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Epstein‒Barr virus (EBV) is a human herpes virus that is saliva-transmissible and universally asymptomatic. It has been confirmed that more than 90% of the population is latently infected with EBV for life. EBV can cause a variety of related cancers, such as nasopharyngeal carcinoma, diffuse large B-cell lymphoma, and Burkitt lymphoma. Currently, many clinical studies have demonstrated that EBV-specific cytotoxic T lymphocytes and other cell therapies can be safely and effectively transfused to prevent and treat some diseases caused by EBV. This review will mainly focus on discussing EBV-specific cytotoxic T lymphocytes and will touch on therapeutic EBV vaccines and chimeric antigen receptor T-cell therapy briefly. [ABSTRACT FROM AUTHOR]

Chimeric antigen receptor (CAR) T cells have demonstrated remarkable efficacy against hematological malignancies; however, they have not experienced the same success against solid tumors such as glioblastoma (GBM). There is a growing need for high-throughput functional screening platforms to measure CAR T-cell potency against solid tumor cells. We used real-time, label-free cellular impedance sensing to evaluate the potency of anti-disialoganglioside (GD2) targeting CAR T-cell products against GD2+ patient-derived GBM stem cells over a period of 2 days and 7 days in vitro. We compared CAR T products using two different modes of gene transfer: retroviral transduction and virus-free CRISPR-editing. Endpoint flow cytometry, cytokine analysis and metabolomics data were acquired and integrated to create a predictive model of CAR T-cell potency. Results indicated faster cytolysis by virus-free CRISPR-edited CAR T cells compared with retrovirally transduced CAR T cells, accompanied by increased inflammatory cytokine release, CD8+ CAR T-cell presence in co-culture conditions and CAR T-cell infiltration into three-dimensional GBM spheroids. Computational modeling identified increased tumor necrosis factor α concentrations with decreased glutamine, lactate and formate as being most predictive of short-term (2 days) and long-term (7 days) CAR T cell potency against GBM stem cells. These studies establish impedance sensing as a high-throughput, label-free assay for preclinical potency testing of CAR T cells against solid tumors. [ABSTRACT FROM AUTHOR]

This prospective clinical study aimed to determine the efficacy and prognostic factors of adoptive activated αβT lymphocyte immunotherapy for various refractory cancers. The primary endpoint was overall survival (OS), and the secondary endpoint was radiological response. The authors treated 96 patients. Activated αβT lymphocytes were infused every 2 weeks for a total of six times. Prognostic factors were identified by analyzing clinical and laboratory data obtained before therapy. Median survival time (MST) was 150 days (95% confidence interval, 105–191), and approximately 20% of patients achieved disease control (complete response + partial response + stable disease). According to the multivariate Cox proportional hazards model with Akaike information criterion–best subset selection, sex, concurrent therapy, neutrophil/lymphocyte ratio, albumin, lactate dehydrogenase, CD4:CD8 ratio and T helper (Th)1:Th2 ratio were strong prognostic factors. Using parameter estimates of the Cox analysis, the authors developed a response scoring system. The authors then determined the threshold of the response score between responders and non-responders. This threshold was able to significantly differentiate OS of responders from that of non-responders. MST of responders was longer than that of non-responders (317.5 days versus 74 days). The validity of this response scoring system was then confirmed by internal validation. Adoptive activated αβT lymphocyte immunotherapy has clinical efficacy in certain patients. The authors' scoring system is the first prognostic model reported for this therapy, and it is useful for selecting patients who might obtain a better prognosis through this modality. [ABSTRACT FROM AUTHOR]

The overuse and misuse of the macrophage 'M1–M2 paradigm' and the terms 'inflammatory' and 'anti-inflammatory' belie accurate description of macrophages and have hindered experimental design. The precise description of derived and isolated macrophages, including source, growth factors, stimuli, and culture conditions, is essential for developing macrophage-based immunotherapies. In vitro and in vivo studies are providing abundant proof-of-principle of the potential to harness macrophage activation to treat certain immune-related diseases. New advances, including derivation of macrophages from iPSCs, chimeric antigen receptor (CAR) macrophages, and Phase 1 clinical trials of ex vivo -derived macrophages might enable novel macrophage-based immunotherapies. While recognizing the heterogeneity of macrophages as well as the need for consistent nomenclature and characterization of macrophage sub-types, there is significant potential for harnessing macrophages for putative adoptive transfer therapies in various diseases. As an essential component of immunity, macrophages have key roles in mammalian host defense, tissue homeostasis, and repair, as well as in disease pathogenesis and pathophysiology. A source of fascination and extensive research, in this Opinion we challenge the utility of the M1–M2 paradigm, and discuss the importance of accurate characterization of human macrophages. We comment on the application of single cell analytics to define macrophage subpopulations and how this could advance therapeutic options. We argue that human macrophage cell therapy can be used to alleviate many diseases, and offer a viewpoint on the knowledge gaps that must be filled to render such a therapeutic approach a reality and, ideally, a common future practice in precision medicine. [ABSTRACT FROM AUTHOR]

