bluuebunny/crossref_metadata_2025 huggingface dataset:
DNA-Microarray for Simultaneous Detection of Herpesviruses and Adenovirus in Patients Undergoing Allogeneic Stem Cell Transplantation.
Authors: Ditzen Anette K. , Rutzka Anke , Stichling Markus , Mueller Rene , Ehricht Ralf , Illmer Thomas , Bornhaeuser Martin , Jacobs Enno , Rohayem Jacques
Published: October 2019

Herpesviruses and adenovirus are an important cause of end-organ disease (EOD) and are associated with graft versus host disease (GVHD) in patients undergoing allogeneic stem cell transplantation (SCT). Detection and monitoring of those viruses is routinely performed by simplex quantitative real-time PCR. Since more than one herpes- or adenovirus are reported to be present shortly after transplantation, detection of all possible (sub-)clinical viral infections is still time-consuming and cost-intense. We therefore designed a DNA-microarray for the simultaneous detection of herpes-simplex-virus-1 and -2 (HSV-1/2), cytomegalovirus (CMV), varicella-zoster-virus (VZV), Epstein-Barr-virus (EBV), human herpesvirus-6 (HHV-6) and adenovirus. The DNA-microarray was validated in comparison to simplex quantitative PCR routinely used in our diagnostic laboratory. We retrospectively screened 608 samples of 35 patients undergoing allogeneic SCT on a regular basis until day 100 post-transplantation. The detection limit of the DNA-microarray is 10 genome equivalents (GE)/ml. We detected one or more viruses in 367 samples (60,2%). Of the positive samples, CMV was detected in 272 (74,1%), EBV in 103 (28,1%), HHV-6 in 81 (22,1%), adenovirus in 35 (9,5%), HSV-1/2 in 21 (5,7%) and VZV in 14 samples (5,2%), respectively. Of the samples containing two viruses (n=62), we detected CMV/EBV in 19 (30,6%), CMV/HHV-6 in 27 (43,6%) and HHV-6/EBV in 7 (11,3%) samples, respectively. CMV/EBV/HHV-6 was detected in 4 (36,4%) of the samples containing 3 viruses (n=11). Results of the DNA-microarray showed good overall agreement in comparison to the simplex quantitative PCR. We conclude that the DNA-microarray is a highly sensitive and specific method for the simultaneous detection of up to six viruses in a single sample. By screening and monitoring SCT-patients for viruses known to be associated with EOD or GVHD we herewith introduce an innovative molecular technique that can simultaneously detect multiple viral infections on a large scale. The assay is currently used to determine the impact of multi-virus detection on clinical end-points like GVHD and infectious complications in patients undergoing allogeneic stem cell transplantation.
Oral Busulfan and Cyclophosphamide or IV Busulfan and Fludarabine for Allogeneic HSCT in Hematologic Malignancies?.
Authors: Altman J. , Venepalli N. , Monreal J. , Rashid K. , Gordon L. , Winter J. , Williams S. , Tallman M. , Frankfurt O. , Evens A. , Singhal S. , Mehta J.
Published: October 2019

Pharmacokinetic properties of IV busulfan are more predictable than those of oral. While this results in lower toxicity, it is unclear if clinical results are superior. 55 consecutive patients with hematologic malignancies (35 AML/MDS, 10 lymphoma/CLL, 7 CML/MPD, 3 ALL) allografted after oral busulfan (14 mg/kg) with cyclophosphamide (120 mg/kg) (BuCy; n=24; cyclosporine-MTX) or IV busulfan (520 mg/m2) with fludarabine (160 mg/m2) (BuFlu; n=31; tacrolimus-MTX) were studied. The donors were HLA-identical sibs (n=41), 10/10 allele-matched unrelated (n=10), or 9/10 allele-matched individuals (n=4; 2 unrelated). Busulfan levels were not monitored. G/GM-CSF were not given routinely. Supportive care was uniform. 11 patients died of transplant-related causes, 21 relapsed (12 dead), and 32 were alive at the last follow-up (23 in continuous CR). The actuarial 2-y overall (OS) and event-free (EFS) survival was 50% (95% CI: 34–67%) and 34% (95% CI: 19–49%). The following pre-HSCT prognostic factors affected OS adversely in univariate analysis: refractory disease, ECOG performance status (PS) 2/3, elevated LDH, albumin <3, CD34+ cell dose ≤6 × 106/kg, HLA mismatch, and platelets <100 × 109/L. The table shows comparison of the BuCy and BuFlu by patient and donor age, type of donors, and the factors found to influence survival. The two groups were comparable.

In a Cox model that included the conditioning regimen and the above variables, low platelets (RR 0.285; P=0.013) and poor PS (RR 0.244; P=0.023) affected OS adversely. EFS was adversely affected by poor PS (RR=0.304; P=0.032), with borderline effect of low platelets (RR=0.451; P=0.062) and refractory disease (RR=0.502; P=0.095). The figure below shows OS by the presence or absence of the 2 adverse variables.

The figure below shows OS by the conditioning regimen. These curves have been generated from the Cox model and have been adjusted for the other variables in the model. While the difference is not statistically significant, BuFlu appears superior. However, similarly generated EFS curves for BuCy and BuFlu are superimposable.

We conclude that with limited patient numbers and follow-up, BuCy and BuFlu appear identical in efficacy although the appearance of the OS curves suggest that differences may emerge with loner follow-up and bigger patient numbers.

