New insights into risk factors for transplant-associated thrombotic microangiopathy in pediatric HSCT

Introduction

Transplant-associated thrombotic microangiopathy (TA-TMA) represents a diagnostic challenge for clinicians and can lead to multiorgan failure and death.^1,2 There is a wide variation in the prevalence of TA-TMA across different reports ranging between 0.5% and 76%^3-8 with a mortality rate reaching as high as 80%.^7,9,10 This variation probably reflects the lack of consistency in the diagnostic criteria for TA-TMA. A previous review in 2004 pointed to the presence of 38 different diagnostic criteria being used in 35 studies, including 447 patients with TA-TMA.⁶ Moreover, the pathophysiology of TA-TMA is poorly understood, and histopathological features overlap with other small vessel injury diseases.¹¹ Recent studies have focused on genetic predisposition and abnormal activation of the complement cascade to achieve a more refined definition of this condition.^12-15 Different risk factors have been associated with a higher risk of developing TA-TMA¹⁶: calcineurin inhibitors (eg, tacrolimus) and sirolimus have been blamed for increasing the risk of endothelial damage,^7,17,18 although the beneficial effect of early discontinuation of these agents remains to be proven¹⁶ and must be balanced with the risk of inducing or worsening graft-versus-host disease (GVHD). Screening of patients for risk factors of TA-TMA before transplant has been problematic, and close monitoring for early signs of microangiopathy is necessary to recognize this complication before severe manifestations occur¹ since early intervention can provide a better outcome.^19,20 Recently reviewed criteria for diagnosis of TA-TMA have included clinical and laboratory findings¹ and should help clinicians to identify patients with early-stage TA-TMA and those at a higher risk of a poor outcome. Recognition of pretransplant variables that could identify high-risk patients might permit prophylaxis or even tailoring of the transplant procedure (eg, conditioning, GVHD prophylaxis). Risk factors have been variably reported in adults,²¹ and data on children remain scarce. We aimed to identify a risk profile for the development of TA-TMA in children undergoing hematopoietic stem cell transplantation (HSCT) and to explore potential correlation with other posttransplant complications that could improve our comprehension of TA-TMA.

Methods

Results

Discussion

This is the first report to our knowledge that has identified comorbidity as a potential risk profile for a pediatric patient’s post-HSCT. Besides that, other risk factors, including aGVHD and a second transplant, were shown to be significant driving risk factors for TA-TMA. The identification of risk factors for the occurrence of TMA will conceivably allow early diagnosis and, perhaps, the timely introduction of therapeutic or preventive measurements.

Since the earliest report of TA-TMA in 1980,³⁰ reports since 2004 showed a variable incidence of TA-TMA between 4% and 35%.^16,31-36 Our report showed an incidence between 5% and 6%. Both centers in the United Kingdom reported similar incidences in the pediatric cohort. This consistency reflects the use of a uniform criterion to define TA-TMA and gives more insight into the true incidence of TA-TMA. Moreover, other studies that used similar criteria³⁷ to define TA-TMA reported an incidence of between 7% and 8% among adults.^16,35 To identify patients who developed TA-TMA, Uderzo et al²¹ used 6 laboratory criteria, including raised LDH >600 IU/L, red cell fragments >2%, unexplained drop of hemoglobin, and a drop in platelets <20 × 10⁹/L after engraftment, elevated TA-TMA index (LDH/platelet) together with 3 clinical criteria: renal dysfunction with raised creatinine, central nervous system involvement, and consistent DIC, and reported an incidence of TA-TMA of 11% among adults and 13% among children. This report confirms the fact that use of a uniform criterion shows a more consistent incidence of TA-TMA.