• The superior antitumor function of fourth-generation CAR T compared to second-generation. • We developed a fourth-generation CAR targeting Trop2 (referred to as anti-Trop2 CAR4), which includes three co-stimulatory domains: CD28, 4-1BB, and CD27. • Anti-Trop2 CAR4-T cells exhibited potent cytotoxicity against breast cancer cells in both 2D and 3D cell cultures. • Anti-Trop2 CAR4-T cell proliferation and cytokine secretion were enhanced when exposed to Trop2-expressing cells. • Anti-Trop2 CAR4-T cells hold promise for improving the effectiveness of breast cancer treatment. The treatment of breast cancer (BC) remains a formidable challenge due to the emergence of drug resistance, necessitating the exploration of innovative strategies. Chimeric antigen receptor (CAR)-T cell therapy, a groundbreaking approach in hematologic malignancies, is actively under investigation for its potential application in solid tumors, including BC. Trophoblast cell surface antigen 2 (Trop2) has emerged as a promising immunotherapeutic target in various cancers and is notably overexpressed in BC. To enhance therapeutic efficacy in BC, a fourth-generation CAR (CAR4) construct was developed. This CAR4 design incorporates an anti-Trop2 single-chain variable fragment (scFv) fused with three costimulatory domains –CD28/4-1BB/CD27, and CD3ζ. Comparative analysis with the conventional second-generation CAR (CAR2; 28ζ) revealed that anti-Trop2 CAR4 T cells exhibited heightened cytotoxicity and interferon-gamma (IFN-γ) production against Trop2-expressing MCF-7 cells. Notably, anti-Trop2 CAR4-T cells demonstrated superior long-term cytotoxic functionality and proliferative capacity. Crucially, anti-Trop2 CAR4-T cells displayed specific cytotoxicity against Trop2-positive BC cells (MDA-MB-231, HCC70, and MCF-7) in both two-dimensional (2D) and three-dimensional (3D) culture systems. Following antigen-specific killing, these cells markedly secreted interleukin-2 (IL-2), tumor necrosis factor-alpha (TNF-α), IFN-γ, and Granzyme B compared to non-transduced T cells. This study highlights the therapeutic potential of anti-Trop2 CAR4-T cells in adoptive T cell therapy for BC, offering significant promise for the advancement of BC treatment strategies. [ABSTRACT FROM AUTHOR]

The acidic metabolic byproducts within the tumor microenvironment (TME) hinder T cell effector functions. However, their effects on T cell infiltration remain largely unexplored. Leveraging the comprehensive The Cancer Genome Atlas dataset, we pinpoint 16 genes that correlate with extracellular acidification and establish a metric known as the "tumor acidity (TuAci) score" for individual patients. We consistently observe a negative association between the TuAci score and T lymphocyte score (T score) across various human cancer types. Mechanistically, extracellular acidification significantly impedes T cell motility by suppressing podosome formation. This phenomenon can be attributed to the reduced expression of methyltransferase-like 3 (METTL3) and the modification of RNA N 6-methyladenosine (m6A), resulting in a subsequent decrease in the expression of integrin β1 (ITGB1). Importantly, enforced ITGB1 expression leads to enhanced T cell infiltration and improved antitumor activity. Our study suggests that modulating METTL3 activity or boosting ITGB1 expression could augment T cell infiltration within the acidic TME, thereby improving the efficacy of cell therapy. [Display omitted] • Tumor acidic score correlates negatively with CD8 infiltration score in human cancers • Extracellular acidification impedes T cell infiltration • Extracellular acidification hinders METTL3-mediated m6A modification in integrin β1 • Modification of CAR-T cells with integrin β1 improves their antitumor activity Wang et al. have identified a correlation between extracellular acidification and limited infiltration of T cells into solid tumors. Elevated extracellular acidification impairs T cell podosome formation by suppressing METTL3-mediated m6A modification, which in turn regulates integrin β1 expression. Integrin β1-modified T cells demonstrated increased infiltration and improved antitumor activities. [ABSTRACT FROM AUTHOR]

Highlights • Cytokine release syndrome (CRS) and neurotoxicity are common after immune effector cell (IEC) therapy. • Neurotoxicity is now termed IEC-associated neurotoxicity syndrome (ICANS). • A consensus grading system for CRS and ICANS has been developed by experts in the field. • The grading system is designed for IEC therapies in clinical trials and commercial use. • Standardized reporting through the Center for International Blood and Marrow Transplant Research will meet commercial Risk Evaluation and Mitigation Strategies requirements. ABSTRACT Chimeric antigen receptor (CAR) T cell therapy is rapidly emerging as one of the most promising therapies for hematologic malignancies. Two CAR T products were recently approved in the United States and Europe for the treatment ofpatients up to age 25years with relapsed or refractory B cell acute lymphoblastic leukemia and/or adults with large B cell lymphoma. Many more CAR T products, as well as other immunotherapies, including various immune cell- and bi-specific antibody-based approaches that function by activation of immune effector cells, are in clinical development for both hematologic and solid tumor malignancies. These therapies are associated with unique toxicities of cytokine release syndrome (CRS) and neurologic toxicity. The assessment and grading of these toxicities vary considerably across clinical trials and across institutions, making it difficult to compare the safety of different products and hindering the ability to develop optimal strategies for management of these toxicities. Moreover, some aspects of these grading systems can be challenging to implement across centers. Therefore, in an effort to harmonize the definitions and grading systems for CRS and neurotoxicity, experts from all aspects of the field met on June 20 and 21, 2018, at a meeting supported by the American Society for Transplantation and Cellular Therapy (ASTCT; formerly American Society for Blood and Marrow Transplantation, ASBMT) in Arlington, VA. Here we report the consensus recommendations of that group and propose new definitions and grading for CRS and neurotoxicity that are objective, easy to apply, and ultimately more accurately categorize the severity of these toxicities. The goal is to provide a uniform consensus grading system for CRS and neurotoxicity associated with immune effector cell therapies, for use across clinical trials and in the postapproval clinical setting. [ABSTRACT FROM AUTHOR]

