Deucrictibant for angioedema due to acquired C1-inhibitor deficiency: A randomized-controlled trial.

BACKGROUND: Angioedema due to acquired C1-inhibitor deficiency is a very rare but serious disease, with an estimated prevalence of 1 per 500,000 persons. There are no approved therapies to treat or prevent angioedema swelling in patients with this condition. Deucrictibant is a specific, orally bioavailable, competitive antagonist of the bradykinin B2 receptor currently under investigation for hereditary angioedema. OBJECTIVE: Our aim was to assess the efficacy and safety of deucrictibant as acute and prophylactic treatment for angioedema due to acquired C1-inhibitor deficiency. METHODS: A 2-part, randomized, double-blind, placebo-controlled crossover study was conducted. In Part 1, 4 consecutive angioedema attacks were treated with 3 doses of deucrictibant (10 mg, 20 mg, and 30 mg) or placebo. In Part 2, deucricibant, 20 mg, or placebo was administered twice daily for 2 treatment periods of 8 weeks. RESULTS: Three patients were enrolled; of those 3 patients, 1 completed both study parts and 2 completed only Part 2. In Part 1, a reduction in attack severity was observed in the 3 attacks treated with deucrictibant as opposed to an increase in severity of the attack treated with placebo. In Part 2, the individual mean monthly attack rates were 2.0, 0.6, and 1.0 during the placebo period and 0.0 across all patients during treatment with deucrictibant. There were no severe adverse events and 1 self-limiting treatment-emergent adverse event (abdominal pain). CONCLUSIONS: Deucrictibant has the potential to effectively and safely treat and prevent angioedema attacks due to acquired C1-inhibitor deficiency.

Management of hereditary angioedema in Japan: Focus on icatibant for the treatment of acute attacks.

Hereditary angioedema (HAE) is characterized by unpredictable, recurring and painful swelling episodes that can be disabling or even life-threatening. Awareness of HAE has progressively grown worldwide, and options for treatment of acute attacks and prevention of future attacks continue to expand; however, unmet needs in diagnosis and treatment remain. In Japan, recognition of HAE within the medical community remains low, and numerous obstacles complicate diagnosis and access to treatment. Importance of timely treatment of HAE attacks with on-demand therapies is continually demonstrated; recommended agents per the WAO/EAACI treatment guidelines published in 2018 include C1 inhibitor (C1-INH) concentrate, ecallantide, and icatibant. In Japan, multiple factors contribute to delayed HAE treatment (potentially leading to life-threatening consequences), including difficulties in finding facilities at which C1-INH agents are readily available. Recognition of challenges faced in Japan can help promote efforts to address current needs and expand access to effective therapies. Icatibant, a potent, selective bradykinin B2 receptor antagonist, has demonstrated inhibition of various bradykinin-induced biological effects in preclinical studies and has shown efficacy in treating attacks in various clinical settings (e.g. clinical trials, real-world studies), and HAE patient populations (e.g. with C1-INH deficiency, normal C1-INH). Icatibant was approved in Japan for the treatment of HAE attacks in September 2018; its addition to the HAE treatment armamentarium contributes to improved patient care. In Japan, disease awareness and education campaigns are warranted to further advance the management of HAE patients in light of the unmet needs and the emerging availability of modern diagnostic approaches and therapies.

Efficacy, pharmacokinetics, and safety of icatibant for the treatment of Japanese patients with an acute attack of hereditary angioedema: A phase 3 open-label study.

BACKGROUND: Hereditary angioedema (HAE) is a genetic disease characterized by recurrent swelling episodes affecting the skin, gastrointestinal mucosa, and upper respiratory tract. METHODS: A phase 3, single-arm, open-label study was performed to evaluate a selective bradykinin B2 receptor antagonist, icatibant, for the treatment of acute attacks in Japanese patients with HAE Type I or II. After the onset of an acute attack, icatibant 30 mg was administered by the patient or a healthcare professional via subcutaneous injection in the abdomen. RESULTS: Eight patients who had an attack affecting the skin (n = 4), abdomen (n = 3), or larynx (n = 1) were treated with icatibant (3 of the injections were self-administered). The median time to onset of symptom relief was 1.75 h (95% confidence interval, 1.00-2.50), and all patients had symptom relief within 5 h after administration. The time to maximum plasma concentration of icatibant was 1.79 h, and the maximum plasma concentration was 405 ng/ml. Seven patients experienced an injection site reaction, and 3 patients had adverse events (2 patients had a worsening or repeat HAE attack 29.0 and 18.3 h after icatibant administration, respectively, and 1 had headache). CONCLUSIONS: Although the number of patients is small, the efficacy and tolerability of icatibant for acute attacks were demonstrated in Japanese patients with HAE, regardless of self-administration or administration by healthcare professional.

