TK216 targets microtubules in Ewing sarcoma cells

Juan Manuel Povedano, Vicky Li, Katherine E. Lake, Xin Bai, Rameshu Rallabandi, Jiwoong Kim, Yang Xie, Jef K. De Brabander, David G. McFadden

Research output: Contribution to journalArticlepeer-review

Abstract

Ewing sarcoma (EWS) is a pediatric malignancy driven by the EWSR1-FLI1 fusion protein formed by the chromosomal translocation t(11; 22). The small molecule TK216 was developed as a first-in-class direct EWSR1-FLI1 inhibitor and is in phase II clinical trials in combination with vincristine for patients with EWS. However, TK216 exhibits anti-cancer activity against cancer cell lines and xenografts that do not express EWSR1-FLI1, and the mechanism underlying cytotoxicity remains unresolved. We apply a forward-genetics screening platform utilizing engineered hypermutation in EWS cell lines and identify recurrent mutations in TUBA1B, encoding ⍺-tubulin, that prove sufficient to drive resistance to TK216. Using reconstituted microtubule (MT) polymerization in vitro and cell-based chemical probe competition assays, we demonstrate that TK216 acts as an MT destabilizing agent. This work defines the mechanism of cytotoxicity of TK216, explains the synergy observed with vincristine, and calls for a reexamination of ongoing clinical trials with TK216.

Original languageEnglish (US)
Pages (from-to)1325-1332.e4
JournalCell Chemical Biology
Volume29
Issue number8
DOIs
StatePublished - Aug 18 2022

Keywords

  • Ewing sarcoma
  • microtubules
  • ONCT-216
  • target identification
  • TK216
  • YK-4-279

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Medicine
  • Molecular Biology
  • Pharmacology
  • Drug Discovery
  • Clinical Biochemistry

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