Nonmyeloablative Hematopoietic Cell Transplantation Research Articles

Post-Transplant Cyclophosphamide Attenuates the Adverse Effect of HLA Disparity and a Major ABO Mismatch on Graft Failure in Non-Myeloablative Allogeneic HCT

Post-Transplant Cyclophosphamide Attenuates the Adverse Effect of HLA Disparity and a Major ABO Mismatch on Graft Failure in Non-Myeloablative Allogeneic HCT

Significant impact of antibiotic exposure on GI-GVHD, NRM, and GRFS following allogeneic HCT with non-myeloablative Flu-TBI conditioning

Background Acute gastro-intestinal graft-versus-host disease (GI-GVHD) and non-relapse mortality (NRM) after allogeneic HCT are closely related to loss of microbial diversity and intestinal dominance by single taxa resulting from the use of antibiotics, dietary changes, and mucosal barrier injury. There is a paucity of data on the impact of use of antibiotics in HCT after Flu-TBI-based non-myeloablative (NMA) conditioning where there is absence of mucositis and limited malnutrition. Methods We did a retrospective single-center analysis of patients receiving Flu-TBI-based NMA HCT for a high-grade myeloid malignancy, mostly AML, and MDS, or acute lymphoblastic leukemia (ALL). We analyzed the impact of pre-engraftment antibiotic exposure, prophylactic ciprofloxacin, and or treatment with broad-spectrum cephalosporin/carbapenem, on HCT outcomes, with a focus on the incidence of acute GI-GVHD by day 180 and NRM at 1 year. Results A total of 150 patients were evaluable with a median age of 62 years. Antibiotics were used in 90 patients; 60 prophylactic use only and 30 therapeutic use with or without previous prophylaxis. Antibiotic use resulted in a significant higher incidence of GI-GVHD Stage 1–4; 29% (26/90) versus 5% (3/60) in those not receiving antibiotics (OR 8.1 (95% CI 2.3–28.3), p = 0.001). Use of antibiotics resulted in higher 1-year NRM (19% (17/90) versus 10% (6/60), HR 2.3, p = 0.06), and decreased 2-year GRFS (42% (38/90) versus 55% (33/60), HR 1.7, p = 0.04), but did not impact RFS or OS. Conclusions Use of antibiotics was related to the occurrence of GI-GVHD, NRM, and GRFS in patients receiving truly NMA HCT. Therefore, in the absence of mucositis and low incidence of bacteremia, antibiotics can and should be used restrictively in this setting.

Early hematopoietic cell transplantation for familial hemophagocytic lymphohistiocytosis in a regional treatment network in Japan.

Familial hemophagocytic lymphohistiocytosis (FHLH) is a fatal hyperinflammation syndrome arising from the genetic defect of perforin-mediated cytolysis. Curative hematopoietic cell transplantation (HCT) is needed before development of central nervous system (CNS) disease. We studied treatment outcomes of 13 patients (FHLH2 n = 11, FHLH3 n = 2) consecutively diagnosed from 2011 to 2022 by flow cytometric screening for non-myeloablative HCT in a regional treatment network in Kyushu, Japan. One patient with a novel PRF1 variant escaped screening, but all patients with FHLH2 reached diagnosis and 8 of them received HCT until 3 and 9months of age, respectively. The earliest HCT was conducted 65days after birth. Three pretransplant deaths occurred in newborns with liver failure at diagnosis. Ten posttransplant patients have remained disease-free, 7 of whom had no neurological involvement. Time from first etoposide infusion to HCT was shorter in patients without CNS disease or bleeding than in patients with those factors (median [range] days: 62 [50-81] vs. 122 [89-209], p = 0.016). Six of 9 unrelated patients had a PRF1 c.1090_1091delCT variant. These results suggest that the critical times to start etoposide and HCT are within 3months after birth and during etoposide control, respectively. Newbornscreening may increase the percentage of disease-free survivors without complications.

SCD Dyslipidemia Normalizes Long-Term Following Nonmyeloablative Hematopoietic Cell Transplantation

