Altered Expression of Functional Proteins in CD4 Regulatory T Cells during Therapy with Idelalisib

Blood ◽  
2015 ◽  
Vol 126 (23) ◽  
pp. 1735-1735 ◽  
Author(s):  
Tiago R Matos ◽  
Benjamin L Lampson ◽  
Masahiro Hirakawa ◽  
Siddha Kasar ◽  
Stacey M. Fernandes ◽  
...  
Keyword(s):  
T Cells ◽  
Regulatory T Cells ◽  
Cd8 T Cells ◽  
Cd4 T Cells ◽  
Drug Exposure ◽  
Cell Surface Expression ◽  
Effector Memory ◽  
Functional Markers ◽  
Altered Expression ◽  
And Function

Abstract Introduction: Idelalisib, a specific small molecule inhibitor of the delta isoform of PI3 kinase (PI3Kd), has recently been FDA approved for the treatment of relapsed CLL. Expression of PI3Kd is predominantly restricted to leukocytes, and inhibition of PI3Kd promotes apoptosis of neoplastic CLL cells. Patients who receive idelalisib often experience toxicities that appear to be immune mediated. In an ongoing phase II study of idelalisib-ofatumumab at DFCI, 16 of 21 patients experienced grade 3 or greater autoimmune toxicities (14 transaminitis, 2 enteritis/colitis and 3 presumed drug-induced pneumonitis). Methods: To identify potential causes of autoimmunity in patients receiving idelalisib, we used single cell mass cytometry (CyTOF) with a panel of 26 surface membrane and 9 intracellular markers to provide a comprehensive phenotypic and functional analysis of all peripheral blood lymphocytes. Blood samples were obtained at baseline prior to therapy and at onset of autoimmune symptoms, which occurred at a median of 28 days from start of therapy. Results in 5 severely affected patients were compared with 3 patients who did not develop autoimmune toxicities and 15 healthy donors. The second sample from unaffected patients was matched in time to the affected patient samples. Results: Within CD3 T cells, idelalisib therapy led to an increased percentage of CD4 T cells and decreased percentage of CD8 T cells. Within CD4 T cells, CD25high Foxp3+ regulatory T cells (Treg) were reduced, resulting in a marked increase of the ratio of conventional CD4 T (Tcon)/Treg and increased CD8/Treg ratio. These findings were similar in patients who developed autoimmune toxicities and those who did not. Analysis of effector memory (EM; 45RA- CCR7-), central memory (CM; 45RA- CCR7+), terminal effector (TEMRA; 45RA+ CCR7-) and naive (45RA+ CCR7+) subsets within Treg, Tcon and CD8 T cells was comparable before and after idelalisib therapy and with healthy individual controls. To evaluate the heterogeneity of each population we utilized viSNE, which allows visualization of complex high-dimensional cytometry data on a two-dimensional plot. Treg in patients who developed autoimmune phenomena were compared with Treg from patients who did not. In patients who developed autoimmunity, baseline Treg had higher expression of PD-1 and lower expression of functional markers, such as GITR, Tbet, CXCR3, PDL-1, granzyme-β and TIM-3. In some cases, expression of these functional markers declined further with idelalisib therapy (Figure 1). Cytotoxic T lymphocyte antigen 4 (CTLA4) is constitutively expressed by Treg, and its deficiency has been shown to limit suppressive function of Treg. Inducible co-stimulator (ICOS) also mediates the suppressive functions of CD4 Treg by regulating cell surface expression of CTLA-4. Interestingly, expression of CTLA-4 and ICOS did not change during idelalisib therapy (Figure 1). Susceptibility to apoptosis was monitored by expression of pro-survival Bcl-2 and pro-apoptotic CD95 (FAS). Treg from patients developing autoimmunity expressed lower levels of Bcl-2 and higher levels of CD95 compared to those who did not. Conversely, Treg of non-affected patients retained higher expression of functional markers and became more active, assessed by increased expression of HLA-DR. The use of CyTOF combined with our comprehensive panel allowed us to delineate up to 13 distinct Treg sub-populations among healthy controls. In contrast, CLL patients often lacked such broad Treg heterogeneity. Expression of functional markers by Tcon and CD8 T cells from patients who would develop side effects, were also slightly lower, suggesting that autoimmune toxicities were not due to expansion or activation of CD8 or Tcon effector cells but due to the defect in both number and function of Tregs, which became insufficient to maintain the immune tolerance. Conclusion: These studies allowed us to identify defects in both Treg number and function, which may be responsible for the autoimmune toxicities in patients receiving idelalisib therapy. Further studies may allow the development of a reliable predictor of these toxicities by evaluation of the Treg markers prior to drug exposure which could be used to guide therapy. Moreover, the identification of idelalisib as a Treg inhibitor suggests that it could be used as an immunomodulatory agent to target the Treg pathway, in patients with solid tumors. Disclosures No relevant conflicts of interest to declare.

