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1.
Mol Carcinog ; 62(2): 145-159, 2023 02.
Artigo em Inglês | MEDLINE | ID: mdl-36218231

RESUMO

Doublecortin like kinase 1 (DCLK1) plays a crucial role in several cancers including colon and pancreatic adenocarcinomas. However, its role in squamous cell carcinoma (SCC) remains unknown. To this end, we examined DCLK1 expression in head and neck SCC (HNSCC) and anal SCC (ASCC). We found that DCLK1 is elevated in patient SCC tissue, which correlated with cancer progression and poorer overall survival. Furthermore, DCLK1 expression is significantly elevated in human papilloma virus negative HNSCC, which are typically aggressive with poor responses to therapy. To understand the role of DCLK1 in tumorigenesis, we used specific shRNA to suppress DCLK1 expression. This significantly reduced tumor growth, spheroid formation, and migration of HNSCC cancer cells. To further the translational relevance of our studies, we sought to identify a selective DCLK1 inhibitor. Current attempts to target DCLK1 using pharmacologic approaches have relied on nonspecific suppression of DCLK1 kinase activity. Here, we demonstrate that DiFiD (3,5-bis [2,4-difluorobenzylidene]-4-piperidone) binds to DCLK1 with high selectivity. Moreover, DiFiD mediated suppression of DCLK1 led to G2/M arrest and apoptosis and significantly suppressed tumor growth of HNSCC xenografts and ASCC patient derived xenografts, supporting that DCLK1 is critical for SCC growth.


Assuntos
Carcinoma de Células Escamosas , Neoplasias de Cabeça e Pescoço , Humanos , Apoptose , Carcinoma de Células Escamosas/tratamento farmacológico , Carcinoma de Células Escamosas/genética , Linhagem Celular Tumoral , Quinases Semelhantes a Duplacortina , Pontos de Checagem da Fase G2 do Ciclo Celular , Peptídeos e Proteínas de Sinalização Intracelular/genética , Proteínas Serina-Treonina Quinases/metabolismo , Carcinoma de Células Escamosas de Cabeça e Pescoço/genética , Animais
3.
Cells ; 10(7)2021 06 26.
Artigo em Inglês | MEDLINE | ID: mdl-34206989

RESUMO

Honokiol (HNK) is a biphenolic compound that has been used in traditional medicine for treating various ailments, including cancers. In this study, we determined the effect of HNK on colon cancer cells in culture and in a colitis-associated cancer model. HNK treatment inhibited proliferation and colony formation while inducing apoptosis. In addition, HNK suppressed colonosphere formation. Molecular docking suggests that HNK interacts with reserve stem cell marker protein DCLK1, with a binding energy of -7.0 Kcal/mol. In vitro kinase assays demonstrated that HNK suppressed the DCLK1 kinase activity. HNK also suppressed the expression of additional cancer stem cell marker proteins LGR5 and CD44. The Hippo signaling pathway is active in intestinal stem cells. In the canonical pathway, YAP1 is phosphorylated at Ser127 by upstream Mst1/2 and Lats1/2. This results in the sequestration of YAP1 in the cytoplasm, thereby not allowing YAP1 to translocate to the nucleus and interact with TEAD1-4 transcription factors to induce gene expression. However, HNK suppressed Ser127 phosphorylation in YAP1, but the protein remains sequestered in the cytoplasm. We further determined that this occurs by YAP1 interacting with PUMA. To determine if this also occurs in vivo, we performed studies in an AOM/DSS induced colitis-associated cancer model. HNK administered by oral gavage at a dose of 5mg/kg bw for 24 weeks demonstrated a significant reduction in the expression of YAP1 and TEAD1 and in the stem marker proteins. Together, these data suggest that HNK prevents colon tumorigenesis in part by inducing PUMA-YAP1 interaction and cytoplasmic sequestration, thereby suppressing the oncogenic YAP1 activity.


Assuntos
Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Compostos de Bifenilo/farmacologia , Carcinogênese/patologia , Neoplasias do Colo/patologia , Lignanas/farmacologia , Células-Tronco Neoplásicas/patologia , Fatores de Transcrição/metabolismo , Animais , Apoptose/efeitos dos fármacos , Proteínas Reguladoras de Apoptose/metabolismo , Biomarcadores Tumorais/metabolismo , Carcinogênese/efeitos dos fármacos , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Colite/complicações , Quinases Semelhantes a Duplacortina , Regulação para Baixo/efeitos dos fármacos , Via de Sinalização Hippo , Humanos , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Masculino , Camundongos Endogâmicos ICR , Modelos Biológicos , Células-Tronco Neoplásicas/efeitos dos fármacos , Ligação Proteica/efeitos dos fármacos , Proteínas Serina-Treonina Quinases/metabolismo , Proteínas Proto-Oncogênicas/metabolismo , Transdução de Sinais/efeitos dos fármacos , Ensaio Tumoral de Célula-Tronco , Proteínas de Sinalização YAP
4.
Biomolecules ; 11(7)2021 06 25.
Artigo em Inglês | MEDLINE | ID: mdl-34202286

RESUMO

A new and readily available pentafluorothiophenyl-substituted N-methyl-piperidone curcuminoid 1a was prepared and investigated for its anti-proliferative, pro-apoptotic and cancer stem cell-differentiating activities against a panel of human tumor cell lines derived from various tumor entities. The compound 1a was highly anti-proliferative and reached IC50 values in the nanomolar concentration range. 1a was superior to the known anti-tumorally active curcuminoid EF24 (2) and its known N-ethyl-piperidone analog 1b in all tested tumor cell lines. Furthermore, 1a induced a noticeable increase of intracellular reactive oxygen species in HT-29 colon adenocarcinoma cells, which possibly leads to a distinct increase in sub-G1 cells, as assessed by cell cycle analysis. A considerable activation of the executioner-caspases 3 and 7 as well as nuclei fragmentation, cell rounding, and membrane protrusions suggest the triggering of an apoptotic mechanism. Yet another effect was the re-organization of the actin cytoskeleton shown by the formation of stress fibers and actin aggregation. 1a also caused cell death in the adherently cultured glioblastoma cell lines U251 and Mz54. We furthermore observed that 1a strongly suppressed the stem cell properties of glioma stem-like cell lines including one primary line, highlighting the potential therapeutic relevance of this new compound.


