scholarly journals Results from a First-in-Human Study with Bnz-1: A Novel Peptide Inhibitor of IL-2, IL-9 and IL-15 for the Treatment of T-Cell Malignancies That Safely and Selectively Decreases Regulatory T-Cells, Natural Killer Cells, and CD8+ Central Memory T-Cells

Blood ◽  
2017 ◽  
Vol 130 (Suppl_1) ◽  
pp. 695-695
Author(s):  
Paul Frohna ◽  
Yutaka Tagaya ◽  
Anoshie Ratnayake ◽  
Nick Doerr ◽  
Asjad Basheer ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Regulatory T Cells ◽  
Nk Cells ◽  
Memory T Cells ◽  
Human Study ◽  
Central Memory ◽  
Post Dose ◽  
Central Memory T Cells ◽  
T Cell Malignancies

Abstract Background: The γc-family of cytokines (IL-2, IL-4, IL-7, IL-9, IL-15, IL-21) control major immune responses and lymphocyte development. However, aberrant γc cytokine activity contributes to or pathologically drives human diseases including T-cell malignancies, graft-versus-host disease, and numerous autoimmune diseases (e.g., alopecia areata), where IL-2, IL-9, and IL-15 have been specifically implicated. Importantly, these multi-cytokine diseases are not effectively and safely treated by the currently available anti-cytokine approaches. Methods: BNZ-1 is the first of a novel class of rationally-designed, multi-cytokine inhibitors that selectively inhibits IL-2 and IL-15, and to a lesser degree, IL-9 signaling through the γc receptor, without affecting IL-4, IL-7, or IL-21. Preclinical proof-of-concept has been demonstrated in several animal models of T-cell malignancies (Nata et al., 2015 J Biol Chem), including Large Granular Lymphocyte Leukemia (LGL) and Adult T-cell Leukemia. In this open-label, single-dose, dose-escalation, first-in-human study conducted at a single center in the United States (NCT03046459), 18 healthy adults (n=3/cohort) received a single intravenous dose of 0.2, 0.4, 0.8, 1.6, 3.2 or 6.4 mg/kg infused over ≤5 minutes on Day 1 and were followed for safety and sample collection for 30 days. Results: All subjects completed the study. BNZ-1 was considered well-tolerated with a good safety profile with no serious or severe adverse events (AEs), no dose-limiting toxicities, and no clinically-significant changes on clinical labs (serum biochemistry, hematology, liver enzymes), vital signs or electrocardiograms. Headache was the only AE reported in ≥1 subject (n=3). BNZ-1 exposure was generally dose proportional with an elimination half-life of ~5 days across the range of doses tested, which is consistent with other PEGylated peptides and supports weekly or every other week dosing. The pharmacodynamic activity of BNZ-1 was characterized by flow cytometry of PBMCs obtained on Days 4, 15 and 31 and used to calculate the maximum change from baseline (Emax) for regulatory T-cells (Tregs; IL-2 effect), NK cells (IL-15 effect) and CD8+ central memory T-cells (Tcm; IL-2 & IL-15 effect). For Tregs, a ~50-60% decrease was observed on Day 4 after doses of 0.4 to 1.6mg/kg, while doses of 3.2 and 6.4mg/kg produced an 80-93% decrease from baseline that peaked on Day 15. NK cells were decreased from baseline on Day 4 by 20%, 40% and 60% at 0.2, 0.4 and 0.8mg/kg, respectively, and plateaued at 70 to 80% decrease at doses ≥1.6 mg/kg. Tcm were decreased at Day 4 for the three highest dose cohorts that continued to decline to Day 15 when all doses, except 0.2 mg/kg, showed a mean decrease ranging from 10 to 81% that generally trended with dose. Tregs, NK cells, and Tcm returned to/toward baseline by 30 days post dose. Post-dose counts of CD4+ and CD8+ T-cells, B-cells, and monocytes were unchanged at all time points studied. Conclusions: These preliminary clinical data suggest that BNZ-1 is a highly-active, selective immunomodulator that safely decreases Tregs, NK cells and Tcm, while leaving the major leukocyte populations unaffected. These data support the planned Phase 1/2 dose-ranging study in LGL and Cutaneous T-cell Lymphoma. Disclosures Frohna: Bioniz Therapeutics: Employment. Tagaya: Bioniz Therapeutics: Equity Ownership. Ratnayake: Bioniz Therapeutics: Consultancy. Doerr: Bioniz Therapeutics: Employment. Basheer: Bioniz Therapeutics: Employment. Al-Mawsawi: Bioniz Therapeutics: Employment. Kim: Bioniz Therapeutics: Employment. Azimi: Bioniz Therapeutics: Employment.

