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1.
Radiat Oncol ; 15(1): 94, 2020 May 06.
Artigo em Inglês | MEDLINE | ID: mdl-32375814

RESUMO

BACKGROUND: To explore the efficacy and safety of Transcatheter rectal arterial chemoembolization with oxaliplatin and S-1 concurrent chemoradiotherapy as neoadjuvant therapy for locally advanced rectal cancer. METHODS: This s a prospective, monocentric, non-randomized clinical study, a total of 95 patients were enrolled and assigned to two groups: an investigational group (n = 50) receiving transcatheter rectal arterial chemoembolization (TRACE) with oxaliplatin and preoperative radiotherapy plus S-1 concurrent chemotherapy (NATRACE-CRT), followed by surgery, a control group (n = 45) receiving standard fluorouracil-based combined modality treatment, consisting of preoperative radiotherapy plus capecitabine based chemotherapy (NA-CRT), followed by surgery. The primary endpoint was postoperative pathological regression rate which evaluated by tumor regression grade (TRG) according to the 7th edition of the American Joint Committee on Cancer (AJCC) standard, and the secondary endpoints included objective response rate (ORR) and toxicity, as well as surgical complications, and postoperative tumor downstaging. RESULTS: Compared with NA-CRT group (17.78% (95% confidence interval (CI): 6.2-29.4)), the TRG0 was 30% (95% CI 16.8-43.2) in the NATRACE-CRT group (P = 0.231). The TRG0 + 1 rate was 60% (95% CI: 45.9-74.1) and 33.33% (95% CI: 19-47.7) in NATRACE-CRT group and NA-CRT group, respectively (P = 0.013). The ORR of the NATRACE-CRT group was 84% and that of the NA-CRT group was 66.67% (p = 0.058). Incidence of preoperative toxic side effects and surgical complications was similar between the two groups. CONCLUSION: TRACE with oxaliplatin plus concurrent S-1 chemoradiotherapy as a neoadjuvant therapy provided better pathological remission rate versus standard treatment with a similar safety profile. TRIAL REGISTRATION: NCT03601156.

2.
Chemistry ; 26(20): 4489-4495, 2020 Apr 06.
Artigo em Inglês | MEDLINE | ID: mdl-32073730

RESUMO

It is challenging to design metal catalysts for in situ transformation of endogenous biomolecules with good performance inside living cells. Herein, we report a multifunctional metal catalyst, ruthenium-coordinated oligo(p-phenylenevinylene) (OPV-Ru), for intracellular catalysis of transfer hydrogenation of nicotinamide adenine dinucleotide (NAD+ ) to its reduced format (NADH). Owing to its amphiphilic characteristic, OPV-Ru possesses good self-assembly capability in water to form nanoparticles through hydrophobic interaction and π-π stacking, and numerous positive charges on the surface of nanoparticles displayed a strong electrostatic interaction with negatively charged substrate molecules, creating a local microenvironment for enhancing the catalysis efficiency in comparison to dispersed catalytic center molecule (TOF value was enhanced by about 15 fold). OPV-Ru could selectively accumulate in the mitochondria of living cells. Benefiting from its inherent fluorescence, the dynamic distribution in cells and uptake behavior of OPV-Ru could be visualized under fluorescence microscopy. This work represents the first demonstration of a multifunctional organometallic complex catalyzing natural hydrogenation transformation in specific subcellular compartments of living cells with excellent performance, fluorescent imaging ability, specific mitochondria targeting and good chemoselectivity with high catalysis efficiency.

