TFEB-mediated lysosomal biogenesis and lysosomal drug sequestration confer resistance to MEK inhibition in pancreatic cancer

Abstract

Oncogenic KRAS mutations are encountered in more than 90% of pancreatic ductal adenocarcinomas. MEK inhibition has failed to procure any clinical benefits in mutant RAS-driven cancers including pancreatic ductal adenocarcinoma (PDAC). To identify potential resistance mechanisms underlying MEK inhibitor (MEKi) resistance in PDAC, we investigated lysosomal drug accumulation in PDAC models both in vitro and in vivo. Mouse PDAC models and human PDAC cell lines as well as human PDAC xenografts treated with the MEK inhibitor trametinib or refametinib led to an enhanced expression of lysosomal markers and enrichment of lysosomal gene sets. A time-dependent, increase in lysosomal content was observed upon MEK inhibition. Strikingly, there was a strong activation of lysosomal biogenesis in cell lines of the classical compared to the basal-like molecular subtype. Increase in lysosomal content was associated with nuclear translocation of the Transcription Factor EB (TFEB) and upregulation of TFEB target genes. siRNA-mediated depletion of TFEB led to a decreased lysosomal biogenesis upon MEK inhibition and potentiated sensitivity. Using LC-MS, we show accumulation of MEKi in the lysosomes of treated cells. Therefore, MEK inhibition triggers lysosomal biogenesis and subsequent drug sequestration. Combined targeting of MEK and lysosomal function may improve sensitivity to MEK inhibition in PDAC.

Lysosomal Biogenesis and Implications for Hydroxychloroquine Disposition

“…Many drugs have been shown to activate TFEB in vitro in recent publications (Lu et al, 2017;Zhitomirsky et al, 2018;Zhao et al, 2020), most of which are weakly basic lipophilic agents that would be physicochemically characterized as lysosomotropic. If lysosomotropism is a major factor in the PK of these drugs, as it is for HCQ, then the distribution of these drugs might be altered by their effect on the lysosome as we have observed for HCQ.…”

“…2C). Recent reports suggest that many weakly basic lipophilic compounds, including CQ, activate lysosomal biogenesis by activating the transcription factor EB (TFEB) (Lu et al, 2017;Zhitomirsky et al, 2018;Zhao et al, 2020), which ultimately increases the lysosomal volume fraction of treated cells and should allow them to sequester even more drug than their original baseline (Ruzickova et al, 2019). CQ has been observed to activate TFEB and increase lysosomal content of the cell, so we investigated the capability of HCQ to do the same.…”

Targeting Lysosomes in Cancer as Promising Strategy to Overcome Chemoresistance—A Mini Review

“…Supporting the hypothesis of lysosomal exocytosis, treatment with cytostatics leads to redistribution of basally perinuclear lysosomes to the plasma membrane. Furthermore, lysosomal content is increasingly released into the extracellular compartment, such as cathepsin D and V-ATPase (41,71). Enhanced membrane fusion and subsequent exocytosis could be mediated by lysosomal calcium release via activation of lysosomal cation channels, such as TRPML1 (42).…”

“…In particular, an increase of lysosomal volume after treatment with the cytostatic weak base sunitinib, but not with the non-basic cytostatics 5-fluorouracil and pemetrexed, positively correlates with drug resistance. In detail, treatment with cytostatic weak bases, such as trametinib, vincristine, sunitinib and doxorubicin leads to increased lysosomal volume in several cancer cell lines (39,46,71). Furthermore, a different study shows that LAMP1, a lysosomal marker, is upregulated in breast cancer cells upon development of resistance by continuous exposure to doxorubicin (47).…”

Getting Lost in the Cell–Lysosomal Entrapment of Chemotherapeutics

“…CaN dephosphorylates TFEB, permitting TFEB translocation into the nucleus to initiate lysosomal biogenesis in an effort to supply cellular nutrients through the degradation and recycling of biomolecules [75]. Considering the importance of mTORC-TFEB signaling axis in maintaining lysosomal homeostasis and cellular metabolism, it is unsurprising to see TFEB signaling circuitry become sabotaged during the acquisition of chemo-resistance and that the dysregulated recruitment and activity of mTORC substrates serves as the underlying etiology for various pathological conditions [76,77].…”