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Colorectal tumorigenesis involves the development of aberrant crypt foci (ACF) or preneoplastic lesions, representing the earliest morphological lesion visible in colon cancer. The purpose of this study was to determine changes in protein expression in carcinogen-induced ACF as they mature and transform into adenomas. Protein expression profiles of azoxymethane (AOM)-induced F344 rat colon ACF and adenomas were compared at four time points, 4 (control), 8, 16, and 24 weeks post AOM administration (n = 9/group), with time points correlating with induction and transformation events. At each time point, micro-dissected ACF and/or adenoma tissues were analyzed across multiple quantitative two-dimensional (2D-DIGE) gels using a Cy-dye labeling technique and a pooled internal standard to quantify expression changes with statistical confidence. Western blot and subsequent network pathway mapping were used to confirm and elucidate differentially expressed (p ≤ 0.05) proteins, including changes in vinculin (Vcl; p = 0.007), scinderin (Scin; p = 0.02), and profilin (Pfn1; p = 0.01), By determining protein expression changes in ACF as they mature and transform into adenomas, a "baseline" of altered regulatory proteins associated with adenocarcinoma development in this model has been elucidated. These data will enable future studies aimed at biomarker identification and understanding the molecular biology of intestinal tumorigenesis and adenocarcinoma maturation under varying intestinal conditions.
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Iron supplements are widely consumed. However, excess iron may accelerate intestinal tumorigenesis. To determine the effect of excess iron on intestinal tumor burden and protein expression changes between tumor and normal tissues, ApcMin/+ mice were fed control (adequate) and excess iron (45 and 450 mg iron/kg diet, respectively; n = 9/group) for 10 wk. Tumor burden was measured, and two-dimensional fluorescence difference gel electrophoresis was used to identify differentially expressed proteins in tumor and normal intestinal tissues. There was a significant increase (78.3%; p ≤ 0.05) in intestinal tumor burden (mm2/cm) with excess iron at wk 10. Of 980 analyzed protein spots, 69 differentially expressed (p ≤ 0.05) protein isoforms were identified, representing 55 genes. Of the isoforms, 56 differed (p ≤ 0.05) between tumor vs. normal tissues from the adequate iron group and 23 differed (p ≤ 0.05) between tumors from the adequate vs. excess iron. Differentially expressed proteins include those involved in cell integrity and adaptive response to reactive oxygen species (including, by gene ID: ANPEP, DPP7, ITGB1, PSMA1 HSPA5). Biochemical pathway analysis found that iron supplementation modulated four highly significant (p ≤ 0.05) functional networks. These findings enhance our understanding of interplay between dietary iron and intestinal tumorigenesis and may help develop more specific dietary guidelines regarding trace element intake.
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Suplementos Nutricionais , Modelos Animais de Doenças , Neoplasias Intestinais , Carga Tumoral , Animais , Camundongos , Humanos , Neoplasias Intestinais/metabolismo , Neoplasias Intestinais/patologia , Neoplasias Intestinais/genética , Ferro/metabolismo , Ferro da Dieta/administração & dosagem , Camundongos Endogâmicos C57BL , Masculino , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Proteômica/métodosRESUMO
Therapeutic management and prognostication for patients with B-acute lymphoblastic leukaemia (B-ALL) require appropriate disease subclassification. BCR::ABL1-like B-ALL is unique in that it is defined by a gene expression profile similar to BCR::ABL1+ B-ALL rather than a unifying recurrent translocation. Current molecular/cytogenetic techniques to identify this subtype are expensive, not widely accessible, have long turnaround times and/or require an adequate liquid biopsy. We have studied a total of 118 B-ALL cases from three institutions in two laboratories to identify surrogates for BCR::ABL1+/like B-ALL. We report that immunoglobulin joining chain (IGJ) and spermatogenesis associated serine-rich 2-like (SPATS2L) immunohistochemistry (IHC) sensitively and specifically identify BCR::ABL1+/like B-ALL. IGJ IHC positivity has a sensitivity of 83%, a specificity of 95%, a positive predictive value (PPV) of 89% and a negative predictive value (NPV) of 90%. SPATS2L staining has similar sensitivity and NPV but lower specificity (85%) and PPV (70%). The presence of either IGJ or SPATS2L staining augments the sensitivity (93%) and NPV (95%). While these findings would need to be validated in larger studies, they suggest that IGJ and/or SPATS2L IHC may be utilized in identifying BCR::ABL1-like B-ALL or in selecting B-ALL cases for confirmatory molecular/genetic testing, particularly in resource-limited settings.
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Leucemia-Linfoma Linfoblástico de Células Precursoras B , Leucemia-Linfoma Linfoblástico de Células Precursoras , Masculino , Humanos , Imuno-Histoquímica , Proteínas de Fusão bcr-abl/genética , Proteínas de Fusão bcr-abl/metabolismo , Leucemia-Linfoma Linfoblástico de Células Precursoras/tratamento farmacológico , Leucemia-Linfoma Linfoblástico de Células Precursoras B/diagnóstico , Leucemia-Linfoma Linfoblástico de Células Precursoras B/genética , Translocação GenéticaRESUMO
Pancreatic Ductal Adenocarcinoma (PDAC) is highly resistant to chemotherapy. Effective alternative therapies have yet to emerge, as chemotherapy remains the best available systemic treatment. However, the discovery of safe and available adjuncts to enhance chemotherapeutic efficacy can still improve survival outcomes. We show that a hyperglycemic state substantially enhances the efficacy of conventional single- and multi-agent chemotherapy regimens against PDAC. Molecular analyses of tumors exposed to high glucose levels reveal that the expression of GCLC (glutamate-cysteine ligase catalytic subunit), a key component of glutathione biosynthesis, is diminished, which in turn augments oxidative anti-tumor damage by chemotherapy. Inhibition of GCLC phenocopies the suppressive effect of forced hyperglycemia in mouse models of PDAC, while rescuing this pathway mitigates anti-tumor effects observed with chemotherapy and high glucose.
