Plasma exchange for COVID‐19 thrombo‐inflammatory disease

Abstract Severe COVID‐19 disease is a hyperinflammatory, pro‐thrombotic state. We undertook plasma exchange (PEX) to determine its effects on organ function and thrombo‐inflammatory markers. Seven critically ill adults with severe COVID‐19 respiratory failure (PaO2:FiO2 ratio < 200 mm Hg) requiring invasive or noninvasive ventilatory support and elevated thrombo‐inflammatory markers (LDH >800 IU/L and D‐dimer >1000 μg/L (or doubling from baseline) received PEX, daily, for a minimum of 5 days. No other immunomodulatory medications were initiated during this period. Seven patients matched for age and baseline biochemistry were a comparator group. Coagulation screening revealed no evidence of coagulopathy. However, von Willebrand Factor (VWF) activity, antigen and VWF antigen: ADAMTS13 ratio, Factor VIII and D‐dimers were all elevated. Following 5 days of PEX, plasma levels of all the above, and ferritin levels, were significantly reduced (P < .05) while lymphocyte counts normalized (P < .05). The PaO2:FiO2 ratio increased from a median interquartile range (IQR) of 11.6 (10.8‐19.7) kPa to 18.1 (16.0‐25.9) kPa (P < .05). Similar improvements were not observed in controls. Acute kidney injury (AKI) occurred among five patients in the control arm but not in patients receiving PEX. PEX improved oxygenation, decreased the incidence of AKI, normalized lymphocyte counts and reduced circulating thrombo‐inflammatory markers including D‐Dimer and VWF Ag:ADAMTS13 ratio.

mediated by the metalloproteinase, ADAMTS13 (a disintegrin-like and metalloprotease with thrombospondin type 1 motif 13). We noted a high VWf Ag:ADAMTS13 ratio in our COVID-19 patients, which may contribute to the inflammatory, prothrombotic phenotype, comparable to parameters seen in thrombotic microangiopathy (TMA), with evidence of micro and macro thrombosis, primarily affecting the lungs.
We therefore performed a preliminary exploration of the impact of plasma exchange (PEX), a therapy successfully used in TMA conditions [6], on inflammatory and coagulation markers and organ function in a critically ill cohort of COVID-19 patients with ARDS.

METHODS
Critically ill adults (≥18 years) patients with proven COVID-19 disease had markers of coagulation measured on admission to the intensive care unit (ICU). Subsequently, a prospective case-controlled, nonblinded descriptive study was performed to assess the effectiveness and safety of PEX. Institutional agreement was confirmed, and written informed consent was obtained from patients or their next-of-kin.
Respiratory failure criteria for treatment included bilateral infiltrates on chest imaging, worsening respiratory function, based on a ratio of arterial oxygen partial pressure to fractional inspired oxy-

RESULTS
Levels of VWF activity, VWF antigen, VWF antigen:ADAMTS13 ratio, and D-dimers were significantly increased in critically ill patients with COVID-19 disease (Table 1). However, other markers of coagulation, including endogenous anticoagulant levels, were all within the normal range. Baseline demographics and laboratory parameters were similar between PEX and control group patients ( Table 2). At the end of the 5-day course, PEX reduced VWF antigen and activity, VWF antigen/ADAMTS13 ratio and Factor VIII levels; all P < .05 ( Figure 1A and B; Table 3). ADAMTS13 activity remained unchanged over this period (P = .375). There was a significant fall of inflammatory markers including ferritin (P = .016) and D-dimers (P = .039), and improvement in lymphopenia (P = .047) (Figure 2; Table 3). There was no statistically significant change in CRP (P = .109), albeit this was skewed by one patient who developed a secondary Klebsiella pneumoniae sepsis. No significant changes were seen in serum cytokines or platelet count (P = .297). Serum cytokines were not elevated at baseline, and PEX had no effect on circulating levels of serum cytokines (Figure 3).

The PaO 2 :FiO 2 ratio increased in all patients from 87 (81-148) mm
Hg at baseline to 136 (120-194) mm Hg after 5 days of PEX (Figure 2; Table 3). No patient developed acute kidney injury (AKI) during PEX treatment; the one patient requiring renal replacement therapy at baseline had this discontinued at day 4.
In contrast, there was no significant fall in CRP, recovery of lymphocytes, or improvement in PaO 2 :FiO 2 ratio among control group patient ( Figure 2). Furthermore, there was an increase in the serum creatinine (P < .05), with five patients in the control group developing new onset AKI.