Activation of bradykinin B2 receptor induced the inflammatory responses of cytosolic phospholipase A2 after the early traumatic brain injury.

Phospholipase A2 is a known aggravator of inflammation and deteriorates neurological outcomes after traumatic brain injury (TBI), however the exact inflammatory mechanisms remain unknown. This study investigated the role of bradykinin and its receptor, which are known initial mediators within inflammation activation, as well as the mechanisms of the cytosolic phospholipase A2 (cPLA2)-related inflammatory responses after TBI. We found that cPLA2 and bradykinin B2 receptor were upregulated after a TBI. Rats treated with the bradykinin B2 receptor inhibitor LF 16-0687 exhibited significantly less cPLA2 expression and related inflammatory responses in the brain cortex after sustaining a controlled cortical impact (CCI) injury. Both the cPLA2 inhibitor and the LF16-0687 improved CCI rat outcomes by decreasing neuron death and reducing brain edema. The following TBI model utilized both primary astrocytes and primary neurons in order to gain further understanding of the inflammation mechanisms of the B2 bradykinin receptor and the cPLA2 in the central nervous system. There was a stronger reaction from the astrocytes as well as a protective effect of LF16-0687 after the stretch injury and bradykinin treatment. The protein kinase C pathway was thought to be involved in the B2 bradykinin receptor as well as the cPLA2-related inflammatory responses. Rottlerin, a Protein Kinase C (PKC) δ inhibitor, decreased the activity of the cPLA2 activity post-injury, and LF16-0687 suppressed both the PKC pathway and the cPLA2 activity within the astrocytes. These results indicated that the bradykinin B2 receptor-mediated pathway is involved in the cPLA2-related inflammatory response from the PKC pathway.

Bifunctional fusion proteins containing the sequence of the bradykinin homologue maximakinin: activities at the rat bradykinin B2 receptor.

To support bradykinin (BK) B2 receptor (B2R) detection and therapeutic stimulation, we developed and characterized fusion proteins consisting of the BK homolog maximakinin (MK), or variants, positioned at the C-terminus of functional proteins (enhanced green fluorescent protein (EGFP), the peroxidase APEX2, or human serum albumin (HSA)). EGFP-MK loses its reactivity with anti-BK antibodies and molecular mass as it progresses in the endosomal tract of cells expressing rat B2Rs (immunoblots, epifluorescence microscopy). APEX2-(NG)15-MK is a bona fide agonist of the rat, but not of the human B2R (calcium and c-Fos signaling) and is compatible with the cytochemistry reagent TrueBlue (microscopy), a luminol-based reagent, or 3,3',5,5'-tetramethylbenzidine (luminescence or colourimetric B2R detection, cell well plate format). APEX2-(NG)15-MK is a non-isotopic ligand suitable for drug discovery via binding competition. Affinity-purified secreted forms of HSA fused with peptides possessing the C-terminal MK or BK sequence failed to stimulate the rat B2R in the concentration range of 50-600 nmol/L. However, the non-secreted construction myc-HSA-MK is a B2R agonist, indicating that protein denaturation made the C-terminal sequence available for receptor binding. Fusion protein ligands of the B2R are stable but subjected to slow intracellular inactivation, strong species specificity, and possible steric hindrance between the receptor and large proteins.

[EFFICACY, PHARMACOKINETICS, AND SAFETY OF ICATIBANT FOR THE TREATMENT OF JAPANESE PATIENTS WITH AN ACUTE ATTACK OF HEREDITARY ANGIOEDEMA: A PHASE 3 OPEN-LABEL STUDY].

BACKGROUND: Hereditary angioedema (HAE) is characterized by paroxysmal edema of the skin, gastrointestinal mucosa, and upper respiratory tract. PURPOSE: This study investigated icatibant, a selective bradykinin B2 receptor antagonist, as treatment for Japanese patients with an acute HAE attack. METHODS: This was an open-label, single-arm, Phase 3 study of Japanese adults with HAE type I or II. Icatibant (30 mg) was administered (by a healthcare professional [HCP] or self-administered) as a subcutaneous injection in the abdomen. RESULTS: Eight patients (4 cutaneous, 3 abdominal, 1 laryngeal) were treated with icatibant (all single injection; 3 self-administered, 5 HCP-administered). The median time to onset of symptom relief was 1.75 hours (95% confidence interval, 1.00 to 2.50); all patients had onset of relief within 5 hours. The estimated time to maximum icatibant concentration in the circulation was 1.79 hours and the maximum concentration was 405 ng/mL. There were 3 patients who experienced 3 adverse events (2 HAE attacks and 1 headache); 7 patients experienced an injection site reaction. CONCLUSION: Although our study was limited by the small number of patients, we found that icatibant was an effective and well-tolerated treatment for Japanese patients with acute HAE attacks, regardless of whether it was administered by a HCP or self-administered.