scholarly journals Expression of the Butyrate/Niacin Receptor, GPR109a on T Cells Plays an Important Role in a Mouse Model of Graft Versus Host Disease

Blood ◽  
2018 ◽  
Vol 132 (Supplement 1) ◽  
pp. 61-61 ◽  
Author(s):  
Melissa D Docampo ◽  
Christoph K. Stein-Thoeringer ◽  
Amina Lazrak ◽  
Marina D Burgos da Silva ◽  
Justin Cross ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Regulatory T Cells ◽  
Cd8 T Cells ◽  
Cd4 T Cells ◽  
Effector Memory ◽  
Knock Out ◽  
Graft Versus Host

Abstract INTRODUCTION: The intestinal microbiota is essential for the fermentation of fibers into the short-chain fatty acids (SCFA) butyrate, acetate and propionate. SCFA can bind to G-protein-coupled receptors GPR41, GPR43 and GPR109a to activate downstream anti-inflammatory signaling pathways. In colitis or graft versus host disease (GVHD), GPR43 signaling has been reported as an important regulator of intestinal homeostasis by increasing the pool of regulatory T cells. In contrast to GPR43, which binds preferentially propionate and acetate, GPR109a is the major receptor for butyrate. We and others have demonstrated that butyrate can ameliorate gastrointestinal injury during GVHD through enterocyte protection. Therefore, we hypothesized that GPR109a plays an important role in the pathophysiology of intestinal GVHD, focusing specifically on alloreactive T cells. METHODS AND RESULTS: Using mouse models of GVHD, we examined the role of the butyrate/niacin receptor, GPR109a in allogeneic hematopoietic cell transplantation (allo-HCT). First, we studied whether a genetic knock-out (KO) of GPR109a in transplant recipient mice affected GVHD, but GPR109a-KO recipient mice did not exhibit increased mortality from GVHD compared to wild type (WT) mice. We next investigated the role of GPR109a in the donor compartment by transplanting either BM or T cells from WT or GPR109a-KO mice into major MHC mismatched BALB/c host mice. Mice transplanted with B6 BM, with T cells from a GPR109a-KO mouse into BALB/c hosts displayed a lower incidence of lethal GVHD (Fig. 1A). To determine whether the attenuation of GVHD is intrinsic to GPR109a-KO T cells, we established BM chimeras and performed a secondary transplant by transplanting B6 BM + (B6 à Ly5.1) or (GPR109a à Ly5.1) T cells into BALB/c hosts. We observed the same improvement in survival in mice that received GPR109a-KO T cells. This indicates an intrinsic role for GPR109a specifically in the donor hematopoietic compartment. Having identified a T-cell specific requirement for GPR109a we next examined expression of GPR109a on WT T cells in vitro at baseline and following stimulation with CD3/28 and found GPR109a significantly upregulated on both CD4+ and CD8+ T cells after 72 hours of stimulation (Fig 1B). At steady state in vivo, we observed the same numbers and percentages of splenic effector memory, central memory, and naïve CD4+ T cells as well as regulatory T cells in WT B6 mice and GPR109a-KO mice, suggesting normal T cell development in the knockout mice. In an in vitro mixed lymphocyte reaction (MLR), GPR109a-KO CD4+ T cells become activated, proliferate, polarize and secrete cytokine (specifically IFNg) to the same level as WT CD4+ T cells, suggesting normal functional capacity. However, after allo-HCT in mice we observed significantly fewer CD4+ and CD8+ T cells, and specifically fewer effector memory CD4+ T cells (Fig. C), in the small and large intestines of mice that received GPR109a-KO T cells at day 7 post transplant. In contrast, we found significantly more regulatory T cells in the intestines (Fig. 1D) and the spleen of GPR1091-KO T cell recipients, while numbers and percentages of polarized Th1 and Th17 T cells were similar between the two groups. We further 16S rRNA sequenced the gut microbiota of mice at day 7 after transplant and observed an increased relative abundance of bacteria from the genus Clostridium (Fig. 1D) along with an increased concentration of cecal butyrate as measured by GC-MS (Fig. 1E). In a preliminary graft versus tumor (GVT) experiment, we found that mice that received A20 tumor cells and GPR109a-KO T cells exhibited increased survival compared to mice that received A20 tumor cells and WT T cells. These preliminary findings suggest that GPR109a-KO T cells maintain their graft versus tumor response while causing less GVHD, and exclude a defective functional capacity. CONCLUSIONS: We report a novel role of the butyrate/niacin receptor GPR109a on donor T cells in allo-HCT as a genetic knock-out on T cells attenuates lethal GVHD. As these T cells are tested as functionally intact, we propose that the reduction in overall T cells of KO T cell recipients may underlie the attenuation in GVHD. Furthermore, such a reduction in allograft-induced gut injury is accompanied by maintenance of the gut commensal Clostridium and butyrate production, which is known to protect the intestinal epithelium and increases the regulatory T cell pool. Disclosures No relevant conflicts of interest to declare.

