Therapy For Hematologic Malignancies Research Articles

Strategies for Reducing Toxicity and Enhancing Efficacy of Chimeric Antigen Receptor T Cell Therapy in Hematological Malignancies.

Chimeric antigen receptor T cell (CAR-T) therapy in hematologic malignancies has made great progress, but there are still some problems. First, T cells from tumor patients show an exhaustion phenotype; thus, the persistence and function of the CAR-Ts are poor, and achieving a satisfactory curative effect is difficult. Second, some patients initially respond well but quickly develop antigen-negative tumor recurrence. Thirdly, CAR-T treatment is not effective in some patients and is accompanied by severe side effects, such as cytokine release syndrome (CRS) and neurotoxicity. The solution to these problems is to reduce the toxicity and enhance the efficacy of CAR-T therapy. In this paper, we describe various strategies for reducing the toxicity and enhancing the efficacy of CAR-T therapy in hematological malignancies. In the first section, strategies for modifying CAR-Ts using gene-editing technologies or combining them with other anti-tumor drugs to enhance the efficacy of CAR-T therapy are introduced. The second section describes some methods in which the design and construction of CAR-Ts differ from the conventional process. The aim of these methods is to enhance the anti-tumor activity of CAR-Ts and prevent tumor recurrence. The third section describes modifying the CAR structure or installing safety switches to radically reduce CAR-T toxicity or regulating inflammatory cytokines to control the symptoms of CAR-T-associated toxicity. Together, the knowledge summarized herein will aid in designing better-suited and safer CAR-T treatment strategies.

Combination therapy of DKK1 inhibition and NKG2D chimeric antigen receptor T cells for the treatment of gastric cancer.

Despite the successful application of chimeric antigen receptor (CAR)-T cell therapy in hematological malignancies, the treatment efficacy in solid tumors remains unsatisfactory, largely due to the highly immunosuppressive tumor microenvironment and low density of specific tumor antigens. Natural killer group 2 member D (NKG2D) CAR-T cells have shown promising treatment effects on several cancers such as lymphoma and multiple myeloma. However, the application and efficacy of NKG2D-CAR-T cells in gastric cancer (GC) still needs further exploration. This study identified a novel combination immunotherapy strategy with Dickkopf-1 (DKK1) inhibition and NKG2D-CAR-T cells, exerting synergistic and superior antitumor effect in GC. We show that the baseline expression of NKG2D ligands (NKG2DLs) is at low levels in GC tissues from The Cancer Genome Atlas and multiple GC cell lines including NCI-N87, MGC803, HGC27, MKN45, SGC7901, NUGC4, and AGS. In addition, DKK1 inhibition by WAY-262611 reverses the suppressive tumor immune microenvironment (TIME) and upregulates NKG2DL expression levels in both GC cell lines and GC tissues from a xenograft NCG mouse model. DKK1 inhibition in GC cells markedly improves the immune-activating and tumor-killing ability of NKG2D-CAR-T cells as shown by cytotoxicity assays in vitro. Moreover, the combination therapy of NKG2D-CAR-T and WAY-262611 triggers superior antitumor effects in vivo in a xenograft NCG mouse model. In sum, our study reveals the role of DKK1 in remodeling GC TIME and regulating the expression levels of NKG2DLs in GC. We also provide a promising treatment strategy of combining DKK1 inhibition with NKG2D-CAR-T cell therapy, which could bring new breakthroughs for GC immunotherapy.

Facing challenges with hope: universal immune cells for hematologic malignancies.

