Orally efficacious broad-spectrum allosteric inhibitor of paramyxovirus polymerase

Cox, Robert; Sourimant, Julien; Toots, Märt; Yoon, Jeong-Joong; Ikegame, Satoshi; Govindarajan, Mugunthan; Watkinson, Ruth E.; Thibault, Patricia A.; Makhsous, Negar; Lin, Michelle J.; Marengo, Jose R.; Sticher, Zachary M.; Kolykhalov, Alexander A.; Natchus, Michael G.; Greninger, Alexander L.; Lee, Benhur; Plemper, Richard K.

doi:10.1038/s41564-020-0752-7

Cited by 22 publications

(59 citation statements)

References 72 publications

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“…We have previously identified signature resistance hot-spots for ERDRP-0519 in MeV and CDV L through viral adaptation (10,12). Escape sites locate to the polymerase and capping domains of L, but notably do not overlap with the resistance profile of recently developed GHP-88309 (28), the only other well-characterized small molecule inhibitor of MeV L (figure 1A).…”

Section: Resultsmentioning

confidence: 99%

“…Of these, two (AZ-27 (34) and AVG-233 (27)) are RSV Lspecific, one (GHP-88309 (28)) inhibits paramyxoviruses of the respiro-and morbillivirus genera, and ERDRP-0519 specifically blocks morbillivirus polymerases (10,12). Remarkably, all of these compounds have been demonstrated to interfere with de novo polymerase initiation at the promoter and backpriming, albeit in the case of AZ-27, AVG-233, and GHP-88309, after incorporation of an additional 2-4 nucleotides (27,28,34). This delayed polymerase arrest demonstrates that these compounds do not directly block phosphodiester bond formation.…”

Section: Discussionmentioning

confidence: 99%

“…The ERDRP-0519 resistance fingerprint is unique among all small-molecule mononegavirus polymerase inhibitors characterized to date, including small molecule inhibitors of closely related respiratory syncytial virus (RSV) such as AZ-27 (24)(25)(26) and AVG-233 (27), and a structurally distinct inhibitor of morbillivirus and respirovirus polymerases, GHP-88309, that we have recently described (28). These features identified the ERDRP-0519 chemotype as a mechanistically first-in-class compound that may have illuminated a conserved, yet currently uncharted, druggable site in mononegavirus polymerases.…”

Section: Cox Et Almentioning

confidence: 99%

“…However, none of the ERDRP-0519 resistance mutations altered polymerase susceptibility to GHP-88309 (28) affected MeV L inhibition by ERDRP-0519 (figure 1F). The lack of cross-resistance between ERDRP-0519 and GHP-88309 confirms that pharmacophores of these inhibitor classes are distinct.…”

Section: Pharmacophore Of Erdrp-0519 Class Is Uniquementioning

confidence: 99%

“…All allosteric paramyxovirus and pneumovirus polymerase inhibitors subjected to in vitro RdRP assays to date were found unable to block the formation of the first phosphodiester bond during backpriming and/or primer extension (27,28,34). Backpriming refers to the spontaneous formation of a circular hairpin structure of the non-encapsidated synthetic templates (35) that allows extension of the resulting paired 3'-ends beyond the actual length of the template (figure 4A).…”

Section: Erdrp-0519 Blocks All Phosphodiester Bond Formationmentioning

confidence: 99%

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Therapeutic Targeting of Measles Virus Polymerase with ERDRP-0519 Suppresses All RNA Synthesis Activity

Cox

Sourimant

Govindarajan

et al. 2020

Preprint

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Morbilliviruses, such as measles virus (MeV) and canine distemper virus (CDV), are highly infectious members of the paramyxovirus family. MeV is responsible for major morbidity and mortality in non-vaccinated populations. ERDRP-0519, a pan-morbillivirus small molecule inhibitor for the treatment of measles, targets the morbillivirus RNA-dependent RNA-polymerase (RdRP) complex and displayed unparalleled oral efficacy against lethal infection of ferrets with CDV, an established surrogate model for human measles. Resistance profiling identified the L subunit of the RdRP, which harbors all enzymatic activity of the polymerase complex, as the molecular target of inhibition. Here, we examined binding characteristics, physical docking site, and the molecular mechanism of action of ERDRP-0519 through label-free biolayer interferometry, photoaffinity cross-linking, and in vitro RdRP assays using purified MeV RdRP complexes and synthetic templates. Results demonstrate that unlike all other mononegavirus small molecule inhibitors identified to date, ERDRP-0519 inhibits all phosphodiester bond formation in both de novo initiation of RNA synthesis at the promoter and RNA elongation by a committed polymerase complex. Photocrosslinking and resistance profiling-informed ligand docking revealed that this unprecedented mechanism of action of ERDRP-0519 is due to simultaneous engagement of the L protein polyribonucleotidyl transferase (PRNTase)-like domain and the flexible intrusion loop by the compound, pharmacologically locking the polymerase in pre-initiation conformation. This study informs selection of ERDRP-0519 as clinical candidate for measles therapy and identifies a previously unrecognized druggable site in mononegavirus L polymerase proteins that can silence all synthesis of viral RNA.ImportanceThe mononegavirus order contains major established and recently emerged human pathogens. Despite the threat to human health, antiviral therapeutics directed against this order remain understudied. The mononegavirus polymerase complex represents a promising drug target due to its central importance for both virus replication and viral mitigation of the innate host antiviral response. In this study, we have mechanistically characterized a clinical candidate small-molecule MeV polymerase inhibitor. The compound blocked all phosphodiester bond formation activity, a unique mechanism of action unlike all other known mononegavirus polymerase inhibitors. Photocrosslinking-based target site mapping demonstrated that this class-defining prototype inhibitor stabilizes a pre-initiation conformation of the viral polymerase complex that sterically cannot accommodate template RNA. Function-equivalent druggable sites exist in all mononegavirus polymerases. In addition to its direct anti-MeV impact, the insight gained in this study can therefore serve as a blueprint for indication spectrum expansion through structure-informed scaffold engineering or targeted drug discovery.

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Section: Resultsmentioning

confidence: 99%