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1.
Clin Cancer Res ; 2021 Sep 15.
Artigo em Inglês | MEDLINE | ID: mdl-34526363

RESUMO

PURPOSE: Targeted therapy and immunotherapy are transforming the treatment approach for intrahepatic cholangiocarcinoma (ICC). However, little is known about the intertumor heterogeneity (ITH) of multifocal ICC and its impacts on patient response to these treatments. We aimed to characterize the immunogenomic and epigenomic heterogeneity of multifocal ICC to guide treatment decision making. EXPERIMENTAL DESIGN: We obtained 66 tumor samples from 16 patients with multifocal ICC and characterized the tumor and immune heterogeneity using whole-exome sequencing, bulk and single-cell RNA sequencing, methylation microarray, and multiplex immunostaining. Patients were divided into high- or low-ITH groups according to the median ITH index. Two independent cohorts were used to validate findings. Responses to anti-PD-1 therapy were assessed. RESULTS: Multifocal ICC presented considerable intertumor genomic, transcriptional, and epigenomic heterogeneity within a patient in high ITH group. The immune profile among multiple tumors within a patient was relatively less heterogeneous in high- or low-ITH group, and consistent responses of multiple tumors to anti-PD-1 immunotherapy were observed. Unsupervised clustering of immune markers identified one low and one high immune subtype, with higher immune cell infiltration, closer tumor-immune cell interactions, and upregulated IFN-signature expression in high-immune subtype. Determining expression levels of CD8B and ICOS facilitated this immune classification and prediction of patient prognosis. Finally, promoter DNA methylation contributed to different immune profiles of two subtypes by regulating immune-gene expression. CONCLUSIONS: There is comprehensive heterogeneity in the genome, transcriptome, and epigenome of multifocal ICC. On the basis of the less heterogeneous immune profile of ICC, we suggest an immune classification that stratifies patients' prognosis and may support personalized immunotherapy.

2.
Hepatology ; 74(3): 1339-1356, 2021 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-33638162

RESUMO

BACKGROUND AND AIMS: The dynamic N6-methyladenosine (m6 A) mRNA modification is essential for acute stress response and cancer progression. Sublethal heat stress from insufficient radiofrequency ablation (IRFA) has been confirmed to promote HCC progression; however, whether m6 A machinery is involved in IRFA-induced HCC recurrence remains open for study. APPROACH AND RESULTS: Using an IRFA HCC orthotopic mouse model, we detected a higher level of m6 A reader YTH N6-methyladenosine RNA binding protein 1-3 (YTHDF1) in the sublethal-heat-exposed transitional zone close to the ablation center than that in the farther area. In addition, we validated the increased m6 A modification and elevated YTHDF1 protein level in sublethal-heat-treated HCC cell lines, HCC patient-derived xenograft (PDX) mouse model, and patients' HCC tissues. Functionally, gain-of-function/loss-of-function assays showed that YTHDF1 promotes HCC cell viability and metastasis. Knockdown of YTHDF1 drastically restrains the tumor metastasis evoked by sublethal heat treatment in tail vein injection lung metastasis and orthotopic HCC mouse models. Mechanistically, we found that sublethal heat treatment increases epidermal factor growth receptor (EGFR) m6 A modification in the vicinity of the 5' untranslated region and promotes its binding with YTHDF1, which enhances the translation of EGFR mRNA. The sublethal-heat-induced up-regulation of EGFR level was further confirmed in the IRFA HCC PDX mouse model and patients' tissues. Combination of YTHDF1 silencing and EGFR inhibition suppressed the malignancies of HCC cells synergically. CONCLUSIONS: The m6 A-YTHDF1-EGFR axis promotes HCC progression after IRFA, supporting the rationale for targeting m6 A machinery combined with EGFR inhibitors to suppress HCC metastasis after RFA.

