Post Transplantation Fludarabine and Cyclophosphamide Selected and Promoted Low Dose of Unrelated UCB Implanation in Combined Transplantation of Haploid and UCB Stem Cells in Childhood Leukemia: 40 Cases Report in Double Center

Abstract Background: For patients without HLA matched donor, Haploid stem cell transplantation with PTCY method has relatively higher risk of relapse and graft-versus-host disease (GVHD). Umbilical cord blood (UCB) is readily available and has helped expand the donor pool to almost all patients requiring a transplant. Meanwhile UCB transplant improves EFS in leukaemia with lower risk of relapse and GVHD. However, the clinical application of UCB is limited due to lower implantation rate because of its low dose of stem cells than other sources of hematopoietic stem cells (HSC). Aims: This study is to explore the feasibility and efficacy of post-transplantation fludarabine and cyclophosphamide to select and promote the unrelated UCB to engraft in combined transplantation of haploid and UCB stem cells for the treatment of childhood and adolescent leukemia. Methods: Total 40 children and adolescent patients with leukemia in Nanfang Hospital and Shenzhen Children's Hospital enrolled this study from Sept.2019 to Jun.2021. These patients were diagnosed as AML (20 HR, 5 IR, 2 relapsed AML), ALL (5 HR, 2 IR, 2 relapsed ALL), AL (1), JMML (2), MDS (1) and BPDCN (1) with a median age of 80 months (range 7 months -17 years). The biggest body weight reached to 77kg. The haploid stem cell was 5/10-9/10 mismatched and UCB was selected as 6/10-10/10 HLA matched or mismatched. The condition regimen was busulfan+fludarabine+CY+Ara-C. Haploid PBSC was infused in day0. All received PTCy 50 mg/kg and post-transplantation fludarabine 40mg/m 2 on days 3 and 4 along with tacrolimus or cyclosporine and mycophenolate mofetil for prophylaxis of acute GVHD. UCB was infused in day6. A median of haploid stem cells of 20.00×10 8/kg (13.00-32.10) of mononuclear cells was infused while a median of UCB cells of 4.32×10 7/kg (1.48-22.78) of total nucleated cells was infused. A median of CD34+ cells of haploid stem cells 13.00×10 6/kg (1.51-32.00) was infused while a median of CD34+ cells of UCB cells 1.74×10 5/kg (0.26-4.80) was infused. The survival rate, umbilical cord blood implantation rate, hematopoietic recovery and the rate of transplant-related complications were analyzed. Results: At a median follow-up of 8 months (range 1-21 months), there were 2/40（5%）cases of transplant-related death. All surviving patients were leukemia free, with one-year overall survival rate 92.8±5.0%. Among these patients, 37/40(92.5%, 95%CI: 84.0%~100.0%) of patients achieved complete chimerism of unrelated UCB cells and 3/40(7.5%, 95%CI: -1.0%~16.0%) of patients achieved mixed chimerism of unrelated UCB cells and haploid cells. In these transpltantation, the CD34+ cell dose of UCB less than 1.0×10 5/kg accounted for 10/40 (25.0%), and less than 2.0×10 5/kg accounted for 23/40 (57.5%).Two patients had primary poor graft function. Neutrophil reconstitution was achieved in 39/40 patients with a median time of 29 days (range 17 - 44 days) without G-CSF after transplantation. Platelet recovery was achieved in 37/40 patients with a median time of 37 days (range 8-92 days). There was a significant linear relationship between platelet recovery time and the dose of total nucleated cells and CD34+ cells in UCB（r=-0.368, P=0.025; r=-0.355, P=0.031）.The incidence of gradeⅠand gradeⅡGVHD was 32.5%（95%CI:17.3%-47.7%）and 42.5%（95%CI:26.5%-58.5%）, respectively. There was no grade Ⅲ-Ⅳ aGVHD and only 1/40(2.5%) case of extensive chronic GVHD. The incidence of chronic limited GVHD was 22.5% (95%CI: 9.0%-36.0%). Twenty-four of 40(60.0%, 95%CI:44.1%-75.9%) patients experienced clinically significant CMV reactivations or infections. One of 40（2.5%, 95%CI: -2.6%-7.6%）patients experienced EB virus reactivation. Two of 40（5.0%, 95%CI: -2.1%-12.1%）patients experienced human herpesvirus 6 infection. Thirteen of 40（32.5%, 95%CI: 17.3%-47.7%）patients presented with hemorrhagic cystitis. Conclusion: In our primary clinical study, post-transplantation fludarabine and cyclophosphamide could effectively select and promote the unrelated UCB to implant rather than haploid cells in combined transplantation of haploid and UCB stem cells even if the CD34+ cells of UCB less than 1.0×10 5/kg. Although acute GVHD was common but just milder degree and with lower incidence of EB virus reactivation. This new strategy has the potential to promote the wilder clinical use of unrelated UCB in the treatment of leukemia. Figure 1 Figure 1. Disclosures No relevant conflicts of interest to declare.

Download Full-text

Radiation Dose Determines Degree of Donor Chimerism after Nonmyeloablative Hematopoietic Cell Transplantation.

Blood ◽

10.1182/blood.v104.11.1828.1828 ◽

2004 ◽

Vol 104 (11) ◽

pp. 1828-1828 ◽

Cited By ~ 2

Author(s):

Christoph Kahl ◽

Marco Mielcarek ◽

Mineo Iwata ◽

Michael Harkey ◽

Barry Storer ◽

...

