AbstractAllogeneic Stem Cell Transplantation Research Articles

Transplant in AML with measurable residual disease: proceed or defer?

Allogeneic stem cell transplantation plays a central role in the management of fit adults with high-risk acute myeloid leukemia (AML) in first complete morphologic remission (CR1). Advances in both donor selection and transplant technology have both dramatically increased accessibility of transplant and led to significant reductions in transplant-related mortality over the past 2 decades. There has, however, been no concomitant reduction in the risk of disease relapse, which remains the major cause of transplant failure. Pivotal to the design of innovative strategies with the potential to reduce relapse risk is accurate identification of patients at the highest risk of disease recurrence. Multiple retrospective studies have identified an increased risk of disease relapse in patients allografted for AML in CR1 with evidence of pretransplant measurable residual disease (MRD). The prognostic significance of pretransplant MRD has been confirmed recently in prospective analyses. The optimal management of patients with evidence of pretransplant MRD remains a matter of conjecture with regard to 2 key issues. First, should the presence of pretransplant MRD delay a decision to proceed to transplant, allowing time for delivery of additional MRD-directed therapy prior to transplant? Second, to what extent can the intensity of the conditioning regimen or the magnitude of the graft-vs-leukemia effect be manipulated to improve the outcome of such patients?

CD34+ -selected hematopoietic stem cell transplant conditioned with a myeloablative regimen in patients with advanced myelofibrosis.

Allogeneic hematopoietic stem cell transplantation (Allo-HCT) remains the only curative treatment for myelofibrosis (MF). Transplantation in patients with MF is mostly done using a reduced intensity conditioning regimen with calcineurin inhibitors for graft versus host disease (GVHD) prophylaxis. Here we sought to evaluate outcomes of patients who underwent an ex vivo CD34+ -selected allo-HCT using myeloablative conditioning (MAC). Twenty-seven patients were included in this retrospective analysis. All patients were conditioned with busulfan, melphalan and fludarabine and antithymocyte globulin to prevent graft rejection. G-CSF mobilized peripheral blood stem cell grafts were depleted of T-cells using immunomagnetic CD34+ selection by CliniMACS device. Median follow-up among survivors was 50.6 months. The estimated 3-year overall survival, relapse free survival and the combined endpoint of GVHD/relapse free survival were 88% (95% CI, 75-100%), 80% (95% CI, 66-98%) and 74% (95% CI, 59-93%), respectively. The cumulative incidence of grade II-IV acute GVHD at day 100 was 33.3% (95% CI 16.4-51.3%), and two patients suffered chronic GVHD. There were no cases of primary graft failure. However, delayed graft failure occurred in two patients. We conclude that CD34+ selected allo-HCT with a MAC resulted in high survival rates in this cohort of patients with MF.

The role of γδ T cells in the context of allogeneic stem cell transplantation

Allogeneic stem cell transplantation is currently the only curative approach for a variety of malignant and non-malignant diseases. In the early transplant era, the intent of this treatment was to apply an intensive myeloablative regimen to eliminate residual malignant cells followed by the hematopoietic rescue of the patients with donor hematopoietic stem cells. However, the focus has shifted over time and allogeneic transplantation is nowadays seen as a cellular therapy in which the donor-derived immune system mounts an anti-infectious and especially an anti-tumor effect in the posttransplant phase. In order to further augment the anti-tumor effect, various approaches have been developed, including the manipulation of the donor-derived immune system in vivo or the adoptive transfer of ex vivo-expanded donor-derived effector cells. Based on their lack of alloreactivity, γδ+ T cells are shifting into the spotlight of research in the context of allogeneic transplantation. Their exploitation with regard to their anti-infectious and anti-tumor properties and their in vivo and ex vivo manipulation will lead to new therapeutic approaches to improve the outcome of patients after allogeneic stem cell transplantation. In this review, the important role of γδ+ T cells in allogeneic matched and mismatched transplantation is summarized and an outlook is discussed on how to best make use of this unique cell population.