Figure Figure Figure Figure Characteristic BuCy BuFlu P Patient age 44 (27–62) 40 (20–61) 0.23 Donor age 46 (25–65) 39 (25–66) 0.25 MSD 22/24 (92%) 19/31 (61%) 0.013 Refractory disease 12/24 (50%) 16/31 (52%) 0.91 ECOG PS 2/3 6/24 (25%) 4/31 (13%) 0.30 Elevated LDH 6/24 (25%) 4/31 (13%) 0.30 Albumin < 3 7/24 (29%) 8/31 (26%) 0.78 CD34+ cell dose ≤6 × 106/kg 21/24 (88%) 21/31 (68%) 0..12 HLA mismatch 1/24 (4%) 3/31 (10%) 0.62 Platelets <100 × 109/L 9/24 (38%) 14/31 (45%) 0.57
Systematic Review of Controlled Clinical Trials on the Use of Ursodeoxycholic Acid for the Prevention of Hepatic Sinusoidal Obstruction Syndrome in Hematopoietic Stem Cell Transplantation.
Authors: Tay Jason , Tinmouth Alan T. , Fergusson Dean , Huebsch Lothar B. , Allan David S.
Published: October 2019

Background: Hepatic sinusoidal obstruction syndrome (HSOS) is a serious life-threatening complication of hematopoietic stem cell transplantation (HSCT). Currently, there is no optimal therapeutic strategy and preventive measures are ill-defined. Ursodeoxycholic acid (UA) is well-tolerated, inexpensive oral medication that has been associated with possible benefit as a prophylactic agent. We sought to summarize and quantify the clinical effects of prophylactic ursodeoxycholic acid in the context of HSCT. We undertook a systematic review of studies addressing the use of UA as monotherapy or in combination with other agents in patients undergoing HSCT.

Methods: The Search Strategy included MEDLINE (1966 to 4th week of March 2006), EMBASE (1980 to 4th week of March 2006), all EBM Reviews (4th quarter of 2005), Ovid Healthstar (1966 to 4th week of March 2006) and Google Scholar on March 20/2006. A random effects model was used to summarize outcome data.

Results: Six studies representing 824 patients were included in the review; 4 randomized clinical trials and 2 historical controlled studies. Three randomized clinical trials comparing prophylactic UA to no treatment demonstrated reduced proportion of HSOS (Relative Risk (RR) 0.34; 95% CI 0.17–0.66). When limited to higher quality studies, the effects remain (RR 0.36; 95%CI 0.15–0.90). Transplant related mortality also appears to be reduced (RR 0.58; 95% CI 0.35–0.95. However, UA did not significantly improve the outcomes of acute graft versus host disease (RR 0.76; 95% CI 0.53–1.09), relapse (RR 0.77; 95% CI 0.46–1.31) or overall survival (RR 1.22; 95 % CI 0.96–1.54).

Conclusion: UA is effective for HSOS prophylaxis in patients undergoing HSCT. HSCT centers should consider UA as standard therapy and controlled trials of novel treatments for HSOS should consider the use of UA as a comparator.
Risk Factor of Hepatic Veno-Occlusive Disease: Analysis of Clinical Data of 5024 Patients Extracted from the Database of the Japan Marrow Donor Program.
Authors: Sakai Miwa , Ohashi Kazuteru , Yamashita Takuya , Akiyama Hideki , Sakamaki Hisashi
Published: October 2019

Hepatic veno-occlusive disease (VOD) is one of the most serious complication of hematopoietic stem cell transplantation (HSCT). Various factors have been identified as increasing the risk of hepatic VOD, but few of them have been associated with a significantly increased risk. We retrospectively analyzed the clinical data of 5024 transplant recipients (median age28, range 0–68) which extracted from the Japan Marrow Donar Program. The diagnosis of VOD was made according to the McDonald’s criteria, and 324 out of 4833 patients (6.7%) were eventually diagnosed with VOD. The possible risk factors based on the previous studies were counted on an initial univariate analysis, and cumulated significant factors were further analyzed for their potential value for VOD development in multivariate analysis. Variables correlated with an increased risk of VOD were: time of transplant >2 times (relative risk (RR) 2.7; p=0.006), pretransplant disease status (RR 2.3; p=0.000), prior liver disease (RR 2.1; p=0.017), ABO blood type mismatch (RR1.7; p=0.000). In patients receiving either busulfan or melphalan for conditioning increased VOD risk (RR 1.5 and 1.8; p=0.007 and 0.002, respectively). In our multivariate analysis, stem cell source, and prophylactic use of heparin and Ursodiol had no significant effect on VOD development. This analysis might contribute to revise the previously reported risk factors for VOD and the data could be used to know which patients might be suitable subjects for new trials for VOD prevention.
Role of Zoledronic Acid in Treating Osteoporosis in Post-Allogeneic Bone Marrow Transplant Patients - A Phase II Trial.
Authors: Chamarthy Usha Sree , Rao D. Sudhaker , Divine George , Khan Shahzad , Janakiraman Nalini
Published: October 2019

Introduction: Multiple risk factors exist for the development of osteoporosis in patients undergoing allogeneic hematopoietic stem cell transplant. Bone loss is greatest during the first year, predominantly the first 40 days. Bisphosphonate therapy combined with calcium and Vitamin D supplementation in allogeneic stem cell transplant patients may be one useful strategy to prevent the accelerated bone resorption.

Study Design: A Phase II clinical trial studying the role of zoledronic acid (Zometa, Novartis Pharmcaeutical Corporation) in treating post-allogeneic bone marrow transplant(BMT) osteoporosis was undertaken. The study included adult patients undergoing allogeneic BMT for malignant and non-malignant disorders. Patients with plasma cell dyscrasias, thyrotoxicosis, primary and secondary hyperparathyroidism were excluded. Patients received zoledronic acid (4 mg IV over 15 minutes) pre-transplant and every three months thereafter for a total of 4 doses. All patients concomitantly received daily calcium (1000mg) and Vitamin D (400IU) supplements (oral or intravenous). Serial bone density(BMD) measurements at lumbar spine, total hip and forearm using DEXA were scheduled to be done at baseline (pre-transplant), 6 weeks, 6 months, 12 months, and 24 months. Urine NTX was a measured as a surrogate marker for bone turnover.