Dyslipidemia is common in patients with sickle cell disease (SCD). Individuals with SCD have a unique type of dyslipidemia. This SCD dyslipidemia (hypocholesterolemia subtype) is characterized by low total cholesterol (TC), low-density lipoprotein cholesterol (LDL-c), and high-density lipoprotein cholesterol (HDL-c), and normal triglycerides (TG). This SCD dyslipidemia state is theorized to be cardioprotective regarding atherosclerotic risk due to the hypolipidemic condition. Hematopoietic cell transplantation (HCT) is a potentially curative therapy for SCD. Nonmyeloablative HCT has been employed at the National Institutes of Health (NIH) and internationally with good results. Long-term survivors of HCT, specifically for hematologic malignancies, are at increased risk for dyslipidemia (hypercholesterolemia subtype), thus increasing atherosclerotic risk long-term. There are no studies investigating the short or long-term effects of HCT on SCD dyslipidemia. Therefore, we report a novel study to characterize lipid profiles at baseline and after HCT for SCD. This cohort study analyzes data from patients who received their first nonmyeloablative HLA-matched sibling or haploidentical HCT for SCD at the NIH from May 2009 to December 2021. All patients received sirolimus as GVHD prophylaxis. Total cholesterol, HDL-c, LDL-c, and triglycerides were collected pre-HCT, one-year post-HCT, and annually thereafter via a fasting serum-based lipid panel (median follow-up of 4 years). Data were analyzed using linear generalized estimating equation regression models to measure the influence of factors such as year of HCT, sirolimus use, gender, age, and graft failure on TC, LDL-c, HDL-c, and TG levels. A total of 116 patients were included in this analysis. There were 72 male and 44 female patients. The median age was 31 (range 10 to 64) years. 90% of the patients had Hb-SS, 3% Hb-SC, 5% Hb-Sβ 0, and 2% Hb-Sβ +. Overall survival is 87%, and 19% experienced graft failure. The median time on sirolimus post-HCT was one year. Baseline, one-year, and two-year mean (25th, 75th percentile) values, respectively, for the TC, LDL-c, HDL-c, and TG are as follows: TC [130.7 (107.0, 147.0), 183.2 (144.5, 220.0), 175.2 (144.5, 209.0)], LDL-c [72.5 (50.0, 87.0), 109 (73.5, 130), 104.2 (75.5, 128.0)], HDL-c [32.3 (23.8, 39.3), 49.4 (38.0, 55.5), 49.2 (41.5, 58.0)] and TG [127.3 (88.5, 156.5), 125.9 (87.0, 153.5), 112.6 (72.0, 133.5)]. Generally, the first-year post-HCT changes were maintained annually thereafter. (Figure 1). According to linear regression modeling, the change in TC, LDL-c, HDL-c, and TG at one-year post-HCT was +50.3 mg/dL (p<0.0001), +36.4 mg/dL (p <0.0001), +20.4 mg/dL (p<0.0001) and -26.5 mg/dL (p<0.0001), respectively. For failed grafts, the estimated changes after one-year post-HCT in TC, LDL-c, HDL-c, and TG were +6.5 mg/dL (p=0.45), -2.1 mg/dL (p=0.78), +14.8 mg/dL (p<0.0001), and -28.0 (p=0.03), respectively. The use of sirolimus only resulted in a significant change in TG (+32.8 mg/dL (p = 0.004)). Age had statistically significant changes only on TC (+1.08 mg/dL for a one-year change in age, p=0.005) and LDL-c (+0.73 mg/dL for a one-year change in age, p=0.03). There were no significant differences in TC, LDL-c, HDL-c, and TG levels (p>0.05) based on gender and HCT type. To summarize, HCT with sustained grafts led to significant rises in TC, LDL-c, and HDL-c and a decline in TG from baseline to one-year post-HCT, resulting in the normalization of lipid profiles. These changes were maintained annually thereafter. Graft failure most notably resulted in TC and LDL-c level declines back to pre-HCT baseline (Figure 2), while HDL-c and TG levels did not completely revert to baseline values. Sirolimus effects were significant on TG levels alone. These findings suggest that the mechanism underlying SCD dyslipidemia is corrected with the reversal of SCD. Long-term follow-up studies are indicated to assess patients' risk for atherosclerosis and adverse cardiac outcomes.

Improvement in Cardiac Morphology Demonstrated by Cardiac Magnetic Resonance Imaging and Echocardiography after Haploidentical Hematopoietic Cell Transplantation in Adults with Sickle Cell Disease

Cardiopulmonary complications account for approximately 40% of deaths in patients with sickle cell disease (SCD). Diffuse myocardial fibrosis, elevated tricuspid regurgitant jet velocity (TRV) and iron overload are all associated with early mortality. While HLA-matched sibling hematopoietic cell transplant (HCT) offers a potential cure, fewer than 20% of patients have a suitable donor. Haploidentical HCT allows for an increased donor pool and has recently demonstrated improved safety and efficacy. Our group has reported improved cardiac morphology via echocardiography at one year after HCT. Herein, we describe the first use of cardiac magnetic resonance imaging (CMR), the gold standard for measuring volume, mass, and ventricular function, to evaluate changes in cardiac morphology post-HCT in adults with SCD. We analyzed baseline and 1-year data from 12 adults with SCD who received nonmyeloablative haploidentical peripheral blood HCT at the NIH. Patients underwent non-contrast CMR at 3T, echocardiography, and laboratory studies. One year after HCT, patients showed marked improvement in cardiac chamber morphology by CMR, including left ventricular (LV) mass and volume (70.2 to 60.1 g/m2, p = 0.02; 114.5 to 90.6 mL/m2, p = 0.001). Furthermore, mean TRV normalized by 1 year, suggesting that HCT may offer a survival benefit. Fewer patients had pathologically prolonged native myocardial T1 times, an indirect marker of myocardial fibrosis at 1 year; these data showed a trend toward significance. In this small sample, CMR was very sensitive in detecting cardiac mass and volume changes after HCT and provided complementary information to echocardiography. Notably, post-HCT improvement in cardiac parameters can be attributed only in part to the resolution of anemia; further studies are required to determine the role of myocardial fibrosis reversal, improved blood flow, and survival impact after HCT for SCD.

Thyroid and Adrenal Dysfunction in Hemoglobinopathies Before and After Allogeneic Hematopoietic Cell Transplant.

To determine the rate and clinical characteristics associated with abnormal thyroid and adrenal function in recipients of nonmyeloablative hematopoietic cell transplantation (HCT) for sickle cell disease (SCD) and beta-thalassemia. We retrospectively reviewed patients who enrolled in 4 nonmyeloablative HCT regimens with alemtuzumab and total body irradiation (TBI). Baseline and annual post-HCT data were compared, which included age, sex, sickle phenotype, thyroid panel (total T3, free T4, thyroid stimulating hormone, antithyroid antibodies), cortisol level, ACTH stimulation testing, ferritin, medications, and other relevant medical history. Among 43 patients in haploidentical transplant and 84 patients in the matched related donor protocols with mostly SCD, the rate of any thyroid disorder pre-HCT was 3.1% (all subclinical hypothyroidism) and post-HCT was 29% (10 hypothyroidism, 4 Grave's disease, and 22 subclinical hypothyroidism). Ninety-two (72%) patients had ferritin >1000 ng/dL, of which 33 patients (35.8%) had thyroid dysfunction. Iron overload was noted in 6 of 10 patients with hypothyroidism and 12 of 22 patients with subclinical hypothyroidism.Sixty-one percent were on narcotics for pain control. With respect to adrenal insufficiency (AI) pre-HCT, 2 patients were maintained on corticosteroids for underlying rheumatologic disorder and 8 had AI diagnosed during pre-HCT ACTH stimulation testing (total 10, 7.9%). Post-HCT, an additional 4 (3%) developed AI from corticosteroid use for acute graft vs host disease, Evans syndrome, or hemolytic anemia. Although iron overload was common in SCD, thyroid dysfunction pre-HCT related to excess iron was less common. Exposure to alemtuzumab or TBI increased the rates of thyroid dysfunction post-HCT. In contrast, AI was more common pre-HCT, but no risk factor was identified. AI post-HCT was infrequent and associated with corticosteroid use for HCT-related complications.