Reliability and validity of ultrasound to measure of muscle mass following allogeneic hematopoietic stem cell transplantation

Allogeneic hematopoietic stem cell transplantation (allo-HSCT) recipients frequently show physical dysfunction due to loss of muscle mass. This study aimed to clarify the reliability and validity of ultrasound in evaluating muscle mass and to analyze the patterns of change in muscle mass before and after allo-HSCT. We conducted a prospective observational study using data from 68 patients who had undergone their first allo-HSCT. We evaluated the thickness of the quadriceps, biceps, and suprahyoid muscle. Three individual evaluators underwent this examination for each muscle before transplantation and on days 30, 90, and 180 after allo-HSCT. Inter-rater reliability was calculated using the interclass correlation (ICC), and the level of correlation between muscle mass measured by ultrasound and psoas muscle mass assessed using computed tomography (CT) was assessed using Pearson correlation. ICC values ranged from 0.897 to 0.977 in the measurement. The correlation scores were 0.730, 0.546 and 0.579 between psoas muscle and the biceps, quadriceps, and suprahyoid muscle. The thickness of the biceps and quadriceps muscle were both significantly decreased after allo-HSCT from baseline. These results showed that the ultrasound technique was a reliable tool for evaluating muscle mass and detecting changes in muscle mass following allo-HSCT.

Long-Term Effects of Allogeneic Hematopoietic Stem Cell Transplantation on Systemic Inflammation in Sickle Cell Disease Patients.

Allogeneic hematopoietic stem cell transplantation (allo-HSCT) is the only currently available curative treatment for sickle cell disease (SCD). However, the effects of HSCT on SCD pathophysiology are poorly elucidated. Here, we assessed red blood cell (RBC) adhesiveness, intensity of hemolysis, vascular tone markers and systemic inflammation, in SCD patients treated with allogeneic HSCT. Thirty-two SCD patients were evaluated before and on long-term follow-up after HSCT. Overall survival was 94% with no severe (grade III-IV) graft-vs-host disease and a 22% rejection rate (graft failure). Hematological parameters, reticulocyte counts, and levels of lactate dehydrogenase (LDH), endothelin-1 and VCAM-1 normalized in SCD patients post-HSCT. Expression of adhesion molecules on reticulocytes and RBC was lower in patients with sustained engraftment. Levels of IL-18, IL-15 and LDH were higher in patients that developed graft failure. Increased levels of plasma pro-inflammatory cytokines, mainly TNF-α, were found in SCD patients long-term after transplantation. SCD patients with sustained engraftment after allo-HSCT showed decreased reticulocyte counts and adhesiveness, diminished hemolysis, and lower levels of vascular tonus markers. Nevertheless, systemic inflammation persists for at least five years after transplantation, indicating that allo-HSCT does not equally affect all aspects of SCD pathophysiology.

A novel Iowa\u2013Mayo validated composite risk assessment tool for allogeneic stem cell transplantation survival outcome prediction

Allogeneic hematopoietic stem cell transplantation (HSCT) is a curative option for many hematologic conditions and is associated with considerable morbidity and mortality. Therefore, prognostic tools are essential to navigate the complex patient, disease, donor, and transplant characteristics that differentially influence outcomes. We developed a novel, comprehensive composite prognostic tool. Using a lasso-penalized Cox regression model (n = 273), performance status, HCT-CI, refined disease-risk index (rDRI), donor and recipient CMV status, and donor age were identified as predictors of disease-free survival (DFS). The results for overall survival (OS) were similar except for recipient CMV status not being included in the model. Models were validated in an external dataset (n = 378) and resulted in a c-statistic of 0.61 and 0.62 for DFS and OS, respectively. Importantly, this tool incorporates donor age as a variable, which has an important role in HSCT outcomes. This needs to be further studied in prospective models. An easy-to-use and a web-based nomogram can be accessed here: https://allohsctsurvivalcalc.iowa.uiowa.edu/.

Late Post-Operative Occurrence of Dentin Hypersensitivity in Adult Patients Following Allogeneic Hematopoietic Stem Cell Transplantation-A Preliminary Report.

Allogeneic hematopoietic stem cell transplantation (alloHSCT) is one of the most commonly performed transplantation procedures nowadays. Despite the significant progress made in the treatment, alloHSCT is still associated with numerous complications also affecting the oral cavity. One of them is dentin hypersensitivity (DH)—a sharp, short-term pain that occurs when stimuli act on exposed dentin. Various authors point out that DH may result in a significantly lower quality of life, among other things by impeding the consumption of food as well as causing difficulties in daily oral hygiene. The aim of the study was a preliminary analysis of the incidence rate and severity of DH pain in adult patients during late period after alloHSCT. The impact of chronic graft-versus-host disease (cGvHD) and time after alloHCT were also considered. A total of 80 patients were examined. cGvHD was identified in 52 participants. The incidence rate and severity of DH pain was assessed on the basis of a questionnaire and a clinical examination. DH pain appeared a serious problem in late period after alloHSCT regardless of post-transplant time. DH primarily affected cGvHD patients. The prevention-treatment protocol for DH should be developed for this group.

