisms and are recognised as self and therefore do not illicit an immune response. Cancer cells can alter their behaviour in many ways to avoid detection and deletion. Firstly, they can overcome programmed cell death (apoptosis) mechanisms that cause cells to die when they have acquired mutations that inappropriately signal the cell to cycle and proliferate. Furthermore, many chemotherapy agents work by triggering apoptotic pathways in cycling cells and thus some cancers are resistant to these types of chemotherapeutic agents due to alterations in their apoptotic machinery. Secondly, cancer cells can evade detection of the immune system by altering the expression of cell surface molecules. MHC molecule expression is essential to trigger an immune response by activating T lymphocytes through the T cell receptor (TCR). Therefore, it is common for cancer cells to down-regulate expression of its MHC molecules2. Finally, cells can secrete immunosuppressive soluble cytokines such as IL-103 and TGF-b4 that can down-regulate the immune response. Generally, these cytokines act as brakes on the immune system to control the immune response in order to prevent damage that can be caused by the immune system when unregulated. Therefore, when immunosuppressive cytokines are inappropriately expressed, it can dampen immune responses and allow for cancer cells to avoid attack by the immune system.
Malignant cells can also up-regulate the expression of certain cell surface molecules that may not be innately antigenic but may be useful as tumour associated antigens (TAA) in future therapies such as prostate specific antigen (PSA). An example of well-studied tumour-associated antigens is the MAGE and GAGE families of genes5. While these antigens were initially described in melanoma, they have been demonstrated to be present in a variety of tumour types including lung and bladder carcinoma, sarcomas, and head and neck tumours6. They are, however, non-detectable in a large range of normal tissues, including brain, bone marrow and peripheral blood. Therefore, they may be used as potential targets for future therapies.
The aim of immunotherapy approaches is to prime the immune system to target these cancer cells specifically and without creating an autoimmune response. Immunotherapy can refer to any method in which the immune system is being altered to become more effective. Generally, there are three modes of immunotherapy that are currently being utilised – antibodies, cytokines and cellular immunotherapy.
Antibodies have been used in a variety of ways to affect cellular behaviour. They can be administered to replace naturally-occurring ligating events. When antibodies bind to cell surface molecules they can have activating, inhibiting or null effects on cell signalling. It is possible to use activating antibodies to ligate death receptors on cancer cells in order to cause these cells to die (ie Fas)7. It is also possible to ligate lymphocyte cell surface receptors in order to induce lymphocytes to expand and activate an immune response (B7.1, LFA-3, ICAM-1)8. In addition, blocking antibodies can be used to interfere with naturally-occurring ligation events that are activating. Using a blocking antibody to the epidermal growth factor receptor (EGF-r) has been effective in reducing the growth of several tumour types that have amplified EGF receptor expression9. Further, as described in this paper by Dr Frazer, anti-viral vaccines can be administered to produce neutralising antibodies against the papilloma virus, which is responsible for cervical cancer. Finally, antibodies can be used to carry cytotoxic drugs to specific cells that express the ligand10. This method aims to target only the cancer cells to receive cytotoxic agents by conjugating the toxic agent to the antibody that has been shown to be specific for only the cancer cell using TAAs. In each method, antibodies can be used to specifically target cancer cells and can be exploited by choosing the appropriate antibody to achieve altered cellular outcomes.
Cytokines are soluble factors that direct and modulate the nature of an immune response. Granulocyte-Colony Stimulating Factor (G-CSF) is regularly administered to chemotherapy patients to boost neutrophil counts following treatment. Furthermore, cytokines (ie FLT-3 Ligand) can also be used to differentiate cells in vivo for later harvesting for more invasive immunotherapy applications11. Finally, it has been shown that cytokines can have local toxic effects in high doses and some direct applications of cytokines to tumours can cause tumour regression (IL-2)12,13. Therefore, cytokines can have both supportive and therapeutic roles in treating cancer patients.