Circulating Memory T Cells Isolated from Hodgkin Lymphoma Patients Display Evidence of Exhaustion and Chronic Activation

Blood ◽  
2014 ◽  
Vol 124 (21) ◽  
pp. 4400-4400
Author(s):  
Catherine S. Diefenbach ◽  
Bruce G. Raphael ◽  
Kenneth B. Hymes ◽  
Tibor Moskovits ◽  
David Kaminetzky ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Memory T Cells ◽  
Central Memory ◽  
Effector Memory ◽  
T Cell Exhaustion ◽  
Differentiation Markers ◽  
Cell Markers ◽  
Cell Exhaustion ◽  
Central Memory T Cells

Abstract Background: In Hodgkin lymphoma (HL) the malignant Hodgkin Reed-Sternberg (HRS) cells comprise only a small fraction of the total cellular tumor population. These HRS cells orchestrate an inflammatory microenvironment of reactive cells that propagate a permissive milieu for HL growth, contributing to an ineffective local anti-tumor immune response. The peritumoral CD4 and CD8 T cells in HL patients show high expression of the receptor programmed death-1 (PD-1), involved in the functional impairment and “exhaustion” of T cells. Growing data suggests that this HL-mediated immune suppression may have effects that extend beyond the tumor microenvironment. High systemic levels of inflammatory cytokines and chemokines in HL patients has been reported. We characterized the systemic immune profile of HL patients with both newly diagnosed (ND) and relapsed (R) disease. Methods: Informed consent for correlative blood testing was obtained from patients with ND (n=8) or R (n=5) HL treated at the NYU Perlmutter Cancer Center or NY Presbyterian/Weil Cornell since January of 2013. Blood samples were drawn pre-treatment, and at sequential timepoints during and after therapy. Peripheral blood mononuclear cells (PBMC) were isolated using Ficoll separation method and cells were frozen for subsequent analysis. The frozen PBMC were then stained with fluorescent-conjugated antibodies against T cell surface molecules in 10-color FACS analysis. The analyses were performed after gating live cells for CD4, CD8 and memory and effector T cell markers. Patient samples were compared to normal controls matched for age and sex (n=18). Results: The median HL patient age was 32 (22-72), and 8 subjects were male. All ND HL patients were treated with ABVD (range 4-6 cycles) +/- consolidative radiation; R patients had median of 3 prior therapies. One patient out of 5 had prior autologous stem cell transplant (SCT), and 1 had prior allogeneic SCT, but was not on immunosuppression. Eight patients (6ND, 2R) responded to therapy (8 CR); 5 patients (1ND, 4R) progressed on therapy or had stable disease. HL patients displayed a high frequency of the exhaustion marker PD-1 on CD4 central memory T cells (CD4+CD45RO+CD27+) compared to normal matched controls (NC): mean 41, standard error (SE) 4.8 for HL patients vs. mean 22.2, SE 1.3 for NC (p = 0.0002) (Figure 1A). PD-1 expression was similarly elevated on CD8 central memory T cells (CD8+CD45RO+CD27+) of HL patients: mean 55, SE 3.3 vs. NC: mean 40, SE 3.3 (p = 0.003) (Figure 1B). HL patients also displayed an increased frequency of PD-1 expression on CD27 negative CD4 effector T cells: mean 43, SE 4, vs. NC: mean 28.5, SE 2.4 (p = 0.003) (Figure 2). In 4 of the HL patients who responded to therapy, PD-1 expression on central memory CD4+ cells declined after therapy: mean 30.1 vs. mean increase of +2.67 in 3 patients who progressed on therapy (p< 0.009). A higher number of subjects in prospective analysis is underway, to confirm whether a response to therapy may be correlated with a reversal of the suppressed phenotype of T cells in these patients. Conclusion: HL patients have evidence of chronic activation/exhaustion in their central memory and effector T cells, suggesting that ineffective immune clearance of the HRS cells may be a systemic rather than local phenomenon. In patients with progressive disease for whom this phenotype persists it is worthy of investigation whether this immune dysfunction is a cause or consequence of resistance to therapy. This may be rationale for immune targeted therapy in patients with relapsed or resistant disease. Figure 1. Evidence for increased levels of T cell exhaustion in central memory T cells of HL patients. PBMC were stained with specific fluorescent conjugated antibodies against T cell markers (CD3, CD4, CD8) together with differentiation markers (CD45RO, CD27) and PD1 and analyzed using FACS (LSR-II). The proportion of PD1+ T cells were determined in: A) CD4+CD45RO+CD27+ and B) CD8+CD45RO+CD27+ T cells. Figure 1. Evidence for increased levels of T cell exhaustion in central memory T cells of HL patients. PBMC were stained with specific fluorescent conjugated antibodies against T cell markers (CD3, CD4, CD8) together with differentiation markers (CD45RO, CD27) and PD1 and analyzed using FACS (LSR-II). The proportion of PD1+ T cells were determined in: A) CD4+CD45RO+CD27+ and B) CD8+CD45RO+CD27+ T cells. Figure 2. Evidence for increased levels of T cell exhaustion in effector memory CD 4+ T cells of HL patients. PBMC were stained with specific fluorescent conjugated antibodies against T cell markers (CD3, CD4) together with differentiation markers (CD45RO, CD27) and PD1 and analyzed using FACS (LSR-II). The proportion of PD1+ T cells was determined in CD4+RO+CD27- T cells Figure 2. Evidence for increased levels of T cell exhaustion in effector memory CD 4+ T cells of HL patients. PBMC were stained with specific fluorescent conjugated antibodies against T cell markers (CD3, CD4) together with differentiation markers (CD45RO, CD27) and PD1 and analyzed using FACS (LSR-II). The proportion of PD1+ T cells was determined in CD4+RO+CD27- T cells Figure 3 Figure 3. Disclosures No relevant conflicts of interest to declare.

