Myeloablative Allogeneic Hematopoietic Stem Cell Research Articles

Development and Validation of a Novel LC-MS/MS Method for a TDM-Guided Personalization of HSCT Conditioning with High-Dose Busulfan in Children

Personalization of busulfan (Bu) exposure via therapeutic drug monitoring (TDM) is recommended for patients treated with high-dose conditioning regimens. Several laboratories' developed methods are available in the literature with a lack of standardization. The aim of this study is to develop a new standardized LC-MS/MS method and validate it according to the international ICH M10 (EMA) guidelines. Our method is based on rapid protein precipitation from 50 µL plasma followed by separation on a reversed-phase C-18 UHPLC column after the addition of deuterated internal standard and has been tested on real samples from pediatric patients treated with myeloablative conditioning regimens, including Bu, before autologous or allogeneic hematopoietic stem cell transplantation (HSCT). The validated LC-MS/MS method is linear over wide concentration ranges (125-2000 ng/mL), accurate, and reproducible in the absence of matrix effects, allowing for the specific and rapid quantification of Bu and allowing next-dose recommendations to be made in a timely fashion to answer clinicians' needs. Given the lack of data on the stability of Bu in real clinical samples, stability was assessed both on quality controls and on real samples to set up a robust protocol in real-life conditions. This novel LC-MS/MS method is suitable for application to the TDM-guided personalization of conditioning treatments with high-dose busulfan in pediatric patients undergoing HSCT.

Rapid Immune Reconstitution and Elevated Regulatory T Cell Frequencies in Patients Treated with Orca-T

BACKGROUND Immune reconstitution following myeloablative allogeneic hematopoietic stem cell transplant (MA-alloHSCT) is significantly delayed for T cell depleted allografts when compared to T-cell replete allografts, a feature that has been implicated in higher rates and grades of infection, less GVT, and worse overall survival. Orca-T is a high precision cell therapy currently being investigated for the treatment of certain hematological malignancies otherwise treated with MA-alloHSCT. The cellular drug products of Orca-T (HSPCs, Tregs, and Tcons) are administered at high purity, in controlled doses, and on an established schedule with the intent to reconstitute the blood and immune system while controlling GVHD. Here, we present data on the immune reconstitution in 100 adult patients who received Orca-T. METHODS In the context of an ongoing multicenter Phase Ib clinical trial of Orca-T in recipients with hematologic malignancies (NCT04013685), we performed longitudinal measurements (days -28, 28, 56, 100, 180, and 365 post-transplant) of immune reconstitution in the first 100 consecutive patients. With fresh whole blood, clinical 5-part leukocyte differentials were performed at clinical sites, and lymphocyte subset frequencies were measured by flow cytometry in a central lab. Principal component analysis (PCA) was performed to investigate potential associations with recipient sex (male vs. female) and donor relation (related vs. unrelated). RESULTS Longitudinal peripheral blood counts of platelets, WBCs, neutrophils, lymphocytes, monocytes, T cells, B cells, and NK cells are presented in Table 1. T cell and B cell counts were readily observed at days 28 and 56 respectively, and increased with each subsequent time point. Median NK cell levels were observed to be in the normal range at all post-transplant time points. CD4+ T cell and Treg cell counts exhibited similar post-transplant patterns with both being appreciably present at d28 and increasing with each subsequent time point. Strikingly, relative to the level measured in 75 corresponding PBSC donors, the Treg frequency among CD4+ T cells was significantly elevated at all time points post-transplant (Figure 1). Median CD8+ T cell counts increased for the first 6 months post-transplant and then plateaued in the normal range. Upon PCA, very few significant differences were observed in 2-group comparisons of recipient sex and donor relation. CONCLUSIONS Orca-T patients exhibit early immune reconstitution of each of the major leukocyte and lymphocyte subsets hypothesized to control relapse and infection. Concomitantly, elevated Treg frequencies were also observed. This feature of immune reconstitution profiles of Orca-T recipients may be correlated to the reduced occurrence and severity of acute and chronic GVHD in these patients (Meyer et al., EHA 2022, #S237). Similar immune reconstitution profiles were observed in patients of disparate sex and regardless of donor relation. Prospective comparisons of immune reconstitution between Orca-T and standard-of-care patients will be performed in our ongoing phase 3 clinical trial (NCT05316701). Figure 1View largeDownload PPTFigure 1View largeDownload PPT Close modal

CT-524 Orca-T, A Precision Engineered Allograft, Results in High GVHD-Free and Relapse-Free Survival Following Myeloablative Conditioning for Hematological Malignancies

