Verwimp Sam, Wagoner Jessica, Arenas Elijah Gabriela, De Coninck Lander, Abdelnabi Rana, Hyde Jennifer L, Schiffer Joshua T, White Judith M, Matthijnssens Jelle, Neyts Johan, Polyak Stephen J, Delang Leen
bioRxiv. 2025 Mar 11:2025.01.24.633564. doi: 10.1101/2025.01.24.633564.
Alphaviruses, including chikungunya virus (CHIKV), pose a significant global health threat, yet specific antiviral therapies remain unavailable.
We evaluated combinations of three oral directly acting antiviral drugs (sofosbuvir (SOF), molnupiravir (MPV), and favipiravir (FAV)), which are approved for other indications, against CHIKV, Semliki Forest virus (SFV), Sindbis virus (SINV), and Venezuelan Equine Encephalitis virus (VEEV) and . We assessed antiviral efficacy in human skin fibroblasts and liver cells, as well as in a mouse model of CHIKV-induced arthritis.
In human skin fibroblasts, synergistic antiviral effects were observed for combinations of MPV + SOF and FAV + SOF against CHIKV, and for FAV + SOF against SFV. In human liver cells, FAV + MPV conferred additive to synergistic activity against VEEV and SINV, while SOF synergized with FAV against SINV. In mice, MPV improved CHIKV-induced foot swelling and reduced systemic infectious virus titres. Combination treatment with MPV and SOF significantly reduced swelling and infectious virus titres compared to monotherapies of each drug. Sequencing of CHIKV RNA from joint tissue revealed that MPV caused dose- dependent increases in mutations in the CHIKV genome. Upon combination therapy of MPV with SOF, the number of mutations was significantly lower compared to monotherapy with several higher doses of MPV.
Combining these approved oral nucleoside analogues confers potent suppression of multiple alphaviruses and with enhanced control of viral genetic evolution in face of antiviral pressure. These drug combinations may ultimately lead to the development of potent combinations of pan-family alphavirus inhibitors.
This work was supported by a PhD fellowship granted to S.V. by the Research Foundation - Flanders (FWO) (11D5923N). L.D.C. was also supported by Research Foundation - Flanders (FWO) PhD fellowship (11L1325N). Dr. Polyak and Schiffer are partially supported by R01AI121129.
Alphaviruses such as chikungunya virus (CHIKV), Sindbis virus (SINV), and Venezuelan Equine Encephalitis virus (VEEV) are a major threat for global health. Alphaviruses are responsible for debilitating diseases with major public health implications, yet no antiviral drugs are currently approved for treating these virus infections. Existing treatment options are limited to supportive care and are unlikely to provide protection against future outbreaks of other alphaviruses. Previous studies have shown that oral approved nucleoside analogues such as favipiravir (FAV), molnupiravir (MPV), and sofosbuvir (SOF) have antiviral activity against certain RNA viruses, including alphaviruses. However, systematic evaluations of these drugs and testing of drug combinations in both and settings are limited. This study provides a comprehensive evaluation of combinations of FAV, MPV and SOF against multiple alphaviruses in two human cell lines and a CHIKV mouse model. We demonstrate that certain combinations of these drugs confer synergistic antiviral activity, effectively suppressing CHIKV, SFV, SINV, and VEEV replication . Moreover, , we show for the first time that MPV treatment results in reduced CHIKV-induced foot swelling and systemic virus replication. Combining MPV with SOF enhances antiviral activity in mice as compared to monotherapy. By sequencing the viral genome, we show that MPV increases the number of mutations in a dose-dependent manner. Combination therapy of MPV and SOF reduces the number of mutations compared to higher doses of MPV. These findings highlight the potential of nucleoside analogue combinations as a promising therapeutic strategy against alphavirus infections. The results of this study suggest that combination therapy with approved nucleoside analogues could provide an effective treatment strategy for alphavirus infections. The observed increased efficacy of drug combinations supports the potential for dose optimization to enhance efficacy while reducing toxicity and development of resistance. Future research should focus on clinical evaluation of these drug combinations, pharmacokinetic studies, and further exploration of their impact on viral evolution. Given the expanding geographical distribution of alphaviruses and the lack of available treatments, these findings provide a foundation for developing pan-alphavirus antiviral therapies that could improve patient outcomes and global outbreak preparedness.
