Our Science – Restifo Website
Nicholas P. Restifo, M.D.
His work focuses on the iterative development of T cell-based adoptive immunotherapies by simultaneously exploring novel aspects of mouse and human T cell immunobiology.
Designing Potent New Cancer Immunotherapies
Our goal is to design new immunotherapies for patients with advanced cancer. Our strategy is based on the use of animal models and human in vitro assays to test hypotheses. We then translate the most promising of these therapies into human clinical trials, which often generate new questions to be tested experimentally. The process is an iterative one that involves close collaboration with basic researchers, biotech scientists and experimental clinicians.
Our work focuses on immunotherapy based on the adoptive transfer of naturally-occurring and gene-engineered tumor-specific T cells. We explore what T cells 'see' in tumor masses, what T cells must 'do' to trigger tumor cell death, and how T cells experience maturational changes that dramatically affect their phenotypes and functions. We also seek methods to best combine anti-tumor T cells with other therapies in order to treat patients with cancer.
Tumor-infiltrating lymphocytes can eradicate tumor masses.
Treatment of patients with cells that have been expanded ex vivo is called adoptive cell transfer (ACT). Cells that are infused back into a patient after expansion can traffic to the tumor and mediate its destruction. The addition of 'preparative lymphodepletion,' the temporary ablation of a cancer patient's immune system can be accomplished using chemotherapy alone or in combination with total-body irradiation, and is associated with enhanced persistence of the transferred T cells. The combination of a lymphodepleting preparative regimen, ACT and administration of the T cell growth factor interleukin-2 (IL-2) can lead to prolonged tumor eradication in patients with metastatic melanoma or other tumor histologies, including leukemias and synovial cell sarcomas, who have exhausted other treatment options.
How T cells target tumors.
Naturally-occurring or gene-engineered tumor-specific MHC class I-restricted T cells are, theoretically, capable of directly recognizing many types of tumor cells. Tumor immunologists now have a deeper understanding of the molecular structures that T cells can recognize. These include: unaltered tissue-differentiation antigens on tumors; Anti-tumor T cells can see the products of mutated genes; viral antigens; products of developmental genes turned on by epigenetic changes in cancers; and antigens expressed by non-transformed tumor vasculature and stroma.
Emerging findings from both mouse studies and clinical trials indicate that intrinsic properties related to the differentiation state of the adoptively transferred T cell populations are crucial to the success of ACT-based approaches.
CD8+ T cells trigger tumor rejection in both mice and humans and can be categorized into distinct memory subsets based on their differentiation states. The available data suggests that CD8+ T cells follow a progressive pathway of differentiation from naive T cells, into central memory (TCM) T cell and effector memory (TEM) T cell populations. The extent of differentiation is determined by the strength of TCR signal and the cytokine environment that the T cell encounters during antigen-specific activation. Experiments using TCR transgenic mice have revealed that CD8+ T cells that are stimulated multiple times with specific antigen and IL-2 show decreased survival and proliferation compared with cells that are stimulated only once or not at all. Although these CD8+ T cells acquired the ability to lyse target cells and to produce IFN-γ, qualities thought to be important in their anti-tumor efficacy, their actual anti-tumor efficacy declined following adoptive transfer. This decline in the function of anti-tumor T cells has been linked with a variety of factors, including cell intrinsic counter-regulation. T cell differentiation is associated with increased production of granzymes and reactive oxygen species, but also with the loss of the ability to produce IL-2, to home to lymph nodes, and to resist apoptotic death, leading to the hypothesis that the differentiation state of CD8+ T cells is inversely related to their capacity to proliferate and persist.
In terms of efficacy in ACT for cancer, naive T cells have been shown to be more effective than memory cells, and within memory pools TCM show increased anti-tumor activity compared with TEM cells in mice. It is clinically relevant that 'younger' T cells can be gene-engineered with high efficiency, although it remains unclear whether the increased efficacy of these T cells depends upon them being separated from fully differentiated effector cells prior to adoptive transfer.