Rates of graft-versus-host disease (GVHD) and non-relapse mortality (NRM) following myeloablative allogeneic hematopoietic stem cell transplant (MA-alloHSCT) remain unacceptably high. Orca-T is a high-precision, allogeneic investigational cell therapy product comprised of stem and immune cells that leverages highly purified, polyclonal donor regulatory T cells to control alloreactive immune responses, reducing the need for pharmacologic GVHD prophylaxis. Orca-T is produced in a central GMP facility and has been successfully scaled to clinical centers throughout the U.S. The aim of these studies was to evaluate the safety and efficacy of Orca-T in patients with hematologic malignancies. Data are reported for 138 patients with high-risk hematologic malignancies who received Orca-T in a single-center Phase 1/2 study (n=41) and a multicenter Phase 1b study (n=97) and had ≥ 100 days of follow-up as of 2/28/22. Orca-T was produced from G-CSF-mobilized peripheral blood (PB) from MRD (n=72), MUD (n=62), or MMUD (n=4). Median follow-up for recipients was 300 days (range: 27-1941). Median age was 49 years, and diagnoses included AML (43%), ALL (27%), MDS (10%), myelofibrosis (7%), and CML (6%). Patients received MAC (busulfan-based, n=109; TBI-based, n=27; BCNU, n=2) followed by GVHD prophylaxis with either single-agent tacrolimus (n=127), sirolimus (n=7), or tacrolimus plus mycophenolate (n=4, MMUD). A contemporaneous CIBMTR-based control arm was obtained that consisted of patients with similar diagnoses who received MA-alloHSCT from a PB source followed by tacrolimus/methotrexate prophylaxis. Orca-T was successfully manufactured, distributed, and infused for all patients. Overall time from donor centers to recipient centers was under 60 hours in all cases. Median time to neutrophil engraftment was 13 days. Rates of grade ≥ 3 acute GVHD in the first 180 days and moderate to severe chronic GVHD through 1 year were low at 4% and 5%, respectively. NRM was infrequent at 4% through 1 year. Orca-T exhibited GVHD-free relapse-free survival (GRFS) of 71% & OS of 90% at 1 year. No formal comparison to CIBMTR cohort was performed. Results from patients treated with Orca-T, a high-precision Treg-engineered donor product, suggest a reduction in cGVHD, improved GRFS, and low toxicity relative to historic data. A multicenter randomized-control phase 3 trial comparing Orca-T to SOC is enrolling.

S237: ORCA-T, AN ENGINEERED ALLOGRAFT, RESULTS IN HIGH GVHD-FREE AND RELAPSE-FREE SURVIVAL FOLLOWING MYELOABLATIVE CONDITIONING FOR HEMATOLOGICAL MALIGNANCIES

Background: Rates of graft versus host disease (GVHD) and non-relapse mortality (NRM) following myeloablative allogeneic hematopoietic stem cell transplant (MA-alloHSCT) remain unacceptably high. Strategies to reduce GVHD and NRM have been compromised by limited efficacy or increased risk of infection and relapse, emphasizing the need for new approaches that holistically improve outcomes. Orca-T is a high-precision, allogeneic investigational cell therapy product comprised of stem and immune cells that leverages highly purified, polyclonal donor regulatory T cells to control alloreactive immune responses, reducing the need for pharmacologic GVHD prophylaxis. Orca-T is produced in a central GMP facility and has been successfully scaled to clinical centers throughout the U.S. Aims: The aim of these studies was to evaluate the safety and efficacy of Orca-T in patients with hematologic malignancies. Methods: As of 28 February 2022, 138 patients with high-risk hematologic malignancies have received Orca-T in a single-center Phase 1-2 study (NCT01660607, n=41) and a multicenter Phase 1b study (NCT04013685, n=97) and have ≥ 100 days of follow-up. Informed consent was obtained from all transplant recipients and donors, and the studies received IRB approval from participating institutions. Orca-T was produced from G-CSF-mobilized peripheral blood (PB) from matched related donors (n=72), matched unrelated donors (n=62), or mismatched unrelated donors (MMUD, n=4). Median follow-up for recipients was 300 days (range: 27-1941). Median age was 49 years, and diagnoses included AML (43%), ALL (27%), MDS (10%), myelofibrosis (7%), and CML (6%). Patients received myeloablative conditioning (busulfan-based, n=109; TBI-based, n=27; BCNU, n=2) followed by GVHD prophylaxis with either single-agent tacrolimus (tac, n=127), sirolimus (n=7), or tac plus mycophenolate (n=4, MMUD). A contemporaneous CIBMTR-based control arm was obtained that consisted of patients with similar diagnoses who received myeloablative alloHSCT from a PB source followed by tac/methotrexate PPX. Results: Orca-T was successfully manufactured, distributed, and infused for all patients enrolled. Overall time from donor centers to recipient centers was under 60 hours in all cases. Median time to neutrophil engraftment was 13 days. The rates of grade ≥ 3 acute GVHD in the first 180 days and moderate to severe chronic GVHD through 1 year were low with Orca-T at 4% and 5%, respectively. NRM was infrequent at 4% through 1 year. Orca-T exhibited GRFS of 71% & OS of 90% at 1 year. No formal comparison to the CIBMTR cohort was performed, and a Phase 3 study has been initiated to confirm these findings. Longitudinal immune reconstitution data was collected and will be presented. Clinical data is summarized in Table 1. *MAGIC Criteria **NIH Consensus Grading Image:Summary/Conclusion: Results from patients treated with Orca-T, a high-precision Treg-engineered donor product, suggest a reduction in cGVHD, improved GRFS, and low toxicity relative to historic data. Orca-T manufacturing was accomplished with consistent and reliable cell manufacturing and distribution across a wide geographic area. A multicenter randomized-control trial phase 3 trial comparing Orca-T to SOC has been initiated.

