Memoirs of a Reincarnated T Cell

Abstract

In two studies published in this issue of Cell Stem Cell, Nishimura et al. (2013) and Vizcardo et al. (2013) reprogram mature, antigen-specific cytotoxic T cells into induced pluripotent stem cells (iPSCs). The antigen-specific iPSCs can be redifferentiated into “rejuvenated” proliferative T cells and have broad applications for adoptive immunotherapy.

One method to confirm the identity of the reincarnated Dalai Lama involves placing several artifacts, only a few of which belonged to the previous Dalai Lama, in front of a candidate child. If the child recognizes an artifact which belonged to the previous Dalai Lama, this is interpreted as a sign that the child is indeed the reincarnated Lama. In the recent work by Nishimura (Nishimura et al., 2013) and Vizcardo (Vizcardo et al., 2013) presented in this issue of Cell Stem Cell, we are also witnesses to a majestic reincarnation of sorts. Both groups reprogrammed human antigen-specific CD8⁺ mature T cells into induced pluripotent stem cells (iPSCs). Furthermore, they showed that redifferentiation of these cells produced CD8⁺ T cells that recognize their original cognate antigen and have features of rejuvenated cells so critically needed to improve the efficacy of T-cell-based therapies for the treatment of cancer and viral-associated diseases (Fearon, 2007).

Needless to say, the regeneration of antigen-specific T cells from iPSCs derived from mature CD8⁺ T cells provides yet another example of the reach of Yamanaka’s discovery that induction of four transcription factors—Oct4, Sox2, Klf4, and c-Myc—can reprogram well-differentiated somatic cells to pluripotency (Takahashi and Yamanaka, 2006). The field of regenerative medicine and the stem cell community continue to profit from Yamanaka’s finding that was fittingly celebrated in Stockholm last month. Amidst immense enthusiasm over the therapeutic potential of iPSCs, however, remain obstacles in translating this discovery to improve the care of patients (Wu and Hochedlinger, 2011). By focusing on the reprogramming of the immune system in adoptive cell transfer immunotherapy, the work of Nishimura et al. and Vizcardo et al. may provide an opportunity to hasten the bench-to-bedside translation of iPSC technology.

Adoptive cell transfer immunotherapy (ACT) of T cells shows great promise in treating patients with advanced cancer, graft-versus-host disease, and viral-associated diseases. As a therapy to treat cancer, for example, ACT relies on the natural ability of CD8⁺ T cells (called tumor-infiltrating lymphocytes, or TILs) to attack tumors (Restifo et al., 2012). The dogged persistence of cancer, however, often results in CD8⁺ T cells that are both exhausted and senescent (Klebanoff et al., 2006). With ACT, the tumor-specific TILs are harvested from a patient’s tumor, expanded ex vivo with the immunostimulatory cytokine IL-2, and infused back into the same patient (Figure 1). Remarkably, up to 20% of patients with advanced melanoma have had complete and durable regression of their metastatic disease (Rosenberg, 2011). In trying to understand why this cohort of patients had such an extraordinary response, it was found that transfer of less-differentiated memory or stem-cell-like CD8⁺ T cells correlated with a more robust eradication of tumor. Therefore, there is a critical need to improve the efficacy of ACT by developing methods such as iPSC technology to expand the pool of memory or stem-cell-like CD8⁺ T cells (Gattinoni et al., 2011).

Figure 1. Reprogramming an Exhausted Immune System for Treatment of Cancer.

(A) Generalized model of T cell differentiation. An immune response is initiated when naive CD8⁺ T cells (T_N) are activated by their cognate antigen to undergo clonal expansion and differentiation into subsets characterized here by a reciprocally graded capacity for self-renewal and cytolytic effector function. Although eradication of tumor ultimately requires robust cytolytic function of terminally differentiated effector (T_EFF) cells, therapeutic efficacy of ACT is improved by transfer of less-differentiated cells because of their ability for self renewal and survival and their ability to generate T_EFF progeny.