scholarly journals Mobilization of CD8+ Central Memory T-Cells with Enhanced Reconstitution Potential in Mice By a Combination of G-CSF and GMI-1271-Mediated E-Selectin Blockade

Blood ◽  
2015 ◽  
Vol 126 (23) ◽  
pp. 512-512 ◽  
Author(s):  
Ingrid G Winkler ◽  
Valerie Barbier ◽  
Kristen J Radford ◽  
Julie M Davies ◽  
Jean-Pierre Levesque ◽  
...  
Keyword(s):  
Bone Marrow ◽  
T Cells ◽  
T Cell ◽  
Research Funding ◽  
Memory T Cells ◽  
Central Memory ◽  
Donor Blood ◽  
Central Memory T Cells

Abstract T-cells are critical mediators of immune defense against pathogens and cancer. Adoptive T cell immunotherapy and T-cell engineering have promising clinical applications but T cell survival and exhaustion are current limitations. Central memory cells (TCM CD62L+ CCR7+) and their precursors, stem central memory T-cells (TSCM) possess the stem-like properties needed to reconstitute and prolong an effective immune response long-term. These cells have been shown to significantly improve therapeutic efficacy of adoptive T-cell therapy. The challenge remains to harvest good quality TCM-cells for these immunotherapy approaches. The bone marrow (BM) is the major reservoir of CD8+ TCM and their precursors. We have previously shown that E-selectin is expressed in the BM vasculature and drives activation and differentiation of hematopoietic stem cells during G-CSF induced mobilization to the blood. We find therapeutic blockade of E-selectin promotes HSC self-renewal and reconstitution in vivo. We now examine the impact of E-selectin blockade on CD8+ T cell mobilization from the bone marrow to the blood and hypothesize that E-selectin blockade may also dampen the activation/differentiation of this subset. First we administered a standard G-CSF regime (filgastim 250ug/kg/day for 3 days) to mice and then dosed some cohorts with GMI-1271 (40mg/kg BID) from 12 to 72 hours within this 3 day period. Administration of G-CSF alone results in a near complete disappearance of bone marrow resident CD8+ TCM cells, and their apparent migration (increase in numbers) to the blood, while CD8+ subsets in the lymph nodes and spleen were barely affected by G-CSF. Furthermore among T-cell subsets, CD8+ but not CD4+ TCM were specifically mobilized into the blood when GMI-1271 was co-administered for the last 12 to 24 hours of G-CSF. These findings are consistent with reports demonstrating the bone marrow to be a major reservoir for CD8+ but not CD4+ central memory T-cells. Administration of GMI-1271 caused a marked enhancement in mobilization into the blood of CD8+ TCM/SCM (CD62Lhi, CCR7+) cells over treatment with G-CSF alone (p<0.05). To determine the functional consequences of this skewed mobilization following GMI-1271 co-administration, 25 uL of mobilized blood was transplanted into irradiated congenic B6.SJL recipients together