RESUMEN
Molecular alterations that confer phenotypic advantages to tumors can also expose specific therapeutic vulnerabilities. To search for potential treatments that would selectively affect metastatic cells, we examined the sensitivity of lineage-related human bladder cancer cell lines with different lung colonization abilities to chloroquine (CQ) or bafilomycin A1, which are inhibitors of lysosome function and autophagy. Both CQ and bafilomycin A1 were more cytotoxic in vitro to highly metastatic cells compared with their less metastatic counterparts. Genetic inactivation of macroautophagy regulators and lysosomal proteins indicated that this was due to greater reliance on the lysosome but not upon macroautophagy. To identify the mechanism underlying these effects, we generated cells resistant to CQ in vitro. Surprisingly, selection for in vitro CQ resistance was sufficient to alter gene expression patterns such that unsupervised cluster analysis of whole-transcriptome data indicated that selection for CQ resistance alone created tumor cells that were more similar to the poorly metastatic parental cells from which the metastatic cells were derived; importantly, these tumor cells also had diminished metastatic ability in vivo. These effects were mediated in part by differential expression of the transcriptional regulator ID4 (inhibitor of DNA binding 4); depletion of ID4 both promoted in vitro CQ sensitivity and restored lung colonization and metastasis of CQ-resistant cells. These data demonstrate that selection for metastasis ability confers selective vulnerability to lysosomal inhibitors and identify ID4 as a potential biomarker for the use of lysosomal inhibitors to reduce metastasis in patients.
Asunto(s)
Cloroquina/farmacología , Resistencia a Antineoplásicos/efectos de los fármacos , Neoplasias Pulmonares , Lisosomas/metabolismo , Macrólidos/farmacología , Neoplasias de la Vejiga Urinaria , Animales , Línea Celular Tumoral , Regulación Neoplásica de la Expresión Génica/efectos de los fármacos , Humanos , Proteínas Inhibidoras de la Diferenciación/biosíntesis , Neoplasias Pulmonares/tratamiento farmacológico , Neoplasias Pulmonares/metabolismo , Neoplasias Pulmonares/patología , Neoplasias Pulmonares/secundario , Lisosomas/patología , Ratones , Metástasis de la Neoplasia , Proteínas de Neoplasias/biosíntesis , Neoplasias de la Vejiga Urinaria/tratamiento farmacológico , Neoplasias de la Vejiga Urinaria/genética , Neoplasias de la Vejiga Urinaria/metabolismo , Neoplasias de la Vejiga Urinaria/patologíaRESUMEN
Porcupine O-acyltransferase (PORCN) is considered essential for Wnt secretion and signaling. However, we observed that PORCN inhibition does not phenocopy the effects of WNT4 knockdown in WNT4-dependent breast cancer cells. This suggests a unique relationship between PORCN and WNT4 signaling. To examine the role of PORCN in WNT4 signaling, here we overexpressed WNT4 or WNT3A in breast cancer, ovarian cancer, and fibrosarcoma cell lines. Conditioned media from these lines and co-culture systems were used to assess the dependence of Wnt secretion and activity on the critical Wnt secretion proteins PORCN and Wnt ligand secretion (WLS) mediator. We observed that WLS is universally required for Wnt secretion and paracrine signaling. In contrast, the dependence of WNT3A secretion and activity on PORCN varied across the cell lines, and WNT4 secretion was PORCN-independent in all models. Surprisingly, WNT4 did not exhibit paracrine activity in any tested context. Absent the expected paracrine activity of secreted WNT4, we identified cell-autonomous Wnt signaling activation by WNT4 and WNT3A, independent of PORCN or Wnt secretion. The PORCN-independent, cell-autonomous Wnt signaling demonstrated here may be critical in WNT4-driven cellular contexts or in those that are considered to have dysfunctional Wnt signaling.