3.
Cell Physiol Biochem ; 48(3): 891-904, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-30032137

RESUMO

BACKGROUND/AIMS: Long non-coding RNAs (lncRNAs) have emerged as new regulators and biomarkers in several cancers. However, few lncRNAs have been well characterized in clear cell renal cell carcinoma (ccRCC). METHODS: We investigated the lncRNA expression profile by microarray analysis in 5 corresponding ccRCC tissues and adjacent normal tissues. Lung cancer-associated transcript 1 (LUCAT1) expression was examined in 90 paired ccRCC tissues by real-time PCR and validated by The Cancer Genome Atlas (TCGA) database. Kaplan-Meier analysis was used to examine the prognostic value of LUCAT1 and CXCL2 in ccRCC patients. Loss and gain of function were performed to explore the effect of LUCAT1 on proliferation and invasion in ccRCC cells. Western blotting was performed to evaluate the underlying mechanisms of LUCAT1 in ccRCC progression. Chemokine stimulation assay was performed to investigate possible mechanisms controlling LUCAT1 expression in ccRCC cells. Enzyme-linked immunosorbent assays were performed to determine serum CXCL2 in ccRCC patients and healthy volunteers. Receiver operating characteristic curve analysis was performed to examine the clinical diagnostic value of serum CXCL2 in ccRCC. RESULTS: We found that LUCAT1 was significantly upregulated in both clinical ccRCC tissues (n = 90) and TCGA ccRCC tissues (n = 448) compared with normal tissues. Statistical analysis revealed that the LUCAT1 expression level positively correlated with tumor T stage (P < 0.01), M stage (P < 0.01), and TNM stage (P < 0.01). Overall survival and disease-free survival time were significantly shorter in the high-LUCAT1-expression group than in the low-LUCAT1-expression group (log-rank P < 0.01). LUCAT1 knockdown inhibited ccRCC cell proliferation and colony formation, induced cell cycle arrest at G1 phase, and inhibited cell migration and invasion. Overexpression of LUCAT1 promoted proliferation, migration, and invasion of ccRCC cells. Mechanistic investigations showed that LUCAT1 induced cell cycle G1 arrest by regulating the expression of cyclin D1, cyclin-dependent kinase 4, and phosphorylated retinoblastoma transcriptional corepressor 1. Moreover, LUCAT1 promoted proliferation and invasion in ccRCC cells partly through inducing the phosphorylation of AKT and suppressing the phosphorylation of GSK-3ß. We also revealed that chemokine CXCL2, upregulated in ccRCC, induced LUCAT1 expression and might be a diagnostic and prognostic biomarker in ccRCC. CONCLUSIONS: LUCAT1 was upregulated in ccRCC tissues and renal cancer cell lines, and significantly correlated with malignant stage and poor prognosis in ccRCC. LUCAT1 promoted proliferation and invasion in ccRCC cells through the AKT/GSK-3ß signaling pathway. We also revealed that LUCAT1 overexpression was induced by chemokine CXCL2. These findings indicate that the CXCL2/LUCAT1/AKT/GSK-3ß axis is a potential therapeutic target and molecular biomarker for ccRCC.


Assuntos
Carcinoma de Células Renais/diagnóstico , Glicogênio Sintase Quinase 3 beta/metabolismo , Neoplasias Renais/diagnóstico , Invasividade Neoplásica/genética , Proteínas Proto-Oncogênicas c-akt/metabolismo , RNA Longo não Codificante/metabolismo , Transdução de Sinais/genética , Biomarcadores Tumorais/genética , Carcinoma de Células Renais/metabolismo , Carcinoma de Células Renais/mortalidade , Linhagem Celular Tumoral , Movimento Celular , Proliferação de Células/genética , Quimiocina CXCL2/sangue , Feminino , Humanos , Estimativa de Kaplan-Meier , Neoplasias Renais/metabolismo , Neoplasias Renais/mortalidade , Masculino , Pessoa de Meia-Idade , Estadiamento de Neoplasias , Interferência de RNA , RNA Longo não Codificante/antagonistas & inibidores , RNA Longo não Codificante/genética , RNA Interferente Pequeno/metabolismo
4.
Oncol Lett ; 15(5): 6770-6776, 2018 May.
Artigo em Inglês | MEDLINE | ID: mdl-29725414