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Carcinoma Ductal Pancreático , Neoplasias Pancreáticas , Animais , Camundongos , Neoplasias Pancreáticas/tratamento farmacológico , Neoplasias Pancreáticas/genética , Carcinoma Ductal Pancreático/tratamento farmacológico , Carcinoma Ductal Pancreático/genética , Administração Cutânea , Glucose , Neoplasias PancreáticasRESUMO
Mucin 4 (MUC4) is a transmembrane mucin that, like most mucins, is not expressed in normal hematopoietic cells, but little is known about its expression in malignant hematopoiesis. B-acute lymphoblastic leukemia (B-ALL) consists of genetically distinct disease subtypes with similarities and differences in gene expression most frequently studied at the mRNA level, which is less amenable to widespread routine clinical use. Here, we demonstrate using immunohistochemistry (IHC) that MUC4 protein is expressed in less than 10% of B-ALL, with expression restricted to BCR::ABL1+ and BCR::ABL1-like (CRLF2 rearranged) subtypes of B-ALL (4/13, 31%). None (0/36, 0%) of the remaining B-ALL subtypes expressed MUC4. We compare clinical and pathologic features of MUC4+ and MUC4- BCR::ABL1+/like cases and most significantly report a possible shorter time to relapse for MUC4+ BCR::ABL1 B-ALL that would need to be validated in larger studies. In conclusion, MUC4 is a specific, albeit insensitive, marker for these high-risk subtypes of B-ALL. We propose that MUC4 IHC may be used diagnostically to rapidly identify these B-ALL subtypes, particularly in resource-limited settings or when an aspirate sample is not available for ancillary genetic studies.
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Mucina-4 , Leucemia-Linfoma Linfoblástico de Células Precursoras B , Leucemia-Linfoma Linfoblástico de Células Precursoras , Humanos , Proteínas de Fusão bcr-abl/genética , Mucina-4/genética , Leucemia-Linfoma Linfoblástico de Células Precursoras B/genética , Leucemia-Linfoma Linfoblástico de Células Precursoras/genética , RecidivaRESUMO
The canonical model of "small cell lung cancer" (SCLC) depicts tumors arising from dual inactivation of TP53 and RB1. However, many genomic studies have persistently identified tumors with no RB1 mutations. Here, we examined RB1 protein expression and function in SCLC. RB1 expression was examined by IHC analysis of 62 human SCLC tumors. These studies showed that â¼14% of SCLC tumors expressed abundant RB1 protein, which is associated with neuroendocrine gene expression and is enriched in YAP1 expression, but no other lineage proteins that stratify SCLC. SCLC cells and xenograft tumors with RB1 protein expression were sensitive to growth inhibition by the CDK4/6 inhibitor palbociclib, and this inhibition was shown to be dependent on RB1 expression by CRISPR knockout. Furthermore, a patient with biopsy-validated wild-type RB1 SCLC who received the CDK4/6 inhibitor abemaciclib demonstrated a dramatic decrease in mutant TP53 ctDNA allelic fraction from 62.1% to 0.4% and decreased tumor mass on CT scans. Importantly, IHC of the diagnostic biopsy specimen showed RB1 positivity. Finally, we identified a transcriptomics-based RB1 loss-of-function signature that discriminates between SCLC cells with or without RB1 protein expression and validated it in the patient who was responsive to abemaciclib, suggesting its potential use to predict CDK4/6 inhibitor response in patients with SCLC. Our study demonstrates that RB1 protein is an actionable target in a subgroup of SCLC, a cancer that exhibits no currently targetable mutations.
