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Immunology and Allergy Clinics of North... Aug 2024The role of contact system activation has been clearly established in the pathogenesis of hereditary angioedema due to C1 inhibitor deficiency (HAE-C1INH). C1 inhibitor... (Review)
Review
The role of contact system activation has been clearly established in the pathogenesis of hereditary angioedema due to C1 inhibitor deficiency (HAE-C1INH). C1 inhibitor (C1INH)-protease complexes, levels of functional C1INH, plasma kallikrein activation, and cleavage of high-molecular-weight kininogen have each been associated with disease activity. More recently, HAE with normal levels of C1INH (HAE-nl-C1INH) has been recognized. Six genetic mutations have been identified which are linked to HAE-nl-C1INH phenotypes. The majority of individuals with HAE-nl-C1INH fall into the unknown category. There is substantial evidence that bradykinin generation underlies the recurrent attacks of swelling in some of these cohorts.
Topics: Humans; Bradykinin; Biomarkers; Complement C1 Inhibitor Protein; Angioedema; Angioedemas, Hereditary; Mutation
PubMed: 38937015
DOI: 10.1016/j.iac.2024.03.011 -
Toxins Jun 2024Despite their evolutionary novelty, lizard venoms are much less studied in comparison to the intense research on snake venoms. While the venoms of helodermatid lizards...
Despite their evolutionary novelty, lizard venoms are much less studied in comparison to the intense research on snake venoms. While the venoms of helodermatid lizards have long been assumed to be for defensive purposes, there is increasing evidence of toxic activities more useful for predation than defence (such as paralytic neurotoxicity). This study aimed to ascertain the effects of , , and lizard venoms on the coagulation and cardiovascular systems. Anticoagulant toxicity was demonstrated for the species studied, with the venoms prolonging clotting times in human and bird plasma due to the destructive cleavage of fibrinogen. In contrast, thromboelastographic analyses on human and bird plasmas in this study demonstrated a procoagulant bioactivity for venoms. A previous study on venom using factor-depleted plasmas as a proxy model suggested a procoagulant factor was present that activated either Factor XI or Factor XII, but could not ascertain the precise target. Our activation studies using purified zymogens confirmed FXII activation. Comparisons of neonate and adult , revealed the neonates to be more potent in the ability to activate FXII, being more similar to the venom of the smaller species than the adult This suggests potent FXII activation a basal trait in the genus, present in the small bodied last common ancestor. This also indicates an ontogenetic difference in prey preferences in the larger species paralleing the change in venom biochemistry. In addition, as birds lack Factor XII, the ability to clot avian plasma suggested an additional procoagulant site of action, which was revealed to be the activation of Factor VII, with being the most potent. This study also examined the effects upon the cardiovascular system, including the liberation of kinins from kininogen, which contributes to hypotension induction. This form of toxicity was previously described for venoms, and was revealed in this study was to also be a pathophysiological effect of and venoms. This suggests that this toxic activity was present in the venom of the last common ancestor of the anguimorph lizards, which is consistent with kallikrein enzymes being a shared toxin trait. This study therefore uncovered novel actions of anguimorph lizard venoms, not only contributing to the evolutionary biology body of knowledge but also revealing novel activities to mine for drug design lead compounds.
Topics: Animals; Lizards; Blood Coagulation; Humans; Anticoagulants; Birds; Venoms; Cardiotoxins; Thrombelastography; Cardiotoxicity
PubMed: 38922177
DOI: 10.3390/toxins16060283 -
Journal of Thrombosis and Haemostasis :... Jun 2024Factor XII (FXII) triggers contact activation by binding to foreign surfaces, with the EGF-1 domain being the primary binding site. Blocking FXII surface-binding might...
BACKGROUND
Factor XII (FXII) triggers contact activation by binding to foreign surfaces, with the EGF-1 domain being the primary binding site. Blocking FXII surface-binding might hold therapeutic value to prevent medical device-induced thrombosis.
OBJECTIVES
To unravel and prevent EGF-1 mediated FXII surface-binding with VH.
METHODS
FXII variants with glutamine substitutions of two positively-charged amino acid patches within the EGF-1 domain were created. Their role in FXII contact activation was assessed using kaolin pull-down experiments, amidolytic activity assays, and clotting assays. FXII EGF-1 domain specific VHHs were raised to inhibit EGF-1 mediated FXII contact activation while preserving quiescence.
RESULTS
Two unique, positively-charged patches in the EGF1 domain were identified (upstream: 73K74K76K78H81K82H; downstream: 87K113K). Neutralizing the charge of both patches led to a 99% reduction in FXII kaolin binding, subsequent decrease in auto-activation of 94% and prolongation of clot formation in aPTT assays from 36 (±2) to 223 (±13) seconds. Three FXII EGF-1 specific VHHs were developed, that are capable of inhibiting kaolin binding and subsequent contact system activation in plasma. The most effective V HH 'F2' binds the positively-charged patches and thereby dose-dependently extends aPTT clotting times from 29 (±2) to 43 (±3) seconds without disrupting FXII quiescence.
