Influencing factors and solution strategies of chimeric antigen receptor T-cell therapy(CAR–T)cell immunotherapy

Chimeric antigen receptor T-cesll therapy(CAR–T)has achieved groundbreaking advancements in clinical application,ushering in a new era for innovative cancer treatment.However,the challenges associated with implementing this novel targeted cell therapy are increasingly significant.Particularly in the clinical management of solid tumors,obstacles such as the immunosuppressive effects of the tumor microenvironment,limited local tumor infiltration capability of CAR–T cells,heterogeneity of tumor targeting antigens,uncertainties surrounding CAR–T quality,control,and clinical adverse reactions have contributed to increased drug resistance and decreased compliance in tumor therapy.These factors have significantly impeded the widespread adoption and utilization of this therapeutic approach.In this paper,we comprehensively analyze recent preclinical and clinical reports on CAR–T therapy while summarizing crucial factors influencing its efficacy.Furthermore,we aim to identify existing solution strategies and explore their current research status.Through this review article,our objective is to broaden perspectives for further exploration into CAR–T therapy strategies and their clinical applications.

Chimeric antigen receptors:On the road to realising their full potential

Chimeric antigen receptors （CARs） are fusion molecules that may be genetically delivered ex-vivo to T-cells and other immune cell populations, thereby conferring specifcity for native target antigens found on the surface of tumour and other target cell types. Antigen recognition by CARs is neither restricted by nor dependent upon human leukocyte antigen antigen expression, favouring widespread use of this technology across transplantation barriers. Signalling is delivered by a designer endodomain that provides a tailored and target-dependent activation signal to polyclonal circulating T-cells. Recent clinical data emphasise the enormous promise of this emer-ging immunotherapeutic strategy for B-cell malignancy, notably acute lymphoblastic leukaemia. In that context, CARs are generally targeted against the ubiquitous B-cell antigen, CD19. However, CAR T-cell immunotherapy is limited by potential for severe ontarget toxicity, notably due to cytokine release syndrome. Furthermore, effcacy in the context of solid tumours remains unproven, owing in part to lack of availability of safe tumourspecific targets, inadequate CAR T-cell homing and hostility of the tumour microenvironment to immune effector deployment. Manufacture and commercial development encountered with more traditional drug products. Finally, there is increasing interest in the application of this technology to the treatment of non-malignant disease states, such as autoimmunity, chronic infection and in the suppression of allograft rejection. Here, we consider the background and direction of travel of this emerging and highly promising treatment for malignant and other disease types.

Tumor-derived exosomes induce initial activation by exosomal CD19 antigen but impair the function of CD19-specific CAR T-cells via TGF-βsignaling

Tumor-derived exosomes (TEXs) enriched in immune suppressive molecules predominantly drive T-cell dysfunction and impair antitumor immunity. Chimeric antigen receptor (CAR) T-cell therapy has emerged as a promising treatment for refractory and relapsed hematological malignancies, but whether lymphoma TEXs have the same impact on CAR T-cell remains unclear. Here, we demonstrated that B-cell lymphoma-derived exosomes induce the initial activation of CD19-CAR T-cells upon stimulation with exosomal CD19. However, lymphoma TEXs might subsequently induce CAR T-cell apoptosis and impair the tumor cytotoxicity of the cells because of the upregulated expression of the inhibitory receptors PD-1, TIM3, and LAG3 upon prolonged exposure. Similar results were observed in the CAR T-cells exposed to plasma exosomes from patients with lymphoma. More importantly, single-cell RNA sequencing revealed that CAR T-cells typically showed differentiated phenotypes and regulatory T-cell (Treg) phenotype conversion. By blocking transforming growth factor β (TGF-β)-Smad3 signaling with TGF-β inhibitor LY2109761, the negative effects of TEXs on Treg conversion, terminal differentiation, and immune checkpoint expression were rescued. Collectively, although TEXs lead to the initial activation of CAR T-cells, the effect of TEXs suppressed CAR T-cells, which can be rescued by LY2109761. A treatment regimen combining CAR T-cell therapy and TGF-β inhibitors might be a novel therapeutic strategy for refractory and relapsed B-cell lymphoma.

Diversity of yO T-cell antigens

In the last two decades, it has become clear that yo T cells recognize a diverse array of antigens including self and foreign, large and small, and peptidic and non-peptidic molecules. In this respect, 78 antigens as a whole resemble more the antigens recognized by antibodies than those recognized by T cells. Because of this antigenic diversity, no single mechanism--such as the major histocompatibility complex （MHC） restriction of ap T cells--is likely to provide a basis for all observed T-cell antigen receptor （TCR）-dependent 78 T-cell responses. Furthermore, available evidence suggests that many individual 78 T cells are poly-specific, probably using different modes of ligand recognition in their responses to unrelated antigens. While posing a unique challenge in the maintenance of self-tolerance, this broad reactivity pattern might enable multiple overlapping uses of 78 T-cell populations, and thus generate a more efficient immune response.

