Post-allogeneic Hematopoietic Stem Cell Transplantation Research Articles

High-Dose Melphalan-Based Sequential Conditioning Chemotherapy Followed By Allogeneic Hematopoietic Stem Cell Transplantation (HSCT) in Adult Patients with Relapsed or Refractory Acute Myeloid Leukemia (AML)

Introduction: The majority of patients with newly diagnosed AML fail to achieve long-lasting remission, even after initial complete remission (CR). Response rates to common salvage regimens reach up to 50% in patients with late relapse and decrease to 10-15% in patients with refractory AML or early relapse. Without allogeneic HSCT the long-term overall survival (OS) for patients with refractory AML hardly reaches 5%. Survival rates of AML patients post allogeneic HSCT are significantly higher for patients transplanted in CR as compared to those transplanted with active leukemia. Common conditioning regimens followed by allogeneic HSCT of patients with active AML have shown disappointing results. Therefore, others and we developed the concept of sequential therapy as a preparative regimen for allogeneic HSCT to optimize the balance between treatment-related toxicity and antileukemic efficacy.Methods: From April 2005 to May 2016, 292 patients with R/R-AML received an allogeneic HSCT at the BMT centers of the German university hospitals of Muenster and Essen. The median age was 56 years (range 17 -74). The remission status prior to transplant was primary refractory (144 patients), secondary refractory (97 patients) or relapsed AML (51 patients). All patients were treated with a single infusion of high dose melphalan, followed by dose-adapted conditioning consisting of fludarabine (120 mg/m2) and 8 Gy TBI or followed by fludarabine (150 mg/m2) and treosulfan (42 g/m2). Median time between melphalan and continuation of the therapy was 5 days (range 1-18 days), resulting in an overall duration of conditioning prior to HSCT of 11 days in most patients. Additional pretreatment with antithymocyte globulin (ATG) was applied to 157 of 222 patients with an unrelated donor and in 44 of 70 patients with a HLA-identical sibling donor. Most patients (except 2) received cyclosporine as acute GvHD prophylaxis in combination with methotrexate (89 patients) or mycophenolate mofetil (197 patients) or both (4 patients).Results: Median follow-up after allogeneic HSCT of all patients alive at last follow-up was 4.8 years. OS rates at 3 years were 34% for primary refractory patients, 29% for secondary refractory patients, and 41% for patients with untreated relapsed AML. Factors associated with an inferior OS were older age, a time interval of 5-9 months between first diagnosis and allogeneic HSCT, and transplantation from an HLA-mismatched unrelated donor. Most importantly, leukemic burden was associated inversely with a 3-year OS of 51%, 31%, and 22% for patients with leukemic blast infiltration of <20%, 20-50%, and >50%, respectively (p <.001). Particularly patients transplanted from HLA-mismatched unrelated donors had a significantly higher non-relapse-related mortality (NRM) rate of 55% (95%CI 27 to 56%) at one year, compared to those transplanted from HLA-matched related (34%, 95%CI 24 to 48%) or unrelated donors (28%, 95%CI 22 to 37%). In addition, adverse risk factors for NRM were older age, pre-existing infection prior to start of conditioning therapy and a higher comorbidity index (HCT-CI >3). Multivariate analysis of risk factors revealed that high disease burden before conditioning (p <.001), longer time interval between first diagnosis and transplantation (p .039), and transplantation from an HLA-mismatched unrelated donor (p .049) were independently associated with inferior overall survival. High risk disease according to cytogenetic and molecular genetic aberrations as an adverse risk factor showed a trend in this analysis (p .052).Conclusion: Allogeneic HSCT remains the most potent treatment option for patients with R/R-AML. High-dose melphalan based sequential conditioning chemotherapy followed by an allogeneic HSCT allows for achieving long-term remission in a substantial proportion of patients with active R/R-AML with otherwise detrimental prognosis. Since allogeneic HSCT is superior to other alternative treatment strategies evaluated in the past, direct allogeneic HSCT should be evaluated prospectively against remission induction therapies followed by an allogeneic HSCT in patients with R/R AML. DisclosuresLenz:Celgene, JJ, Hexal, Roche, BMS, Novartis, Bayer: Consultancy; AstraZeneca, Gilead, JJ, Celgene, Novartis, Bayer: Research Funding.

Effects of Gram-negative Rod Blood Stream Infection on Acute GVHD in Allogeneic Hematopoietic Stem Cell Transplantation: A Single-institute Analysis.

A bloodstream infection (BSI) is the most common serious infectious complication of hematopoietic stem cell transplantation (HSCT). BSI promotes an inflammatory state, which exacerbates acute graft-versus-host disease (GVHD). We investigated whether a Gram-negative rod bloodstream infection (GNR-BSI), which develops early after allo-HSCT, affected the onset or exacerbated acute GVHD in 465 patients who underwent allo-HSCT from 1995 through 2015 at a single institution. Eighty-eight patients (19%) developed BSI during the study period. Among the cultures, 50 (57%) were Gram-positive cocci (GPC) and 31 (35%) were GNR. Of the 465 patients, 187 (40%) developed acute GVHD of grade II or higher within the first 100 days post-allogeneic HSCT: 124 (27%) had acute GVHD grade II, 47 (10%) had grade III, and 16 (3%) had grade IV. Multivariate analysis revealed that GNR-BSI was a significant risk factor for grade II-IV acute GVHD (grade II-IV: hazard ratio [HR] 1.75, 95% confidence interval [CI] 1.03-2.97; grade III-IV: HR 2.37, 95% CI 1.03-5.43). These results suggest that GNR-BSI may predict the onset and exacerbation of acute GVHD.

