BIT1 as an Effector of EGFR-TKI-induced Apoptosis via TLE1 Inhibition in Lung Adenocarcinoma Cells

Abstract

Background/aim: Acquired resistance to epidermal growth factor receptor (EGFR)-tyrosine kinase inhibitors (TKIs) remains a substantial clinical obstacle in treating lung adenocarcinoma (LUAD). Identifying pro-survival pathways that allow tumor cells to evade TKI-induced apoptosis is critical for overcoming this resistance. The transcriptional repressor transducin-like enhancer of split 1 (TLE1) was previously identified as a crucial oncogenic factor that promotes survival in resistant cells. This study investigates the mitochondrial protein Bcl-2 inhibitor of transcription 1 (Bit1) as a key pro-apoptotic signal that overrides the TLE1-mediated survival program.

Materials and methods: EGFR-TKI sensitive, resistant, and drug-tolerant persister LUAD models were utilized. Bit1 release and TLE1 translocation were assessed via subcellular fractionation. Cell fate following genetic manipulation was determined through viability and apoptosis assays, while RNA-sequencing identified TLE1-regulated transcriptional changes.

Results: In sensitive EGFR-mutant non-small cell lung cancer (NSCLC) cells, TKI treatment triggers the rapid cytosolic release of the mitochondrial outer membrane permeabilization (MOMP)-tethered protein Bit1. This early mobilization precedes cytochrome C release and occurs independently of full MOMP, identifying the Bit1 pathway as a novel, early-response death signal in lung cancer. While Bit1 downregulation attenuated EGFR-TKI-induced apoptosis in EGFR-mutant lung adenocarcinoma cells, ectopic mitochondrial Bit1 expression restored TKI sensitivity in resistant cells. Mechanistically, TKI exposure triggered cytosolic Bit1-AES complex formation, resulting in the nuclear exclusion and sequestration of TLE1. This axis is also pivotal in adaptive resistance; TLE1 was upregulated in drug-tolerant persister (DTP) cells and required for their survival, while Bit1 activation attenuated DTP formation. Transcriptomic analysis revealed that TLE1 coordinates a resistance program enriched for EMT and Notch signaling.

Conclusion: The Bit1/TLE1 axis is a pivotal regulatory switch dictating apoptotic outcomes following EGFR-TKI treatment. This mechanism identifies a therapeutic vulnerability, suggesting that pharmacological Bit1 pathway activation could be an effective strategy to overcome acquired resistance in LUAD.

Keywords: BIT1; EGFR-TKI resistance; HES1; Lung adenocarcinoma; TLE1; drug tolerant persisters.