Partial T-Cell Depletion By Low-Dose Anti-Thymocyte Globulin to Reduce Severe Acute and Chronic Graft-Versus-Host Disease after Allogeneic Stem Cell Transplantation

Abstract Background: T cells from a stem cell source are inevitably contaminated, and over 5.0×104/kg T cells are thought to induce graft-versus-host disease (GVHD) in HLA-mismatched or haplo-identical stem cell transplantations (SCTs) [4]. To suppress GVHD reactions, a procedure for T-cell depletion (TCD) was developed over the past several decades, especially for HLA-mismatched and haplo-identical SCTs, which are at high risk for GVHD. To reduce the incidence of GVHD, a potentially effective agent is anti-thymocyte globulin (ATG), which is generally administered at a dose of ≥ 5-10 mg/kg. Based on data regarding the use of ATG for the treatment of aplastic anemia, we hypothesized that ATG might accommodate engraftment and inhibit GVHD. We attempted to use a lower dose of ATG to decrease non-relapse mortality (NRM) in Japanese patients undergoing an HLA-matched SCT. Patients and method: We treated patients with hematological diseases who underwent an allogeneic SCT after March 2010 without or with 2.5 mg/kg ATG. The inclusion criteria for underlying disease included both hematological malignancies and bone marrow failures. All consecutive patients transplanted from an allogeneic related or unrelated donor were included. Cord blood transplantations were omitted from this analysis. The patients who underwent an SCT before February 2010 (n=20) were examined as the control group without ATG treatment. ATG was administered 1 day prior to the transplantation day at 2.5 mg/kg with 500 mg/body methylpredonisolone as a preconditioning procedure. GVHD prophylaxis, tacrolimus 0.03 mg/kg and short-term MTX (10-7-7 mg/m2) was adapted for both the ATG group and the control group. Results: Thirty-nine (21 male, 18 female) recipients were recruited (median age 49 yrs, range 19-64 yrs). Their underlying diseases were acute myeloid leukemia (n=14), acute lymphoblastic leukemia (n=10), myelodysplastic syndrome (n=5), lymphoma (n=7), and myeloma, aplastic anemia, and other malignancy (n=1 each). Preparation regimens were myeloablative for 17 patients (14 cyclophosphamide [CY]/total body irradiation [TBI], two busulfan [BU]/CY, and another) and non-myeloablative for the other 22 patients (14 fludarabine/melphalan [Flu/Mel] and eight Flu/BU). All but one patient achieved engraftment, and one secondary graft failure was observed. The overall incidences of acute and chronic GVHD were 63.2% and 15.8% for the ATG-treated patients (40.0% and 25.0% for the control cohort), respectively. Acute GVHD (grades II to IV and III to IV) in the recipients who received ATG occurred in 21.1% and 0.0% (control cohort, 10.0% and 5.0%), respectively. The estimated probability of overall survival (OS) 2.5 yrs after transplantation was 77.8% for the ATG group (controls, 57.1%). The relapse rate 2.5 yrs after transplantation was 21.1% and 20.0% in the ATG and control groups, respectively. The NRM rate was decreased after ATG treatment: 25.0% vs. 10.5% (not significant). The causes of mortality with or without ATG were recurrent diseases (n=1 and 2), infection (n=1 and 0), and adverse events caused by transplant-related complication (n=1 and 5), respectively. No deaths due to acute or chronic GVHD occurred. Discussion: Low-dose ATG could suppress the incidence of severe acute GVHD and chronic GVHD without increasing the NRM, although our study design did not have enough power to make a conclusion about the efficacy of low-dose ATG. However, partial T-cell depletion may be effective for HLA-matched SCT recipients. Our results show that ATG at 2.5 mg/kg can be used safely for the Japanese transplant population of HLA-matched donors. Low-dose ATG is a potential treatment to partially disempower T cells from a stem cell source, which are inevitably contaminated. Recent developments in the prophylaxis for GVHD, such as selective cytotoxic T-cell depletion by using a post-transplant CY regimen, are promising strategies to fully suppress T cells as the GVHD enhancer. Previous studies revealed the clinical efficacy of GVHD prophylaxis but did not clarify the significance of its survival benefit. Likewise, our present findings indicated a lack of survival benefit by ATG treatment in this small study. However, the low-dose ATG contributed to a reduction of severe GVHD. Although early mortality after transplantation is decreasing, late-onset comorbidity including chronic GVHD remains a significant problem. Disclosures No relevant conflicts of interest to declare.

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Transplantation Of TcRαβ/CD19 Depleted Stem Cells From Haploidentical Donors In Children: Current Results

Blood ◽

10.1182/blood.v122.21.692.692 ◽

2013 ◽

Vol 122 (21) ◽

pp. 692-692 ◽

Cited By ~ 1

Author(s):

Peter Lang ◽

Tobias Feuchtinger ◽

Heiko-Manuel Teltschik ◽

Michael Schumm ◽

Patrick Schlegel ◽

...

Keyword(s):

