Subsequent Malignancies after CD19-Targeted Chimeric Antigen Receptor T cells in Patients with Lymphoma.

Chimeric antigen receptor (CAR) T cells are an established treatment for B cell non-Hodgkin lymphomas (B-NHL). With the remarkable success in improving survival, understanding the late effects of CAR T cell therapy is becoming more relevant. The aim of this study is to determine the incidence of subsequent malignancies in adult patients with B-NHL. We retrospectively studied 355 patients from two different medical centers treated with four different CAR T cell products from 2016 to 2022. The overall cumulative incidence for subsequent malignancies at 36 months was 14% (95% CI: 9.2%, 19%). Subsequent malignancies were grouped into three primary categories: solid tumor, hematologic malignancy, and dermatologic malignancy with cumulative incidences at 36 months of 6.1% (95% CI: 3.1%-10%), 4.5% (95% CI: 2.1%-8.1%) and 4.2% (95% CI: 2.1%-7.5%) respectively. Notably, no cases of T cell malignancies were observed. In univariable analysis, increasing age was associated with higher risk for subsequent malignancy. While the overall benefits of CAR T products continue to outweigh their potential risks, more studies and longer follow ups are needed to further demonstrate the risks, patterns, and molecular pathways that lead to the development of subsequent malignancies.

Engineering strategies to safely drive CAR T-cells into the future.

Chimeric antigen receptor (CAR) T-cell therapy has proven a breakthrough in cancer treatment in the last decade, giving unprecedented results against hematological malignancies. All approved CAR T-cell products, as well as many being assessed in clinical trials, are generated using viral vectors to deploy the exogenous genetic material into T-cells. Viral vectors have a long-standing clinical history in gene delivery, and thus underwent iterations of optimization to improve their efficiency and safety. Nonetheless, their capacity to integrate semi-randomly into the host genome makes them potentially oncogenic via insertional mutagenesis and dysregulation of key cellular genes. Secondary cancers following CAR T-cell administration appear to be a rare adverse event. However several cases documented in the last few years put the spotlight on this issue, which might have been underestimated so far, given the relatively recent deployment of CAR T-cell therapies. Furthermore, the initial successes obtained in hematological malignancies have not yet been replicated in solid tumors. It is now clear that further enhancements are needed to allow CAR T-cells to increase long-term persistence, overcome exhaustion and cope with the immunosuppressive tumor microenvironment. To this aim, a variety of genomic engineering strategies are under evaluation, most relying on CRISPR/Cas9 or other gene editing technologies. These approaches are liable to introduce unintended, irreversible genomic alterations in the product cells. In the first part of this review, we will discuss the viral and non-viral approaches used for the generation of CAR T-cells, whereas in the second part we will focus on gene editing and non-gene editing T-cell engineering, with particular regard to advantages, limitations, and safety. Finally, we will critically analyze the different gene deployment and genomic engineering combinations, delineating strategies with a superior safety profile for the production of next-generation CAR T-cell.

A bibliometric and knowledge-map study on the treatment of hematological malignancies with CAR-T cells from 2012 to 2023.

Recently, CAR-T cell therapy in hematological malignancies has received extensive attention. The objective of this study is to gain a comprehensive understanding of the current research status, development trends, research hotspots, and emerging topics pertaining to CAR-T cells in the treatment of hematological malignancies. Articles pertaining to CAR-T cell therapy for hematological malignancies from the years 2012 to 2023 were obtained and assessed from the Web of Science Core Collection (WoSCC). A bibliometric approach was employed to conduct a scientific, comprehensive, and objective quantitative analysis, as well as a visual analysis, of this particular research domain. A comprehensive analysis was conducted on a corpus of 3643 articles, which were collaboratively authored by 72 countries and various research institutions. CAR-T cell research in treating hematological malignancies shows an increasing trend each year. Notably, the study identified the countries and institutions displaying the highest level of activity, the journals with the most citations and output, as well as the authors who garnered the highest frequency of citations and co-citations. Furthermore, the analysis successfully identified the research hotspots and highlighted six emerging topics within this domain. This study conducted a comprehensive exploration and analysis of the research status, development trends, research hotspots, and emerging topics about CAR-T cells in the treatment of hematological malignancies from 2012 to 2023. The findings of this study will serve as a valuable reference and guide for researchers seeking to delve deeper into this field and determine the future direction of their research.

