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Targeting cell surface molecules using radioligand and antibody-based therapies has yielded considerable success across cancers. However, it remains unclear how the expression of putative lineage markers, particularly cell surface molecules, varies in the process of lineage plasticity, wherein tumor cells alter their identity and acquire new oncogenic properties. A notable example of lineage plasticity is the transformation of prostate adenocarcinoma (PRAD) to neuroendocrine prostate cancer (NEPC)-a growing resistance mechanism that results in the loss of responsiveness to androgen blockade and portends dismal patient survival. To understand how lineage markers vary across the evolution of lineage plasticity in prostate cancer, we applied single-cell analyses to 21 human prostate tumor biopsies and two genetically engineered mouse models, together with tissue microarray analysis on 131 tumor samples. Not only did we observe a higher degree of phenotypic heterogeneity in castrate-resistant PRAD and NEPC than previously anticipated but also found that the expression of molecules targeted therapeutically, namely PSMA, STEAP1, STEAP2, TROP2, CEACAM5, and DLL3, varied within a subset of gene-regulatory networks (GRNs). We also noted that NEPC and small cell lung cancer subtypes shared a set of GRNs, indicative of conserved biologic pathways that may be exploited therapeutically across tumor types. While this extreme level of transcriptional heterogeneity, particularly in cell surface marker expression, may mitigate the durability of clinical responses to current and future antigen-directed therapies, its delineation may yield signatures for patient selection in clinical trials, potentially across distinct cancer types.
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Análise de Célula Única , Masculino , Humanos , Análise de Célula Única/métodos , Animais , Camundongos , Neoplasias da Próstata/genética , Neoplasias da Próstata/metabolismo , Neoplasias da Próstata/patologia , Neoplasias da Próstata/tratamento farmacológico , Antígenos de Superfície/metabolismo , Antígenos de Superfície/genética , Antígenos de Neoplasias/metabolismo , Antígenos de Neoplasias/genética , Antígenos de Neoplasias/imunologia , Biomarcadores Tumorais/metabolismo , Biomarcadores Tumorais/genética , Adenocarcinoma/genética , Adenocarcinoma/patologia , Adenocarcinoma/metabolismo , Adenocarcinoma/tratamento farmacológico , Carcinoma Neuroendócrino/genética , Carcinoma Neuroendócrino/patologia , Carcinoma Neuroendócrino/metabolismo , Carcinoma Neuroendócrino/tratamento farmacológico , Regulação Neoplásica da Expressão Gênica , Neoplasias de Próstata Resistentes à Castração/metabolismo , Neoplasias de Próstata Resistentes à Castração/patologia , Neoplasias de Próstata Resistentes à Castração/genética , Neoplasias de Próstata Resistentes à Castração/tratamento farmacológicoRESUMO
Specific chromatin marks keep master regulators of differentiation silent yet poised for activation by extracellular signals. We report that nodal TGF-ß signals use the poised histone mark H3K9me3 to trigger differentiation of mammalian embryonic stem cells. Nodal receptors induce the formation of companion Smad4-Smad2/3 and TRIM33-Smad2/3 complexes. The PHD-Bromo cassette of TRIM33 facilitates binding of TRIM33-Smad2/3 to H3K9me3 and H3K18ac on the promoters of mesendoderm regulators Gsc and Mixl1. The crystal structure of this cassette, bound to histone H3 peptides, illustrates that PHD recognizes K9me3, and Bromo binds an adjacent K18ac. The interaction between TRIM33-Smad2/3 and H3K9me3 displaces the chromatin-compacting factor HP1γ, making nodal response elements accessible to Smad4-Smad2/3 for Pol II recruitment. In turn, Smad4 increases K18 acetylation to augment TRIM33-Smad2/3 binding. Thus, nodal effectors use the H3K9me3 mark as a platform to switch master regulators of stem cell differentiation from the poised to the active state.
