TP53 mutations are frequent in esophageal squamous cell carcinoma (ESCC) and other SCCs and are a... more TP53 mutations are frequent in esophageal squamous cell carcinoma (ESCC) and other SCCs and are associated with a proclivity for metastasis. Here, we report that colony-stimulating factor-1 (CSF-1) expression is upregulated significantly in a p53-R172H–dependent manner in metastatic lung lesions of ESCC. The p53-R172H–dependent CSF-1 signaling, through its cognate receptor CSF-1R, increases tumor cell invasion and lung metastasis, which in turn is mediated in part through Stat3 phosphorylation and epithelial-to-mesenchymal transition (EMT). In Trp53R172H tumor cells, p53 occupies the Csf-1 promoter. The Csf-1 locus is enriched with histone 3 lysine 27 acetylation (H3K27ac), which is likely permissive for fostering an interaction between bromodomain-containing domain 4 (BRD4) and p53-R172H to regulate Csf-1 transcription. Inhibition of BRD4 not only reduces tumor invasion and lung metastasis but also reduces circulating CSF-1 levels. Overall, our results establish a novel p53-R172H–d...
Cancer-associated fibroblasts (CAF) can promote tumor growth, metastasis, and therapeutic resista... more Cancer-associated fibroblasts (CAF) can promote tumor growth, metastasis, and therapeutic resistance in esophageal squamous cell carcinoma (ESCC), but the mechanisms of action remain elusive. Our objective was to identify secreted factor(s) that mediate the communication between CAFs and ESCC tumor cells with the aim of identifying potential druggable targets. Through unbiased cytokine arrays, we have identified CC motif chemokine ligand 5 (CCL5) as a secreted factor that is increased upon co-culture of ESCC cells and CAFs, which we replicated in esophageal adenocarcinoma (EAC) with CAFs. Loss of tumor-cell-derived CCL5 reduces ESCC cell proliferation in vitro and in vivo and we propose this is mediated, in part, by a reduction in ERK1/2 signaling. Loss of tumor-derived CCL5 reduces the percentage of CAFs recruited to xenograft tumors in vivo. CCL5 is a ligand for the CC motif receptor 5 (CCR5), for which a clinically approved inhibitor exists, namely Maraviroc. Maraviroc treatment reduced tumor volume, CAF recruitment, and ERK1/2 signaling in vivo, thus, mimicking the effects observed with genetic loss of CCL5. High CCL5 or CCR5 expression is associated with worse prognosis in low-grade esophageal carcinomas. Implications: These data highlight the role of CCL5 in tumorigenesis and the therapeutic potential of targeting the CCL5–CCR5 axis in ESCC.
Mdm2 and its homologue MdmX form an E3-ligase complex that is best understood as the major regula... more Mdm2 and its homologue MdmX form an E3-ligase complex that is best understood as the major regulator of p53. Yet Mdm2 and MdmX have functions in cells that are independent of their ability to degrade p53. Amongst the functions regulated by Mdm2 is cell migration, although the molecular mechanism(s) involved have not been well characterized. We show, in a p53 null model, that either siRNA knockdown of Mdm2 or MdmX as well as pharmacological inhibition of the Mdm2/X complex E3-ligase can reduce migration of cells grown as monolayer or invasion of cells from pre-formed spheroids into collagen-based matrices. This is consistent with our observation that Mdm2 ablation or inhibition leads to decreased cell spreading and attachment of cells to the extracellular matrix. In line with these findings, we found that modulation of Mdm2, MdmX or the Mdm2/X complex impacts focal adhesion (FA) formation, a main step in cell attachment, spreading and migration. Physiologically, Mdm2 silencing leads to decreased metastatic burden in mouse models. Mechanistically, we have discovered that Mdm2 modulates the RNA levels of Sprouty4 and that Sprouty4 is needed for the effects of Mdm2 knockdown on cell migration, FA formation and metastasis. Taken together, we have discovered a pathway by which Mdm2, through the activity of the Mdm2/X complex, mitigates FA formation, migration and ultimately metastasis by regulation of Sprouty4 independently of p53. Our findings suggest that blocking Mdm2 or the Mdm2/X complex might be a potential target to prevent metastasis. Citation Format: Rafaela Muniz de Queiroz, Gizem Efe, Asja Guzman, Naoko Hashimoto, Yusuke Kawashima, Tomoaki Tanaka, Anil K. Rustgi, Carol Prives. Mdm2 regulates metastasis and associated cellular processes through modulation of Sprouty4 in a p53-independent manner [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 2 (Clinical Trials and Late-Breaking Research); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(8_Suppl):Abstract nr LB160.