Tissue kallikrein protects SH-SY5Y neuronal cells against oxygen and glucose deprivation-induced injury through bradykinin B2 receptor-dependent regulation of autophagy induction.

Previous studies have demonstrated that tissue kallikrein (TK) protects against cerebral ischemia injury mainly through inhibition of apoptosis via bradykinin B2 receptor (B2R). In this study, we proposed that autophagy induction contributed to the neuroprotective mechanism of TK. To validate this hypothesis, we investigated TK-induced autophagy and its signaling mechanisms in human SH-SY5Y cells exposed to oxygen and glucose deprivation (OGD). We found that TK treatment enhanced autophagy induction, reflected by augmented LC3 conversion and Beclin1 expression, decreased p62 levels and increased monomeric red fluorescent protein-LC3 puncta formation. Green fluorescent protein-monomeric red fluorescent protein-LC3 adenovirus assay indicated that TK maintained autophagic flux. Moreover, bafilomycin A1 (Baf.A1) caused obvious LC3-II accumulation either in the presence or absence of TK. Autophagy inhibition by Beclin1 knockdown or Baf.A1 treatment abrogated the neuroprotective effects of TK. Mitogen-activated protein kinase kinase 1/2 (MEK1/2)/extracellular signal-regulated kinase (ERK)1/2 and AMP-activated protein kinase (AMPK)/tuberous sclerosis complex 2 (TSC2)/mammalian target of rapamycin (mTOR) signaling were induced by OGD stress and enhanced by TK. MEK/ERK inhibitor U0126 alone elevated autophagy in OGD conditions, but impaired TK-induced autophagy. Blockade of AMPK/TSC2/mTOR signaling by AMPK inhibitor compound C and shRNA mediated the knockdown of AMPK α1 and TSC2 but abolished autophagy in SH-SY5Y cells exposed to OGD treated either with or without TK. Moreover, B2R expression was up-regulated by OGD exposure. B2R knockdown attenuated autophagy and suppressed MEK1/2/ERK1/2 and AMPK/TSC2/mTOR signaling in OGD conditions in either the presence or absence of TK. In sum, we revealed the significance of B2R-mediated MEK/ERK and AMPK signaling in autophagy induction under OGD stress, and proposed novel mechanisms involved in the neuropotective function of TK through B2R-dependent regulation of autophagy. We propose the depicted model for the neuroprotective mechanism of tissue kallikrein (TK) during OGD stress: TK enhances bradykinin B2 receptor (B2R)-mediated MEK1/2/ERK1/2 and AMPK/TSC2/mTOR signaling, thus inducing protective autophagy. The findings reported in this study should provide new evidence for the pro-survival role of B2R-mediated autophagy in cerebral ischemia.

Bradykinin B2 receptor expression in the bronchial mucosa of allergic asthmatics: the role of NF-kB.

BACKGROUND: Bradykinin (BK) mediates acute allergic asthma and airway remodelling. Nuclear factor-kappa B (NF-kB) is potentially involved in BK B2 receptor (B2R) regulation. OBJECTIVE: In this observational cross-sectional study, B2R and NF-kB expression was evaluated in bronchial biopsies from mild asthmatics (after diluent/allergen challenge) and healthy controls, examining the role of NF-kB in B2R expression in primary human fibroblasts from normal and asthmatic subjects (HNBFb and HABFb). METHODS: B2R and NF-kB (total and nuclear) expression was analysed by immunohistochemistry in biopsies from 10 mild intermittent asthmatics (48 h after diluent/allergen challenge) and 10 controls undergoing bronchoscopy. B2R co-localization in 5B5(+) and αSMA(+) mesenchymal cells was studied by immunofluorescence/confocal microscopy, and B2R expression in HABFb/HNBFb incubated with interleukin (IL)-4/IL-13 with/without BK, and after NF-kB inhibitor, by Western blotting. RESULTS: Bronchial mucosa B2R and nuclear NF-kB expression was higher in asthmatics after diluent (B2R only) and allergen challenge than in controls (P < 0.05), while B2R and NF-kB (total and nuclear) increased after allergen compared with after diluent (P < 0.05). Allergen exposure increased B2R expression in 5B5(+) and αSMA(+) cells. Constitutive B2R protein expression was higher in HABFb than in HNBFb (P < 0.05) and increased in both cell types after IL-13 or IL-4/IL-13 and BK treatment. This increase was suppressed by a NF-kB inhibitor (P < 0.05). CONCLUSIONS & CLINICAL RELEVANCE: Bronchial B2R expression is constitutively elevated in allergic asthma and is further increased after allergen exposure together with NF-kB expression. NF-kB inhibitor blocked IL-4/IL-13-induced increase in B2R expression in cultured fibroblasts, suggesting a role as potential anti-asthma drug.