Results: 22 patients were enrolled between January 2002 and August 2005. BMD data were available at baseline on 20 patients, at 6 weeks on 11 patients, at 6 months on 11 patients, at 1 year on 9, and on 5 patients on 2 years. Random coefficient analyses were performed. A statistically significant reduction in total hip BMD was observed (−3.6%, p<0.001). The reduction for the lumbar spine (−1.1%) was not significant (p=0.416). The observed BMD change was positive at the forearm (+2.4%) and statistically significant (p=0.047). Post-transplant urine N-telopeptide (NTX) measurements were higher than baseline. Severe hypocalcemia (12%) was the only adverse effect attributable to the study drug.

Discussion: Review of recent literature implies that the degree of increased bone resorption pre- and post-BMT is highest at the femoral neck/total hip and can be up to 7.8% in the absence of bisphosphonate therapy. Our data indicates that zoledronic acid helps curtail the expected reduction in bone loss. Bisphoshonate therapy may be needed more frequently and for longer duration. Calcium/Vitamin D supplementation may need replenished more rigorously. Using urine NTX measurements concurrently with BMD to monitor bone turnover and response to treatment is indicated until standards of therapy are established. In conclusion, bisphosphonate therapy with zoledronic acid does impact standard of care for patients after allogeneic stem cell transplantation.
Costs of Imatinib, Costs of Transplants and Gross National Income Per Capita Impact on Transplant Rates for Chronic Myeloid Leukemia in European Countries.
Authors: Gratwohl Alois , Baldomero Helen , Schwendener Alvin , Gratwohl Michael , Frauendorfer Karl , Urbano-Ispizua Alvaro
Published: October 2019

Hematopoietic stem cell transplants (HSCT) are among the most expensive medical procedures. As a consequence, transplants are preferentially done in more wealthy countries. This significant dependency between transplant rates (number of transplants per 10 million inhabitants) and gross national income (GNI) per capita has been well described. We noticed a different pattern in HSCT for chronic myeloid leukemia (CML) in recent years. We did investigate the evolution of transplant rates for leukemias in Europe in respect to the economy of the participating 38 countries. Using World Bank definitions, countries were grouped according to their GNI per capita into low (A), middle (B), and high income (C) countries and trends of transplant rates for CML and acute myeloid leukemia (AML) were computed with regression analyses from 1991 to 2004. Transplant rates increased for AML in significant dependency with GNI per capita from 0.1 (A), 0.3 (B) and 12 (C) in 1991 to 2.8 (A), 22 (B) and 47 (C) in 2004. Transplant rates were distinct by World Bank category and the increase presented in all three groups with a near linear clear predictability as illustrated by the R2 ’s of 93.55, 94.34 or 98.76 (Groups A, B, C). A different pattern was observed for CML. Initially transplant rates did increase from 0.1 (A), 0.7 (B) and 13 (C) in 1991 to 0.28 (A), 15 (B) and 28 (C) in 1999, distinct by World Bank category and with a similar linear prediction pattern as for AML (R2 96.39, 95.75, 95.35 for groups A, B, C). Transplant rates then declined for CML in high income countries (C), they remained at the same level in middle income countries to the extent that comparable transplant rates (15, 15) were observed for middle and high income countries in recent years. We compared costs. Costs for one year treatment with Imatinib (400 mg daily dose) in the different participating European countries ranged between 28 000 and 44 000 Euros with a median of 30 411 Euros (mean 32 417 Euros). Costs for an allogeneic HSCT from an HLA identical sibling ranged from 26 515 Euros to 180 000 Euros with a median of 63 450 Euros (mean 72 173 Euros). Cost comparisons indicate that cost for an allogeneic HSCT correspond to 0·9 to 5·9 (median of 2·0, mean 2·3) years of drug treatment. Data suggest that Imatinib has replaced HSCT preferentially in countries with high income. HSCT remains a cost effective procedure in countries with middle income. Cost considerations appear to impact on the choice between a once in a lifetime procedure or continuous drug treatment. Integration of cost analyses into the decision algorythms will become of increasing importance.
Allogeneic Hematopoietic Stem Cell Transplantation Cures CLL: A Retrospective Analysis from the SFGM-TC Registry.
Authors: Michallet Mauricette , Lê Quoc-Hung , Vernant Jean-Paul , Nicolini Franck E. , Harousseau Jean-Luc , Colombat Philippe , Deconninck Eric , Cazin Bruno , Blaise Didier
Published: October 2019

This retrospective study concerned 471 B-CLL patients registered in the SFGM-TC registry from Apr 1,984 to Feb 2,005, who underwent either autologous transplantation (n=313, 138 F and 175 M, median age = 54, 236 PBSC and 77 BM) or allogeneic transplantation (n=158, 78 F and 80 M, median age = 49, 76 PBSC, 81 BM and 1 cord blood cell transplant from 17 related and 141 unrelated donors). Among alloT patients, 50 were ABO incompatible and 70 sex-mismatched. The median interval diagnosis-transplantation was 32 months for autoT and 51 months for alloT. Just before conditioning 302 autoT and 143 alloT were evaluated for the disease status: 100 and 26 patients were in CR, 170 and 55 were in PR, 4 and 13 in stable disease (SD), 28 and 49 in progressive disease (PD) for autoT and alloT respectively. Among alloT patients, 73 received reduced intensity conditioning (RIC) and 85 standard conditioning (72 Cyt+TBI, 33 Fluda+TBI, 23 Fluda+Bu+ATG, 8 Cyt+Bu and 21 other). Before autoT the conditioning consisted of 224 Cyt+TBI, 45 BEAM and 44 other. After alloT, 71 patients developed an aGVHD ≥ grade II and 60 developed a cGVHD (25 limited and 35 extensive). The non-relapse mortality at 1 year was 29%. With a mean follow-up of 28 months for autoT and 40 months for alloT, the probabilities of 3-year, 5-year and 8-year overall survival were 80%, 66%, 45.5% after autoT and 52%, 48% and 35% after alloT respectively. An analysis aimed to determine the percentage of long-term survivors, or patients censored on the final plateau of survival curves was performed on alloT and autoT groups. A mixture model, gfcure with Splus statistical package determined the percentages of long-term survivors and its adequacy was verified graphically. The percentage of long-term survivors for the autoT group was 1.2%, with a mean survival length for uncured population of 160 months. Fig A shows that both curves were close and consequently shows good adequacy and the absence of a final plateau. The percentage of long-term survivors for alloT was 34.03% (figure1). Fig B shows rather good adequacy. The study of the impact of usual prognosis factors (age, time diagnosis-transplant, sex match, HLA match, CMV status, type of conditioning, BM or PBSC, ABO compatibility and disease status before transplantation) on the percentage of long-term survivors showed that only the status of disease at transplant had a significant impact: (CR vs SD or PD, HR: 0.11 [0.02–0.5] p=0.01 and PR vs SD or PD, HR: 0.30 [0.09–0.96] p=0.04). This study pointed out the possibility of curing B-CLL patients who responded to conventional chemotherapy with allogeneic transplantation rather than with autologous transplantation.