Treating Relapsed/Refractory Acute Myeloid Leukemia with Chidamide, Fludarabine, Cytarabine and Granulocyte-Colony Stimulating Factor with Subsequent Bridging to Myeloablative Allogeneic Hematopoietic Stem Cell Transplantation.

Treating Relapsed/Refractory Acute Myeloid Leukemia with Chidamide, Fludarabine, Cytarabine and Granulocyte-Colony Stimulating Factor with Subsequent Bridging to Myeloablative Allogeneic Hematopoietic Stem Cell Transplantation.

Radiation-Free myeloablative allogeneic hematopoietic stem cell transplantation for adult acute lymphoblastic leukemia: A comparison of outcomes between patients with and without central nervous system involvement

For patients with acute lymphoblastic leukemia (ALL) undergoing allogeneic hematopoietic stem cell transplantation (allo-HSCT), total body irradiation (TBI) has been particularly advocated as a part of the conditioning regimen in case of extramedullary involvement in sanctuary sites such as the central nervous system (CNS), to ensure greater tissue penetration. In resource-limited countries lacking TBI facilities; however, ALL patients undergo radiation-free myeloablative conditioning, though its impacts on post-HSCT outcomes of the patients with pre-HSCT CNS involvement have not been analyzed. In this 14-year series of 278 adult (> 18 y) ALL patients undergoing TBI-free busulfan/cyclophosphamide conditioning allo-HSCT, we found that the long-term probabilities of overall survival, disease free survival, relapse and non-relapse mortality were not significantly different between CNS-involved and CNS-spared patients. Moreover, there was no statistically significant difference in the incidence of post-HSCT CNS relapse between CNS-involved and CNS-spared patients. Pre-HSCT cranial radiation therapy (CRT) showed no significant preventive effect on the likelihood of post-HSCT CNS relapse. Through multivariable regression analysis, grade III-IV acute graft-versus-host disease (GvHD), extensive chronic GvHD and post-HSCT relapse were ascertained as independent determinants of mortality (Adj.R2 = 53.9 %, F(12,265) = 28.1, P < 0.001), while other parameters including Philadelphia translocation, pre-HSCT CNS involvement and CRT were found to have no independent effect. Although this study was not an attempt to compare TBI-based vs. non-TBI conditioning, the TBI-free myeloablative allo-HSCT was shown to be feasible and an option for adult ALL patients with CNS involvement, considering the comparable outcomes between patients with and without CNS involvement.

Ex Vivo T Cell-Depleted and In Vivo T Cell-Depleted Allogeneic Stem Cell Transplantation in Patients with Acute Myeloid Leukemia in First Complete Remission Resulted in Similar Overall Survival Survival: On Behalf of the ALWP of the EBMT and the MSKCC

Background. Myeloablative allogeneic hematopoietic stem cell transplantation (allo-HSCT) is a well-established therapy for adult patients with acute myeloid leukemia (AML) in first complete remission (CR1) with intermediate or high-risk cytogenetics. However regimen-related toxicities, in particular graft-versus-host disease (GVHD) have limited its widespread use. Therefore several strategies have been developed to deplete T cells in order to prevent GVHD. In vivo T-cell depletion (TCD) using antithymocyte globulin (ATG) with an unmanipulated graft is widely used in some centers in Europe, while other centers have develop an effective platform based on ex-vivo TCD CD34 selected (CD34+) graft. The aim of the current study was to compare these two strategies for GVHD prevention.Patients and methods. This retrospective analysis assessed the outcome of all consecutives patients over 18 years at time of transplant with an AML in CR1 with intermediate or high risk cytogenetics who received an in-vivo or ex-vivo T cell depleted myeloablative allo-HSCT using busulfan or total body irradiation based conditioning regimen between 2005 and 2011 and reported to the EBMT ALWP registry centers or at Memorial Sloan Kettering Cancer Center (MSK), respectively. Patients treated in EBMT centers received in vivo TCD with either thymoglobuline or grafalon ATG. In addition, all patients in the ATG group received GVHD prophylaxis consisting of a calcineurin inhibitor in combination with another drug. Patients treated at MSK received an ex-vivo TCD graft (CD34+). TCD was performed by immunomagnetic CD34+ cell selection of peripheral blood grafts. No additional GVHD prophylaxis was administered in those patients. Main transplant outcomes including refined GVHD-free/relapse-free survival (GRFS) were evaluated.Results. A total of 525 patients were included (363 ATG, 162 CD34+) with a median age of 46 (19-77) and 58 (range, 20-73) years, respectively (p<0.001). The ATG cohort had more unrelated donors (67.5% vs. 54.9%, p=0.006). Median follow-up was significantly longer in the CD34+ cohort [58.0 (range, 6-139.0), vs. 24.5, 51.9-131.0) months, p<0.001], therefore all patients were censored at 2 years for comparison between groups. There were no additional differences between cohorts including donor/recipient gender, CMV status and cytogenetic risk. The CD34+ cohort had less relapse (21.6% vs. 30%, p=0.03), grade II-IV aGVHD (11.3% vs. 21.2%, p=0.006), III-IV aGVHD (1.3% vs. 6.2%, p=0.01), cGVHD (2.5% vs. 27.6%, p<0.0001) and extensive cGVHD (0.6% vs. 11.3%, p=0<0.0001) and a higher GRFS (61% vs. 47%, p=0.0007) while there was no difference regarding OS (67.6% vs. 56.4%, p=0.31), LFS (61% vs. 57.9%, p=0.29) and NRM (17.4%, vs 12.1%, p=0.16). In multivariate analysis, the cumulative incidence of grade II-IV aGVHD and chronic GVHD were higher in the ATG cohort [HR 2.0 (95% CI 1.1-3.7), p=0.02 and HR 15.1 (95% CI 5.3-42.2), p<0.0001]. There were no statistical differences between both approach in terms of relapse, NRM, OS, and LFS. Parameters associated with a lower GRFS were ATG group, adverse cytogenetic and use of an unrelated donor [HR 1.7 (95% CI 1.2-2.5) p=0.001; HR 1,7 (95% CI 1.3-2.2) p=0.0002 and HR 1.5 (95% CI 1.1-2.0) p=0.01]. Cytogenetic status (high risk versus intermediate risk) also has a significant impact on relapse incidence [HR 2.1 (95% CI 1.4-3.0), p<0.0001] and LFS [HR 1.7 (95% CI 1.2-2.3) p=0.0006]. Finally there was significantly more death related to infection in the CD34 group (16/52, vs. 19/112, p=0.04).Conclusion. The cumulative incidence of acute (total and severe) and chronic GVHD were higher after allo-HSCT with ATG, leading to a lower GRFS in those patients. In contrast, enhance immunosuppression in the CD34+ group lead to a higher incidence of infectious related death. Overall NRM, OS and LFS are similar after ex-vivo CD34 selected and in-vivo ATG TCD allo-HSCT from related/unrelated donors in patients with AML in CR1 and intermediate/high-risk cytogenetic. Both approaches are associated with a good disease control and a high OS, and should be compared in a prospective randomized trial. DisclosuresMohty:Sanofi: Honoraria, Speakers Bureau.