3.
Invest New Drugs ; 38(1): 148-159, 2020 02.
Artigo em Inglês | MEDLINE | ID: mdl-31399906

RESUMO

Background The 5-year survival rate for extensive-disease small-cell lung carcinoma (ED-SCLC) is only 1%. Recently, apatinib exerted promising effects on cancer patients after failure of first-line chemotherapy. Methods This study enrolled 24 ED-SCLC patients to study the efficacy and toxicity of apatinib in combination with chemotherapy and maintenance therapy. The primary endpoints were overall survival (OS) and progression-free survival (PFS). The secondary endpoints included toxicity and safety. Apatinib was given 250 mg/day during the chemotherapy interval, and as maintenance therapy after 4-6 cycles until the patient progressed, died, or was intolerant to drug toxicity. The study further evaluated the cytotoxicity, cell-cycle arrest and apoptotic induction of apatinib in A549 and H446 cells. Results There was no difference in short-term efficacy between combined and chemotherapy groups. Long-term efficacy showed that the median PFS was 7.8 months and 4.9 months in combination and chemotherapy groups, respectively [p = 0.002, HR(95%CI): 0.18(0.06-0.60)]. The median OS was 12.1 months and 8.2 months in combination and chemotherapy groups, respectively [p = 0.023, HR(95%CI): 0.38 (0.16-0.90)]. Multivariate Cox regression analysis showed that apatinib combined with chemotherapy was an independent prognostic factor for OS and PFS. The ECOG score was an independent prognostic factor affecting OS. In vitro analysis showed that apatinib inhibited cell proliferation and caused cell-cycle arrest and apoptosis. Conclusion Apatinib combination/maintenance therapy showed promising efficacy and safety to extend OS/PFS in ED-SCLC and will be a potent therapeutic option in future practice. Although the scale of this study is small, further research on large sample sizes is needed.

4.
Nat Genet ; 52(1): 48-55, 2020 01.
Artigo em Inglês | MEDLINE | ID: mdl-31844323

RESUMO

R-loops are nucleic acid structures formed by an RNA:DNA hybrid and unpaired single-stranded DNA that represent a source of genomic instability in mammalian cells1-4. Here we show that N6-methyladenosine (m6A) modification, contributing to different aspects of messenger RNA metabolism5,6, is detectable on the majority of RNA:DNA hybrids in human pluripotent stem cells. We demonstrate that m6A-containing R-loops accumulate during G2/M and are depleted at G0/G1 phases of the cell cycle, and that the m6A reader promoting mRNA degradation, YTHDF2 (ref. 7), interacts with R-loop-enriched loci in dividing cells. Consequently, YTHDF2 knockout leads to increased R-loop levels, cell growth retardation and accumulation of γH2AX, a marker for DNA double-strand breaks, in mammalian cells. Our results suggest that m6A regulates accumulation of R-loops, implying a role for this modification in safeguarding genomic stability.


Assuntos
Adenosina/análogos & derivados , DNA/química , Instabilidade Genômica , Células-Tronco Pluripotentes/metabolismo , Estabilidade de RNA/efeitos dos fármacos , Proteínas de Ligação a RNA/fisiologia , RNA/química , Adenosina/farmacologia , Animais , DNA/efeitos dos fármacos , DNA/genética , Dano ao DNA , Humanos , Camundongos , Camundongos Knockout , Mitose , Células-Tronco Pluripotentes/citologia , RNA/efeitos dos fármacos , RNA/genética , RNA Mensageiro/química , RNA Mensageiro/genética , RNA Mensageiro/metabolismo
5.
Angew Chem Int Ed Engl ; 58(50): 17978-17985, 2019 Dec 09.
Artigo em Inglês | MEDLINE | ID: mdl-31589803

RESUMO

The synthesis of conjugated polymers with ionic substituents directly bound to their main chain repeat units is a strategy for generating strongly electron-accepting conjugated polyelectrolytes, as demonstrated through the synthesis of a series of ionic azaquinodimethane (iAQM) compounds. The introduction of cationic substituents onto the quinoidal para-azaquinodimethane (AQM) core gives rise to a strongly electron-accepting building block, which can be employed in the synthesis of ionic small molecules and conjugated polyelectrolytes (CPEs). Electrochemical measurements alongside theoretical calculations indicate notably low-lying LUMO values for the iAQMs. The optical band gaps measured for these compounds are highly tunable based on structure, ranging from 2.30 eV in small molecules down to 1.22 eV in polymers. The iAQM small molecules and CPEs showcase the band gap reduction effects of combining the donor-acceptor strategy with the bond-length alternation reduction strategy. As a demonstration of their utility, the iAQM CPEs so generated were used as active agents in photothermal therapy.

