scholarly journals Lentiviral vector induces high-quality memory T cells via dendritic cells transduction

2021 ◽  
Vol 4 (1) ◽  
Author(s):  
Min Wen Ku ◽  
Pierre Authié ◽  
Fabien Nevo ◽  
Philippe Souque ◽  
Maryline Bourgine ◽  
...  
Keyword(s):  
T Cells ◽  
Lentiviral Vector ◽  
Adenovirus Type ◽  
Effector Memory ◽  
High Quality ◽  
Memory Phenotype ◽  
Β2 Microglobulin ◽  
Clear Cut ◽  
Tumor Protection

AbstractWe report a lentiviral vector harboring the human β2-microglobulin promoter, with predominant expression in immune cells and minimal proximal enhancers to improve vector safety. This lentiviral vector efficiently transduces major dendritic cell subsets in vivo. With a mycobacterial immunogen, we observed distinct functional signatures and memory phenotype in lentiviral vector- or Adenovirus type 5 (Ad5)-immunized mice, despite comparable antigen-specific CD8+ T cell magnitudes. Compared to Ad5, lentiviral vector immunization resulted in higher multifunctional and IL-2-producing CD8+ T cells. Furthermore, lentiviral vector immunization primed CD8+ T cells towards central memory phenotype, while Ad5 immunization favored effector memory phenotype. Studies using HIV antigens in outbred rats demonstrated additional clear-cut evidence for an immunogenic advantage of lentiviral vector over Ad5. Additionally, lentiviral vector provided enhance therapeutic anti-tumor protection than Ad5. In conclusion, coupling lentiviral vector with β2-microglobulin promoter represents a promising approach to produce long-lasting, high-quality cellular immunity for vaccinal purposes.

Targeting Lewis Y-Positive Multiple Myeloma and Acute Myeloid Leukemia with Gene-Modified T Cells Demonstrating Memory Phenotype

Blood ◽  
2008 ◽  
Vol 112 (11) ◽  
pp. 3900-3900
Author(s):  
Stefan Peinert ◽  
David S. Ritchie ◽  
Dirk Hoenemann ◽  
Simon J. Harrison ◽  
Preethi Guru ◽  
...  
Keyword(s):  
Bone Marrow ◽  
T Cells ◽  
T Cell ◽  
Tumor Cells ◽  
T Cell Subsets ◽  
Effector Memory ◽  
Memory Phenotype ◽  
Lewis Y ◽  
Active Proliferation

Abstract AML and MM are sensitive to immune control as evidenced by T cell mediated allogeneic graft versus leukemia/myeloma effect. Adoptive immunotherapy with gene modified T cells has shown clinical activity in some solid tumors and B cell non Hodgkin lymphomas. The carbohydrate antigen Lewis Y (LeY) is a tumor associated antigen expressed on numerous epithelial cancers. Our aim was to generate gene-modified clinical-grade T cells directed against LeY positive hematological malignancies. Moreover, we aimed to produce cells that possessed T cell memory, essential for in vivo T cell persistence and long-term control of tumor cell targets. MM and AML cell lines were found to express differing levels of the LeY antigen ranging from negative (median fluorescence intensity (MFI) equal to mature lymphocytes (lymph) as internal control) to strongly positive (up to10×MFI lymph). Furthermore, 25/46 (54%) and 15/29 (52)% of primary MM and AML bone marrow samples were LeY-positive (≥5×MFI lymph), respectively. Lewis Y expression did not correlate with patient age, gender, clinical risk status, cytogenetic abnormalities, extent of previous chemotherapy, degree of bone marrow infiltrate, disease subtype, cytopenias in peripheral blood (MM and AML), LDH, WBC > or ≤ 30×109/L (AML), beta2 microglobulin, albumin or pretreatment with bortezomib, thalidomide or lenalidomide (MM). We manufactured a novel retroviral vector construct enabling efficient transduction of PBMC-derived T cells with resultant high expression (up to 65%) of a single-chain anti-LeY chimeric T cell receptor comprising T cell activation via CD3zeta and CD28 signaling domains. Under GMP conditions, we achieved >100-fold expansion of T cells using a 12 day culture protocol. Anti-LeY T cells lysed LeY-positive tumor cells in vitro while sparing LeY-negative control tumor cells and LeY expressing neutrophils (moderate LeY expression, >3×MFI lymph). Similar transduction rates were achieved in CD4 and CD8 T cell subsets. Twelve-day culture T cells showed low expression levels of CD45RA and CCR7, moderate levels of the co-stimulatory molecules CD27 and CD28, and active proliferation in response to IL-2 and IL-15, suggesting an effector memory phenotype. On re-exposure to LeY expressing tumor cells, anti-LeY T cells displayed active proliferation and IFN-gamma production. In vivo efficacy was demonstrated in three independent experiments of a MM xenograft mouse model showing improved disease free survival of mice receiving anti LeY T cells compared to control mice treated with untransduced T cells. Transplantation of syngeneic murine anti LeY T cells into sublethally irradiated Balb/C mice subsequently monitored for up to two years was found to be safe without generation of lymphoproliferative disorders arising from the anti LeY T cells and no impact on OS compared to irradiated control mice not receiving adoptive T cell transfer. Consequently, we have developed a first in human phase I trial of autologous anti LeY T cells for patients with LeY-positive MM or AML. LeY is a promising and immunologically relevant target for T cell immunotherapy and our product is likely to lead to persistence of anti-LeY T cells in patients, an outcome which will be specifically addressed in our upcoming study.

