BACKGROUND Immune-mediated hemolytic anemia (IMHA) is a common disease that affects all breeds of dogs and is associated with significant morbidity and mortality. Intravascular hemolysis of erythrocytes in IMHA is caused by complement activation and is often fatal. No current treatments target complement activation in canine IMHA. Human C1 esterase (C1-INH) reduces canine complement-mediated hemolysis in vitro, and a recent pharmacokinetic analysis of an FDA licensed formulation of C1-INH in dogs confirmed that a 50 IU/kg dose of C1-INH is safe to administer to dogs, and effectively inhibits canine complement mediated hemolysis ex-vivo. The C1INCH randomized controlled trial will evaluate the efficacy of this drug in dogs with intravascular hemolysis. METHODS We will conduct a multicenter, placebo-controlled double-blind randomized clinical trial of C1-INH in dogs with intravascular hemolysis due to IMHA. We will randomize 18 dogs to receive three doses of intravenous C1-INH or saline in 24 h. Immunosuppressive and antithrombotic therapies will be standardized. Primary outcome measures will be changes in plasma free hemoglobin, serum concentrations of LDH, bilirubin, and haptoglobin. Using patient samples, we will evaluate complement activation in canine IMHA using a novel C5b-9 ELISA assay, flow cytometric detection of C3b on RBC, and by measurement of residual plasma complement activity. Secondary outcome measures will be survival to hospital discharge, duration of hospitalization, number and volume of red blood cell transfusions, and rescue therapy requirements. We will monitor dogs for adverse drug reactions. Sample size was estimated from pilot data on LDH and hemolysis index (HI) in dogs with IMHA. To detect 2-way differences between the upper and lower 50% of the LDH and HI values of equivalent size with 80% power at P < 0.05 will require 9 dogs in each arm. DISCUSSION We anticipate that IV administration of C1-INH will significantly inhibit complement mediated hemolysis in dogs with intravascular IMHA, as determined by blood biomarker measurements (decreased plasma hemoglobin, LDH and bilirubin, increased haptoglobin). We expect this will translate into significant reductions in transfusion requirements and duration of hospitalization. TRIAL REGISTRATION This trial has been prospectively registered with the AVMA registry (AAHSD005025).

中文翻译：

---

犬血管内溶血（C1INCH）中的C1抑制剂：一项随机对照试验的研究方案。

背景技术免疫介导的溶血性贫血（IMHA）是一种影响所有犬种的常见疾病，并且与明显的发病率和死亡率相关。IMHA中红细胞的血管内溶血是由补体激活引起的，通常是致命的。目前尚无针对犬IMHA补体激活的治疗方法。人C1酯酶（C1-INH）可以在体外减少犬补体介导的溶血作用，最近对FDA许可的C1-INH制剂在犬体内进行药代动力学分析，证实50 IU / kg剂量的C1-INH可安全地用于犬，并有效地抑制犬补体介导的体外溶血。C1INCH随机对照试验将评估该药物在血管内溶血犬中的功效。方法我们将进行一个多中心研究，C1-INH的安慰剂对照双盲随机临床试验在IMHA引起的血管内溶血犬中的应用。我们将随机分配18只狗在24小时内接受三剂静脉注射C1-INH或生理盐水。免疫抑制和抗血栓形成疗法将被标准化。主要的预后指标将是血浆游离血红蛋白，LDH，胆红素和触珠蛋白的血清浓度的变化。使用患者样品，我们将使用新型C5b-9 ELISA分析，RBC上的C3b的流式细胞仪检测以及通过测量残留血浆补体活性来评估犬IMHA中的补体激活。次要结果指标是出院存活率，住院时间，红细胞输血数量和量以及抢救治疗的要求。我们将监视狗的药物不良反应。样本量是根据IMHA犬LDH和溶血指数（HI）的初步数据估算得出的。要检测大小相等的LDH和HI值的上限和下限的50％之间的2向差异，且P <0.05时具有80％的功效，每只手臂需要9条狗。讨论我们预计，通过血液生物标志物测定（血浆血红蛋白，LDH和胆红素减少，触珠蛋白增加），静脉注射C1-INH可以显着抑制血管内IMHA犬的补体介导的溶血。我们希望这将大大减少输血需求和住院时间。试验注册该试验已在AVMA注册中心（AAHSD005025）进行了前瞻性注册。要检测大小相等的LDH和HI值的上限和下限的50％之间的2向差异，且P <0.05时具有80％的功效，每只手臂需要9条狗。讨论我们预计，静脉注射C1-INH可以显着抑制血管内IMHA犬的补体介导的溶血，这是通过血液生物标志物测定（血浆血红蛋白，LDH和胆红素减少，触珠蛋白增加）确定的。我们希望这将大大减少输血需求和住院时间。试验注册该试验已在AVMA注册中心（AAHSD005025）进行了前瞻性注册。要检测大小相等的LDH和HI值的上限和下限的50％之间的2向差异，且P <0.05时具有80％的功效，每只手臂需要9条狗。讨论我们预计，静脉注射C1-INH可以显着抑制血管内IMHA犬的补体介导的溶血作用，这是通过血液生物标志物测定确定的（血浆血红蛋白，LDH和胆红素减少，触珠蛋白增加）。我们希望这将大大减少输血需求和住院时间。试验注册该试验已在AVMA注册中心（AAHSD005025）进行了前瞻性注册。讨论我们预计，通过血液生物标志物测定（血浆血红蛋白，LDH和胆红素减少，触珠蛋白增加），静脉注射C1-INH可以显着抑制血管内IMHA犬的补体介导的溶血。我们希望这将大大减少输血需求和住院时间。试验注册该试验已在AVMA注册中心（AAHSD005025）进行了前瞻性注册。讨论我们预计，通过血液生物标志物测定（血浆血红蛋白，LDH和胆红素减少，触珠蛋白增加），静脉注射C1-INH可以显着抑制血管内IMHA犬的补体介导的溶血。我们希望这将大大减少输血需求和住院时间。试验注册该试验已在AVMA注册中心（AAHSD005025）进行了前瞻性注册。