6.
J Biol Chem ; 294(48): 18220-18231, 2019 Nov 29.
Artigo em Inglês | MEDLINE | ID: mdl-31640989

RESUMO

Single-cell RNA-Seq (scRNA-Seq) has led to an unprecedented understanding of gene expression and regulation in individual cells. Many scRNA-Seq approaches rely upon the template switching property of Moloney murine leukemia virus (MMLV)-type reverse transcriptases. Template switching is believed to happen in a sequential process involving nontemplated addition of three protruding nucleotides (+CCC) to the 3'-end of the nascent cDNA, which can then anneal to the matching rGrGrG 3'-end of the template-switching oligo (TSO), allowing the reverse transcriptase (RT) to switch templates and continue copying the TSO sequence. In this study, we present a detailed analysis of template switching biases with respect to the RNA template, specifically of the role of the sequence and nature of its 5'-end (capped versus noncapped) in these biases. Our findings confirmed that the presence of a 5'-m7G cap enhances template switching efficiency. We also profiled the composition of the nontemplated addition in the absence of TSO and observed that the 5'-end of RNA template influences the terminal transferase activity of the RT. Furthermore, we found that designing new TSOs that pair with the most common nontemplated additions did little to improve template switching efficiency. Our results provide evidence suggesting that, in contrast to the current understanding of the template switching process, nontemplated addition and template switching are concurrent and competing processes.

7.
Neuroreport ; 30(16): 1074-1080, 2019 11 06.
Artigo em Inglês | MEDLINE | ID: mdl-31503209

RESUMO

OBJECTIVE: We previously showed differences in brain grey matter volume changes between patients with early-onset adult depression (EOD) and late-onset adult depression (LOD). Here, we aim to identify whether cortical thickness (CT) is affected by the age of onset in patients with depression. METHODS: High-resolution MRI images were obtained for 54 major depressive disorder (MDD) patients with EOD, 58 patients with LOD, 57 young healthy controls (HCs), and 58 aged HCs. Depression severity was assessed using the Hamilton Depression Rating Scale 17-item (HDRS17). Associations between CT of patients and clinical scores were analyzed. RESULTS: There was a significant main effect of diagnosis for the left rostal anterior cingulate (rACC), right inferior temporal, right lateral orbitofrontal cortex (lOFC), and bilateral pericalcarine. A remarkable onset age-group effect on CT was observed in the rACC and bilateral caudal anterior cingulate (cACC). The diagnosis-by-onset age interaction effect was found in bilateral rACC and right lOFC. Thinning CT in bilateral rACC was observed in EOD patients compared to young HCs. Compared to older HCs, thicker CT in lOFC was seen in the LOD patient group. Compared with the LOD group, the EOD group showed cortical thinning of the right cACC and posterior cingulate cortex (PCC). There were no significant associations between CT in right cACC or PCC with symptom severity or illness duration. CONCLUSIONS: MDD patients with different age at onset show distinct CT alterations, suggesting potentially divergent pathological mechanisms of EOD and LOD.

8.
Sci Rep ; 9(1): 8594, 2019 Jun 13.
Artigo em Inglês | MEDLINE | ID: mdl-31197197

RESUMO

Eukaryotic mRNAs are modified at their 5' end early during transcription by the addition of N7-methylguanosine (m7G), which forms the "cap" on the first 5' nucleotide. Identification of the 5' nucleotide on mRNA is necessary for determination of the Transcription Start Site (TSS). We explored the effect of various reaction conditions on the activity of the yeast scavenger mRNA decapping enzyme DcpS and examined decapping of 30 chemically distinct cap structures varying the state of methylation, sugar, phosphate linkage, and base composition on 25mer RNA oligonucleotides. Contrary to the generally accepted belief that DcpS enzymes only decap short oligonucleotides, we found that the yeast scavenger decapping enzyme decaps RNA transcripts as long as 1400 nucleotides. Further, we validated the application of yDcpS for enriching capped RNA using a strategy of specifically tagging the 5' end of capped RNA by first decapping and then recapping it with an affinity-tagged guanosine nucleotide.

