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1.
bioRxiv ; 2023 Sep 17.
Artigo em Inglês | MEDLINE | ID: mdl-37745587

RESUMO

Breast cancer is categorized by the molecular and histologic presentation of the tumor, with the major histologic subtypes being No Special Type (NST) and Invasive Lobular Carcinoma (ILC). ILC are characterized by growth in a single file discohesive manner with stromal infiltration attributed to their hallmark pathognomonic loss of E-cadherin ( CDH1 ). Few ILC cell line models are available to researchers. Here we report the successful establishment and characterization of a novel ILC cell line, WCRC-25, from a metastatic pleural effusion from a postmenopausal Caucasian woman with metastatic ILC. WCRC-25 is an ER-negative luminal epithelial ILC cell line with both luminal and Her2-like features. It exhibits anchorage independent growth and haptotactic migration towards Collagen I. Sequencing revealed a CDH1 Q706* truncating mutation, together with mutations in FOXA1, CTCF, BRCA2 and TP53 , which were also seen in a series of metastatic lesions from the patient. Copy number analyses revealed amplification and deletion of genes frequently altered in ILC while optical genome mapping revealed novel structural rearrangements. RNA-seq analysis comparing the primary tumor, metastases and the cell line revealed signatures for cell cycle progression and receptor tyrosine kinase signaling. To assess targetability, we treated WCRC-25 with AZD5363 and Alpelisib confirming WCRC-25 as susceptible to PI3K/AKT signaling inhibition as predicted by our RNA sequencing analysis. In conclusion, we report WCRC-25 as a novel ILC cell line with promise as a valuable research tool to advance our understanding of ILC and its therapeutic vulnerabilities. Financial support: The work was in part supported by a Susan G Komen Leadership Grant to SO (SAC160073) and NCI R01 CA252378 (SO/AVL). AVL and SO are Komen Scholars, Hillman Foundation Fellows and supported by BCRF. This project used the UPMC Hillman Cancer Center and Tissue and Research Pathology/Pitt Biospecimen Core shared resource which is supported in part by award P30CA047904. This research was also supported in part by the University of Pittsburgh Center for Research Computing, RRID:SCR_022735, through the resources provided. Specifically, this work used the HTC cluster, which is supported by NIH award number S10OD028483. Finally, partial support was provided by the Magee-Womens Research Institute and Foundation, The Shear Family Foundation, and The Metastatic Breast Cancer Network.

2.
bioRxiv ; 2023 Aug 14.
Artigo em Inglês | MEDLINE | ID: mdl-37645906

RESUMO

Nitro fatty acids (NO 2 -FAs) are endogenously generated lipid signaling mediators from metabolic and inflammatory reactions between conjugated diene fatty acids and nitric oxide or nitrite-derived reactive species. NO 2 -FAs undergo reversible Michael addition with hyperreactive protein cysteine thiolates to induce posttranslational protein modifications that can impact protein function. Herein, we report a novel mechanism of action of natural and non-natural nitroalkenes structurally similar to ( E ) 10-nitro-octadec-9-enoic acid (CP-6), recently de-risked by preclinical Investigational New Drug-enabling studies and Phase 1 and Phase 2 clinical trials and found to induce DNA damage in a TNBC xenograft by inhibiting homologous-recombination (HR)-mediated repair of DNA double-strand breaks (DSB). CP-6 specifically targets Cys319, essential in RAD51-controlled HR-mediated DNA DSB repair in cells. A nitroalkene library screen identified two structurally different nitroalkenes, a non-natural fatty acid [( E ) 8-nitro- nonadec-7-enoic acid (CP-8)] and a dicarboxylate ester [dimethyl ( E )nitro-oct-4-enedioate (CP- 23)] superior to CP-6 in TNBC cells killing, synergism with three different inhibitors of the poly ADP-ribose polymerase (PARP) and γ-IR. CP-8 and CP-23 effectively inhibited γ-IR-induced RAD51 foci formation and HR in a GFP-reported assay but did not affect benign human epithelial cells or cell cycle phases. In vivo, CP-8 and CP-23's efficacies diverged as only CP-8 showed promising anticancer activities alone and combined with the PARP inhibitor talazoparib in an HR-proficient TNBC mouse model. As preliminary preclinical toxicology analysis also suggests CP-8 as safe, our data endorse CP-8 as a novel anticancer molecule for treating cancers sensitive to homologous recombination-mediated DNA repair inhibitors.