Alemtuzumab and CXCL9 levels predict likelihood of sustained engraftment after reduced-intensity conditioning HCT.

Overall survival after reduced-intensity conditioning (RIC) allogeneic hematopoietic cell transplantation (HCT) using alemtuzumab, fludarabine, and melphalan is associated with high rates of mixed chimerism (MC) and secondary graft failure (GF). We hypothesized that peritransplantation alemtuzumab levels or specific patterns of inflammation would predict these risks. We assessed samples from the Bone Marrow Transplant Clinical Trials Network 1204 (NCT01998633) to study the impact of alemtuzumab levels and cytokine patterns on MC and impending or established secondary GF (defined as donor chimerism <5% after initial engraftment and/or requirement of cellular intervention). Thirty-three patients with hemophagocytic lymphohistiocytosis (n= 25) and other IEIs (n= 8) who underwent HCTs with T-cell-replete grafts were included. Patients with day 0 alemtuzumab levels ≤0.32μg/mL had a markedly lower incidence of MC, 14.3%, vs 90.9% in patients with levels >0.32μg/mL (P= .008). Impending or established secondary GF was only observed in patients with day 0 alemtuzumab levels >0.32μg/mL (P= .08). Unexpectedly, patients with impending or established secondary GF had lower CXCL9 levels. The cumulative incidence of impending or established secondary GF in patients with a day 14+ CXCL9 level ≤2394 pg/mL (day 14+ median) was 73.6% vs 0% in patients with a level >2394 pg/mL (P= .002). CXCL9 levels inversely correlated with alemtuzumab levels. These data suggest a model in which higher levels of alemtuzumab at day 0 deplete donor T cells, inhibit the graft-versus-marrow reaction (thereby suppressing CXCL9 levels), and adversely affect sustained engraftment in the nonmyeloablative HCT setting. This trial was registered at www.clinicaltrials.gov as #NCT01998633.

Comparison of reduced intensity and nonmyeloablative conditioning for adults with acute myeloid leukemia undergoing allogeneic hematopoietic cell transplantation in first or second remission.

Reduced intensity conditioning (RIC) and nonmyeloablative (NMA) conditioning regimens have expanded use of allogeneic hematopoietic cell transplantation (HCT) in AML to include older and medically less-fit patients, but relative efficacies and toxicities remain poorly defined. Here, we analyzed outcomes from 343 adults transplanted in remission after RIC (n = 137) or NMA (n = 206) conditioning between 2006 and 2021. The characteristics of RIC and NMA HCT patients were similar except that RIC patients were younger and their time between most recent remission achievement and allografting was shorter. There were no significant differences in relapse risk, relapse-free survival (RFS), overall survival (OS), and non-relapse mortality (NRM) between RIC and NMA HCT patients, both overall (relapse: hazard ratio [HR] = 0.80, P = 0.27; RFS: HR = 0.93, P = 0.61; OS: HR = 0.93, P = 0.66; NRM: HR = 1.13, P = 0.59) and when patients were stratified by pre-HCT measurable residual disease (MRD) status. After multivariable adjustment, there was no statistically significant association between conditioning intensity and relapse (HR = 0.69, P = 0.088), RFS (HR = 0.86, P = 0.37), OS (HR = 0.89, P = 0.49), or NRM (HR = 1.37, P = 0.19). In this non-randomized cohort of adults undergoing allografting for AML in first or second remission at our center, we could not detect statistically significantdifferences in outcomes between those assigned to RIC and those assigned to NMA conditioning.

Pulmonary Function before and after Hematopoietic Cell Transplant in People Living with Sickle Cell Disease

Background People living with sickle cell disease (SCD) have increasing access to hematopoietic cell transplant (HCT). However, there are insufficient data on the effects of HCT, and other curative therapies, on pulmonary function in this population. Unlike patients with cancer, individuals with SCD have a lifelong genetic condition with recurring vaso-occlusive events and inflammatory processes known to cause significant pulmonary morbidity. Therefore, studies that report lung function after transplant in cancer survivors may not apply to individuals with SCD. Indeed, data are needed to evaluate the late lung health effects of HSCT in the SCD population. We hypothesized that percent predicted forced expiratory volume in one second (FEV-1), a pulmonary function outcome measure which has been found to be associated with early death in SCD, would be stable one-year post HCT. Methods People living with SCD who underwent first-time HCT, including HLA-matched sibling and haploidentical HCT, at the NIH Clinical Center from 2004 to 2019 with Hgb SS, SC, Sbeta+ thalassemia, and Sbeta0 thalassemia with available pulmonary function data were included in this study. Global Lung Function Initiative reference equations were used, and descriptive statistics were calculated for FEV-1, forced vital capacity (FVC), FEV-1/FVC, total lung capacity (TLC), and diffusing capacity of the lungs for carbon monoxide (DLCO) for patients with pre- and one-year post-HCT. Generalized estimating equations were employed to evaluate inter-individual changes in percent-predicted FEV-1 pre- and one-year post-HCT. Results Seventy-seven people living with SCD (HgbSS n=69 [90%], HgbSC 2 [3%], HbSbeta+ 2 [3%], HbSbeta0 4 [5%]) underwent non-myeloablative HCT (HLA-matched sibling n=53 [69%%] and haplo-identical n=24 [31%]) . Of these, 42 (42%) were female with median (25th, 75th percentile) age 31.0 (24.0, 37.0) years; there were four patients less than 18 years old. Median (25th, 75th) percent predicted forced expiratory volume in one second (FEV-1) was 68.4% (61.3%, 79.1%) at baseline and 69.9% (60.5%, 78.2%) one-year post-HCT; baseline median (25th, 75th) percent predicted DLCO was 48.0 (43.0, 54.0) and 48.0 (45.0, 56.0) one-year post-HCT. There was no significant change in percent predicted FEV-1 (0.88 (0.78, 2.55; p=0.30) one-year post-HCT. Conclusion Pulmonary function remained stable in people living with SCD one-year post-HCT. Allogeneic HCT for SCD may cease the cycle of vaso-occlusive injury to the lung tissue and airway system and prevent continued damage. In addition, it is possible that without the ongoing injury, lung healing may also occur over time. Multicenter studies with longer follow-up are needed to evaluate the long-term lung health effects of HCT on individuals with SCD. Figure 1View largeDownload PPTFigure 1View largeDownload PPT Close modal

