• ホーム
  • 検索結果
  • ALK inhibition activates LC3B-independent, protective autophagy in EML4-ALK positive lung cancer cells.

ALK inhibition activates LC3B-independent, protective autophagy in EML4-ALK positive lung cancer cells.

Scientific reports (2021-04-29)
Anna M Schläfli, Igor Tokarchuk, Sarah Parejo, Susanne Jutzi, Sabina Berezowska, Nikolai Engedal, Mario P Tschan
要旨

ALK inhibitors effectively target EML4-ALK positive non-small cell lung cancer, but their effects are hampered by treatment resistance. In the present study, we asked whether ALK inhibition affects autophagy, and whether this may influence treatment response. Whereas the impact of targeted therapies on autophagic activity previously have been assessed by surrogate marker proteins such as LC3B, we here thoroughly examined effects on functional autophagic activity, i.e. on the sequestration and degradation of autophagic cargo, in addition to autophagic markers. Interestingly, the ALK inhibitor Ceritinib decreased mTOR activity and increased GFP-WIPI1 dot formation in H3122 and H2228 EML4-ALK+ lung cancer cells, suggesting autophagy activation. Moreover, an mCherry-EGFP-LC3B based assay indicated elevated LC3B carrier flux upon ALK inhibition. In accordance, autophagic cargo sequestration and long-lived protein degradation significantly increased upon ALK inhibition. Intriguingly, autophagic cargo flux was dependent on VPS34 and ULK1, but not LC3B. Co-treating H3122 cells with Ceritinib and a VPS34 inhibitor or Bafilomycin A1 resulted in reduced cell numbers. Moreover, VPS34 inhibition reduced clonogenic recovery of Ceritinib-treated cells. In summary, our results indicate that ALK inhibition triggers LC3B-independent macroautophagic flux in EML4-ALK+ cells to support cancer cell survival and clonogenic growth.

材料
製品番号
ブランド
製品内容

Sigma-Aldrich
ペニシリン-ストレプトマイシン, Solution stabilized, with 10,000 units penicillin and 10 mg streptomycin/mL, 0.1 μm filtered, BioReagent, suitable for cell culture
Sigma-Aldrich
トリプシン-EDTA 溶液, 0.25%, sterile-filtered, BioReagent, suitable for cell culture, 2.5 g porcine trypsin and 0.2 g EDTA • 4Na per liter of Hanks′ Balanced Salt Solution with phenol red
Sigma-Aldrich
HEPES, ≥99.5% (titration)
Sigma-Aldrich
ウシ胎児血清, non-USA origin, sterile-filtered, suitable for cell culture
Sigma-Aldrich
ダルベッコ改変イーグル培地 - 低グルコース, With 1000 mg/L glucose, L-glutamine, and sodium bicarbonate, liquid, sterile-filtered, suitable for cell culture
Sigma-Aldrich
ダルベッコ改変イーグル培地/栄養混合物F-12ハム, With L-glutamine, 15 mM HEPES, and sodium bicarbonate, liquid, sterile-filtered, suitable for cell culture
Sigma-Aldrich
ジメチルスルホキシド, ≥99.5% (GC), suitable for plant cell culture
Sigma-Aldrich
クロロキン 二リン酸塩, powder or crystals, 98.5-101.0% (EP)
Sigma-Aldrich
MRT68921 dihydrochloride, ≥98% (HPLC)
Sigma-Aldrich
Monoclonal Anti-SQSTM1 antibody produced in mouse, clone 2C11, purified immunoglobulin, buffered aqueous solution