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Neoplasias Pulmonares , Neoplasias da Retina , Retinoblastoma , Carcinoma de Pequenas Células do Pulmão , Humanos , Neoplasias Pulmonares/tratamento farmacológico , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/metabolismo , Carcinoma de Pequenas Células do Pulmão/tratamento farmacológico , Carcinoma de Pequenas Células do Pulmão/genética , Carcinoma de Pequenas Células do Pulmão/metabolismo , Proteína do Retinoblastoma/genética , Mutação , Quinase 4 Dependente de Ciclina/genéticaRESUMO
BACKGROUND & AIMS: Mechanisms contributing to the onset and progression of Barrett's (BE)-associated esophageal adenocarcinoma (EAC) remain elusive. Here, we interrogated the major signaling pathways deregulated early in the development of Barrett's neoplasia. METHODS: Whole-transcriptome RNA sequencing analysis was performed in primary BE, EAC, normal esophageal squamous, and gastric biopsy tissues (n = 89). Select pathway components were confirmed by quantitative polymerase chain reaction in an independent cohort of premalignant and malignant biopsy tissues (n = 885). Functional impact of selected pathway was interrogated using transcriptomic, proteomic, and pharmacogenetic analyses in mammalian esophageal organotypic and patient-derived BE/EAC cell line models, in vitro and/or in vivo. RESULTS: The vast majority of primary BE/EAC tissues and cell line models showed hyperactivation of EphB2 signaling. Transcriptomic/proteomic analyses identified EphB2 as an endogenous binding partner of MYC binding protein 2, and an upstream regulator of c-MYC. Knockdown of EphB2 significantly impeded the viability/proliferation of EAC and BE cells in vitro/in vivo. Activation of EphB2 in normal esophageal squamous 3-dimensional organotypes disrupted epithelial maturation and promoted columnar differentiation programs, notably including MYC. EphB2 and MYC showed selective induction in esophageal submucosal glands with acinar ductal metaplasia, and in a porcine model of BE-like esophageal submucosal gland spheroids. Clinically approved inhibitors of MEK, a protein kinase that regulates MYC, effectively suppressed EAC tumor growth in vivo. CONCLUSIONS: The EphB2 signaling is frequently hyperactivated across the BE-EAC continuum. EphB2 is an upstream regulator of MYC, and activation of EphB2-MYC axis likely precedes BE development. Targeting EphB2/MYC could be a promising therapeutic strategy for this often refractory and aggressive cancer.
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Esôfago de Barrett , Carcinoma de Células Escamosas , Neoplasias Esofágicas , Suínos , Animais , Esôfago de Barrett/patologia , Efrina-B2/genética , Proteômica , Neoplasias Esofágicas/patologia , Carcinoma de Células Escamosas/patologia , Proto-Oncogenes , Proteínas Tirosina Quinases/genética , Quinases de Proteína Quinase Ativadas por Mitógeno/genética , Mamíferos/genéticaRESUMO
OBJECTIVE: To investigate the effect of superovulation with human chorionic gonadotropin (hCG) or gonadotropin-releasing hormone agonist (GnRHa) trigger on leukocyte density and expression of leukocyte-specific genes in the peri-implantation period in the mouse uterus. DESIGN: Laboratory research. SETTING: University laboratory facility. INTERVENTIONS: Female mice were mated to fertile male mice in one of three protocols: (1) natural mating or mating following injection with pregnant mare serum gonadotropin followed by trigger with (2) GnRHa or (3) hCG. Female mice were killed prior to implantation, 3 days after ovulation (E3.5), and the ovaries and uterine tissue were collected. Total RNA was isolated and assayed using quantitative reverse transcription polymerase chain reaction, and the uterine tissue was stained for histologic analysis of immune cell markers. MAIN OUTCOME MEASURES: Endometrial leukocyte (CD45) and vessel density (CD31) by immunohistochemical staining; expression of leukocyte markers CD11b, CD335, and CD22, by quantitative reverse transcription polymerase chain reaction in the whole uterine tissue. RESULTS: Superovulation decreased (compared with controls) the endometrial leukocyte density, based on the number of cells staining for CD45, and endometrial vessel density, based on the number of cells staining for CD31. Leukocyte density was additionally decreased in the GnRHa trigger group compared with that in the hCG trigger group. Superovulation with hCG and GnRHa triggers decreased the uterine expression of the B-cell marker CD22 compared with controls. The expression of the natural killer cell marker CD11b was decreased by the hCG trigger but not by the GnRHa. Abundance of mRNA encoding the CD335 natural killer cell marker was not affected by superovulation or trigger agent. CONCLUSIONS: In mice, superovulation with the GnRHa trigger compared with that with the hCG trigger differentially alters key immunologic factors in the uterine peri-implantation. These altered immunologic factors have roles in angiogenesis that may assist in elucidating the effects of assisted reproductive technologies on implantation efficiency and fetal growth and development.
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Gonadotropina Coriônica , Hormônio Liberador de Gonadotropina , Leucócitos , Superovulação , Animais , Gonadotropina Coriônica/farmacologia , Feminino , Hormônio Liberador de Gonadotropina/agonistas , Cavalos , Humanos , Fatores Imunológicos , Leucócitos/citologia , Camundongos , Gravidez , Útero/metabolismoRESUMO
Small-cell lung cancer (SCLC) can be subgrouped into common 'pure' and rare 'combined' SCLC (c-SCLC). c-SCLC features a mixed tumor histology of both SCLC and non-small-cell lung cancer (NSCLC). We performed targeted exome sequencing on 90 patients with SCLC, including two with c-SCLC, and discovered RUNX1T1 amplification specific to small cell tumors of both patients with c-SCLC, but in only 2 of 88 'pure' SCLC patients. RUNX1T1 was first identified in the fusion transcript AML1/ETO, which occurs in 12%-15% of acute myelogenous leukemia (AML). We further show higher expression of RUNX1T1 in the SCLC component of another c-SCLC tumor by in situ hybridization. RUNX1T1 expression was enriched in SCLC compared with all other cancers, including NSCLC, in both cell lines and tumor specimens, as shown by mRNA level and western blotting. Transcriptomic analysis of hallmark genes decreased by stable RUNX1T1 overexpression revealed a significant change in E2F targets. Validation experiments in multiple lung cancer cell lines showed that RUNX1T1 overexpression consistently decreased CDKN1A (p21) expression and increased E2F transcriptional activity, which is commonly altered in SCLC. Chromatin immunoprecipitation (ChIP) in these overexpressing cells demonstrated that RUNX1T1 interacts with the CDKN1A (p21) promoter region, which displayed parallel reductions in histone 3 acetylation. Furthermore, reduced p21 expression could be dramatically restored by HDAC inhibition using Trichostatin A. Reanalysis of ChIP-seq data in Kasumi-1 AML cells showed that knockdown of the RUNX1T1 fusion protein was associated with increased global acetylation, including the CDKN1A (p21) promoter. Thus, our study identifies RUNX1T1 as a biomarker and potential epigenetic regulator of SCLC.