Differential regulation of endosomal GPCR/ß-arrestin complexes and trafficking by MAPK.

ß-Arrestins are signaling adaptors that bind to agonist-occupied G protein-coupled receptors (GPCRs) and target them for endocytosis; however, the mechanisms regulating receptor/ß-arrestin complexes and trafficking in endosomes, remain ill defined. Here we show, in live cells, differential dynamic regulation of endosomal bradykinin B2 receptor (B2R) complexes with either ß-arrestin-1 or -2. We find a novel role for MAPK in the B2R/ß-arrestin-2 complex formation, receptor trafficking and signaling mediated by an ERK1/2 regulatory motif in the hinge domain of the rat ß-arrestin-2 (PET(178)P), but not rat ß-arrestin-1 (PER(177)P). While the ERK1/2 regulatory motif is conserved between rat and mouse ß-arrestin-2, it is surprisingly not conserved in human ß-arrestin-2 (PEK(178)P). However, mutation of lysine 178 to threonine is sufficient to confer MAPK sensitivity to the human ß-arrestin-2. Furthermore, substitution for a phosphomimetic residue in both the rat and the human ß-arrestin-2 (T/K178D) significantly stabilizes B2R/ß-arrestin complexes in endosomes, delays receptor recycling to the plasma membrane and maintains intracellular MAPK signaling. Similarly, the endosomal trafficking of ß2-adrenergic, angiotensin II type 1 and vasopressin V2 receptors was altered by the ß-arrestin-2 T178D mutant. Our findings unveil a novel subtype specific mode of MAPK-dependent regulation of ß-arrestins in intracellular trafficking and signaling of GPCRs, and suggest differential endosomal receptor/ß-arrestin-2 signaling roles among species.

Repeat treatment of acute hereditary angioedema attacks with open-label icatibant in the FAST-1 trial.

Hereditary angioedema (HAE) is characterized by potentially life-threatening recurrent episodes of oedema. The open-label extension (OLE) phase of the For Angioedema Subcutaneous Treatment (FAST)-1 trial (NCT00097695) evaluated the efficacy and safety of repeated icatibant exposure in adults with multiple HAE attacks. Following completion of the randomized, controlled phase, patients could receive open-label icatibant (30 mg subcutaneously) for subsequent attacks. The primary end-point was time to onset of primary symptom relief, as assessed by visual analogue scale (VAS). Descriptive statistics were reported for cutaneous/abdominal attacks 1-10 treated in the OLE phase and individual laryngeal attacks. Post-hoc analyses were conducted in patients with ≥ 5 attacks across the controlled and OLE phases. Safety was evaluated throughout. During the OLE phase, 72 patients received icatibant for 340 attacks. For cutaneous/abdominal attacks 1-10, the median time to onset of primary symptom relief was 1·0-2·0 h. For laryngeal attacks 1-12, patient-assessed median time to initial symptom improvement was 0·3-1·2 h. Post-hoc analyses showed the time to onset of symptom relief based on composite VAS was consistent across repeated treatments with icatibant. One injection of icatibant was sufficient to treat 88·2% of attacks; rescue medication was required in 5·3% of attacks. No icatibant-related serious adverse events were reported. Icatibant provided consistent efficacy and was well tolerated for repeated treatment of HAE attacks.

Antihypertensive and renoprotective effect of the kinin pathway activated by potassium in a model of salt sensitivity following overload proteinuria.

The albumin overload model induces proteinuria and tubulointersitial damage, followed by hypertension when rats are exposed to a hypersodic diet. To understand the effect of kinin system stimulation on salt-sensitive hypertension and to explore its potential renoprotective effects, the model was induced in Sprague-Dawley rats that had previously received a high-potassium diet to enhance activity of the kinin pathway, followed with/without administration of icatibant to block the kinin B2 receptor (B2R). A disease control group received albumin but not potassium or icatibant, and all groups were exposed to a hypersodic diet to induce salt-sensitive hypertension. Potassium treatment increased the synthesis and excretion of tissue kallikrein (Klk1/rKLK1) accompanied by a significant reduction in blood pressure and renal fibrosis and with downregulation of renal transforming growth factor-ß (TGF-ß) mRNA and protein compared with rats that did not receive potassium. Participation of the B2R was evidenced by the fact that all beneficial effects were lost in the presence of the B2R antagonist. In vitro experiments using the HK-2 proximal tubule cell line showed that treatment of tubular cells with 10 nM bradykinin reduced the epithelial-mesenchymal transdifferentiation and albumin-induced production of TGF-ß, and the effects produced by bradykinin were prevented by pretreatment with the B2R antagonist. These experiments support not only the pathogenic role of the kinin pathway in salt sensitivity but also sustain its role as a renoprotective, antifibrotic paracrine system that modulates renal levels of TGF-ß.

