Refractory Severe Aplastic Anemia Research Articles

Real-World Effectiveness and Safety of Eltrombopag in Patients with Aplastic Anemia: A Chinese Nationwide Survey

Immunosuppressive therapy (IST) based on antithymoglobin (ATG) and cyclosporin (CsA) is the first-line treatment for severe aplastic anemia (SAA) patients who are unfit for transplantation,and the overall response rate (ORR) is about 70%.The utility of eltrombopag (EPAG),a TPO receptor agonist, achieved robust hematologic response in refractory and treatment-naïve SAA patients in clinical trials and some other studies. However, only a few data came from Asia countries where higher incidence of AA has been reported. A retrospective study on the use of EPAG in AA was conducted in mainland China. Aplastic anemia (transfusion dependent non-severe, severe, and very severe) patients who started eltrombopag before Feb 2019 and continued for at least 3 months either as first-line treatment or as rescue treatment, were enrolled. The maximum daily dosage of EPAG used continuously for at least 2 weeks is defined as the stable dosage. Response criteria were referred to that used in previous reports (Townsley DM, NEJM 2017; BCSH, BJH 2016). Fifty-six patients from eleven centers were enrolled in this study, including 26 males and 30 females at the median age of 39 (7-80) years. All patients were transfusion-dependent by the time of EPAG administration, and there were 14 VSAA, 24 SAA and 18 transfusion dependent non-severe aplastic anemia (TD-NSAA). Nineteen treatment-naïve patients received EPAG and IST (ATG+CsA, n=10; CsA/CsA+androgen, n=9) as first-line treatment. Thirty-seven patients were refractory to IST. Eltrombopag was administered at a median dose of 75 (25-150) mg per day for 7 (3-31) month. The median follow-up time was 9 (3-40) months. The overall response rate in patients receiving EPAG as first-line therapy was 78.9% (15/19), and most patients achieved complete response (CR) (10/15). Among the 10 patients receiving ATG+CsA, 6 patients achieved hematologic response (HR) at 3 months post-treatment, including 3 CR. Six patients were diagnosed as VSAA and three achieved HR. For the 9 patients treated with CsA/CsA+androgen, 8 achieved HR (88.9%) and 4 were CR (44.4%) at 3 months. By last follow-up, the cumulative HR rate was 70% in ATG+CsA group and 89% in CsA/CsA+ androgen group. Among the 14 responders, 11 patients receiving EPAG at a stable dosage ≤75mg/d and achieved HR at 3 months. The overall response rate in IST-refractory patients was 46% (17/37), with trilineage response in 27% patients at 3 months. For the 18 ATG+CsA refractory SAA patients,trilineage HR occurred in 4 patients (22.2%, 4/18), bi-lineage HR in one patient and single lineage HR in one patient. Thus, the total HR was 33.3% (6/18) at 3 months and increased to 44% (8/18) by last follow-up. Among the 19 CsA/CsA+ androgen refractory patients, 6 (31.5%, 6/19) achieved trilineage HR, one achieved bi-lineage HR and 4 achieved single lineage response. Total HR rate was 57.9% (11/19) at 3 months after EPAG initiation and 68% (13/19) by last follow-up, including 9 patients with trilineage HR. Among 17 responders, 13 received a stable EPAG dose of≤75mg/d. Most patients tolerated EPAG well. Adverse events occurred in 29 patients (52%) and most were mild to moderate, including gastrointestinal symptom (n=15, e.g. abdominal pain, nausea), impaired liver function (n=5), skin changes (n=7, e.g. skin pruritus and rash) and musculoskeletal pain (n=6), and venous thrombus (n=2). Eltrombopag dosage was reduced in 2 patients due to severe digestive symptoms at 100 mg/d and discontinued in one patient who suffered from upper limb venous thrombus. In conclusion, EPAG is effective and safe in treating Chinese AA patients at a daily dose of 75mg and less. The real-world result of EPAG in Chinese patients is similar to those reported in Western countries. Disclosures No relevant conflicts of interest to declare.

PRO99 MULTI-CRITERIA DECISION ANALYSIS (MCDA) OF ELTROMBOPAG OLAMINE IN THE TREATMENT OF PATIENTS WITH REFRACTORY SEVERE APLASTIC ANEMIA UNDER THE PRIVATE HEALTHCARE SYSTEM IN BRAZIL: A HEALTH TECHNOLOGY ASSESSMENT (HTA) ALTERNATIVE MODEL

In Brazil, cost-effectiveness analysis is not the only indicator to orient the decision-making process, especially due to the absence of a willingness-to-pay threshold. Using an alternative health technology assessment (HTA) model, this study aimed to evaluate the incorporation of eltrombopag olamine for the treatment of refractory severe aplastic anemia (SAA) in the Brazilian supplementary healthcare system (SHS). An alternative HTA model, consisting of a Multi-Criteria Decision Analysis (MCDA) presented the following flow: problem context description, criteria selection, process structuring, criteria rationale construction, score attribution, mathematical model elaboration and software development. Criteria and score scales were based on literature review and validated by specialists in a second moment. The criteria were organized according to relevance level in a proposed HTA diagram, and they were classified as eliminators, decision-making and complementary. Eliminators criteria were the minimum requirements to enable the technology assessment on the next steps. Decision-making criteria represented the main aspects/indicators during the HTA processes, including clinical benefit and economic impact of eltrombopag. Complementary criteria were only evaluated in case the previous result demonstrated any uncertainty. Scores were indicated by a scale of 0 to 10. A sensitivity analysis was performed to verify the result robustness. Eltrombopag attended all requirements of eliminators criteria. Regarding the decision-making criteria, the new technology indicated to have an assertive clinical evidence (7.00), with very high drug effect (8.50) and very low budget impact (0.02) due to very small use expectation. Final score was 7.69, which indicated recommendation of eltrombopag incorporation. There was no need to evaluate the complementary criteria. Sensitivity analysis results demonstrated consistency to the base case. Considering the context of refractory SAA in Brazil, the incorporation of eltrombopag demonstrated to be favorable, being an option with high overall clinical benefit accompanied by a very low budget impact in the Brazilian SHS.

The role of mesenchymal stem cells in allogeneic hematopoietic stem cell transplantation for patients with refractory severe aplastic anemia

目的评价脐血间充质干细胞（UC-MSC）在难治性重型再生障碍性贫血（RSAA）异基因造血干细胞移植（allo-HSCT）中的疗效及安全性。方法回顾性分析郑州大学附属肿瘤医院2010年3月至2018年7月间接受UC-MSC联合同胞相合供者及无关供者）外周血干细胞移植的25例RSAA患者，其中同胞相合供者组10例，无关供者组15例。结果25例患者中男13例，女12例。1例患者原发性植入失败，1例患者血小板植入不良，其余23例患者成功植入。粒细胞、血小板植入的中位时间分别为12.5（10~23）d、15（11~25）d。Ⅰ/Ⅱ度急性GVHD、慢性GVHD发生率分别为37.5%（9/24）、21.7%（5/23），未发生重度GVHD和严重移植相关并发症。中位随访22.9（1.6~107.8）个月，25例患者中4例死亡，21例（84%）存活，移植后5年总生存率为（83.6±7.5）%。结论UC-MSC联合allo-HSCT治疗RSAA疗效确切、安全性好。

Treatment optimization and genomic outcomes in refractory severe aplastic anemia treated with eltrombopag

AbstractEltrombopag (EPAG) received approval from the US Food and Drug Administration for the treatment of refractory severe aplastic anemia (rSAA) based on treatment of 43 patients with doses escalating from 50 to 150 mg daily for 12 weeks. Response kinetics suggested that more prolonged administration of EPAG at a dose of 150 mg could speed and improve response rates. We enrolled 40 patients with rSAA in a study of EPAG 150 mg daily, with a primary end point of response at 24 weeks. Twenty (50%) of 40 patients responded at 24 weeks; 5 (25%) of 20 would have been deemed nonresponders at 12 weeks, the end point of the previous study. Fifteen of the 19 responding patients continuing on EPAG had drug discontinued for robust response; 5 of the 15 required EPAG re-initiation for relapse, with all recovering response. To analyze risk of clonal progression, we combined long-term data from the 83 patients with rSAA enrolled in both studies. Evolution to an abnormal karyotype occurred in 16 (19%), most within 6 months of EPAG initiation. Targeted deep sequencing/whole-exome sequencing was performed pre-EPAG and at primary response end point and/or time of clonal evolution or longest follow-up. Cytogenetic evolution did not correlate with mutational status, and overall mutated allele fractions of myeloid cancer genes did not increase on EPAG. In summary, extended administration of EPAG at a dose of 150 mg for 24 weeks rescued responses in some patients with rSAA not responding at 12 weeks. The temporal relationship between clonal evolution and drug exposure suggests that EPAG may promote expansion of dormant preexisting clones with an aberrant karyotype. The studies were registered at www.clinicaltrials.gov as #NCT00922883 and #NCT01891994.