Isolated Extramedullary Relapse After Human Leukocyte Antigen-Matched Myeloablative Allogeneic Hematopoietic Stem Cell Transplantation in Acute Myeloid Leukemia Patients: Case Reports and Literature Review

Isolated extramedullary relapse (iEMR) of acute myeloid leukemia (AML) after allogeneic hematopoietic stem cell transplantation (allo-HSCT) is rare and has a dismal prognosis. Among 67 patients with AML after allo-HSCT, iEMR and bone marrow relapse occurred in 6% and 20.9%, respectively, with a median time to relapse of 11.5 and 6.5 months, respectively. Here, we presented 4 iEMR-AML cases. Common relapse locations occurred in the central nervous system, skin, and lymph nodes. We also report a rare case of cardiac iEMR that responded to chemoradiotherapy. Two cases responded to local/systemic treatments, which resulted in prolonged survival. Another case had iEMR in the presence of chronic graft-versus-host disease. Bone marrow relapse occurring after iEMR was typical and found in three-fourths of the cases. In conclusion, iEMR-AML occurrence after allo-HSCT is not rare in Thai patients. Its unpredictability and lack of graft-versus-leukemia effect highlight the importance of monitoring EMR carefully and promptly providing treatments once it is detected.

Trudności w leczeniu chorego na ostrą białaczkę szpikową z mutacją FLT3-ITD i wysokim stosunkiem allelicznym — oporność na standardową chemioterapię indukującą w połączeniu z midostauryną

The development of targeted therapies in AML patients enabling treatment individualization, such as new FLT3 tyrosine kinase inhibitors, is a promising option for improving treatment outcomes and prolonging patient survival. However, the treatment of patients with a high FLT3-ITD allelic ratio (FLT3-ITD high ) associated with an extremely unfavourable prognosis remains a major clinical problem. The study presents a case of a 20-year-old patient with FLT3-ITD high extramedullary AML at diagnosis. Individualized chemotherapy according to the DA ‘3 + 7’ regimen combined with midostaurin was administered. After the induction treatment, complete remission (CR) was not achieved. After second induction chemotherapy, CR1 was achieved with the presence of residua disease. One cycle of consolidation chemotherapy was then administered, and after myeloablative conditioning allogeneic hematopoietic stem cell transplantation from an unrelated donor was performed. The patient has remained in CR with no residual disease for 18 months.

