The role of chimeric antigen receptor T cells targeting more than one antigen in the treatment of B-cell malignancies.

Several products containing chimeric antigen receptor T cells targeting CD19 (CART19) have been approved for the treatment of patients with relapsed/refractory non-Hodgkin's lymphoma (NHL) and acute lymphoblastic leukaemia (ALL). Despite very impressive response rates, a significant percentage of patients experience disease relapse and die of progressive disease. A major cause of CART19 failure is loss or downregulation of CD19 expression in tumour cells, which has prompted a myriad of novel strategies aimed at targeting more than one antigen (e.g. CD19 and CD20 or CD22). Dual targeting can the accomplished through co-administration of two separate products, co-transduction with two different vectors, bicistronic cassettes or tandem receptors. In this manuscript, we review the pros and cons of each strategy and the clinical results obtained so far.

Academic challenges on advanced therapy medicinal products' development: a regulatory perspective.

Advanced therapy medicinal products (ATMPs) are becoming the new kid on the block for the treatment of a variety of indications with promising results. Despite the academic contribution to the basic and clinical research of ATMPs, undertaking a full product development process is extraordinarily challenging and demanding for academic institutions. Meeting regulatory requirements is probably the most challenging aspect of academic development, considering the limited experience and resources compared with pharmaceutical companies. This review aims to outline the key aspects to be considered when developing novel ATMPs from an academic perspective, based on the results of our own experience and interaction with the Spanish Agency of Medicines and Medical Devices (AEMPS) and European Medicine Agency (EMA) related to a number of academic ATMP initiatives carried out at our center during the last 5 years. Emphasis is placed on understanding the regulatory requirements during the early phases of the drug development process, particularly for the preparation of a Clinical Trial Application. Academic centers usually lack expertise in product-related documentation (such as the Investigational Medicinal Product Dossier), and therefore, early interaction with regulators is crucial to understand their requirements and receive guidance to comply with them. Insights are shared on managing quality, nonclinical, clinical, and risk and benefit documentation, based on our own experience and challenges. This review aims to empower academic and clinical settings by providing crucial regulatory knowledge to smooth the regulatory journey of ATMPs.

NK cells enhance CAR-T cell antitumor efficacy by enhancing immune/tumor cells cluster formation and improving CAR-T cell fitness.

BACKGROUND: Chimeric antigen receptor (CAR)-T cell immunotherapy has modified the concept of treatment in hematological malignancies. In comparison with pediatric patients, where responses are maintained over many years, older patients, such as those with non-Hodgkin's lymphoma (NHL) and multiple myeloma (MM), present lower persistence of CAR-T cells that might be due to decreased fitness of T cells acquired with aging. Moreover, cord blood derived-NK cells (CB-NKs) and CAR-NK cells derived from CB-NK can be used 'off-the-shelf' as immune cells with antitumor properties for the treatment of cancer patients. However, to date, clinical studies have only demonstrated the safety of these therapies but not optimal efficacy. To confront the shortcomings of each therapy, we devised a novel approach consisting of simultaneous (CAR-)NK cell and CAR-T cell administration. In this setting, NK cells demonstrate an important immunoregulation of T cells that could be exploited to enhance the efficacy of CAR-T cells. METHODS: A combinatorial treatment based on either CAR-T and CAR-NK cells or CB-NK and CAR-T cells in two models of NHL and MM was performed. Antitumor efficacy was analyzed in vitro and in vivo, and parameters related to early activation, exhaustion and senescence of T cells were analyzed. RESULTS: We show that CAR-NK cells derived from CB-NK are only effective at high doses (high E:T ratio) and that their activity rapidly decreases over time in comparison with CAR-T cells. In comparison and to exploit the potential of 'off-the-shelf' CB-NK, we demonstrate that a low number of CB-NK in the CAR-T cell product promotes an early activation of CAR-T cells and their migration to MM cells leading to enhanced anti-MM efficacy. Moreover, cytokines related to CRS development were not increased, and importantly, CB-NK enhanced the fitness of both CARpos and CARneg T cells, promoting lower levels of exhaustion and senescence. CONCLUSION: This study demonstrates a relevant immunoregulatory role of CB-NK collaborating with CAR-T cells to enhance their antitumor activity. A novel and different approach to consider in CAR-T cell immunotherapy studies is presented here with the goal to enhance the efficacy of the treatment.

IL-15 Enhances the Persistence and Function of BCMA-Targeting CAR-T Cells Compared to IL-2 or IL-15/IL-7 by Limiting CAR-T Cell Dysfunction and Differentiation.