Many patients have achieved a favorable overall survival rate since allogenic hematopoietic stem cell transplantation (allo-HSCT) has been widely implemented to treat hematologic malignancies. However, graft-versus-host disease (GVHD) and complications of immunosuppressive drugs after allo-HSCT are the main causes of non-relapse mortality and a poor quality of life. In addition, GVHD and infusion-induced toxicity still occur with donor lymphocyte infusions (DLIs) and chimeric antigen receptor (CAR) T-cell therapy. Because of the special immune tolerance characteristics and anti-tumor ability of universal immune cells, universal immune cell therapy may strongly reduce GVHD, while simultaneously reducing tumor burden. Nevertheless, widespread application of universal immune cell therapy is mainly restricted by poor expansion and persistence efficacy. Many strategies have been applied to improve universal immune cell proliferation and persistence efficacy, including the use of universal cell lines, signaling regulation and CAR technology. In this review we have summarized current advances in universal immune cell therapy for hematologic malignancies with a discussion of future perspectives.

CAR T-cells to treat brain tumors.

Tremendous success using CAR T therapy in hematological malignancies has garnered significant interest in developing such treatments for solid tumors, including brain tumors. This success, however, has yet to be mirrored in solid organ neoplasms. CAR T function has shown limited efficacy against brain tumors due to several factors including the immunosuppressive tumor microenvironment, blood-brain barrier, and tumor-antigen heterogeneity. Despite these considerations, CAR T-cell therapy has the potential to be implemented as a treatment modality for brain tumors. Here, we review adult and pediatric brain tumors, including glioblastoma, diffuse midline gliomas, and medulloblastomas that continue to portend a grim prognosis. We describe insights gained from different preclinical models using CAR T therapy against various brain tumors and results gathered from ongoing clinical trials. Furthermore, we outline the challenges limiting CAR T therapy success against brain tumors and summarize advancements made to overcome these obstacles.

Pathological evaluation of renal complications in children following allogeneic hematopoietic stem cell transplantation: a retrospective cohort study

BackgroundAllogeneic hematopoietic stem cell transplantation (allo-HSCT) is a curative therapy for hematologic malignancies and non-malignant disorders, such as aplastic anemia, fanconi anemia, and certain immune deficiencies. Post-transplantation kidney injury is a common complication and involves a wide spectrum of structural abnormalities, including glomerular (MSPGN, mesangial proliferative glomerulonephritis; FSGS, focal segmental glomerulosclerosis; MPGN, membranoproliferative glomerulonephritis; MCD, minimal change disease), vascular (TMA, thrombotic microangiopathy), and/or tubulointerstitial (TIN, tubulointerstitial nephritis; ATI, acute tubular injury). Renal biopsy is the gold-standard examination for defining multiple etiologies of kidney impairment. Although kidney injury following HSCT has been studied, little is known about the effects of allo-HSCT on renal pathology in pediatric patients.MethodsWe retrospectively analyzed renal biopsy specimens from children with kidney injury after allo-HSCT and correlated results with clinical data in the last 10 years.ResultsAmong 25 children (18 males and 7 females), three patients had proteinuria indicating nephrotic syndrome (24-hour urinary total protein/weight > 50 mg/kg/d), nine patients had severely reduced estimated glomerular filtration rate (eGFR < 30 ml/min/1.73 m2) and four patients received kidney replacement therapy (KRT). The main pathologies identified from kidney biopsies were MSPGN (n = 12), FSGS (n = 12), MPGN (n = 5), TMA (n = 4), MCD (n = 3), diffuse glomerular fibrosis (DGF, n = 2), ATI and TIN, in isolation or combined with other pathologies. The median follow-up time was 16.5 (0.5 ~ 68.0) months. Three patients died of recurrent malignancy and/or severe infection, one child developed to end-stage renal disease (ESRD), six patients (24%) had elevated serum creatinine (SCr > 100µmol/l) and nine patients (36%) still had proteinuria.ConclusionsThis study evaluates histomorphologic findings from kidney biopsies of pediatric recipients following allo-HSCT. Detailed evaluation of renal biopsy samples is helpful to elucidate the nature of renal insult, and may potentially identify treatable disease processes.

2021-2022 Drug Updates in Hematologic Malignancies.

