Severe Chronic GVHD Research Articles

Oral acute graft‐versus‐host disease

AbstractOral acute graft‐versus‐host disease (aGVHD) is rare and with no diagnostic criteria. We report a case of oral aGVHD with three clinical phases. A self‐limited prodrome of largely subjective oral symptoms was followed by concurrent oral and upper gastrointestinal aGVHD. Six months after transplantation, the patient was diagnosed with severe oral and upper gastrointestinal chronic GVHD. We compared the salivary microbiota of our patient at the time of diagnosis of aGVHD with 50 contemporaneous transplant recipients and found no evidence for oral microbiota involvement in pathogenesis. This in‐depth N‐of‐1 analysis reveals novel aspects of oral aGVHD pathogenesis.

Selected donor CD34(+) cell boosts for salvage treatment of poor graft function following allogeneic hematopoietic stem cell transplantation in primary myelofibrosis: 3 cases report

A retrospective analysis was conducted on three patients with primary myelofibrosis who underwent allogeneic hematopoietic stem cell transplantation (allo-HSCT) at Shanghai Zhaxin Traditional Chinese and Western Medicine Hospital from 2020 to 2023. They subsequently developed poor graft function. The patients received selected donor CD34(+) cell boosts as salvage therapy. There were two male patients and one female patient, with a median age of 68 (39-69) years. The median time from allo-HSCT to the selected donor CD34(+) cell boost was 83 (56-154) days. The median infusion of selected donor CD34(+) cells was 7.67 (7.61-9.06) ×10(6)/kg, with a CD34(+) cell purity of 97.76% (96.50%-97.91%) and a recovery rate of 70% (42%-75%) . Hematological recovery was achieved in two cases. No acute GVHD was observed in any of the three patients. One case of moderate oral chronic GVHD was noted. Selected donor CD34(+) cell boosts for the treatment of poor graft function after allo-HSCT in primary myelofibrosis was effective and no severe acute or chronic GVHD was observed.

Efficacy and safety of outpatient fludarabine, cyclophosphamide, and rituximab based allogeneic hematopoietic cell transplantation in adults with severe aplastic anemia

The age effect in severe aplastic anemia (SAA) following allogeneic hematopoietic cell transplantation (HCT) favors the use of reduced intensity conditioning (RIC) regimens in older adults. We implemented a non-myeloablative regimen consisting of fludarabine, cyclophosphamide, and rituximab (FCR) to improve HCT outcomes in SAA. Patients who underwent first HCT for SAA utilizing an FCR regimen between January 2016 and May 2022 were included. Outcomes analyzed included time to engraftment, incidence of graft failure, GVHD, viral reactivation, disease recurrence, and GVHD-free, relapse-free survival (GRFS). Among 24 patients included, median age was 43.5 years (22–62) and a variety of donor types and stem cell sources were represented. At median follow-up of 26.9 months (2.4–72.7), no cases of grade III-IV acute (aGVHD) or severe chronic GVHD (cGVHD) were recorded. Viral reactivation was minimal, and there were no cases of graft failure or PTLD, with 100% disease-free and overall survival at last follow up. The estimate of 1-year GRFS was 86.3% (95% CI: 72.8–100%), with moderate cGVHD accounting for all events. The FCR regimen in SAA was well tolerated, even in older adults, with 100% disease-free survival with low GVHD and infection rates. These encouraging findings should be validated in larger prospective trials.

The metabolic profiles of patients with autologous hematopoietic stem cell transplantation with graft versus host disease.

e24010 Background: Graft versus host disease (GvHD) is a serious inflammatory complication of hematopoietic stem cell transplant (HSCT) and a leading contributor to morbidity and mortality in HSCT patients. Previous studies have identified increased risk of cardiovascular death in HSCT patients who develop GvHD and characterized echocardiographic changes associated with chronic GvHD. However, details of the metabolic derangements in HSCT patients who develop GvHD remain poorly studied. Methods: This is a retrospective study of patients who underwent HSCT at H. Lee Moffitt Cancer Center and later developed severe chronic GVHD at 1 year. Metabolic data including weight, blood pressure, and lipid profile were recorded at baseline and at 1 year after HSCT. Results: Between January 1, 2010, and December 31, 2020, 1805 patients who received allogeneic HSCT were reviewed. At 1 year, 371 patients (20.5%) had severe GVHD. Compared to baseline, patients had higher total cholesterol (209 vs 183 mg/dL, P < 0.001), high-density lipoprotein (61 vs 47 mg/dL, P < 0.001), and triglycerides (211 vs 163 mg/dL, P < 0.001) at 1 year of follow up. Low-density lipoprotein values were not significantly different (107 vs 102 mg/dL, p = 0.15). Patient weight was significantly less at 1 year (163 vs 183 lbs, p < 0.001). Although blood pressure was not significantly different between baseline and 1 year, more patients were placed on antihypertensive medications at 1 year (50% vs 31%). Conclusions: This study finds that patients who undergo allogeneic HSCT and subsequently develop severe GVHD are at risk for metabolic derangements that may worsen cardiovascular health. We specifically demonstrate an increased need for antihypertensive therapy and elevations in total cholesterol and serum triglycerides. Further research is needed on the impact of these derangements on major adverse cardiac events and the role of cardiovascular risk factor management in these patients.

Improved Outcome of Allogeneic Transplantation in Older Patients Treated for Myeloid Malignancies Using Post-Transplantation Cyclophosphamide and Reduced Duration of Immune Suppression

BackgroundAllogeneic stem cell transplantation (alloSCT) offers curative potential for older patients with myeloid malignancies. We evaluated the efficacy and safety of alloSCT using post-transplantation cyclophosphamide (PTCy) in combination with a very short duration of immune suppression (IS) in this population. MethodsWe retrospectively analyzed 92 consecutive patients aged 65 years and older who underwent an alloSCT for myeloid malignancies between February 2018 and December 2022 at our institution. Data on patient characteristics, treatment modalities, and outcomes were collected. ResultsNinety-two patients received an alloSCT with PTCy-based GVHD prophylaxis. The majority had minimal comorbidities and were diagnosed with acute myeloid leukemia (AML). Patients mostly received conditioning regimens with low to intermediate TCI scores. In 43% of patients, IS could be permanently stopped at day +90, resulting in a median time of IS of 2.93 months in high-risk patients. At a median follow-up of 21.3 months, the 1- and 2-year overall survival rates were 89% and 87%, respectively. Relapse-free survival rates were 88% and 84% at 1 and 2 years, respectively. The 1- and 2-year cumulative incidences of relapse were 8% and 13%, while transplant-related mortality (TRM) estimates were 9% at both time points. Acute GVHD grade 3-4 occurred in 7% within the first 180 days and severe chronic GVHD in 6% of patients. This all resulted in a 1- and 2-year graft versus host and relapse free survival (GRFS) of 74% and 70%, respectively. ConclusionAlloSCT using PTCy in combination with a short duration of IS in older patients with myeloid malignancies demonstrates favorable survival outcomes due to low relapse rates and a low TRM. The low incidence of relapse and acceptable rates of graft-versus-host disease suggest the efficacy and safety of this approach. Further studies are warranted to validate these findings and optimize transplant strategies for older patients with myeloid malignancies.