3.
Redox Biol ; 66: 102856, 2023 10.
Artigo em Inglês | MEDLINE | ID: mdl-37633047

RESUMO

Nitro fatty acids (NO2-FAs) are endogenously generated lipid signaling mediators from metabolic and inflammatory reactions between conjugated diene fatty acids and nitric oxide or nitrite-derived reactive species. NO2-FAs undergo reversible Michael addition with hyperreactive protein cysteine thiolates to induce posttranslational protein modifications that can impact protein function. Herein, we report a novel mechanism of action of natural and non-natural nitroalkenes structurally similar to (E) 10-nitro-octadec-9-enoic acid (CP-6), recently de-risked by preclinical Investigational New Drug-enabling studies and Phase 1 and Phase 2 clinical trials and found to induce DNA damage in a TNBC xenograft by inhibiting homologous-recombination (HR)-mediated repair of DNA double-strand breaks (DSB). CP-6 specifically targets Cys319, essential in RAD51-controlled HR-mediated DNA DSB repair in cells. A nitroalkene library screen identified two structurally different nitroalkenes, a non-natural fatty acid [(E) 8-nitro-nonadec-7-enoic acid (CP-8)] and a dicarboxylate ester [dimethyl (E)nitro-oct-4-enedioate (CP-23)] superior to CP-6 in TNBC cells killing, synergism with three different inhibitors of the poly ADP-ribose polymerase (PARP) and γ-IR. CP-8 and CP-23 effectively inhibited γ-IR-induced RAD51 foci formation and HR in a GFP-reported assay but did not affect benign human epithelial cells or cell cycle phases. In vivo, CP-8 and CP-23's efficacies diverged as only CP-8 showed promising anticancer activities alone and combined with the PARP inhibitor talazoparib in an HR-proficient TNBC mouse model. As preliminary preclinical toxicology analysis also suggests CP-8 as safe, our data endorse CP-8 as a novel anticancer molecule for treating cancers sensitive to homologous recombination-mediated DNA repair inhibitors.


Assuntos
Neoplasias de Mama Triplo Negativas , Animais , Camundongos , Humanos , Neoplasias de Mama Triplo Negativas/tratamento farmacológico , Neoplasias de Mama Triplo Negativas/genética , Dióxido de Nitrogênio , Recombinação Homóloga , Apoptose , Alcenos , DNA , Rad51 Recombinase
4.
Redox Biol ; 56: 102443, 2022 10.
Artigo em Inglês | MEDLINE | ID: mdl-36058112

RESUMO

RAD51 is a critical recombinase that functions in concert with auxiliary mediator proteins to direct the homologous recombination (HR) DNA repair pathway. We show that Cys319 RAD51 possesses nucleophilic characteristics and is important for irradiation-induced RAD51 foci formation and resistance to inhibitors of poly (ADP-ribose) polymerase (PARP). We have previously identified that cysteine (Cys) oxidation of proteins can be important for activity and modulated via binding to peroxiredoxin 1 (PRDX1). PRDX1 reduces peroxides and coordinates the signaling actions of protein binding partners. Loss of PRDX1 inhibits irradiation-induced RAD51 foci formation and represses HR DNA repair. PRDX1-deficient human breast cancer cells and mouse embryonic fibroblasts display disrupted RAD51 foci formation and decreased HR, resulting in increased DNA damage and sensitization of cells to irradiation. Following irradiation cells deficient in PRDX1 had increased incorporation of the sulfenylation probe DAz-2 in RAD51 Cys319, a functionally-significant, thiol that PRDX1 is critical for maintaining in a reduced state. Molecular dynamics (MD) simulations of dT-DNA bound to a non-oxidized RAD51 protein showed tight binding throughout the simulation, while dT-DNA dissociated from an oxidized Cys319 RAD51 filament. These novel data establish RAD51 Cys319 as a functionally-significant site for the redox regulation of HR and cellular responses to IR.


Assuntos
Inibidores de Poli(ADP-Ribose) Polimerases , Rad51 Recombinase , Difosfato de Adenosina/metabolismo , Animais , Cisteína/metabolismo , DNA/metabolismo , Reparo do DNA , Fibroblastos/metabolismo , Recombinação Homóloga , Humanos , Camundongos , Oxirredução , Peróxidos , Peroxirredoxinas/genética , Peroxirredoxinas/metabolismo , Inibidores de Poli(ADP-Ribose) Polimerases/farmacologia , Poli(ADP-Ribose) Polimerases/genética , Rad51 Recombinase/genética , Rad51 Recombinase/metabolismo , Ribose
5.
Br J Cancer ; 125(8): 1146-1157, 2021 10.
Artigo em Inglês | MEDLINE | ID: mdl-34389806

RESUMO

BACKGROUND: Peroxiredoxin 1 (PRDX1) belongs to an abundant family of peroxidases whose role in cancer is still unresolved. While mouse knockout studies demonstrate a tumour suppressive role for PRDX1, in cancer cell xenografts, results denote PRDX1 as a drug target. Probably, this phenotypic discrepancy stems from distinct roles of PRDX1 in certain cell types or stages of tumour progression. METHODS: We demonstrate an important cell-autonomous function for PRDX1 utilising a syngeneic mouse model (BALB/c) and mammary fibroblasts (MFs) obtained from it. RESULTS: Loss of PRDX1 in vivo promotes collagen remodelling known to promote breast cancer progression. PRDX1 inactivation in MFs occurs via SRC-induced phosphorylation of PRDX1 TYR194 and not through the expected direct oxidation of CYS52 in PRDX1 by ROS. TYR194-phosphorylated PRDX1 fails to bind to lysyl oxidases (LOX) and leads to the accumulation of extracellular LOX proteins which supports enhanced collagen remodelling associated with breast cancer progression. CONCLUSIONS: This study reveals a cell type-specific tumour suppressive role for PRDX1 that is supported by survival analyses, depending on PRDX1 protein levels in breast cancer cohorts.


