The Story So Far
The molecules of interest came from a recent publication doi: 10.1016/j.antiviral.2023.105697. The paper describes a chemotype containing an indole core and a diketo acid branch, with three different substituents on the indole nitrogen (Figure below).
In the paper, the six molecules were tested against SARS-CoV-2 nsp13 in unwindase and ATPase assays. Results showed that the molecules inhibit nsp13 in the low μM range and no cytoxicity was observed in Vero E6 cells.
The first step was to confirm these results, including in biophysical assays not used in the original work, such as SPR. For this reason, two of the molecules, and their ethyl esters, were selected for synthesis. In the original poster, these molecules showed interesting activity in the unwinding assay and antiviral assay, and also their ethyl esters showed activity in the unwinding and NTPase assay. The figure below shows the selected compounds and their reported activities, from the paper recently published.
Details of the synthetic chemistry can be found in the dedicated wiki page.
The three molecules below were registered with a RA number and tested on the FRET unwinding assay vs SARS-CoV-2 nsp13. Results show low consistency between the triplicates and, also, at different magnitude to the literature. To evaluate the possible origin of these discrepancies, we decided to purchase SSYA100-01 and Licoflavone C, which were used as controls in the paper above.
SSYA100-01 was purchased and tested on nsp13 FRET unwinding assay and also ADP-glo assay. The molecule showed inhibition of nsp13, however, not to the same potency described in the paper above. Upon further look in the literature, SSYA100-01 has been described to inhibit nsp13 in a large range of IC50 values. Data can be found here.
It was requested that the unwinding assay and ADP-glo assay to be run with these four compounds. So far, we have data regarding the unwinding assay. SSYA100-01 showed an IC50 closer to reported in the paper above. Licoflavone C showed an IC50 3x higher than the reported. The three indoles showed inhibition similar to the previous results. All results are shown in a table below. Data can be found here.
Two of the three indoles were tested for antiviral activity and showed to be inactive (higher concentration tested 20 uM). The kinetic solubility of these is below. Data can be find here.
RA-0003628 will also be tested on antiviral assay and kinetic solubility.
We are finalising collecting data on these molecules. A live master file of data can be found here.