Long-term follow-up of adult hematopoietic stem cell transplant recipients diagnosed with COVID-19.

e19028 Background: Long-term complications of COVID-19 in hematopoietic stem cell transplant (HCT) recipients are unknown. Recent studies have described short term outcomes of COVID-19 infection post allogeneic (allo) and autologous (auto) HCT. In this study we provide long term follow-up of the outcomes of COVID-19 infection in allo and auto HCT recipients. Methods: We performed a retrospective study of adult patients who have received allo or auto HCT and were subsequently diagnosed with COVID-19 infection between March-December 2020. We summarized patient characteristics, laboratory and treatment data related to COVID-19 infection in these patients. Results: In this study, we provide long-term follow-up of over 7 months. Fifteen patients were identified for inclusion (allo n = 12, auto n = 3). Median follow-up was 7.8 months (range 1.9-10.7) for surviving patients. Median age at COVID-19 diagnosis was 55 years (range 24-77). Most patients were &gt; 1 year out from transplant (allo n = 10, auto n = 1, 73%). Two patients (allo n = 1, auto n = 1, 13%) had undergone transplant within the preceding month. Most allo patients (n = 11, 73%) had received myeloablative conditioning. At the time of COVID-19 diagnosis, 9 allo patients (75%) were on immunosuppression (IS) (n = 7 for chronic graft-versus-host-disease (GVHD); n = 2 for GVHD prophylaxis). Eleven patients (73%) required hospitalization (allo n = 9, auto n = 2). Per the National Institutes of Health definitions of COVID-19 illness severity, 3 patients had critical disease (allo n = 2, auto n = 1, 20%), 5 severe (allo n = 5, 33%), 3 moderate (allo n = 2, auto n = 1, 20%), and 4 mild (allo n = 3, auto n = 1, 27%). All patients with chronic GVHD required hospitalization. Two patients died (allo n = 1, auto n = 1, 13%)—both had critical COVID-19 infections, were &gt; 65 years old, &gt; 3 years out from transplant, and had significant comorbidities. The allo patient was receiving prednisone &lt; 1 mg/kg for chronic lung GVHD at COVID-19 diagnosis. Two allo patients developed either acute GVHD or chronic GVHD exacerbation within 3 months of their infection. One patient developed biopsy-proven acute GVHD (max grade III) 3 weeks after her infection and another patient developed a severe exacerbation of chronic GVHD within 3 months—both continue to require multi-modal IS. The remaining 7 patients with chronic GVHD have been maintained on either stable or tapered IS. Conclusions: Given the effect of COVID-19 infection, its impact on HCT recipients is important to define. The majority of HCT patients who contracted moderate-critical COVID-19 infections in our study were either on IS or had significant comorbidities. Our observational data points to the importance of long-term follow-up in HCT patients. Future studies are needed to delineate whether there is a relationship between COVID-19 infection and GVHD development or exacerbation. The role of vaccination in HCT recipients remains to be explored.

Clinical Characteristics of Myositis Associated with Graft-Versus-Host Disease.

Recipients of allogeneic hematopoietic stem cell transplantation (HSCT) are at increased risk for inflammatory myositis; histological subsets reported include dermatomyositis, necrotising myopathy and chronic graft-versus-host disease (cGVHD)-related myositis. Though corticosteroids and various immunosuppressive therapies have been used, there is a lack of consensus guidelines dictating therapy. Recent evidence suggests the fascia as a preferential target in cGVHD myositis, with conditioning regimens promoting fascial microtrauma. Positron emission tomography (PET) can be a useful diagnostic tool, and case reports suggest that the Bruton's tyrosine kinase inhibitor ibrutinib may have therapeutic potential. Emerging therapies include targeted B cell depletion with rituximab, and extracorporeal photophoresis. Clinicians need to be vigilant for the development of inflammatory myositis post-allogeneic HSCT as most patients respond to treatment. Advances in immunohistochemistry to determine the dominant cell type and cytokine profile may enable targeted and individualised therapies.

Sexuality- and Fertility-Related Issues in Women after Allogeneic Hematopoietic Stem Cell Transplantation

Sexual dysfunction and fertility related issues appear as major post-allogeneic hematopoietic stem cell transplantation (HSCT) late effects in young women, with a heavy impact on quality of life. The objective of the present study was to evaluate the impact of disease and treatments on sexual quality of life, ovarian function, and family planning initiatives in the context of allogeneic HSCT. Between January 2014 and January 2016, adult female patientswho underwent HSCT before age 35 and had been followed formore than 2 years in our centerwere offered participation inthe study through a self-reported surveyand/or ovarian function assessment if age <40 at inclusion. A total of 63 patients were included, with a median age of 23.4 years at transplantation and 30.9 years at inclusion. Twenty-nine patients (46%) underwent HSCT for acute leukemia and 16 (25%) underwent HSCT for aplastic anemia (AA). The conditioning regimen was myeloablative conditioning (MAC) in 37 patients (59%) and reduced-intensity conditioning (RIC) in 26 (41%). Fifty-eight patients completed the survey, and 34 were evaluated for ovarian function. Symptoms of hypoestrogenism were reported by 86% of the patients and changes in sexual life were reported by 76%, due mainly to low sex drive, negative impact of infertility problems, physical sequelae, and loss of self-confidence. Premature ovarian failure (POF) occurred in 74% of patients and was significantly associated with conditioning regimen (MACversusRIC; P=.001) and baseline disease (bone marrow failureversusacute leukemiaversusothers; P < .001). However, one-half of the patients developed a POF despite the use of a RIC regimen. For 27 patients (47%), disease and treatments modified their desire for pregnancy, due mainly to fear of relapse and of disease transmission to offspring. Thirteen pregnancies were reported (21%), of which 8 were spontaneous and 5 were obtained through assisted reproductive technologies, mainly oocyte donation. With a median post-transplantation follow-up of 12.2 years, the 10-year cumulative incidence of first pregnancy was 16.6% (95% CI, 8.8-30.0). Among 20 patients (32%) who engaged in a family planning initiative, 13 (65%) succeeded in having children: 11 got pregnant and 2 adopted. Sixteen patients benefited from fertility preservation techniques consisting of ovarian tissue cryopreservation, and a single autologous ovarian tissue transplantation had been performed at the time of this report. This study shows a strong impact of disease and treatments on sexual quality of life, ovarian function, and family planning initiatives in the context of HSCT. It demonstrates the need to improve clinicians' awareness of sexual health- and fertility-related issues after HSCT. The difficulty of predicting ovarian function and fertility issues after RIC supports wide indications of pretransplantation fertility preservation. Evaluation of the use of cryopreserved ovarian tissues is warranted.