Keyword(s):

Stem Cells ◽

Stem Cell ◽

Radiation Dose ◽

Low Dose ◽

Donor Chimerism ◽

Stem Cell Source ◽

Cd34 Cells ◽

Cell Source ◽

Total Body

Abstract Efforts to replace total body irradiation (TBI) used for transplant conditioning with agents that have less acute and long-term toxicities require a better understanding of the biological effects of low dose TBI. We therefore retrospectively analyzed the role of radiation dose, stem cell source, and type of immunosuppression on both the stability and degree of donor chimerism in canine recipients of matched littermate hematopoietic cell transplants. Recipients were prepared with 200 cGy (n=26), 100 cGy (n=76) or 50 cGy (n=19) total body irradiation (TBI) at 7 cGy/min. Stem cell sources included bone marrow (BM) alone (n=58), BM plus G-CSF mobilized peripheral blood mononuclear cells (G-PBMC) (n=42), BM and CD14-depleted G-PBMC (n=13), or BM and T-cell-depleted G-PBMC (n=8). Posttransplant immunosuppression consisted of cyclosporin (CSP) only (n=53), CSP plus mycophenolate mofetil (MMF) (n=23), CSP and rapamycin (n=12), CSP, MMF and rapamycin (n=5); or CSP and MMF in combination with pretransplant immunosuppression (n=28). The percentage of donor granulocytes in the peripheral blood, as determined by PCR amplification of variable numbers of tandem repeats (VNTR), served as a marker for engraftment. TBI dose and stem cell source were both significantly associated with long-term (>26 weeks) stable engraftment in multivariate analysis (p=0.0001 and p=0.004, respectively). Among the 39 dogs with stable engraftment, however, TBI dose was the only factor examined that was associated with the degree of donor chimerism (mean % of donor granulocytes after 200 cGy, 100 cGy and 50 cGy of TBI: 65%, 52%, and 24%, respectively; p=0.008). To determine whether low-dose irradiation directly affected recipient stem/progenitor cell numbers and thereby conferred a competitive disadvantage to donor cells, CD34+ cells were isolated from two normal human donors. One preparation of CD34 cells was ex vivo irradiated (=200 cGy) and then injected into NOD/SCID beta2m-/- mice in combination with an equal number of unirradiated CD34 cells from the second donor. The contributions of each donor to human engraftment were assessed at 10 weeks by VNTR. After 200 cGy, the irradiated population contributed 74% less than expected, 24% less after 100 cGy, but only 6% less after 50 cGy. Flow analysis of Caspase-3 activation indicated that a significant percentage of cells irradiated with 200 cGy were apoptotic, and that this was associated with the loss of L-selectin and P-selectin glycoprotein ligand-1. In conclusion, our findings suggest that TBI, in addition to its well-characterized immunosuppressive effects, determines the degree of donor cell engraftment by directly compromising recipient stem cells, thereby providing a competitive advantage to donor stem cells.

Download Full-text

Delayed Platelet Recovery Following Cord Blood Transplantation: Insights from a Mouse Model.

Blood ◽

10.1182/blood.v106.11.1727.1727 ◽

2005 ◽

Vol 106 (11) ◽

pp. 1727-1727

Author(s):

Tao Du ◽

Camelia Iancu-Rubin ◽

George F. Atweh ◽

Rona Singer Weinberg

Keyword(s):

Stem Cells ◽

Stem Cell ◽

Hematopoietic Stem Cells ◽

Cord Blood ◽

Lower Number ◽

Post Transplantation ◽

Hematopoietic Stem ◽

Platelet Recovery ◽

Platelet Counts ◽

Polyploid Cells

Abstract Umbilical cord blood (CB) is an important source of hematopoietic stem cells for stem cell transplantation and is being used with increasing frequency. A major concern related to clinical CB transplantation is the long delay in platelet recovery. Megakaryopoiesis is characterized by the acquisition of lineage-specific markers (e.g. CD41) during the early stages that is followed by polyploidization (DNA content > 4N) during the later stages of megakaryopoiesis. Using a newborn blood (NB) model of CB transplantation that was previously developed in our laboratory, we asked whether the delay in platelet recovery is a result of a decrease in the rate of megakaryocyte production or a delay in their maturation. C57BL/6 mice were transplanted with either murine adult bone marrow (BM) cells or murine new-born blood (NB) cells following lethal irradiation. We had previously shown that the concentration of Lin-Sca-1+c-Kit+ stem cells in murine adult BM was approximately 3 times higher than that in NB. To correct for this difference in stem cell concentration, irradiated mice were transplanted with either 0.5 ×106 BM cells or 2×106 NB cells. Platelet counts at 2 and 4 weeks were lower in mice transplanted with NB cells than in mice transplanted with BM cells (Table 1). Interestingly, the platelet counts became comparable in NB and BM recipients at 8 weeks post-transplantation. We compared the ability of BM cells from both NB and BM recipients to form CFU-Meg colonies in methylcellulose. At 2 and 4 weeks post-transplantation, BM cultures derived from NB recipients generated fewer CFU-Meg colonies than cultures from BM recipients (Table 1). Interestingly, after 8 weeks, the numbers of CFU-Meg from both stem cell sources were similar. We also compared the ability of BM cells from NB and BM recipients to differentiate into megakaryocytes in liquid culture, using CD41 expression and polyploidy as markers of megakaryocytic maturation. At 2 weeks post-transplantation, cultures from NB recipients generated 13% CD41+ cells whereas cultures from BM recipients generated 22% CD41+ cells. However, by 4 weeks post-transplantation, the numbers of CD41+ cells were similar in cultures derived from NB recipients and BM recipients (Table 1). Moreover, at 2 and 4 weeks post-transplantation, there were fewer polyploid cells in liquid cultures from NB recipients compared to BM recipients (Table 1). The lower number of polyploid cells was commensurate with the lower number of CD41+ cells. This suggests that the rate of maturation of megakaryocyte is similar in NB and BM. In conclusion, our studies show that CB stem cells generate megakaryocytic progenitors at a slower rate than BM stem cells and that the delay in platelet recovery is not a result of a delay in the maturation of megakaryocytic progenitors. Thus, in order to increase the rate of platelet recovery following CB transplantation, the focus should be on enhancing the rate of production of megakaryocytic progenitors from hematopoietic stem cells. Table 1. Platelet(×103/μl) CFU-Meg(Colonies/1 × 105 Cells) CD41(%) Ploidy(% >4N DNAContent) normal BM(n=5) 1200±120 35.5±5 55±5 27±3 2 weeks NBT-2M(n=5) 190± 38 10±2 13±2 7.5±1 BMT-0.5M(n=5) 400±50 17±4 22±1 11±2 4 weeks NBT-2M(n=5) 550±59 18±2 26±2 14.5±2 BMT-0.5M(n=5) 730±110 23±3 27±1 18±3 8 weeks NBT-2M(n=5) 930±115 39±7 N/A N/A BMT-0.5M(n=5) 970±130 40±4 N/A N/A