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Montagem e Desmontagem da Cromatina , Células-Tronco Embrionárias/metabolismo , Histonas/metabolismo , Proteínas Smad/metabolismo , Fatores de Transcrição/metabolismo , Sequência de Aminoácidos , Animais , Cristalografia por Raios X , Proteína Goosecoid/genética , Proteínas de Homeodomínio/genética , Humanos , Camundongos , Modelos Moleculares , Dados de Sequência Molecular , Regiões Promotoras Genéticas , Alinhamento de SequênciaRESUMO
TGF-beta and BMP receptor kinases activate Smad transcription factors by C-terminal phosphorylation. We have identified a subsequent agonist-induced phosphorylation that plays a central dual role in Smad transcriptional activation and turnover. As receptor-activated Smads form transcriptional complexes, they are phosphorylated at an interdomain linker region by CDK8 and CDK9, which are components of transcriptional mediator and elongation complexes. These phosphorylations promote Smad transcriptional action, which in the case of Smad1 is mediated by the recruitment of YAP to the phosphorylated linker sites. An effector of the highly conserved Hippo organ size control pathway, YAP supports Smad1-dependent transcription and is required for BMP suppression of neural differentiation of mouse embryonic stem cells. The phosphorylated linker is ultimately recognized by specific ubiquitin ligases, leading to proteasome-mediated turnover of activated Smad proteins. Thus, nuclear CDK8/9 drive a cycle of Smad utilization and disposal that is an integral part of canonical BMP and TGF-beta pathways.
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Proteínas Morfogenéticas Ósseas/metabolismo , Quinase 8 Dependente de Ciclina/metabolismo , Quinase 9 Dependente de Ciclina/metabolismo , Proteínas Smad/genética , Ativação Transcricional , Fator de Crescimento Transformador beta/metabolismo , Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Animais , Proteínas de Ciclo Celular , Linhagem Celular , Inibição de Contato , Embrião de Mamíferos/citologia , Humanos , Camundongos , Tamanho do Órgão , Fosfoproteínas/metabolismo , Fosforilação , Estrutura Terciária de Proteína , Transdução de Sinais , Proteínas Smad/química , Proteína Smad1/genética , Proteínas de Sinalização YAPRESUMO
The DNA damage response (DDR) is activated by oncogenic stress, but the mechanisms by which this occurs, and the particular DDR functions that constitute barriers to tumorigenesis, remain unclear. We established a mouse model of sporadic oncogene-driven breast tumorigenesis in a series of mutant mouse strains with specific DDR deficiencies to reveal a role for the Mre11 complex in the response to oncogene activation. We demonstrate that an Mre11-mediated DDR restrains mammary hyperplasia by effecting an oncogene-induced G2 arrest. Impairment of Mre11 complex functions promotes the progression of mammary hyperplasias into invasive and metastatic breast cancers, which are often associated with secondary inactivation of the Ink4a-Arf (CDKN2a) locus. These findings provide insight into the mechanism of DDR engagement by activated oncogenes and highlight genetic interactions between the DDR and Ink4a-Arf pathways in suppression of oncogene-driven tumorigenesis and metastasis.
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Neoplasias da Mama/genética , Carcinogênese , Proteínas de Ligação a DNA/metabolismo , Oncogenes , Animais , Neoplasias da Mama/metabolismo , Neoplasias da Mama/patologia , Inibidor p16 de Quinase Dependente de Ciclina/genética , Inibidor p16 de Quinase Dependente de Ciclina/metabolismo , Dano ao DNA/genética , Proteínas de Ligação a DNA/genética , Feminino , Regulação Neoplásica da Expressão Gênica , Humanos , Hiperplasia/genética , Proteína Homóloga a MRE11 , Glândulas Mamárias Animais/crescimento & desenvolvimento , Glândulas Mamárias Animais/metabolismo , Glândulas Mamárias Animais/patologia , Camundongos , Metástase Neoplásica/genéticaRESUMO
Purpose To establish the prognostic value of biopsy of the central and marginal ablation zones for time to local tumor progression (LTP) after radiofrequency (RF) ablation of colorectal cancer liver metastasis (CLM). Materials and Methods A total of 47 patients with 67 CLMs were enrolled in this prospective institutional review board-approved and HIPAA-compliant study between November 2009 and August 2012. Mean tumor size was 2.1 cm (range, 0.6-4.3 cm). Biopsy of the center and margin of the ablation zone was performed immediately after RF ablation (mean number of biopsy samples per ablation zone, 1.9) and was evaluated for the presence of viable tumor cells. Samples containing tumor cells at morphologic evaluation were further interrogated with immunohistochemistry and were classified as either positive, viable tumor (V) or negative, necrotic (N). Minimal ablation margin size was evaluated in the first postablation CT study performed 4-8 weeks after ablation. Variables were evaluated as predictors of time to LTP with the competing-risks model (uni- and multivariate analyses). Results Technical effectiveness was evident in 66 of 67 (98%) ablated lesions on the first contrast material-enhanced CT images at 4-8-week follow-up. The cumulative incidence of LTP at 12-month follow-up was 22% (95% confidence interval [CI]: 12, 32). Samples from 16 (24%) of 67 ablation zones were classified as viable tumor. At univariate analysis, tumor size, minimal margin size, and biopsy results were significant in predicting LTP. When these variables were subsequently entered in a multivariate model, margin size of less than 5 mm (P < .001; hazard ratio [HR], 6.7) and positive biopsy results (P = .008; HR, 3.4) were significant. LTP within 12 months after RF ablation was noted in 3% (95% CI: 0, 9) of necrotic CLMs with margins of at least 5 mm. Conclusion Biopsy proof of complete tumor ablation and minimal ablation margins of at least 5 mm are independent predictors of LTP and yield the best oncologic outcomes. (©) RSNA, 2016.
