scholarly journals A Long-Acting Pharmacological Grade Interleukin-7 Molecule Logarithmically Accelerates Ucart Proliferation, Differentiation, and Tumor Killing

Blood ◽  
2018 ◽  
Vol 132 (Supplement 1) ◽  
pp. 2199-2199
Author(s):  
Matt L Cooper ◽  
Karl W. Staser ◽  
Julie Ritchey ◽  
Jessica Niswonger ◽  
Byung Ha Lee ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Cell Lymphoma ◽  
B Cell Lymphoma ◽  
Effector Memory ◽  
Photon Flux ◽  
Interleukin 7 ◽  
Tumor Killing ◽  
Long Acting

Abstract Background: Chimeric antigen receptor T cell (CART) therapy is revolutionizing modern cancer therapy, with two anti-CD19 CARTs FDA-approved for relapsed/refractory B cell lymphoma/leukemia and many other CARTs for solid and liquid tumors currently undergoing clinical trials. Our group recently demonstrated multiplexed CRISPR/Cas9 gene-editing of anti-CD7 CARTs to produce CD7 and T cell receptor alpha constant (TRAC)-deleted "off-the-shelf" universal (U)CART7s that effectively kill CD7+ T cell lymphoma in vivo without causing GVHD or fratricide (Cooper et al, Leukemia, 2018). However, in current clinical practice, suboptimal CART persistence and tumor killing permit tumor cell escape and, ultimately, disease relapse. Reasoning that a pro-lymphoid growth factor could promote CART efficacy, we supplemented UCART infusion with subcutaneous injections of the long-acting form of recombinant human interleukin-7 fused with hybrid Fc (rhIL-7-hyFc, NT-I7) in vivo using a CD19+ lymphoma xenograft model. Methods: To create anti-CD19 universal CARTs (UCART19), we activated human T cells on CD3/CD28 beads, electroporated the T cells with Cas9 mRNA and a TRAC-targeted gRNA, and virally transduced an anti-CD19 scFv 3rd generation CAR containing a peptidase 2A-cleaved human CD34 construct for both purification and tracking in vivo. Residual TRAC+ cells were depleted using magnetic selection. For xenograft tumor modeling in vivo, we injected NOD-scid IL2Rgammanull (NSG) mice with 5x105 RamosCBR-GFP cells four days prior to UCART19 (2x106 cells). Mice were treated with NT-I7 (10mg/kg SC) on days +1, +15 and +29 post UCART19 infusion. Results:RamosCBR-GFP mice receiving NT-I7 without UCART19 (NT-I7 only group) survived marginally longer (24 day med survival) than mice receiving RamosCBR-GFP cells alone (No tx group) (21 day medium survival, p=0.018, NT-I7 only vs. No Tx). While RamosCBR-GFP mice treated with UCART19 alone (UCART19 group) survived 33 days, 100% of RamosCBR-GFP mice treated with UCART19 and NT-I7 (UCART19+NT-I7 group) were alive at 80 days (Fig 1a), with no mouse showing signs of xenogeneic GVHD (p<0.0001, UCART19+NT-I7 vs. UCART19). At three weeks post UCART19 infusion, bioluminescent imaging (BLI) revealed minimal tumor signal in UCART19+NT-I7 treated mice (108 vs. 1010 photon flux/s, p<0.05, UCART19+NT-I7 vs. UCART19) and near-undetectable photon flux/s at four weeks (107 vs 1011 photon flux/s, p<0.0001, UCART19+NT-I7 vs. UCART19). Quantitative 17-parameter flow cytometric analyses of the blood, bone marrow, and spleens revealed an up to ~8000-fold increase in UCART19 cells in NT-I7-treated mice four weeks post UCART19 infusion (Fig 1a). These UCART19 cells demonstrated a predominantly effector and effector memory phenotype. Discussion: CARTs engineered to express interleukin-7 and CCL19 showed increased migration to and killing of solid tumors (Adachi et al, Nature Biotechnology, 2018). However, genetically engineered potentiation strategies lack "off-switches" and may preclude additional genetic enhancements required for universal "off-the-shelf" CART development. Here, we demonstrate that a pharmacological grade long-acting interleukin-7 agonist can potentiate adoptive cellular therapies. Specifically, NT-I7 can dramatically enhance gene modified T cell proliferation, persistence and tumor killing in vivo, resulting in enhanced survival, providing a tunable clinic-ready adjuvant for reversing suboptimal CART activity in vivo. Disclosures Cooper: WUGEN: Consultancy, Equity Ownership. Lee:NeoImmuneTech: Employment. Park:NeoImmuneTech: Employment.