Clinically applicable CD34+-derived blood dendritic cell subsets exhibit key subset-specific features and potently boost anti-tumor T and NK cell responses

Allogeneic stem cell transplantation (alloSCT), following induction chemotherapy, can be curative for hemato-oncology patients due to powerful graft-versus-tumor immunity. However, disease recurrence remains the major cause of treatment failure, emphasizing the need for potent adjuvant immunotherapy. In this regard, dendritic cell (DC) vaccination is highly attractive, as DCs are the key orchestrators of innate and adaptive immunity. Natural DC subsets are postulated to be more powerful compared with monocyte-derived DCs, due to their unique functional properties and cross-talk capacity. Yet, obtaining sufficient numbers of natural DCs, particularly type 1 conventional DCs (cDC1s), is challenging due to low frequencies in human blood. We developed a clinically applicable culture protocol using donor-derived G-CSF mobilized CD34+ hematopoietic progenitor cells (HPCs) for simultaneous generation of high numbers of cDC1s, cDC2s and plasmacytoid DCs (pDCs). Transcriptomic analyses demonstrated that these ex vivo-generated DCs highly resemble their in vivo blood counterparts. In more detail, we demonstrated that the CD141+CLEG9A+ cDC1 subset exhibited key features of in vivo cDC1s, reflected by high expression of co-stimulatory molecules and release of IL-12p70 and TNF-α. Furthermore, cDC1s efficiently primed alloreactive T cells, potently cross-presented long-peptides and boosted expansion of minor histocompatibility antigen-experienced T cells. Moreover, they strongly enhanced NK cell activation, degranulation and anti-leukemic reactivity. Together, we developed a robust culture protocol to generate highly functional blood DC subsets for in vivo application as tailored adjuvant immunotherapy to boost innate and adaptive anti-tumor immunity in alloSCT patients.

Umbilical Cord Blood as an Alternate Donor Sources for High Risk Elderly Patients Undergoing Allogeneic Stem Cell Transplantation for Hematological Malignancies

Allogeneic stem cell transplantation remains the only curative option for many hematological malignancies. Umbilical cord blood (UCB) is an alternate donor source with potentially increased morbidity in elderly patients. We evaluated outcomes in alternate donor sources, prior to the initiation of haploidentical transplantation at our institution, of matched unrelated donor (MUD) and UCB in elderly patients (mean age 64, range 60-75). One hundred and eighty-four patients were included (MRD: 57; MUD: 69; UCB: 58). There was no difference in acute or chronic graft versus host disease among donor sources (all p > .05). In this high-risk population, 128 (70%) had either a high disease risk index or high (>2) comorbidity index. Median progression free survival (PFS) was lowest among UCB (5.5 months; 95% CI 2.8-9.4) and MUD (5.6 months; 95% CI 3.7-17.7) as compared to MRD (18.3 months; 95% CI 8.4-64.8). On multivariable analysis, the rate of mortality was higher for UCB than MRD patients (1.88, 95% CI 1.04 – 3.37), but there was no difference between UCB and MUD (p = .61) or between MUD and MRD (p = .25). Conclusions were similar for PFS. Our experience shows that even in this high-risk elderly cohort, UCB continues to be a viable alternate donor source.

Gene expression signatures associated with sensitivity to azacitidine in myelodysplastic syndromes

Allogeneic stem cell transplantation is currently the only curative treatment option for myelodysplastic syndromes (MDS). Pre-transplant debulking treatment have been employed for advanced MDS and we previously reported that marrow response (blast ≤ 5%) following the bridging therapy with hypomethylating agent was an independent favorable factor for survival; however, it is still not clear which patients will respond to hypomethylating agent and which genomic features can predict the response. In this study, we performed RNAseq for 23 MDS patients among which 14 (61%) and 9 (39%) patients showed marrow complete remission and primary resistance to azacitidine, respectively. Differential expression-based analyses of treatment-naive, baseline gene expression profiles revealed that molecular functions representing mitochondria and apoptosis were up-regulated in responders. In contrast, we identified genes involved in the Wnt pathway were relatively up-regulated in non-responders. In independent validation cohorts of MDS patients, the expression of gene sets specific to non-responders and responders distinguished the patients with favorable prognosis and those responded to azacitidine highlighting the prognostic and predictive implication. In addition, a systems biology approach identified genes involved in ubiquitination, such as UBC and PFDN2, which may be key players in the regulation of differential gene expression in treatment responders and non-responders. Taken together, identifying the gene expression signature may advance our understanding of the molecular mechanisms of azacitidine and may also serve to predict patient responses to drug treatment.