Repeat treatment with icatibant for multiple hereditary angioedema attacks: FAST-2 open-label study.

BACKGROUND: The For Angioedema Subcutaneous Treatment (FAST)-2, a phase III, double-blind, randomized, multicenter, placebo-controlled study (ClinicalTrials.gov identifier: NCT00500656), established the efficacy and safety of single injections of icatibant, a bradykinin B2 receptor antagonist, in the treatment of hereditary angioedema (HAE) attacks. Here, we evaluate the efficacy and safety of repeated treatment with icatibant in adult patients experiencing HAE attacks during the FAST-2 open-label extension (OLE) phase. METHODS: Patients completing the controlled phase were eligible to participate in the OLE phase and receive open-label icatibant (30 mg subcutaneously) for the treatment of cutaneous, abdominal, and/or laryngeal HAE attack(s) severe enough to warrant treatment. Time to onset of symptom relief was calculated for each attack. Descriptive analyses (median, 95% CIs) were performed for all attacks; post hoc analyses were conducted in patients with at least five icatibant-treated attacks throughout the FAST-2 OLE phase. Safety was also monitored. RESULTS: Fifty-four patients received icatibant for 374 attacks (176 cutaneous, 168 abdominal, and 30 laryngeal). For cutaneous and/or abdominal attacks (attacks 2-5), the median times to onset of symptom relief ranged between 2.0 and 2.5 h. For all laryngeal attacks, the median times to regression (start of improvement) of symptoms ranged between 0.3 and 4.0 h. Post hoc analyses showed that the overall median time to onset of symptom relief was 2.0 h. Overall, 89.8% of attacks resolved with a single icatibant injection. No drug-related serious adverse events were reported. CONCLUSIONS: These findings have demonstrated the efficacy and safety of repeated icatibant treatment for HAE attacks.

Exercise Induced-Cytokines Response in Marathon Runners: Role of ACE I/D and BDKRB2 +9/-9 Polymorphisms.

Renin-angiotensin system (RAS) and kallikrein-kinin system (KKS) have a different site of interaction and modulate vascular tone and inflammatory response as well on exercise adaptation, which is modulated by exercise-induced cytokines. The aim of the study was to evaluate the role of ACE I/D and BDKRB2 +9/-9 polymorphism on exercise-induced cytokine response. Seventy-four male marathon finishers, aged 30 to 55 years, participated in this study. Plasma levels of exercise-induced cytokines were determined 24 h before, immediately after, and 24 h and 72 h after the São Paulo International Marathon. Plasma concentrations of MCP-1, IL-6 and FGF-21 increased after marathon in all genotypes of BDKRB2. IL-10, FSTL and BDNF increased significantly after marathon in the genotypes with the presence of the -9 allele. FSTL and BDNF concentrations were higher in the -9/-9 genotype compared to the +9/+9 genotype before (p = 0.006) and after the race (p = 0.023), respectively. Apelin, IL-15, musclin and myostatin concentrations were significantly reduced after the race only in the presence of -9 allele. Marathon increased plasma concentrations of MCP1, IL-6, BDNF and FGF-21 in all genotypes of ACE I/D polymorphism. Plasma concentrations of IL-8 and MIP-1alpha before the race (p = 0.015 and p = 0.031, respectively), of MIP-1alpha and IL-10 after the race (p = 0.033 and p = 0.047, respectively) and VEGF 72 h after the race (p = 0.018) were lower in II homozygotes compared to runners with the presence of D allele. One day after the race we also observed lower levels of MIP-1alpha in runners with II homozygotes compared to DD homozygotes (p = 0.026). Before the marathon race myostatin concentrations were higher in DD compared to II genotypes (p = 0.009). Myostatin, musclin, IL-15, IL-6 and apelin levels decreased after race in genotypes with the presence of D allele. After the race ACE activity was negatively correlated with MCP1 (r = -56, p < 0.016) and positively correlated with IL-8, IL-10 and MIP1-alpha (r = 0.72, p < 0.0007, r = 0.72, p < 0.0007, r = 0.47, p < 0.048, respectively). The runners with the -9/-9 genotype have greater response in exercise-induced cytokines related to muscle repair and cardioprotection indicating that BDKRB2 participate on exercise adaptations and runners with DD genotype have greater inflammatory response as well as ACE activity was positively correlated with inflammatory mediators. DD homozygotes also had higher myostatin levels which modulates protein homeostasis.

