RESUMEN
BACKGROUND: Triple-negative breast cancer (TNBC) affects young women and is the most aggressive subtype of breast cancer (BC). TNBCs disproportionally affect women of African-American (AA) descent compared to other ethnicities. We have identified DNA repair gene RAD51 as a poor prognosis marker in TNBC and its posttranscriptional regulation through microRNAs (miRNAs). This study aims to delineate the mechanisms leading to RAD51 upregulation and develop novel therapeutic combinations to effectively treat TNBCs and reduce disparity in clinical outcomes. METHODS: Analysis of TCGA data for BC cohorts using the UALCAN portal and PrognoScan identified the overexpression of RAD51 in TNBCs. miRNA sequencing identified significant downregulation of RAD51-targeting miRNAs miR-214-5P and miR-142-3P. RT-PCR assays were used to validate the levels of miRNAs and RAD51, and immunohistochemical and immunoblotting techniques were used similarly for RAD51 protein levels in TNBC tissues and cell lines. Luciferase assays were performed under the control of RAD51 3'-UTR to confirm that miR-214-5P regulates RAD51 expression. To examine the effect of miR-214-5P-mediated downregulation of RAD51 on homologous recombination (HR) in TNBC cells, Dr-GFP reporter assays were performed. To assess the levels of olaparib-induced DNA damage responses in miR-214-5P, transfected cells, immunoblots, and immunofluorescence assays were used. Furthermore, COMET assays were used to measure DNA lesions and colony assays were performed to assess the sensitivity of BRCA-proficient TNBC cells to olaparib. RESULTS: In-silico analysis identified upregulation of RAD51 as a poor prognostic marker in TNBCs. miRNA-seq data showed significant downregulation of miR-214-5P and miR-142-3P in TNBC cell lines derived from AA women compared to Caucasian-American (CA) women. miR-214-5P mimics downregulated RAD51 expression and induces HR deficiency as measured by Dr-GFP assays in these cell lines. Based on these results, we designed a combination treatment of miR-214-5P and olaparib in HR-proficient AA TNBC cell lines using clonogenic survival assays. The combination of miR-214-5P and olaparib showed synergistic lethality compared to individual treatments in these cell lines. CONCLUSIONS: Our studies identified a novel epigenetic regulation of RAD51 in TNBCs by miR-214-5P suggesting a novel combination therapies involving miR-214-5P and olaparib to treat HR-proficient TNBCs and to reduce racial disparity in therapeutic outcomes.
Asunto(s)
MicroARNs , Neoplasias de la Mama Triple Negativas , Humanos , Femenino , Neoplasias de la Mama Triple Negativas/tratamiento farmacológico , Neoplasias de la Mama Triple Negativas/genética , Neoplasias de la Mama Triple Negativas/metabolismo , Epigénesis Genética , Factores Raciales , Línea Celular Tumoral , MicroARNs/genética , Regulación Neoplásica de la Expresión Génica , Proliferación Celular/genética , Recombinasa Rad51/genética , Recombinasa Rad51/metabolismoRESUMEN
A novel nitrogen mustard CBISC has been synthesized and evaluated as an anticancer agent. CBISC has been shown to exhibit enhanced cell proliferation inhibition properties against mutant p53 cell lines colorectal cancer WiDr, pancreatic cancer (MIAPaCa-2 and PANC-1), and triple negative breast cancer (MDA-MB-231 and MDA-MB-468). In vitro mechanism of action studies revealed perturbations in the p53 pathway and increased cell death as evidenced by western blotting, immunofluorescent microscopy and MTT assay. Further, in vivo studies revealed that CBISC is well tolerated in healthy mice and exhibited significant in vivo tumor growth inhibition properties in WiDr and MIAPaCa-2 xenograft models. These studies illustrate the potential utility of CBISC as an anticancer agent.