with 2x105 congenic BM cells to analyze long-term donor T-cell engraftment in the recipient mice. We found G-CSF mobilized donor blood did not contribute CD8+ TCM cells that can persist post-transplant (<0.5% at 20 weeks post-transplant). In contrast when donor mice were mobilized with G-CSF together with E-selectin blockade (GMI-1271), we found elevated levels of donor blood derived CD8+ T-cells demonstrating robust long-term CD8+ T-cell persistence / regeneration (5.3 ±3.2% of total recipient T-cells, p=0.04). This dramatic boost in donor CD8+ T-cell reconstitution in mobilized blood following GMI-1271 co-administration is likely to be due to the long-term persistence and in vivo amplification of CD8+ TCM cells from donor mobilized blood. Similar in vivo enhancing effects of GMI-1271 were also observed with other mobilizing agents such as combined CXCR4 and VLA-4 blockade and GM-CSF resulting in a significant 4.9-fold boost in donor CD8+ reconstitution with GMI-1271. Importantly, only 12 hours of E-selectin blockade was sufficient to achieve this boost in CD8+ TCM numbers in the blood following G-CSF. In a previous report we have shown that therapeutic blockade of E-selectin promotes HSC self-renewal in vivo. Thus, it is possible that E-selectin blockade boosts mobilization of CD8+ TCM/SCM with stem-like properties into the blood by loosening factors retaining CD8+ TCM/SCM in the bone marrow and/or blocking the E-selectin-mediated activation and differentiation of this T-cell subset. In summary, our studies identify E-selectin blockade as a novel target to improve harvesting of CD8+ TCM/SCM cells with stem-like properties. Blockade of this target with GMI-1271 significantly improves the in vivo reconstitution potential and regenerative properties of CD8+ T-cells from donor blood allowing a valuable source of desired T-cells for use in adoptive immunotherapy and T-cell engineering. Disclosures Winkler: GlycoMimetics Inc: Research Funding. Barbier:GlycoMimetics Inc: Research Funding. Davies:GlycoMimetics Inc: Research Funding. Smith:GlycoMimetics, Inc.: Employment. Fogler:GlycoMimetics, Inc.: Employment. Magnani:GlycoMimetics Inc: Employment, Equity Ownership, Membership on an entity's Board of Directors or advisory committees.

scholarly journals Targeted in-vitro-stimulation reveals highly proliferative multi-virus-specific human central memory T cells as candidates for prophylactic T cell therapy

PLoS ONE ◽  
2019 ◽  
Vol 14 (9) ◽  
pp. e0223258 ◽  
Author(s):  
Benjamin Faist ◽  
Fabian Schlott ◽  
Christian Stemberger ◽  
Kevin M. Dennehy ◽  
Angela Krackhardt ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Cell Therapy ◽  
Memory T Cells ◽  
Central Memory ◽  
T Cell Therapy ◽  
Central Memory T Cells

Activation of T and NK Cells Following Lenalidomide Therapy in Patients with Follicular Lymphoma.