CONCLUSION
The two unique, positively-charged patches in FXII EGF-1 cooperatively mediate FXII surface-binding making both patches crucial for contact activation. Targeting these with FXII EGF-1 specific VHHs can exclusively decrease FXII surface-binding and subsequent contact activation, while preserving zymogen quiescence. These patches thus have potential as druggable target in preventing medical device-induced thrombosis.
PubMed: 38897387
DOI: 10.1016/j.jtha.2024.06.005 -
British Journal of Pharmacology Jun 2024Psoriasis is an autoimmune inflammatory skin disease, featuring microvascular abnormalities and elevated levels of bradykinin. Contact activation of Factor XII can...
BACKGROUND AND PURPOSE
Psoriasis is an autoimmune inflammatory skin disease, featuring microvascular abnormalities and elevated levels of bradykinin. Contact activation of Factor XII can initiate the plasma kallikrein-kinin cascade, producing inflammation and angioedema. The role of Factor XII in psoriasis is unknown.
EXPERIMENTAL APPROACH
The effects of deficiency of Factor XII or its enzymatic substrate, prekallikrein, were examined in the imiquimod-induced mouse model of psoriasis. Skin microcirculation was assessed using intravital confocal microscopy and laser Doppler flowmeter. A novel antibody blocking Factor XII activation was evaluated for psoriasis prevention.
KEY RESULTS
Expression of Factor XII was markedly up-regulated in human and mouse psoriatic skin. Genetic deletion of Factor XII or prekallikrein, attenuated imiquimod-induced psoriatic lesions in mice. Psoriatic induction increased skin microvascular blood perfusion, causing vasodilation, hyperpermeability and angiogenesis. It also promoted neutrophil-vascular interaction, inflammatory cytokine release and enhanced Factor XII / prekallikrein enzymatic activity with elevated bradykinin. Factor XII or prekallikrein deficiency ameliorated these microvascular abnormalities and abolished bradykinin increase. Antagonism of bradykinin B receptors reproduced the microvascular protection of Factor XII / prekallikrein deficiency, attenuated psoriatic lesions, and prevented protection by Factor XII / prekallikrein deficiency against psoriasis. Furthermore, treatment of mice with Factor XII antibody alleviated experimentally induced psoriasis and suppressed microvascular inflammation.
CONCLUSION AND IMPLICATIONS
Activation of Factor XII promoted psoriasis via prekallikrein-dependent formation of bradykinin, which critically mediated psoriatic microvascular inflammation. Inhibition of contact activation represents a novel therapeutic strategy for psoriasis.
PubMed: 38872396
DOI: 10.1111/bph.16428 -
Journal of Neuroinflammation Jun 2024Activation of the kallikrein-kinin system promotes vascular leakage, inflammation, and neurodegeneration in ischemic stroke. Inhibition of plasma kallikrein (PK) - a key...
Activation of the kallikrein-kinin system promotes vascular leakage, inflammation, and neurodegeneration in ischemic stroke. Inhibition of plasma kallikrein (PK) - a key component of the KKS - in the acute phase of ischemic stroke has been reported to reduce thrombosis, inflammation, and damage to the blood-brain barrier. However, the role of PK during the recovery phase after cerebral ischemia is unknown. To this end, we evaluated the effect of subacute PK inhibition starting from day 3 on the recovery process after transient middle artery occlusion (tMCAO). Our study demonstrated a protective effect of PK inhibition by reducing infarct volume and improving functional outcome at day 7 after tMCAO. In addition, we observed reduced thrombus formation in cerebral microvessels, fewer infiltrated immune cells, and an improvement in blood-brain barrier integrity. This protective effect was facilitated by promoting tight junction reintegration, reducing detrimental matrix metalloproteinases, and upregulating regenerative angiogenic markers. Our findings suggest that PK inhibition in the subacute phase might be a promising approach to accelerate the post-stroke recovery process.
Topics: Animals; Recovery of Function; Male; Plasma Kallikrein; Mice; Mice, Inbred C57BL; Infarction, Middle Cerebral Artery; Blood-Brain Barrier; Stroke; Thrombosis; Ischemic Stroke; Inflammation
PubMed: 38872149
DOI: 10.1186/s12974-024-03149-w -
Cell Biochemistry and Biophysics Jun 2024Human plasma kallikrein (PKa) is a member of the serine protease family and serves as a key mediator of the kallikrein-kinin system (KKS), which is known for its...
