Pentostatin Is Advantageous Relative to Fludarabine for in Vivo Murine T Cell Depletion, Suppression of T Cell Cytokine Secretion, and Inhibition of HVGR

Blood ◽  
2008 ◽  
Vol 112 (11) ◽  
pp. 3483-3483
Author(s):  
Jacopo Mariotti ◽  
Jason Foley ◽  
Kaitlyn Ryan ◽  
Nicole Buxhoeveden ◽  
Daniel Fowler
Keyword(s):  
T Cells ◽  
T Cell ◽  
Cd8 T Cells ◽  
Cytokine Secretion ◽  
Cell Depletion ◽  
T Cell Depletion ◽  
T Cell Suppression ◽  
Ifn Γ ◽  
Cell Suppression

Abstract Although fludarabine and pentostatin are variably utilized for conditioning prior to clinical allogeneic transplantation, limited data exists with respect to their relative efficacy in terms of host immune T cell depletion and T cell suppression. To directly compare these agents in vivo in a murine model, we compared a regimen of fludarabine plus cyclophosphamide (FC) similar to one that we previously developed (Petrus et al, BBMT, 2000) to a new regimen of pentostatin plus cyclophosphamide (PC). Cohorts of mice (n=5–10) received a three-day regimen consisting of P alone (1 mg/kg/d), F alone (100 mg/kg/d), C alone (50 mg/kg/d), or combination PC or FC. Similar to our previous data, administration of P, F, or C alone yielded minimal host T cell depletion (as measured by enumeration of splenic CD4+ and CD8+ T cells) and minimal T cell suppression (as determined by CD3, CD28 co-stimulation of a constant number of remaining splenic T cells and measuring resultant cytokine secretion by multi-analyte assay). The PC and FC regimens were similar in terms of myeloid suppression (p=.2). However, the PC regimen was more potent in terms of depleting host CD4+ T cells (remaining host CD4 number [× 10^6/spleen], 2.1±0.3 [PC] vs. 4.4±0.6 [FC], p<0.01) and CD8+ T cells (remaining host CD8 number, 1.7±0.2 [PC] vs. 2.4±0.5 [FC], p<0.01). Moreover, the PC regimen yielded greater T cell immune suppression than the FC regimen (cytokine values are pg/ml/0.5×10^6 cells/ml; all comparisons p<0.05) with respect to capacity to secrete IFN-γ (13±5 [PC] vs. 48±12 [FC]), IL-2 (59±44 [PC] vs. 258±32 [FC]), IL-4 (34±10 [PC] vs. 104±12 [FC]), and IL-10 (15±3 [PC] vs. 34±5 [FC]). In light of this differential in both immune T cell depletion and suppression of T cell effector function, we hypothesized that T cells from PC-treated recipients would have reduced capacity to mediate a host-versus-graft rejection response (HVGR) relative to FC-treated recipients. To directly test this hypothesis, we utilized a host T cell add-back model of rejection whereby BALB/c hosts were lethally irradiated (1050 cGy; day -2), reconstituted with host-type T cells from PC- or FC-treated recipients (day -1; 0.1 × 10^6 T cells transferred), and finally challenged with fully MHC-disparate transplantation (B6 donor bone marrow cells, 10 × 10^6 cells; day 0). In vivo HVGR was quantified by the following method at day 7 post-BMT: harvest of splenic T cells, stimulation with host- or donor-type dendritic cells, and use of six-color flow cytometry to detect host T cells, CD4 and CD8 subsets, and cytokine secretion by capture method. Consistent with our hypothesis, PC-treated cells acquired greatly reduced alloreactivity in vivo relative to FC-treated cells: the percentage of host CD4+ T cells secreting IFN-γ in an allospecific manner was 2.3±0.8% in recipients of PC-treated T cells and 62.7±13.4% in recipients of FC-treated cells (p<0.001). Similarly, the percentage of host CD8+ T cells secreting IFN-γ in an allospecific manner was 8.6±2.8% in recipients of PC-treated T cells and 92.7±4.1% in recipients of FC-treated T cells (p<0.001). We therefore conclude that at similar levels of myeloid suppression, the PC regimen is superior to the FC regimen in terms of murine T cell depletion, suppression of global T cell cytokine secretion, and inhibition of in vivo capacity to acquire allospecificity in response to fully genetically disparate marrow allografts. These data provide a rationale to develop PC regimens as an alternative to currently utilized FC regimens.