Assuntos
Antineoplásicos Fitogênicos/síntese química , Antineoplásicos Fitogênicos/farmacologia , Diarileptanoides/síntese química , Diarileptanoides/farmacologia , Ensaios de Seleção de Medicamentos Antitumorais/métodos , Animais , Animais Geneticamente Modificados , Apoptose/efeitos dos fármacos , Apoptose/fisiologia , Proliferação de Células/efeitos dos fármacos , Proliferação de Células/fisiologia , Curcumina/síntese química , Curcumina/farmacologia , Relação Dose-Resposta a Droga , Células HCT116 , Células HT29 , Humanos , Células MCF-7 , Peixe-Zebra
5.
FASEB J ; 35(5): e21533, 2021 05.
Artigo em Inglês | MEDLINE | ID: mdl-33826787

RESUMO

Polycystic kidney disease (PKD) is a genetic disorder characterized by aberrant renal epithelial cell proliferation and formation and progressive growth of numerous fluid-filled cysts within the kidneys. Previously, we showed that there is elevated Notch signaling compared to normal renal epithelial cells and that Notch signaling contributes to the proliferation of cystic cells. Quinomycin A, a bis-intercalator peptide, has previously been shown to target the Notch signaling pathway and inhibit tumor growth in cancer. Here, we show that Quinomycin A decreased cell proliferation and cyst growth of human ADPKD cyst epithelial cells cultured within a 3D collagen gel. Treatment with Quinomycin A reduced kidney weight to body weight ratio and decreased renal cystic area and fibrosis in Pkd1RC/RC ; Pkd2+/- mice, an orthologous PKD mouse model. This was accompanied by reduced expression of Notch pathway proteins, RBPjk and HeyL and cell proliferation in kidneys of PKD mice. Quinomycin A treatments also normalized cilia length of cyst epithelial cells derived from the collecting ducts. This is the first study to demonstrate that Quinomycin A effectively inhibits PKD progression and suggests that Quinomycin A has potential therapeutic value for PKD patients.


Assuntos
Antibacterianos/farmacologia , Cistos/tratamento farmacológico , Modelos Animais de Doenças , Equinomicina/farmacologia , Doenças Renais Policísticas/complicações , Canais de Cátion TRPP/fisiologia , Animais , Cistos/etiologia , Cistos/metabolismo , Cistos/patologia , Progressão da Doença , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout
6.
JCI Insight ; 6(8)2021 03 30.
Artigo em Inglês | MEDLINE | ID: mdl-33784251

RESUMO

Despite the recent launch of tolvaptan, the search for safer polycystic kidney disease (PKD) drugs continues. Ciclopirox (CPX) or its olamine salt (CPX-O) is contained in a number of commercially available antifungal agents. CPX is also reported to possess anticancer activity. Several mechanisms of action have been proposed, including chelation of iron and inhibition of iron-dependent enzymes. Here, we show that CPX-O inhibited in vitro cystogenesis of primary human PKD cyst-lining epithelial cells cultured in a 3D collagen matrix. To assess the in vivo role of CPX-O, we treated PKD mice with CPX-O. CPX-O reduced the kidney-to-body weight ratios of PKD mice. The CPX-O treatment was also associated with decreased cell proliferation, decreased cystic area, and improved renal function. Ferritin levels were markedly elevated in cystic kidneys of PKD mice, and CPX-O treatment reduced renal ferritin levels. The reduction in ferritin was associated with increased ferritinophagy marker nuclear receptor coactivator 4, which reversed upon CPX-O treatment in PKD mice. Interestingly, these effects on ferritin appeared independent of iron. These data suggest that CPX-O can induce ferritin degradation via ferritinophagy, which is associated with decreased cyst growth progression in PKD mice. Most importantly these data indicate that CPX-O has the potential to treat autosomal dominant PKD.


Assuntos
Antifúngicos/farmacologia , Ciclopirox/farmacologia , Cistos , Ferritinas/metabolismo , Rim/efeitos dos fármacos , Doenças Renais Policísticas , Animais , Antifúngicos/uso terapêutico , Proliferação de Células , Ciclopirox/uso terapêutico , Colágeno , Células Epiteliais/efeitos dos fármacos , Células Epiteliais/patologia , Humanos , Camundongos Endogâmicos C57BL , Coativadores de Receptor Nuclear/metabolismo , Tamanho do Órgão , Doenças Renais Policísticas/tratamento farmacológico , Doenças Renais Policísticas/metabolismo , Doenças Renais Policísticas/patologia , Rim Policístico Autossômico Dominante
7.
Chem Biol Drug Des ; 97(2): 325-340, 2021 02.
Artigo em Inglês | MEDLINE | ID: mdl-32909340

RESUMO

Spiropyrans have been extensively investigated because of their thermo- and photochromic characteristics, but their biotherapeutic properties have not been explored much. We report anti-proliferative properties of a novel 3,3'-azadimethylene dinaphthospiropyran 11. Dibenzospiropyrans and dinaphthospiropyrans were synthesized by a simple and expedient method using acid-catalyzed aldol condensation of salicylaldehyde and 2-hydroxy-1-naphthaldehyde, respectively, with cyclic ketones. Together with structural elucidation by 2D NMR and X-ray crystallography studies, we provide a putative mechanism for their formation. Compound 11 showed solvatochromism and exhibited altered spectral characteristics depending on the pH. In acidic conditions, 11 remains in open form, whereas upon alkalinization it reverts back to closed form. Based on the in vitro anti-proliferative activity in H441, HCT-116, MiaPaCa-2, and Panc-1 cancer cell lines, 11 was submitted to further investigation. It reduced HCT116 colonosphere formation and demonstrated induction of caspase cascade, suggesting apoptosis. In vitro proliferation assays also suggested that HCl and trifluoroacetate salts of 11 are more effective. Treatment of mice carrying HCT-116 xenografts with 11 (5 µg/day, intraperitoneal for 3 weeks) suppressed tumor growth by 62%. Overall, the results reveal a new series of structurally complex, but relatively easy to synthesize molecules of which compound 11 represents a lead for anticancer development.