Asunto(s)
Aciltransferasas/metabolismo , Proteínas de la Membrana/metabolismo , Vía de Señalización Wnt , Proteína Wnt3A/metabolismo , Proteína Wnt4/metabolismo , Aciltransferasas/antagonistas & inhibidores , Aciltransferasas/genética , Línea Celular Tumoral , Proliferación Celular , Técnicas de Cocultivo , Medios de Cultivo Condicionados/química , Fulvestrant/farmacología , Humanos , Péptidos y Proteínas de Señalización Intracelular/antagonistas & inhibidores , Péptidos y Proteínas de Señalización Intracelular/genética , Péptidos y Proteínas de Señalización Intracelular/metabolismo , Proteínas de la Membrana/antagonistas & inhibidores , Proteínas de la Membrana/genética , Comunicación Paracrina , Transporte de Proteínas , Interferencia de ARN , ARN Interferente Pequeño/metabolismo , Receptores Acoplados a Proteínas G/antagonistas & inhibidores , Receptores Acoplados a Proteínas G/genética , Receptores Acoplados a Proteínas G/metabolismo , Vía de Señalización Wnt/efectos de los fármacos , Proteína Wnt3A/antagonistas & inhibidores , Proteína Wnt3A/genética , Proteína Wnt4/antagonistas & inhibidores , Proteína Wnt4/genéticaRESUMEN
Amylo-α-1,6-glucosidase,4-α-glucanotransferase (AGL) is an enzyme primarily responsible for glycogen debranching. Germline mutations lead to glycogen storage disease type III (GSDIII). We recently found AGL to be a tumor suppressor in xenograft models of human bladder cancer (BC) and low levels of AGL expression in BC are associated with poor patient prognosis. However, the impact of low AGL expression on the susceptibility of normal bladder to carcinogenesis is unknown. We address this gap by developing a germline Agl knockout (Agl-/-) mouse that recapitulates biochemical and histological features of GSDIII. Agl-/- mice exposed to N-butyl-N-(4-hydroxybutyl) nitrosamine (BBN) had a higher BC incidence compared with wild-type mice (Agl+/+). To determine if the increased BC incidence observed was due to decreased Agl expression in the urothelium specifically, we developed a urothelium-specific conditional Agl knockout (Aglcko) mouse using a Uroplakin II-Cre allele. BBN-induced carcinogenesis experiments repeated in Aglcko mice revealed that Aglcko mice had a higher BC incidence than control (Aglfl/fl) mice. RNA sequencing revealed that tumors from Agl-/- mice had 19 differentially expressed genes compared with control mice. An 'Agl Loss' gene signature was developed and found to successfully stratify normal and tumor samples in two BC patient datasets. These results support the role of AGL loss in promoting carcinogenesis and provide a rationale for evaluating Agl expression levels, or Agl Loss gene signature scores, in normal urothelium of populations at risk of BC development such as older male smokers.
Asunto(s)
Sistema de la Enzima Desramificadora del Glucógeno/fisiología , Neoplasias de la Vejiga Urinaria/etiología , Animales , Butilhidroxibutilnitrosamina , Ingeniería Genética , Sistema de la Enzima Desramificadora del Glucógeno/genética , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Análisis de Secuencia de ARNRESUMEN
Osteosarcoma (OSA) is the most common primary bone tumor in humans. However, the cell of origin of OSA is not clearly defined although there is evidence that osteoblasts may serve as OSA progenitors. The role of osteocytes, terminally differentiated osteoblasts, as OSA progenitors has yet to be described. Analysis of patient cDNA from publicly available microarray data revealed that patients with OSA have increased expression of dentin matrix phosphoprotein 1 (DMP1), a marker of osteocytes. Analysis of multiple murine, human, and canine OSA cell lines revealed DMP1 expression. To test the tumorigenic potential of osteocytes, MLO-Y4, a SV-40 immortalized murine osteocyte cell line, was injected into subcutaneous and orthotopic (intratibial) sites of mice. Tumor growth occurred in both locations. Orthotopic MLO-Y4 tumors produced mixed osteoblastic/osteolytic radiographic lesions; a hallmark of OSA. Together, these data demonstrate for the first time that osteocytes can serve as OSA progenitors.
