USP10/GSK3β-mediated inhibition of PTEN drives resistance to PI3K inhibitors in breast cancer
Nishi Kumari, Sarah C.E. Wright, Christopher M. Witham, Laia Monserrat, Marta Palafox, John L.C. Richard, Carlotta Costa, Moshe Elkabets, Mark Agostino, Theresa Klemm, Melissa Eccles, Alex Garnham, Ting Wu, Jonas A. Nilsson, Nikita Walz, Veena Venugopal, Anthony Cerra, Natali Vasilevski, Stephanie Bridgeman, Sona Bassi, Azad Saei, Moutaz Helal, Philipp Neundorf, Angela Riedel, Mathias Rosenfeldt, Jespal Gill, Nikolett Pahor, Oliver Hartmann, Jacky Chung, Sachdev S. Sidhu, Nina Moderau, Sudhakar Jha, Jordi Rodon, Markus E. Diefenbacher, David Komander, Violeta Serra, Pieter Johan Adam Eichhorn
Nishi Kumari, Sarah C.E. Wright, Christopher M. Witham, Laia Monserrat, Marta Palafox, John L.C. Richard, Carlotta Costa, Moshe Elkabets, Mark Agostino, Theresa Klemm, Melissa Eccles, Alex Garnham, Ting Wu, Jonas A. Nilsson, Nikita Walz, Veena Venugopal, Anthony Cerra, Natali Vasilevski, Stephanie Bridgeman, Sona Bassi, Azad Saei, Moutaz Helal, Philipp Neundorf, Angela Riedel, Mathias Rosenfeldt, Jespal Gill, Nikolett Pahor, Oliver Hartmann, Jacky Chung, Sachdev S. Sidhu, Nina Moderau, Sudhakar Jha, Jordi Rodon, Markus E. Diefenbacher, David Komander, Violeta Serra, Pieter Johan Adam Eichhorn
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Research Article Cell biology Oncology

USP10/GSK3β-mediated inhibition of PTEN drives resistance to PI3K inhibitors in breast cancer

Abstract

Activating mutations in PIK3CA, the gene encoding the catalytic p110α subunit of PI3K, are some of the most frequent genomic alterations in breast cancer. Alpelisib, a small-molecule inhibitor that targets p110α, is a recommended drug for patients with PIK3CA-mutant advanced breast cancer. However, clinical success for PI3K inhibitors (PI3Kis) has been limited by their narrow therapeutic window. The lipid phosphatase PTEN is a potent tumor suppressor and a major negative regulator of the PI3K pathway. Unsurprisingly, inactivating mutations in PTEN correlate with tumor progression and resistance to PI3K inhibition due to persistent PI3K signaling. Here, we demonstrate that PI3K inhibition leads rapidly to the inactivation of PTEN. Using a functional genetic screen, we show that this effect is mediated by a USP10-GSK3β signaling axis, in which USP10 stabilizes GSK3β, resulting in GSK3β-mediated phosphorylation of the C-terminal tail of PTEN. This phosphorylation inhibits PTEN dimerization and thus prevents its activation. Downregulation of GSK3β or USP10 resensitizes PI3Ki-resistant breast cancer models and patient-derived organoids to PI3K inhibition and induces tumor regression. Our study establishes that enhancing PTEN activity is a new strategy to treat PIK3CA mutant tumors and provides a strong rationale for pursuing USP10 inhibitors in the clinic.

Authors

Nishi Kumari, Sarah C.E. Wright, Christopher M. Witham, Laia Monserrat, Marta Palafox, John L.C. Richard, Carlotta Costa, Moshe Elkabets, Mark Agostino, Theresa Klemm, Melissa Eccles, Alex Garnham, Ting Wu, Jonas A. Nilsson, Nikita Walz, Veena Venugopal, Anthony Cerra, Natali Vasilevski, Stephanie Bridgeman, Sona Bassi, Azad Saei, Moutaz Helal, Philipp Neundorf, Angela Riedel, Mathias Rosenfeldt, Jespal Gill, Nikolett Pahor, Oliver Hartmann, Jacky Chung, Sachdev S. Sidhu, Nina Moderau, Sudhakar Jha, Jordi Rodon, Markus E. Diefenbacher, David Komander, Violeta Serra, Pieter Johan Adam Eichhorn

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Figure 5

PI3Kis induce PTEN T366 phosphorylation, regulating its dimerization.

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PI3Kis induce PTEN T366 phosphorylation, regulating its dimerization. (A...
(A) Schematic of PTEN in an open, active, unphosphorylated state (top) and PTEN phosphorylation in the C-terminal tail inducing a closed, inactive conformation (bottom). (B) Co-IP assay from total lysate of HEK293T cells, transfected with GFP-PTEN or FLAG-PTEN, with or without shRNA USP10L1. IP with anti-FLAG antibody. Western blot was probed with the indicated antibodies. (C) Co-IP assay from total lysate of HEK293T cells or HEK293T USP10KO1 cells transfected with GFP-PTEN and FLAG-PTEN. IP with anti-FLAG antibody. Western blot was probed with indicated antibodies. (D) Co-IP assay from total lysate of HEK293T, HEK293T USP10KO1, or HEK293T USP10KO1 cells ectopically expressing USP10, transfected with GFP-PTEN and FLAG -PTEN. IP with anti-FLAG antibody. Western blot was probed with the indicated antibodies. (E) Initially prepared model of unphosphorylated PTEN dimer or phosphorylated PTEN dimer (pT366) prior to molecular dynamics simulation. Phosphorylated residues are shown as spheres. Blue indicates monomer 1; red indicates monomer 2; teal/magenta indicates phosphothreonine residues. (F) Molecular mechanics — general born surface area binding energies for PTEN dimers with differential phosphorylation status. (G) Co-IP assay from total lysate of HEK293T cells, transfected with GFP-PTEN, HA-PTEN, or HA-PTEN T366A. IP with anti-GFP antibody. Western blot was probed with the indicated antibodies. (H) MCF7 cells treated for 24 hours with 1 μM indicated PI3Kis or AKT inhibitors (AKTi). Whole-cell lysates were collected and probed with the indicated antibodies. (I) MCF7 cells transfected with GFP-PTEN and FLAG-PTEN were treated with 10 μM BYL719, as indicated. IP with anti-FLAG antibody. Western blot was probed with indicated antibodies.