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Epigênese Genética , Neoplasias Pulmonares/genética , Proteína 1 Parceira de Translocação de RUNX1/metabolismo , Carcinoma de Pequenas Células do Pulmão/genética , Acetilação , Linhagem Celular Tumoral , Inibidor de Quinase Dependente de Ciclina p21/genética , Inibidor de Quinase Dependente de Ciclina p21/metabolismo , Fatores de Transcrição E2F/metabolismo , Regulação Neoplásica da Expressão Gênica , Histonas/metabolismo , Humanos , Regiões Promotoras Genéticas , Proteína 1 Parceira de Translocação de RUNX1/genética , Regulação para Cima/genéticaRESUMO
BACKGROUND & AIMS: Esophageal adenocarcinoma (EAC) is resistant to standard chemoradiation treatments, and few targeted therapies are available. We used large-scale tissue profiling and pharmacogenetic analyses to identify deregulated signaling pathways in EAC tissues that might be targeted to slow tumor growth or progression. METHODS: We collected 397 biopsy specimens from patients with EAC and nonmalignant Barrett's esophagus (BE), with or without dysplasia. We performed RNA-sequencing analyses and used systems biology approaches to identify pathways that are differentially activated in EAC vs nonmalignant dysplastic tissues; pathway activities were confirmed with immunohistochemistry and quantitative real-time polymerase chain reaction analyses of signaling components in patient tissue samples. Human EAC (FLO-1 and EsoAd1), dysplastic BE (CP-B, CP-C, CP-D), and nondysplastic BE (CP-A) cells were incubated with pharmacologic inhibitors or transfected with small interfering RNAs. We measured effects on proliferation, colony formation, migration, and/or growth of xenograft tumors in nude mice. RESULTS: Comparisons of EAC vs nondysplastic BE tissues showed hyperactivation of transforming growth factor-ß (TGFB) and/or Jun N-terminal kinase (JNK) signaling pathways in more than 80% of EAC samples. Immunohistochemical analyses showed increased nuclear localization of phosphorylated JUN and SMAD proteins in EAC tumor tissues compared with nonmalignant tissues. Genes regulated by the TGFB and JNK pathway were overexpressed specifically in EAC and dysplastic BE. Pharmacologic inhibition or knockdown of TGFB or JNK signaling components in EAC cells (FLO-1 or EsoAd1) significantly reduced cell proliferation, colony formation, cell migration, and/or growth of xenograft tumors in mice in a SMAD4-independent manner. Inhibition of the TGFB pathway in BE cell lines reduced the proliferation of dysplastic, but not nondysplastic, cells. CONCLUSIONS: In a transcriptome analysis of EAC and nondysplastic BE tissues, we found the TGFB and JNK signaling pathways to be hyperactivated in EACs and the genes regulated by these pathways to be overexpressed in EAC and dysplastic BE. Inhibiting these pathways in EAC cells reduces their proliferation, migration, and formation of xenograft tumors. Strategies to block the TGFB and JNK signaling pathways might be developed for treatment of EAC.
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Adenocarcinoma/genética , Adenocarcinoma/metabolismo , Neoplasias Esofágicas/genética , Neoplasias Esofágicas/metabolismo , Sistema de Sinalização das MAP Quinases/genética , RNA Neoplásico/análise , Fator de Crescimento Transformador beta/metabolismo , Animais , Esôfago de Barrett/genética , Esôfago de Barrett/metabolismo , Esôfago de Barrett/patologia , Benzamidas/farmacologia , Linhagem Celular Tumoral , Movimento Celular , Proliferação de Células , Dioxóis/farmacologia , Feminino , Regulação Neoplásica da Expressão Gênica , Humanos , Masculino , Camundongos , Transplante de Neoplasias , Análise de Sequência com Séries de Oligonucleotídeos , Testes Farmacogenômicos , Proteínas Proto-Oncogênicas c-jun/metabolismo , Pirazóis/farmacologia , Quinolinas/farmacologia , Receptores de Fatores de Crescimento Transformadores beta/antagonistas & inibidores , Proteínas Smad/genética , Proteínas Smad/metabolismo , Biologia de Sistemas , Transcriptoma , Fator de Crescimento Transformador beta/antagonistas & inibidores , Fator de Crescimento Transformador beta/genética , Ensaio Tumoral de Célula-TroncoRESUMO
The majority of small cell lung cancer (SCLC) patients demonstrate initial chemo-sensitivity, whereas a distinct subgroup of SCLC patients, termed chemo-refractory, do not respond to treatment. There is little understanding of how to distinguish these patients prior to disease treatment. Here we used gene expression profiling to stratify SCLC into subgroups and characterized a molecular phenotype that may identify, in part, chemo-refractive SCLC patients. Two subgroups of SCLC were identified in both cell lines and tumors by the reciprocal expression of two genes; INSM1, a neuroendocrine transcription factor, and YAP1, a key mediator of the Hippo pathway. The great majority of tumors expressed INSM1, which was prognostic for increased progression-free survival and associated with chemo-sensitivity in cell lines. YAP1 is expressed in a minority of SCLC tumors and was shown in cell lines to be downstream of the retinoblastoma protein (RB1) and associated with decreased drug sensitivity. RB1 expression in SCLC cell lines sensitizes them to CDK4/6 inhibitors. Wild-type RB1 mutation status, used as a surrogate marker of YAP1 expression, was prognostic for decreased patient survival and increased chemo-refractory tumor response. Thus, the reciprocal expression of INSM1 and YAP1 appears to stratify SCLC into distinct subgroups and may be useful, along with RB1 mutation status, to identify chemo-refractory SCLC patients.
