RB expression confers sensitivity to CDK4/6 inhibitor–mediated radiosensitization across breast cancer subtypes
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
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
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 2

CDK4/6 inhibition suppresses HR in RB WT TNBC.

Options: View larger image (or click on image) Download as PowerPoint
CDK4/6 inhibition suppresses HR in RB WT TNBC. (A–C) A GFP reporter syst...
(A–C) A GFP reporter system was used to measure relative HR repair efficiency in RB WT MCF-7 (A) and MDA-MB-231 (B and C) cells after treatment with in IC50 concentration of palbociclib (gray), ribociclib (blue), or abemaciclib (green). A CHK1/2 inhibitor (200 nM AZD7762) was used as a positive control, and a DNAPK inhibitor (1.5 μM NU7441) was used as the negative control. For the reporter assay a 1-way ANOVA with Dunnett’s post hoc test was used to compare treatment groups to DMSO-treated cells. *P < 0.05; **P < 0.01; ***P < 0.001, ****P < 0.0001. (D, E, G, and H) For RAD51 immunofluorescence, cells were pretreated for 1 hour with palbociclib, and coverslips were fixed 6 hours and 16 hours after 4 Gy radiation in RB WT MDA-MB-231 (D) and CAL-120 (E) cells, as well as RB-null MDA-MB-468 (G) and CAL-851 (H) TNBC cells. Two-tailed t tests were performed between radiation and combination treated groups at each RAD51 time point, and correction was performed for multiple comparisons. (F and I) Western blots were used to assess RAD51 protein expression at the same time points. All experiments represent the average of 3 independent experiments and bar graphs display the average ± SEM.