Immunotherapy for Osteosarcoma: Genetic Modification of T cells Overcomes Low Levels of Tumor Antigen Expression

Center for Cell and Gene Therapy, Baylor College of Medicine, Texas Children's Hospital, Houston, Texas 77030, USA.
Molecular Therapy (Impact Factor: 6.23). 07/2009; 17(10):1779-87. DOI: 10.1038/mt.2009.133
Source: PubMed


Human epidermal growth factor receptor 2 (HER2) is expressed by the majority of human osteosarcomas and is a risk factor for poor outcome. Unlike breast cancer, osteosarcoma cells express HER2 at too low, a level for patients to benefit from HER2 monoclonal antibodies. We reasoned that this limitation might be overcome by genetically modifying T cells with HER2-specific chimeric antigen receptors (CARs), because even a low frequency of receptor engagement could be sufficient to induce effector cell killing of the tumor. HER2-specific T cells were generated by retroviral transduction with a HER2-specific CAR containing a CD28.zeta signaling domain. HER2-specific T cells recognized HER2-positive osteosarcoma cells as judged by their ability to proliferate, produce immunostimulatory T helper 1 cytokines, and kill HER2-positive osteosarcoma cell lines in vitro. The adoptive transfer of HER2-specific T cells caused regression of established osteosarcoma xenografts in locoregional as well as metastatic mouse models. In contrast, delivery of nontransduced (NT) T cells did not change the tumor growth pattern. Genetic modification of T cells with CARs specific for target antigens, expressed at too low a level to be effectively recognized by monoclonal antibodies, may allow immunotherapy to be more broadly applicable for human cancer therapy.

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Available from: Laszlo Perlaky, Oct 06, 2015
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    • "The construction of the HER2-specific CAR (pSFG.FRP5.CD28.ζ) has been previously described.26,27,28 The construction of the IL-13Rα2-specific CAR is described in detail elsewhere (Krebs et al. submitted for publication). "
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    ABSTRACT: Preclinical and early clinical studies have demonstrated that chimeric antigen receptor (CAR)-redirected T cells are highly promising in cancer therapy. We observed that targeting HER2 in a Glioblastoma cell line results in the emergence of HER2-null tumor cells that maintain the expression of non-targeted tumor associated antigens (TAA). Combinational targeting of these TAAs could thus offset this escape mechanism. We studied the single-cell co-expression patterns of HER2, IL-13Rα2 and EphA2 in primary Glioblastoma samples using multicolor flow cytometry and immunofluorescence, and applied a binomial routine to the permutations of antigen expression and the related odds of complete tumor elimination. This mathematical model demonstrated that co-targeting HER2 and IL-13Rα2 could maximally expand the therapeutic reach of the T cell product in all primary tumors studied. Targeting a third antigen did not predict an added advantage in the tumor cohort studied. We thus generated bispecific T cell products from healthy donors and from GBM patients by pooling T cells individually expressing HER2 and IL-13Rα2-specific CARs and by making individual T cells to co-express both molecules. Both HER2/IL-13Rα2-bispecific T cell products offset antigen escape, producing enhanced effector activity in vitro immunoassays (against autologous glioma cells in the case of GBM patient products) and in an orthotopic xenogeneic murine model. Further, T cells co-expressing HER2- and IL-13Rα2-CARs exhibited accentuated yet antigen-dependent downstream signaling and a particularly enhanced antitumor activity.Molecular Therapy (2013); doi:10.1038/mt.2013.185.
    Molecular Therapy 08/2013; 21(11). DOI:10.1038/mt.2013.185 · 6.23 Impact Factor
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    • "In our study, we have also demonstrated that B7-H3 expression is negatively associated with the intensity of infiltrating CD8+ T lymphocytes in tumor sites, suggesting that one of the most important contributions of B7-H3 expression in this malignancy is the impairment of host T cell-mediated immunity by negatively regulating T lymphocyte infiltration. Decreased numbers of tumor-infiltrating immune cells, including T, B and natural killer (NK) cells, have been shown to correlate with decreased survival times in human osteosarcoma [35], [36]. Moreover, a recent study has shown that the ratio of CD8+ T cells to regulatory T cells is inversely associated with the outcome of osteosarcoma in dogs [37]. "
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    ABSTRACT: B7-H3 is a member of the B7-family of co-stimulatory molecules, which has been shown to be broadly expressed in various tumor tissues, and which plays an important role in adaptive immune responses. The role of B7-H3 in osteosarcoma, however, remains unknown. In this study we used immunohistochemistry to analyze B7-H3 expression in 61 primary osteosarcoma tissues with case-matched adjacent normal tissues, and 37 osteochondroma and 20 bone fibrous dysplasia tissues. B7-H3 expression was expressed in 91.8% (56/61) of the osteosarcoma lesions, and the intensity of B7-H3 expression in osteosarcoma was significantly increased compared with adjacent normal tissues, osteochondroma and bone fibrous dysplasia tissues (p<0.001). Patients with high tumor B7-H3 levels had a significantly shorter survival time and recurrence time than patients with low tumor B7-H3 levels (p<0.001). Moreover, tumor B7-H3 expression inversely correlated with the number of tumor-infiltrating CD8(+) T cells (p<0.05). In vitro, increasing expression of B7-H3 promotes osteosarcoma cell invasion, at least in part by upregulating matrix metalloproteinase-2 (MMP-2). In conclusion, our study provides the first evidence of B7-H3 expression in osteosarcoma cells as a potential mechanism controlling tumor immunity and invasive malignancy, and which is correlated with patients' survival and metastasis.
    PLoS ONE 08/2013; 8(8):e70689. DOI:10.1371/journal.pone.0070689 · 3.23 Impact Factor
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    • "Furthermore, in the context of modest expression of target antigens, antibodies exhibit limited efficacy.31,39 T cells expressing antibody-derived CARs have been shown to overcome such limitations.31,40 "
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    ABSTRACT: Targeted T cells are emerging as effective non-toxic therapies for cancer. Multiple elements, however, contribute to the overall pathogenesis of cancer through both distinct and redundant mechanisms. Hence, targeting multiple cancer-specific markers simultaneously could result in better therapeutic efficacy. We created a functional chimeric antigen receptor-the TanCAR, a novel artificial molecule that mediates bispecific activation and targeting of T cells. We demonstrate the feasibility of cumulative integration of structure and docking simulation data using computational tools to interrogate the design and predict the functionality of such a complex bispecific molecule. Our prototype TanCAR induced distinct T cell reactivity against each of two tumor restricted antigens, and produced synergistic enhancement of effector functions when both antigens were simultaneously encountered. Furthermore, the TanCAR preserved the cytolytic ability of T cells upon loss of one of the target molecules and better controlled established experimental tumors by recognition of both targets in an animal disease model. This proof-of-concept approach can be used to increase the specificity of effector cells for malignant versus normal target cells, to offset antigen escape or to allow for targeting the tumor and its microenvironment.Molecular Therapy-Nucleic Acids (2013) 2, e105; doi:10.1038/mtna.2013.32; published online 9 July 2013.
    Molecular Therapy - Nucleic Acids 07/2013; 2(7):e105. DOI:10.1038/mtna.2013.32 · 4.51 Impact Factor
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