Asunto(s)
Antineoplásicos/síntesis química , Antineoplásicos/farmacología , Daño del ADN , Proteínas Mutantes/metabolismo , Proteína p53 Supresora de Tumor/genética , Animales , Apoptosis/efectos de los fármacos , Sistemas CRISPR-Cas/genética , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , Clorambucilo/química , Clorambucilo/farmacología , Cloranfenicol/química , Cloranfenicol/farmacología , Femenino , Ratones Desnudos , Poli(ADP-Ribosa) Polimerasas/metabolismo , Proteína p53 Supresora de Tumor/metabolismo , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
A series of nitric oxide (NO) donor furoxan conjugates of N, N-dialkylcarboxy coumarins have been synthesized as potential anticancer agents. The synthesized compounds have been tested for their in vitro antiproliferative activities on various cancer and noncancerous cell lines. The candidate derivatives exhibit selectivity towards cancer cells with excellent activities in low nM to µM concentrations. In vitro mechanistic studies indicate that the candidate compounds generate substantial NO, inhibit colony formation, and cause apoptosis in cancer cells. A preliminary in vivo tolerance study of the lead candidate 10 in mice indicates that it is well-tolerated, evidenced by zero mortality and normal body weight gains in treated mice. Further translation of the lead derivative 10 using MDA-MB-231 based tumor xenograft model shows good tumor growth reduction.
Asunto(s)
Antineoplásicos/farmacología , Cumarinas/farmacología , Óxido Nítrico/farmacología , Animales , Antineoplásicos/síntesis química , Antineoplásicos/química , Apoptosis/efectos de los fármacos , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , Cumarinas/química , Relación Dosis-Respuesta a Droga , Ensayos de Selección de Medicamentos Antitumorales , Femenino , Humanos , Ratones , Estructura Molecular , Óxido Nítrico/química , Relación Estructura-ActividadRESUMEN
Several phosphonium derivatives have been synthesized from Baylis-Hillman (BH) reaction derived allyl bromides and aryl phosphines as mitochondria targeting anticancer agents. In vitro cell proliferation inhibition studies on various solid tumor cell lines indicate that most of the compounds exhibit IC50 values in µM concentrations. Further studies reveal that ß-substituted BH bromide derived phosphonium derivatives enhance the biological activity to low µM IC50 values. In vitrometabolic studies show that the lead candidate compound 16 inhibits the production of mitochondrial ATP, increases the proton leak within the mitochondrial membrane and abolishes the spare respiratory capacity in a concentration dependent manner.
Asunto(s)
Antineoplásicos/farmacología , Ácidos Carboxílicos/farmacología , Ésteres/farmacología , Compuestos Organofosforados/farmacología , Animales , Antineoplásicos/síntesis química , Antineoplásicos/química , Ácidos Carboxílicos/síntesis química , Ácidos Carboxílicos/química , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , Supervivencia Celular/efectos de los fármacos , Relación Dosis-Respuesta a Droga , Ensayos de Selección de Medicamentos Antitumorales , Ésteres/síntesis química , Ésteres/química , Femenino , Humanos , Ratones , Estructura Molecular , Compuestos Organofosforados/síntesis química , Compuestos Organofosforados/química , Relación Estructura-ActividadRESUMEN
BACKGROUND: Breast cancer remains as one of the most lethal types of cancer in women. Among various subtypes, triple-negative breast cancer (TNBC) is the most aggressive and hard to treat type of breast cancer. Mechanistically, increased DNA repair and cell cycle checkpoint activation remain as the foremost reasons behind TNBC tumor resistance to chemotherapy and disease recurrence. METHODS: We evaluated the mechanism of prexasertib-induced regulation of homologous recombination (HR) proteins using 20S proteasome inhibitors and RT-PCR. HR efficiency and DNA damages were evaluated using Dr-GFP and comet assays. DNA morphology and DNA repair focus studies were analyzed using immunofluorescence. UALCAN portal was used to evaluate the expression of RAD51 and survival probability based on tumor stage, subtype, and race in breast cancer patients. RESULTS: Our results show that prexasertib treatment promotes both post-translational and transcriptional mediated regulation of BRCA1 and RAD51 proteins. Additionally, prexasertib-treated TNBC cells revealed over 55% reduction in HR efficiency compared to control cells. Based on these results, we hypothesized that prexasertib treatment induced homologous recombination deficiency (HRD) and thus should synergize with PARP inhibitors (PARPi) in TNBC cells. As predicted, combined treatment of prexasertib and PARPi olaparib increased DNA strand breaks, γH2AX foci, and nuclear disintegration relative to single-agent treatment. Further, the prexasertib and olaparib combination was synergistic in multiple TNBC cell lines, as indicated by combination index (CI) values. Analysis of TCGA data revealed elevated RAD51 expression in breast tumors compared to normal breast tissues, especially in TNBC subtype. Interestingly, there was a discrepancy in RAD51 expression in racial groups, with African-American and Asian breast cancer patients showing elevated RAD51 expression compared to Caucasian breast cancer patients. Consistent with these observations, African-American and Asian TNBC patients show decreased survival. CONCLUSIONS: Based on these data, RAD51 could be a biomarker for aggressive TNBC and for racial disparity in breast cancer. As positive correlation exists between RAD51 and CHEK1 expression in breast cancer, the in vitro preclinical data presented here provides additional mechanistic insights for further evaluation of the rational combination of prexasertib and olaparib for improved outcomes and reduced racial disparity in TNBC.