Pulmonary function in children and adolescents with sickle cell disease after nonmyeloablative hematopoietic cell transplantation.

Pulmonary complications are common in sickle cell disease (SCD). The use of standard myeloablative conditioning regimens may increase the risk of lung injury. We report serial pulmonary function testing (PFT) outcomes in children with SCD who underwent a matched-sibling donor hematopoietic cell transplantation (HCT) using nonmyeloablative (NMA) protocol. This is a retrospective chart review describing pulmonary outcomes in pediatric patients post HCT. The conditioning regimen consisted of alemtuzumab and a single fraction of 300cGy of total body irradiation (TBI), and sirolimus for graft-versus-host disease (GVHD) prophylaxis. Serial PFT testing was performed pre and post HCT. The evaluated pulmonary measures included: forced vital capacity (FVC), forced expiratory volume in the first second (FEV1 ), FEV1 /FVC, and forced expiratory flow (FEF25-75 ). Twelve subjects were included in the analysis. All had HbSS genotype, and five of the 12 patients had one or more episodes of acute chest syndrome prior to HCT. Serial PFT measures were completed per patient. No patient was diagnosed with chronic GVHD of any organ post HCT. The baseline median FVC, FEV1 , FEV1 /FVC, and FEF25-75 were within the normal range and remained relatively unchanged post HCT. A linear mixed effects model, adjusting for gender and time from HCT, suggested no significant relationship between HCT and PFT parameters, including FVC, FEV1 , and FEV1 /FVC. Interestingly, the FEF25-75 results exhibited a shift in the means post HCT (pre-HCT 86.2% predicted and post-HCT 93.05% predicted, p-value = .018). Our study suggests that HCT in children with SCD may prevent the anticipated decline in pulmonary function over time.

439 - Stable Pulmonary Function in Children and Adolescents with Sickle Cell Disease after Nonmyeloablative Hematopoietic Cell Transplantation- the Calgary Experience

439 - Stable Pulmonary Function in Children and Adolescents with Sickle Cell Disease after Nonmyeloablative Hematopoietic Cell Transplantation- the Calgary Experience

Consolidative CD8+ Memory T-Cell Donor Lymphocyte Infusion Augments Donor Chimerism with a Low Risk of Gvhd Following Matched Related Donor Hematopoietic Cell Transplantation

IntroductionMixed chimerism (MC) after allogeneic hematopoietic cell transplantation (HCT) is associated with an increased risk of disease relapse. An unmanipulated donor lymphocyte infusion (DLI) can convert MC to full donor chimerism (FC) but can also precipitate severe acute graft-versus-host disease (GVHD). We recently showed in a phase I trial that a subset of purified CD8+CD44hi memory T cells derived from HLA-matched sibling donors may induce graft-versus-tumor reactions in patients with relapsed hematologic malignancies following allogeneic HCT with objective responses in 25% of patients and a low incidence of GVHD (Muffly et al, 2016 ASH Abstract #4615). Here we report on the preliminary results of a phase II study using CD8+ memory T-cell DLI as consolidative therapy early after transplant for patients with MC to assess the rate of conversion to FC, the risk of GVHD, and the prevention of disease relapse.MethodsPatients: Eligible patients were 18-75 years of age with a high-risk or refractory hematologic malignancy who underwent nonmyeloablative allogeneic HCT from a matched sibling donor using total lymphoid irradiation and antithymocyte globulin conditioning, and who had MC (<95% donor CD3+ cells) at day +30. Patients were without acute GVHD at the time of the CD8+ memory T-cell DLI and were on tacrolimus or cyclosporine monotherapy for GVHD prophylaxis at the time of DLI.CD8+ memory T cells: Phenotypic CD8+ memory T cells were isolated from matched sibling donors following two days of unmobilized apheresis using a tandem immunomagnetic selection strategy including CD45RA depletion followed by CD8+ enrichment on the CliniMACS Plus instrument. Cell purity was determined by flow cytometry analysis. CD8+ memory T cells were infused between day +30 and day +70 at a dose of 5 x 106 cells/kg, which was determined in the prior phase I trial to be the maximum dose that could be collected from two apheresis products.Study Endpoints: Following DLI, patients were assessed for the development of acute and chronic GVHD, conversion from MC to FC (≥95% donor CD3+ chimerism) within three months, and for disease relapse. Secondary outcomes included the incidence of adverse events, invasive infections, non-relapse mortality, overall survival, and disease-free survival.ResultsPatient characteristics and outcomes after DLI are summarized in Table 1. Nine patients received CD8+ memory T-cell DLI at a median of 42 days post-HCT (range 33-62 days). The cell product was highly enriched for CD8+/CD45RA-/CD45RO+ cells, and flow analysis confirmed the selected cells were phenotypically consistent with the CD8+ T effector memory subset with a post enrichment purity of ≥96% (range 96.4-98.8%) in all patients. Median age of the patients was 60 years (range 30-71 years) with a median follow up of 306 days post-HCT (range 131-747 days). Diagnoses included non-Hodgkin lymphoma (n=5), acute myeloid leukemia (n=2), myelodysplastic syndrome (n=1), and Hodgkin lymphoma (n=1). Median donor CD3+ chimerism at day +30 was 88% (range 23-93%).CD8+ memory T-cell DLI was well tolerated with no adverse events. Six of the nine patients (67%) converted from MC to FC within three months post-DLI with a median donor CD3+ chimerism of 97% (range 95-100%). Two patients developed grade II acute GVHD (skin only) that was treated to resolution. One patient developed mild chronic GVHD. No patients developed severe acute GVHD (grade III-IV) or extensive chronic GVHD. No patients developed invasive infections, and non-relapse mortality was 0%. Of the three patients who failed to convert from MC to FC, two had residual disease at the time of transplant, one of whom had early disease relapse at day +88. Four of the six patients who converted to FC were in complete remission (CR) at the last follow up. One of the three patients with persistent MC was in CR at the last follow up. For the entire study cohort, the median disease-free survival is 1.1 years and the median overall survival is not yet reached.ConclusionsConsolidative CD8+ memory T-cell DLI following nonmyeloablative matched related donor HCT is well tolerated, converted six of nine patients (67%) with MC to FC, and was associated with a low rate of GVHD with no cases of severe acute or extensive chronic GVHD in this study. Patients continue to accrue to trial, and further accrual and longer follow up are needed to determine if this strategy will reduce relapse rates for high-risk patients with MC. [Display omitted] DisclosuresNo relevant conflicts of interest to declare.