包括基孔肯雅病毒（CHIKV）在内的甲病毒对全球健康构成重大威胁，但目前仍没有特效抗病毒疗法。
我们评估了三种已获其他适应症批准的口服直接作用抗病毒药物（索磷布韦（SOF）、莫努匹韦（MPV）和法匹拉韦（FAV））对CHIKV、塞姆利基森林病毒（SFV）、辛德毕斯病毒（SINV）和委内瑞拉马脑炎病毒（VEEV）的联合作用。我们在人皮肤成纤维细胞和肝细胞以及CHIKV诱导的关节炎小鼠模型中评估了抗病毒效果。
在人皮肤成纤维细胞中，观察到MPV + SOF和FAV + SOF联合用药对CHIKV有协同抗病毒作用，FAV + SOF对SFV有协同抗病毒作用。在人肝细胞中，FAV + MPV对VEEV和SINV具有相加至协同活性，而SOF与FAV对SINV具有协同作用。在小鼠中，MPV改善了CHIKV诱导的足部肿胀并降低了全身感染性病毒滴度。与每种药物的单药治疗相比，MPV和SOF联合治疗显著减轻了肿胀并降低了感染性病毒滴度。对关节组织中的CHIKV RNA进行测序发现，MPV导致CHIKV基因组中的突变呈剂量依赖性增加。与使用几种较高剂量MPV的单药治疗相比，MPV与SOF联合治疗时的突变数量显著更低。
联合使用这些已获批准的口服核苷类似物可有效抑制多种甲病毒，并在抗病毒压力下增强对病毒基因进化的控制。这些药物组合最终可能会促成强效的泛甲病毒抑制剂组合的开发。
这项工作得到了弗拉芒研究基金会（FWO）授予S.V.的博士奖学金（11D5923N）的支持。L.D.C.也得到了弗拉芒研究基金会（FWO）博士奖学金（11L1325N）的支持。Polyak博士和Schiffer博士部分得到了R01AI121129的支持。
基孔肯雅病毒（CHIKV）、辛德毕斯病毒（SINV）和委内瑞拉马脑炎病毒（VEEV）等甲病毒是全球健康的主要威胁。甲病毒可导致使人衰弱的疾病，具有重大的公共卫生影响，但目前尚无抗病毒药物被批准用于治疗这些病毒感染。现有的治疗选择仅限于支持性护理，不太可能预防未来其他甲病毒的爆发。先前的研究表明，已获批准的口服核苷类似物，如法匹拉韦（FAV）和莫努匹韦（MPV）以及索磷布韦（SOF），对某些RNA病毒具有抗病毒活性，包括甲病毒。然而，对这些药物的系统评估以及在体内和体外环境中对药物组合的测试都很有限。 本研究全面评估了FAV、MPV和SOF联合用药在两种人类细胞系和CHIKV小鼠模型中对多种甲病毒的作用。我们证明，这些药物的某些组合具有协同抗病毒活性，可有效抑制CHIKV、SFV、SINV和VEEV的复制。此外，我们首次表明，MPV治疗可减轻CHIKV诱导的足部肿胀和全身病毒复制。与单药治疗相比，MPV与SOF联合使用可增强小鼠的抗病毒活性。通过对病毒基因组进行测序，我们表明MPV以剂量依赖性方式增加突变数量。与较高剂量的MPV相比，MPV和SOF联合治疗可减少突变数量。这些发现突出了核苷类似物组合作为一种有前景的抗甲病毒感染治疗策略的潜力。 本研究结果表明，使用已获批准的核苷类似物进行联合治疗可为甲病毒感染提供一种有效的治疗策略。观察到的药物组合疗效提高支持了通过剂量优化来提高疗效同时降低毒性和耐药性发展的可能性。未来的研究应侧重于对这些药物组合的临床评估、药代动力学研究以及进一步探索它们对病毒进化的影响。鉴于甲病毒的地理分布不断扩大且缺乏可用的治疗方法，这些发现为开发可改善患者预后和全球疫情防范能力的泛甲病毒抗病毒疗法奠定了基础。