Value of pre-transplant consolidation chemotherapy in adults with acute lymphoblastic leukemia undergoing allogeneic hematopoietic stem cell transplantation without minimal residual disease in first complete remission

Allogeneic hematopoietic stem cell transplantation (allo-HSCT) is recommended for adults acute lymphoblastic leukemia (ALL) with minimal residual disease (MRD) negative during their first complete remission (CR1). However, the role of pre-transplant consolidation chemotherapy remains unclear. We evaluated 78 CR1/MRD-negative patients, the consolidation and non-consolidation groups had similar 5-year OS (74.8% [95% CI: 62.2–87.3%] vs. 74.2% [95% CI: 53.2–95.1%], p = .894), RFS (72.2% [95% CI: 59.6–84.7%] vs. 73.1% [95% CI: 54.2–91.9%], p = .942), CIR (9.4% [95% CI: 9.1–9.7%] vs. 18.9% [95% CI: 17.3–20.4%], p = .376), and NRM (18.4% [95% CI: 17.7–19.0%] vs. 8.0% [95% CI: 7.3–8.6%], p = .375). Multivariable analysis confirmed that high cytogenetic risk independently predicted poor OS and RFS, although pre-transplant consolidation chemotherapy did not predict the prognosis. Based on these findings, we recommend performing transplantation immediately for adult ALL patients after they have achieved CR1/MRD-negative status when there are readily available donors.

Hematological Complications of Donor Origin Following Allogeneic Hematopoietic Stem Cell Transplantation

Allogeneic hematopoietic stem cell transplant (HSCT) offers curative therapy for some malignant hematological disease and marrow failure states. Post-transplant donor cell pathology, which might become more prevalent in long term survivors of allogeneic HSCT, warrants further study. We report five recipients of allogeneic HSCT from our institution who developed donor cell abnormalities, discuss potential mechanisms underlying these disorders and propose potential avenues for future study. Case 1 involves a 43-year-old female who developed leukopenia and adenopathy and was subsequently found to have diffuse large B-cell lymphoma. She received a matched unrelated donor (MUD) SCT from a 23-year-old male and subsequently developed a reactive lymphocytosis. In case 2, a 44-year-old male with myelodysplastic syndrome (MDS) received an allogenic peripheral blood (PB) SCT from his 50-year-old sister and later developed donor-cell derived acute myeloid leukemia (AML) with chimerism studies demonstrating 100% donor cells in CD3, CD33/66 and CD56. Case 3 involved a 72-year-old female with AML who received a PBSCT from her 65-year-old brother and subsequently developed MDS with peripheral blood chimerism studies demonstrating 100% donor cells in all three compartments. Case 4 involved a 40-year-old female with AML who achieved molecular remission following chemotherapy and underwent a double umbilical cord transplant, subsequently developing donor-cell derived AML with chimerism studies negative for host cells. Finally, case 5 highlighted a 54-year-old male with AML who underwent a MUDSCT from a 44-year-old male and passed away from progressive leukemia. Peripheral blood demonstrated only donor cells with near identity of phenotypic and molecular studies. It remains possible that no malignant cells exited the marrow. Nonetheless, this case in particular highlights the difficulty in precisely diagnosing donor cell leukemia (DCL). DCL correlates with increasing donor age, suggesting that age-related mutation in hematopoietic stem cells might be the cause. Notably, the average donor age in our case studies was 53 years old. Hematopoiesis is maintained by both hematopoietic stem and progenitor cells and during the normal aging process becomes clonally restricted. Clonal hematopoiesis of indeterminate potential (CHIP) describes the presence of a clonal blood cell population associated with a driver mutation at a VAF ≥ 2%, in the absence of severe cytopenia or a WHO defined disorder. These mutations are found in 5% of individuals greater than 50 years of age, with the most common gene mutations being found inDNMT3A, ASXL1,andTET2. The rate of development of overt neoplasia in patients with CHIP is 0.5% to 1% per year and these patients are 4 to 15 times more likely to develop a hematologic malignancy. CHIP mutations might confer a survival advantage allowing these clones to expand under the influence of a selection pressure such as chemotherapy or immunosuppression. These surviving clones might be the origin of DCL. DCL is estimated to account for approximately 1-5% of all post-transplant leukemia. When transplanted, expansion of these clones as required for repopulation of the hematopoietic stem cell niche might accelerate the acquisition of secondary mutations providing surviving mutated clones a survival advantage leading to malignancy. An open question remains particularly as it concerns older donors >55 years, as to whether they should be screened by next generation sequencing (NGS) for the presence of mutations known to be associated with CHIP or frank hematologic malignancy. If the incidence of DCL increases, consideration should be given to subjecting older donors to these screening methods. When multiple donors of different age are available preference can be assigned to younger donors to avoid possibly transmitting CHIP. Disclosures Fazal: BMS:Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Speakers Bureau;Jansen:Speakers Bureau;Novartis:Consultancy, Speakers Bureau;Celgene:Speakers Bureau;Gilead/Kite:Consultancy, Speakers Bureau;Glaxosmith Kline:Consultancy, Speakers Bureau;Agios:Consultancy, Speakers Bureau;Takeda:Consultancy, Speakers Bureau;Incyte Corporation:Consultancy, Honoraria, Speakers Bureau;Karyopham:Speakers Bureau;Amgen:Consultancy, Speakers Bureau;Stemline:Consultancy, Speakers Bureau;Jazz Pharma:Consultancy, Speakers Bureau.

