Treatment of severe aplastic anaemia with combined immunosuppression: anti-thymocyte globulin, ciclosporin and mycophenolate mofetil

Severe aplastic anaemia (SAA) can be successfully treated with immunosuppressive therapies or haematopoietic stem cell transplantation (HSCT). Response rates with horse anti-thymocyte globulin (h-ATG) plus ciclosporin (CsA) are about 60-70%, and robust responders have an excellent long-term survival. We introduced a third immunosuppressive agent to standard h-ATG/CsA, mycophenolate mofetil (MMF), in an attempt to improve the response rate and survival, and to decrease the relapse rate and clonal evolution to myelodysplasia. A total of 104 consecutive patients with SAA were treated with h-ATG/CsA/MMF between May 1999 and June 2003 at the National Institutes of Health Clinical Center. The overall response rate at 6 months was 62%, with 24 (37% of responders) patients relapsing at a median of 389 d from ATG. Nine patients showed evidence of clonal evolution following ATG. After a median follow up of 42 months, the median survival among responders was not reached and among non-responders was 58 months. Over half of the relapses occurred during MMF administration. Despite a strong theoretical rationale for its use, MMF did not result in the improvement of response or relapse rates when compared with historical standard h-ATG/CsA.     

Efficacy of rabbit anti-thymocyte globulin in severe aplastic anemia

Background

A combination of horse anti-thymocyte globulin and cyclosporine produces responses in 60–70% of patients with severe aplastic anemia. We performed a phase II study of rabbit anti-thymocyte globulin and cyclosporine as first-line therapy for severe aplastic anemia.

Design and Methods

Twenty patients with severe aplastic anemia treated with rabbit anti-thymocyte globulin were compared to 67 historical control cases with matched clinical characteristics treated with horse anti-thymocyte globulin.

Results

Response rates at 3, 6 and 12 months were similar for patients treated with rabbit anti-thymocyte globulin or horse anti-thymocyte globulin: 40% versus 55% (P=0.43), 45% versus 58% (P=0.44) and 50% versus 58% (P=0.61), respectively. No differences in early mortality rates or overall survival were observed. We then performed multivariable analyses of response at 6 months and overall survival and identified the presence of a paroxysmal nocturnal hemoglobinuria clone (P=0.01) and a pretreatment absolute reticulocyte count greater than 30×10⁹/L (P=0.007) as independent predictors of response and younger age (P=0.003), higher pretreatment absolute neutrophil (P=0.02) and absolute lymphocyte counts (P=0.03) as independent predictors of overall survival. None of the immunogenetic polymorphisms studied was predictive of response to immunosupressive therapy.

Conclusions

Despite reports suggesting differences in biological activity of different anti-thymocyte globulin preparations, rabbit and horse anti-thymocyte globulin appear to have a similar efficacy for up-front treatment of severe aplastic anemia. Clinicaltrial.gov: {"type":"clinical-trial","attrs":{"text":"NCT01231841","term_id":"NCT01231841"}}NCT01231841)     

A third course of anti-thymocyte globulin in aplastic anaemia is only beneficial in previous responders

This retrospective study evaluated the outcome of 18 patients with aplastic anaemia treated with a third course of anti-thymocyte globulin (ATG)-containing immunosuppressive therapy (IST). Of the 18 patients, seven had responded to one of the previous courses of ATG and 11 were refractory to both the previous courses. Self-limiting grade >/=3 liver toxicity was observed in three patients. Two patients had to discontinue ATG because of severe systemic side effects. The incidence and manifestations of serum sickness did not appear to be different during the three courses. All of the seven patients that previously responded to one of the courses responded to a third course. In contrast, of 11 patients refractory to the previous courses, only two had a transient partial response. The 3-yr event-free survival for the patients who had responded to one of the previous courses of ATG was significantly superior to that of patients refractory to both the previous courses of ATG (83% vs. 0%, P = 0.0001). For aplastic anaemia patients, a third course of ATG-containing IST is a reasonable option in previous responders. Patients refractory to previous two courses of ATG have a much lower response rate and may be suitable candidates for novel therapeutic options.     