Assuntos
Antineoplásicos/uso terapêutico , Benzopiranos/química , Neoplasias do Colo/tratamento farmacológico , Indóis/química , Nitrocompostos/química , Animais , Antineoplásicos/química , Antineoplásicos/farmacologia , Apoptose/efeitos dos fármacos , Benzopiranos/farmacologia , Benzopiranos/uso terapêutico , Pontos de Checagem do Ciclo Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Neoplasias do Colo/metabolismo , Neoplasias do Colo/patologia , Cristalografia por Raios X , Ensaios de Seleção de Medicamentos Antitumorais , Humanos , Indóis/farmacologia , Indóis/uso terapêutico , Camundongos , Conformação Molecular , Nitrocompostos/farmacologia , Nitrocompostos/uso terapêutico , Transplante Heterólogo
8.
Nutr Cancer ; 73(11-12): 2249-2271, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-33305598

RESUMO

Bladder cancer is the 9th most prevalent cancer worldwide and carries a protracted treatment course with significant patient expense, morbidity, and mortality. Over 95% of bladder cancers arise from the urothelium and invade into the underlying muscle layer before metastasizing. Trans-urethral resection and BCG therapy is the current first-line treatment for non-muscle invasive bladder cancer but carries a high rate of tumor recurrence and progression. The poor outcomes associated with advanced disease indicate the urgent need for new and improved treatment strategies. There is increasing investigation into the molecular signaling pathways involved in bladder cancer pathogenesis with the goal of uncovering potential therapeutic targets. This article reviews the major signaling pathways implicated in bladder cancer, including PI3K/AKT/mTOR, Ras/Raf/MEK/MAPK, NF-κB, Wnt/ß-catenin, Notch, Hedgehog, Hippo, JAK/STAT, and TGF-ß as well as major cellular receptors central to cancer pathophysiology, including EGFR, Her2, FGFR, and VEGF. We also discuss various naturally occurring phytochemicals that show evidence of targeting these molecular pathways including curcumin, resveratrol, green tea polyphenols, sulforaphane, erucin, genistein, genipin, baicalein, quercetin, isoquercitin, vitamin E, parthenolide, dioscin, triptolide, kaempferol, pterostilbene, isoliquiritigenin, and escin. This review highlights the potential use of these compounds in treatment of bladder cancer.


Assuntos
Neoplasias da Bexiga Urinária , Genisteína , Humanos , Fosfatidilinositol 3-Quinases , Compostos Fitoquímicos/farmacologia , Compostos Fitoquímicos/uso terapêutico , Transdução de Sinais , Neoplasias da Bexiga Urinária/tratamento farmacológico
9.
Cell Death Dis ; 11(2): 149, 2020 02 24.
Artigo em Inglês | MEDLINE | ID: mdl-32094348

RESUMO

Osteosarcoma (OS) is the most common primary bone tumor that primarily affects children and adolescents. Studies suggested that dysregulation JAK/STAT signaling promotes the development of OS. Cells treated with pimozide, a STAT5 inhibitor suppressed proliferation and colony formation and induced sub G0/G1 cell cycle arrest and apoptosis. There was a reduction in cyclin D1 and CDK2 expression and Rb phosphorylation, and activation of Caspase-3 and PARP cleavage. In addition, pimozide suppressed the formation of 3-dimensional osteospheres and growth of the cells in the Tumor in a Dish lung organoid system. Furthermore, there was a reduction in expression of cancer stem cell marker proteins DCLK1, CD44, CD133, Oct-4, and ABCG2. More importantly, it was the short form of DCLK1 that was upregulated in osteospheres, which was suppressed in response to pimozide. We further confirmed by flow cytometry a reduction in DCLK1+ cells. Moreover, pimozide inhibits the phosphorylation of STAT5, STAT3, and ERK in OS cells. Molecular docking studies suggest that pimozide interacts with STAT5A and STAT5B with binding energies of -8.4 and -6.4 Kcal/mol, respectively. Binding was confirmed by cellular thermal shift assay. To further understand the role of STAT5, we knocked down the two isoforms using specific siRNAs. While knockdown of the proteins did not affect the cells, knockdown of STAT5B reduced pimozide-induced necrosis and further enhanced late apoptosis. To determine the effect of pimozide on tumor growth in vivo, we administered pimozide intraperitoneally at a dose of 10 mg/kg BW every day for 21 days in mice carrying KHOS/NP tumor xenografts. Pimozide treatment significantly suppressed xenograft growth. Western blot and immunohistochemistry analyses also demonstrated significant inhibition of stem cell marker proteins. Together, these data suggest that pimozide treatment suppresses OS growth by targeting both proliferating cells and stem cells at least in part by inhibiting the STAT5 signaling pathway.