Analysis of determinant factors of liver fibrosis progression in ex-thalassemic patients.

Allogeneic hematopoietic stem cell transplantation (allo-HSCT) potentially renders thalassemia patients disease-free with presumably cessation of associated complications. This study analyzes the liver fibrosis status and the determinants of its progression in ex-thalassemic patients. The liver fibrosis status of 108 pediatric transfusion-dependent β-thalassemia major patients was evaluated before and one year after allo-HSCT using transient elastography (TE). All patients achieved normal hematopoiesis. In univariate analyses, not in all, but in patients developing significant post-HSCT iron overload or hepatic graft-versus-host disease (GvHD), as well as recipients of bone marrow stem cells (BMSC), significant TE increment occurred. In multivariable analyses, through a model with large effect size (Adj.R2 = 26%, F(3,104) = 13.53, P < 0.001), post-HSCT serum ferritin and hepatic GvHD were ascertained as independent determinants of significant TE increase, and the effect of stem cell graft source approached the level of significance. Excluding the patients with intermediate/high Lucarelli risk classes, the TE increase was significantly greater only in BMSC recipients (P = 0.033). Although the risk impact of allograft source on liver fibrosis progression requires further evaluation; hepatic status of ex-thalassemic patients can be preserved after HSCT, if hepatic GvHD is controlled and adequate post-transplantation iron depletion is ensured.

2020 – ACTIVATION OF THE RECEPTOR TYROSINE KINASE (RET) BY GDNF/GFRa1 IMPROVES CORD BLOOD-DERIVED HSC IN VITRO EXPANSION AND IN VIVO ENGRAFTMENT

Allogeneic hematopoietic stem cell (HSC) transplantation is the gold standard therapy for numerous hematological disorders, but limitations exist in HSC number available per donor and our relative inability to successfully expand transplantable HSCs in vitro. In recent years, promising new molecules have been discovered, which are able to expand HSCs in vitro and improve transplantation in vivo in immunodeficient mouse models and non-human primates (e.g. SR1 and UM171). Here we demonstrate that acute activation of the receptor tyrosine kinase, RET, by its key ligand and co-receptor (GDNF/GFRa1), significantly improves the outgrowth of HSCs in vitro and long-term engraftment in immunodeficient Kit-mutant (KitW41/W41) mice. The expression of RET at the cell surface of cord blood-derived hematopoietic stem and progenitor cells (HSPCs, CD34+CD38-REThi) enriches for stem cell frequency in limiting dilution transplantation assays (∼1 in 135), with CD34+CD38-RETlow cells showing low stem cell frequency with limited multilineage engraftment (∼1 in 531). Activation of RET in vitro with a single dose of GDNF/GFRa1 reduces apoptosis at early time points, leading to increased HSC expansion over a 7-day period (>4-fold increase vs control). Transplantation of GDNF/GFRa1 treated HSPCs significantly improves engraftment of primary mice, comparable to the previously published SR1/UM171 treatment, and combination treatment of GDNF/GFRa1 with SR1/UM171 improves engraftment compared to individual GDNF/GFRa1 or SR1/UM171 treatments alone. RET is a single pass transmembrane receptor tyrosine kinase, governing a variety of downstream signalling pathways. Interrogation of the dynamic kinome changes governed by RET in HSPCs, using PAMGene kinome array kinetic technology and single cell mass cytometry revealed key phosphorylation cascades responsive to GDNF/GFRa1 treatment. Key cascades including anti-apoptotic p53 phosphorylation, NFkB activation and reduced reactive oxygen species, revealed important cellular programs for HSC survival, outgrowth and engraftment activated by GDNF/GFRa1. Our results provide a promising new target to improve the outgrowth of transplantable HSCs for clinical use, with multiple downstream functions activated for successful transplantation.

