Multiple studies have implicated the depletion of
isoprenoid biosynthetic pathway intermediates in the induction of autophagy. However, the exact mechanism by which
isoprenoid biosynthesis inhibitors induce autophagy has not been well established. We hypothesized that inhibition of
farnesyl diphosphate synthase (FDPS) and
geranylgeranyl diphosphate synthase (GGDPS) by
bisphosphonates would induce autophagy by depleting cellular
geranylgeranyl diphosphate (GGPP) and impairing protein geranylgeranylation. Herein, we show that an inhibitor of FDPS (
zoledronate) and an inhibitor of GGDPS (
digeranyl bisphosphonate, DGBP) induce autophagy in PC3
prostate cancer and MDA-MB-231
breast cancer cells as measured by accumulation of the autophagic marker LC3-II. Treatment of cells with lysosomal
protease inhibitors [(2S,3S)-trans-epoxysuccinyl-L-leucylamido-3-methylbutane ethyl
ester (E-64d) and
pepstatin A] in combination with
zoledronate or
digeranyl bisphosphonate further enhances the formation of LC3-II, indicating that these compounds induce autophagic flux. It is noteworthy that the addition of exogenous GGPP prevented the accumulation of LC3-II and impairment of Rab6 (a
GGTase II substrate) geranylgeranylation by
isoprenoid pathway inhibitors (
lovastatin,
zoledronate, and DGBP). However, exogenous GGPP did not restore
isoprenoid pathway inhibitor-induced impairment of Rap1a (a
GGTase I substrate) geranylgeranylation. In addition, specific inhibitors of farnesyl
transferase and geranylgeranyl
transferase I are unable to induce autophagy in our system. Furthermore, the addition of
bafilomycin A1 (an inhibitor of autophagy processing) enhanced the antiproliferative effects of
digeranyl bisphosphonate. These results are the first to demonstrate that
bisphosphonates induce autophagy. Our study suggests that induction of autophagy in PC3 cells with these agents is probably dependent upon impairment of geranylgeranylation of
GGTase II substrates.