Assuntos
Osteossarcoma/tratamento farmacológico , Pimozida/farmacologia , Fator de Transcrição STAT5/farmacologia , Proteínas Supressoras de Tumor/farmacologia , Animais , Apoptose/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Pontos de Checagem da Fase G1 do Ciclo Celular/efeitos dos fármacos , Humanos , Células-Tronco Neoplásicas/efeitos dos fármacos , Células-Tronco Neoplásicas/metabolismo , Osteossarcoma/metabolismo , Fator de Transcrição STAT5/efeitos dos fármacos , Fator de Transcrição STAT5/metabolismo , Transdução de Sinais/efeitos dos fármacos
10.
Sci Rep ; 10(1): 1290, 2020 Jan 28.
Artigo em Inglês | MEDLINE | ID: mdl-31992775

RESUMO

Cancer stem cells (CSCs) have the ability to self-renew and induce drug resistance and recurrence in colorectal cancer (CRC). As current chemotherapy doesn't eliminate CSCs completely, there is a need to identify novel agents to target them. We investigated the effects of cucurbitacin B (C-B) or I (C-I), a natural compound that exists in edible plants (bitter melons, cucumbers, pumpkins and zucchini), against CRC. C-B or C-I inhibited proliferation, clonogenicity, induced G2/M cell-cycle arrest and caspase-mediated-apoptosis of CRC cells. C-B or C-I suppressed colonosphere formation and inhibited expression of CD44, DCLK1 and LGR5. These compounds inhibited notch signaling by reducing the expression of Notch 1-4 receptors, their ligands (Jagged 1-2, DLL1,3,4), γ-secretase complex proteins (Presenilin 1, Nicastrin), and downstream target Hes-1. Molecular docking showed that C-B or C-I binds to the ankyrin domain of Notch receptor, which was confirmed using the cellular thermal shift assay. Finally, C-B or C-I inhibited tumor xenograft growth in nude mice and decreased the expression of CSC-markers and notch signaling proteins in tumor tissues. Together, our study suggests that C-B and C-I inhibit colon cancer growth by inhibiting Notch signaling pathway.


Assuntos
Neoplasias do Colo/tratamento farmacológico , Simulação de Acoplamento Molecular , Receptores Notch , Transdução de Sinais/efeitos dos fármacos , Triterpenos , Animais , Neoplasias do Colo/química , Neoplasias do Colo/metabolismo , Neoplasias do Colo/patologia , Células HCT116 , Humanos , Masculino , Camundongos , Camundongos Nus , Proteínas de Neoplasias/química , Proteínas de Neoplasias/metabolismo , Domínios Proteicos , Receptores Notch/química , Receptores Notch/metabolismo , Triterpenos/química , Triterpenos/farmacologia , Ensaios Antitumorais Modelo de Xenoenxerto
11.
J Biol Chem ; 295(4): 1021-1035, 2020 01 24.
Artigo em Inglês | MEDLINE | ID: mdl-31836665

RESUMO

Wnt signaling regulates immunomodulatory functions during infection and inflammation. Employing NCCIT and HCT116 cells, having high endogenous Wnt signaling, we observed elevated levels of low-density lipoprotein receptor-related protein 5/6 (LRP5/6) and Frizzled class receptor 10 (FZD10) and increases in ß-catenin, doublecortin-like kinase 1 (DCLK1), CD44 molecule (CD44), and aldehyde dehydrogenase 1 family member A1 (ALDH1A1). siRNA-induced knockdown of these receptors antagonized TOPflash reporter activity and spheroid growth in vitro and elevated Wnt-inhibitory factor 1 (WIF1) activity. Elevated mRNA and protein levels of LRP5/6 and FZD10 paralleled expression of WNT2b and WNT4 in colonic crypts at days 6 and 12 post-infection with Citrobacter rodentium (CR) and tended to decline at days 20-34. The CR mutant escV or the tankyrase inhibitor XAV939 attenuated these responses. A three-dimensional organoid assay in colonic crypts isolated from CR-infected mice revealed elevated levels of LRP5/6 and FZD10 and ß-catenin co-localization with enhancer of zeste 2 polycomb repressive complex 2 subunit (EZH2). Co-immunoprecipitation in the membrane fraction revealed that axin associates with LRP5/6 in CR-infected crypts, and this association was correlated with increased ß-catenin. Colon tumors from either CR-infected ApcPMin/+ or azoxymethane/dextran sodium sulfate (AOM/DSS)-treated mice had high LRP5/6 or FZD10 levels, and chronic Notch blockade through the γ-secretase inhibitor dibenzazepine down-regulated LRP5/6 and FZD10 expression. In CR-responsive CT-26 cells, siRNA-induced LRP5/6 or FZD10 knockdown antagonized TOPflash reporter activity. Elevated miR-153-3p levels correlated with LRP5/6 and FZD10, and miR-153-3p sequestration via a plasmid-based miR inhibitor system attenuated Wnt signaling. We conclude that infection-induced signals from the plasma membrane epigenetically regulate Wnt signaling.


Assuntos
Membrana Celular/metabolismo , Citrobacter rodentium/fisiologia , Infecções por Enterobacteriaceae/genética , Epigênese Genética , Via de Sinalização Wnt/genética , Receptores Frizzled/genética , Receptores Frizzled/metabolismo , Células HCT116 , Células HEK293 , Humanos , Ligantes , Proteína-5 Relacionada a Receptor de Lipoproteína de Baixa Densidade/genética , Proteína-5 Relacionada a Receptor de Lipoproteína de Baixa Densidade/metabolismo , Proteína-6 Relacionada a Receptor de Lipoproteína de Baixa Densidade/genética , Proteína-6 Relacionada a Receptor de Lipoproteína de Baixa Densidade/metabolismo , Masculino , MicroRNAs/genética , MicroRNAs/metabolismo , Organoides/patologia , Receptores Notch/metabolismo
12.
Gastroenterology ; 158(5): 1433-1449.e27, 2020 04.
Artigo em Inglês | MEDLINE | ID: mdl-31786131