Unveiling therapeutic frontiers: DON/DRP-104 as innovative Plasma kallikrein inhibitors against carcinoma-associated hereditary angioedema shocks - a comprehensive molecular dynamics exploration.
Human plasma kallikrein (PKa) is a member of the serine protease family and serves as a key mediator of the kallikrein-kinin system (KKS), which is known for its regulatory roles in inflammation, vasodilation, blood pressure, and coagulation. Genetic dysregulation of KKS leads to Hereditary Angioedema (HAE), which is characterized by spontaneous, painful swelling in various body regions. Importantly, HAE frequently coexists with various cancers. Despite substantial efforts towards the development of PKa inhibitors for HAE, there remains a need for bifunctional agents addressing both anti-cancer and anti-HAE aspects, especially against carcinoma-associated comorbid HAE conditions. Consequently, we investigated the therapeutic potential of the anti-glutamine prodrug, isopropyl(S)-2-((S)-2-acetamido-3-(1H-indol-3-yl)-propanamido)-6-diazo-5-oxo-hexanoate (DRP-104), and its active form, 6-Diazo-5-oxo-l-norleucine (DON), recognized for their anti-cancer properties, as novel PKa inhibitors. Utilizing structure-based in silico methods, we conducted a comparative analysis with berotralstat, a clinically approved HAE prophylactic, and sebetralstat, an investigational HAE therapeutic agent, in Phase 3 clinical trials. Inhibiting PKa with DON resulted in relatively heightened structural stability, rigidity, restricted protein folding, and solvent-accessible loop exposure, contributing to increased intra-atomic hydrogen bond formation. Conversely, PKa inhibition with DRP-104 induced restricted residue flexibility and significantly disrupted the critical SER195-HIS57 arrangement in the catalytic triad. Both DON and DRP-104, along with the reference drugs, induced strong cooperative intra-residue motion and bidirectional displacement in the PKa architecture. The results revealed favorable binding kinetics of DON/DRP-104, showing thermodynamic profiles that were either superior or comparable to those of the reference drugs. These findings support their consideration for clinical investigations into the management of carcinoma-associated HAE.
PubMed: 38869687
DOI: 10.1007/s12013-024-01266-0 -
Journal of Biotechnology Jun 2024Ecallantide comprises Kunitz Domain 1 of Tissue Factor Pathway Inhibitor, mutated at seven amino acid positions to inhibit plasma kallikrein (PK). It is used to treat...
Ecallantide comprises Kunitz Domain 1 of Tissue Factor Pathway Inhibitor, mutated at seven amino acid positions to inhibit plasma kallikrein (PK). It is used to treat acute hereditary angioedema (HAE). We appended hexahistidine tags to the N- or C-terminus of recombinant Ecallantide (rEcall) and expressed and purified the resulting proteins, with or without fusion to human serum albumin (HSA), using Pichia pastoris. The inhibitory constant (K) of rEcall-H6 or H6-rEcall for PK was not increased by albumin fusion. When I-labelled rEcall proteins were injected intravenously into mice, the area under the clearance curve (AUC) was significantly increased, 3.4- and 3.6-fold, for fusion proteins H6-rEcall-HSA and HSA-rEcall-H6 versus their unfused counterparts but remained 2- to 3-fold less than that of HSA-H6. The terminal half-life of H6-rEcall-HSA and HSA-H6 did not differ, although that of HSA-rEcall-H6 was significantly shorter than either other protein. Receptor Associated Protein (RAP), a Low-density lipoprotein Receptor-related Protein (LRP1) antagonist, competed H6-rEcall-HSA clearance more effectively than intravenous immunoglobulin (IVIg), a neonatal Fc receptor (FcRn) antagonist. HSA fusion decreases rEcall clearance in vivo, but LRP1-mediated clearance remains more important than FcRn-mediated recycling for rEcall fusion proteins. The properties of H6-rEcall-HSA warrant investigation in a murine model of HAE.
PubMed: 38844246
DOI: 10.1016/j.jbiotec.2024.06.002 -
Journal of Pharmacokinetics and... May 2024Hereditary angioedema (HAE) due to C1-inhibitor deficiency is a rare, debilitating, genetic disorder characterized by recurrent, unpredictable, attacks of edema. The...