Low Incidence of Post-Transplant EBV-Related Disease After Alemtuzumab-Mediated T Cell Depletion Is Explained by the Differential Susceptibility to Alemtuzumab of B Cells and Protective CD8 and CD4 T Cells

Blood ◽  
2010 ◽  
Vol 116 (21) ◽  
pp. 2330-2330
Author(s):  
Constantijn J.M. Halkes ◽  
Inge Jedema ◽  
Judith Olde Wolbers ◽  
Esther M van Egmond ◽  
Peter A. Von Dem Borne ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
B Cells ◽  
B Cell ◽  
Cd8 T Cells ◽  
Cd4 T Cells ◽  
Cell Depletion ◽  
T Cell Depletion

Abstract Abstract 2330 In vivo T cell depletion with anti-thymocyte globulin (ATG) or alemtuzumab (anti-CD52) before reduced intensity allogeneic stem cell transplantation (alloSCT) in combination with in vitro T cell depletion with alemtuzumab reduces the risk of GVHD. Detectable levels of circulating antibodies are present up to several months after the alloSCT, leading to a delayed immune reconstitution which is associated with an increased incidence of opportunistic infections and early relapses. Prior to 2007, combined in vitro (Alemtuzumab 20 mg added “to the bag”) and in vivo T cell depletion with horse-derived ATG (h-ATG) resulted in good engraftment without GVHD in the absence of GVHD prophylaxis after reduced intensity alloSCT using conditioning with fludarabine and busulphan. Due to the unavailability of h-ATG, rabbit-derived ATG (r-ATG) 10–14 mg/kg was introduced in the conditioning regimen in 2007. Strikingly, in this cohort of patients, early EBV reactivation and EBV-associated post-transplantation lymphoproliferative disease (PTLD) was observed in 10 out of 18 patients at a median time of 6 weeks after alloSCT (range 5 to 11 weeks) in the absence of GVHD or immunosuppressive treatment. Analysis of T and B cell recovery early after transplantation revealed preferential depletion of T cells as compared to B cells, thereby allowing unrestricted proliferation of EBV infected B cells. Due to this unacceptable high incidence of EBV-related complications, in the conditioning regimen r-ATG was replaced by low dose alemtuzumab (15 mg i.v. day -4 and -3) in 2008. In this cohort of 60 patients, only 2 patients experienced transient EBV reactivation during the first 3 months after alloSCT and one patient developed an EBV-associated lymphoma 4 weeks after alloSCT. To investigate the mechanisms underlying the low incidence of EBV reactivation using alemtuzumab for T cell depletion, we studied the in vivo and in vitro effects of alemtuzumab on different lymphocyte subsets. First, lineage-specific reconstitution was studied in 20 patients from the alemtuzumab cohort with known CD52 negative diseases (11 AML and 9 multiple myeloma) to exclude the confounding effect of antibody absorption by malignant cells. Whereas at 3 weeks after alloSCT detectable numbers of circulating NK cells and T cells were observed (medians 71 (range 6–378), and 12 (range 1–1164)E6/L, respectively), no circulating B cells could be detected (median 0, range 0–1 E6/L). At 6 weeks after alloSCT, NK and T cell numbers further increased (medians 212 (52-813), and 130 (range 25–1509)E6/L, respectively), whereas B cell numbers still remained low in the majority of patients (median 15, range 0–813E6/L). In all patients, T cells were detectable before the appearance of circulating B cells. Furthermore, the expression of CD52 and the sensitivity to alemtuzumab-mediated complement-dependent cell lysis (CDC) of B cells, T cells and NK cells was measured in vitro. The highest CD52 expression was observed on B cells (mean fluorescence intensity (MFI) 120), resulting in 95% lysis after incubation with 10ug/mL alemtuzumab and rabbit complement. NK cells showed a significantly lower CD52 expression (MFI 41), which was also reflected by a lower susceptibility to alemtuzumab-mediated CDC (62% lysis). Interestingly, differential expression of CD52 was observed on CD4 and CD8 T cells (MFI 120 and 101, respectively). Cytotoxicity analysis revealed relative protection of CD8 compared to CD4 T cells against alemtuzumab-mediated CDC, resulting in 52% and 90% lysis, respectively. Based on these results, we investigated in detail the presence and phenotype of the CD4 and CD8 subsets and EBV-specific CD8 T cells using tetramer staining at 6 weeks after alloSCT. In accordance with the in-vitro expression and susceptibility data, circulating CD52+ CD8 T cells including EBV-specific T cells were detectable. Interestingly, the majority of circulating CD4 T cells (64-93%, n=4) lacked CD52 expression, explaining their capacity to persist in the presence of alemtuzumab. We conclude that in vivo and in vitro T cell depletion with alemtuzumab is associated with a relatively low risk of EBV-associated PTLD because of efficient B cell depletion and persistent EBV immunity allowed by the relative insusceptibility for alemtuzumab of CD8 T cells and the development of CD52 negative escape variants of CD4 T cells. Disclosures: No relevant conflicts of interest to declare.