Therapeutic Efficacy of Engineered T Cells Targeting CD19 in a B-Cell Lymphoma Model.

Blood ◽  
2006 ◽  
Vol 108 (11) ◽  
pp. 5486-5486
Author(s):  
Robert E. Hawkins ◽  
David E. Gilham ◽  
Fiona C. Thistlethwaite ◽  
John A. Radford ◽  
Eleanor J. Cheadle
Keyword(s):  
T Cells ◽  
T Cell ◽  
B Cell ◽  
Cell Lymphoma ◽  
B Cell Lymphoma ◽  
Phase I Clinical Trials ◽  
Specific Receptor ◽  
Cell Engraftment ◽  
Engineered T Cells

Abstract The success of monoclonal antibodies in the treatment of certain cancers demonstrates that immune based therapies can work and are particularly effective in B cell malignancies. However, tumours can still avoid antibody mediate mechanisms of attack and there is currently no estalbished method of effectively recruiting T cells to harness their potential anti-tumour effects. We are exploring gene therapy approaches to endow T cells with antibody type specificity in order to more efficiently target and lyse tumours and thereby improve the overall immune therapy of cancer T cells grafted with a CD19 specific receptor consisting of a CD19 scFv linked to human CD3zeta (CD19z) were tested for their potency against B cell lymphoma lines in vivo. T cells were engineered using retroviral vestors to possess a CD19 specific receptor which endows the T cells with specificity for B cell lymphoma. The vector incorporates a truncated hCD34 gene as a marker to facilitate assessment of transduced cells using as clinically applicable, non-immunogenic marker gene. Mice bearing B cell lymphoma were treated with a systemic infusion of targeted T cells with or without non-myeloablative chemotherapy. Human T cells targeting CD19 cured 40% of SCID/beige mice with 6 day established metastatic tumour but only in conjunction with a single dose of cyclophosphamide. Murine T cells expressing the CD19z receptor were also effective with cure of 24hr established s.c human CD19+ tumour in SCID/beige and immunocompetent mice. Pretreatment with cyclophosphamide did not affect T cell engraftment or efficacy in immuno-compromised animals but was necessary for T cell engraftment in immuno-competent animals. These results which parallel the approach successfully used with tumour infiltrating lymhocytes in melanoma patients conclusively demonstrate that the combination of engineered T cells with “pre-conditioning” chemotherapy significantly impacts upon tumour growth in vivo and this evidence supports the development of phase I clinical trials targeting B cell lymphoma.

Enforced Expression of Lin28b Drives Development of Peripheral T Cell Lymphoma In Vivo

Blood ◽  
2011 ◽  
Vol 118 (21) ◽  
pp. 1392-1392 ◽  
Author(s):  
Sarah H Beachy ◽  
Masahiro Onozawa ◽  
Yang Jo Chung ◽  
Christopher Slape ◽  
Sven Bilke ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Transgenic Mice ◽  
Cell Lymphoma ◽  
Regulatory Elements ◽  
T Cell Lymphoma ◽  
Effector Memory ◽  
Peripheral T Cell Lymphoma ◽  
Chronic Inflammatory Condition