Hereditary angioedema (HAE) due to C1-inhibitor deficiency is a rare, debilitating, genetic disorder characterized by recurrent, unpredictable, attacks of edema. The clinical symptoms of HAE arise from excess bradykinin generation due to dysregulation of the plasma kallikrein-kinin system (KKS). A quantitative systems pharmacology (QSP) model that mechanistically describes the KKS and its role in HAE pathophysiology was developed based on HAE attacks being triggered by autoactivation of factor XII (FXII) to activated FXII (FXIIa), resulting in kallikrein production from prekallikrein. A base pharmacodynamic model was constructed and parameterized from literature data and ex vivo assays measuring inhibition of kallikrein activity in plasma of HAE patients or healthy volunteers who received lanadelumab. HAE attacks were simulated using a virtual patient population, with attacks recorded when systemic bradykinin levels exceeded 20 pM. The model was validated by comparing the simulations to observations from lanadelumab and plasma-derived C1-inhibitor clinical trials. The model was then applied to analyze the impact of nonadherence to a daily oral preventive therapy; simulations showed a correlation between the number of missed doses per month and reduced drug effectiveness. The impact of reducing lanadelumab dosing frequency from 300 mg every 2 weeks (Q2W) to every 4 weeks (Q4W) was also examined and showed that while attack rates with Q4W dosing were substantially reduced, the extent of reduction was greater with Q2W dosing. Overall, the QSP model showed good agreement with clinical data and could be used for hypothesis testing and outcome predictions.
PubMed: 38734778
DOI: 10.1007/s10928-024-09919-6 -
Annals of Allergy, Asthma & Immunology... Apr 2024Hereditary angioedema (HAE) is a rare genetic condition causing unpredictable and severe episodes of angioedema that are debilitating and life-threatening. Moreover, HAE... (Review)
Review
Hereditary angioedema (HAE) is a rare genetic condition causing unpredictable and severe episodes of angioedema that are debilitating and life-threatening. Moreover, HAE can be classified into HAE due to C1-esterase inhibitor deficiency (HAE-C1INH) or HAE with normal C1INH. Moreover, HAE-C1INH is subcategorized as types I and II based on deficient or dysfunctional circulating C1INH protein resulting from inherited or spontaneous mutations in the SERPING1 gene leading to uncontrolled factor XII/plasma kallikrein activation and excessive bradykinin production. Bradykinin-2 receptor activation leads to vasodilation, increased vascular permeability, and smooth muscle contractions, resulting in subcutaneous or submucosal fluid extravasation that can affect the face, extremities, airway, and gastrointestinal and genitourinary systems. Furthermore, HAE with normal C1INH is caused by either a known or unknown genetic mutation, and the mechanisms are less well-established but most forms are thought to be related to bradykinin signaling with a similar presentation as HAE-C1INH despite normal levels of C1INH protein and function. Current HAE management strategies include on-demand and prophylactic treatments which replace C1INH, reduce kallikrein activity, or block bradykinin binding to the bradykinin B2 receptor. With the advent of additional small molecule inhibitors, monoclonal antibodies, RNA-targeted therapies, gene therapies, and gene modification approaches, preclinical studies and human clinical trials are underway to further expand therapeutic options in HAE. This review article will briefly summarize current HAE treatments and provide an overview of potential future therapies for HAE.
PubMed: 38679158
DOI: 10.1016/j.anai.2024.04.029 -
Clinica Chimica Acta; International... Jun 2024Using lectins to target cancer-associated modifications of PSA glycostructure for identification of clinically significant prostate cancers, e.g., Gleason score...
BACKGROUND
Using lectins to target cancer-associated modifications of PSA glycostructure for identification of clinically significant prostate cancers, e.g., Gleason score (GS) ≥ 7, from benign and indolent cancers (GS 6), is highly promising yet technically challenging. From previous findings to quantify increased PSA fucosylation in urine, we set out to construct a robust, specific test concept suitable for plasma samples.
METHODS
Macrophage galactose-binding lectin (MGL) coupled to 100 nm Eu3 + -nanoparticles was used to probe PSA captured from cancer cell lines, seminal plasma, and plasma samples from 249 patients with a clinical suspicion of prostate cancer onto 3 mm dense spots of free PSA antibody fab fragments. Results were compared to four kallikrein tests: tPSA, fPSA, iPSA and hK2.
RESULTS
The fPSAglycovariant provided superior discrimination of the GS ≥ 7 and benign + GS 6 groups (p 0.0003) compared to fPSA (NS). The corresponding AUC in ROC analysis was 0.70 compared to 0.66 for tPSA. In contrast to all four kallikrein tests, the fPSAGV was independent of prostate gland volume. Using a logistic regression analysis the fPSAGV significantly improved on the four-kallikrein model.
CONCLUSIONS
Due to Eu-nanoparticles and a dense fPSA capture spot, the fPSA glycovariant identifies an fPSA subform with the highest cancer specificity compared to the four conventional kallikreins.
Topics: Humans; Male; Prostate-Specific Antigen; Prostatic Neoplasms; Nanoparticles; Lectins; Aged; Glycosylation
PubMed: 38677453
DOI: 10.1016/j.cca.2024.119689