Sara Ghorashian

Molecular remission of infant B-ALL after infusion of universal TALEN gene-edited CAR T cells

Universal gene-edited CAR19 T cells eliminate infant leukemia. CAR sharing Chimeric antigen receptor (CAR) T cells can be very effective in treating acute lymphocytic leukemia. Unfortunately, these therapeutic cells have to be custom-made for each patient, and this is not always feasible, especially for patients who do not have sufficient healthy T cells. Qasim et al. demonstrate that there may be another option for these patients. By using gene editing to simultaneously introduce the CAR and disrupt TCR and CD52 in T cells, the authors generated functional CAR T cells that could evade host immunity for use in unmatched recipients. These “off-the-shelf” CAR T cells were then used to treat two infants with relapsed refractory acute lymphocytic leukemia and bridge them to allogeneic stem cell transplantation. Autologous T cells engineered to express chimeric antigen receptor against the B cell antigen CD19 (CAR19) are achieving marked leukemic remissions in early-phase trials but can be difficult to manufacture, especially in infants or heavily treated patients. We generated universal CAR19 (UCART19) T cells by lentiviral transduction of non–human leukocyte antigen–matched donor cells and simultaneous transcription activator-like effector nuclease (TALEN)–mediated gene editing of T cell receptor α chain and CD52 gene loci. Two infants with relapsed refractory CD19+ B cell acute lymphoblastic leukemia received lymphodepleting chemotherapy and anti-CD52 serotherapy, followed by a single-dose infusion of UCART19 cells. Molecular remissions were achieved within 28 days in both infants, and UCART19 cells persisted until conditioning ahead of successful allogeneic stem cell transplantation. This bridge-to-transplantation strategy demonstrates the therapeutic potential of gene-editing technology.

Nonhematopoietic antigen blocks memory programming of alloreactive CD8 + T cells and drives their eventual exhaustion in mouse models of bone marrow transplantation

Allogeneic blood or BM transplantation (BMT) is the most commonly applied form of adoptive cellular therapy for cancer. In this context, the ability of donor T cells to respond to recipient antigens is coopted to generate graft-versus-tumor (GVT) responses. The major reason for treatment failure is tumor recurrence, which is linked to the eventual loss of functional, host-specific CTLs. In this study, we have explored the role of recipient antigen expression by nonhematopoietic cells in the failure to sustain effective CTL immunity. Using clinically relevant models, we found that nonhematopoietic antigen severely disrupts the formation of donor CD8+ T cell memory at 2 distinct levels that operate in the early and late phases of the response. First, initial and direct encounters between donor CD8+ T cells and nonhematopoietic cells blocked the programming of memory precursors essential for establishing recall immunity. Second, surviving CD8+ T cells became functionally exhausted with heightened expression of the coinhibitory receptor programmed death-1 (PD-1). These 2 factors acted together to induce even more profound failure in long-term immunosurveillance. Crucially, the functions of exhausted CD8+ T cells could be partially restored by late in vivo blockade of the interaction between PD-1 and its ligand, PD-L1, without induction of graft-versus-host disease, suggestive of a potential clinical strategy to prevent or treat relapse following allogeneic BMT.

OX40- and CD27-Mediated Costimulation Synergizes with Anti–PD-L1 Blockade by Forcing Exhausted CD8+ T Cells To Exit Quiescence

Exhaustion of chronically stimulated CD8+ T cells is a significant obstacle to immune control of chronic infections or tumors. Although coinhibitory checkpoint blockade with anti–programmed death ligand 1 (PD-L1) Ab can restore functions to exhausted T cell populations, recovery is often incomplete and dependent upon the pool size of a quiescent T-bethigh subset that expresses lower levels of PD-1. In a model in which unhelped, HY-specific CD8+ T cells gradually lose function following transfer to male bone marrow transplantation recipients, we have explored the effect of shifting the balance away from coinhibition and toward costimulation by combining anti–PD-L1 with agonistic Abs to the TNFR superfamily members, OX40 and CD27. Several weeks following T cell transfer, both agonistic Abs, but especially anti-CD27, demonstrated synergy with anti–PD-L1 by enhancing CD8+ T cell proliferation and effector cytokine generation. Anti-CD27 and anti–PD-L1 synergized by downregulating the expression of multiple quiescence-related genes concomitant with a reduced frequency of T-bethigh cells within the exhausted population. However, in the presence of persistent Ag, the CD8+ T cell response was not sustained and the overall size of the effector cytokine-producing pool eventually contracted to levels below that of controls. Thus, CD27-mediated costimulation can synergize with coinhibitory checkpoint blockade to switch off molecular programs for quiescence in exhausted T cell populations, but at the expense of losing precursor cells required to maintain a response.

