The HDAC inhibitor AR42 interacts with pazopanib to kill trametinib/dabrafenib-resistant melanoma cells in vitro and in vivo.

Laurence Booth,Andrew Poklepovic,John Lee,Cindy Sander,Jane L Roberts,Paul Dent,John M Kirkwood

doi:10.18632/oncotarget.14829

Laurence Booth, Andrew Poklepovic + Show 5 more

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https://doi.org/10.18632/oncotarget.14829

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Abstract

Studies focused on the killing of activated B-RAF melanoma cells by the histone deacetylase (HDAC) inhibitor AR42. Compared to other tumor cell lines, PDX melanoma isolates were significantly more sensitive to AR42-induced killing. AR42 and the multi-kinase inhibitor pazopanib interacted to activate: an eIF2α–Beclin1 pathway causing autophagosome formation; an eIF2α–DR4/DR5/CD95 pathway; and an eIF2α-dependent reduction in the expression of c-FLIP-s, MCL-1 and BCL-XL. AR42 did not alter basal chaperone activity but increased the ability of pazopanib to inhibit HSP90, HSP70 and GRP78. AR42 and pazopanib caused HSP90/HSP70 dissociation from RAF-1 and B-RAF that resulted in reduced ‘RAF’ expression. The drug combination activated a DNA-damage-ATM-AMPK pathway that was associated with: NFκB activation; reduced mTOR S2448 and ULK-1 S757 phosphorylation; and increased ULK-1 S317 and ATG13 S318 phosphorylation. Knock down of PERK, eIF2α, Beclin1, ATG5 or AMPKα, or expression of IκB S32A S36A, ca-mTOR or TRX, reduced cell killing. AR42, via lysosomal degradation, reduced the protein expression of HDACs 2/5/6/10/11. In vivo, a 3-day exposure of dabrafenib/trametinib resistant melanoma cells to the AR42 pazopanib combination reduced tumor growth and enhanced survival from ∼25 to ∼40 days. Tumor cells that had adapted through therapy exhibited elevated HGF expression and the c-MET inhibitor crizotinib enhanced AR42 pazopanib lethality in this evolved drug-resistant population.

Highlights

Prior studies have demonstrated that histone deacetylase inhibitors such as valproate, vorinostat and AR42 enhance the cytotoxic potential of multi-kinase inhibitors such as sorafenib, regorafenib and pazopanib [1,2,3,4,5,6]
The standard of care treatment for mutant B-RAF melanoma is the combination of the MEK1/2/5 inhibitor trametinib and the mutant B-RAF specific inhibitor dabrafenib
In PDX models of mutant B-RAF melanoma AR42 and pazopanib, as single agents, and in combination were more effective at killing the melanoma cells than the combination of trametinib/dabrafenib at their individual 100% C max plasma concentrations (Figure 1D)

Summary

Introduction

Prior studies have demonstrated that histone deacetylase inhibitors such as valproate, vorinostat and AR42 enhance the cytotoxic potential of multi-kinase inhibitors such as sorafenib, regorafenib and pazopanib [1,2,3,4,5,6]. In addition to being inhibitors of protein kinases, were recently discovered to be potent inhibitors of chaperone ATPase activities. It has been reported that acetylation can alter the activity of chaperones, generally thought to be an inhibitory effect, with the regulatory acetylation of HSP90 controlled by HDAC6 [9, 10]. We determined that that phosphodiesterase 5 inhibitors such as sildenafil (Viagra®), via PKG signaling, did not significantly alter basal chaperone ATPase activities www.impactjournals.com/oncotarget but instead facilitated sorafenib and pazopanib to cause further inhibition of chaperone ATPase activities and a more rapid NH2-terminus conformational change [11]. Whether chaperone acetylation facilitates the chaperone inhibitory effects of a multi-kinase inhibitor is unknown

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