9.
Front Psychiatry ; 10: 418, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31249539

RESUMO

Background: Neuroimaging studies have shown that the high synchrony of spontaneous neural activity in the homotopic regions between hemispheres is an important functional structural feature of normal human brains, and this feature is abnormal in the patients with various mental disorders. However, little is known about this feature in obsessive-compulsive disorder (OCD). This study aimed to further analyze the underlying neural mechanisms of OCD and to explore whether clinical characteristics are correlated with the alerted homotopic connectivity in patients with OCD. Methods: Using voxel-mirrored homotopic connectivity (VMHC) during resting state, we compared 46 OCD patients and 46 healthy controls (HCs) matched for age, gender, and education level. A partial correlation analysis was used to investigate the relationship between altered VMHC and clinical characteristics in patients with OCD. Results: Patients with OCD showed lower VMHC than HCs in fusiform gyrus/inferior occipital gyrus, lingual gyrus, postcentral gyrus/precentral gyrus, putamen, and orbital frontal gyrus. A significant positive correlation was observed between altered VMHC in the angular gyrus/middle occipital gyrus and illness duration in patients. Conclusions: Interhemispheric functional imbalance may be an essential aspect of the pathophysiological mechanism of OCD, which is reflected not only in the cortico-striato-thalamo-cortical (CSTC) loop but also elsewhere in the brain.

10.
Cancer Manag Res ; 11: 2605-2616, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31114327

RESUMO

Background: Postoperative recurrence is the main cause of a poor prognosis in early-stage lung adenocarcinoma (LUAD). Factors that can predict recurrence risk are critically needed. Materials and methods: In this study, we designed a screening procedure based on gene profile data and performed validation using TCGA and Daping hospital's cohorts. Differentially expressed genes (DEGs) between patients with recurrence-free survival (RFS) <1 year and RFS >3 years were identified, overlapping genes among these DEGs were selected as candidate biomarkers. A Cox proportional hazards model, immunohistochemistry and Kaplan-Meier survival analysis were performed to validate these biomarkers in two distinct validation sets. Results: SFTPB, SFTPD, SFTA1P, HLA-DQB1, ITGB8, ANLN, and LRRN1 were overlapped both in TCGA and Daping discovery sets. The Cox proportional hazards model analysis of the TCGA validation set showed that HLA-DQB1 was an independent prognostic factor for RFS (HR=0.686, 95% CI, 0.542-0.868). Immunohistochemistry and Kaplan-Meier analysis in Daping validation sets confirmed HLA-DQB1 expression on tumor cells (not interstitial cells) to be an effective predictor of postoperative recurrence. Further examination revealed that the level of HLA-DQB1 expression on tumor cells was positively correlated with CD4- and CD8-positive lymphocyte infiltration into the tumor. Conclusion: All results indicate that high expression of HLA-DQB1 on tumor cells is a good prognostic marker in early-stage LUAD, and the mechanism may be related to anti-tumor immune activity.

11.
Comput Struct Biotechnol J ; 17: 324-332, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-30906512

RESUMO

Angiogenesis is essential for tumor growth. Vascular endothelial growth factor (VEGF), a crucial factor in tumor angiogenesis, has been reported to be transcriptionally regulated by hypoxia-inducible factor-1 (HIF-1). An 8-oxo-G or apurinic/apyrimidinic (AP) site, which is frequently associated with DNA damage, has been identified in the promoter region of VEGF. However, the detailed molecular mechanisms by which AP sites regulate VEGF gene transcription are largely unknown. The dual functional protein apurinic/apyrimidinic endonuclease 1 (APE1) is both the key enzyme in DNA base excision repair and the redox factor shown to regulate HIF-1 DNA-binding activity. In the present study, we tested the involvement of both the AP endonuclease and redox activity of APE1 in regulating HIF-1 DNA binding and VEGF transcription in HUVECs. By employing two APE1 activity-specific inhibitors and AP-site-containing reporter constructs, we confirmed that both activities of APE1 were involved in regulating VEGF expression under hypoxic conditions. Furthermore, we found that the interaction between APE1 and its downstream repair enzyme, DNA polymerase ß, was compromised when the N-terminal structure of APE1 was distorted under oxidative conditions. Our data suggest that the DNA repair and redox activity of APE1 can play a collaborative role in regulating the transcriptional initiation of the AP-site-containing promoter.