Recently, we have found in a distinct population of CD8+ T memory stem (TSCM) cells. These cells express high levels of Sca-1, Bcl2 and IL-2Rβ and were shown in mice to have superior anti-tumor properties compared with other memory T cell subsets. In humans, TSCM resemble naive T cells, in that they are CD45RA+ CD45RO-. They express the IL-7 receptor alpha-chain, which can facilitate their survival131, and high levels of molecules that facilitate homing to lymph nodes, such as CD62L and CCR7. In addition, TSCM cells express co-stimulatory receptors, including CD27 and CD28, which may facilitate their proliferative capacities, and high levels of CD95 and IL-2Rβ.
TSCM express a gene program that enables them to proliferate extensively and they can further differentiate into TCM and TEM cells. Importantly, human TSCM show increased anti-tumor activity compared with TCM and TEM cells, suggesting that they will be more effective for ACT in patients with cancer.
Arresting or reversing T cell differentiation might be desirable in the setting of ACT for cancer under some circumstances. Future ACT-based immunotherapies might rely more on the phenotype, telomere length, IL-2-producing capacity and TCR affinity of T cells, when determining which cells should be transferred.
The differentiation state of CD8+ T cells found in TILs isolated from patients is not yet clear. It is possible that in some patients, available tumor-specific T cells are already terminally differentiated into TEM cells. In addition, the culture conditions currently used for preparing TILs for ACT include two weeks or more of culture with high doses of IL-2, CD3-specific antibody and, in some cases, irradiated allogeneic stimulator cells may cause further differentiation of isolated TILs.
It is not clear if limiting these powerful differentiation factors will improve the efficacy of TILs, or if any improvements in T cell anti-tumor functions (if any such improvement occurs) will compensate for the reduced TIL viability that might result from restricting these factors. However, because IL-2 promotes T cell effector differentiation and susceptibility to apoptosis, it will be important to explore whether reducing IL-2 concentrations in TIL culture conditions may be beneficial for ACT purposes. Other cytokines worth investigating are IL-15, which promotes TCM cell differentiation and IL-21, which has been shown to 'arrest' T cell differentiation and render 'younger' cells at the end of culture, as well as pharmacologic modulators of the Wnt signalling pathway.
CD4+ T cells can also efficiently promote tumor rejection, in part through their ability to secrete IL-2 and recruit and sustain anti-tumor CD8+ cells. CD4+ T cells can also alter the function of APCs, especially DCs. In addition, CD4+ T cells do not merely enhance CD8+ T cell function, but they can play a more direct role in tumor elimination.
The roles that CD4+ T cells play in the anti-tumor immune response crucially depends on their polarization, which is determined by their expression of key transcription factors and has been reviewed elsewhere. As already mentioned, TReg cells have potent immunoinhibitory functions and may contribute to the immunosuppressive tumor microenvironment. The role of TH2 cells in the anti-tumor immune response remains unclear, but TH1 cells can mediate anti-tumor responses in mouse models, in part through their production of IFN-γ.
New evidence suggests that adoptively transferred TH17 cells can promote long-lived anti-tumor immunity. Although IL-17 itself has been linked to carcinogenesis, it is clear that 'type 17' polarization conditions can also augment the anti-tumor functions of CD8+ T cells. TH17 cells are capable of inducing efficient tumor rejection, but only if they are able to differentiate into effector T cells with Th1-like properties. Interestingly, TH17 cells show similarities to stem cells in terms of their gene expression profiles, resistance to apoptosis, and in their capacities of self-renewal and multipotency.
In summary, ACT after lymphodepletion has emerged as a promising advance in cancer immunotherapy. Emerging data from preclinical and clinical studies have increased our understanding of the mechanisms underlying successful immunotherapies and helped us to identify the most effective T cell populations. In addition, gene engineering has expanded the potential target population that could benefit from ACT-based immunotherapies.
ACT-based therapies are not FDA-approved and are only available in a handful of locations worldwide although this approach to cancer treatment is growing as an experimental treatment option. The immune system is capable of sterilizing immunity and the induction of long-term durable complete responses that are probably curative. The use of adoptive T cell-based therapies to eradicate cancer is at a rare nexus of basic immunology and clinically meaningful therapy.
This page was last updated on 2/22/2013.