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.
View: Text | PDF
Research Article Oncology

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

×

Figure 4

RB is required for the radiosensitization of TNBC cell lines.

Options: View larger image (or click on image) Download as PowerPoint
RB is required for the radiosensitization of TNBC cell lines. (A–D) Clon...
(A–D) Clonogenic survival assays were performed in breast cancer cell lines with transient knockdown of RB1 (A and B) or CRISPR RB1-KO (C and D). (E) Average radiation enhancement ratios (rER) were compared between parental MDA-MB-231 cells and MDA-MB-231 RB1 CRISPR cells. (F–H) The efficiency of siRNA mediated knockdown (F), CRISPR RB1-KO (G), and RB overexpression (H) were assessed using Western blots, where transfected samples were harvested 48 hours after transfection. (I and J) RB was transiently overexpressed in MDA-MB-231 (I) and MCF-7 (J) RB1 CRISPR cell lines, and clonogenic survival assays were used to assess rescue of the radiosensitization phenotype. For CRISPR cells, a 1-way ANOVA with Dunnett’s post hoc test was used to compare CDK4/6 inhibitor–treated cells against vehicle-treated cells. For transfected cells, treatment pairs were compared with a 2-tailed Student’s t test and corrected for multiple comparisons. *P < 0.05; ***P < 0.001. All clonogenics represent the average of 3 experiments and are graphed as average ± SEM; Western blots are representative blots from n = 3 experiments.