Genetic ablation of adhesion ligands mitigates rejection of allogeneic cellular immunotherapies.

Allogeneic cellular immunotherapies hold promise for broad clinical implementation but face limitations due to potential rejection of donor cells by the host immune system. Silencing of beta-2 microglobulin (B2M) expression is commonly employed to evade T cell-mediated rejection by the host, although the absence of B2M is expected to trigger missing-self responses by host natural killer (NK) cells. Here, we demonstrate that genetic deletion of the adhesion ligands CD54 and CD58 in B2M-deficient chimeric antigen receptor (CAR) T cells and multi-edited induced pluripotent stem cell (iPSC)-derived CAR NK cells reduces their susceptibility to rejection by host NK cells in vitro and in vivo. The absence of adhesion ligands limits rejection in a unidirectional manner in B2M-deficient and B2M-sufficient settings without affecting the antitumor functionality of the engineered donor cells. Thus, these data suggest that genetic ablation of adhesion ligands effectively alleviates rejection by host immune cells, facilitating the implementation of universal immunotherapy.

CAR-T Cells in Chronic Lymphocytic Leukemia.

The treatment outcomes of patients with chronic lymphocytic leukemia (CLL) have considerably improved with the introduction of targeted therapies based on Bruton kinase inhibitors (BTKIs), venetoclax, and anti-CD20 monoclonal antibodies. However, despite these consistent improvements, patients who become resistant to these agents have poor outcomes and need new and more efficacious therapeutic strategies. Among these new treatments, a potentially curative approach consists of the use of chimeric antigen receptor T (CAR-T) cell therapy, which achieved remarkable success in various B-cell malignancies, including B-cell Non-Hodgkin Lymphomas (NHLs) and B-acute lymphoblastic Leukemia (ALL). However, although CAR-T cells were initially used for the treatment of CLL, their efficacy in CLL patients was lower than in other B-cell malignancies. This review analyses possible mechanisms of these failures, highlighting some recent developments that could offer the perspective of the incorporation of CAR-T cells in treatment protocols for relapsed/refractory CLL patients.

Bispecific CAR-T cells targeting FAP and GPC3 have the potential to treat hepatocellular carcinoma.

Chimeric antigen receptor (CAR) T cell therapy has demonstrated robust efficacy against hematological malignancies, but there are still some challenges regarding treating solid tumors, including tumor heterogeneity, antigen escape, and an immunosuppressive microenvironment. Here, we found that SNU398, a hepatocellular carcinoma (HCC) cell line, exhibited high expression levels of fibroblast activation protein (FAP) and Glypican 3 (GPC3), which were negatively correlated with patient prognosis. The HepG2 HCC cell line highly expressed GPC3, while the SNU387 cell line exhibited high expression of FAP. Thus, we developed bispecific CAR-T cells to simultaneously target FAP and GPC3 to address tumor heterogeneity in HCC. The anti-FAP-GPC3 bispecific CAR-T cells could recognize and be activated by FAP or GPC3 expressed by tumor cells. Compared with anti-FAP CAR-T cells or anti-GPC3 CAR-T cells, bispecific CAR-T cells achieved more robust activity against tumor cells expressing FAP and GPC3 in vitro. The anti-FAP-GPC3 bispecific CAR-T cells also exhibited superior antitumor efficacy and significantly prolonged the survival of mice compared with single-target CAR-T cells in vivo. Overall, the use of anti-FAP-GPC3 bispecific CAR-T cells is a promising treatment approach to reduce tumor recurrence caused by tumor antigen heterogeneity.

Severe anaphylaxis after chimeric antigen receptor T-cell injection: a case report.

Anaphylactic reactions at the time of chimeric antigen receptor T (CAR-T) cell infusion are adverse events that have not been reported in pivotal clinical trials or in real-world series. We report the case of patient with severe anaphylaxis with cardiac arrest after tisagenlecleucel injection for Diffuse Large B cell Lymphoma, who recovered after resuscitation and intensive care treatment; we also conducted a Food and Drug Administration Adverse Event Reporting System database analysis and found several cases of severe anaphlyaxis after CAR-T cell injection. Although not reported in pivotal CAR-T cell studies, anaphylaxis can occur after CAR-T cell injection, highlighting the need to include anaphylaxis as a possible side effect in future studies.