Proliferation and Activation Patterns of Naïve, Memory and Regulatory T Cells in Patients with Multiple Myeloma During Thalidomide, Interferon-α and Bortezomib Maintenance Therapy.

Blood ◽  
2009 ◽  
Vol 114 (22) ◽  
pp. 3880-3880
Author(s):  
Michael Hundemer ◽  
Isabelle Herth ◽  
Tobias Meissner ◽  
Dirk Hose ◽  
Anthony D Ho ◽  
...  
Keyword(s):  
Multiple Myeloma ◽  
T Cells ◽  
Regulatory T Cells ◽  
Maintenance Therapy ◽  
Cd8 T Cells ◽  
Cd4 T Cells ◽  
Granzyme B ◽  
Effector Memory ◽  
Control Group ◽  
Ifn Γ

Abstract Abstract 3880 Poster Board III-816 Hundemer and Herth (Contributed equally) Introduction In patients with Multiple myeloma, maintenance therapy after high-dose chemotherapy and autologous stem cell transplantation is performed with the aim to prolong remission duration and survival. Beside IFN-α, thalidomide and bortezomib are increasingly applied in maintenance protocols. In this prospective study we have analysed the implication of the various types of maintenance therapy on the patients T cell pattern and activation status. Patients and Methods T cells from 63 patients in clinical remission were analyzed. The median duration of remission was 38,6 months. Eighteen patients were treated with IFN-α, 22 with thalidomide, 7 with bortezomib and 16 patients received no maintenance therapy (control group). Peripheral blood mononuclear cells were isolated and stimulated with CD3/CD28 beads. Activated and nonactivated T cells were analyzed by flow cytometry (CD45RA, CD45RO, CCR7, CD28, CD200R, CD95, CD279, CD69, CD134 and TCRγ/δ) and ELISA (IFN-γ, perforine and granzym B). Furthermore the rate of IFN-γ-producing and regulatory T cells were analyzed by intracytoplasmatic staining and flow cytometry. Results All groups including the control group showed an up-regulation of CD69 and CD134 on CD4+ and CD8+ T cells after activation (p<0,001), on CD8+ T cells in the bortezomib-group only CD69 was upregulated (p=0,008). Patients treated with IFN-α showed a high rate of naïve T cells (CD45RA- and CCR7-positive), while in the thalidomide-group a high rate of effector memory T-cells (CD45RA- and CCR7-negative) were observed (CD45RA on CD8+ and CD4+ T cells: p<0,001, CCR7 on CD8+ T cells: p=0,03, CCR7 on CD4+ T cells: p=0,003). Regarding the surface marker CD28 on CD8+ T cells the IFN-α-group demonstrated a significant higher expression than the control-group (p=0,04) and the bortezomib-group a significant lower expression than the IFN-α- and the thalidomide-group (p=0,006 and p=0,02). Furthermore the rate of IFN-γ-producing CD4+ T cells was significant higher in the thalidomide-group than in the IFN-α-group after activation (p=0,02). On the basis of the cytoplasmatic staining of Foxp3 there was a trend to a higher amount of regulatory T cells in the thalidomide-group compared to the IFN-α-group (p=0,07). Analysis of IFN-y secretion by ELISA, an increases IFN-γ secretion could be demonstrated in all groups after activation (control group: p=0,002, IFN-α-group: p<0,001, thalidomide-group: p<0,001, bortezomib group: p=0,01), furthermore in all groups despite the bortezomib-group an increase of the granzyme B-production can be observed (control group: p=0,003, IFN-α-group: p=0,03, thalidomide-group: p<0,001). Regarding the activated state of the T cells the production of IFN-γ, perforine and granzyme B was significant higher in the thalidomide-group than in the IFN-α-group (IFN-γ: p=0,05, perforine: p=0,02, granzyme B: p=0,04). Furthermore the nonactivated and the activated T cells of the patients treated with thalidomide showed a significant higher production of granzyme B than the T cells of the control group (p=0,0003 and p=0,006). Conclusion During maintenance therapy, thalidomide promotes maturation and proliferation of effector memory T cells and regulatory T cells, while IFN-α treatment increases the number of naïve T cells and subsequently, the T cell activation in the thalidomide group was significantly higher than in the IFN-α group. These results have profound impact on the development of novel immunomodulating therapy strategies in the treatment of multiple myeloma. Disclosures: No relevant conflicts of interest to declare.

scholarly journals Elevated frequencies of highly activated CD4+ T cells in HIV+ patients developing immune reconstitution inflammatory syndrome

Blood ◽  
2010 ◽  
Vol 116 (19) ◽  
pp. 3818-3827 ◽  
Author(s):  
Lis R. V. Antonelli ◽  
Yolanda Mahnke ◽  
Jessica N. Hodge ◽  
Brian O. Porter ◽  
Daniel L. Barber ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Regulatory T Cells ◽  
Immune Reconstitution Inflammatory Syndrome ◽  
Cd4 T Cells ◽  
Immune Reconstitution ◽  
Interferon Γ ◽  
Effector Memory ◽  
Hiv Patients ◽  
Inflammatory Syndrome