Outcomes with allogeneic stem cell transplant using cryopreserved versus fresh hematopoietic progenitor cell products

BackgroundAllogeneic hematopoietic stem cell transplant (alloHSCT) is a mainstay of treatment for hematologic malignancies such as acute leukemias and aggressive lymphomas. Historically, fresh hematopoietic progenitor cell (HPC) products have been preferred to cryopreserved products (cryo-HPC) due to concerns of loss of stem cell viability and number with the cryopreservation procedure. ObjectiveWe aimed to analyze the outcomes of patients who received cryo-HPCs during the COVID-19 pandemic and compare this against historical cohorts that received fresh HPC. Study DesignA retrospective chart review was conducted on all adult patients who received a peripheral blood alloHSCT in British Columbia, Canada between June 2017 and November 2021. Baseline characteristics, Kaplan-Meier (KM) overall survival (OS), engraftment, and incidences of acute and chronic graft versus host disease were compared between patients who received cryo-HPCs and fresh HPCs. Univariable analysis followed by multivariable analysis was performed using a backward stepwise selection procedure to generate predictors of OS, cumulative incidence of relapse (CIR), nonrelapse mortality (NRM), and primary and secondary graft failure. ResultsThree hundred eighty-three patients were included in the analysis, with cryo-HPC representing 40%. Median viability was higher in the fresh-HPC group at 99.2% (IQR 98.3–99.5) versus cryo-HPCs at 97.0% (96.0, 98.6) (P < 0.01). The 12-month actuarial survivals were 77% in the fresh HPC and 75% in the cryo-HPC groups (P = 0.21). There were no differences between cryo-HPCs and fresh HPCs on univariable analysis of OS, CIR, or NRM. There was a shorter median time to platelet engraftment in patients receiving fresh HPC at 17 days (IQR 16, 20) versus cryo-HPC at 21 days (IQR 18, 29), P < 0.001. There was a shorter median time to neutrophil engraftment in the fresh HPC group at 17 days (IQR 14, 20) versus 20 days (17, 23), P < 0.001. Cryo-HPC accounted for 5 out of 6 cases of primary graft failure (P = 0.04), and 3 out of five cases of secondary graft failure (P = 0.39). There were no significant differences in acute GVHD between the fresh HPC and cryo-HPC groups (P = 0.34). The incidence of moderate or severe chronic GVHD was 32% in the fresh-HPC group and 17% in the cryo-HPC group (P < 0.001).In multivariable analysis, cryopreservation did not emerge as an independent predictor of OS, CIR, NRM, primary GF or secondary GF. However, viability <90% on arrival at our center was a significant predictor of OS (HR 5.3, 2.3–12.3, P < 0.01), primary graft failure (OR 36.3, 5.4–210.2, P < 0.01), and secondary graft failure (OR 18.4, 1.7–121.1, P < 0.01). ConclusionsPatients who received cryo-HPCs had similar OS and relapse rates to those who received fresh-HPCs but typically took 2–3 days longer to achieve engraftment of platelets or neutrophils and were associated increased primary graft failure. However, after accounting for multiple variables, cryopreservation was no longer a significant predictor of survival or engraftment while viability <90% emerged as an important predictor of OS, primary graft failure, and secondary graft failure. If confirmed, this suggests that viability on arrival at the infusion center may be a good quality control indicator used to identify HPC products that may warrant recollection if the risk of graft failure is sufficiently increased.

Long term results of a prospective multicenter observational study on the use of anti-human T-lymphocyte immunoglobulin (ATLG) in unrelated donor transplantation (ATOS study)

ATOS is a prospective observational study evaluating the outcome of patients receiving anti-human T-lymphocyte immunoglobulin (ATLG) in unrelated donor transplantation. Primary endpoint was severe GvHD and relapse-free survival (SGRFS). GvHD prophylaxis consisted of ATLG and CSA/ MTX or MMF. Outcome was compared to the ATLG arm of our prospective randomized phase III multicenter trial trial (RCT) [1, 2]. 165 patients, median age 54 (18; 77) years, with haematological malignancies with early (45.5%), intermediate (17.6%), and advanced (37.0%) disease were included. ATLG dose differed between centers according to local practise (median total ATLG dose of 46 (IQR 32–60, range 15–91) mg/kg). Median follow-up was 70 months. Estimated probabilities at 5 years follow up were for SGRFS 0.27, OS 0.52, DFS 0.43, NRM 0.23, relapse 0.34, acute GvhD °III/IV 0.13, severe chronic GvHD 0.27. OS rates differed dependent on disease status. An effect of the given ATLG dose could not be separated from potential center effects. Despite higher age and more advanced disease in ATOS, outcome was similar to the ATLG arm of our RCT. This long-term, multicenter, experience in routine clinical practice confirms the GvHD-protective effect of ATLG without compromising relapse and non-relapse mortality rates.Clinical Trial Registry: German clinical trials register DRKS00004581.

Efficacy of allogeneic hematopoietic stem cell transplantation based on a total body irradiation conditioning treatment regimen for adult acute lymphocytic leukemia

Objective: To analyze the efficacy of allo-HSCT with total body irradiation (TBI) and chemotherapy alone in the treatment of adult ALL and to explore the factors affecting prognosis. Methods: The clinical data of 95 adult patients with ALL who underwent allo-HSCT from January 2015 to August 2022 were included. According to the conditioning regimen, the patients were divided into two groups: the TBI plus cyclophosphamide (TBI/Cy) group (n=53) and the busulfan plus cyclophosphamide (Bu/Cy) group (n=42). Hematopoietic reconstitution after transplantation, GVHD, transplantation-related complications, relapse rate (RR), non-relapse mortality (NRM), OS, and LFS were compared, and the factors related to prognosis were analyzed. Results: The median time of neutrophil engraftment was 14 (10-25) days in the TBI/Cy group and 14 (10-24) days in the Bu/Cy group (P=0.106). The median time of megakaryocyte engraftment was 17 (10-42) days in the TBI/Cy group and 19 (11-42) days in the Bu/Cy group (P=0.488). The incidence of grade Ⅱ-Ⅳ acute GVHD (aGVHD) in the TBI/Cy and Bu/Cy groups was 41.5% and 35.7%, respectively (P=0.565). The incidence of grade Ⅲ-Ⅳ aGVHD in these two groups was 24.5% and 4.8%, respectively (P=0.009). The incidence of severe chronic GVHD in the two groups was 16.7% and 13.5%, respectively (P=0.689). The incidence of cytomegalovirus infection, Epstein-Barr virus infection, severe infection, and hemorrhagic cystitis in the two groups was 41.5% and 35.7% (P=0.565), 34.0% and 35.7% (P=0.859), 43.4% and 33.3% (P=0.318), and 20.8% and 50.0% (P=0.003), respectively. The median follow-up time was 37.1 months and 53.3 months in the TBI/Cy and Bu/Cy groups, respectively. The 2-year cumulative RR was 17.0% in the TBI/Cy group and 42.9% in the Bu/Cy group (P=0.017). The 2-year cumulative NRM was 24.5% and 7.1%, respectively (P=0.120). The 2-year LFS was 58.5% and 50.0%, respectively (P=0.466). The 2-year OS rate was 69.8% and 64.3%, respectively (P=0.697). In the multivariate analysis, the conditioning regimen containing TBI was a protective factor for relapse after transplantation (HR=0.304, 95% CI 0.135-0.688, P=0.004), whereas the effect on NRM was not significant (HR=1.393, 95% CI 0.355-5.462, P=0.634). Infection was an independent risk factor for OS after allo-HSCT in adult patients with ALL. Conclusion: allo-HSCT based on TBI conditioning regimen had lower relapse rate and lower incidence of hemorrhagic cystitis for adult ALL, compared with chemotherapy regimen. While the incidence o grade Ⅲ/Ⅳ aGVHD was hgher in TBI conditioning regimen than that in chemotherapy regimen.