Post-allogeneic hematopoietic stem cell transplantation viral reactivations and viremias: a focused review on human herpesvirus-6, BK virus and adenovirus.

Human cytomegalovirus and Epstein–Barr virus have been recognized as potential drivers of morbidity and mortality of patients undergoing allogeneic stem cell transplantation for years. Specific protocols for monitoring, prophylaxis and pre-emptive therapy are in place in many transplant settings. In this review, we focus on the next three most frequent viruses, human herpesvirus-6, BK virus and adenovirus, causing reactivation and/or viremia after allogeneic transplant, which are increasingly detected in patients in the post-transplant period owing to emerging techniques of molecular biology, recipients’ characteristics, treatment modalities used for conditioning and factors related donors or stem cell source. Given the less frequent detection of an illness related to these viruses, there are often no specific protocols in place for the management of affected patients. While some patients develop significant morbidity (generally older), others may not need therapy at all (generally younger or children). Furthermore, some of the antiviral therapies used are potentially toxic. With the addition of increased risk of secondary infections, risk of graft failure or increased risk of graft-versus-host disease as well as the relationship with other post-transplant complications, the outcomes of patients with these viremias remain unsatisfactory and even long-term survivors experience increased morbidity.

Prognostic impact of circulating tumor DNA status post-allogeneic hematopoietic stem cell transplantation in acute myeloid leukemia and myelodysplastic syndromes

Relapse is an issue of concern for patients with acute myeloid leukemia and myelodysplastic syndromes (AML/MDS) who underwent allogeneic hematopoietic stem cell transplantation (allo HSCT). The conventional minimal residual disease (MRD) test of the bone marrow has serious limitations regarding invasiveness and applicability. To address this problem, we investigated the utility of a novel MRD test using tumor-derived fragmentary DNA, or circulating tumor DNA (ctDNA) for the identification of patients with high risk of AML/MDS relapse after undergoing myeloablative allo HSCT. We retrospectively collected tumor specimens and available matched serum samples at diagnosis and at 1 and 3 months post-allo HSCT from 53 patients with AML/MDS. After identifying driver mutations in 51 patients using next-generation sequencing, we designed at least one 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 allo HSCT. Both the mutation persistence in the bone marrow at 1 and 3 months post allo HSCT and the corresponding ctDNA persistence in the matched serum were comparably associated with higher 3-year cumulative incidence of relapse rates. These results demonstrate, for the first time, the utility of ctDNA as a noninvasive prognostic biomarker in patients with AML/MDS who underwent allo HSCT.

Evaluation of Next-Generation Sequencing and Crystal Digital PCR for Chimerism Monitoring of Post-Allogeneic Hematopoietic Stem Cell Transplantation

Hematopoietic stem cell transplantation (HSCT) is a curative treatment for most hematologic diseases. To evaluate the level of donor engraftment, chimerism must be carefully monitored after HSCT. Short tandem repeats, quantitative PCR (qPCR), and, more recently, digital PCR (dPCR) are widely used to determine the proportions of donor and recipient cells after HSCT. The screening and quantification of chimerism have been evaluated by 2 new methods: a ready-to-use next-generation sequencing (NGS)-based method using the Devyser ChimerismNGS kit and an original combination of the Stilla crystal digital PCR (cdPCR) platform with 3-color multiplexing capacity using GenDX KMRtrack reagents. The genotyping of 4 HSCT pairs by cdPCR using 11 triplex mixes of the GenDX KMRtype kit was consistent at 98.8% with qPCR. Informative samples (n=20) from 6 donor-recipient pairs and 1 external proficiency test demonstrated the reliability of the results (0.1% to 50%) for the 2 methods. The methods are also highly sensitive (0.1%) and accurate. The chimerism values of the 2 methods are correlated and concordant with those of the reference methods. In addition, the ADVYSER software (Devyser) is user-friendly and well adapted to chimerism monitoring. In conclusion, these 2 innovative methods are easy to perform and user-friendly in all molecular, hematology, and immunogenetic laboratories and allow the genotyping and monitoring of chimerism with high performance and sensitivity.

The impact of chimerism on DNA-based human identification from skin surface cells of post-allogenic hematopoietic stem cell transplantation (HCST) patients

The use of biological traces recovered from touched or handled items increased with the advance of the forensic analysis system. Thus, DNA profiles obtained from touch DNA became a useful tool in forensic investigation. However, a chimeric person with more than one chromosomal population can be challenging for a forensic analyst. We investigated the genetic profile in blood, buccal swab, and skin swabs from twenty-four recipients aged 21–63 years who underwent a matched sibling allogeneic hematopoietic stem cell transplantation with no sign of skin graft versus host disease. Autosomal short tandem repeats genotyping was performed to evaluate chimerism status at 15 loci along with gender marker Amelogenin. According to our results, donor chimerism was detected in all recipient's blood samples, while in buccal swabs, five recipients showed no presence of donor-derived cells in their genotype. Epithelial cells swabbed from hand fingertips were not devoid of donor-derived cells since all recipients showed high chimerism (39.69%–96.66%) in their genotypes. A significant change in chimerism was seen among various types of biological samples (p<0.05). No correlations were observed between chimerism and recipient age, gender, or time after transplant (p> 0.05). The loci D21S11, D8S1179, and FGA were the most informative, whereas D13S317, Vwa, and TOPX were the least informative STR markers. We concluded that touch DNA from a person who has undergone a successful allogeneic HSCTs should not be considered as reliable evidence for human identifications. Therefore, necessary precautions must be taken to avoid false identification and miscarriage of justice.