Impact of disease status at allogeneic stem cell transplantation on adolescent and young adult patients with early T‐cell precursor acute lymphoblastic leukemia

Early T-cell precursor (ETP) acute lymphoblastic leukemia (ALL) is an aggressive subset of T-cell ALL, and the role of allogeneic hematopoietic stem cell transplantation (allo-HSCT) in adolescent and young adult (AYA) patients has not been sufficiently described. We retrospectively analysed the data of 30 AYA patients (19 in first complete remission [CR1], 3 in CR2, and 8 with active disease) with ETP-ALL who underwent myeloablative allogeneic hematopoietic stem cell transplantation (allo-HSCT) from HLA-matched related, n=2, unrelated, n=5, or HLA-haploidentical related, n=23 donors with an emphasis on the impact of disease status on the outcomes of transplant. The stem cell source was unmanipulated G-CSF mobilized bone marrow or peripheral blood stem cells. All patients achieved neutrophil engraftment with full donor chimerism. The cumulative incidences of grade II to IV acute graft-versus-host disease (GVHD) and chronic GVHD at 2 years were 37% and 33%, respectively. Overall, 16 patients died. The causes of death were relapse (8 patients), infection (4 patients) and GVHD (4 patients). The estimated 2-year overall survival (OS) and leukemia-free survival (LFS) for the whole cohort were 47.8% and 46.2%, respectively. Patients transplanted in CR1/2 had significantly better 2-year OS and LFS than patients with active disease (61.7% vs. 12.5%, p=0.02; and 58.3% vs. 12.5%, p=0.04, respectively). There was a trend toward an inferior OS rate in those patients in CR1 with chemoresistance or in CR2 compared with patients in CR1 with chemosensitivity, although this did not reach statistical significance. Our data support allo-HSCT, especially from HLA-haploidentical donors as an effective therapeutic strategy in AYA patients with ETP-ALL and disease status was significantly associated with survival in these patients.

Decitabine induction with myeloablative conditioning and allogeneic hematopoietic stem cell transplantation in high-risk patients with myeloid malignancies is associated with a high rate of infectious complications

Patients with high-risk myelodysplastic syndrome or acute myeloid leukemia have an increased risk of death following allogeneic hematopoietic stem cell transplantation (allo-HSCT). Decitabine has minimal non-hematologic toxicity and proven efficacy in myeloid diseases, and post-transplant cyclophosphamide (PTCy) has reduced rates of graft-versus-host-disease (GVHD). We hypothesized that decitabine induction with allo-HSCT and PTCy would improve outcomes in a high-risk myeloid disease population. We performed a phase-II trial of decitabine at 20 mg/m2 for 10 days followed by allo-HSCT using a myeloablative regimen of fludarabine, IV busulfan and 4 Gy total body irradiation with PTCy for GVHD prophylaxis. Twenty patients underwent decitabine induction and 17 patients proceeded to transplant per protocol. Median overall survival from decitabine induction was 210 days (95 % CI 122-not reached). All patients developed grade 4 neutropenia after decitabine, eleven patients (55 %) developed grade 3−4 infections, and 5 cases were fatal. There were 5/20 (25 %) long-term survivors with a median follow-up of 3.6 years. Decitabine induction followed by myeloablative allo-HSCT in a high-risk population was associated with a high risk of infection and mortality related to enhanced immunosuppression. Further exploration of decitabine conditioning on reduced intensity platforms and improved infectious prophylaxis and screening may better mitigate toxicity (ClinicalTrials.gov (NCT01707004)).

Ruxolitinib in children with steroid-refractory acute graft-versus-host disease: A retrospective multicenter study of the pediatric group of SFGM-TC.

We conducted a national multicenter retrospective study in France to evaluate the efficacy and tolerance of ruxolitinib in children with steroid-refractory acute graft-versus-host disease (aGVHD) after allogeneic hematopoietic stem cell transplant. Patients were recruited from the 15 pediatric transplantation centers. Transplanted patients were eligible if they met the following criteria: aged ≤ 18 years at transplantation, receiving a myeloablative allogeneic hematopoietic stem cell transplant, having an aGVHD of grade ≥2, and treated with ruxolitinib for steroid-refractory aGVHD. Twenty-nine patients received ruxolitinib for steroid-refractory aGVHD. Six patients achieved a complete response at day 28 after the start of treatment but finally 19 patients (65.5%) achieved a complete response (CR) with a median delay of 41 days (5-93 days). Two patients had a partial response. All patients who achieved CR or partial response discontinued corticosteroid treatment. Eight patients showed treatment failure. The overall response rate was 72.4%. Twenty-three of 29 patients were alive at a median follow-up of 685 days (177-1042 days) after the hematopoietic stem cell transplantation. Viral replication was observed in 41.4% of cases. We did not observe severe hematological adverse events and cytopenia requiring a modification of ruxolitinib doses always resolved. The median initial dose of ruxolitinib was 12.6mg/m2 /day with an important range. We could not demonstrate any relationship between initial dose and effectiveness. Ruxolitinib may constitute a promising second-line treatment for children with steroid-refractory aGVHD that should be validated in a prospective large-scale pharmacokinetic and efficacy trial.

A gut microbiota score predicting acute graft-versus-host disease following myeloablative allogeneic hematopoietic stem cell transplantation.

