Sorafenib Enhances Pemetrexed Cytotoxicity through an Autophagy-Dependent Mechanism in Cancer Cells

Abstract

Pemetrexed (ALIMTA) is a folate anti-metabolite that has been approved for the treatment of non-small cell lung cancer, and has been shown to stimulate autophagy. In the present study, we sought to further understand the role of autophagy in the response to pemetrexed and to test if combination therapy could enhance the level of toxicity through altered autophagy in tumor cells. The multi-kinase inhibitor sorafenib (NEXAVAR), used in the treatment of renal and hepatocellular carcinoma, suppresses tumor angiogenesis and promotes autophagy in tumor cells. We found that sorafenib interacted in a greater than additive fashion with pemetrexed to increase autophagy and to kill a diverse array of tumor cell types. Tumor cell types that displayed high levels of cell killing after combination treatment showed elevated levels of AKT, p70 S6K and/or phosphorylated mTOR, in addition to Class III RTKs such as PDGFRβ and VEGFR1, known in vivo targets of sorafenib. In xenograft and in syngeneic animal models of mammary carcinoma and glioblastoma, the combination of sorafenib and pemetrexed suppressed tumor growth without deleterious effects on normal tissues or animal body mass. Taken together, the data suggest that premexetred and sorafenib act synergistically to enhance tumor killing via the promotion of a toxic form of autophagy that leads to activation of the intrinsic apoptosis pathway, and predict that combination treatment represents a future therapeutic option in the treatment of solid tumors.

Sorafenib and pemetrexed toxicity in cancer cells is mediated via SRC-ERK signaling

“…11,12,15 Inhibition of mTOR stimulates autophagy by enabling the association of ATG proteins required to initiate formation of the autophagosome. [6][7][8][9][10] We have demonstrated previously that pemetrexed induces tumor cell death and autophagy in a dose-dependent fashion. 6 Receptor tyrosine kinases (RTKs) are important regulators of a variety of cellular signaling cascades, e.g., ERK1/2, which stimulate cell growth and survival.…”

“…[6][7][8][9][10] We have demonstrated previously that pemetrexed induces tumor cell death and autophagy in a dose-dependent fashion. 6 Receptor tyrosine kinases (RTKs) are important regulators of a variety of cellular signaling cascades, e.g., ERK1/2, which stimulate cell growth and survival. 16,17 For this reason, many diverse pharmacological inhibitors have been developed to target RTKs thereby blocking activation of signal transduction pathways, in turn, to promote growth inhibitory and pro-apoptotic effects in cells.…”

“…6 During our studies examining PDGFRβ we also surveyed the expression and activity of other kinases whose functions can be modulated by this receptor, most notably SRC family non-receptor tyrosine kinases. Knockdown of PDGFRβ increased SRC Y416 phosphorylation almost 2-fold ( Fig.…”

“…30 Based on our prior data showing that fulvestrant resistant MCF7 cells (MCF7F) are more aggressive than parental MCF7 cells, we next determined whether PP2A activity levels and the expression of an endogenous inhibitor of PP2A, SET/ I2PP2A influenced the ERK1/2-pathway-dependent regulation of pemetrexed and sorafenib toxicity. 6,31 PP2A activity was lower ZMP leads to the activation of AMP-activated protein kinase (AMPK) and downstream inhibition of mammalian target of rapamycin (mTOR). 11,12,15 Inhibition of mTOR stimulates autophagy by enabling the association of ATG proteins required to initiate formation of the autophagosome.…”

“…In these tumor cell models, combination treatment resulted in a profound enhancement of autophagy and autophagy-dependent tumor cell killing. 6 The present studies sought to define in further detail how pemetrexed (PTX) and sorafenib (SOR) interact to kill mammary carcinoma cells. More specifically, based on prior data linking sorafenib, PDGFRβ and SRC, we initially determined whether drug-induced SRC activation plays any role in the regulation of ERK1/2 and in pemetrexed/sorafenib toxicity.…”