(B) Rejuvenating T cells with Yamanaka factors. Adoptive T-cell-based immunotherapy (ACT) for cancer involves isolating CD8⁺ lymphocytes from a surgically excised tumor and expanding the cells ex vivo with T cell growth factor IL-2. Lymphocytes are subsequently infused back into the patient to attack the remaining tumor sites. Lymphocytes isolated from the tumor generally have a terminally differentiated effector (T_EFF) or effector memory (T_EM) phenotype, and the IL-2 driven expansion results in further differentiation. There is considerable evidence that transfer of less-differentiated lymphocytes such as stem-cell memory T cells (T_SCM) or central memory T cells (T_CM) results in substantially greater antitumor immunity. Therefore, rejuvenating lymphocytes to a less-differentiated state using induced pluripotent stem cell (iPSC) technology shows great promise in substantially improving the efficacy of ACT. Reprogramming with Yamanaka factors and redifferentiation with Delta-like 1 expressing OP9 (OP9/DLL1) stromal cells rejuvenates lymphocytes and preserves less-differentiated T_SCM and T_CM CD8⁺ T cell subsets that may result in improved eradication of tumor when transferred into cancer patients.

In their study, Vizcardo et al. attempt to address this need by obtaining TILs from one of our patients suffering from metastatic melanoma at the Surgery Branch of the National Cancer Institute. The TILs have an antigen specificity for MART-1, which is commonly expressed on melanoma tumors. The MART-1-specific TILs were transduced with Yamanaka factors and reprogrammed to iPSCs as evidenced by ESC-like morphology, capacity for teratoma formation, endogenous expression of OCT3/4, SOX2, KLF4, and c-MYC, and disappearance of T cell markers CD3 and CD8. Subsequent coculture with Delta-like 1 expressing OP9 feeder cells resulted in redifferentation to CD8⁺ T cells (Schmitt and Zúñiga-Pflücker, 2002). When cultured with antigen presenting cells and challenged with the cognate antigen MART-1, reprogrammed cells produced interferon gamma, demonstrating functional integrity.

Although Vizcardo and colleagues show that reincarnated CD8⁺ T cells have a memory for their original tumor antigen, it is not clear if reincarnated cells are indeed rejuvenated. That is, does the reprogramming process confer greater replicative capacity, multipotency, and antitumor immunity that is characteristic of memory or stem-cell-like T cells? We know that TILs are generally characterized by a state of exhaustion and terminal differentiation. Thus, the lingering question is whether the reincarnated cells have preserved a memory or stem-cell-like phenotype so critical to improving the efficacy of ACT for cancer or viral infection.

Nishimura et al. provide additional insight into whether terminally differentiated CD8⁺ T cells can be rejuvenated through reprogramming by showing that redifferentiated CD8⁺ T cells have elongated telomeres and greater proliferative capacity. Using an HIV-1-epitope specific clone of CD8⁺ T cells, they demonstrate reprogramming through an iPSC intermediate into functional CD8⁺ T cells capable of cytolytic activity and interferon gamma release with recognition of their cognate peptide. Importantly, the cells also show enhanced expression of CCR7, CD27, and CD28, revealing characteristic stem-cell-like traits compared to the original T cell clone from which the cells were derived (Gattinoni et al., 2011). In fact, reincarnated CD8⁺ T cells have even longer telomeres than their original counterparts, perhaps imparted by their sojourn through the iPSC state where telomerase activity runs hot. The authors duly recognize that the clinical translation of rejuvenating cytotoxic T cells to treat patients with HIV is complicated, and with good reason, but we feel this remains a remarkable demonstration of rejuvenated T cells and is an important first step in broadening the therapeutic application of ACT to treat maladies whose common denominator is a terminally differentiated and exhausted immune response.

Taken together, the work of the two groups led by Vizcardo and Nishimura demonstrate that antigen-experienced CD8⁺ T cells can be reincarnated through an iPSC intermediate into CD8⁺ T cells that preserve memory for their original epitope, and moreover, develop a rejuvenated memory phenotype upon redifferentiation. This has important clinical implications for ACT because it provides proof of concept that terminally differentiated lymphocytes can be reincarnated from an exhausted state to a memory phenotype that may have demonstrably superior antitumor immunity (Figure 1). We look forward to further preclinical work investigating the efficacy of reprogrammed T cells in eradicating tumors and chronic viral diseases and feel this may be a particularly fruitful means to hasten the bench-to-bedside translation of iPSC technology and regenerative medicine for the treatment of patients with viral-associated diseases and cancer.