During JADPRO Live 2022, Jenni Tobin, PharmD, reviewed the indications of newly approved therapies in hematologic malignancies (including those for multiple myeloma, lymphoma, and acute leukemia) approved from late 2021 to late 2022. Dr. Tobin discussed their unique mechanisms of action, administration, and how to monitor for and manage side effects associated with these new therapeutics.

CAR T-cell Therapy of Hematologic Malignancies: An Update in Targeted Antigens

Immunotherapy with genetically engineered T-cells that express the chimeric antigen receptor (CAR) has raised hopes for the treatment of pediatric malignancies. Although CAR T-cell development is on a fast-moving pace and evolution, the context of exploring novel targetable antigens has been neglected. In this review study, we analyze the prominent hematologic antigens targeted by engineered T-cells in both preclinical and clinical aspects. Furthermore, we discuss the outcomes of CAR-based therapy in hematologic cancers from different viewpoints of treatment and provide some critical features for additional considerations. Almost certainly, most of the engineered T-cells redirected against hematologic disorders aim at conventional target antigens rather than targeting an ideal target antigen that is exclusively expressed on cancerous cells and restricted to normal tissues. CAR-based clinical trials in hematologic cancers have often dealt with CD19, followed by BCMA, CD22, and CD20 antigens. Besides, most of the scFvs used in the CAR structure are derived from murine antibodies, which may raise the concern about immunogenicity by reducing the persistence of modified T-cells. Nevertheless, short- and long-term life-threatening toxicities and the development of escape mechanisms that result in resistance and antigen loss are not thoroughly understood yet. The ultimate goal of using modified CAR T-cells is to make them effective and curative. Therefore, a better understanding of all the features pertaining to target antigens is imperative. Also, the methods to identify candidate target antigens and manage the associated obstacles of CAR T-cells should be evaluated and prioritized.

Mucosal-associated invariant T cells in hematological malignancies: Current knowledge, pending questions.

Non-classical HLA restricted T cell subsets such as γδ T and NK-T cells are showing promises for immune-based therapy of hematological malignancies. Mucosal-Associated Invariant T cells (MAIT) belong to this family of innate-like T cell subsets and are the focus of many studies on infectious diseases, owing to their unusual recognition of bacterial/fungal metabolites. Their ability to produce type 1 cytokines (IFNγ, TNFα) as well as cytotoxic effector molecules endows them with potential anti-tumor functions. However, their contribution to tumor surveillance in solid cancers is unclear, and only few studies have specifically focused on MAIT cells in blood cancers. In this review, we wish to recapitulate our current knowledge on MAIT cells biology in hematological neoplasms, at diagnosis and/or during treatment, as well as tentative approaches to target them as therapeutic tools. We also wish to take this opportunity to briefly elaborate on what we think are important question to address in this field, as well as potential limitations to overcome in order to make MAIT cells the basis of future, novel therapies for hematological cancers.

Cardiotoxicities of Novel Therapies in Hematologic Malignancies: Chimeric Antigen Receptor T-Cell Therapy and Bispecific T-Cell Engager Therapy.

The field of malignant hematology is transforming with novel immunotherapeutic approaches. Unfortunately, quality of life, treatment efficacy, and life expectancy are negatively affected by cardiotoxic side effects of treatment. To date, the exact mechanism and incidence of cardiotoxicity associated with these therapies is unclear. These events are believed to be triggered or occur concurrently with cytokine release syndrome. Furthermore, there are no formal guidelines to provide evaluation, treatment, and surveillance. We aim to synthesize available literature with updates on the cardiotoxic effects of novel therapies used in malignant hematologic disorders, with a focus on chimeric antigen receptor T-cell therapy and bispecific T-cell engager therapy, along with a proposed algorithm that may guide pretreatment evaluation, monitoring during treatment, and post-treatment surveillance.

Challenges and optimal strategies of CAR T therapy for hematological malignancies.