Although studies have reported that intestinal microbiota are associated with acute graft‐versus‐host disease (aGVHD), they lacked a satisfactory method for predicting aGVHD. We collected stool and blood samples at day 15 posttransplant from 150 patients from two centers who underwent myeloablative conditioning allogeneic hematopoietic stem cell transplantation (allo‐HSCT). Stool microbiota were detected by 16S ribosomal RNA gene sequencing; inflammatory factors and T lymphocytes were detected by multiplex immunoassays and flow cytometry, respectively. A gut microbiota score (GMS) from a LASSO (least absolute shrinkage and selection operator) model was developed and validated to predict aGVHD. In the discovery cohort, the GMS could predict II‐IV aGVHD (area under the receiver operating characteristic [ROC] curve [AUC] = 0.904, P < .0001). Furthermore, the validation model was consistent with the discovery set (AUC = 0.887, P < .0001). Regulatory T/T‐helper 17 (Treg/Th17) cells ratio in the low GMS subgroup was higher compared with the high GMS (P = .012), and the validation set is consistent with the discovery set (P = .003). In addition, high cytokine levels were associated with high GMS. In conclusion, the GMS at neutrophil engraftment could predict aGVHD, and it was a potential and novel method. The GMS was associated with the inflammatory factor and Treg/Th17 balance.

Myeloablative and Reduced-Intensity Conditioned Allogeneic Hematopoietic Stem Cell Transplantation in Myelofibrosis: A Retrospective Study by the Chronic Malignancies Working Party of the European Society for Blood and Marrow Transplantation.

This retrospective study by the European Society for Blood and Marrow Transplantation analyzed the outcome of 2224 patients with myelofibrosis (MF) who underwent allogeneic stem cell transplantation (allo-SCT) between 2000 and 2014; 781 (35%) underwent myeloablative conditioning (MAC) and 1443 (65%) reduced-intensity conditioning (RIC). Median patient age was 52.9 years (range, 18 to 74 years) and 57.5 years (range, 21 to 76 years) in the MAC and RIC cohorts, respectively. Donor type was similar: matched sibling donors (MAC, 317 [41%]; RIC, 552 [38%]) and unrelated donors (MAC, 464 [59%]; RIC, 891 [62%]). Median time to both neutrophil and platelet (>20 × 109/L) engraftment did not differ between cohorts. Rates of grade II to IV acute GVHD were 28% (MAC) and 31% (RIC; P = NS). Cumulative chronic GVHD rates (limited/extensive) were 22%/27% (MAC) and 19%/31% (RIC; P = .10). Cumulative incidences of nonrelapse mortality (NRM) at 1, 3, and 5 years were 25.5%, 32.2%, and 34.6% (MAC) and 26.3%, 32.8%, and 34.4% (RIC), respectively. There was a trend toward a higher relapse rate with RIC regimens compared with MAC (P = .08); rates at 1, 3, and 5 years were 10.9%, 17.2%, and 20.1% (MAC) and 14%, 19.7%, and 23.2% (RIC), respectively. No significant difference in 5-year probabilities of overall survival (OS) was noted: MAC (53.0%; 95% confidence interval [CI], 49.1% to 56.9%) and RIC (51.0%; 95% CI, 48.3% to 53.7%); P = .78. Regarding the composite end point of GVHD-free/relapse-free survival (GRFS), the unadjusted Kaplan-Meier estimate of 5-year GRFS was 32.4% (95% CI, 29.0% to 36.1%) in the MAC group and 26.1% (95% CI, 23.9% to 28.2%) in the RIC group (P = .001). In the MAC cohort, multivariable analysis confirmed worse OS and NRM with older age (>50 years), using an unrelated donor and a Karnofsky Performance Status of 80 or less. For the RIC cohort, worse OS and NRM were associated with age 60 to 70 years compared with younger recipients, use of a mismatched donor, and poor performance status. In conclusion, although similar OS rates existed for both cohorts overall, this study suggests that MAC should still be used for younger individuals suitable for such an approach due to a trend toward less relapse and an overall suggested advantage of improved GRFS, albeit this should be examined in a more homogeneous cohort. RIC allo-SCT still offers significant survival advantage in the older, fitter MF allograft patient, and optimization to reduce significant relapse and NRM rates is required.

Prognostic impact of circulating tumor DNA status post–allogeneic hematopoietic stem cell transplantation in AML and MDS

AbstractThis study was performed to assess the utility of tumor-derived fragmentary DNA, or circulating tumor DNA (ctDNA), for identifying high-risk patients for relapse of acute myeloid leukemia and myelodysplastic syndrome (AML/MDS) after undergoing myeloablative allogeneic hematopoietic stem cell transplantation (alloSCT). We retrospectively collected tumor and available matched serum samples at diagnosis and 1 and 3 months post-alloSCT from 53 patients with AML/MDS. After identifying driver mutations in 51 patients using next-generation sequencing, we designed at least 1 personalized digital polymerase chain reaction assay per case. Diagnostic ctDNA and matched tumor DNA exhibited excellent correlations with variant allele frequencies. Sixteen patients relapsed after a median of 7 months post-alloSCT. Both mutation persistence (MP) in bone marrow (BM) at 1 and 3 months post-alloSCT and corresponding ctDNA persistence (CP) in the matched serum (MP1 and MP3; CP1 and CP3, respectively) were comparably associated with higher 3-year cumulative incidence of relapse (CIR) rates (MP1 vs non-MP1, 72.9% vs 13.8% [P = .0012]; CP1 vs non-CP1, 65.6% vs 9.0% [P = .0002]; MP3 vs non-MP3, 80% vs 11.6% [P = .0002]; CP3 vs non-CP3, 71.4% vs 8.4% [P < .0001]). We subsequently evaluated whether subset analysis of patients with 3 genes associated with clonal hematopoiesis, DNMT3A, TET2, and ASXL1 (DTA), could also be helpful in relapse prediction. As a result, CP based on DTA gene mutations also had the prognostic effect on CIR. These results, for the first time, support the utility of ctDNA as a noninvasive prognostic biomarker in patients with AML/MDS undergoing alloSCT.

