Immune Reconstitution after Haploidentical Hematopoietic Cell Transplantation: Impact of Reduced Intensity Conditioning and CD3/CD19-Depleted Grafts

Blood ◽  
2008 ◽  
Vol 112 (11) ◽  
pp. 1175-1175
Author(s):  
Birgit Federmann ◽  
Matthias Haegele ◽  
Christoph Faul ◽  
Wichard Vogel ◽  
Lothar Kanz ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Nk Cells ◽  
Cell Transplantation ◽  
Immune Reconstitution ◽  
Hematopoietic Cell Transplantation ◽  
Nk Cell ◽  
Reduced Intensity Conditioning ◽  
Haploidentical Hematopoietic Cell Transplantation ◽  
Reduced Intensity

Abstract Haploidentical hematopoietic cell transplantation (HHCT) using CD3/CD19 depleted grafts may lead to faster engraftment and immune reconstitution since grafts contain also graft-facilitating-cells, CD34− progenitors, NK cells, and dendritic cells. Reduced intensity conditioning may also have a positive impact on immune reconstitution following HHCT. 26 adults received CD3/CD19 depleted HHCT after RIC (150–200 mg/m2 fludarabine, 10mg/kg thiothepa, 120 mg/m2 melphalan and 5mg/day OKT-3 (day −5 to +14)) at our institution between 2005–2008. We prospectively evaluated engraftment and immune reconstitution. B-, NK-, T- and T-cell subsets (CD3/8, CD4/8, CD4/45RA/RO), TCR-Vβ repertoire and NK-cell receptors (NKP30, NKP44, NKP46, NKG2D, CD158a/b/e, CD85j, NKG2A, CD161) were analyzed by FACS. Grafts contained 8.8×106 CD34+ (range, 4.3–18.0 ×106), 2.9×104 CD3+ (range, 1.2–9.2×104) and 3.6×107 CD56+ (range, 0.02–23.0 ×107) cells/kg. Engraftment was rapid with a median time to >500 granulocytes/μl of 11 days (range, 9–15) and a median time to >20 000 platelets/μl of 11 days (range, 8–23). Full chimerism was reached on day 14 (median; range, 6–26). NK-cell engraftment was rapid, reaching normal values on day 20 (median of 247 CD16+CD56+CD3− cells/μl (range, 1–886)) with NK cells comprising up to 70% of lymphocytes. B-cell reconstitution was delayed with 81 (range, 0–280) and 335 (range, 11–452) CD19+20+ cells/μl on days 150 and 400, respectively. T-cell reconstitution was impaired with 49 (range, 0–586) and 364 (range, 35–536) CD3+ cells/μl on day 60 and day 150, respectively. We observed an increase of CD3+CD8+ cells in contrast to CD3+CD4+ cells early after HHCT with a median of 24 (range, 0–399) vs 16 (range, 0–257) and 159 (range, 1–402) vs 96 (range, 18–289) cells/μl on day 50 and day 200, respectively. CD4+CD45RA+ T cells increased slowly while CD4+CD45RO+ T cells reconstituted faster with a median of 61 CD4+CD45RO+ cells/μl (range, 0–310) vs 24 CD4+CD45RA+ (range, 0 to 152) on day 100. Within the CD4+CD25+ regulatory T cells there was a slow regeneration with median of 14 CD4+CD25+ cells/μl (range, 0–96) on day 100 and 28 CD4+CD25+ cells/μl (range, 19–160) on day 200. CD14+CD45+ monocytes did not reach normal values within the time of observation with 7 CD14+CD45+ cells/μl (range, 0–21) on day 120 and 7 CD14+CD45+ cells (range, 2–381) on day 400. TCR-Vβ repertoire and NK-cell receptor reconstitution was analyzed so far in 7 and 8 patients, respectively. We found a skewed T-cell repertoire with oligoclonal T-cell expansions to day 100 and normalization after day 200. An increased natural cytotoxicity receptor (NKP30, NKP44, NKP46) and NKG2A, but decreased NKG2D and KIR-expression was observed on NK-cells until day 100. In conclusion, T- and B-cell reconstitution is delayed after HHCT using CD3/CD19 depleted grafts and RIC. However, T-cell reconstitution is faster compared to data published with CD34 selected grafts and myeloablative conditioning. A fast NK-cell reconstitution early after HHCT was observed. Thus a combination of reduced intensity conditioning with CD3/CD19 depleted grafts appears to accelerate the immune recovery after haploidentical stem cell transplantation.