Download Full-text

Rapid Mobilization Of CD34+ Progenitor Cells With TG0054-03, a novel CXC Chemokine Receptor 4 (CXCR4) Antagonoist

Blood ◽

10.1182/blood.v122.21.905.905 ◽

2013 ◽

Vol 122 (21) ◽

pp. 905-905 ◽

Cited By ~ 4

Author(s):

Michael W. Schuster ◽

Nabil Hagog ◽

Bita Jalilizeinali ◽

Sharon Funkhauser ◽

Mary Sophy Yohannan ◽

...

Keyword(s):

Stem Cells ◽

Stem Cell ◽

Chemokine Receptor ◽

Peripheral Blood ◽

Mononuclear Cells ◽

Single Agent ◽

Platelet Recovery ◽

Cd34 Cells ◽

Stem Cell Collection ◽

Historical Controls

Abstract Background TG-0054 (burixafor) is a potent and specific antagonist of the human CXCR4 chemokine receptor. TG-0054 blocks the interaction between CXCR4 and stromal cell-derived factor-1 (SDF-1), thus causing a rapid mobilization of stem cells from the bone marrow into peripheral blood within 1-3 hours of intravenous administration of the drug. Materials and Methods: An early phase II trial was conducted in patients with multiple myeloma (MM), non-Hodgkin lymphoma (NHL) or Hodgkin disease (HD) to evaluate the safety and stem cell mobilization of TG-0054 alone or in combination with G-CSF. 12 patients (1 HL, 7 MM, and 4 NHL patients) received an i.v. dose of 3.14 mg/kg TG-0054, and peripheral blood CD34 counts were assessed at 2, 4 and 6 hours post drug infusion. Patients who achieved at least 10 CD34 cell/ul in the peripheral blood were collected by large volume leukapheresis (24L) for 1-4 days to obtain a predetermined target of >2.5 x 106 CD34 cells /kg. Results: Seven patients (1 HD, 6 MM) were successfully mobilized with TG-0054 as a single agent achieving a cumulative CD34+ stem cell collection of 4.0 to 10.4 x106 cells /kg over 2-4 leukapheresis sessions. Patients who failed to mobilize with TG0054 as a single agent on day +1 were placed on the second arm of the study, where they received 5 doses of granulocyte colony stimulating factor (G-CSF) at a dose of 10 ug/kg beginning on day +4 with TG-0054added back in combination on day +8. These remaining five patients (1 MM, 4 NHL) who did not mobilize with TG0054 alone on day +1 and received G-CSF plus TG-0054 were leukapheresed on day +8. Those patients achieved a cumulative CD34 peripheral blood stem cell collection of of 3.2-21.0 x106 cells /kg over 1-4 leukaphereses sessions. Conclusion: TG-0054 exhibited potent and rapid mobilization of CD34+ stem cells, with favorable safety profile in patients. Engraftment All 12 patients received conditioning regimens (BEAM for the lymphoma patients and melphalan for the myeloma patients) followed by a stem cell infusion of at least 3.0 x106 CD34+ cells/kg. All patients engrafted without delay compared to historical controls. Median days to WBC engraftment were 12. Median days to platelet recovery of 20,000 and 50,000 were 20 and 20.5 days, respectively. Engraftment results of TG-0054 mobilized patients are similar to those seen in a matched group of historical controls. We have observed that mobilization with TG-0054 caused a preferential mobilization of mononuclear cells (MNC) component of the graft. Percent MNC in TG-0054 mobilized patients‘ grafts were 78.9+15.2 (median 81.5) as compared to percent of MNC in G-CSF mobilized patients‘ grafts of 62.6+27.7 (median 69.6). Disclosures: Schuster: TaiGen Biotechnology Co., Ltd: Research Funding. Tsai:TaiGen Biotechnology Co., Ltd: Employment. Hsu:TaiGen Biotechnology Co., Ltd: Employment, Equity Ownership. Chang:TaiGen Biotechnology Co., Ltd: Employment. Hsu:TaiGen Biotechnology Co., Ltd: Employment.

Download Full-text

Risk Factors of BK Virus Cystitis in Haematopoietic Stem Cell Transplantation-a Retrospective Monocentric Study

Blood ◽

10.1182/blood-2018-99-115321 ◽

2018 ◽

Vol 132 (Supplement 1) ◽

pp. 3361-3361

Author(s):

Eléonore Kaphan ◽

Raphaele Germi ◽

Martin Carre ◽

Claude-Eric Bulabois ◽

Sebastien Bailly ◽

...