Chimeric antigen receptor (CAR)-T cell immunotherapy has revolutionized the treatment of B-lymphoid malignancies. For multiple myeloma (MM), B-cell maturation antigen (BCMA)-targeted CAR-T cells have achieved outstanding complete response rates, but unfortunately, patients often relapse within a year of receiving the therapy. Increased persistence and reduced dysfunction are crucial features that enhance the durability of CAR-T cell responses. One of the factors that influence CAR-T cell in vivo longevity and loss of function, but which has not yet been extensively studied for BCMA-directed CAR-T cells, are the cytokines used during their production. We here compared the impact of IL-2, IL-15 and a combination of IL-15/IL-7 on the phenotype and function of ARI2h, an academic BCMA-directed CAR-T cell that is currently being administered to MM patients. For this study, flow cytometry, in vitro cytotoxicity assays and analysis of cytokine release were performed. In addition, ARI2h cells expanded with IL-2, IL-15, or IL-15/IL-7 were injected into MM tumor-bearing mice to assess their in vivo efficacy. We demonstrated that each of the cytokine conditions was suitable for the expansion of ARI2h cells, with clear in vitro activity. Strikingly, however, IL-15-produced ARI2h cells had improved in vivo efficacy and persistence. When explored further, it was found that IL-15 drove a less-differentiated ARI2h phenotype, ameliorated parameters related to CAR-T cell dysfunction, and lowered the release of cytokines potentially involved in cytokine release syndrome and MM progression. Moreover, we observed that IL-15 was less potent in inducing T cell senescence and DNA damage accumulation, both of which may contribute to an unfavorable CAR-T cell phenotype. These findings show the superiority of IL-15 to IL-2 and IL-15/IL-7 in the quality of anti-BCMA CAR-T cells, particularly their efficacy and persistence, and as such, could improve the duration of responses if applied to the clinical production of CAR-T cells for patients.

Natural Killer Cells in Immunotherapy: Are We Nearly There?

Natural killer (NK) cells are potent anti-tumor and anti-microbial cells of our innate immune system. They are equipped with a vast array of receptors that recognize tumor cells and other pathogens. The innate immune activity of NK cells develops faster than the adaptive one performed by T cells, and studies suggest an important immunoregulatory role for each population against the other. The association, observed in acute myeloid leukemia patients receiving haploidentical killer-immunoglobulin-like-receptor-mismatched NK cells, with induction of complete remission was the determinant to begin an increasing number of clinical studies administering NK cells for the treatment of cancer patients. Unfortunately, even though transfused NK cells demonstrated safety, their observed efficacy was poor. In recent years, novel studies have emerged, combining NK cells with other immunotherapeutic agents, such as monoclonal antibodies, which might improve clinical efficacy. Moreover, genetically-modified NK cells aimed at arming NK cells with better efficacy and persistence have appeared as another option. Here, we review novel pre-clinical and clinical studies published in the last five years administering NK cells as a monotherapy and combined with other agents, and we also review chimeric antigen receptor-modified NK cells for the treatment of cancer patients. We then describe studies regarding the role of NK cells as anti-microbial effectors, as lessons that we could learn and apply in immunotherapy applications of NK cells; these studies highlight an important immunoregulatory role performed between T cells and NK cells that should be considered when designing immunotherapeutic strategies. Lastly, we highlight novel strategies that could be combined with NK cell immunotherapy to improve their targeting, activity, and persistence.

Senescence in the Development and Response to Cancer with Immunotherapy: A Double-Edged Sword.

Cellular senescence was first described as a physiological tumor cell suppressor mechanism that leads to cell growth arrest with production of the senescence-associated secretory phenotype known as SASP. The main role of SASP in physiological conditions is to attract immune cells to clear senescent cells avoiding tumor development. However, senescence can be damage-associated and, depending on the nature of these stimuli, additional types of senescence have been described. In the context of cancer, damage-associated senescence has been described as a consequence of chemotherapy treatments that were initially thought of as a tumor suppressor mechanism. However, in certain contexts, senescence after chemotherapy can promote cancer progression, especially when immune cells become senescent and cannot clear senescent tumor cells. Moreover, aging itself leads to continuous inflammaging and immunosenescence which are responsible for rewiring immune cells to become defective in their functionality. Here, we define different types of senescence, pathways that activate them, and functions of SASP in these events. Additionally, we describe the role of senescence in cancer and its treatments, including how aging and chemotherapy contribute to senescence in tumor cells, before focusing on immune cell senescence and its role in cancer. Finally, we discuss potential therapeutic interventions to reverse cell senescence.