Remarkable improvement relative to traditional approaches in the treatment of hematological malignancies by chimeric antigen receptor (CAR) T-cell therapy has promoted sequential approvals of eight commercial CAR T products within last 5 years. Although CAR T cells' productization is now rapidly boosting their extensive clinical application in real-world patients, the limitation of their clinical efficacy and related toxicities inspire further optimization of CAR structure and substantial development of innovative trials in various scenarios. Herein, we first summarized the current status and major progress in CAR T therapy for hematological malignancies, then described crucial factors which possibly compromise the clinical efficacies of CAR T cells, such as CAR T cell exhaustion and loss of antigen, and finally, we discussed the potential optimization strategies to tackle the challenges in the field of CAR T therapy.

Successful Autologous Bone Marrow Transplantation in Active COVID-19 Patients: Case Report

BackgroundPatients with hematologic malignancies are considered at high risk for COVID-19 infection either from the disease or the treatment. Hematopoietic stem cell transplantation, one of the approved therapies for hematologic malignancies, was performed worldwide during the COVID-19 era with some regulations, such as COVID-19 testing, before proceeding with transplantation or cellular therapy. To the authors’ knowledge, none have reported the result of autologous hematopoietic stem cell transplantation in an active COVID-19 patient. Case PresentationWe describe a successful clinical course of autologous bone marrow transplantation for 2 lymphoma patients who tested positive for COVID-19. A thorough discussion was conducted between multidisciplinary hemato-oncology, intensive care, and infectious diseases teams. The decision was to proceed toward bone marrow transplantation with some modifications in the transplantation protocol and close patient monitoring. ConclusionOur cases lend credence that successful autologous bone marrow transplantation is possible among active COVID-19 patients. The obstacles we faced could be overcome with collaboration between a highly qualified multidisciplinary team. Despite the potential complications, the benefits of bone marrow transplantation among patients with a high risk of relapse and who are still COVID-19-positive outweigh the risks. However, further studies are still recommended to support our inference.

Immunogenic Cell Death in Hematological Malignancy Therapy.

Although the curative effect of hematological malignancies has been improved in recent years, relapse or drug resistance of hematological malignancies will eventually recur. Furthermore, the microenvironment disorder is an important mechanism in the pathogenesis of hematological malignancies. Immunogenic cell death (ICD) is a unique mechanism of regulated cell death (RCD) that triggers an intact antigen-specific adaptive immune response by firing a set of danger signals or damage-associated molecular patterns (DAMPs), which is an immunotherapeutic modality with the potential for the treatment of hematological malignancies. This review summarizes the existing knowledge about the induction of ICD in hematological malignancies and the current research on combining ICD inducers with other treatment strategies for hematological malignancies.

Identifying effect modifiers of CAR-T cell therapeutic efficacy: a systematic review and individual patient data meta-analysis protocol

BackgroundChimeric antigen receptor T cell therapy (CAR-T) represents a promising and exciting new therapy for hematologic malignancies, where prognosis for relapsed/refractory patients remains poor. Encouraging results from clinical trials have often been tempered by heterogeneity in response to treatment among patients, as well as safety concerns including cytokine release syndrome. The identification of specific patient or treatment-specific factors underlying this heterogeneity may provide the key to the long-term sustainability of this complex and expensive therapy. An individual patient data meta-analysis (IPMDA) may provide potential explanations for the high degree of heterogeneity. Therefore, our objective is to perform a systematic review and IPDMA of CAR-T cell therapy in patients with hematologic malignancies to explore potential effect modifiers of CAR-T cell therapy.Methods and analysisWe will search MEDLINE, Embase, and the Cochrane Central Register of Controlled Clinical Trials. Studies will be screened in duplicate at the abstract level, then at the full-text level by two independent reviewers. We will include any prospective clinical trial of CAR-T cell therapy in patients with hematologic malignancies. Our primary outcome is complete response, while secondary outcomes of interest include overall response, progression-free survival, overall survival, and safety. IPD will be collected from each included trial and, in the case of missing data, corresponding authors/study sponsors will be contacted. Standard aggregate meta-analyses will be performed, followed by the IPD meta-analysis using a one-stage approach. A modified Institute of Health Economics tool will be used to evaluate the risk of bias of included studies.Ethics and disseminationIdentifying characteristics that may act as modifiers of CAR-T cell efficacy is of paramount importance and can help shape future clinical trials in the field. Results from this study will be submitted for publication in a peer-reviewed scientific journal, presented at relevant conferences and shared with relevant stakeholders.