Asunto(s)
Antineoplásicos/farmacología , Recombinación Homóloga/efectos de los fármacos , Ftalazinas/farmacología , Piperazinas/farmacología , Inhibidores de Poli(ADP-Ribosa) Polimerasas/farmacología , Pirazinas/farmacología , Pirazoles/farmacología , Neoplasias de la Mama Triple Negativas/patología , Proteína BRCA1/genética , Biomarcadores de Tumor/genética , Línea Celular Tumoral , Quinasa 1 Reguladora del Ciclo Celular (Checkpoint 1)/genética , Daño del ADN/efectos de los fármacos , Resistencia a Antineoplásicos , Sinergismo Farmacológico , Femenino , Regulación Neoplásica de la Expresión Génica/efectos de los fármacos , Humanos , Recombinasa Rad51/genética , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
The normal female reproductive hormone estrogen has been linked with increased risk of breast and many other forms of cancer. This is largely due to metabolic conversion of estrogens into highly reactive catechol estrogen quinones which can interact with DNA and cause a variety of DNA adducts and lesions. Detection and analysis of these adducts and their associated cellular responses involve complex chemical, enzymatic, and LC-MS based methods, which are both laborious and require specialized expertise and instrumentation. Herein, we show that using a biotin-labeled estradiol allows immunodetection of estrogen-induced DNA adducts by slot blot and single-cell molecular combing and proximity ligation assays. The biotinylated and unlabeled estradiols induced similar levels of DNA single and double strand breaks as measured by comet assays. Using biotinylated estrogen, we further show that estrogens are able to activate the Fanconi anemia-BRCA tumor suppressor pathway and cause DNA strand breaks and oxidatively modified DNA bases as well as gross chromosomal aberrations. Utilization of biotin-labeled estrogens could be a powerful tool to detect estrogen adducts and associated DNA damage, and to track estrogen adduct-induced cellular responses and carcinogenic mechanisms in cultured cells. The techniques presented here allow simple and rapid detection and quantitation of estrogen adducts by slot blot as well as direct visualization on the DNA strand and could pave the way for developing new treatments to protect the genome from the effects of reactive estrogen metabolites. © 2016 Wiley Periodicals, Inc.
Asunto(s)
Carcinógenos/metabolismo , Aductos de ADN/metabolismo , Estradiol/química , Estrógenos/toxicidad , Proteínas del Grupo de Complementación de la Anemia de Fanconi/metabolismo , Biotinilación , Células Cultivadas , Aberraciones Cromosómicas , Estrógenos/química , Femenino , Regulación de la Expresión Génica/efectos de los fármacos , Humanos , Células MCF-7 , Análisis de la Célula IndividualRESUMEN
Aberrant expression of hedgehog molecules, particularly Gli1, is common in cancers of many tissues and is responsible for their aggressive behavior and chemoresistance. Here we demonstrate a novel and tumor-specific role for aberrant Gli1 in the regulation of the S-phase checkpoint that suppresses replication stress and resistance to chemotherapy. Inhibition of Gli1 in tumor cells induced replication stress-mediated DNA damage response, attenuated their clonogenic potential, abrogated camptothecin (CPT)-induced Chk1 phosphorylation, and potentiated its cytotoxicity. However, in normal fibroblasts, Gli1 siRNAs showed no significant changes in CPT-induced Chk1 phosphorylation. Further analysis of ataxia telangiectasia and Rad3-related protein (ATR)/Chk1 signaling cascade genes in tumor cells revealed an unexpected mechanism whereby Gli1 regulates ATR-mediated Chk1 phosphorylation by transcriptional regulation of the BH3-only protein Bid. Consistent with its role in DNA damage response, Bid down-regulation in tumor cells abolished CPT-induced Chk1 phosphorylation and sensitized them to CPT. Correspondingly, Gli1 inhibition affected the expression of Bid and the association of replication protein A (RPA) with the ATR- interacting protein (ATRIP)-ATR complex, and this compromised the S-phase checkpoint. Conversely, complementation of Bid in Gli1-deficient cells restored CPT-induced Chk1 phosphorylation. An in silico analysis of the Bid promoter identified a putative Gli1 binding site, and further studies using luciferase reporter assays confirmed Gli1-dependent promoter activity. Collectively, our studies established a novel connection between aberrant Gli1 and Bid in the survival of tumor cells and their response to chemotherapy, at least in part, by regulating the S-phase checkpoint. Importantly, our data suggest a novel drug combination of Gli1 and Top1 inhibitors as an effective therapeutic strategy in treating tumors that expresses Gli1.
