Assaf Marcus

Recognition of tumors by the innate immune system and natural killer cells.

In recent years, roles of the immune system in immune surveillance of cancer have been explored using a variety of approaches. The roles of the adaptive immune system have been a major emphasis, but increasing evidence supports a role for innate immune effector cells such as natural killer (NK) cells in tumor surveillance. Here, we discuss some of the evidence for roles in tumor surveillance of innate immune cells. In particular, we focus on NK cells and other immune cells that express germline-encoded receptors, often labeled NK receptors. The impact of these receptors and the cells that express them on tumor suppression is summarized. We discuss in detail some of the pathways and events in tumor cells that induce or upregulate cell-surface expression of the ligands for these receptors, and the logic of how those pathways serve to identify malignant, or potentially malignant cells. How tumors often evade tumor suppression mediated by innate killer cells is another major subject of the review. We end with a discussion on some of the implications of the various findings with respect to possible therapeutic approaches.

Immunosurveillance and immunotherapy of tumors by innate immune cells.

Increasing evidence supports a role for innate immune effector cells in tumor surveillance. Natural killer (NK) cells and myeloid cells represent the two main subsets of innate immune cells possessing efficient but quite different tumor suppressive abilities. Here, we describe the germline-encoded NK cell receptors that play a role in suppressing tumor development and describe briefly the cellular pathways leading to the upregulation of their ligands in tumor cells. We also describe mechanisms underlying the elimination of tumor cells by macrophages and a recently characterized mechanism dedicated to sensing cytosolic DNA that is implicated in antitumor immune responses.

Evidence for Natural Killer Cell Memory

Natural killer (NK) cells are generally considered to be part of the innate immune system. Over the past few years, however, evidence has accumulated suggesting that NK cells have certain features that are characteristic of the adaptive immune system. NK cells reportedly respond in an antigen-specific manner to a variety of small molecules and certain viruses, and mediate enhanced responses to these antigens upon secondary exposure. In infections with mouse cytomegalovirus (MCMV), MCMV-specific NK cells undergo clonal expansion, and display increased effector function after the resolution of the infection. In addition, inflammatory conditions resulting from exposure to certain cytokines seem to promote prolonged effector function in NK cells in an antigen-non-specific fashion. Taken together, these studies reveal new aspects of NK biology, and suggest that NK cells, like T and B cells, may carry out memory responses and may also exhibit greater capacity to distinguish antigens than was previously recognized.

Allogeneic chimeric antigen receptor-modified cells for adoptive cell therapy of cancer

Introduction: Chimeric antigen (or antibody) receptors (CAR) are fusion proteins typically combining an antibody-derived targeting fragment with signaling domains capable of activating immune cells. Recent clinical trials have shown the tremendous potential of adoptive cell transfer (ACT) of autologous T cells engineered to express a CD19-specific CAR targeting B-cell malignancies. Building on this approach, ACT therapies employing allogeneic CAR-expressing cytotoxic cells are now being explored. Areas covered: The basic principles of CAR-ACT are introduced. The potential benefits as well as problems of using allogeneic CAR-modified cells against tumor antigens are discussed. Various approaches to allogeneic CAR therapy are presented, including donor leukocyte infusion, CAR-redirected γδ T cells and natural killer cells, strategies to avoid graft-versus-host disease, modulation of lymphocyte migration, and exploitation of graft-versus-host reactivity. Expert opinion: CAR-modified allogeneic cells have the potential to act as universal effector cells, which can be administered to any patient regardless of MHC type. Such universal effector cells could be used as an ‘off-the-shelf’ cell-mediated treatment for cancer.

Dysregulated cellular functions and cell stress pathways provide critical cues for activating and targeting natural killer cells to transformed and infected cells

Natural killer (NK) cells recognize and kill cancer cells and infected cells by engaging cell surface ligands that are induced preferentially or exclusively on these cells. These ligands are recognized by activating receptors on NK cells, such as NKG2D. In addition to activation by cell surface ligands, the acquisition of optimal effector activity by NK cells is driven in vivo by cytokines and other signals. This review addresses a developing theme in NK cell biology: that NK‐activating ligands on cells, and the provision of cytokines and other signals that drive high effector function in NK cells, are driven by abnormalities that arise from transformation or the infected state. The pathways include genomic damage, which causes self DNA to be exposed in the cytosol of affected cells, where it activates the DNA sensor cGAS. The resulting signaling induces NKG2D ligands and also mobilizes NK cell activation. Other key pathways that regulate NKG2D ligands include PI‐3 kinase activation, histone acetylation, and the integrated stress response. This review summarizes the roles of these pathways and their relevance in both viral infections and cancer.