Among the patient-derived risk factors, sex, and underlying disease did not influence the risk for TA-TMA, similar to data reported from previous studies.^16,35,38 On the contrary, others reported increased risk of TA-TMA among females more than males, whereas underlying disease had no impact.^21,39 In our cohort of patients, active comorbidity stood as a significant factor associated with TA-TMA. Although the pathophysiology of TA-TMA is not clearly understood, most studies reported endothelial injury to be the first and initiating step in the development of TA-TMA. Activation of endothelial cells to produce a procoagulant state leads to the activation of antigen-presenting immune cells and lymphocytes with production of multiple cytokines driving complement activation ending in microthrombus formation.⁴⁰ Here, we defined active comorbidity using factors that are likely inducers of endothelial damage, including uncontrolled infections, the use of steroids to suppress active inflammation, cardiovascular instability, and pulmonary compromise.^41-44

Reports from adults using hematopoietic cell transplantation-specific comorbidity index score to define comorbidity, however, failed to demonstrate a link between comorbidity and TA-TMA.¹⁶ However, this finding needs to be taken with caution in the pediatric population, where risk factors such as obesity, peptic ulcer, depression/anxiety, and coronary artery disease are seldom seen. Furthermore, the authors do not detail the risk profile for each patient. A current study in GOSH (N.B., R.E., P.A., G.L., R.C., P.V., and K.R., manuscript in preparation) demonstrated the failure of hematopoietic cell transplantation-specific comorbidity index score to be valid among 950 children who received a HSCT from 2006 to 2016.

Within transplant-related factors, factors that can potentially induce endothelial damage were analyzed. In our cohort, intensity of the conditioning regimen and use of TBI did not influence the development of TA-TMA with a risk between 5% and 6% with myeloablative, nonmyeloablative-conditioning regimen, or TBI-based conditioning. Nakamae et al⁴⁵ showed in multivariate analysis that the use of Bu-based MAC conditioning was an independent risk factor for TA-TMA. Interestingly, in the same report, the risk of TA-TMA risk was 13% among all MAC and was not lowered (25%) among patients who received nonmyeloablative protocols: 77% being Flu-based conditioning. Eissner et al⁴⁶ reported that Flu induces apoptosis, activation, alloreactivity of human endothelial cells and causes damage to dermal and alveolar epithelial cells. Flu is a major constituent of non-MAC, and the effect of Flu on endothelial cells was thought to contribute to the pathogenesis of TA-TMA after non-MAC.^31,47 However, it was clear in the Elliott et al⁴⁷ report that the patients who developed TA-TMA after Flu-based conditioning had undergone prior autologous transplantation and therefore were exposed to 2 transplant procedures. A subsequent conditioned HSCT was identified in our study cohort as a risk factor for TA-TMA with a risk factor of 15% vs 5% for patients who received only 1 graft. This is similar to previous studies in adults with a risk ranging between 12%^16,35 and 52%³⁰ with subsequent transplant vs an average of 7% in patients receiving only 1 graft.¹⁶ The effect of TBI has been controversial with some reports showing increased incidence of TA-TMA,^21,39 whereas others including ours showing no effect on the development of TA-TMA.^16,35

The use of a mismatched graft was considered a potential risk factor for TA-TMA in adults.^21,45,48,49 We reported a significantly higher incidence of TA-TMA of 17% with haploidentical HSCT vs <10% among other donor sources. Within this group, grade III to IV aGVHD and the presence of active comorbidity were the main driving risk factors for increased TA-TMA in our study.

The occurrence of aGVHD was the most important risk factor to TA-TMA in our series. These results are concordant with other studies that reported a strong link between GVHD and TA-TMA. Some considered the link as being related to the use of more immunosuppressive medications, in particular, CSA or high levels of tacrolimus, combination of CSA with sirolimus as prophylaxis, or therapy for GVHD, ^18,45,50, with existing guidelines recommending discontinuation of calcineurin inhibitors as the main intervention after initial diagnosis.^1,7,40 The mechanism appears to be partly due to decreased levels of nitric oxide and vascular endothelial growth factor.⁴⁰ In vitro studies have shown that both CSA and tacrolimus + sirolimus increased expression of intracellular adhesion molecule-1 with CSA alone having a further detrimental effect on the endothelium through increasing reactivity of extracellular matrix to platelets inducing a procoagulant status.⁵¹