Assuntos
Neoplasias da Mama/patologia , Matriz Extracelular/metabolismo , Peroxirredoxinas/metabolismo , Proteína-Lisina 6-Oxidase/metabolismo , Tirosina/metabolismo , Animais , Neoplasias da Mama/metabolismo , Linhagem Celular Tumoral , Colágeno/metabolismo , Feminino , Regulação Neoplásica da Expressão Gênica , Células HEK293 , Humanos , Camundongos , Fosforilação , Prognóstico , Análise de Sobrevida
6.
Mol Cancer Ther ; 20(6): 1210-1219, 2021 06.
Artigo em Inglês | MEDLINE | ID: mdl-33785649

RESUMO

Disease models, including in vitro cell culture and animal models, have contributed significantly to developing diagnostics and treatments over the past several decades. The successes of traditional drug screening methods were generally hampered by not adequately mimicking critical in vivo features, such as a 3D microenvironment and dynamic drug diffusion through the extracellular matrix (ECM). To address these issues, we developed a 3D dynamic drug delivery system for cancer drug screening that mimicks drug dissemination through the tumor vasculature and the ECM by creating collagen-embedded microfluidic channels. Using this novel 3D ECM microsystem, we compared viability of tumor pieces with traditionally used 2D methods in response to three different drug combinations. Drug diffusion profiles were evaluated by simulation methods and tested in the 3D ECM microsystem and a 2D 96-well setup. Compared with the 2D control, the 3D ECM microsystem produced reliable data on viability, drug ratios, and combination indeces. This novel approach enables higher throughput and sets the stage for future applications utilizing drug sensitivity predicting algorithms based on dynamic diffusion profiles requiring only minimal patient tissue. Our findings moved drug sensitivity screening closer to clinical implications with a focus on testing combinatorial drug effects, an option often limited by the amount of available patient tissues.


Assuntos
Antineoplásicos/uso terapêutico , Avaliação Pré-Clínica de Medicamentos/métodos , Imageamento Tridimensional/métodos , Dispositivos Lab-On-A-Chip/normas , Animais , Modelos Animais de Doenças , Matriz Extracelular , Feminino , Humanos , Camundongos , Camundongos Nus
7.
BMC Cancer ; 19(1): 812, 2019 Aug 16.
Artigo em Inglês | MEDLINE | ID: mdl-31419957

RESUMO

BACKGROUND: Reactive oxygen species (ROS), including hydrogen peroxide, drive differentiation of normal fibroblasts into activated fibroblasts, which can generate high amounts of hydrogen peroxide themselves, thereby increasing oxidative stress in the microenvironment. This way, activated fibroblasts can transition into cancer-associated fibroblasts (CAFs). METHODS: Mammary fibroblasts from either female 8 weeks old PRDX1 knockout and wildtype mice or Balb/c mice were studied for characteristic protein expression using immunofluorescence and immunoblotting. Cancer-associated fibroblasts was examined by transwell migration and invasion assays. The binding of PRDX1 to JNK1 was assessed by co-immuneprecipitation and JNK regulation of CAF phenotypes was examined using the JNK inhibitor SP600125. Extracellular hydrogen peroxide levels were measured by chemiluminescence via the reaction between hypochlorite and luminol. Statistical analyses were done using Students t-test. RESULTS: We show here PRDX1 activity as an essential switch in regulating the activated phenotype as loss of PRDX1 results in the development of a CAF-like phenotype in mammary fibroblasts. We also show that PRDX1 regulates JNK kinase signaling thereby inhibiting CAF-like markers and CAF invasion. Inhibition of JNK activity reduced these behaviors. CONCLUSIONS: These data suggest that PRDX1 repressed the activated phenotype of fibroblasts in part through JNK inhibition which may present a novel therapeutic option for CAF-enriched cancers such as breast cancer.


Assuntos
Fibroblastos/metabolismo , Glândulas Mamárias Animais/citologia , Proteína Quinase 8 Ativada por Mitógeno/metabolismo , Peroxirredoxinas/genética , Peroxirredoxinas/metabolismo , Fenótipo , Actinas/metabolismo , Animais , Antracenos/farmacologia , Feminino , Técnicas de Inativação de Genes , Células HEK293 , Humanos , Peróxido de Hidrogênio/metabolismo , Camundongos , Camundongos Endogâmicos BALB C , Proteína Quinase 8 Ativada por Mitógeno/antagonistas & inibidores , Estresse Oxidativo , Espécies Reativas de Oxigênio/metabolismo , Transfecção , Microambiente Tumoral
8.
Redox Biol ; 21: 101104, 2019 02.
Artigo em Inglês | MEDLINE | ID: mdl-30690320

RESUMO

Transcription factors control the rate of transcription of genetic information from DNA to messenger RNA, by binding specific DNA sequences in promoter regions. Transcriptional gene control is a rate-limiting process that is tightly regulated and based on transient environmental signals which are translated into long-term changes in gene transcription. Post-translational modifications (PTMs) on transcription factors by phosphorylation or acetylation have profound effects not only on sub-cellular localization but also on substrate specificity through changes in DNA binding capacity. During times of cellular stress, specific transcription factors are in place to help protect the cell from damage by initiating the transcription of antioxidant response genes. Here we discuss PTMs caused by reactive oxygen species (ROS), such as H2O2, that can expeditiously regulate the activation of transcription factors involved in the oxidative stress response. Part of this rapid regulation are proteins involved in H2O2-related reduction and oxidation (redox) reactions such as redoxins, H2O2 scavengers described to interact with transcription factors. Redoxins have highly reactive cysteines of rate constants around 6-10-1 s-1 that engage in nucleophilic substitution of a thiol-disulfide with another thiol in inter-disulfide exchange reactions. We propose here that H2O2 signal transduction induced inter-disulfide exchange reactions between redoxin cysteines and cysteine thiols of transcription factors to allow for rapid and precise on and off switching of transcription factor activity. Thus, redoxins are essential modulators of stress response pathways beyond H2O2 scavenging capacity.


