Toll-Like Receptor 2 and 4 Are Expressed on Transplanted T-Cell Subsets and Influence Outcome after Allogeneic Unrelated Peripheral Blood Stem Cell Transplantation.

Blood ◽  
2007 ◽  
Vol 110 (11) ◽  
pp. 1066-1066
Author(s):  
Uwe Platzbecker ◽  
Jan Stoehlmacher ◽  
Eray Goekkurt ◽  
Caroline Pabst ◽  
Christian Thiede ◽  
...  
Keyword(s):  
T Cells ◽  
Stem Cell ◽  
T Cell ◽  
Stem Cell Transplantation ◽  
Cd8 T Cells ◽  
Peripheral Blood ◽  
Peripheral Blood Stem Cell ◽  
Toll Like Receptor ◽  
Clinical Endpoints ◽  
Blood Stem Cell Transplantation

Abstract Introduction: Recently, Toll-like receptor (TLR) 2 and 4 have been identified as the most important receptors for LPS, which is contained in the cell wall of gram-negative bacteria and is known to be a main inducer of graft versus host disease (GVHD). The role of TLR expressing T-cells within the graft for the induction of GVHD in patients after unrelated peripheral blood stem cell transplantation (PBSCT) is unknown. Methods and patients: We therefore determined by flow cytometry expression of TLR 2 and 4 on T-cells within the graft of 63 patients receiving unrelated PBSCT after intensive conditioning followed by cyclosporine A and methotrexate as GVHD prophylaxis. Additionally, donor specific single nucleotide polymorphisms (SNP) for TLR2 (R753Q), TLR4 (D299G) and TLR4 (Y135A) were determined. The data were finally correlated with clinical endpoints. Results: As expected, TLRs were not expressed on T-cells in peripheral blood of healthy donors (TLR 2: <1.0%, TLR4 <0.5% of T-cells, n=6). In contrast we detected a distinct up-regulation of these receptors on T-cells within the grafts. TLR2 and TLR4 expression on CD4+ T-cells ranged from 1.2%–12.3% (median 3.1%) for TLR2 and from 1.2%–12.0% (median 3.7%) for TLR4. Among the CD8+ T-cells 0.9%–16.1% (median 3.3%) expressed TLR2 and 0.7%–13.3% (median 3.5%) expressed TLR4. The SNP for TLR2 (R753Q) and TLR4 (D299G) was found in 8.6% and 10.3% of the allogeneic donors, respectively but did neither correlate with the expression levels of TLR on T-cells nor with clinical endpoints. Treatment-related mortality from infections was observed in 10 patients (16%). Interestingly, higher expression of TLR2 and 4 on CD4+ but not CD8+ T-cells was significantly associated with an increased cumulative incidence of fatal infections (38% vs. 8% p=0.03 for TLR2 and 37% vs. 9% p=0.007 for TLR4). Neither the overall CD3+, CD4+ and CD8+ cell dose nor the expression of TLR2 and 4 on CD4+ and CD8+ T-cells showed a significant association with the incidence of acute or chronic GVHD or relapse. Conclusion: These data suggest a previously unrecognised up-regulation of TLR, on CD4+ and CD8+ T-cells contained in G-CSF mobilized apheresis products. Whether these phenotypic changes impact on T cell function or patient outcome warrants further investigation.

Intensive Immunosuppression Plus T-Cell Depleted Autologous Peripheral Blood Stem Cell Transplantation for Systemic Sclerosis.

Blood ◽  
2004 ◽  
Vol 104 (11) ◽  
pp. 5205-5205
Author(s):  
Diane BuchBarker ◽  
Albert D. Donnenberg ◽  
Thomas A. Medsger ◽  
Michael P. Carroll ◽  
Deborah L. Griffin ◽  
...  
Keyword(s):  
T Cells ◽  
Stem Cell ◽  
Systemic Sclerosis ◽  
T Cell ◽  
Stem Cell Transplantation ◽  
Cell Transplantation ◽  
Peripheral Blood ◽  
Peripheral Blood Stem Cell ◽  
The Mean ◽  
Blood Stem Cell Transplantation