Figure Figure Figure Figure
Lack of Predictive Value of KIR-HLA Genotypes for Autologous Transplant Outcome for Multiple Myeloma.
Authors: Hsu Katharine C. , Tricot Guido , Haessler Jeff , Pinto-Agnello Clara , van Rhee Frits , Fox Michelle , Dupont Bo
Published: October 2019

In allogeneic hematopoietic cell transplantation (HCT), donor alloreactive natural killer (NK) cells can prevent leukemia relapse and prolong survival. Donor NK alloreactivity has been shown to be influenced by the killer Ig-like receptor (KIR) repertoire of the donor and the HLA class I KIR ligand phenotype of the recipient. In T-cell depleted HLA-identical sibling HCT, it has been shown that lack of recipient class I ligand for donor inhibitory KIR is associated with improved outcome in acute myelogenous leukemia. In the normal population, more than 60% of individuals lack class I ligand for one or more of their inhibitory KIR. Because autoreactive NK cells are known to occur in normal individuals with low frequency, we sought to determine if these same NK clones driven by autologous KIR-HLA interactions might expand and play a role following treatment involving autologous stem cell infusion for hematologic malignancy. 233 patients with multiple myeloma who underwent high-dose chemotherapy with autologous stem cell infusion were evaluated with HLA and KIR genotyping. KIR-HLA genotype combinations were then correlated with event-free survival (EFS) and overall survival (OS) following autologous HCT. Cox regression models were fit to examine the association of KIR-HLA combinations with the hazards of mortality and relapse. We found that there was no association between lack of KIR ligand for inhibitory KIR and improved EFS (p=0.69) or OS (p=0.36), nor was there any association between presence of specific activating KIRs and transplant outcome. Furthermore, the cumulative effect of more than one activating KIR did not affect outcome, including the specific combination of KIR2DS1 and KIR2DS2 (EFS p=0.86 and OS p=0.67), which have been previously reported to be associated with lower leukemia relapse in allogeneic HLA-identical HCT. The combination of activating KIR with their known or putative HLA ligands was also not associated with improvement in transplant outcome. Patients were also evaluated for KIR haplotype (AA versus AB and BB). There was no association between KIR haplotype and DFS (p=0.59) or OS (p=0.96). These data suggest that KIR genotypes and KIR-HLA genotype combinations are not associated with improved outcome in multiple myeloma patients undergoing autologous HCT. This implies that NK cells in autologous HCT do not significantly impact outcome, either because they are not expanded in the post-HCT period or because they have no activity against autologous multiple myeloma cells.
Selective Depletion of CD25+ Host-Reactive Donor Lymphocytes from Allografts Preserves a CD25− CD4+foxp3+ Fraction of T Cells and Thereby Provides a Source for Efficient Reconstitution of Regulatory T Cells and Additional GVHD Control.
Authors: Mielke Stephan , Rezvani Katayoun , Solomon Scott R. , Savani Bipin N. , Nunes Raquel , Yong Agnes , Schindler John , Read Elizabeth J. , Vitetta Ellen S. , Barrett A. John
Published: October 2019

We previously showed that removal of host-reactive donor T cells from allografts by anti-CD25 immunotoxin (IT) is clinically feasible and reduces the frequency of severe graft-versus-host-disease (GVHD) in a high-risk group of elderly patients undergoing matched-sibling transplantation (Blood, 2005,106:1123). However, it is possible that the concurrent removal of CD4+CD25+ regulatory T cells (Tregs) with alloactivated CD25+ T cells could precipitate more GVHD if depletion of alloreacting cells were incomplete. We, therefore, studied the recovery of Tregs in 16 patients receiving CD25-IT-treated stem cell grafts during the first 150 days after transplantation. Tregs were characterized by surface phenotyping for CD3, CD4, CD25 and CD27, intracellular staining for forkhead protein3 (foxp3) and quantitative real time PCR for foxp3 gene. We also measured Tregs in 13/16 donors (Don), 10/16 SD products and 13/16 patients pretransplant (Pre). Patients received a median of 1.0 (0.2–1.5) ×108/kg selectively depleted (CD25−) CD3+ T cells and a stem cell product containing 0.25 ×104/kg (0.1–5.0) residual unselected CD3+ T cells. Lymphocyte recovery was prompt with a median lymphocyte count of 613/μL (215–2883/μL) and median 97% donor T cell chimerism at day+30. Despite CD25-depletion, patients reconstituted sufficient levels of CD4+foxp3+ T cells at day+30. These CD4+foxp3+ T cells underwent further expansion and reached their highest median levels 150 days post transplantation (Figure A). Interestingly, all SD-products contained a significant fraction of CD4+foxp3+ Tregs that persisted after complete CD25-depletion. These cells were exclusively CD25− CD4+foxp3+ Tregs and represented 1.5–4.8% of the CD4+ cells in the SD product. Of note, the Treg content of the donor was associated with the risk of GVHD development posttransplant. Acute grade II GVHD was restricted to 5 patients whose donors had significantly fewer Tregs compared to those with no or grade I GVHD (Figure B). In this study we show efficient Treg reconstitution despite giving a CD25-depleted SD allograft. We identified a population of CD25− CD4+foxp3+ Tregs in the SD product. Such cells have been shown to exert regulatory function in mice. Our results suggest that Treg recovery following SD is derived from a major contribution of CD25− Tregs which escape the depletion process in addition to a residual fraction of CD25− and CD25+ Tregs delivered with the stem cell product. All these Treg fractions may undergo marked expansion post-transplant and provide additional GVHD protection if the removal of alloreacting T cells by the SD procedure were incomplete.