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Ablação por Cateter/métodos , Neoplasias Colorretais/patologia , Neoplasias Hepáticas/diagnóstico por imagem , Neoplasias Hepáticas/secundário , Neoplasias Hepáticas/cirurgia , Tomografia Computadorizada por Raios X , Adulto , Idoso , Idoso de 80 Anos ou mais , Biópsia , Meios de Contraste , Progressão da Doença , Feminino , Humanos , Imuno-Histoquímica , Masculino , Pessoa de Meia-Idade , Prognóstico , Estudos Prospectivos , Ondas de Rádio , Resultado do Tratamento , Carga TumoralRESUMO
Tunneling nanotubes (TnTs) are long, non-adherent, actin-based cellular extensions that act as conduits for transport of cellular cargo between connected cells. The mechanisms of nanotube formation and the effects of the tumor microenvironment and cellular signals on TnT formation are unknown. In the present study, we explored exosomes as potential mediators of TnT formation in mesothelioma and the potential relationship of lipid rafts to TnT formation. Mesothelioma cells co-cultured with exogenous mesothelioma-derived exosomes formed more TnTs than cells cultured without exosomes within 24-48 h; and this effect was most prominent in media conditions (low-serum, hyperglycemic medium) that support TnT formation (1.3-1.9-fold difference). Fluorescence and electron microscopy confirmed the purity of isolated exosomes and revealed that they localized predominantly at the base of and within TnTs, in addition to the extracellular environment. Time-lapse microscopic imaging demonstrated uptake of tumor exosomes by TnTs, which facilitated intercellular transfer of these exosomes between connected cells. Mesothelioma cells connected via TnTs were also significantly enriched for lipid rafts at nearly a 2-fold higher number compared with cells not connected by TnTs. Our findings provide supportive evidence of exosomes as potential chemotactic stimuli for TnT formation, and also lipid raft formation as a potential biomarker for TnT-forming cells.
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Transporte Biológico/fisiologia , Comunicação Celular/fisiologia , Exossomos/metabolismo , Microdomínios da Membrana/metabolismo , Mesotelioma/metabolismo , Técnicas de Cultura de Células , Linhagem Celular Tumoral , Humanos , Nanotubos , Transdução de Sinais , Microambiente TumoralRESUMO
BRAFV600E mutation occurs in 46% of melanomas and drives high levels of ERK activity and ERK-dependent proliferation. However, BRAFV600E is insufficient to drive melanoma in GEMM models, and 82% of human benign nevi harbor BRAFV600E mutations. We show here that BRAFV600E inhibits mesenchymal migration by causing feedback inhibition of RAC1 activity. ERK pathway inhibition induces RAC1 activation and restores migration and invasion. In cells with BRAFV600E, mutant RAC1, overexpression of PREX1, PREX2, or PTEN inactivation restore RAC1 activity and cell motility. Together, these lesions occur in 48% of BRAFV600E melanomas. Thus, although BRAFV600E activation of ERK deregulates cell proliferation, it prevents full malignant transformation by causing feedback inhibition of cell migration. Secondary mutations are, therefore, required for tumorigenesis. One mechanism underlying tumor evolution may be the selection of lesions that rescue the deleterious effects of oncogenic drivers.