Haploidentical donor peripheral blood stem cell transplantation using third-party cord blood compared with matched unrelated donor transplantation for patients with hematologic malignancies

Objectives: To assess the efficacy of cord blood-assisted haploid peripheral blood stem cell transplantation (haplo-cord-PBSCT) versus unrelated donor peripheral blood stem cell transplantation (UD-PBSCT) in the treatment of malignant hematological diseases. Methods: A retrospective analysis was performed on one hundred and four patients with malignant hematological diseases who underwent haplo-cord-PBSCT and fifty-two patients who underwent UD-PBSCT at Xiangya Hospital of Central South University between January 2016 and December 2021. Results: ①The median implantation time for neutrophils in the haplo-cord-PBSCT and UD-PBSCT groups was 13 (9-22) days and 13 (10-24) days, respectively (P=0.834), whereas the median implantation time for platelets was 15 (7-103) days and 14 (8-38) days, respectively (P=0.816). The cumulative implantation rate of neutrophils at 30 days after transplantation in the haplo-cord-PBSCT group and the UD-PBSCT group was 100% (P=0.314), and the cumulative platelet implantation rate at 100 days after transplantation was 95.2% (95% CI 88.3% - 98.1% ) and 100% (P=0.927), respectively. 30 days after transplantation, both groups of patients achieved complete donor chimerism, and no umbilical cord blood stem cells were implanted. ②The cumulative incidence rates of grade Ⅱ-Ⅳ acute GVHD within 100 days after transplantation in the haplo-cord-PBSCT group and the UD-PBSCT group were 29.1% (95% CI 20.1% -38.1% ) and 28.8% (95% CI 17.2% -41.6% (P=0.965), respectively. The cumulative incidence rates of grade Ⅲ/Ⅳ acute GVHD were 7.8% (95% CI 3.6% -14.0% ) and 9.6% (95% CI 3.5% -19.5% ) (P=0.725). The cumulative incidence rates of 2-year chronic GVHD in the haplo-cord-PBSCT group and the UD-PBSCT group were 45.3% (95% CI 36.1% -56.1% ) and 35.1% (95% CI 21.6% -44.1% ), respectively (P=0.237). The cumulative incidence rates of severe chronic GVHD at 2 years after transplantation were 13.6% (95% CI 7.6% -21.3% ) and 12.9% (95% CI 5.1% -24.3% ), respectively (P=0.840). ③The 2-year CIR after transplantation in the haplo-cord-PBSCT group and UD-PBSCT group were 12.8% (95% CI 7.0% -20.5% ) and 10.0% (95% CI 3.6% -20.2% ), respectively (P=0.341), and the NRM were 14.7% (95% CI 8.4% -22.6% ) and 16.2% (95% CI 7.4% -28.0% ), respectively (P=0.681). ④The 2-year OS rates in the haplo-cord-PBSCT and UD-PBSCT groups after transplantation were 82.2% (95% CI 74.8% -90.3% ) and 75.5% (95% CI 64.2% -88.7% ), respectively (P=0.276). The 2-year DFS rates were 69.9% (95% CI 61.2% -79.8% ) and 73.8% (95% CI 62.4% -87.3% ), respectively (P=0.551). The 2-year rates of GVHD-free/recurrence-free survival (GRFS) were 55.3% (95% CI 44.8% -64.8% ) and 64.7% (95% CI 52.8% -79.3% ), respectively (P=0.284) . Conclusion: The findings of this study indicate that haplo-cord-PBSCT and UD-PBSCT have comparable efficacy and safety in the treatment of malignant hematological diseases and can be used as an alternative treatment options.

Severity and organ distribution of chronic graft-versus-host disease with posttransplant cyclophosphamide-based versus methotrexate/calcineurin inhibitor-based allogeneic hematopoietic cell transplantation.

The reduced risk of chronic graft-versus-host-disease (GVHD) with posttransplant cyclophosphamide (ptCy) in the setting of haploidentical related donor and more recently, with HLA-matched related and matched and mismatched unrelated donor allogeneic transplantation has been established. There is, however, paucity of data to show if ptCy impacts chronic GVHD pathogenesis, its phenotype and evolution after HCT regardless of the donor status. We examined the differences in chronic GVHD incidence and presentation in 314 consecutive patients after receiving their first allogeneic transplantation (HCT) using ptCy-based GVHD prophylaxis (ptCy-HCT; n = 120; including 95 with haploidentical related donor) versus conventional calcineurin inhibitor-based prophylaxis (CNI-MUD; n = 194) between 2012 and 2019. The 1-year cumulative incidence of all-grade chronic GVHD and moderate/severe chronic GVHD was 24% and 12%, respectively, after ptCy-HCT and 40% and 23% in the CNI-MUD recipients (p = 0.0003 and 0.007). Multivariable analysis confirmed that use of CNI-based GVHD prophylaxis and peripheral blood stem cell graft as the risk factors for chronic GVHD. The cumulative incidence of visceral (involving ≥1 of the following organs: liver, lungs, gastrointestinal tract, serous membranes) chronic GVHD was significantly higher with CNI-MUD vs. ptCy-HCT (27% vs. 15% at 1 year, p = 0.009). The incidence of moderate/severe visceral chronic GVHD was 20% in CNI-MUD group vs. 7.7% in the ptCy-HCT group at 1 year (p = 0.002). In addition, significantly fewer ptCy-HCT recipients developed severe chronic GVHD in ≥3 organs (0.8%) vs. 8.8% in the CNI-MUD group at 1-year posttransplant (p = 0.004). There was no significant different in relapse, non-relapse mortality, and relapse-free and overall survival between the two groups. Further investigation is needed to confirm that reduced risk and severity of chronic GVHD, less visceral organ distribution with ptCy-HCT leads to improved quality of life.

Checkpoint Inhibitors and Allo-SCT in Hodgkin Lymphoma: A Matter of Time - a Study By the SFGM-TC

Introduction: Allogeneic stem cell transplantation (allo-SCT) for patients with relapsed/refractory Hodgkin lymphoma (r/R HL) after autologous transplantation is now an established option. Many patients receive Check-Point Inhibitors (CPI) as relapse treatment. Nevertheless, concerns have been previously reported on higher risk of acute graft versus host disease (aGVHD) after CPI exposure, without any control group to elucidate risk factors. Methods: We conducted a multicentric retrospective case control study on behalf of the Francophone Society for Bone Marrow Transplantation and Cell Therapy to compare outcomes after allo-SCT for patients with r/R HL who were exposed or not exposed to CPI. Results: Between 2015 and 2018, 149 patients underwent allo-SCT for r/R HL in 21 centers. CPI was used for 50 patients (n=48 Nivolumab, n=2 Pembrolizumab). We did not identify any differences regarding baseline characteristics. Reduced intensity conditioning (RIC) was commonly used (90.2%), with Peripheral Blood Stem Cells (75.7%). A majority of patients had haploidentical (44.6%), followed by siblings (28.4%), matched unrelated (24.3%) and mismatched unrelated donors (2.7%). Cumulative incidence (CI) of aGVHD grade ≥II in the whole cohort was 39.6% and 14.8% developed a severe form (grade III-IV). There were no differences between CPI (n=50) and no CPI cohorts (n=99), regarding CI of aGVHD of any grade (respectively 58% and 57.6%, p=1.00), grade ≥II (respectively 36% and 41.4%, p=0.65), severe aGVHD (grade III-IV, respectively 12% and 16.2%, p=0.67) and chronic GVHD (respectively 26% and 34.3%, p=0.24), as described in Table 1. We did not observe excess of mortality from GVHD between both groups. The time interval between the last CPI injection and allo-SCT was found to be the only significant factor associated with a higher incidence and severity of aGVHD, as depicted in Figure 1. The median time from the last CPI injection and allo-SCT was 50 days [33;103]. Specifically, within 30 days, the incidence of severe aGVHD was significantly increased at 41.7% compared to only 2.7% in the control group (p=0.002). No severe forms were observed after 60 days, but it persisted a higher risk of aGVHD grade ≥II even after 60 days of delay. Thus, a higher percentage of patients required systemic therapy for aGVHD within 120 days compared to after 120 days (45% vs 0% respectively, p=0.03). The delay of CPI treatment did not impact the CI and severity of chronic GVHD or CI of relapse after allo-SCT. Of note, post-transplant cyclophosphamide did not reduce CI of aGVHD. With a median follow-up of 34.7 months, the 2-year Overall Survival, Relapse Free Survival and GVHD free, Relapse Free Survival (GRFS) were respectively 69.9% (IC95 41.1-78.5), 67.1% (IC95 58.8-76.7) and 48.5% (IC95 40.3-58.4), without differences between CPI and no CPI cohorts. Two-year non relapse mortality was 17.4% in whole cohort, without hepatic sinusoidal obstruction syndrome nor steroid-requiring febrile syndrome in CPI cohort. Conclusion: CPI use before allo-SCT within 60 days before allo-SCT can lead to significant severe aGVHD, with persisting higher risk of aGVHD grade ≥II even after 60 days of delay. This highlights the importance of exercising caution when administering CPI injections within 60 days before allo-SCT. There is need to develop GVHD prophylaxis strategies in this population at risk. The findings from this study may help inform clinical decision-making and management of HL patients undergoing allo-SCT.