Efficacy and safety of chimeric antigen receptor T-cell (CAR-T) therapy in hematologic malignancies: a living systematic review on comparative studies.

Chimeric antigen receptor T-cell (CAR-T) cell therapies have been claimed to be curative in responsive patients. Nonetheless, response rates can vary according to different characteristics, and these therapies are associated with important adverse events such as cytokine release syndrome, neurologic adverse events, and B-cell aplasia. This living systematic review aims to provide a timely, rigorous, and continuously updated synthesis of the evidence available on the role of CAR-T therapy for the treatment of patients with hematologic malignancies. A systematic review with meta-analysis of randomized controlled trials (RCTs) and comparative non-randomized studies of interventions (NRSI), evaluating the effect of CAR-T therapy versus other active treatments, hematopoietic stem cell transplantation, standard of care (SoC) or any other intervention, was performed in patients with hematologic malignancies. The primary outcome is overall survival (OS). Certainty of the evidence was determined using the Grading of Recommendations, Assessment, Development and Evaluation (GRADE) approach. Searches were performed in the Epistemonikos database, which collates information from multiple sources to identify systematic reviews and their included primary studies, including Cochrane Database of Systematic Reviews, MEDLINE, EMBASE, CINAHL, PsycINFO, LILACS, DARE, HTA Database, Campbell database, JBI Database of Systematic Reviews and Implementation Reports, EPPI-Centre Evidence Library. A manual search was also carried out. We included the evidence published up to 1 July 2022. We included the evidence published up to 1 July 2022. We considered 139 RCTs and 1725 NRSI as potentially eligible. Two RCTs (N = 681) comparing CAR-T therapy with SoC in patients with recurrent/relapsed (R/R) B-cell lymphoma were included. RCTs did not show statistical differences in OS, serious adverse events, or total adverse events with grade ⩾ 3. Higher complete response with substantial heterogeneity [risk ratio = 1.59; 95% confidence interval (CI) = (1.30-1.93); I 2 = 89%; 2 studies; 681 participants; very low certainty evidence] and higher progression-free survival [hazard ratio for progression or death = 0.49; 95% CI = (0.37-0.65); 1 study; 359 participants; moderate certainty evidence] were reported with CAR-T therapies. Nine NRSI (N = 540) in patients with T or B-cell acute lymphoblastic leukemia or R/R B-cell lymphoma were also included, providing secondary data. In general, the GRADE certainty of the evidence for main outcomes was mostly low or very low. So far, assuming important limitations in the level of certainty due to scarce and heterogenous comparative studies, CAR-T therapies have shown some benefit in terms of progression-free survival, but no overall survival, in patients with R/R B-cell lymphoma. Despite one-arm trials have already facilitated approval of CAR-T cell treatments, additional evidence from large comparative studies is still needed to better characterize the benefit-harm ratio of the use of CAR-T in a variety of patient populations with hematological malignancies. https://doi.org/10.12688/openreseurope.14390.1. 10.17605/OSF.IO/V6HDX.

CAR designs for solid tumors: overcoming hurdles and paving the way for effective immunotherapy.