Advances in the development of chimeric antigen receptor-T-cell therapy in B-cell acute lymphoblastic leukemia

CD19-targeted chimeric antigen receptor T-cell(CAR-T)therapy is effective in refractory/relapsed(R/R)B-cell acute lymphoblastic leukemia(B-ALL).This review focuses on achievements,current obstacles,and future directions in CAR-T research.A high complete remission rate of 68%to 93%could be achieved after anti-CD19 CAR-T treatment for B-ALL.Cytokine release syndrome and CAR-T-related neurotoxicity could be managed.In view of difficulties collecting autologous lymphocytes,universal CAR-T is a direction to explore.Regarding the high relapse rate after anti-CD19 CAR-T therapy,the main solutions have been developing new targets including CD22 CAR-T,or CD19/CD22 dual CAR-T.Additionally,some studies showed that bridging into transplant post-CAR-T could improve leukemia-free survival.Some patients who did not respond to CAR-T therapy were found to have an abnormal conformation of the CD19 exon or trogocytosis.Anti-CD19 CAR-T therapy for R/R B-ALL is effective.From individual to universal CAR-T,from one target to multi-targets,CAR-T-cell has a chance to be off the shelf in the future.

Efficacy and safety of chimeric antigen receptor-T cells in the treatment of B cell lymphoma:a systematic review and meta-analysis

Background:Conventional treatment has limited efficacy in relapsed/refractory B-cell lymphoma.Since chimeric antigen receptor T-cell(CAR-T)technology has shown high safety and results in high remission rates,we investigated its efficacy and safety in B-cell lymphoma treatment and analyzed potential affecting factors to provide evidence for therapeutic strategies and applications.Methods:We searched databases including PubMed,Embase,and Cochrane up to July 2019.Meta-analysis 1 was conducted to study the efficacy of CAR-T cell for treating B-cell lymphoma,measuring the response rate and complete remission rate as outcomes.Sub-group analysis was performed for age,pathological type,target antigen,co-stimulatory molecule,and conditioning chemotherapy.Meta-analysis 2 was undertaken on the safety of the treatment with the incidence rate of toxicity(cytokine-releasing syndrome[CRS],neurotoxicity)as an outcome.Results:Seventeen studies were included in the systematic review and meta-analysis.It was found that CAR-T cells had good therapeutic effects in the following cases:B-cell lymphoma(patients≥65 years old);diffuse large B-cell lymphoma pathological type;patients with treatment target antigen other than CD19;patients treated with co-stimulatory molecules other than CD28,including 4-1BB+CD28 or 4-1BB;and patients treated with cyclophosphamide/fludarabine pre-treatment protocol conditioning chemotherapy.Although the CRS and neurotoxicity incidences were high,most were reversible with minimal risk of death.Conclusion:CAR-T cell treatment is safe for clinical application;however,toxicity effects should be monitored.

Features of intestinal T-cell lymphomas in Chinese population without evidence of celiac disease and their close association with Epstein-Barr virus infection

Background Intestinal T-cell lymphoma （ITCL） is a heterogeneous lymphoid neoplastic group with variable clinical and pathological features. ITCL in oriental countries is different from enteropathy-type intestinal T-cell lymphoma （ETCL） in relation to celiac disease and Epstein-Barr virus （EBV）. The objective of this study was to investigate the clinicopathological features, immunophenotype, expression of cytotoxic molecule （TIA-1）, T-cell receptor （TCR）-γ gene rearrangement, and Epstein-Barr virus （EBV） latent infection in primary ITCL without celiac disease in Chinese. Methods The clinical data of 42 patients were analyzed, and the patients were followed up. Compared with human reactive lymphoid tissues, in situ hybridization for EBER1/2, polymerase chain reaction for TCR-~/gene rearrangement, and immunohistochemical staining for immunophenotypes, TIA-1 and EBV latent membrane proteins （LMP-1） were investigated. Survival curves of different clinicopathological features, immunophenotypes, expression of LMPI , TCR-γ/gene rearrangement and therapy were analyzed. Results Three fourths of the patients suffered from ITCL in China were men with a peak age incidence in the 4th decade. Common presenting features included fever and hemotochezia. The prognosis was poor with a median survival of 3.0 months. The lesions were mostly localized in the ileocecum and colon. About 38/42 （90. 5% ） patients demonstrated pleomorphic medium-sized on large seen. All 42 patients with ITCL revealed CD45RO cells. Histological features of celiac disease were rarely positive. Neoplastic cells partially expressed T-cell differentiated antigens （CD3ε, CD4, CD8） and NK cell associated antigen （CD56）. The positive frequency of CD3e, CIM, CD8 and CD56 was 28/42 （66.7%） 7/42 （16.7%）, 10/42 （23.8%） and 12/42 （28.6%） respectively. Thirty-nine cells （92. 9% ） expressed TIA-1, but none expressed CD20 and CD68. More than half of the patients （64. 3% , 64.3% and 59.5% ） revealed TCR-γ primers respectively. EBER1/2 was detected in 41 （97.6%） of LMP-1 was 38. 1% （16/42）. gene rearrangement by three different TCR-γ the 42 patients. The expression frequency ofConclusions Primary ITCL without celiac disease in Chinese is a special highly EBV-associated clinicopathological entity. There are few similarities in patients with celiac disease in western countries. A small proportion of primary ITCLs in Chinese and extranodal NK/T-eell lymphoma of nasal type belong to the same speetrum.