PB2338 LONG‐TERM OUTCOMES OF ALLOGENEIC HEMATOPOIETIC CELL TRANSPLANTATION IN SEVERE APLASTIC ANEMIA: PNH CLONES MATTER

Background:Severe aplastic anemia (SAA) is a disease of young people, in which allogeneic hematopoietic cell transplantation (HCT) provides curative therapy. Recent studies have investigated the role of peripheral grafts, complement inhibition in patients with paroxysmal nocturnal hemoglobinuria (PNH) clones and long‐term complications.Aims:We aimed to investigate long‐term outcomes and prognostic factors in HCT for SAA.Methods:We retrospectively reviewed charts from consecutive patients transplanted for SAA in our center and analyzed: age, gender, ECOG performance status, time from diagnosis to transplant, pre‐transplant lines of treatment, pre‐transplant transfusions, hemoglobin and neutrophils at transplant, PNH (paroxysmal nocturnal hemoglobinuria) clone, graft source (bone marrow/peripheral), donor type (sibling/unrelated), HLA matching, conditioning (Cyclophospamide/Anti‐thymocyte globulin with/without fludarabine), days of neutrophil and platelet engraftment, acute (grade 2‐4) and extensive chronic GVHD (graft‐versus‐host disease), TMA (thrombotic microangiopathy), post‐transplant lymphoproliferative disease (PTLD), infections, relapse, late complications and survival. Multivariate analyses were performed to determine independent predictors of outcomes.Results:We studied 24 patients aged 20 years (range 9‐46) with severe (18) or very severe (6) AA. PNH clones (≥60%) were detected in 3 patients without signs of hemolysis or thrombosis. The majority of patients (14/24) were transplanted at refractory disease following a median of 2 (range 1‐4) previous lines of immunosuppression according to current recommendations. Bone marrow grafts (19/24) and sibling donors (15/24) were preferred when available. Matched unrelated donors were chosen in 7 patients without sibling donors; while 2 patients were transplanted with mismatched unrelated donors before the era of alternative transplantations. Fludarabine was added to the conditioning in 13/24 patients.Regarding complications, TMA developed in 3 patients and resolved in 2/3 with cyclosporine cessation; whereas PTLD in 2: one involving the central nervous system with a fatal outcome and one that resolved after Rituximab treatment. With a follow‐up of 59 months (0.9‐245), CI (cumulative incidence) of acute GVHD was 22.2% and was independently predicted by peripheral grafts and TMA (p<0.001). Ten‐year CI of chronic GVHD was 31.6% and was independently predicted by hemoglobin at transplant (p = 0.007), HLA mismatch (p<0.001), PNH clones (p<0.001) and TMA. Lastly, 10‐year OS reached 70.6% and was independently associated with a PNH clone (p = 0.034, Figure). No secondary malignancy or fatal long‐term complication was observed.Summary/Conclusion:Our study confirms that high cure rates are achieved in heavily immunosuppressed refractory SAA patients, even in older ones or post unrelated transplants. Peripheral grafts increase morbidity but not mortality in HCT for AA. More than that, our data suggest that novel approaches are needed in patients with PNH clones in order to improve morbidity and mortality in the era of effective complement inhibition.image

Ex Vivo Nicotinamide-Expanded (NAM-Expanded) Unrelated Cord Bloodtransplantation (UCB) for Refractory Severe Aplastic Anemia Results in Rapid Engraftment and Expedites Immune Recovery

Introduction Allogeneic hematopoietic stem cell transplantation using UCB (UCBT) is an alternative approach for severe aplastic anemia (SAA) patients (pts) lacking an HLA matched donor. However, UCBT is associated with delayed engraftment, high graft failure and mortality rates. Ex-vivo expanded UCB using nicotinamide (NAM) can engraft in NOD/SCID mice, and in pts with hematological malignancies in pilot studies. Here we investigated whether NAM-expanded UCBT would accelerate engraftment and immune reconstitution in refractory SAA pts. Methods Eligible SAA pts with severe neutropenia who failed immunosuppressive therapy underwent a NAM-expanded UCBT at a single center in a phase II trial. Pts were conditioned with cyclophosphamide (60 mg/kg × 2), horse ATG (40 mg/kg × 4), fludarabine (25 mg/m2 × 5) and 200cGy of TBI. GVHD prophylaxis included tacrolimus and MMF. Cohort 1 is designed to transplant six pts with a single NAM-expanded unit combined with 3 × 106 CD34+ cells/kg from a haploidentical donor as a stem cell backup. Once adequate cord engraftment is established, Cohort 2 will transplant a NAM-expanded unit alone. Engraftment and immune recovery were assessed and compared with a conventional UCBT combined with haploCD34+ cells using the same conditioning and GVHD prophylaxis. Results From 2017 to 2018, two SAA pts (22 years male and 45 years female, pre-transplant ANC ≤300/uL, failed ATG/CSA/Eltrombopag) successfully underwent a single 6/8 HLA-matched NAM-expanded UCBT combined with haplo CD34+ cells. UCB units before expansion contained a median total nucleated cell (TNC) dose of 2.8 × 107/kg and 1.7 × 105 CD34+ cells /kg. At transplant, NAM-expanded units contained a median 6.0 × 107 TNCs/kg and 96.4 × 105 CD34+ cells /kg, representing a median TNC and CD34+ cell expansion of 2-fold and 52-fold, respectively. At 14 and 7 months post-transplant, both pts were GVHD-free and survived with stable engraftment and transfusion independence (Figure 1). The median time to neutrophil and platelet recovery was only 6.5 days (range 6-7) and 35.5 days (31-40), with >95% cord donor myeloid and T-cell chimerism occurring at days 6.5 and 23.5, respectively. Immune recovery was brisk (Figure 2): absolute CD4+ > 200 cells/μL occurred at 17 and 60 days; at day 100, mean CD4+ cells were 382/μL and IGA 92 mg/dL. Compared to 16 SAA pts who received a single unexpanded UCBT with haplo CD34+ cells at our institute from 2013 to 2016, engraftment and immune recovery were superior in SAA pts with NAM-expanded UCBT (P Conclusion These results show for the first time that NAM-expanded UCB results in rapid cord engraftment, sustained hematopoiesis and accelerated immune recovery in treatment refractory, neutropenic SAA pts. The high numbers of CD34+ progenitor cells in NAM-expanded grafts could potentially overcome graft failure which limits conventional UCBT for SAA.

