Cancer immunotherapy: insights from transgenic animal models
Introduction
Cancer is the second cause of death in the Western industrialized world [1]. Although there has been success in curing non-metastatic cancer [2], [3], most forms of metastatic cancer are on the long run incurable with conventional treatment modalities such as surgery, radio- and chemotherapy. A major limitation of these modalities is the narrow therapeutic window between killing neoplastic while preserving normal cells. In the search for more tumor-specific therapies that are less toxic to normal cells, tumor immunotherapy strategies have gained interest [4]. Fundamental to the immunotherapeutical approach of cancer is the assumption that a tumor differs from normal tissues by tumor antigens, which are either unique (tumor specific antigen, TSA) or relatively restricted to tumor tissue (tumor associated antigen, TAA) [5], [6]. As a consequence of the presence of these TSA or TAA the tumor is capable of inducing a specific immune response existing of a complex of integrated actions of a variety of immune cells, endothelial cells and a wide range of cytokines, growth factors, and antibodies. However, most tumors have developed mechanisms to escape immune surveillance, e.g. by down regulation of MHC and/or costimulatory molecules [7]. Upregulation of molecules which induce anergy and/or apoptosis in the attacking immune effector cells is also a common feature of tumor cells [8], [9].
In the last decade different strategies have been developed for experimental immunotherapy of cancer. To elicit an effective anti-tumor response either the immune response can be potentiated or the tumor cells can be modified to become more immunogenic [10], [11]. Roughly these tumor immunotherapy strategies can be divided into two categories, active and passive immunization strategies. Active specific immunotherapy aims to prime the naı̈ve immune T cells in vivo by presenting tumor antigens via antigen presenting cells, in the context of MHC along with the necessary costimulatory molecules. This has been attempted using intact irradiated tumor cells, gene-modified tumor cells, viral oncolysates, tumor peptides conjugated to an immunogenic carrier molecule or administered in combination with an immune adjuvant, and recombinant viral vectors containing the tumor antigen encoding gene [11]. Another approach is to use ex vivo loaded professional antigen presenting cells such as dendritic cells [12], [13]. In passive immunotherapy strategies, immune system components are added systemically or at the site of the tumor. Adoptive immunotherapy, for example, whereby the patient's autologous immune effector cells are enriched for a subpopulation of anti-tumor immune cells by sorting or expanding the effector cells of interest ex vivo [12]. Also cytokines like tumor necrosis factor α (TNF-α), either alone or fused to anti-tumor antibodies can induce tumor regression. In addition, antibodies fused to drugs or prodrug-activating enzymes can also lower the tumor burden [11]. Antibodies can also mediate the activity of the various non-specific (active non-specific) effector systems. Tumor-specific monoclonal antibodies (Mabs) can mediate cytolysis either by engaging NK, monocytes or granulocytes via Fc receptors (ADCC) or by complement activation. Originally Mabs were of mouse origin and could induce human anti-mouse antibody (HAMA) responses when used in patients. By reducing the immunogenicity of these xenogeneic antibodies, i.e. by ‘humanizing’ the constant regions by recombinant technology, these antibodies can be used repeatedly [14], [15]. Furthermore, elegant bispecific antibody constructs are designed to bring immune effector cells into contact with tumor cells and to simultaneously stimulate their cytotoxic activity. Examples include antibodies that recognize a tumor surface antigen on the one and CD16 on the other hand to activate NK cells [16], or CD3 to activate T cells [17], [18]. Cytokines at the site of the tumor can also recruit immune effector cells [11], [18]. The different strategies employed for experimental immunotherapy of cancer are depicted in Fig. 1.