SummaryAlthough the E3 ligase Mdm2 and its homologue and binding partner MdmX are the major regul... more SummaryAlthough the E3 ligase Mdm2 and its homologue and binding partner MdmX are the major regulators of the p53 tumor suppressor protein, it is now evident that Mdm2 and MdmX have multiple functions that are independent of p53. For example, Mdm2 can regulate cell migration, although mechanistic insight into this function is still lacking. Here we show in cells lacking p53 expression that knockdown of Mdm2 or MdmX, as well as pharmacological inhibition of the Mdm2/MdmX complex, not only reduces cell migration and invasion, but also impairs cell spreading and focal adhesion formation. In addition, Mdm2 knockdown decreases metastasisin vivo. Remarkably, Mdm2 modulates the expression of Sprouty4, which is required for the Mdm2 mediated effects on cell migration, focal adhesion formation and metastasis. Our findings describe a molecular mechanism by which the Mdm2-X complex, through Sprouty4, regulates cellular processes leading to decreased metastatic capability independent of p53.
Targeted CCR5 inhibition replicates CCL5 loss with reduced tumor cell proliferation in vitro and ... more Targeted CCR5 inhibition replicates CCL5 loss with reduced tumor cell proliferation in vitro and in vivo.
Mdm2 and its homologue MdmX form an E3-ligase complex that is best understood as the major regula... more Mdm2 and its homologue MdmX form an E3-ligase complex that is best understood as the major regulator of p53. Yet Mdm2 and MdmX have functions in cells that are independent of their ability to degrade p53. Amongst the functions regulated by Mdm2 is cell migration, although the molecular mechanism(s) involved have not been well characterized. We show, in a p53 null model, that either siRNA knockdown of Mdm2 or MdmX as well as pharmacological inhibition of the Mdm2/X complex E3-ligase can reduce migration of cells grown as monolayer or invasion of cells from pre-formed spheroids into collagen-based matrices. This is consistent with our observation that Mdm2 ablation or inhibition leads to decreased cell spreading and attachment of cells to the extracellular matrix. In line with these findings, we found that modulation of Mdm2, MdmX or the Mdm2/X complex impacts focal adhesion (FA) formation, a main step in cell attachment, spreading and migration. Physiologically, Mdm2 silencing leads ...
Cancer-associated fibroblasts (CAF) can promote tumor growth, metastasis, and therapeutic resista... more Cancer-associated fibroblasts (CAF) can promote tumor growth, metastasis, and therapeutic resistance in esophageal squamous cell carcinoma (ESCC), but the mechanisms of action remain elusive. Our objective was to identify secreted factor(s) that mediate the communication between CAFs and ESCC tumor cells with the aim of identifying potential druggable targets. Through unbiased cytokine arrays, we have identified CC motif chemokine ligand 5 (CCL5) as a secreted factor that is increased upon co-culture of ESCC cells and CAFs, which we replicated in esophageal adenocarcinoma (EAC) with CAFs. Loss of tumor-cell-derived CCL5 reduces ESCC cell proliferation in vitro and in vivo and we propose this is mediated, in part, by a reduction in ERK1/2 signaling. Loss of tumor-derived CCL5 reduces the percentage of CAFs recruited to xenograft tumors in vivo. CCL5 is a ligand for the CC motif receptor 5 (CCR5), for which a clinically approved inhibitor exists, namely Maraviroc. Maraviroc treatment ...