Acute Leukemia and MDS Patients Older Than 70 Years Have Inferior Survival Following Allogeneic Stem Cell Transplant Compared to Those with 60-69 Years of Age

Introduction: Increasing number of older patients are undergoing allogeneic stem cell transplants (alloSCT) for acute leukemia and MDS. We and others have previously reported alloSCT outcomes of older patients. However, majority of these studies included very few patients older than 70 years. In addition, the information on the influence of donor source on alloSCT outcomes among older patients is limited. In this study, we aim to answer two questions: Is there any difference in long-term survival between patients 60-69 years and those older than 70 years? Does donor source affect alloSCT outcomes of these two groups? Methods: We retrospectively evaluated alloSCT outcomes of 467 patients older than 60 years with acute leukemia (AML and ALL) and MDS. We compared outcomes between two groups: 60-69 years of age and those older than 70 years. The primary objective was to evaluate long-term overall survival (OS). Multivariable analyses included age, comorbidity index, KPS, disease risk index, donor relation, CMV serotype, conditioning regimen, GVD prophylaxis, and thymoglobulin use as variables. Results: Between January 2010 and December 2022, 335 patients with 60-69 years and 132 older than 70 years underwent alloSCT. Median age of the population was 66 years (range, 60-80); 6% (N=30) were African American and 22% (N=101) were of ethnic minority; 53% (N=249) had AML, 39% (N=184) had MDS, and 7% (N=34) had ALL; donor was matched unrelated in 60% (N=279), matched related and haploidentical in 14% each, and mismatched unrelated in 12% (N=55). Source of allograft was peripheral blood stem cells in all patients. Higher proportion of patients older than 70 years received matched unrelated donor compared to those with 60-69 years (69% vs 56%; p=0.004). Reduced intensity conditioning (RIC) regimen was frequently used among patients older than 70 years compared to 60-69 years (97% vs 79%, p&amp;lt;0.001). Neutrophil and platelet engraftment time was similar in both groups. Graft failure rate was higher for patients older than 70 years compared to 60-69 years (5.3% vs. 1.2%; p=0.014). The cumulative incidence of grade 3-4 acute GVHD was similar (13.5% vs 7.6%, p=0.2), while severe chronic GVHD was higher for the 60-69 group compared to the 70+ group (20.1% vs 10%, p=0.009). Median follow up for OS was 4.46 years. At 3 years, OS was 48% for the 60-69 group and 38.23% for the 70+ group (HR 1.34, p=0.02); RFS was 44% for the 60-69 group and 36.3% for the 70+ group (HR 1.32, p=0.02); relapse rate was 26.2% for the 60-69 group and 27.2% for the 70+ group (HR 1.13, p=0.51); and NRM was 30% for the 60-69 group and 37% for the 70+ group (sHR 1.26, p=0.19). In the multivariable analyses, patients older than 70 years experienced poor survival (HR 1.41, p=0.01) and relapse-free survival (RFS) (HR 1.4, p=0.01), and numerically higher non-relapse mortality (NRM) (sHR 1.38, p=0.08) compared to those with 60-69 years. Compared to haploidentical donors, matched related (sHR 0.39, p=0.04) and unrelated donors (sHR 0.28, p=0.005) were associated with lower NRM, while a trend for increased relapse rate and NRM was associated with the use of thymoglobulin and post-transplant cyclophosphamide (PTCy) containing GVHD prophylaxis, respectively. Conclusion: Our results indicate that although rates of acute GVHD and relapse were similar between both groups, patients older than 70 had worse OS and RFS compared to patients between 60-69. The NRM was numerically higher in patients older than 70 years of age. Haploidentical donor was associated with increased NRM, most likely secondary to toxicity associated with PTCy based GVHD prophylaxis. Detailed analysis of toxicity will be presented at the conference.

Patients with High-Risk Myelodysplastic Neoplasms Do Not Benefit from Remission Induction with Azacitidine Prior to Hematopoietic Cell Transplantation: A Retrospective, Single-Center Analysis with Extended Follow-up

Background: The value of remission induction in high-risk myelodysplastic neoplasms (MDS) with increased blasts prior to allogeneic hematopoietic cell transplantation (HCT) remains controversial. The new ELN recommendations include the re-designation of patients with 10-20% blasts as AML/MDS, raising the question of whether these patients should receive induction chemotherapy before HCT. An alternative is treatment with hypomethylating agents (HMA) such as azacitidine, for which allo-sensitizing effects have been proposed. In contrast, bridging to HCT with HMA may increase drop-out rate for allogeneic HCT, while at the same time not further benefiting patients who subsequently receive HCT. Methods: We report on 135 patients diagnosed with MDS, chronic myelomonocytic leukemia or myelodysplastic/myeloproliferative neoplasm who received HCT at our center between 2010 and 2022. Median age was 60 years (range 26 to 77 years) and 44.4% of patients were female. IPSS-R score was high in 25.8% of patients and very high in 44.8% of patients. 33.3% of patients received remission induction prior to HCT, 28.8% of all patients with HMA. Conditioning regimens were myeloablative in 8.1%, FLAMSA-based reduced intensity conditioning (RIC) in 26.7%, and RIC in 60.7% of cases. Donors were matched (8/8) related in 23.7%, matched (8/8) unrelated in 53.3%, and mismatched (7/8 or haploidentical) in 23%. Relationships between outcomes and patient, treatment and disease parameters were analyzed using multivariate Cox proportional hazards regression and, in the case of relapse, non-relapse mortality (NRM) and graft-versus-host disease (GVHD), competing risk analysis. P-values were adjusted for multiple comparisons using false discovery rate adjustment. Results: Median follow-up after HCT was 46 months. Kaplan-Meier estimates for OS and RFS at 5 years were 56.6% and 43%, respectively. The cumulative incidences of relapse, NRM, acute GVHD grade II to IV, and chronic GVHD at 5 years were 39%, 17.8%, 36.4%, and 40%, respectively. OS was significantly correlated with the IPSS-R cytogenetic risk category but not with the percentage of blasts in bone marrow before treatment initiation. In multivariate regression analysis, OS was significantly worse in patients with poor or very poor IPSS-R cytogenetic category (HR 1.56; p&amp;lt;0.001) but not in patients with a pre-treatment bone marrow blast count exceeding 10% (HR 1.25; p=0.19). Similarly, cytogenetic risk, but not bone marrow blast count, was significantly associated with worse RFS (HR 1.5 vs. 1.2; p&amp;lt;0.001 vs. 0.199) and higher cumulative hazard of relapse (HR 1.74 vs. 1.36; &amp;lt;0.0001 vs. p=0.31). Bridging or induction treatment with azacitidine before HCT by trend showed a higher cumulative hazard of relapse (HR 2.3; p=0.057) and worse RFS (HR 1.7; p=0.088). In exploratory subgroup analyses of patients with a bone marrow blast count above 10% (n=69), OS was not significantly different between patients who received remission induction prior to HCT and those who did not (HR 1.3; p=0.5), while the cumulative hazard of relapse by trend even increased in the first group (HR 1.85; p=0.13). Furthermore, there was no significant difference in OS between patients with a bone marrow blast count above 10% who received intensified sequential conditioning with FLAMSA-RIC versus fludarabine/treosulfan (n=44; HR 1.45; p=0.52). Donor mismatch was the only parameter significantly associated with higher NRM (HR 2.4 and p=0.044), and GVHD prophylaxis with anti-thymocyte globulin and/or tacrolimus the only parameters associated with a significant reduction of chronic GVHD severity (f²=0.05 and 0.06; p=0.03 and 0.02), while no single parameter was associated with a significant difference in acute GVHD severity. Conclusions: This study reports promising long-term outcomes for MDS patients undergoing HCT in the real-world setting. In our analysis, cytogenetic risk, but not bone marrow blast count, was the main determinant of transplantation outcomes. Furthermore, remission induction prior to HCT with azacitidine, which is commonly used in patients with increased bone marrow blasts, was not associated with a survival benefit but by trend even with higher relapse risk and worse RFS in multivariate analysis. This suggests that the perceived role of azacitidine as a bridge-to-transplant should be questioned. Rather, eligible patients should receive HCT as soon as possible.

