In this study, we demonstrate for the first time that the betaine-type alkaloid Lycobetaine (LYC) preferentially binds to c-MYC and BCL-2 promoter G-quadruplexes (G4s). Biophysical experiments including absorption ti-trations, CD melting and NMR studies showed that LYC had the ability to bind and stabilize both c-MYC and BCL-2 G4s through an end-stacking mode. Additional biological assays including RT-PCR and Western blotting indicated that LYC simultaneously inhibited the expression of c-MYC and BCL-2 oncogenes in A549/DDP cells, which thereby led to cell cycle arrest, apoptosis, as well as inhibition on cell proliferation and migration. Taken together, these results revealed a new potential anticancer mechanism of LYC with potential application of LYC in drug-resistant cancers.