scholarly journals Rapamycin Improves the Response of Effector and Memory CD8+ T Cells Induced by Immunization With ASP2 of Trypanosoma cruzi

2021 ◽  
Vol 11 ◽  
Author(s):  
Barbara Ferri Moraschi ◽  
Isaú Henrique Noronha ◽  
Camila Pontes Ferreira ◽  
Leonardo M. Cariste ◽  
Caroline B. Monteiro ◽  
...  
Keyword(s):  
T Cells ◽  
Trypanosoma Cruzi ◽  
Vaccine Development ◽  
Human Adenovirus ◽  
Adenovirus Type ◽  
T Cell Response ◽  
Effector Memory ◽  
Mtor Inhibition ◽  
Beneficial Effects

Deficiency in memory formation and increased immunosenescence are pivotal features of Trypanosoma cruzi infection proposed to play a role in parasite persistence and disease development. The vaccination protocol that consists in a prime with plasmid DNA followed by the boost with a deficient recombinant human adenovirus type 5, both carrying the ASP2 gene of T. cruzi, is a powerful strategy to elicit effector memory CD8+ T-cells against this parasite. In virus infections, the inhibition of mTOR, a kinase involved in several biological processes, improves the response of memory CD8+ T-cells. Therefore, our aim was to assess the role of rapamycin, the pharmacological inhibitor of mTOR, in CD8+ T response against T. cruzi induced by heterologous prime-boost vaccine. For this purpose, C57BL/6 or A/Sn mice were immunized and daily treated with rapamycin for 34 days. CD8+ T-cells response was evaluated by immunophenotyping, intracellular staining, ELISpot assay and in vivo cytotoxicity. In comparison with vehicle-injection, rapamycin administration during immunization enhanced the frequency of ASP2-specific CD8+ T-cells and the percentage of the polyfunctional population, which degranulated (CD107a+) and secreted both interferon gamma (IFNγ) and tumor necrosis factor (TNF). The beneficial effects were long-lasting and could be detected 95 days after priming. Moreover, the effects were detected in mice immunized with ten-fold lower doses of plasmid/adenovirus. Additionally, the highly susceptible to T. cruzi infection A/Sn mice, when immunized with low vaccine doses, treated with rapamycin, and challenged with trypomastigote forms of the Y strain showed a survival rate of 100%, compared with 42% in vehicle-injected group. Trying to shed light on the biological mechanisms involved in these beneficial effects on CD8+ T-cells by mTOR inhibition after immunization, we showed that in vivo proliferation was higher after rapamycin treatment compared with vehicle-injected group. Taken together, our data provide a new approach to vaccine development against intracellular parasites, placing the mTOR inhibitor rapamycin as an adjuvant to improve effective CD8+ T-cell response.

Allogeneic Memory T Cells Derived from Host DC-Activated CD44loCD8+ T Precursors Sustain Host Tissue Injury of Ongoing Graft-versus-Host Disease.

Blood ◽  
2004 ◽  
Vol 104 (11) ◽  
pp. 3040-3040
Author(s):  
Yi Zhang ◽  
Gerard Joe ◽  
Elizabeth Hexner ◽  
Jiang Zhu ◽  
Stephen G. Emerson
Keyword(s):  
T Cells ◽  
T Cell ◽  
Cd8 T Cells ◽  
Memory T Cells ◽  
Effector Memory ◽  
Memory Phenotype ◽  
Memory T Cell ◽  
Cell Responses ◽  
With Memory