Figure Figure
Cord Blood Transplantation from Unrelated Donor for Treatment of Childhood Acute Myeloid Leukemia: A Japan Cord Blood Bank Network Analysis.
Authors: Isoyama Keiichi , Kato Koji , Takanashi Minoko , Azuma Hiroshi , Nagamura-Inoue Tokiko , Kai Shunro , Oda Megumu , Nakabayashi Masao , Kato Shunichi
Published: October 2019

Results of cord blood transplantation (UCBT) from unrelated donor in childhood acute myeloid leukemia (AML) in Japan have not been previously reported. We analyzed 162 children receiving UCB transplants for AML [78 in low risk group (first or second complete remission; CR1 or CR2 and 84 in more advanced stage (high risk group)). Poor prognosis cytogenetic abnormalities were identified in 32 cases. Most patients received a 1 or 2 HLA antigens-mismatched UCB transplants. The median number of collected nucleated cells (NCs) was 4.7 x 107/kg. Cumulative incidence (CI) of neutrophil recovery was 79.5% +/− 3.2%%(71.5% +/− 5% in high risk group, and 88.3% +/− 3% in low risk group). The median time to neutrophil recovery was 27 day. In multivariate analysis, the significant factor influencing neutrophil engraftment was low risk group. Platelet recovery was 58.7% +/− 4.0%, acute graft-versus-host disease (GVHD) was 50% +/− 5%, and transplantation-related mortality (TRM) was 29.9% +/− 5%(13.6% +/− 4.4% in low risk group, and 51.4% +/− 10% for children in high risk group). The 5-year CI of relapse was 29% +/− 3% and was associated with disease status. The 5-year event-free survival (EFS) was 38.5% +/− 3% (55.6% +/− 7.9% in low risk group, and 22.8% +/− 5.8% for high risk group). The 5-year over all survival (OS) was 39.8% +/− 3% (65.6% +/− 8% in low risk group, and 17.4% +/− 5% for high risk group). In multivariable analysis, a low risk group was associated with good 5-year survival rates. Poor cytogenetic features had no significant influence on OS and EFS, respectively. We conclude that UCBT is a therapeutic option for children with very poor-prognosis AML and who lack an HLA-identical sibling.
Impact of HLA Allele Mismatch on the Outcome of Patients with Reduced-Intensity Allogeneic Haematopoietic Stem Cell Transplantation from Unrelated Donors: Analysis of 103 Patients of the French Registry.
Authors: Toma Andrea , Balere-Appert Marie-Lorraine , Chir Zina , Boiron Jean-Michel , Bordigoni Pierre , Socie Gerard , Vernant Jean-Paul , Renaud Marc , Milpied Noel-Jean , Michallet Mauricette , Deconninck Eric , Guyotat Denis , Garban Frederic , Sirvent Anne , Contentin Nathalie , Blaise Didier , Raffoux Colette , Buzyn Agnes
Published: October 2019

Reduced-intensity conditioning regimens (RIC) had become a classical strategy of allogeneic hematopoietic stem cell transplantation (HSCT) and many patients are now transplanted with unrelated donor. The aim of this restrospective study was to evaluate the impact of HLA mismatches between donor (D) and recipient (R) at the allelic level on survival after RIC. We analyzed 103 patients registered in France from Jan 1999 to Dec 2003 with a median age of 46 years (18–67). All patients had hematologic malignancies: AL (n=35), MM (n=18), CLL (n=5), NHL (n=11), HD (n=9), CML (n=12), MDS (n=9), and MPS (n=4). 39% of the patients were in an advanced phase of the disease at time of HSCT. Anti-thymocytes globulins (ATG) were part of the conditioning regimen for 77% of patients. The main source of stem cells was PBSC (n=65). Seventy-one D/R pairs (69%) were 10/10 HLA match at the allelic level. Mismatches concerned 5, 6, 15, 2 and 7 D/R pairs for HLA-A, -B, -C, -DRB1 and -DQB1, respectively. The results showed that 96% of patients engrafted. Acute GVHD grade II to IV and grade III/IV occurred in 46% and 19% of patients, respectively. The risk of developing cGvHD was 45% at 2 years. Overall survival (OS) was 42% at five years. Among the 47 patients alive, the median disease free survival (DFS) was 28 months. Among non-HLA parameters studied, the only factor associated with a good OS was the diagnosis of lymphoid disease (HD or NHL or CLL) (p=0.003). Recipient age <46y was only associated with less acute GvHD grade II to IV (p=0.008). Among HLA mismatches, we found that HLA-A and/or -B allelic mismatches had a negative impact on OS (p=0.006), DFS (p=0.006), acute GvHD grade II to IV (p=0.05). On the other hand, HLA-C or -DQB1 mismatches did not impact on OS, DFS, acute or chronic GvHD. We could not analyze DRB1 mismatch since there was only 2 patients reported. In conclusion, HSCT following RIC, with match or mismatch unrelated donors, is a feasible approach with best results observed for patients with lymphoid malignancies (NHL, CLL or HD). Among allelic HLA mismatches, HLA-A and/or -B seemed to be deleterious as compared to HLA-C or DQB1. These results help to identify most suitable donors and patients who are likely to benefit from RIC with unrelated donors when there is not a fully HLA match donor available.
Hematopoietic Stem Cell Transplantation (HSCT) for Benign Indications Using Plasma Depleted Umbilical Cord Blood Units (UCB) That Were Not Depleted of Red Blood Cells.
Authors: Rosenthal Joseph , Jaing Tang-Her , Nademanee Auayporn , Karanes Chatchada , Graham Michael , Chan Lee Lee , Eames Gretchen , Tan Ah Moy , Tan Poh-Lin , Wang Brian , Tseng Peggy , Chow Tiffany , Wu Tina , Lin Hai-Peng , Gjertson David , Petz Lawrence , Tan Patrick , Chow Robert
Published: October 2019

