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In this study, we assessed the antiparasitic activity and mechanism of action of milciclib (MCB), a cyclin-dependent kinase inhibitor from the Medicines for Malaria Venture Pathogen Box, against Toxoplasma gondii using in vitro and ex vivo models of the maternal\u2013fetal interface. MCB exhibited no cytotoxic effects in human trophoblastic BeWo cells or placental villous explants at concentrations up to 1.62 \u03bcM. The compound significantly reduced intracellular parasite proliferation in a dose-dependent manner, achieving approximately 39.72, 73.22, and 91.52% of inhibition at 0.40, 0.81, and 1.62 \u03bcM, respectively, and maintained a persistent antiproliferative effect following drug withdrawal. Importantly, MCB also reduced parasite burden in human placental explants (approximately 47.69% of inhibition) while preserving tissue viability and architecture. Mechanistically, MCB induced G1 phase arrest in T. gondii without affecting parasite adhesion or invasion, suggesting a specific action on intracellular replication. Ultrastructural analyses revealed marked morphological alterations in treated parasites, supporting a direct antiparasitic effect. In addition, MCB modulated host cell responses by reducing macrophage migration inhibitory factor (MIF) and reactive oxygen species (ROS) production accompanied by decrease in parasite burden, without triggering a pro-inflammatory profile. In silico analyses indicated potential interactions with parasite cyclin-dependent kinase-related proteins, particularly CRK2. Together, these findings identify MCB as a promising candidate for drug repositioning in congenital toxoplasmosis and support further investigation of its efficacy and safety in vivo.<\/jats:p>","DOI":"10.1021\/acsomega.6c05747","type":"journal-article","created":{"date-parts":[[2026,9,28]],"date-time":"2026-09-28T16:05:18Z","timestamp":1790611518000},"page":"61168-61182","source":"Crossref","is-referenced-by-count":0,"title":["Repurposing of\nMilciclib as a Potential Inhibitor\nof\n                    <i>Toxoplasma gondii<\/i>\n                    in Human Trophoblasts\nand Placental Explants"],"prefix":"10.1021","volume":"11","author":[{"given":"Rafael Martins","family":"de Oliveira","sequence":"first","affiliation":[{"name":"Universidade Federal de Uberla\u0302ndia , , , ,","place":["Uberla\u0302ndia, Minas Gerais, Brazil, 38400-694"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Nata\u0301lia Carine","family":"Lima dos 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38400-694"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Guilherme","family":"de Souza","sequence":"additional","affiliation":[{"name":"Universidade Federal de Uberla\u0302ndia , , , ,","place":["Uberla\u0302ndia, Minas Gerais, Brazil, 38400-694"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Izadora","family":"Santos Damasceno","sequence":"additional","affiliation":[{"name":"Universidade Federal de Uberla\u0302ndia , , , ,","place":["Uberla\u0302ndia, Minas Gerais, Brazil, 38400-694"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Muriel Pereira","family":"Souto","sequence":"additional","affiliation":[{"name":"Universidade Federal de Uberla\u0302ndia , , , ,","place":["Uberla\u0302ndia, Minas Gerais, Brazil, 38400-694"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Rosiane Nascimento","family":"Alves","sequence":"additional","affiliation":[{"name":"Universidade do Estado de Minas Gerais , , , ,","place":["Ituiutaba, Minas Gerais, Brazil, 38302-192"]},{"name":"Universidade Federal de Uberla\u0302ndia , , , ,","place":["Uberla\u0302ndia, Minas Gerais, Brazil, 38400-902"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Angelica Oliveira","family":"Gomes","sequence":"additional","affiliation":[{"name":"Universidade Federal do Tria\u0302ngulo Mineiro , , , ,","place":["Uberaba, Minas Gerais, Brazil, 38025-350"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Ingrid de Oliveira","family":"Dias","sequence":"additional","affiliation":[{"name":"Faculdade de Medicina de Jundiai\u0301 , , , ,","place":["Jundiai\u0301, Sa\u0303o Paulo, Brazil, 13202-550"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-4418-0838","authenticated-orcid":true,"given":"Juliana Quero","family":"Reima\u0303o","sequence":"additional","affiliation":[{"name":"Faculdade de Medicina de Jundiai\u0301 , , , ,","place":["Jundiai\u0301, 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