Treatment of hereditary angioedema-single or multiple pathways to the rescue.

Hereditary angioedema (HAE) is a rare disease caused by mutations in the SERPING1 gene. This results in deficient or dysfunctional C1 esterase inhibitor (C1-INH) and affects multiple proteases involved in the complement, contact-system, coagulation, and fibrinolytic pathways. Current options for the treatment and prevention of HAE attacks include treating all affected pathways via direct C1-INH replacement therapy; or specifically targeting components of the contact activation system, in particular by blocking the bradykinin B2 receptor (B2R) or inhibiting plasma kallikrein, to prevent bradykinin generation. Intravenously administered plasma-derived C1-INH (pdC1-INH) and recombinant human C1-INH have demonstrated efficacy and safety for treatment of HAE attacks, although time to onset of symptom relief varied among trials, specific agents, and dosing regimens. Data from retrospective and observational analyses support that short-term prophylaxis with intravenous C1-INH products can help prevent HAE attacks in patients undergoing medical or dental procedures. Long-term prophylaxis with intravenous or subcutaneous pdC1-INH significantly decreased the HAE attack rate vs. placebo, although breakthrough attacks were observed. Pathway-specific therapies for the management of HAE include the B2R antagonist icatibant and plasma kallikrein inhibitors ecallantide, lanadelumab, and berotralstat. Icatibant, administered for treatment of angioedema attacks, reduced B2R-mediated vascular permeability and, compared with placebo, reduced the time to initial symptom improvement. Plasma kallikrein inhibitors, such as ecallantide, block the binding site of kallikrein to prevent cleavage of high molecular weight kininogen and subsequent bradykinin generation. Ecallantide was shown to be efficacious for HAE attacks and is licensed for this indication in the United States, but the labeling recommends that only health care providers administer treatment because of the risk of anaphylaxis. In addition to C1-INH replacement therapy, the plasma kallikrein inhibitors lanadelumab and berotralstat are recommended as first-line options for long-term prophylaxis and have demonstrated marked reductions in HAE attack rates. Investigational therapies, including the activated factor XII inhibitor garadacimab and an antisense oligonucleotide targeting plasma prekallikrein messenger RNA (donidalorsen), have shown promise as long-term prophylaxis. Given the requirement of lifelong management for HAE, further research is needed to determine how best to individualize optimal treatments for each patient.

The Bradykinin B2 Receptor Agonist (NG291) Causes Rapid Onset of Transient Blood-Brain Barrier Disruption Without Evidence of Early Brain Injury.

Background: The blood-brain barrier (BBB) describes the brain's highly specialized capillaries, which form a dynamic interface that maintains central nervous system (CNS) homeostasis. The BBB supports the CNS, in part, by preventing the entry of potentially harmful circulating molecules into the brain. However, this specialized function is challenging for the development of CNS therapeutics. Several strategies to facilitate drug delivery into the brain parenchyma via disruption of the BBB have been proposed. Bradykinin has proven effective in disrupting mechanisms across the blood-tumor barrier. Unfortunately, bradykinin has limited therapeutic value because of its short half-life and the undesirable biological activity elicited by its active metabolites. Objective: To evaluate NG291, a stable bradykinin analog, with selective agonist activity on the bradykinin-B2 receptor and its ability to disrupt the BBB transiently. Methods: Sprague Dawley rats and CD-1 mice were subjected to NG291 treatment (either 50 or 100 µg/kg, intravenously). Time and dose-dependent BBB disruption were evaluated by histological analysis of Evans blue (EB) extravasation. Transcellular and paracellular BBB leakage were assessed by infiltration of 99mTc-albumin (66.5 KDa) and 14C-sucrose (340 Da) radiolabeled probes into the brains of CD-1 mice treated with NG291. NG291 influence on P-glycoprotein (P-gp) efflux pump activity was evaluated by quantifying the brain accumulation of 3H-verapamil, a known P-gp substrate, in CD-1 mice. Results: NG291-mediated BBB disruption was localized, dose-dependent, and reversible as measured by EB extravasation. 99mTc-albumin leakage was significantly increased by 50 µg/kg of NG291, whereas 100 µg/kg of NG291 significantly augmented both 14C-sucrose and 99mTc-albumin leakage. NG291 enhanced P-gp efflux transporter activity and was unable to increase brain uptake of the P-gp substrate pralidoxime. NG291 did not evoke significant short-term neurotoxicity, as it did not increase brain water content, the number of Fluoro-Jade C positive cells, or astrocyte activation. Conclusion: Our findings strongly suggest that NG291 increases BBB permeability by two different mechanisms in a dose-dependent manner and increases P-gp efflux transport. This increased permeability may facilitate the penetration into the brain of therapeutic candidates that are not P-gp substrates.