• Targeting TGFβR2 and TIM3 signaling pathways in the tumor microenvironment of prostate cancer shows potential for enhancing the efficacy of PSMA-CAR-T cell therapy. • Understanding and effectively targeting immunosuppressive pathways that are specific to the tumor microenvironment not only offers potential for the treatment of advanced cancers but is also essential for the success of advanced cellular immunotherapy. • This study provides insights into the therapeutic potential of targeting immune checkpoint molecules in the context of prostate cancer. The immune checkpoint molecules, Transforming growth factor beta receptor II (TGFβRII) and T cell immunoglobulin and mucin domain 3 (TIM3), have been identified as contributors to T cell immune suppression in prostate cancer. The objective of this investigation was to improve the tumor killing capability of prostate-specific membrane antigen (PSMA)-chimeric antigen receptor T (CAR-T) cells by targeting TIM3 and TGFβRII simultaneously. To generate dnTGFβRII-trTIM3-PSMA-CAR-T (DT-PSMA-CAR-T) cells, the surface of PSMA-CAR-T cells was overexpressed with dominant negative TGFβRII (dnTGFβRII) and truncated extracellular TIM3 (trTIM3). The efficacy of DT-PSMA-CAR-T cells was assessed through in vitro killing experiments and animal experiments. The DT-PSMA-CAR-T cells demonstrated the ability to eradicate PSMA-positive prostate cancer cells, even in the presence of exogenous TGF-β and/or TIM3 activating antibodies. In addition, the cells demonstrated the ability to eliminate tumor tissue in an immunodeficient mouse model transplanted with GAL9-PSMA-PC3 cells in vitro, prolonging survival without significant toxic side effects. This study emphasizes that upregulating dnTGFβRII and trTIM3 on the surface of T cells can potentially diminish the inhibitory effects of TGFβRII and TIM3. [ABSTRACT FROM AUTHOR]

Hematopoietic stem cell transplantation (HSCT) is a potentially curative treatment for many pediatric diseases. Cellular immunotherapy, such as the use of chimeric antigen receptor T cells, has emerged recently as a highly effective modality to treat refractory lymphoid malignancies. Although HSCT and cellular immunotherapy are now considered essential parts of the current therapeutic armamentarium, both are associated with a wide range of acute complications, many of which are unique to these treatment modalities. These acute complications are seen not only in hospitalized patients but also in patients in the ambulatory setting, including the emergency department. Emergency care providers will need to be able to recognize these complications and provide appropriate initial assessment and management. In this review, we describe the acute complications associated with HSCT and cellular immunotherapy, with the emphasis on their recognition, workup, and treatment. [ABSTRACT FROM AUTHOR]

Background Adoptive transfer of donor-derived T cells can be applied to improve immune reconstitution in immune-compromised patients after allogeneic stem cell transplantation. The separation of beneficial T cells from potentially harmful T cells can be achieved by using the major histocompatibility complex (MHC) I- Strep tamer isolation technology, which has proven its feasibility for the fast and pure isolation of T-cell populations with a single specificity. We have analyzed the feasibility of the simultaneous isolation of multiple antigen-specific T-cell populations in one procedure by combining different MHC I- Strep tamers. Methods First, the effect of combining different amounts of MHC I- Strep tamers used in the isolation procedure on the isolation efficacy of target antigen-specific T cells and on the number of off-target co-isolated contaminating cells was assessed. The feasibility of this approach was demonstrated in large-scale validation procedures targeting both high and low frequent T-cell populations using the Good Manufacturing Practice (GMP)-compliant CliniMACS Plus device. Results T-cell products targeting up to 24 different T-cell populations could be isolated in one, simultaneous MHC I- Strep tamer procedure, by adjusting the amount of MHC I- Strep tamers per target antigen-specific T-cell population. Concurrently, the co-isolation of potentially harmful contaminating T cells remained below our safety limit. This technology allows the reproducible isolation of high and low frequent T-cell populations. However, the expected therapeutic relevance of direct clinical application without in vitro expansion of these low frequent T-cell populations is questionable. Discussion This study provides a feasible, fast and safe method for the generation of highly personalized MHC I- Strep tamer isolated T-cell products for adoptive immunotherapy. [ABSTRACT FROM AUTHOR]

