Yilong Zou

Therapy-induced tumour secretomes promote resistance and tumour progression.

Drug resistance invariably limits the clinical efficacy of targeted therapy with kinase inhibitors against cancer. Here we show that targeted therapy with BRAF, ALK or EGFR kinase inhibitors induces a complex network of secreted signals in drug-stressed human and mouse melanoma and human lung adenocarcinoma cells. This therapy-induced secretome stimulates the outgrowth, dissemination and metastasis of drug-resistant cancer cell clones and supports the survival of drug-sensitive cancer cells, contributing to incomplete tumour regression. The tumour-promoting secretome of melanoma cells treated with the kinase inhibitor vemurafenib is driven by downregulation of the transcription factor FRA1. In situ transcriptome analysis of drug-resistant melanoma cells responding to the regressing tumour microenvironment revealed hyperactivation of several signalling pathways, most prominently the AKT pathway. Dual inhibition of RAF and the PI(3)K/AKT/mTOR intracellular signalling pathways blunted the outgrowth of the drug-resistant cell population in BRAF mutant human melanoma, suggesting this combination therapy as a strategy against tumour relapse. Thus, therapeutic inhibition of oncogenic drivers induces vast secretome changes in drug-sensitive cancer cells, paradoxically establishing a tumour microenvironment that supports the expansion of drug-resistant clones, but is susceptible to combination therapy.

Selection of Bone Metastasis Seeds by Mesenchymal Signals in the Primary Tumor Stroma

How organ-specific metastatic traits arise in primary tumors remains unknown. Here, we show a role of the breast tumor stroma in selecting cancer cells that are primed for metastasis in bone. Cancer-associated fibroblasts (CAFs) in triple-negative (TN) breast tumors skew heterogeneous cancer cell populations toward a predominance of clones that thrive on the CAF-derived factors CXCL12 and IGF1. Limiting concentrations of these factors select for cancer cells with high Src activity, a known clinical predictor of bone relapse and an enhancer of PI3K-Akt pathway activation by CXCL12 and IGF1. Carcinoma clones selected in this manner are primed for metastasis in the CXCL12-rich microenvironment of the bone marrow. The evidence suggests that stromal signals resembling those of a distant organ select for cancer cells that are primed for metastasis in that organ, thus illuminating the evolution of metastatic traits in a primary tumor and its distant metastases.

Metastatic Latency and Immune Evasion through Autocrine Inhibition of WNT.

Metastasis frequently develops years after the removal of a primary tumor, from a minority of disseminated cancer cells that survived as latent entities through unknown mechanisms. We isolated latency competent cancer (LCC) cells from early stage human lung and breast carcinoma cell lines and defined the mechanisms that suppress outgrowth, support long-term survival, and maintain tumor-initiating potential in these cells during the latent metastasis stage. LCC cells show stem-cell-like characteristics and express SOX2 and SOX9 transcription factors, which are essential for their survival in host organs under immune surveillance and for metastatic outgrowth under permissive conditions. Through expression of the WNT inhibitor DKK1, LCC cells self-impose a slow-cycling state with broad downregulation of ULBP ligands for NK cells and evasion of NK-cell-mediated clearance. By expressing a Sox-dependent stem-like state and actively silencing WNT signaling, LCC cells can enter quiescence and evade innate immunity to remain latent for extended periods.

TGF-β Tumor Suppression through a Lethal EMT

TGF-β signaling can be pro-tumorigenic or tumor suppressive. We investigated this duality in pancreatic ductal adenocarcinoma (PDA), which, with other gastrointestinal cancers, exhibits frequent inactivation of the TGF-β mediator Smad4. We show that TGF-β induces an epithelial-mesenchymal transition (EMT), generally considered a pro-tumorigenic event. However, in TGF-β-sensitive PDA cells, EMT becomes lethal by converting TGF-β-induced Sox4 from an enforcer of tumorigenesis into a promoter of apoptosis. This is the result of an EMT-linked remodeling of the cellular transcription factor landscape, including the repression of the gastrointestinal lineage-master regulator Klf5. Klf5 cooperates with Sox4 in oncogenesis and prevents Sox4-induced apoptosis. Smad4 is required for EMT but dispensable for Sox4 induction by TGF-β. TGF-β-induced Sox4 is thus geared to bolster progenitor identity, whereas simultaneous Smad4-dependent EMT strips Sox4 of an essential partner in oncogenesis. Our work demonstrates that TGF-β tumor suppression functions through an EMT-mediated disruption of a lineage-specific transcriptional network.