Asunto(s)
Proteína Proapoptótica que Interacciona Mediante Dominios BH3/metabolismo , Regulación Neoplásica de la Expresión Génica , Proteínas Quinasas/metabolismo , Fase S , Inhibidores de Topoisomerasa I/metabolismo , Factores de Transcripción/metabolismo , Antineoplásicos/química , Sitios de Unión , Camptotecina/química , Ciclo Celular , Línea Celular Tumoral , Supervivencia Celular , Quinasa 1 Reguladora del Ciclo Celular (Checkpoint 1) , Reparación del ADN , Regulación hacia Abajo , Resistencia a Medicamentos , Fibroblastos/metabolismo , Humanos , Fosforilación , Regiones Promotoras Genéticas , Interferencia de ARN , ARN Interferente Pequeño/metabolismo , Proteína de Replicación A/metabolismo , Transducción de Señal , Inhibidores de Topoisomerasa I/química , Proteína con Dedos de Zinc GLI1RESUMEN
Combination chemotherapy regimens that include fluoropyrimidine (FP) drugs, e.g., 5-fluorouracil (5-FU), are central to the treatment of colorectal cancer liver metastases (CRLMs), a major cause of cancer mortality. We tested a second-generation FP polymer, CF10, in a CC531/WAGRij syngeneic orthotopic rat model of liver metastasis to determine if CF10 improved response relative to 5-FU. CF10 displayed increased potency relative to 5-FU in CC531 rat colorectal cancer cells based on clonogenic assay results and caused increased apoptosis, as shown using a live/dead assay. The increased potency of CF10 to CC531 cells was associated with increased replication stress, as assessed by Western blot for biomarkers of ATR/Chk1 and ATM/Chk2 pathway activation. CF10 dosed to deliver equivalent FP content as an established dose of 5-FU in rats (50 mg/kg) did not cause weight loss in WAGRij rats even when combined with ethynyl uracil (EU), an inhibitor of dihydropyrimidine dehydrogenase, the enzyme primarily responsible for 5-FU degradation in the liver. In contrast, 5-FU caused significant weight loss that was exacerbated in combination with EU. Importantly, CF10 was significantly more effective than 5-FU at inhibiting tumor progression (~90% reduction) in the CC531/WAG/Rij CRLM model. Our results reveal strong potential for CF10 to be used for CRLM treatment.
RESUMEN
Chemoresistance contributes to the majority of deaths in women with ovarian cancer (OC). Altered DNA repair and metabolic signaling is implicated in mediating therapeutic resistance. DNA damage checkpoint kinase 1 (CHK1) integrates cell cycle and DNA repair in replicating cells, and its inhibition causes replication stress, repair deficiency and cell cycle dysregulation. We observed elevated Poly-ADP-ribosylation (PAR) of proteins (PARylation) and subsequent decrease in cellular NAD+ levels in OC cells treated with the CHK1 inhibitor prexasertib, indicating activation of NAD+ dependent DNA repair enzymes poly-ADP-ribose polymerases (PARP1/2). While multiple PARP inhibitors are in clinical use in treating OC, tumor resistance to these drugs is highly imminent. We reasoned that inhibition of dePARylation by targeting Poly (ADP-ribose) glycohydrolase (PARG) would disrupt metabolic and DNA repair crosstalk to overcome chemoresistance. Although PARG inhibition (PARGi) trapped PARylation of the proteins and activated CHK1, it did not cause any significant OC cell death. However, OC cells deficient in CHK1 were hypersensitive to PARGi, suggesting a role for metabolic and DNA repair crosstalk in protection of OC cells. Correspondingly, OC cells treated with a combination of CHK1 and PARG inhibitors exhibited excessive replication stress-mediated DNA lesions, cell cycle dysregulation, and mitotic catastrophe compared to individual drugs. Interestingly, increased PARylation observed in combination treatment resulted in depletion of NAD+ levels. These decreased NAD+ levels were also paralleled with reduced aldehyde dehydrogenase (ALDH) activity, which requires NAD+ to maintain cancer stem cells. Furthermore, prexasertib and PARGi combinations exhibited synergistic cell death in OC cells, including an isogenic chemoresistant cell line and 3D organoid models of primary patient-derived OC cell lines. Collectively, our data highlight a novel crosstalk between metabolism and DNA repair involving replication stress and NAD+-dependent PARylation, and suggest a novel combination therapy of CHK1 and PARG inhibitors to overcome chemoresistance in OC.
