PROTACs go macrocyclic

PROTACs (for proteolysis-targeting chimeras) are double-headed molecules composed of a ligand for a target protein and a ligand for an E3 ubiquitin ligase, chemically joined by a flexible linker. The PROTAC simultaneously recruits the target protein and the E3 ligase into close proximity, and this leads to the target protein being destroyed inside the cell.

PROTACs are conventionally described as containing a single linker. Previous work from the group had shown that the PROTAC can be “sandwiched” between the ligase and the target while in action. This allows the two proteins to make tight contact with each other –an important feature of how a PROTAC works.

This observation inspired the team’s idea to freeze the PROTAC’s flexibility, “locking” the molecule in its bioactive conformation. To do this, guided by the three-dimensional picture of how the PROTAC binds within the ternary complex, the team designed a chemical strategy to add a cyclizing linker to the progenitor molecule. 

The resulting macrocyclic PROTAC, called “macroPROTAC”, was remarkably potent at degrading its target Brd4 and at killing cancer cell lines that depend on the activity of Brd4. They also found that the macroPROTAC was more selective among similar target proteins that it engages with.

The research was conducted by a multidisciplinary team composed of organic chemist Dr Andrea Testa, biophysicist crystallographer Dr Scott Hughes, computational chemist Dr Xavier Lucas, and cell biologist Dr Jane Wright. Andrea designed, synthesized and biophysically characterized the macroPROTAC; Scott crystallized the ternary complex between the macroPROTAC, Brd4 and the E3 ligase VHL, and contributed biophysical data; Xavi performed computational and molecular dynamics simulations that guided molecular design; and Jane performed bioassays to assess the biological activity of the compound in cancer cells.

The team’s findings support macrocyclization as an attractive strategy to boost PROTAC design. It  opens new directions and untaps previously unrealized potential of this class of molecules for drug discovery.

The research was conducted within the School of Life Sciences at Dundee, funded by a European Research Council starting grant and a European Commission Marie-Curie Fellowship.