Abstract Immune reconstitution inflammatory syndrome (IRIS) is a considerable problem in the treatment of HIV-infected patients. To identify immunologic correlates of IRIS, we characterized T-cell phenotypic markers and serum cytokine levels in HIV patients with a range of different AIDS-defining illnesses, before and at regular time points after initiation of antiretroviral therapy. Patients developing IRIS episodes displayed higher frequencies of effector memory, PD-1+, HLA-DR+, and Ki67+ CD4+ T cells than patients without IRIS. Moreover, PD-1+ CD4+ T cells in IRIS patients expressed increased levels of LAG-3, CTLA-4, and ICOS and had a Th1/Th17 skewed cytokine profile upon polyclonal stimulation. Elevated PD-1 and Ki67 expression was also seen in regulatory T cells of IRIS patients. Furthermore, IRIS patients displayed higher serum interferon-γ, compared with non-IRIS patients, near the time of their IRIS events and higher serum interleukin-7 levels, suggesting that the T-cell populations are also exposed to augmented homeostatic signals. In conclusion, our findings indicate that IRIS appears to be a predominantly CD4-mediated phenomenon with reconstituting effector and regulatory T cells showing evidence of increased activation from antigenic exposure. These studies are registered online at http://clinicaltrials.gov as NCT00557570 and NCT00286767.

The Role of IL-12 and IL-23 in the Generation and Maintenance of Memory CD4+ T Cells.

Blood ◽  
2004 ◽  
Vol 104 (11) ◽  
pp. 2664-2664
Author(s):  
Aileen M. Cleary ◽  
David B. Lewis
Keyword(s):  
T Cells ◽  
Cd4 T Cells ◽  
Chemokine Receptors ◽  
Human Memory ◽  
Surface Expression ◽  
Effector Memory ◽  
Memory Cells ◽  
Peripheral Lymph ◽  
And Function ◽  
Memory Cd4 T Cells

Abstract Human memory CD4+ T cells can be distinguished from antigenically-naïve CD4+ T cells based on their CD45RAlowCD45R0high and CD45RAhighCD45R0low surface phenotypes, respectively. Memory CD4+ T cells from adult peripheral blood can be further divided based on surface expression of the CCR7 chemokine receptor. Th1 memory CD4+ T cells that are CCR7high (putative central memory cells or Tcm) are expected to be preferentially targeted to peripheral lymph nodes where the ligands for CCR7 are expressed in high amounts. These cells have been reported to lack expression of the CCR3 and CCR5 chemokine receptors, which facilitate entry into inflamed tissues, and produce little or no interferon (IFN)-γ after stimulation via the αβ-TCR/CD3 complex. CD45RAlowCD45R0highCCR7low CD4+ T cells account for virtually all IFN-g production by human CD4 T cells after ab-TCR/CD3 stimulation using monoclonal antibodies, and for this reason were termed effector memory cells (Tem). These findings, as well as the observation of shorter telomere lengths for memory CD4+ T cells that are CCR7low compared to those that CCR7high suggest that the Tcm population may be an intermediate between naïve CD4+ T cells and Tem. It has recently been proposed that the level of signal strength and γc containing cytokines play a role in memory T cell generation. However, little is known whether IL-12 or IL-23 are necessary and for this differentiation and/or maintenance. Our laboratory has previously described a patient with IL-12Rβ1 deficiency, which ablates both IL-12 and IL-23 signaling. This patient had a deficiency in Tem number and function, unexpectedly suggesting that IL-12 and/or IL-23 may play a key role in this process. We therefore hypothesized that signaling through IL-12Rβ1 plays a key role in the latter stages of generation and/or maintenance of human memory CD4+ T cells. Preliminary data thus far show CCR7 expression to be slightly decreased on activated Tcm in response to incubation with IL-2 or IL-12 alone, and to a greater extent with IL-2 and IL-12 incubated together. In addition, spontaneous apoptosis of both Tcm and Tem is decreased upon incubation with IL-12. Taken together, these data suggest that IL-12 may play a role in both generation of Tem and maintenance of both Tcm and Tem.

Ex Vivo Fludarabine Selectively Depletes Human Naive T-Cell Subsets: A Step Toward Modifying Donor Lymphocyte Infusion.