RESUMO

BACKGROUND & AIMS: Prolactin (PRL) signaling is up-regulated in hormone-responsive cancers. The PRL receptor (PRLR) is a class I cytokine receptor that signals via the Janus kinase (JAK)-signal transducer and activator of transcription and mitogen-activated protein kinase pathways to regulate cell proliferation, migration, stem cell features, and apoptosis. Patients with pancreatic ductal adenocarcinoma (PDAC) have high plasma levels of PRL. We investigated whether PRLR signaling contributes to the growth of pancreatic tumors in mice. METHODS: We used immunohistochemical analyses to compare levels of PRL and PRLR in multitumor tissue microarrays. We used structure-based virtual screening and fragment-based drug discovery to identify compounds likely to bind PRLR and interfere with its signaling. Human pancreatic cell lines (AsPC-1, BxPC-3, Panc-1, and MiaPaCa-2), with or without knockdown of PRLR (clustered regularly interspaced short palindromic repeats or small hairpin RNA), were incubated with PRL or penfluridol and analyzed in proliferation and spheroid formation. C57BL/6 mice were given injections of UNKC-6141 cells, with or without knockdown of PRLR, into pancreas, and tumor development was monitored for 4 weeks, with some mice receiving penfluridol treatment for 21 days. Human pancreatic tumor tissues were implanted into interscapular fat pads of NSG mice, and mice were given injections of penfluridol daily for 28 days. Nude mice were given injections of Panc-1 cells, xenograft tumors were grown for 2 weeks, and mice were then given intraperitoneal penfluridol for 35 days. Tumors were collected from mice and analyzed by histology, immunohistochemistry, and immunoblots. RESULTS: Levels of PRLR were increased in PDAC compared with nontumor pancreatic tissues. Incubation of pancreatic cell lines with PRL activated signaling via JAK2-signal transducer and activator of transcription 3 and extracellular signal-regulated kinase, as well as formation of pancospheres and cell migration; these activities were not observed in cells with PRLR knockdown. Pancreatic cancer cells with PRLR knockdown formed significantly smaller tumors in mice. We identified several diphenylbutylpiperidine-class antipsychotic drugs as agents that decreased PRL-induced JAK2 signaling; incubation of pancreatic cancer cells with these compounds reduced their proliferation and formation of panco spheres. Injections of 1 of these compounds, penfluridol, slowed the growth of xenograft tumors in the different mouse models, reducing proliferation and inducing autophagy of the tumor cells. CONCLUSIONS: Levels of PRLR are increased in PDAC, and exposure to PRL increases proliferation and migration of pancreatic cancer cells. Antipsychotic drugs, such as penfluridol, block PRL signaling in pancreatic cancer cells to reduce their proliferation, induce autophagy, and slow the growth of xenograft tumors in mice. These drugs might be tested in patients with PDAC.


Assuntos
Antipsicóticos/farmacologia , Carcinoma Ductal Pancreático/tratamento farmacológico , Neoplasias Pancreáticas/tratamento farmacológico , Penfluridol/farmacologia , Prolactina/metabolismo , Receptores da Prolactina/antagonistas & inibidores , Animais , Antipsicóticos/uso terapêutico , Autofagia/efeitos dos fármacos , Carcinoma Ductal Pancreático/sangue , Carcinoma Ductal Pancreático/patologia , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Descoberta de Drogas , Técnicas de Silenciamento de Genes , Humanos , Injeções Intraperitoneais , Janus Quinase 2/metabolismo , Masculino , Camundongos , Pâncreas/patologia , Neoplasias Pancreáticas/sangue , Neoplasias Pancreáticas/patologia , Penfluridol/uso terapêutico , Prolactina/sangue , Receptores da Prolactina/genética , Receptores da Prolactina/metabolismo , Fator de Transcrição STAT3/metabolismo , Transdução de Sinais/efeitos dos fármacos , Esferoides Celulares , Análise Serial de Tecidos , Ensaios Antitumorais Modelo de Xenoenxerto
13.
Oncotarget ; 10(54): 5576-5591, 2019 Sep 24.
Artigo em Inglês | MEDLINE | ID: mdl-31608135

RESUMO

Given that colon cancer is the third most common cancer in incidence and cause of death in the United States, and current treatment modalities are insufficient, there is a need to develop novel agents. Towards this, here we focus on γ-Mangostin, a bioactive compound present in the Mangosteen (Garcinia mangostana) fruit. γ-Mangostin suppressed proliferation and colony formation, and induced cell cycle arrest and apoptosis of colon cancer cell lines. Further, γ-Mangostin inhibited colonosphere formation. Molecular docking and CETSA (Cellular thermal shift assay) binding assays demonstrated that γ-Mangostin interacts with transcription factor TCF4 (T-Cell Factor 4) at the ß-catenin binding domain with the binding energy of -5.5 Kcal/mol. Moreover, γ-Mangostin treatment decreased TCF4 expression and reduced TCF reporter activity. The compound also suppressed the expression of Wnt signaling target proteins cyclin D1 and c-Myc, and stem cell markers such as LGR5, DCLK1 and CD44. To determine the effect of γ-Mangostin on tumor growth in vivo, we administered nude mice harboring HCT116 tumor xenografts with 5 mg/Kg of γ-Mangostin intraperitoneally for 21 days. γ-Mangostin treatment significantly suppressed tumor growth, with notably lowered tumor volume and weight. In addition, western blot analysis revealed a significant decrease in the expression of TCF4 and its downstream targets such as cyclin D1 and c-Myc. Together, these data suggest that γ-Mangostin inhibits colon cancer growth through targeting TCF4. γ-Mangostin may be a potential therapeutic agent for colon cancer.