12.
Psychiatry Res Neuroimaging ; 286: 4-10, 2019 04 30.
Artigo em Inglês | MEDLINE | ID: mdl-30822678

RESUMO

Considerable evidence suggests that the tryptophan hydroxylase-2 (TPH2) gene is associated with the pathophysiology of major depressive disorder (MDD). In the present study, we investigated alterations of white matter (WM) integrity and the impact of TPH2 polymorphism on WM in a sample of 118 first-episode, medication-naïve, MDD patients and 118 well-matched healthy controls. Whole brain analyses of fractional anisotropy (FA) were performed using tract-based spatial statistics (TBSS). The results showed that the MDD group had significantly reduced FA values for the genu and body of the corpus callosum (CC) and the bilateral anterior corona radiate (ACR). In the MDD patient group, the GG homozygote subgroup exhibited a widespread reduction of FA (uncorrected) and significantly reduced FA in the left retrolenticular portion of the internal capsule and left superior longitudinal fasciculus (SLF) compared with those of the T carriers (GT/TT) (FWE corrected). No significant correlation was found between the FA values in any brain region and the patients' clinical variables. Our findings demonstrate the presence of abnormal white matter integrity in untreated patients with first-episode depression. TPH2-rs4570625 polymorphisms may be involved in the pathological mechanism of WM microarchitecture in patients.


Assuntos
Corpo Caloso/diagnóstico por imagem , Transtorno Depressivo Maior/diagnóstico por imagem , Transtorno Depressivo Maior/genética , Polimorfismo de Nucleotídeo Único/genética , Triptofano Hidroxilase/genética , Substância Branca/diagnóstico por imagem , Adulto , Corpo Caloso/enzimologia , Transtorno Depressivo Maior/enzimologia , Imagem de Tensor de Difusão/métodos , Feminino , Humanos , Cápsula Interna/diagnóstico por imagem , Cápsula Interna/enzimologia , Masculino , Pessoa de Meia-Idade , Triptofano Hidroxilase/metabolismo , Substância Branca/enzimologia , Adulto Jovem
13.
Chin Med J (Engl) ; 131(24): 2938-2946, 2018 Dec 20.
Artigo em Inglês | MEDLINE | ID: mdl-30539906

RESUMO

Background: Decision-making concerning the treatment of choroid plexus tumor (CPT) in pediatric patients remains a topic of considerable debate. The aim of this work was to describe clinical features and prognostic risk factors of CPT in the pediatric population and to provide theoretical opinions regarding clinical decisions for CPT. Methods: The data of 96 patients with CPT and younger than 14 years were retrospectively analyzed. Clinical characteristics such as pathological type of CPTs, rate and severity of hydrocephalus, treatment and outcome, and recurrence were investigated. For categorical variables, the Pearson's Chi-square test was performed. The Mann-Whitney U-test was used for comparisons between nonnormally distributed parameters. Log-rank test was used for progression-free survival (PFS). Results: The study included 70 choroid plexus papilloma (CPP) cases, 17 atypical choroid plexus papilloma (aCPP) cases, and 9 choroid plexus carcinoma (CPC) cases. Compared with patients with CPP or aCPP, patients with CPC had a shorter disease course (median: CPP, 4 months; aCPP, 2 months; CPC, 1 month; H: 23.5, P < 0.001), higher rate of acute hydrocephalus (CPP, 27.1%; aCPP, 52.9%; CPC, 77.8%; χ2 = 10.9, P < 0.05), and lower incidence of cure rate (CPP, 85.7%; aCPP, 70.5%; CPC, 33.3%; χ2 = 13.5, P < 0.05). The severity of hydrocephalus with tumor in the lateral or third ventricle was significantly higher than that with tumors in the fourth ventricle (severe hydrocephalus: lateral ventricle, 51.7%; third ventricle, 47.0%; fourth ventricle, 11.1%; χ2 = 26.0, P < 0.001). Patients with gross total surgical resection had no better PFS than those with partial resection because of the use of adjuvant therapy in the latter (χ2 = 4.0, P > 0.05). Patients with CPC experienced shorter time for recurrence than those with CPP or aCPP (χ2 = 40.1, P < 0.0001). Conclusions: Our results indicated that CPP in the fourth ventricle could trigger serious clinical symptoms at an early stage, requiring early intervention. Adjuvant treatment might be necessary for patients with partially resected CPP, aCPP, and CPC to achieve a favorable outcome.


