circEZH2E2 /E3 is a dual suppressor of miR363/miR708 to promote EZH2 expression and prostate cancer progression.

The histone methyltransferase enhancer of zeste homolog 2 (EZH2) is overexpressed in a variety of malignancies including prostate cancer (PCa) and may play important roles in tumor progression. Gene copy number gains, enhanced transcription, and a few circRNAs have been reported to upregulate EZH2. It was not known whether EZH2 itself generates circRNAs that promote its own expression. We here report the identification of circEZH2E2/E3 that is derived from exons 2 and 3 of the EZH2 gene and overexpressed in PCa. We show that circEZH2E2/E3 functions as a dual inhibitor for both miR363 and miR708 that target the EZH2 3'UTR and CDS, respectively, resulting in the upregulation of EZH2 expression and hence the downregulation of EZH2-repressed genes (e.g., CDH1 and DAB2IP), and enhancement of PCa cell proliferation, migration, invasion, and xenograft PCa growth. Overexpression of circEZH2E2/E3 is significantly correlated with higher tumor grade, tumor progression, and unfavorable progression-free and disease-specific survival in PCa patients. These findings show a novel autoenhancing EZH2-circEZH2E2/E3 -miR363/miR708-EZH2 regulatory loop, by which circEZH2E2/E3 plays important roles in PCa tumorigenesis and progression by upregulating EZH2, and may have potential diagnostic, prognostic, and therapeutic uses in PCa management.

SOX9 and HMGB3 co-operatively transactivate NANOG and promote prostate cancer progression.

BACKGROUND: The homeodomain-containing transcription factor NANOG is overexpressed in prostate adenocarcinoma (PCa) and predicts poor prognosis. The SOX family transcription factor SOX9, as well as the transcription co-activator HMGB3 of the HMGB family, are also overexpressed and may play pivotal roles in PCa. However, it is unknown whether SOX9 and HMGB3 interact with each other, or if they regulate NANOG gene transcription. METHODS: We identified potential SOX9 responsive elements in NANOG promoter, and investigated if SOX9 regulated NANOG transcription in co-operation with HMGB3 by experimental analysis of potential SOX9 binding sites in NANOG promoter, reporter gene transcription assays with or without interference or artificial overexpression of SOX9 and/or HMGB3, and protein-binding assays of SOX9-HMGB3 interaction. Clinicopathologic and prognostic significance of SOX9-HMGB3 overexpression in PCa was analyzed. RESULTS: SOX9 activated NANOG gene transcription by preferentially binding to a highly conserved consensus cis-regulatory element (-573 to -568) in NANOG promoter, and promoted the expression of NANOG downstream oncogenic genes. Importantly, HMGB3 functioned as a partner of SOX9 to co-operatively enhance transactivation of NANOG by interacting with SOX9, predominantly via the HMG Box A domain of HMGB3. Overexpression of SOX9 and/or HMGB3 enhanced PCa cell survival and cell migration and were significantly associated with PCa progression. Notably, Cox proportional regression analysis showed that co-overexpression of both SOX9 and HMGB3 was an independent unfavorable prognosticator for both CRPC-free survival (relative risk [RR] = 3.779，95% confidence interval [CI]: 1.159-12.322, p = 0.028) and overall survival (RR = 3.615，95% CI: 1.101-11.876, p = 0.034). CONCLUSIONS: These findings showed a novel SOX9/HMGB3/NANOG regulatory mechanism, deregulation of which played important roles in PCa progression.

A novel intronic circular RNA circFGFR1int2 up-regulates FGFR1 by recruiting transcriptional activators P65/FUS and suppressing miR-4687-5p to promote prostate cancer progression.

