Resistance to abemaciclib is associated with increased metastatic potential and lysosomal protein deregulation in breast cancer cells

Abstract

Cyclin dependent kinase 4 and 6 inhibitors (CDK4/6i) such as abemaciclib are routinely used to treat metastatic estrogen receptor positive (ER+)/HER2-negative breast cancer. However, adaptive mechanisms inhibit their effectiveness and allow for disease progression. Using murine metastatic ER+ breast cancer cells, we show that acquired resistance to abemaciclib is accompanied by increase in metastatic potential. Mass spectrometry-based proteomics from abemaciclib sensitive and resistant cells showed that lysosomal proteins including CTSD (cathepsin D), CTSA (cathepsin A) and CD68 were significantly increased in resistant cells. Combination of abemaciclib and a lysosomal destabilizer, such as hydroxychloroquine (HCQ) or bafilomycin A1, re-sensitized resistant cells to abemaciclib. Also, combination of abemaciclib and HCQ decreased migration and invasive potential and increased lysosomal membrane permeability (LMP) in resistant cells. Pro-survival BCL2 protein levels were elevated in resistant cells, and a triple treatment with abemaciclib, HCQ, and BCL2 inhibitor, venetoclax, significantly inhibited cell growth compared to treatment with abemaciclib and HCQ. Furthermore, resistant cells showed increased levels of TFEB (Transcription Factor EB), a master regulator of lysosomal-autophagy genes, and siRNA mediated knockdown of TFEB decreased invasion in resistant cells. TFEB gene was found to be mutated in a subset of invasive human breast cancer samples, and overall survival analysis in ER+, lymph node-positive breast cancer showed that increased TFEB expression correlated with decreased survival. Collectively, we show that prolonged exposure to abemaciclib in ER+ breast cancer cells leads to resistance accompanied by an aggressive phenotype that is partly supported by deregulated lysosomal function. Implications: Our data implicate that resistance to abemaciclib is associated with deregulation of lysosomes and augmented metastatic potential, and therefore, the lysosomal pathway could be a therapeutic target in advanced ER+ breast cancer. GRAPHICAL ABSTRACT O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=164 SRC="FIGDIR/small/537215v1_ufig1.gif" ALT="Figure 1"> View larger version (45K): org.highwire.dtl.DTLVardef@1bd5e3aorg.highwire.dtl.DTLVardef@175559borg.highwire.dtl.DTLVardef@473d17org.highwire.dtl.DTLVardef@fbf0bf_HPS_FORMAT_FIGEXP M_FIG C_FIG