UCB is an attractive source for unrelated HSCT of benign indications; however, cell dosage is a critical factor for UCB HSCT. The red cell depletion (RD) and post-thaw wash techniques that are widely used incur significant nucleated cell loss; therefore, two strategies to minimize cell loss are to deplete plasma, but not the red blood cells (PD) during processing and forego post-thaw wash. A retrospective audited analysis was performed on 61 patients with benign disorders who were transplanted with 68 PD UCB units (8 double cords) with 29 thalassemias, 8 AA, 5 WAS, 5 SCID, 2 osteoporosis, 2 sickle cell disease, 2 Hemophagocytic Lymphohistiocytosis, 2 Hurler Syndrome, 1 CGD, 1 Fanconi’s Anemia, 1 Leroy I-Cell Disease, 1 Lymphohistiocytosis, 1 OIMD, and 1 Alpha Mannosidosis. Transplant characteristics: patient median age 4.25 years old (range 0.3–39); median weight 17 kg (range 5–76); male 56%; median # HLA ABDR matches of 5.0 (12–6/6; 19–5/6; 23–4/6; 5–3/6, 1–2/6); median pre-freeze TNC dose 8.1 x 107/kg; median post-thaw TNC dose as reported by TC 7.7 x 107/kg; median pre-freeze CD34 dose 3.1 x 105/kg; transplants outside of U.S.− 32 (52%); non-myeloablative − 9; 44% post-thaw washed (W), 56% infused without post-thaw wash (NW). The Kaplan-Meier estimates of 3-month ANC500 and 6-month platelet 20K and 50K engraftment are 87±5%, 83±6%, and 84±6% respectively. The median time to engraftment for ANC 500, platelet 20K, and 50K are 20 days (range 11–64), 46 days (range 13–153), and 61 days (range 21–171) respectively. No major adverse event was observed in either the W or the NW group, and the median time to engraftment for ANC 500, platelet 20K and 50K for W vs. NW were 21 vs. 19 days, 55 vs. 44.5 days, and 76 vs. 59 days respectively. The incidence of reported grade II–IV acute GVHD was 29%, and 10% had grade III–IV acute GVHD. 33% developed limited chronic GVHD, and 15% developed extensive chronic GVHD. With a median follow-up of 219 days (range 4–1402 days), the Kaplan-Meier estimates of 1-year TRM, OS and disease-free survival were 20±6%, 78±6% and 72±6% respectively. These results demonstrate that HSCT using unrelated PD UCB can be performed safely with outstanding results in patients with benign disorders, and post-thaw washing may delay engraftment of HSCT using PD UCB.
High-Dose Methotrexate (MTX) as Part of a Novel Immunosuppressive Regimen in Canine MHC-Haploidentical Hematopoietic Cell Transplantation (HCT).
Authors: Thakar Monica S. , Gooley Ted A. , Santos Erlinda B. , Storb Rainer F. , Sandmaier Brenda M.
Published: October 2019

In a preclinical canine model using major histocompatibility (MHC)-haploidentical littermate peripheral blood stem cell (PBSC) grafts, initial donor chimerism could be achieved after a nonmyeloablative conditioning regimen consisting of monoclonal antibody S5 (anti-CD44) from days −7 to −2, 200 cGy TBI on day 0, and MMF and cyclosporine for postgrafting immunosuppression. However, sustainable donor chimerisms could not be achieved in many of these dogs, with 25 of 44 (57%) rejecting their grafts at a median of 7 weeks after transplant. In order to improve the engraftment rate, a novel strategy to overcome the immunological barrier created by this MHC-haploidentical setting was developed. By adding single, progressively higher doses of MTX on day+3 after transplantation to the above regimen, we hoped to control host T and NK cells, both of which contribute to graft rejection in MHC-haploidentical HCT, and to target alloreactive donor T cells which are responsible for graft-versus-host disease (GVHD). Once dose toxicity is established, this system will also allow the study of transplants using gene-modified, MTX-resistant CD34 selected PBSC, which can provide additional intrinsic graft protection on exposure to MTX. To date, 11 dogs have been transplanted using the above regimen and adding either 50 (n=5), 100 (n=3), or 200 (n=3) mg/m2 MTX on day +3 followed by leucovorin rescue on days +4 and +5. Early results showed that 4 of 5 dogs experienced graft rejection in the MTX50 group, 0 of 3 dogs rejected in MTX100 (p=0.047, Monte Carlo), and all 3 dogs in MTX200 have continued to exhibit stable mixed chimerisms from +42 to +76 days after transplant. Dogs in all 3 cohorts had rapid engraftment within 1 week, with similar average peak mononuclear cell (MNC) chimerisms at comparable timepoints (83% at 5 wks, 88% at 6 wks, and 80% at 5 wks at MTX50, 100, and 200 respectively). None of the 11 dogs have shown evidence of GVHD or MTX toxicity, and all dogs cleared MTX within 48 hours. All dogs exhibited early donor tolerance on mixed lymphocyte culture assays within 2 weeks and preserved NK activity posttransplant. While these data are preliminary, adding single, progressively larger doses of MTX on day +3 to a novel, nonmyeloablative conditioning regimen appeared to improve engraftment among MHC-haploidentical canine recipients with no signs of GVHD. There was high initial donor chimerism without undue early toxicity. Further, this study will aid in our establishment of high-dose, posttransplant MTX as a way of promoting sustained donor chimerism after transplanting CD34 selected, MTX-resistant canine PBSC in the MHC-haploidentical setting. This preclinical data serve as a basis for developing nonmyeloablative transplant options for patients without HLA-identical donors.
Oral Administration of PBI-1402 Significantly Reduces Erythropenia and Accelerates Peripheral Blood Recovery in Myeloablated Mice Post Transplantation.
Authors: Morin Marie-Josée , Krosl Gorazd , Lagraoui Mouna , Grouix Brigitte , Penney Christopher , Gagnon Lyne , Roy Denis-Claude
Published: October 2019