Fibroblast growth factor receptor 1 (FGFR1) is a core component of the FGFs/FGFR pathway that activates multiple signalling pathways, including ERK1/2, PI3K/AKT, PLCγ, and NF-κB. Aberrant expression of FGFR1 due to gene amplification, chromosome rearrangement, point mutation, and epigenetic deregulations, have been reported in various cancers. FGFR1 overexpression has also been reported in prostate cancer (PCa), but the underlining mechanisms are not clear. Here we report a novel circular RNA, circFGFR1int2, derived from intron 2 of FGFR1 gene, which is overexpressed in PCa and associated with tumor progression. Importantly, we show that circFGFR1int2 facilitates FGFR1 transcription by recruiting transcription activators P65/FUS and by interacting with FGFR1 promoter. Moreover, we show that circFGFR1int2 suppresses post-transcriptional inhibitory effects of miR-4687-5p on FGFR1 mRNA. These mechanisms synergistically promote PCa cell growth, migration, and invasion. Overexpression of circFGFR1int2 is significantly correlated with higher tumor grade, Gleason score, and PSA level, and is a significant unfavorable prognosticator for CRPC-free survival (CFS) (RR = 3.277, 95% confidence interval: 1.192-9.009; P = 0.021). These findings unravelled novel mechanisms controlling FGFR1 gene expression by intronic circRNA and its potential clinicopathological utility as a diagnostic or therapeutic target.

The roles of mutated SPINK1 gene in prostate cancer cells.

SPINK1-positive prostate cancer (PCa) has been identified as an aggressive PCa subtype. However, there is a lack of definite studies to elucidate the underlying mechanism of the loss of SPINK1 expression in most PCa cells except 22Rv1 cells, which are derived from a human prostatic carcinoma xenograft, CWR22R. The aim of this study was to investigate the mechanisms of SPINK1 protein positive/negative expression and its biological roles in PCa cell lines. SPINK1 mRNA was highly expressed in 22Rv1 cells compared with LNCaP, C4-2B, DU145, and PC-3 cells, and the protein was only detected in 22Rv1 cells. Among these cell lines, the wild-type SPINK1 coding sequence was only found in 22Rv1 cells, and two mutation sites, the c.194G>A missense mutation and the c.210T>C synonymous mutation, were found in other cell lines. Our further research showed that the mutations were associated with a reduction in SPINK1 mRNA and protein levels. Functional experiments indicated that SPINK1 promoted PC-3 cell proliferation, migration, and invasion, while knockdown of SPINK1 attenuated 22Rv1 cell proliferation, migration, and invasion. The wild-type SPINK1 gene can promote the malignant behaviors of cells more than the mutated ones. Cell cycle analysis by flow cytometry showed that SPINK1 decreased the percentage of cells in the G0/G1 phase and increased the percentage of S phase cells. We demonstrated that the c.194G>A and c.210T>C mutations in the SPINK1 gene decreased the mRNA and protein levels. The wild-type SPINK1 gene is related to aggressive biological behaviors of PCa cells and may be a potential therapeutic target for PCa.

High-resolution temporal dynamic transcriptome landscape reveals a GhCAL-mediated flowering regulatory pathway in cotton (Gossypium hirsutum L.).

The transition from vegetative to reproductive growth is very important for early maturity in cotton. However, the genetic control of this highly dynamic and complex developmental process remains unclear. A high-resolution tissue- and stage-specific transcriptome profile was generated from six developmental stages using 72 samples of two early-maturing and two late-maturing cotton varieties. The results of histological analysis of paraffin sections showed that flower bud differentiation occurred at the third true leaf stage (3TLS) in early-maturing varieties, but at the fifth true leaf stage (5TLS) in late-maturing varieties. Using pairwise comparison and weighted gene co-expression network analysis, 5312 differentially expressed genes were obtained, which were divided into 10 gene co-expression modules. In the MElightcyan module, 46 candidate genes regulating cotton flower bud differentiation were identified and expressed at the flower bud differentiation stage. A novel key regulatory gene related to flower bud differentiation, GhCAL, was identified in the MElightcyan module. Anti-GhCAL transgenic cotton plants exhibited late flower bud differentiation and flowering time. GhCAL formed heterodimers with GhAP1-A04/GhAGL6-D09 and regulated the expression of GhAP1-A04 and GhAGL6-D09. GhAP1-A04- and GhAGL6-D09-silenced plants also showed significant late flowering. Finally, we propose a new flowering regulatory pathway mediated by GhCAL. This study elucidated the molecular mechanism of cotton flowering regulation and provides good genetic resources for cotton early-maturing breeding.