Integration of molecular testing for the personalized management of patients with diffuse large B-cell lymphoma and follicular lymphoma

Diffuse large B-cell lymphoma(DLBCL)and follicular lymphoma(FL)are the most common forms of aggressive and indolent lymphoma,respectively.The majority of patients are cured by standard R-CHOP immunochemotherapy,but 30%–40%of DLBCL and 20%of FL patients relapse or are refractory(R/R).DLBCL and FL are phenotypically and genetically hereterogenous B-cell neoplasms.To date,the diagnosis of DLBCL and FL has been based on morphology,immunophenotyping and cytogenetics.However,next-generation sequencing(NGS)is widening our understanding of the genetic basis of the B-cell lymphomas.In this review we will discuss how integrating the NGS-based characterization of somatic gene mutations with diagnostic or prognostic value in DLBCL and FL could help refine B-cell lymphoma classification as part of a multidisciplinary pathology work-up.We will also discuss how molecular testing can identify candidates for clinical trials with targeted therapies and help predict therapeutic outcome to currently available treatments,including chimeric antigen receptor T-cell,as well as explore the application of circulating cell-free DNA,a non-invasive method for patient monitoring.We conclude that molecular analyses can drive improvements in patient outcomes due to an increased understanding of the different pathogenic pathways affected by each DLBCL subtype and indolent FL vs R/R FL.

Anti-BCMA CAR-T Cell Therapy in Relapsed or Refractory Multiple Myeloma Patients with Impaired Renal Function

Anti-B cell maturation antigen(BCMA)chimeric antigen receptor(CAR)T-cell therapyis effective and well-tolerated for refractory or relapsed multiple myeloma(RRMM).The purposcof the present study was to analyze efficacy in RRMM patients with renal impairment treated byanti-BCMA CAR-T cell therapy.A total of 59 RRMM patients were selected,and divided intoimpaired renal function(lRF)group[basclinc cstimated giomerular filtration rate(eSFR)<90 m/min/1.73 m^2(n=18)]and normal renal function(NRF)group(baseline eGFR≥90 mL/min/1.73 m,n=41).For patients with IRF,eGFR at the 6th month post-CAR-T cells infusion was significantlyhigher than the baseline(P<0.05).The multivariate analysis showed that light chain type and beta-2 micro-globulin(bcta-2M)were associated factors with the decrease of serum creatinine.Medianprogression-free survival(PFS)in the NRF group and IRF group was 266 days and 181 daysrespectively.Overall survival(OS)in the NRF group and lRF group was 877 days and 238 daysrespectively.There was no significant difference in the objective response rate(ORR)between thelRF group and the NRF group.It is suggested that CAR-T cells therapy could improve the renalfunction during the treatment of RRMM.The renal function could be more significantly improvedin RRMM patients with light chain type than with other types.

T-cell receptor-engineered T cells for cancer treatment： current status and future directions

T-cell receptor （TCR）-engineered T cells are a novel option for adoptive cell therapy used for the treatment of several advanced forms of cancer. Work using TCR- engineered T cells began more than two decades ago, with numerous preclinical studies showing that such cells could mediate tumor lysis and eradication. The success of these trials provided the foundation for clinical trials, including recent clinical successes using TCR- engineered T cells to target New York esophageal squamous cell carcinoma （NY-ESO-1）. These successes demonstrate the potential of this approach to treat cancer. In this review, we provide a perspective on the current and future applications of TCR-engineered T cells for the treatment of cancer. Our summary focuses on TCR activation and both pre-clinical and clinical applications of TCR-engineered T cells. We also discuss how to enhance the function of TCR-engineered T cells and prolong their longevity in the tumor microenvironment.