Allogeneic transplantation of hematopoetic stem cells in acute leukemia in children, adolescents and young adults in the Republic of Belarus

Allogeneic hematopoietic stem cell transplantation (HSCT) is a recognized method for treating children with a very high risk group for acute lymphoblastic leukemia (ALL) and a high risk group for acute myeloid leukemia (AML). The use of allogeneic HSCT for certain risk groups of acute leukemia significantly improves the survival of these patients compared to chemotherapeutic regimens. The aim of this study was to identify the causes of failure of HSC transplantation in children with acute leukemia in a homogeneous group of patients and the possibility of further improvement in survival rates. The study was approved by the Independent Ethics Committee and the Scientific Council of the Belarusian Research Center for Pediatric Oncology, Hematology and Immunology (Republic of Belarus). The study included 101 patients with ALL and 65 patients with AML who underwent the first HSCT, in accordance with the first-line treatment protocol or relapse for 2 consecutive time periods (1998–2008 and 2009–2018). For the entire group of patients, an increase in overall (by 13%) and event-free survival (by 7%) was revealed due to a decrease in post-transplant mortality not related to relapse by 16% (p = 0.077). Significant improvement in survival over time occurred in the group of patients with acute or chronic “graft versus host” disease. The data obtained indicate that all patients with acute leukemia who have indications for HSCT in the first line of treatment or relapse should be transplanted from any available donor, as this will significantly increase their chances of recovery.

The influence of various doses of busulfan in conditioning regimes on outcome of allogeneic hematopoietic stem cell transplantation in children with acute myeloid leukemia

Allogeneic hematopoietic stem cell transplantation (allo-HSCT) is a potentially curative therapy for patients with acute myeloid leukemia (AML). The conditioning regimen administered for this patient based on busulfan (Bu) combined with cyclophosphamide (Cy), fludarabine (Flu) or some other agents. Comparisons of myeloablative conditioning (MAC) versus reduced intensity conditioning (RIC) have demonstrated a various results between relapse and toxicity in a few reports. We suppose, that dose intensity of Bu across regimens may affect treatment outcomes. Aim of this retrospective study was to evaluate the impact dose of busulfan to overall survival (OS), transplant-related mortality (TRM), relapse-free survival (RFS), toxicity, the incidence of primary graft failure and acute "graft versus host" disease (GvHD) in transplantation in children and adolescents with AML. The study was approved by the Independent Ethics Committee and the Scientific Council of the I.P. Pavlov First Saint Petersburg State Medical University. We analyzed 110 AML pediatric patients with the median age 9 (range 1–19) y.o., who underwent first allo-HSCT with Bu based conditioning in R.M. Gorbacheva Memorial Institute from 2002 to 2018. Patients were divided into 3 groups: Bu1 – patients, who received Bu at the dose 8–10 mg/kg, n = 34 (31%), in Bu2 – dose of Bu was 12 mg/kg, n = 35 (32%), in Bu3 – dose of Bu was &gt; 12 mg/kg, n = 41 (37%). In Bu1 Bu was combined with Flu in 31 (91%) pts and Cy in 3 (9%); in Bu2 – with Flu in 12 (34%), Cy in 7 (20%) and other agents in 16 (46%); in Bu3 – with Cy in 32 (78%), with Flu in 7 (17%) and other agents in 2 pts (5%) (p &lt; 0.001). Patients in Bu2 received more Cy based GvHD prophylaxis regimens (69% vs 44% in Bu1, vs 29% in Bu3, p = 0.003) and more haplo grafts (51% vs 29% in Bu1, vs 15% in Bu3, p = 0.003). The complete remission at the HSCT was observed in 79 % in Bu1, 49% in Bu2, 61% in Bu3 (p = 0.02). Probabilities of OS, RFS, TRM were estimated by using the Kaplan–Meier method. Incidence of toxicity, acute GvHD and primary graft failure – by using Mann–Whitney U-test. Transplant engraftment was achieved in 95 (86%) of patients. Graft failure occurs in the 5 patients of Bu1 group (15%), in the 6 pts of Bu2 (17%) and in the 4 pts of Bu3 (10%) (p = 0.7). Median follow-up was 2 years for Bu1 and Bu3, 1 year for Bu2. Two-year OS was similar (Bu1 = 59% vs Bu2 = 60% vs Bu3 51%, p = 0.7). Two-year OS of pts with CR before HSCT was 70% in Bu1, 82% in Bu2, 60% in Bu3, p = 0,3 and 14%, 39%, 38% for pts with progression disease (PD), respectively (p = 0.5). Two-year RFS was 74% in Bu1, 82% in Bu2, 64% in Bu3 at CR (p = 0.4); 43%, 39% and 38% in pts with progression, respectively (p = 0.9). Median of RFS were also similar for the pts in PD (4 months in Bu1, 5 months in Bu2 and Bu3, p = 0.9) and not achieved for pts at CR. Drug related toxicity grade III–IV 4 experienced in 35% pts in Bu1, 29% in Bu2, in 54% in Bu3 (p = 0.04). Mucositis and toxic hepatitis were the most common adverse events. Sinusoidal obstruction syndrome (SOS) experienced in 8 pts from different group: 4 from Bu2 (11%), 3 from Bu3 (7%) and only pts from Bu1 (3%) with previously treated of inotuzumab (p = 0.4). The most pts with VOD (3/5) had PD at the HSCT. Cumulative incidence of acute GvHD grade 2 (15% vs 14% vs 10%, p = 0.8) were not different. Acute GvHD grade III–IV was observed a bit more often in Bu3 (34%), than in Bu1 (18%) and Bu2 (17%) (p = 0.09). TRM up to D+100 was also higher in Bu3 (15%), than in Bu2 (6%) and Bu1 (0%) (p = 0.05). The transplant results of children with similar disease status of AML, received MAC or RIC conditioning with various dose of Bu, were not associated with significant differences in overall outcomes. The higher dose Bu may increase incidence of toxicity grade III–IV (p = 0.04) and acute GvHD grade III–IV (p = 0.09) with increasing of early TRM (p = 0.05).