Rapid Engraftment and Immune Recovery in Treatment Refractory Severe Aplastic Anemia Patients Undergoing Ex Vivo Nicotinamide-Expanded (NAM-Expanded) Unrelated Cord Blood Transplantation

Introduction. Allogeneic hematopoietic stem cell transplantation using UCB is an alternative approach for pts (pts) with hematological malignancies lacking an HLA matched donor. However, for pts with severe aplastic anemia (SAA), UCB transplants are associated with delayed engraftment, high graft failure rates and poor survival. Ex-vivo expanded UCB using nicotinamide (NAM) can engraft in NOD/SCID mice, and in pilot studies in pts with hematological malignancies, results in rapid engraftment and durable hematopoiesis. Here we investigated a novel transplantation approach using NAM-expanded UCB in refractory SAA pts who lacked an HLA matched donor, hypothesizing this regimen would accelerate engraftment and immune reconstitution compared to conventional UCB transplants.PTS AND Methods. Eligible pts had SAA with severe neutropenia (ANC<1000) unresponsive to immunosuppressive therapy (IST) underwent a NAM-expanded UCB-transplant at a single center in a phase II trial (NCT03173937). Pts were conditioned with cyclophosphamide (60 mg/kg x 2), horse ATG (40 mg/kg x 4), fludarabine (25 mg/m2 x5) and 200cGy of TBI. GVHD prophylaxis included tacrolimus and MMF. Cohort I is designed to transplant six pts with a single NAM-expanded unit combined with 3 x 106 CD34+ cells/kg from a haploidentical donor as a backup stem cell source. Once adequate cord engraftment is established in Cohort 1, the study will proceed to Cohort 2 to transplant a NAM-expanded unit alone without haplo-CD34+ cells. Engraftment, chimerism, and immune recovery were assessed and compared with SAA pts who received a conventional non-expanded UCB transplant combined with haplo-CD34+ cells using the identical conditioning and GVHD prophylaxis.Results. From 2017 to 2018, two SAA pts (22 years male and 45 years female, pre-transplant ANC ≤300/uL, and had failed ATG/CSA/Eltrombopag) were successfully transplanted with a single ≥ 6/8 HLA-matched NAM-expanded UCB unit combined with haplo CD34+ cells from a relative. The UCB units before expansion contained a median total nucleated cell (TNC) dose of 2.8 x 107/kg and 1.7 x 105 CD34+ cells/kg. At transplant, the cultured NAM-expanded units contained a median 6.0 x 107 TNCs/kg and 96.4 x105 CD34+ cells/kg, representing a median post TNC and CD34+ cell expansion of 2-fold and 52-fold, respectively. At 12 months and 5 months post-transplant, both pts survive with stable engraftment, transfusion independence, and without acute or chronic GVHD. The median time to neutrophil recovery (ANC > 500/ μL) was only 6.5 days (range 6-7), and platelet recovery was 35.5 days (31-40); chimerism studies showed that both pts achieved >95% cord donor myeloid chimerism and T-cell chimerism at a median 6.5 (6-7) and 23.5 days (21-26) respectively . Immune recovery in both pts receiving NAM-expanded UCB was brisk (Figure 1): absolute CD4+ count > 200 cells/μL occurred at 17 and 60 days; at day 100, median CD4+ numbers was 382/μL and median IGA was 92 mg/dL. In comparison to 16 SAA pts transplanted sequentially at our institute from 2013-2016 using a single unexpanded CBU combined with haplo CD34+ cells, median cord graft doses were 3.6 x 107 TNCs/kg and 1.2 x105 CD34+ cells/kg; b) median time to ANC and platelet recovery was 10 and 51 days; c) median time to >95% cord donor myeloid chimerism was 63 days; d) at day 100, only 3/16 (19%) unexpanded UCB recipients had CD4+ count > 200 cells/μL, and the median CD4+ number was only 74 cells/μL and the median IGA was only 31 mg/dL. This first in human transplant trial suggests neutrophil engraftment, platelet recovery, and post-transplant immune recovery are superior inSAA pts transplanted with NAM expanded UCB compared to conventional nonexpanded UCB (all P<0.05, Figure 1).Conclusion. These encouraging preliminary results show for the first time that NAM-expanded UCB results in rapid cord engraftment, sustained hematopoiesis and accelerated immune recovery in treatment refractory, neutropenic SAA pts. The high numbers of transplanted CD34+ progenitor cells in NAM-expanded grafts could potentially overcome graft failure associated with conventional UCB transplantation for SAA, obviating the need for co-transplanting haplo CD34+ cells as a stem cell back-up. DisclosuresNo relevant conflicts of interest to declare.

Eltrombopag for Moderate Aplastic Anemia and Unilineage Cytopenias: Dosing, Long-Term Follow-up, Clonal Evolution and Somatic Mutation Profiling

▪Eltrombopag (EPAG) received FDA approval for treatment of refractory severe aplastic anemia (rSAA) in 2014, based on our phase I/II dose escalation trial of single agent EPAG for patients failing one or more treatment cycles with ATG/cyclosporine (Olnes NEJM 2012; Desmond Blood 2014). There is no standard treatment for patients with moderate aplastic anemia (MAA) or hypo-productive uni-lineage cytopenias (MAA/UC), conditions that can also impact on morbidity, mortality and quality of life. To explore the safety and effectiveness of EPAG in MAA/UC, we conducted a phase II study of EPAG given at escalating doses from 50-300mg/day (25-150mg/day for East Asians) through a primary hematologic response endpoint at 16-20 weeks (NCT 01328587). 34 patients enrolled between February 2012 and March 2017. 27 had never been treated with ATG/CSA IST. Responding patients could continue EPAG treatment on an extension arm.The drug was well-tolerated in 33/34 patients, with 1 patient coming off study at 10 weeks for nausea and vomiting. 25 patients reached the maximal dose. The median duration of follow-up in all patients was 16 months, and 27 months in responding patients. 17 of 34 (50%) of patients met criteria for response at the primary endpoint in at least one initially protocol-qualifying lineage (Hb <8.5 or RBC transfusion-dependence and/or platelets < 30,000 or platelet transfusion-dependence). 12 of 23 with severe anemia had an erythroid response (> 1.5 gr/dL increase in Hb or > 50% reduction in transfusions), including a patient with RSP19-mutated Diamond-Blackfan anemia. 7/24 with severe thrombocytopenia had a platelet response (>20,000/ul or transfusion-independence). 2/13 with both severe anemia and thrombocytopenia had bi-lineage responses at the primary endpoint, and an additional 5 went on to bi-lineage responses during the extension period. 3 patients without a response to EPAG were later treated with ATG/CSA and responded.EPAG was discontinued in 11/17 (65%) responding patients upon achievement of robust blood counts (Hb > 10mg/dL, platelets > 50,000/uL, and ANC > 1000) or stable blood counts for 6 months in 1 patient (6%) after a median duration of drug administration of 8 months (2-14 months) (Fig 1). In contrast to our prior experience in rSAA, the majority of patients still being followed on study after drug discontinuation (8/10) needed to have EPAG re-initiated for declining counts at a median of 6 months (2-38) later. All 8 responded again, with 4/8 able to discontinue EPAG after a second robust or stable response.2/34 patients (6%) developed marrow cytogenetic abnormalities while on drug in contrast to 16/83 (16%) rSAA patients treated with EPAG in our prior studies. Both MAA patients were responders and neither had dysplastic changes or increased blasts. Patient # 5 had +8 in 7/20 metaphases at 22m, went off drug and relapsed, and EPAG was restarted off protocol. Repeat cytogenetics on EPAG were normal. Patient #20 had del13q in 5/20 metaphases at 9.5m, EPAG was stopped, and repeat cytogenetics off drug were normal. Of note, patient #12 had a robust response and had been off drug for 25m when counts declined and BM revealed dysplastic changes with no increase in blasts and normal cytogenetics.The acquisition and selection of somatic mutations, particularly in myeloid candidate genes (MCG) recurrently mutated in MDS/AMLhas been proposed to be an initiating step in clonal evolution. We performed targeted next generation exome sequencing (NGS) of 66 MCG and additional genes previously found to be somatically-mutated in SAA (Yoshizato, NEJM, 2015) pre-EPAG treatment and at the primary response endpoint of 16-20 weeks. Only 5 patients showed somatic mutations in these genes at baseline (SETBP1, CBL, SF3B1, PPMID, EP300). There were no significant changes in VAF. 2 mutations became detectable on EPAG (BCOR, and DNMT3A transiently), and 1 disappeared (SF3B1).In summary, administration of EPAG at escalating doses up to of 300mg/day was well-tolerated and 50% of patients with MAA/UC had clinically-meaningful responses, including those not previously treated with IST. The responses were durable, often robust, although frequently required ongoing EPAG treatment, in contrast to the experience in rSAA. Clonal cytogenetic evolution was rare, with no instances of chromosome 7 abnormalities, and there was no consistent expansion of MCG somatically-mutated clone size in patients during EPAG treatment. DisclosuresWinkler:National Institute of Health: Research Funding. Young:CRADA with Novartis: Research Funding; GlaxoSmithKline: Research Funding; National Institute of Health: Research Funding. Dunbar:National Institute of Health: Research Funding.