Stem Cells ◽

T Cells ◽

Stem Cell ◽

T Cell ◽

T Lymphocytes ◽

Nk Cells ◽

Chronic Gvhd ◽

Cell Depletion ◽

T Cell Depletion ◽

Αβ T Cells

Abstract T-cell depletion of the graft is an effective method to prevent or completely avoid Graft-versus-Host Disease (GvHD) in haploidentical stem cell transplantation. In order to increase the T-cell depletion efficacy while maintaining the anti-tumor and anti-infectious properties of the graft, we have investigated a new T-cell depletion method which removes αβ+ T-lymphocytes via a biotinylated anti-TcRαβ antibody followed by an anti-biotin antibody conjugated to magnetic microbeads while retaining γδ+ T-lymphocytes, Natural killer (NK) cells and other cells in the graft. In addition, CD19+ B-lymphocytes were concomitantly depleted for the prevention of posttransplant EBV-associated lymphoproliferative disease. The CliniMACS system was used for manipulation of peripheral stem cell grafts from full haplotype mismatched family donors in 35 patients. Results The overall depletion of αβ+ T-cells was highly effective with 4.6 log (range 3.8–5.0). Patients received a median number of only 14 x 103/kg residual αβ+ T-cells. Recovery of CD34+ stem cells was 72%, and the median number of infused CD34+ stem cells was 12 x 106/kg (range 5-38 x 106/kg). Additionally, the patients received 2 types of potential antileukemic effector cells: 107 x 106/kg (range 35 -192 x 106/kg) CD56+ NK-cells and 11 x 106/kg (range 5–30 x 106/kg) γδ+ T-lymphocytes. Diagnoses were ALL (n=20), AML/MDS/JMML (n=9), nonmalignant diseases (n=4), solid tumors (n=2); disease status: CR2-CR6 (n=17), active disease (n=18). 23 patients received a second or third SCT (65%). A toxicity reduced conditioning regimen (fludarabin 40mg/m² or clofarabin 50mg/m² (day -8 to d -5), thiotepa 10mg/kg (d -4), melphalan 70mg/m² (d -3 and d -2) was used. The anti CD3 specific OKT3 antibody was used as rejection prophylaxis from day -8 to day -1 without affecting cotransfused effector cells because of its short half-life period in the first 7 patients. However, due to its restricted availability, the substance was substituted since 2011 by a reduced ATG-F dose (15mg/kg) given at start of the conditioning regimen in order not to impair NK and γδ+ T-cells of the grafts (1 mg/kg d -12, 4 mg/kg d -11, 5 mg/kg d -10 and -9; n=28 patients). Short course MMF (until day +30) was given in 25 patients. Graft rejection occurred in 14% of the patients. However, after reconditioning and second stem cell donation, final engraftment was achieved in all patients. The median time to reach neutrophil and platelet recovery in patients with primary engraftment was 10 and 11 days respectively. All patients showed a rapid immune reconstitution with 250 (OKT3 conditioning) and 273 (ATG conditioning) CD3+ T-cells/µl, 30 (OKT3) and 47 (ATG) CD3+4+/µl and 300 (OKT3) and 382 (ATG) CD56+ NK-cells/µl at day +30 posttransplant. γδ+ T-cells started to expand faster than αβ+ T-cells in the early post-transplant period (156 vs. 82 cells/µl at day +30) whereas at day +90, αβ+ T-cells were predominant (170 vs. 134 cells/µl). Acute GvHD grade 0-I occurred in 25 patients (71%); 6 patients had GvHD II (17%), 3 patients had GvHD III (9%) and one patients experienced GvHD grade IV (3%). 3 patients experienced chronic GvHD (8%). Incidence of acute GvHD was not influenced by the number of residual T cells or by the type of serotherapy. 1 year EFS for patients with acute leukemias was 66% (any CR) and 14% (active disease).TRM at 1 year was 20%. Conclusions These data indicate that transplantation of TcR αβ+/CD19 depleted cells from a haploidentical donor results in sustained engraftment, remarkably fast immune reconstitution and low incidence of both acute and chronic GvHD. OKT3 could be substituted by ATG without negative effects. The anti-leukemic efficacy of this approach in comparison to other methods of T-cell depletion needs to be evaluated with a longer patient follow-up. Disclosures: No relevant conflicts of interest to declare.

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HLA-Mismatch Is Associated With Worse Outcomes After Unrelated Donor Reduced Intensity Conditioning Hematopoietic Cell Transplantation: A Cibmtr Analysis

Blood ◽

10.1182/blood.v122.21.547.547 ◽

2013 ◽

Vol 122 (21) ◽

pp. 547-547 ◽

Cited By ~ 2

Author(s):

John Koreth ◽

Kwang Woo Ahn ◽

Joseph Pidala ◽

James L. Gajewski ◽

Hailin Wang ◽

...

Keyword(s):

T Cell ◽

Acute Gvhd ◽

Hematopoietic Cell Transplantation ◽

Chronic Gvhd ◽

Cell Depletion ◽

Unrelated Donor ◽

T Cell Depletion ◽

Gvhd Prophylaxis ◽

Conditioning Intensity

Abstract In myeloablative unrelated donor allogeneic hematopoietic cell transplantation (HCT) a 1-locus HLA-mismatch (-A, -B, -C, -DRB1) is associated with lower survival compared to fully matched pairs. However data in reduced-intensity and non-myeloablative conditioning (together called RIC) HCT are limited. We analyzed adult AML/ALL/CML/MDS recipients of first 8/8 HLA-matched or 1-locus mismatched unrelated donor (MUD, MMUD) RIC HCT performed in the period 1999-2011 and registered in the CIBMTR. HLA-A, -B, -C and -DRB1 loci were typed in all pairs at high resolution; -DQB1 and -DPB1 loci could not be evaluated in all pairs. Transplants involving ex-vivo T-cell depletion, CD34+ selection, or post-transplant cyclophosphamide were excluded. Overall survival (OS) was the primary outcome. Secondary outcomes included non-relapse mortality (NRM), relapse, disease-free survival (DFS) and acute and chronic GVHD. Individual locus mismatch was also assessed. Apart from HLA matching, variables related to patient (age, race, sex, KPS, diagnosis, disease-risk), donor (age, parity), both (sex match/ABO match/CMV match) treatment (conditioning intensity, TBI use, in-vivo T-cell depletion (ATG), graft source (PB, BM) and GVHD prophylaxis (CyA-, Tac-based)) were considered. 2588 RIC HCT (8/8 MUD: 2025; 7/8 MMUD: 563) from 144 centers and 12 countries were analyzed. Median follow up in 8/8 MUD and 7/8 MMUD was 38 and 48 months respectively. Diagnoses were AML (65%), ALL (8%), CML (7%), MDS (20%). Conditioning intensity was RIC (79%), NMA (21%). 58% received in-vivo T-cell depletion. Graft source was PBSC (85%), BM (15%). GVHD prophylaxis was Tac-based (70%), CyA-based (27%). Mismatches involved HLA-A (188), -B (81), -C (219), and -DRB1 (75); with -DPB1 and -DQB1 typing available in 1382 and 2502 cases respectively. Compared to 8/8 MUD, 7/8 MMUD recipients were more likely to be younger and ethnic minorities and to have older and parous donors. In univariate analyses DQB1- and -DPB1 mismatch was not associated with worse OS, DFS, or NRM and was not further evaluated. There was a trend toward more grade II-IV acute GVHD in -DPB1 double (p=0.02) but not single mismatches. In multivariate models 7/8 MMUD RIC HCT had worse grade II-IV and III-IV acute GVHD, NRM, DFS and OS, but not relapse or chronic GVHD (Table). No significant interactions were identified between degree of HLA matching and other clinical variables. Adjusted 1- and 3-year NRM for 8/8 MUD vs. 7/8 MMUD was 20.4% vs. 28.9% (p<0.0001) and 29.2% vs. 38.1% (p<0.0007) respectively. Adjusted 1- and 3-year OS was 54.7% vs. 48.8% (p=0.01) and 37.4% vs. 30.9% (p=0.005) respectively (Figure). There was no difference between allele and antigen mismatches. HLA-A, -B, -C, and -DRB1 locus mismatches were each associated with 1 or more impaired outcomes (acute GVHD, NRM, DFS, and/or OS). Table 1 7/8 vs. 8/8 HLA HR (95% CI) p-value Acute GVHD II-IV 1.29 (1.09-1.53) 0.003 Acute GVHD III-IV 1.69 (1.00-3.36) 0.05 Chronic GVHD 1.11 (0.96-1.28) 0.15 Relapse 1.01 (0.87-1.17) 0.92 NRM 1.52 (1.29-1.79) <0.0001 DFS 1.20 (1.07-1.34) 0.0015 OS 1.25 (1.11-1.40) 0.0001 Compared to 8/8 MUD, both 7/8 allele and antigen MMUD RIC HCT have greater treatment toxicity and worse survival, of a magnitude similar to that seen in myeloablative transplantation. An isolated mismatch at HLA-A, -B, -C, or -DRB1 was associated with 1 or more adverse outcomes. In unrelated donor RIC HCT, matching for all alleles of HLA-A, -B, -C and -DRB1 loci results in superior outcomes. Disclosures: No relevant conflicts of interest to declare.