High Disease-Free Survival in Patients with High-Risk MDS Treated with Orca-T

BACKGROUND Allogeneic hematopoietic cell transplant (alloHCT) may be a curative treatment option for patients with myelodysplastic syndrome (MDS). The use of investigational therapies that potentially reduce transplant toxicity could improve outcomes in this patient population with difficult to treat disease. Per CIBMTR, survival after alloHCT for MDS using matched donors is approximately 65% at 1 year and relapse-free survival at 1 year was ~69% in MDS patients in the RICMAC trial who received myeloablative conditioning (MAC). Orca-T is a high-precision cell therapy biologic that includes stem and immune cells, derived from allogeneic donors, that leverages highly purified, polyclonal donor regulatory T cells to control alloreactive immune responses. In this phase 1b study, Orca-T was evaluated in MDS patients who were eligible for transplant per 2017 International Expert Panel recommendations. METHODS As of 03/31/23, 16 patients with intermediate to very high risk (HR) MDS with ≤10% blasts in the bone marrow at the time of transplant had received Orca-T in the phase 1b study. IPSS-M prognostic scores were Low (6%), Moderate Low (6%), Moderate High (19%), High (44%), and Very High (25%). Baseline HCT-CI scores were 0 (n=6), 1 (n=3), 2 (n=2), 3 (n=3), and 4 (n=2). The median f/u was 19.1 months (range 6.3 - 32.2). Patients were aged 43 - 65 (median 59) and 56% female. Donors were 8/8 HLA-matched related (n=7) or unrelated (n=9). Patients received a busulfan-based myeloablative chemotherapy regimen prior to Orca-T, followed by single-agent GvHD prophylaxis with tacrolimus. RESULTS Orca-T was successfully manufactured in a single, centralized GMP manufacturing facility for all patients enrolled, distributed, and infused throughout the U.S. Orca-T was delivered within &amp;lt; 72 hours vein-to-vein (i.e. time between end of donor apheresis to start of recipient's Orca-T infusion). Disease-free survival (RFS) at 12 months was 94% (Figure). At 1 year, no patients had experienced grade ≥ 3 acute GvHD and moderate to severe chronic GvHD had occurred in 2 patients. Modified GRFS (survival free of Grade 3-4 aGvHD, moderate-to-severe cGvHD and relapse) at 12 months was 75%. Non-relapse mortality (NRM) and overall survival (OS) were 0% and 94%, respectively, at 12 months. Additionally, one MDS patient with very poor prognosis, characterized by mutated TP53 and complex karyotype, remains in CR without active signs of acute or cGvHD at 500 days post-alloHCT. CONCLUSIONS Orca-T demonstrated promising results in this HR-MDS patient population, including high RFS, low incidence of GvHD, and very high OS at 1 year. A randomized controlled phase 3 multi-center trial comparing Orca-T to SOC in patients with MDS, AML, and ALL is currently enrolling across the US (NCT05316701).

MSCs-IL33-ILC2s-AREG-Tecs Axis Promotes the Remodeling of Thymic Function in Acute Gvhd

Background: Previous studies suggested that mesenchymal stromal cells (MSCs) administrated at the acute graft versus host disease (aGVHD) phase effectively decreased the incidence and severity of chronic GVHD (cGVHD) by boosting thymic regeneration. Our research has found that MSCs expressing interleukin 33 (IL33) may participate in thymus regeneration from aGVHD damage through type 2 innate lymphoid cells (ILC2s)-mediated tissue recovery. However, the mechanism for ILC2s repairing the damaged thymus caused by aGVHD is unclear. Here, we further explored the mechanisms on how ILC2s promote thymus recovery post-BMT. Methods: We carried out studies in a aGVHD mice model of fully MHC-mismatched myeloablative bone marrow transplantation (C57BL/6 to BALB/c). IL33 knocked-down (MSC/IL33 -) and empty-load MSCs (MSC/Control) were generated by lentivirus transfection, and then administrated intravenously at day 7 post-BMT to intervene in aGVHD and assess the effect of IL33 from MSCs on ILC2s (Lin-CD45+iCD3-CD127+GATA3+) in the thymus by flow cytometery at day 14 post-BMT. Clinical scores and survival were recorded once every days. Furthermore, we isolated, purified and amplified ILC2s from C57BL/6 mouse thymus by immunomagnetic bead sorting and flow cytometry for aGVHD intervention and studied whether ILC2s can home to thymus by fluorescence tracing and the effects of ILC2s on TECs by immunofluorescence. To study the potential mechanism of ILC2s affecting TECs by using transcriptome sequencing (RNA-seq) and related molecular biology experiments. Finally, the potential target amphiregulin (AREG) was found from RNA-seq analyses and the murine recombinant AREG was used as an intervention to evaluate its effect on aGVHD mouse. Results: In this study, we found that knocking down the expression of IL33 in MSCs partially blocked the repair effect of MSCs on thymus and decreased the expansion of thymic ILC2s in aGVHD mice compared with the MSC/Control group. Then, we found that exogenous infusion of thymic ILC2s is very effective in treating thymic tissue injury, and can home to the recipient thymus and further migrate to the thymic medulla region. Furthermore, we found that ILC2s may home to the thymus and migrate to the thymic medulla region through CCR7 and MSCs promote the high expression of AREG in ILC2s through RNA-seq. Importantly, we found that the expression level of intrathymic AREG in No-GVHD, MSCs-GVHD and ILC2s-GVHD group was significantly increased than aGVHD group on day 14 post-BMT and exogenous administration of AREG after transplantation can significantly increase the proliferation of thymic epithelial cells, especially the regeneration of thymic medulla and the generation of new thymus-derived CD4/CD8 double-positive T cells. Conclusion: In conclusion, we demonstrate that MSCs infusion may rely on the MSCs-IL33-ILC2s-AREG-TECs axis mediated by ILC2s to remodeling the thymic microenvironment homeostasis, thus promoting the reconstruction of T cell immune homeostasis. Importantly, the results presented in this report identify a previously unrecognized role for the epidermal growth factor family member AREG in promoting the remodeling of thymus tissue. and providing new insights for the mechanism of MSCs treatment for GVHD.

