Aims: Characterization of a small anticancer compound.
Background: The development of small molecules as new anti-cancer therapeutics is necessary to
improve anti-tumor efficacy and reduce toxicities, especially for the treatment of brain tumors,
where only small molecules can effectively cross the brain-blood barrier. Several novel hits were
previously selected by concurrently screening colon and glioma cancer cell lines with a sensorconjugated
reporter system. Here, we focused on one of them.
Objective: Elucidating the potential target(s) of a novel anticancer compound.
Methods: Computer-assisted structural and motif analysis (least absolute shrinkage and selection
operator or LASSO score) was used to assess compound’s targets, then direct kinase activity assays
were used for the confirmation; Western blot of phosphorylated kinases, as well as FACS and
caspase 3/7 activity assays, were used to decipher the action mechanisms. Finally, the expression
profiling of proteins involved in various G-protein pathways by real-time PCR was performed.
Results: The small chemical, (4E)-4-[2-(9-ethyl-9H-carbazol-3-yl)hydrazin-1-ylidene]-3-methyl-
4,5-dihydro-1H-pyrazol-5-one, with a formula C18H17N5O and MW of 319.36, designated as
VUGX01, was predicted to be a ligand/inhibitor to receptor tyrosine kinases (RTKs) by computer
analysis (least absolute shrinkage and selection operator or LASSO score). However, direct analysis
with recombinant kinases showed that it is not an effective inhibitor to the popular receptor kinases
at 1μM concentration. This compound can activate caspases in some tumor cell lines but has minimal
effects on the cell cycle. Drug treatments lead to the changes in phosphorylation of AKT and c-
RAF, as well as the expression level of MAP2K, suggesting this compound may interact with Gprotein
coupled receptors (GPCRs). The expression profiling of 82 proteins involved in various Gprotein
pathways by real-time PCR showed that the treatment up-regulates the expression of several
proteins, including angiotensinogen, angiotensin II receptor, and IP3-kinase catalytic subunit
Conclusion: VUGX01 can effectively block proliferation and induce apoptosis of certain types of
cancer cells, even it is predicted by high LASSO score, but it is not an effective RTKs inhibitor, it
may inhibit cell growth through acting as a novel ligand to one or several GPCRs.