Blood ◽  
2005 ◽  
Vol 106 (11) ◽  
pp. 1071-1071
Author(s):  
Melody M. Smith ◽  
Cynthia R. Giver ◽  
Edmund K. Waller ◽  
Christopher R. Flowers
Keyword(s):  
T Cells ◽  
T Cell ◽  
Cd8 T Cells ◽  
Cd4 T Cells ◽  
Memory T Cells ◽  
Ex Vivo ◽  
T Cell Subsets ◽  
Effector Memory ◽  
Naïve T Cell ◽  
Naive T Cell

Abstract Ex vivo modification of donor lymphocytes with purine analogs (mDL) may help to minimize graft versus host disease (GvHD) while providing beneficial graft versus leukemia (GvL) effects. In a murine model system, we have shown that allogeneic donor splenocytes, treated with fludarabine ex vivo have significantly reduced GvHD activity when transferred to irradiated recipient mice, and retain anti-viral and GvL activities (Giver, 2003). This effect appears to be mediated by relative depletion of donor CD4 CD44low, “naive” T-cells. As a first step toward developing mDL for use in patients, we sought to evaluate the effects of ex vivo fludarabine exposure on human T-cell subsets, and to determine the minimum dose of fludarabine required to achieve this effect. Methods: Peripheral blood mononuclear cell samples from 6 healthy volunteers were evaluated at 0, 24, 48, and 72 hour time points after ex vivo incubation in varying dosages of fludarabine: 2, 5, and 10(n=3) mcg/ml. Fludarabine incubated samples were compared to samples that received no fludarabine (untreated). The total viable cell number was determined and the fractions and absolute numbers of viable CD4 and CD8 naïve and memory T-cells were determined using flow cytometry after incubation with 7-AAD (dead cell stain), CD4, CD8, CD45RA, CD62L, and CCR7 antibodies, and measuring the total viable cells/ml. Results: The numbers of viable CD4 and CD8 T-cells remained relatively stable in control cultures. Without fludarabine, the average viability at 72 hr of naive and memory T-cells were 92% and 77% for CD4 and 86% and 63% for CD 8 (Fig. 1A). Naive CD4 T-cells were more sensitive to fludarabine-induced death than memory CD4 cells. At 72 hr, the average viability of fludarabine-treated naive CD4 T-cells was 33% at 2 mcg/ml (8.2X the reduction observed in untreated cells) and 30% at 5 mcg/ml, while memory CD4 T-cells averaged 47% viability at 2 mcg/ml (2.3X the reduction observed in untreated cells) (Fig. 1B) and 38% at 5 mcg/ml. The average viability of naive CD8 T-cells at 72 hr was 27% at 2 mcg/ml and 20% at 5 mcg/ml, while memory CD8 T-cell viability was 22% at 2 mcg/ml and 17% at 5 mcg/ml. Analyses on central memory, effector memory, and Temra T-cells, and B-cell and dendritic cell subsets are ongoing. The 5 and 10 mcg/ml doses also yielded similar results in 3 initial subjects, suggesting that 2 mcg/ml or a lower dose of fludarabine is sufficient to achieve relative depletion of the naive T-cell subset. Conclusions: Future work will determine the minimal dose of fludarabine to achieve this effect, test the feasibility of using ex vivo nucleoside analog incubation to reduce alloreactivity in samples from patient/donor pairs, and determine the maximum tolerated dose of mDL in a phase 1 clinical trial with patients at high risk for relapse and infectious complications following allogeneic transplantation. Figure Figure

Altered Naive and Memory T Cell Homeostasis and Immunosenescence Characterize Younger Patients with Immunosuppressive-Responsive Myelodysplastic Syndrome

Blood ◽  
2008 ◽  
Vol 112 (11) ◽  
pp. 3647-3647
Author(s):  
JianXiang Zou ◽  
Dana E Rollison ◽  
David Boulware ◽  
Elaine M. Sloand ◽  
Loretta Pfannes ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Immunosuppressive Therapy ◽  
Cd8 T Cells ◽  
Cd4 T Cells ◽  
Effector Memory ◽  
Cell Populations ◽  
T Cell Homeostasis ◽  
Cell Homeostasis ◽  
Younger Patients