Keyword(s):

Risk Factors ◽

Stem Cells ◽

Renal Failure ◽

Acute Renal Failure ◽

Stem Cell ◽

Acute Gvhd ◽

Clinical Symptoms ◽

Bk Virus ◽

Post Transplantation ◽

Peripheral Stem Cells

Abstract Background: BK virus (BKV) is a human polyomavirus. Reactivation occurs during deep immunosuppression as in hematopoietic stem cell transplantation (HSCT) and renal transplantation, leading to hemorrhagic cystitis (HC) and nephropathy respectively. In HSCT, systematic PCR for BKV in urine is positive for 50 to 100% of patients (pts), but only 5 to 40% develop a BKV HC. Thus, BKV PCR is usefull to confirm a diagnostic of BKV HC but not to predict its occurrence. Several risk factors to develop BKV HC have been studied, especially mismatched HLA and haploidentical HSCT. Objectives: The aim of this retrospective study was to ascertain the risk factors to develop BKV HC. Methods: A retrospective study was performed by considering data from Grenoble University Hospital in the national retrospective register ProMISe, from the SFGM-TC. The period of the study covered from January 2014 to January 2018. PCR BKV in urine was performed when pts presented hematuria grade 2 or higher with clinical symptoms of cystitis. Viral nucleic acid was extracted from the urine samples with the EasyMag platform (Biomérieux) and the qPCR with BK Virus R-GENE®kit (ARGENE) on a LightCycler 480 (ROCHE). BKV HC is defined by the association of clinical symptoms of cystitis, haematuria grade 2 or higher and a BKV viruria >7 log10 copies/mL. Univariable and multivariable logistic regression model were used to identify risk factors for BK cystitis. Results: 188 HSCT were performed during the study period. After exclusion of 13 pts for early mortality (<30 days) and 4 for engraftment failure, 171 pts were finally considered for analysis, from whom 43 (25.1%) developed a BKV HC. The median age of patients presenting with BKV HC was 44 years (23-62) and males represented 67.4 %. Acute leukemia was the most common indication of HSCT (74.4%), followed by myelodysplastic syndromes (9.3%), lymphoma (6.9%), myeloproliferative neoplasms (4.7%) and aplastic anemia (4.7%). In most cases, pts were not in complete response at transplant (51.2%). First autologous or allogenic HSCT had previously been performed for 30.2% of pts. The majority of pts had a transplant with peripheral stem cells as graft source (76.6%), and had a transplant with mismatched HLA (9/10, n=9, 20.9%) or haploidentical donors (n=13, 30.2%). Twenty-nine pts (67.4%) received reduced-intensity conditioning and twenty-two pts (51.2%) received cyclophosphamide post allograft to prevent Graft Versus Host Disease (GVHD). BKV HC prophylaxis relied on hyperhydratation and mesna during the conditioning regimen. The median time to develop HC was 42 days post-transplantation (30-55) mainly with a grade 3 HC (53.5%). The median viruria was 9 log (9-10). Cidofovir was administered as curative treatment to 20 pts (46.5%) and 25 pts (58%) needed bladder irrigation and forced diuresis. The median level of platelets at diagnosis was 58 G/L (29-123). At diagnosis of BKV HC, 32.6% of pts presented a bacterial cystitis and 62.8% an acute renal failure. Allogenic HSCT was complicated by an acute GVHD in 88.4% of pts and 69.8% were treated by corticosteroids. CMV reactivation was observed in 39.5% of pts, and HHV6 in 18.6%. In univariate analysis, post-transplant cyclophosphamide (p<0.001), age below 40 years (p<0.001), history of previous auto or allograft (p=0.007), allograft with mismatched HLA (9/10 and haploidentical) (p<0.001), use of peripheral stem cells (p=0.047), engraftment of platelets >100 days (p=0.016), acute GVHD (p=0.007), corticotherapy (p<0.001), co-infection by HHV6 (p=0.006), association to bacterial cystitis (p=0.002), acute renal failure (p=0.009) and platelets below 50G/L (p<0.001) were significantly associated with increased risk of BKV HC. After logistic regression, the risk factors associated with BKV HC were reduced to: exposition post-transplantation to cyclophosphamide (OR 4.1, 1.5-10.7, p=0.004), age below 40 years (OR 4.1, 1.6-10.9, p=0.004), corticosteroids therapy (OR 3.9, 1.6-9.5, p=0.033), acute renal failure (OR 3.8, 1.5-9.6, p=0.0056), bacterial cystitis (OR 3.3, 1.2-8.7, p=0.0175), and platelets below 50G/L (OR 3.8, 1.382-10.486, p=0.097). Conclusion: BKV HC was observed in 25.1% of patients. Exposition to cyclophosphamide, young age, corticosteroids therapy and bacterial cystitis are potential risk factors of BKV HC. Surprisingly, young age was not expected as risk factor. Disclosures No relevant conflicts of interest to declare.

Download Full-text

Mesenchymal Stem Cells Promote the Engraftment of Cord Blood CD34+ Cells but Do Not Accelarate Platelet Recovery NOD/SCID Mice.

Blood ◽

10.1182/blood.v106.11.1267.1267 ◽

2005 ◽

Vol 106 (11) ◽

pp. 1267-1267

Author(s):

Yvette van Hensbergen ◽

Helen de Boer ◽

Manon C. Slot ◽

Laurus F. Schipper ◽

Anneke Brand ◽

...

