RB expression confers sensitivity to CDK4/6 inhibitor–mediated radiosensitization across breast cancer subtypes
Andrea M. Pesch, … , James M. Rae, Corey W. Speers
Andrea M. Pesch, … , James M. Rae, Corey W. Speers
Published December 21, 2021
Citation Information: JCI Insight. 2022;7(3):e154402. https://doi.org/10.1172/jci.insight.154402.
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RB expression confers sensitivity to CDK4/6 inhibitor–mediated radiosensitization across breast cancer subtypes

Abstract

Standard radiation therapy (RT) does not reliably provide locoregional control for women with multinode-positive breast cancer and triple-negative breast cancer (TNBC). We hypothesized that CDK4/6 inhibition (CDK4/6i) would increase the radiosensitivity not only of estrogen receptor–positive (ER+) cells, but also of TNBC that expresses retinoblastoma (RB) protein. We found that CDK4/6i radiosensitized RB WT TNBC (n = 4, radiation enhancement ratio [rER]: 1.49–2.22) but failed to radiosensitize RB-null TNBC (n = 3, rER: 0.84–1.00). RB expression predicted response to CDK4/6i + RT (R2 = 0.84), and radiosensitization was lost in ER+/TNBC cells (rER: 0.88–1.13) after RB1 knockdown in isogenic and nonisogenic models. CDK4/6i suppressed homologous recombination (HR) in RB WT cells but not in RB-null cells or isogenic models of RB1 loss; HR competency was rescued with RB reexpression. Radiosensitization was independent of nonhomologous end joining and the known effects of CDK4/6i on cell cycle arrest. Mechanistically, RB and RAD51 interact in vitro to promote HR repair. CDK4/6i produced RB-dependent radiosensitization in TNBC xenografts but not in isogenic RB1-null xenografts. Our data provide the preclinical rationale for a clinical trial expanding the use of CDK4/6i + RT to difficult-to-control RB-intact breast cancers (including TNBC) and nominate RB status as a predictive biomarker of therapeutic efficacy.

Authors

Andrea M. Pesch, Nicole H. Hirsh, Anna R. Michmerhuizen, Kassidy M. Jungles, Kari Wilder-Romans, Benjamin C. Chandler, Meilan Liu, Lynn M. Lerner, Charles A. Nino, Connor Ward, Erin F. Cobain, Theodore S. Lawrence, Lori J. Pierce, James M. Rae, Corey W. Speers

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Figure 6

RB is required for efficient repair of dsDNA breaks through HR.

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RB is required for efficient repair of dsDNA breaks through HR. (A–D) RA...
(A–D) RAD51 foci formation was used to assess HR competency with transient (A and B) knockdown or CRISPR-KO (C and D) of RB1 in MCF-7 and MDA-MB-231 cells at 6 hours after radiation. Cells transfected with a control siRNA or cells expressing Cas9 with control guides (AAVS1) were used for comparison. (E–G) Immunoprecipitation of GFP-RB and myc-RAD51 was performed 24 hours after transfection of HEK-293T cells (E), MCF-7 cells (F), or MDA-MB-231 cells (G) using myc-trap beads (black boxes), GFP-trap beads (green boxes), or control beads (white boxes). Expression of myc-RAD51 and GFP-RB was assessed in protein inputs and IP lysates with Western blotting. In siRNA transfected cells, a 1-way ANOVA was used to compare treatment groups. For CRISPR experiments performed across multiple cell lines, 2-tailed t tests were performed between paired radiation, and combination-treated groups within each cell line and correction was performed for multiple comparisons. *P < 0.05; **P < 0.01; ***P < 0.001, ****P < 0.0001. All experiments represent the mean of n = 3 independent experiments.