Chimeric antigen receptor T cell (CAR-T) therapy has revolutionized immunotherapy by modifying patients' immune cells genetically. By expressing CARs, these modified cells can specifically identify and eliminate tumor cells. The success of CAR-T therapy in hematological malignancies, such as leukemia and lymphoma, has been remarkable. Numerous studies have reported improved patient outcomes and increased survival rates. However, the application of CAR-T therapy in treating solid tumors faces significant challenges. Solid tumors possess complex microenvironments containing stromal cells, extracellular matrix components, and blood vessels. These factors can impede the infiltration and persistence of CAR-T cells within the tumor. Additionally, the lack of target antigens exclusively expressed on tumor cells raises concerns about off-target effects and potential toxicity. This review aims to discuss advancements achieved by CAR-T therapy in solid tumors and the clinical outcomes in the realm of solid tumors.

Chimeric antigen receptor T (CAR-T) cells: Novel cell therapy for hematological malignancies.

Over the last decade, the emergence of several novel therapeutic approaches has changed the therapeutic perspective of human malignancies. Adoptive immunotherapy through chimeric antigen receptor T cell (CAR-T), which includes the engineering of T cells to recognize tumor-specific membrane antigens and, as a result, death of cancer cells, has created various clinical benefits for the treatment of several human malignancies. In particular, CAR-T-cell-based immunotherapy is known as a critical approach for the treatment of patients with hematological malignancies such as acute lymphoblastic leukemia (ALL), multiple myeloma (MM), chronic lymphocytic leukemia (CLL), acute myeloid leukemia (AML), Hodgkin lymphoma (HL), and non-Hodgkin's lymphoma (NHL). However, CAR-T-cell therapy of hematological malignancies is associated with various side effects. There are still extensive challenges in association with further progress of this therapeutic approach, from manufacturing and engineering issues to limitations of applications and serious toxicities. Therefore, further studies are required to enhance efficacy and minimize adverse events. In the current review, we summarize the development of CAR-T-cell-based immunotherapy and current clinical antitumor applications to treat hematological malignancies. Furthermore, we will mention the current advantages, disadvantages, challenges, and therapeutic limitations of CAR-T-cell therapy.

Leveraging Natural Killer Cell Innate Immunity against Hematologic Malignancies: From Stem Cell Transplant to Adoptive Transfer and Beyond

Numerous recent advancements in T-cell based immunotherapies have revolutionized the treatment of hematologic malignancies. In the race towards the first approved allogeneic cellular therapy product, there is growing interest in utilizing natural killer (NK) cells as a platform for off-the-shelf cellular therapies due to their scalable manufacturing potential, potent anti-tumor efficacy, and superior safety profile. Allogeneic NK cell therapies are now being actively explored in the setting of hematopoietic stem cell transplantation and adoptive transfer. Increasingly sophisticated gene editing techniques have permitted the engineering of chimeric antigen receptors, ectopic cytokine expression, and tumor recognition signals to improve the overall cytotoxicity of NK cell therapies. Furthermore, the enhancement of antibody-dependent cellular cytotoxicity has been achieved through the use of NK cell engagers and combination regimens with monoclonal antibodies that act synergistically with CD16-expressing NK cells. Finally, a greater understanding of NK cell biology and the mechanisms of resistance have allowed the preclinical development of NK checkpoint blockade and methods to modulate the tumor microenvironment, which have been evaluated in early phase trials. This review will discuss the recent clinical advancements in NK cell therapies in hematologic malignancies as well as promising avenues of future research.

Chimeric Antigen Receptor T-cell Therapy in Hematologic Malignancies and Patient-reported Outcomes: A Scoping Review.