Outcomes of HMA and Venetoclax in Relapsed AML Post-Allogenic Hematopoietic Stem Cell Transplant

Background: There is no standard of care treatment for Acute Myeloid Leukemia (AML) in relapse post-allogeneic hematopoietic stem cell transplant (post-HSCT), with overall 5-year survival about 5-10%. Venetoclax (Ven) is a novel BCL2 inhibitor approved by the FDA for treatment of newly diagnosed AML in combination with hypomethylating agents (HMA) or low dose cytarabine for patients unfit for intensive induction. However, data in relapsed/refractory AML are limited, especially in the post-HSCT setting. In this retrospective study, we reviewed outcomes of patients with AML relapse post-HSCT who received Ven in combination with HMA in a single center. Methods: Charts of 17 patients who had AML relapse post-HSCT treated with combination of Ven and HMA between November 2018 - March 2020 at the University of Kansas Medical Center were reviewed. We utilized descriptive statistics for baseline characteristics and outcomes, and Kaplan-Meier log-rank test for calculating overall survival. Results Seventeen patients received Ven+HMA for AML relapse post-HSCT in our center. At the time of SCT, patients were in first complete remission (CR) (n=15); second CR (n=1) and primary failure induction (1). Median age was 62 at time of relapse (31-71) years, and 8 patients were female (47%). 9 patients (53%) had adverse risk AML (ELN 2017) 8 of them were in CR1 and 1 with primary induction failure. Common mutations included DNMT3a, ASLX-1, TET2 (3); TP53 (2); IDH1/2 (2); NPM1/FLT3 (1); NPM1/IDH2 (1); NPM1 (1 transplanted in CR2); FLT3 (1). 2/17 had received Ven+HMA prior to SCT; 4 patients received HMA alone prior to SCT. 11 patients (65%) were naïve to either Ven or HMA prior to relapse. Median time to relapse was 181 (44-851) days post-HSCT. 9 (53%) patients received Azacitidine+Ven and 8 (47%) received Decitabine+Ven. HMA+Ven was the first line of therapy post-HSCT relapse in 14 patients. 2 had donor lymphocyte infusion (DLI) after either MEC or dacogen but relapsed prior to Ven+HMA. 1 had IDH (2) inhibitor. Patients received median of 2 (1-10) cycles of HMA+Ven. Six (35%) patients achieved complete remission/complete remission with incomplete hematologic recovery (CR/CRi), and 2/6 patients had negative measurable residual disease by multiparameter flow cytometry. Median overall survival was 361 days from relapse (Figure 1). 3/14 patients received subsequent DLI with Ven+HMA. Disease progression was the most common cause of death in 8/9 of patients who died during the follow up period. Most common side effects included neutropenic fever (n=8, 47%) and acute graft versus host disease (aGVHD) (n=5, 30%). 2/5 developed new aGVHD on HMA+Ven with no prior history of aGVHD. However, aGVHD was mainly grade I-II and responsive to therapy. Discussion HMA+Venetoclax demonstrates potential activity in patients with AML relapse post-HSCT with a CR/CRi rate of 35%, comparable to other salvage therapies. There were no unexpected side effect in this high-risk population. Larger studies are needed to confirm efficacy and toxicity in this setting. Disclosures McGuirk: Pluristem Ltd: Research Funding; Kite Pharmaceuticals: Consultancy, Honoraria, Research Funding, Speakers Bureau; Gamida Cell: Research Funding; Bellicum Pharmaceutical: Research Funding; Allo Vir: Consultancy, Honoraria, Research Funding; Juno Therapeutics: Consultancy, Honoraria, Research Funding; Astellas: Research Funding; Fresenius Biotech: Research Funding; Novartis: Research Funding. Lin:Aptevo: Research Funding; Abbvie: Research Funding; Bio-Path Holdings: Research Funding; Celgene: Research Funding; Gilead Sciences: Research Funding; Incyte: Research Funding; Jazz: Research Funding; Mateon Therapeutics: Research Funding; Ono Pharmaceutical: Research Funding; Pfizer: Research Funding; Prescient Therapeutics: Research Funding; Seattle Genetics: Research Funding; Tolero Pharmaceuticals: Research Funding; Trovagene: Research Funding; Genetech-Roche: Research Funding; Celyad: Research Funding; Astellas Pharma: Research Funding.

Salvage Therapy with Low-Dose Ruxolitinib Led to a Significant Improvement in Bronchiolitis Obliterans Syndrome in Patients with cGvHD after Allogeneic Hematopoietic Stem Cell Transplantation

Background Bronchiolitis obliterans syndrome (BOS) is a life-threatening pulmonary manifestation of chronic graft versus host disease (cGVHD) post-allogeneic hematopoietic stem cell transplantation (HSCT), without a clear standard of care. Systemic corticosteroids remain the backbone of therapy for BOS, but their side effect profile makes them disadvantageous for prolonged use. Ruxolitinib (RUXO), a Janus-associated kinase (JAK) 1/2 inhibitor approved for the treatment of patients with intermediate-2 or high-risk primary myelofibrosis,has shown excellent response rates in steroid-refractory/intolerant cGVHD. This study seeks to investigate the efficacy of ruxolitinib on steroid-refractory BOS, and additionally, determine the efficacy of ruxolitinib in these patients and the clinical factors affecting the response to ruxolitinib. Method This study involved adult patients suffering BOS after an allogeneic HSCT for a hematological malignancy between January 2014 and June 2018 at the First Affiliated Hospital of Zhejiang University School of Medicine and Ningbo Hospital of Zhejiang. Univariable analyses for treatment response risk factors were performed using the Fisher exact test and chi-square test. Logistic regression analysis was used to perform multivariable analysis to calculate the hazard ratio (HR) with a 95% confidence interval (CI). Overall survival (OS) was defined as the time from stem cell infusion to death from any cause and calculated by the Kaplan-Meier method. Results Twenty-five patients were retrospectively enrolled in this analysis and patient characteristics were shown in Table 1. At the time of BOS diagnosis, most patients presented with moderate or severe obstruction. Besides, all patients had worsening lung function or worsening clinical symptoms under systemic prednisone treatment. After a median duration of RUXO therapy of 8.5 (1.5-20) months, the overall response rate was 68.0%: two patients (8.0%) achieved complete remission, and 15 (60.0%) achieved partial remission. The median time from initiation of RUXO to achieve the best responses was 3 months (range, 1.5-8). Since initiating RUXO, predicted forced expiratory volume in 1 s (FEV1) % increased after 3 months and 6 months compared with measurements before RUXO in responders. Of note, an average reduction in prednisone dose of 39.3% after 3 months of ruxolitinib therapy and 59.5% after 6 months of ruxolitinib therapy was reported. Only FEV1 mild decline before ruxolitinib with a ratio ≤15% was an independent predictor to achieve a response to RUXO (HR = 0.063, 95% CI 0.006-0.624, P = 0.018). As for safety profile, hematological adverse effect included that two patients harboring cytopenia with grade 2-3. Regarding the nonhematological toxicities, two patients developed grade 3 hypertriglyceridemia. Referring to infection events, 11 patients (44%) had a severe pulmonary infection of ≥3 grade. Of note, cytomegalovirus (CMV) DNAemia was observed in 3 patients (12%) and Epstein-Barr virus (EBV) DNAemia was observed in 10 patients (40%). Following a median follow-up of 255 days after RUXO, the 2-year incidence of nonrelapse mortality and 2-year overall survival rate after RUXO among patients with BOS was 27.8% and 62.6%, respectively. Conclusion The present study showed that in heavily pretreated patients with BOS, ruxolitinib induced a relatively high response, leading to an increased 2-year OS rate in patients with BOS. Although a small number of subjects and a retrospective nonrandomized design of this study might make a bias, our data still indicated RUXO as a promising treatment option to improve the prognosis of post-HSCT BOS, with pulmonary function stabilization or even improvement and steroid sparing. Also, the substantial clinical benefits and efficacy of ruxolitinib on BOS need to be balanced against infection risks. Disclosures No relevant conflicts of interest to declare. OffLabel Disclosure: Ruxolitinib (RUX, Jakafi, Novartis, Basel, Switzerland)