Abstract Autologous peripheral blood stem cell transplantation (PBSCT) may be therapeutic in patients (pts) with severe autoimmune disorders. A pre-PBSCT regimen that optimizes immunosuppression without excessive myelotoxicity would potentially have favorable safety and efficacy profiles in these pts. To test this hypothesis we are conducting a phase I trial of cyclophosphamide (CY) dose escalation (initial CY dose 200 mg/m2/day x 5, days −7 to −3) plus fludarabine (25 mg/m2/day x 5, days −7 to −3) and rabbit anti-thymocyte globulin (ATG; 2.5 mg/kg/day x 3, days −5 to −3) followed by T-cell depleted (TCD) autologous PBSCT in adults with systemic sclerosis (SSc). To date, 5 pts (2 male, 3 female; median age, 44 yr; range, 39–59 yr) have undergone TCD PBSCT. Each pt had satisfactory PBSC collection with one 4-hr large-volume leukapheresis (LVL) after mobilization with CY (2.0 g/m2) plus G-CSF. Mean yields of CD34+ and CD3+ cells by LVL were 18.8 (range, 10.5–33.0) x 106/kg and 1.23 (range, 0.37–2.45) x 108/kg, respectively. To deplete T cells in the PBSC product we used a combination of CD34+ selection (Isolex 300i v 2.5; Baxter Oncology) and ex vivo incubation with anti-CD3 antibody (OKT3; Ortho Biotech), which resulted in a mean 5.2-log reduction of T cells (range, 4.7–6.3 log). The final PBSC products contained a mean of 9.94 (range, 4.8–13.0) x 106 CD34+ cells/kg and 1.34 (range, 0.06–4.54) x 103 CD3+ cells/kg. The daily dose of CY was 200 mg/m2 in the first 3 pts and 400 mg/m2 in the next 2 pts. Using rare-event flow cytometry, we found that the mean half-life (t 1/2) of CD3+ cells in these pts was biphasic; t 1/2 was 0.8 days from day −7 to −5 and 0.2 days from −5 to −3 days, correlating with ATG administration. In contrast, the mean t1/2 of CD3+ cells was 2.5 days in pts with hematologic malignancies receiving a myeloablative preparative regimen of CY, busulfan and etoposide. After PBSCT, the median nadir of WBC, neutrophil and platelet levels were 1.2 (range, 0.4–2.0), 0.9 (range, 0.1–1.7) and 135 (range, 114–185) x 109/L, respectively. One pt at the second CY dose level developed pericardial effusion and tamponade at day +5 and required surgical intervention. No pts developed opportunistic infections. All pts are alive at a median of 6.5+ months (range, 4.9+–15.5+ months) after PBSCT. Compared with pre-PBSCT levels, Rodnan total skin score (TSS) decreased by 10, 11 and 15 points, respectively, in 3 pts and increased by 5 and 8 points in 2 pts. One pt with worsening TSS at 12 months after PBSCT developed SSc renal crisis and requires hemodialysis. Of 3 pts with improved TSS, 1 has worsening polymyositis and 1 has recurrence of palpable tendon friction rubs. Even at the lowest CY dose, this immunosuppressive regimen provides significantly greater and more rapid T-cell kill than a conventional myeloablative regimen. The efficacy of this regimen and TCD-PBSCT in SSc is encouraging but requires longer followup and experience with a larger number of pts.

Impact of gamma/delta TCR+, Foxp3+ and Toll-Like Receptor Expressing T-Cells on the Outcome of Patients Receiving Unrelated Peripheral Blood Stem Cell Transplantation.

Blood ◽  
2006 ◽  
Vol 108 (11) ◽  
pp. 2899-2899
Author(s):  
Uwe Platzbecker ◽  
Caroline Pabst ◽  
Holger Schirutschke ◽  
Ines Wagner ◽  
Cathrin Theuser ◽  
...  
Keyword(s):  
T Cells ◽  
Stem Cell ◽  
Stem Cell Transplantation ◽  
Peripheral Blood Stem Cell ◽  
T Cell Subsets ◽  
Toll Like Receptor ◽  
Gamma Delta ◽  
Cell Dose ◽  
Increased Risk ◽  
Blood Stem Cell Transplantation

Abstract Introduction: The role of gamma/delta (GD), foxp3 (regulatory, TREGs) and Toll-like receptor (TLR) expressing T-cells contained in the graft for the induction of graft versus host disease (GVHD) in patients after unrelated peripheral blood stem cell transplantation (PBSCT) is unknown. Methods and patients: We therefore determined by flowcytometry and PCR the content of these T-cell subsets within the graft of 63 patients with hematological malignancies receiving unrelated PBSCT followed by cyclosporine A and methotrexate as GVHD prophylaxis and correlated these data with clinical endpoints. Patient and donor pairs were matched at least in nine out of ten characteristics (n=17), whereas a complete match (10 out of 10) was observed in 46 (73%) patients. Results: The grafts contained a median of 11.25 x 106/kg (2.7–33.5) CD4+foxp3 T-cells and 9.79 x 106/kg (1.4–36.2) GD T-cells. Patients receiving lower amounts of CD4+foxp3+ cells had an increased risk of acute GVHD II–IV (65% vs. 44%, p=0.03). Whereas the overall CD3+ cell dose was not associated with the incidence of GVHD, increased counts of gdTCR expressing CD3+ cells showed significant correlation with an increased risk of GVHD II–IV (66% vs. 40%, p=0.028). A lower CD34+ cell dose was also significantly associated with an increased risk of GVHD II–IV (63% vs. 38%, p=0.04). Interestingly, there was a positive correlation between CD34+ and CD4+foxp3 cell dose (r=0.34, p=0.007). In contrast to the blood of healthy donors, TLR2 and 4 were both expressed on up to 11 % of CD4+ and CD8+ cells within the graft but had no influence on the severity of GVHD. Interestingly, a higher expression of TLR2 and 4 on CD4+ but not CD8+ T-cells was associated with a higher treatment-related mortality (CD4: 17% for TLR2low vs. 45% for TLR2high, 13% for TLR4low vs. 52% for TLR4high, p=0.002). Conclusion: Strategies aiming at selective manipulation of graft composition or add-back of distinct T-cell subsets after unrelated PBSCT might be a promising strategy in order to modulate the extent of clinical GVHD. Furthermore, up-regulation of TLR2 and 4 on T-cells and their role for non-relapse mortality warrants further investigation.

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