Inital hits
Please go to Researchers involved for a list of researchers who contributed to this work
2023-11-16
The selected compounds were screened against DENV2 Rdrp picogreen assay to determine the IC50. Unfortunately all the tested compounds showed an IC50 higher than 150 µM. The compounds were subsequently tested in a DENV2 and DENV4 phenotypic assays using Huh7 cells. Of these, the 9 compounds below showed promising antiviral inhibition and were analyzed for EC50.
Filtering the compounds through the STOPLIGHT calculator and checking them in an online PAINs remover program showed one compound is potentially a PAIN (RA-3301).
2023-12-11
Determining EC50 in a luciferase inhibition assay showed three compounds with antiviral effect below 10 μM: RA-3272, RA-3301, and RA-3318. For each hit outlined below, 10 compounds were purchased as an initials SAR exploration. For RA-3301, that had been previously flagged as a possible PAIN, that element was removed from the structures purchased for the expansion.
2024-02-22
Whole cell toxicity was determined to be above 10 μM, and while this does not rule out RA-3301 as a PAIN, it indicates the compound does not hit on many other targets. Further more, WaveDelta tests showed the three hits interact with DENV2 RdRp, despite initially not in the picogreen assay. The picogreen assay does have limitations, and it is possible these compounds are outside of these limitations. While the Kd for the three compounds are moderate, this is encouraging for future tests. Solubility has been measured in a single-point buffer-based assay - solubility is low but validity and usefulness of this data is questionable.
2024-04-09
DLS data shows that aggregation is best for RA-3301, moderate for RA-3318, but poor for RA-3272 (data 1, data 2).
2024-04-16
To confirm RA-3272 and RA-3301 are blocking replication of the virus, and not just luciferase expression, EC90 for the two compounds was tested under these conditions which showed that RA-3272 only expressed moderate inhibition of viral replication, while RA-3301 showed good inhibition (2.3 μM on DENV2 and 2.4 μM on DENV4).
2024-06-04
The initial hits RA-3272 and RA-3301 have now been tested several times in the nano-luciferase and antiviral replication assays, as well as for toxicity on the Huh7 cell line. This table summarizes the data:
The raw data can be found in the following files:
Coming soon!