14.
Gastroenterology ; 157(6): 1646-1659.e11, 2019 12.
Artigo em Inglês | MEDLINE | ID: mdl-31442435

RESUMO

BACKGROUND & AIMS: The histone lysine demethylase 3A (KDM3A) demethylates H3K9me1 and H3K9Me2 to increase gene transcription and is upregulated in tumors, including pancreatic tumors. We investigated its activities in pancreatic cancer cell lines and its regulation of the gene encoding doublecortin calmodulin-like kinase 1 (DCLK1), a marker of cancer stem cells. METHODS: We knocked down KDM3A in MiaPaCa-2 and S2-007 pancreatic cancer cell lines and overexpressed KDM3A in HPNE cells (human noncancerous pancreatic ductal cell line); we evaluated cell migration, invasion, and spheroid formation under hypoxic and normoxic conditions. Nude mice were given orthotopic injections of S2-007 cells, with or without (control) knockdown of KDM3A, and HPNE cells, with or without (control) overexpression of KDM3A; tumor growth was assessed. We analyzed pancreatic tumor tissues from mice and pancreatic cancer cell lines by immunohistochemistry and immunoblotting. We performed RNA-sequencing analysis of MiaPaCa-2 and S2-007 cells with knockdown of KDM3A and evaluated localization of DCLK1 and KDM3A by immunofluorescence. We analyzed the cancer genome atlas for levels of KDM3A and DCLK1 messenger RNA in human pancreatic ductal adenocarcinoma (PDAC) tissues and association with patient survival time. RESULTS: Levels of KDM3A were increased in human pancreatic tumor tissues and cell lines, compared with adjacent nontumor pancreatic tissues, such as islet and acinar cells. Knockdown of KDM3A in S2-007 cells significantly reduced colony formation, invasion, migration, and spheroid formation, compared with control cells, and slowed growth of orthotopic tumors in mice. We identified KDM3A-binding sites in the DCLK1 promoter; S2-007 cells with knockdown of KDM3A had reduced levels of DCLK1. HPNE cells that overexpressed KDM3A formed foci and spheres in culture and formed tumors and metastases in mice, whereas control HPNE cells did not. Hypoxia induced sphere formation and increased levels of KDM3A in S2-007 cells and in HPNE cells that overexpressed DCLK1, but not control HPNE cells. Levels of KDM3A and DCLK1 messenger RNA were higher in human PDAC than nontumor pancreatic tissues and correlated with shorter survival times of patients. CONCLUSIONS: We found human PDAC samples and pancreatic cancer cell lines to overexpress KDM3A. KDM3A increases expression of DCLK1, and levels of both proteins are increased in human PDAC samples. Knockdown of KDM3A in pancreatic cancer cell lines reduced their invasive and sphere-forming activities in culture and formation of orthotopic tumors in mice. Hypoxia increased expression of KDM3A in pancreatic cancer cells. Strategies to disrupt this pathway might be developed for treatment of pancreatic cancer.


Assuntos
Carcinogênese/genética , Carcinoma Ductal Pancreático/genética , Regulação Neoplásica da Expressão Gênica , Peptídeos e Proteínas de Sinalização Intracelular/genética , Histona Desmetilases com o Domínio Jumonji/metabolismo , Neoplasias Pancreáticas/genética , Proteínas Serina-Treonina Quinases/genética , Animais , Carcinoma Ductal Pancreático/mortalidade , Carcinoma Ductal Pancreático/patologia , Linhagem Celular Tumoral , Metilação de DNA , Conjuntos de Dados como Assunto , Quinases Semelhantes a Duplacortina , Feminino , Técnicas de Silenciamento de Genes , Humanos , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Histona Desmetilases com o Domínio Jumonji/genética , Masculino , Camundongos , Pessoa de Meia-Idade , Neoplasias Pancreáticas/mortalidade , Neoplasias Pancreáticas/patologia , Regiões Promotoras Genéticas/genética , Proteínas Serina-Treonina Quinases/metabolismo , Análise de Sobrevida , Regulação para Cima , Ensaios Antitumorais Modelo de Xenoenxerto
15.
Mol Carcinog ; 58(8): 1400-1409, 2019 08.
Artigo em Inglês | MEDLINE | ID: mdl-31020708

RESUMO

We previously reported that ionizing radiation (IR) mediates cell death through the induction of CUGBP elav-like family member 2 (CELF2), a tumor suppressor. CELF2 is an RNA binding protein that modulates mRNA stability and translation. Since IR induces autophagy, we hypothesized that CELF2 regulates autophagy-mediated colorectal cancer (CRC) cell death. For clinical relevance, we determined CELF2 levels in The Cancer Genome Atlas (TCGA). Role of CELF2 in radiation response was carried out in CRC cell lines by immunoblotting, immunofluorescence, autophagic vacuole analyses, RNA stability assay, quantitative polymerase chain reaction and electron microscopy. In vivo studies were performed in a xenograft tumor model. TCGA analyses demonstrated that compared to normal tissue, CELF2 is expressed at significantly lower levels in CRC, and is associated with better overall 5-year survival in patients receiving radiation. Mechanistically, CELF2 increased levels of critical components of the autophagy cascade including Beclin-1, ATG5, and ATG12 by modulating mRNA stability. CELF2 also increased autophagic flux in CRC. IR significantly induced autophagy in CRC which correlates with increased levels of CELF2 and autophagy associated proteins. Silencing CELF2 with siRNA, mitigated IR induced autophagy. Moreover, knockdown of CELF2 in vivo conferred tumor resistance to IR. These studies elucidate an unrecognized role for CELF2 in inducing autophagy and potentiating the effects of radiotherapy in CRC.