Mechanisms of bone marrow progenitor cell apoptosis in aplastic anaemia and the effect of anti-thymocyte globulin: examination of the role of the Fas-Fas-L interaction

The mechanism of action of anti-thymocyte globulin (ATG) in aplastic anaemia (AA) is complex. Bone marrow (BM) CD34(+) cells in AA have been shown to be more apoptotic and have a higher expression of Fas antigen (Fas-ag) than in normal donors. The aims of this study were to delineate further the mechanism for increased bone marrow progenitor cell apoptosis in AA and investigate the effects of ATG on apoptosis and Fas-ag expression. BM was obtained from six normal donors and 10 untreated AA patients. We confirmed that AA BM CD34(+) cells were more apoptotic than normal donor cells (P = 0.002). Following treatment with ATG, the mean percentage reduction of apoptosis was 34% (9.2-65.9%). BM from 30 AA and 10 normal donors was then stained for CD34, Fas-ag and 7-AminoActinomycin D. The proportion of CD34(+) Fas(+) cells was higher in untreated AA (P = 0.0001) than in normal donors. Results also showed that the majority of CD34(+) Fas(+) cells were apoptotic/dead in normal donors (mean 81%) and AA (88%), indicating that Fas is involved in apoptosis of CD34(+) cells. In contrast, the majority of CD34(+) Fas(-) cells in normal donors were live (mean 91%), while two patterns emerged in untreated AA. In seven patients, the majority of cells were live, however, in the remaining eight patients, the majority of cells were apoptotic/dead, suggesting an alternative mechanism for apoptosis in addition to Fas-ag. Finally, we have shown that in vivo ATG treatment reduced the expression of Fas-ag on AA BM CD34(+) cells.     

Prospective randomized trial comparing two doses of rabbit anti-thymocyte globulin in patients with severe aplastic anaemia

The treatment of choice for patients with severe aplastic anaemia (SAA) includes immunosuppressive therapy (IST) with anti-thymocyte globulin (ATG) and ciclosporin A. However, the optimal dose for rabbit ATG has yet to be established. We herein report the first prospective, randomized, multicentre study comparing two doses of rabbit ATG in patients with SAA. Patients with SAA who required initial IST in Japan (n = 89), China (n = 85) and Korea (n = 48) were enrolled between May 2012 and October 2017. A 1:1 block randomization was employed for two doses of rabbit ATG. In total, 222 patients were randomized, with 112 patients receiving 2·5 mg/kg and 110 receiving 3·5 mg/kg of rabbit ATG for 5 days. The primary endpoint was the haematological response at day 180. After 6 months, no significant difference in response rates was observed between the 2·5 and 3·5 mg/kg groups (49% vs. 48%, P = 0·894). Overall survival at 3 years was similar between the two groups [85% (95% confidence interval [CI], 76%−91%) vs. 91% (95% CI, 82%−96%); P = 0·107]. The current study revealed no significant differences in the efficacy and safety between the 2·5 and 3·5 mg/kg doses of rabbit ATG in patients with SAA. Trial registration: UMIN000011134.     

Repeated treatment with horse antilymphocyte globulin for severe aplastic anaemia

In a single-centre study the feasibility and efficacy of repeated antilymphocyte globulin (ALG) for patients with severe aplastic anaemia (SAA) not responding to an initial ALG treatment or relapsing after initial response to ALG was evaluated. 139 consecutive patients with newly diagnosed SAA were treated with ALG between 1976 and 1995. 89 patients responded to a first course; 50 patients did not become transfusion independent. Of the 89 responders, 66 remained in remission, 23 relapsed. 43 patients received a second or subsequent course of ALG for failure to respond ( n  = 25) or relapse ( n  = 18) and were given a total of 53 courses. Acute reactions in the multiply exposed patients occurred during the first ALG treatment in 11 (26%) and during subsequent exposures in 16/53 courses (30%; P  > 0.2). Incidence of serum sickness was 63% (27/43) after the initial course compared to 57% (30/53) after subsequent courses ( P  > 0.2), but clinical signs of serum sickness occurred earlier after repeated (median 6 d) as compared to initial exposure (13 d; P  = 0.008). Transfusion-independent haemopoiesis was achieved in 27/43 (63%) and survival probabilities for the 43 patients receiving multiple courses of ALG was 52 ± 8% at 10 years. The probability of developing a late clonal disorder was 53 ± 10% after multiple, as compared to 34 ± 7% after single exposure ( P  = 0.15). No difference in results was observed between patients retreated for failure to first ALG or for relapse. ALG of the same species can be repeated without increased risks of side-effects in patients with SAA. A second or subsequent course of ALG from the same source can be effective when the first course has failed.     

Restoration of immunity and reactivation of hepatitis B virus after immunosuppressive therapy in a patient with severe aplastic anaemia

We recently treated a patient with severe aplastic anaemia (SAA) who also had chronic hepatitis B virus (HBV) infection. The HBV serological status at the time of diagnosis of SAA was HBsAg(+) and HBeAg(+). Subsequent analysis of the precore region of HBV DNA showed wild-type. He received anti-thymocyte globulin (ATG) and cyclosporin A (CsA) therapy twice. After each course of ATG infusion and during CsA therapy he developed lymphopenia for 1 and 2.5 months, respectively. His serum alanine aminotransferase (ALT) became normalized during the period of lymphopenia, but the serum HBV viral load increased. When his peripheral lymphocytes count recovered, his ALT became elevated again. Lamivudine was effective to normalize his elevated ALT and suppress viral replication. The phenomenon observed in this case supports the prevailing notion that hepatitis B flare-up in HBV carriers after chemotherapy is caused by an immune-mediated mechanism. Meanwhile, this is the first documented case of SAA who developed HBV reactivation upon recovery of lymphopenia after immunosuppressive therapy. This also highlights the necessity of pre-emptive therapy with lamivudine in SAA/HBsAg(+) patients to receive immunosuppressive therapy with ATG/CsA.     