Assuntos
Oxirredução , Estresse Oxidativo/genética , Transcrição Gênica , Animais , Cisteína/metabolismo , Humanos , Processamento de Proteína Pós-Traducional , Espécies Reativas de Oxigênio/metabolismo , Transdução de Sinais , Compostos de Sulfidrila/metabolismo , Fatores de Transcrição/metabolismo
9.
Antioxidants (Basel) ; 8(2)2019 Jan 23.
Artigo em Inglês | MEDLINE | ID: mdl-30678096

RESUMO

Reactive oxygen and nitrogen species have cell signaling properties and are involved in a multitude of processes beyond redox homeostasis. The peroxiredoxin (Prdx) proteins are highly sensitive intracellular peroxidases that can coordinate cell signaling via direct reactive species scavenging or by acting as a redox sensor that enables control of binding partner activity. Oxidation of the peroxidatic cysteine residue of Prdx proteins are the classical post-translational modification that has been recognized to modulate downstream signaling cascades, but increasing evidence supports that dynamic changes to phosphorylation of Prdx proteins is also an important determinant in redox signaling. Phosphorylation of Prdx proteins affects three-dimensional structure and function to coordinate cell proliferation, wound healing, cell fate and lipid signaling. The advent of large proteomic datasets has shown that there are many opportunities to understand further how phosphorylation of Prdx proteins fit into intracellular signaling cascades in normal or malignant cells and that more research is necessary. This review summarizes the Prdx family of proteins and details how post-translational modification by kinases and phosphatases controls intracellular signaling.

10.
J Biol Chem ; 294(2): 397-404, 2019 01 11.
Artigo em Inglês | MEDLINE | ID: mdl-30478172

RESUMO

Homologous recombination (HR)-directed DNA double-strand break (DSB) repair enables template-directed DNA repair to maintain genomic stability. RAD51 recombinase (RAD51) is a critical component of HR and facilitates DNA strand exchange in DSB repair. We report here that treating triple-negative breast cancer (TNBC) cells with the fatty acid nitroalkene 10-nitro-octadec-9-enoic acid (OA-NO2) in combination with the antineoplastic DNA-damaging agents doxorubicin, cisplatin, olaparib, and γ-irradiation (IR) enhances the antiproliferative effects of these agents. OA-NO2 inhibited IR-induced RAD51 foci formation and enhanced H2A histone family member X (H2AX) phosphorylation in TNBC cells. Analyses of fluorescent DSB reporter activity with both static-flow cytometry and kinetic live-cell studies enabling temporal resolution of recombination revealed that OA-NO2 inhibits HR and not nonhomologous end joining (NHEJ). OA-NO2 alkylated Cys-319 in RAD51, and this alkylation depended on the Michael acceptor properties of OA-NO2 because nonnitrated and saturated nonelectrophilic analogs of OA-NO2, octadecanoic acid and 10-nitro-octadecanoic acid, did not react with Cys-319. Of note, OA-NO2 alkylation of RAD51 inhibited its binding to ssDNA. RAD51 Cys-319 resides within the SH3-binding site of ABL proto-oncogene 1, nonreceptor tyrosine kinase (ABL1), so we investigated the effect of OA-NO2-mediated Cys-319 alkylation on ABL1 binding and found that OA-NO2 inhibits RAD51-ABL1 complex formation both in vitro and in cell-based immunoprecipitation assays. The inhibition of the RAD51-ABL1 complex also suppressed downstream RAD51 Tyr-315 phosphorylation. In conclusion, RAD51 Cys-319 is a functionally significant site for adduction of soft electrophiles such as OA-NO2 and suggests further investigation of lipid electrophile-based combinational therapies for TNBC.


Assuntos
Antineoplásicos/administração & dosagem , Dano ao DNA/efeitos dos fármacos , Ácidos Graxos/administração & dosagem , Rad51 Recombinase/metabolismo , Neoplasias de Mama Triplo Negativas/enzimologia , Neoplasias de Mama Triplo Negativas/fisiopatologia , Alquilação , Antineoplásicos/química , Proliferação de Células/efeitos dos fármacos , Cisplatino/administração & dosagem , Reparo do DNA , Doxorrubicina/administração & dosagem , Quimioterapia Combinada , Ácidos Graxos/química , Humanos , Ligação Proteica/efeitos dos fármacos , Proto-Oncogene Mas , Proteínas Proto-Oncogênicas c-abl/genética , Proteínas Proto-Oncogênicas c-abl/metabolismo , Rad51 Recombinase/química , Rad51 Recombinase/genética , Neoplasias de Mama Triplo Negativas/genética , Neoplasias de Mama Triplo Negativas/metabolismo
11.
Free Radic Biol Med ; 120: 170-180, 2018 05 20.
Artigo em Inglês | MEDLINE | ID: mdl-29545070