Haploidentical TCRab Depleted Hematopoietic Cell Transplantation Leads To Rapid Immune Reconstitution and Promising Clinical Outcome In Adults

Blood ◽  
2013 ◽  
Vol 122 (21) ◽  
pp. 5475-5475
Author(s):  
Sebastian P. Haen ◽  
Michael Schumm ◽  
Wichard Vogel ◽  
Christoph Faul ◽  
Rupert Handgretinger ◽  
...  
Keyword(s):  
T Cells ◽  
Body Weight ◽  
Clinical Outcome ◽  
Cell Transplantation ◽  
Immune Reconstitution ◽  
Hematopoietic Cell Transplantation ◽  
Reduced Intensity Conditioning ◽  
Virus Pneumonia ◽  
Reduced Intensity ◽  
Observation Period

Abstract Immunomagnetic TCRab-depletion and reduced intensity conditioning (RIC) for haploidentical hematopoietic cell transplantation (HHCT) has been reported to result in improved immune reconstitution and clinical outcome in children. We here report the first clinical results in 10 adult patients. HHCT using TCRab-depleted grafts was performed in 7 men and 3 women (median age at transplantation 35 years, range 21-68 years). Patients were treated for ALL (n=1), AML (n=5), CLL (n=1), NHL (n=2) or SAA (n=1). Disease status prior transplantation was CR in two patients, minimal residual disease but cytologic CR in 1 patient, conditioning out of aplasia after salvage chemotherapie in 4 patients, and active disease in 3 patients. Haploidentical donors were parents (n=3), siblings (n=4) and children (n=3). Seven patients were treated with HHCT due to relapse after first (n=6) or second (n=1) allogeneic HCT. In 3 patients, HHCT was performed as primary transplantation. Conditioning regimen was myeloablative (12 Gy TBI/high dose cyclophosphamide) in 1 patient and reduced intensity conditioning (fludarabine/clofarabine, thiotepa, melphalan) in the remaining 9 patients. All conditioning regimens included ATG and mycophenolate mofetil as postgrafting immunosuppression until day +30. Patients received a median of 6.18 x 106 (range 3.4-20.4 x106) CD34+,5.92 x 106 (range 2.26-14.84) CD3+and 36.77 x 106 (range 15.75-51.84) CD56+ cells per kilogram body weight. TCRab-depletion of the grafts was a median of 4.5 logs with 0.38 104 TCRab+ CD3+ (range 0.06-1.17) cells per kilogram body weight in the graft. Median engraftment of neutrophils (> 500/µl) and thrombocytes (> 25.000/µl) occurred on days 12 (range day 8-15) and 14 (range day 10-23), respectively, with one patient not achieving thrombocyte engraftment until death. Patients were followed for a median of 276 days after HHCT (range 41-723 days). To date, 6 patients remain alive and in CR (60%). Four patients died due to viral infections (n=4, 40%) at a median of 92 days after HHCT (range 41-100 days) comprising adenovirus viremia, varicella zoster virus encephalitis, respiratory syncytial virus pneumonia and herpes simplex virus pneumonia. One patient also suffered from severe graft versus host disease (GvHD) of the liver. Acute GvHD ≥II occurred in a total of 5 patients with development of chronic GvHD not being observed. None of the patients experienced relapse of the underlying hematologic disease, the non-relapse mortality was 40%. Median overall and disease free survival were not reached during the observation period. Immune reconstitution was studied by flow cytometry at least every week for a median follow-up of 122 days after transplantation (range 25-558 days). Detailed information on immune reconstitution data is provided in the table. Median T cell (CD3+) engraftment (> 200/µl) occurred on day 42 (range 21-397 days). Two patients did not reach > 200/µl CD3+ cells until death. Engraftment of CD4+ T cells > 200/µl was only achieved in 3 patients during the observation period. Our data indicate that in comparison to approaches applying CD34 selected or CD3/CD19 depleted grafts, depletion of TCRab T cells in HHCT might lead to more rapid immune reconstitution. A prospective study evaluating the role of TCRab depletion in HHCT in adults is presently ongoing. Table Peripheral cell counts of leukocyte subsets. NK cells T cells T helper cells Cytotoxic T cells B cells CD56+ CD16+ CD3- CD3+ CD3+ CD4+ CD3+ CD8+ CD19+ Day 30 (median [/µl]) 435 28 3 12 1 range [/µl] 113-642 5-1,655 0-292 2-509 0-212 Day 60 (median [/µl]) 433 457 54 342 213 range [/µl] 188-504 1-1,367 0-175 0-368 0-556 Day 100 (median [/µl]) 319 74 9 39 82 range [/µl] 118-1,223 0-2,292 0-229 0-1,289 19-344 Peak (median [/µl]) 466 416 186 221 309 range [/µl] 134-2,650 5-4,529 3-787 2-4,029 1-3,065 Day Peak (median [day]) 52 62 59 62 63 range [days] 19-337 25-397 8-558 25-344 19-344 Disclosures: No relevant conflicts of interest to declare.