Blood ◽  
2012 ◽  
Vol 120 (21) ◽  
pp. 2766-2766
Author(s):  
Seema Rawal ◽  
Nathan Fowler ◽  
Min Zhang ◽  
Zhiqiang Wang ◽  
Tariq Muzzafar ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
B Cell ◽  
Nk Cells ◽  
Research Funding ◽  
Memory T Cells ◽  
T Cell Subsets ◽  
Central Memory T Cells

Abstract Abstract 2766 Background: Lenalidomide plus rituximab therapy is a highly effective and well-tolerated therapy in patients (pts) with follicular lymphoma (FL). In a Phase II trial, this combination induced a complete remission rate of 87% in pts with advanced stage untreated FL (Fowler et al, Ann Oncol, 2011; 22; suppl 4:137). A randomized Phase III trial was recently initiated to compare this combination with current standard of care therapies in pts with FL. Although lenalidomide is known to be an immunomodulatory drug with effects on a variety of immune cells in vitro, its effects have not been well studied in vivo in humans. Understanding the in vivo effects of lenalidomide could lead to novel combination strategies to enhance the efficacy and improve clinical outcome in FL and other malignancies. Methods: Pts received lenalidomide 20 mg/day on days 1–21 of each 28-day cycle and rituximab was given at 375 mg/m2on day 1 of each cycle. Peripheral blood mononuclear cells (PBMC) were phenotyped by multiparametric flow cytometry at baseline, on cycle 2 day 15 (C2D15), and at the end of cycle 6. In addition, peripheral blood (PB) samples were collected in PAXgene Blood RNA tubes at baseline and on C2D15 for whole genome gene expression profiling (GEP). Results: Immunophenotyping of baseline and end of cycle 6 PBMC (n=17) showed that the percentages and absolute numbers of CD3+, CD4+, CD8+, TCRgd, and Foxp3+ regulatory T cells; and NK, NKT, and myeloid dendritic cells were not significantly different between the two time points. However, a significant increase in CD4+CD45RO+ (p<0.01) and CD8+CD45RO+ (p=0.04) memory T cells was observed post-therapy. Further characterization of CD4+ T cells showed a significant increase in central memory T cells (p<0.001) and a decrease in naïve (p<0.01) and terminally differentiated (p<0.01) T cells, but no change in effector memory T cells. The increase in CD8+ central memory T cells was marginally significant (p=0.06). Plasmacytoid dendritic cells (PDC) were also significantly increased (p=0.02). In contrast, no such changes in T cell subsets or PDC were observed in FL pts (n=9) treated with 6 cycles of R-CHOP chemotherapy that received equal number of rituximab doses and analyzed at similar time points (baseline and end of cycle 6). To understand lenalidomide-induced changes on a molecular level, we compared GEP data at C2D15 vs. baseline for 7 pairs of PB samples. The paired significance analysis of microarrays method, based on Student's t test, identified 1,748 differentially expressed genes (DEG; 713 up, 1035 down), without a fold-change threshold, in C2D15 samples vs. baseline. Results were influenced by rituximab-induced depletion of B cells in C2D15 samples, but there were many changes that suggested altered PBMC physiology. Noteworthy up-regulated genes (>1.5 fold) included genes associated with T and NK cell activation including BATF, CCR2, CD1B, CD2, CD160, CTLA4, CXCR3, ICOS, and LAG3; and CD163 and CD209, phagocytic receptors expressed on monocytes/macrophages. Down-regulated genes (>1.5 fold) included CXCR5, which mediates B cell migration into follicles; and IL1B and TNFSF13B (BAFF), which are produced by activated macrophages and induce B cell proliferation. Gene set enrichment analysis of all GEP results, and Ingenuity Pathway Analysis of DEGs, indicated up regulation of multiple pathways and processes including ribosomal and mitochondrial components involved in translation and oxidative phosphorylation, CTLA4 signaling in cytotoxic T cells, and differentiation and signaling by ICOS and CD28 in T helper cells. We confirmed up regulation of CTLA4, ICOS, and LAG3 at the protein level in C2D15 PBMC by flow cytometry. Furthermore, treatment of PBMC derived from untreated FL pts with lenalidomide in vitro resulted in up regulation of these molecules in T and/or NK cells consistent with our in vivo results. Conclusions: In FL pts, lenalidomide induced multiple changes in the immune system including increases in PDC and memory T cell subsets, activation of T and NK cells, and down-regulation of certain genes mediating B cell migration and proliferation. These results provide insights into the mechanism of action of lenalidomide and suggest that it can be combined with other immunostimulatory agents such as therapeutic vaccines, adoptive T cell therapy strategies, and immune checkpoint inhibitors to further enhance its efficacy in FL and other malignancies. Disclosures: Fowler: Celgene: Research Funding. Heise:Celgene Corporation: Employment, Equity Ownership. Lacerte:Celgene: Honoraria. Samaniego:Celgene: Research Funding. Neelapu:Celgene Corporation: Research Funding.