Advancing CAR T-cell therapy for chronic lymphocytic leukemia: exploring resistance mechanisms and the innovative strategies to overcome them.

Chimeric antigen receptor (CAR) T-cell therapy has ushered in substantial advancements in the management of various B-cell malignancies. However, its integration into chronic lymphocytic leukemia (CLL) treatment has been challenging, attributed largely to the development of very effective chemo-free alternatives. Additionally, CAR T-cell responses in CLL have not been as high as in other B-cell lymphomas or leukemias. However, a critical void exists in therapeutic options for patients with high-risk diseases who are resistant to the current CLL therapies, underscoring the urgency for adoptive immunotherapies in these patients. The diminished CAR T-cell efficacy within CLL can be traced to factors such as compromised T-cell fitness due to persistent antigenic stimulation inherent to CLL. Resistance mechanisms encompass tumor-related factors like antigen escape, CAR T-cell-intrinsic factors like T-cell exhaustion, and a suppressive tumor microenvironment (TME). New strategies to combat CAR T-cell resistance include the concurrent administration of therapies that augment CAR T-cell endurance and function, as well as the engineering of novel CAR T-cells targeting different antigens. Moreover, the concept of "armored" CAR T-cells, armed with transgenic modulators to modify both CAR T-cell function and the tumor milieu, is gaining traction. Beyond this, the development of readily available, allogeneic CAR T-cells and natural killer (NK) cells presents a promising countermeasure to innate T-cell defects in CLL patients. In this review, we explore the role of CAR T-cell therapy in CLL, the intricate tapestry of resistance mechanisms, and the pioneering methods studied to overcome resistance.

Immunotherapeutic Strategies Targeting Breast Cancer Stem Cells.

Breast cancer is the most commonly diagnosed cancer in women and is a leading cause of cancer death in women worldwide. Despite the implementation of multiple treatment options, including immunotherapy, breast cancer treatment remains a challenge. In this review, we aim to summarize present challenges in breast cancer immunotherapy and recent advancements in overcoming treatment resistance. We elaborate on the inhibition of signaling cascades, such as the Notch, Hedgehog, Hippo, and WNT signaling pathways, which regulate the self-renewal and differentiation of breast cancer stem cells and, consequently, disease progression and survival. Cancer stem cells represent a rare population of cancer cells, likely originating from non-malignant stem or progenitor cells, with the ability to evade immune surveillance and develop resistance to immunotherapeutic treatments. We also discuss the interactions between breast cancer stem cells and the immune system, including potential agents targeting breast cancer stem cell-associated signaling pathways, and provide an overview of the emerging approaches to breast cancer stem cell-targeted immunotherapy. Finally, we consider the development of breast cancer vaccines and adoptive cellular therapies, which train the immune system to recognize tumor-associated antigens, for eliciting T cell-mediated responses to target breast cancer stem cells.

Generating universal anti-CD19 CAR T cells with a defined memory phenotype by CRISPR/Cas9 editing and safety evaluation of the transcriptome.

Introduction: Chimeric antigen receptor-expressing T cells (CAR T cells) have revolutionized cancer treatment, particularly in B cell malignancies. However, the use of autologous T cells for CAR T therapy presents several limitations, including high costs, variable efficacy, and adverse effects linked to cell phenotype. Methods: To overcome these challenges, we developed a strategy to generate universal and safe anti-CD19 CAR T cells with a defined memory phenotype. Our approach utilizes CRISPR/Cas9 technology to target and eliminate the B2M and TRAC genes, reducing graft-versus-host and host-versus-graft responses. Additionally, we selected less differentiated T cells to improve the stability and persistence of the universal CAR T cells. The safety of this method was assessed using our CRISPRroots transcriptome analysis pipeline, which ensures successful gene knockout and the absence of unintended off-target effects on gene expression or transcriptome sequence. Results: In vitro experiments demonstrated the successful generation of functional universal CAR T cells. These cells exhibited potent lytic activity against tumor cells and a reduced cytokine secretion profile. The CRISPRroots analysis confirmed effective gene knockout and no unintended off-target effects, validating it as a pioneering tool for on/off-target and transcriptome analysis in genome editing experiments. Discussion: Our findings establish a robust pipeline for manufacturing safe, universal CAR T cells with a favorable memory phenotype. This approach has the potential to address the current limitations of autologous CAR T cell therapy, offering a more stable and persistent treatment option with reduced adverse effects. The use of CRISPRroots enhances the reliability and safety of gene editing in the development of CAR T cell therapies. Conclusion: We have developed a potent and reliable method for producing universal CAR T cells with a defined memory phenotype, demonstrating both efficacy and safety in vitro. This innovative approach could significantly improve the therapeutic landscape for patients with B cell malignancies.