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PURPOSE: We conducted research on CDK4/6 inhibitors (CDK4/6i) simultaneously in the preclinical and clinical spaces to gain a deeper understanding of how senescence influences tumor growth in humans. PATIENTS AND METHODS: We coordinated a first-in-kind phase II clinical trial of the CDK4/6i abemaciclib for patients with progressive dedifferentiated liposarcoma (DDLS) with cellular studies interrogating the molecular basis of geroconversion. RESULTS: Thirty patients with progressing DDLS enrolled and were treated with 200 mg of abemaciclib twice daily. The median progression-free survival was 33 weeks at the time of the data lock, with 23 of 30 progression-free at 12 weeks (76.7%, two-sided 95% CI, 57.7%-90.1%). No new safety signals were identified. Concurrent preclinical work in liposarcoma cell lines identified ANGPTL4 as a necessary late regulator of geroconversion, the pathway from reversible cell-cycle exit to a stably arrested inflammation-provoking senescent cell. Using this insight, we were able to identify patients in which abemaciclib induced tumor cell senescence. Senescence correlated with increased leukocyte infiltration, primarily CD4-positive cells, within a month of therapy. However, those individuals with both senescence and increased TILs were also more likely to acquire resistance later in therapy. These suggest that combining senolytics with abemaciclib in a subset of patients may improve the duration of response. CONCLUSIONS: Abemaciclib was well tolerated and showed promising activity in DDLS. The discovery of ANGPTL4 as a late regulator of geroconversion helped to define how CDK4/6i-induced cellular senescence modulates the immune tumor microenvironment and contributes to both positive and negative clinical outcomes. See related commentary by Weiss et al., p. 649.
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Aminopiridinas , Lipossarcoma , Humanos , Aminopiridinas/farmacologia , Aminopiridinas/uso terapêutico , Benzimidazóis/farmacologia , Benzimidazóis/uso terapêutico , Lipossarcoma/tratamento farmacológico , Lipossarcoma/patologia , Senescência Celular , Quinase 4 Dependente de Ciclina , Microambiente TumoralRESUMO
Targeting cell surface molecules using radioligand and antibody-based therapies has yielded considerable success across cancers. However, it remains unclear how the expression of putative lineage markers, particularly cell surface molecules, varies in the process of lineage plasticity, wherein tumor cells alter their identity and acquire new oncogenic properties. A notable example of lineage plasticity is the transformation of prostate adenocarcinoma (PRAD) to neuroendocrine prostate cancer (NEPC)--a growing resistance mechanism that results in the loss of responsiveness to androgen blockade and portends dismal patient survival. To understand how lineage markers vary across the evolution of lineage plasticity in prostate cancer, we applied single cell analyses to 21 human prostate tumor biopsies and two genetically engineered mouse models, together with tissue microarray analysis (TMA) on 131 tumor samples. Not only did we observe a higher degree of phenotypic heterogeneity in castrate-resistant PRAD and NEPC than previously anticipated, but also found that the expression of molecules targeted therapeutically, namely PSMA, STEAP1, STEAP2, TROP2, CEACAM5, and DLL3, varied within a subset of gene-regulatory networks (GRNs). We also noted that NEPC and small cell lung cancer (SCLC) subtypes shared a set of GRNs, indicative of conserved biologic pathways that may be exploited therapeutically across tumor types. While this extreme level of transcriptional heterogeneity, particularly in cell surface marker expression, may mitigate the durability of clinical responses to novel antigen-directed therapies, its delineation may yield signatures for patient selection in clinical trials, potentially across distinct cancer types.
