Measurement of Bradykinin Formation and Degradation in Blood Plasma: Relevance for Acquired Angioedema Associated With Angiotensin Converting Enzyme Inhibition and for Hereditary Angioedema Due to Factor XII or Plasminogen Gene Variants

Abstract

Bradykinin (BK)-mediated angioedema (AE) states are rare acquired or hereditary conditions involving localized edema of the subcutaneous and submucosal tissues. Citrated plasma from healthy volunteers or patients with hereditary angioedema (HAE) with normal level of C1-inhibitor (C1-INH) was used to investigate pathways of BK formation and breakdown relevant to AE physiopathology. The half-life of BK (100 nM) added to normal plasma was 34 s, a value that was increased ~12-fold when the angiotensin converting enzyme (ACE) inhibitor enalaprilat (130 nM) was added (enzyme immunoassay measurements). The BK half-life was similarly increased ~5-fold following 2 daily oral doses of enalapril maleate in healthy volunteers, finding of possible relevance for the most common form of drug-associated AE. We also addressed the kinetics of immunoreactive BK (iBK) formation and decline, spontaneous or under three standardized stimuli: tissue kallikrein (KLK-1), the particulate material Kontact-APTT™ and tissue plasminogen activator (tPA). Relative to controls, iBK production was rapid (10–20 min) and very intense in response to tPA in plasma of female heterozygotes for variants in gene F12 coding for factor XII (FXII) (p.Thr328Lys, 9 patients; p.Thr328Arg, one). An increased response to Kontact-APTT™ and an early tPA-induced cleavage of anomalous FXII (immunoblots) were also observed. Biotechnological inhibitors showed that the early response to tPA was dependent on plasmin, FXIIa and plasma kallikrein. Results from post-menopausal and pre-menopausal women with HAE-FXII were indistinguishable. The iBK production profiles in seven patients with the plasminogen p.Lys330Glu variant (HAE-PLG) did not significantly differ from those of controls, except for an unexpected, rapid and lanadelumab-resistant potentiation of KLK-1 effect. This enzyme did not cleave plasminogen or factor XII, suggesting a possible idiosyncratic interaction of the plasminogen pathogenic variant with KLK-1 activity. KLK-1 abounds in salivary glands and human saliva, hypothetically correlating with the clinical presentation of HAE-PLG that includes the swelling of the tongue, lips and contiguous throat tissues. Samples from HAE patients with normal C1-INH levels and F12 gene did not produce excessive iBK in response to stimuli. The ex vivo approach provides physiopathological insight into AE states and supports the heterogeneous physiopathology of HAE with normal C1-INH.

In Vitro Modeling of Bradykinin-Mediated Angioedema States

“…The half-life of BK was 27 sec, ACE accounting for most of the clearance activity, as the t 1/2 value increased nine-fold in the presence of the ACE inhibitor enalaprilat. We essentially confirmed these results using the commercial EIA, a lower initial BK concentration and human plasma [ 18 ] ( Figure 3 A). Arginine carboxypeptidases (kininase I), producing des-Arg 9 -BK from BK, was a very minor pathway of metabolism (11%), but more important when ACE was blocked (50%) [ 16 ].…”

“…A perhaps unexpected result was that the low background of iBK formation observed in control plasma in the absence of stimulant is not modified in patients with HAE-C1-INH, HAE-FXII or HAE-PLG [ 18 , 44 ] ( Figure 5 A). HAE patients are not symptomatic most of the time: the buffering action of peptidases and protease inhibitors may protect patients from a possible low-grade BK formation during remissions.…”

“…The most common cause of acquired AE is drug-induced, in the context of treatment with angiotensin-I converting enzyme (ACE) inhibitors for cardiovascular conditions [ 14 , 15 ]. The basis of this disorder may be an insufficient clearance of BK, as ACE is by far the major peptidase that inactivates kinins in blood/plasma/serum and in vivo [ 16 , 17 , 18 ]. However, what triggers kinin formation in affected patients is unclear [ 19 ].…”

“…Thus, Adam et al [ 33 ] used ERK1/2 signaling in cells expressing rabbit B 2 receptors to confirm iBK in extracts of plasma stimulated with contaminated heparin. Later, the accumulation of c-Fos in HEK 293a cells expressing the human B 2 receptor was successfully used as a discriminative bioassay to confirm measurements of iBK in extracts of human blood or plasma [ 12 , 18 ]. Lys-BK and BK are approximately equipotent as agonists of the B 2 receptor, so they add up in EIA results as “iBK” and in bioassay results as well, if either one or both are present in a given extract.…”

“…Human healthy subjects who took two daily doses of enalapril maleate, the prodrug of enalaprilat, exhibited a ~5-fold increase of BK t 1/2 added ex vivo to their plasma relative to the pre-treatment value [ 18 ]. Altogether, the findings illustrate that BK is highly unstable and may contribute to the acquired AE associated with ACE inhibitors if, for some reason, important generation of kinins is triggered.…”