scholarly journals Chemokine Receptor Expression Identifies Pre–T Helper (Th)1, Pre–Th2, and Nonpolarized Cells among Human CD4+ Central Memory T Cells

2004 ◽  
Vol 200 (6) ◽  
pp. 725-735 ◽  
Author(s):  
Laura Rivino ◽  
Mara Messi ◽  
David Jarrossay ◽  
Antonio Lanzavecchia ◽  
Federica Sallusto ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
T Cell Receptor ◽  
Memory T Cells ◽  
Cell Receptor ◽  
Central Memory ◽  
Th2 Cells ◽  
T Helper ◽  
Central Memory T Cells ◽  
Th 1

We previously reported that central–memory T cells (TCM cells), which express lymph node homing receptors CCR7 and CD62L, are largely devoid of effector functions but acquire characteristics of effector–memory T cells (TEM cells) (i.e., CCR7− T helper [Th]1 or Th2 cells) after stimulation with T cell receptor agonists or homeostatic cytokines. Here we show that three chemokine receptors identify functional subsets within the human CD4+ TCM cell pool. TCM cells expressing CXCR3 secreted low amounts of interferon γ, whereas CCR4+ TCM cells produced some interleukin (IL)-4, but not IL-5. In response to IL-7 and IL-15, CXCR3+ TCM and CCR4+ TCM cells invariably generated fully differentiated CCR7− Th1 and Th2 cells, respectively, suggesting that they represent pre-Th1 and pre-Th2 cells. Conversely, CXCR5+ TCM cells lacking CXCR3 and CCR4 remained nonpolarized and retained CCR7 and CD62L expression upon cytokine-driven expansion. Unlike naive cells, all memory subsets had a low T cell receptor rearrangement excision circle content, spontaneously incorporated bromodeoxyuridine ex vivo, and contained cells specific for tetanus toxoid. Conversely, recall responses to cytomegalovirus and vaccinia virus were largely restricted to CXCR3+ TCM and TEM cells. We conclude that antigen-specific memory T cells are distributed between TEM cells and different subsets of TCM cells. Our results also explain how the quality of primary T cell responses could be maintained by TCM cells in the absence of antigen.

scholarly journals Low-dose controlled release of mTOR inhibitors maintains T cell plasticity and promotes central memory T cells

2017 ◽  
Vol 263 ◽  
pp. 151-161 ◽  
Author(s):  
Joshua M. Gammon ◽  
Emily A. Gosselin ◽  
Lisa H. Tostanoski ◽  
Yu-Chieh Chiu ◽  
Xiangbin Zeng ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Controlled Release ◽  
Low Dose ◽  
Memory T Cells ◽  
Mtor Inhibitors ◽  
Central Memory ◽  
Cell Plasticity ◽  
Central Memory T Cells

scholarly journals Lenalidomide Enhances the Function of CS1 Chimeric Antigen Receptor Redirected-T Cells Against Multiple Myeloma

Blood ◽  
2016 ◽  
Vol 128 (22) ◽  
pp. 812-812 ◽  
Author(s):  
Xiuli Wang ◽  
Ryan Urak ◽  
Miriam Walter ◽  
Lihong Weng ◽  
Laura Lim ◽  
...  
Keyword(s):  
Multiple Myeloma ◽  
T Cells ◽  
T Cell ◽  
Memory T Cells ◽  
Central Memory ◽  
Central Memory T Cells ◽  
Car T Cells ◽  
Car T Cell ◽  
Car T