Rapid restoration of virus-specific T immunity via adoptive transfer of ex vivo generated T cells has been proven as a powerful therapy for patients with advanced cancers and refractory viral infections such as cytomegalovirus (CMV) and Epstein-Barr virus (EBV). BK virus (BKV), John Cunningham virus (JCV), and Merkel cell carcinoma virus (MCV) are the members of the rapidly growing human polyomavirus (hPyV) family that commonly infects most healthy humans. These viruses have a clearly established potential for causing severe end-organ damage or malignant transformation, especially in individuals with weakened immunity who are unable to mount or regain endogenous T-cell responses as a result of underlying leukemia or iatrogenic immunosuppression in autoimmunity, bone marrow and solid organ transplant settings. Here we will discuss recent advances in using T-cell–based immunotherapies to save patients suffering from PyV-associated diseases including hemorrhagic cystitis, BKV virus–associated nephropathy, and JC-associated progressive multifocal leukoencephalopathy (PML). We will also review progress in the understanding of Merkel cell carcinoma (MCC) as a virally driven tumor that is amenable to immune intervention and can be targeted with adoptively transferred T cells specific for viral oncoproteins. [ABSTRACT FROM AUTHOR]

Blood transfusion is a clinical replacement therapy with many successes with some benefit and, also, some harm. Cancer is a multifaceted disease potentially associated with the immune system's weakness where the cancerous tumor cells escape from the immune system. Allogeneic blood transfusion, through five major mechanisms including the lymphocyte-T set, myeloid-derived suppressor cells (MDSCs), tumor-associated macrophages (TAMs), natural killer cells (NKCs), and dendritic cells (DCs) can help the recipient's defense mechanisms. On the other hand, the role for each of the listed items includes activation of the antitumor CD8+ cytotoxic T lymphocytes (CD8+/CTL), temporal inactivation of Tregs, inactivation of the STAT3 signaling pathway, the use of bacteria to enhance the antitumor immune response and cellular immunotherapy. The above issues are concisely addressed in this manuscript based on a literature survey on this topic carried out by the first author. [ABSTRACT FROM AUTHOR]

Background aims. Despite the availability of multiple treatment strategies, patients with gastric carcinoma (GC) have a dismal prognosis. The aim of this study was to evaluate the efficacy and safety of cellular immunotherapy (CIT) with the use of autologous natural killer cells, y5T cells and cytokine-induced killer cells in combination with chemotherapy in patients with GC. Methods. In this open-label pilot cohort study, patients were treated with the combination therapy (chemo/CIT group) or chemotherapy alone (control group). Progression-free survival (PFS), overall survival (OS), quality of life (QOL) and adverse events were investigated. Results. Fifty-eight patients were analyzed, 30 in the chemo/CIT group and 28 in the control group. The median PFS of the chemo/CIT group was significantly longer compared with the control group (P = 0.021). In subgroup analysis, in patients with stage III GC, node-positive metastasis or poorly differentiated carcinoma, the 2-year PFS rate in chemo/CIT versus control groups was 62.5% versus 26.7% (P = 0.022), 50% versus 27.3% (P = 0.016) and 56.3% versus 28.6% (P = 0.005), respectively. The median OS in either group has not yet been reached, and there was no significant difference in OS between the groups. The QOL was improved in the patients treated with chemo/CIT compared with the control group. CIT was well tolerated and not related to any significant adverse events. Conclusions. A combination of CIT and chemotherapy for patients with GC was safe, improved QOL, and might prevent recurrence, especially in GC patients with advanced stage, poorly differentiated carcinoma or lymph node metastasis. [ABSTRACT FROM AUTHOR]

Background aims. Natural killer (NK) cell-based immunotherapy is a promising treatment for a variety of malignancies. However, generating sufficient cell numbers for therapy remains a challenge. T o achieve this, optimization of protocols is required. Methods. Mature NK cells were expanded from peripheral blood mononuclear cells PBMCs in the presence of anti-CD3 monoclonal antibody and interleukin-2. Additionally, NK-cell progenitors were generated from CD34+ hematopoietic stem cells or different T/NK-cell progenitor populations. Generated NK cells were extensively phenotyped, and functionality was determined by means of cytotoxicity assay. Results. Addition of ascorbic acid (AA) resulted in more proliferation of NK cells without influencing NK-cell functionality. In more detail, PBMC-derived NK cells expanded 2362-fold (median, range: 90-31,351) in the presence of AA and were capable of killing tumor cells under normoxia and hypoxia. Moreover, hematopoietic stem cell--derived progenitors appeared to mature faster in the presence of AA, which was also observed in the NK-cell differentiation from early T/NK-cell progenitors. Conclusions. Mature NK cells proliferate faster in the presence of phospho-L-AA, resulting in higher cell numbers with accurate functional capacity, which is required for adoptive immunotherapy. [ABSTRACT FROM AUTHOR]

Passive immunotherapy approaches in COVID-19 treatment. [Display omitted] • Challenges regarding the SARS-CoV-2 vaccination have remained unanswered. • At this time, there is no specific treatment for COVID-19-induced cytokine storm. • Passive immunotherapy is a promising way to manage COVID-19. • Inflammatory molecules are appropriate targets to alleviate COVID-19. In late 2019, severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) emerged, causing a global pandemic called COVID-19. Currently, there is no definitive treatment for this emerging disease. Global efforts resulted in developing multiple platforms of COVID-19 vaccines, but their efficacy in humans should be wholly investigated in the long-term clinical and epidemiological follow-ups. Despite the international efforts, COVID-19 vaccination accompanies challenges, including financial and political obstacles, serious adverse effects (AEs), the impossibility of using vaccines in certain groups of people in the community, and viral evasion due to emerging novel variants of SARS-CoV-2 in many countries. For these reasons, passive immunotherapy has been considered a complementary remedy and a promising way to manage COVID-19. These approaches are based on reduced inflammation due to inhibiting cytokine storm phenomena, immunomodulation, preventing acute respiratory distress syndrome (ARDS), viral neutralization, and decreased viral load. This article highlights passive immunotherapy and immunomodulation approaches in managing and treating COVID-19 patients and discusses relevant clinical trials (CTs). [ABSTRACT FROM AUTHOR]