RESUMEN
To determine whether the bleomycin (BLM)-induced bystander response occurs in human brain glioblastoma (BMG-1) cells, the BMG-1 cells were exposed to two different concentrations of BLM. The co-culture methodology was adopted to study the in vitro bystander effects. DNA damage was measured using the micronucleus (MN) and γ-H2AX assays. Cytotoxicity was measured using the trypan blue assay. Cell cycle kinetics was analyzed using flow cytometry. The overall results did not show any significant increase in either genotoxicity or cytotoxicity or a delay in the cell cycle kinetics in BMG-1 bystander cells co-cultured with BLM-exposed cells, suggesting that BLM did not induce a bystander response in the BMG-1 cells. Furthermore, the MN results of the BLM-exposed BMG-1 cells co-cultured with unexposed bystander human lung adenocarcinoma (A549 and NCI-H460) cells and vice versa suggested that the BMG-1 cells do not secrete bystander signals but do respond to those signals. Analyzing the underlying mechanism and pathways involved in preventing the cells from secreting bystander signals will provide new insights that can be applied to inhibit these mechanisms in other cell types, thereby preventing and controlling the bystander response and genomic instability and increasing the therapeutic gain in chemotherapy.
Asunto(s)
Adenocarcinoma/metabolismo , Antibióticos Antineoplásicos/farmacología , Bleomicina/farmacología , Neoplasias Encefálicas/metabolismo , Efecto Espectador/efectos de los fármacos , Glioblastoma/metabolismo , Neoplasias Pulmonares/metabolismo , Adenocarcinoma/tratamiento farmacológico , Adenocarcinoma/patología , Antibióticos Antineoplásicos/uso terapéutico , Bleomicina/uso terapéutico , Neoplasias Encefálicas/tratamiento farmacológico , Neoplasias Encefálicas/patología , Línea Celular Tumoral , Técnicas de Cocultivo , Femenino , Inestabilidad Genómica/efectos de los fármacos , Glioblastoma/tratamiento farmacológico , Glioblastoma/patología , Histonas/metabolismo , Humanos , Neoplasias Pulmonares/tratamiento farmacológico , Neoplasias Pulmonares/patología , Masculino , Proteínas de Neoplasias/metabolismo , Transducción de Señal/efectos de los fármacosRESUMEN
Triple negative breast cancer (TNBC), an aggressive and highly metastatic subtype of breast cancer. Glioma-associated oncogene 1 (GLI1) is a transcription factor and effector of the Hedgehog (Hh) signaling pathway, and is predictive of poor survival for TNBC patients. A nanostring DNA Damage Response (DDR) mRNA panel was used to identify GLI1-induced regulation of DDR genes. Western blots, immunohistochemistry and immunofluorescence were used to evaluate protein expression. Colony assays and mammosphere formation assays were utilized to assess survival of cancer cells. Flow cytometry analyses were employed to evaluate changes in the cell cycle profile, and DNA fiber assays were used to analyze alterations in replication dynamics in TNBC cells. The UALCAN portal and Ensemble programs were used for computational analysis of TCGA data. CompuSyn software was used to calculate combination index (CI) values to assess synergism in drug combination experiments. Inhibition of GLI1 in TNBC cells transcriptionally downregulate expression of FANCD2 and its foci formation, and causes a homologous recombination repair (HR) deficiency. As HR-deficient cancer cells are sensitive to PARP-targeted therapies, we evaluated a combination of the GLI1 inhibitor, GANT61, and a PARP inhibitor (olaparib) in TNBC cells. Combination of GANT61 and olaparib elevated DNA damage levels and these drug combinations caused synergistic lethality to TNBC cells. Aberrantly activated GLI1 regulates HR-mediated DNA repair by transcriptionally regulating FANCD2 to overcome chemotherapy-induced replication stress and DNA damage, and it contributes to resistance of TNBC cells to therapeutics.