Alemtuzumab/Low Dose TBI Conditioning Facilitates Stable Long-Term Donor Hematopoietic Cell Engraftment from Sibling Donors in Children with Sickle Cell Disease

BackgroundHematopoietic stem cell transplant (HCT) is the only established curative approach for sickle cell disease (SCD). Due to the high toxicity profile of myeloablative regimens, some adult patients with severe sickle cell anemia are not eligible for such therapy. However, a new adult nonmyeloablative protocol (National Institutes of Health, NIH protocol) described by Hsieh et al. (2014) was able to achieve a curative degree of mixed donor chimerisms with minimal transplant-related complications. The NIH protocol includes alemtuzumab, low dose total-body irradiation (TBI) of 300 cGy, and prolonged sirolimus for graft-versus-host disease (GVHD) prophylaxis for those recipients with a matched-sibling donor (MSD).The Alberta Children's Hospital has adopted this published regimen for MSD HCT in adolescents and younger children with SCD since the NIH data was presented in 2013.To our knowledge, there is no published literature describing the application of the NIH protocol in a pediatric population; the youngest subject in the referenced adult study was 16 years of age.MethodsThis retrospective cohort describes outcomes of nonmyeloablative allogeneic MSD HCT in children and adolescents with SCD who underwent HCT between 2013-2017. Fourteen patients with SCD under 18 years of age at the time of HCT received the NIH conditioning regimen. HLA-matched siblings had either normal hemoglobin or sickle cell trait and of the appropriate weight/height to donate peripheral blood stem cells (PBSCs) without blood priming of the apheresis circuit.The conditioning regimen consisted of alemtuzumab 1.0 mg/kg administered subcutaneously daily divided over five days (Days -7 to Day -3). Patients received a single TBI dose of 300 cGy on day -2, with testicular shielding for male recipients. GVHD prophylaxis consisted of a sirolimus loading dose of 3mg/m2/dose (PO) on day -1, followed by 1mg/m2/dose once daily starting on Day 0. PBSCs were typically collected on Day -3 to allow for cancellation of the HCT before TBI in the event of failed mobilization. Unmanipulated PBSCs were infused fresh on Day 0. Patients were eligible for early discharge post-transplant even prior to neutrophil engraftment. Weaning of sirolimus was initiated no earlier than 1-year post-HCT and if donor T-cell chimerisms were greater than 50%. Treatment failure was defined as clinical manifestation of SCD, failure to start weaning sirolimus by 2-years post-HSCT, or a failure to complete a sirolimus wean by 2.5 years post-HCT.ResultsFourteen patients (3 M,11 F) are included on this retrospective study. Median age at HCT was 11 years. Post-HCT follow-up ranges from 1 to 48 months (median 17). There were no failed donor stem cell mobilizations. Patients received a median of 11.4x106 CD34 cells per kilogram of body weight.All patients were discharged home prior to neutrophil engraftment with a median hospital stay of 7 days (range 5-19). All patients had donor neutrophil engraftment at a median of 21 days (mean= 22, SD= 4) and platelet engraftment at median of 14 days (mean= 12, SD= 14). Six patients never required platelet transfusion. Readmission after initial discharge was required in 9/14 patients with an average of 1 readmission/patient (range 1-4), typically for fever or CMV pre-emptive therapy. One patient developed septic shock with Acinetobacter ursingii related to poor central line care at home.All patients are currently alive. Post-HCT, there have been no cases of graft failure, sickling crises, veno-occlusive disease, idiopathic pneumonia syndrome, cerebral hemorrhage, PRES, or PTLD. Although 6/14 patients had CMV reactivation, only 3 patients required pre-emptive therapy. None had CMV/EBV disease.There were no treatment failures as defined by protocol. Currently, 8 patients are off sirolimus, 1 is currently weaning, with no secondary graft failure events. Only 2 patients over 1-year post-HCT have not initiated sirolimus weaning. There have been no cases of acute or chronic graft-versus-host disease.ConclusionsNonmyeloablative conditioning regimen is safe and effective as curative therapy for children with SCD. Outpatient management pre-engraftment was feasible in this population. In contrast to adult data, sirolimus weaning was possible in most patients by one-year post-HCT. Graft integrity was maintained after discontinuation of sirolimus in all eligible patients. [Display omitted] DisclosuresGuilcher:Patient Access Solutions: Consultancy; Canada Market Research: Consultancy; Jazz pharmaceuticals: Consultancy. Bruce:Bristol-Meyers-Squibb - consultant 2016-2017: Consultancy; ovartis consult and ad board 2015-2016: Consultancy; Apopharma 2015: Consultancy.