Asunto(s)
Neoplasias Óseas/metabolismo , Proteínas de la Matriz Extracelular/biosíntesis , Osteosarcoma/metabolismo , Fosfoproteínas/biosíntesis , Animales , Neoplasias Óseas/etiología , Neoplasias Óseas/patología , Diferenciación Celular/genética , Línea Celular Tumoral , Perros , Proteínas de la Matriz Extracelular/genética , Regulación Neoplásica de la Expresión Génica , Humanos , Ratones , Osteoblastos/metabolismo , Osteoblastos/patología , Osteocitos/metabolismo , Osteocitos/patología , Osteosarcoma/etiología , Osteosarcoma/patología , Fosfoproteínas/genética , Células Madre/metabolismo , Células Madre/patologíaRESUMEN
BACKGROUND: Angiogenesis contributes to the progression of urothelial carcinoma (UC). In the current study, the authors investigated the role of maintenance sunitinib in patients with advanced UC. METHODS: Patients with locally recurrent/metastatic UC and adequate organ function who achieved stable disease or a partial or complete response after 4 to 6 chemotherapy cycles were randomized to sunitinib at a dose of 50 mg/day (28 days on and 14 days off) or placebo. The primary endpoint was the 6-month progression rate. Secondary endpoints were safety, survival, change in serum vascular endothelial growth factor (VEGF)/soluble VEGF receptor-2 (sVEGFR2), and the activity of sunitinib in patients who developed disease progression while receiving placebo. A total of 38 eligible patients per treatment arm were required to select better therapy with 90% probability (α = .05). RESULTS: A total of 54 eligible patients were randomized to either the sunitinib arm (26 patients) or the placebo arm (28 patients). The median number of cycles received was 2 cycles per treatment arm. The most common grade 3 to 4 adverse events (graded according to version 3.0 of the National Cancer Institute Common Terminology Criteria for Adverse Events) among patients receiving sunitinib were thrombocytopenia, diarrhea, mucositis, fatigue, and hypertension. There were no grade 3 or 4 adverse events noted among > 5% of patients receiving placebo. The 6-month progression rate was 72% versus 64%. The median progression-free survival (PFS) was 2.9 months (range, 0.5 months-32.5 months) versus 2.7 months (range, 0.8 months -65 months) for the sunitinib versus placebo arms, respectively. Patients receiving placebo were found to have no changes in their serum VEGF/sVEGFR2 levels over time. Patients treated with sunitinib had no significant change in their VEGF level, but the sVEGFR2 level significantly decreased after cycles 1 and 2 (P < .0001) and at the time of disease progression (P = .0002). A baseline VEGF level that was at or greater than the median was found to be correlated with a longer PFS. Sixteen patients who were receiving placebo received sunitinib at the time of disease progression, with the best responses being 1 partial response (6.3%), 6 cases of stable disease (37.5%), and 5 cases of progressive disease (31.3%); 4 patients were not evaluable for response. The median PFS was 3.7 months (range, 0.1 months-22 months). CONCLUSIONS: The current multicenter study was limited by premature closure and a small sample size. Maintenance sunitinib did not appear to improve the 6-month progression rate. Open-label sunitinib was found to have only modest activity. The sVEGFR2 level decreased among patients receiving sunitinib.
Asunto(s)
Inhibidores de la Angiogénesis/uso terapéutico , Antineoplásicos/uso terapéutico , Carcinoma de Células Transicionales/tratamiento farmacológico , Indoles/uso terapéutico , Pirroles/uso terapéutico , Neoplasias Urológicas/tratamiento farmacológico , Receptor 2 de Factores de Crecimiento Endotelial Vascular/antagonistas & inhibidores , Adulto , Anciano , Inhibidores de la Angiogénesis/administración & dosificación , Factores de Confusión Epidemiológicos , Progresión de la Enfermedad , Supervivencia sin Enfermedad , Método Doble Ciego , Esquema de Medicación , Terminación Anticipada de los Ensayos Clínicos , Femenino , Humanos , Indoles/administración & dosificación , Estimación de Kaplan-Meier , Masculino , Persona de Mediana Edad , Pirroles/administración & dosificación , Tamaño de la Muestra , Sunitinib , Resultado del Tratamiento , Factor A de Crecimiento Endotelial Vascular/sangre , Receptor 2 de Factores de Crecimiento Endotelial Vascular/sangreRESUMEN