The goal of this study was to determine the potential of PBI-1402, a compound that was shown to provide radio- and chemoprotection, to ameliorate the myeloablative therapy induced cytopenia and/or accelerate blood cell recovery of myeloablated hosts following autologous bone marrow transplantation in mouse model. Mice were exposed to irradiation and transplanted the following day with 5×105 bone marrow cells from syngeneic donors. Following transplantation, mice were divided into two groups that received two different doses of PBI-1402 orally and a control group that received PBS for 14 days post transplantation. The results showed that PBI-1402 treatment significantly diminished post-transplant erythropenia in a dose dependent manner. Similarly, determination of haemoglobin (Hb) demonstrated a significant attenuation of Hb loss and additionally, significantly accelerated post-transplantation recovery with the highest dose of PBI-1402. A significantly higher number of platelets was observed with the highest dose of PBI-1402. In addition, similar results were obtained with analysis of white blood cell (WBC) recovery. To determine if the observed effect of PBI-1402 treatment on red blood cell cytopenia and Hb recovery is the consequence of increased erythrocyte progenitor response to Epo, we performed an Epo dose response experiment on CFU-E bone marrow samples obtained from treated and control mice at different time points post transplantation. The results indicated that erythroid progenitors from treated and control mice responded similarly to Epo. However, PBI-1402 treated mice contained significantly higher number of CFU-E. Interestingly, analysis of more primitive Sca1+ progenitors revealed a significant increase in the number of Sca1+ cells from the bone marrow of mice treated with PBI-1402 (p < 0.05). These results demonstrate that PBI-1402 treatment significantly reduced irradiation induced erythropenia with concomitant acceleration in erythroid and Hb recovery. Additionally, treatment with PBI-1402 accelerated WBC recovery and platelet recovery and increased more primitive Sca1+ progenitors. PBI-1402 may offer protection from myeloabaltive therapy or accelerate recovery from remaining resident or transplanted stem cells.
Establishment of Chimerism in Rhesus Macaques through Nonmyeloablative Hematopoietic Stem Cell Transplant and Costimulation Blockade-Based Immunosuppression.
Authors: Kean Leslie , Adams Andrew , Strobert Elizabeth , Pearson Thomas , Larsen Christian
Published: October 2019

We established a strategy for producing high level hematopoietic chimerism after non-myeloablative conditioning in the Rhesus macaque. This strategy relies on hematopoietic stem cell transplantation after induction with a non-myeloablative dose of busulfan and blockade of the IL2-receptor in the setting of mTOR inhibition with rapamycin and combined CD28/CD154 costimulation blockade. Hematopoietic stem cells derived from bone marrow and leukopheresis products both were found to be successful in inducing high level chimerism. Mean peripheral blood peak donor chimerism was 81% with a median chimerism duration of 145 days. This prolonged chimerism occurred in the setting of full MHC disparity in unrelated transplant donors and recipients. Additional immune modulation strategies such as pre-transplant CD8 depletion, donor-specific transfusion, recipient thymectomy or peri-transplant deoxyspergualin treatment did not improve the level or durability of chimerism. Recipient immunologic assessment suggested that chimerism occurred amidst donor-specific down-regulation of alloreactive T cells, and the reappearance of vigorous T-mediated alloreactivity accompanied rejection of the transplants. Furthermore, viral reactivation constituted a significant transplant-related toxicity and may have negatively impacted the ability to achieve indefinite survival of transplanted stem cells. Nevertheless, this T cell costimulation blockade-based chimerism-induction regimen induced the longest-lived stem cell chimerism reported to date for non-human primates and thus represents an important platform upon which to evaluate emerging tolerance-induction strategies. Future nonmyeloablative transplant strategies based on immunomodulation via T cell costimulation blockade may have significant clinical relevence for nonmalignant hematologic diseases such as sickle cell disease and beta-thalassemia.
A Radiation-Free Immune and Epigenetics Based Conditioning Regimen for Allogeneic HCT: Combination of Anti-CD3 and Romidepsin.
Authors: Li Nainong , Zhang Chunyan , Lin Chia-Lei , McCulloch Bill , Wright John , Kirschbaum Mark , Forman Stephen , Zeng Defu
Published: October 2019

Hematopoietic cell transplantation (HCT) is a curative therapy for hematological malignancies as well as refractory autoimmune diseases. However, graft versus host disease (GVHD) remains a major obstacle in classical HCT, where recipients are usually conditioned with total body irradiation or high dose chemotherapy. We recently reported that donor CD8+ T cells facilitated engraftment and mediated graft versus leukemia (GVL) without causing graft versus host disease (GVHD) in young (6–8 weeks old) MHC-mismatched mouse recipients conditioned with anti-CD3 mAb (Blood 2005). Thereafter, we observed that anti-CD3 conditioning alone was not sufficient for induction of chimerism in old (>12 weeks) recipients, due to the higher percentage of residual host CD8+ T cells in the old recipients. Romidepsin (Desipeptide), a histone deacetylase inhibitor, was reported to induce apoptosis of human T cell lymphoma lines. We hypothesize that depsipeptide will induce the apoptosis of anti-CD3 activated proliferating T cells, and that conditioning with a combination of anti-CD3 and depsipeptide will markedly reduce the residual host T cells and allow donor stem cell engraftment.

