TK216 targets microtubules in Ewing sarcoma cells

Cell Chem Biol. 2022 Aug 18;29(8):1325-1332.e4. doi: 10.1016/j.chembiol.2022.06.002. Epub 2022 Jul 7.

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.

Keywords: Ewing sarcoma; ONCT-216; TK216; YK-4-279; microtubules; target identification.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Antineoplastic Agents* / pharmacology
  • Antineoplastic Agents* / therapeutic use
  • Cell Line, Tumor
  • Child
  • Gene Expression Regulation, Neoplastic
  • Humans
  • Microtubules / metabolism
  • Sarcoma, Ewing* / drug therapy
  • Sarcoma, Ewing* / genetics
  • Sarcoma, Ewing* / pathology
  • Vincristine / pharmacology
  • Vincristine / therapeutic use

Substances

  • Antineoplastic Agents
  • Vincristine