Effect of androgens on the response to antithymocyte globulin in patients with aplastic anaemia

30 patients with aplastic anaemia (18/30 with severe aplastic anaemia) were prospectively randomized to be treated with 100 mg/kg ATG with or without the oral androgen Methenolone (3 mg/kg). 15 of 30 patients responded. Among the 15 patients receiving ATG plus androgen, 11 patients (73%) responded, including 8 complete and 3 partial responses. 4 of the 15 patients (31%) receiving ATG only responded, including 2 complete and 2 partial responses. The difference in response rate was statistically significant (p = 0.01). The survival rate in the total population of 30 patients was 64%. The survival rate in the group receiving ATG plus androgen was 87%; in the group receiving ATG only it was 43%. The difference in survival rates between both groups did not reach statistical significance (p = 0.15). Toxicity of ATG and androgens was considerable but manageable. These data support the result of the recent European reevaluation of a large pool of patients by the EBMT (39), that androgens in addition to ATG increase survival in patients with aplastic anaemia. They are, however, in contradiction to a controlled American study showing no benefit of a combined treatment with androgens as compared to ATG only. Further controlled studies on a larger number of patients are indicated to determine the therapeutic efficacy of androgens in addition to immunosuppression in aplastic anaemia.     

The incidence and significance of fevers during treatment with antithymocyte globulin for aplastic anaemia

Antithymocyte globulin (ATG) is a foreign protein used widely to treat aplastic anaemia (AA). Febrile reactions occurring during its administration may be impossible to distinguish clinically from fever due to sepsis, and are therefore routinely treated with intravenous antibiotics after collection of blood cultures. A statistically highly significant difference was found in positive blood cultures between 39 AA patients who developed fever during ATG therapy, and 38 febrile neutropenic patients with acute leukaemia, suggesting that most fevers developing during ATG treatment are not due to infection. It may therefore be reasonable to consider early discontinuation of intravenous antibiotics in patients who are clinically stable and have no proven sepsis.     

Hepatitis‐associated aplastic anaemia treated successfully with antilymphocyte globulin

Aplastic anaemia may occur following an acute attack of hepatitis. This is a rare condition, which if not recognized and promptly treated, may be fatal. Antilymphocyte globulin and allogeneic bone marrow transplantation have been used in the treatment of this condition. We report the case of a young man who developed severe aplastic anaemia following nonviral hepatitis.     

Unrelated donor marrow transplantation in children with severe aplastic anaemia using cyclophosphamide, anti-thymocyte globulin and total body irradiation

We report a favourable outcome in 15 patients with severe aplastic anaemia (SAA) who were < 20 years of age and who underwent bone marrow transplantation (BMT) from a human leucocyte antigen (HLA)-matched unrelated donor. All patients were non-responders to intensive immunosuppressive therapy (IST) and were multiply transfused. The conditioning regimen consisted of cyclophosphamide (60 mg/kg/d, on d -4 and -3), anti-thymocyte globulin (2.5 mg/kg/d, on d -5 to -2) and total body irradiation (2.5 Gy x 2/d, on d -2 and -1). Patients received cyclosporine and methotrexate for prophylaxis of graft-versus-host disease (GVHD), except for the last four who received tacrolimus instead of cyclosporine. Donor/recipient pairs were identical for HLA class I and II antigens by serological typing, but four pairs were found to have a mismatch at the HLA-A, -B or -DRB1 locus by high-resolution typing. All patients achieved rapid engraftment and are alive at 2-86 months after transplantation (median follow-up, 51 months). Moderate to severe acute GVHD occurred in 5 out of 15 patients (33%); only one patient developed extensive chronic GVHD. Considering our encouraging results, unrelated donor transplantation for SAA is recommended as a salvage therapy in non-responders to IST.     

Eltrombopag in the management of aplastic anaemia: real-world experience in a non-trial setting

Objective: The thrombopoietin mimetic eltrombopag has been used in clinical trials for the frontline and salvage treatment of aplastic anaemia (AA). Eltrombopag was investigated in AA patients on a non-trial all-comer basis.

Methods: Consecutive newly diagnosed and relapsed/refractory AA patients were treated with eltrombopag.