Wnt ligand WNT4 is critical in female reproductive tissue development, with WNT4 dysregulation linked to related pathologies including breast cancer (invasive lobular carcinoma, ILC) and gynecologic cancers. WNT4 signaling in these contexts is distinct from canonical Wnt signaling yet inadequately understood. We previously identified atypical intracellular activity of WNT4 (independent of Wnt secretion) regulating mitochondrial function, and herein examine intracellular functions of WNT4. We further examine how convergent mechanisms of WNT4 dysregulation impact cancer metabolism. In ILC, WNT4 is co-opted by estrogen receptor α (ER) via genomic binding in WNT4 intron 1, while in gynecologic cancers, a common genetic polymorphism (rs3820282) at this ER binding site alters WNT4 regulation. Using proximity biotinylation (BioID), we show canonical Wnt ligand WNT3A is trafficked for secretion, but WNT4 is localized to the cytosol and mitochondria. We identified DHRS2, mTOR, and STAT1 as putative WNT4 cytosolic/mitochondrial signaling partners. Whole metabolite profiling, and integrated transcriptomic data, support that WNT4 mediates metabolic reprogramming via fatty acid and amino acid metabolism. Furthermore, ovarian cancer cell lines with rs3820282 variant genotype are WNT4 dependent and have active WNT4 metabolic signaling. In protein array analyses of a cohort of 103 human gynecologic tumors enriched for patient diversity, germline rs3820282 genotype is associated with metabolic remodeling. Variant genotype tumors show increased AMPK activation and downstream signaling, with the highest AMPK signaling activity in variant genotype tumors from non-White patients. Taken together, atypical intracellular WNT4 signaling, in part via genetic dysregulation, regulates the distinct metabolic phenotypes of ILC and gynecologic cancers. SIGNIFICANCE: WNT4 regulates breast and gynecologic cancer metabolism via a previously unappreciated intracellular signaling mechanism at the mitochondria, with WNT4 mediating metabolic remodeling. Understanding WNT4 dysregulation by estrogen and genetic polymorphism offers new opportunities for defining tumor biology, precision therapeutics, and personalized cancer risk assessment.
Asunto(s)
Neoplasias de la Mama , Neoplasias de los Genitales Femeninos , Humanos , Femenino , Ligandos , Proteínas Quinasas Activadas por AMP/metabolismo , Neoplasias de los Genitales Femeninos/genética , Transducción de Señal , Neoplasias de la Mama/genética , Proteína Wnt4/genética , Carbonil Reductasa (NADPH)/metabolismoRESUMEN
Invasive lobular carcinoma (ILC) is the most common special histologic subtype of breast cancer, and nearly all ILC tumors express estrogen receptor alpha (ER). However, clinical and laboratory data suggest ILC are strongly estrogen-driven but not equally antiestrogen-sensitive. We hypothesized ILC-specific ER coregulators mediate ER functions and antiestrogen resistance in ILC, and profiled ER-associated proteins by mass spectrometry. Three ER+ ILC cell lines (MDA MB 134VI, SUM44PE, and BCK4) were compared with ER+ invasive ductal carcinoma (IDC) line data, and we examined whether siRNA of identified proteins suppressed ER-driven proliferation in ILC cells. This identified mediator of DNA damage checkpoint 1 (MDC1), a tumor suppressor in DNA damage response (DDR), as a novel ER coregulator in ILC. We confirmed ER:MDC1 interaction was specific to ILC versus IDC cells, and found MDC1 knockdown suppressed ILC cell proliferation and tamoxifen resistance. Using RNA-sequencing, we found in ILC cells MDC1 knockdown broadly dysregulates the ER transcriptome, with ER:MDC1 target genes enriched for promoter hormone response elements. Importantly, our data are inconsistent with MDC1 tumor suppressor functions in DDR, but suggest a novel oncogenic role for MDC1 as an ER coregulator. Supporting this, in breast tumor tissue microarrays, MDC1 protein was frequently low or absent in IDC, but MDC1 loss was rare in ER+ ILC. ER:MDC1 interaction and MDC1 coregulator functions may underlie ER function in ILC and serve as targets to overcome antiestrogen resistance in ILC. IMPLICATIONS: MDC1 has novel ER coregulator activity in ILC, which may underlie ILC-specific ER functions, estrogen response, and antiestrogen resistance.