Abstract Multiple myeloma (MM) is an incurable malignancy of plasma cells even with great advances in treatment. Chimeric Antigen Receptor (CAR) directed T cell therapy, which can specifically recognize tumor associated antigens and kill tumor cells in an MHC independent manner, is a promising approach for hematological malignancy. There are several candidate antigens for CAR T cell targeting of multiple myeloma, including BCMA and CS1. Our goal is to develop novel CARs for the treatment of MM and explore the potential benefits of combinatorial therapy of CAR T cells and immunomodulatory drugs (IMiDs) such as lenalidomide. In the present study, we redirected central memory T cells to express second-generation CARs specific for either CS1 or BCMA that incorporate CD28 signaling moieties. Central memory T cells were activated by CD3/CD28 bead stimulation, transduced with lentivirus encoding the CAR construct, and expanded ex vivo. The engineered and expanded CS1 and BCMA CAR T cells exhibited similar phenotypes and comparable in vitro effector function. However, once adoptively transferred into MM tumor-bearing NOD/Scid IL2RγCnull (NSG) mice by intravenous injection of 1x10^6 CAR T cells, CS1 CAR T cells exhibited superior antitumor activity over BCMA CART cells and significantly prolonged mouse survival (P<0.01). To further improve the anti-MM activity of CAR T cell therapy, we investigated the effects of lenalidomide on CS1 CAR T cell function against MM. Central memory T cells were activated and transduced with lentivirus encoding CS1 CAR and then expanded in vitro in the presence of 0, 1 or 10mM lenalidomide for 3-4 weeks and then effector function was evaluated. We found that CD8+ CAR T cells were preferentially expanded over CD4+ CAR T cells in a dose-dependent manner. Lenalidomide-treated CAR T cells secreted higher levels of Th1 cytokines such as IFN-gamma, TNF-alpha, and IL-2, but reduced Th2 cytokines such as IL-4 and IL-10 upon antigen stimulation as compared with untreated CAR T cells. Meanwhile we observed that lenalidomide greatly improved the maintenance of T cell memory markers (CD62L, CD28, and CD27) in the culture and enhanced the formation of immune synapses between CAR T cells and MM cells. RNA-seq analysis revealed that more than 600 genes were differentially expressed among the lenalidomide treated and un-treated CD8+CAR+ T cells. Among those, expression of immune synapse related genes such as cell junction and biological assembly is significantly increased with lenalidomide treatment. Moreover, lenalidomide results in elevated gene transcrips characteristic of memory, homing and cytolytic function of CAR T cells. To test the synergistic effects, MM bearing mice were treated with a single infusion of 1x10^6 CS1 CAR T cells (i.v) on day 0 and/or 5-7.5mgkg-1 of lenalidomide daily (i.p.) initiating on day 0 for 30 days. CS1 CAR T cells and lenalidomide exhibited synergistic anti-MM activity in vivo when MM mice received combinatorial treatment. The combination therapy significantly inhibited tumor growth in vivo, prolonged mouse survival (P<0.01) and improved CAR T cell persistence in mice as compared to single-agent treatment. Taken together, these findings indicate that lenalidomide plays a co-stimulatory role in immune modulation of CAR T cells and strengthens the anti-tumor activity of CS1 CAR T cells in vivo. Rational combination of these immunotherapeutic regimens is an effective strategy and the planned clinical trial will use a combination of lenalidomide and CS1 CAR T cells for increasing treatment efficacy. Disclosures No relevant conflicts of interest to declare.

Imatinib Mesylate Has a Stage-Specific Inhibitory Role in Generation of CD26highCD8+ Central Memory T Cells in Vitro and in Vivo.