Asunto(s)
Replicación del ADN , Sinergismo Farmacológico , Recombinación Homóloga , Inhibidores de Poli(ADP-Ribosa) Polimerasas/farmacología , Piridinas/farmacología , Pirimidinas/farmacología , Neoplasias de la Mama Triple Negativas/tratamiento farmacológico , Proteína con Dedos de Zinc GLI1/antagonistas & inhibidores , Apoptosis , Ciclo Celular , Movimiento Celular , Proliferación Celular , Quimioterapia Combinada , Proteína del Grupo de Complementación D2 de la Anemia de Fanconi/genética , Proteína del Grupo de Complementación D2 de la Anemia de Fanconi/metabolismo , Humanos , Estrés Oxidativo , Pronóstico , Tasa de Supervivencia , Neoplasias de la Mama Triple Negativas/metabolismo , Neoplasias de la Mama Triple Negativas/patología , Células Tumorales CultivadasRESUMEN
BACKGROUND: DNA damage accumulation and mitochondrial abnormalities are elevated in neurons during aging and may contribute to neurodegenerative pathologic conditions such as Alzheimer's disease. BRCA1 interacting protein 1 or BRIP1 is a 5' to 3' DNA helicase that catalyzes many abnormal DNA structures during DNA replication, gene transcription, and recombination, and contribute to genomic integrity. OBJECTIVE: BRIP1 functions were reasonably well studied in DNA repair; however, there is limited data on its role and regulation during aging and neurodegenerative diseases. METHODS: We used immunohistochemistry, western blot, and qRT-PCR assays to analyze the expression of BRIP1. Immunofluorescence studies were performed to study the formation of R-loops, reactive oxygen species (ROS) generation, and mitochondrial morphology. Flow cytometry and transmission electron microscopy were used to evaluate mitochondrial ROS and mitochondrial structures, respectively. Oxygen consumption rate was measured using Seahorse, and the Presto Blue™ assays were used to evaluate cell viability. RESULTS: Our results demonstrate the expression of BRIP1 in mouse and human brain tissues and in neuronal cell lines. BRIP1 levels were elevated in the hippocampal regions of the brains, specifically in the dentate gyrus. BRIP1 downregulation in neuronal cells caused increased R-loop formation basally and in response to H2O2 treatment. Furthermore, BRIP1 deficient cells exhibited elevated levels of excitotoxicity induced by L-Glutamic acid exposure as evidenced by (mitochondrial) ROS levels, deteriorated mitochondrial health, and cell death compared to BRIP1 proficient neuronal cells. CONCLUSION: Overall, our results indicate an important role for BRIP1 in maintaining neuronal cell health and homeostasis by suppressing cellular oxidative stress.
Asunto(s)
Encéfalo/patología , Daño del ADN , Proteínas del Grupo de Complementación de la Anemia de Fanconi/genética , Neuronas/metabolismo , ARN Helicasas/genética , Animales , Línea Celular , Supervivencia Celular , Humanos , Ratones , Ratones Endogámicos C57BL , Enfermedades Neurodegenerativas/genética , Neuronas/patología , Estrés Oxidativo/genéticaRESUMEN
Clear cell renal cell carcinoma is the most common and deadly type of cancer affecting the kidney, and is characterized histologically by large intracellular lipid deposits. These deposits are thought to result from lipid metabolic reprogramming occurring in tumor cells, but the exact mechanisms and implications of these metabolic alterations are incompletely understood. Obesity is an independent risk factor for clear cell renal cell carcinoma, and is also associated with lipid accumulation in noncancerous epithelial cells of the proximal tubule, where clear cell renal cell carcinoma originates. This article explores the potential link between obesity-associated renal lipid metabolic disturbances and lipid metabolic reprogramming in clear cell renal cell carcinoma, and discusses potential implications for future research.