The inclusion of patient-reported outcome (PRO) measures in chimeric antigen receptor (CAR) T-cell therapy research is critical for understanding the impact of this novel approach from a unique patient standpoint. We performed a scoping review to map the available literature on the use of PRO measures in CAR T-cell therapy studies of patients with hematologic malignancies published between January 2015 and July 2022. Fourteen studies were identified, of which 7 (50%) were investigational early-phase trials, 6 (42.9%) were observational studies, and 1 (7.1%) was a pilot study. The EQ-5D and the PROMIS-29 were the 2 most frequently used PRO measures, being included in 6 (42.9%) and 5 (35.7%) studies, respectively. Despite differences in study designs, there seems to be evidence of improvements over time since CAR T-cell infusion in important domains such as physical functioning and fatigue, at least in patients who respond to therapy. Overall, the studies identified in our review have shown the added value of PRO assessment in CAR T-cell therapy research by providing novel information that complements the knowledge on safety and efficacy. However, there are several questions which remain to be answered in future research. For example, limited evidence exists regarding patient experience during important phases of the disease trajectory as only 4 (28.6%) and 5 (35.7%) studies provided information on PROs during the first 2 weeks from CAR T-cell infusion and after the first year, respectively. Time is ripe for a more systematic implementation of high-quality PRO assessment in future clinical trials and in real-life settings of patients treated with CAR T-cell therapy.

Pre-Clinical Evaluation of Allogeneic 'Off-the-Shelf' CAR-iNKT Cells Targeting CD19 (ALA-101)-Expressing Hematological Malignancies

Background Arovella Therapeutics is a preclinical stage company developing allogeneic 'off-the-shelf' human invariant natural killer T (iNKT) cell-based therapies for hematological malignancies and solid tumors. iNKT cells are a unique and rare subset of lymphocytes that express an invariant T cell receptor (iTCR) and are restricted by the glycolipid-presenting, non-polymorphic CD1d. Unlike allogeneic conventional T cells that can cause acute graft versus host disease (aGVHD) following administration, iNKT cells exert a protective effect against aGVHD after allogeneic stem cell transplantation in humans. Early clinical studies suggest that non-engineered and CAR-engineered allogeneic iNKT cells are safe in humans and represent a promising therapeutic approach, bypassing the requirement to delete the endogenous TCR. Their inherent biological properties enable iNKT cells to mount a swift and robust antitumor immune response when engineered with a Chimeric Antigen Receptor (CAR). The use of iNKT cells from healthy donors has many advantages over allogeneic T cells and NK cells, including recognition of CD1d+ on tumor cells through the iTCR in addition to the CAR mediated recognition of a tumor specific antigen. Here, we report results from early preclinical studies assessing the functional profile of iNKT cells engineered to express an optimized CAR targeting CD19 (ALA-101) for allogeneic use. Methods iNKT cells were purified from the peripheral blood of healthy donors followed by lentiviral vector transduction (development construct) to express a CD19-targeting CAR. After transduction, cells were expanded to generate ALA-101. The CD19 CAR functionality of ALA-101 was compared against non-transduced (NT) iNKT cells or with CAR19-T cells transduced with the same lentiviral vector. Flow cytometry was used to assess the expression of the CAR and ALA-101 phenotypes and to determine CD19 and CD1d expression levels in the primary patient samples; marginal zone lymphoma (MZL) cells and two chronic lymphocytic leukemia (CLL) samples. In vitro cytotoxicity assays and intracellular cytokine staining were used to evaluate the CAR-dependent and -independent anti-tumor activities using B cell lymphoblastic leukemia (B-ALL) SEM cell line and primary tumor cells from patients. In vivo efficacy testing of ALA-101 is ongoing in a disseminated NSG mouse model of SEM cells expressing luciferase. Results The purity of iNKT cells following selection from peripheral blood mononuclear cells of healthy donors was greater than 99%. ALA-101 was optimized for CD19 CAR functionality using a pre-clinical lentiviral vector. Using multiple independent donors, we confirmed that iNKT cells engineered with lentiviral vector encoding CD19 CAR (ALA-101) maintained the surface CAR expression over 35 days and expressed variable frequencies of CD27, CD62L and CCR7. Compared to day 11 CAR19-T cells, day 17 and day 35 ALA-101 cells demonstrated higher cytotoxicity against the CD19+CD1d- B-ALL SEM cells; similarly, ALA-101 demonstrated higher cytotoxic activity against CD19+CD1d+ malignant B cells from patients with CLL and MZL. Of note, when tested at different time points, ALA-101 cells expanded for 17 and 35 days showed similar cytotoxicity against primary patient samples and SEM cells. Intracellular cytokine staining from tumor cell co-cultures demonstrated that the majority of ALA-101 expressed TNFα, Perforin and Granzyme B. Conclusions Here we report the preclinical activity of ALA-101, a novel allogeneic CAR19-iNKT cell product for the treatment of CD19+ B-cell malignancies. Our data shows the successful generation of polyfunctional allogeneic iNKT cells in terms of cytotoxicity and cytokine profile when engineered to express a CD19-targeting CAR. We have initiated the manufacturing of ALA-101 with the intention to enter clinic trials in CD19 positive malignancies. Taken together, the data indicates that iNKT cells are a potent allogeneic, 'off-the-shelf' cell therapy to treat a broad spectrum of hematological malignancies and solid tumors.