Background aims. Natural killer (NK) cells, either naive or genetically engineered, are increasingly considered for cellular therapy of patients with malignancies. When using NK cells from peripheral blood, the number of expanded NK cells can be highly variable and the need for NK cell enrichment can make the process expensive. The NK-92 cell line (CD56+/CD3-) that was isolated from a patient with lymphoma has predictable high cytotoxic activity and can be expanded under good manufacturing practice conditions in recombinant interleukin-2. Methods. Fifteen patients (age, 9-71 years) with advanced, treatment-resistant malignancies, either solid tumors/sarcomas (n = 13) or leukemia/lymphoma (n = 2), received two infusions of NK-92 cells, given 48 h apart. Three cohorts of patients were treated with escalating doses of NK-92 cells (n = 7 at 1 × 109, n = 6 at 3 × 109 and n = 2 at 1 × 1010 cells/m2). Results. No infusion-related or long-term side effects were observed. The dose of 1010 cells/m2 was considered the maximum expandable cell dose with the use of an established culture bag system. Three fourths of patients with lung cancer had some anti-tumor response. Only one patient of seven had development of human leukocyte antigen antibodies. The persistence of NK-92 cells (male origin) in die circulation was confirmed by Y chromosome-specific polymerase chain reaction in two female patients. Conclusions. Infusions of NK-92 cells up to 1010 cells/m2 were well tolerated. Despite the allogeneic nature of NK-92, development of human leukocyte antigen antibodies in these patients with cancer appears to be rare. The cells can persist in the recipient's circulation for at least 48 h. Some encouraging responses were seen in patients with advanced lung cancer. [ABSTRACT FROM AUTHOR]

Abstract: Human telomerase reverse transcriptase (hTERT) has been identified as an ideal tumor-associated antigen (TAA). Use of a synthetic hTERT epitope peptide to pulse dendritic cells can induce autologous T cell anti-tumor immune responses, but such responses induced by a single epitope peptide have been shown to be weak and a narrow-spectrum. Here, we designed dendritic tandem multiple antigenic peptides (MAPs) containing the following three hTERT epitope peptides: I540, V461 and L766, which are HLA-A*02-, HLA-A*24- and HLA-RDB1*04/11/15-restricted, respectively. The MAPs and their three single-epitope peptides were obtained through solid-phase synthesis. Healthy volunteers that were HLA-A*02+/HLA-DRB1*04+ and HLA-A*24+/HLA-DRB1*15+ were recruited. Myeloid dendritic cells were isolated by magnetic activated cell sorting and were divided into a MAP-stimulated group (MAP-DC), a group in which the three epitope peptides were mixed and used to stimulate the DCs (MixP-DC) and a no peptide-stimulated group (NoP-DC, control group). All of the DCs were cultured in serum-free medium, pulsed with the corresponding peptides on the 3rd, 5th and 7th days, and co-cultured with autologous lymphocytes when they were mature. The related cytokines were measured via ELISA. The killing effects of cytotoxic T lymphocytes (CTLs) on SW480/A549 tumor cells expressing HLA-A*02+, HepG2/SMMC-7721 cells expressing HLA-A*24+ and SKOV3 cells negative for HLA-A*02/A*24 were detected by flow cytometry. Our results indicated that the CTLs induced by the MAP-DCs had the greatest anti-tumor effect. Therefore, the dendritic tandem multiple antigenic hTERT epitope peptides combined with MDCs may represent a powerful, broad-spectrum anti-tumor vaccine. [Copyright &y& Elsevier]

Abstract: Establishment of a leukemia plasmacytoid dendritic cell line (PMDC05) and intra-lineage transformation from pDCs to mDCs in PMDC05 has been reported. In this paper, we show the applicability of PMDC05 for cellular immunotherapy. By stimulation with LPS, PMDC05 showed enhancement in expression of antigen presentation-associated surface molecules and production of cytokines (IL-12p70 and TNF-α). The antigen presenting ability was markedly increased in PMDC05 stimulated with LPS. By co-culturing of CD8+ T cells with LPS-stimulated and WT1/CMVpp65 peptide-pulsed PMDC05, WT1/CMVpp65 tetramer+ cytotoxic T lymphocytes were efficiently generated. These findings reveal the applicability of PMDC05 in cellular immunotherapy for tumor and severe viral infections. [Copyright &y& Elsevier]