Cellular immunotherapy involves the alteration of autologous immune cells ex vivo, which are then administered to the patient to produce a specific anti-tumour effect. Current models are focusing on the use of dendritic cells (DC), which are the most potent antigen-presenting cells and therefore the best candidates to introduce tumour-specific antigens. The enormous potential for exploiting DC for immunotherapy has been hindered until recently by the rarity of this cell type in the human body (less than 1.0% of mononuclear cells14,15) and the lack of methods to generate DC in vitro. There are two major ways of isolating human DC:
The fundamental role of DC in orchestrating the different arms of the immune system defines them as important mediators of the immune response. Cell-mediated responses include adaptive responses facilitated by CD4+ and CD8+ T cells and the innate response facilitated by natural killer (NK) and natural killer T cells (NKT) cells. DCs engage and activate these lymphocyte subsets via separate mechanisms in order to control and define the nature of the immune response generated. Each lymphocyte subset has a unique mechanism for killing target cells, but they all produce the anti-tumour cytokine IFN-g in response to activation. DCs present peptide antigens within the context of MHC class I and II to CD8+ and CD4+ T cells respectively. CD8+ cytolytic T cells kill quickly via granzymes when activated after ligation with MHC I and co-stimulation molecules such as a B7 family member. CD4+ T helper cells, when activated – also through co-stimulation molecules and ligation with peptide-MHC II – can kill target cells through Fas-FasL interactions. NKT cells recognise lipid and glycolipid antigens within the context of CD1a expressed on DC while NK cells recognise and kill cells that do not express MHC I molecules. This dynamic arrangement allows the interplay between initiators and effectors to produce a multi-pronged attack against antigen-bearing cells.
Immunisations using autologous dendritic cells loaded with tumour antigens should overcome two of the major issues in cancer therapies today – donor suitability and engraftment complications (attaining suitable donors, graft rejections and graft versus host disease), and the specificity of action of the therapeutic agent against the tumour alone. Several successful studies in murine models of malignancy have increased the potential of DC vaccination as a possible form of therapy in humans. These studies have shown that DC pulsed with specific antigens induces both protective and therapeutic tumour immunity in immunocompetent mice [A Porgador and A Gilboa (1995), C Celluzzi, et al (1996), P Paglia, et al (1996)]. Various methods have been used to load DC with antigen that result in anti-tumour immunity. Tumour material in the form of peptides18–20, cell sonicates21 and RNA22,23 have all been used in an effort to mount a specific anti-tumour response by the immune system.
Preliminary studies in humans using DC alone or DC loaded with antigen have been conducted in patients with advanced malignant disease. A feasibility and toxicity study was performed using monocyte-derived DC (MoDC), which demonstrated that sufficient numbers of MoDC could be generated in vitro to fulfil dose requirements. Cryopreserved MoDC were then administered to patients without any adverse effects24. This study paved the way for further DC studies where DCs loaded with tumour lysate were injected into patients and anti-tumour immunity was shown by tumour-specific T cell activation and IgM/IgG antibody production25,26. Taken together, these studies demonstrate the potential beneficial use of DC-based vaccines. The results of recent clinical trials and recent advances in DC vaccination are further discussed by Dr Hart and his group below.
The status of immune system function in cancer patients has recently been of interest with respect to new approaches of anti-cancer therapies. However, these studies have focused on immunocompentency following chemotherapy, and baseline data prior to chemotherapy is lacking. Patients with acute lymphoblastic leukaemia (ALL), Hodgkin’s disease, or solid tumours were examined for immune function after successful chemotherapy (with or without radiotherapy)27. Immune responses to specific antigens were lower than normal for both the humoral and cellular arms of the immune system in most patients, with only 19% demonstrating normal responses 12 months post-chemotherapy27. Further studies involving the cytokine profiles of cancer patients have shown that IL-2 is deficient in PBSC after traditional chemotherapy and bone marrow transplantation28, and mononuclear cells in patients with advanced cancer show deficiency in T helper 1 responses (decreased IFN-g, IL-10, IL-12 and increased IL-4)29. While it is important to assess immune function following chemotherapy, baseline immunologic data is necessary to draw meaningful conclusions as to the status of immune function in cancer patients prior to therapy.
There have been many advances recently in the field of cancer immunotherapy, and with the forthcoming publication of clinical trials underway we are certain to refine and improve our methods and efficacy for future therapies.
2. F Garrido, F Ruiz-Cabello, T Cabrera, J J Perez-Villar, M Lopez-Botet, M Duggan-Keen, P L Stern. “Implications for immunosurveillance of altered HLA class I phenotypes in human tumours.” Immunol Today,18, 2 (Feb 1997): 89-95.
5. T F Logan, W Shannon, J Bryant, P Kane, N Wolmark, M Posner, J M Kirkwood, M S Ernstoff, J W Futrell, L D Straw, et al. “Preparation of viable tumour cell vaccine from human solid tumours: relationship between tumour mass and cell yield.” Melanoma Res, 3, 6 (Dec 1993): 451-5.