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Synergy of Unrelated Donor and Full Intensity Conditioning Breaks the Control of Graft Versus Host Disease By Alemtuzumab

Blood ◽

10.1182/blood.v124.21.1206.1206 ◽

2014 ◽

Vol 124 (21) ◽

pp. 1206-1206

Author(s):

Olivia Laverick ◽

Amy Publicover ◽

Laura Jardine ◽

Kile Green ◽

Alan Potter ◽

...

Keyword(s):

T Cell ◽

Acute Gvhd ◽

Chronic Gvhd ◽

Cell Depletion ◽

Reduced Intensity Conditioning ◽

T Cell Depletion ◽

Gvhd Prophylaxis ◽

Full Intensity ◽

Conditioning Intensity ◽

Reduced Intensity

Abstract Many variables influence the risk of graft versus host disease following hematopoietic stem cell transplantation. Comparison between preparative regimens is hampered by the use of many different combinations of chemotherapy and radiotherapy, varying intensity of conditioning, use of T cell depletion and donors who are either siblings or unrelated volunteers. Many reduced intensity regimens also incorporate enhanced GVHD prophylaxis with in vivo T cell depletion. Here we describe a cohort of patients prepared in a modular fashion with either reduced or full intensity conditioning combined with a uniform GVHD prophylaxis regimen for all transplants with sibling donors (alemtuzumab 30mg) and for all with unrelated donors (UD; alemtuzumab 60mg). Thus it was possible to dissect independently the effect of conditioning intensity and sibling or UD type upon GVHD risk in this settig of in vivo T cell depletion. Patients and analysis: the study was a retrospective analysis of 258 sequential transplants performed in adults with hematological malignancy between September 2005 and September 2013 at a single UK institution. Reduced intensity conditioning (n = 221) included fludarabine 150mg/m2 plus melphalan 140mg/m2 or fludarabine 150mg/m2 plus busulfan 9.6mg/kg. Full intensity transplants (n = 37) received 12Gy TBI plus melphalan 140mg/m2, 12Gy TBI plus cyclophosphamide 120mg/kg, or busulfan 16mg/kg plus cyclophosphamide 120mg/kg. All patients with sibling donors received 30mg alemtuzumab and those with UD received a 60mg of alemtuzumab. UD matching was similar in both reduced intensity and full intensity cohorts (92.2% and 86.5% 10/10 matches, respectively) but patients receiving reduced intensity were older than those receiving full intensity conditioning (median age 51 vs 31; p < 0.001). Outcome was analyzed according to EBMT guidelines. Relapse, non-relapse mortality and cGVHD were treated as competing risks and analysed as cumulative incidence. Outcome: the incidence of acute GVHD grades I-IV was comparable between reduced intensity and full intensity sibling transplants (45% vs 45%; p = NS) indicating a lack of effect of conditioning intensity upon GVHD risk in this setting. There was a slight increase in the risk of GVHD between reduced intensity UD compared with reduced intensity sibling donor transplants (57% vs 45%; p = NS) but a marked synergistic increase between UD transplants performed with full intensity compared with reduced intensity conditioning (100% vs 57%; p = < 0.001). The incidence of grades III-IV acute GVHD was also higher in full intensity UD transplants (16%) compared with reduced intensity UD transplants (5%). The incidence of chronic GVHD was also highest in full intensity UD transplants but both conditioning intensity and UD contributed in an additive manner: the rate of chronic GVHD progressed from 33% to 44% in reduced intensity and full intensity sibling transplants respectively and from 57% to 75% for reduced and full intensity UD transplants, respectively. Two year overall survival was comparable in all groups, ranging from 55% to 70%. In keeping with the higher rates of acute GVHD in full intensity transplants performed with UD, this group experienced the lowest relapse risk (15% vs 29% for all the other groups combined; p = 0.04) but the highest non-relapse mortality, reaching 41% at 2 years compared with 28% for all the other groups combined (p = 0.08). Conclusion: these results show that alemtuzumab provides good protection from acute GVHD in reduced intensity transplantation from sibling and UD. In sibling transplants given identical GVHD prophylaxis, full intensity conditioning does not increase the risk of GVHD. In contrast, a slight increase in GVHD risk with UD transplants seen with reduced intensity conditioning, is amplified in a synergistic manner by full intensity conditioning. This is associated with a high non-relapse mortality, even though the median age of full intensity patients is more than 20 years younger than those receiving reduced intensity conditioning. Disclosures No relevant conflicts of interest to declare.

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Chronic Graft-Versus-Host Disease after Allogeneic Stem Cell Transplant with in Vivo T-Cell Depletion By Alemtuzumab: Incidence, Outcome and Pattern of Organ Involvement

Blood ◽

10.1182/blood.v124.21.3912.3912 ◽

2014 ◽

Vol 124 (21) ◽

pp. 3912-3912 ◽

Cited By ~ 1

Author(s):

Maria Chiara Finazzi ◽

Cristina Boschini ◽

Janice Ward ◽

Charles Craddock ◽

Alessandro Rambaldi ◽

...