Real-Life Experience of Sorafenib Maintenance after Allogeneic Hematopoietic Stem Cell Transplantation for FLT3-ITD AML Reveals High Rates of Toxicity-Related Treatment Interruption

Introduction: FLT3-Internal Tandem Duplication (FLT3-ITD) mutations of the gene encoding the FLT3 tyrosine kinase receptor are found in 25-30% of AML patients and are associated with a poor prognosis despite intensive chemotherapy and allogeneic hematopoietic stem cell transplantation (HSCT). FLT3 tyrosine kinase inhibitors (TKI) significantly improve survival of FLT3 mutated AML patients in large prospective trials (Stone, NEJM 2017; doi:10.1056/NEJMoa1614359). Post-HSCT maintenance with sorafenib - a broad-spectrum TKI with strong activity against FLT3 - was recently shown to reduce the risk of relapse and improve survival in the phase II SORMAIN trial (Burchert, JCO 2020; doi: 10.1200/JCO.19.03345). Importantly, clinical trials reported a low rate of toxicities requiring sorafenib discontinuation (Chen et al, BBMT 2014;doi: 10.1016/j.bbmt.2014.09.007; Burchert et al., J Clin Oncol. 2020; doi: 10.1200/JCO.19.03345). The aim of our single-center retrospective analysis was to evaluate the real-life experience in patients treated with post-allogeneic HSCT sorafenib maintenance therapy for FLT3-ITD AML with a particular focus on tolerability and toxicity-related treatment interruption. Methods: We conducted a single-center retrospective study on 33 FLT3-ITD AML patients undergoing allogeneic HSCT in complete remission between 2013 and 2020, including 20 patients who received sorafenib maintenance and 13 who did not. Sorafenib was started at hematologic reconstitution at 200 mg BID and increased at 400 mg BID after a week in case of good tolerance. Treatment was maintained until two years if tolerated. In case of toxicity, the drug was either reduced to 200mg BID, or stopped. Among the 13 patients in the untreated group, 8 were transplanted between 2013 and 2016 before we started to routinely use sorafenib maintenance, and 5 were transplanted in 2017 or later. Among the latter, sorafenib maintenance was not started because of aGVHD and insufficient hematologic reconstitution (n=3), low-allelic ratio of FLT3-ITD (n=1), early death due to PTLD (n=1). Results: In the sorafenib group (n=20), median time from transplant to sorafenib initiation was 68 days (range 40-213). Overall, 16 patients (80%) experienced toxicities leading to dose reduction (4 patients, 20%) or interruption (12 patients, 60%). In these patients, the average time on sorafenib before toxicity occurrence was 98 days (range 1-717; only one patient stopped treatment after 1 day because of severe diarrhea). Most common toxicities were skin (7, grade II), gastrointestinal (5, grade II), and hematologic (5, grade II and III). One patient experienced uveitis (grade III) and pneumonia (grade IV), both resolutive after sorafenib interruption. One patient experienced severe hypertension (grade III), 1 had hepatitis (grade III) and another one a PRESS syndrome (grade IV) possibly related to sorafenib. Other causes of sorafenib interruption included FLT3-ITD negative relapse in 1 patient, relapse in 1 patient, and end of scheduled maintenance in 1 patient. Among the 12 patients who interrupted sorafenib because of toxicities, 3 patients were re-challenged with good tolerance in 2 cases, while 7 were switched to midostaurine. Among these 7, 4 completed the 2-y treatment and 3 interrupted midostaurine because of toxicities (2 patients experienced digestive intolerance and 1 pneumonia). The average time of sorafenib exposure in the whole sorafenib group was 180 days (range 1-765). Importantly, the analysis of patients' outcome confirmed the previously reported positive impact of sorafenib maintenance on overall survival (Figure 1B) despite high rates of treatment interruption. Conclusion: Our real-life analysis reveals higher rates of toxicity-related interruption of sorafenib maintenance after allogeneic HSCT than previously reported in clinical trials. Interestingly, our results confirm the favorable impact of sorafenib maintenance on clinical outcome despite a high rate of toxicity-related treatment interruption, suggesting the feasibility of switching to other maintenance approaches in case of low tolerability. Disclosures No relevant conflicts of interest to declare. OffLabel Disclosure: sorafenib as a maintenance treatment to prevent FLT3-ITD mutated AML reccurence after allogeneic transplantation

Clinical prevalence and outcome of cardiovascular events in the first 100 days postallogeneic hematopoietic stem cell transplant.