Augmenting the landscape of chimeric antigen receptor T-cell therapy.

INTRODUCTION: The inception of recombinant DNA technology and live cell genomic alteration have paved the path for the excellence of cell and gene therapies and often provided the first curative treatment for many indications. The approval of the first Chimeric Antigen Receptor (CAR) T-cell therapy was one of the breakthrough innovations that became the headline in 2017. Currently, the therapy is primarily restricted to a few nations, and the market is growing at a CAGR (current annual growth rate) of 11.6% (2022-2032), as opposed to the established bio-therapeutic market at a CAGR of 15.9% (2023-2030). The limited technology democratization is attributed to its autologous nature, lack of awareness, therapy inclusion criteria, high infrastructure cost, trained personnel, complex manufacturing processes, regulatory challenges, recurrence of the disease, and long-term follow-ups. AREAS COVERED: This review discusses the vision and strategies focusing on the CAR T-cell therapy democratization with mitigation plans. Further, it also covers the strategies to leverage the mRNA-based CAR T platform for building an ecosystem to ensure availability, accessibility, and affordability to the community. EXPERT OPINION: mRNA-guided CAR T cell therapy is a rapidly growing area wherein a collaborative approach among the stakeholders is needed for its success.

Fully equipped CARs to address tumor heterogeneity, enhance safety, and improve the functionality of cellular immunotherapies.

Although adoptive transfer of chimeric antigen receptor (CAR)-engineered T cells has achieved unprecedented response rates in patients with certain hematological malignancies, this therapeutic modality is still far from fulfilling its remarkable potential, especially in the context of solid cancers. Antigen escape variants, off-tumor destruction of healthy tissues expressing tumor-associated antigens (TAAs), poor CAR-T cell persistence, and the occurrence of functional exhaustion represent some of the most prominent hurdles that limit CAR-T cell ability to induce long-lasting remissions with a tolerable adverse effect profile. In this review, we summarize the main approaches that have been developed to face such bottlenecks, including the adapter CAR (AdCAR) system, Boolean-logic gating, epitope editing, the modulation of cell-intrinsic signaling pathways, and the incorporation of safety switches to precisely control CAR-T cell activation. We also discuss the most pressing issues pertaining to the selection of co-stimulatory domains, with a focus on strategies aimed at promoting CAR-T cell persistence and optimal antitumor functionality.

Small molecule-regulated switches to provide functional control of CAR T cells within the patient.

INTRODUCTION: CAR T cells have generated great excitement due to their remarkable clinical response rates in selected hematologic malignancies. However, these engineered immune cells are living drugs which are hard to control after administration. AREAS COVERED: We discuss small molecule-regulated switch systems which can potentially be used to control CAR T cell function within the patient, as well as the most important obstacles in the CAR T cell field, which might be overcome with those switch systems. EXPERT OPINION: There is an urgent need to develop advanced switch systems. Once available, we expect that they will open up new avenues for future CAR T cell generations.

Pediatric Cancer Immunotherapy and Potential for Impact on Fertility: A Need for Evidence-Based Guidance.

