NAPRT, the rate-limiting enzyme of the Preiss–Handler NAD biosynthetic pathway, has emerged as a key biomarker for the clinical success of NAMPT inhibitors in cancer treatment. Previous studies found that high protein levels of NAPRT conferred resistance to NAMPT inhibition in several tumor types whereas the simultaneous blockade of NAMPT and NAPRT results in marked anti-tumor effects. While research has mainly focused on NAMPT inhibitors, the few available NAPRT inhibitors (NAPRTi) have a low affinity for the enzyme and have been scarcely characterized. In this work, a collection of diverse compounds was screened in silico against the NAPRT structure, and the selected hits were tested through cell-based assays in the NAPRT-proficient OVCAR-5 ovarian cell line and on the recombinant hNAPRT. We found different chemotypes that efficiently inhibit the enzyme in the micromolar range concentration and for which direct engagement with the target was verified by differential scanning fluorimetry. Of note, the therapeutic potential of these compounds was evidenced by a synergistic interaction between the NAMPT inhibitor FK866 and the new NAPRTi in terms of decreasing OVCAR-5 intracellular NAD levels and cell viability. For example, compound IM29 can potentiate the effect of FK866 of more than two-fold in reducing intracellular NAD levels. These results pave the way for the development of a new generation of human NAPRTi with anticancer activity.

We report the discovery of several highly potent small molecules with low-nM potency against severe acute respiratory syndrome coronavirus (SARS-CoV; lowest half-maximal inhibitory concentration (IC₅₀: 13 nM), SARS-CoV-2 (IC₅₀: 23 nM), and Middle East respiratory syndrome coronavirus (MERS-CoV; IC₅₀: 76 nM) in pseudovirus-based assays with excellent selectivity index (SI) values (>5000), demonstrating potential pan-coronavirus inhibitory activities. Some compounds showed 100% inhibition against the cytopathic effects (CPE; IC₁₀₀) of an authentic SARS-CoV-2 (US_WA-1/2020) variant at 1.25 µM. The most active inhibitors also potently inhibited variants of concern (VOCs), including the UK (B.1.1.7) and South African (B.1.351) variants and the Delta variant (B.1.617.2) originally identified in India in pseudovirus-based assay. Surface plasmon resonance (SPR) analysis with one potent inhibitor confirmed that it binds to the prefusion SARS-CoV-2 spike protein trimer. These small-molecule inhibitors prevented virus-mediated cell–cell fusion. The absorption, distribution, metabolism, and excretion (ADME) data for one of the most active inhibitors, NBCoV1, demonstrated drug-like properties. An in vivo pharmacokinetics (PK) study of NBCoV1 in rats demonstrated an excellent half-life (t_1/2) of 11.3 h, a mean resident time (MRT) of 14.2 h, and oral bioavailability. We expect these lead inhibitors to facilitate the further development of preclinical and clinical candidates.