Recent advances in allogeneic hematopoietic stem cell transplant (HSCT) have allowed us to offer HSCT to older, advanced disease patients with more co-morbidities. Cardiovascular toxicity post-transplant is a major concern due to the increased risk of mortality. Few studies have examined the prevalence of CV events including CAD (MI, angina, PCI, CABG, CHF, arrhythmias), HTN, stroke/TIA, and death in the first 100days post-transplant. We assessed the impact of pretransplant MUGA results in predicting postallogeneic HSCT CV events and overall survival in the first 100days, and whether or not transient anthracycline-induced cardiomyopathy or cumulative anthracycline dose affected overall survival. This retrospective, cohort study included 665 patients with a median age of 52years who underwent HSCT from 2009 to 2015. The most frequent CV event in the first 100days post-HSCT was arrhythmia seen in 2.9% of patients followed up by CHF (12.3%), MI (9%), and angina (8%). Two patients had PCI, and both survived the first 100days. Cardiovascular risk factors predict for a poor MUGA scan but not survival. Higher dose anthracycline pretransplant predicted for a poor outcome. A history of CV disease, MI, or CAD was the most important predictive of CV events, P-value=.00002. 88.6% survived the first 100days. Patients with an EF<50% had a significant likelihood of having a CV event compared to patients with an EF>60% (OR=5.3, 95% CI [1.6-18.1], P=.0219). Cumulative anthracycline dose did not have a significant impact on overall survival.

The Transplant Evaluation Rating Scale predicts overall survival after allogeneic hematopoietic stem cell transplantation

AbstractTo evaluate the impact of psychosocial risks on post–hematopoietic stem cell transplantation (HSCT) outcomes, we prospectively conducted psychosocial assessment of 556 consecutive allogeneic HSCT patients who received their first allogeneic transplant at our center between 2003 and 2017. The Transplant Evaluation Rating Scale (TERS) score was prospectively assessed by a psychologist before transplantation, and patients were categorized as low, intermediate, or high risk based on their TERS score. Patients in the high-risk TERS group had significantly longer hospital stays during the first 180 days and 1 year post–allogeneic HSCT compared with the low-risk group (16 vs 13 and 21 vs 16 days; P = .05 and .02, respectively). The survival estimates for low-, intermediate-, and high-risk TERS groups at 3 year were as follows: overall survival (OS), 73%, 60%, and 65%; disease-free survival (DFS), 63%, 55%, and 60%; nonrelapse mortality (NRM), 11%, 20%, and 17%; and relapse, 26%, 25%, and 23%, respectively. In a multivariable analysis, intermediate- and high-risk TERS scores predicted for inferior OS, similar DFS, and higher NRM compared with low-risk TERS score. In a subset analysis of patients with low/intermediate risk per Disease Risk Index, multivariable analysis showed that high- and intermediate-risk TERS scores predicted for significantly worse OS, worse DFS, higher NRM, and similar relapse rates compared with low-risk TERS score. Our findings show that psychosocial factors as measured by TERS score are strong predictors of morbidity and mortality after HSCT among patients with low/intermediate disease risk.

Study protocol of a pilot study evaluating feasibility and acceptability of a psychosexual intervention for couples postallogeneic haematopoietic stem cell transplantation

IntroductionSexual dysfunction is one of the most common side effects of allogeneic haematopoietic stem cell transplantation (HSCT) for haematological cancers. Problems can persist between 5 and 10 years post-transplant and...

Is immunological recovery clinically relevant at 100 days after allogeneic transplantation?

Background/AimsImmune reconstitution following allogeneic hematopoietic stem cell transplantation (HSCT) is affected by multiple variables during the transplantation.MethodsWe assessed the clinical factors contributing to immune function reconstitution at 100 days post-allogeneic HSCT in 114 patients receiving fludarabine-based conditioning. Immunophenotypic analysis using flow cytometry was performed to evaluate the percentage and the absolute numbers of T-cell subsets, natural killer cells, and B-cells as clinical outcomes.ResultsTacrolimus-based graft-versus-host disease (GVHD) prophylaxis, T-cell depletion, and acute GVHD were significantly associated with delayed immune reconstitution of T-cell subsets. The incidence of chronic GVHD was significantly increased in the normal recovery group compared to the abnormal group (p = 0.01). Epstein-Barr virus reactivation was more frequently observed in the abnormal group of T-cell subsets (p = 0.045). All viral reactivation events including cytomegalovirus reactivation appeared to be more frequent in the abnormal group of T-cell subsets.ConclusionsThe immune recovery status post-allogeneic HSCT was affected by GVHD prophylactic regimens, especially in cases receiving tacrolimus-based GVHD prophylaxis, T-cell depletion, and possibly those manifesting acute GVHD. Delayed immune reconstitution might increase the morbidity due to viral reactivation. Treatment strategies are needed to prevent infectious complications and enhance immune reconstitution based on the immune recovery status following allogeneic HSCT with fludarabine-based conditioning.

Role of Post-Allogeneic Hematopoietic Stem Cell Transplantation Low-Dose Azacitidine for Prevention of Relapse in Patients with Acute Myeloid Leukemia and Myelodysplastic Syndrome: A Retrospective Analysis

Introduction Relapse after allogeneic hematopoietic stem cell transplantation (HSCT) for acute myeloid leukemia (AML) and myelodysplastic syndrome (MDS) remains a major cause of treatment failure with associated poor prognosis. Low-dose post-HSCT maintenance azacitidine (AZA) has proved to be beneficial in preventing relapses (de Lima et al. Cancer 2010). Herein, we retrospectively evaluated outcomes of patients (pts) treated with low-dose AZA post-HSCT. Methods Pts with AML and MDS who underwent allogeneic HSCT between 2012-2018 at two centers were evaluated and categorized into AZA group (with low-dose AZA maintenance) and control group (without AZA maintenance). In control group, pts who died or relapsed prior to day +100, had grade III-IV aGVHD or received maintenance therapy other than AZA (e.g. FLT3 inhibitors) were excluded. Using Kaplan-Meier method, event (defined as relapse or death) free survival (EFS) and overall survival (OS) were computed and compared using log-rank test. Results A total of 107 patients were included; 53 (49.5%) in AZA and 54 (50.5%) in control groups. Disease and transplant-related characteristics are shown in table 1. Both groups were comparable with respect to age, diagnosis, disease risk stratification, disease status at HSCT, and MRD status at day +30 and day +100 (table 1). More pts received reduced intensity conditioning (RIC) in the AZA group (71.6% vs 37%, p= Conclusions In our cohort, low-dose AZA maintenance was generally well tolerated. The results of this study indicate that AZA maintenance is feasible and associated with improved EFS and OS. In addition, AZA has the potential of improving relapse rates even in pts who are unable to receive myeloablative conditioning as a significant number of pts in AZA group in this study received RIC. The optimal timing, duration of treatment and selection of pts most suitable for AZA for maximum benefit, however, requires further studies.