Immunotherapeutic ablation of lymphoma is a conceptually attractive treatment strategy that is the subject of intense translational research. Cytotoxic T lymphocytes (CTLs) that are genetically modified to express CD19- or CD20-specific, single-chain antibody–derived chimeric antigen receptors (CARs) display HLA-independent antigen-specific recognition/killing of lymphoma targets. Here, we describe our initial experience in applying CAR-redirected autologous CTL adoptive therapy to patients with recurrent lymphoma. Using plasmid vector electrotransfer/drug selection systems, cloned and polyclonal CAR+ CTLs were generated from autologous peripheral blood mononuclear cells and expanded in vitro to cell numbers sufficient for clinical use. In 2 FDA-authorized trials, patients with recurrent diffuse large cell lymphoma were treated with cloned CD8+ CTLs expressing a CD20-specific CAR (along with NeoR) after autologous hematopoietic stem cell transplantation, and patients with refractory follicular lymphoma were treated with polyclonal T cell preparations expressing a CD19-specific CAR (along with HyTK, a fusion of hygromycin resistance and HSV-1 thymidine kinase suicide genes) and low-dose s.c. recombinant human interleukin-2. A total of 15 infusions were administered (5 at 108cells/m2, 7 at 109cells/m2, and 3 at 2 × 109cells/m2) to 4 patients. Overt toxicities attributable to CTL administration were not observed; however, detection of transferred CTLs in the circulation, as measured by quantitative polymerase chain reaction, was short (24 hours to 7 days), and cellular antitransgene immune rejection responses were noted in 2 patients. These studies reveal the primary barrier to therapeutic efficacy is limited persistence, and provide the rationale to prospectively define T cell populations intrinsically programmed for survival after adoptive transfer and to modulate the immune status of recipients to prevent/delay antitransgene rejection responses. [Copyright &y& Elsevier]

Abstract: Dendritic cells (DCs) are the most potent antigen presenting cells of the immune system as they can act as initiators, stimulators and regulators of the immune response. Human DCs are most commonly generated for clinical use by in vitro differentiation of monocytes with exogenous cytokines. Here, we investigate the effect of LCOS 1013 on the production of mature Mo-DCs. LCOS 1013 is a new bacterial component from walls of gram+ Klebsellia pneumoniae bacteria that contain some OmpA glycoproteins. Purified peripheral blood monocytes were cultured for 6 days with IL-4 and GM-CSF in order to obtain immature dendritic cells (Im-MoDCs). On day six, Im-MoDCs were matured with either LCOS 1013, TNF alpha, LPS or CD40-Ligand. LCOS 1013 matured Mo-DCs (LCO-DCs) showed a higher expression of DC-LAMP, CD80, CD83, CD54 and CD40 than TNF alpha, LPS and CD40L matured Mo-DCs. Interestingly, LCO-DCs exhibited high expression of full competent CCR7 and high secretion of IL-12 during their maturation. Functionally, LCO-DCs have equivalent potency to trigger mixed leukocyte reaction and antigen-specific reaction and polarize immune response towards Th1 way. Moreover, we found that LCOS 1013 activates DCs through TLR2. LCOS 1013 represents an attractive therapeutic maturation agent of DCs allowing the production of Mo-DCs with high capacity to migrate and to induced Th1 immune responses. [Copyright &y& Elsevier]

Abstract: Introduction: Ewing sarcoma (EWS) is a malignant bone-associated sarcoma, with poor prognosis in case of metastasis or relapse. To explore the feasibility of natural killer (NK) cell mediated immunotherapy and to identify molecular mechanisms involved, the susceptibility of EWS to NK cells was investigated. Methods and results: All EWS cell lines tested (n =7) were lysed by purified allogeneic NK cells from healthy donors, and the efficacy of lysis was increased by activating NK cells with interleukin-15 (IL-15). FACS analysis and immunohistochemistry revealed that EWS cell lines as well as primary tumor cells expressed ligands for the activating NK cell receptors NKG2D and DNAM-1. NK cell cytotoxicity to EWS cells critically depended on the combination of NKG2D and DNAM-1 signaling, since blocking either of these receptors abrogated lysis by resting NK cells. Cytokine-activated NK cells more efficiently recognized EWS cells, since only combined, but not single blockade of NKG2D and DNAM-1 by antibodies inhibited lysis of EWS cells. Induction or blockade of HLA class I on EWS cells did not significantly influence lysis. This suggests that predominantly activating, rather than inhibitory signals on EWS cells determined susceptibility to NK cell cytotoxicity. NK cell cytotoxicity to EWS cells and K562 was reduced in EWS patients at diagnosis (n =11) compared to age matched controls, despite normal NK cell numbers and increased expression of NKG2D. The impaired function of these NK cells was restored after activation with IL-15 in vitro. Conclusion: These results demonstrate that EWS cells are potentially susceptible to NK cell cytotoxicity due to the expression of activating NK cell receptor ligands. The use of cytokine-activated NK cells rather than resting NK cells in immunotherapy may be instrumental to optimize NK cell reactivity to EWS. [Copyright &y& Elsevier]

