Interferon-complement loop in transplant-associated thrombotic microangiopathy

Abstract

Transplant-associated thrombotic microangiopathy (TA-TMA) is an important cause of morbidity and mortality after hematopoietic stem cell transplantation (HSCT). The complement inhibitor eculizumab improves TA-TMA, but not all patients respond to therapy, prompting a search for additional targetable pathways of endothelial injury. TA-TMA is relatively common after HSCT and can serve as a model to study mechanisms of tissue injury in other thrombotic microangiopathies. In this work, we performed transcriptome analyses of peripheral blood mononuclear cells collected before HSCT, at onset of TA-TMA, and after resolution of TA-TMA in children with and without TA-TMA after HSCT. We observed significant upregulation of the classical, alternative, and lectin complement pathways during active TA-TMA. Essentially all upregulated genes and pathways returned to baseline expression levels at resolution of TA-TMA after eculizumab therapy, supporting the clinical practice of discontinuing complement blockade after resolution of TA-TMA. Further analysis of the global transcriptional regulatory network showed a notable interferon signature associated with TA-TMA with increased STAT1 and STAT2 signaling that resolved after complement blockade. In summary, we observed activation of multiple complement pathways in TA-TMA, in contrast to atypical hemolytic uremic syndrome (aHUS), where complement activation occurs largely via the alternative pathway. Our data also suggest a key relationship between increased interferon signaling, complement activation, and TA-TMA. We propose a model of an "interferon-complement loop" that can perpetuate endothelial injury and thrombotic microangiopathy. These findings open opportunities to study novel complement blockers and combined anti-complement and anti-interferon therapies in patients with TA-TMA and other microangiopathies like aHUS and lupus-associated TMAs.

Graphical abstract

Figure 1.

Gene expression changes between clinical time points of active TA-TMA and TA-TMA resolution in 4 autologous HSCT recipients with TA-TMA. The figure shows a section of the heatmap that illustrates the change in complement and interferon gene expression profiles in patients with TA-TMA. “TMA diagnosis” indicates gene expression changes from pretransplant baseline compared with when TA-TMA was diagnosed posttransplant, but before initiating complement-blocking therapy with eculizumab. “TMA resolution” indicates gene expression changes from when TA-TMA was diagnosed compared with clinical resolution of TA-TMA, after completing eculizumab therapy. Genes shown are upregulated at the time of TA-TMA diagnosis (FDR < 0.1) and belong to at least of 1 of the enriched pathway or gene lists shown in Table 1. Green bars on the right side indicate statistically significant changes at the time of TA-TMA resolution (FDR < 0.1). Data indicate that complement pathways are highly upregulated at TA-TMA diagnosis in subjects with TA-TMA and normalize after resolution of TA-TMA.

Figure 2.

Complement gene expression in 4 patients with TA-TMA at time of clinical diagnosis. This figure illustrates gene expression in alternative, classical, and lectin complement pathways among cases with TA-TMA at the time of clinical diagnosis but before initiation of complement-blocking therapy with eculizumab. Red, orange, and yellow colors indicate upregulated pathways and the degree of upregulation (red > orange > yellow). Blue colors indicate downregulated and the degree of downregulation (dark blue > light blue). Red letters and red outlines indicate statistically significant changes in gene expression from pretransplant baseline to the time of TA-TMA clinical diagnosis.

Figure 3.

Changes in interferon gene expression between clinical time points in 4 cases with TA-TMA and 3 controls without TA-TMA after autologous HSCT. The figure illustrates changes in interferon gene expression profiles in autologous HSCT recipients with TA-TMA and without TA-TMA. In cases with TA-TMA, “TMA diagnosis” indicates gene expression changes at time of clinical diagnosis of TA-TMA compared with pretransplant baseline, but before the start of complement-blocking therapy with eculizumab. “TMA resolution” illustrates gene expression change at the time of TA-TMA clinical diagnosis compared with TA-TMA resolution, after completing eculizumab therapy. In controls without TA-TMA, this evaluation was performed at matched timepoints after transplant. Genes shown were upregulated in the “TMA diagnosis” comparison for patients with or without TA-TMA and are members of interferon-related gene sets (Hallmark interferon-γ response and hallmark interferon-α response) and transcription factor target gene lists (STAT1 and STAT2 targets K562 IFN-α 30 minutes, STAT1 and STAT2 targets K562 IFN-α 6 hours). Green bars indicate statistically significant differences between time points. Data indicate that interferon pathways are highly upregulated at clinical diagnosis of TA-TMA in cases with TA-TMA and normalize after therapy with eculizumab. Interferon pathways are not upregulated in controls without TA-TMA at matched time points after transplant.

Figure 4.

Interferon-complement loop. This figure displays the relationship between interferon and complement activation that perpetuates vascular endothelial injury in clinical conditions presenting with thrombotic microangiopathy where both interferon and complement pathways are activated. CFP, CFD, CFB, CFH: complement factors P, D, B and H.