Comparing Costs and Consequences of Eltrombopag Plus IST Versus IST Alone for First-Line Treatment in Severe Aplastic Anemia: Results from a Responder Analysis

Background: Severe Aplastic Anemia (SAA) is a rare bone marrow disorder characterized by inadequate levels of peripheral, multi-lineage blood cells.Of the two available first-line treatments for SAA, allogeneic hematopoietic stem cell transplantation is limited by patient eligibility and donor availability, and immunosuppressive therapy (IST) is characterized by a significant proportion of non-responders, toxicity, and risk of transformation to diseases such as acute myelogenous leukemia. Patients who do not respond to treatments become transfusion dependent, which has a significant impact on patients' quality of life as well as healthcare costs.Eltrombopag is the only TPO-R agonist approved for the treatment of refractory SAA. In a Phase I/II clinical trial, eltrombopag, given in association with IST, showed efficacy in patients with naive-SAA, and offers a significantly improved first-line alternative to patients affected by SAA (Townsley, et al 2017). In the US healthcare environment, there is a need to compare costs and consequences to understand value.Objective: Evaluate eltrombopag as a first-line treatment versus IST alone for SAA from the American private healthcare system perspective.Methods: A responder model for newly diagnosed SAA patients was created to assess the treatment pathway and economic impact of including eltrombopag in addition to IST (antithymocyte globulin and cyclosporine A) as a first-line treatment. A simulated cohort with two treatment arms underwent 6 months of treatment with either eltrombopag in addition to IST or with IST alone and were followed for a 3-year time period. Each arm received a diagnostic test measuring response at 6 months. Patients who achieved complete or partial response in either arm received low-dose cyclosporine A as maintenance therapy for an additional 6 months of treatment. Patients who did not respond in either arm continued with eltrombopag monotherapy as a second-line therapy for an additional 6 months.First-line therapy (eltrombopag with IST, IST alone), maintenance therapy (low-dose cyclosporine A), second-line therapy (eltrombopag monotherapy), administration, routine care, mortality and adverse event costs were included in the analysis. Workplace productivity related costs were not considered. Response rates, mortality, dosing, treatment duration and adverse event rates for each arm were based on a phase I/II trial (Townsley, et al 2017). Drug costs were obtained from a large online database (REDBOOK Online). Administration costs were based on the 2017 CMS Medical Fee Schedule. Routine care rates (visits, hospitalizations, tests and transfusions) were based on published data (Peffault De Latour, et al, 2017). Routine care, mortality and adverse event costs were based on CPT codes from the American Medical Association, HCUPnet and published data (Toner, et al 2011). All cost data are reported in 2018 US dollars. See figure 1 for details.Results: In a simulated cohort with a population of one million, the annual incidence of aplastic anemia was 0.000234% and SAA accounted for 83.8% of those cases. The two treatment paths were compared for their consequences. Based on the clinical trial data, in the treatment arm with eltrombopag and IST, 94% of patients experienced treatment response relative to the IST arm where only 66% of patients experienced treatment response. Further, in the treatment arm with eltrombopag and IST, the patients experienced a reduced annual risk of mortality by 0.3% relative to the IST arm. Use of eltrombopag therapy as a first-line therapy produced a cost increase of $77,442 over 3 years. First-line drug costs accounted for an increase of $109,147, while improvements in response rates led to cost offsets for second-line drugs and produced $29,663 in savings. Adverse event, routine care and mortality costs had relatively negligible effects on either treatment arm over a 3-year time period. Sensitivity analyses confirmed the robustness of the analysis.Conclusion: When following treatment approaches specified in clinical studies, high response rates combined with reduced risk of mortality and less usage of rescue medication, and a low disease incidence are likely to lead to manageable economic consequences with eltrombopag + IST therapy from the American private healthcare system perspective. In a simulated cohort with a population of one million, this was estimated to be $77,442 over 3 years. [Display omitted] DisclosuresSaid:Novartis: Employment. Cai:Novartis: Employment. Forsythe:Novartis: Consultancy. Roy:Novartis: Employment.

Unrelated cord blood transplantation in patients with idiopathic refractory severe aplastic anemia: a nationwide phase 2 study

AbstractOutcomes remain poor for refractory severe aplastic anemia (SAA) patients. Alternative donor transplantation may be considered, but results from previous studies are not encouraging. We conducted a prospective nationwide phase 2 study to assess unrelated cord blood (CB) transplantation (CBT) efficacy and safety in refractory SAA patients (Aplastic Anemia and Cord Blood Transplantation protocol). To demonstrate a significant difference in 1-year survival from 20% (null hypothesis) to 50% (alternative hypothesis), we needed to include 25 transplanted patients and therefore included 26 (median age, 16 years). Eligibility criteria required 1 or 2 unrelated CB units, containing separately or together >4 × 107 frozen nucleated cells (NCs) per kilogram of recipient body weight. Conditioning regimen comprised fludarabine (FLU), cyclophosphamide (CY), antithymocyte globulin (ATG), and 2-Gy total body irradiation (TBI). With a median follow-up of 38.8 months, engraftment occurred in 23 patients (88%); cumulative incidences of grade II-IV acute and chronic graft-versus-host disease were 45.8% and 36%, respectively. Twenty-three patients were alive at 1 year, with an 88.5% overall survival (OS) rate, differing significantly from the expected 20% (P < .0001; 84% OS at 2 years). CBT with units containing ≥4 × 107 frozen NCs per kilogram is therefore a valuable curative option for young adults with refractory SAA and no available matched unrelated donors. This trial was registered at www.clinicaltrials.gov as #NCT01343953.

Eltrombopag for Refractory Severe Aplastic Anemia: Dosing Regimens, Long-Term Follow-up, Clonal Evolution and Somatic Mutation Profiling