RESUMO

Increased reactive species (RS; reactive oxygen and nitrogen species) are a byproduct of both enzymatic and non-enzymatic systems, and critical in cancer development, including breast tumorigenesis. To investigate the role of RS-related genes in breast cancer, expression levels of the most common annotated genes involved in regulating cellular RS levels and proteins that are substrates of RS in specific subtypes of breast cancer 9 were evaluated using public data bases. Based on the premise that increased RS promote tumor formation, and breast cancer subtypes vary in aggressiveness, we hypothesized that specific RS gene expression signatures are associated with breast cancer aggressiveness and patient survival. We identified a group of genes (GSTK1, PRDX2, PRDX3 and SLC36A1) that differentiate Luminal B tumors in two clusters and predict survival of patients with Luminal B breast cancers. Furthermore, network analyses of these four genes revealed an overlap of known LumB related pathways with those of RS-related signaling, which included regulation of M-phase and mitochondrial functions.


Assuntos
Neoplasias da Mama/genética , Neoplasias da Mama/metabolismo , Neoplasias da Mama/mortalidade , Espécies Reativas de Oxigênio/metabolismo , Transcriptoma , Sistemas de Transporte de Aminoácidos/genética , Análise por Conglomerados , Feminino , Glutationa Transferase/genética , Humanos , Estimativa de Kaplan-Meier , Oxirredução , Peroxirredoxina III/genética , Peroxirredoxinas/genética , Simportadores/genética
12.
J Biol Chem ; 293(4): 1120-1137, 2018 01 26.
Artigo em Inglês | MEDLINE | ID: mdl-29158255

RESUMO

Triple-negative breast cancer (TNBC) comprises ∼20% of all breast cancers and is the most aggressive mammary cancer subtype. Devoid of the estrogen and progesterone receptors, along with the receptor tyrosine kinase ERB2 (HER2), that define most mammary cancers, there are no targeted therapies for patients with TNBC. This, combined with a high metastatic rate and a lower 5-year survival rate than for other breast cancer phenotypes, means there is significant unmet need for new therapeutic strategies. Herein, the anti-neoplastic effects of the electrophilic fatty acid nitroalkene derivative, 10-nitro-octadec-9-enoic acid (nitro-oleic acid, NO2-OA), were investigated in multiple preclinical models of TNBC. NO2-OA reduced TNBC cell growth and viability in vitro, attenuated TNFα-induced TNBC cell migration and invasion, and inhibited the tumor growth of MDA-MB-231 TNBC cell xenografts in the mammary fat pads of female nude mice. The up-regulation of these aggressive tumor cell growth, migration, and invasion phenotypes is mediated in part by the constitutive activation of pro-inflammatory nuclear factor κB (NF-κB) signaling in TNBC. NO2-OA inhibited TNFα-induced NF-κB transcriptional activity in human TNBC cells and suppressed downstream NF-κB target gene expression, including the metastasis-related proteins intercellular adhesion molecule-1 and urokinase-type plasminogen activator. The mechanisms accounting for NF-κB signaling inhibition by NO2-OA in TNBC cells were multifaceted, as NO2-OA (a) inhibited the inhibitor of NF-κB subunit kinase ß phosphorylation and downstream inhibitor of NF-κB degradation, (b) alkylated the NF-κB RelA protein to prevent DNA binding, and (c) promoted RelA polyubiquitination and proteasomal degradation. Comparisons with non-tumorigenic human breast epithelial MCF-10A and MCF7 cells revealed that NO2-OA more selectively inhibited TNBC function. This was attributed to more facile mechanisms for maintaining redox homeostasis in normal breast epithelium, including a more favorable thiol/disulfide balance, greater extents of multidrug resistance protein-1 (MRP1) expression, and greater MRP1-mediated efflux of NO2-OA-glutathione conjugates. These observations reveal that electrophilic fatty acid nitroalkenes react with more alkylation-sensitive targets in TNBC cells to inhibit growth and viability.


Assuntos
Movimento Celular , Ácidos Graxos/metabolismo , Transdução de Sinais , Neoplasias de Mama Triplo Negativas/metabolismo , Animais , Sobrevivência Celular , Ácidos Graxos/genética , Feminino , Humanos , Células MCF-7 , Camundongos , Camundongos Nus , Invasividade Neoplásica , Proteínas de Neoplasias/genética , Proteínas de Neoplasias/metabolismo , Neoplasias de Mama Triplo Negativas/genética , Neoplasias de Mama Triplo Negativas/patologia
13.
Antioxid Redox Signal ; 28(7): 591-608, 2018 03 01.
Artigo em Inglês | MEDLINE | ID: mdl-29237274