To test our hypothesis, we first added Romidepsin (1.25–10 ng/ml) to cultures of T cells with or without anti-CD3 stimulation. We found that, although Romidepsin showed no effect on un-stimulated T cells, it augmented apoptosis of anti-CD3 activated T cells in a dose dependent manner, and the maximum augmentation was 5 fold. In addition, when Romidepsin (1.25–10 ng/ml) was added to a culture of mixed lymphocyte reaction (MLR), we found that it suppressed MLR in a dose dependent manner also, and the maximum suppression was greater than 98%. Second, old (> 12 weeks) BALB/c recipients were conditioned with one I.V. injection of anti-CD3 (20μg/g) and three I.P. injections (every other day) of Romidepsin at a dose of 0.4 μg/g. 7 days after anti-CD3 injection, recipients were injected with donor bone marrow cells (100×106) and CD4+- T depleted spleen (CD4−-SPL) cells (100×106). CD4−-SPL cells were injected again 7 days later. We found that, 4 weeks after HCT, 7/8 of the recipients conditioned with a combination of anti-CD3 and Romidepsin but only 1/8 of the recipients conditioned with anti-CD3 alone became chimeric. The recipients showed healthy appearance without signs of GVHD. The results are combined from two replicate experiments. This radiation free and GVHD preventive conditioning regimen may provide a novel approach for clinical HCT.
A PCR-Based Assay for the Detection of Langerhans-Cell Chimerism after Allogeneic Stem Cell Transplantation.
Authors: Meyer Ralf G. , Bakhtiar Shahrzad , Bender Klaus , Schmitt Timo , Konur Abdo , Huber Christoph , Herr Wolfgang
Published: October 2019

Early acute GVHD of the skin frequently occurs in patients after allogeneic hematopoietic stem cell transplantation. Although T cell depletion reduces the incidence and severity, it does not completely prevent skin GVHD. This leads to a prolonged need for immunosuppressive medication in a significant number of patients. For the induction of acute GVHD, the stimulation of donor T cells by residing host antigen presenting cells such as Langerhans cells of the skin (LCs) plays a central role. The absence of donor T cells after depletion, however, seems to hamper an early switch of LCs from host to donor origin. Therefore, the monitoring of LC chimerism is of great interest. We and others have provided evidence for a delayed switch in LC chimerism after T cell depleted reduced intensity stem cell transplantation. However, the assays used so far either are imprecise when applying low numbers of isolated cells or they depend on the detection of the Y-chromosome in skin sections of sex-mismatched transplants. In an attempt to set up a more sensitive assay of general applicability, we combined the detection of donor chimerism and tissue specific markers in a single multiplex PCR. We established PCRs for 10 different cDNA regions of constitutively expressed genes containing single nucleotide polymorphisms (SNPs). The SNP-containing products of the multiplex PCR were subsequently analyzed by the primer extension method (minisequencing) and subsequently analyzed by capillary electrophoresis. All SNP-containing cDNAs were expressed in peripheral blood mononuclear cells (PBMCs) as well as in isolated CD4- and CD8-positive T-lymphocytes, in myeloid dendritic cells, LCs, and keratinocytes. When we tested this approach on PBMCs of 10 patients and their HLA-matched sibling donors, the assay distinguished all pairs in 1 to 6 out of 10 systems. In a subsequent step, the 10plex PCR was combined with the tissue specific markers langerin for LCs and cytokeratin 10 to distinguish LCs from keratinocytes. Their expression was detected using gene-specific probes in the same minisequencing reaction used for the detection of SNPS. The resulting 12plex assay distinguished sibling donors from the patients with the same specificity and, in the same reaction, detected Langerin as well as cytokeratin 10 in purified LCs and keratinocytes, respectively. In summary, we established a sensitive assay allowing the simultaneous detection of donor chimerism together with the tissue specificity of isolated LCs that is independent of sex-mismatched donors. The addition of further tissue specific markers might allow performing chimerism studies on other tissue resident antigen presenting cells.
Crucial Role of FLT3 Ligand in Immune Reconstitution Following Bone Marrow Transplantation and High Dose Chemotherapy.
Authors: Buza-Vidas Natalija , Cheng Min , Duarte Sara , Jacobsen Sten Eirik W. , Sitnicka Ewa
Published: October 2019

Almost five decades after the first clinical transplantations, delayed immune reconstitution mechanisms regulating immune reconstitution post-transplantation remain to be established. While early recovery of innate immunity (myeloid and NK cells) results in reconstitution of protective immunity against bacterial pathogens, the levels of functional lymphocytes frequently remain abnormal for months or years. Low levels of B cells, immunoglobulins and CD4 T cells, predispose transplant recipients to severe bacterial and viral infections. Whereas the identification of granulocyte colony stimulating factor (G-CSF) and granulocyte/monocyte stimulating factor (GM-CSF) has been used to successfully promote clinical granulocyte recovery, no cytokine treatment has been developed to accelerate B and T cell reconstitution following myeloablation and BMT. While adult fms-like tyrosine kinase 3 (FLT3) ligand (FL) deficient (FL−/−) mice have reduced numbers of early B and T cell progenitors, they sustain close to normal levels of mature B and T cells (McKenna, H.J. et al. Blood 2000, Sitnicka, E. et al. Immunity 2002, Sitnicka, E. et al. J Exp Med 2003).

Herein, we demonstrate that adult BM cells fail to reconstitute B cell progenitors and conventional B cells in lethally irradiated FL−/− recipients, which also display delayed kinetics of T cell reconstitution. Similarly, FL is essential for B cell regeneration following chemotherapy-induced myeloablation. In contrast, fetal progenitors reconstitute B lymphopoiesis in FL−/− mice albeit at reduced levels. A critical role of FL in adult B lymphopoiesis is further substantiated by an age-progressive decline in peripheral conventional B cells in FL−/− mice, whereas fetally and early postnatally derived B1 and marginal zone B cells are sustained in a FL-independent manner. Thus, FL plays a crucial role in sustaining conventional B lymphopoiesis in adult mice, and as a consequence, our finding implicate a critical role of FL in promoting immune reconstitution following myeloablation and BM transplantation.