Keyword(s):

Stem Cells ◽

Mesenchymal Stem Cells ◽

Cord Blood ◽

Ex Vivo ◽

Human Platelets ◽

Scid Mice ◽

Post Transplantation ◽

Platelet Recovery ◽

Cd34 Cells ◽

Cord Blood Cd34

Abstract Aim: Delayed platelet reconstitution in the peripheral blood (PB) remains a problem in transplantation with umbilical cord blood (CB)-derived stem cells. Previously, we have shown that transplantation with ex-vivo expanded CB CD34+ cells (CD34exp) with thrombopoietin for 10 days, results in an accelerated platelet reconstitution in NOD/SCID mice. It has been shown that mesenchymal stem cells (MSC) are able to enhance the overall engraftment when co-transplanted with CB CD34+ cells. Therefore, we investigated whether co-transplantation of MSC with CD34+ cells or CD34exp cells may have an additive effect in shortening the time to platelet recovery and on the total number of platelets in the PB at 6 weeks after transplantation. Methods: To evaluate the time to platelet recovery and the total number of platelets at 6 weeks after transplantation, we used 4 groups of irradiated NOD/SCID mice, divided according to the transplant received: 1) CD34+ 2) MSC+CD34+ 3) CD34exp 4) MSC+CD34exp. Human platelet recovery was measured twice a week for the first three weeks and once a week thereafter, using an assay that reliably detects 1x106plt/L. The percentage of human CD45+ cells in the bone marrow (BM) was evaluated at 6 weeks after transplantation. Results: In accordance with previous experiments, platelet recovery started earlier in mice transplanted with CD34exp cells compared to CD34+ cells (Table 1). Co-transplantation of MSC with CD34+ cells did not result in an accelerated platelet recovery during the first 2 weeks after transplantation, as was observed for expanded cells. However, co-transplantation of MSC did enhance the number of platelets at 6 weeks after transplantation (454.2±264.5 plt/μ l for MSC+CD34+ vs. 101.9±78.4 plt/μ l for CD34+). MSC had no affect on either the time to platelet recovery nor the total number of human platelets at 6 weeks after transplantation when co-transplanted with CD34exp cells. To assess the overall efficacy of the MSC on the engraftment of human CB cells, we evaluated the percentage of human CD45+ cells in the BM of the NOD/SCID mice at 6 weeks after transplantation. In mice transplanted with MSC+CD34+, the percentage of human CD45+ cells was higher compared to controls transplanted with CD34+ cells only (30.4% for MSC+CD34+ vs. 17.8% for CD34+). No further engraftment enhancing effect of MSC was observed following transplantation of CD34exp cells only (32.1% for CD34exp vs. 35.7% for MSC+CD34exp). Conclusion: Our results show that transplantation with CD34exp cells results in an accelerated platelet recovery in NOD/SCID mice, an effect that can not be achieved by co-transplantation of MSC+CD34+ cells. However, at 6 weeks after transplantation co-transplantation with MSC+CD34+ cells results in a higher number of platelets in the PB. In addition, the level of engraftment of human CD45+ cells in the BM of NOD/SCID mice is increased by co-transplantation of MSC+CD34+ cells. In contrast, MSC did not affect the time to platelet recovery, the number of human platelets at 6 weeks after transplantation, or the engraftment of human CD45+ cells in the BM when co-transplanted with CD34exp. Table 1: % of mice with ≥ 1x106 platelets/L in the PB Days post transplantation 6 9 13 16 CD34+ 0% 20% 67% 100% MSC+CD34+ 20% 0% 80% 100% CD34exp 83% 100% 100% 100% MSC+CD34exp 60% 100% 100% 100%

Download Full-text

The Use Of Tevagrastim (Biosimilar Filgrastim XMO2) For Hematopoietic Stem Cell Mobilization In HLA Matched Sibling Donors For Allogeneic Stem Cell Transplantation To AML/MDS Patients

Blood ◽

10.1182/blood.v122.21.3275.3275 ◽

2013 ◽

Vol 122 (21) ◽

pp. 3275-3275 ◽

Cited By ~ 1

Author(s):

Ivetta Danylesko ◽

Rina Sareli ◽

Nira Bloom-Varda ◽

Ronit Yerushalmi ◽

Noga Shem-Tov ◽

...

Keyword(s):