Assuntos
Autofagia/fisiologia , Proteínas CELF/metabolismo , Sobrevivência Celular/efeitos da radiação , Neoplasias Colorretais/patologia , Neoplasias Colorretais/radioterapia , Proteínas do Tecido Nervoso/metabolismo , Animais , Proteína 12 Relacionada à Autofagia/metabolismo , Proteína 5 Relacionada à Autofagia/metabolismo , Proteína Beclina-1/metabolismo , Proteínas CELF/genética , Linhagem Celular Tumoral , Sobrevivência Celular/genética , Células HCT116 , Humanos , Masculino , Camundongos , Transplante de Neoplasias , Proteínas do Tecido Nervoso/genética , Prognóstico , Interferência de RNA , RNA Mensageiro/genética , RNA Interferente Pequeno/genética , Radiação Ionizante , Transplante Heterólogo
16.
Oncotarget ; 10(16): 1554-1571, 2019 Feb 22.
Artigo em Inglês | MEDLINE | ID: mdl-30899425

RESUMO

Super-enhancers (SEs) are unique areas of the genome which drive high-level of transcription and play a pivotal role in the cell physiology. Previous studies have established several important genes in cancer as SE-driven oncogenes. It is likely that oncogenes may hack the resident tissue regenerative program and interfere with SE-driven repair networks, leading to the specific pancreatic ductal adenocarcinoma (PDAC) phenotype. Here, we used ChIP-Seq to identify the presence of SE in PDAC cell lines. Differential H3K27AC marks were identified at enhancer regions of genes including c-MYC, MED1, OCT-4, NANOG, and SOX2 that can act as SE in non-cancerous, cancerous and metastatic PDAC cell lines. GZ17-6.02 affects acetylation of the genes, reduces transcription of major transcription factors, sonic hedgehog pathway proteins, and stem cell markers. In accordance with the decrease in Oct-4 expression, ChIP-Seq revealed a significant decrease in the occupancy of OCT-4 in the entire genome after GZ17-6.02 treatment suggesting the possible inhibitory effect of GZ17-6.02 on PDAC. Hence, SE genes are associated with PDAC and targeting their regulation with GZ17-6.02 offers a novel approach for treatment.

17.
Cancer Res ; 79(7): 1681-1695, 2019 04 01.
Artigo em Inglês | MEDLINE | ID: mdl-30674533

RESUMO

Metastasis is a major cause of cancer-related deaths. A dearth of preclinical models that recapitulate the metastatic microenvironment has impeded the development of therapeutic agents that are effective against metastatic disease. Because the majority of solid tumors metastasize to the lung, we developed a multicellular lung organoid that mimics the lung microenvironment with air sac-like structures and production of lung surfactant protein. We used these cultures, called primitive lung-in-a-dish (PLiD), to recreate metastatic disease using primary and established cancer cells. The metastatic tumor-in-a-dish (mTiD) cultures resemble the architecture of metastatic tumors in the lung, including angiogenesis. Pretreating PLiD with tumor exosomes enhanced cancer cell colonization. We next tested the response of primary and established cancer cells to current chemotherapeutic agents and an anti-VEGF antibody in mTiD against cancer cells in two-dimensional (2D) or 3D cultures. The response of primary patient-derived colon and ovarian tumor cells to therapy in mTiD cultures matched the response of the patient in the clinic, but not in 2D or single-cell-type 3D cultures. The sensitive mTiD cultures also produced significantly lower circulating markers for cancer similar to that seen in patients who responded to therapy. Thus, we have developed a novel method for lung colonization in vitro, a final stage in tumor metastasis. Moreover, the technique has significant utility in precision/personalized medicine, wherein this phenotypic screen can be coupled with current DNA pharmacogenetics to identify the ideal therapeutic agent, thereby increasing the probability of response to treatment while reducing unnecessary side effects. SIGNIFICANCE: A lung organoid that exhibits characteristics of a normal human lung is developed to study the biology of metastatic disease and therapeutic intervention.


Assuntos
Neoplasias Pulmonares/secundário , Organoides/patologia , Animais , Antineoplásicos/uso terapêutico , Linhagem Celular , Linhagem Celular Tumoral , Ensaios de Seleção de Medicamentos Antitumorais , Feminino , Humanos , Neoplasias Pulmonares/tratamento farmacológico , Metástase Neoplásica , Neoplasias Ovarianas/tratamento farmacológico , Neoplasias Ovarianas/patologia , Microambiente Tumoral
18.
Leukemia ; 33(7): 1675-1686, 2019 07.
Artigo em Inglês | MEDLINE | ID: mdl-30664664

RESUMO

p97 is an ATPase that works in concert with histone deacetylase 6 (HDAC6), to facilitate the degradation of misfolded proteins by autophagosomes. p97 has also been implicated in DNA repair and maintaining genomic stability. In this study, we determined the effect of combined inhibition of p97 and HDAC6 activities in mantle cell lymphoma (MCL) cells. We report that treatment with p97 inhibitors induces dose-dependent apoptosis in MCL cells. The p97 inhibitor CB-5083 induces ER stress markers GRP78 and CHOP and results in the accumulation of polyubiquitylated proteins. Co-treatment with CB-5083 and the HDAC6 inhibitor ACY-1215 result in marked downregulation of CDK4, Cyclin D1, and BRCA1 levels without inhibiting autophagic flux. Consequently, treatment with CB-5083 accentuates DNA damage in response to treatment with ACY-1215 resulting in enhanced accumulation of H2AX-γ and synergistic apoptosis. Furthermore, ATM loss severely impairs phosphorylation of 53BP1 following co-treatment with CB-5083 and ACY-1215 in response to gamma irradiation. Finally, co-treatment CB-5083 and ACY-1215 results in reduced tumor volumes and improves survival in Z138C and Jeko-1 xenografts in NSG mice. These observations suggest that combined inhibition of p97 and HDAC6 abrogates resolution of proteotoxic stress and impairs DNA repair mechanisms in MCL cells.


