Thrombotic microangiopathies assessment: mind the complement

Abstract

When faced with microangiopathic haemolytic anaemia, thrombocytopenia and organ dysfunction, clinicians should suspect thrombotic microangiopathy (TMA). The endothelial damage that leads to this histological lesion can be triggered by several conditions or diseases, hindering an early diagnosis and aetiological treatment. However, due to systemic involvement in TMA and its low incidence, an accurate early diagnosis is often troublesome. In the last few decades, major improvements have been made in the pathophysiological knowledge of TMAs such as thrombotic thrombocytopenic purpura [TTP, caused by ADAMTS-13 (a disintegrin and metalloproteinase with a thrombospondin Type 1 motif, member 13) deficiency] and atypical haemolytic uraemic syndrome (aHUS, associated with dysregulation of the alternative complement pathway), together with enhancements in patient management due to new diagnostic tools and treatments. However, diagnosis of aHUS requires the exclusion of all the other entities that can cause TMA, delaying the introduction of terminal complement blockers, which have shown high efficacy in haemolysis control and especially in avoiding organ damage if used early. Importantly, there is increasing evidence that other forms of TMA could present overactivation of the complement system, worsening their clinical progression. This review addresses the diagnostic and therapeutic approach when there is clinical suspicion of TMA, emphasizing complement evaluation as a potential tool for the inclusive diagnosis of aHUS, as well as for the improvement of current knowledge of its pathophysiological involvement in other TMAs. The development of both new complement activation biomarkers and inhibitory treatments will probably improve the management of TMA patients in the near future, reducing response times and improving patient outcomes.

Keywords: C5b-9 deposits; complement system; endothelial cells (ECs); membrane attack complex (C5b-9); soluble C5b-9; thrombotic microangiopathies.

FIGURE 1

Pathological findings of renal biopsy in acute and chronic TMA. (A) A thrombus is identified within glomerular capillaries (arrow head) (Masson’s trichrome, 60×). (B) Segmental mesangiolysis is indicative of active lesion (arrow head) (silver, 40×). (C) Chronic lesions show reduplication of GBM (arrow head) (silver, 60×). (D) Necrosis of the arterial wall (arrow head) and glomeruli of shrunken appearance (Masson’s trichrome, 20×). (E) Artery show intimal thickening with concentric appearance (‘onion-skin’) (haematoxylin and eosin, 60×) (F) Acute tubular necrosis (arrow head) is a finding resulting of ischemic changes (haematoxylin and eosin, 20×).

FIGURE 2

TMA management algorithm. When there are TMA signs, clinical and laboratory evaluation is needed to achieve an aetiological diagnosis and specific treatment. ADAMTS-13 activity assessment is urgent, and if <10%, a TTP should be diagnosed, followed by specific treatment. If Stx is detected, then diagnosis of STEC-HUS should be followed by supportive therapy. This supportive therapy, among early start of aetiological treatment, follows the differential diagnosis of all secondary TMA forms. When aHUS is suspected (by exclusion of other TMA forms), a complete evaluation of complement should be performed, eculizumab being the first-line treatment. Complement assessment though both in vitro evaluation of C5b-9 deposits on ECs and soluble C5b-9 levels is strongly encouraged in all cases of TMA, when possible. Especially in secondary forms, where if the patient shows complement overactivation and no response to the treatment of choice, a second-line treatment consisting of administration of complement terminal blockers may be contemplated. If complement assessment is not available—in secondary TMA forms—a therapeutic approach with eculizumab may be also considered (in the absence of response to aetiological treatment).ADAMTS-13, a disintegrin and metalloproteinase with a thrombospondin Type 1 motif, member 13; AF-ab, Anti-phospholipid antibodies; HAV, hepatitis A virus; HBV, hepatitis B virus; HCV, hepatitis C virus; CMV, cytomegalovirus; EBV, Epstein–Barr virus; HIV, human immunodeficiency virus.

FIGURE 3

Complement system. The complement system is the first line of physical defence against pathogens and altered host cells and is composed of a cascade of several plasmatic and membrane proteins expressed on the cell surface. There are three different pathways through which the complement system can be activated: the classical, the lectin and the alternative pathway (upper part of the image). Each of the pathways leads to the activation of C3 convertase that cleaves C3 into C3a and C3b, which binds Factor B and generates active convertases on targeted surfaces. The terminal complement cascade is then initiated by the cleavage of protein C5 by C5 convertase, followed by C5a release (potent anaphylotoxin) and C5b-9 molecule assembly, forming the MAC. This structure becomes inserted into the target cell lipid bilayer and induces cell lysis through proton influx. Dysfunction of the complement cascade may be mediated by aberrations in the pathways of activation, complement regulatory proteins or complement deficiencies, whether genetic or acquired. Soluble C5b-9 levels and C5b-9 deposition induced by patient activated plasma on ECs in vitro are two of the most promising complement monitoring tools used to provide insight into the integrity of the entire complement reaction cascade.

FIGURE 4

Clinical case—complement assessment in TMA for early aetiological treatment. A 52-year-old man, with a past medical history of pancreatic ductal adenocarcinoma (cT1cN0M0), was treated with surgery and concomitant gemcitabine from March to August 2017. After 2 months, he showed nephrotic syndrome, MAHA, thrombocytopenia and progressive kidney failure. A kidney biopsy confirmed TMA both in the glomerulus and arterioles with negative immunofluorescence. To make a rapid differential diagnosis, we ruled out TTP (ADAMTS-13 activity: 87%), STEC-HUS (negative Stx detection), infections (negative cultures) and autoimmune diseases, among other causes of TMA. Concomitantly, we performed a complement assessment with plasma levels of complement components (C3, C4, CH50 and C5b-9) and also with C5b-9 deposition evaluation on ECs culture [57]. Complement levels were within the normal range, but 5 days after admission, we identified significant deposition of C5b-9 on EC (3.2 ± 0.2-fold increase, P < 0.05) (Figure 4) (A). Despite these results, we spent 4 weeks in the initial management of TMA and especially ruling out tumour recurrence (tumour markers, PET-CT) and gemcitabine-induced TMA. MAHA, thrombocytopenia, kidney failure and nephrotic syndrome persisted despite supportive therapy and plasma exchange (18 sessions). Finally, we reached a clinical diagnosis of aHUS by exclusion. At that point (November 2017), we started eculizumab following the data sheet, with TMA haematological response (normalization of platelet count, LDH, haptoglobin and reticulocytes) after 2 weeks, and normal kidney function with complete remission of nephrotic syndrome after 4 weeks, as well as a negativization of C5b-9 deposits (Figure 4) (B). Three months after admission, genetic study revealed a CFI mutation (heterozygous variant in exon 5, c.739T>G [p.Cys247Gly] considered as pathogenic) and risk haplotypes in heterozygosis for CFH and MCP. After 29 months, the patient maintained haematological and kidney remission on eculizumab. In conclusion, complement assessment in all TMA cases (the proposed algorithm in Figure 2) could critically reduce diagnostic times for aHUS (supporting physician suspicion) and establish the therapeutic potential of complement blockade in secondary TMA forms when aetiological treatment—if available—fails.