Project description:For many breast cancer (BCa) patients, symptomatic bone metastases appear after years or even decades of latency. How metastatic cells disseminate, and how micrometastatic lesions remain dormant and undetectable yet initiate colonization, are major questions in cancer research. Here we identify and functionally analyse a molecular mechanism involved in bone metastatic latency of estrogen receptor–positive (ER)+ BCa. We developed an in vivo loss-of-function, genome-wide shRNA screening to identify genes relevant for long-latent relapse in BCa. This screen revealed the kinase MSK1 as an important regulator of metastatic dormancy. Importantly, low MSK1 expression associates with early metastasis in ER+ BCa patients. Reduced MSK1 levels impaired cellular differentiation and increased the bone homing and growth capacity of metastatic cells. MSK1 modulates chromatin status at promoters to regulate the expression of luminal differentiation genes, including those for the GATA-3 and FOXA1 transcription factors, which prevent the progression of ER+ BCa towards metastasis. Our results identify the regulation of luminal cell differentiation by MSK1 via modulation of chromatin remodelling to be a key mechanism for controlling metastatic dormancy in BCa. We propose that MSK1 could be a useful marker for stratifying BCa patients as high- or low-risk for early relapse, allowing patients to receive appropriate treatments.
Project description:The transcription factor GATA3 is essential for luminal cell differentiation during mammary gland development and critical for formation of the luminal subtypes of breast cancer. Ectopic expression of GATA3 promoted global alterations of the transcriptome of basal triple-negative breast cancer cells resulting in molecular and cellular changes associated with a more differentiated, luminal tumor subtype and a concomitant reduction in primary tumor growth, lung metastasis, and macrophage recruitment at the metastatic site. Importantly, we demonstrate that the inhibition of metastases by GATA3 results from the suppression of lysyl oxidase (LOX) expression, a metastasis promoting matrix protein that affects cell proliferation, cross-linking of extracellular collagen types, and establishment of the metastatic niche. There are 2 samples sent in triplicates.
Project description:Despite widespread knowledge that bone marrow-resident breast cancer cells (BMRCs) affect tumor progression, signaling mechanisms of BMRCs implicated in maintaining long-term dormancy have not been characterized. To overcome these hurdles, we developed a novel experimental model of tumor dormancy employing circulating tumor cells (CTCs) derived from metastatic breast cancer patients (de novo CTCs), transplanted them in immunocompromised mice, and re-isolated these cells from xenografted mice bone marrow (ex vivo BMRCs) and blood (ex vivo CTCs) to perform downstream transcriptomic analyses. Here we report that the balance between mTORC1 vs mTORC2 signaling regulates BMRC mitotic and/or dormancy characteristics.
Project description:The mechanisms regulating breast cancer differentiation state are poorly understood. Of particular interest are molecular regulators controlling the highly aggressive and poorly differentiated traits of basal-like breast carcinomas. Here we show that the Polycomb factor EZH2 maintains the differentiation state of basal-like breast cancer cells, and promotes the expression of progenitor-associated and basal-lineage genes. Specifically, EZH2 regulates the composition of basal-like breast cancer cell populations by promoting a M-bM-^@M-^\bi-lineageM-bM-^@M-^] differentiation state, in which cells co-express basal- and luminal-lineage markers. We show that human basal-like breast cancers contain a subpopulation of bi-lineage cells, and that EZH2-deficient cells give rise to tumors with a decreased proportion of such cells. Bi-lineage cells express genes that are active in normal luminal progenitors, and possess increased colony formation capacity, consistent with a primitive differentiation state. We found that GATA3, a driver of luminal differentiation, performs a function opposite to EZH2, acting to suppress bi-lineage identity and luminal progenitor gene expression. GATA3 levels increase upon EZH2 silencing, leading to the observed decrease in bi-lineage cell numbers. Our findings reveal a novel role for EZH2 in controlling basal-like breast cancer differentiation state and intra-tumoral cell composition. Total of four treatments (HCC70 cells stably expressing shEZH2, shEED, or EZH2 cDNA, and MDA-MB-468 cells stably expressing shEZH2) were done in duplicates, each with its own control.