Asunto(s)
Proteínas Adaptadoras Transductoras de Señales/genética , Neoplasias de la Mama/genética , Carcinoma Lobular/genética , Proteínas de Ciclo Celular/genética , Receptores de Estrógenos/genética , Mama/patología , Neoplasias de la Mama/tratamiento farmacológico , Neoplasias de la Mama/patología , Carcinoma Lobular/tratamiento farmacológico , Carcinoma Lobular/patología , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , Proliferación Celular/genética , Femenino , Humanos , Células MCF-7 , Regiones Promotoras Genéticas/efectos de los fármacos , Regiones Promotoras Genéticas/genética , Transducción de Señal/genética , Tamoxifeno/uso terapéutico , Transcriptoma/efectos de los fármacos , Transcriptoma/genéticaRESUMEN
The androgen receptor (AR) is important in the development of both experimental and human bladder cancer. However, the role of AR in bladder cancer growth and progression is less clear, with literature indicating that more advanced stage and grade disease are associated with reduced AR expression. To determine the mechanisms underlying these relationships, we profiled AR-expressing human bladder cancer cells by AR chromatin immunoprecipitation sequencing and complementary transcriptomic approaches in response to in vitro stimulation by the synthetic androgen R1881. In vivo functional genomics consisting of pooled shRNA or pooled open reading frame libraries was employed to evaluate 97 genes that recapitulate the direction of expression associated with androgen stimulation. Interestingly, we identified CD44, the receptor for hyaluronic acid, a potent biomarker and driver of progressive disease in multiple tumor types, as significantly associated with androgen stimulation. CRISPR-based mutagenesis of androgen response elements associated with CD44 identified a novel silencer element leading to the direct transcriptional repression of CD44 expression. In human patients with bladder cancer, tumor AR and CD44 mRNA and protein expression were inversely correlated, suggesting a clinically relevant AR-CD44 axis. Collectively, our work describes a novel mechanism partly explaining the inverse relationship between AR and bladder cancer tumor progression and suggests that AR and CD44 expression may be useful for prognostication and therapeutic selection in primary bladder cancer. SIGNIFICANCE: This study describes novel AREs that suppress CD44 and an expected inverse correlation of AR-CD44 expression observed in human bladder tumors.
Asunto(s)
Biomarcadores de Tumor/metabolismo , Regulación Neoplásica de la Expresión Génica , Receptores de Hialuranos/metabolismo , Receptores Androgénicos/metabolismo , Neoplasias de la Vejiga Urinaria/patología , Animales , Apoptosis , Biomarcadores de Tumor/genética , Proliferación Celular , Perfilación de la Expresión Génica , Humanos , Receptores de Hialuranos/genética , Masculino , Ratones , Ratones Desnudos , Pronóstico , Receptores Androgénicos/genética , Células Tumorales Cultivadas , Neoplasias de la Vejiga Urinaria/genética , Neoplasias de la Vejiga Urinaria/metabolismo , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
The development of a new, less invasive, and more rapidly implemented method of quantifying endothelial cell density in tumors could facilitate experimental and clinical studies of angiogenesis. Therefore, we evaluated the utility of tumor fine needle aspiration (FNA) coupled with flow cytometry for assessment of tumor angiogenesis. Samples were obtained from cutaneous tumors of mice using FNA, then immunostained and assessed by flow cytometry to determine the number of CD31(+) endothelial cells. Results of the FNA/flow cytometry technique were compared with quantification of tumor microvessel density using immunohistochemistry. The ability of the FNA/cytometry technique to quantify the effects of anti-angiogenic therapy and to monitor changes in tumor angiogenesis over time in individual tumors was also determined. We found that endothelial cell percentages determined in tumor tissue aspirates by flow cytometry correlated well with the percentages of endothelial cells determined in whole tumor digests by flow cytometry and with tumor microvessel density measurements by immunohistochemistry. Moreover, we found that repeated FNA sampling of tumors did not induce endothelial cell changes. Interestingly, by employing repeated FNA sampling of the same tumors we were able to observe a sudden and marked decline in tumor angiogenesis triggered when tumors reached a certain size. Thus, we conclude that the FNA/flow cytometry technique is an efficient, reproducible, and relatively non-invasive method of rapidly assessing tumor angiogenesis, which could be readily applied to evaluation of tumor angiogenesis in clinical settings in humans.