Assuntos
Neoplasias do Plexo Corióideo/mortalidade , Criança , Pré-Escolar , Neoplasias do Plexo Corióideo/patologia , Neoplasias do Plexo Corióideo/cirurgia , Feminino , Humanos , Hidrocefalia/etiologia , Lactente , Masculino , Metástase Neoplásica , Recidiva Local de Neoplasia , Prognóstico , Fatores de Risco
14.
Cancer Med ; 7(9): 4406-4419, 2018 09.
Artigo em Inglês | MEDLINE | ID: mdl-30109782

RESUMO

BACKGROUND: Epithelial-to-mesenchymal transition (EMT) plays a pivotal role in resistance to EGFR tyrosine kinase inhibitors (TKIs) in non-small-cell lung cancer (NSCLC). Our previous study revealed that in osteosarcoma, human apurinic/apyrimidinic endonuclease 1 (APE1) regulates transforming growth factor-ß (TGF-ß), an important player in EMT. We therefore hypothesized a link between APE1 and EGFR-TKI responsiveness in NSCLC. METHODS: The protein levels of APE1 were analyzed in tumors of NSCLC patients receiving EGFR-TKI treatment. The correlation between APE1 expression and progression-free survival (PFS), overall survival (OS), or response rate were analyzed. The impact of APE1 on the response to EGFR-TKIs was measured by exogenous manipulation of APE1 in EGFR-TKI-sensitive and EGFR-TKI-resistant cells. RESULTS: We indicate that low expression of APE1 in tumors is associated with a significantly longer PFS (20.8 months vs 8.4 months, P = 0.008) and a preferential OS (39.0 months vs 17.0 months, P = 0.001), with no difference in initial response rate to EGFR-TKIs. We observed that APE1 protein level was significantly increased in EGFR-TKI-resistant cells and was associated with downregulated E-cadherin and upregulated vimentin. The EMT phenotype, as well as the levels of TGF-ß, was suppressed in APE1 knockdown HCC827/IR and PC-9/ER cells, while the EMT phenotype was promoted in APE1-overexpressed HCC827 and PC-9 cells. Furthermore, a specific APE1 redox inhibitor (ie, E3330) effectively reversed the EMT phenotype and further sensitized the cells to EGFR-TKIs. CONCLUSION: This study revealed a significant role of APE1 in EGFR-TKI resistance via novel regulatory effects on the EMT phenotype in NSCLC.