RESUMO

SIGNIFICANCE: It has been proposed that cancer cells are heavily dependent on their antioxidant defenses for survival and growth. Peroxiredoxins are a family of abundant thiol-dependent peroxidases that break down hydrogen peroxide, and they have a central role in the maintenance and response of cells to alterations in redox homeostasis. As such, they are potential targets for disrupting tumor growth. Recent Advances: Genetic disruption of peroxiredoxin expression in mice leads to an increased incidence of neoplastic disease, consistent with a role for peroxiredoxins in protecting genomic integrity. In contrast, many human tumors display increased levels of peroxiredoxin expression, suggesting that strengthened antioxidant defenses provide a survival advantage for tumor progression. Peroxiredoxin inhibitors are being developed and explored as therapeutic agents in different cancer models. CRITICAL ISSUES: It is important to complement peroxiredoxin knockout and expression studies with an improved understanding of the biological function of the peroxiredoxins. Although current results can be interpreted within the context that peroxiredoxins scavenge hydroperoxides, some peroxiredoxin family members appear to have more complex roles in regulating the response of cells to oxidative stress through protein interactions with constituents of other signaling pathways. FUTURE DIRECTIONS: Further mechanistic information is required for understanding the role of oxidative stress in cancer, the function of peroxiredoxins in normal versus cancer cells, and for the design and testing of specific peroxiredoxin inhibitors that display selectivity to malignant cells. Antioxid. Redox Signal. 28, 591-608.


Assuntos
Peróxido de Hidrogênio/metabolismo , Neoplasias/metabolismo , Estresse Oxidativo/genética , Peroxirredoxinas/metabolismo , Progressão da Doença , Humanos , Neoplasias/genética , Neoplasias/patologia , Oxirredução , Peroxidases/metabolismo , Transdução de Sinais
14.
Antioxid Redox Signal ; 28(1): 62-77, 2018 Jan 01.
Artigo em Inglês | MEDLINE | ID: mdl-28398822

RESUMO

Precision in redox signaling is attained through posttranslational protein modifications such as oxidation of protein thiols. The peroxidase peroxiredoxin 1 (PRDX1) regulates signal transduction through changes in thiol oxidation of its cysteines. We demonstrate here that PRDX1 is a binding partner for the tumor suppressive transcription factor FOXO3 that directly regulates the FOXO3 stress response. Heightened oxidative stress evokes formation of disulfide-bound heterotrimers linking dimeric PRDX1 to monomeric FOXO3. Absence of PRDX1 enhances FOXO3 nuclear localization and transcription that are dependent on the presence of Cys31 or Cys150 within FOXO3. Notably, FOXO3-T32 phosphorylation is constitutively enhanced in these mutants, but nuclear translocation of mutant FOXO3 is restored with PI3K inhibition. Here we show that on H2O2 exposure, transcription of tumor suppressive miRNAs let-7b and let-7c is regulated by FOXO3 or PRDX1 expression levels and that let-7c is a novel target for FOXO3. Conjointly, inhibition of let-7 microRNAs increases let-7-phenotypes in PRDX1-deficient breast cancer cells. Altogether, these data ascertain the existence of an H2O2-sensitive PRDX1-FOXO3 signaling axis that fine tunes FOXO3 activity toward the transcription of gene targets in response to oxidative stress. Antioxid. Redox Signal. 28, 62-77.


Assuntos
Proteína Forkhead Box O3/genética , Regulação da Expressão Gênica , MicroRNAs/genética , Oxirredução , Peroxirredoxinas/metabolismo , Interferência de RNA , Sítios de Ligação , Linhagem Celular , Movimento Celular , Dissulfetos , Humanos , Modelos Biológicos , Estresse Oxidativo , Peroxirredoxinas/genética , Regiões Promotoras Genéticas , Ligação Proteica , Transporte Proteico
15.
Oncotarget ; 8(54): 92667-92681, 2017 Nov 03.
Artigo em Inglês | MEDLINE | ID: mdl-29190947

RESUMO

Peroxiredoxin-1 (Prdx1), a member of the thioredoxin (Txn) system, is overexpressed and correlates with poor prognosis in pancreatic cancer patients and can suppress Kras signaling through redox-mediated inhibition of ERK and AKT in lung and breast cancer. Its redox function is maintained by Txn and sulfiredoxin (Srxn), and its tumor promoting functions are activated by post-translational modification. We studied the role of the Txn system in pancreatic neoplasia and cancer by determining how it regulates the phosphorylation of Kras effectors and by determining its association with patient survival. We found that elevated Prdx1 nuclear localization significantly correlated with better patient survival. Our data also demonstrate that the expression of the Txn system is dysregulated, with elevated Prdx1 expression and significantly decreased Txn and Srxn expression in pancreatic lesions of targeted mutant Kras mouse models. This correlated with distinct differences in the interconversion of Prdx1 oligomers that affect its ability to regulate ERK and AKT phosphorylation. Our data also suggest that Prdx1 post-translational modification and oligomerization suppress Prdx1 mediated redox regulation of ERK phosphorylation. We observed distinct differences in Txn expression and in the ability of pTyr-Prdx1 to bind to pERK in a PanIN model of pancreatic neoplasia as compared to an IPMN model, indicating a distinct difference in the function of post-translationally modified Prdx1 in cells with less Txn expression. Modified Txn system function and post-translational regulation may therefore play a significant role in pancreatic tumorigenesis by altering Kras effector phosphorylation and inhibiting the tumor suppressive redox functions of Prdx1.