Chimerism Monitoring with Highly Sensitive and Precise Next-Generation Sequencing Assay in Patients Post-Allogeneic Hematopoietic Stem Cell Transplantation

<h3>Introduction</h3> Chimerism testing of peripheral blood and bone marrow aspirates following allogeneic hematopoietic cell transplantation (HCT) is important to evaluate engraftment kinetics and potentially early detection of disease relapse¹. NGS-based chimerism assays use SNP panels and provide standardized workflow with automated data analyses. <h3>Objectives</h3> We aim to evaluate performance of an NGS-based chimerism assay and compare it to existing laboratory method based on Short Tandem Repeat (STR) analysis. <h3>Methods</h3> Genomic DNA (gDNA) derived from PBMCs of unrelated donors were mixed at fractions ranging from 0.1% - 50% to evaluate sensitivity and specificity of the NGS-based chimerism assay in artificial genomic chimeras. 3-10 ng gDNA was used to prepare libraries with the assay kit and sequenced on Illumina MiSeq. <h3>Results</h3> Linear regression analyses of gDNA fractions (expected, observed) detected a linear signal at a minimum of 0.1% of minor gDNA in a two-gDNA chimeric mix (R²=1, average CV = 4.5%, range = 0.2-12.4). Chimeric samples with three distinct gDNAs gave linear signal as low as 0.5% of minor gDNA fraction (CV < 2%, range = 0.01-5.5%), a scenario that is clinically relevant in double-cord HCT or second HCT from different donors. The NGS-based chimerism assay was also tested on post-transplant samples derived from three HCT recipients. Results were compared longitudinally and between the two test methods at various time points (days) post transplantation. While majority results were consistent with previous data from STR, some discrepancies were observed. Patient 1, day+128, showed microchimerism (0.1%) that was undetected by STR (0%). Patient 2, day+287, showed low level (2.42%) of recipient DNA while STR reported full donor engraftment. Such clinical cases underscore the need for high sensitivity assays to more accurately quantitate donor-recipient fractions in post-HCT samples. <h3>Conclusions</h3> Using the test NGS-based chimerism assay on two and three gDNA mixes, we observe 10X greater sensitivity of minor fraction quantitation compared to current STR method. In our experience, high sensitivity, precision over a wide range of detection and robust performance with low input DNA may qualify this NGS-based chimerism assay as a potential method of choice for routine engraftment monitoring and potential early detection of disease relapse.

Single-Cell RNA-Seq Identifies Potentially Pathogenic B Cell Populations That Uniquely Circulate in Patients with Chronic Gvhd

While B cells are known to contribute to the pathogenesis of chronic graft-versus-host disease (cGVHD) in mice, it has been challenging to elucidate intrinsic mechanisms of tolerance loss in patients. To identify distinct and potentially targetable B-cell subsets in cGVHD, we employed single-cell RNA-Seq along with an unsupervised hierarchical clustering analysis, targeting 10,000 single B cells from each of eight patients who were &amp;gt;12 months post-allogeneic hematopoietic stem cell transplantation (HCT) and either had active cGVHD manifestations (n=4) or never developed cGVHD (n=4). Bioinformatics analysis of pooled cell data (using R with Seurat extension package) identified 6 major B cell clusters common to all patients (Figure 1A). "Intra-cluster" gene comparison (using R package DESeq2, false-discovery rate 0.05) revealed numerous differentially expressed genes between patient groups. The greatest number of differentially-expressed genes occurred in a cluster referred to herein as 'Cluster 6' (Figure 1A, in yellow with asterisk). Within Cluster 6, B cells from active cGVHD patients expressed significantly increased ITGAX (CD11c, Padj =0.007), TNFRSF13B (TACI, a receptor for BAFF, Padj =0.003), IGHG1 (IgG1, Padj =9.3e-06) and IGHG3 (IgG3, Padj =1.7e-12), along with 44 additional genes (to be discussed). Thus, Cluster 6 in cGVHD patients may represent a CD11cpos, BAFF-responsive B cell subset primed to undergo isotype switching in response to alloantigen. Flow cytometry analysis on PBMCs from an independent HCT patient cohort (n=10) confirmed that CD11cpos B cells were indeed significantly expanded in cGVHD (P &amp;lt; 0.01, Figure 1B), and revealed these B cells were also TACIpos, CD19high, forward scatter high (FSChigh) blast-like cells (Figure 1C). We found that these CD11cpos B cells had mixed expression of CD21, CD27, IgD and CD24 (Figure 1C). Remarkably, other recent studies on bulk patient B cells have suggested that similar CD11cposCD21negCD19highT-BETpos cells are critical drivers of humoral autoimmunity in diseases including systemic lupus erythematosus (SLE; Scharer et al. 2019; Rubtsova et al. 2017; Rubtsov et al. 2011). This subset now identified by single-cell RNA-Seq is consistent with a population of TACIhigh B cells that produced IgG in response to BAFF treatment ex vivo (Sarantopoulos 2009). Data suggest we have identified functionally distinct and potentially targetable B cell subpopulations. We are employing functional assays to determine whether the additional molecular pathways now elucidated account for our previous work showing greater ex vivo B cell survival rates and hyper-responsiveness to surrogate antigen (Allen et al. 2012, 2014), certain TLR agonists (Suthers et al. 2017), and NOTCH ligand (Poe et al. 2017). In addition to more deeply characterizing B-cell subsets in cGVHD, our single-cell RNA-Seq analyses identified several genes significantly altered across multiple B cell clusters in the cGVHD group, implicating more broad alterations of some genes in this disease. Among these is CKS2, a critical cell cycle regulator, which was significantly increased in cGVHD B cells (Padj 1.0e-10 to 0.018, depending on the cluster evaluated). Increased CKS2 expression was validated by qPCR analysis on B cells from a separate HCT patient cohort with or without cGVHD (P &amp;lt; 0.001, Figure 1D), suggesting that the majority of cGVHD B cells are primed to enter the cell cycle at multiple stages of differentiation when exposed to the proper stimuli. In summary, we used an unbiased approach to identify and further characterize an extrafollicular CD11cposTACIposCD19high B cell population in cGVHD patients that appears to be activated and undergoing active IgG isotype switching. This plasmablast-like B cell population is potentially amenable to therapeutic intervention to prevent pathogenic antibody production. Importantly, we also identify gene alterations across the cGVHD peripheral B cell compartment that potentially underpin promotion of hyperactivated B cells in this disease. Therapeutic strategies to target these pathways will also be discussed. This work was supported by a National Institutes of Health grant, R01HL129061. Disclosures Ho: Omeros Corporation: Membership on an entity's Board of Directors or advisory committees; Jazz Pharmaceuticals: Research Funding; Jazz Pharmaceuticals: Consultancy. Horwitz:Abbvie Inc: Membership on an entity's Board of Directors or advisory committees. Rizzieri:Celgene, Gilead, Seattle Genetics, Stemline: Other: Speaker; AbbVie, Agios, AROG, Bayer, Celgene, Gilead, Jazz, Novartis, Pfizer, Sanofi, Seattle Genetics, Stemline, Teva: Other: Advisory Board; AROG, Bayer, Celgene, Celltron, Mustang, Pfizer, Seattle Genetics, Stemline: Consultancy; Stemline: Research Funding.