Abstract Abstract 1392 Lin-28 was first identified as a heterochronic gene in C. elegans and is expressed at high levels during early larval stages with dramatically decreased expression in subsequent developmental stages. Interestingly, recent work demonstrated that induced expression of LIN28 can reprogram human fibroblasts to acquire pluripotency (in combination with NANOG, OCT4 and SOX2), providing additional evidence for a positive correlation between LIN28 expression and maintenance of a more immature and stem cell-like developmental state. Lin28a and Lin28b, the mammalian homologues of lin-28, have been implicated in oncogenic transformation in a variety of tumor types, in part by their ability to promote the degradation of the let-7 family of microRNAs (miRs), which are known to target oncogenes such as Myc and Ras. We recently noted that Lin28b was markedly overexpressed in hematopoietic tissues of NUP98-HOXD13 (NHD13) transgenic mice that develop a myelodysplastic syndrome (MDS) that subsequently transforms to acute myeloid leukemia (AML) or pre-T lymphoblastic leukemia/lymphoma (pre-T LBL). In order to elucidate the contribution of Lin28b overexpression to the differentiation block and malignant phenotype observed in the NHD13 mice, we designed a Lin28b transgenic mouse by targeting the expression of the transgene to hematopoietic tissues with Vav regulatory elements. In this model, clinically healthy Lin28b mice exhibited aberrant thymic architecture and retention of thymocytes that was correlated with peripheral blood lymphopenia (a 2.6-fold decrease in circulating lymphocytes). The lymphopenia was principally due to decreased numbers of CD4+ and CD8+ cells, although there was a significant increase in the number of CD4 and CD8 effector memory T cells (CD44hiCD62Llo) compared with wild type mice. Additionally, deep sequencing of thymic miRs from clinically healthy transgenic mice revealed a 2–5-fold downregulation of let-7 family members, including let-7d, g, f, i and miR-98. Importantly, with age, the Lin28b mice developed an aggressive, lethal, peripheral T cell lymphoma (PTCL), characterized by widespread infiltration of parenchymal organs with a mixed infiltrate of inflammatory cells and malignant CD4+ T cells. Clonal Tcrb gene rearrangements were observed in the lymphomas and the malignant cells engrafted and formed tumors in immunodeficient Scid mice. The Lin28b transgenic mice also had clinical signs consistent with a chronic inflammatory condition, such as eosinophilia, anemia, pleural effusions and ascites. The lymphomas overexpressed Il6 and Myc, and activated Nfκb, demonstrating in vivo involvement of a previously reported pathway that links Lin28b expression with inflammation and malignant transformation. Analysis of a publically available dataset indicated that Lin28b was overexpressed by 8-fold in a set of PTCL patient samples compared with activated CD4+ cells. Taken together, these findings demonstrate in vivo evidence for an oncogenic function of Lin28b and provide a model for further study of both the biology and identification of new therapeutic targets for PTCL, a heterogenous disease with poor prognosis. Disclosures: No relevant conflicts of interest to declare.

Direct Comparison of In Vivo Fate of Second and Third-Generation CD19-Specific Chimeric Antigen Receptor (CAR)-T Cells in Patients with B-Cell Lymphoma: Reversal of Toxicity from Tonic Signaling

Blood ◽  
2016 ◽  
Vol 128 (22) ◽  
pp. 1851-1851 ◽  
Author(s):  
Diogo Gomes da Silva ◽  
Malini Mukherjee ◽  
Madhuwanti Srinivasan ◽  
Olga Dakhova ◽  
Hao Liu ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
B Cell ◽  
Cell Lymphoma ◽  
B Cell Lymphoma ◽  
2Nd Generation ◽  
Car T Cells ◽  
Car T ◽  
Equity Ownership