Results: In a 4.5-year period, 20 consecutive AA patients (newly diagnosed, N?=?10; relapsed/refractory, N?=?10) at a median age of 47 (22–84) years were treated with eltrombopag. For newly diagnosed patients, the frontline use of eltrombopag (concomitant medications: anti-thymocyte globulin, ATG, and ciclosporin, N?=?4; ciclosporin, N?=?5; nil, N?=?1) at a median maximum dose of 150 (50–300)?mg/day led to an overall response rate (ORR) of 90% (trilineage: 60%; neutrophil: 20%; platelet: 10%). After a median follow-up of 47 (14–179) weeks, responses were maintained in all cases. In relapsed/refractory patients, eltrombopag at a median maximum dose of 150 (50–300)?mg/day led to an ORR of 50% (trilineage: 40%; neutrophil: 10%), with responses maintained after a median follow-up of 115 (53–253) weeks. Adverse effects included reversible skin pigmentation (observed in all patients taking eltrombopag at ≥150?mg/day), dyspepsia, and liver function derangement.

Conclusion: In a routine haematological practice, the use of eltrombopag in AA patients was feasible, safe, and associated with very favourable responses.     

Outcomes of older patients (≥60 years) with acquired aplastic anaemia treated with immunosuppressive therapy

We report a single center experience in treating 24 consecutive patients ≥60 years old for aplastic anaemia (AA) with immunosuppressive therapy (IST). The main outcomes of interest were the tolerability and toxicity of IST, response rate and survival. Median age was 70 years (range 61–78). Seven patients received standard IST consisting of standard dose antithymocyte‐globulin (ATG) with or without ciclosporin (CsA), and 17 patients received attenuated IST consisting of at least a 50% dose reduction of ATG with CsA or CsA alone. Six patients (25%) had early deaths, mostly due to infection. Early mortality appeared higher in the standard IST group although this was not statistically significant (43% vs. 18%, P = 0·4). The 2‐year cumulative incidence of response was 42% [95% confidence interval (CI) 26–69%]. Responders had significantly better survival than non‐responders (P = 0·0002). The 3‐year probability of overall survival was 49% (95% CI 27–68%). Moderate or severe co‐morbidities (P = 0·03) as determined by Charlson Co‐morbidity Index and very severe AA (P = 0·007) were associated with significantly inferior overall survival while age was not. Nine out of 14 evaluable patients in the attenuated IST group had durable responses to treatment, suggesting that patients not suitable for standard IST can still benefit from attenuated IST.     

Pregnancy and severe aplastic anaemia: causal relation or coincidence?

The relationship between aplastic anaemia (AA) and pregnancy remains uncertain. To assess whether an association between pregnancy and severe aplastic anaemia (SAA) exists, we compared the frequency of pregnancy in 35 young women with newly diagnosed SAA with the expected frequency in the general population. The observed pregnancy rate in the SAA group was 3–6%. This percentage approximates the expected pregnancy rate of 4.4% in the general population and is not compatible with a strong association between pregnancy and SAA.     

Direct binding of antithymoctye globulin to haemopoietic progenitor cells in aplastic anaemia

Antithymocyte globulin (ATG) is widely used in the treatment of aplastic anaemia (AA) and a response occurs in 60-80% of patients. However, its exact mechanism of action in the treatment of AA has yet to be determined. Previously, we have shown that ATG increases colony growth from purified bone marrow CD34+ cells of AA patients in vitro, and decreases stem cell apoptosis and the expression of soluble Fas receptor after ATG therapy in vivo. The aim of this study was to further examine the association of ATG with AA haemopoietic progenitor cells. We describe here that ATG bound directly to CD34+ cells. Forty-six patients and 20 normal control subjects were studied. ATG bound to CD34+ cells in normal control subjects (mean 90.38%) as determined by flow cytometry. The mean percentage of CD34+ cells binding to ATG was 59.90% in untreated aplastic patients, 83.24% in partial responders, 58.3% in non-responders and 62.73% in relapsed patients. In completely recovered patients, ATG binding was indistinguishable from control subjects. The functionality of AA patients' haemopoietic progenitor cells was assessed using colony assays. These results demonstrate the direct binding of ATG to CD34+ cells and suggest that differences in its binding to AA CD34+ cells could reflect functional differences in the haemopoietic stem cell compartment throughout the disease process.     

Epstein-Barr virus reactivation in a patient treated with anti-thymocyte globulin for severe aplastic anemia

Epstein-Barr virus (EBV) infection and reactivation is an increasing complication in immune deficient patients, particularly after allogeneic hematopoietic stem cell transplantation (HSCT). Therapy with anti-thymocyte globulin (ATG) is associated with higher incidence of EBV-related disease in HSCT patients, but this risk is not documented in patients receiving ATG for severe aplastic anemia (SAA). We describe the case of a patient who developed an EBV infection, with the clinical features of an infectious mononucleosis, after immune suppression with cyclosporine and two courses of ATG for SAA.