CD19 chimeric antigen receptor T cell therapy for haematological malignancies.

T cells can be redirected to recognize tumour antigens by genetic modification to express a chimeric antigen receptor (CAR). These consist of antibody‐derived antigen‐binding regions linked to T cell signalling elements. CD19 is an ideal target because it is expressed on most B cell malignancies as well as normal B cells but not on other cell types, restricting any ‘on target, off tumour’ toxicity to B cell depletion. Recent clinical studies involving CD19 CAR‐directed T cells have shown unprecedented responses in a range of B cell malignancies, even in patients with chemorefractory relapse. Durable responses have been achieved, although the persistence of modified T cells may be limited. This therapy is not without toxicity, however. Cytokine release syndrome and neurotoxicity appear to be frequent but are treatable and reversible. CAR T cell therapy holds the promise of a tailored cellular therapy, which can form memory and be adapted to the tumour microenvironment. This review will provide a perspective on the currently available data, as well as on future developments in the field.

Human MHC Class I-restricted high avidity CD4+ T cells generated by co-transfer of TCR and CD8 mediate efficient tumor rejection in vivo

In this study, we generated human MHC Class I-restricted CD4+ T cells specific for Epstein-Barr virus (EBV) and cytomegalovirus (CMV), two herpesviridae associated with lymphoma, nasopharyngeal carcinoma and medulloblastoma, respectively. Retroviral transfer of virus-specific, HLA-A2-restricted TCR-coding genes generated CD4+ T cells that recognized HLA-A2/peptide multimers and produced cytokines when stimulated with MHC Class II-deficient cells presenting the relevant viral peptides in the context of HLA-A2. Peptide titration revealed that CD4+ T cells had a 10-fold lower avidity than CD8+ T cells expressing the same TCR. The impaired avidity of CD4+ T cells was corrected by simultaneously transferring TCR- and CD8-coding genes. The CD8 co-receptor did not alter the cytokine signature of CD4+ T cells, which remained distinct from that of CD8+ T cells. Using the xenogeneic NOD/SCID mouse model, we demonstrated that human CD4+ T cells expressing a specific TCR and CD8 can confer efficient protection against the growth of tumors expressing the EBV or CMV antigens recognized by the TCR. In summary, we describe a robust approach for generating therapeutic CD4+ T cells capable of providing MHC Class I-restricted immunity against MHC Class II-negative tumors in vivo.

CD8 T Cell Tolerance to a Tumor-Associated Self-Antigen Is Reversed by CD4 T Cells Engineered To Express the Same T Cell Receptor

Ag receptors used for cancer immunotherapy are often directed against tumor-associated Ags also expressed in normal tissues. Targeting of such Ags can result in unwanted autoimmune attack of normal tissues or induction of tolerance in therapeutic T cells. We used a murine model to study the phenotype and function of T cells redirected against the murine double minute protein 2 (MDM2), a tumor-associated Ag that shows low expression in many normal tissues. Transfer of MDM2-TCR–engineered T cells into bone marrow chimeric mice revealed that Ag recognition in hematopoietic tissues maintained T cell function, whereas presentation of MDM2 in nonhematopoietic tissues caused reduced effector function. TCR-engineered CD8+ T cells underwent rapid turnover, downmodulated CD8 expression, and lost cytotoxic function. We found that MDM2-TCR–engineered CD4+ T cells provided help and restored cytotoxic function of CD8+ T cells bearing the same TCR. Although the introduction of the CD8 coreceptor enhanced the ability of CD4+ T cells to recognize MDM2 in vitro, the improved self-antigen recognition abolished their ability to provide helper function in vivo. The data indicate that the same class I–restricted TCR responsible for Ag recognition and tolerance induction in CD8+ T cells can, in the absence of the CD8 coreceptor, elicit CD4 T cell help and partially reverse tolerance. Thus MHC class I–restricted CD4+ T cells may enhance the efficacy of therapeutic TCR-engineered CD8+ T cells and can be readily generated with the same TCR.