RESUMEN
Ovarian cancer (OC) is one of the most lethal type of cancer in women due to a lack of effective targeted therapies and high rates of treatment resistance and disease recurrence. Recently Poly (ADP-ribose) polymerase inhibitors (PARPi) have shown promise as chemotherapeutic agents; however, their efficacy is limited to a small fraction of patients with BRCA mutations. Here we show a novel function for the Hedgehog (Hh) transcription factor Glioma associated protein 1 (GLI1) in regulation of key Fanconi anemia (FA) gene, FANCD2 in OC cells. GLI1 inhibition in HR-proficient OC cells induces HR deficiency (BRCAness), replication stress and synergistic lethality when combined with PARP inhibition. Treatment of OC cells with combination of GLI1 and PARP inhibitors shows enhanced DNA damage, synergy in cytotoxicity, and strong in vivo anticancer responses.
Asunto(s)
Proteína del Grupo de Complementación D2 de la Anemia de Fanconi/metabolismo , Proteínas Hedgehog/metabolismo , Recombinación Homóloga/fisiología , Neoplasias Ováricas/metabolismo , Inhibidores de Poli(ADP-Ribosa) Polimerasas/uso terapéutico , Proteína con Dedos de Zinc GLI1/metabolismo , Animales , Línea Celular Tumoral , Relación Dosis-Respuesta a Droga , Proteína del Grupo de Complementación D2 de la Anemia de Fanconi/genética , Femenino , Proteínas Hedgehog/antagonistas & inhibidores , Proteínas Hedgehog/genética , Humanos , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones Desnudos , Ratones Transgénicos , Neoplasias Ováricas/tratamiento farmacológico , Neoplasias Ováricas/genética , Ftalazinas/farmacología , Ftalazinas/uso terapéutico , Piperazinas/farmacología , Piperazinas/uso terapéutico , Inhibidores de Poli(ADP-Ribosa) Polimerasas/farmacología , Piridinas/farmacología , Piridinas/uso terapéutico , Pirimidinas/farmacología , Pirimidinas/uso terapéutico , Transcripción Genética/efectos de los fármacos , Transcripción Genética/fisiología , Ensayos Antitumor por Modelo de Xenoinjerto/métodos , Proteína con Dedos de Zinc GLI1/antagonistas & inhibidores , Proteína con Dedos de Zinc GLI1/genéticaRESUMEN
AraC-FdUMP[10] (CF10) is a second-generation polymeric fluoropyrimidine that targets both thymidylate synthase (TS), the target of 5-fluorouracil (5-FU), and DNA topoisomerase 1 (Top1), the target of irinotecan, two drugs that are key components of FOLFIRNOX, a standard-of-care regimen for pancreatic ductal adenocarcinoma (PDAC). We demonstrated that F10 and CF10 are potent inhibitors of PDAC cell survival (in multiple cell lines including patient-derived lines) with IC50s in the nanomolar range and are nearly 1,000-fold more potent than 5-FU. The increased potency of CF10 relative to 5-FU correlated with enhanced TS inhibition and strong Top1 cleavage complex formation. Furthermore, CF10 displayed single-agent activity in PDAC murine xenografts without inducing weight loss. Through a focused drug synergy screen, we identified that combining CF10 with targeting the DNA repair enzyme, poly (ADP-ribose) glycohydrolase, induces substantial DNA damage and apoptosis. This work moves CF10 closer to a clinical trial for the treatment of PDAC. IMPLICATIONS: CF10 is a promising polymeric fluoropyrimidine with dual mechanisms of action (i.e., TS and Top1 inhibition) for the treatment of PDAC and synergizes with targeting of DNA repair. VISUAL OVERVIEW: http://mcr.aacrjournals.org/content/molcanres/19/4/565/F1.large.jpg.
