Harisha Kalenahalli Rajappa, Kailaje Varun, Kondreddi Ravinder Reddy, Gudla Chandra Sekhar, Singh Shraddha, Ramakrishnaiah Sharada, Isloor Shrikrishna, Narayanan Shridhar, Shandil Radha Krishan, Rudramurthy Gudepalya Renukaiah
Foundation for Neglected Disease Research (FNDR), Bengaluru 561203, India.
Veterinary College, Karnataka Veterinary, Animal and Fisheries Sciences University (KVAFSU), Hebbal, Bengaluru 560024, India.
Viruses. 2025 Jul 4;17(7):945. doi: 10.3390/v17070945.
Rabies, a viral encephalitis caused by rabies virus (RABV), is 100% fatal upon the onset of symptoms. Effective post-exposure prophylaxis (PEP) measures are available, but they are often difficult to access in low-income countries. WHO estimates about 59,000 deaths due to rabies globally, and the majority are contributed by developing countries. Hence, developing drugs for the treatment of post-symptomatic rabies is an urgent and unmet demand. It is worth noting that previous efforts regarding antiviral strategies, such as small-interfering RNA, antibodies and small-molecule inhibitors, against the rabies virus have failed to show efficacy in pre-clinical studies, especially when the virus has reached the central nervous system (CNS). Therefore, drug repurposing seems to be an alternative tool for the development of new anti-rabies drugs. We validated and used a high-throughput, FITC-conjugated antibody-based flow cytometry assay to expedite the identification of repurposable new drug candidates against the RABV. The assay was validated using ribavirin and salinomycin as reference compounds, which showed EC50 values of 10.08 µM and 0.07 µM, respectively. We screened a SelleckChem library comprising 3035 FDA-approved compounds against RABV (CVS-11) at 10 µM concentration. Five compounds (clofazimine, tiamulin, difloxacin, harringtonine and homoharringtonine) were active against RABV, with greater than 90% inhibition. Homoharringtonine (HHT) identified in the present study is active against laboratory-adapted RABV (CVS-11) and clinical isolates of RABV, with an average EC50 of 0.3 µM in both BHK-21 and Neuro-2a cell lines and exhibits post-entry inhibition.
狂犬病是由狂犬病病毒(RABV)引起的病毒性脑炎,一旦出现症状,致死率为100%。虽然有有效的暴露后预防(PEP)措施,但在低收入国家往往难以获得。世界卫生组织估计,全球每年约有59000人死于狂犬病,其中大部分来自发展中国家。因此,开发治疗症状出现后狂犬病的药物是一项迫切且未得到满足的需求。值得注意的是,此前针对狂犬病病毒的抗病毒策略,如小分子干扰RNA、抗体和小分子抑制剂,在临床前研究中均未显示出疗效,尤其是当病毒已到达中枢神经系统(CNS)时。因此,药物再利用似乎是开发新型抗狂犬病药物的一种替代工具。我们验证并使用了一种基于异硫氰酸荧光素(FITC)偶联抗体的高通量流式细胞术检测方法,以加速鉴定可再利用的抗RABV新药候选物。该检测方法使用利巴韦林和沙利霉素作为参考化合物进行了验证,它们的半数有效浓度(EC50)值分别为10.08 μM和0.07 μM。我们以10 μM的浓度筛选了一个包含3035种美国食品药品监督管理局(FDA)批准化合物的SelleckChem文库,以检测其对RABV(CVS-11)的作用。有五种化合物(氯法齐明、泰妙菌素、双氟沙星、高三尖杉酯碱和高同型半胱氨酸)对RABV有活性,抑制率大于90%。在本研究中鉴定出的高三尖杉酯碱(HHT)对实验室适应的RABV(CVS-11)和RABV临床分离株均有活性,在BHK-21和Neuro-2a细胞系中的平均EC50均为0.3 μM,并表现出病毒进入后的抑制作用。