Abstract BACKGROUND: A subset of patients with Myelodysplastic Syndrome (MDS) responds well to immunosuppressive therapy (IST) and the only validated predictor of response is age, with younger patients faring much better than older patients. Hematologic improvement on immunosuppressive therapy is associated with a survival benefit with response rates ranging from 15% to 50%, clearly comparable or better than results with other existing therapies in MDS. Despite progress in the basic understanding of immune pathobiology of MDS and a clear therapeutic value, including improved long-term survival, IST including anti-thymocyte globulin (ATG) and/or cyclosporine A (CyA) is rarely offered to MDS patients in the U.S. due to uncertain criteria for selection of patients and potential toxicities. In addition, there is an underlying concern that inappropriate use of immunosuppressive therapy may negatively impact risk for leukemia progression, which occurs in 30–40% of MDS cases. The long-term goal of this study is to identify an immune signature that has postive predictive power for IST responsiveness. METHODS: To determine the effect of age on T-cell homeostasis and function and IST response, we performed a study of 54 MDS patients compared to 37 healthy controls. In a pilot study, T cell abnormalities associated with response to equine anti-lymphocyte globulin (eATG, lymphoglobulin, Pfizer, Inc) and/or CyA was studied in 12 younger MDS patients composed of 6 responders and 6 non-responders. RESULTS: CD4+ T-cells are normally present in the peripheral blood lymphocyte pool at 2 to 4 times greater than that of CD8+ T-cells, and diminished CD4:CD8 ratio has been previously shown to correlate with poor survival outcome in MDS. Similar to previous reports, we found that the age-adjusted CD4:CD8 ratio was reduced in MDS patients compared to healthy controls (p-value <0.0001) Interestingly, our analysis revealed that inadequate CD4+ rather than expansion of CD8+ T-cells was associated with a lower ratio in this group of MDS patients that included both lower and higher risk MDS patients defined by the International Prognostic Scoring System (IPSS). Analysis of the percentage of T-cells with naïve and memory phenoytpes using CD45RA and CD62L display, demonstrated positive correlations between age and both % CD62L positive naïve cells and central memory CD4+ T-cells (naïve: slope=0.39, p=0.12; central memory: slope=1.26, p=0.005). Furthermore, the proportions of CD62L- CD4+ T-cell populations, including effector memory and terminal effector memory T-cells, were greater in younger MDS patients (slope=−0.82, p=0.08 and slope=−0.83, p=0.015, respectively) suggesting a possible relationship to IST responsiveness. Specific characteristics associated with response to eATG in the pilot study of 12 younger patients included altered distribution of T cell populations (i.e., lower CD4/CD8 ratio, p<0.001) and higher constitutive proliferative index of the T cell populations (p=0.03 CD4+ and p=0.02 CD8+ T-cells, respectively). We also found that hematological response was associated with blockade of homeostatic proliferation of T cells associated with reconstitution of the naïve T cell pool. Reduction in CD4+ T-cells and expansion of autoreactive CD8+ T-cells suggests that apoptotic conditions may drive the expansion of cells through homeostatic cytokines such as IL-7, IL-15, and/or IL-21, which are all cytokines of the IL-2Rγc family that control homeostatic proliferation. Comparisons of the IL-7Ra, IL-15Ra, IL-2Ra, and IL-21Ra subunit demonstrated overexpression of IL-21Ra in patients 35.4% ± 3.4 in CD4+ T-cells and 31.8% ± 4.3 in CD8+ T-cells compared to healthy donors 0.9% ± 0.5 and 0.5% ± 0.5 (p<0.0001). CONCLUSIONS: Association between the T-cell abnormalities reported in this study and response to IST strongly suggests that aberrant T-cell homeostasis may represent a critical determinant of autoimmunity in MDS that may have positive predictive power for response to IST.

Functional T Cell Array Reveals Pre-Treatment Expanded Terminal Effector Memory Response Is Associated with Lenalidomide Failure in Non-Del(5q) Myelodysplastic Syndrome.

Blood ◽  
2009 ◽  
Vol 114 (22) ◽  
pp. 1759-1759
Author(s):  
P.K. Epling-Burnette ◽  
JianXiang Zou ◽  
Jeffrey S. Painter ◽  
Dung-Tsa Chen ◽  
Jimmy Fulp ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Myelodysplastic Syndrome ◽  
Cd8 T Cells ◽  
Cd4 T Cells ◽  
Erythroid Differentiation ◽  
T Cell Response ◽  
Effector Memory ◽  
Terminal Effector

Abstract Abstract 1759 Poster Board I-785 Lenalidomide (LEN) is a thalidomide derivative with proven efficacy for the treatment of patients with myelodysplastic syndrome (MDS). High rates of erythroid and cytogenetic response in patients with chromosome 5q deletion [del(5q)] produced the first FDA-approved karyotype-specific treatment for this disease. Transfusion-independence rates of approximately 25% have been reported previously for patients with non-[del(5q)] and efficacy in this population has been linked to the promotion of erythroid differentiation. Because impaired erythroid differentiation in lower-risk MDS may occur through several pathophysiological mechanisms, the identification of additional factors with predictive value for both response and failure to LEN are needed to optimize success of treatment. In addition to affecting erythroid differentiation, LEN has well-known potential for immune modulation and generates highly potent effector T cell responses in vitro and in vivo by potentiating T cell receptor signaling. Immune deregulation mediated by autoreactive effector T cells has been linked to impaired erythropoiesis and granulopoiesis in a distinct subset of MDS patients raising the question of whether LEN impacts the disease process in this subset of patients. To understand the relationship between T cell deregulation and LEN response, we conducted a pilot study of 13 low/INT-1-risk non-del (5q) MDS patients (7 responders and 6 non-responders) treated with LEN and determined 23 covariates related to functional T cell response measured prior to treatment and then correlated to treatment outcome. Of these 23 covariates, multiple T cell immune parameters were analyzed but were not associated with response including interferon-g (IFNg) production by CD4+ T cells (p=0.9) and CD8+ T cells (p=0.27), Tumor Necrosis Factor (TNF)-a production by CD4+ T cells (p=1.0) and CD8+ T cells (p=0.8), TCR-associated proliferation within the CD4+ (p=1.0) and CD8+ (p=0.4) compartment, CD4/CD8 ratio (p=0.3), percentage of CD4+ (p=0.5) and CD8+ (p=0.5) T lymphocytes, and the percentage of naïve and three different types of memory CD4+ T cells. Analysis was performed using two-group comparison statistical tests (two-sample t-test and Wilcoxon rank sum test) to compare responders (R) vs non-responders (NR). Only one factor was independently linked to LEN response. Results showed that a higher percentage of CD8 T cells (mean 56% in NR vs 32% in R) with a Terminal Effector Memory [TEM]) phenotype (CD45RA+/CD62L-) was associated with LEN failure (p=0.02). This population of T cells occurs at a low frequency in healthy individuals but can be induced to differentiate in vitro under constant exposure to long-term antigen and cytokine stimulation. We have shown previously that CD8+ TEM T cells are expanded in patients with impaired myelopoiesis due to immune dysregulation in Large Granular Lymphocyte (LGL) leukemia. In conclusion, these results suggest that CD8+ terminal effector memory expansion may be linked to immune deregulation in MDS and represents an important biomarker with negative predictive importance for LEN response in non-del(5q) low-risk MDS. Disclosures No relevant conflicts of interest to declare.