Abstract Although adoptive transfer of T cells with second-generation CD19-specific CARs containing CD28 or 4-1BB costimulatory endodomains shows remarkable clinical efficacy against B cell malignancies, the optimal choice of costimulatory domains in these and other CARs remains controversial. Depending on the precise CAR structure and specificity, individual endodomains may be associated with deleterious ligand-independent tonic signaling in the transduced T cell. Long et al. (Nat Med 2015) established the CD28 co-stimulatory endodomain can have a toxic tonic signaling effect, but it is unclear if tonic 4-1BB signaling may have deleterious consequences as well, and if such effects can be reversed. We therefore modeled tonic CAR signaling in T cells by transducing them with gammaretroviral vectors expressing 2nd-generation CD19.CAR constructs containing either the CD28 or 4-1BB costimulatory endodomain (in addition to the CD3-ζ chain endodomain). Compared to CAR-T cells with the CD28 endodomain alone, those with 4-1BB alone expanded 70% more slowly following transduction. Impaired expansion of 4-1BB CD19.CAR-T cells was coupled with a 4-fold increase in apoptosis and a gradual downregulation of CAR expression, and was a consequence of 4-1BB-associated tonic TRAF2-dependent signaling, leading to activation of NF-κB, upregulation of Fas and augmented Fas-dependent activation-induced T cell death (AICD). Moreover, expression of 4-1BB CAR from a gammaretroviral vector increased tonic signaling through a self-amplifying/positive feedback effect on the retroviral LTR promoter. Because of the toxicity of 4-1BB in our gammaretroviral CAR.CD19 construct (manifest by delayed expansion and increased apoptosis) we could not directly compare the in vivo fate of T cells expressing CAR.CD19 4-1BB with that of co-administered CAR.CD19 CD28 T cells in patients with lymphoma. We found, however, that the adverse effects of tonic 4-1BB costimulation could be overcome in a 3rd-generation CAR.CD19 vector, containing both CD28 and 4-1BB costimulatory molecules in tandem. We thus compared the fate of a 3rd-generation vector containing both CD28 and 4-1BB costimulatory domains with that of a 2nd-generation vector containing CD28 alone. Six patients with refractory/relapsed diffuse large B-cell lymphoma received 2 cell populations, one expressing 2nd and one expressing 3rd generation vectors. To determine whether CD28 alone was optimal (which would suggest 4-1BB is antagonistic) or whether 4-1BB had an additive or synergistic effect contributing to superior persistence and expansion of the CD28-41BB combination, patients were simultaneously infused with 1-20×106 of both 2nd and 3rd generation CAR+ T cells/m2 48-72 hours after lymphodepletion with cyclophosphamide (500 mg/m2/d) and fludarabine (30 mg/m2/d) × 3. Persistence of infused T cells was assessed in blood by CD19.CAR qPCR assays specific for each CAR. Molecular signals peaked approximately 2 weeks post infusion, remaining detectable for up to 6 months. The 3rd-generation CAR-T cells had a mean 23-fold (range 1.1 to 109-fold) higher expansion than 2nd-generation CAR-T cells and correspondingly longer persistence. Two patients had grade 2 cytokine release syndrome, with elevation of proinflammatory cytokines, including IL-6, at the time of peak expansion of T cells. Of the 5 patients evaluable for response, 2 entered complete remission (the longest ongoing for 9 months), 1 has had continued complete remission after autologous stem cell transplantation, 1 had a partial response, and 1 progressed. In conclusion, our data indicate that infusion of T cells carrying a CD19.CAR containing CD28 and 4-1BB endodomains is safe and can have efficacy at every dose level tested. Additionally, in a side-by-side comparison, the 3rdgeneration vector produced greater in vivo expansion and persistence than an otherwise identical CAR-T cell population with CD28 alone. Disclosures Rooney: Cell Medica: Membership on an entity's Board of Directors or advisory committees, Patents & Royalties; Viracyte: Equity Ownership. Heslop:Celgene: Patents & Royalties, Research Funding; Chimerix: Other: Endpoint adjudication committee; Viracyte: Equity Ownership; Cell Medica: Patents & Royalties: Licensing agreement EBV-specific T cells.

scholarly journals Modeling Sézary Syndrome for Immunophenotyping and Anti-Tumor Effect of Ucart and Long-Acting Interleukin-7 Combination Therapy

Blood ◽  
2018 ◽  
Vol 132 (Supplement 1) ◽  
pp. 340-340
Author(s):  
Karl W. Staser ◽  
Matthew L. Cooper ◽  
Jaebok Choi ◽  
Anand Chukka ◽  
Kidist Ashami ◽  
...  
Keyword(s):  
T Cells ◽  
Tumor Burden ◽  
Photon Flux ◽  
Sézary Syndrome ◽  
Sezary Syndrome ◽  
Preclinical Models ◽  
Nsg Mice ◽  
Interleukin 7 ◽  
Long Acting

