Therapeutic response to CDK4/6 inhibition in breast cancer defined by ex vivo analyses of human tumors

Abstract

To model the heterogeneity of breast cancer as observed in the clinic, we employed an ex vivo model of breast tumor tissue. This methodology maintained the histological integrity of the tumor tissue in unselected breast cancers, and importantly, the explants retained key molecular markers that are currently used to guide breast cancer treatment (e.g., ER and Her2 status). The primary tumors displayed the expected wide range of positivity for the proliferation marker Ki67, and a strong positive correlation between the Ki67 indices of the primary and corresponding explanted tumor tissues was observed. Collectively, these findings indicate that multiple facets of tumor pathophysiology are recapitulated in this ex vivo model. To interrogate the potential of this preclinical model to inform determinants of therapeutic response, we investigated the cytostatic response to the CDK4/6 inhibitor, PD-0332991. This inhibitor was highly effective at suppressing proliferation in approximately 85% of cases, irrespective of ER or HER2 status. However, 15% of cases were completely resistant to PD-0332991. Marker analyses in both the primary tumor tissue and the corresponding explant revealed that cases resistant to CDK4/6 inhibition lacked the RB-tumor suppressor. These studies provide important insights into the spectrum of breast tumors that could be treated with CDK4/6 inhibitors, and defines functional determinants of response analogous to those identified through neoadjuvant studies.

Figures

Figure 1. Explants retain tissue architecture and clinical biomarkers. (A) Schematic of the explant procedure utilized in this study. (B) Representative hematoxylin/eosin, Her2 and ER staining between explant and surgical specimens. (C) Histological scoring of staining of primary tumor tissue and explants with Her2 and ER show concordance of staining.

Figure 2. Proliferative index in explants is consistent with that observed in the surgical specimen. (A) Representative Ki67 staining from surgical specimen and associated explants. (B) Quantification of Ki67 staining from matched explant and surgical specimens demonstrating concordance between the two. Data was analyzed by paired t-test. Correlation coefficient 0.9385 and p < 0.0001 for concordance in the measures.

Figure 3. Differential response to CDK4/6 inhibition in breast cancer explants. (A) Representative Ki67 of explants treated with DMSO (vehicle) or PD-0332991. (B) Quantification of Ki67 from PD-0332991 treated cultures. (C) Example of normal tissue, matched tumor and lymph-node metastasis stained for Ki67.

Figure 4. p16ink4a and RB status are associated with the response to PD-0332991 in tumor specimens. (A) Representative staining of p16ink4a and RB in surgical specimens and explants. (B) Association of p16ink4a and RB status in response to PD-0332991 among explanted tissue cultures. Cases that did not respond to PD-0332991 are boxed in red.