Project description:The transcription factor GATA3 is essential for luminal cell differentiation during mammary gland development and critical for formation of the luminal subtypes of breast cancer. Ectopic expression of GATA3 promoted global alterations of the transcriptome of basal triple-negative breast cancer cells resulting in molecular and cellular changes associated with a more differentiated, luminal tumor subtype and a concomitant reduction in primary tumor growth, lung metastasis, and macrophage recruitment at the metastatic site. Importantly, we demonstrate that the inhibition of metastases by GATA3 results from the suppression of lysyl oxidase (LOX) expression, a metastasis promoting matrix protein that affects cell proliferation, cross-linking of extracellular collagen types, and establishment of the metastatic niche.
Project description:Molecular signaling that regulates differentiation, survival and proliferation of the prostate luminal epithelial cells has not been thoroughly understood. Herein, we show that increased canonical Notch1 activity suppresses terminal differentiation of prostate luminal epithelial cells but is insufficient to transform. Augmented Notch1 activity delays anoikis of luminal epithelial cells by augmenting NF-κB activity independent of Hes-1, stimulates luminal cell proliferation by potentiating the PI3K-AKT signaling, and rescues the capacities of a fraction of prostate luminal epithelial cells for unipotent differentiation in vivo and short-term self-renewal in vitro. Epithelial cell-autonomous AR signaling is dispensable for the Notch-mediated cellular survival and proliferation. This study reveals a previously unappreciated role of Notch in prostatic luminal epithelial cell differentiation, supports the presence of a lineage hierarchy within the prostate luminal epithelial cells, and implies a pro-metastatic function of Notch signaling during prostate cancer progression. Two group comparison (WT and Mut)
Project description:Although the estrogen receptor (ER) positive variant of breast cancer is touted as the most indolent and favorable, the majority of breast cancer deaths are in fact from this subtype. There are several features of this category of breast cancers that likely account for this outcome. The first is that metastatic relapse can occur many years after initial diagnosis of primary disease. The second is that once the cancer cells awaken into full-blown metastatic disease, they are largely resistant to ER-directed therapies (i.e. hormonal therapy, HT). The third is that when metastases do occur, they are invariably in many locations. This observation suggests that these dormant/sleeping metastatic cells are “globally” awakened as if by a “systemic” infection. We suggest that these three processes be not only linked, but underlie the lethal features of metastatic disease. We hypothesized that mtDNA is necessary for the escape from therapy induced tumor dormancy of luminal breast cancer cells
Project description: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 develop new therapeutic strategies to treat and prevent metastatic disease. Prevailing theories hypothesize that metastases are seeded by rare tumor cells with unique properties, which may function like stem cells in their ability to initiate and propagate metastatic tumors.2 3-5 However, the identity of metastasis-initiating cells in human breast cancer remains elusive, and whether metastases are hierarchically organized is unknown.2 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 (PDX) models of human breast cancer, we developed a highly sensitive 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 vs. high metastatic burden. Metastatic cells from low-burden tissues were distinct due to their increased expression of stem cell, EMT, pro-survival, and dormancy-associated genes. In contrast, metastatic cells from high-burden tissues were similar to primary tumor 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 significant tumor-initiating capacity, and differentiate to produce luminal-like cancer cells. Progression to high metastatic burden was associated with increased proliferation and cMYC expression, which could be attenuated by treatment with cyclin dependent kinase (CDK) inhibitors. These findings support a hierarchical model for metastasis, where metastases are initiated by stem-like cells that proliferate and differentiate to produce advanced metastatic disease.
Project description: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 develop new therapeutic strategies to treat and prevent metastatic disease. Prevailing theories hypothesize that metastases are seeded by rare tumor cells with unique properties, which may function like stem cells in their ability to initiate and propagate metastatic tumors.2 3-5 However, the identity of metastasis-initiating cells in human breast cancer remains elusive, and whether metastases are hierarchically organized is unknown.2 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 (PDX) models of human breast cancer, we developed a highly sensitive 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 vs. high metastatic burden. Metastatic cells from low-burden tissues were distinct due to their increased expression of stem cell, EMT, pro-survival, and dormancy-associated genes. In contrast, metastatic cells from high-burden tissues were similar to primary tumor 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 significant tumor-initiating capacity, and differentiate to produce luminal-like cancer cells. Progression to high metastatic burden was associated with increased proliferation and cMYC expression, which could be attenuated by treatment with cyclin dependent kinase (CDK) inhibitors. These findings support a hierarchical model for metastasis, where metastases are initiated by stem-like cells that proliferate and differentiate to produce advanced metastatic disease.