Keyword(s):

Stem Cell ◽

T Cell ◽

Acute Gvhd ◽

Stem Cell Transplant ◽

Chronic Gvhd ◽

Cell Depletion ◽

Organ Involvement ◽

Cell Transplant ◽

Allogeneic Stem Cell Transplant ◽

T Cell Depletion

Abstract Introduction Graft-versus-Host Disease (GvHD) is one of the leading causes of mortality and morbidity following allogeneic stem cell transplant. In vivo T cell depletion by alemtuzumab as part of the transplant conditioning is an effective strategy to reduce the risk of GvHD. While it is recognised that the overall incidence of GvHD is reduced by alemtuzumab, the incidence of chronic GvHD as defined by the National Institute of Health (NIH) consensus criteria, the impact on outcome, and the pattern of organ involvement have not been defined yet in this transplant setting. Methods Consecutive patients (n = 323) undergoing allogeneic stem cell transplantation at the Queen Elizabeth Hospital, Birmingham, between January 1 2008 and June 30 2012 were reviewed in this retrospective, single centre study. Medical records were examined and data regarding the development of GvHD were collected; NIH consensus criteria for diagnosis and staging of chronic GvHD were stringently applied. Clinical characteristics of GvHD occurring in patients transplanted following T cell depletion by alemtuzumab administration (n=248) were compared with those of patients transplanted with a T cell replete graft (n=75). Patients receiving alemtuzumab were mainly treated with reduced-intensity conditioning protocols, while patients in the no-T-cell depletion group were mainly treated with a myeloablative, sibling transplant. Results After a median follow up of 38.4 months, the cumulative incidence (CI) of grade II-IV classic acute GvHD was 35% and 48% for patients transplanted respectively with or without T cell depletion by alemtuzumab (p= 0.041, Figure 1); with a CI of grade III-IV classic acute GvHD of 13% and 27% (p=0.007). The 2-years CI of grade II-IV late acute GvHD was not significantly different in the two groups (20% and 23% for patients respectively treated with or without alemtuzumab, p=0.589, Figure 2). T cell depletion by alemtuzumab significantly reduces the 3 years cumulative incidence of classic chronic GvHD (5% versus 31%, p<0.0001, Figure 3.A), but without a significant difference in the incidence of overlap syndrome between patients with and without T cell depletion (3 years CI respectively 6% and 7%, p=0.839, Figure 3.B). The pattern of organ involvement by classic acute GvHD was similar in patients with and without T cell depletion. The pattern of organ involvement by late acute GvHD in the alemtuzumab group was, however, significantly different compared to the T cell replete group (skin-gut-liver involvement reported respectively in 83%-28%-4% of patients and 56%-48%-20% of patients, p=0.003). Distribution of organ involvement by classic chronic and overlap syndrome was similar in the two groups; however, it seems that alemtuzumab prevents the development of lung GvHD (lung GvHD developed in 4 patients over the 75 patients of the no-T-cell depletion group, while none of the 248 patients transplanted with alemtuzumab experienced lung GvHD). In a multivariate analysis, the development of chronic GvHD was an independent predictor of higher mortality risk (HR 1.66, p = 0.04) and severe NIH global score at peak was confirmed as a poor prognostic factor for survival (HR 2.27, p=0.02). The negative impact of chronic GvHD and of the severe forms of chronic GvHD was independent of age and alemtuzumab administration. Conclusion This retrospective analysis provides for the first time data on the incidence rates of NIH-defined GvHD categories in patients transplanted after T cell depletion by alemtuzumab. Patients transplanted with alemtuzumab experienced a lower incidence of classic acute and classic chronic GvHD compared to patients not receiving T cell depletion. In contrast, alemtuzumab conditioning appeared to have no effect on the incidence of late acute GvHD or overlap syndrome, suggesting that these two entities of GvHD are driven by different immunological mechanisms as compared to classic acute and classic chronic GvHD. We also confirmed the utility of the NIH classification of GvHD and of the NIH global severity score to predict survival in alemtuzumab-conditioned allogeneic stem cell transplant. Disclosures No relevant conflicts of interest to declare.

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Decreased Incidence of GvHD after Reduced Intensity Conditioning and a Fixed T-CELL Dose in Hematological Malingnancy Patients

Blood ◽

10.1182/blood.v124.21.5889.5889 ◽

2014 ◽

Vol 124 (21) ◽

pp. 5889-5889 ◽

Cited By ~ 1

Author(s):

Audrey Simon ◽

Eddy Roosnek ◽

Yordanka Tirefort ◽

Yan Beauverd ◽

Carole Dantin ◽

...

Keyword(s):