Modified cladribine, cytarabine, and G-CSF as a salvage regimen in patients with relapsed/refractory acute myeloid leukemia: a bridge to myeloablative allogeneic hematopoietic stem cell transplantation

Patients with primary refractory or early relapsed acute myeloid leukemia (AML) have a dismal prognosis, and the treatment options for these patients are limited. The present study retrospectively examined the efficacy and toxicities of the combination of cladribine 5mg/m2 per day and intermediate-dose cytarabine 1g/m2 per day for 5days and granulocyte colony-stimulating factor (G-CSF) as a salvage treatment in 36 patients with relapsed/refractory AML. Among these, 32 patients had de novo AML, and the remaining 4 patients had secondary AML. The median age for the study cohort was 45.8years. According to the European LeukemiaNet prognostic index, 5 patients had favorable risk, 18 had intermediate risk, and 11 had poor risk. The complete remission was achieved in 58% of the patients with tolerable toxicities. Fifteen patients underwent stem cell transplantation later. Patients who underwent allogeneic hematopoietic stem cell transplantation had a significantly improved 1-year overall survival compared with those who did not (73% vs. 29%, P < 0.001). The results suggested that, as a salvage regimen, modified cladribine, cytarabine, and G-CSF were effective and well tolerated for patients with relapsed/refractory AML, especially for patients who underwent subsequent stem cell transplantation.

Concomitant Use of Blinatumomab and Donor Lymphocyte Infusion for Mixed-Phenotype Acute Leukemia: A Case Report With Literature Review

Blinatumomab and donor lymphocyte infusion (DLI) combination is a promising cancer therapy, whereby blinatumomab might achieve an initial reduction in leukemic-cell burden using T cells, and after tumor clearance, DLI can potentially stimulate the donor immune system to achieve longer lasting remission. Here, we present a 51-year-old female with mixed phenotype acute leukemia who had a hematologic relapse 3 months after she received total body irradiation-based myeloablative allogeneic hematopoietic stem cell transplantation from an unrelated human leukocyte antigen matched (10/10) donor and achieved complete remission with minimal residual disease negativity by multi-parameter flow cytometry using the combination of blinatumomab and DLI. To the best of our knowledge, this is the first report to describe the use of blinatumomab and DLI combination therapy in the treatment of B/myeloid mixed phenotype acute leukemia.

Peritoneal Cavity Cells Ameliorate Graft-Versus-Host Disease of Mice after MHC Mismatched Bone Marrow Transplantation

Background: Peritoneal cavity cells are a group of isolated cells with unique functions, they have great anti-inflammatory, immune modulation and tissue recovering abilities. Several studies have independently reported the therapeutic function of peritoneal cavity cells on mice models with ulcerative colitis or other inflammatory bowel diseases. In the current study, we further investigated the effect of peritoneal cells on treatment of GVHD on mice models.Methods: We selected a dose of 7.0Gy of TBI for BABL/c mice. And 4 hours after TBI, BABL/c mice were infused with 5*106BMCs cells or 1*106spleen cells of C57BL/6 mice through tail veins to construct MHC mismatched myeloablative allogeneic hematopoietic stem cell transplantation model. The mice were randomly separated into two groups, which were infused with normal saline and 5* 106peritoneal cavity cells on day 0, 3, and 5. Survival time, weight changes, GVHD score were evaluated. Peritoneal cavity cells of GFP C57BL/6 transgenic mice were injected into mice recipients at day 5 after transplantation to observe the distribution of GFP positive cells in mice recipients. Flow cytometry was used to test proportion of CD4/CD8 cells and proportions of effector T cells and Naïve T cells in CD4 and CD8 cells of spleen and bone marrow in BMT group and peritoneal cavity cells group at different time points after transplantation. TNF-α, IFN-γ, IL-2, IL-17, IL-15, IL-4, IL-10 and TGF-β in plasma were studied with the method of Luminex at different time points in two groups. Expression of IL-10 and TGF-β in colons, intestines, and livers were assessed with immunofluorescence staining.Results: Mice in peritoneal cavity cells group had significant longer survival time and rapidly weight loss recover. Mice in peritoneal group had better performance in activity, unhairing, and skin changes as well. Counts of blood cells and chimeric status at day 7, 14, 21, 28 after transplantation showed that blood count recovered and stable chimerism in both groups. Small living animal imaging technology found that peritoneal cavity cells concentrate in colons and intestines after injection of GFP C57BL/6 transgenic mice peritoneal cavity cells. Fluorescence microscope showed that large amounts of green fluorocyte distributed mostly in colons and intestines, with few in liver. Flow cytometry proved that many GFP positive cells in intestines and colons (30%, and 15%, respectively), and a few in livers and lungs (approximately 5%), while negative in control group. We analyzed the lymphocyte subsets of spleen and bone marrow in two groups with flow cytometry and found that peritoneal cells treatment could increase the proportion of CD4 cells and decrease CD8 cells. In CD4 subsets, proportion of effective T cell decreased apparently 3 weeks after transplantation, and count of naïve T cells increased, which is not found in BMT group. Flow cytometry also showed that proportion of Treg cells, Th2 cells and NK cells were significantly higher in peritoneal cavity cells group, while proportion of Th1 cells were lower. TGF-β, IL-10 and IL-4 were significantly higher in peritoneal cavity cell treatment group, while TNF-α, IFN-γ, IL-15, IL-2 were lower. Immunofluorescence staining also showed that TGF-β and IL-10 were strongly expressed in colons and intestines, but not in BMT group.Conclusion: These results demonstrate that peritoneal cavity cells could ameliorate graft-versus-host disease of mice after MHC mismatched bone marrow transplantation. Survival time was prolonged, and weight loss, GVHD score, and pathologic injuries to tissues improved after infusion of 5* 106peritoneal cavity cells into BMT mice. Peritoneal cavity cells injected to BMT mice concentrate mainly in colons and intestines, which functioned as anti-inflammation and tissue repairing cells. These cells could modulate differentiation of T lymphocytes of mice recipients, decrease proportion of CD8 cells and increase CD4 cells, increase proportion of Tn cells and decrease that of Te cells in CD4 subsets, and increase proportion of Tregs, Th2 and NK cells and decrease that of Th1 cells. Peritoneal cavity cells could influence levels of cytokines by increasing anti-inflammatory factors including TGF-β, IL-10, IL-4, and significantly decreasing inflammatory factors like IFN-γ, TNF-α, and IL-15. DisclosuresLiu:West China Hospital of Sichuan University: Employment.

