Flavonoids from the Leaves of Monanthotaxis Filipes Modulate Pcsk9 and Ldlr

Proprotein convertase subtilisin/kexin type 9 (PCSK9) plays a crucial role in the regulation of blood cholesterol level. Its inhibition attenuates hypercholesterolemia, and hence is a viable approach for the management of atherosclerotic cardiovascular disease (ASCVD). We evaluated the flavonoids of the leaves of Monanthotaxis filipes P.H. Hoekstra (Annonaceae) for their effect on PCSK9 and low-density lipoprotein receptor (LDLR) expression at mRNA level in HepG2 cells using quantitative real-time PCR analysis, and for their influence on protein expression by ELISA. Six natural products were isolated by chromatographic separation, and were identified by spectroscopic (NMR, IR, UV, MS) analyses. 2',3',4',6'-Tetramethoxychalcone (1) reduced the PCSK9 protein amount and altered LDLR both on mRNA and protein levels, 6,7,8-trimethoxyflavone (2) inhibited PCSK9 both on mRNA and protein levels but did not change the amount of LDLR in HepG2 cells, whereas 2՛,4՛dihydroxy-6՛-methoxy-3՛,5՛-dimethylchalcone (5) decreased PCSK9 and upregulated LDLR protein expression. The above activities along with low cytotoxicities suggest the suitability of these flavonoids for the management of cardiovascular diseases. Molecular dynamics simulations revealed that an allosteric mechanism underlies the inhibitory effect of 2 on PCSK9, whereas that the pronounced activity of 5 is due to the interaction of its benzene ring with the Cys358, Pro438, Val460 and Trp461 residues of the catalytic site, as proposed by Molecular Mechanics Poisson Boltzmann surface area (MM-PBSA) analyses. Our results reveal that natural flavonoids may provide an affordable alternative to monoclonal antibodies for the management of atherosclerotic cardiovascular disease (ASCVD).