Phase I / II Study on Infusion of Alloreactive or Ex Vivo il-15 Stimulated Natural Killer Cells after Haploidentical Stem Cell Transplantation in Pediatric Patients with Acute Leukemia (PHINK): A Study of the Spanish Hematopoietic Stem Cell Transplantation Group (GETH)

Background: Haploidentical hematopoietic stem cell transplantation (haplo-HSCT) is a highly complicated but effective procedure for high-risk pediatric leukemia in the absence of an HLA-identical donor. Post-transplant relapse is the main problem for patient survival. According to literature, Natural Killer (NK) cells play a potent role as inducers of the graft-versus-leukemia (GvL) effect in haplo-HSCT. The presence of alloreactive NK cells correlates with a lower relapse rate, improved engraftment, decreasing graft-versus-recipient disease (GvHD) and viral infection risk. In the absence of donor-recipient NK cell alloreactivity, other NK cell activation strategies are necessary. We propose a phase I/II clinical trial with dose escalation, multicenter, framed in the GETH, to determine dose-limiting toxicity (DLT) and the maximum tolerated dose (MTD) of a single infusion of alloreactive NK cells (Allo-NK) or NK cells stimulated ex vivo with IL-15 (IL15-NK) after haplo-HSCT in pediatric patients with high-risk leukemias. Methods: Eighteen pediatric patients with high-risk leukemia were recruited between September 16, 2020, and May 12, 2023, at 5 hospitals in Spain, Hospital Universitario La Paz, Hospital Universitario Gregorio Marañón, Hospital Universitario Niño Jesús, Hospital Universitario Central of Asturias and Hospital Santa Creu and Sant Pau. Patients were stratified in 2 treatment groups: Allo-NK (n=9) or IL15-NK (n=9), depending on the KIR disparity between donor and recipient. Our phase I/II clinical trial presents a dose-escalation design (3+3). The treatment cohorts are divided into cohort 1 (up to 2x10 7 cells/kg), cohort 2 (2-5x10 7 cells/kg) and cohort 3 (5-10x10 7 cells/kg). We have monitored infections reactivation, as well as, chimerism and engraftment after transplantation. In Allo-NK group, the KIR clone that provides alloreactivity in the NK cell was also tracked. The phenotypic profile and functional capacity of NK cells present in the patient up to 360 days after transplantation have also been analyzed. Results: Nine patients (4 in Allo-NK and 5 in IL-15NK) were included in cohort 1, 7 patients (4 in Allo-NK and 3 in IL-15NK) in cohort 2 and 2 patients (1 in Allo-NK and 1 in IL-15NK) in cohort 3. We were unable to complete the dose escalation design initially proposed due to the high weight of some patients and the unavailability of ex vivo expanded NK cells. In IL15-NK group patients presented a higher prevalence of B-ALL (p=0.018) and had received a previous transplant (p=0.082). NK cell infusions of IL15-NK group showed higher expression of CD25 and CD69 receptors and higher functional capacity. In the Allo-NK group, the most prevalent KIR-reactive clone was KIR2DL1 (55.6%). We also observed a tendency for KIR2DL1 alloreactive clone to increase from 30 (8,18%) to 360 (25,63%) days after transplantation. Regarding clinical outcomes in 2 study arms, the incidence of severe (Grades III-IV) acute GvHD was 22.2% and severe chronic GvHD was only observed in 1 (5.6%) patient (Table 1). The incidence any infection or endothelial vascular complications were 72% and 33% respectively. No statistical differences were observed in GvHD, infections and endothelial vascular complications. We found 2 relapses, 1 in each study arm (11.1%). Five patients died, 1 (5.6%) because of disease progression and 4 (22.2%) because transplant related complications (TRM) (Table 1). Dividing patients into the 3 dose-escalation cohorts, we found no significant differences between cohorts in the primary clinical outcomes analysed. One-years overall survival (OS) and disease-free survival (DFS) was 72.22% (CI95% 51.93-100) and GvHD-free/relapse-free survival (GRSF) was 64.1% (CI95% 44.62-92.1), finding no significant differences between either 2 study arms or the 3 dose-escalation cohorts. Regarding the analysis of the NK phenotype, we observed a tendency to increase the expression of CD16 and CD57 receptors between 30 to 360 days after transplantation in both groups. Conclusions: Two study arms shows similar data respect to clinical outcome. Patients infused with IL-15-stimulated NK cells showed similar toxicity to the alloreactive NK group. According to dose escalation, we observed no association between increased toxicity and increased number of infused NK cells. We were unfortunately unable to complete cohort 3 because of the cells/kg ratio in some patients.

Delayed Initiation of Calcineurin Inhibitor Is Critical for Tolerance Induction By Posttransplant Cyclophosphamide

[Introduction] We have previously shown that T-cell exhaustion of donor alloreactive T cells plays a critical role in tolerance induction after mouse allogeneic hematopoietic cell transplantation (allo-HCT) (Asakura S: JCI 2010). Emerging evidence indicates that T-cell exhaustion is a multistep process, including precursors of exhausted T cells (pTex) with stem cell-like properties, pTex-derived transitory exhausted T cells (transitory-Tex) with potent effector-like functions, and terminally exhausted T cells (terminal-Tex) with the expression of multiple inhibitory receptors and severely impaired functions. We have demonstrated, for the first time, that calcineurin inhibitors (CNIs) inhibit tolerance induction by suppressing the terminal exhaustion of donor T cells while inducing transitory-Tex after allo-HCT in mice (Senjo H: Blood 2023). In contrast, post-transplantation cyclophosphamide (PTCy) efficiently induces tolerance with low incidence of chronic GVHD. In the current study, we evaluated the mechanisms by which PTCy could induce tolerance even with the use of CNIs. [Methods] In mouse haploidentical HCT model,B6D2F1 recipients (H-2 b/d) were lethally irradiated and transplanted with 2 × 10 6 T cells together with T-cell-depleted bone marrow cells from C57BL/6 (H-2 b) donors on day 0. CSP (25 mg/kg) was administered from either day 0 (early-CSP) or day 5 (late-CSP), and PTCy (50 mg/kg) was i.p. injected on day 3 after HCT. In clinical study, peripheral blood mononuclear cells were prospectively collected from patients who underwent PTCy-based haplo-PBSCT at our institutions after approval of institutional review. [Results] We have shown that Ly6C is a specific marker for transitory-Tex in mice (Senjo H: Blood 2023). In mice, early-CSP followed by PTCy (CSP→PTCy) significantly expanded Ly6C +PD-1 +TOX low CD8 + T cells and Ly6C +PD-1 +GZMB + CD4 + T cells on day 14, while late-CSP following PTCy (PTCy→CSP) induced T-cell differentiation towards Ly6C -PD-1 +TOX high terminally exhausted T cells. When CSP→PTCy recipients were injected with anti-PD-L1 monoclonal antibodies, transitory-Tex demonstrated vigorous proliferation, while terminal-Tex did not respond to PD-1 blockade. These findings indicate that CSP administration after PTCy promotes tolerance induction, whereas CSP administered before PTCy inhibits tolerance induction. Next, we explored the specific markers for human transitory-Tex using scRNA-seq of T cells from the patients who underwent PTCy haplo-PBSCT, in which tacrolimus (TAC) was started either from day -1 (early-TAC) or day 5 (late-TAC). UMAP analysis showed that early-TAC expanded cluster 5 (C5) out of six CD8 + clusters (C1-C6) and C8 out of three CD4 + clusters (C7-C9) (Fig.1). C5 highly expressed exhaustion markers such as Tox and Pdcd1, together with effector molecules such as Prf1, Gzmb and Cx3cr1, while C8 was characterized with high expression of Gzmb, indicating that C5 and C8 are the human counterparts of transitory-Tex. Analysis of differentially expressed genes showed that Fcgr3a (CD16A) was selectively expressed in C5, while Gzmb and Cd74 were selectively expressed in C8. These two populations showed robust proliferative responses to TCR stimulation in vitro. ScTCR-seq demonstrated that C5 and C8 clusters showed less TCR diversity compared to other clusters, suggesting that C5 and C8 were enriched with alloreactive donor T cells that clonally expanded post-transplant. FCM confirmed that both CX3CR1 +CD16 + CD8 + T cells and GZMB +CD74 +CD4 + T cells were significantly increased in the early-TAC group compared to the late-TAC group on day 28 (Fig. 2). In the early-TAC group, 2/9 patients developed moderate and severe chronic GVHD, and only 1/9 patient was off-immunosuppressants (Fig 2). In sharp contrasts, in the late-TAC group, no patient developed chronic GVHD, and 7/9 patients were off-immunosuppressants. Importantly, two patients who developed chronic GVHD had a particularly high proportions of CX3CR1 +CD16 +CD8 + and GZMB +CD74 +CD4 + transitory-Tex on day 28 after HCT (Fig.2). [Conclusion] Our study demonstrated that early initiation of CNIs before PTCy induced transitory-Tex and inhibited tolerance induction after PTCy haplo-PBSCT. Administration of CNIs after PTCy is critical for tolerance reduction without chronic GVHD.