TP53 mutations are frequent in esophageal squamous cell carcinoma (ESCC) and other SCCs and are a... more TP53 mutations are frequent in esophageal squamous cell carcinoma (ESCC) and other SCCs and are associated with a proclivity for metastasis. Here, we report that colony-stimulating factor-1 (CSF-1) expression is upregulated significantly in a p53-R172H–dependent manner in metastatic lung lesions of ESCC. The p53-R172H–dependent CSF-1 signaling, through its cognate receptor CSF-1R, increases tumor cell invasion and lung metastasis, which in turn is mediated in part through Stat3 phosphorylation and epithelial-to-mesenchymal transition (EMT). In Trp53R172H tumor cells, p53 occupies the Csf-1 promoter. The Csf-1 locus is enriched with histone 3 lysine 27 acetylation (H3K27ac), which is likely permissive for fostering an interaction between bromodomain-containing domain 4 (BRD4) and p53-R172H to regulate Csf-1 transcription. Inhibition of BRD4 not only reduces tumor invasion and lung metastasis but also reduces circulating CSF-1 levels. Overall, our results establish a novel p53-R172H–d...
Cancer-associated fibroblasts (CAF) can promote tumor growth, metastasis, and therapeutic resista... more Cancer-associated fibroblasts (CAF) can promote tumor growth, metastasis, and therapeutic resistance in esophageal squamous cell carcinoma (ESCC), but the mechanisms of action remain elusive. Our objective was to identify secreted factor(s) that mediate the communication between CAFs and ESCC tumor cells with the aim of identifying potential druggable targets. Through unbiased cytokine arrays, we have identified CC motif chemokine ligand 5 (CCL5) as a secreted factor that is increased upon co-culture of ESCC cells and CAFs, which we replicated in esophageal adenocarcinoma (EAC) with CAFs. Loss of tumor-cell-derived CCL5 reduces ESCC cell proliferation in vitro and in vivo and we propose this is mediated, in part, by a reduction in ERK1/2 signaling. Loss of tumor-derived CCL5 reduces the percentage of CAFs recruited to xenograft tumors in vivo. CCL5 is a ligand for the CC motif receptor 5 (CCR5), for which a clinically approved inhibitor exists, namely Maraviroc. Maraviroc treatment reduced tumor volume, CAF recruitment, and ERK1/2 signaling in vivo, thus, mimicking the effects observed with genetic loss of CCL5. High CCL5 or CCR5 expression is associated with worse prognosis in low-grade esophageal carcinomas. Implications: These data highlight the role of CCL5 in tumorigenesis and the therapeutic potential of targeting the CCL5–CCR5 axis in ESCC.
Mdm2 and its homologue MdmX form an E3-ligase complex that is best understood as the major regula... more Mdm2 and its homologue MdmX form an E3-ligase complex that is best understood as the major regulator of p53. Yet Mdm2 and MdmX have functions in cells that are independent of their ability to degrade p53. Amongst the functions regulated by Mdm2 is cell migration, although the molecular mechanism(s) involved have not been well characterized. We show, in a p53 null model, that either siRNA knockdown of Mdm2 or MdmX as well as pharmacological inhibition of the Mdm2/X complex E3-ligase can reduce migration of cells grown as monolayer or invasion of cells from pre-formed spheroids into collagen-based matrices. This is consistent with our observation that Mdm2 ablation or inhibition leads to decreased cell spreading and attachment of cells to the extracellular matrix. In line with these findings, we found that modulation of Mdm2, MdmX or the Mdm2/X complex impacts focal adhesion (FA) formation, a main step in cell attachment, spreading and migration. Physiologically, Mdm2 silencing leads to decreased metastatic burden in mouse models. Mechanistically, we have discovered that Mdm2 modulates the RNA levels of Sprouty4 and that Sprouty4 is needed for the effects of Mdm2 knockdown on cell migration, FA formation and metastasis. Taken together, we have discovered a pathway by which Mdm2, through the activity of the Mdm2/X complex, mitigates FA formation, migration and ultimately metastasis by regulation of Sprouty4 independently of p53. Our findings suggest that blocking Mdm2 or the Mdm2/X complex might be a potential target to prevent metastasis. Citation Format: Rafaela Muniz de Queiroz, Gizem Efe, Asja Guzman, Naoko Hashimoto, Yusuke Kawashima, Tomoaki Tanaka, Anil K. Rustgi, Carol Prives. Mdm2 regulates metastasis and associated cellular processes through modulation of Sprouty4 in a p53-independent manner [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 2 (Clinical Trials and Late-Breaking Research); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(8_Suppl):Abstract nr LB160.