The antihypertensive effect of MK on spontaneously hypertensive rats through the AMPK/Akt/eNOS/NO and ERK1/2/Cx43 signaling pathways.

We investigated the antihypertensive effects of maximakinin (MK) on spontaneously hypertensive rats (SHRs). The effects of MK on arterial blood pressure in SHRs were observed, and flow cytometry and 4,5-diaminofluorescein-2 staining were used to examine MK-induced nitric oxide (NO) release in human umbilical vein endothelial cells (HUVECs). Western blotting was used to analyze the effects of MK on the expression of AMP-activated protein kinase (AMPK), Akt, Connexin 43, ERK1/2, p38, and p-eNOS in HUVECs. The results showed that MK induced a more significant antihypertensive effect on SHRs than bradykinin (BK). MK induced significant increases in endothelial nitric oxide synthase (eNOS) phosphorylation and NO release in HUVECs. MK also significantly increased the phosphorylation of Akt and AMPK in HUVECs. The AMPK inhibitor compound C blocked the effect of MK on the generation of NO. MK induced the phosphorylation of ERK1/2, p38, and Connexin 43. The expression of p-Connexin 43 was significantly decreased in the presence of the ERK1/2 inhibitor U0126 but not the p38 inhibitor SB203580. The effects of MK on the phosphorylation of AMPK and ERK1/2 were significantly decreased by the BK B2 receptor inhibitor HOE-140. In summary, MK can significantly reduce blood pressure in SHRs. The antihypertensive effect might be mediated through the activation of the BK B2 receptor, while the downstream AMPK/PI3K/Akt/eNOS/NO and ERK1/2/Connexin 43 signaling pathways play additional roles.

Bradykinin Activates the Bradykinin B2 Receptor to Ameliorate Neuronal Injury in a Rat Model of Spinal Cord Ischemia-Reperfusion Injury.

Bradykinin and bradykinin B2 receptors (B2R) play important roles in both the peripheral and central nervous systems. The aim of this study was to explore the changes of bradykinin and B2R in spinal cord ischemic injury (SCII) and whether bradykinin treatment would improve the neurologic function of SCII rats. The rats were divided into the sham group, the SCII group, and three doses of bradykinin (50, 100, 150 µg/kg) groups. The neurologic function was assessed by the Basso, Beattie, and Bresnahan (BBB) score at -1, 1, 3, 5, and 7 days postsurgery. Bradykinin concentration in serum and IL-6, TNF-α, and MCP-1 levels in the spinal cord were detected by ELISA. The mRNA expressions of B2R, IL-6, TNF-α, MCP-1, COX-2, and iNOS in the spinal cord were determined by RT-PCR. The protein expressions of B2R, COX-2, iNOS, p65, and p-p65 were detected by Western blot. Immunohistochemical staining was used to examine B2R expression in the L4-6 segments of the spinal cord. Bradykinin levels in serum and B2R expression in the spinal cord were downregulated in SCII rats. Bradykinin treatment significantly improved the hind limb motor function of SCII rats and increased B2R expression, inhibiting COX-2, iNOS, and p-p65 expression in the spinal cord of SCII rats together with a decrease of the inflammatory mediators of IL-6, TNF-α, and MCP-1 levels. Bradykinin administration activated B2R in the spinal cord of SCII rats, which may improve hind limb locomotor recovery by regulating the NF-κB signaling pathway to inhibit the inflammatory response. These findings may provide a theoretical basis for the clinical application of bradykinin in SCII.

Hereditary angioedema due to C1 inhibitor deficiency: real-world experience from the Icatibant Outcome Survey in Spain.