Asunto(s)
Antimetabolitos Antineoplásicos/uso terapéutico , Citarabina/uso terapéutico , Proteínas Activadoras de GTPasa/antagonistas & inhibidores , Neoplasias/tratamiento farmacológico , Animales , Antimetabolitos Antineoplásicos/farmacología , Citarabina/farmacología , Femenino , Humanos , Ratones , Ratones DesnudosRESUMEN
Chemotherapy regimens that include 5-fluorouracil (5-FU) are central to colorectal cancer treatment; however, risk/benefit concerns limit 5-FU's use, necessitating development of improved fluoropyrimidine (FP) drugs. In our study, we evaluated a second-generation nanoscale FP polymer, CF10, for improved antitumor activity. CF10 was more potent than the prototype FP polymer F10 and much more potent than 5-FU in multiple colorectal cancer cell lines including HCT-116, LS174T, SW480, and T84D. CF10 displayed improved stability to exonuclease degradation relative to F10 and reduced susceptibility to thymidine antagonism due to extension of the polymer with arabinosyl cytidine. In colorectal cancer cells, CF10 strongly inhibited thymidylate synthase (TS), induced Top1 cleavage complex formation and caused replication stress, while similar concentrations of 5-FU were ineffective. CF10 was well tolerated in vivo and invoked a reduced inflammatory response relative to 5-FU. Blood chemistry parameters in CF10-treated mice were within normal limits. In vivo, CF10 displayed antitumor activity in several colorectal cancer flank tumor models including HCT-116, HT-29, and CT-26. CF10's antitumor activity was associated with increased plasma levels of FP deoxynucleotide metabolites relative to 5-FU. CF10 significantly reduced tumor growth and improved survival (84.5 days vs. 32 days; P < 0.0001) relative to 5-FU in an orthotopic HCT-116-luc colorectal cancer model that spontaneously metastasized to liver. Improved survival in the orthotopic model correlated with localization of a fluorescent CF10 conjugate to tumor. Together, our preclinical data support an early-phase clinical trial of CF10 for treatment of colorectal cancer.
Asunto(s)
Neoplasias Colorrectales/tratamiento farmacológico , Fluorouracilo/metabolismo , Polímeros/metabolismo , Animales , Neoplasias Colorrectales/patología , Humanos , Ratones , Ratones DesnudosRESUMEN
Stem cells are a sub population of cell types that form the foundation of our body, and have the potential to replicate, replenish and repair limitlessly to maintain the tissue and organ homeostasis. Increased lifetime and frequent replication set them vulnerable for both exogenous and endogenous agents-induced DNA damage compared to normal cells. To counter these damages and preserve genetic information, stem cells have evolved with various DNA damage response and repair mechanisms. Furthermore, upon experiencing irreparable DNA damage, stem cells mostly prefer early senescence or apoptosis to avoid the accumulation of damages. However, the failure of these mechanisms leads to various diseases, including cancer. Especially, given the importance of stem cells in early development, DNA repair deficiency in stem cells leads to various disabilities like developmental delay, premature aging, sensitivity to DNA damaging agents, degenerative diseases, etc. In this review, we have summarized the recent update about how DNA repair mechanisms are regulated in stem cells and their association with disease progression and pathogenesis.
Asunto(s)
Envejecimiento Prematuro/metabolismo , Apoptosis , Daño del ADN , Reparación del ADN , Discapacidades del Desarrollo/metabolismo , Neoplasias/metabolismo , Envejecimiento Prematuro/patología , Animales , Senescencia Celular , Discapacidades del Desarrollo/patología , Humanos , Neoplasias/patologíaRESUMEN
Phosphofurin acidic cluster sorting protein-1 (PACS-1) is a multifunctional membrane traffic regulator that plays important roles in organ homeostasis and disease. In this study, we elucidate a novel nuclear function for PACS-1 in maintaining chromosomal integrity. PACS-1 progressively accumulates in the nucleus during cell cycle progression, where it interacts with class I histone deacetylases 2 and 3 (HDAC2 and HDAC3) to regulate chromatin dynamics by maintaining the acetylation status of histones. PACS-1 knockdown results in the proteasome-mediated degradation of HDAC2 and HDAC3, compromised chromatin maturation, as indicated by elevated levels of histones H3K9 and H4K16 acetylation, and, consequently, increased replication stress-induced DNA damage and genomic instability.
Asunto(s)
Cromatina/fisiología , Inestabilidad Genómica , Histona Desacetilasa 1/metabolismo , Histona Desacetilasas/metabolismo , Proteínas de Transporte Vesicular/fisiología , Ciclo Celular , Línea Celular Tumoral , Núcleo Celular/metabolismo , Citosol/metabolismo , Replicación del ADN , Técnicas de Silenciamiento del Gen , Células HeLa , Humanos , Complejo de la Endopetidasa Proteasomal/metabolismo , Estabilidad Proteica , Proteínas de Transporte Vesicular/genéticaRESUMEN
Mortality in ovarian cancer is predominantly due to acquired chemoresistance and tumor recurrence. UBIQUITIN CONJUGATING ENZYME E2 or RAD6 expression increases in cell lines and patient tumors in response to platinum-based chemotherapy and promotes both activation of DNA damage response pathways and expression of stemness genes and a stem cell-like phenotype driving ovarian cancer chemoresistance.