No abstract

Adoptive transfer of T cell receptor (TCR)-transduced T cells may be an attractive strategy to target both hematological malignancies and solid tumors. By introducing a TCR, large numbers of T cells with defined antigen (Ag) specificity can be obtained. However, by introduction of a TCR, mixed TCR dimers can be formed. Besides the decrease in TCR expression of the introduced and endogenous TCR, these mixed TCR dimers could harbor potentially harmful specificities. In this study, we demonstrate that introduction of TCRs resulted in formation of neoreactive mixed TCR dimers, composed of the introduced TCR chains pairing with either the endogenous TCR α or β chain. Neoreactivities observed were HLA class I or class II restricted. Most neoreactive mixed TCR dimers were allo-HLA reactive; however, neoreactive mixed TCR dimers with autoreactive activity were also observed. We demonstrate that inclusion of an extra disulfide bond between the constant domains of the introduced TCR markedly reduced neoreactivity, whereas enhanced effectiveness of the introduced TCR was observed. In conclusion, TCR transfer results in the formation of neoreactive mixed TCR dimers with the potential to generate off-target effects, underlining the importance of searching for techniques to facilitate preferential pairing.

Purpose: In human leukocyte antigen (HLA)–matched stem cell transplantation (SCT), it has been shown that beneficial immune response mediating graft-versus-tumor (GVT) responses can be separated from graft-versus-host disease (GVHD) immune responses. In this study, we investigated whether it would be possible to dissect the beneficial immune response of allo-HLA–reactive T cells with potent antitumor reactivity from GVHD-inducing T cells present in the detrimental immune response after HLA-mismatched SCT. Experimental Design: The presence of specific tumor-reactive T cells in the allo-HLA repertoire was analyzed at the time of severe GVHD after HLA-mismatched SCT, using tetramers composed of different tumor-associated antigens (TAA). Results: High-avidity allo-HLA-restricted T cells specific for the TAA preferentially expressed antigen on melanomas (PRAME) were identified that exerted highly single-peptide–specific reactivity. The T cells recognized multiple different tumor cell lines and leukemic cells, whereas no reactivity against a large panel of nonmalignant cells was observed. These T cells, however, also exerted low reactivity against mature dendritic cells (DC) and kidney epithelial cells, which was shown to be because of low PRAME expression. Conclusions: On the basis of potential beneficial specificity and high reactivity, the T-cell receptors of these PRAME-specific T cells may be effective tools for adoptive T-cell therapy. Clinical studies have to determine the significance of the reactivity observed against mature DCs and kidney epithelial cells. Clin Cancer Res; 17(17); 5615–25. ©2011 AACR.

Retroviral transfer of T-cell receptors (TCR) to peripheral blood-derived T cells generates large numbers of T cells with the same antigen specificity, potentially useful for adoptive immunotherapy. One drawback of this procedure is the formation of mixed TCR dimers with unknown specificities due to pairing of endogenous and introduced TCR chains. We investigated whether ;D T cells can be an alternative effector population for TCR gene transfer because the ;DTCR is not able to form dimers with the ABTCR. Peripheral bloodderived ;D T cells were transduced with human leukocyte antigen (HLA) class I-or HLA class II-restricted minor histocompatibility antigen (mHag) or virus-specific TCRs. Because most ;D T cells do not express CD4 and CD8, we subsequently transferred these coreceptors. The TCR-transduced ;D T cells exerted high levels of antigen-specific cytotoxicity and produced IFN-; and IL-4, particularly in the presence of the relevant coreceptor. ;D T cells transferred with a TCR specific for the hematopoiesis-specific mHag HA-2 in combination with CD8 displayed high antileukemic reactivity against HA-2-expressing leukemic cells. These data show that transfer of ABTCRs to ;D T cells generated potent effector cells for immunotherapy of leukemia, without the expression of potentially hazardous mixed TCR dimers. (Cancer Res 2006; 66(6): 3331-7) IntroductionCellular immunotherapy is a promising strategy for the treatment of cancer (1). However, adoptive transfer of sufficient numbers of antigen-specific T cells requires complex isolation methods and laborious and time-consuming tissue culture procedures. An alternative method to obtain large numbers of T cells with a defined antigen specificity is the retroviral transfer of a T-cell receptor (TCR). Because T-cell specificity is exclusively determined by the TCR, T-cell specificity can be functionally transferred to other T lymphocytes by retroviral TCR gene transfer. We and others have shown that transfer of human leukocyte antigen (HLA) class I-and HLA class II-restricted TCRs to CD8 + and CD4 + T cells, respectively, generated T cells with converted antigen-specific cytolytic activity and cytokine production (2-10).The potential in vivo efficacy of TCR-transferred T cells was shown in mouse models (8,9). The TCR-transferred T cells were activated in vivo, homed to effector sites, and contributed to tumor clearance.A potential disadvantage of TCR gene transfer to other ah T cells is the formation of mixed TCR dimers. Chains of the introduced TCR can pair with the endogenous TCR chains naturally expressed by the TCR-transferred T cells. The specificity of these mixed TCR dimers is unknown and, therefore, autoreactivity cannot be excluded. To limit the number of T cells with different TCR chains and thus the chance to generate autoreactive T cells, T cells with defined antigen specificity and, therefore, with a limited TCR repertoire can be selected as host cells for TCR gene transfer. We previously showed the reprogramming of cytomegalovirus (CMV)-specifi...

Non-small cell lung cancer (NSCLC) is the second most prevalent type of cancer. With the current treatment regimens, the mortality rate remains high. Therefore, better therapeutic approaches are necessary. NSCLCs generally possess many genetic mutations and are well infiltrated by T cells (TIL), making TIL therapy an attractive option. Here we show that T cells from treatment naive, stage I-IVa NSCLC tumors can effectively be isolated and expanded, with similar efficiency as from normal lung tissue. Importantly, 76% (13/17) of tested TIL products isolated from NSCLC lesions exhibited clear reactivity against primary tumor digests, with 0.5%-30% of T cells producing the inflammatory cytokine Interferon (IFN)-γ. Both CD4+ and CD8+ T cells displayed tumor reactivity. The cytokine production correlated well with CD137 and CD40L expression. Furthermore, almost half (7/17) of the TIL products contained polyfunctional T cells that produced Tumor Necrosis Factor (TNF)-α and/or IL-2 in addition to IFN-γ, a hallmark of effective immune responses. Tumor-reactivity in the TIL products correlated with high percentages of CD103+CD69+CD8+ T cell infiltrates in the tumor lesions, with PD-1hiCD4+ T cells, and with FoxP3+CD25+CD4+ regulatory T cell infiltrates, suggesting that the composition of T cell infiltrates may predict the level of tumor reactivity. In conclusion, the effective generation of tumor-reactive and polyfunctional TIL products implies that TIL therapy will be a successful treatment regimen for NSCLC patients.