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Technologies for staining and imaging multiple antigens in single tissue sections are developing rapidly due to their potential to uncover spatial relationships between proteins with cellular resolution. Detections are performed simultaneously or sequentially depending on the approach. However, several technologies can detect limited numbers of antigens or require expensive equipment and reagents. Another serious concern is the lack of flexibility. Most commercialized reagents are validated for defined antibody panels, and introducing any changes is laborious and costly. In this chapter, we describe a method where we combine, for the first time, multiplexed IF followed by sequential immunohistochemistry (IHC) with AEC chromogen on Leica Bond staining processors with paraffin tissue sections. We present data for successful detection of 10 antigens in a single tissue section with preserved tissue integrity. Our method is designed for use with any combination of antibodies of interest, with images collected using whole slide scanners. We include an image viewing and image analysis workflow using nonlinear warping to combine all staining passes in a single full-resolution image of the entire tissue section, aligned at the single cell level.
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Biomarcadores Tumorais , Proteínas , Imuno-Histoquímica , Biomarcadores Tumorais/metabolismo , Imunofluorescência , Coloração e Rotulagem , Antígenos/análiseRESUMO
BACKGROUND: Thermal ablation is a definitive local treatment for selected colorectal liver metastases (CLM) that can be ablated with adequate margins. A critical limitation has been local tumor progression (LTP). METHODS: This prospective, single-group, phase 2 study enrolled patients with CLM < 5 cm in maximum diameter, at a tertiary cancer center between November 2009 and February 2019. Biopsy of the ablation zone center and margin was performed immediately after ablation. Viable tumor in tissue biopsy and ablation margins < 5 mm were assessed as predictors of 12-month LTP. RESULTS: We enrolled 107 patients with 182 CLMs. Mean tumor size was 2.0 (range, 0.6-4.6) cm. Microwave ablation was used in 51% and radiofrequency ablation in 49% of tumors. The 12- and 24-month cumulative incidence of LTP was 22% (95% confidence interval [CI]: 17, 29) and 29% (95% CI: 23, 36), respectively. LTP at 12 months was 7% (95% CI: 3, 14) for the biopsy tumor-negative ablation zone with margins ≥ 5 mm vs. 63% (95% CI: 35, 85) for the biopsy-positive ablation zone with margins < 5 mm (p < 0.001). CONCLUSIONS: Biopsy-proven complete tumor ablation with margins of at least 5 mm achieves optimal local tumor control for CLM, regardless of the ablation modality used.
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Drug resistance in cancer is often linked to changes in tumor cell state or lineage, but the molecular mechanisms driving this plasticity remain unclear. Using murine organoid and genetically engineered mouse models, we investigated the causes of lineage plasticity in prostate cancer and its relationship to antiandrogen resistance. We found that plasticity initiates in an epithelial population defined by mixed luminal-basal phenotype and that it depends on increased Janus kinase (JAK) and fibroblast growth factor receptor (FGFR) activity. Organoid cultures from patients with castration-resistant disease harboring mixed-lineage cells reproduce the dependency observed in mice by up-regulating luminal gene expression upon JAK and FGFR inhibitor treatment. Single-cell analysis confirms the presence of mixed-lineage cells with increased JAK/STAT (signal transducer and activator of transcription) and FGFR signaling in a subset of patients with metastatic disease, with implications for stratifying patients for clinical trials.
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Plasticidade Celular , Resistencia a Medicamentos Antineoplásicos , Receptores ErbB , Janus Quinases , Neoplasias da Próstata , Fatores de Transcrição STAT , Antagonistas de Androgênios , Animais , Humanos , Inibidores de Janus Quinases/uso terapêutico , Janus Quinases/genética , Janus Quinases/metabolismo , Masculino , Camundongos , Neoplasias Experimentais , Organoides , Neoplasias da Próstata/tratamento farmacológico , Neoplasias da Próstata/patologia , Fatores de Transcrição STAT/genética , Fatores de Transcrição STAT/metabolismo , Transdução de SinaisRESUMO
Colorectal cancer (CRC) is a leading cause of cancer mortality. Creatine metabolism was previously shown to critically regulate colon cancer progression. We report that RGX-202, an oral small-molecule SLC6A8 transporter inhibitor, robustly inhibits creatine import in vitro and in vivo, reduces intracellular phosphocreatine and ATP levels, and induces tumor apoptosis. RGX-202 suppressed CRC growth across KRAS wild-type and KRAS mutant xenograft, syngeneic, and patient-derived xenograft (PDX) tumors. Antitumor efficacy correlated with tumoral expression of creatine kinase B. Combining RGX-202 with 5-fluorouracil or the DHODH inhibitor leflunomide caused regressions of multiple colorectal xenograft and PDX tumors of distinct mutational backgrounds. RGX-202 also perturbed creatine metabolism in patients with metastatic CRC in a phase 1 trial, mirroring pharmacodynamic effects on creatine metabolism observed in mice. This is, to our knowledge, the first demonstration of preclinical and human pharmacodynamic activity for creatine metabolism targeting in oncology, thus revealing a critical therapeutic target.