Stem Cell ◽

T Cell ◽

Disease Risk ◽

Risk Index ◽

Chronic Gvhd ◽

Univariate Analysis ◽

Cell Depletion ◽

Unrelated Donor ◽

T Cell Depletion ◽

Two Factors

Abstract Introduction: To decrease graft versus host disease (GvHD), the Geneva transplantation team has performed allogeneic hematopoietic stem cell (alloHSCT) with reduced intensity conditioning (RIC) and T cell depletion (TCD) to treat hematological malignancies for older or non fit for myeloablative conditioning patients. This is a new approach of engineering stem cell products that lowers the risk of GvHD while preserving graft versus leukemia (GvL) as much as possible. Patient and methods: We report a retrospective study of 73 patients who received alloHSCT with RIC and TCD between 2001-2013. The median age was 59 years (21-70), 60% were male. Disease at transplant time was acute leukaemia for 45%, Hodgkin lymphoma and non-Hodgkin lymphoma for 24%, myelodysplastic disorders for 13%, myeloproliferative disorders for 9,3 % and multiple myeloma for 8%. Source of stem cell was peripheral in 96% of the cases. 41% of the donors were matched related donor, 37% matched unrelated donor, 19% mismatched unrelated donor and 3% mistmatched related donor. The conditioning regimen consisted on fludarabine with busulfan or melphalan and ATG. Extensive T-cell depletion was done using Campath in the bag followed by washing procedures to remove free antibody. Fixed number of CD3+ T-cell addback was given on d+1 to preserve GvL with minimal residual disease (MRD) assessment and early donor lymphocyte infusions (DLI) given if MRD positive. Doses of DLI were preserved and frozen at the time of stem cell harvest. GvHD prophylaxis was with ciclosporine and mycophenolate mofetil. Results: With a median follow up of 5 (0.5-11) years, the 5-year overall survival (OS), disease free survival (DFS), current disease free survival, relapse rate and non relapse mortality (NRM) were 41.7% (95%CI 30.7-53.7%), 38.8% (95%CI 28.8-50.8%), 39,5% (95%CI 27.7-51.7%), 45.3% (95%CI 32.7-57.2%) and 15.8% (95%CI 8.3-25.4%) respectively. The main cause of death was relapse 38.7 % followed by GvHD 17% and infection 1.3%. In this cohort, the cumulative incidence (CI) of acute GvHD was 15.1% (95% CI: 8.0-24.3%) as well as for acute GvHD grade II-IV. CI of chronic GvHD was 14.7% (95%CI:7.2-23.6%) with extensive chronic GvHD CI being 5.9% (95% CI: 1.9-13.4%). Five patients received DLI for relapses, 27 for mixed chimerism and 8 for both causes. The average number of DLI was 2. Twenty-eight patients entered CR, 4 PR and 13 did not respond to DLI. In univariate analysis, two factors GvHD before DLI and GvHD after first DLI have a tendency for favorable impact on OS respectively p=0.093 and 0.071. For DFS, two factors are significant: disease risk index and GvHD after first DLI respectively p=0.013 and 0.044. For NRM disease risk index is the only factor which is statistically significant p=0.005. For relapse no factors were significant. Discussion: Our study showed a lower rate of acute and chronic GvHD as compared to other studies with unmanipulated stem cells. However, we describe a high rate of relapse incidence and relapse mortality. We have found in univariate analysis two factors statistically significant for DFS GvHD before and after first DLI. Our cohort is a heterogeneous group with different diseases at different stages, which can explain those results. It’s a monocentric study and small number of patient can be a limit for this work. Of note, since 2009 we have changed our strategy introducing a day +100 preemptive DLI infusion in the absence of GvHD, with escalading doses of lymphocytes every 8 weeks up to 5x 107 CD3/kg in the absence of GvHD to improve response. We don’t have enough patients and follow up to draw any conclusion regarding this new strategy. To improve the outcomes, the selection of patients who may receive partial T-cell depletion should be refined, avoiding transplanting patients with high risk of relapse with this strategy. To help decision making, the revised disease risk index as presented by Armand et al. (Blood 2014;123:3664) may be useful. Disclosures No relevant conflicts of interest to declare.

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In Vivo T Cell Depletion with Thymoglobulin or Alemtuzumab Is Associated with Worse Outcome Following Allogeneic Hematopoietic Stem Cell Transplantation for Acute Myeloid Leukemia Patients Transplanted in Remission

Blood ◽

10.1182/blood.v126.23.3115.3115 ◽

2015 ◽

Vol 126 (23) ◽

pp. 3115-3115

Author(s):

Scott R. Solomon ◽

Melissa Sanacore ◽

Xu Zhang ◽

Katelin Connor ◽

Melhem Solh ◽

...

Keyword(s):

Stem Cell ◽

T Cell ◽

Hematopoietic Stem Cell Transplantation ◽

Hematopoietic Stem Cell ◽

Myeloid Leukemia ◽

Chronic Gvhd ◽

Cell Depletion ◽

T Cell Depletion ◽

Hematopoietic Stem

Abstract In vivo T Cell Depletion with Thymoglobulin or Alemtuzumab Is Associated With Worse Outcome Following Allogeneic Hematopoietic Stem Cell Transplantation for Acute Myeloid Leukemia Patients Transplanted in Remission. Allogeneic hematopoietic stem cell transplantation (HSCT) reduces relapse risk in adults with acute myeloid leukemia (AML) due in large part to the potent graft-versus-leukemia effect of donor lymphocytes. However, this benefit must be balanced by the increased morbidity and mortality associated with graft-versus-host disease (GVHD). Serotherapy, in the form of thymoglobulin or alemtuzumab, has been used for in vivo T cell depletion as a strategy to reduce GVHD. We analyzed 144 consecutive AML patients transplanted in remission (CR1 - 111, CR≥2 - 33) from either a matched related (MRD, n=44), unrelated (MUD, n=62), or haploidentical (haplo, n=38) marrow of PBSC donor, in order to analyze the effect of serotherapy, in relation to other disease-, patient- and transplant-related risk factors, on post-transplant outcomes. Patients were transplanted at a single institution between 3/15/06 to 12/19/14. Baseline characteristics of the patient cohort included age >50 in 88 (61%), KPS<90 in 93 (65%), CMI ≥3 in 61 (42%) of patients. Disease risk index (DRI) was defined as low, intermediate, and high in 5 (4%), 110 (76%), and 29 (20%) patients respectively per the revised Dana Farber/CIBMTR criteria. Myeloablative chemotherapy was given in 96 (67%) patients, and PBSC was the source of stem cells in 120 (83%) patients. Serotherapy was utilized in 21 (15%) patients [thymoglobulin - 8, alemtuzumab - 13]. Serotherapy patients were more likely to be older (median age 59 vs. 52 years, p=0.013) and have a MUD (81% vs. 37%, p<0.001), but otherwise had similar baseline characteristics in regards to disease status, DRI, regimen intensity. Acute GVHD grade II-IV occurred in 38% of patients, whereas chronic GVHD was seen in 44%. Chronic GVHD occurred less often in patients receiving serotherapy (19% vs. 49%, p=0.016). Estimated one year non-relapse mortality (NRM) at 1 and 3 years was 4% and 13% respectively and was statistically similar in serotherapy and non-serotherapy patients. The estimated 3 year OS, DFS, and relapse was 58%, 51%, and 37% respectively for the whole cohort; 64%, 55%, and 33% in non-serotherapy patients vs. 29%, 27%, and 57% in serotherapy patients (figure 1). Cox analysis was performed utilizing the following variables: age, disease status, DRI, KPS, CMI, transplant type (MRD, MUD, haplo), conditioning intensity, stem cell source, use of serotherapy, year of transplant, acute and chronic GVHD. Variables were selected by a 10% threshold. Acute and chronic GVHD were modeled as time-dependent variables. In multivariate analysis, unfavorable risk factors for survival included only two variables: the use of serotherapy (HR 3.11, p<0.001) and high risk DRI (HR 1.89, p=0.038). Use of serotherapy also had a negative effect on relapse (HR 2.69, p=0.003) and DFS (HR 2.73, p<0.001), with no effect on NRM. Following allogeneic HSCT for AML patients in remission, the use of serotherapy for in vivo T cell depletion had a major negative impact on survival due to increased relapse risk. Figure 1. Figure 1. Disclosures No relevant conflicts of interest to declare.