However, 2 factors seem not to support this hypothesis. First, in previous reports, the withdrawal of calcineurin inhibitors failed to reverse or even to halt the process of TA-TMA.^16,21,31 Uderzo et al²¹ demonstrated clearly that the withdrawal of immunosuppressive medications did not prevent mortality, where 11 out of 14 (78%) patients who stopped immunosuppressive therapy died vs 21 out of 49 (42%) patients who continued on immunosuppression. Within another report on 22 patients with TA-TMA, 18 of them had either discontinued CSA or reduced the dose in addition to other therapeutic measures: plasma infusion and/or therapeutic plasma exchange. A response was only recorded among 5 out of 18 patients. Of note, nonrelapse mortality was 77% (14 out of 18 patients) with GVHD being the cause of death among 9 of them. A recent report by Li et al¹⁶ showed similar results where 60 patients with TA-TMA either discontinued CSA or tacrolimus or sirolimus (31 patients) or shifted from one to another (29 patients) with no evidence that withdrawal of either calcineurin inhibitors or sirolimus was beneficial, with a clear demonstration that GVHD grades III to IV were the main driving factor for TA-TMA. Second, in a pilot study carried out by Ringdén et al including 24 HSCT patients, comparing the use of tacrolimus/sirolimus to use of CSA with or without methotrexate, none of the patients developed grade III to IV aGVHD or TA-TMA. Hence, being on calcineurin inhibitors or sirolimus with the absence of high-grade GVHD, TA-TMA was not reported.⁵² In our series, when using multivariate analysis, we noted that neither the use of calcineurin inhibitors nor the use of mismatched grafts, including haplo-HSCT, drove the development of TA-TMA, with aGVHD grades II to IV and III to IV being the main risk factors for TA-TMA. A growing body of literature supports the role of endothelial dysfunction in the development of GVHD^53-56 with evidence of complement activation as a downstream effect of endothelial dysfunction.^57,58 Recent studies in adults showed a strong association between aGVHD grades III to IV and renal TA-TMA,⁵⁴ steroid refractory GVHD and TA-TMA,⁵⁸ and gastrointestinal aGVHD and TA-TMA³⁸ with dismal outcomes across most of the reports. In our series, more than half of the patients who were diagnosed with TA-TMA with concomitant aGVHD died. This is also similar to previous reports from adults where Zhang et al³⁸ reported poor survival of 52% (n = 50) among patients who developed TA-TMA vs 77% in patients who had only aGVHD (n = 83) and 86.6% among patients who neither developed aGVHD nor TA-TMA and aGVHD (n = 67) with higher nonrelapse mortality but low risk of relapse. Another group reported a drop in 90-day survival among patients with TA-TMA from 96% in patients with no documented driving factor to 50% in patients with concomitant GVHD. All these studies, including ours, demonstrate that vascular GVHD might be a specific form of TA-TMA that requires intensified treatment of GVHD to control TA-TMA rather than discontinuation of immune suppressive medications, which might worsen GVHD leading to increased mortality.

To date, current therapeutic recommendations for TA-TMA are controversial with no approved medicine by regulatory authorities. Therapeutic plasma exchange, in addition to being invasive, has shown no benefit in treating adults with TA-TMA^2,19,31 possibly due to the lack of autoantibodies against factor H or other endothelial proteins.⁴⁰ With more insight into TA-TMA and evidence of terminal complement activation, complement blockers like eculizumab represent an appealing therapeutic intervention for TA-TMA; however, a failure rate of 48% was recorded in our series, which is similar to other reports with 30% to 40% response failure.^59,60

In our studied cohort, the choice on the use of complement blockade vs defibrotide was made based on the accessibility to treatment, the analysis of complement activation, the presence of bleeding, and the center experience.

In patients treated at GOSH, P10 to P25 had evidence of complement activation as shown by elevated sC5b-C9. With access to a complement blockade, either eculizumab or Coversin was used among all patients except P24. P24 was the only patient from GOSH whose complement was not checked at the start of proteinuria. Diagnosis of TA-TMA came after renal biopsy confirming microangiopathy. On the contrary, the GNCH BMT center with easier access to defibrotide and previous experience with the use of defibrotide therapy in TA-TMA²⁹ used defibrotide in most patients who developed TA-TMA.