Recovery of a fully functional immune system is a slow and often incomplete process following allogeneic stem cell transplantation. While innate immunity reconstitutes quickly, adaptive B- and especially T-cell lymphopoeisis may be compromised for years following transplantation. In large part, these immune system deficits are due to the decrease, or even absence, of thymopoiesis following transplantation. Thereby, T-cell reconstitution initially relies upon expansion of mature donor T cells; a proliferation driven by high cytokine levels and the presence of allo-reactive antigens. This peripheral mechanism of T-cell generation may have important clinical consequences. By expanding tumouricidal T cells, it may provide a venue to enhance T-cellular immunotherapy following transplantation. Alternatively, decreased thymic function may impair long-term anti-tumour immunity and increase the likelihood of graft-versus-host disease. [Copyright &y& Elsevier]

Multiple myeloma is a life-threatening haematological malignancy for which standard therapy is inadequate. Autologous stem cell transplantation results in effective cytoreduction, but patients subsequently relapse from sites of chemotherapy-resistant disease. Allogeneic transplantation may result in durable responses due to anti-tumour immunity mediated by donor lymphocytes. However, morbidity and mortality related to graft-vs-host disease remains a challenge. A promising area of investigation is the development of immunotherapeutic approaches that target and eliminate myeloma cells more selectively. A variety of tumour-associated antigens have been identified in myeloma cells that may be targeted selectively by host immunity. Vaccination strategies targeting single antigens and whole-cell approaches, which have the advantage of presenting patient-specific and potentially unidentified antigens to immune effector cells, have shown promise in clinical studies. In addition, the use of monoclonal antibodies has been evaluated in preclinical and clinical studies. Effector cell dysfunction and the increased number of regulatory T cells in patients with malignancy may limit the efficacy of immunotherapeutic approaches. Strategies to improve upon immunotherapy for multiple myeloma involve the depletion of T regulatory cells, combining active and passive immunotherapy, the use of cytokine adjuvants, and using immunotherapy in conjunction with autologous and allogeneic transplantation. [Copyright &y& Elsevier]

Abstract: Prostate cancer is currently the most commonly diagnosed malignancy, and the second leading cause of cancer-related death, in men in the United States. Most deaths from prostate cancer are due to metastatic disease that no longer responds to androgen deprivation. There have been several advances within the last decade leading to the approval of new agents for treating patients with castration-resistant prostate cancer. However, these agents can have significant side effects, and the average survival benefit from chemotherapy has been modest. Results from several clinical trials have suggested that immune-based therapies may have clinical benefits in patients with prostate cancer with significantly fewer adverse events. These new therapies, however, pose new potential benefits and risks. We review here two cellular immunotherapies furthest in clinical development for prostate cancer, and discuss the potential challenges to the treating oncologist with the possible advent of these new therapies. [Copyright &y& Elsevier]

Abstract: New approaches to therapeutics of advanced prostate cancer are urgently needed. GVAX® (granulocyte-macrophage colony-stimulating factor [GM-CSF] gene transduced irradiated prostate cancer vaccine cells) offers the possibility that “host versus prostate cancer” immune responses can be generated in prostate cancer patients. Critical components involve the dendritic cell loading of candidate prostate cancer lymph node metastasis and candidate bone metastasis antigens derived from irradiated prostate cancer whole cells. GM-CSF acts at the vaccination site to enhance activation dendritic cells and antigen presentation to both the B-cell and T-cell arms of the immune system. GVAX® preclinically—in both rat and transgenic prostate cancer models—has antitumor activity which has informed early clinical trial designs. Clinical investigations reviewed in this report suggest that vaccination is safe and immune tolerance can be broken against prostate cancer. Multi-institutional phase III investigation is currently underway to evaluate the impact of allogeneic prostate GVAX® cellular immunotherapy on time to progression and overall survival in hormone refractory prostate cancer. [Copyright &y& Elsevier]

Cellular immunotherapy (CI), as we now know it, began in the early 1980s with the use of lymphokine-activated killer cells (LAK) and progressed to the use of the immunologically specific, tumor-infiltrating lymphocytes (TIL). TIL were shown to be particularly effective against melanoma and it was in these trials that we learned the importance of immunologic specificity for tumor. With the identification and characterization of tumor antigens recognized by TIL, we now see the use of these antigens in various forms constituting vaccines. Investigators are using tumor antigens alone or in combination with dendritic cells (DCs), the body’s most efficient and powerful antigen-presenting cell. Therapies are being delivered to many patients with different types of cancer in order to combat bulky disease, eliminate micro-metastatic disease, and provide a memory mechanism to fight tumor recurrence. This review will detail the past 18 years and present the developments that have been made in this therapy. Many believe that with continued development, immunotherapy will provide a fourth modality of cancer therapy. [Copyright &y& Elsevier]