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Comparable Outcomes after Allogeneic Stem-Cell Transplantation with Intravenous Busulfan or Treosulfan-Based Reduced-Intensity and Reduced Toxicity Regimens in Acute Myeloid Leukemia. a Study on Behalf of the Acute Leukemia Working Party of EBMT

Blood ◽

10.1182/blood.v128.22.2280.2280 ◽

2016 ◽

Vol 128 (22) ◽

pp. 2280-2280

Author(s):

Avichai Shimoni ◽

Myriam Labopin ◽

Bipin N. Savani ◽

Rose-Marie Hamladji ◽

Dietrich W. Beelen ◽

...

Keyword(s):

Stem Cell ◽

T Cell ◽

Acute Gvhd ◽

Advanced Disease ◽

Chronic Gvhd ◽

Working Party ◽

Cell Depletion ◽

T Cell Depletion ◽

Reduced Toxicity

Abstract Allogeneic stem cell transplantation (SCT) is a potentially curative therapy for patients with acute myeloid leukemia (AML). Myeloablative conditioning (MAC) is associated with prohibitive rates of non-relapse mortality (NRM) in older and less medically fit patients. Several reduced intensity conditioning regimens (RIC) and more recently the more dose-intensive reduced toxicity myeloablative (RTC) regimens were designed to replace MAC in this setting. The backbone of these regimens is usually fludarabine with busulfan and more recently also with treosulfan, but there is no clear data on the comparative outcomes with these different regimens in the different SCT settings. The current study included 3561 patients with AML given a first allogeneic SCT from an HLA-matched sibling (n=1683) or a 10/10 matched unrelated donor (n=1878) between the years 2000-2014 and reported to the acute leukemia working party (ALWP) of EBMT. Only patients given fludarabine with either intravenous busulfan (ivBu), (FB, n=2990) or treosulfan (FT, n=571) alone were analyzed. Fludarabine and ivBu at 6.4 mg/kg (n=1457) or treosulfan at 30-36 gr/m2 (n=168) were considered RIC regimens while fludarabine with ivBu at a total dose of 9.6-12.8 mg/kg (n=1533) or treosulfan at 42 gr/m2(n=403) were considered RTC regimens according to EBMT criteria. The median age of FB and FT recipients was 55.5 and 58.3 years, respectively (P< 0.0001). The status at SCT was 72.5% CR1, 15.0% CR2 and 12.5% advanced disease in the FB group compared to 55.0%, 20.3% and 24.7% in the FT group, respectively (P<0.0001). More FT recipients had SCT from unrelated donors (64.8% Vs. 50.4%, P<0.0001) but less had in-vivo T-cell depletion (58.4%Vs 70.5%, P<0.0001). Cytogenetic subgroup distribution was similar between the groups. Ninety percent had peripheral blood stem cell grafts in both groups. The median follow-up was 19 and 43 months after FB and FT, respectively. Using univariant analysis, the 2-year relapse incidence (RI) was 32.7% and 35.5%, respectively (P=0.49). NRM was 17.6% and 19.4%, respectively (P=0.09). Leukemia-free survival (LFS) and overall survival (OS) were 49.5% and 54.8% after FB and 45.1% and 52.6% after FT, respectively (P=0.04, P=0.17). Acute GVHD grade II-IV and chronic GVHD were 23.1% and 35.7% after FB and 18.8% and 39.8% after FT, respectively (P=0.03, P=0.04). In all, the GVHD/ relapse-free survival (GRFS) was 36.5% and 31.5%, respectively (P=0.08). After adjusting for the differences in patient characteristics, there was no difference between the FB and FT groups in RI, NRM, LFS, OS and GRFS. However, acute GVHD grade (II-IV) was higher after FB (HR, 1.49, P=0.0004). The same observations were seen when the analysis was limited to RIC or RTC regimens only, or when only patients in remission were analyzed. However, when analyzing only the 516 patients with advanced disease at SCT, 2-year OS was 29.7% and 43.0% after FB and FT (P=0.002) and this difference remained significant in the multivariant analysis (HR, 1.50, p=0.003). Among the entire group, the factors associated with reduced survival were advanced age (HR 1.01, P<0.0001), secondary AML (HR 1.19, P=0.005), CR2 (HR 1.21, P=0.007) and advanced disease (HR 2.02, P<0.0001) compared to CR1, and female donor to male recipient (HR 1.15, P=0.03). Conditioning type and intensity, donor type, CMV status and in vivo T-cell depletion were not significant. Relapse was lower and NRM was higher with RTC compared with RIC, but OS was similar. The same factors predicted for GRFS, a surrogate for quality of life, with the only difference been the positive role of in vivo T-cell depletion (HR 0.8, P=0.0002). In conclusion, RIC and RTC regimens with ivBu or treosulfan-based regimens are associated with similar transplantation outcomes. OS is primarily affected by disease factors such as status of disease at SCT and secondary leukemia. Treosulfan- based conditioning is associated with a lower rate of acute GVHD, but with similar rates of chronic GVHD, NRM and GRFS. Treosulfan conditioning may have some advantage in patients with advanced disease at SCT. Disclosures No relevant conflicts of interest to declare.

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NaïVe T Cell Depletion of PBSC Grafts Results in Very Low Rates of Chronic Gvhd and High Survival

Blood ◽

10.1182/blood.v128.22.668.668 ◽

2016 ◽

Vol 128 (22) ◽

pp. 668-668

Author(s):

Marie Bleakley ◽

Ted A. Gooley ◽

Barbara Hilzinger ◽

Stanley R Riddell ◽

Warren D Shlomchik

Keyword(s):