16.
Oncol Rep ; 35(3): 1409-16, 2016 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-26707944

RESUMO

Breastfeeding can reduce breast cancer risk; however, unknown factors modify this protective effect. Zinc (Zn) modulates an array of cellular functions including oxidative stress, cell proliferation, motility and apoptosis. Marginal Zn intake is common in women and is associated with breast cancer. We reported that marginal Zn intake in mice leads to mammary gland hypoplasia and hallmarks of pre-neoplastic lesions. In the present study, we tested the hypothesis that marginal Zn intake confounds the protective effect of lactation on breast cancer. Nulliparous mice fed control (ZA, 30 mg Zn/kg) or a marginal Zn diet (ZD, 15 mg Zn/kg), were bred and offspring were weaned naturally. Post-involution, mice were gavaged with corn oil or 7,12-dimethylbenz(a)anthracene (DMBA, 1 mg/wk for 4 weeks) and tumor development was monitored. A ZD diet led to insufficient involution, increased fibrosis and oxidative stress. Following DMBA treatment, mice fed ZD had higher oxidative stress in mammary tissue that correlated with reduced levels of peroxiredoxin-1 and p53 and tended to have shorter tumor latency and greater incidence of non-palpable tumors. In summary, marginal Zn intake creates a toxic mammary gland microenvironment and abrogates the protective effect of lactation on carcinogenesis.


Assuntos
Neoplasias da Mama/dietoterapia , Transformação Celular Neoplásica/efeitos dos fármacos , Glândulas Mamárias Animais/efeitos dos fármacos , Estresse Oxidativo/efeitos dos fármacos , Zinco/administração & dosagem , 9,10-Dimetil-1,2-benzantraceno/toxicidade , Animais , Apoptose/efeitos dos fármacos , Neoplasias da Mama/induzido quimicamente , Neoplasias da Mama/metabolismo , Neoplasias da Mama/patologia , Carcinogênese/efeitos dos fármacos , Movimento Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Feminino , Humanos , Lactação/genética , Glândulas Mamárias Animais/patologia , Camundongos , Peroxirredoxinas/biossíntese , Proteína Supressora de Tumor p53/biossíntese
17.
Nucleic Acids Res ; 43(13): 6334-47, 2015 Jul 27.
Artigo em Inglês | MEDLINE | ID: mdl-26082495

RESUMO

Cellular DNA is organized into chromosomes and capped by a unique nucleoprotein structure, the telomere. Both oxidative stress and telomere shortening/dysfunction cause aging-related degenerative pathologies and increase cancer risk. However, a direct connection between oxidative damage to telomeric DNA, comprising <1% of the genome, and telomere dysfunction has not been established. By fusing the KillerRed chromophore with the telomere repeat binding factor 1, TRF1, we developed a novel approach to generate localized damage to telomere DNA and to monitor the real time damage response at the single telomere level. We found that DNA damage at long telomeres in U2OS cells is not repaired efficiently compared to DNA damage in non-telomeric regions of the same length in heterochromatin. Telomeric DNA damage shortens the average length of telomeres and leads to cell senescence in HeLa cells and cell death in HeLa, U2OS and IMR90 cells, when DNA damage at non-telomeric regions is undetectable. Telomere-specific damage induces chromosomal aberrations, including chromatid telomere loss and telomere associations, distinct from the damage induced by ionizing irradiation. Taken together, our results demonstrate that oxidative damage induces telomere dysfunction and underline the importance of maintaining telomere integrity upon oxidative damage.


Assuntos
Dano ao DNA , Estresse Oxidativo , Encurtamento do Telômero , Telômero/metabolismo , Morte Celular , Linhagem Celular , Senescência Celular , Reparo do DNA , Proteínas de Fluorescência Verde/genética , Guanina/análogos & derivados , Guanina/metabolismo , Humanos , Sequências Repetitivas de Ácido Nucleico , Telômero/química , Proteína 1 de Ligação a Repetições Teloméricas/genética
18.
Adv Cancer Res ; 119: 107-25, 2013.
Artigo em Inglês | MEDLINE | ID: mdl-23870510

RESUMO

Breast cancer is a highly complex tissue composed of neoplastic and stromal cells. Carcinoma-associated fibroblasts (CAFs) are commonly found in the cancer stroma, where they promote tumor growth and enhance vascularity in the microenvironment. Upon exposure to oxidative stress, fibroblasts undergo activation to become myofibroblasts. These cells are highly mobile and contractile and often express numerous mesenchymal markers. CAF activation is irreversible, making them incapable of being removed by nemosis. In breast cancer, almost 80% of stromal fibroblasts acquire an activated phenotype that manifests by secretion of elevated levels of growth factors, cytokines, and metalloproteinases. They also produce hydrogen peroxide, which induces the generation of subsequent sets of activated fibroblasts and tumorigenic alterations in epithelial cells. While under oxidative stress, the tumor stroma releases high energy nutrients that fuel cancer cells and facilitate their growth and survival. This review describes how breast cancer progression is dependent upon oxidative stress activated stroma and proposes potential new therapeutic avenues.


