The use of thymoglobulin after allogeneic hematopoietic stem cell transplantation (HCT) has been implicated in increased the risk of relapse and infections. Recent data have suggested the use of haplo-transplant as second choice alternative to MUD after first choice related donor (RD). We hypothesized that outcomes after MUD transplant with use of thymoglobulin similar to RD HCT would justify the use of MUD HCT with addition of thymoglobulin as best second donor choice after RD. We compared the outcomes of 98 (29 matched RD/ 69 matched MUD) consecutive patients with AML/MDS who underwent HCT with Fludarabine/ Busulfan conditioning between 7/2009-12/2014. Patients Characteristics are shown in Table-1. Patients received full intensity conditioning (FIC) with Fludarabine 40 mg/m2 for 4 days and Busulfan 3.2 mg/Kg single daily dose for 4 days or reduced intensity conditioning with Fludarabine and Busulfan 3.2 mg/Kg single daily dose for 2 days. GVHD prophylaxis for FIC was Tacrolimus/mini Methotrexate and Tacrolimus/Mycophenolate for RIC. Thymoglobulin was added at dose of 1.5 mg/Kg for 3 days for MUD patients only. With median follow of 772 days( RD 809 days, MUD 756 days), the overall survival at 1 year was 76% +/- 8% and 72% +/- 5% for RD and MUD respectively, P=0.89. All patient engrafted except two patients in FIC MUD group (Primary graft failure). The cumulative incidence of relapse at 1 year was 38% +/- 8% and 24% +/- 5% for RD & MUD respectively, while the treatment related mortality (TRM) at 2 years were 16% +/- 3% and 20% +/- 3% for RD and MUD HCT recipients. Cumulative Incidence acute GVHD grade II-IV was 34% in RD at day 100 and day 180 while it was 28% & 38% at day 100 and day 180 in MUD respectively. The cumulative incidence of mild to moderate chronic GVHD was observed in 49% of MUD recipients and 53% of RD recipients at 2 years. The addition of thymoglobulin to MUD HCT results in outcomes comparable to RD HCT in AML/MDS patients. Our data suggest the use of MUD donors as second choice for patient who lacks matched siblings while reserving other alternative donors for patients who lacks full match MUD donors.Tabled 1Patients CharacteristicAllo-RelatedUnrelated DonorPNumber ( AML/MDS)29 (22/7)69 (61/8)Mean age ( Range )53.2 ( 34-73)55.95 (26-73)0.32Status at Transplant0.81CR1 (Complete remission -1)17 (59%)44 (64%)CR2 (Complete remission -2)3 (10%)5 (7%)PIF, REL (primary refractory, Relapse)9 (31%)20 (29%)Conditioning Regimen Intensity0.41FIC (Full Intensity)/Flu-Bu422 (75%)44 (64%)RIC ( Reduced Intensity)Flu-Bu27 (25%)25 (36%)CD34 Infused X million/Kg (Range)Mean (million/Kg)5.86.830.075Median to WBC engraftment ( Days)12.5711.640.93Median to Plat engraftment (Days)19.27180.39 Open table in a new tab
2,3,7,8-Tetrachlorodibenzo-p-dioxin (TCDD) is known to cause a large number of adverse effects, mediated largely by its binding to the aryl-hydrocarbon receptor (AhR) and subsequent modulation of gene expression. It is thought that AhR mediates these effects through the untimely and disproportionate expression of specific genes. However, the exact mechanism, or the genes involved, through which TCDD leads to these effects is still unknown. This study reports the discovery of a novel target gene, epiregulin, which is regulated by TCDD-activated AhR. Epiregulin is a growth regulator which belongs to the epidermal growth factor (EGF) family. Using real time quantitative PCR (qPCR), it was established that TCDD upregulates epiregulin gene expression. The promoter region of epiregulin has a dioxin responsive element (DRE) 56 nucleotides upstream of the transcription start site, along with three potential Sp1 binding sites. Chromatin immunoprecipitation (ChIP) assays with an anti-AhR antibody showed promoter occupancy upon TCDD treatment. Luciferase reporter assays using a vector harboring the first 125 base pairs of the epiregulin rat promoter revealed an increase in signal on TCDD treatment, which was lost upon mutation of the DRE. Epiregulin and TCDD treatment mediated a dose-dependent increase in primary mouse keratinocyte growth. These results demonstrate that AhR directly increases epiregulin expression, which could play an important role in TCDD mediated tumor promotion observed in rodent models.