Prednisolone Intake Differentially Regulates IL12p40 and IL18 - Impact on Severe Chronic Gvhd, Relapse and Mortality

Background: Chronic graft-versus-host disease (cGVHD) is an immune-mediated complication after allogeneic stem cell transplantation (alloSCT). It is associated with graft-versus-leukemia effects and lower relapse rates, but in its severe form also with increased morbidity and mortality. The early interferon-gamma (IFNγ)-response marker CXCL9 is increased in mild and severe cGVHD and strongly correlates with IDO activity (1). Interleukin (IL)-18 is an innate, IFNγ inducing cytokine that associates with poor outcome of allografted patients (2), and with severe fibrosing cGVHD (1). The adaptive immunological arm of IFNγ induction relies on IL12 that is produced during cognate interactions between dendritic cells and T cells. IL12 kinetics is poorly characterized after alloSCT. We hypothesized that adaptive (IL12p40) and innate (IL18) immune mediators and their modulation by immunosuppressive drugs (ATG, prednisolone) impact cGVHD severity and outcome. Methods: Patients before alloSCT were included in an observational study allowing collection of clinical data and blood before and for one year after alloSCT. The training cohort included 350 patients transplanted before 2018 at our institution. The validation cohort consisted of 205 patients transplanted after 2018. Patients were eligible if they were not relapsed and did not receive donor leukocyte infusions (DLI) until d180, and did not develop refractory acute GVHD. Cytokines that belonged to the IFNγ pathway (IFNγ, CXCL9, CXCL10, IL12p40, IL12EBI3, IL17, IL18, IL23) or to innate inflammatory pathways (TNFa, IFNa, IL1b, activin, follistatin) were measured by R&amp;D ELISAs. Cumulative (equivalent) prednisolone dosages between d0 and d100 after alloSCT were calculated from electronic patient records. Flow cytometry of PBMC of 100 patients on day+180/onset cGVHD were performed using 24-gate multicolor staining. Mild, severe fibrosing and severe gastro-intestinal (GI) cGVHD were diagnosed according to simplified NIH criteria (training cohort only). Overall survival, non-relapse mortality and time to relapse from time of blood sampling were calculated by multivariable Cox regression analyses. Results: Median time of blood sampling at onset of cGVHD (n=180) was day +168 (range 41-723). Control patients (n=169) were collected on day +180 (171-183). Mild cGVHD was recorded in 133 patients, severe fibrosing cGVHD in 39 patients, and isolated GI-cGVHD in 8 patients. Receiver-operator characteristic (ROC) curves showed that only CXCL9, IL18 and IL12p40 were significantly linked to severe cGVHD. IL18 was associated with severe fibrosing cGVHD, whereas CXCL9 and IL12p40 were correlated to any grade cGVHD. In multivariable Cox analyses including IL12p40, IL18, age, mismatch, recipient and donor gender, disease stage, ATG, MTX, and diagnosis, IL18 associated with poor OS (HR per log2 increase 1.81 (1.19-2.74, p=0.005), whereas IL12p40 associated with superior OS (HR 0.53 (0.35-0.81), p=0.003). This could be validated in the independent cohort. Time course analyses of serum levels of CXCL9, IL12p40 and IL18 revealed a strong increase of CXCL9 and IL12p40 toward onset of cGVHD, but no stimulation of IL18 levels. Cumulative prednisolone intake&amp;gt;100 mg until day+100 significantly reduced IL12p40 serum levels on d180/at onset of cGVHD, without influencing IL18 production (Figure 1). In contrast, ATG reduced both, IL18 and IL12p40 at day180/onset of cGVHD. This might explain the reduced incidence of severe fibrosing cGVHD in patients receiving ATG prophylaxis (days-3 to -1 before alloSCT, 4% vs. 23%, p&amp;lt;0.001, n=343), whereas no effect of early prednisolone intake was noticed (p=0.916, n=137). Multicolor flow cytometry showed that IL12p40, but not IL18 correlated with blood counts of T-lymphocyte populations, including CD4+, CD8+ and γδ-Tcells. IL12p40 and γδ-Tcells were associated with reduced relapse rates. In summary, CXCL9, IL12p40 and IL18 are associated with severe cGVHD. IL18 correlated with severe fibrosing cGVHD and increased mortality, whereas IL12p40 was associated with reduced NRM, reduced relapse risk, improved lymphocyte recovery and γδ-T-cells, and was affected by prednisolone intake but not ATG. Conclusion: These results may help to understand and potentially separate the GVL effects from mortality/morbidity that associate with cGVHD. 1)PMID34755129 2)PMID31680001

Impact of Allograft T-Cell (CD3) Dose on Outcomes When CD34 Cell Dose Is Capped: Analysis of 811 Consecutive Allogeneic PBSC Transplants from a Single Center

