Despite major advances in understanding the molecular and genetic basis of cancer, metastasis remains the cause of >90% of cancer-related mortality1. Understanding metastasis initiation and progression is critical to developing new therapeutic strategies to treat and prevent metastatic disease. Prevailing theories hypothesize that metastases are seeded by rare tumour cells with unique properties, which may function like stem cells in their ability to initiate and propagate metastatic tumours2–5. However, the identity of metastasis-initiating cells in human breast cancer remains elusive, and whether metastases are hierarchically organized is unknown2. Here we show at the single-cell level that early stage metastatic cells possess a distinct stem-like gene expression signature. To identify and isolate metastatic cells from patient-derived xenograft models of human breast cancer, we developed a highly sensitive fluorescence-activated cell sorting (FACS)-based assay, which allowed us to enumerate metastatic cells in mouse peripheral tissues. We compared gene signatures in metastatic cells from tissues with low versus high metastatic burden. Metastatic cells from low-burden tissues were distinct owing to their increased expression of stem cell, epithelial-to-mesenchymal transition, pro-survival, and dormancy-associated genes. By contrast, metastatic cells from high-burden tissues were similar to primary tumour cells, which were more heterogeneous and expressed higher levels of luminal differentiation genes. Transplantation of stem-like metastatic cells from low-burden tissues showed that they have considerable tumour-initiating capacity, and can differentiate to produce luminal-like cancer cells. Progression to high metastatic burden was associated with increased proliferation and MYC expression, which could be attenuated by treatment with cyclin-dependent kinase (CDK) inhibitors. These findings support a hierarchical model for metastasis, in which metastases are initiated by stem-like cells that proliferate and differentiate to produce advanced metastatic disease.

Hepatoblastoma, the most common pediatric liver cancer, is tightly linked to excessive Wnt/beta-catenin signaling. Here, we used microarray analysis to identify two tumor subclasses resembling distinct phases of liver development and a discriminating 16-gene signature. beta-catenin activated different transcriptional programs in the two tumor types, with distinctive expression of hepatic stem/progenitor markers in immature tumors. This highly proliferating subclass was typified by gains of chromosomes 8q and 2p and upregulated Myc signaling. Myc-induced hepatoblastoma-like tumors in mice strikingly resembled the human immature subtype, and Myc downregulation in hepatoblastoma cells impaired tumorigenesis in vivo. Remarkably, the 16-gene signature discriminated invasive and metastatic hepatoblastomas and predicted prognosis with high accuracy.

Summary Protein misfolding stimulates a signaling pathway involving noncoding RNAs to promote cell death. The endoplasmic reticulum (ER) is the primary organelle for folding and maturation of secretory and transmembrane proteins. Inability to meet protein-folding demand leads to “ER stress,” and activates IRE1α, an ER transmembrane kinase-endoribonuclease (RNase). IRE1α promotes adaptation through splicing Xbp1 mRNA or apoptosis through incompletely understood mechanisms. Here we found that sustained IRE1α RNase activation caused rapid decay of select microRNAs (miRs -17, -34a, -96, -125b) that normally repress translation of Caspase-2 mRNA, and thus sharply elevates protein levels of this initiator protease of the mitochondrial apoptotic pathway. In cell-free systems, recombinant IRE1α endonucleolytically cleaved microRNA precursors at sites distinct from DICER. Thus, IRE1α regulates translation of a proapoptotic protein through terminating microRNA biogenesis, and noncoding RNAs are part of the ER stress response.

Enhancing chemotherapeutic efficiency through improved drug delivery would facilitate treatment of chemoresistant cancers, such as recurrent mammary tumors and liver cancer. One way to improve drug delivery is through the use of nanodiamond (ND) therapies, which are both scalable and biocompatible. Here, we examined the efficacy of an ND-conjugated chemotherapeutic in mouse models of liver and mammary cancer. A complex (NDX) of ND and doxorubicin (Dox) overcame drug efflux and significantly increased apoptosis and tumor growth inhibition beyond conventional Dox treatment in both murine liver tumor and mammary carcinoma models. Unmodified Dox treatment represents the clinical standard for most cancer treatment regimens, and NDX had significantly decreased toxicity in vivo compared to standard Dox treatment. Thus, ND-conjugated chemotherapy represents a promising, biocompatible strategy for overcoming chemoresistance and enhancing chemotherapy efficacy and safety.