Multi-Omics Reveal Plasma Molecular Signatures of B-ALL Patients during CAR-T Cell Therapy

Background: The Chimeric antigen receptor (CAR) T cell therapy for hematological malignancies has revolutionized anticancer therapy. However, CAR-T cells can cause immunity-related adverse events, such as cytokine release syndrome and neurotoxicity; the efficacy of CAR-T cell therapy also varies substantially with different patients. Poor understanding of plasma proteomic and metabolomic features during chimeric antigen receptor (CAR) T cell therapy may affect the exploitation of therapeutic potential in the management of hematological malignancies. Methods: We longitudinally performed proteomic and metabolomic profiling of plasma from 20 patients with relapsed/refractory B-cell acute lymphoblastic leukemia (B-ALL) on CAR-T cell therapy. Blood samples were collected from B-ALL patients at different time points, including before fludarabine/cyclophosphamide lymphodepletion (Pre-FC), Day 0 (D0), Day 7 (D7), Day 14 (D14), Day 28 (D28), and Peak-IL-6 during CAR-T cell therapy. The plasma samples from 22 age- and gender-matched healthy controls (HC) were used as controls. Results: Compared with HC, 239, 217, 282, 260 and 261 proteins were markedly different in terms of expression level in these five groups (Pre-FC, D0, D7, D14, D28 versus HC), respectively (Figure 1). The pathways of differentially expressed proteins (DEPs) were highly enriched in processes implicated in actin binding, cadherin binding, regulation of ion transport, platelet aggregation and regulation of actin cytoskeleton, with actin binding obtaining the most significant change (Figure 1). 104, 139, 175, 223 and 87 metabolites were significantly different in these five groups, respectively (Figure 1). Interestingly, the alterations of plasma metabolites became more extensive on the 14th day after CAR-T cell infusion, indicating that the change of metabolites may lag behind that of proteins. Furthermore, CAR-T cell therapy reshaped plasma proteome and metabolome with discrete kinetics, as represented by upregulated acute-phase response and triglycerides metabolism and downregulated actin cytoskeleton organization, retrograde endocannabinoid signaling and glycosylphosphatidylinositol (GPI)-anchor biosynthesis during the therapy. We also identified sets of covarying molecules that were involved in acute inflammatory response, epithelial mesenchymal transition, glutathione metabolism and tryptophan metabolism in concert with the dynamics of circulating IL-6 in patients with cytokines storm. Subgroup analysis further revealed numerous prognostic biomarkers, such as proteins related to allograft rejection, which were down-regulated in patients who achieved stable complete remission. Conclusions: This study provided a dynamic landscape of plasma proteome and metabolome in B-ALL patients during CAR-T cell therapy and offered novel insights into CAR-T cell therapy-related toxicities and prognostic heterogeneity. Figure 1View largeDownload PPTFigure 1View largeDownload PPT Close modal