▪Eltrombopag (EPAG) received FDA approval for treatment of refractory severe aplastic anemia (rSAA) in 2014, based on our phase I/II dose escalation trial of single agent EPAG at 50-150 mg daily over a period of 12 weeks (Olnes NEJM 2012; Desmond Blood 2014).Two observations warranted further investigation of EPAG in this unique patient population. First, cell count kinetics and lack of acute toxicities suggested that extended administration of EPAG at a fixed dose of 150mg could speed and improve response rates. Second, 19% of patients developed new cytogenetic abnormalities on EPAG, raising concerns that EPAG might promote progression to MDS/AML.We conducted a subsequent phase II study of EPAG given at a fixed daily dose of 150mg for 6 months in patients with rSAA (NCT01891994). Thirty-nine patients enrolled between July 2013 and April 2017. Primary endpoint was hematologic response at 6 months. Responding participants could continue EPAG treatment. Secondary endpoints included response at 3 months and the rate of clonal cytogenetic evolution.Nineteen of 39 (49%) patients met criteria for hematologic response at 6 months. Of these, 5/19 (26%) patients would have been deemed non-responders at 3 months of treatment. EPAG was continued in 18 patients on the extension arm. EPAG was discontinued for robust response in 13/18 (72%) after a median duration of drug administration of 12 months (6-27.5 months). EPAG was re-initiated for relapse in 3/13 patients, and all 3 recovered response.At median follow up of 6 months (range 2 - 39 m), 6/39 patients (15%) developed marrow cytogenetic abnormalities, a rate comparable to our previous cohort.Given the similar rates of response and clonal evolution in our two consecutive studies, we analyzed the relationship between outcomes and cytogenetic progression for all patients (n=83) at up to 8 years of follow-up. Sixteen of 83 (18%) patients clonally evolved (Table 1). Clonal evolution was an early event after EPAG initiation. Evolution occurred within 6 months in 13/16 evolvers (81%), and in 6/6 evolvers with high risk chromosome 7 abnormalities (5/6 within 3 months). The frequency of high risk clonal evolution 24 months post intervention is comparable to historic controls with rSAA. However, direct temporal comparisons of evolution events are limited by differences in the sequence of cytogenetic tests. Non-chromosome 7 cytogenetic abnormalities were often transient, and not associated with dysplasia. Two evolvers continued EPAG off the rSAA protocol and cytogenetics normalized (UPN 71,14).The acquisition and selection of somatic mutations, particularly of myeloid candidate genes recurrently mutated in MDS/AML, has been proposed to be an initiating step in clonal evolution. We performed whole exome sequencing (WES) on samples obtained pre-EPAG treatment and at the primary response endpoint and/or time of clonal evolution in 21 responding patients and in 11 patients with cytogenetic evolution. Candidate gene mutations were detected in patients who responded (6/21) and in those with cytogenetic evolution (4/11). Clonal hematopoiesis without an identifiable driver mutation was common. In post-EPAG samples, additional myeloid candidate gene somatic mutations were detected in 2 cytogenetic evolvers and in 4 responding patients, all at low variant allele frequencies (VAF) that were close to the 2.5% detection threshold. There was no significant change in VAF in either candidate or in non-candidate genes in responders and in cytogenetic evolvers. Only one early cytogenetic evolver (UPN 01) showed an expansion of mutated clones (SETBP1 and RUNX1), from VAF of 5% to 40%, when 80% of bone marrow metaphases showed chromosome 7q deletion.In summary, extended administration of EPAG at a fixed dose of 150 mg daily for 6 instead of 3 months induces additional responses in a subgroup of patients with rSAA. After EPAG was discontinued, most patients maintained durable robust responses. The temporal relationship between clonal evolution and drug exposure suggests that in a subgroup of patients, EPAG may promote expansion of dormant pre-existing clones with an aberrant karyotype. No clinical or laboratory findings prior to therapy, including WES, predicted response or risk of clonal evolution. Careful monitoring of refSAA patients treated with EPAG is indicated, particularly in the first 6 months of treatment. [Display omitted] DisclosuresWinkler:Novartis/GSK to institute: Research Funding. Cooper:Novartis/GSK to institute: Research Funding. Townsley:Novartis/GSK to institute: Research Funding. Grasmeder:Novartis/GSK to institute: Research Funding. Young:Novartis/GSK to institute: Research Funding. Dunbar:Novartis/GSK to institute: Research Funding.

Treatment for the relapsed or refractory severe aplastic anaemia

Severe aplastic anaemia is a marrow failure syndrome characterized by pancytopenia and hypocellular bone marrow.Immunosuppressive therapy(IST)consisting of antithymocyte or antilymphocyte globulin(ATG)and cyclosporine has been recommended for children lacking HLA-matched sibling donor.For children unresponsive to initial immunosuppressive therapy or those who relapsed after first immunosuppressive therapy, they can choose repeat course of immunosuppressive therapy, allogeneic hematopoietic stem cell transplantation, new drugs or clinical trials.And the prognosis has been greatly improved in recent years, dueing to the development of allogeneic hematopoietic stem cell transplantation and the application of new drugs such as eltrombopag.This article reviews researches advance in different therapies for children with relapsed or refractory severe aplastic anaemia, aimed at guaiding treatment methods. Key words: Severe aplastic anaemia; Children

Mesenchymal Stem Cell Benefits Observed in Bone Marrow Failure and Acquired Aplastic Anemia.

Acquired aplastic anemia (AA) is a type of bone marrow failure (BMF) syndrome characterized by partial or total bone marrow (BM) destruction resulting in peripheral blood (PB) pancytopenia, which is the reduction in the number of red blood cells (RBC) and white blood cells (WBC), as well as platelets (PLT). The first-line treatment option of AA is given by hematopoietic stem cell (HSCs) transplant and/or immunosuppressive (IS) drug administration. Some patients did not respond to the treatment and remain pancytopenic following IS drugs. The studies are in progress to test the efficacy of adoptive cellular therapies as mesenchymal stem cells (MSCs), which confer low immunogenicity and are reliable allogeneic transplants in refractory severe aplastic anemia (SAA) cases. Moreover, bone marrow stromal cells (BMSC) constitute an essential component of the hematopoietic niche, responsible for stimulating and enhancing the proliferation of HSCs by secreting regulatory molecules and cytokines, providing stimulus to natural BM microenvironment for hematopoiesis. This review summarizes scientific evidences of the hematopoiesis improvements after MSC transplant, observed in acquired AA/BMF animal models as well as in patients with acquired AA. Additionally, we discuss the direct and indirect contribution of MSCs to the pathogenesis of acquired AA.

Alternative Donor Transplantation with High-Dose Post-Transplantation Cyclophosphamide for Refractory Severe Aplastic Anemia

Severe aplastic anemia (SAA) is a life-threatening hematopoietic stem cell disorder that is treated with bone marrow transplantation (BMT) or immunosuppressive therapy (IST). The management of patients with refractory SAA after IST is a major challenge. Alternative donor BMT is the best chance for cure in refractory SAA, but morbidity and mortality from graft failure and complications of graft-versus-host disease (GVHD) have limited enthusiasm for this approach. Here, we employed post-transplantation high-dose cyclophosphamide in an effort to safely expand the donor pool in 16 consecutive patients with refractory SAA who did not have a matched sibling donor. Between July 2011 and August 2016, 16 patients underwent allogeneic (allo) BMT for refractory SAA from 13 haploidentical donors and 3 unrelated donors. The nonmyeloablative conditioning regimen consisted of antithymocyte globulin, fludarabine, low-dose cyclophosphamide, and total body irradiation. Post-transplantation cyclophosphamide 50 mg/kg/day i.v. on days +3 and +4 was administered for GVHD prophylaxis. Additionally, patients received mycophenolate mofetil on days +5 through 35 and tacrolimus from day +5 through 1 year. The median age of the patients at the time of transplantation was 30 (range, 11 to 69) years. The median time to neutrophil recovery over 1000 × 103/mm3 for 3 consecutive days was 19 (range, 16 to 27) days, to red cell engraftment was 25 (range, 2 to 58) days, and to last platelet transfusion to keep platelets counts over 50 × 103/mm3 was 27.5 (range, 22 to 108) days. Graft failure, primary or secondary, was not seen in any of the patients. All 16 patients are alive, transfusion independent, and without evidence of clonality. The median follow-up is 21 (range, 3 to 64) months. Two patients had grade 1 or 2 skin-only acute GVHD. These same 2 also had mild chronic GVHD of the skin/mouth requiring systemic steroids. One of these GVHD patients was able to come off all IST by 15 months and the other by 17 months. All other patients stopped IST at 1 year. Nonmyeloablative alloBMT using post-transplantation cyclophosphamide allowed for safe expansion of the donor pool to include HLA-haploidentical donors. This approach appears promising in refractory SAA patients. Importantly, engraftment was 100%, pre-existing clonal disease was eradicated, and the risk of GVHD was low.