RESUMO

Previous studies have demonstrated that long noncoding RNAs (lncRNAs) exhibit critical regulatory roles in cancer biology. However, few lncRNAs have been well characterized in bladder cancer. In the previous study, we demonstrated that gastric adenocarcinoma associated, positive CD44 regulator, long intergenic noncoding RNA (GAPLINC) was significantly upregulated in bladder cancer tissues compared with normal tissues in The Cancer Genome Atlas (TCGA) cohort (P=0.039) and a validated cohort of 80 patients with bladder cancer (P=0.021). Statistical analysis revealed that GAPLINC expression level was associated with tumor stage in the validated cohort (P=0.017). Kaplan-Meier analysis demonstrated that patients in the high GAPLINC expression group had a worse overall survival (P=0.0386), indicating that GAPLINC may be a sensitive prognostic biomarker for patients with bladder cancer. Furthermore, knockdown of GAPLINC inhibited cell proliferation and colony formation, promoted cells cycle arrest at G1 phase and suppressed cells migration and invasion. The findings of the present study suggest that GAPLINC exhibits an oncogenic role in bladder cancer and may be a potential prognostic biomarker and therapeutic target.

5.
Oncol Lett ; 15(5): 7855-7863, 2018 May.
Artigo em Inglês | MEDLINE | ID: mdl-29725475

RESUMO

Previous years have witnessed the importance of long non-coding RNAs (lncRNAs) in cancer research. The lncRNA Pvt1 oncogene (non-protein coding) (PVT1) was revealed to be upregulated in various cancer types. The aim of the present study was to investigate the function of PVT1 in clear cell renal cell carcinoma (ccRCC). The expression of PVT1 in ccRCC was analyzed using reverse transcription-quantitative polymerase chain reaction, and it was revealed that PVT1 expression was upregulated in ccRCC tissues compared with that in normal adjacent tissues. Next, PVT1 expression from The Cancer Genome Atlas datasets was validated, and it was also revealed that the high expression of PVT1 was associated with advanced disease stage and a poor prognosis. Furthermore, the knockdown of PVT1 induced apoptosis by increasing the expression of poly ADP ribose polymerase and Bcl-2-associated X protein, and promoted cell cycle arrest at the G1 phase by decreasing the expression of cyclin D1. Study of the mechanism involved indicated that PVT1 promoted the progression of ccRCC partly through activation of the epidermal growth factor receptor pathway. Altogether, the results of the present study suggested that PVT1 serves oncogenic functions and may be a biomarker and therapeutic target in ccRCC.

6.
Front Oncol ; 8: 682, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-30723697

RESUMO

The chemokine ligands and their receptors play critical roles in cancer progression and patients outcomes. We found that CXCL13 was significantly upregulated in ccRCC tissues compared with normal tissues in both The Cancer Genome Atlas (TCGA) cohort and a validated cohort of 90 pairs ccRCC tissues. Statistical analysis showed that high CXCL13 expression related to advanced disease stage and poor prognosis in ccRCC. We also revealed that serum CXCL13 levels in ccRCC patients (n = 50) were significantly higher than in healthy controls (n = 40). Receiver operating characteristic (ROC) curve revealed that tissue and serum CXCL13 expression might be a diagnostic biomarker for ccRCC with an area under curve (AUC) of 0.809 and 0.704, respectively. CXCL13 was significantly associated with its receptor, CXCR5, in ccRCC tissues, and ccRCC patients in high CXCL13 high CXCR5 expression group have a worst prognosis. Functional and mechanistic study revealed that CXCL13 promoted the proliferation and migration of ccRCC cells by binding to CXCR5 and activated PI3K/AKT/mTOR signaling pathway. These results suggested that CXCL13/CXCR5 axis played a significant role in ccRCC and might be a therapeutic target and prognostic biomarker.