Stem Cells ◽

Stem Cell ◽

Side Effects ◽

Conventional Therapy ◽

Post Transplantation ◽

Hematopoietic Stem ◽

Cd34 Cells ◽

Grade Ii ◽

Historical Controls ◽

Biosimilar Filgrastim

Abstract Introduction Human recombinant G-CSF Filgrastim (Neupogen) has been widely used for the mobilization of CD34+ hematopoietic stem cells (HSC) from healthy donors. The experience with biosimilar G-CSF agents is limited and the only publication, to our knowledge, is the recent study by Schmitt M and co-authors (BMT 2013; 48, 922–925). This study involved 11donors and pts with various disease categories. The Biosimilar G-CSF was found to be comparable in efficacy and safety to Neupogen. Developing new cost effective mobilizing reagents if important. We, therefore, initiated a prospective study assessing Tevagrastim (biosimilar Filgrastim XMO2) for mobilization of CD34+ PB HSC in healthy sibling donors (MRD) for transplantation in pts with AML/MDS (NCT01542944). Materials and methods 24 pts with AML or high-risk MDS undergoing allo-SCT from MRD were investigated. The study was approved by the National Regulatory Authorities and both patients and donors signed an informed consent. The donors, median age 46 years (range, 25–64), F- 14; M- 10 received Tevagrastim in a standard dose of 10 μg/kg BW s.c. BID for 4 days. On the morning of the 5th day they underwent conventional leukapheresis. The target yields of CD 34 cell was 5 × 106 CD34+ cells/kg BW of the recipient. If one leukapheresis was insufficient a second was performed and, the last dose of Tevagrastim was administered on the evening of the 5th day. The conditioning was myeloablative Bu/Cy (n=10), reduced toxicity Flu/Treo (n=7), Flu/Bu4 (n=3) or RIC Flu/Bu2 (n=4). The study parameters were: a CD34+ cell count,both absolute numbers and the CD34+ cells per kg BW of the recipient, the number of leukapheresis procedures, the number of CD3+ T lymphocytes in the graft, post transplantation engraftment including the WBC, the neutrophils and the platelets, as well as side effects. Follow up was 100 days post allo-SCT. Results Efficacy 77-1982 × 106 (median 749 × 106) CD34+ were collected. The number of CD34+ cell per kg BW of the pts was 0.93-35.4 × 106 (median 10.2 × 106). Collections contained 144-709 × 108 (median 299 × 108) CD3+ T-cells, 1.74-11.6 ×108 (median 4.4 ×108) per kg BW of the pts. The mean number of leukapheresis procedures was 1.3. Engraftment was: ANC >0.5× 109/L and >1× 109/L within a median of 13 days (range, 10–21) and 13.5 days (range, 10–22), respectively. PLT reached counts of >20× 109/L and >50× 109/L within a median of 16 days (range, 12–33) and 17 days (range, 12–33) from allo-SCT, respectively. The median days of isolation was 10 (range, 6-21). As for blood support, the median number of PC and PLT transfusions was 5 (range, 2-20) and 21 (range, 0-180), respectively. 18/22 (81.8%) pts achieved full donor chimerism at 1 month after transplantation (2 pts are too early to evaluate). Safety Overall Tevagrastim was found to be safe with minimal transient side effects. Neither allergic reactions nor severe adverse events were observed in the donors. 12/24 donors reported transient arthralgias and 2 developed flu-like syndrome while receiving Tevagrastim. As for transplantation related toxicity in the 24 pts transplanted with Tevagrastim mobilized HSC grafts, side effects were not different than those we observed in historical controls. The main side effects were mucositis (n-15, grade II -9, grade III-IV- 6), infection complications (n-20) and fluid retention (n-8). One pt suffered from VOD (Grade-I) that resolved with conventional therapy. Five pts developed acute GVHD (grade II-III) that responded to conventional therapy. In total TRM was 1/24 at d 100. 2 pts died from leukemia progression. Conclusions Our study with 24 AML/MDS pts, indicates that the G-CSF biosimilar XM02, Tevagrastim is safe and efficient for stem cells mobilization in HLA matched normal sibling donors. The CD34 yield and post transplantation engraftment are similar to those achieved with the human recombinant G-CSF Filgrastim (Neupogen). We have not seen significant differences in the graft CD34+ and CD3+ T lymphocytes cell count, the number of leukapheresis procedures and the regeneration of WBC, neutrophils and platelets in comparison with our historical controls. All patients promptly engrafted, and the donors developed only expected side effects like arthralgias and flu-like syndrome. Neither graft rejection nor side effects occurred more frequently than expected from the standard G-CSF. Disclosures: Nagler: Teva : Consultancy, Honoraria, Research Funding.

Download Full-text

Myeloablative Umbilical Cord Blood Transplantation for Hematologic Malignancies Is Comparable to Unrelated Donor Transplantation: A Retrospective Single-Center Study

Blood ◽

10.1182/blood.v120.21.1995.1995 ◽

2012 ◽

Vol 120 (21) ◽

pp. 1995-1995

Author(s):

Filippo Milano ◽

Theodore A. Gooley ◽

Paul O'Donnell ◽

Boglarka Gyurkocza ◽

H. Joachim Deeg ◽

...

Keyword(s):

Stem Cells ◽

Cord Blood ◽

Acute Gvhd ◽

Disease Risk ◽

Unrelated Donor ◽

Platelet Recovery ◽

Risk Of Death ◽

Increased Risk ◽

The Difference ◽

Risk Of Relapse

Abstract Abstract 1995 Background: The number of cord blood transplants (CBT) is rapidly increasing with suggestion of outcomes comparable to those obtained after unrelated donor transplantation (URD). We conducted, a retrospective analysis comparing post-transplant outcomes between myeloablative CBT and myeloablative URD at our Institution. Methods: Between January 2006 and December 2011 a total of 488 patients received either a CBT (n=88) or URD (n=400). Of these 400, 358 (90%) received a 10/10 HLA-matched (MURD) and 42 (10%) a ≤9/10 HLA-mismatched (MMURD) graft. All patients received a double CB graft except for 12 patients (13%) who received a single CB unit. In addition, 25 (28%) patients received an ex vivo expanded graft as part of either a single or double CBT. Mycophenolate mofetil and cyclosporine were used for graft-versus-host disease (GVHD) prophylaxis in all CBT recipients, while FK506 + methotrexate was preferentially used among URD patients (n=339, 84%). Conditioning regimens for both groups are summarized in Table 1. Time-to-event outcomes were compared between groups using Cox regression, and logistic regression was used for acute GvHD. All models were adjusted for age, disease risk and CMV serostatus. Results: Patient characteristics are shown in Table 1. Differences between groups included higher median age in URD recipients and a higher proportion of non-caucasian and CMV seropositivity in CB recipients. Disease risk was similar between the 2 groups. Peripheral blood stem cells (PBSC) was used for the majority of URD grafts (61%). The median time to neutrophil [URD 19 days vs CBT 23 days; hazard ratio (HR) 1.91 (1.46–2.51, p<0.0001)] and platelet recovery [URD 19 days vs CBT 45 days; HR=2.76 (2.05–3.71, p<0.0001)] was significantly shorter for URD recipients. In multivariate analysis, the risk of mortality was similar in URD vs CBT (HR=1.11 (0.71–1.73, p=0.64)). When HLA-match status was considered in the URD group, the risk of death was higher in the MMURD group compared to CBT, although the difference was not statistically significant (HR=1.37 (0.83–2.26, p=0.22)). The risk of relapse was suggestively higher in the URD group overall relative to CBT (HR=1.90 (0.94–3.84, p=0.07)), and this difference was enhanced when HLA matching and source of stem cells in URD were considered. In particular, recipients of unrelated (matched or mismatched) PBSC had a higher risk of relapse relative to CBT (HR=2.33 (1.11–4.91, p=0.03)), as did the MMURD (BM or PBSC) group (HR=2.34 (1.07–5.11, p=0.03)). Furthermore, unrelated recipients of matched or mismatched PBSC each had a higher risk of relapse relative to CBT (matched PBSC: HR=2.44 (1.11–5.38, p=0.03); mismatched PBSC: HR=3.89 (1.63–9.30, p=0.002)). The combined results for mortality and relapse led to an increased risk of relapse-free survival (RFS) failure (earliest of relapse or death) for patients receiving PBSC from a mismatched URD (HR=1.88 (1.08–3.27, p=0.03)); the risk of failure was also increased for PBSC recipients from a matched URD, but the difference was not statistically significant (HR=1.42 (0.87–2.32, p=0.16)). The risk of non-relapse mortality (NRM) was similar between URD and CBT (HR=0.89 (0.52–1.53, p=0.67)), and while there was less chronic GvHD in the URD group, the difference was not statistically significant, and this slight reduction was largely due to the effect in BM recipients (URD BM vs CBT, HR=0.59 (0.35–0.98, p=0.04); URD PBSC vs CBT, HR=1.01 (0.62–1.66, p=0.95)). The risks of grades 2–4 and 3–4 acute GvHD were similar between URD and CBT groups (odds ratio (OR) 1.10 (0.57–2.11, p=0.78)), and (OR=0.70 (0.38–1.31, p=0.26)), respectively. Conclusions: Our data suggest that OS, RFS and NRM after CBT are not inferior to those observed after URD transplantation, and OS and RFS might be higher when the URD group is restricted to recipients of PBSC, particularly those who are mismatched with their unrelated donor. Relapse occurred less frequently for CBT recipients especially when compared to MMURD or URD with PBSC. The retrospective nature of this study and the heterogeneity of the population do not allow us to draw definitive conclusions, however, our results reinforce the need for a randomized study to definitively address these comparisons. Disclosures: No relevant conflicts of interest to declare.