Humoral immunotherapy using the monoclonal anti-CD20 antibody rituximab induces remissions in ∼60% of patients with relapsed follicular lymphoma; however, most patients eventually relapse despite continued expression of CD20 on lymphoma cells. We have hypothesized that cellular immunotherapy targeting CD20+ cells might provide a more effective mechanism for eliminating lymphoma cells than anti-CD20 antibodies and are therefore investigating the utility of cytotoxic T lymphocytes (CTL) genetically modified to target the CD20 antigen. Peripheral blood mononuclear cells were activated with anti-CD3 antibody (OKT3) and recombinant human interleukin-2 and electroporated with a plasmid containing a CD20-specific scFvFc:ζ chimeric T cell receptor gene and a neomycin phosphotransferase gene (neoR). Transfected cells were selected using the antibiotic G418 and cloned by limiting dilution. Using this approach, we have generated CD8+ CTL clones with CD20-specific cytotoxicity, which specifically lysed CD20+ target cells, including actual tumor cells from patients with follicular lymphoma, small lymphocytic lymphoma, splenic marginal zone lymphoma, diffuse large B cell lymphoma, and chronic lymphocytic leukemia. The CTL clones have been expanded to numbers sufficient for therapy (∼109 cells). Our data indicate the feasibility of generating and expanding CD20-specific CTL and, for the first time, demonstrate that such CTL exhibit specific cytotoxicity against actual tumor cells isolated from patients with a variety of B lymphoid malignancies. In view of these promising findings, a Phase I clinical trial for relapsed follicular lymphoma is being initiated. [Copyright &y& Elsevier]

• Regulatory macrophage (Mreg) have immunosuppressive and anti-inflammatory properties. • Mreg could delete activated T cells, induce Tregs and secrete effector molecules. • Mreg act as a negative immunoregulatory in transplantation. • Mreg combined with low-dose immunosuppressants may induce transplant immune tolerance. Regulatory macrophages (Mregs) are a subtype of macrophages that are involved in regulating immune responses and inhibiting activated T lymphocyte proliferation. With advances in our basic understanding of Mregs and the revelation of their biological characteristics, Mregs have become a focus of research. In addition to promoting malignant tumor progression, Mregs also play an immunosuppressive role in inflammatory diseases and transplantation. Recent studies have shown that Mregs are closely associated with the induction of transplantation immune tolerance. Immune regulatory cell treatment as an adjunct immunosuppressive therapy offers new insights into the mechanism by which transplantation immune tolerance is established. The application of Mreg-based cellular immunotherapy has shown promise in clinical solid organ transplantation. Here, we provide a comprehensive overview of Mreg morphology, phenotype, induction and negative immunoregulatory function and discuss the role of Mregs in different transplantation models as well as their potential application value in clinical organ transplantation. [ABSTRACT FROM AUTHOR]

Breast cancer is the most common malignancy in women worldwide, with a relatively high proportion of patients experiencing resistance to standard treatments. Cellular immunotherapy (CI), which is based on the extraction, modification, and re-infusion of the patient's immune cells, is showing promising results in these patients. Among CI possible approaches, adoptive cell therapy (ACT) and dendritic cell (DC) vaccination are the most comprehensively explored in both primary/translational research studies and clinical trials. ACT may include the use of tumor-infiltrating lymphocytes (TILs), T cell receptor (TCR)-, or chimeric antigen receptor (CAR)-engineered T-cells. There are indications suggesting that a biomarker-based approach might be beneficial in effectively selecting breast cancer patients for CI. Here, we sought to provide the current knowledge of CI in breast cancer, focusing on candidate biomarkers, ongoing clinical trials, limitations, and immediate future perspectives. [ABSTRACT FROM AUTHOR]

• Combination of gemcitabine and immunotherapy was tested in cholangiocarcinoma (CCA). • Cytotoxic T-lymphocyte (CTL) activated by self-differentiated dendritic cell (SD-DC). • Pretreatment of chemo-resistant CCA with gemcitabine enhanced CTL killing ability. • Gemcitabine sensitized CCA cells via upregulation of HLA class I and Fas receptor. • This study supported a combination of chemotherapy and CTLs for chemo-resistant CCA. Cholangiocarcinoma (CCA) can resist chemotherapy resulting in treatment failure. Gemcitabine, a chemotherapeutic drug, can sensitize cancer cells to become susceptible to cytotoxic T-lymphocytes (CTLs). We, therefore, hypothesized that a combination of gemcitabine and CTLs would be more effective for CCA treatment than individual therapy. To test this hypothesis, we conducted an in vitro study using gemcitabine combined with CTLs to treat gemcitabine-resistant CCA (KKU-213) cells. KKU-213 cells were pretreated with gemcitabine and tested for killing by CTLs activated by dendritic cells that were prepared by three different methods, including: (i) monocyte-derived dendritic cells pulsed with cancer cell lysate (Mo-DC + Lys), (ii) self-differentiated dendritic cells pulsed with cancer-cell lysate (SD-DC + Lys), and (iii) SD-DC presenting tumor-associated antigen PRKAR1A (SD-DC-PR). KKU-213 cells pretreated with gemcitabine were killed by CTLs activated by either SD-DC + Lys or SD-DC-PR more efficiently than those activated by Mo-DC + Lys. Furthermore, KKU-213 cells pretreated with a low dose (2 µM) of gemcitabine significantly enhanced the cytotoxic activity of CTLs activated by either SD-DC + Lys or SD-DC-PR at all evaluated effector (E) to target cell (T) ratios. At an E:T ratio of 5:1, KKU-213 cells pretreated with gemcitabine enhanced the cytotoxic activity of CLTs by approximately 2.5-fold (greater than 50% cell death) compared to untreated condition. The upregulation of HLA class I upon pretreatment of KKU-213 cells with gemcitabine may suggest a mechanism that leads to alteration of the antigen presentation process to promote CTL functions. These findings support the concept of combination therapy for overcoming chemo-resistant CCA. [ABSTRACT FROM AUTHOR]