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Antineoplásicos , Neoplasias do Colo , Neoplasias Colorretais , Animais , Antineoplásicos/farmacologia , Linhagem Celular Tumoral , Proliferação de Células , Neoplasias do Colo/tratamento farmacológico , Neoplasias do Colo/genética , Neoplasias Colorretais/patologia , Creatina/metabolismo , Creatina/farmacologia , Creatina/uso terapêutico , Humanos , Proteínas de Membrana Transportadoras , Camundongos , Camundongos Nus , Mutação , Proteínas do Tecido Nervoso/metabolismo , Proteínas da Membrana Plasmática de Transporte de Neurotransmissores/genética , Proteínas da Membrana Plasmática de Transporte de Neurotransmissores/farmacologia , Proteínas Proto-Oncogênicas p21(ras)/metabolismoRESUMO
gC1qR is highly expressed in breast cancer and plays a role in cancer cell proliferation. This study explored therapy with gC1qR monoclonal antibody 60.11, directed against the C1q binding domain of gC1qR, in a murine orthotopic xenotransplant model of triple negative breast cancer. MDA231 breast cancer cells were injected into the mammary fat pad of athymic nu/nu female mice. Mice were segregated into three groups (n = 5, each) and treated with the vehicle (group 1) or gC1qR antibody 60.11 (100 mg/kg) twice weekly, starting at day 3 post-implantation (group 2) or when the tumor volume reached 100 mm3 (group 3). At study termination (d = 35), the average tumor volume in the control group measured 895 ± 143 mm3, compared to 401 ± 48 mm3 and 701 ± 100 mm3 in groups 2 and 3, respectively (p < 0.05). Immunohistochemical staining of excised tumors revealed increased apoptosis (caspase 3 and TUNEL staining) in 60.11-treated mice compared to controls, and decreased angiogenesis (CD31 staining). Slightly decreased white blood cell counts were noted in 60.11-treated mice. Otherwise, no overt toxicities were observed. These data are the first to demonstrate an in vivo anti-tumor effect of 60.11 therapy in a mouse model of triple negative breast cancer.
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INTRODUCTION: Spread through air spaces (STAS) is a method of invasion in lung adenocarcinoma and is associated with tumor recurrence and poor survival. The spatial orientation of STAS cells in the lung alveolar parenchyma is not known. The aim of this study was to use high-resolution and high-quality three-dimensional (3D) reconstruction of images from immunohistochemical (IHC) and multiplex immunofluorescence (IF) experiments to understand the spatial architecture of tumor cell clusters by STAS in the lung parenchyma. METHODS: Four lung adenocarcinomas, three micropapillary-predominant and one solid predominant adenocarcinoma subtypes, were investigated. A 3D reconstruction image was created from formalin-fixed, paraffin-embedded blocks. A total of 350 serial sections were obtained and subjected to hematoxylin and eosin (100 slides), IHC (200 slides), and multiplex IF staining (50 slides) with the following antibodies: cluster of differentiation 31, collagen type IV, thyroid transcription factor-1, and E-cadherin. Whole slide images were reconstructed into 3D images for evaluation. RESULTS: Serial 3D image analysis by hematoxylin and eosin, IHC, and IF staining revealed that the micropapillary clusters and solid nests of STAS are focally attached to the alveolar walls, away from the main tumor. CONCLUSIONS: Our 3D reconstructions found that STAS tumor cells can attach to the alveolar walls rather than appearing free floating, as seen on the two-dimensional sections. This suggests that the tumor cells detach from the main tumor, migrate through air spaces, and reattach to the alveolar walls through vessel co-option, allowing them to survive and grow. This may explain the higher recurrence rate and worse survival of patients with STAS-positive tumors who undergo limited resection than those who undergo lobectomy.