HLA-Haploidentical Familial Donor Hematopoietic Cell Transplantation without Ex Vivo- T Cell Depletion after Reduced-Intensity Conditioning of Busulfan, Fludarabine, and Anti-Thymocyte Globulin: A Preliminary Report.

Blood ◽  
2007 ◽  
Vol 110 (11) ◽  
pp. 3077-3077
Author(s):  
Kyoo-Hyung Lee ◽  
Seong-Jun Choi ◽  
Jung-Hee Lee ◽  
Ho-Jin Shin ◽  
Young-Shin Lee ◽  
...  
Keyword(s):  
T Cell ◽  
High Risk ◽  
Cell Transplantation ◽  
Acute Gvhd ◽  
Hematopoietic Cell Transplantation ◽  
Hematopoietic Cell ◽  
Cell Depletion ◽  
Reduced Intensity Conditioning ◽  
T Cell Depletion ◽  
Reduced Intensity

Abstract Animal hematopoietic cell transplantation (HCT) models and several small clinical trials showed that successful engraftment can be achieved across HLA-haplotype difference after reduced-intensity conditioning (RIC). Furthermore, decreased graft-versus-host disease (GVHD) and transplantation-related mortality (TRM) after RIC was shown in a swine leukocyte antigen-haploidentical HCT experiment. Therefore, a protocol investigating the role of RIC in HLA-haploidentical familial donor HCT was initiated in April 2004 and 20 patients [13 male and 7 female; median age 26.5 years (16–65)] without HLA-matched donor enrolled until June 2007. The diagnosis were AML (n=9), ALL (n=4), acute biphenotypic leukemia (n=1), MDS (n=4), and SAA (n=2), and all patients had high-risk features, i.e. first complete remission (CR) but with high-risk chromosomal abnormality (n=1), first CR after salvage (n=1), second CR (n=6), recurrent/refractory state (n=7), immunotherapy failure (n=4), and high-risk MDS (RAEB-1, n=1). The RIC included iv busulfan 3.2 mg/kg × 2, fludarabine 30 mg/m2 × 6, plus anti-thymocyte globulin [Thymoglobuline 3 mg/kg (n=17) or Lymphoglobuline 15 mg/kg (n=3)] × 4. After receiving G-CSF, the donors (13 mothers; 5 offsprings; and 2 HLA-haploidentical siblings) underwent 2 or 3 daily leukapheresis, and the collected cells were given to patients without T cell depletion [medians of; 7.9 (3.7–12.1)×108/kg MNC, 6.9 (3.6–73.5)×106/kg CD34+ cells, and 4.6 (1.8–8.5)×108/kg CD3+ cells]. GVHD prophylaxis was cyclosporine 3 mg/kg/day iv from day -1 and a short course