Assuntos
Carcinoma Pulmonar de Células não Pequenas/genética , Carcinoma Pulmonar de Células não Pequenas/patologia , DNA Liase (Sítios Apurínicos ou Apirimidínicos)/genética , Transição Epitelial-Mesenquimal/genética , Receptores ErbB/antagonistas & inibidores , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/patologia , Adulto , Idoso , Animais , Antineoplásicos/farmacologia , Biomarcadores , Carcinoma Pulmonar de Células não Pequenas/tratamento farmacológico , Receptores ErbB/genética , Feminino , Deleção de Genes , Genômica/métodos , Sequenciamento de Nucleotídeos em Larga Escala , Humanos , Neoplasias Pulmonares/tratamento farmacológico , Masculino , Camundongos , Pessoa de Meia-Idade , Estadiamento de Neoplasias , Inibidores de Proteínas Quinases/farmacologia , Transdução de Sinais
15.
PeerJ ; 6: e5119, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-30002967

RESUMO

Insects provide an accessible system to study the contribution of DNA methylation to complex epigenetic phenotypes created to regulate gene expression, chromatin states, imprinting and dosage compensation. The members of genus Drosophila have been used as a model system to study aspects of biology like development, behaviour and genetics. Despite the popularity of Drosophila melanogaster as a genetic and epigenetic model organism, DNA methylation studies are limited due to low levels of genomic 5-methylcytosine. Our study employs a sensitive liquid chromatography-mass spectrometry (LCMS) based method to quantify the levels of 5-methylcytosine from the genomic DNA in different members of the genus Drosophila. Our results reveal that, despite being phylogenetically related, there is a marked variation in the levels of 5-methylcytosine between the genomes of the members of genus Drosophila. Also, there is a change in the genomic levels of 5-methylcytosine through each life cycle stage of holometabolous development in D. melanogaster.

16.
Oncotarget ; 9(39): 25414-25426, 2018 May 22.
Artigo em Inglês | MEDLINE | ID: mdl-29875998

RESUMO

Radiotherapy in osteosarcoma patients is problematic due to radioresistance; therefore, understanding the mechanism of radioresistance is integral to providing effective radiotherapeutic regimens for osteosarcoma. We now report the activity of an miRNA, miR-513a-5p, in stimulating radiosensitivity of osteosarcoma cells in vitro and in vivo. MiR-513a-5p expression is decreased in osteosarcoma tissue from patients and cultured osteosarcoma cell lines. However, exogenous re-expression of this miRNA in osteosarcoma cell lines, including HOS, U2OS and 9901, can induce sensitization to ionizing radiation. We also confirm that miR-513a-5p suppresses APE1 expression, and that both the redox and DNA repair activity of APE1 were decreased in miR-513a-5p expressing cell lines. By suppressing APE1, miR-513a-5p induces the DNA damage response which stimulates apoptosis after irradiation. Our report establishes miR-513a-5p as a radiosensitizing miRNA and identifies its activity in the suppression of APE1, which could directly lead to radiosensitization.

17.
Nucleic Acids Res ; 46(11): 5753-5763, 2018 06 20.
Artigo em Inglês | MEDLINE | ID: mdl-29750267

RESUMO

Ribonucleic acid (RNA) is capable of hosting a variety of chemically diverse modifications, in both naturally-occurring post-transcriptional modifications and artificial chemical modifications used to expand the functionality of RNA. However, few studies have addressed how base modifications affect RNA polymerase and reverse transcriptase activity and fidelity. Here, we describe the fidelity of RNA synthesis and reverse transcription of modified ribonucleotides using an assay based on Pacific Biosciences Single Molecule Real-Time sequencing. Several modified bases, including methylated (m6A, m5C and m5U), hydroxymethylated (hm5U) and isomeric bases (pseudouridine), were examined. By comparing each modified base to the equivalent unmodified RNA base, we can determine how the modification affected cumulative RNA polymerase and reverse transcriptase fidelity. 5-hydroxymethyluridine and N6-methyladenosine both increased the combined error rate of T7 RNA polymerase and reverse transcriptases, while pseudouridine specifically increased the error rate of RNA synthesis by T7 RNA polymerase. In addition, we examined the frequency, mutational spectrum and sequence context of reverse transcription errors on DNA templates from an analysis of second strand DNA synthesis.