Anti-GvHD Effect of Antithymocyte Globulin Is Mediated by Antibodies Binding to T Cells and Regulatory NK Cells, and Less So or Not at All by Antibodies Binding to Other Mononuclear Cell Subsets

Blood ◽  
2010 ◽  
Vol 116 (21) ◽  
pp. 2346-2346
Author(s):  
Mette Hoegh-Petersen ◽  
Minaa Amin ◽  
Yiping Liu ◽  
Alejandra Ugarte-Torres ◽  
Tyler S Williamson ◽  
...  
Keyword(s):  
T Cells ◽  
Dendritic Cells ◽  
B Cells ◽  
Nk Cells ◽  
Cd8 T Cells ◽  
Cd4 T Cells ◽  
Antithymocyte Globulin ◽  
Chronic Gvhd ◽  
Wilcoxon Test ◽  
Effector Memory

Abstract Abstract 2346 Introduction: Polyclonal rabbit-anti-human T cell globulin may decrease the likelihood of graft-vs-host disease (GVHD) without increasing the likelihood of relapse. We have recently shown that high levels of antithymocyte globulin (ATG) capable of binding to total lymphocytes are associated with a low likelihood of acute GVHD grade 2–4 (aGVHD) as well as chronic GVHD needing systemic therapy (cGVHD) but not increased likelihood of relapse (Podgorny PJ et al, BBMT 16:915, 2010). ATG is polyclonal, composed of antibodies for antigens expressed on multiple cell subsets, including T cells, B cells, NK cells, monocytes and dendritic cells. These cell subsets may play a role in the pathogenesis of GVHD. The anti-GVHD effect of ATG may be mediated through killing/inhibition of one or several of these cell subsets (eg, T cells) or their subsets (eg, naïve T cells as based on mouse experiments naïve T cells are thought to play a major role in the pathogenesis of GVHD). To better understand the mechanism of action of ATG on GVHD, we set out to determine levels of which ATG fraction (capable of binding to which cell subset) are associated with subsequent development of GVHD. Patients and Methods: A total of 121 patients were studied, whose myeloablative conditioning included 4.5 mg/kg ATG (Thymoglobulin). Serum was collected on day 7. Using flow cytometry, levels of the following ATG fractions were determined: capable of binding to 1. naïve B cells, 2. memory B cells, 3. naïve CD4 T cells, 4. central memory (CM) CD4 T cells, 5. effector memory (EM) CD4 T cells, 6. naïve CD8 T cells, 7. CM CD8 T cells, 8. EM CD8 T cells not expressing CD45RA (EMRA-), 9. EM CD8 T cells expressing CD45RA (EMRA+), 10. cytolytic (CD16+CD56+) NK cells, 11. regulatory (CD16-CD56high) NK cells, 12. CD16+CD56− NK cells, 13. monocytes and 14. dendritic cells/dendritic cell precursors (DCs). For each ATG fraction, levels in patients with versus without aGVHD or cGVHD were compared using Mann-Whitney-Wilcoxon test. For each fraction for which the levels appeared to be significantly different (p<0.05), we determined whether patients with high fraction level had a significantly lower likelihood of aGVHD or cGVHD than patients with low fraction level (high/low cutoff level was determined from ROC curve, using the point with maximum sum of sensitivity and specificity). This was done using log-binomial regression models, ie, multivariate analysis adjusting for recipient age (continuous), stem cell source (marrow or cord blood versus blood stem cells), donor type (HLA-matched sibling versus other), donor/recipient sex (M/M versus other) and days of follow up (continuous). Results: In univariate analyses, patients developing aGVHD had significantly lower levels of the following ATG fractions: binding to naïve CD4 T cells, EM CD4 T cells, naïve CD8 T cells and regulatory NK cells. Patients developing cGVHD had significantly lower levels of the following ATG fractions: capable of binding to naïve CD4 T cells, CM CD4 T cells, EM CD4 T cells, naïve CD8 T cells and regulatory NK cells. Patients who did vs did not develop relapse had similar levels of all ATG fractions. In multivariate analyses, high levels of the following ATG fractions were significantly associated with a low likelihood of aGVHD: capable of binding to naïve CD4 T cells (relative risk=.33, p=.001), EM CD4 T cells (RR=.30, p<.001), naïve CD8 T cells (RR=.33, p=.002) and regulatory NK cells (RR=.36, p=.001). High levels of the following ATG fractions were significantly associated with a low likelihood of cGVHD: capable of binding to naïve CD4 T cells (RR=.59, p=.028), CM CD4 T cells (RR=.49, p=.009), EM CD4 T cells (RR=.51, p=.006), naïve CD8 T cells (RR=.46, p=.005) and regulatory NK cells (RR=.55, p=.036). Conclusion: For both aGVHD and cGVHD, the anti-GVHD effect with relapse-neutral effect of ATG appears to be mediated by antibodies to antigens expressed on naïve T cells (both CD4 and CD8), EM CD4 T cells and regulatory NK cells, and to a lesser degree or not at all by antibodies binding to antigens expressed on B cells, cytolytic NK cells, monocytes or DCs. This is the first step towards identifying the antibody(ies) within ATG important for the anti-GVHD effect without impacting relapse. If such antibody(ies) is (are) found in the future, it should be explored whether such antibody(ies) alone or ATG enriched for such antibody(ies) could further decrease GVHD without impacting relapse. Disclosures: No relevant conflicts of interest to declare.