of methotrexate. As a part of separate phase 1 study, the two most-recently enrolled patients received additional donor CD34+ cell-derived NK cells 6 weeks after HCT. Except one patients with SAA who died due to K. pneumoniae sepsis on day 18, all 19 evaluable patients engrafted with ANC> 500/μl median 17 days (12–53) and platelet> 20,000/μl median 23 days (12–100) after HCT. Eight patients experienced acute GVHD (grades I, II, III, and IV; 2, 3, 2, and 1, respectively). Cumulative incidences (CI) of overall and grade II-IV acute GVHD were 40 and 30%, respectively. Eight patients experienced chronic GVHD (limited, 4; extensive, 4; CI, 51%). Fourteen showed positive CMV antigenemia, while 2 suffered CMV colitis, which resolved after treatment. As early as 2 weeks after HCT, 15 of 16 evaluable patients, and, by 4 weeks, all of 17 evaluable patients showed donor chimerism ≥95% on STR-PCR, which was maintained until 24 weeks in all 11 patients tested. Thirteen patients are alive after median follow-up of 13.6 months (1.5–37.9; Kaplan-Meier survival, 55.6%). Of 16 patients with acute leukemia and high-risk MDS, 8 remain alive without recurrence (event-free-survival, 40.9%). Two patients died of K. pneumoniae sepsis and grade IV acute GVHD, respectively (CI of TRM, 11%). Immune recovery in 10 patients without relapse for > 6 months showed robust lymphocyte contents and immunoglobulin levels at 6 months (means of; 1,060/ul CD3+, 222/ul CD4+, 767/ul CD8+ cells, and 1,317 mg/dl IgG) and 12 months. After RIC, consistent engraftment and durable complete donor hematopoietic chimerism can be achieved from HLA-haploidentical familial donor. The frequencies of GVHD and TRM were low.

Natural Killer Cell Immune Reconstitution Predicts Outcomes for Patients with Chronic Lymphocytic Leukemia Undergoing Allogeneic Stem Cell Transplantation

Blood ◽  
2008 ◽  
Vol 112 (11) ◽  
pp. 3300-3300
Author(s):  
Don Benson ◽  
Leslie Andritsos ◽  
Mehdi Hamadani ◽  
Thomas Lin ◽  
Joseph Flynn ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Nk Cells ◽  
Cd8 T Cells ◽  
Immune Reconstitution ◽  
Nk Cell ◽  
Disease Status ◽  
Lymphocytic Leukemia ◽  
T Cell Subsets ◽  
Cell Recovery