Loss of the multifunctional RNA-binding protein RBM47 as a source of selectable metastatic traits in breast cancer

The mechanisms through which cancer cells lock in altered transcriptional programs in support of metastasis remain largely unknown. Through integrative analysis of clinical breast cancer gene expression datasets, cell line models of breast cancer progression, and mutation data from cancer genome resequencing studies, we identified RNA binding motif protein 47 (RBM47) as a suppressor of breast cancer progression and metastasis. RBM47 inhibited breast cancer re-initiation and growth in experimental models. Transcriptome-wide HITS-CLIP analysis revealed widespread RBM47 binding to mRNAs, most prominently in introns and 3′UTRs. RBM47 altered splicing and abundance of a subset of its target mRNAs. Some of the mRNAs stabilized by RBM47, as exemplified by dickkopf WNT signaling pathway inhibitor 1, inhibit tumor progression downstream of RBM47. Our work identifies RBM47 as an RNA-binding protein that can suppress breast cancer progression and demonstrates how the inactivation of a broadly targeted RNA chaperone enables selection of a pro-metastatic state. DOI: http://dx.doi.org/10.7554/eLife.02734.001

Pericyte-like spreading by disseminated cancer cells activates YAP and MRTF for metastatic colonization

Metastatic seeding by disseminated cancer cells principally occurs in perivascular niches. Here, we show that mechanotransduction signalling triggered by the pericyte-like spreading of disseminated cancer cells on host tissue capillaries is critical for metastatic colonization. Disseminated cancer cells employ L1CAM (cell adhesion molecule L1) to spread on capillaries and activate the mechanotransduction effectors YAP (Yes-associated protein) and MRTF (myocardin-related transcription factor). This spreading is robust enough to displace resident pericytes, which also use L1CAM for perivascular spreading. L1CAM activates YAP by engaging β1 integrin and ILK (integrin-linked kinase). L1CAM and YAP signalling enables the outgrowth of metastasis-initiating cells both immediately following their infiltration of target organs and after they exit from a period of latency. Our results identify an important step in the initiation of metastatic colonization, define its molecular constituents and provide an explanation for the widespread association of L1CAM with metastatic relapse in the clinic.Massagué and colleagues show that disseminated cancer cells use L1CAM to spread on capillaries and to achieve their outgrowth through activating YAP signalling.

Structural basis for genome wide recognition of 5-bp GC motifs by SMAD transcription factors.

Smad transcription factors activated by TGF-β or by BMP receptors form trimeric complexes with Smad4 to target specific genes for cell fate regulation. The CAGAC motif has been considered as the main binding element for Smad2/3/4, whereas Smad1/5/8 have been thought to preferentially bind GC-rich elements. However, chromatin immunoprecipitation analysis in embryonic stem cells showed extensive binding of Smad2/3/4 to GC-rich cis-regulatory elements. Here, we present the structural basis for specific binding of Smad3 and Smad4 to GC-rich motifs in the goosecoid promoter, a nodal-regulated differentiation gene. The structures revealed a 5-bp consensus sequence GGC(GC)|(CG) as the binding site for both TGF-β and BMP-activated Smads and for Smad4. These 5GC motifs are highly represented as clusters in Smad-bound regions genome-wide. Our results provide a basis for understanding the functional adaptability of Smads in different cellular contexts, and their dependence on lineage-determining transcription factors to target specific genes in TGF-β and BMP pathways.Smad transcription factors are part of the TGF-β signal transduction pathways and are recruited to the genome by cell lineage-defining factors. Here, the authors identify specific Smad binding GC-rich motifs and provide structural information showing Smad3 and Smad4 bound to these motifs.