G-CSF mobilized PBSC allografts contain higher T-cell doses than bone marrow grafts and have been associated with a significantly higher risk of chronic GVHD. Higher CD34+ cell content in PBSC grafts has also been shown to predict higher rates of chronic GVHD and associated mortality 1,2 To mitigate this risk our center adopted a policy of capping CD34+ cell dose at 5 x 10e6/kg recipient weight for unmanipulated PBSC grafts. For this study, we assessed the effect of T-cell (CD3) dose administered under this capped CD34 strategy, on GVHD and other transplant outcomes. Consecutive first PBSC allografts for hematologic malignancy performed using the CD34 cap strategy between 2010 and 2022 (n=811) were included in this analysis. GVHD prophylaxis was tacrolimus/methotrexate for MRD and MUD and post-transplant cyclophosphamide based for Haplo. Patients receiving anti-T cell serotherapy or ex-vivo T-depletion were excluded. GVHD was uniformly assessed and graded by a dedicated practitioner for all cases. Patient characteristics were: median age 55 (18-80); 57% male; race -white 74%, black 21%, Asian 5%; diagnosis AML 38%, ALL 18%, MDS 18%, NHL 10%; donor type haplo 42%, MRD 32%, MUD 26%, %; high/v. high 25%, low/intermediate 73%; HCT-CI (0-2) 43%, (&amp;gt;3)57%, regimen - myeloablative 48%, RIC 31%, NMA 21%). Median follow-up for survivors was 64 (16-151) months. Median CD34 and CD3 doses infused were 5.0 x 10e6/kg (min 1.59) and 17.3 x 10e7/kg (range 1.2-83.3) respectively. Median CD3 dose infused were higher with female donors (20.5 vs 15.5 x 10e7/kg, p&amp;lt;0.001) and older donors (18.6 vs 15.9 x 10e7 for donors aged &amp;gt;38 vs &amp;lt;38, p&amp;lt;0.001). The effect of Infused CD3 dose was assessed in terciles (ranges 1.2-13.5, 13.6-22, 22.1-83.3 x 10e7/kg). Median peripheral blood donor CD3 chimerism was 100% for all CD3 dose terciles at d 30, through 180 post-transplant. On univariate analysis - estimated 3-year survival, DFS, NRM, relapse and acute GVHD incidence were not significantly different between the low, middle and high CD3 terciles. In contrast, the cumulative incidence (CI) of moderate-to-severe chronic GVHD at 3 years was significantly higher in the highest vs lowest tercile (36.8% vs 28.6%, p=0.042) (Fig 1), while severe chronic GVHD (14.5% vs 13%) was not significantly different. On stratified Cox multivariable analysis comparing the highest to the lowest CD3 dose terciles - survival, DFS, NRM, relapse and acute GVHD were not significantly different. Moderate-to-severe chronic GVHD was significantly higher in the highest versus lowest tercile (HR 1.53, p=0.005). In order to address specific effects of CD3 dose for particular donor groups, we analyzed incidence of chronic GVHD by CD3 dose terciles separately for the three donor types (MRD n=258, MUD n=210 and Haplo n=343). Whereas the CI of moderate-to-severe chronic GVHD was not significantly different for lowest versus highest CD3 tercile in MRD and Haplo transplant recipients, it was significantly higher for MUD recipients (50% vs 26%, p=0.008, GRFS 29% vs 9%, p=0.005). On multivariable analysis, risk of moderate-to-severe chronic GVHD was significantly higher for the highest versus lowest CD3 dose terciles (HR 2.62, p=0.002) and for the middle vs lowest terciles (HR1.77, p=0.043) in MUD patients (Fig 1). In contrast, CD3 dose had no significant impact on chronic GVHD for MRD and Haplo recipients. This large retrospective analysis of PBSC allografts using a CD34 cap (&amp;lt;5 x 10e6/kg) shows that this approach produced infused CD3 doses ranging between 1.2 and 83.3 x 10e6 /kg. The median and upper limit of infused CD3 dose was lower than previously reported by studies with no CD34 cap 2,3. CD3 dose had a significant impact on the rate of moderate-to-severe chronic GVHD in MUD recipients. A prospective comparison of capped versus uncapped CD34 dosing may help determine the value of this strategy in mitigating GVHD risk of PBSC grafts. These data suggest the need for more aggressive GVHD prophylaxis in patients infused with higher CD3 doses in MUD patients. 1. Mohty M, Bilger K, Jourdan E, et al. Leukemia. 2003;17(5):869-875. 2. Zaucha JM, Gooley T, Bensinger WI, et al. Blood. 2001;98(12):3221-3227. 3. Saad A, Lamb L, Wang T, et al. Biol Blood Marrow Transplant. 2019;25(9):1875-1883.

The Outcomes of Cord Blood Transplantations Infused with Higher Total Number of Nucleated Cells Were Equivalent to Unrelated Bone Marrow Transplantations

Introduction: Choosing a suitable donor is essential to achieve a good response for allogeneic hematopoietic cell transplantation (allo-HCT). Several previous studies have compared unrelated bone marrow transplantation (UR-BMT) and cord blood transplantation (CBT), but the results differed among each study, and their superiority has been controversial. The total number of nucleated cells (TNC) in cord blood unit is crucial for CBT to avoid early complications, including engraftment failure. The outcome of CBT may improve by choosing cord blood unit with a higher TNC. Methods: We retrospectively compared the outcomes of CBT with higher TNC (hCBT) and UR-BMT conducted in our institute from January 1st, 2013, to May 31st, 2023. Cut off point of TNC was the median value. The primary endpoint was overall survival (OS), and the secondary endpoints were non-relapse mortality (NRM), cumulative incidence of relapse, and cumulative incidence of complications. Results: We analyzed a total of 316 patients, including 139 patients who underwent CBT and 177 patients who received UR-BMT. The median infused TNC was 2.71 (range, 0.027-6.42) ×10^7/kg. hCBT group included 69 patients. Significantly more patients in the hCBT group had prior allo-HCTs (hCBT: 18.8%, UR-BMT: 5.6%, p=0.003), and hCBT procedures were more frequently conducted since 2018 (hCBT: 66.7%, UR-BMT: 36.7%, p&amp;lt;0.001). For each group, most patients received tacrolimus and short-term methotrexate as graft-versus-host disease prophylaxis (hCBT: 94.2%, UR-BMT: 95.5%, p=0.277). ABO matched donors were more commonly chosen for UR-BMT (hCBT: 24.6%, UR-BMT: 46.3%, p=0.006). Patients with myelodysplastic syndrome underwent UR-BMT more frequently (hCBT: 7.2%, UR-BMT: 20.9%). Patients with a high or very high disease risk index were more frequently included in the hCBT group (hCBT: 37.5%, UR-BMT: 27.9%, p=0.060). Patient sex, age, days from diagnosis to transplantation, and hematopoietic cell transplantation comorbidity index, conditioning intensity were well balanced between each group. The two-year OS of hCBT was equivalent to UR-BMT (hCBT: 53.6%, UR-BMT: 58.9%, p=0.56) (Figure 1). There was no significant difference in the two-year NRM between the two groups (hCBT: 24.9%, UR-BMT: 25.4%, p=0.87). The hCBT group had a lower neutrophil engraftment rate achieved at day 28 (hCBT: 83.0%, UR-BMT: 92.3%, p&amp;lt;0.001). The two-year relapse rate was similar between the hCBT and UR-BMT groups (hCBT: 26.1%, UR-BMT: 18.7%, p=0.34). The cumulative incidence of acute GVHD tended to be lower in the hCBT group (hCBT: 18.2%, UR-BMT: 33.6% at 100 days, p=0.067), and the same results were observed when analyzing grade 2-4 or grade 3-4 acute GVHD. The cumulative incidence of chronic GVHD was significantly lower in the hCBT group (hCBT: 4.7%, UR-BMT: 48.3%, p&amp;lt;0.001), and the cumulative incidence of severe chronic GVHD was also lower in hCBT group (hCBT: 0.0%, UR-BMT: 13.9%, p&amp;lt;0.001). To reduce the impact of patients' characteristics, propensity score matching was conducted. The covariates included prior allo-HCTs, disease risk index, and allo-HCT conducted before 2018 or not. After propensity score matching, a total of 116 patients were included in the analysis with 58 patients for each group. The two-year OS of hCBT was also equivalent to UR-BMT (hCBT: 61.8%, UR-BMT: 55.2%, p=0.43) (Figure 2). The two-year NRM was slightly lower in hCBT group, but it is not significant (hCBT: 19.1%, UR-BMT: 30.1%, p=0.21). The hCBT group had a late neutrophil engraftment (hCBT: 87.0%, UR-BMT: 89.1% at day 28, p&amp;lt;0.001). The two-year relapse rate was similar between the hCBT and UR-BMT groups (hCBT: 25.0%, UR-BMT: 20.0%, p=0.54). The cumulative incidence of acute GVHD was tended to be lower in hCBT group (hCBT: 19.8%, UR-BMT: 38.7% at 100 days, p=0.057) and the cumulative incidence of chronic GVHD was significantly lower in hCBT group (hCBT: 5.3%, UR-BMT: 56.3%, p&amp;lt;0.001). Discussion: This was the first study to validate that the outcomes of CBT with higher TNC were equivalent to UR-BMT. Although the hCBT group experienced later neutrophil engraftment, the rate of neutrophil engraftment achieved at day 28 was comparable to UR-BMT, and a lower incidence rate of chronic GVHD might be attributed to lower NRM. From these findings, hCBT may be a worth considering option for patients requiring allo-HCT.