Target Discovery and Validation Reviews and Protocols

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termed luminal subtype A (dark blue branches), demonstrated the highest expression
of ER, estrogen-regulated protein LIV-1, and the transcription factors
hepatocyte nuclear factor 3, alpha (HNF3A), X-box binding protein 1, and
GATA-binding protein 3 (GATA3) (Fig. 3G). The second, smaller group of
tumors with luminal epithelial characteristics, termed luminal subtype B (light
blue branches), showed low expression of the ER cluster, but it was further distinguished
from luminal subtype A by the high expression of a novel set of
genes such as GGH, LAPTMB4, NSEP1, and CCNE1, whose coordinated function
is unknown (Fig. 3D). Among the three ER-negative groups, the basal
epithelial-like subtype (red branches) was characterized by high expression of
KRT5 and KRT17, annexin 8, CX3CL1, and TRIM29 and was completely
negative for the luminal/ER cluster of genes (Fig. 3E), whereas the ERBB2+
subtype (pink) was characterized by high expression of several genes in
the ERBB2 amplicon at 17q22.24, including ERBB2, GRB7, and TRAP100
(Fig. 3C), suggesting that much of the genetic influences are because of gene
amplification. A normal breast tissue-like group (green branches) was identified
that showed the highest expression of many genes known to be expressed by
adipose tissue and other nonepithelial cell types (Fig. 3F). These tumors also
showed strong expression of basal epithelial genes and low expression of luminal
epithelial genes. However, it is unclear whether these tumors represent
poorly sampled tumor tissue or a distinct, clinically important group. In conclusion,
the molecular portraits defined by gene expression patterns seem to be
stable and homogenous and represent the tumor itself, not merely the particular
tumor sample, because we could recognize the distinctive expression profile of
a tumor in independent samples. The inherent properties of the tumors thus
seem to be sustained throughout chemotherapy as well as between a primary
tumor and its lymph node metastasis, and these properties could be represented
by a relatively small number of genes whose variation in expression formed a
platform for classification.
3.3. Validation
The robustness of the tumor subtypes has been validated in independent
data sets by us (22) as well as by others (24–29). When conducting a similar
analysis of a breast tumor data set published by van’t Veer et al. (30), to display
the expression pattern of 460 of the intrinsic genes previously mentioned
across their 97 tumor samples, the same subtypes were noted. In particular,
the basal-like subtype was the most homogenous group of tumors, tightly
clustered together with high correlation to the intrinsic expression centroid
representing this phenotype (22). This relationship has been true for all
data sets analyzed this far, even though they have comprised different patient
populations, indicating that the substantial differences in the characteristics of