Metastatic breast cancer (BC) has a poor prognosis and is largely considered incurable. A better understanding of the molecular determinants of BC metastasis could facilitate development of improved prevention and treatment strategies. We used lentiviral barcoding coupled to single-cell RNA sequencing to trace clonal and transcriptional evolution during BC metastasis and showed that metastases derive from rare pro-metastatic clones that are under-represented in primary tumors. Both low clonal fitness and high metastatic potential were independent of clonal origin. Differential expression and classification analyses revealed that the pro-metastatic phenotype was acquired by rare cells characterized by the concomitant hyper-activation of extracellular matrix remodeling and dsRNA-interferon signaling pathways. Notably, genetic silencing of key genes in these pathways (KCNQ1OT1 or IFI6, respectively) significantly impaired migration in vitro and metastasis in vivo, with marginal effects on cell proliferation and tumor growth. Gene expression signatures derived from the identified pro-metastatic genes predict metastatic progression in BC patients, independently of known prognostic factors. This study elucidates previously unknown mechanisms of BC metastasis and provides prognostic predictors and therapeutic targets for metastasis prevention.
<div>Abstract<p>Metastatic breast cancer has a poor prognosis and is largely considered incurable. A better understanding of the molecular determinants of breast cancer metastasis could facilitate development of improved prevention and treatment strategies. We used lentiviral barcoding coupled to single-cell RNA sequencing to trace clonal and transcriptional evolution during breast cancer metastasis and showed that metastases derive from rare prometastatic clones that are underrepresented in primary tumors. Both low clonal fitness and high metastatic potential were independent of clonal origin. Differential expression and classification analyses revealed that the prometastatic phenotype was acquired by rare cells characterized by the concomitant hyperactivation of extracellular matrix remodeling and dsRNA-IFN signaling pathways. Notably, genetic silencing of key genes in these pathways (KCNQ1OT1 or IFI6, respectively) significantly impaired migration <i>in vitro</i> and metastasis <i>in vivo</i>, with marginal effects on cell proliferation and tumor growth. Gene expression signatures derived from the identified prometastatic genes predict metastatic progression in patients with breast cancer, independently of known prognostic factors. This study elucidates previously unknown mechanisms of breast cancer metastasis and provides prognostic predictors and therapeutic targets for metastasis prevention.</p>Significance:<p>Transcriptional lineage tracing coupled with single-cell transcriptomics defined the transcriptional programs underlying metastatic progression in breast cancer, identifying prognostic signatures and prevention strategies.</p></div>
The molecular determinants of breast cancer (BC) pro-metastatic phenotype are largely unknown. Here, we leveraged lentiviral barcoding coupled to single-cell RNA sequencing to trace clonal and transcriptional evolution during BC metastatization. We showed that metastases derive from rare pro-metastatic clones that are under-represented in primary tumors. Both low clonal-fitness and high metastatic-potential are independent of clonal origin. Differential expression and classification analyses revealed that the pro-metastatic phenotype is acquired in rare cells by concomitant hyper-activation of extracellular-matrix remodeling, dsRNA-interferon signaling, and stress-response pathways. Notably, genetic silencing of single pro-metastatic genes from different pathways significantly impairs migration in vitro and metastatization in vivo, with negligible effects on cell proliferation and tumor growth. In addition, gene-expression signatures from identified pro-metastatic genes predicts metastatic progression in BC patients, independently of known prognostic factors. This study elucidates previously unknown mechanisms of BC metastatization, and provides novel prognosis predictors and therapeutic targets for metastasis prevention.
<p>Supplementary Table including information about the predictive power of genes and signatures identified from metastatic clones and patient cohorts GEO accession numbers</p>
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