Identification and evaluation of small-molecule inhibitors against the dNTPase SAMHD1 via a comprehensive screening funnel

Si Min Zhang* (Main author), Cynthia B.J. Paulin, Huazhang Shu, Miriam Yagüe-Capilla, Maurice Michel, Petra Marttila, Florian Ortis, Henri Colyn Bwanika, Cristopher Dirks, Rajagopal Papagudi Venkatram, Elisée Wiita, Ann-Sofie Jemth, Ingrid Almlöf, Olga Loseva, Femke M. Hormann, Tobias Koolmeister, Erika Linde, Sun Lee, Sabin Llona-Miguez, Martin HaraldssonHanna Axelsson, Kia Strömberg, Evert J. Homan, Martin Scobie, Thomas Lundbäck, Thomas Helleday, Sean G. Rudd*

*Corresponding author for this work

Research output: Contribution to journalArticleResearchpeer-review

1 Citation (Scopus)


SAMHD1 is a dNTP triphosphohydrolase governing nucleotide pool homeostasis and can detoxify chemotherapy metabolites controlling their clinical responses. To understand SAMHD1 biology and investigate the potential of targeting SAMHD1 as neoadjuvant to current chemotherapies, we set out to discover selective small-molecule inhibitors. Here, we report a discovery pipeline encompassing a biochemical screening campaign and a set of complementary biochemical, biophysical, and cell-based readouts for rigorous characterization of the screen output. The identified small molecules, TH6342 and analogs, accompanied by inactive control TH7126, demonstrated specific, low μM potency against both physiological and oncology-drug-derived substrates. By coupling kinetic studies with thermal shift assays, we reveal the inhibitory mechanism of TH6342 and analogs, which engage pre-tetrameric SAMHD1 and deter oligomerization and allosteric activation without occupying nucleotide-binding pockets. Altogether, our study diversifies inhibitory modes against SAMHD1, and the discovery pipeline reported herein represents a thorough framework for future SAMHD1 inhibitor development.
Original languageEnglish
Article number108907
Issue number2
Publication statusPublished - 16 Feb 2024
Externally publishedYes


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