SummaryAlthough the E3 ligase Mdm2 and its homologue and binding partner MdmX are the major regul... more SummaryAlthough the E3 ligase Mdm2 and its homologue and binding partner MdmX are the major regulators of the p53 tumor suppressor protein, it is now evident that Mdm2 and MdmX have multiple functions that are independent of p53. For example, Mdm2 can regulate cell migration, although mechanistic insight into this function is still lacking. Here we show in cells lacking p53 expression that knockdown of Mdm2 or MdmX, as well as pharmacological inhibition of the Mdm2/MdmX complex, not only reduces cell migration and invasion, but also impairs cell spreading and focal adhesion formation. In addition, Mdm2 knockdown decreases metastasisin vivo. Remarkably, Mdm2 modulates the expression of Sprouty4, which is required for the Mdm2 mediated effects on cell migration, focal adhesion formation and metastasis. Our findings describe a molecular mechanism by which the Mdm2-X complex, through Sprouty4, regulates cellular processes leading to decreased metastatic capability independent of p53.
Targeted CCR5 inhibition replicates CCL5 loss with reduced tumor cell proliferation in vitro and ... more Targeted CCR5 inhibition replicates CCL5 loss with reduced tumor cell proliferation in vitro and in vivo.
Mdm2 and its homologue MdmX form an E3-ligase complex that is best understood as the major regula... more Mdm2 and its homologue MdmX form an E3-ligase complex that is best understood as the major regulator of p53. Yet Mdm2 and MdmX have functions in cells that are independent of their ability to degrade p53. Amongst the functions regulated by Mdm2 is cell migration, although the molecular mechanism(s) involved have not been well characterized. We show, in a p53 null model, that either siRNA knockdown of Mdm2 or MdmX as well as pharmacological inhibition of the Mdm2/X complex E3-ligase can reduce migration of cells grown as monolayer or invasion of cells from pre-formed spheroids into collagen-based matrices. This is consistent with our observation that Mdm2 ablation or inhibition leads to decreased cell spreading and attachment of cells to the extracellular matrix. In line with these findings, we found that modulation of Mdm2, MdmX or the Mdm2/X complex impacts focal adhesion (FA) formation, a main step in cell attachment, spreading and migration. Physiologically, Mdm2 silencing leads ...
Cancer-associated fibroblasts (CAF) can promote tumor growth, metastasis, and therapeutic resista... more Cancer-associated fibroblasts (CAF) can promote tumor growth, metastasis, and therapeutic resistance in esophageal squamous cell carcinoma (ESCC), but the mechanisms of action remain elusive. Our objective was to identify secreted factor(s) that mediate the communication between CAFs and ESCC tumor cells with the aim of identifying potential druggable targets. Through unbiased cytokine arrays, we have identified CC motif chemokine ligand 5 (CCL5) as a secreted factor that is increased upon co-culture of ESCC cells and CAFs, which we replicated in esophageal adenocarcinoma (EAC) with CAFs. Loss of tumor-cell-derived CCL5 reduces ESCC cell proliferation in vitro and in vivo and we propose this is mediated, in part, by a reduction in ERK1/2 signaling. Loss of tumor-derived CCL5 reduces the percentage of CAFs recruited to xenograft tumors in vivo. CCL5 is a ligand for the CC motif receptor 5 (CCR5), for which a clinically approved inhibitor exists, namely Maraviroc. Maraviroc treatment ...
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