Posttransplant cyclophosphamide as a platform for immunotherapy after allogeneic stem cell transplantation for multiple myeloma

Abstract Allogeneic stem cell transplantation (alloSCT) is a highly effective, curative therapy for leukemia yet does not provide a survival benefit above autologous SCT in patients with multiple myeloma (MM). To explore this, we developed preclinical models of SCT using C57Bl/6 recipient mice and either C57Bl/6 (ASCT) or C3H.SW (alloSCT) donor grafts. Importantly, these models recapitulated the clinical setting whereby alloSCT provided superior outcomes, compared to ASCT, in recipients bearing MLL-AF9-driven acute myeloid leukemia (AML) but not in those bearing Vk*MYC-MM. Interestingly, we found that MM-specific, T cell-mediated immunity was generated after ASCT, which failed due to MM-induced T cell exhaustion. MM relapse after ASCT could be prevented by TIGIT or PD-1 targeted immune checkpoint inhibition or via depletion of suppressive CSF1-R+ myeloid cells. Conversely, after alloSCT, T cell exhaustion was driven principally by alloantigen and CD8 T cells expressed high levels of PD-1, TIGIT and TIM-3. Furthermore, Vk*MYC myeloma exploited this alloantigen-driven T cell exhaustion via expression of high levels of PD-L1 and CD155 (the cognate ligands for PD-1 and TIGIT), which were expressed minimally on MLL-AF9 AML. To exploit alloSCT in MM, we used post-transplant cyclophosphamide (PT-Cy) to delete high affinity alloreactive T cells that generate the exhausted donor T cell pool. Subsequent administration of CD137 agonists enhanced T cell activation and cytolytic activity within bone marrow, without exacerbating GVHD. Thus, PT-Cy provides a platform for optimizing immunotherapy after alloSCT.

Diverse IL-6 signalling modalities drive pathogenic T cell differentiation and graft-versus-host-disease after allotransplantation

