Comparative oncogenomics identifies breast tumors enriched in functional tumor-initiating cells

Abstract

The claudin-low subtype is a recently identified rare molecular subtype of human breast cancer that expresses low levels of tight and adherens junction genes and shows high expression of epithelial-to-mesenchymal transition (EMT) genes. These tumors are enriched in gene expression signatures derived from human tumor-initiating cells (TICs) and human mammary stem cells. Through cross-species analysis, we discovered mouse mammary tumors that have similar gene expression characteristics as human claudin-low tumors and were also enriched for the human TIC signature. Such claudin-low tumors were similarly rare but came from a number of distinct mouse models, including the p53 null transplant model. Here we present a molecular characterization of 50 p53 null mammary tumors compared with other mouse models and human breast tumor subtypes. Similar to human tumors, the murine p53 null tumors fell into multiple molecular subtypes, including two basal-like, a luminal, a claudin-low, and a subtype unique to this model. The claudin-low tumors also showed high gene expression of EMT inducers, low expression of the miR-200 family, and low to absent expression of both claudin 3 and E-cadherin. These murine subtypes also contained distinct genomic DNA copy number changes, some of which are similarly altered in their cognate human subtype counterpart. Finally, limiting dilution transplantation revealed that p53 null claudin-low tumors are highly enriched for TICs compared with the more common adenocarcinomas arising in the same model, thus providing a unique preclinical mouse model to investigate the therapeutic response of TICs.

Fig. 1.

Intrinsic gene set clustering analysis of 50 p53 null tumors and 117 samples from 13 GEMM previously published in Herschkowitz et al. (13). (A) Overview of the complete cluster diagram. (B) Experimental sample-associated dendrogram. Boxes indicate the p53 null tumor subtypes based on SigClust analysis. (C) Luminal epithelial expression pattern that is highly expressed in luminal p53 null tumors, MMTV-Neu, and MMTV-PyMT tumors (D). Basal epithelial expression pattern including K5 and ID4, which are highly expressed in basal-like p53 null tumors. (E) Mesenchymal genes, including snail homolog 1. (F) Genes expressed at low levels in claudin-low tumors, including CLDN3, CLDN7, and ELF5. (G) Proliferation signature. (H) Individual genes discussed within the text.

Fig. 2.

p53 null claudin-low tumors have features of EMT. These tumors (A) express core EMT signature, EMT markers, and (B) showed marked down-regulation of miRNAs involved in negative regulation of stemness and EMT. Three technical replicates were averaged for each. 200a (P < 0.0001), 200b (P = 0.0002), 200c (P < 0.0001), 141 (P = 0.0005), 429 (P < 0.0001), 205 (P = 0.004), 182 (P = 0.004), 96 (P = 0.005), 183 (P < 0.0001), 203 (P = 0.04).

Fig. 3.

Tumor genomic DNA copy number landscape plots for mouse p53 null tumor classes. At top is the overall pattern for all 34 tumors considered together, and then below are the landscape plots for each of the five expression-defined subtypes. Gray shading indicates the overall frequency of aberrations seen in that subtype, and the black shading indicates the group-specific CNA (P value threshold 0.05).

Fig. 4.

p53 null claudin-low tumors express markers of stem cells and are enriched for tumor-initiating ability. (A) Claudin-low tumors have high percentages of double-positive (CD29⁺/CD24⁺) cells compared with (B) other p53 null tumors. (C) Limiting dilution transplantation of claudin-low vs. adenocarcinoma cells. Sample sizes are implied by the sizes of the circles (area is proportional to sample size).