6. B Van den Eynde, O Peeters, O De Backer, B Gaugler, S Lucas, T Boon. “A new family of genes coding for an antigen recognised by autologous cytolytic T lymphocytes on a human melanoma.” J Exp Med,182, 3 (Sep 1995): 689-98.
11. M Sivanandham, C I Stavropoulos, E M Kim, B Mancke, M K Wallack. “Therapeutic effect of colon tumor cells expressing FLT-3 ligand plus systemic IL-2 in mice with syngeneic colon cancer.” Cancer Immunol Immunother, 51, 2, (2002): 63-71.
12. S M Dubinett, L Patrone, J Tobias, A J Cochran, D R Wen, W H McBride. “Intratumoral interleukin-2 immunotherapy: activation of tumor-infiltrating and splenic lymphocytes in vivo.” Cancer Immunol Immunother, 36, 3 (1993): 156-62.
15. S Patterson, M Helbert, N R English, A J Pinching, S C Knight. “The effect of AZT on dendritic cell number and provirus load in the peripheral blood of AIDS patients: a preliminary study.” Res Virol, 147, 2-3 (Mar-Jun 1996): 109-14.
16. N Romani, S Gruner, D Brang, E Kampgen, A Lenz, B Trockenbacher, G Konwalinka, P O Fritsch, R M Steinman, G Schuler. “Proliferating dendritic cell progenitors in human blood.” J Exp Med, 180, 1 (Jul 1994): 83-93.
17. D Strunk, K Rappersberger, C Egger, H Strobl, E Kromer, A Elbe, D Maurer, G Stingl. “Generation of human dendritic cells/Langerhans cells from circulating CD34+ hematopoietic progenitor cells.” Blood, 87, 4 (Feb 1996): 1292-302.
19. C M Celluzzi, J I Mayordomo, W J Storkus, M T Lotze, L D Falo Jr. “Peptide-pulsed dendritic cells induce antigen-specific CTL-mediated protective tumor immunity.” J Exp Med, 183, 1 (Jan 1996): 283-7.
20. P Paglia, C Chiodoni, M Rodolfo, M P Colombo. “Murine dendritic cells loaded in vitro with soluble protein prime cytotoxic T lymphocytes against tumor antigen in vivo.” J Exp Med, 183, 1 (Jan 1996): 317-22.
23. D M Ashley, B Faiola, S Nair, L P Hale, D D Bigner, E Gilboa. “Bone marrow-generated dendritic cells pulsed with tumor extracts or tumor RNA induce antitumor immunity against central nervous system tumors.” J Exp Med, 186, 7 (Oct 1997): 1177-82.
24. M A Morse, Y Deng, D Coleman, S Hull, E Kitrell-Fisher, S Nair, J Schlom, M E Ryback, H K Lyerly. “A Phase I study of active immunotherapy with carcinoembryonic antigen peptide (CAP-1)-pulsed, autologous human cultured dendritic cells in patients with metastatic malignancies expressing carcinoembryonic antigen.” Clin Cancer Res, 5, 6 (Jun 1999): 1331-8.
26. L Holtl, C Rieser, C Papesh, R Ramoner, M Herold, H Klocker, C Radmayr, A Stenzl, G Bartsch, M Thurnher. “Cellular and humoral immune responses in patients with metastatic renal cell carcinoma after vaccination with antigen pulsed dendritic cells.” J Urol, 161, 3 (Mar 1999): 777-82.
27. M M Mustafa, G R Buchanan, N J Winick, G H McCracken, I Tkaczewski, M Lipscomb, Q Ansari, M S Agopian. “Immune recovery in children with malignancy after cessation of chemotherapy.” J Pediatr Hematol Oncol, 20, 5 (Sep-Oct 1998): 451-7.
28. K Welte, N Ciobanu, M A Moore, S Gulati, R J O’Reilly, R Mertelsmann. “Defective interleukin 2 production in patients after bone marrow transplantation and in vitro restoration of defective T lymphocyte proliferation by highly purified interleukin 2.” Blood, 64, 2 (Aug 1984): 380-5.
29. S Goto, M Sato, R Kaneko, M Itoh, S Sato, S Takeuchi. “Analysis of Th1 and Th2 cytokine production by peripheral blood mononuclear cells as a parameter of immunological dysfunction in advanced cancer patients.” Cancer Immunol Immunother, 48, 8 (Nov 1999): 435-42.