BACKGROUND: The Icatibant Outcome Survey (IOS) is an international registry monitoring the use of icatibant, a bradykinin B2 receptor antagonist indicated for the acute treatment of hereditary angioedema (HAE) attacks. Our goal was to assess disease characteristics and icatibant treatment outcomes in patients with HAE due to C1 inhibitor deficiency (HAE type 1 or 2 (HAE-1/2)) from Spain relative to other countries participating in IOS. METHODS: Descriptive retrospective analyses of data are reported from 10 centers in Spain vs 51 centers in 12 other participating countries (July 2009 to January 2019). RESULTS: No meaningful differences were identified between patients in Spain (n = 119) and patients across other countries (n = 907) regarding median age at symptom onset (15.0 vs 12.0 years) or diagnosis (22.3 vs 20.5 years). Overall HAE attack rates (total attacks/total years of follow-up) were 2.66 in Spain and 1.46 across other countries. Patients in Spain reported fewer severe/very severe HAE attacks before treatment (41.0% vs 45.9%; P < 0.0001) and, for icatibant-treated attacks, longer median time to treatment (2.9 vs 1.0 h), time to attack resolution (18.0 vs 5.5 h), and total attack duration (24.6 vs 8.0 h). Use of androgens for long-term prophylaxis was higher in Spain (51.2% vs 26.7%). CONCLUSION: Patients with HAE-1/2 in Spain reported fewer severe/very severe attacks, administered icatibant later, and had longer-lasting attacks than did patients across other countries in IOS. These differences may indicate varying disease management practices (e.g., delayed icatibant treatment) and reporting. Efforts to raise awareness on the benefits of early on-demand treatment may be warranted. TRIAL REGISTRATION: NCT01034969.

Polymorphism of the bradykinin type 2 receptor gene modulates blood pressure profile and microvascular function in prepubescent children.

Previous reports reveal that +9/-9 polymorphism of the bradykinin B2 receptor (BDKRB2) is suggestive of cardiometabolic diseases. The aim of this study was to examine the impact of BDKRB2 + 9/-9 polymorphism genotypes on the blood pressure parameters and microvascular function in prepubescent children. We screened for BDKRB2 + 9/-9 polymorphism in the DNA of 145 children (86 boys and 59 girls), and its association with body composition, blood pressure levels, biochemical parameters, and endothelial function was determined. No significant association of the BDKRB2 genotypes with gender (P=0.377), race (P=0.949) or family history of cardiovascular disease (CVD) (P=0.858) was observed. Moreover, we did not identify any interaction between BDKRB2 genotypes with a phenotype of obesity (P=0.144). Children carrying the +9/+9 genotype exhibited a significant linear trend with higher levels of systolic blood pressure and pulse pressure (P<0.001). Moreover, the presence of +9 allele resulted in a decrease of reactive hyperemia index, showing a decreasing linear trend from -9/-9 to +9/+9, wherein this parameter of endothelial function was the lowest in the +9/+9 children, intermediate in the +9/-9 children, and the highest in the -9/-9 children (P<0.001). There was a significant inverse correlation between reactive hyperemia index and systolic blood pressure (r= - 0.348, P< 0.001) and pulse pressure (r= - 0.399, P< 0.001). Our findings indicate that the +9/+9 BDKRB2 genotype was associated with high blood pressure and microvascular dysfunction in prepubescent Brazilian children.

Survey of actual conditions of erythema marginatum as a prodromal symptom in Japanese patients with hereditary angioedema.

BACKGROUND: Hereditary angioedema (HAE) is a rare but life-threatening condition. HAE types I and II (HAE-1/2) result from C1-inhibitor (C1-INH) deficiency. However, recent genetic analysis has established a new type of HAE with normal C1-INH (HAEnC1-INH). The mutations of factor XII, plasminogen, angiopoietin 1, and kininogen 1 genes may be the cause of HAEnC1-INH. Nevertheless, other causative molecules (HAE-unknown) may be involved. The Japanese therapeutic environment for HAE has been improving owing to the self-subcutaneous injection of icatibant, which was approved for the treatment of acute attack and enables early therapy. Erythema marginatum (EM) is a visible prodromal symptom which occasionally occurs prior to an angioedema attack; hence, recognizing the risk of an acute attack is important for early treatment. However, the detailed characteristics of EM remain unclear. In this study, we first investigated the clinical manifestations of EM in Japanese patients with HAE. METHODS: A 20-point survey was developed and distributed to 40 physicians to gather clinical data on EM from patients with HAE. RESULTS: Data on 68 patients with HAE (58 patients with HAE-1/2 and 10 patients with HAE-unknown) were collected. Of the patients with HAE-1/2, 53.4% experienced EM, whereas 43.1% did not. The forearm was the most frequent area of EM (64.5%), followed by the abdomen (29.0%) and upper arm and precordium (19.3%). Of the HAE-1/2 patients with EM, 41.9% always had angioedema following EM, while 29.0% always had colocalization of EM with angioedema. Moreover, 3.2% showed a correlation between the awareness of EM and severity of an angioedema attack. In 60.9% of HAE-1/2 patients with EM, the interval between the awareness of EM and appearance of angioedema was <3 h. Of the patients with HAE-unknown, 30.0% also experienced EM. CONCLUSION: We confirmed that more than one-half of Japanese patients with HAE-1/2 and one-third of those with HAE-unknown develop EM as the prodromal symptom of an angioedema attack. Physicians should communicate the significance of EM to patients with HAE to prepare them for possible imminent attacks.