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BACKGROUND: Vitamin D deficiency may increase esophageal cancer risk. Vitamin D affects genes regulating proliferation, apoptosis, and differentiation and induces the tumor suppressor 15-hydroxyprostaglandin dehydrogenase (PGDH) in other cancers. This nonrandomized interventional study assessed effects of vitamin D supplementation in Barrett's esophagus (BE). We hypothesized that vitamin D supplementation may have beneficial effects on gene expression including 15-PGDH in BE. METHODS: BE subjects with low grade or no dysplasia received vitamin D3 (cholecalciferol) 50,000 international units weekly plus a proton pump inhibitor for 12 weeks. Esophageal biopsies from normal plus metaplastic BE epithelium and blood samples were obtained before and after vitamin D supplementation. Serum 25-hydroxyvitamin D was measured to characterize vitamin D status. Esophageal gene expression was assessed using microarrays. RESULTS: 18 study subjects were evaluated. The baseline mean serum 25-hydroxyvitamin D level was 27 ng/mL (normal ≥30 ng/mL). After vitamin D supplementation, 25-hydroxyvitamin D levels rose significantly (median increase of 31.6 ng/mL, p<0.001). There were no significant changes in gene expression from esophageal squamous or Barrett's epithelium including 15-PGDH after supplementation. CONCLUSION: BE subjects were vitamin D insufficient. Despite improved vitamin D status with supplementation, no significant alterations in gene expression profiles were noted. If vitamin D supplementation benefits BE, a longer duration or higher dose of supplementation may be needed.
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Esôfago de Barrett , Colecalciferol/sangue , Regulação Enzimológica da Expressão Gênica/efeitos dos fármacos , Hidroxiprostaglandina Desidrogenases/biossíntese , Vitamina D , Idoso , Esôfago de Barrett/tratamento farmacológico , Esôfago de Barrett/enzimologia , Esôfago de Barrett/patologia , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Projetos Piloto , Inibidores da Bomba de Prótons/administração & dosagem , Inibidores da Bomba de Prótons/farmacocinética , Vitamina D/administração & dosagem , Vitamina D/farmacocinéticaRESUMO
Small cell lung cancer (SCLC) is an aggressive cancer that represents ~15% of all lung cancers. Currently there are no targeted therapies to treat SCLC. Our genomic analysis of a metastatic SCLC cohort identified recurrent RICTOR amplification. Here, we examine the translational potential of this observation. RICTOR was the most frequently amplified gene observed (~14% patients), and co-amplified with FGF10 and IL7R on chromosome 5p13. RICTOR copy number variation correlated with RICTOR protein expression in SCLC cells. In parallel, cells with RICTOR copy number (CN) gain showed increased sensitivity to three mTOR inhibitors, AZD8055, AZD2014 and INK128 in cell growth assays, with AZD2014 demonstrating the best inhibition of downstream signaling. SCLC cells with RICTOR CN gain also migrated more rapidly in chemotaxis and scratch wound assays and were again more sensitive to mTOR inhibitors. The overall survival in SCLC patients with RICTOR amplification was significantly decreased (p = 0.021). Taken together, our results suggest that SCLC patients with RICTOR amplification may constitute a clinically important subgroup because of their potential response to mTORC1/2 inhibitors.