Assuntos
Adenosina Trifosfatases/antagonistas & inibidores , Reparo do DNA/efeitos dos fármacos , Sinergismo Farmacológico , Desacetilase 6 de Histona/antagonistas & inibidores , Ácidos Hidroxâmicos/farmacologia , Indóis/farmacologia , Linfoma de Célula do Manto/tratamento farmacológico , Linfoma de Célula do Manto/genética , Proteínas Nucleares/antagonistas & inibidores , Pirimidinas/farmacologia , Animais , Apoptose , Autofagia , Proliferação de Células , Dano ao DNA/efeitos dos fármacos , Quimioterapia Combinada , Chaperona BiP do Retículo Endoplasmático , Regulação Neoplásica da Expressão Gênica , Inibidores de Histona Desacetilases/farmacologia , Humanos , Linfoma de Célula do Manto/patologia , Camundongos , Camundongos Endogâmicos NOD , Camundongos SCID , Células Tumorais Cultivadas , Ensaios Antitumorais Modelo de Xenoenxerto
19.
ChemMedChem ; 13(11): 1115-1123, 2018 06 06.
Artigo em Inglês | MEDLINE | ID: mdl-29570947

RESUMO

A series of readily available curcuminoids with a halogenated bis(4-methoxy/4,5-dimethoxybenzylidene)-4-piperidone structure were prepared and analyzed for their cytotoxic impact on eight human cancer cell lines of five different entities. The known 3,4,5-trimethoxybenzylidene curcuminoid 2 a and the new bis-(3-bromophenyl) and bis-(3,5-dibromophenyl) derivatives 3 c and 3 d proved to be more strongly antiproliferative than the known curcuminoid EF24 against six of these cell lines. Compounds 2 a and 3 c caused a distinct increase of reactive oxygen species, which eventually elicited apoptosis in 518A2 melanoma cells. Compound 2 a arrested 518A2 melanoma cells in G1 phase of the cell cycle and had no effect on the expression of pro-metastatic matrix metalloproteinases MMP-2 and MMP-9, whereas 3 c led to an accumulation of 518A2 cells in the G2 /M phase and to a downregulation of MMP-2 expression. In addition, treatment with 2 a and 3 c resulted in significant inhibition of colony formation in HCT116 cells. Both 2 a and 3 c showed antiangiogenic activity, for example, by inhibiting the formation of sub-intestinal veins (SIV) in zebrafish embryos. Compound 3 c was also well tolerated by mice and inhibited the growth of HCT116 colon cancer xenografts.


Assuntos
Antineoplásicos/farmacologia , Curcumina/análogos & derivados , Curcumina/farmacologia , Piperidonas/farmacologia , Inibidores da Angiogênese/síntese química , Inibidores da Angiogênese/química , Inibidores da Angiogênese/farmacologia , Animais , Antineoplásicos/síntese química , Antineoplásicos/química , Apoptose/efeitos dos fármacos , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Curcumina/síntese química , Pontos de Checagem da Fase G1 do Ciclo Celular/efeitos dos fármacos , Halogenação , Humanos , Masculino , Camundongos Nus , Estrutura Molecular , Piperidonas/síntese química , Piperidonas/química , Espécies Reativas de Oxigênio/metabolismo , Relação Estrutura-Atividade , Ensaios Antitumorais Modelo de Xenoenxerto , Peixe-Zebra
20.
Curr Med Chem ; 25(22): 2585-2594, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-28137215

RESUMO

Pancreatic ductal adenocarcinoma is one of the deadliest cancers worldwide and the fourth leading cause of cancer-related deaths in United States. Regardless of the advances in molecular pathogenesis and consequential efforts to suppress the disease, this cancer remains a major health problem in United States. By 2030, the projection is that pancreatic cancer will be climb up to be the second leading cause of cancer-related deaths in the United States. Pancreatic cancer is a rapidly invasive and highly metastatic cancer, and does not respond to standard therapies. Emerging evidence supports that the presence of a unique population of cells called cancer stem cells (CSCs) as potential cancer inducing cells and efforts are underway to develop therapeutic strategies targeting these cells. CSCs are rare quiescent cells, and with the capacity to self-renew through asymmetric/symmetric cell division, as well as differentiate into various lineages of cells in the cancer. Studies have been shown that CSCs are highly resistant to standard therapy and also responsible for drug resistance, cancer recurrence and metastasis. To overcome this problem, we need novel preventive agents that target these CSCs. Natural compounds or phytochemicals have ability to target these CSCs and their signaling pathways. Therefore, in the present review article, we summarize our current understanding of pancreatic CSCs and their signaling pathways, and the phytochemicals that target these cells including curcumin, resveratrol, tea polyphenol EGCG (epigallocatechin- 3-gallate), crocetinic acid, sulforaphane, genistein, indole-3-carbinol, vitamin E δ- tocotrienol, Plumbagin, quercetin, triptolide, Licofelene and Quinomycin. These natural compounds or phytochemicals, which inhibit cancer stem cells may prove to be promising agents for the prevention and treatment of pancreatic cancers.


Assuntos
Neoplasias Pancreáticas/prevenção & controle , Compostos Fitoquímicos/uso terapêutico , Catequina/análogos & derivados , Catequina/farmacologia , Catequina/uso terapêutico , Quinases Semelhantes a Duplacortina , Proteínas Hedgehog/antagonistas & inibidores , Proteínas Hedgehog/metabolismo , Humanos , Peptídeos e Proteínas de Sinalização Intracelular/antagonistas & inibidores , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Proteínas Quinases Ativadas por Mitógeno/antagonistas & inibidores , Proteínas Quinases Ativadas por Mitógeno/metabolismo , Células-Tronco Neoplásicas/citologia , Células-Tronco Neoplásicas/efeitos dos fármacos , Células-Tronco Neoplásicas/metabolismo , Neoplasias Pancreáticas/metabolismo , Neoplasias Pancreáticas/patologia , Compostos Fitoquímicos/farmacologia , Proteínas Serina-Treonina Quinases/antagonistas & inibidores , Proteínas Serina-Treonina Quinases/metabolismo , Transdução de Sinais/efeitos dos fármacos , Proteínas Wnt/antagonistas & inibidores , Proteínas Wnt/metabolismo
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