The aryl hydrocarbon receptor (AhR) is a ligand-activated transcription factor responsible for mediating the cellular response to the toxic compound 2,3,7,8,-tetrachlorodibenzo-p-dioxin. An essential role for the AhR in cellular biology has been established previously, but no high-affinity endogenous ligand has yet been identified. We have confirmed the presence of a putative endogenous ligand(s) in CV-1 cells through transient transfection with various cytochrome P450 isoforms. Expression of cytochromes P450 1A1, 1A2, or 1B1 reduced AhR-mediated luciferase reporter activity, whereas cytochrome P450 2E1 exhibited no significant effect. Studies with 2,4,3′,5′-tetramethoxystilbene, a potent and specific inhibitor of cytochrome P450 1B1, was able to partially block cytochrome P450 1B1-mediated reduction in reporter gene activity. These results provide evidence of the existence of a possible feedback mechanism in which AhR-regulated cytochromes P450 from the CYP1A and CYP1B families are able to metabolically alter putative endogenous ligand(s). Several experiments were performed to provide initial characterization of these putative endogenous ligands, including electrophoretic mobility shift assay analyses, which demonstrated that these ligands directly activate the AhR. Soluble extracts from various C57BL/6J and Ahr-null mouse tissues were also analyzed for the presence of AhR activators. Studies revealed that Ahr-null mouse lung tissue had a 4-fold increase in AhR-mediated reporter activity in cells. Quantitative polymerase chain reaction analysis revealed that lung tissue exhibits relatively high constitutive CYP1A1 mRNA levels. These results suggest that there is an autoregulatory feedback loop between the AhR and cytochrome P450 1A1 in mouse lung.
Haploidentical stem cell transplantation (HaploSCT) using post-transplant cyclophosphamide (PTCY) has been performed primarily with non-myeloablative conditioning. We are exploring a melphalan-based, myeloablative yet reduced-intensity conditioning (RIC), in an ongoing phase II clinical trial. Outcomes of the first 84 pts treated after 01/2009 are reported. 47(56%) pts were males. 74 (88%) pts had their first transplant, 10 (12%) as second transplant. The median age was 46 years (range 19-67). 57 (67.9%) pts were protocol eligible. The conditioning regimen was melphalan 100-140mg/m2 with fludarabine, +/- thiotepa previously described by us (Ciurea SO, BBMT 2012;18:1835). All had a bone marrow graft except 4 pts (95%). Diagnoses were AML/MDS 49 (58.3%), CML 9 (11%), ALL 10 (12%), and lymphoma/CLL 13 (15%) (4 Hodgkin's, 4 NHL, 5 CLL), other 3 (3.6%). 28/46 (61%) pts with myeloid diseases were in CR at transplant, and 15/21 pts with AML in remission had poor-risk cytogenetics. Donors were siblings (N=36), children (N=35), parents (N=12), cousin (N=1). All pts achieved engraftment except 3 (96.4%), 91.6% with full donor chimerism. Median time to neutrophil engraftment was 18 days (11-43 days). The cumulative incidence of acute and cGVHD for different groups is presented in Table 1. Overall, for the entire cohort, the NRM was 25.7%, relapse rate was 32% and PFS was 42.3%. The median OS for first transplants was 25.6 months (mo) and 6.5 mo for second transplant pts. For pts receiving their first transplant, PFS was similar for those who received full and RIC. Of the 49 pts who had first transplant for AML/MDS, 27 (55.1%) were in complete remission prior to transplant. NRM for these pts was 9%, relapse rate 24.3% and PFS 66.8% at 50 mo of median follow-up (Table 1, Figure 1B). Melphalan-based conditioning for HaploSCT offers good disease control with low treatment-related mortality. Factors associated with survival were protocol eligibility and remission status for myeloid diseases while the melphalan dose did not impact outcomes. A low relapse rate was observed for lymphoma/CLL pts.Table 1Outcomes overallAll patients(N=84) (%)Myeloid in CR (N=27) (%)ALL(N=10) (%)Lymphoma/CLL(N=14) (%)PNRM25.78.933.325.00.17Relapse Rate32.024.325.021.40.96PFS42.366.841.753.60.22aGVHD II-IV32.626.950.035.70.46aGVHD III-IV7.8037.57.10.0038cGVHD Lim.+Ext.21.321.757.117.90.11cGVHD Ext. only10.217.717.100.36 Open table in a new tab
Azacitidine (AZA) maintenance following allogeneic hematopoietic cell transplantation (HCT) for acute myeloid leukemia (AML) and myelodysplastic syndrome (MDS) may reduce relapse risk and improve survival. Given logistic and toxicity-related challenges, identifying subgroups appropriate for this approach is an unmet need. Using data from two centers, we retrospectively compared event-free survival (EFS) and overall survival (OS) of AML and MDS patients who received AZA maintenance (n = 59) with historic controls (n = 90). Controls were selected according to the following criteria: no death, relapse, or Grade III–IV acute GVHD for 100 days after transplant. In multivariable analysis, AZA maintenance yielded significantly improved EFS (p = 0.019) and OS (p = 0.011). Outcomes differed according to regimen intensity. For reduced-intensity transplant, EFS (p = 0.004) and OS (p = 0.004) were significantly improved and equivalent to myeloablative transplant. A significant benefit following myeloablative transplant was not observed. Within the limitation of its retrospective nature, this study suggests that AZA maintenance improves outcomes following reduced-intensity HCT, comparable to myeloablative HCT.
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