Efficacy and Safety of Unrelated Cord Blood Transplantation in Patients with Acquired Refractory Aplastic Anemia: A Phase II Study on Behalf of Eurocord and the Francophone Society of Bone Marrow Transplantation and Cellular Therapy

▪Background: While therapeutic outcomes in patients (pts) with acquired severe aplastic anemia (SAA) has greatly improved in recent years, results remain poor for pts who are refractory to first-line immunosuppressive therapy (IST) and lack a matched unrelated donor. We conducted a prospective multi-center phase II uncontrolled trial to assess the efficacy and safety of cord blood transplantation (CBT) in refractory SAA pts (NCT 01343953).Patients and methods: Pts with SAA refractory to IST with anti-thymocyte globuline (ATG) and ciclosporin (CsA) were eligible in case of no existing matched unrelated donor but available one or two unrelated CB units containing alone or both together more than 4 x 107/Kg frozen nucleated cells/Kg (no more than 2 out of 6 HLA mismatches allowed between each CB unit and the pts). Pts with isolated bone marrow cytogenetic abnormality (absence of morphologic evidence of myelodysplastic syndrome) were also eligible. The conditioning regimen consisted of fludarabine 30mg/m2 (D-6 to D-3), cyclophosphamide 30mg/kg (D-6 to D-3), ATG (thymoglobulin) 2.5mg/kg at D-3 and D-2 (5mg/Kg total dose) and total body irradiation (2 Gray) on D-2. All pts received one injection of anti-CD20 (150 mg/m2) to prevent EBV reactivation (D+5). CsA was given alone as prophylaxis for graft versus host disease (GVHD). The trial used the Fleming's Single Stage Phase II Design, with sample size computed to demonstrate an improved overall survival (OS) rate at one year from 20% up to 50%, with type I and type II error rates set at 0.05. A minimum sample of 25 pts was required, with a minimum number of 9 pts alive at one year needed to indicate treatment efficacy. Secondary endpoints included cumulative incidences (CumI) of engraftment, acute and chronic GVHD (aGVHD and cGVHD), infections, relapse and late cause of death.Results: 29 pts were included between June 2011 and October 2015. One pt was diagnosed with dyskeratosis congenita and a matched unrelated donor was identified for another pt between inclusion and anticipated date of CBT (exclusion criteria). One pt died before CBT due to a septic shock (D+4 after inclusion). Analyses were conducted in the remaining 26 pts. Median time between SAA diagnosis and CBT was 12 months (interquartile range (IQR) 8.7 to 17.8). All pts received at least one course of IST before CBT (2 courses, n=8). Twenty-three pts received 20 or more transfusions before CBT. BM karyotyping showed 2 pts respectively with monosomy 7 in 4 and 15 mitoses as well as one pt with trisomy 8 in 4 mitoses before CBT. Eight pts (31%) have a PNH clone (median 4.5% [IQR: 3.1-9.7]). Median age at time of CBT was 16 years (IQR 8.25 to 23) and median follow-up is 13.2 months (IQR 4.1 - 23.5). Three out of 26 pts incorrectly received twice the dose of ATG (10mg/Kg total dose) due to protocol violation. Sixteen pts received one CB and 10 received two CB units. Median number of nucleated cells infused was 3.7 x 107/Kg (IQR, 3-4.9) and CD34+ cells infused was 1.4 x 105/Kg (IQR, 1.0-1.9). Myeloid engraftment occurred in 23 pts, with a 60 day-CumI of neutrophil engraftment of 88.5% with full chimerism for all of them. Three pts did not engraft (2 deaths at D71 and D92 and one successful second HSCT). The 100 day-CumI of grade II-IV acute GVHD was 40% (95% CI, 20-60) (8 grade II; 0 grade III; 2 grade IV). Among the 23 pts alive at day 100, five developed cGVHD leading to a one-year CumI of cGVHD at 27% (95% CI, 7-47) (severe cGvHD in 2 pts). During follow-up, 8 pts experienced a total of 18 grade III infections with a 6-month CumI of 32% (95% CI, 13-51). None of the pts presented post-transplant EBV-related lymphoproliferative disorder. Immune reconstitution is shown in Figure 1. Three pts died before 1 year due to non-engraftment (n=2) and infection (n=1, one of those who received twice the dose of ATG). With 23 pts alive at 1 year, the primary objective of the study was thus reached. The one-year treatment related mortality (TRM) was 15% (95%CI, 4-35). One additional pt who had also received twice the dose of ATG died (severe cGvHD) after one year (at 13.6 months), resulting in a 14-month OS of 81% (95%CI, 66-100) (Figure 2).Conclusion: CBT with at least 4 x 107/Kg frozen nucleated cells/Kg after a FLU-CY-TBI-ATG conditioning regimen gave rise to very encouraging results in this particular high risk SAA refractory population. Pediatric and young adult SAA pts who are refractory to first-line IST should now be considered either way for matched unrelated donor or CBT. [Display omitted] [Display omitted] DisclosuresPeffault de Latour:Novartis: Consultancy, Honoraria, Research Funding; Pfizer: Consultancy, Honoraria, Research Funding; Alexion: Consultancy, Honoraria, Research Funding; Amgen: Research Funding.

Interferon-γ Perturbs Key Signaling Pathways Induced By Thrombopoietin, but Not Eltrombopag, in Human Hematopoietic Stem/Progenitor Cells

▪Thrombopoietin (TPO) is the main regulator of hematopoietic stem and progenitor cell (HSPC) self-renewal and survival. Upon binding to its receptor, c-MPL, TPO activates cell signaling, through JAK-STAT and other pathways, which is tightly balanced by negative regulatory signaling processes. Recent studies indicate that chronic exposure of HSPCs to IFNγ, as exemplified in subjects with severe aplastic anemia (SAA), impairs self-renewal by perturbing TPO signaling pathways. Despite elevated levels of TPO in subjects with SAA, the TPO receptor agonist Eltrombopag (Epag) improves trilineage hematopoiesis in refractory SAA, suggesting that it may activate signaling within HSPC in a way that is distinct from TPO under inflammatory conditions. To address the paradox of Epag efficacy despite high endogenous TPO levels in bone marrow failure, G-CSF mobilized human CD34+ cells from 6 healthy donors were cultured in the presence of SCF, FLT3 and either TPO 5 ng/ml (TPO5) or Epag 3 μg/ml (Epag), with or without IFNγ 100 ng/ml. After 7 days in culture, cells were characterized via flow cytometry, CFU assay and transplantation in immunodeficient (NSG) mice. The percentages of CD34+ cells in cultures containing TPO5 or Epag alone were similar (83.3 ± 9.7% and 87.6 ± 7.1%, respectively), but were better preserved with Epag than TPO5 in the presence of IFNγ (46.7 ± 16.1% and 24.6 ± 15.0% respectively, p<0.05). Accordingly, when comparing 7-day cultures with and without IFNγ, the absolute numbers of CD34+ cells were markedly reduced with TPO5 (average 7.6-fold, p<0.005) but only minimally decreased with Epag (average 1.6-fold, p = n.s.). The adjusted numbers of CFUs after 7 days in the presence of IFNγ similarly decreased 2.7-fold with TPO5 but remained unchanged with Epag compared to cultures without IFNγ. When the 7-day expanded progeny of an equal starting number of CD34+ cells was transplanted in NSG mice, human cell engraftment was superior with Epag (34 ± 3.8% human CD45+ cells) than with TPO5 (21 ± 1.8% human CD45+ cells, p<0.05) cultures in the presence of IFNγ, suggesting an impact of Epag on the most primitive long-term repopulating HSPCs. To investigate potential mechanisms by which Epag positively affects maintenance of HSPCs under inflammatory conditions, we examined cell signaling pathways induced upon binding of TPO, Epag and IFNγ to their respective receptors in human CD34+ cells. At a concentration of 5ng/mL, TPO induced a rapid (peak < 1 hour) and high potency rise in STAT5 phosphorylation followed by a rapid (< 2 hours) decay in signal. In contrast, Epag induced a slow (peak 4 hours) low potency rise in STAT5 phosphorylation, and the signal persisted for at least 10 hours. The difference in cell signaling potency and kinetics between TPO and Epag is likely related to their binding to distinct regions of c-MPL, resulting in alternate receptor conformational changes. We next investigated the impact of IFNγ on TPO and Epag-induced STAT5 phosphorylation at the signal peak (<1 and 4 hours, respectively). As previously shown in murine HSPCs, IFNγ impaired TPO signaling in human HSPCs (Figure, panels A, C). In contrast, Epag-induced STAT5 phosphorylation was preserved or increased in the presence of IFNγ (Figure, panels B, C). When Epag and TPO were combined, inhibition of TPO signaling by IFNγ was partially restored (Figure, panel D). By reducing the dose of TPO from 5 to 1ng/mL, and therefore reducing the potency of signaling to levels similar to Epag, the inhibitory effect of IFNγ on TPO signaling was abolished (Figure, panel E). Activation of IFNγ receptor by its ligand induces phosphorylation of STAT1 and subsequent expression of suppressor of cytokine signaling-1 (SOCS-1), a negative regulator of both IFNγ and c-MPL receptors via inhibition of STAT1 and STAT5 phosphorylation, respectively. We found that IFNγ-induced phosphorylation of STAT1 was increased in the presence of TPO 5ng/mL (1.5-fold increase, p<0.05) but unaffected by Epag (1.1-fold increase, p = n.s.) or TPO 1ng/mL (1.1-fold increase, p = n.s.). Our data indicate that Epag counters IFNγ-induced perturbation of TPO signaling in human HSPCs. Epag produces an unopposed low potency, slow kinetic positive signal and activates c-Mpl above a threshold level critical for HSPC self-renewal. Epag’s evasion of IFN blockade of a critical pathway of growth factor cell signaling may explain its efficacy in improving hematopoiesis in SAA. [Display omitted] DisclosuresCheng:Novartis: Research Funding. Cheruku:Novartis: Research Funding. Alvarado:Novartis: Research Funding. Cash:Novartis: Research Funding. Dunbar:Novartis: Research Funding. Young:Novartis: Research Funding. Larochelle:Novartis: Research Funding.