Abstract Introduction: Chronic lymphocytic leukemia (CLL), the most common form of leukemia in the Western hemisphere, is associated with severe innate, adaptive and humoral immune dysregulation. CLL remains essentially incurable, with the potential exception of allogeneic stem cell transplantation (ASCT). Natural killer (NK) cells are CD56(+), CD3(−) large granular lymphocytes that comprise a key cellular subset of the innate immune system. Preliminary in vitro data suggest an NK cell versus CLL effect exists, similar to that observed in acute myeloid leukemia (AML) and other blood cancers. Novel immune therapies for CLL (e.g., rituximab, alemtuzumab) likely exert anti-tumor effect, in part, through NK cells, in fact. Although NK cells contribute to the graft-versus-tumor effect following ASCT for other blood cancers, little is known regarding the potential role NK cells may play in the clinical allogeneic transplant setting for CLL. Herein, we provide, to our knowledge, the first report regarding NK cell immune reconstitution following ASCT for CLL. Methods: 27 CLL patients underwent reduced intensity conditioning (RIC) with ASCT. Median age was 52 years (43–69), median number of prior therapies was 3 (2–11). 55% had chemotherapy-refractory disease, and 55% had “high-risk” cytogenetics by FISH (deletion 17p or 11q22-23 abnormality). 14 patients had sibling donors, 15 had volunteerunrelated donors. Conditioning regimens included Fludarabine/TBI/Alemtuzumab (n=8), Fludarabine/Busulfan with (n=9) or without ATG (n=6), and Fludarabine/Cyclophosphamide (n=4). GVHD prophylaxis consisted of tacrolimus/MMF (n=8) or tacrolimus/methotrexate (n=19). Patients underwent bone marrow assessment prior to day +75 following ASCT. Marrow was studied for engraftment, donor chimerism, and disease status as well as lymphoid immune reconstitution by percentage of total lymphocytes and absolute lymphocyte counts by multi-color flow cytometry. Results: NK cell immune reconstitution was predicted by disease status at transplantation. Patients in complete or partial remission at the time of ASCT had more robust NK cell recovery (mean = 45% of total lymphocytes +/− SEM 5%) as compared to patients entering transplant with refractory disease (16% +/− 1, p < 0.01). No differences were observed in CD4(+) or CD8(+) T cells and no lymphocyte subset recovery was associated with CD34(+) or CD3(+) cell dosage. Achieving complete donor chimerism by day +60 was associated with robust NK cell recovery (55% +/− 1 versus 7% +/−1, p = 0.02), recovery of CD4 and CD8 T cells was not associated with chimerism status, however. Patients who went onto exhibit a complete response to ASCT had greater early NK cell reconstitution (31% +/− 3) as compared to those who had no response (8% +/− 1, p = 0.01). No differences in T cell subsets were associated with response. Patients who ultimately achieved complete remission following transplant had a lower CLL:NK cell ratio in marrow (0.35 +/− 0.07) than those who did not (8.1 +/− 1, p = 0.01). However, differences in CLL:CD4(+) and CLL:CD8(+) T cells were not predictive of response. Trends to improvement in progression free survival and overall survival were observed for patients with NK cell reconstitution above the median for the group as compared to those below; no such trends were observed regarding T cell subsets. Greater NK cell reconstitution trended towards ultimate eradication of minimal residual disease following ASCT, but no such trends were observed for T cell subsets. Conclusions: Early NK cell recovery predicts survival following autologous and allogeneic SCT in a number of hematologic malignancies; however, little is known regarding this phenomenon in CLL. To our knowledge, these are the first findings to implicate a potentially important therapeutic role for early NK cell compartment recovery in CLL following ASCT. Further research into restoring and augmenting NK cell function following RIC/ASCT for CLL is warranted.

scholarly journals Method of Mobilization: Implication on Cell Subsets in the Graft and Immune Reconstitution Post Autologous Hematopoietic Cell Transplantation (AHCT)

Blood ◽  
2014 ◽  
Vol 124 (21) ◽  
pp. 1132-1132
Author(s):  
Melhem M. Solh ◽  
Rathmann Kristin ◽  
Sauvi chang-Fong ◽  
Jeremiah Oyer ◽  
Wesam B. Ahmed ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Cell Transplantation ◽  
Peripheral Blood ◽  
Immune Reconstitution ◽  
Hematopoietic Cell Transplantation ◽  
Hematopoietic Cell ◽  
P Value ◽  
Autologous Hematopoietic Cell Transplantation ◽  
Graft Composition