In Vivo T Cell Depletion Using Rabbit Derived ATG Leads to An Increased EBV-PTLD Risk Due to An Induced Imbalance Between B and T Cell Recovery Which Is Not Seen After Horse Derived ATG or Alemtuzumab

Blood ◽  
2011 ◽  
Vol 118 (21) ◽  
pp. 1979-1979 ◽  
Author(s):  
C.J.M. Halkes ◽  
J.H.F. Falkenburg ◽  
H.M. van Egmond ◽  
J. Olde Wolbers ◽  
C.W.J. Starrenburg ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
B Cells ◽  
Cd8 T Cells ◽  
Cell Depletion ◽  
Cell Recovery ◽  
Post Transplantation ◽  
T Cell Depletion

Abstract Abstract 1979 Control of replication of endogenous viruses like CMV and EBV is fully dependent on CMV or EBV specific T cells after allogeneic stem cell transplantation (alloSCT). In the absence of specific CD8 T cell control, proliferation of EBV infected B cells can lead to post transplantation lymphoproliferative disease (PTLD). In an initial cohort of patients treated with horse derived anti thymocyte globulin (h-ATG), no early PTLD was observed. However, due to unavailability in Europe, h-ATG had to be replaced by rabbit derived ATG (r-ATG), leading to an unacceptable high incidence of EBV-PTLD (26% during first 3 months after alloSCT). Replacement of r-ATG by alemtuzumab (ALT) significantly reduced the incidence of EBV-PTLD (3 months incidence of EBV-PTLD 2%). To determine the immunological basis of these findings we performed a detailed analysis of immune reconstitution in these three cohorts of transplanted patients. The first cohort (41 patients) received h-ATG (Lymphoglobulin) 10 mg/kg/day for 4 days. The second cohort (19 patients) received r-ATG (Thymoglobulin) 2.0 or 3.5 mg/kg/day for 4 days and the third cohort (60 patients) received ALT, 15 mg/day for 2 days. All grafts consisted of PBSC to which 20 mg of ALT was added for in vitro T cell depletion. All patients received a fludarabin and busulphan based conditioning regimen. No standard post transplantation immunosuppressive treatment was given. In the r-ATG cohort, early EBV-PTLD occurred after a median of 7 weeks (range 4–12 weeks) post alloSCT. Three r-ATG treated patients died while high levels of circulating EBV-DNA were present (> log 4.0 copies/mL). Incidence of CMV disease was not significantly different in the three cohorts (5%, 6% and 0%, respectively). In contrast to the other 2 cohorts, immune reconstitution in the r-ATG cohort was characterized by an imbalance between recovery of B cells and CD8 T cells. Already 3 weeks after alloSCT, the majority (67%) of r-ATG patients showed a more rapid reconstitution of B cells than CD8 T cells, leading to B cells outnumbering CD8 T cells. This was seen in only a small minority of patients after h-ATG and ALT (17% and 6%, respectively, p<0.01 versus r-ATG). Because rapid recovery of T cells in the alemtuzumab patients was frequently found in the presence of circulating ALT (mean concentration 0.43 μg/mL and 0.12 μg/mL after 3 and 6 weeks, respectively), the phenotype of circulating CD4 and CD8 T cells at 6 weeks after ALT was analyzed. The majority of circulating CD8 and