Download Full-text

A Stemness Screen Reveals C3ORF54/INKA1 As a Gate-Keeper of Human Stem Cell Latency

Blood ◽

10.1182/blood-2018-99-115065 ◽

2018 ◽

Vol 132 (Supplement 1) ◽

pp. 325-325

Author(s):

Kerstin B. Kaufmann ◽

Laura Garcia Prat ◽

Shin-Ichiro Takayanagi ◽

Jessica McLeod ◽

Olga I. Gan ◽

...

Keyword(s):

Stem Cells ◽

Stem Cell ◽

Fold Increase ◽

Steady Increase ◽

Post Transplantation ◽

Self Renewal ◽

Hematopoietic Stem ◽

Cd34 Cells

Abstract The controversy generated from recent murine studies as to whether hematopoietic stem cells (HSC) contribute to steady-state hematopoiesis emphasizes how limited our knowledge is of the mechanisms governing HSC self-renewal, activation and latency; a problem most acute in the study of human HSC and leukemia stem cells (LSC). Many hallmark stem cell properties are shared by HSC and LSC and therefore a better understanding of stemness regulation is crucial to improved HSC therapies and leukemia treatments targeting LSC. Our previous work on LSC subsets from >80 AML patient samples revealed that HSC and LSC share a transcriptional network that represent the core elements of stemness (Eppert, Nature Med 2011; Ng, Nature 2016). Hence, to identify the key regulators of LSC/HSC self-renewal and persistence we selected 64 candidate genes based on expression in functionally validated LSC vs. non-LSC fractions and assessed their potential to enhance self-renewal in a competitive in vivo screen. Here, we transduced cord blood CD34+CD38- cells with 64 barcoded lentiviral vectors to assemble 16 pools, each consisting of 8 individual gene-transduced populations, for transplantation into NSG mice. Strikingly, individual overexpression (OE) of 5 high scoring candidates revealed delayed repopulation kinetics of human HSC/progenitor cells (HSPC): gene-marking of human CD45+ and lin-CD34+ cells was reduced relative to input and control at 4w post transplantation, whereas by 20w engraftment of marked cells reached or exceeded input levels. For one of these candidates, C3ORF54/INKA1, we found that OE did not alter lineage composition neither in in vitro nor in vivo assays but increased the proportion of primitive CD34+ cells at 20w in vivo; moreover, secondary transplantation revealed a 4.5-fold increase in HSC frequency. Of note, serial transplantation from earlier time points (2w, 4w) revealed superior engraftment and hence greater self-renewal capacity upon INKA1-OE. Since we observed a 4-fold increase of phenotypic multipotent progenitors (MPP) relative to HSC within the CD34+ compartment (20w) we assessed whether INKA1-OE acts selectively on either cell population. The observation of latency in engraftment was recapitulated with sorted INKA1-OE HSC but not MPP. Likewise, liquid culture of HSPC and CFU-C assays on sorted HSC showed an initial delay in activation and colony formation upon INKA1-OE that was completely restored by extended culture and secondary CFU-C, respectively. INKA1-OE MPP showed a slight increase in total colony count in primary CFU-C and increased CDK6 levels in contrast to reduced CDK6 levels in INKA1-OE HSC emphasizing opposing effects of INKA1 on cell cycle entry and progression in either population. Taken together, this suggests that INKA1-OE preserves self-renewal capacity by retaining HSC preferentially in a latent state, however, upon transition to MPP leads to enhanced activation. Whilst INKA1 has been described as an inhibitor of p21(Cdc42/Rac)-activated kinase 4 (PAK4), no role for PAK4 is described in hematopoiesis. Nonetheless, its regulator Cdc42 is implicated in aging of murine HSPC by affecting H4K16 acetylation (H4K16ac) levels and polarity and has recently been described to regulate AML cell polarity and division symmetry. In our experiments immunostaining of HSPC subsets cultured in vitro and from xenografts indicates that INKA1-OE differentially affects epigenetics of these subsets linking H4K16ac to the regulation of stem cell latency. In AML, transcriptional upregulation of INKA1 in LSC vs. non-LSC fractions and at relapse in paired diagnosis-relapse analysis (Shlush, Nature 2017) implicates INKA1 as a regulator of LSC self-renewal and persistence. Indeed, INKA1-OE in cells derived from a primary human AML sample (8227) with a phenotypic and functional hierarchy (Lechman, Cancer Cell 2016) revealed a strong latency phenotype: In vitro and in vivo we observed label retention along with a steady increase in percentage of CD34+ cells, transient differentiation block, reduced growth rate, G0 accumulation and global reduction of H4K16ac. In summary, our data implicates INKA1 as a gate-keeper of stem cell latency in normal human hematopoiesis and leukemia. Studying the detailed pathways involved will shed light upon the mechanisms involved in HSC activation and latency induction and will help to harness these for novel therapeutic approaches. Disclosures Takayanagi: Kyowa Hakko Kirin Co., Ltd.: Employment.