Blood ◽  
2008 ◽  
Vol 112 (11) ◽  
pp. 3243-3243
Author(s):  
Kazuaki Yokoyama ◽  
Tokiko Nagamura-Inoue ◽  
Shin Nakayama ◽  
Ikuo Ishige ◽  
Kazuo Ogami ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Cd8 T Cells ◽  
Memory T Cells ◽  
Central Memory ◽  
T Cell Subsets ◽  
Effector Memory ◽  
Short Term ◽  
Central Memory T Cells

Abstract CD26 is a transmembrane glycoprotein with intrinsic dipeptidyl peptidase IV (DPPIV) activity as well as costimulatory activity of mitotic signals triggered by the CD3/TCR complex. Based on the expression level of CD26, CD4+ and CD8+ T cells can be divided into 3 (high/intermediate/low or negative) subsets. The significance of CD26 has been studied mainly on CD4+ T cells, and CD26highCD4+ T cells are considered to represent effector memory T cells of a typical Th1 phenotype producing IL2 and IFNg. Furthermore, we reported a significant decrease of this subset in CML patients under imatinib therapy in comparison to those under IFNa therapy and normal volunteers. In contrast, the role of each subset of CD8+ T cells has not yet been clarified. Multi-parameter flow cytometry analysis was performed to characterize CD8+ T cells differentially expressing CD26 in combination with intracellular detection of effector molecules such as perforin (P) and granzyme B (Gr). The capacity to secrete effector cytokines such as IFNg following short-term stimulation was also assessed. As a result, according to the expression level of CD26, we could clearly categorize CD8+ T cells as follows: CD26highCD8+ T cells are defined as central memory T cells which has a phenotype of CD45RO+CD28+CD27+ IFNg+Gr−P+/−, CD26intCD8+ T cells as naïve T cells of CD45ROCD28+ CD27+ IFNg−Gr−P−, and CD26lowCD8+ T cells as effector memory/effector T cells of CD45RO−/+ CD28−CD27−IFNg++Gr++P++, respectively. We next investigated the effects of imatinib on 3 distinct subsets during CD8+ T cell differentiation program. Peripheral blood mononuclear cells were primed with anti-CD3/CD28 MAb and subjected to the grading doses of imatinib for short term culture, followed by flow cytometory. CFSE labeling was used for monitoring cell proliferation. Intriguingly, we found that imatinib dose-dependently inhibits activation, cytokine production and proliferation of CD26highCD8+ central memory T cell subsets in a differentiation stage-specific manner. Finally, we compared the absolute number of peripheral blood CD26highCD8+ T cell subsets between 20 patients with CML in imatinib-induced CCR and 20 normal volunteers, clearly indicating a significant decrease of this subset in CML patients (22.30/ml vs 45.60/ml, p<0.01). The present study offers another evidence for immunomodulatory effects of imatinib or the critical role of Abl (-related) kinase in T cell development, and draws special attention to susceptibility to viral infection of CML patients under long-term imatinib therapy. Figure Figure

Increased CD8+ central memory T cells in patients with multiple sclerosis

2007 ◽  
Vol 13 (2) ◽  
pp. 149-155 ◽  
Author(s):  
Guang-Zhi Liu ◽  
Li-Bo Fang ◽  
Peter Hjelmström ◽  
Xu-Guang Gao
Keyword(s):  
Multiple Sclerosis ◽  
T Cells ◽  
T Cell ◽  
Peripheral Blood ◽  
Memory T Cells ◽  
Central Memory ◽  
Skewed Distribution ◽  
T Cell Subsets ◽  
Healthy Controls ◽  
Central Memory T Cells

AT-cell-mediated autoimmune process against central nervous system myelin is believed to underlie the pathogenesis of multiple sclerosis (MS). Formation of immunological memory is based on the differentiation of naïve T cells to memory T cells after exposure to antigens and specific cytokines. The aim of this study was to analyse peripheral blood mononuclear cells in patients with MS for different T-cell subsets including naïve and memory T cells. Flow cytometry and enzyme-linked immunosorbent assay were used to analyse memory T-cell subsets and plasma concentration of interleukin-15 (IL-15) in peripheral blood of MS patients, patients with other neurological disorders and healthy controls. MS patients had a skewed distribution of T cells with an increased level of CD8 + /CCR7 + /CD45RA— central memory T cells (TCM) compared to healthy controls. In addition, MS patients showed significantly higher levels of plasma IL-15 than healthy controls did. Upregulated CD8+ TCM in MS patients may reflect a persistent chronic inflammatory response that may have been induced during early stages of the disease. This derangement may be important for maintaining chronic inflammation in MS. Multiple Sclerosis 2007; 13: 149–155. http://msj.sagepub.com

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