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Neoplasias Pulmonares , Adenocarcinoma de Pulmão/patologia , Imunofluorescência , Humanos , Neoplasias Pulmonares/patologia , Invasividade Neoplásica/patologia , Recidiva Local de Neoplasia/patologia , Estadiamento de Neoplasias , PrognósticoRESUMO
Mesothelioma is an aggressive cancer of the serous membranes with poor prognosis despite combination therapy consisting of surgery, radiotherapy, and platinum-based chemotherapy. Targeted therapies, including immunotherapies, have reported limited success, suggesting the need for additional therapeutic targets. This study investigates a potential new therapeutic target, gC1qR/HABP1/p32 (gC1qR), which is overexpressed in all morphologic subtypes of mesothelioma. gC1qR is a complement receptor that is associated with several cellular functions, including cell proliferation and angiogenesis. In vitro and in vivo experiments were conducted to test the hypothesis that targeting gC1qR with a specific gC1qR monoclonal antibody 60.11 reduces mesothelioma tumor growth, using the biphasic mesothelioma cell line MSTO-211H (MSTO). In vitro studies demonstrate cell surface and extracellular gC1qR expression by MSTO cells, and a modest 25.3 ± 1.8% (n = 4) reduction in cell proliferation by the gC1qR blocking 60.11 antibody. This inhibition was specific for targeting the C1q binding domain of gC1qR at aa 76-93, as a separate monoclonal antibody 74.5.2, directed against amino acids 204-218, had no discernable effect. In vivo studies, using a murine orthotopic xenotransplant model, demonstrated an even greater reduction in MSTO tumor growth (50% inhibition) in mice treated with the 60.11 antibody compared to controls. Immunohistochemical studies of resected tumors revealed increased cellular apoptosis by caspase 3 and TUNEL staining, in 60.11 treated tumors compared to controls, as well as impaired angiogenesis by decreased CD31 staining. Taken together, these data identify gC1qR as a potential new therapeutic target against mesothelioma with both antiproliferative and antiangiogenic properties.
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Optimal functioning of neuronal networks is critical to the complex cognitive processes of memory and executive function that deteriorate in Alzheimer's disease (AD). Here we use cellular and animal models as well as human biospecimens to show that AD-related stressors mediate global disturbances in dynamic intra- and inter-neuronal networks through pathologic rewiring of the chaperome system into epichaperomes. These structures provide the backbone upon which proteome-wide connectivity, and in turn, protein networks become disturbed and ultimately dysfunctional. We introduce the term protein connectivity-based dysfunction (PCBD) to define this mechanism. Among most sensitive to PCBD are pathways with key roles in synaptic plasticity. We show at cellular and target organ levels that network connectivity and functional imbalances revert to normal levels upon epichaperome inhibition. In conclusion, we provide proof-of-principle to propose AD is a PCBDopathy, a disease of proteome-wide connectivity defects mediated by maladaptive epichaperomes.
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Doença de Alzheimer/metabolismo , Hipocampo/metabolismo , Plasticidade Neuronal/fisiologia , Proteoma/metabolismo , Doença de Alzheimer/patologia , Animais , Encéfalo/patologia , Mapeamento Encefálico , Disfunção Cognitiva/metabolismo , Função Executiva/fisiologia , Feminino , Hipocampo/patologia , Humanos , Masculino , Memória/fisiologia , Camundongos , Vias NeuraisRESUMO
PURPOSE: SMAD4 has shown promise in identifying patients with colorectal cancer at high risk of recurrence or death.Experimental Design: A discovery cohort and independent validation cohort were classified by SMAD4 status. SMAD4 status and immune infiltrate measurements were tested for association with recurrence-free survival (RFS). Patient-derived xenografts from SMAD4-deficient and SMAD4-retained tumors were used to examine chemoresistance. RESULTS: The discovery cohort consisted of 364 patients with stage I-IV colorectal cancer. Median age at diagnosis was 53 years. The cohort consisted of 61% left-sided tumors and 62% stage II/III patients. Median follow-up was 5.4 years (interquartile range, 2.3-8.2). SMAD4 loss, noted in 13% of tumors, was associated with higher tumor and nodal stage, adjuvant therapy use, fewer tumor-infiltrating lymphocytes (TIL), and lower peritumoral lymphocyte aggregate (PLA) scores (all P < 0.04). SMAD4 loss was associated with worse RFS (P = 0.02). When stratified by SMAD4 and immune infiltrate status, patients with SMAD4 loss and low TIL or PLA had worse RFS (P = 0.002 and P = 0.006, respectively). Among patients receiving 5-fluorouracil (5-FU)-based systemic chemotherapy, those with SMAD4 loss had a median RFS of 3.8 years compared with 13 years for patients with SMAD4 retained. In xenografted mice, the SMAD4-lost tumors displayed resistance to 5-FU. An independent cohort replicated our findings, in particular, the association of SMAD4 loss with decreased immune infiltrate, as well as worse disease-specific survival. CONCLUSIONS: Our data show SMAD4 loss correlates with worse clinical outcome, resistance to chemotherapy, and decreased immune infiltrate, supporting its use as a prognostic marker in patients with colorectal cancer.