Improving TCR Gene Therapy for Treatment of Haematological Malignancies

Adoptive immunotherapy using TCR gene modified T cells may allow separation of beneficial Graft versus tumour responses from harmful GvHD. Improvements to this include methods to generate high avidity or high affinity TCR, improvements in vector design and reduction in mispairing. Following adoptive transfer, TCR transduced T cells must be able to survive and persist in vivo to give most effective antitumour responses. Central memory or naive T cells have both been shown to be more effective than effector cells at expanding and persisting in vivo. Lymphodepletion may enhance persistence of transferred T cell populations. TCR gene transfer can be used to redirect CD4 helper T cells, and these could be used in combination with CD8+ tumour specific T cells to provide help for the antitumour response. Antigen specific T regulatory T cells can also be generated by TCR gene transfer and could be used to suppress unwanted alloresponses.

T cell gene-engineering to enhance GVT and suppress GVHD

Gene-engineering of T cells offers the possibility of uniformly changing the characteristics of their immune responses. The ability to direct polyclonal T cells to a single antigenic specificity is a powerful tool with which to probe anti-tumour immune responses, and in turn ask which tumour antigens represent the best targets for immune therapy. The intracellular components of TCR signalling pathways can also be manipulated to optimise anti-tumour responses. Such manipulated T cells may in the future represent an important adjunctive therapy alongside allogeneic haematopoietic stem cell transplantation (ASCT), in order to specifically boost a graft versus leukaemia (GVL) effect. In addition, the ability to confer a suppressive phenotype to CD4 cells or to engineer a susceptibility gene into effector cells provides new therapeutic avenues for graft versus host disease (GVHD), the most challenging adverse effect arising post ASCT. Within this review, mechanisms of GVL and GVHD are discussed and we consider how they may be separated through T cell gene engineering. In addition, we highlight recently investigated safety issues which will impact the future clinical application of gene-manipulated immune cells.

Redirection to the bone marrow improves T cell persistence and antitumor functions

A key predictor for the success of gene-modified T cell therapies for cancer is the persistence of transferred cells in the patient. The propensity of less differentiated memory T cells to expand and survive efficiently has therefore made them attractive candidates for clinical application. We hypothesized that redirecting T cells to specialized niches in the BM that support memory differentiation would confer increased therapeutic efficacy. We show that overexpression of chemokine receptor CXCR4 in CD8+ T cells (TCXCR4) enhanced their migration toward vascular-associated CXCL12+ cells in the BM and increased their local engraftment. Increased access of TCXCR4 to the BM microenvironment induced IL-15–dependent homeostatic expansion and promoted the differentiation of memory precursor–like cells with low expression of programmed death-1, resistance to apoptosis, and a heightened capacity to generate polyfunctional cytokine-producing effector cells. Following transfer to lymphoma-bearing mice, TCXCR4 showed a greater capacity for effector expansion and better tumor protection, the latter being independent of changes in trafficking to the tumor bed or local out-competition of regulatory T cells. Thus, redirected homing of T cells to the BM confers increased memory differentiation and antitumor immunity, suggesting an innovative solution to increase the persistence and functions of therapeutic T cells.

Open access? Widening access to chimeric antigen receptor (CAR) therapy for ALL

T cells that are genetically modified to express chimeric antigen receptors (CARs) specific for CD19 show great promise for the treatment of relapsed/refractory acute lymphoblastic leukemia (ALL). The first U.S. Food and Drug Administration approval of a cellular cancer therapy in 2017, Novartiss CD19-targeting CAR T-cell product Kymriah™ within the context of relapsed/refractory pediatric ALL, followed rapidly by approval of Kites Yescarta™ and, more recently, Kymriah™ for diffuse large B-cell indications in adults, highlights the pace of progress made in this field. In this review, we will consider the latest evidence from CAR T-cell therapy for B-lineage ALL. We discuss the barriers to CAR T-cell therapy for ALL patients and give a perspective on the strategy we have taken to date to widen access to CAR T-cell therapy for UK pediatric patients with high-risk ALL.