Asunto(s)
Biopsia con Aguja Fina/métodos , Citometría de Flujo/métodos , Neoplasias/irrigación sanguínea , Neoplasias/patología , Neovascularización Patológica/diagnóstico , Inhibidores de la Angiogénesis/farmacología , Inhibidores de la Angiogénesis/uso terapéutico , Animales , Línea Celular Tumoral , Células Endoteliales/efectos de los fármacos , Células Endoteliales/patología , Inmunohistoquímica , Ratones , Microvasos/efectos de los fármacos , Microvasos/patología , Neovascularización Patológica/tratamiento farmacológico , Factores de TiempoRESUMEN
Clinical studies over the past several years have reported that metastasis-free survival times in humans and dogs with osteosarcoma are significantly increased in patients that develop chronic bacterial osteomyelitis at their surgical site. However, the immunological mechanism by which osteomyelitis may suppress tumor growth has not been investigated. Therefore, we used a mouse model of osteomyelitis to assess the effects of bone infection on innate immunity and tumor growth. A chronic Staphylococcal osteomyelitis model was established in C3H mice and the effects of infection on tumor growth of syngeneic DLM8 osteosarcoma were assessed. The effects of infection on tumor angiogenesis and innate immunity, including NK cell and monocyte responses, were assessed. We found that osteomyelitis significantly inhibited the growth of tumors in mice, and that the effect was independent of the infecting bacterial type, tumor type, or mouse strain. Depletion of NK cells or monocytes reversed the antitumor activity elicited by infection. Moreover, infected mice had a significant increase in circulating monocytes and numbers of tumor associated macrophages. Infection suppressed tumor angiogenesis but did not affect the numbers of circulating endothelial cells. Therefore, we concluded that chronic localized bacterial infection could elicit significant systemic antitumor activity dependent on NK cells and macrophages.
Asunto(s)
Osteomielitis/complicaciones , Osteosarcoma/complicaciones , Osteosarcoma/inmunología , Infecciones Estafilocócicas/complicaciones , Animales , Enfermedad Crónica , Modelos Animales de Enfermedad , Inmunidad Innata , Células Asesinas Naturales/inmunología , Macrófagos/inmunología , Ratones , Ratones Endogámicos C3H , Ratones Endogámicos , Monocitos/inmunología , Neoplasias , Neovascularización Patológica , Osteosarcoma/patologíaRESUMEN
Invasive lobular carcinoma of the breast (ILC) is strongly estrogen-driven and represents a unique context for estrogen receptor (ER) signaling. In ILC, ER controls the expression of the Wnt ligand WNT4, which is critical for endocrine response and anti-estrogen resistance. However, signaling mediated by WNT4 is cell type- and tissue-specific, and has not been explored in ILC. We utilized reverse phase protein array (RPPA) to characterize ER and WNT4-driven signaling in ILC cells and identified that WNT4 mediates downstream mTOR signaling via phosphorylation of S6 Kinase. Additionally, ER and WNT4 control levels of MCL-1, which is associated with regulation of mitochondrial function. In this context, WNT4 knockdown led to decreased ATP production and increased mitochondrial fragmentation. WNT4 regulation of both mTOR signaling and MCL-1 were also observed in anti-estrogen resistant models of ILC. We identified that high WNT4 expression is associated with similar mTOR pathway activation in ILC and serous ovarian cancer tumors, suggesting that WNT4 signaling is active in multiple tumor types. The identified downstream pathways offer insight into WNT4 signaling and represent potential targets to overcome anti-estrogen resistance for patients with ILC.
RESUMEN
RhoGDI2 specifically suppresses bladder cancer metastasis but not primary tumor growth, which involves tumor-associated macrophages. We report that macrophage-secreted osteopontin binds to CD44s on the tumor cells and promotes invasion and clonal growth. These effects are RhoGDI2-sensitive and require CD44s binding to the Rac GEF TIAM1. Osteopontin expression correlates with tumor aggressiveness and poor clinical outcome in patients. Inhibiting this pathway potently blocked lung and lymph node metastasis; however, primary tumors and established metastasis were less sensitive. Osteopontin-CD44s-TIAM1 promotes clonal growth in vitro but not at high cell density. These data identify osteopontin-CD44-TIAM1-Rac1 axis as a RhoGDI2-sensitive pathway and potential therapeutic target in bladder cancer metastasis. They also elucidate the mechanism behind RhoGDI2 specificity for metastasis over established tumors.