Assuntos
RNA Polimerases Dirigidas por DNA/metabolismo , DNA Polimerase Dirigida por RNA/metabolismo , RNA/química , Proteínas Virais/metabolismo , Sequência de Bases , DNA Complementar/biossíntese , DNA Complementar/química , RNA/biossíntese , Transcrição Reversa , Ribonucleotídeos/química , Ribonucleotídeos/metabolismo , Transcrição Genética
18.
Nucleic Acids Res ; 46(11): 5664-5677, 2018 06 20.
Artigo em Inglês | MEDLINE | ID: mdl-29750271

RESUMO

Base excision repair (BER) handles many forms of endogenous DNA damage, and apurinic/apyrimidinic endonuclease 1 (APE1) is central to this process. Deletion of both alleles of APE1 (a.k.a. Apex1) in mice leads to embryonic lethality, and deficiency in cells can promote cell death. Unlike most other BER proteins, APE1 expression is inversely correlated with cellular senescence in primary human fibroblasts. Depletion of APE1 via shRNA induced senescence in normal human BJ fibroblasts, a phenotype that was not seen in counterpart cells expressing telomerase. APE1 knock-down in primary fibroblasts resulted in global DNA damage accumulation, and the induction of p16INK4a and p21WAF1 stress response pathways; the DNA damage response, as assessed by γ-H2AX, was particularly pronounced at telomeres. Conditional knock-out of Apex1 in mice at post-natal day 7/12 resulted in impaired growth, reduced organ size, and increased cellular senescence. The effect of Apex1 deletion at post-natal week 6 was less obvious, other than cellular senescence, until ∼8-months of age, when premature aging characteristics, such as hair loss and impaired wound healing, were seen. Low APE1 expression in patient cancer tissue also correlated with increased senescence. Our results point to a key role for APE1 in regulating cellular senescence and aging features, with telomere status apparently affecting the outcome.


Assuntos
Senescência Celular , DNA Liase (Sítios Apurínicos ou Apirimidínicos)/metabolismo , Senilidade Prematura/genética , Animais , Células Cultivadas , Dano ao DNA , Reparo do DNA , DNA Liase (Sítios Apurínicos ou Apirimidínicos)/deficiência , DNA Liase (Sítios Apurínicos ou Apirimidínicos)/genética , Fibroblastos/metabolismo , Camundongos Knockout , Telomerase/metabolismo , Telômero/metabolismo
19.
Proc Natl Acad Sci U S A ; 115(14): E3116-E3125, 2018 04 03.
Artigo em Inglês | MEDLINE | ID: mdl-29555775

RESUMO

Certain viruses of bacteria (bacteriophages) enzymatically hypermodify their DNA to protect their genetic material from host restriction endonuclease-mediated cleavage. Historically, it has been known that virion DNAs from the Delftia phage ΦW-14 and the Bacillus phage SP10 contain the hypermodified pyrimidines α-putrescinylthymidine and α-glutamylthymidine, respectively. These bases derive from the modification of 5-hydroxymethyl-2'-deoxyuridine (5-hmdU) in newly replicated phage DNA via a pyrophosphorylated intermediate. Like ΦW-14 and SP10, the Pseudomonas phage M6 and the Salmonella phage ViI encode kinase homologs predicted to phosphorylate 5-hmdU DNA but have uncharacterized nucleotide content [Iyer et al. (2013) Nucleic Acids Res 41:7635-7655]. We report here the discovery and characterization of two bases, 5-(2-aminoethoxy)methyluridine (5-NeOmdU) and 5-(2-aminoethyl)uridine (5-NedU), in the virion DNA of ViI and M6 phages, respectively. Furthermore, we show that recombinant expression of five gene products encoded by phage ViI is sufficient to reconstitute the formation of 5-NeOmdU in vitro. These findings point to an unexplored diversity of DNA modifications and the underlying biochemistry of their formation.


Assuntos
Bactérias/metabolismo , Infecções Bacterianas/microbiologia , Proteínas de Bactérias/metabolismo , Bacteriófagos/genética , DNA Viral/biossíntese , Timidina/química , Uridina/química , Bacteriófagos/crescimento & desenvolvimento , Bacteriófagos/metabolismo , Genoma Viral
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