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Amplificação de Genes , Neoplasias Pulmonares/genética , Inibidores de Proteínas Quinases/farmacologia , Proteína Companheira de mTOR Insensível à Rapamicina/genética , Carcinoma de Pequenas Células do Pulmão/genética , Adulto , Idoso , Idoso de 80 Anos ou mais , Benzamidas , Benzoxazóis/farmacologia , Feminino , Fator 10 de Crescimento de Fibroblastos/genética , Humanos , Subunidade alfa de Receptor de Interleucina-7/genética , Neoplasias Pulmonares/tratamento farmacológico , Masculino , Pessoa de Meia-Idade , Terapia de Alvo Molecular , Morfolinas/farmacologia , Pirimidinas/farmacologia , Carcinoma de Pequenas Células do Pulmão/tratamento farmacológico , Análise de Sobrevida , Resultado do TratamentoRESUMO
IMPORTANCE: Esophageal adenocarcinoma and its precursor lesion Barrett esophagus have seen a dramatic increase in incidence over the past 4 decades yet marked genetic heterogeneity of this disease has precluded advances in understanding its pathogenesis and improving treatment. OBJECTIVE: To identify novel disease susceptibility variants in a familial syndrome of esophageal adenocarcinoma and Barrett esophagus, termed familial Barrett esophagus, by using high-throughput sequencing in affected individuals from a large, multigenerational family. DESIGN, SETTING, AND PARTICIPANTS: We performed whole exome sequencing (WES) from peripheral lymphocyte DNA on 4 distant relatives from our multiplex, multigenerational familial Barrett esophagus family to identify candidate disease susceptibility variants. Gene variants were filtered, verified, and segregation analysis performed to identify a single candidate variant. Gene expression analysis was done with both quantitative real-time polymerase chain reaction and in situ RNA hybridization. A 3-dimensional organotypic cell culture model of esophageal maturation was utilized to determine the phenotypic effects of our gene variant. We used electron microscopy on esophageal mucosa from an affected family member carrying the gene variant to assess ultrastructural changes. MAIN OUTCOMES AND MEASURES: Identification of a novel, germline disease susceptibility variant in a previously uncharacterized gene. RESULTS: A multiplex, multigenerational family with 14 members affected (3 members with esophageal adenocarcinoma and 11 with Barrett esophagus) was identified, and whole-exome sequencing identified a germline mutation (S631G) at a highly conserved serine residue in the uncharacterized gene VSIG10L that segregated in affected members. Transfection of S631G variant into a 3-dimensional organotypic culture model of normal esophageal squamous cells dramatically inhibited epithelial maturation compared with the wild-type. VSIG10L exhibited high expression in normal squamous esophagus with marked loss of expression in Barrett-associated lesions. Electron microscopy of squamous esophageal mucosa harboring the S631G variant revealed dilated intercellular spaces and reduced desmosomes. CONCLUSIONS AND RELEVANCE: This study presents VSIG10L as a candidate familial Barrett esophagus susceptibility gene, with a putative role in maintaining normal esophageal homeostasis. Further research assessing VSIG10L function may reveal pathways important for esophageal maturation and the pathogenesis of Barrett esophagus and esophageal adenocarcinoma.
Assuntos
Antígenos de Neoplasias/genética , Esôfago de Barrett/genética , Glicoproteínas de Membrana/genética , Adenocarcinoma/genética , Adulto , Idoso , Sequência de Aminoácidos , Células Cultivadas , Células Epiteliais/fisiologia , Neoplasias Esofágicas/genética , Feminino , Estudos de Associação Genética , Predisposição Genética para Doença , Mutação em Linhagem Germinativa , Humanos , Masculino , Pessoa de Meia-Idade , Adulto JovemRESUMO
CD30 is a cytokine receptor belonging to the TNF superfamily (TNFRSF8) that acts as a regulator of apoptosis. The presence of CD30 antigen is important in the diagnosis of Hodgkin disease and anaplastic large cell lymphoma. There have been sporadic reports of CD30 expression in nonlymphoid tumors, including malignant mesothelioma. Given the remarkable success of brentuximab vedotin, an antibody-drug conjugate directed against CD30 antigen, in lymphoid malignancies, we undertook a study to examine the incidence of CD30 in mesothelioma and to investigate the ability to target CD30 antigen in mesothelioma. Mesothelioma tumor specimens (N = 83) were examined for CD30 expression by IHC. Positive CD30 expression was noted in 13 mesothelioma specimens, primarily those of epithelial histology. There was no significant correlation of CD30 positivity with tumor grade, stage, or survival. Examination of four mesothelioma cell lines (H28, H2052, H2452, and 211H) for CD30 expression by both FACS analysis and confocal microscopy showed that CD30 antigen localized to the cell membrane. Brentuximab vedotin treatment of cultured mesothelioma cells produced a dose-dependent decrease in cell growth and viability at clinically relevant concentrations. Our studies validate the presence of CD30 antigen in a subgroup of epithelial-type mesothelioma tumors and indicate that selected mesothelioma patients may derive benefit from brentuximab vedotin treatment.
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Antígeno Ki-1/metabolismo , Neoplasias Pulmonares/metabolismo , Mesotelioma/metabolismo , Antineoplásicos/farmacologia , Apoptose/efeitos dos fármacos , Brentuximab Vedotin , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Humanos , Imunoconjugados/farmacologia , Neoplasias Pulmonares/tratamento farmacológico , Mesotelioma/tratamento farmacológico , Mesotelioma Maligno , Carcinoma de Pequenas Células do Pulmão/tratamento farmacológico , Carcinoma de Pequenas Células do Pulmão/metabolismoRESUMO
Unlike lung adenocarcinoma, little progress has been made in the treatment of squamous cell lung carcinoma (SCC). The Cancer Genome Atlas (TCGA) has recently reported that receptor tyrosine kinase signaling pathways are altered in 26% of SCC tumors, validating the importance of downstream Signal Transducers and Activators of Transcription 3 (STAT3) activity as a prime therapeutic target in this cancer. In the present report we examine the status of an endogenous inhibitor of STAT3, called Protein Inhibitor of Activated STAT3 (PIAS3), in SCC and its potential role in this disease. We examine PIAS3 expression in SCC tumors and cell lines by immunohistochemistry of a tissue microarray and western blotting. PIAS3 mRNA expression and survival data are analyzed in the TCGA data set. SCC cell lines are treated with curcumin to regulate PIAS3 expression and cell growth. PIAS3 protein expression is decreased in a majority of lung SCC tumors and cell lines. Analysis of PIAS3 mRNA transcript levels demonstrated that low PIAS3 levels predicted poor survival; Cox regression analysis revealed a hazard ratio of 0.57 (95% CI: 0.37-0.87), indicating a decrease in the risk of death by 43% for every unit elevation in PIAS3 gene expression. Curcumin treatment increased endogenous PIAS3 expression and decreased cell growth and viability in Calu-1 cells, a model of SCC. Our results implicate PIAS3 loss in the pathology of lung SCC and raise the therapeutic possibility of upregulating PIAS3 expression as a single target that can suppress signaling from the multiple receptor tyrosine kinase receptors found to be amplified in SCC.