Abstract Allogeneic stem cell transplantation (alloSCT) and graft-versus-host disease (GVHD) are characterized by systemic interleukin 6 (IL-6) dysregulation, which plays a significant role in shaping donor immune responses and T cell polarization. GVHD is a T cell-mediated disease and the severity and tissue distribution is heavily influenced by T cell-derived cytokines, therefore it is critical to understand the factors that drive T cell polarization in this context to inform therapeutic strategies. IL-6 has a unique receptor system composed of IL-6Ra and the signal transducing molecule gp130, in which signaling occurs via multiple pathways either directly (classical), indirectly via a soluble IL-6 receptor (trans), or presented via antigen presenting cells (cluster). We examined the influence of IL-6 signaling modalities on T cell polarization following allotransplantation, where we found specific targeting of these pathways modulates GVHD outcomes. Using donor grafts composed of IL-6Ra deficient T cells resulted in a profound loss of pathogenic Th17/Th22 differentiation and increased GVHD survival, demonstrating these populations are highly dependent upon classical IL-6 signaling post-transplant. Whilst targeting cluster signaling through IL-6Ra deficient DC had no effect on T cell cytokine responses, trans-signaling inhibition via soluble gp130-Fc resulted in severe skin GVHD. This effect was due to significant expansion of pathogenic donor Th22 and was prevented by donor IL-22 deficiency. These data demonstrate an important role for IL-6 trans signaling in regulating pathogenic T cell polarization pathways following allotransplantation and support IL-6 classical signaling as an important target for GVHD prevention.

Cellular therapies

Cellular therapies Abstract. Transfusion medicine and allogeneic stem cell transplantation are well known and established cellular therapies in hematology. Since decades many efforts have been made, in order to re-program the patient's own immune system in order to clear malignancies. A breakthrough was achieved with the manufacturing and optimizing of so-called chimeric antigen receptor (CAR) T-cells, genetically engineered cells, specifically directed against tumor antigens. In this review we discuss the structure of CAR T-cells, their manufacturing and the different steps of a CAR T-cell treatment according to the current licensing. Furthermore, we give an outlook on future prospects of cellular therapies including the major issues in the field.

Long-Term Follow-up of Hematopoietic Stem Cell Transplantation with the Modified Peking Regimen for the Treatment of Acute Leukemia

Allogeneic hematopoietic stem cell transplantation (allo-HSCT) is the only curative treatment strategy for patients with acute leukemia. The ATG-based transplantation system initiated by Peking University people's hospital, known as the Peking regimen, has become a mainstream transplant system worldwide. Here, based on the Peking regimen, we report a modified protocol:(1)add Fludarabine and replace ATG with ATG-F in the conditioning regimen; (2)transfuse higher-dose cell collections from granulocyte-colony stimulating factor(G-CSF) primed bone marrow and peripheral blood samples; (3) add Basiliximab (a CD25-antibody) on day +3 for acute GVHD prophylaxis. In this study, 265 patients (158 patients with haplo-SCT and 107 patients with sibling-SCT) underwent allo-HSCT with our modified protocols. All patients achieved sustained full-donor chimerism. The incidence of grade II-IV and III-IV acute GVHD in haplo-SCT comparing with sibling-SCT was 36.1%(57/158) vs 17.8%(19/107)(P=0.001) and 13.3% (21/158) vs 9.3%(10/107)(P&gt;0.05) respectively. The 2-year cumulative incidence of total chronic GVHD and extensive chronic GVHD in haplo-SCT was 41% (65/158) and 15% (24/158) respectively. The 3-year cumulative incidence of non-relapse mortality (NRM) in haplo-SCT and sibling-SCT was 6.3% (10/158) and 4.7%(5/107) respectively(P&gt;0.05). The 100-day cumulative incidence of CMV viremia in haplo-SCT and sibling-SCT was 35.5% (56/158) and 23.4%(25/107) respectively(P=0.036). A total of 36 patients in haplo-SCT group and 24 patients in haplo-SCT group had recurrent disease, reaching a cumulative incidence of relapse of 20.8% in haplo-SCT and 23.4% in sibling-SCT at 3 years respectively(P&gt;0.05). The relapse ratio of haplo-SCT and sibing-SCT in the 1st year, between the 1st and the 2nd year and after 2 years was 21.5% vs 14.1%(P&gt;0.05), 1.3%(2/158) vs 0%(P&gt;0.05) and 0% vs 6.5%(P=0.009) respectively. The 3-year overall survival(OS) and leukemia-free survival(LFS) rates in haplo-SCT and sibling-SCT was 78.8% vs 74.2% and 76.8% vs 75.04% respectively(P&gt;0.05) by the Kaplan-Meier estimate. The 3-year GVHD-free and leukemia-free survival rates (GRFS) in haplo-SCT and sibling-SCT were 43.4% vs 69.5%(P=0.045) respectively. Lower OS in haplo-SCT was associated with III-IV aucte GVHD and lower MNC(&lt;19×10^8/L) in grafts by Cox regression analysis. In a word, the results from our experience showed that the modified protocol based on the Peking Regimen is safe and reliable for acute leukemia patients and brings on a long-stage survival post transplantation. Disclosures Zheng: Pfizer: Research Funding.