The use of immunotherapies for the treatment of cancer in children, adolescents, and young adults has become common. As the use of immunotherapy has expanded, including in earlier lines of therapy, it has become evident that several aspects of how these immunotherapies impact longer-term outcomes among survivors are understudied. Traditional cancer therapies like alkylating and platin agents carry the greatest risk of infertility, but little is known about the impact of novel immunotherapies on fertility. This topic is of great interest to patients, patient advocates, and clinicians. In this article, we review immunotherapeutic agents used to treat childhood and young adult cancers and discuss potential mechanisms by which they may impact fertility based on the known interplay between the immune system and reproductive organs. We highlight the relative paucity of high-quality literature examining these late effects. We discuss interventions to optimize fertility preservation (FP) for our patients. Conducting longitudinal, collaborative, and prospective research on the fertility outcomes of pediatric and young adult patients with cancer who receive immunotherapy is critical to learn how to effectively counsel our patients on long-term fertility outcomes and indications for FP procedures. Collection of patient-level data will be necessary to draft evidence-based guidelines on which providers can make therapy recommendations.

Photo-metallo-immunotherapy: Fabricating Chromium-Based Nanocomposites to Enhance CAR-T Cell Infiltration and Cytotoxicity against Solid Tumors.

The infiltration and cytotoxicity of chimeric antigen receptor (CAR)-T cells are crucial for effective elimination of solid tumors. While metallo-immunotherapy is a promising strategy that can activate the antitumor immunity, its role in promoting CAR-T cell therapy remains elusive. The first single-element nanomaterial based on chromium nanoparticles (Cr NPs) for cancer photo-metallo-immunotherapy has been reported previously. Herein, an extended study using biodegradable polydopamine as a versatile carrier for these nanoparticles, enabling synergistic CAR-T cell therapy, is reported. The results show that these nanocomposites with or without further encapsulation of the anticancer drug alpelisib can promote the CAR-T cell migration and antitumor effect. Upon irradiation with near-infrared light, they caused mild hyperthermia that can "warm" the "cold" tumor microenvironment (TME). The administration of B7-H3 CAR-T cells to NOD severe combined immunodeficiency gamma mice bearing a human hepatoma or PIK3CA-mutated breast tumor can significantly inhibit the tumor growth after the induction of tumor hyperthermia by the nanocomposites and promote the secretion of serum cytokines, including IL-2, IFN-γ, and TNF-α. The trivalent Cr3+ ions, which are the major degradation product of these nanocomposites, can increase the CXCL13 and CCL3 chemokine expressions to generate tertiary lymphoid structures (TLSs) in the tumor tissues, facilitating the CAR-T cell infiltration.

Consolidation with First and Second Allogeneic Transplants in Adults with Relapsed/Refractory B-ALL Following Response to CD19CAR T Cell Therapy.

CD19-targeted chimeric antigen receptor T cell (CAR-T) therapy has led to unprecedented rates of complete remission (CR) in children and adults with relapsed/refractory (r/r) B-cell acute lymphoblastic leukemia (B-ALL), yet the majority of adults relapse after initial response. One proposed method to extend the durability of remission in adults following response to CAR-T therapy is consolidation with allogeneic hematopoietic cell transplantation (alloHCT). Considering the limited published data for the utility of post CAR-T therapy consolidative alloHCT in r/r B-ALL, especially data related to patients receiving a second alloHCT, we sought to describe outcomes of patients with r/r B-ALL at our institution who received their first or second alloHCT following response to CAR-T therapy. We performed a retrospective analysis of adult patients with r/r B-ALL who responded to either investigational or standard of care (SOC) CD19-targeted CAR-T therapy and underwent consolidation with alloHCT while in CR without interim therapy. We identified 45 patients, of whom 26 (58%) and 19 (42%) received their first and second alloHCT as consolidation post CAR-T therapy, respectively. The median age was 31 years (range: 19-67) and 31 (69%) patients were Hispanic. Ph-like was the most common genetic subtype and comprised over half of cases (53%; n = 24). The median number of prior therapies pre-transplant was 5 (range: 2-7), and disease status at the time of alloHCT was CR1, CR2 or ≥CR3 in 7 (16%), 22 (49%) and 16 (35%) patients, respectively. The median time from CAR-T therapy until alloHCT was 93 (range: 42-262) days. The conditioning regimen was radiation-based myeloablative (MAC) in 22 (49%) patients. With a median follow-up of 2.47 years (range: 0.13-6.93), 2-year overall survival (OS), relapse free survival (RFS), cumulative incidence of relapse (CIR) and non-relapse mortality (NRM) were 57.3% (95% CI: 0.432-0.760), 56.2% (95% CI: 0.562-0.745), 23.3% (95% CI: 0.13-0.42), and 20.4% (95% CI: 0.109-0.384), respectively. Two-year OS (52% vs. 68%, P = .641), RFS (54% vs. 59%, P = .820), CIR (33.5% vs. 8.5%, P = .104), and NRM (12.5% vs. 32.2%, P = .120) were not significantly different between patients who underwent their first vs. second transplant, respectively. In univariate analysis, only Ph-like genotype was associated with inferior RFS (P = .03). AlloHCT post CAR-T response is associated with a relatively low early mortality rate and encouraging survival results in high-risk adults with r/r B-ALL, extending to the second alloHCT for fit and eligible patients.