CD4 T cells lacked CD52 expression (56% (range 0–99%) and 81% (range 0–93%), respectively). Using tetramer staining, cytotoxicity assays and analysis of cytokine production, we demonstrated the presence of functional CD52 negative as well as CD52 positive CMV and EBV specific CD8 T cells. Based on FLAER negativity, it was demonstrated that the CD52 negative T cells are GPI anchor deficient, representing a PNH-like clone escaping ALT induced cell lysis. Because almost half of the circulating CD8 T cells were CD52 positive, we examined expression of CD52 and the in-vitro sensitivity to ALT-mediated complement-dependent cell lysis (CDC) of B cells, CD4 and CD8 T cells of healthy donors. The highest CD52 expression was observed on B cells (mean fluorescence intensity (MFI) 120), resulting in 95% lysis after incubation with ALT and complement. Differential expression of CD52 was observed on CD4 and CD8 T cells, MFI 120 and 101 respectively, resulting in relative protection of CD52 positive CD8 compared to CD4 T cells against ALT-mediated CDC (52% and 90% lysis). We conclude that the high incidence of EBV-PTLD after in-vivo T cell depletion with r-ATG is caused by an induced imbalance between B and T cell recovery, which is not seen after h-ATG or ALT. In-vivo T cell depletion with ALT is associated with a relatively low risk of EBV disease because of efficient B cell depletion and persistent EBV immunity due to the relative insusceptibility for ALT of CD8 T cells and the development of functional CD52 negative escape variants of CD4 and CD8 T cells. Disclosures: Off Label Use: Alemtuzumab and Anti Thymocyte Globulin used for in vivo T cell depletion prior to allogeneic stem cell transplantation.

scholarly journals Gamma Interferon-Induced T-Cell Loss in Virulent Mycobacterium avium Infection

2005 ◽  
Vol 73 (6) ◽  
pp. 3577-3586 ◽  
Author(s):  
Manuela Flórido ◽  
John E. Pearl ◽  
Alejandra Solache ◽  
Margarida Borges ◽  
Laura Haynes ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Mycobacterium Avium ◽  
Cell Loss ◽  
Gamma Interferon ◽  
Cell Depletion ◽  
Mycobacterial Antigen ◽  
T Cell Depletion ◽  
Avium Infection ◽  
Ifn Γ

ABSTRACT Infection by virulent Mycobacterium avium caused progressive severe lymphopenia in C57BL/6 mice due to increased apoptosis rates. T-cell depletion did not occur in gamma interferon (IFN-γ)-deficient mice which showed increased T-cell numbers and proliferation; in contrast, deficiency in nitric oxide synthase 2 did not prevent T-cell loss. Although T-cell loss was IFN-γ dependent, expression of the IFN-γ receptor on T cells was not required for depletion. Similarly, while T-cell loss was optimal if the T cells expressed IFN-γ, CD8+ T-cell depletion could occur in the absence of T-cell-derived IFN-γ. Depletion did not require that the T cells be specific for mycobacterial antigen and was not affected by deficiencies in the tumor necrosis factor receptors p55 or p75, the Fas receptor (CD95), or the respiratory burst enzymes or by forced expression of bcl-2 in hematopoietic cells.