Although, the number of reports documenting successful immunotherapy in tumor patients increases, specifically when used in minimal residual disease situations [19], [20], there is clearly a need to enhance our knowledge and to evaluate existing and novel immunotherapeutical strategies. Tolerance induction to specific tumor-antigens is a major hindrance for effective immune responses to tumors. To study this, a number of animal models have proven to be of great value, despite this still relevant, more human resembling immunocompetent animal models are needed. Since the first transgenic mice were generated in 1982 [21], transgenic animal models have been used extensively to investigate biomedically important mechanisms underlying a variety of diseases. For cancer, transgenic mouse models promoting tumorigenesis have advanced our understanding of the mechanisms by which cancer initiates and progresses [22]. The last decade, however, transgenic animal models are no longer used solely to understand the pathogenesis of disease but also to develop and evaluate new therapies. To evaluate tumor-immunotherapeutical strategies transgenic animals have been generated which express tumor associated antigens, human HLA, oncogenes, mutated tumor suppressor genes, and also human immune-effector cell molecules (Table 1). At first non-specific promoters were used to express the genes of interest, which resulted in expression in all tissues. However, with the growing availability of the genomic sequences of genes, transgenic animal models have now been generated expressing the transgene accurately in a cell and tissue specific manner. Although, now valuable immunocompetent transgenic mice can be generated a lot of tumor models have been established and evaluated in the past in animal strains that are not suitable for the development of transgenic animals. Therefore, frequently a lot of time-consuming backcrossing is necessary to finally obtain a transgenic animal model in which tumors can be induced. To circumvent this problem, transgenic animals expressing the transgene of interest are crossed with transgenic animal models generated to develop ‘spontaneously’ certain tumors, e.g. expressing the SV40 large T antigen oncogene in a tissue specific manner [23]. A disadvantage of the latter approach is the fact that previously obtained knowledge from tumor bearing animal models generated before the era of transgenesis can not be implemented in the evaluation of immunotherapeutical strategies in transgenic animal models. For the same reason it is sometimes desirable to express a transgene in different species than the mouse, although the mouse remains the species of choice for many experiments involving the introduction of foreign DNA into the genome, simply because of technical limitations regarding other species [24].
Since so many different transgenic animals have been generated to evaluate a plethora of different anti-tumor immunotherapeutical strategies with variable results, this review aims to give a survey on the existing models and to discuss their potential regarding anti-cancer immunotherapeutical strategies.
Section snippets
Transgenic rodent models expressing tumor associated antigens
Clinical studies can be difficult to implement, particularly when clear understanding of the potential efficacy, limitations, and safety of an immunotherapeutic strategy is not available from relevant animal investigations. TAA as opposed to TSA are besides present on the tumor also present on normal tissue. Therefore, immune-mediated therapies directed at tumor associated antigens deal with a balance between desirable anti-tumor responses and unwanted autoimmune reactions. Mice carrying a
HLA transgenic mouse models
Among the numerous genes studied for their role in disease development, polymorphisms associated with HLA class I and class II loci has been known for many years to play a significant role in predisposition to disease. HLA molecules are encoded by MHC genes on the short arm of chromosome 6. Structure analysis of the HLA molecules revealed that it contained a peptide binding cleft in which the variable region of the HLA molecules is situated [94]. Genetic polymorphism at the MHC locus determines
Oncogene transgenic mice to study immunotherapeutical strategies
The generation of mice designed to overexpress activated forms of oncogenes has allowed scientists to causally link the function of these genes with specific tumor cell functions, such as proliferation, apoptosis, angiogenesis or metastasis [125]. In addition, these mice can be used to develop and test new therapies, such as tumor immunotherapy.
Transgenic mice expressing immune effector cell molecules
Immune effector cells can recognize tumor cells and initiate their elimination. For this the immune cell uses specific molecules. To further investigate the mechanisms involved in this immune activating transgenic mice have been generated expressing modified human immune effector molecules.
Concluding remarks
Transgenic mouse models to study anti-cancer immunotherapeutical strategies have allowed one to study the effects of different immunotherapy approaches on cancer. The mouse models used can be divided into four groups, the TSA or TAA, the HLA, the oncogene, and the immune effector molecule transgenic mice, which are each of particular interest to a specific immunotherapeutical approach. Results obtained using these mice have already found their ways to the patient, as they are currently being
Reviewer
Professor Jacques Robert and Dr Alain Ravaud, Institut Bergonié, 180, rue de Saint-Genès/229, cours de l'Argonne, F-33076 Bordeaux cedex, France.
Acknowledgments
We apologize to those whose work has not been cited due to space limitations.
Dr Pamela M.J. McLaughlin graduated from the University of Groningen, The Netherlands, obtaining her doctorate in Medical Sciences and is currently gaining Post-Doctoral experiences at the Laboratory of Tumor Immunology in collaboration with Dr Bart-Jan Kroesen and Dr Martin C. Harmsen under supervision of Professor Dr Lou F.M.H. de Leij.
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Dr Pamela M.J. McLaughlin graduated from the University of Groningen, The Netherlands, obtaining her doctorate in Medical Sciences and is currently gaining Post-Doctoral experiences at the Laboratory of Tumor Immunology in collaboration with Dr Bart-Jan Kroesen and Dr Martin C. Harmsen under supervision of Professor Dr Lou F.M.H. de Leij.
Dr B.J. Kroesen is an immunologist at the Department of Pathology and Laboratory Medicine at the University Hospital Groningen and has special research interest in immune homeostasis in relation to pathology and immunotherapeutical treatment.