T Cells ◽

T Cell ◽

Acute Leukemia ◽

Chronic Gvhd ◽

Hematopoietic Cell ◽

T Cell Subsets ◽

Cell Depletion ◽

Effector Memory ◽

Phase Ii Trials ◽

T Cell Depletion

Abstract Background Graft-versus-host disease (GVHD) frequently causes morbidity and mortality after allogeneic hematopoietic cell transplantation (HCT) as a result of organ damage and infections. In HLA-identical HCT, GVHD results from recognition by donor T cells of minor histocompatibility (H) antigens on recipient tissues. Complete T cell depletion (TCD) of donor hematopoietic cell products is more effective than pharmacologic immunosuppression for preventing GVHD, but is complicated by delayed immune reconstitution and consequent life-threatening infections.Approaches to HCT which preferentially deplete the T cells that primarily cause GVHD and preserve pathogen-specific T cells may improve HCT outcomes. Mature CD3+ CD8+ and CD3+ CD4+ T cells can be classified into CD45RA+ CD62L+ naïve (TN) and CD45RO+ memory (TM) subsets, the latter of which includes effector memory (TEM) and central memory (TCM) cells. Murine studies in which allogeneic TCD bone marrow (BM) is transplanted with purified T cells from individual T cell subsets to irradiated minor H antigen disparate recipients have demonstrated that the most severe GVHD results from transplanting T cells of the TN subset. Purified TCM causes mild GVHD and TEM do not cause detectable GVHD and can transfer immunity to pathogens.In vitro studies have similarly demonstrated that human donor CD8+ T cells specific for recipient minor H antigens are found predominantly within the TN cell subset, suggesting selective TN cell depletion may alter the GVHD incidence and/or severity in human HCT. Methods and results We developed an effective process for engineering human peripheral blood stem cell (PBSC) grafts that depletes CD45RA+ TN cells and retains CD34+ stem cells and functional CD45RO+ TM cells specific for a broad range of opportunistic pathogens (Bleakley BBMT 2014). We are conducting clinical trials to evaluate the selective depletion of TN cells from HLA-matched allogeneic PBSC grafts for the prevention of GVHD in patients with acute leukemia, the first of which has been published (Bleakley JCI 2015, N=35). Seventy patients have now been treated on three consecutive phase II trials. The median age was 34 years (1-56 years), 56% of patients had a diagnosis of ALL, 46% had previously relapsed or had detectable disease (MRD or relapse) at the time of HCT, and 23% had unrelated donor (URD) grafts. Intensive myeloablative, TBI-containing (13.2Gy) conditioning was used for 63 patients, whilst 7 patients received a medium intensity 'midi' preparative regimen, including 4Gy of TBI. The TN-depletion procedure was successfully performed on URD PBSC products shipped overnight from donor centers throughout the US, as well as on MRD PBSC collected at our centers. Reliable engraftment with high-level donor chimerism was observed in recipients of 'midi' as well as intensive myeloablative conditioning. The 2-year estimates of overall survival, disease-free survival, survival free of relapse and chronic GVHD (CRFS) and survival free of relapse, grade II-IV acute GVHD, and chronic GVHD (GRFS) are 79%, 73%, 69% and 63% respectively. Median follow-up among survivors is 26 months. The frequency and severity of chronic GVHD is remarkably low (5%) compared to historical rates of 40-60% chronic GVHD in HLA-matched PBSC transplantation with conventional calcineurin inhibitor-based immunosuppression. Relapse and non-relapse mortality (NRM) are acceptably low at 19% and 8%, respectively. No NRM occurred in patients <40 years. Updated results will be presented. Conclusions The outcomes of recipients of TN-depleted PBSC grafts compare very favorably to published results of HCT for patients with acute leukemia. For example, the 69% incidence of CRFS at 2 years in TN-depleted recipients compares with reported 2-year GRFS rates of 37% and 17% in recipients of allogeneic PBSC from HLA-matched related donors with or without ATG (Kroger et al. NEJM 2016). Our results suggest that TN-depletion of PBSC grafts may reduce the risk of chronic GVHD without negatively impacting other important HCT outcomes. Disclosures Riddell: Juno Therapeutics: Equity Ownership, Patents & Royalties, Research Funding; Cell Medica: Consultancy, Honoraria; Adaptive Biotechnologies: Consultancy, Honoraria.

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Post-Transplant Immune Suppressive Therapy Is Not Necessary In Related and Unrelated Reduced Intensity Conditioning Stem Cell Transplantation Using Low Dose Alemtuzumab In Vivo T Cell Depletion Combined with Alemtuzumab-Mediated In Vitro T Cell Depletion of the Graft

Blood ◽

10.1182/blood.v116.21.2328.2328 ◽

2010 ◽

Vol 116 (21) ◽

pp. 2328-2328

Author(s):

P.A. Von Dem Borne ◽

C.J.M. Halkes ◽

C.W.J. Starrenburg ◽

W.A.F. Marijt ◽

J.H.F. Falkenburg

Keyword(s):