Abstract Background: Sézary syndrome (SS) is a highly-morbid T cell leukemic lymphoma with no widely-effective treatments and few preclinical models. We demonstrated effective T cell lymphoma therapy with allogeneic gene-edited anti-CD7 CARTs (Cooper et al, Leukemia, 2018). However, SS T cells typically lose CD7 but maintain ubiquitous high CD2 expression. Thus, we generated CD2- and TRAC-deleted anti-CD2 universal CARTs (UCART2) and multiple SS xenograft models (PDXs) as preclinical UCART2 testing platforms. We further tested a stable homodimeric interleukin-7 molecule, the long-acting form of recombinant human interleukin-7 fused with hybrid Fc (rhIL-7-hyFc, NT-I7), to potentiate UCART2 killing of an SS xenograft in vivo. Methods: To generate SS PDX models, we injected NOD scid IL2Rgammanull (NSG) mice expressing SCF, GM-CSF, and IL-3 (NSG-SGM3) with ~2x106 mononuclear cells derived from SS patients. We immunophenotyped SS patient blood and PDX engraftment with two 21-color flow cytometry panels assessing major immune subsets, CTCL, and exhaustion markers (Staser et al, Cytometry A, 2018). To generate UCART2s, we activated human T cells on CD3/CD28 beads, electroporated the T cells with Cas9, a TRAC-targeted gRNA, and a CD2-targeted gRNA followed by viral transduction with an anti-CD2 scFv 3rd generation CAR. For initial UCART2 testing, we injected NSG mice with 5x105 cells from a human Sézary cell line transduced with click beetle red luciferase (HHCBR-GFP) four days prior to UCART2 treatment. Mice were treated with NT-I7 (10mg/kg SC) on days +1, +15 and +29 post UCART2 infusion. Results: SS patient blood showed specific defects in monocyte, monocytic dendritic cell, and natural killer cell differentiation, increased skewing toward granulocytes and non-classical CD16+ monocytes (p<0.01, SS vs. normal PBMCs), and loss of effector memory CD4 cells (8% vs 34%, p<0.001, SS vs. normal PBMCs). SS cells were CD3+CD4+CD2+CD5+CD8- with variable CD7 loss and PD1 gain. Four of six unique human SS samples injected in NSG-SGM3 mice engrafted within ~6 weeks with no signs of xenogeneic GVHD. Following engraftment, SS cells showed near ubiquitous PD1 expression (>90% vs ~20%, p<0.001, SS vs. normal PBMCs), CD7 loss, and increased CD30 and CD26 expression. Immunohistochemistry further revealed atypical CD3+CD4+CD8-CD7- lymphocytes lining the dermo-epidermal junction. Second generation PDXs showed infiltration of the spleens, blood, and bone marrow with CD2+CD7- human cells and developed alopecia, scaling, and subcutaneous and intraperitoneal masses, with immunophenotyping, sequencing, and UCART treatment studies ongoing. To test UCART2's efficacy in killing SS cells in vivo, we injected NSG mice with HHCBR-GFP+ cells. UCART2-treated HHCBR-GFP mice showed dramatically reduced tumor burden as compared to control UCART19-treated HHCBR-GFP mice (BLI; 10^7 vs. 10^11 photon flux/s at 3 weeks, p<0.0001, UCART2 vs. UCART19). Moreover, UCART2-treated HHCBR-GFP mice survived ~40 days as compared to ~21 days in the UCART19 group. Remarkably, UCART2-treated HHCBR-GFP mice receiving NT-I7 showed virtually no tumor burden (maximum 106 photon flux/s vs. 1010 photon flux/s, UCART2+NT-I7 vs. UCART2 only groups) with 100% of UCART2- and NT-I7-treated HHCBR-GFP mice surviving beyond 49 days (Figure 1). Discussion: We describe the generation of physiologically-relevant SS preclinical models, comprehensive immunophenotyping of patient SS samples, clonal SS PDX outgrowth, and the highly effective anti-tumor activity of UCART2- plus NT-I7-mediated killing of SS cells in vivo using an NSG xenograft model. Ongoing studies involve treating primary SS and CTCL PDX models with UCART2 and NT-I7. These preclinical data validate the use of allogeneic "off-the-shelf" adoptive immunotherapy for the treatment of Sézary syndrome, while demonstrating the dramatic enhancement of CART efficacy using a dose-adjustable, clinic-ready long-acting interleukin-7 agonist given in an adjuvant setting. Disclosures Park: NeoImmuneTech: Employment. Lee:NeoImmuneTech: Employment. Musiek:Seattle Genetics: Honoraria; Actelion: Other: Scientific Advisory Committee ; Kyowa Kirin: Honoraria.

scholarly journals Recombinant Interleukin-7 Induces Proliferation of Naive Macaque CD4+ and CD8+ T Cells In Vivo

2004 ◽  
Vol 78 (18) ◽  
pp. 9740-9749 ◽  
Author(s):  
Marcin Moniuszko ◽  
Terry Fry ◽  
Wen-Po Tsai ◽  
Michel Morre ◽  
Brigitte Assouline ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Rhesus Macaques ◽  
Memory Cell ◽  
T Cell Subsets ◽  
Effector Memory ◽  
Phenotypic Change ◽  
Necrosis Factor Alpha ◽  
Interleukin 7