γδT cells:Major advances in basic and clinical research in tumor immunotherapy

γδT cells are a kind of innate immune T cell.They have not attracted sufficient attention because they account for only a small proportion of all immune cells,and many basic factors related to these cells remain unclear.However,in recent years,with the rapid development of tumor immunotherapy,γδT cells have attracted increasing attention because of their ability to exert cytotoxic effects on most tumor cells without major histocompatibility complex(MHC)restriction.An increasing number of basic studies have focused on the development,antigen recognition,activation,and antitumor immune response ofγδT cells.Additionally,γδT cell-based immunotherapeutic strategies are being developed,and the number of clinical trials investigating such strategies is increasing.This review mainly summarizes the progress of basic research and the clinical application ofγδT cells in tumor immunotherapy to provide a theoretical basis for further the development ofγδT cell-based strategies in the future.

EBV-associated lymphoproliferative disease post-CAR-T cell therapy

Epstein–Barr virus(EBV)-associated lymphoproliferative diseases(EBV-LPDs)are common complications that occur after solid organ transplantation or allogeneic hematopoietic stem-cell transplantation(HSCT).However,their occurrence and treatment post-chimeric antigen receptor-modified T(CAR-T)cell therapy has not been reported.Two patients had been diagnosed with EBV-positive aggressive B-cell lymphoma and experienced relapses after multiple lines of treatment.After receiving CAR-T cell therapy in tandem with autologous HSCT,the patients achieved complete remission.However,with a median time of 38.5 months after CAR-T cell therapy,B-cell-derived EBV-LPDs were diagnosed,and they were relieved through the administration of immune checkpoint inhibitor or B-cell-depleting agents.Collectively,our report suggests that EBV-LPDs may represent a long-term adverse event after CAR-T cell therapy,especially in patients who previously had EBV-positive disorders,and they can be resolved by immune normalization strategy or B-cell depleting therapy.

CAR-T cell therapy:Where are we now,and where are we heading?

Chimeric antigen receptor(CAR)-T-cell therapies have exhibited remarkable efficacy in the treatment of hematologic malignancies,with 9 CAR-T-cell products currently available.Furthermore,CAR-T cells have shown promising potential for expanding their therapeutic applications to diverse areas,including solid tumors,myocardial fibrosis,and autoimmune and infectious diseases.Despite these advancements,significant challenges pertaining to treatment-related toxic reactions and relapses persist.Consequently,current research efforts are focused on addressing these issues to enhance the safety and efficacy of CAR-T cells and reduce the relapse rate.This article provides a comprehensive overview of the present state of CAR-T-cell therapies,including their achievements,existing challenges,and potential future developments.

Revisiting mechanisms of resistance to immunotherapies in metastatic clear-cell renal-cell carcinoma

Renal-cell carcinoma(RCC)remains a leading cause of cancer-related mortality worldwide.Though newer therapeutic combinations of immune checkpoint inhibitors and targeted therapies have greatly improved outcomes,resistance to these therapies is becoming a challenge for long-term control.Mechanisms of resistance have been explored in a variety of solid tumors,including RCC.Based upon our review of the current literature on the mechanisms of resistance to immunotherapies for the management of metastatic clear-cell renal cell carcinomas(mccRCC),the ensuing conclusions have been made:The management of mccRCC has progressed substantially with the advent of checkpoint inhibitors and targeted oral therapies,alone and/or in combination.Nevertheless,innate or developed resistance to these therapies remains an ongoing challenge,particularly to immune checkpoint inhibitors(ICIs).Several of the known mechanisms of resistance have been well defined,but recent progression in cellular therapies helps to expand the armamentarium of potential combination options that may overcome these modes of resistance and improve long-term disease control and survival for an otherwise dismal disease.In the ensuing review and update of the literature on the mechanisms of resistance to immunotherapies in mccRCC,we have revisited the known resistance mechanisms of immunotherapies in metastatic clear-cell RCC and explored ongoing and future strategies to overcome them.

Adoptive therapy with CAR redirected T cells for hematological malignancies

The survival of patients with hematological malignancies has been significantly improved due to the development of new therapeutic agents. However, relapse remains a major matter for concern. Recently, T cells engineered with chimeric antigen receptor(CAR) were reported to show unprecedented responses in a range of hematological malignancies. The persistence of the CAR-T cell can last for years and tends toward long-term antitumor memory by which relapses can be effectively prevented. The primary side effects that appear in most clinical trials are cytokine release syndrome and neurotoxicity. However, these symptoms can be treated and reversed. In this review, we describe CAR structure and function and summarize recent advances in CAR-T cell therapy in hematological malignancies.

Study on Cellular Differentiation and Genes Rearrangement of Malignant Lymphoproliferative Disorders

Malignant lymphoproliferative disorders are common diseases characterized by the clonal proliferation of cells derived from the lymphocytes of different developmental stages. The analyses of routine morphology and cytochemistry fail to determine the origin and differentiation stages. The cell differentiation origin of most of malignant lymphoproliferative disorders can be defined by using monoclonal antibodies(McAbs) against the differentiation antigens of lymphocytes, but it is unable to determine the cell origin of a minority of undifferentiated malignancies and also fails to distinguish the malig-