Successful use of anti-CD19 CAR T cells in severe treatment-refractory stiff-person syndrome.

Treatment with autologous chimeric antigen receptor (CAR) T cells has emerged as a highly effective approach in neuroimmunological disorders such as myasthenia gravis. We report a case of successful anti-CD19 CAR T cell use in treatment-refractory stiff-person syndrome (SPS). To investigate clinical and immunological effects of anti-CD19 CAR T cell use in treatment-refractory SPS, a 69-y-old female with a 9-y history of treatment-refractory SPS with deteriorating episodes of stiffness received an infusion of autologous anti-CD19 CAR T cells (KYV-101) and was monitored clinically and immunologically for more than 6 mo. CAR T cell infusion resulted in reduced leg stiffness, drastic improvement in gait, walking speed increase over 100%, and daily walking distance improvement from less than 50 m to over 6 km within 3 mo. GABAergic medication (benzodiazepines) was reduced by 40%. KYV-101 CAR T cells were well tolerated with only low-grade cytokine release syndrome. This report of successful use of anti-CD19 CAR T cells in treatment-refractory SPS supports continued exploration of this approach in SPS and other B cell-related autoimmune disorders.

CAR-T Cells in Acute Myeloid Leukemia: Where Do We Stand?

Despite recent advances, the prognosis of acute myeloid leukemia (AML) remains unsatisfactory due to disease recurrence and the development of resistance to both conventional and novel therapies. Engineered T cells expressing chimeric antigen receptors (CARs) on their cellular surface represent one of the most promising anticancer agents. CAR-T cells are increasingly used in patients with B cell malignancies, with remarkable clinical results despite some immune-related toxicities. However, at present, the role of CAR-T cells in myeloid neoplasms, including AML, is extremely limited, as specific molecular targets for immune cells are generally lacking on AML blasts. Besides the paucity of dispensable targets, as myeloid antigens are often co-expressed on normal hematopoietic stem and progenitor cells with potentially intolerable myeloablation, the AML microenvironment is hostile to T cell proliferation due to inhibitory soluble factors. In addition, the rapidly progressive nature of the disease further complicates the use of CAR-T in AML. This review discusses the current state of CAR-T cell therapy in AML, including the still scanty clinical evidence and the potential approaches to overcome its limitations, including genetic modifications and combinatorial strategies, to make CAR-T cell therapy an effective option for AML patients.

The Challenging Approach to Multiple Myeloma: From Disease Diagnosis and Monitoring to Complications Management.

The outcome of multiple myeloma (MM) has significantly improved in the last few decades due to several factors such as new biological discoveries allowing to better stratify disease risk, development of more effective therapies and better management of side effects related to them. However, handling all these aspects requires an interdisciplinary approach involving multiple knowledge and collaboration of different specialists. The hematologist, faced with a patient with MM, must not only choose a treatment according to patient and disease characteristics but must also know when therapy needs to be started and how to monitor it during and after treatment. Moreover, he must deal not only with organ issues related to MM such as bone disease, renal failure or neurological disease but also with adverse events, often very serious, related to novel therapies, particularly new generation immunotherapies such as CAR T cell therapy and bispecific antibodies. In this review, we provide an overview on the newer MM diagnostic and monitoring strategies and on the main side effects of MM therapies, focusing on adverse events occurring during treatment with CAR T cells and bispecific antibodies.