{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,9,10]],"date-time":"2026-09-10T12:34:47Z","timestamp":1789043687553,"version":"build-2803163510"},"update-to":[{"DOI":"10.1371\/journal.ppat.1013560","type":"new_version","label":"New version","source":"publisher","updated":{"date-parts":[[2025,10,10]],"date-time":"2025-10-10T00:00:00Z","timestamp":1760054400000}}],"reference-count":98,"publisher":"Public Library of Science (PLoS)","issue":"10","license":[{"start":{"date-parts":[[2025,10,3]],"date-time":"2025-10-03T00:00:00Z","timestamp":1759449600000},"content-version":"vor","delay-in-days":0,"URL":"http:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"content-domain":{"domain":["www.plospathogens.org"],"crossmark-restriction":false},"short-container-title":["PLoS Pathog"],"abstract":"<jats:p>West Nile virus (WNV) poses a global public health threat. This study demonstrates that the WNV RNA tolerates CpG and UpA dinucleotide enrichment in different genomic regions resulting in attenuation of CpG- and CpG\/UpA-enriched variants. Attenuation was zinc finger antiviral protein 1 (ZAP)-dependent, and ZAP knockout (ZAP-KO) cells were used to generate high-titer stocks. Ten enriched variants, with permuted control and wild-type (WT) viruses, were screened in immunocompetent mice upon intraperitoneal injection. In contrast to lethal WNV-WT and permuted viruses, the E-MAX variant, with the RNA region encoding envelope (E) protein enriched both with CpG and UpA, caused no mortality. E-MAX was immunogenic and protective against lethal challenge. Stability of enriched dinucleotides was confirmed upon serial passaging in ZAP-WT and ZAP-KO cells, with only minor (17\u201321%) reversion at a single site in ZAP-WT condition. E-MAX upregulated interferon (IFN) signaling genes in human cells, suggesting that the combination of CpG\/UpA-mediated attenuation, and concurrent activation of IFN responses potentially driven by CpG\/UpA enrichment, may contribute to E-MAX immunogenicity. Evaluation using footpad injection in mice showed E-MAX had a promising safety and immunogenicity profile, although brain infection was occasionally detected. Then, we developed the E-MAX+\u2009FR variant by combining CpG\/UpA enrichment with two amino acid substitutions in functional domains of the E protein. This strategy eliminated neuroinvasion while maintaining immunogenicity and protection. Altogether, CpG\/UpA dinucleotide enrichment in the genomic E region in combination with amino acid substitutions in the E protein yields a promising platform for vaccine development against WNV and potentially other flaviviruses.<\/jats:p>","DOI":"10.1371\/journal.ppat.1013560","type":"journal-article","created":{"date-parts":[[2025,10,3]],"date-time":"2025-10-03T17:36:12Z","timestamp":1759512972000},"page":"e1013560","update-policy":"https:\/\/doi.org\/10.1371\/journal.ppat.corrections_policy","source":"Crossref","is-referenced-by-count":3,"title":["West Nile virus vaccine candidates attenuated by dinucleotide enrichment are immunogenic and protective against lethal infection"],"prefix":"10.1371","volume":"21","author":[{"given":"Nguyen Phuong Khanh","family":"Le","sequence":"first","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Prince 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