scholarly journals CD4 T Cell Depletion Substantially Augments the Rescue Potential of PD-L1 Blockade for Deeply Exhausted CD8 T Cells

2015 ◽  
Vol 195 (3) ◽  
pp. 1054-1063 ◽  
Author(s):  
Pablo Penaloza-MacMaster ◽  
Nicholas M. Provine ◽  
Eryn Blass ◽  
Dan H. Barouch
Keyword(s):  
T Cells ◽  
T Cell ◽  
Cd8 T Cells ◽  
Cd4 T Cell ◽  
Cell Depletion ◽  
T Cell Depletion

scholarly journals IL-1 enhances expansion, effector function, tissue localization, and memory response of antigen-specific CD8 T cells

2013 ◽  
Vol 210 (3) ◽  
pp. 491-502 ◽  
Author(s):  
Shlomo Z. Ben-Sasson ◽  
Alison Hogg ◽  
Jane Hu-Li ◽  
Paul Wingfield ◽  
Xi Chen ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Cd8 T Cells ◽  
Granzyme B ◽  
Interleukin 1 ◽  
Cd8 T Cell ◽  
Cell Receptor ◽  
In Vivo Models ◽  
Ifn Γ

Here, we show that interleukin-1 (IL-1) enhances antigen-driven CD8 T cell responses. When administered to recipients of OT-I T cell receptor transgenic CD8 T cells specific for an ovalbumin (OVA) peptide, IL-1 results in an increase in the numbers of wild-type but not IL1R1−/− OT-I cells, particularly in spleen, liver, and lung, upon immunization with OVA and lipopolysaccharide. IL-1 administration also results in an enhancement in the frequency of antigen-specific cells that are granzyme B+, have cytotoxic activity, and/ or produce interferon γ (IFN-γ). Cells primed in the presence of IL-1 display enhanced expression of granzyme B and increased capacity to produce IFN-γ when rechallenged 2 mo after priming. In three in vivo models, IL-1 enhances the protective value of weak immunogens. Thus, IL-1 has a marked enhancing effect on antigen-specific CD8 T cell expansion, differentiation, migration to the periphery, and memory.

Reconstitution of CD52-Negative CD4 T Cells after Alemtuzumab-Based T Cell Depleted Allogeneic Hematopoietic Stem Cell Transplantation

Blood ◽  
2008 ◽  
Vol 112 (11) ◽  
pp. 1182-1182
Author(s):  
Eva M Wagner ◽  
Aline N Lay ◽  
Sina Wenzel ◽  
Timo Schmitt ◽  
Julia Hemmerling ◽  
...  
Keyword(s):  
Flow Cytometry ◽  
T Cells ◽  
T Cell ◽  
Cd4 T Cells ◽  
Lymphocytic Leukemia ◽  
Cell Depletion ◽  
Down Regulation ◽  
T Cell Depletion ◽  
Hematopoietic Stem