Anti-thymocyte globulin with CsA and MMF as GVHD prophylaxis in nonmyeloablative HLA-mismatched allogeneic HCT.

Nonmyeloablative regimens are used for allogeneic hematopoietic cell transplantation (HCT) of older or medically unfit patients, but successful outcome is still hindered by graft-versus-host disease (GVHD), especially in the setting of HLA-mismatched HCT. New GVHD prophylaxis strategies are emerging, including the triple drug strategy, that improve the GVHD-free and relapse-free survival (GRFS). Because the impact of ATG in HLA-mismatched Flu-TBI-based nonmyeloablative HCT has not been investigated, we did a retrospective analysis in three Dutch centers. 67 patients were evaluable, with a median age of 56 years. Overall survival, relapse-free survival and GRFS at 4 years were 52%, 43%, and 38%, respectively. NRM findings and cumulative incidence of relapse at 4 years were 26% and 31%, respectively. At 1-year grade II-IV had occurred in 40% of the patients, and the incidence of moderate-severe chronic GVHD incidence was 16%. Acknowledging the limitations of retrospective analyses, we conclude that the use of ATG for HLA-mismatched truly nonmyeloablative Flu-TBI HCT is feasible and results in acceptable long term outcomes, especially with regards to GRFS. We consider ATG in combination with cyclosporin and mycophenolate mofetil as an alternative for the triple drug strategy that uses sirolimus for GVHD prophylaxis in this particular setting.

Safety and efficacy of a nonmyeloablative pretransplant conditioning regimen using total lymphoid irradiation with volumetric modulated arc therapy in healthy dogs: A pilot study.

Allogeneic hematopoietic cell transplantation (HCT) has been an effective treatment for human patients with haematological malignancies (Baron & Storb, 2006; Bair et al., 2020; Copelan et al., 2019). However, the optimal pretransplant conditioning treatment is unclear in canine allogeneic HCT. This pilot study aimed to evaluate the safety and efficacy of total lymphoid irradiation (TLI) with volumetric modulated arc therapy (VMAT) for a nonmyeloablative HCT conditioning. Six healthy dogs were treated with 8 or 12 Gy TLI using VMAT. Haematological and physical changes were recorded over 8 weeks. To assess the effect of peripheral lymphocyte condition, lymphocyte subset and proliferative ability were examined. At the end of the experiment, necropsy was performed. All dogs showed mild‐to‐moderate neutropenia and thrombocytopenia, and these haematological changes resolved spontaneously. One dog treated with 8 Gy TLI developed transient cutaneous infection. No major complication was seen in the other seven dogs. Myelocytes and erythroblast cytopenia of bone marrow were detected in two dogs treated with 12 Gy TLI. This study is the first report of TLI using VMAT in dogs, and results suggest that this regimen is a feasible nonmyeloablative treatment.

Long-term Outcomes with Nonmyeloablative HLA-Identical Related Hematopoietic Cell Transplantation Using Tacrolimus and Mycophenolate Mofetil for Graft-versus-Host Disease Prophylaxis

Nonmyeloablative allogeneic hematopoietic cell transplantation (HCT) from HLA-identical related donors using cyclosporine (CSP) and mycophenolate mofetil (MMF) for postgrafting immunosuppression is effective therapy for hematologic cancers. However, graft-versus-host-disease (GVHD) remains a major cause of morbidity and mortality. Pilot data suggested lower acute GVHD incidence with tacrolimus/MMF compared to historical experience using CSP/MMF after nonmyeloablative HCT. In a phase II multicenter trial, we evaluated the effect of tacrolimus/MMF for GVHD prophylaxis after HLA-identical related donor peripheral blood HCT in patients with hematologic malignancies (n=150) using conditioning with 2 Gy total body irradiation (TBI) for patients with a preceding (within 6 months) planned autologous HCT (n=50) or combined with 90 mg/m2 fludarabine for those without recent autologous HCT (n=100). Oral tacrolimus was given from days -3 to 56 (tapered by day +180 if no GVHD). Oral MMF was given from days 0 to 27. Patient median age was 57 (range, 20 to 74) years. The cumulative incidences (CI) of day 100 grade II to IV and III to IV acute GVHD were 27% and 4%, respectively. With median follow-up of 10.3 (range, 3.1 to 14.5) years, the 5-year CI of chronic extensive GVHD was 48%. One-year and 5-year estimates of nonrelapse mortality, relapse/progression, survival, and progression-free survival were 9% and 13%, 35% and 50%, 73% and 53%, and 56% and 37%, respectively. GVHD prophylaxis with tacrolimus/MMF resulted in a low risk of acute GVHD and compared favorably with results from a concurrent trial using CSP/MMF. A randomized phase III trial to investigate tacrolimus/MMF versus CSP/MMF in nonmyeloablative HCT is warranted.

Sirolimus Combined with Cyclosporine (CSP) and Mycophenolate Mofetil (MMF) As Graft-Vs-Host Disease (GVHD) Prophylaxis after Nonmyeloablative (NMA) Hematopoietic Cell Transplantation (HCT) Using HLA Class I or Class II Antigen Mismatched Donors: Results from a Phase II Multi-Center Trial