SUCCESSFUL TREATMENT OF SEVERE APLASTIC ANEMIA WITH ELTROMBOPAG: A CASE REPORT

Introduction: Aplastic anemia is a clinical syndrome characterized by peripheral pancytopenia and deficiency of hematopoietic precursors in the bone marrow. Allogeneic hematopoietic stem cell transplantation (AHSCT) should be considered as first line treatment for young patients with an available donor. However,alternative therapy options are scant in patients who are not candidates for transplantation. Here, we report the efficacy of eltrombopag in a case of severe aplastic anemia. Case Presentation: Twenty-two years old female patient was admitted to the Marmara University Hospital because of severe aplastic anemia. There was not an available HLA-matched sibling donor and immunosuppressive treatment with horse-derived ATG (40 mg per day for 5 days) and cyclosporine (5 mg/kg per day) was started. At the sixth month of therapy she was still in need of transfusion. Eltrombopag was prescribed at a dose of 50 mg and the dose was increased up to 150 mg per day in 2 months. Successful response was noted within 2 weeks of 150 mg dosage and this response was sustained at the 4th month following discontinuation of drug. Conclusions: Eltrombopag provides good and permanent clinical response in refractory severe aplastic anemia.

Using Haploidentical (haplo) Donors and High-Dose Post-Transplant Cyclophosphamide (PTCy) for Refractory Severe Aplastic Anemia (SAA)

SAA is a life-threatening hematopoietic stem cell disorder that is treated with either bone marrow transplantation (BMT) or immunosuppressive therapy (IST). BMT is the treatment of choice for young patients with a matched sibling donor. Those with acquired SAA lacking a matched sibling donor and older adults (> 40yrs) generally receive IST because of the high toxicity of BMT in these patients. Roughly two-thirds will respond but up to 40% of patients eventually relapse or acquire a secondary clonal disease such as paroxysmal nocturnal hemoglobinuria (PNH) or myelodysplastic syndromes. Thus, a major challenge in treating SAA is the management of patients who are have relapsed after or are refractory to IST or who have acquired a secondary clonal disorder. BMT is the only curative therapy for patients with inherited SAA but problems with graft failure, graft-versus-host disease (GVHD), and lack of matched unrelated donors especially for ethnic minorities have limited the success of alternative donor BMT, especially for adult patients. PTCy after HLA-haplo BMT has been shown to facilitate engraftment and yield rates of GVHD comparable to those of matched sibling BMT in hematologic malignancies. We hypothesized that PTCy would improve outcomes for patients with refractory SAA by reducing the risk of GVHD and expanding the donor pool to include haplo related and mismatched unrelated donors (URD).Patients received an allogeneic BMT (alloBMT) for relapsed or refractory SAA. Relapsed/ refractory disease was defined as having failed previous course of IST. In the majority of the cohort, haplo donors were used due to lack of availability of suitable and timely URDs. Nine patients underwent haploBMT. A mismatched URD was used in one case of no suitably available haplo donor given the inherited nature of her disorder. The nonmyeloablative preparatory regimen included antithymocyte globulin (0.5 mg/kg day-9 and 2 mg/kg on days -8,-7), fludarabine (30mg/m2/day on days - to -2), low dose CY (14.5 mg/kg on days -4, -5 ) and total body irradiation (200cGy day -1). After the unmanipulated marrow graft was infused on day 0, cyclophosphamide 50mg/kg/day IV on days +3,+4 post-transplant was administered. All patients received tacrolimus and mycophenolate mofetil (MMF) beginning day+5 through 35 (MMF) and day +5 through 1 year (tacrolimus) for additional graft versus host disease (GVHD) prophylaxis. Neutrophil engraftment was defined as an ANC >1.0 x109/L measured for three consecutive measurements on different days. Red cell engraftment was defined as days from last red blood cell transfusion. Platelet engraftment was defined as a platelet count >50,000 for 7 days without transfusion.Ten patients were treated according to this IRB approved protocol. Eight patients had documented acquired disease and received grafts from 5/10 related donors. Six had failed ATG-containing regimens previously and two relapsed after high dose cyclophosphamide as IST. Two patients had inherited syndromes: one with Diamond Blackfan who received a graft from 5/10 related donor and one with telomeres less than the first percentile in length with a presumed familial syndrome who received a 9/10 unrelated graft. Median follow-up time is 17.5 months (range 5-49). The median age was 34 years (range 17-54) with 6 patients greater than age 30 years, and 50% were males. Median time to neutrophil engraftment was 18 days (range 16-24). Median time to red cell engraftment was 25 days (range 16-48). Median time to platelet engraftment was 27.5 days (range 22-108). At the time of BMT, six patients had PNH clones, and all were eliminated. All patients are alive and well, fully engrafted with 100% donor chimerism in blood and marrow. Two patients had grade 1-2 skin acute GVHD. These two also had mild chronic GVHD of the skin/mouth requiring systemic steroids. One patient was able to come off all immunosuppression by 15 months and the other by 17 months.Nonmyeloablative alloBMT with PTCy appears promising in patients with refractory acquired and inherited SAA with acceptable rates of engraftment, eradication of pre-existing clonal diseases, and expansion of the donor pool. Given these promising results, this approach is being utilized in the next BMTCTN national trial for acquired refractory SAA. We believe this approach could also be considered as upfront therapy for SAA, and such a trial is in development at Hopkins DisclosuresBrodsky:Alexion Pharmaceuticals: Membership on an entity's Board of Directors or advisory committees.