7.
Int J Mol Sci ; 17(1)2015 Dec 23.
Artigo em Inglês | MEDLINE | ID: mdl-26703590

RESUMO

Basic fibroblast growth factor (bFGF) is a multifunctional factor in acceleration of cell proliferation, differentiation and transference, and therefore widely used in clinical applications. In this study, expression vector pWX-Nt03 harboring a codon-optimized bFGF gene was constructed and introduced into the tobacco chloroplasts by particle bombardment. After four rounds of selection, bFGF was proved to integrate into the chloroplast genome of regenerated plants and two of four transgenic plants were confirmed to be homoplastomic by PCR and Southern hybridization. ELISA assay indicated that bFGF represented approximately 0.1% of total soluble protein in the leaves of transplastomic tobacco plants. This is the first report of bFGF expression via chloroplast transformation in model plant, providing an additional option for the production of chloroplast-produced therapeutic proteins.


Assuntos
Cloroplastos/genética , Fator 2 de Crescimento de Fibroblastos/genética , Plantas Geneticamente Modificadas/genética , Tabaco/genética , Cloroplastos/metabolismo , Fator 2 de Crescimento de Fibroblastos/metabolismo , Genoma de Cloroplastos , Instabilidade Genômica , Humanos , Plantas Geneticamente Modificadas/metabolismo , Recombinação Genética , Tabaco/metabolismo
8.
Genet Mol Biol ; 33(2): 325-7, 2010 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-21637489

RESUMO

A triploid (2n = 3x = 36) rice plant was obtained by screening a twin seedling population in which each seed germinated to two or three sprouts that were then crossed with diploid plants. One diploid plant was chosen among the various F(1) progenies and developed into an F (2) population via self-pollination. Compared with the control variety Shanyou 63, this F (2) population had a stable agronomical performance in field trials, as confirmed by the F-test. The stability of the F (2) population was further substantiated by molecular analysis with simple sequence repeat markers. Specifically, of 160 markers assayed, 37 (covering all 12 chromosomes) were polymorphic between the parental lines. Testing the F (1) hybrid individually with these markers showed that each PCR product had only a single band instead of two bands from each parent. The bands were identical to either maternal (23 markers) or paternal (eight markers) bands or distinct from both parents (six markers). The amplified bands of all 60 randomly selected F (2) plants were uniform and identical to those of the F (1) hybrid. These results suggest that the F (1) plant is a non-segregating hybrid and that a stable F (2) population was obtained. This novel system provides an efficient means for shortening the cycle of hybrid rice seed production.

9.
Genet. mol. biol ; 33(2): 325-327, 2010. ilus, tab
Artigo em Inglês | LILACS-Express | LILACS | ID: lil-548810

RESUMO

A triploid (2n = 3x = 36) rice plant was obtained by screening a twin seedling population in which each seed germinated to two or three sprouts that were then crossed with diploid plants. One diploid plant was chosen among the various F1 progenies and developed into an F2 population via self-pollination. Compared with the control variety Shanyou 63, this F2 population had a stable agronomical performance in field trials, as confirmed by the F-test. The stability of the F2 population was further substantiated by molecular analysis with simple sequence repeat markers. Specifically, of 160 markers assayed, 37 (covering all 12 chromosomes) were polymorphic between the parental lines. Testing the F1 hybrid individually with these markers showed that each PCR product had only a single band instead of two bands from each parent. The bands were identical to either maternal (23 markers) or paternal (eight markers) bands or distinct from both parents (six markers). The amplified bands of all 60 randomly selected F2 plants were uniform and identical to those of the F1 hybrid. These results suggest that the F1 plant is a non-segregating hybrid and that a stable F2 population was obtained. This novel system provides an efficient means for shortening the cycle of hybrid rice seed production.

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