Refinement of breast cancer molecular classification by miRNA expression profiles

Rolf Søkilde, Helena Persson, Anna Ehinger, Anna Chiara Pirona, Mårten Fernö, Cecilia Hegardt, Christer Larsson, Niklas Loman, Martin Malmberg, Lisa Rydén, Lao Saal, Åke Borg, Johan Vallon-Christerson, Carlos Rovira, Rolf Søkilde, Helena Persson, Anna Ehinger, Anna Chiara Pirona, Mårten Fernö, Cecilia Hegardt, Christer Larsson, Niklas Loman, Martin Malmberg, Lisa Rydén, Lao Saal, Åke Borg, Johan Vallon-Christerson, Carlos Rovira

Abstract

Background: Accurate classification of breast cancer using gene expression profiles has contributed to a better understanding of the biological mechanisms behind the disease and has paved the way for better prognostication and treatment prediction.

Results: We found that miRNA profiles largely recapitulate intrinsic subtypes. In the case of HER2-enriched tumors a small set of miRNAs including the HER2-encoded mir-4728 identifies the group with very high specificity. We also identified differential expression of the miR-99a/let-7c/miR-125b miRNA cluster as a marker for separation of the Luminal A and B subtypes. High expression of this miRNA cluster is linked to better overall survival among patients with Luminal A tumors. Correlation between the miRNA cluster and their precursor LINC00478 is highly significant suggesting that its expression could help improve the accuracy of present day's signatures.

Conclusions: We show here that miRNA expression can be translated into mRNA profiles and that the inclusion of miRNA information facilitates the molecular diagnosis of specific subtypes, in particular the clinically relevant sub-classification of luminal tumors.

Keywords: Breast cancer; Differential expression; LINC00478; Mir-4728; Molecular subtypes; Non-coding RNA; miR-99a/let-7c/miR-125b; microRNA.

Conflict of interest statement

The authors declare that they have no competing interests.

Figures

Fig. 1
Fig. 1
Consensus clustering identified three main tumors clusters with distinct sample composition. Enrichment of PAM50 subtypes and clinical parameters within cluster was evaluated using the χ2 test and p-values are given in parenthesis
Fig. 2
Fig. 2
Sample clustering and heatmap for the expression of a set of 73 unique miRNAs collected from the 15 most significantly differentially expressed miRNAs from each comparison between tumor subtypes. Samples are colored by subtype according to the PAM50 classification. Red = Basal-like, purple = HER2-enriched, blue = Luminal A, and cyan = Luminal B. The miRNA expression values were standardized by row mean centering and dividing by row standard deviation in R before distance calculation and clustering
Fig. 3
Fig. 3
a Sample clustering and heatmap for the 15 most significant miRNAs differentially expressed between luminal A (blue) and luminal B (cyan) samples. b Corresponding gene signature activation using absolute inference of patient signatures (AIPS) model. The AIPS model uses gene expression signatures in the mRNA dataset to identify active and inactive biological processes. The AIPS algorithm assigns a random or independent value to samples, which do not show a clear active nor in-active process. Red = active, blue = inactive, white = Random/Independent
Fig. 4
Fig. 4
a Survival analysis based on the mean expression of the microRNAs in the miR-99a/let-7c/miR-125b miRNA cluster. We used the median to split the patients into miR99a high (higher than median expression) and miR-99a low (lower than median expression). These groups were used in a survival analysis with R survival package, stratified based on the pam50 group and plotted using the survminer package in R. b We used the median to split the patients into LINC00478 high (higher than median expression) and LINC00478 low (lower than median expression). These groups were used in a survival analysis with R survival package, stratified based on the pam50 group and plotted using the survminer package in R We found a significant benefit for Luminal A breast cancer patients of having higher than median LINC00478 expression. c A focused analysis was done on the Luminal A samples, with ER+, HER2-, node negative, endocrine treatment, no chemotherapy and no anti-HER2 treatment. This subgroup represents 725 patients

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