Abstract Graft-versus-host disease (GVHD) directed against minor histocompatibility antigen (miHAs) evolves over weeks to months, suggesting a requirement for persistent alloreactive donor T cells. In patients with allogeneic bone marrow transplantation (allo-BMT), persistency of GVHD is accompanied with elevated allogeneic CD4+ T cells with memory phenotype in peripheral blood. In contrast, several other studies have recently shown that T cells with memory phenotype (CD44hiCD62Lhi/lo) from normal donor mice do not induce acute GVHD. While these T cells with memory phenotype may be induced by environmental antigenic stimulation or may represent cells undergoing homeostasis in vivo, we found that early activated donor CD44hiCD8+ T cells with effector/memory phenotype upon ex vivo host DC stimulation are also functional defective in GVHD induction in vivo. However, whether alloreactive memory T cells might develop in vivo in recipient with ongoing GVHD, and if this is the case, whether these in vivo generated alloreactive memory T cells may be responsbile for persistency of GVHD, remain unknown. Using the C3H.SW anti-C57BL/6 (B6) and B6 anti-BALB.B mouse models of human GVHD directed against miHAs, we found that alloreactive CD8+ T cells secreting high levels of IFN-γ in recipient mice receiving C3H.SW CD44loCD8+ T cells + T−BM peaked by day 14, declined by day 28, and increased again after 35 days of transplantation, corresponding to the kinetics of primary and memory T cell responses. Indeed, while donor C3H.SW CD8+ T cells recovered from these B6 mice receiving C3H.SW CD44loCD8+ T cells + T−BM 10 days after allo-BMT, at the peak time of primary allogeneic immune response, upregulated the expression of effector marker CD25, donor CD8+ T cells recovered 42 days after allo-BMT from B6 mice with ongoing GVHD, at the time of memory T cell development, expressed high levels of CD44 and CD122 but down-regulated CD25. However, both d10-CD8+ and d42-CD8+ T cells expressed identical levels of cytotoxic molecules including granzyme B, perforin and FasL and were able to kill B6 mouse-derived EL-4 leukemic cells. Compared to naïve CD44loCD8+ T cells that were lost after cultured in the presence of IL-2+IL-15 for 5 days, d42-donor CD8+ T cells recovered from B6 mice with ongoing GVHD survived over 5 days in the presence of IL-2+IL-15 alone and these surviving d42-CD8+ T cells were able to rapidly proliferate in responding to B6 DCs+IL-2+IL-15 in secondary culture. Flow cytometry analysis showed that d42-CD8+ T cells contained at least two distinct subsets: CD44hiCD62Llo (80% to 85%) and CD44hiCD62LhiCD8+(3% to 6%) T cells, resembling to the phenotype of effector memory and central memory CD8+ T cells, respectively. Administration of irradiated secondary B6 mice with either d42-CD44hiCD62Lhi or d42-CD44hiCD62Llo CD8+ T cell subset recovered at day 42 from primary B6 mice receiving C3H.SW CD44loCD8+ T cells + T−BM caused virulent GVHD. These results indicate that alloreactive memory T cells develop in vivo in recipient mice with acute GVHD where host mHAs persist and may be responsible for the persistence of GVHD. Accordingly, we suggest that in vivo blockade of both alloreactive effector and memory T cell responses will be necessary for GVHD prevention and treatment.

Adoptive Immunotherapy with Memory T Cells.

Blood ◽  
2008 ◽  
Vol 112 (11) ◽  
pp. sci-25-sci-25 ◽  
Author(s):  
Helen E. Heslop
Keyword(s):  
T Cells ◽  
T Cell ◽  
Memory T Cells ◽  
Ex Vivo ◽  
Central Memory ◽  
Effector Memory ◽  
Memory Phenotype ◽  
Effector Memory T Cells ◽  
Effector Memory Cells