scholarly journals CD8+CD44hi but not CD4+CD44hi memory T cells mediate potent graft antilymphoma activity without GVHD

Blood ◽  
2011 ◽  
Vol 117 (11) ◽  
pp. 3230-3239 ◽  
Author(s):  
Suparna Dutt ◽  
Jeanette Baker ◽  
Holbrook E. Kohrt ◽  
Neeraja Kambham ◽  
Mrinmoy Sanyal ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Cd8 T Cells ◽  
Cd4 T Cells ◽  
Memory T Cells ◽  
Cell Subset ◽  
B Cell Lymphoma ◽  
Effector Memory ◽  
T Cell Subset ◽  
Progressive Growth

Abstract Allogeneic hematopoietic cell transplantation can be curative in patients with leukemia and lymphoma. However, progressive growth of malignant cells, relapse after transplantation, and graft-versus-host disease (GVHD) remain important problems. The goal of the current murine study was to select a freshly isolated donor T-cell subset for infusion that separates antilymphoma activity from GVHD, and to determine whether the selected subset could effectively prevent or treat progressive growth of a naturally occurring B-cell lymphoma (BCL1) without GVHD after recipients were given T cell–depleted bone marrow transplantations from major histocompatibility complex–mismatched donors. Lethal GVHD was observed when total T cells, naive CD4+ T cells, or naive CD8+ T cells were used. Memory CD4+CD44hi and CD8+CD44hi T cells containing both central and effector memory cells did not induce lethal GVHD, but only memory CD8+ T cells had potent antilymphoma activity and promoted complete chimerism. Infusion of CD8+ memory T cells after transplantation was able to eradicate the BCL1 lymphoma even after progressive growth without inducing severe GVHD. In conclusion, the memory CD8+ T-cell subset separated graft antilymphoma activity from GVHD more effectively than naive T cells, memory CD4+ T cells, or memory total T cells.

scholarly journals c-Jun NH2-Terminal Kinase (JNK)1 and JNK2 Have Distinct Roles in CD8+ T Cell Activation

2002 ◽  
Vol 195 (7) ◽  
pp. 811-823 ◽  
Author(s):  
Dietrich Conze ◽  
Troy Krahl ◽  
Norman Kennedy ◽  
Linda Weiss ◽  
Joanne Lumsden ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
T Cell Activation ◽  
Cd8 T Cells ◽  
Cd4 T Cells ◽  
Cell Activation ◽  
Cd8 T Cell ◽  
Cell Surface Expression ◽  
Chain Gene ◽  
Α Chain

The c-Jun NH2-terminal kinase (JNK) signaling pathway is induced by cytokines and stress stimuli and is implicated in cell death and differentiation, but the specific function of this pathway depends on the cell type. Here we examined the role of JNK1 and JNK2 in CD8+ T cells. Unlike CD4+ T cells, the absence of JNK2 causes increased interleukin (IL)-2 production and proliferation of CD8+ T cells. In contrast, JNK1-deficient CD8+ T cells are unable to undergo antigen-stimulated expansion in vitro, even in the presence of exogenous IL-2. The hypoproliferation of these cells is associated with impaired IL-2 receptor α chain (CD25) gene and cell surface expression. The reduced level of nuclear activating protein 1 (AP-1) complexes in activated JNK1-deficient CD8+ T cells can account for the impaired IL-2 receptor α chain gene expression. Thus, JNK1 and JNK2 play different roles during CD8+ T cell activation and these roles differ from those in CD4+ T cells.

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