DISCUSSION
Severe infection with COVID-19 presents a unique thromboinflammatory syndrome that remains incompletely characterized.
The very high reported incidence of arterial, venous and pulmonary thromboembolism detected by imaging [2], and the widespread pulmonary macro-and microthrombi noted at post-mortem implicates the prothrombotic component as a major factor underlying the pathophysiology of this disease [3,4].
COVID-19 shares features of TMA but displays a distinct phenotype. Indeed, we found a raised VWF, elevated Factor VIII and an increased VWF antigen:ADAMTS13 ratio indicative of a prothrombotic predisposition. Conversely, ADAMTS13 activity was normal, and there was no thrombocytopenia. Levels of other coagulation factors and endogenous anticoagulants such as antithrombin, protein C and protein S were remarkably normal, especially given the likelihood of markedly raised fibrin generation as suggested by high levels of D-dimers. Fibrinogens were high, behaving as an acute phase marker.
These coagulation abnormalities appear unique to COVID-19 disease.
There were no consumptive features as seen in disseminated intravascular coagulation (DIC), which might be expected with the continuous, severe inflammatory-thrombotic process.
PEX restores hemostatic balance and reduces levels of circulating inflammatory markers in a range of TMA. There was therefore a compelling rationale to explore its utility in COVID-19. We therefore undertook a limited case-controlled study to determine safety and feasibility, and its impact on thrombo-inflammatory markers and respiratory parameters. We were able to demonstrate normalization of prothrombotic and acute phase response markers, such as ferritin, fibrinogen, and CRP following 5 days of PEX. The VWF Ag:ADAMTS13 ratio has been highlighted in other thrombotic conditions with higher ratios being associated with inferior outcomes in stroke and cardiac disease [8,9]. Lymphopenia and elevated D-dimers levels, poor prognostic risk factors in COVID-19, were also reversed [10]. Notably, circulating levels of proinflammatory cytokines were unaffected, but F I G U R E 1 A and B, Comparison of coagulation data of patients before and following five courses of plasma exchange. Grey areas highlight normal ranges (n = 7) Abbreviation: PEX, plasma exchange.
these were not particularly high at baseline, particularly in relation to other causes of ARDS such as bacterial sepsis [11]. Our finding of low levels of circulating cytokines is in keeping with other studies in COVID-19 patients, suggesting the description of "cytokine storm" is a misnomer in this condition.
Our finding of an improvement in PaO 2 :FiO 2 ratio among all treated patients is of particular importance, as mortality in COVID-19 is primarily associated with severe hypoxemic respiratory failure and histological features of pulmonary thrombosis. The fall in D-dimers, as a marker of fibrinolytic activity, likely represents a reduction in fibrin clot formation. No safety concerns were highlighted during the course of PEX, in particular no bleeding or AKI.
Three patients deteriorated once PEX was stopped with worsening gas exchange and a recrudescence in inflammatory marker levels. A summary of the impact of PEX on thrombotic, inflammatory, and cytokine parameters and organ function measured before initiation of and on completion of PEX. n = 7; apart from P:F ratio (n = 6); LDH (n = 5); serum cytokines (n = 5), and serum creatinine (n = 6). Data are expressed as median and interquartile range.

F I G U R E 3
Comparison of serum cytokines before and following five courses of plasma exchange (n = 5). Cytokines measured using a multiplex assay, from PRE the 1st PEX and samples taken post the final PEX on day 5. The increased IL-6 post-PEX in one case represents an increased CRP and severe sepsis. There was a trend for decrease in cytokines, but this was not significant. Other than IL-6, the remaining cytokines were within the normal range understanding of severe later stage COVID-19. It was anticipated that their ongoing severe ARDS picture represented a fibrotic stage of disease however, with PEX, their inflammatory markers improved as did their respiratory parameters and, in one case, renal function.
Despite promising results, we acknowledge a number of limitations.
This was a small case controlled study. There was also variation in the timing of initiation of PEX within the disease course. The subset of patients who are most likely to benefit from PEX and the optimal duration of treatment remains unknown. A longer course, or a repeat course, could be envisaged, recognizing this may create logistic difficulties in terms of the number of patients who can be treated concurrently.

CONCLUSION
PEX in patients with severe COVID-19 disease with ARDS was associated with improvements in oxygenation and reversal of the thromboinflammatory markers and normalization of the lymphocytes, even when used late in the disease process. There is a positive impact on organ function and significant improvement in parameters defined as poor prognosis in severe COVID-19 infection. This provides support for conduct of a formal randomized controlled trial.