Abstract Method of Mobilization: Implication on Cell Subsets in The Graft and Immune Reconstitution post Autologous Hematopoietic Cell Transplantation (AHCT) The optimal mobilization method for either myeloma or lymphoma patients undergoing AHCT is still debatable and strategies for graft collection vary between different institutions. Plerixafor, a CXCR4 antagonist is used for peripheral blood stem cell mobilization in multiple myeloma and non-Hodgkins lymphoma patients requiring AHCT. The effect of plerixafor on graft composition has scarce data that are based mostly on cryopreserved samples. Moreover; the effect of plerixafor on immune reconstitution and hematologic recovery post AHCT has not been well evaluated. The goal of our study was to compare graft composition, hematologic and immune reconstitution recovery among patients mobilized with plerixafor plus G-CSF to those mobilized with G-CSF alone. Methods: 49 patients eligible for AHCT were enrolled on a single arm prospective trial at a single transplant center. All patients were mobilized with G-CSF 10µg/kg/day for 4 consecutive days. A peripheral blood CD34 level of <20/µl on day 4 was used as a cutoff to use plerixafor 0.24mg/kg in addition to G-CSf on 9pm of the fourth day. Peripheral blood collection was started on day 5 and was continued till the target dose is achieved or a minimum CD 34+ cell dose of >2x106 cells/Kg was obtained after 3 collection days. Samples from the freshly collected graft and patients' peripheral blood on days +30 and +60 were analyzed by flow cytometry (BD FACSCanto II) . A single platform assay was used (Beckman-Coulter Stem kit) via a ISHAGE protocol. The antibody cocktail contained the following pre-conjugated monoclonal antibodies: CD56-PE (Miltenyi Biotech, Auburn, CA), CD3-APC, CD16-FITC, (Beckman Coulter, Brea, CA), CD19-PE-CY7 (BD Biosciences, San Jose, CA). Data were acquired using BD FACSCanto II (BD Biosciences) and analyzed with the FACSDiva software (BD Biosciences) to quantify CD3+ T cells, CD3+ CD56+ NK-like T cells, CD56+ CD16+ and CD56+ CD16- NK cells as well as CD19+ B cells. Results: 49 patients with a median age of 58 years (range 21-75) were mobilized with either G-CSF alone (N=16) or plerixafor +G-CSF (G+P)(N=33).The median number of collection days was 1.42 and 1.81 (p=0.2) and the median collected CD34+ dose was 8.28x106/kg and 5.24x106 /kg (p=022) in the G+P and G-CSF alone groups respectively. Both groups had similar times to neutrophil and platelet engraftment. The graft analysis showed a white blood count of 309x109/l and 262x109/l (p=0.38), median percentage of CD34+ cells of 0.75% and 0.73% (p=0.81), percentage of CD3+ T cells of 25.6% and 22% (p=0.6) in the G+P and G-CSF alone groups resepectively. Both groups had similar proportions of CD3+, CD4+,CD8+, NK, NKT and iNKT cells in the mobilized grafts. Peripheral blood samples at day +30 and day +60 were analyzed for T cell markers and hematologic recovery (table 1). There was no significant difference between absolute lymphocyte counts, NK cell counts, T cells and absolute neutrophil count. Conclusion: Plerixafor when combined with G-CSF helps in achieving mobilization goals in patients predicted to be poor mobilizers based on peripheral CD34 levels. The addition of plerixafor doesn't not seem to affect T cell composition of the graft and yields similar hematologic and immune recovery when compared to mobilization with G-CSF alone. Table 1: Immune Reconstitution at Day 30 and Day 60 post Autologous Transplantation Treatment Group G-CSF (N=16) Plerixafor + G-CSF (N=33) P-value G-CSF (N=16) Plerixafor + G-CSF (N=33) P-value Day 30 Day 60 WBC 5.08 5.41 0.873 4.94 5.38 0.654 HGB 10.86 11.19 0.353 11.22 11.17 0.757 HCT 32.35 33.66 0.321 33.36 33.53 0.565 PLT 119.88 161.42 0.068 166.94 173.73 0.949 Abs Lymph 1.09 1.44 0.296 1.41 1.50 0.974 % NK 26.14 30.38 0.277 11.53 20.09 0.095 Abs NK 0.31 0.35 0.186 0.17 0.21 0.470 % T cell 67 60 0.183 76.15 67.39 0.340 Abs T cell 0.72 0.96 0.717 1.35 .82 0.095 NKT%* 5.28 3.33 8.25 3.38 B cell % 2.38 1.52 0.922 2.63 5.58 0.424 Abs. Neut count 2.99 2.64 0.488 2.85 3.01 0.848 Disclosures No relevant conflicts of interest to declare.