The Thrombopoietin Receptor Agonist Eltrombopag Has DNA Repair Activity in Human Hematopoietic Stem and Progenitor Cells

Recent studies have uncovered a specific function of thrombopoietin (TPO) in the regulation of hematopoietic stem/progenitor cell (HSPC) DNA damage response. Eltrombopag, an oral non-immunogenic TPO receptor agonist, has recently received FDA approval for the treatment of patients with refractory severe aplastic anemia, but its mode of action is incompletely understood and a role in HSPC DNA repair has not been investigated. G-CSF mobilized human CD34+ cells from 5 independent healthy donors were cultured in the presence of SCF and Flt3-L (SF), SF and TPO (SFT), or SF and Eltrombopag (SFE) for 24 hours before exposure to 2Gy γ-irradiation, and then cultured for an additional 5 to 24 hours. DNA damage was quantified by flow cytometric determination of γH2AX expression, a marker of irradiation-induced DNA double-strand breaks (DSB), and CD34+ cell survival was measured by flow cytometry using Annexin V and a viability dye. There were significantly fewer γH2AX+ cells 5 hours post-irradiation when the culture included TPO or Eltrombopag than with SF alone (Figure A, n=5). Five hours post-irradiation, cultures containing TPO or Eltrombopag had significantly increased percentages of live cells (Figure B, n=5), as well as decreased percentages of cells undergoing apoptosis compared to cultures with SF alone (SFT 12.6 ± 0.5% p=0.003; SFE 12.4 ± 2.1% p=0.012; SF 21.5 ± 3.7%, n=5). RT-qPCR arrays performed at 5 hours after irradiation on CD34+ cells cultured as above with SFT or SFE showed a significant decrease (p≤0.05) of at least two-fold in several pro-apoptotic or cell cycle arrest genes (BBC3, CCNO, GADD45G, PPM1D) compared to CD34+ cells cultured with SF alone. Twenty-four hours post-irradiation, cells cultured with TPO or Eltrombopag had significantly increased percentages of live cells (Figure B, n=3), and decreased percentages of dead cells compared to cells cultured with SF alone (SFT 9.75 ± 1.0% p=0.013; SFE 16.3 ± 0.6% p=0.032; SF 36.5 ± 6.2%, n=3). Progenitor cell survival was assessed using the CFU assay. The number of colony-forming cells was 5.9 (± 0.4) and 3.6 (± 0.2) fold higher when cultured with TPO or Eltrombopag, respectively, before γ-irradiation than when cultured with SF alone (p=0.005 and 0.006, respectively, n=2). Survival of long-term repopulating HSCs was assessed by quantifying human CD45+ cell engraftment at least 2 months after intravenous injection of NSG mice with irradiated human CD34+CD38- cells pre-cultured for 24 hours with SF, SFT or SFE. Engraftment of cells cultured with TPO or Eltrombopag was significantly higher than engraftment obtained after injection of cells cultured with SF alone before γ-irradiation (Figure C). We conclude that, analogous to TPO, Eltrombopag favors DNA DSB repair and, consequently, survival of both hematopoietic stem and progenitor cells after γ-irradiation. These pre-clinical data suggest that Eltrombopag may be of benefit in the treatment of patients with Fanconi Anemia (FA), an inherited bone marrow failure syndrome in which patients have increased susceptibility to DNA damage due to defects in the FA DNA repair pathway. [Display omitted] DisclosuresNo relevant conflicts of interest to declare.

Eltrombopag Added to Standard Immunosuppression for Aplastic Anemia Accelerates Count Recovery and Increases Response Rates

Immunosuppressive treatment (IST) with horse antithymocyte globulin (hATG) and cyclosporine (CsA) for severe aplastic anemia (SAA) has a 60-65% rate of overall hematologic response (OR) and approximately 10% complete response (CR). These rates have remained stable over the last three decades despite efforts to improve upon this regimen (Scheinberg Blood 2012). Eltrombopag (EPAG), an oral thrombopoietin receptor agonist has activity in refractory SAA producing hematologic responses, the majority multilineage, in about 40% of patients when administered as a single agent (Olnes NEJM 2012; Desmond Blood 2014). We conducted an investigator-initiated, phase II, single-center trial to test the efficacy of adding EPAG to h-ATG/CsA (clinicaltrials.gov, NCT01623167), using a triple cohort design to optimize the regimen.Eighty-eight patients with treatment-naive SAA have been enrolled from July 2012 to October 2015. All received standard h-ATG and CsA. EPAG was administered at 150 mg daily to three consecutive cohorts: in the first (n=30) and second (n=31) cohorts, EPAG was introduced after hATG, from day 14 until 6 months (cohort 1) or 3 months (cohort 2), due to initial concern of hepatotoxicity. In the third cohort, EPAG was concurrent with hATG on day 1 and continued for 6 months (n=27). As analysis of our previous IST trials suggested that CR correlates with better survival and less evolution to MDS/AML, the primary endpoint in the current protocol was CR at 6 months.Baseline enrollment characteristics were similar across cohorts: median age was 31 years (range 3 – 82; 20% were <18 years old), 26% had very severe disease (ANC<200/uL), and 34% had PNH clones >1%. EPAG was well tolerated when combined with CsA and hATG; 2 patients discontinued EPAG for severe cutaneous reactions, and 6 patients withdrew before 6 months due to refractoriness (n=4) or evolution to MDS (n=2). OR and CR were higher at 3 months and 6 months for all cohorts compared to historic rates (Table): OR at 6 months was 80%, 87% and 92%, respectively for the three cohorts; CR at 6 months was 33%, 26%, and 54%, respectively. For all patients evaluable to date, OR at 3 and 6 months of 80% and 85% and CR rates of 28 and 34% were higher when compared to our historical rates (p<0.001 for both 3 and 6 months).Improvements in blood counts among partial responders were robust by 3 months in all cohorts: median ANC 1790/ul; hemoglobin 10 gm/dL; and platelets 60,500/ul. For patients with very severe neutropenia (ANC<200/ul) the median time to ANC >500/uL was 47 days for all cohorts (n=23), and 35 days for cohort 3 (n=7). Among responders, median time to transfusion-independence for platelets was 32 days and for red cells 42 days. Serial BM biopsies showed improved cellularity in 80% without increased fibrosis. Median increase from baseline in BM CD34+ cells, as measured by flow cytometry, was 17-fold and 4-fold at 3 months for cohorts 1 and 2 respectively (p<0.001). Pharmacokinetics at 3 months showed a trend toward higher drug bioavailability with older age and female sex. Previously established baseline predictors for response such as telomere length, reticulocyte count, lymphocyte count, paroxysmal nocturnal hemoglobinuria, were not predictive of outcome. Twelve patients underwent hematopoietic stem cell transplantation (HSCT) due to relapse (n=3), refractoriness (n=6) or evolution to MDS (n=3). Death occurred in 3 subjects.At median follow up 15 months (range 1 – 42 m), cytogenetic abnormalities have occurred in 7 patients: 4 monosomy or partial deletion of chromosome 7, 1 complex (t(3;3)(q21;q26), -7), 1 deletion 13q that later normalized, and 1 subject with trisomy 6 and trisomy 15 in 2 metaphases. We sequenced 54 candidate genes, recurrently mutated in MDS/AML: somatic mutations were not identified in 4 of 7 subjects prior to treatment or at the time of cytogenetic evolution. Clonal evolution to MDS occurred at a similar frequency compared to our historic experience with standard IST.Addition of EPAG to IST markedly increases overall and complete hematologic response rates in treatment-naive SAA. These results suggest that immediate institution of EPAG with IST may salvage and expand residual hematopoietic stem cells in aplastic anemia, accelerating the rate and quality of hematopoietic recovery. Rapid blood count improvement and reduced transfusion burden support early use of EPAG for patients with newly diagnosed severe aplastic anemia. [Display omitted] Disclosures:Townsley:Novartis: Research Funding ; GSK: Research Funding. Off Label Use: Eltrombopag (Promacta) is not FDA approved for treatment naive aplastic anemia. Dumitriu:Novartis: Research Funding ; GSK: Research Funding. Winkler:GSK: Research Funding ; Novartis: Research Funding. Larochelle:Novartis: Research Funding ; GSK: Research Funding. Dunbar:Novartis: Research Funding ; GSK: Research Funding. Young:Novartis: Research Funding ; GSK: Research Funding.