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BAF complex vulnerabilities in cancer demonstrated via structure-based PROTAC design

Abstract

Targeting subunits of BAF/PBAF chromatin remodeling complexes has been proposed as an approach to exploit cancer vulnerabilities. Here, we develop proteolysis targeting chimera (PROTAC) degraders of the BAF ATPase subunits SMARCA2 and SMARCA4 using a bromodomain ligand and recruitment of the E3 ubiquitin ligase VHL. High-resolution ternary complex crystal structures and biophysical investigation guided rational and efficient optimization toward ACBI1, a potent and cooperative degrader of SMARCA2, SMARCA4 and PBRM1. ACBI1 induced anti-proliferative effects and cell death caused by SMARCA2 depletion in SMARCA4 mutant cancer cells, and in acute myeloid leukemia cells dependent on SMARCA4 ATPase activity. These findings exemplify a successful biophysics- and structure-based PROTAC design approach to degrade high profile drug targets, and pave the way toward new therapeutics for the treatment of tumors sensitive to the loss of BAF complex ATPases.

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Data availability

Atomic coordinates and structure factors for new protein structures SMARCA2BD:SMARCABD, SMARCA2BD:PROTAC 1:VCB, SMARCA2BD:PROTAC 2:VCB and SMARCA4BD:PROTAC 2:VCB have been deposited in the PDB under accession numbers 6HAZ, 6HAY, 6HAX and 6HR2, respectively. All data generated and analyzed during this study are included in this published article and its Supplementary Information or are available from the corresponding authors upon reasonable request.

Additional information

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Acknowledgements

This project has received funding from Boehringer Ingelheim. A.C. was partially supported by a European Research Council (ERC) Starting Grant (grant agreement no. ERC-2012-StG-311460 DrugE3CRLs). Biophysics, drug discovery and proteomics/computing activities at Dundee were partially supported by Wellcome Trust strategic awards (nos. 100476/Z/12/Z, 094090/Z/10/Z and 097945/C/11/Z, respectively). We are thankful to D. Covini for synthesis support; G. Glendinning and S. Mayer for compound logistics; D. Haering and A. Weiss for help with cooperativity and protein degradation measurements; S. Winkler and M. Scharnweber for help with SPR measurements; K. Mayr for LogD measurements; S. Doebel, G. Flotzinger, G. Siszler and P. Werni for protein production and purification; P. Fyfe for support with the in-house X-ray facility; M. Gadd for the gift of the VHLR69A construct; D. Lamont for assistance in proteomics and V. Vetma for data analysis. We also thank the Diamond Light Source for beamtime (BAG proposal MX14980) and for beamline support at beamline IO4-1 and I24.

Author information

A.C., M.P. and M.K. conceived the idea. A.C., W.F., M.K., S.S., M.P., D.B.M. and P.E. directed the project. W.F., S.S. and C.D. designed compounds. M.K., A.C., W.F., T.O.H., C.W., N.T., N.W., E.D., H.W., C.D., B.S., S.W. and M.J.R. designed experiments and interpreted data. E.D., N.T., J.K.O., O.P. and P.G. synthesized compounds. Crystallography was performed by M.J.R., G. Bader and A.W. P. G. Boehmelt helped in the design of a cell line used for testing PROTAC-induced protein degradation. Protein production was performed and supervised by M.J.R. and A.Z. T. Gerstberger supervised TR–FRET competition experiments, B.S. performed TR–FRET competition experiments. K.R. supervised SPR experiments, M.G. performed SPR experiments. E.T. and C.W. performed degradation studies supervised by R.S. and M.K. T. Gmaschitz, J.W., K.E.-W. and C.R. performed overexpression studies, proliferation, degradation and apoptosis assays. M.J.R. and D.Z. developed and performed fluorescence polarization cooperativity assays and M.J.R. performed the ITC cooperativity assays. C.W. developed and performed the cell lysate AlphaLISA assay. N.T. and N.W. performed MS proteomics. M.Y.W. developed IP–MS protocols. J.R. and H.A. supervised in vitro drug metabolism and pharmacokinetics experiments. W.F. and M.K. wrote the manuscript with input from all other authors.

Competing interests

A.C. is a scientific founder, director and shareholder of Amphista Therapeutics, a company that is developing targeted protein degradation therapeutic platforms.

Correspondence to Darryl B. McConnell or Alessio Ciulli.

Supplementary information

Supplementary Information

Supplementary Tables 1–6, Supplementary Figures 1–15

Reporting Summary

Supplementary Note

Synthetic Procedures

Source data

Dataset 1

Proteomic analysis of relative protein abundance in MV-4-11 cells using ACBI1.

Dataset 2

Proteomic analysis of relative protein abundance in MV-4-11 cells using PROTAC 2.

Dataset 3

IP–MS data representing protein abundance in MV-4-11 cells using PROTAC 2.

Dataset 4

IP–MS data representing protein abundance in MV-4-11 cells using PROTAC 2, including IgG controls.

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Fig. 1: Rational design and evaluation of a partial degrader of SMARCA2 and SMARCA4, PROTAC 1.
Fig. 2: Ternary co-crystal structure of SMARCA2BD:PROTAC 1:VCB.
Fig. 3: Ternary co-crystal structure of SMARCA2BD:PROTAC 2:VCB and biophysical data validate a rational design strategy.
Fig. 4: ACBI1 is a potent and selective degrader of SMARCA2, SMARCA4 and PBRM1.
Fig. 5: Effects on the proliferation and apoptosis of cancer cells in the presence of ACBI1.