Long-term Effects of Myeloablative Allogeneic Hematopoietic Stem Cell Transplantation in Pediatric Patients with Acute Lymphoblastic Leukemia

Allogeneic hematopoetic stem cell transplantation (HSCT) is a curative option for children and adolescents with high-risk leukemia. Although acute complications were reduced during the last decade, considerable late effects are still limiting the overall success rate. This article emphasizes the specific pediatric aspects of long-term aftercare following myeloablative HSCT and provides an organ-based overview that covers main clinical patterns, incidence, and risk factors enhanced by current references and screening guidelines. In the last years, several attempts were made to separate pediatric outcome data from findings in adults. It turned out that not only the indication for but also the time and the procedures of HSCT substantially differ. Nearly any organ might be affected after the complex transplantation process and includes endocrinopathies, musculoskeletal disorders, cardiopulmonary complications, and secondary malignancies. Patients after HSCT in childhood have a high risk for developing a wide range of late sequelae and may benefit from regular screening and early intervention. The occurrence and patterns of late effects depend on the intensity and severity of conditioning and are strongly associated with patient's age at transplant and beginning of complications.

Upper gastrointestinal acute graft-versus-host disease adds minimal prognostic value in isolation or with other graft-versus-host disease symptoms as currently diagnosed and treated.

Upper gastrointestinal acute graft-versus-host disease is reported in approximately 30% of hematopoietic stem cell transplant recipients developing acute graft-versus-host disease. Currently classified as Grade II in consensus criteria, upper gastrointestinal acute graft-versus-host disease is often treated with systemic immunosuppression. We reviewed the Center for International Blood and Marrow Transplant Research database to assess the prognostic implications of upper gastrointestinal acute graft-versus-host disease in isolation or with other acute graft-versus-host disease manifestations. 8567 adult recipients of myeloablative allogeneic hematopoietic stem cell transplant receiving T-cell replete grafts for acute leukemia, chronic myeloid leukemia or myelodysplastic syndrome between 2000 and 2012 were analyzed. 51% of transplants were from unrelated donors. Reported upper gastrointestinal acute graft-versus-host disease incidence was 12.1%; 2.7% of recipients had isolated upper gastrointestinal acute graft-versus-host disease, of whom 95% received systemic steroids. Patients with isolated upper gastrointestinal involvement had similar survival, disease-free survival, transplant-related mortality, and relapse as patients with Grades 0, I, or II acute graft-versus-host disease. Unrelated donor recipients with isolated upper gastrointestinal acute graft-versus-host disease had less subsequent chronic graft-versus-host disease than those with Grades I or II disease (P=0.016 and P=0.0004, respectively). Upper gastrointestinal involvement added no significant prognostic information when present in addition to other manifestations of Grades I or II acute graft-versus-host disease. If upper gastrointestinal symptoms were reclassified as Grade 0 or I, 425 of 2083 patients (20.4%) with Grade II disease would be downgraded, potentially impacting the interpretation of clinical trial outcomes. Defining upper gastrointestinal acute graft-versus-host disease as a Grade II entity, as it is currently diagnosed and treated, is not strongly supported by this analysis. The general approach to diagnosis, treatment and grading of upper gastrointestinal symptoms and their impact on subsequent acute graft-versus-host disease therapy warrants reevaluation.