ABSTRACT Interleukin-7 (IL-7) regulates T-cell homeostasis, and its availability is augmented in lymphopenic hosts. Naive CD8+ T cells transferred to lymphopenic mice acquire a memory-like phenotype, raising the possibility that IL-7 is the biological mediator of this effect. Here, we provide direct evidence that IL-7 induces the acquisition of memory-cell markers not only in CD8+ T cells but also in CD4+ T-cell subsets in immune-competent Indian rhesus macaques. The increase of these memory-like populations was dependent on the dose of the cytokine, and these cells were found in the blood as well as secondary lymphoid organs. Memory-like CD4+ and CD8+ T cells acquired the ability to secrete tumor necrosis factor alpha and, to a lesser extent, gamma interferon following stimulation with a cognate antigen. The phenotypic change observed in naive T cells was promptly reversed after discontinuation of IL-7. Importantly, IL-7 induced cycling of both CD4+ and CD8+ central memory and effector memory T cells, demonstrating its contribution to the maintenance of the entire T-cell pool. Thus, IL-7 may be of benefit in the treatment of iatrogenic or virus-induced T-cell depletion.

Immunomodulatory Agent Lenalidomide Enhances Antitumor Functions of Chimeric Receptor-Modified T Cells in Vitro and in Vivo

Blood ◽  
2014 ◽  
Vol 124 (21) ◽  
pp. 805-805 ◽  
Author(s):  
Otáhal Pavel ◽  
Dana Prukova ◽  
Vlastimil Král ◽  
Radek Jaksa ◽  
Lucie Lateckova ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
B Cell ◽  
Cell Lymphoma ◽  
B Cell Lymphoma ◽  
Chimeric Receptor ◽  
Car T Cells ◽  
Car T

Abstract Tumor immunotherapy based on the use of Chimeric Receptor Modified T cells (CAR T cells) is a promising approach for the treatment of a refractory hematological cancer. However, a robust response mediated by CAR T cells is observed only in a minority of patients and the expansion and persistence of CAR T cells in vivo is mostly unpredictable. In order to enhance the effectiveness of CAR-based immunotherapy we tested the immunoadjuvant properities of lenalidomide in combination with CAR19 T cells in a mouse model of B cell lymphoma. CAR19 construct which was used is composed of anti-CD19scFv joined with signaling domain of 4-1BB and TCR zeta and was delivered into T cell via lentiviral transduction. Lenalidomide is an immunomodulatory drug used for the treatment of multiple myeloma and selected B-cell malignancies, e.g. mantle cell lymphoma (MCL) or activated B-cell subtype of diffuse large B-cell lymphoma (ABC-DLBCL). However, the precise mechanism of action is not very well understood and it is believed that is mediated by a modulation of activity of E3 ubiquitin ligase cereblon which leads to increased ubiquitinylation of Ikaros and Aiolos transcription factors resulting in changes of expression of various receptors on the surface of tumor cells. To test our hypothesis, immunodeficient NSG mice (NOD-SCID-gamma chain null mice) were s.c. transplanted with various human B cells lymphoma cells (MCL or ABC-DLBCL) followed by i.v treatment with CAR19 T cells with or without daily i.p. lenalidomide. First, when we measured the growth of tumors following treatment with CAR19 T cells plus lenalidomide we found that this combination more effectively suppressed growth of s.c. B-NHL tumors than treatment with only CAR19 T cells or only lenalidomide (Figure 1, 1x10e7 Nemo tumor cell s.c., followed with 2 doses of 1x10(7) CAR19 T cells + Lenalidomide daily, tumor weight was measured 14 days after treatment). Additionally, in this experiment lenalidomide significantly enhanced infiltration of residual tumors by CD8+CAR19 T cells (not shown). Next, we tested the response of CAR19 T cells in vitro to B-NHL cells in the presence or, absence of lenalidomide to determine the costimulatory effect of lenalidomide on signaling via CAR, our data show that lenalidomide significantly enhanced functional response of CAR19 T cells following recognition of B cells in vitro which is demonstrated by enhanced production of IFN-gamma and by increased expression of CD69 by CAR19 T cells, interestingly, this effect was seen only if CAR19 T cells but not B-NHL cells were pre-treated with lenalidomide or, when we activated CAR19 T cell with antibody to CAR but not with antibody to CD3. Thus, our data indicate that lenalidomide might work through direct effects on T cells and specifically enhance signaling via CAR. The biochemical events underlying this costimulatory effect of lenalidomide on signaling by CAR are currently being investigated. In summary, our data support the use of lenalidomide for augmentation CAR-based immunotherapy in clinical settings. Figure 1 Figure 1. Disclosures Klener: Cellgene: Research Funding.