When there is evidence of complement activation, the use of complement blockade should be considered an option to stop the evolution of TA-TMA. However, complement activation is not the sole player for the development and evolution of TA-TMA. Defibrotide, which has broad-spectrum anti-thrombotic, anti-ischemic, and anti-inflammatory activity, may protect the endothelium from damage associated with TA-TMA. Conversely, if TA-TMA is associated with bleeding, then defibrotide may be contraindicated.

A study by IEWP and PDWP EBMT on the use of defibrotide in TA-TMA recorded a resolution rate of 77% among both children (17/22) and adult (13/17) cohorts.²⁹

A recent multicenter retrospective study on behalf of IEWP and PDWP EBMT (P.A., G.L., R.C., P.V., and K.R., submitted for publication 2020) including 29 centers, 80 allo-HSCTs, and 6 auto-HSCTs, 55 patients (68%) received eculizumab as a first-line measure (elevated sC5b-9 were checked in only 41 patients). At 6 months from the start of therapy, the cumulative incidence of TA-TMA resolution was 41%, and overall survival was 50%.

In order to assess the true benefit of defibrotide in TA-TMA patients, a prospective multicenter study is required to allow accurate comparison between defibrotide and eculizumab/Coversin to determine which patients are most likely to benefit from treatment and guide treatment dose and duration.

Given the severity of TA-TMA and the noted high rate of mortality recorded in all studies to date, this requires consideration of potential prophylactic measures to prevent the occurrence of TA-TMA. One of the most appealing prophylactic medications is defibrotide. Defibrotide is a mixture of single-stranded oligonucleotides with endothelial protection properties mediated via inhibition of tumor necrosis factor-α and its pro-fibrinolytic, antithrombotic, anti-inflammatory, and thrombolytic properties. It is US Food and Drug Administration approved for the treatment of VOD, another endothelial damage complication of HSCT.⁴⁰ The use of defibrotide the day prior to conditioning has been demonstrated to effectively provide protection against the development of VOD in high-risk patients.⁶¹ In vitro, defibrotide protects against endothelial injury induced by Flu and calcineurin inhibitors.^46,51 A trial of defibrotide prophylaxis in patients at high risk for development of TA-TMA is currently underway in the United States (NCT#03384693).

To sum, our study has demonstrated a profile for pediatric patients undergoing HSCT who are at greater risk of developing TA-TMA. We have shown that higher-grade aGVHD, multiple transplants, and comorbidity as being the main drivers for pediatric TA-TMA. Given the nonavailability of effective treatments and high nonrelapse mortality associated with TA-TMA, identification of patients at risk and considering prophylactic measures may be worthwhile. The link between GVHD and TA-TMA demonstrated in our study and other recent reports identifies a particular form of TA-TMA that might represent a vascular GVHD. One alternative approach might be to substitute calcinuerine inhibitors with other modalities of immunosuppression; however, with this approach, it is paramount that good control of GVHD is achieved or TA-TMA may be exacerbated.

Supplementary Material

The full-text version of this article contains a data supplement.

Authorship

Contribution: R.E. designed the research and wrote the manuscript; R.E. and J.S. analyzed the data; G.L., S.-H.L., A.B., Z.N., G.O., N.B., R.C., K.R., P.A., S.O., T.F., A.J.C., R.S., and A.W. were actively involved in patients' care, collected data, and helped in designing the research; W.Q., C.B., M.S., A.R.G., and P.V. checked data and worked on editing the writing of the manuscript; H.Y. and B.D., from pharmacy, wrote and analyzed data on defibrotide and eculizumab; D.K., S.G., and P.W., from the genetics laboratory, ran genetics on complement; and S.A. revised the histopathology slides.

Conflict-of-interest disclosure: The authors declare no competing financial interests.

Correspondence: Reem A. Elfeky, Great Ormond Street (GOS) Hospital for Children NHS Foundation Trust, University College London GOS Institute of Child Health, and NIHR GOSH BRC, 30 Guilford St, London WC1N 3JH, United Kingdom; e-mail: [email protected] and [email protected].