两种老药物，NVP-AEW541 和 GSK-J4，被重新用于对抗刚地弓形虫 RH 株。

BACKGROUND: Toxoplasma gondii is a zoonotic pathogen that causes toxoplasmosis and leads to serious public health problems in developing countries. However, current clinical therapeutic drugs have some disadvantages, such as serious side effects, a long course of treatment and the emergence of drug-resistant strains. The urgent need to identify novel anti-Toxoplasma drugs has initiated the effective strategy of repurposing well-characterized drugs. As a principled screening for the identification of effective compounds against Toxoplasma gondii, in the current study, a collection of 666 compounds were screened for their ability to significantly inhibit Toxoplasma growth. METHODS: The inhibition of parasite growth was determined using a luminescence-based β-galactosidase activity assay. Meanwhile, the effect of compounds on the viability of host cells was measured using CCK8. To assess the inhibition of the selected compounds on discrete steps of the T. gondii lytic cycle, the invasion, intracellular proliferation and egress abilities were evaluated. Finally, a murine infection model of toxoplasmosis was used to monitor the protective efficacy of drugs against acute infection of a highly virulent RH strain. RESULTS: A total of 68 compounds demonstrated more than 70% parasite growth inhibition. After excluding compounds that impaired host cell viability, we further characterized two compounds, NVP-AEW541 and GSK-J4 HCl, which had IC values for parasite growth of 1.17 μM and 2.37 μM, respectively. In addition, both compounds showed low toxicity to the host cell. Furthermore, we demonstrated that NVP-AEW541 inhibits tachyzoite invasion, while GSK-J4 HCl inhibits intracellular tachyzoite proliferation by halting cell cycle progression from G1 to S phase. These findings prompted us to analyse the efficacy of the two compounds in vivo by using established mouse models of acute toxoplasmosis. In addition to prolonging the survival time of mice acutely infected with T. gondii, both compounds had a remarkable ability to reduce the parasite burden of tissues. CONCLUSIONS: Our findings suggest that both NVP-AEW541 and GSK-J4 could be potentially repurposed as candidate drugs against T. gondii infection.

背景：刚地弓形虫是一种人畜共患病原体，可引起弓形体病，并在发展中国家导致严重的公共卫生问题。然而，目前的临床治疗药物存在一些缺点，如严重的副作用、治疗过程长以及耐药菌株的出现。因此，迫切需要寻找新的抗弓形虫药物，这就促使人们采用了重新利用特征明确的药物的有效策略。作为鉴定有效抗弓形虫化合物的原则性筛选方法，在本研究中，使用发光β-半乳糖苷酶活性测定法筛选了 666 种化合物，以确定它们抑制弓形虫生长的能力。

方法：使用发光β-半乳糖苷酶活性测定法测定寄生虫生长的抑制情况。同时，使用 CCK8 测定化合物对宿主细胞活力的影响。为了评估所选化合物对弓形虫裂解周期离散步骤的抑制作用，评估了入侵、细胞内增殖和逸出能力。最后，使用弓形虫感染的小鼠模型监测药物对高毒 RH 株急性感染的保护效果。

结果：共有 68 种化合物表现出超过 70%的寄生虫生长抑制。在排除对宿主细胞活力有损害的化合物后，我们进一步表征了两种化合物，NVP-AEW541 和 GSK-J4 HCl，它们对寄生虫生长的 IC 值分别为 1.17 μM 和 2.37 μM。此外，这两种化合物对宿主细胞的毒性都较低。此外，我们证明 NVP-AEW541 抑制速殖子的入侵，而 GSK-J4 HCl 通过阻止细胞周期从 G1 期到 S 期的进展来抑制细胞内速殖子的增殖。这些发现促使我们使用建立的急性弓形虫感染小鼠模型来分析这两种化合物的体内疗效。除了延长急性感染弓形虫的小鼠的存活时间外，这两种化合物都具有显著降低组织寄生虫负荷的能力。

结论：我们的研究结果表明，NVP-AEW541 和 GSK-J4 均可作为抗弓形虫感染的候选药物重新使用。