Asunto(s)
Receptores de Hialuranos/metabolismo , Neoplasias Pulmonares/secundario , Osteopontina/metabolismo , Neoplasias de la Vejiga Urinaria/patología , Inhibidor beta de Disociación del Nucleótido Guanina rho/metabolismo , Animales , Humanos , Neoplasias Pulmonares/patología , Ratones , Ratones Desnudos , Metástasis de la Neoplasia , Neoplasias de la Vejiga Urinaria/metabolismoRESUMEN
Cross-talk between tumor cells and their microenvironment is critical for malignant progression. Cross-talk mediators, including soluble factors and direct cell contact, have been identified, but roles for the interaction of physical forces between tumor cells and the bone microenvironment have not been described. Here, we report preclinical evidence that tumor-generated pressure acts to modify the bone microenvironment to promote the growth of prostate cancer bone metastases. Tumors growing in mouse tibiae increased intraosseous pressure. Application of pressure to osteocytes, the main mechanotransducing cells in bone, induced prostate cancer growth and invasion. Mechanistic investigations revealed that this process was mediated in part by upregulation of CCL5 and matrix metalloproteinases in osteocytes. Our results defined the critical contribution of physical forces to tumor cell growth in the tumor microenvironment, and they identified osteocytes as a critical mediator in the bone metastatic niche.
Asunto(s)
Neoplasias Óseas/patología , Presión , Neoplasias de la Próstata/patología , Microambiente Tumoral , Animales , Neoplasias Óseas/secundario , Proliferación Celular/fisiología , Quimiocina CCL5/biosíntesis , Regulación Neoplásica de la Expresión Génica , Humanos , Masculino , Metaloproteinasas de la Matriz/biosíntesis , Ratones , Metástasis de la Neoplasia , Osteocitos/metabolismo , Osteocitos/patologíaRESUMEN
Men who die of prostate cancer (PCa) do so because of systemic metastases, the most frequent of which are within the skeleton. Recent data suggest that the colonization of the skeleton is mediated in part by collagen type I, the most abundant protein within the bone. We have shown that enhanced collagen I binding through increased expression of integrin α2ß1 stimulated in vitro invasion and promoted the growth of PCa cells within the bone. Accordingly, we sought to determine whether α2ß1 integrin is a potential mediator of skeletal metastasis. To examine whether α2ß1 integrin mediates PCa metastasis, α2 integrin was over-expressed in low-tumorigenic LNCaP PCa cells or selectively knocked-down in highly metastatic LNCaPcol PCa cells. We document that the over-expression of α2 cDNA stimulated whereas α2 shRNA inhibited the ability of transduced cells to bind to or migrate towards collagen in vitro. Correspondingly, α2 integrin knock-down reduced the tumor burden of intra-osseous tumors compared to control-transduced cells. To investigate the clinical significance of α2ß1 expression in PCa, α2ß1 protein was measured in prostatic tissues and in soft tissue and bone metastases. The data demonstrate that α2ß1 protein was elevated in PCa skeletal metastases compared to either PCa primary lesions or soft tissue metastases suggesting that α2ß1 contributes to the selective metastasis to the bone. Taken together, these data support that α2ß1 integrin is needed for the efficient metastasis of PCa cells to the skeleton.
Asunto(s)
Neoplasias Óseas/secundario , Integrina alfa2beta1/fisiología , Neoplasias de la Próstata/patología , Secuencia de Bases , Neoplasias Óseas/metabolismo , Línea Celular Tumoral , Cartilla de ADN , Humanos , Inmunohistoquímica , Masculino , Neoplasias de la Próstata/metabolismo , Reacción en Cadena de la Polimerasa de Transcriptasa InversaRESUMEN
Bone metastasis is a clinically devastating development of progressive cancers including prostate carcinoma, breast carcinoma and multiple myeloma. Bone metastases are typically painful, lead to adverse skeletal-related events, such as fracture, and are highly resistant to therapy. A major contribution to the ability of cancers to successfully establish bone metastases is their ability to exploit mechanisms of normal bone remodeling. Wnts are a large family of morphogenic proteins that are critical for bone development and contribute to maintaining bone mass in the mature organism. Wnt function is balanced by the presence of a variety of endogenous inhibitors, such as the dickkopf family members, secreted frizzled related proteins and sclerostin. Together, these factors contribute to normal bone homeostasis, allowing for dynamic changes in bone to withstand alterations in physical forces and physiological needs. In this review, we describe the role that Wnts and their inhibitors have in normal bone biology and cancer-related bone pathology. An overview of Wnt signaling pathways is discussed and key bone microenvironment cellular players, as they pertain to Wnt biology, are examined. Finally, we describe clinical trials of several Wnt inhibitor antagonists for patients with tumor-related bone disease. As few options currently exist for the treatment of bone-metastatic disease, Wnt proteins and their inhibitors offer promise for the development of novel therapeutics.