NK-Cell Recovery and Immune Reconstitution after Haploidentical Hematopoietic Cell Transplantation Using Either CD34 Selected Grafts and Adoptive NK-Cell Transfer or CD3/CD19 Depleted Grafts: Comparison of Two Strategies for NK Cell Based Immunotherapy.

Blood ◽  
2007 ◽  
Vol 110 (11) ◽  
pp. 2988-2988 ◽  
Author(s):  
Chiara Gentilini ◽  
Matthias Haegele ◽  
Arne Muessig ◽  
Axel Nogai ◽  
Constanze Kliem ◽  
...  
Keyword(s):  
T Cells ◽  
Nk Cells ◽  
Hematopoietic Cell Transplantation ◽  
Nk Cell ◽  
Group Versus ◽  
Cell Group ◽  
Immune Recovery ◽  
Cell Recovery ◽  
Cd34 Cells ◽  
Haploidentical Hematopoietic Cell Transplantation

Abstract NK-cells have been shown to play a pivotal role in haploidentical hematopoietic cell transplantation (HHCT) for engraftment, GvL effects and to combat infectious complications. Different strategies have been employed to hasten NK-cell recovery after HHCT. Here we compare the immune recovery of 17 patients after CD34 selected HHCT receiving additional adoptive CD3-depleted CD56-enriched NK cells 2 days after HHCT (adoptive NK-cells), with 18 patients receiving CD3/CD19 depleted grafts (CD3/CD19) for HHCT. Transplantations were performed at two different institutions with a median follow-up of >1 year. Conditioning consisted of 12 Gy TBI, thiotepa (10mg/kg), fludarabine (150 mg/m2) and OKT3 (day −4 to +2) in the group receiving CD34 selected grafts and adoptive NK-cell transfusions. All patients in the CD3/CD19 group received conditioning with fludarabine (150–200 mg/m2), thiotepa (10 mg/kg), melphalan (120 mg/m2) and OKT-3 (day −5 to +14). No postgrafting immunosuppression was used in both groups. Seven out of the 17 patients in the adoptive NK-cell group received IL-2 activated NK cells. Median age was 37 years in the adoptive NK-cell group compared to 40 years in the CD3/CD19 group. Diagnoses in the adoptive NK-cell group included AML (n=10), ALL (n=3), CML (n=2), and Hodgkin’s disease (n=1) and MDS (n=1). Diagnoses in the CD3/CD19 group were AML (n=10), ALL (n=5), NHL (n=1), CML (n=1) and multiple myeloma (n=1). The grafts contained a median of 12.5x10E6 CD34+ cells/kg and 1.1×10E4 CD3+ cells/kg in the CD34 selected group versus 9.2×10E6 CD34+cells/kg and 2.3×10E4 CD3+cells/kg in the CD3/CD19 group. The number of transferred CD56+ cells was 8.3×10E6/kg in the adoptive NK-cell group and 7.2×10E7/kg cells in the CD3/CD19 group. Hematopoietic recovery was similar in both groups. Among the patients receiving adoptive NK-cells we observed a striking difference in immune recovery between the patients receiving IL2-activated and those treated with non-activated NK cells: patients receiving activated NK cells showed significantly lower numbers of NK- and T cells during the first months post transplant (p=<0.05). In addition, we compared the immune recovery of the patients in the CD3/CD19 group and the 10 patients in the adoptive NK cells group receiving unstimulated NK-cells. There was a significant faster recovery of CD4+ T cells in the adoptive NK-cell group with a median day 40 count of 179 versus 2 cells/μl (p<0.05). However, patients in the CD3/CD19 group showed a significant faster and more sustained recovery of NK-cells with a median day 40 CD56+ count of 1464 versus 254 cells/μL (p<0.05). After day 50 no significant difference between the two groups was observed. The incidence of GvHD≥II was similar with 47% in the adoptive NK-cell group versus 50% in the CD3/CD19 group. In conclusion, patients receiving CD3/19 depleted grafts show a faster and more sustained NK-cell reconstitution but a slower T cell recovery in the early phase after transplant. The clinical impact of these differences warrants evaluation in further prospective studies.

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