Abstract The human CD52 molecule is the target of the monoclonal antibody Alemtuzumab, which is used for treating patients with chemo-refractory chronic lymphocytic leukemia as well as for T cell depletion (TCD) in the context of allogeneic hematopoietic stem cell transplantation (HSCT). The molecule is expressed on the surface of lymphocytes, dendritic cells and to a lesser extent on blood-derived monocytes. Previously, investigators have demonstrated that the surface expression of CD52 on T cells is down-regulated after in vitro incubation with Alemtuzumab. By treating purified human CD4 T cells over 4 hours with 10 μg/mL Alemtuzumab in medium supplemented with 10% human AB serum in vitro, we observed a strong decrease of CD52 expression by flow cytometry with a maximum 3–7 days after incubation. The CD52 down-regulation was also found at weaker intensity on CD8 T cells. From previous studies in chronic lymphocytic leukemia patients, it is known that Alemtuzumab treatment also leads to a down-regulation of CD52 on T cells in vivo. However, similar experiments have not been performed in allogeneic HSCT patients receiving Alemtuzumab in vivo for T cell depletion. We therefore analyzed the expression of CD52 on human peripheral blood mononuclear cells isolated at repeated time points from 22 allogeneic HSCT patients after reduced-intensity conditioning with fludarabine and melphalan and in vivo T cell depletion with Alemtuzumab (100 mg). Half of the patients received prophylactic CD8-depleted donor lymphocyte infusions (DLI) to promote immune reconstitution. By flow cytometry, we observed that the CD52 expression on monocytes, B cells, and natural killer cells remained unaltered after transplantation and was not influenced by the application of DLI. In contrast, the majority of CD4 T cells were CD52-negative (median, 72%) after transplantation and they remained CD52-negative in patients who did not receive DLI throughout the first year after HSCT. The permanent lack of CD52 expression could not be explained by a continuous effect of Alemtuzumab, because earlier studies have shown that the antibody is not present in active plasma concentrations beyond day +60 after HSCT. In contrast, patients receiving CD8-depleted DLI demonstrated a significant increase in the proportion of CD52-positive CD4 T cells. In three of our patients (DLI: n=2, non-DLI: n=1) we analyzed the donor chimerism of CD52-positive and CD52-negative CD4 T cells sorted with high purity by flow cytometry. Three months after HSCT (before DLI), the proportion of donor T cells was clearly higher among the CD52-negative compared to the small proportion of CD52-positive cells in all patients (44% vs. 10%, 83% vs. 0%, and 100% vs. 40%). In the patient who did not receive DLI, the donor T cell chimerism remained mixed in the CD52-negative and CD52-positive fractions on days 200 (CD52-negative: 95%; CD52-positive: 15%) and 350 (CD52-negative: 92%; CD52-positive: 65%). In contrast, the two patients receiving CD8-depleted DLI showed a strong increase in the proportion of CD52-positive CD4 T cells that were of complete donor origin. Altogether, CD52 is permanently down-regulated in reconstituting CD4 T cells following HSCT with an Alemtuzumab-based TCD regimen unless DLI are applied. Our data support the idea of an active mechanism for CD52 down-regulation in CD4 T cells that is not related to B cells and natural killer cells and that appears to differently affect donor and host T cells, respectively.

A Comparative Analysis of Immune Reconstitution Following Reduced Intensity Conditioning with CAMPATH-1H and Total Lymphoid Irradiation/Anti-Thymocyte Globulin Prior to Allogeneic Stem Cell Transplantation.

Blood ◽  
2009 ◽  
Vol 114 (22) ◽  
pp. 1148-1148
Author(s):  
Brett Glotzbecker ◽  
Heidi Mills ◽  
Jacalyn Rosenblatt ◽  
Zekui Wu ◽  
Kerry Wellenstein ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Cd8 T Cells ◽  
Cell Depletion ◽  
T Cell Proliferation ◽  
Reduced Intensity Conditioning ◽  
T Cell Depletion ◽  
Post Transplant ◽  
Initial Cohort ◽  
Reduced Intensity