T Cell ◽

Low Dose ◽

Chronic Gvhd ◽

Cell Depletion ◽

Suppressive Therapy ◽

T Cell Depletion ◽

Post Transplant ◽

Immune Suppressive Therapy

Abstract Abstract 2328 Introduction T cell depletion with alemtuzumab administered in vivo to the patient reduces the risk of graft-versus-host disease (GVHD) and graft rejection following reduced intensity conditioning allogeneic stem cell transplantation (RIC SCT). However, high doses of alemtuzumab can result in delayed immune reconstitution, increased non relapse mortality (NRM) due to infections, and potential loss of graft versus tumor responses. Recently, the feasibility of T cell depletion with low dose in vivo alemtuzumab was demonstrated in HLA-identical related RIC SCT. Dose reduction of alemtuzumab to 30 mg combined with post-transplant immune suppressive therapy with cyclosporine tapered from 3 months after transplantation resulted in a low risk of GVHD, no increase in NRM and improved lymphocyte recovery (Chakraverty et al, Blood pre-published online June 29, 2010). Early immunotherapeutic intervention after SCT with donor lymphocytes may be hampered by the administration of post-transplant immune suppressive therapy. We investigated whether in RIC SCT using low dose in vivo alemtuzumab, post-transplant immune suppressive therapy can be replaced by alemtuzumab-mediated in vitro T cell depletion of the graft just prior to infusion (“Campath in the bag”). Patient and donor characteristics Between 2007 and 2009, 29 patients were transplanted with an unrelated donor, and 28 patients with a related donor using a RIC regimen consisting of fludarabine (50 mg/m2 p.o. day -10 to -5), busulphan (3.2 mg/kg i.v. day -6 and -5) and alemtuzumab (15 mg i.v. day -4 and -3), followed by infusion of the graft after in vitro incubation with 20 mg alemtuzumab. No additional immune suppressive therapy was used after SCT. Unrelated donors were matched for HLA-A, B, C, DR and DQ, three patient donor combinations had one HLA-DQ mismatch. Median patient age was 59 years (range 21–72). Remission status at the time of transplant was: 47% CR, 37% PR, 9% SD, 7% PD. Indications were diverse (19 AML, 16 myeloma, 6 CLL, 5 low grade NHL, 3 aggressive T-NHL, 2 aggressive B-NHL, 2 SAA, 1 CML, 1 myelofibrosis, 1 CMML, 1 ALL). The unrelated and related transplanted group were comparable regarding age, disease and remission status. The median Gratwohl transplantation risk score was 3 in the related group (range 1–5), and 5 in the unrelated group (range 3–6). Results All patients engrafted; platelet numbers of 50 × 109/L were reached after a median of 11 days (range 0–38 days), neutrophil numbers of 0.5 × 109/L were reached after a median of 18 days (range 0–161 days) post transplant. Two patients had secondary graft failure. In patients transplanted with a related donor, grade 1–2 and 3–4 acute GVHD was observed in 36% and 4% of evaluable patients, respectively, resolving in all patients without development of chronic GVHD. NRM was 0% at 3 months and 4% at 1 year. Overall survival was 100% at 3 months and 89% at 1 year. In patients transplanted with an unrelated donor more acute GVHD was observed (59% grade 1–2, 15% grade 3–4 of evaluable patients). 24% of evaluable patients developed chronic GVHD, which was limited in 75% and extensive in 25% of these patients. Chronic GVHD resolved in most patients, one patient has ongoing extensive chronic GVHD. NRM was 7% at 3 months and 24% at 1 year. Overall survival was 93% at 3 months and 55% at 1 year. Conclusions RIC SCT using low dose alemtuzumab in vivo T cell depletion combined with alemtuzumab-mediated in vitro T cell depletion of the graft without additional post-transplant immune suppressive therapy is feasible in patients transplanted with related and unrelated donors. Results are excellent in patients transplanted with related donors with low NRM and high overall survival. Although results are good in patients transplanted with unrelated donors considering the high Gratwohl score in this group, further improvement is being sought by increasing the efficiency of T cell depletion. This RIC SCT regimen without post-transplant immune suppressive therapy is an appropriate platform for early cellular immunotherapeutic interventions including donor lymphocyte infusion after SCT. Disclosures: No relevant conflicts of interest to declare.

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Functionally Altered GPI-Anchor Negative Treg Following Alemtuzumab-Based T-Cell Depletion Are Associated with Acute Gvhd.

Blood ◽

10.1182/blood.v120.21.3059.3059 ◽

2012 ◽

Vol 120 (21) ◽

pp. 3059-3059

Author(s):

Eva M Wagner ◽

Lukas A Schaefer ◽

Tobias Bopp ◽

Matthias Theobald ◽

Wolfgang Herr ◽

...

Keyword(s):

T Cells ◽

T Cell ◽

Acute Gvhd ◽

Chronic Gvhd ◽

Cell Depletion ◽

Gpi Anchor ◽

Activation Markers ◽

T Cell Depletion ◽

Hematopoietic Stem

Abstract Abstract 3059 Introduction: The monoclonal anti-CD52antibody Alemtuzumab is frequently used for T-cell depletion (TCD) in the context of allogeneic hematopoietic stem cell transplantation (HSCT) to prevent graft versus host disease (GVHD). We previously demonstrated the long term persistence of functionally impaired glycosylphosphatidylinositol (GPI)-anchor negative effector T-cells in patients receiving high dose (100mg) Alemtuzumab in combination with a dose reduced conditioning regimen (Fludarabin + Melpahlan) (Meyer, Wagner et al. BMT 2010). Despite of Alemtuzumab-mediated TCD, half of our patients developed acute GVHD. Since regulatory T cells (Treg) play a major role for controlling GVHD, we asked whether GPI-anchor negative Treg are present in patients with or without GVHD. Methods: We analyzed peripheral blood samples of 12 patients with acute GVHD (aGVHD), 7 patients with chronic GVHD (cGVHD), and 10 patients who never developed GVHD after Alemtuzumab-mediated TCD. To analyze Treg-subsets, we stained for CD3, CD4, CD25, CD127, FoxP3, CD52 as well as for the activation-markers GARP, HLA-DR and CD45RA. Treg were identified as CD3+CD4+CD25+CD127- or CD3+CD4+CD25+FoxP3+ cells and subdivided according to their CD52-expression. We used FLAER staining to confirm that the loss of CD52 on Treg resulted from the loss of the GPI-anchors themselves. We were able to study Treg subpopulations in the time course of patients who recovered from acute GVHD in comparison to patients with persisting late acute GVHD. In individual patients, we isolated GPI-anchor positive and negative Treg by FACS-Sort, expanded them and performed Treg suppression assays. Results: GPI-anchor negative Treg were observed in all patients, independent of the development of GVHD. However, the frequency of GPI-anchor negative Treg varied considerably between patients with acute GvHD and those with chronic GVHD or without GvHD. The percentage of GPI-anchor negative Treg was significantly elevated in patients with aGVHD: median 80.35% (range 56,2–96,8%) in comparison to 17,4% (range 0–57,8%) in patients with cGVHD or without GVHD. Activated Treg were almost exclusively detected among GPI-anchor positive Treg-subpopulation. Patients who resolved from aGVHD restored GPI-anchor positive Treg and the amount of activated Treg rose. The percentage of GPI-anchor negative Treg populations remained high in patients with ongoing aGVHD. In addition, these patients had no GARP-positive activated Treg even under long term immunosuppressive treatment. Preliminary experiments with sorted and expanded Treg populations suggest that GPI-anchor negative Treg were unable to suppress T-cell proliferation upon IL-2 stimulation. Summary: We demonstrate for the first time the reconstitution of GPI-anchor negative Treg in patients following Alemtuzumab-mediated TCD. These T cells were functionally altered and were less likely to exhibit an activated phenotype in vivo. Ongoing acute GVHD was associated with high percentages of GPI-negative Treg suggesting that their functional alteration might play a role in aGVHD pathophysiology. This is in line with the finding that only in patients who resolved aGVHD, the frequency of GPI-anchor positive Treg increased significantly. Further functional analyses are ongoing to estimate the cellular consequence of missing GPI-anchored proteins. In addition, correlating the reconstitution of GPI-anchor negative T-cell populations with further clinical events is ongoing. Disclosures: No relevant conflicts of interest to declare.

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