The success of HCT is highly dependent on the availability of compatible donors. Depending on ethnicity, fully HLA-matched donors cannot be identified for 25-84% of patients. In an effort to extend the use of HLA-mismatched donors to patients only eligible for NMA HCT, we previously demonstrated that engraftment and long-term survival can be achieved using CSP and MMF immunosuppression after HLA class I antigen-mismatched HCT (Nakamae et al, 2010). Based on this trial, we designed a phase II multi-center trial to assess the efficacy of GVHD prophylaxis with triple immune suppression with CSP, MMF and sirolimus after HLA class I or class II antigen mismatched NMA HCT (fludarabine 90mg/m2 and 2-3 Gy TBI). Patients ineligible for high-dose conditioning with only HLA class I or class II antigen mismatched donors available were eligible for inclusion. The primary objective was to determine whether the incidence of grades 2-4 acute GVHD can be reduced to less than the historical rate of 70% with the triple-immunosuppression. The evaluation was planned to be carried out separately among HLA class I and class II mismatched patients. Secondary objectives included day 100 non-relapse mortality (NRM) and grades 3-4 acute GVHD. CSP was started on day -3 and continued to day +150 and then tapered off by day +180. Sirolimus was started on day -3 through day +180 and then tapered off by day +365. MMF was given thrice daily from days 0 to +30 then twice daily to day +100 and tapered off by day +150. A total of 76 patients were enrolled and received conditioning with fludarabine and TBI followed by infusion of unmodified G-CSF mobilized peripheral blood stem cell (PBSC) grafts. The median age was 63 (21-76) years, and median follow-up of surviving patients was 47 (4-94) months. 51 (67%) and 25 (33%) of patients had HLA class I or II mismatches, respectively. Transplant demographics are summarized in Table 1. Sustained engraftment was observed in all but 1 (1%) patient with myeloproliferative disease who had autologous recovery after transplant. Median number of days with neutrophils &amp;lt; 500 cells/ml and platelets &amp;lt;20,000 platelets/ml were 14 (0-67) and 2 (0-70), respectively. Full donor T-cell chimerism (&amp;gt;95% donor) was achieved in 48% and 63% at days +28 and 84, respectively. Outcomes are summarized in Table 2.The cumulative incidence of grade 2-4 acute GVHD at day +100 was 34% in all patients (29% and 48% after HLA class I and class II mismatched, respectively; p=0.09) (Figure 1), with only 3% of patients experiencing grade 3 acute GVHD and no patients experienced grade 4 acute GVHD. NRM at day +100, 2 years and 4 years were 4%, 15% and 17%, respectively (HLA class I vs class II: p=0.58) (Figure 2). The 2- and 4-year cumulative incidences of chronic GVHD were 54% and 57% (HLA class I vs. class II: p=0.11). Relapse incidences at 2 and 4 years were 26% and 31% with no differences between the class I and II mismatched patients (p=0.54) (Figure 2). The cumulative incidences at 2 and 4 years of progression-free survival (PFS) were 59% and 52%, and overall survival (OS) were 68% and 62% with no differences between HLA class I and II mismatches for PFS or OS (p=0.64 and 0.90, respectively) (Figure 2). Compared to historical data from our previous trial using MMF and CSP prophylaxis alone in patients with HLA class I mismatches, the addition of sirolimus to GVHD prophylaxis with CSP/MMF improved outcomes both after HLA class I or class II mismatched HCT (Nakamae et al, 2010). In a comparison of HLA class I mismatches only, day +100 grade 2-4 acute GVHD was reduced from 69% in the historical cohort to 29% in the current trial. The relapse incidences were comparable, while 2-year NRM decreased from 47%, in historical controls, to 16%, and translated into improved OS from 26% to 67% at 2 years in the current study. No difference was observed in the incidence of chronic GVHD. Furthermore, in the current trial, similar results were seen using HLA class II mismatched donors, suggesting that the triple drug immunosuppression could be used with either type of mismatch. The current trial demonstrates that triple drug immunosuppression with CSP, MMF, and sirolimus is safe and effective in reducing both acute GVHD and NRM after NMA HCT with either HLA class I or class II antigen mismatched donors. Disclosures Maloney: Juno Therapeutics: Honoraria, Patents &amp; Royalties: patients pending , Research Funding; Celgene,Kite Pharma: Honoraria, Research Funding; BioLine RX, Gilead,Genentech,Novartis: Honoraria; A2 Biotherapeutics: Honoraria, Other: Stock options . OffLabel Disclosure: In the current study cyclosporine, mycophenolate mofetil and sirolimus are used to prevent graft versus host disease after allogeneic hematopoietic cell transplantation. Fludarabine is used together with total body irradiation as at part of the conditioning prior to allogeneic hematopoietic cell transplantation.

Rabbit Antithymocyte Globulin Serum Levels: Factors Impacting the Levels and Clinical Outcomes Impacted by the Levels

Antithymocyte globulin (ATG) levels and clearance vary significantly among patients receiving the same weight-based dose of ATG. To date, ATG area under the curve (AUC), its determinants, and its impact on clinical outcomes have been examined in pediatric hematopoietic cell transplant (HCT) and adult nonmyeloablative HCT. Here we set out to examine ATG AUC in 219 uniformly treated adults undergoing myeloablative allogeneic HCT at our institution. Sera were collected for the determination of pre- or post-HCT ATG AUC. The lowest quintiles of pre- and post-HCT AUC were associated with inferior chronic graft-versus-host disease (GVHD) and relapse-free survival (cGRFS) and a higher risk of acute GVHD, respectively. The highest pre- or post-HCT ATG AUC quintiles were not associated with risk of death, nonrelapse mortality, or relapse. Factors most strongly associated with AUC were day –2 recipient absolute lymphocyte count, body mass index (BMI), and graft lymphocyte content. To achieve ideal pre-HCT AUC (avoiding low AUC to maximize cGRFS) in this HCT setting, ATG dosing will need to take into consideration recipient weight, BMI, and blood and graft lymphocyte counts. Further studies are required to develop a modern ATG dosing schema and to demonstrate that adjusting ATG dose to target a particular AUC is feasible and leads to improved outcomes.