Abstract Abstract 1148 Poster Board I-170 Graft versus host disease (GVHD) remains a significant cause of morbidity and mortality after allogeneic hematopoietic stem cell transplantation (HCT). In vivo quantitative T-cell depletion using CAMPATH-1h (anti-CD52) has been explored in an effort to prevent acute GVHD. More recently, a regimen consisting of total lymphoid irradiation and anti-thymocyte globulin (ATG) has been shown to polarize T cells towards an inhibitory phenotype potentially reducing the associated risk for GVHD. However, these strategies may be associated with impaired post-transplant immune reconstitution, increased risk of tumor relapse and opportunistic infection. In this study we examined the pattern of cellular immune recovery following T cell depletion with CAMPATH-1h and compared results with an initial cohort of patients undergoing reduced intensity conditioning with TLI and ATG. Immunologic analyses were performed on twenty patients undergoing reduced intensity conditioning in conjunction with low dose CAMPATH -1h (50 mg) and an initial cohort of 5 patients treated with TLI/ATG. Conditioning with CAMPATH-1h resulted in the significant depletion of CD3, CD4, and CD8 T cells in the early post-transplant period and persistence of CD4 T cell depletion (< 200 cells /uL) for more than 6 months. Following TLI/ATG, persistent depletion of CD4+ T cells was also observed but no significant decrease in CD8 T cells was seen. A two-fold increase in circulating CD56+ NK cells, 21.8 to 41.6% (p=0.004), was seen following TLI-ATG, which was not noted following Campath conditioning. CAMPATH-1h conditioning was associated with a significant decrease in mean CD45RO+ memory T cells in the early post-transplant period (27.2 to 5.7% of the total population of nonadherent peripheral blood mononuclear cells, p=0.034). Relative percentages of naïve T cells (CD45RA+), central memory (CD45RO+CD62L+CCR7+) (CM), and effector memory (CD45RO+CD62L-CCR7-) (EM) T cells remained stable in the pre- and post-transplantation period. The CM:EM was 0.6 pre-transplant and at day 60, respectively. In contrast, T cell recovery in early post-transplant following the TLI/ATG regimen was associated with no reduction in CD45RO+ memory T cells. A significant rise in the relative percentages of naïve T cells from 39 to 61.3% (p=0.04), CM cells from 12 to 32.8% (p=0.05), a corresponding fall in EM cells from 57.9 to 32.5% (p=0.10), and a significant change in the CM:EM levels (0.2 pre-transplant, 1.0 day 60 post-transplant) was noted after TLI/ATG. The mean percentage of regulatory T cells as defined by the percentage of CD4+/CD25+ cells that express FoxP3 rose in the early post-transplant period following both regimens (8 to 20.7% at Day 30, p=0.003 in the CAMPATH group; 5.6 to 16.9% at Day 30, p=0.03 in the ATG/TLI group). Functional analyses demonstrated that the T cell proliferative response to the mitogen, Phytohemagglutinin (PHA), was profoundly depressed following CAMPATH-1h with mean SI decreasing from 34 pre-transplant to 1.4 at Day 30. In contrast, treatment with TLI/ATG resulted in no significant change in T cell proliferation in response to PHA with SI only decreasing from 45 pre-transplant to 36 at Day 30. Assessment of T cell polarization following stimulation with PHA or phorbol-ester (PMA)/ionomycin, recipient derived dendritic cells (DCs) or third party DCs demonstrated a rise of CD8+ T cells expressing, IL-4 and IL-10 consistent with a suppressor phenotype. Minimal T cell proliferation was observed following stimulation with patient derived DCs, which is consistent with suppression of the expansion of alloreactive T cells. In summary, both CAMPATH and TLI/ATG result in CD4+ T cell depletion but TLI/ATG resulted in relative preservation of CD8+ T cells, persistence of memory cells, relative preservation of central memory as compared to memory effector cells and intact response to mitogens. TLI/ATG therapy was also associated with the polarization of CD8+ T cells towards a Tc2 phenotype and lack of proliferation in response to recipient derived DCs. As such, TLI/ATG appears to be associated with more modest level of functional T cell depletion characterized by Tc2 polarization and suppression of host/donor alloreactivity. Disclosures Spitzer: Genzyme: Consultancy. Avigan:Genzyme: Consultancy.

scholarly journals Effects of in Vivo Cd8+ T Cell Depletion on Virus Replication in Rhesus Macaques Immunized with a Live, Attenuated Simian Immunodeficiency Virus Vaccine