Assuntos
Carcinoma de Células Escamosas/metabolismo , Neoplasias Pulmonares/metabolismo , Chaperonas Moleculares/metabolismo , Proteínas Inibidoras de STAT Ativados/metabolismo , Linhagem Celular , Linhagem Celular Tumoral , Curcumina/farmacologia , Humanos , Chaperonas Moleculares/genética , Prognóstico , Proteínas Inibidoras de STAT Ativados/genética , RNA Mensageiro/metabolismoRESUMO
Non-steroidal anti-inflammatory drugs prevent colorectal cancer by inhibiting cyclooxygenase (COX) enzymes that synthesize tumor-promoting prostaglandins. 15-hydroxyprostaglandin dehydrogenase (15-PGDH) is a tumor suppressor that degrades tumor-promoting prostaglandins. Murine knockout of 15-PGDH increases susceptibility to azoxymethane-induced colon tumors. It also renders these mice resistant to celecoxib, a selective inhibitor of inducible COX-2 during colon neoplasia. Similarly, humans with low colonic 15-PGDH are also resistant to colon adenoma prevention with celecoxib. Here, we used aspirin and sulindac, which inhibit both COX-1 and COX-2, in order to determine if these broader COX inhibitors can prevent colon tumors in 15-PGDH knockout (KO) mice. Unlike celecoxib, sulindac proved highly effective in colon tumor prevention of 15-PGDH KO mice. Significantly, however, aspirin demonstrated no effect on colon tumor incidence in either 15-PGDH wild-type or KO mice, despite a comparable reduction in colonic mucosal Prostaglandin E2 (PGE2) levels by both sulindac and aspirin. Notably, colon tumor prevention activity by sulindac was accompanied by a marked induction of lymphoid aggregates and proximal colonic inflammatory mass lesions, a side effect seen to a lesser degree with celecoxib, but not with aspirin. These findings suggest that sulindac may be the most effective agent for colon cancer prevention in humans with low 15-PGDH, but its use may also be associated with inflammatory lesions in the colon.
Assuntos
Anti-Inflamatórios não Esteroides/farmacologia , Neoplasias do Colo/tratamento farmacológico , Neoplasias do Colo/prevenção & controle , Hidroxiprostaglandina Desidrogenases/genética , Sulindaco/farmacologia , Animais , Antineoplásicos/farmacologia , Aspirina/farmacologia , Azoximetano , Carcinógenos , Celecoxib , Quimioprevenção , Neoplasias do Colo/induzido quimicamente , Ciclo-Oxigenase 1/efeitos dos fármacos , Ciclo-Oxigenase 2/efeitos dos fármacos , Inibidores de Ciclo-Oxigenase 2/farmacologia , Dinoprostona/metabolismo , Inflamação/imunologia , Mucosa Intestinal/patologia , Proteínas de Membrana/efeitos dos fármacos , Camundongos , Camundongos Knockout , Pirazóis/farmacologia , Sulfonamidas/farmacologiaRESUMO
PURPOSE: Deregulation of STAT3 activation is a hallmark of many cancer cells, and the underlying mechanisms are subject to intense investigation. We examined the extent of PIAS3 expression in mesothelioma cells and human tumor samples and determined the functional effects of PIAS3 expression on STAT3 signaling. EXPERIMENTAL DESIGN: We evaluated the expression of PIAS3 in mesothelioma tumors from patients and correlated the expression levels with the course of the disease. We also measured the effects of enhanced PIAS3 activity on STAT3 signaling, cellular growth, and viability in cultured mesothelioma cells. RESULTS: Gene expression databases revealed that mesotheliomas have the lowest levels of PIAS3 transcripts among solid tumors. PIAS3 expression in human mesothelioma tumors is significantly correlated with overall survival intervals (P = 0.058). The high expression of PIAS3 is predictive of a favorable prognosis and decreases the probability of death within one year after diagnosis by 44%. PIAS3 expression is functionally linked to STAT3 activation in mesothelioma cell lines. STAT3 downregulation with siRNA or enhanced expression of PIAS3 both inhibited mesothelioma cell growth and induced apoptosis. Mesothelioma cells are sensitive to curcumin and respond by the induction of PIAS3. Corroborative evidence has been obtained from STAT3 inhibition experiments. Exposure of the cells to a peptide derived from the PIAS3 protein that interferes with STAT3 function resulted in apoptosis induction and the inhibition of cell growth. CONCLUSION: These results suggest that PIAS3 protein expression impacts survival in patients with mesothelioma and that PIAS3 activation could become a therapeutic strategy. Clin Cancer Res; 20(19); 5124-32. ©2014 AACR.
