Rapidly progressive acute chest syndrome is associated with complement-mediated cell injury in adults with sickle cell disease Free

Introduction Rapidly progressive acute chest syndrome (RP-ACS) is a distinct phenotype of acute chest syndrome, characterized by rapid progression of respiratory compromise, with respiratory failure within 24 hours of the onset of respiratory symptoms and frequently complicated by multiorgan failure (Chaturvedi et al. 2016). Thrombocytopenia early in the presentation is the only predictor of developing this phenotype. In an autopsy case series, platelet-rich microthrombi were found in patients with SCD with fatal ACS. These patients were also found to have a severe drop in platelets before death. The phenotype of ischemic multiorgan failure with microthrombi and thrombocytopenia recapitulates other thrombotic microangiopathic processes characterized by complement dysregulation.

Abnormal complement activation has been observed in SCD patients and mice at baseline and during acute crises. Building on this premise, we tested the hypothesis that rapidly progressive acute chest syndrome in adults with sickle cell disease is associated with increased complement activation in a functional assay of complement-mediated cell injury.

Methods Adults with SCD (confirmed by Hb electrophoresis) were prospectively enrolled from Vanderbilt University Medical Center between May 2016 and June 2017 in three groups: (1) RP-ACS - (i) meeting criteria for ACS, defined as a new pulmonary infiltrate on chest X-ray with any one of fever, cough, wheezing, or dyspnea, (ii) and RP-ACS defined as respiratory failure requiring mechanical ventilation <24 hours from onset of respiratory symptoms; (2) ACS without respiratory failure – (i) meeting criteria for ACS, (ii) not meeting criteria for RP-ACS, (iii) no prior history of life threatening ACS; (3) SCD controls without ACS – (i) with no evidence of acute chest syndrome, vaso-occlusive crisis or other acute illness, and (ii) no history of life-threatening ACS. or cases, blood was drawn by venipuncture during the acute episode before starting red cell exchange. For controls, blood was drawn during routine visits (asymptomatic).

The modified Ham (mHam) assay was run as previously described (Gavriilaki et al. 2015). The mHam assay is based on the principle that a PNH cell line (PIGA-null TF-1 cells) lacking the cell surface complement regulators CD55 and CD59 undergoes lysis in serum containing activated complement. Cell death (measured by a cell viability assay) is a measure of complement activation. Data were summarized as counts (percentages) and medians or means for categorical and continuous variables. We compared the proportion of positive mHam assays (>20% non-viable cells) in each group using the Fisher's exact test. Since this threshold is optimized to diagnose complement-mediated TMA, we also compared the proportion of non-viable cells in each group using the Mann-Whitney test.. P < 0.05 was considered significant.

Results We enrolled 19 patients with SCD, including 2 with RP-ACS, 6 with ACS without respiratory failure, and 11 controls with no ACS. At baseline, patients with RP-ACS and ACS (without respiratory failure) had similar mean hemoglobin levels (10.4 g/dl vs 9.8 g/dl), platelet counts (488 x 10⁹/L vs 310 x 10⁹/L), and HbS levels (63.1% vs 61.0%). At presentation, platelet levels were not significantly different between the groups(380 x 10⁹/L in the RP-ACS vs 261 x 10⁹/L) ; however, nadir platelet count was lower in the RP-ACS group (76.5 ±13.4 x 10⁹/L versus 125.3 ± 66.1 x 10⁹/L). Both patients with RP-ACS died within a mean of 2 days (1 day and 3days).

A positive mHam assay was present in 100% (2/2) of RP-ACS group and 0 of the ACS and no-ACS groups (p = 0.005). The mean percentage of nonviable cells was significantly higher in the RP-ACS group (43.4%) compared to the ACS (8.2%, one‐sided p = 0.02), and compared to the no-ACS groups 7.7% (one‐sided p = 0.0148). Additional testing on a single sample by flow cytometry showed that C5b9 deposition was fully blocked by adding eculizumab but only partially blocked by danicopan, suggesting a contribution of the classical complement pathway as a driver.

Conclusion We demonstrate that the RP-ACS patients in our study exhibited increased complement activation. However, due to the small sample size, our results should be considered as hypothesis-generating. Future studies with larger cohorts and newer functional complement assays, like the mHAM 2.0, should be considered to obtain definitive results.