Download Full-text

Long-Term Cryopreservation Does Not Affect Quality of Peripheral Blood Stem Cell Grafts: A Comparative Study of Native, Short-Term and Long-Term Cryopreserved Haematopoietic Stem Cells

Cell Transplantation ◽

10.1177/09636897211036004 ◽

2021 ◽

Vol 30 ◽

pp. 096368972110360

Author(s):

Daniel Lysak ◽

Michaela Brychtová ◽

Martin Leba ◽

Miroslava Čedíková ◽

Daniel Georgiev ◽

...

Keyword(s):

Stem Cells ◽

Stem Cell ◽

Statistical Significance ◽

Extended Period ◽

High Dose ◽

Atp Production ◽

Cd34 Cells ◽

Negative Effect

Cryopreserved haematopoietic progenitor cells are used to restore autologous haematopoiesis after high dose chemotherapy. Although the cells are routinely stored for a long period, concerns remain about the maximum storage time and the possible negative effect of storage on their potency. We evaluated the effect of cryopreservation on the quality of peripheral stem cell grafts stored for a short (3 months) and a long (10 years) period and we compared it to native products.The viability of CD34+ cells remained unaffected during storage, the apoptotic cells were represented up to 10% and did not differ between groups. The clonogenic activity measured by ATP production has decreased with the length of storage (ATP/cell 1.28 nM in native vs. 0.63 in long term stored products, P < 0.05). Only borderline changes without statistical significance were detected when examining mitochondrial and aldehyde dehydrogenase metabolic activity and intracellular pH, showing their good preservation during cell storage. Our experience demonstrates that cryostorage has no major negative effect on stem cell quality and potency, and therefore autologous stem cells can be stored safely for an extended period of at least 10 years. On the other hand, long term storage for 10 years and longer may lead to mild reduction of clonogenic capacity. When a sufficient dose of stem cells is infused, these changes will not have a clinical impact. However, in products stored beyond 10 years, especially when a low number of CD34+ cells is available, the quality of stem cell graft should be verified before infusion using the appropriate potency assays.

Download Full-text

Post-transplantation cyclophosphamide reduces the incidence of acute graft-versus-host disease in patients with acute myeloid leukemia/myelodysplastic syndromes who receive immune checkpoint inhibitors after allogeneic hematopoietic stem cell transplantation

Journal for ImmunoTherapy of Cancer ◽

10.1136/jitc-2020-001818 ◽

2021 ◽

Vol 9 (2) ◽

pp. e001818 ◽

Cited By ~ 1

Author(s):

Chantal Saberian ◽

Noha Abdel-Wahab ◽

Ala Abudayyeh ◽

Hind Rafei ◽

Jacinth Joseph ◽

...

Keyword(s):

Acute Myeloid Leukemia ◽

Stem Cell ◽

Myelodysplastic Syndromes ◽

Myeloid Leukemia ◽

Acute Gvhd ◽

Checkpoint Inhibitors ◽

Post Transplantation ◽

Hematopoietic Stem ◽

Gvhd Prophylaxis ◽

Acute Myeloid

BackgroundImmune checkpoint inhibitors (ICIs) are being used after allogeneic hematopoietic stem cell transplantation (alloHCT) to reverse immune dysfunction. However, a major concern for the use of ICIs after alloHCT is the increased risk of graft-versus-host disease (GVHD). We analyzed the association between GVHD prophylaxis and frequency of GVHD in patients who had received ICI therapy after alloHCT.MethodsA retrospective study was performed in 21 patients with acute myeloid leukemia (n=16) or myelodysplastic syndromes (n=5) who were treated with antiprogrammed cell death protein 1 (16 patients) or anticytotoxic T lymphocyte-associated antigen 4 (5 patients) therapy for disease relapse after alloHCT. Associations between the type of GVHD prophylaxis and incidence of GVHD were analyzed.ResultsFour patients (19%) developed acute GVHD. The incidence of acute GVHD was associated only with the type of post-transplantation GVHD prophylaxis; none of the other variables included (stem cell source, donor type, age at alloHCT, conditioning regimen and prior history of GVHD) were associated with the frequency of acute GVHD. Twelve patients received post-transplantation cyclophosphamide (PTCy) for GVHD prophylaxis. Patients who received PTCy had a significantly shorter median time to initiation of ICI therapy after alloHCT compared with patients who did not receive PTCy (median 5.1 months compared with 26.6 months). Despite early ICI therapy initiation, patients who received PTCy had a lower observed cumulative incidence of grades 2–4 acute GVHD compared with patients who did not receive PTCy (16% compared with 22%; p=0.7). After controlling for comorbidities and time from alloHCT to ICI therapy initiation, the analysis showed that PTCy was associated with a 90% reduced risk of acute GVHD (HR 0.1, 95% CI 0.02 to 0.6, p=0.01).ConclusionsICI therapy for relapsed acute myeloid leukemia/myelodysplastic syndromes after alloHCT may be a safe and feasible option. PTCy appears to decrease the incidence of acute GVHD in this cohort of patients.

Download Full-text