title: Factor XII full and partial null in rat confers robust antithrombotic efficacy with no bleeding.
abstract: This report aims at exploring quantitatively the relationship between FXII inhibition and thromboprotection. FXII full and partial null in rats were established via zinc finger nuclease-mediated knockout and siRNA-mediated knockdown, respectively. The rats were subsequently characterized in thrombosis and hemostasis models. Knockout rats exhibited complete thromboprotection in both the arteriovenous shunt model (∼100% clot weight reduction) and the FeCl3-induced arterial thrombosis model (no reduction in blood flow), without any increase in cuticle bleeding time compared with wild-type control rats. Ex-vivo aPTT and the ellagic acid-triggered thrombin generation assay (TGA) exhibited anticoagulant changes. In contrast, ex-vivo PT or high tissue factor-triggered TGA was indistinguishable from control. Rats receiving single doses (0, 0.01, 0.03, 0.1, 0.3, 1 mg/kg) of FXII siRNA exhibited dose-dependent knockdown in liver FXII mRNA and plasma FXII protein (95 and 99%, respectively, at 1 mg/kg) at day 7 post dosing. FXII knockdown was associated with dose-dependent thromboprotection (maximal efficacy achieved with 1 mg/kg in both models) and negligible change in cuticle bleeding times. Ex-vivo TGA triggered with low-level (0.5 μmol/l) ellagic acid tracked best with the knockdown levels and efficacy. Our findings confirm and extend literature reports of an attractive benefit-to-risk profile of targeting FXII for antithrombotic therapies. Titrating of FXII is instructive for its pharmacological inhibition. The knockout rat is valuable for evaluating both mechanism-based safety concerns and off-target effects of FXII(a) inhibitors. Detailed TGA analyses will inform on optimal trigger conditions in studying pharmacodynamic effects of FXII(a) inhibition.
authors.affiliation: aIn Vivo Pharmacology bCardiometabolic Diseases cGenetically Engineered Models, Merck Sharp & Dohme Corp., Kenilworth, New Jersey dRNA Therapeutics; Merck Sharp & Dohme Corp., West Point, Pennsylvania, USA *Current affiliation: Department of Pharmacology, Vanderbilt University, Nashville, Tennessee, USA.
authors.forename: Tian-Quan
authors.initials: TQ
authors.lastname: Cai
authors.forename: Weizhen
authors.initials: W
authors.lastname: Wu
authors.forename: Myung K
authors.initials: MK
authors.lastname: Shin
authors.forename: Yiming
authors.initials: Y
authors.lastname: Xu
authors.forename: Nina
authors.initials: N
authors.lastname: Jochnowitz
authors.forename: Yuchen
authors.initials: Y
authors.lastname: Zhou
authors.forename: Lizbeth
authors.initials: L
authors.lastname: Hoos
authors.forename: Ross
authors.initials: R
authors.lastname: Bentley
authors.forename: Walter
authors.initials: W
authors.lastname: Strapps
authors.forename: Anil
authors.initials: A
authors.lastname: Thankappan
authors.forename: Joseph M
authors.initials: JM
authors.lastname: Metzger
authors.forename: Martin L
authors.initials: ML
authors.lastname: Ogletree
authors.forename: Marija
authors.initials: M
authors.lastname: Tadin-Strapps
authors.forename: Dietmar A
authors.initials: DA
authors.lastname: Seiffert
authors.forename: Zhu
authors.initials: Z
authors.lastname: Chen
mesh_terms: D000818:Animals; D001166:Arteriovenous Shunt, Surgical; D002712:Chlorides / Q000494:pharmacology; D004195:Disease Models, Animal; D004305:Dose-Response Relationship, Drug; D004610:Ellagic Acid / Q000494:pharmacology; D004706:Endodeoxyribonucleases / Q000235:genetics* / Q000378:metabolism; D005174:Factor XII / Q000037:antagonists & inhibitors* / Q000235:genetics / Q000378:metabolism; D005290:Ferric Compounds / Q000494:pharmacology; D055786:Gene Knockout Techniques; D006470:Hemorrhage / Q000517:prevention & control; D006801:Humans; D008099:Liver / Q000378:metabolism; D008297:Male; D010314:Partial Thromboplastin Time; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D034741:RNA, Small Interfering / Q000008:administration & dosage* / Q000235:genetics / Q000378:metabolism; D051381:Rats; D017207:Rats, Sprague-Dawley; D013917:Thrombin / Q000378:metabolism; D015912:Thrombolytic Therapy / Q000379:methods*; D013927:Thrombosis / Q000139:chemically induced / Q000235:genetics / Q000473:pathology / Q000628:therapy*; D016335:Zinc Fingers / Q000235:genetics
chemical_list: D002712:Chlorides; D005290:Ferric Compounds; D012333:RNA, Messenger; D034741:RNA, Small Interfering; D004610:Ellagic Acid; D005174:Factor XII; D004706:Endodeoxyribonucleases; D013917:Thrombin; C024555:ferric chloride