Defibrotide Inhibits Endothelial Cell Injury Induced By Plasmas of Patients with Thrombotic Microangiopathies

Introduction: Defibrotide is a polydisperse mixture of predominantly single-stranded polydeoxyribonucleotide sodium salts derived from porcine intestinal mucosa. It is currently the only therapy FDA-approved to treat hematopoietic stem cell transplant (HSCT)-associated hepatic veno-occlusive disease (VOD) with multi-organ dysfunction. Its mechanisms of action are complex and incompletely understood. In vitro, defibrotide can: promote endothelial cell (EC) mitogenesis via interaction with basic fibroblast growth factor; inhibit platelet adhesion and aggregation; reduce calcineurin inhibitor-induced EC apoptosis; and block pro-inflammatory cytokine activity, in the absence of anticoagulant effects (Blood Adv 2018;2:1495-1509). Given the critical role of EC injury with impaired regulation of thrombo-fibrinolytic pathways in both transplant-associated VOD and thrombotic microangiopathies (TMA), defibrotide has been used off-label to treat TMAs post-allogeneic HSCT. Significant clinical responses occurred in the majority of cases not complicated by renal failure (Bone Marrow Transplant 2019;54:142-145). It is known that conditioning regimens for autologous HSCT involve activation of pro-inflammatory pathways in EC which can be blocked by defibrotide (Biol Blood Marrow Transplant 2011;17:497-506). We sought to determine whether clinically relevant concentrations of defibrotide could block activation of caspase 8 in EC, an early step in apoptotic injury, as induced by plasmas from patients with TMAs secondary to thrombotic thrombocytopenic purpura (TTP), cancer/chemotherapy, and allogeneic HSCT. Methods: We utilized plasmas from individuals with acute TMAs: 13 with TTP; 5 with an atypical hemolytic-uremic syndrome (HUS)-type of TMA in the setting of cancer chemotherapy, persisting after withdrawal of chemotherapy; and 10 with TMAs following alloHSCT, persisting after withdrawal of GvHD prophylaxis. The in vitro model system was previously published by our lab, documenting the ability of plasmas from acute TTP patients to activate caspase 8 in EC, followed by apoptotic EC death (Blood 2008;112:340-349). This effect was dependent on plasma-based proinflammatory cytokines capable of down-regulating EC cytoprotective factors. Briefly, 5 x 105 primary human neonatal microvascular endothelial cells of dermal origin were plated in 1ml culture medium/well of 12 well polystyrene plates (3.6cm2/well) pre-coated with 1% fibronectin. 2% (v/v) patient or control plasma was added, alone or with varying concentrations of defibrotide (0.5-10μg/ml), for 3hrs at 37ºC. Cells were then harvested and a commercial colorimetric kit (Millipore) was used to assess caspase 8 enzyme activity. Results: Initial dose-response curves showed a plateau effect for defibrotide-mediated inhibition of caspase 8 activation at 5μg/ml; subsequent experiments utilized that dose. To control for the possibility of direct inhibition of caspase 8 by defibrotide, we found that caspase 8 activity initiated by staurosporine (1μM) was unaffected by defibrotide (Fig.) &amp;gt;50% inhibition of caspase 8 was seen for 3/5 cancer/chemotherapy-linked TMA patient plasmas, 7/13 TTP plasmas, and 0/10 TA-TMA plasmas. &amp;gt;20% suppression of such activation was seen for 5/5 cancer/chemotherapy TMA plasmas, 12/13 TTP plasmas, and 4/10 TA-TMA plasmas. Representative examples are shown in the Fig., including defibrotide blocking caspase 8 activity induced by plasmas from a patient with cancer/chemotherapy linked TMA (patient 18) or TTP (patient 41), but not from another TTP patient (patient 17). Conclusions: Interventions with high degrees of therapeutic success are available for TMAs related to TTP and primary aHUS. However, TMAs complicate 20% of alloHSCT, and though half may resolve when GvHD immunoprophylaxis is discontinued, three year survival rates for those in whom the TMA persists are as low as 11%. Based on multiple case reports, defibrotide may represent an important resource in those cases, and in other TMAs refractory to conventional treatment. Further studies are required to see if defibrotide-mediated blockade of plasma-induced EC activation and injury which we demonstrate in vitro correlates with in vivo responses, and if the mechanism involves suppression of pro-inflammatory cytokine-mediated cell activation. Disclosures No relevant conflicts of interest to declare.