First-in-Human CLL1-CD33 Compound CAR T Cell Therapy Induces Complete Remission in Patients with Refractory Acute Myeloid Leukemia: Update on Phase 1 Clinical Trial

▪BackgroundCD19-specific chimeric antigen receptor (CAR) T cell therapy has achieved high efficacy in acute lymphoblastic leukemia patients. However, the treatment of acute myeloid leukemia (AML) has remained a particular challenge due to the heterogeneity of AML bearing cells, which renders single antigen targeting CAR T cell therapy ineffective. CLL1 and CD33 are often used as targets for AML CAR T cell therapy. CLL1 is associated with leukemia stem cells and disease relapse, and CD33 is expressed on the bulk AML disease. Previously, we demonstrated the profound anti-tumor activity of CLL1-CD33 compound CAR (cCAR) T cells. Here we present the efficacy of cCAR in preclinical study and update the success in level 1 dose escalation clinical trial on relapsed/refractory AML patients.MethodsWe engineered a cCAR comprising of an anti-CLL1 CAR linked to an anti- CD33 CAR via a self-cleaving P2A peptide and expressing both functional CAR molecules on the surface of a T-cell cell. We tested the anti-leukemic activities of CLL1-CD33 cCAR using multiple AML cell lines, primary human AML samples, human leukemia cell line (REH cells) expressing either CLL1 or CD33, and multiple mouse models. An alemtuzumab safety switch has also been established to ensure the elimination of CAR T cells following tumor eradication. Children and adults with relapsed/refractory AML were enrolled in our phase 1 dose escalation trial with primary objective to evaluate the safety of cCAR and secondary objective to assess the efficacy of cCAR anti-tumor activity.ResultsCo-culture assays results showed that cCAR displayed profound tumor killing effects in AML cell lines, primary patient samples and multiple mouse model systems. Our preclinical findings suggest that cCAR, targeting two discrete AML antigens: CLL1 and CD33, is an effective two-pronged approach in treating bulk AML disease and eradicating leukemia stem cells. Patients enrolled in the phase 1 dose escalation trial have shown remarkable response to cCAR treatment. Noticeably, a 6-yr-old female patient diagnosed with a complex karyotype AML including FLT3-ITD mutation had achieved complete remission. The patient was diagnosed with Fanconi anemia, which had progressed to juvenile myelomonocytic leukemia and eventually transformed into AML. The patient had been resistant to multiple lines of treatments, including 5 cycles of chemotherapy with FLT3 inhibitor prior to receiving cCAR. Before the treatment, patient's leukemia blasts comprised 73% of the peripheral blood mononuclear cells and 81% of the bone marrow. Patient underwent lymphodepletion therapy (Fludarabine and Cyclophosphamide) prior to cCAR infusion. Two split doses, each consisting of 1x106/kg CAR T cells, were infused on day 1 and day 2 respectively. On day 12, while leukemia blast still counting up to 98% of the bone marrow (Fig. 1A), robust CAR T cell expansion was detected in both peripheral blood and bone marrow. On day 19, patient achieved MRD- complete remission with bone marrow aspirates revealing complete ablation of myeloid cells (Fig. 1B). Flow cytometry confirmed the absence of leukemia blasts and showed that CAR T cells comprised 36% of the PBMC and 60% of the bone marrow. The patient later underwent non-myeloablative hematopoietic cell transplantation with less toxicities compared to conventional total body radiation and high dose chemotherapies. Updated results on other patients enrolled in this clinical trial including adverse events will be presented.ConclusionOur first-in-human clinical trial demonstrates promising efficacy of cCAR therapy in treating patients with relapsed/ refractory AML. cCAR is able to eradicate leukemia blasts and leukemia stem cells, exerting a profound tumor killing effect that is superior to single target CAR T cell therapies. cCAR is also shown to induce total myeloid ablation in bone marrow, suggesting that it may act as a safer alternative to avoid the severe toxicities caused by standard bone marrow ablation regimens without sacrificing the anti-tumor efficacy. This strategy will likely benefit patients who are unable to tolerate total body radiation or high dose chemotherapies. In addition to AML, cCAR also has the potential to treat blast crisis developed from myelodysplastic syndrome, chronic myeloid leukemia, and chronic myeloproliferative neoplasm. DisclosuresPinz:iCell Gene Therapeutics LLC: Employment. Ma:iCAR Bio Therapeutics Ltd: Employment. Wada:iCell Gene Therapeutics LLC: Employment. Chen:iCell Gene Therapeutics LLC: Employment. Ma:iCell Gene Therapeutics LLC: Employment. Ma:iCell Gene Therapeutics LLC, iCAR Bio Therapeutics Ltd: Consultancy, Equity Ownership, Research Funding.

Induction of Tolerance Towards Solid Organ Allografts Using Hematopoietic Cell Transplantation in Large Animal Models.

The application of hematopoietic cell transplantation for induction of immune tolerance has been limited by toxicities associated with conditioning regimens and to graft-versus-host disease (GVHD). Decades of animal studies have culminated into sufficient control of these two problems, making immune tolerance a viable alternative to life-long application of immunosuppressive drugs to prevent allograft rejection. Studies in mice have paved the way for the application of HCT with limited toxicity in large animal models. Resultant studies in the pig, dog, and ultimately the nonhuman primate have led to appropriate methods for achieving nonmyeloablative irradiation protocols, dose, and timing of post-grafting immunosuppressive drugs, monoclonal antibody therapy, and biologicals for costimulatory molecule blockade. The genetics field has been extensively evaluated in appreciation of the ultimate need to obtain organs from MHC-mismatched unrelated donors. Nonmyeloablative conditioning regimens have been shown to be successful in inducing immune tolerance across all three animal models. Postgrafting immunosuppression is also important in assuring sustained donor hematopoiesis for tolerance. Donor chimerism need not be permanent to establish stable engraftment of donor organs, thereby essentially eliminating the risk of GVHD. Using nonmyeloablative HCT with monoclonal antibody immunosuppression, the kidney has been successfully transplanted in MHC-mismatched nonhuman primates. Nonmyeloablative HCT for the establishment of temporary mixed chimerism has led to the establishment of stable tolerance against solid organ allografts in large animal models. The kidney, considered a tolerogenic organ, has been successfully transplanted in the clinic. Other organs such as heart, lung, and vascularized composite allografts (face and hands), remain distant possibilities. Further study in large animal models will be required to improve tolerance against these organs before success can be attained in the clinic.