Silencing VDAC1 Expression by siRNA Inhibits Cancer Cell Proliferation and Tumor Growth In Vivo

Abstract

Alterations in cellular metabolism and bioenergetics are vital for cancer cell growth and motility. Here, the role of the mitochondrial protein voltage-dependent anion channel (VDAC1), a master gatekeeper regulating the flux of metabolites and ions between mitochondria and the cytoplasm, in regulating the growth of several cancer cell lines was investigated by silencing VDAC1 expression using small interfering RNA (siRNA). A single siRNA specific to the human VDAC1 sequence at nanomolar concentrations led to some 90% decrease in VDAC1 levels in the lung A549 and H358, prostate PC-3, colon HCT116, glioblastoma U87, liver HepG2, and pancreas Panc-1 cancer cell lines. VDAC1 silencing persisted 144 hours post-transfection and resulted in profound inhibition of cell growth in cancer but not in noncancerous cells, with up to 90% inhibition being observed over 5 days that was prolonged by a second transfection. Cells expressing low VDAC1 levels showed decreased mitochondrial membrane potential and adenoside triphosphate (ATP) levels, suggesting limited metabolite exchange between mitochondria and cytosol. Moreover, cells silenced for VDAC1 expression showed decreased migration, even in the presence of the wound healing accelerator basic fibroblast growth factor (bFGF). VDAC1-siRNA inhibited cancer cell growth in a Matrigel-based assay in host nude mice. Finally, in a xenograft lung cancer mouse model, chemically modified VDAC1-siRNA not only inhibited tumor growth but also resulted in tumor regression. This study thus shows that VDAC1 silencing by means of RNA interference (RNAi) dramatically inhibits cancer cell growth and tumor development by disabling the abnormal metabolic behavior of cancer cells, potentially paving the way for a more effective pipeline of anticancer drugs.

Novel function of ceramide for regulation of mitochondrial ATP release in astrocytes

“…Previous studies using a similar or the same ATP assay as applied in our study showed that VDAC1 downregulation leads to reduced levels of intracellular ATP (45,46,82). This observation is quite remarkable since this assay does not distinguish between mitochondrial or cytosolic ATP levels, indicating that blocking VDAC1 will reduce both ATP pools.…”

“…There are numerous reports, however, that VDAC1 downregulation profoundly decreases levels of intra-and extracellular (secreted) ATP (45,46,82). Based on our observation that ceramide depletion decreases complex formation between tubulin and VDAC1, we tested if reducing ceramide will up-regulate cellular ATP levels.…”

“…In AD, Aβ has been shown to bind to VDAC1, reduce ATP levels, and cause mitochondrial fragmentation during early stages of the disease (35,43,44). VDAC1 downregulation has been reported to reduce cellular ATP levels and induce mitochondrial fragmentation (45,46), eventually leading to by guest, on May 11, 2018 www.jlr.org Downloaded from neurodegeneration. In astrocytes, VDAC1 is critical for secretion of ATP, a gliotransmitter regulating synaptic plasticity and neuronal function (47)(48)(49)(50).…”

The mitochondrial voltage-dependent anion channel 1 in tumor cells

“…Moreover, the cancer-associated abnormalities in glucose metabolism enhance cellular resistance to apoptosis, with mitochondria being found to play a key role in this process [275][276][277][278][279]. Lung cancer tumors and cell lines Up-regulation Western blot Tumor and healthy tissues from the same patient were compared [421] The high glycolytic rate that is characteristic of hypoxic solid tumors is attributed to marked over-expression of mitochondrial-bound HK that is considered the rate-limiting enzyme of glycolysis [2,[280][281][282]. HK likely plays a key role in cell growth rate and survival of cancer cells and is probably a major factor underlying the Warburg effect [273,281,[283][284][285].…”

“…It has also been shown that VDAC1 silencing potentiated H 2 O 2 -induced apoptosis and impaired mitochondrial Ca 2+ loading, while silencing VDAC2 had the opposite effects [261]. Furthermore, a single siRNA specific to the human VDAC1 sequence (hVDAC1-siRNA) was shown to silence VDAC1 expression by over 90% at low nanomolar concentrations and inhibit the growth of various cancer cell types, both in vitro and in vivo [421]. In a xenograft lung cancer mouse model, chemically modified VDAC1-siRNA not only inhibited tumor growth but also resulted in tumor regression [421].…”