Assuntos
Neoplasias Pulmonares/genética , Neoplasias Pulmonares/metabolismo , Mesotelioma/genética , Mesotelioma/metabolismo , Chaperonas Moleculares/genética , Proteínas Inibidoras de STAT Ativados/genética , Fator de Transcrição STAT3/metabolismo , Apoptose/efeitos dos fármacos , Apoptose/genética , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Curcumina/farmacologia , Expressão Gênica , Humanos , Neoplasias Pulmonares/mortalidade , Mesotelioma/mortalidade , Mesotelioma Maligno , Chaperonas Moleculares/química , Chaperonas Moleculares/metabolismo , Chaperonas Moleculares/farmacologia , Fragmentos de Peptídeos/farmacologia , Prognóstico , Proteínas Inibidoras de STAT Ativados/química , Proteínas Inibidoras de STAT Ativados/metabolismo , Proteínas Inibidoras de STAT Ativados/farmacologia , Interferência de RNA , RNA Mensageiro/genética , RNA Interferente Pequeno/genética , Fator de Transcrição STAT3/genéticaRESUMO
BACKGROUND: There is growing interest in defining the somatic mutations associated with small-cell lung cancer (SCLC). Unfortunately, a serious blockade to genomic analyses of this disease is a limited access to tumors because surgery is rarely performed. We used our clinical/pathologic database of SCLC patients to determine the availability of biopsy specimens that could be used for genomic studies and to identify tumors for initial oncogene analysis. METHODS: DNA was extracted from six tumors, three primary and three metastatic, and analyzed by SEQUENOM platform technology. RESULTS: Primary-resected tumor tissue represents less than 3% of all diagnostic specimens in this disease, highlighting the limited access to tissue sufficient for comprehensive genomic analyses. We identified an activating M918T RET somatic mutation in a metastatic SCLC tumor specimen. Bioinformatic search identified RET mutations in other SCLC studies. Stable overexpression of both mutant M918T and wild-type RET in two SCLC cell lines, H1048 and SW1271, activated ERK signaling, MYC expression, and increased cell proliferation, particularly by mutant RET. Stable cells became sensitized to the RET tyrosine kinase inhibitors, vandetanib and ponatinib. Further analysis of RET mRNA expression in SCLC revealed wide variability in both cells and tumors, and SCLC cells demonstrated significantly higher RET expression compared with adenocarcinoma lung cells. CONCLUSIONS: Our data suggest that a subpopulation of SCLC patients may derive benefit from tyrosine kinase inhibitors targeting RET. Coupled with the presence of RET fusion proteins in non-small-cell lung cancer, our data indicate an emerging role for RET in SCLC.
Assuntos
Regulação Neoplásica da Expressão Gênica , Neoplasias Pulmonares/genética , Mutação , Proteínas Proto-Oncogênicas c-ret/genética , RNA Neoplásico/genética , Carcinoma de Pequenas Células do Pulmão/genética , Idoso , Linhagem Celular Tumoral , Proliferação de Células , Análise Mutacional de DNA , Seguimentos , Humanos , Immunoblotting , Imuno-Histoquímica , Neoplasias Pulmonares/metabolismo , Neoplasias Pulmonares/patologia , Proteínas Proto-Oncogênicas c-ret/metabolismo , Reação em Cadeia da Polimerase em Tempo Real , Estudos Retrospectivos , Transdução de Sinais , Carcinoma de Pequenas Células do Pulmão/metabolismo , Carcinoma de Pequenas Células do Pulmão/patologiaRESUMO
Rational design and develop of targeted contrast agents binding to cancer-related proteins will achieve more accurate cancer diagnosis and prognosis by magnetic resonance (MR) imaging. CREKA is a tumor-homing pentapeptide (Cys-Arg-Glu-Lys-Ala) specifically homes to fibrin-fibronectin complexes abundantly expressed in tumor microenvironment. In this study, we developed and evaluated a CREKA peptide targeted multiplexed Gd-MR probe (CREKA-Tris-Gd(DOTA)3) for MR imaging of breast tumors. CREKA and azide bearing Gd(III) was attached to a maleimide-functional trialkyne scaffold via thiol-maleimide and azide-alkyne click chemistry, respectively. CREKA-Tris-Gd(DOTA)3 has a well-defined structure with a molecular weight of 2914 Da. The T1 relaxivity of CREKA-Tris-Gd(DOTA)3 is 8.06 mM(-1) s(-1) per Gd (24.18 mM(-1) s(-1) per molecule) at room temperature and 3 T. Fluorescence imaging showed high binding specificity of CREKA to a 4T1 breast tumor model in mice while it was not found for the scrambled CREKA (CERAK). The CREKA peptide-targeted contrast agent resulted in greater contrast enhancement than the corresponding CERAK agent and the commercialized contrast agent ProHance(®) in tumor at a dose of 0.1 mmol Gd/kg in female athymic mice bearing 4T1 breast carcinoma xenograft. This small molecular contrast agent was easily excreted from body after imaging indicated low toxicity. The targeted MRI contrast agent has a potential for specific cancer molecular imaging with MRI.