Assuntos
Neoplasias da Mama/patologia , Estresse Oxidativo , Células Estromais/patologia , Microambiente Tumoral , Adulto , Idoso , Neoplasias da Mama/metabolismo , Neoplasias da Mama Masculina/metabolismo , Neoplasias da Mama Masculina/patologia , Caveolina 1/metabolismo , Linhagem Celular Tumoral , Transformação Celular Neoplásica , Progressão da Doença , Feminino , Fibroblastos/citologia , Fibroblastos/patologia , Humanos , Masculino , Pessoa de Meia-Idade , Mitocôndrias/metabolismo , Miofibroblastos/citologia , Fenótipo , Espécies Reativas de Oxigênio , Células Estromais/metabolismo
19.
PLoS One ; 7(11): e47281, 2012.
Artigo em Inglês | MEDLINE | ID: mdl-23144812

RESUMO

Many human diseases are attributable to complex interactions among genetic and environmental factors. Statistical tools capable of modeling such complex interactions are necessary to improve identification of genetic factors that increase a patient's risk of disease. Logic Forest (LF), a bagging ensemble algorithm based on logic regression (LR), is able to discover interactions among binary variables predictive of response such as the biologic interactions that predispose individuals to disease. However, LF's ability to recover interactions degrades for more infrequently occurring interactions. A rare genetic interaction may occur if, for example, the interaction increases disease risk in a patient subpopulation that represents only a small proportion of the overall patient population. We present an alternative ensemble adaptation of LR based on boosting rather than bagging called LBoost. We compare the ability of LBoost and LF to identify variable interactions in simulation studies. Results indicate that LBoost is superior to LF for identifying genetic interactions associated with disease that are infrequent in the population. We apply LBoost to a subset of single nucleotide polymorphisms on the PRDX genes from the Cancer Genetic Markers of Susceptibility Breast Cancer Scan to investigate genetic risk for breast cancer. LBoost is publicly available on CRAN as part of the LogicForest package, http://cran.r-project.org/.


Assuntos
Algoritmos , Neoplasias da Mama/genética , Epistasia Genética , Peroxirredoxinas/genética , Simulação por Computador , Feminino , Predisposição Genética para Doença , Humanos , Modelos Genéticos , Polimorfismo de Nucleotídeo Único
20.
J Biol Chem ; 287(45): 37732-44, 2012 Nov 02.
Artigo em Inglês | MEDLINE | ID: mdl-22989880

RESUMO

Prostate cancer (PCa) is the most frequently diagnosed malignancy in men, and the second highest contributor of male cancer related lethality. Disease mortality is due primarily to metastatic spread, highlighting the urgent need to identify factors involved in this progression. Activation of the genetic epithelial to mesenchymal transition (EMT) program is implicated as a major contributor of PCa progression. Initiation of EMT confers invasive and metastatic behavior in preclinical models and is correlated with poor clinical prognosis. Extracellular Hsp90 (eHsp90) promotes cell motility and invasion in cancer cells and metastasis in preclinical models, however, the mechanistic basis for its widespread tumorigenic function remains unclear. We have identified a novel and pivotal role for eHsp90 in driving EMT events in PCa. In support of this notion, more metastatic PCa lines exhibited increased eHsp90 expression relative to their lineage-related nonmetastatic counterparts. We demonstrate that eHsp90 promoted cell motility in an ERK and matrix metalloproteinase-2/9-dependent manner, and shifted cellular morphology toward a mesenchymal phenotype. Conversely, inhibition of eHsp90 attenuated pro-motility signaling, blocked PCa migration, and shifted cell morphology toward an epithelial phenotype. Last, we report that surface eHsp90 was found in primary PCa tumor specimens, and elevated eHsp90 expression was associated with increased levels of matrix metalloproteinase-2/9 transcripts. We conclude that eHsp90 serves as a driver of EMT events, providing a mechanistic basis for its ability to promote cancer progression and metastasis in preclinical models. Furthermore, its newly identified expression in PCa specimens, and potential regulation of pro-metastatic genes, supports a putative clinical role for eHsp90 in PCa progression.


Assuntos
Transição Epitelial-Mesenquimal/genética , Proteínas de Choque Térmico HSP90/genética , Neoplasias da Próstata/genética , Transdução de Sinais/genética , Anticorpos Bloqueadores/imunologia , Anticorpos Bloqueadores/farmacologia , Western Blotting , Linhagem Celular Tumoral , Movimento Celular/efeitos dos fármacos , Movimento Celular/genética , Dipeptídeos/farmacologia , Progressão da Doença , Regulação Neoplásica da Expressão Gênica , Células HEK293 , Proteínas de Choque Térmico HSP90/imunologia , Proteínas de Choque Térmico HSP90/metabolismo , Humanos , Proteína-1 Relacionada a Receptor de Lipoproteína de Baixa Densidade/genética , Proteína-1 Relacionada a Receptor de Lipoproteína de Baixa Densidade/metabolismo , Masculino , Metaloproteinase 2 da Matriz/genética , Metaloproteinase 2 da Matriz/metabolismo , Metaloproteinase 9 da Matriz/genética , Metaloproteinase 9 da Matriz/metabolismo , Neoplasias da Próstata/metabolismo , Neoplasias da Próstata/patologia , Inibidores de Proteases/farmacologia , Interferência de RNA , Reação em Cadeia da Polimerase Via Transcriptase Reversa
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