Chimeric Antigen Receptor-Engineered T-Cells - A New Way and Era for Lymphoma Treatment

2020 ◽  
Vol 14 (4) ◽  
pp. 312-323
Author(s):  
Romeo G. Mihăilă
Keyword(s):  
T Cells ◽  
T Cell ◽  
Response Rate ◽  
Cell Lymphoma ◽  
B Cell Lymphoma ◽  
Car T Cells ◽  
Future Developments ◽  
Car T Cell ◽  
Large B Cell Lymphoma ◽  
Car T

Background: Patients with refractory or relapsed diffuse large B-cell lymphoma have a poor prognosis with the current standard of care. Objective: Chimeric Antigen Receptor T-cells (CAR T-cells) are functionally reprogrammed lymphocytes, which are able to recognize and kill tumor cells. The aim of this study is to make progress in this area. Method: A mini-review was achieved using the articles published in Web of Science and PubMed in the last year and the new patents were made in this field. Results: The responses to CAR T-cell products axicabtagene ciloleucel and tisagenlecleucel are promising; the objective response rate can reach up to 83%, and the complete response rate ranges between 40 and 58%. About half of the patients may have serious side effects, such as cytokine release syndrome and neurotoxicity. Current and future developments include the improvement of CAR T-cell expansion and polyfunctionality, the combined use of CAR T-cells with a fusion protein between interferon and an anti-CD20 monoclonal antibody, with checkpoint inhibitors or small molecule sensitizers that have apoptotic-regulatory effects. Furthermore, the use of IL-12-expressing CAR T-cells, an improved technology for the production of CAR T-cells based on targeted nucleases, the widespread use of allogeneic CAR T-cells or universal CAR T-cells obtained from genetically engineered healthy donor T-cells are future developments actively considered. Conclusion: CAR T-cell therapy significantly improved the outcome of patients with relapsed or refractory diffuse large B-cell lymphoma. The advances in CAR T-cells production technology will improve the results and enable the expansion of this new immunotherapy.

scholarly journals Use of immune repertoire sequencing to resolve discordant microscopic and immunochemical findings in a case of T cell-rich large B cell lymphoma in a young dog

2021 ◽  
Vol 17 (1) ◽  
Author(s):  
Gary Kwok Cheong Lee ◽  
Dorothee Bienzle ◽  
Stefan Matthias Keller ◽  
Mei-Hua Hwang ◽  
Nikos Darzentas ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
B Cell ◽  
Cell Lymphoma ◽  
B Cell Lymphoma ◽  
Immune Repertoire ◽  
Large B Cell Lymphoma ◽  
Repertoire Sequencing ◽  
Reactive Lymphocytes ◽  
Large B Cell

Abstract Background Lymphocytic neoplasms with frequent reactive lymphocytes are uncommonly reported in dogs, and can pose a diagnostic challenge. Different diagnostic modalities such as cytology, flow cytometry, histopathology, immunohistochemistry, and clonality testing, are sometimes required for a diagnosis. This report illustrates the value of using a multi-modal diagnostic approach to decipher a complex lymphocytic tumor, and introduces immune repertoire sequencing as a diagnostic adjunct. Case presentation A 10-month-old Great Dane was referred for marked ascites. Cytologic analysis of abdominal fluid and hepatic aspirates revealed a mixed lymphocyte population including numerous large lymphocytes, yielding a diagnosis of lymphoma. Flow cytometrically, abdominal fluid lymphocytes were highly positive for CD4, CD5, CD18, CD45, and MHC II, consistent with T cell lymphoma. Due to a rapidly deteriorating clinical condition, the dog was euthanized. Post mortem histologic evaluation showed effacement of the liver by aggregates of B cells surrounded by T cells, suggestive of hepatic T cell-rich large B cell lymphoma. Immune repertoire sequencing confirmed the presence of clonal B cells in the liver but not the abdominal fluid, whereas reactive T cells with shared, polyclonal immune repertoires were found in both locations. Conclusions T cell-rich large B cell lymphoma is a rare neoplasm in dogs that may be challenging to diagnose and classify due to mixed lymphocyte populations. In this case, the results of histopathology, immunohistochemistry and immune repertoire sequencing were most consistent with a hepatic B cell neoplasm and reactive T cells exfoliating into the abdominal fluid. Immune repertoire sequencing was helpful in delineating neoplastic from reactive lymphocytes and characterizing repertoire overlap in both compartments. The potential pitfalls of equating atypical cytomorphology and monotypic marker expression in neoplasia are highlighted.