1999 ◽  
Vol 191 (11) ◽  
pp. 1921-1932 ◽  
Author(s):  
Karin J. Metzner ◽  
Xia Jin ◽  
Fred V. Lee ◽  
Agegnehu Gettie ◽  
Daniel E. Bauer ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Simian Immunodeficiency Virus ◽  
Cd8 T Cells ◽  
Rhesus Macaques ◽  
Cd8 T Cell ◽  
T Cell Depletion ◽  
Challenge Virus ◽  
Immunodeficiency Virus

The role of CD8+ T lymphocytes in controlling replication of live, attenuated simian immunodeficiency virus (SIV) was investigated as part of a vaccine study to examine the correlates of protection in the SIV/rhesus macaque model. Rhesus macaques immunized for &gt;2 yr with nef-deleted SIV (SIVmac239Δnef) and protected from challenge with pathogenic SIVmac251 were treated with anti-CD8 antibody (OKT8F) to deplete CD8+ T cells in vivo. The effects of CD8 depletion on viral load were measured using a novel quantitative assay based on real-time polymerase chain reaction using molecular beacons. This assay allows simultaneous detection of both the vaccine strain and the challenge virus in the same sample, enabling direct quantification of changes in each viral population. Our results show that CD8+ T cells were depleted within 1 h after administration of OKT8F, and were reduced by as much as 99% in the peripheral blood. CD8+ T cell depletion was associated with a 1–2 log increase in SIVmac239Δnef plasma viremia. Control of SIVmac239Δnef replication was temporally associated with the recovery of CD8+ T cells between days 8 and 10. The challenge virus, SIVmac251, was not detectable in either the plasma or lymph nodes after depletion of CD8+ T cells. Overall, our results indicate that CD8+ T cells play an important role in controlling replication of live, attenuated SIV in vivo.

scholarly journals Intestinal Intraepithelial Lymphocytes Are Primed for Gamma Interferon and MIP-1β Expression and Display Antiviral Cytotoxic Activity despite Severe CD4+ T-Cell Depletion in Primary Simian Immunodeficiency Virus Infection

1998 ◽  
Vol 72 (8) ◽  
pp. 6421-6429 ◽  
Author(s):  
Joseph J. Mattapallil ◽  
Zeljka Smit-McBride ◽  
Michael McChesney ◽  
Satya Dandekar
Keyword(s):  
T Cells ◽  
T Cell ◽  
Simian Immunodeficiency Virus ◽  
Intraepithelial Lymphocytes ◽  
Cell Depletion ◽  
T Cell Depletion ◽  
Immunodeficiency Virus ◽  
Siv Infection ◽  
Ifn Γ ◽  
Ctl Activity

ABSTRACT Intraepithelial lymphocytes (IEL) are a critical effector component of the gut-associated lymphoid tissue (GALT) and play an important role in mucosal immunity as well as in the maintenance of the epithelial cell integrity and barrier function. The objective of this study was to determine whether simian immunodeficiency virus (SIV) infection of rhesus macaques would cause alterations in the immunophenotypic profiles of IEL and their mitogen-specific cytokine (gamma interferon [IFN-γ] and MIP-1β) responses (by flow cytometry) and virus-specific cytotoxic T-cell (CTL) activity (by the chromium release assay). Virally infected IEL were detected through the entire course of SIV infection by in situ hybridization. Severe depletion of CD4+ single-positive and CD4+CD8+ double-positive T cells occurred early in primary SIV infection, which was coincident with an increased prevalence of CD8+ T cells. This was in contrast to a gradual depletion of CD4+ T cells in peripheral blood. The CD8+ IEL were the primary producers of IFN-γ and MIP-1β and were found to retain their potential to produce both IFN-γ and MIP-1β through the entire course of SIV infection. SIV-specific CTL activity was detected in primary IEL at 1, 2, and 4 weeks post-SIV infection. These results demonstrated that IEL may be involved in generating antiviral immune responses early in SIV infection and in suppressing viral infection thereafter. Alterations in homeostasis in epithelia due to severe CD4+ T-cell depletion accompanied by changes in the cytokine and chemokine production by IEL may play a role in the enteropathogenesis of SIV infection.

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