RESUMEN
Metastasis is the primary cause of death from many tumors, and novel anti-metastatic therapies are necessary. Recently, we showed that metastatic tumors down-regulate key oxidative phosphorylation (OXPHOS) genes in favor of glycolysis, a further enhancement of the Warburg effect. Therefore, we sought to determine if restriction of glycolysis using 2-deoxy-D-glucose (2DG) would lead to increased utilization of OXPHOS and inhibition of the metastatic phenotype. Noncytotoxic concentrations of 2DG dose-dependently inhibited in vitro migration and invasion in the highly metastatic DLM8-luc-M1 osteosarcoma (OS) cell line, as well as other metastatic human, canine, and murine cancer cells of different histotypes. This was associated with cytoskeletal rearrangement and inhibition of cathepsin L expression. A dose-dependent shift toward OXPHOS was confirmed by demonstrating increased oxygen utilization and decreased lactate production in 2DG treated cells. Finally, 2DG treatment significantly delayed metastasis and prolonged survival in an orthotopic postsurgical OS model. In conclusion, this work suggests that forcing cells away from glycolysis may inhibit key components of the metastatic phenotype, providing a novel avenue for metastasis prevention.
Asunto(s)
Antimetabolitos/farmacología , Neoplasias Óseas/prevención & control , Desoxiglucosa/farmacología , Glucólisis/efectos de los fármacos , Osteosarcoma/tratamiento farmacológico , Fosforilación Oxidativa/efectos de los fármacos , Animales , Western Blotting , Neoplasias Óseas/metabolismo , Neoplasias Óseas/secundario , Catepsina L/metabolismo , Adhesión Celular/efectos de los fármacos , Movimiento Celular/efectos de los fármacos , Proliferación Celular/efectos de los fármacos , Perros , Femenino , Técnica del Anticuerpo Fluorescente , Humanos , Técnicas In Vitro , Ratones , Ratones Endogámicos C3H , Osteosarcoma/metabolismo , Osteosarcoma/patología , Consumo de Oxígeno/efectos de los fármacos , Fenotipo , Tasa de Supervivencia , Células Tumorales CultivadasRESUMEN
The tumor microenvironment (TME) is a very complex niche that consists of multiple cell types, supportive matrix and soluble factors. Cells in the TME consist of both host cells that are present at tumor site at the onset of tumor growth and cells that are recruited in either response to tumor- or host-derived factors. PCa (PCa) thrives on crosstalk between tumor cells and the TME. Crosstalk results in an orchestrated evolution of both the tumor and microenvironment as the tumor progresses. The TME reacts to PCa-produced soluble factors as well as direct interaction with PCa cells. In return, the TME produces soluble factors, structural support and direct contact interactions that influence the establishment and progression of PCa. In this review, we focus on the host side of the equation to provide a foundation for understanding how different aspects of the TME contribute to PCa progression. We discuss immune effector cells, specialized niches, such as the vascular and bone marrow, and several key protein factors that mediate host effects on PCa. This discussion highlights the concept that the TME offers a potentially very fertile target for PCa therapy.
RESUMEN
Osteosarcoma (OS) is the most common bone tumor in humans. Newer, more clinically relevant models of OS are required to investigate novel therapeutics. The ability to study spontaneous micrometastases in the absence of a primary tumor is important since this is the manner in which most patients are treated clinically. Therefore, we have developed a novel model of murine OS using the DLM8 cell line, which is syngeneic to C3H mice. We have engineered these cells to express firefly luciferase so the development of metastases can be followed serially and non-invasively. These cells form osteolytic/osteoproductive lesions and metastasize spontaneously after orthotopic implantation in the proximal tibia, and the development of soft-tissue metastasis can be followed serially by luciferase expression following amputation. We have demonstrated a significant prolongation of disease-free and overall survival in the surgical adjuvant setting following treatment with doxorubicin or carboplatin, drugs which form the mainstay of treatment for human OS. In conclusion, we have developed a novel surgical adjuvant model of metastatic OS in immunocompetent mice that closely recapitulates the clinical situation, allowing the evaluation of novel therapeutics in the context of minimal residual disease.