scholarly journals Interleukin-7 Regulates Bim Proapoptotic Activity in Peripheral T-Cell Survival

2009 ◽  
Vol 30 (3) ◽  
pp. 590-600 ◽  
Author(s):  
Wen Qing Li ◽  
Tad Guszczynski ◽  
Julie A. Hixon ◽  
Scott K. Durum
Keyword(s):  
T Cells ◽  
T Cell ◽  
Cell Survival ◽  
T Cell Development ◽  
Cell Development ◽  
Effector Memory ◽  
Intracellular Location ◽  
Interleukin 7 ◽  
Peripheral T Cells ◽  
T Cell Survival

ABSTRACT Interleukin-7 (IL-7) is critical for T-cell development and peripheral T-cell homeostasis. The survival of pro-T cells and mature T cells requires IL-7. The survival function of IL-7 is accomplished partly through induction of the antiapoptotic protein Bcl-2 and inhibition of proapoptotic proteins Bax and Bad. We show here that the proapoptotic protein Bim, a BH3-only protein belonging to the Bcl-2 family, also plays a role in peripheral T-cell survival. Deletion of Bim partially protected an IL-7-dependent T-cell line and peripheral T cells, especially cells with an effector memory phenotype, from IL-7 deprivation. However, T-cell development in the thymus was not restored in IL-7−/− Rag2−/− mice reconstituted with Bim−/− bone marrow. IL-7 withdrawal altered neither the intracellular location of Bim, which was constitutively mitochondrial, nor its association with Bcl-2; however, a reduction in its association with the prosurvival protein Mcl-1 was observed. IL-7 withdrawal did not increase Bim mRNA or protein expression but did induce changes in the isoelectric point of BimEL and its reactivity with an antiphosphoserine antibody. Our findings suggest that the maintenance of peripheral T cells by IL-7 occurs partly through inhibition of Bim activity at the posttranslational level.

scholarly journals Accelerated, but not conventional, radiotherapy of murine B-cell lymphoma induces potent T cell–mediated remissions

2018 ◽  
Vol 2 (19) ◽  
pp. 2568-2580 ◽  
Author(s):  
Suparna Dutt ◽  
Michelle B. Atallah ◽  
Yoshitaka Minamida ◽  
Alexander Filatenkov ◽  
Kent P. Jensen ◽  
...  
Keyword(s):  
T Cells ◽  
Dendritic Cells ◽  
T Cell ◽  
B Cell ◽  
Cd8 T Cells ◽  
Hematopoietic Cell Transplantation ◽  
Cell Lymphoma ◽  
B Cell Lymphoma ◽  
Tumor Irradiation ◽  
Total Radiation Dose

Abstract Conventional local tumor irradiation (LTI), delivered in small daily doses over several weeks, is used clinically as a palliative, rather than curative, treatment for chemotherapy-resistant diffuse large B-cell lymphoma (DLBCL) for patients who are ineligible for hematopoietic cell transplantation. Our goal was to test the hypothesis that accelerated, but not conventional, LTI would be more curative by inducing T cell–mediated durable remissions. We irradiated subcutaneous A20 and BL3750 lymphoma tumors in mice with a clinically relevant total radiation dose of 30 Gy LTI, delivered in 10 doses of 3 Gy over 4 days (accelerated irradiation) or as 10 doses of 3 Gy over 12 days (conventional irradiation). Compared with conventional LTI, accelerated LTI resulted in more complete and durable tumor remissions. The majority of these mice were resistant to rechallenge with lymphoma cells, demonstrating the induction of memory antitumor immunity. The increased efficacy of accelerated LTI correlated with higher levels of tumor cell necrosis vs apoptosis and expression of “immunogenic cell death” markers, including calreticulin, heat shock protein 70 (Hsp70), and Hsp90. Accelerated LTI–induced remissions were not seen in immunodeficient Rag-2−/− mice, CD8+ T-cell–depleted mice, or Batf-3−/− mice lacking CD8α+ and CD103+ dendritic cells. Accelerated, but not conventional, LTI in immunocompetent hosts induced marked increases in tumor-infiltrating CD4+ and CD8+ T cells and MHCII+CD103+CD11c+ dendritic cells and corresponding reductions in exhausted PD-1+Eomes+CD8+ T cells and CD4+CD25+FOXP3+ regulatory T cells. These findings raise the possibility that accelerated LTI can provide effective immune control of human DLBCL.

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