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Protocolos de Quimioterapia Combinada Antineoplásica/farmacologia , Biomarcadores Tumorais/deficiência , Neoplasias Colorretais/patologia , Recidiva Local de Neoplasia/diagnóstico , Proteína Smad4/deficiência , Adulto , Idoso , Animais , Protocolos de Quimioterapia Combinada Antineoplásica/uso terapêutico , Biomarcadores Tumorais/imunologia , Quimioterapia Adjuvante/métodos , Colo/patologia , Colo/cirurgia , Neoplasias Colorretais/imunologia , Neoplasias Colorretais/mortalidade , Neoplasias Colorretais/terapia , Intervalo Livre de Doença , Resistencia a Medicamentos Antineoplásicos/imunologia , Feminino , Fluoruracila/farmacologia , Fluoruracila/uso terapêutico , Seguimentos , Humanos , Linfócitos do Interstício Tumoral/imunologia , Masculino , Camundongos , Pessoa de Meia-Idade , Recidiva Local de Neoplasia/imunologia , Recidiva Local de Neoplasia/patologia , Recidiva Local de Neoplasia/prevenção & controle , Prognóstico , Estudos Prospectivos , Reto/patologia , Reto/cirurgia , Proteína Smad4/imunologia , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
Rectal cancer (RC) is a challenging disease to treat that requires chemotherapy, radiation and surgery to optimize outcomes for individual patients. No accurate model of RC exists to answer fundamental research questions relevant to patients. We established a biorepository of 65 patient-derived RC organoid cultures (tumoroids) from patients with primary, metastatic or recurrent disease. RC tumoroids retained molecular features of the tumors from which they were derived, and their ex vivo responses to clinically relevant chemotherapy and radiation treatment correlated with the clinical responses noted in individual patients' tumors. Upon engraftment into murine rectal mucosa, human RC tumoroids gave rise to invasive RC followed by metastasis to lung and liver. Importantly, engrafted tumors displayed the heterogenous sensitivity to chemotherapy observed clinically. Thus, the biology and drug sensitivity of RC clinical isolates can be efficiently interrogated using an organoid-based, ex vivo platform coupled with in vivo endoluminal propagation in animals.
Assuntos
Quimiorradioterapia , Organoides/patologia , Neoplasias Retais/tratamento farmacológico , Neoplasias Retais/radioterapia , Animais , Fluoruracila/farmacologia , Humanos , Neoplasias Hepáticas/tratamento farmacológico , Neoplasias Hepáticas/patologia , Neoplasias Hepáticas/radioterapia , Neoplasias Hepáticas/secundário , Neoplasias Pulmonares/tratamento farmacológico , Neoplasias Pulmonares/patologia , Neoplasias Pulmonares/radioterapia , Neoplasias Pulmonares/secundário , Camundongos , Metástase Neoplásica , Organoides/efeitos dos fármacos , Organoides/efeitos da radiação , Neoplasias Retais/patologiaRESUMO
CONTEXT: Colonic complications are rare but lethal events in acute pancreatitis. CASE REPORT: We report the case of a 42-year-old man who suffered from a pancreatico-colonic fistula following a necrosectomy for severe pancreatitis; the fistula was demonstrated by spiral computed tomography using rectal water soluble contrast media. CONCLUSION: Computed tomography with rectal contrast detects pancreatico-colonic fistulas.