Tumor-specific allogeneic cells for cancer therapy

Adoptive cell transfer (ACT) therapy involves transfer of therapeutic lymphocytes to patients mostly for the treatment of cancer and viral infections. One modality to generate therapeutic lymphocytes is to genetically engineer them to express a chimeric antigen receptor (CAR) capable of recognizing the desired target. Current ACT approaches employ the patients own (syngeneic) lymphocytes, which is both economically and technically challenging. Using foreign (allogeneic) lymphocytes in ACT is problematic because of the severe immunological reaction that occurs between genetically mismatched individuals. However, recently our group has developed a protocol, which allows for safe and effective ACT therapy in a murine model of metastatic disease using allogeneic T cells redirected with a human EGFR2/neuregulin (Her2/neu)-specific CAR. Mild preconditioning of the recipient delayed the rejection of the allogeneic donor T cells such that they had enough time to destroy the tumor, but not enough to cause significant damage to the host. By modulating lymphocyte migration using FTY720, we were actually able to exploit the allogeneic anti-host reaction in order to augment therapeutic benefit while concurrently improving the safety of the treatment. Therefore, we suggest that CAR-based allogeneic ACT therapy could be universally used as a safe and potent ‘off-the-shelf’ treatment for cancer.

Adoptive Cell Therapy of Systemic Metastases Using erbB-2-Specific T Cells Redirected with a Chimeric Antibody-Based Receptor

Immunotherapy of cancer using adoptive cell transfer combined with the advent of gene-engineering technologies has become an appealing option for a wide spectrum of cancers. In contrast to T cell receptor-based approaches, which are MHC restricted, chimeric antibody-based receptors (CAR), pioneered by our group, allow for a broader application, which are not restricted to individual tissue types. Here, we describe our studies using T cells redirected with CAR specific to the erbB-2 growth factor proto-oncogene as a common tumor target antigen. In a murine model for lung metastasis, we demonstrate that under defined conditions, CAR-expressing T cells (T-bodies) can eliminate systemic lung metastases, which are generally felt to be incurable. The antitumor effect of systemically injected T-bodies was augmented by using increased injected cell doses and repeated administration cycles as well as by pre-vaccination of the tumor-bearing mice. Most importantly, we were able to establish a protocol enabling the use of MHC mismatched T-bodies in a safe and effective manner. We found that a single dose of allogeneic T-bodies under mild immunosuppressive conditions could cure metastases, demonstrating the efficacy of this modality against disseminated disease. These results provide a proof of principle for using allogeneic erbB-2-specific T-bodies as a standard treatment of erbB-2-expressing tumors.

Tumor-Derived cGAMP Triggers a STING-Mediated Interferon Response in Non-tumor Cells to Activate the NK Cell Response

Graphical Abstract Figure. No caption available. SUMMARY Detection of cytosolic DNA by the enzyme cGAS triggers the production of cGAMP, a second messenger that binds and activates the adaptor protein STING, which leads to interferon (IFN) production. Here, we found that in vivo natural killer (NK) cell killing of tumor cells, but not of normal cells, depends on STING expression in non‐tumor cells. Experiments using transplantable tumor models in STING‐ and cGAS‐deficient mice revealed that cGAS expression by tumor cells was critical for tumor rejection by NK cells. In contrast, cGAS expression by host cells was dispensable, suggesting that tumor‐derived cGAMP is transferred to non‐tumor cells, where it activates STING. cGAMP administration triggered STING activation and IFN‐&bgr; production in myeloid cells and B cells but not NK cells. Our results reveal that the anti‐tumor response of NK cells critically depends on the cytosolic DNA sensing pathway, similar to its role in defense against pathogens, and identify tumor‐derived cGAMP as a major determinant of tumor immunogenicity with implications for cancer immunotherapy. HIGHLIGHTSSting−/− mice, but not Cgas−/− mice, fail to mount optimal NK cell anti‐tumor responsesCgas−/− tumor cells are defective in inducing anti‐tumor responses by NK cellscGAS is constitutively active in tumor cells but not in untransformed cellscGAMP is transferred from tumor cells to other cells in the TME to activate STING &NA; Marcus et al. find that cGAMP produced by tumor cells triggers the activation of the STING pathway in immune cells within the tumor microenvironment. This leads to interferon production by these cells, which in turn activates NK cell anti‐tumor immunity.