Clinical adoptive cellular immunotherapy of malignancy and viral infection should transfer T cells that expand in vivo on exposure to antigen and can enter the memory compartment to persist long-term. A number of factors, including cellular phenotype, influence the behavior of the infused line. Primate studies have shown that antigen-specific CD8+ T cell clones only persisted long-term in vivo if they were derived from central memory T cells, but not from effector memory T cells, reacquiring phenotypic and functional properties of memory T cells.1 Other studies have suggested that adoptive transfer of ex vivo-expanded effector memory T cells will have poor survival and clinical efficacy, reporting instead that less differentiated T cells with longer telomeres exhibit longer persistence. These data imply that prolonged ex vivo expansion, required, for example, for T cell cloning, adversely affects subsequent in vivo expansion and survival. However, our trials administering ex vivo-expanded, polyclonal EBV-specific T cell lines demonstrated that expanded effector memory T cells, infused into a lymphodepleted host, can expand massively in vivo, enter the memory compartment, and persist for up to seven years after infusion. Furthermore, in a study infusing trivirus-specific CTLs with effector memory phenotype, we saw expansion of CTLs specific for the latent viruses CMV and EBV. By contrast, adenoviral-specific CTL persisted only in patients who were acutely infected with the agent2 We recently compared non-specifically activated T cells (ATC) with EBV-specific CTLs derived from the same initial peripheral blood collection and expressing distinguishable chimeric GD2-specific chimeric antigen receptors (CARATC and CAR-CTL). In this study, ATCs were cultured for 14 to 21 days. Between 0.9% and 6.1% retained a central memory (CCR7+, CD62L+) phenotype, up to 30% had an effector memory phenotype (CCR7−, CD62L+), and the remainder had a terminally/fully differentiated effector phenotype. By contrast, EBV-CTL were cultured for 30 to 44 days and expressed no CCR7, but up to 50% were CD62L+, and contained cells that were terminally/fully differentiated effectors and effector memory cells. These EBV-CTLs also all had a CD45RO memory phenotype, while about 13% to 60% of ATCs expressed CD45RA, a marker of naïve T cells. Despite these differences in memory subsets, it was the CAR-CTLs that had the clearly greater persistence and could be shown to retain functionality, while CAR-ATC rapidly disappeared from the circulation and could not be recovered. Hence, factors other than phenotype, such as antigenic stimulation and costimulation almost certainly influence cell fate after infusion, and determine whether or not effector memory cells can re-access the central memory pool. Ultimately, strategies that combine selection of optimal phenotype with the provision of antigen stimulation and co-stimulation and a cytokine milieu that favors homeostatic expansion will likely lead to the most effective outcomes following adoptive T cell transfer.

scholarly journals Reovirus-induced cell-mediated immunity for the treatment of multiple myeloma within the resistant bone marrow niche

2021 ◽  
Vol 9 (3) ◽  
pp. e001803
Author(s):  
Louise M E Müller ◽  
Gemma Migneco ◽  
Gina B Scott ◽  
Jenny Down ◽  
Sancha King ◽  
...  
Keyword(s):  
Multiple Myeloma ◽  
T Cells ◽  
Immune Responses ◽  
Stromal Cells ◽  
Tumor Burden ◽  
Effector Memory ◽  
In Vivo Model ◽  
Bm Niche

BackgroundMultiple myeloma (MM) remains an incurable disease and oncolytic viruses offer a well-tolerated addition to the therapeutic arsenal. Oncolytic reovirus has progressed to phase I clinical trials and its direct lytic potential has been extensively studied. However, to date, the role for reovirus-induced immunotherapy against MM, and the impact of the bone marrow (BM) niche, have not been reported.MethodsThis study used human peripheral blood mononuclear cells from healthy donors and in vitro co-culture of MM cells and BM stromal cells to recapitulate the resistant BM niche. Additionally, the 5TGM1-Kalw/RijHSD immunocompetent in vivo model was used to examine reovirus efficacy and characterize reovirus-induced immune responses in the BM and spleen following intravenous administration. Collectively, these in vitro and in vivo models were used to characterize the development of innate and adaptive antimyeloma immunity following reovirus treatment.ResultsUsing the 5TGM1-Kalw/RijHSD immunocompetent in vivo model we have demonstrated that reovirus reduces both MM tumor burden and myeloma-induced bone disease. Furthermore, detailed immune characterization revealed that reovirus: (i) increased natural killer (NK) cell and CD8+ T cell numbers; (ii) activated NK cells and CD8+ T cells and (iii) upregulated effector-memory CD8+ T cells. Moreover, increased effector-memory CD8+ T cells correlated with decreased tumor burden. Next, we explored the potential for reovirus-induced immunotherapy using human co-culture models to mimic the myeloma-supportive BM niche. MM cells co-cultured with BM stromal cells displayed resistance to reovirus-induced oncolysis and bystander cytokine-killing but remained susceptible to killing by reovirus-activated NK cells and MM-specific cytotoxic T lymphocytes.ConclusionThese data highlight the importance of reovirus-induced immunotherapy for targeting MM cells within the BM niche and suggest that combination with agents which boost antitumor immune responses should be a priority.

scholarly journals Human Effector Memory CD4+ T Cells Directly Recognize Allogeneic Endothelial Cells In Vitro and In Vivo

2007 ◽  
Vol 179 (7) ◽  
pp. 4397-4404 ◽  
Author(s):  
Stephen L. Shiao ◽  
Nancy C. Kirkiles-Smith ◽  
Benjamin R. Shepherd ◽  
Jennifer M. McNiff ◽  
Edward J. Carr ◽  
...  
Keyword(s):  
T Cells ◽  
Endothelial Cells ◽  
Cd4 T Cells ◽  
Effector Memory ◽  
Memory Cd4 T Cells

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.

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