Xenografts faithfully recapitulate breast cancer-specific gene expression patterns of parent primary breast tumors

Abstract

Though xenografts are used extensively for drug development in breast cancer, how well xenografts reflect the breadth of primary breast tumor subtypes has not been well characterized. Moreover, few studies have compared the gene expression of xenograft tumors to the primary tumors from which they were derived. Here we investigate whether the ability of human breast tumors (n = 20) to create xenografts in immune-deficient mice is associated with breast cancer immunohistochemical (IHC) and intrinsic subtype. We also characterize how precisely the gene expression of xenografts reprises that of parent breast tumors, using hierarchical clustering and other correlation-based techniques applied to Agilent 44K gene expression data from 16 samples including four matched primary tumor-xenograft pairs. Of the breast tumors studied, 25 % (5/20) generated xenografts. Receptor and intrinsic subtype were significant predictors of xenograft success, with all (4/4) triple-negative (TN) tumors and no (0/12) HR+Her2- tumors forming xenografts (P = 0.0005). Tumor cell expression of ALDH1, a stem cell marker, trended toward successful engraftment (P = 0.14), though CDK5/6, a basal marker, did not. Though hierarchical clustering across the 500 most variable genes segregated human breast tumors from xenograft tumors, when clustering was performed over the PAM50 gene set the primary tumor-xenograft pairs clustered together, with all IHC subtypes clustered in distinct groups. Greater similarity between primary tumor-xenograft pairs relative to random pairings was confirmed by calculation of the within-pair between-pair scatter ratio (WPBPSR) distribution (P = 0.0269), though there was a shift in the xenografts toward more aggressive features including higher proliferation scores relative to the primary. Triple-negative breast tumors demonstrate superior ability to create xenografts compared to HR+ tumors, which may reflect higher proliferation or relatively stroma-independent growth of this subtype. Xenograft tumors' gene expression faithfully resembles that of their parent tumors, yet also demonstrates a shift toward more aggressive molecular features.

Fig. 1

Hematoxylin and eosin stain of B34 primary tumor (a), xenograft passage 1 (b), and xenograft passage 2 (c) tumors. The morphological characteristics of this tumor were maintained between each passage. Primary tumor consisted of moderately poorly differentiated tumor cells with small regions of human stroma (a). Passaged tumors consisted of highly proliferative moderately poorly differentiated tumor cells with well-vascularized murine stroma (b, c)

Fig. 2

Aldehyde dehydrogenase 1 staining. DAB staining (arrow) is ALDH1 staining in tumor cells, a marker of cancer stem cells. Two cases positive for ALDH1 are shown above

Fig. 3

Heatmap plot of hierarchical clustering result using top 500 most variable genes determined by the standard deviation of log-intensities across all arrays in the experiment. Group A is human breast primary tumors that created xenografts, group B is primary tumors that did not create xenografts, and group C is xenografts

Fig. 4

a Unsupervised clustering of primary tumors and xenografts over PAM-50 genes. Heatmap rows are PAM-50 genes, columns are individual tumors. Top dendrogram depicts relationships among tumors by gene expression. b Two-dimensional representation of distance between all samples by multidimensional scaling of gene expression for PAM-50 genes. Three xenograft-primary pairs are circled in red; the fourth pair (BX-B51 and X-B51) is indicated by red arrows. Color key for the sample annotation strips in (a): Her2pos and ERpos: dark pink = positive, black = negative, light pink = missing; Xeno: orange = xenograft; black = primary tumor; Pairs: xenograft-primary pairs are same color; subtype: red = Basal; dark blue = LumA; light blue = LumB; pink = Her2; green = normal

Fig. 5

a Correlation coefficients of PAM50 gene expression of xenograft-primary pairs compared to all possible pairs of xenografts and xenograft-generating primary tumors. (P = 0.0056). b Within-pair between-pair scatter ratio. Xenograft-primary pairs indicated with red arrow. c Correlation barplot of individual PAM50 genes. The height of each bar represents the Pearson correlation coefficient of xenograft-primary pairs for a gene