Genome-wide identification and expression analysis of the BURP domain-containing genes in Gossypium hirsutum.

BACKGROUND: Many BURP domain-containing proteins, which are unique to plants, have been identified. They performed diverse functions in plant development and the stress response. To date, only a few BURP domain-containing genes have been studied, and no comprehensive analysis of the gene family in cotton has been reported. RESULTS: In this study, 18, 17 and 30 putative BURP genes were identified in G. raimondii (D5), G. arboreum (A2) and G. hirsutum (AD1), respectively. These BURP genes were phylogenetically classified into eight subfamilies, which were confirmed by analyses of gene structures, motifs and protein domains. The uneven distribution of BURPs in chromosomes and gene duplication analysis indicated that segmental duplication might be the main driving force of the GhBURP family expansion. Promoter regions of all GhBURPs contained at least one putative stress-related cis-elements. Analysis of transcriptomic data and qRT-PCR showed that GhBURPs showed different expression patterns in different organs, and all of them, especially the members of the RD22-like subfamily, could be induced by different stresses, such as abscisic acid (ABA) and salicylic acid (SA), which indicated that the GhBURPs may performed important functions in cotton's responses to various abiotic stresses. CONCLUSIONS: Our study comprehensively analyzed BURP genes in G. hirsutum, providing insight into the functions of GhBURPs in cotton development and adaptation to stresses.

MicroRNA181c inhibits prostate cancer cell growth and invasion by targeting multiple ERK signaling pathway components.

BACKGROUND: The ERK signaling pathway is frequently deregulated in tumorigenesis, mostly by classical mechanisms such as gene mutation of its components (eg, RAS and RAF). However, whether and how multiple key components of ERK pathway are regulated by microRNAs are not clear. METHODS: We firstly predicted post-transcriptional regulation of multiple key components of the ERK signaling pathway by miR181c through bioinformatics analysis, and then confirmed the post-transcriptional regulation by dual luciferase reporter gene assays and Western blot analysis. The biological effects of miR181c on prostate cancer cell proliferation, apoptosis, migration, and invasion were measured by CCK-8 assay, flow cytometry, wound scratch assay, transwell cell migration, and invasion assays. RESULTS: miR181c post-transcriptionally regulated multiple key members of the ERK signaling pathway, including extracellular signal-regulated kinase 2 (ERK2), ribosomal S6 kinase 2 (RSK2), serum response factor (SRF), and FBJ murine osteosarcoma viral oncogene homolog (c-Fos). Ectopic expression of miR181c mimics effectively suppressed prostate cancer cell proliferation, migration, and invasion, but promoted cell apoptosis. Furthermore, miR181c treatment combined with the multi-kinase inhibitor sorafenib significantly enhanced these anti-tumor effects. CONCLUSIONS: Downregulation of miR181c results in deregulated ERK signaling and promotes prostate cancer cell growth and metastasis.

[Knocking-out of HIF1α gene by CRISPR/cas9 inhibits proliferation and invasiveness of prostate cancer DU145 cells].

OBJECTIVE: To explore the role of HIF1α gene in prostate cancer cell line DU145 by knocking it out with a novel gene-editing tool CRISPR/cas9 system. METHODS: A CRISPR/cas9 system with two sgRNAs targeting exon 1 of the HIF1α gene was constructed for the knock out experiment. CCK8 assay and transwell experiment were carried out to assess the effect of the knock out on the proliferation, migration and invasiveness of DU145 cells. RESULTS: The efficiency of gene-targeting was measured through a T7E1 assaying and sequence analysis, which confirmed that the partial knock out was successful and has led to a significant decrease in the expression of HIF1α and inhibition of cell proliferation, migration and invasiveness. CONCLUSION: A CRISPR/cas9 system for the knock out of HIF1α has been successfully constructed, which could inhibit the proliferation and migration of DU145 cells. The system can facilitate further studies of the HIF1α gene and its roles in tumorigenesis.

MiR200c targets IRS1 and suppresses prostate cancer cell growth.

BACKGROUND: The downregulation of the tumor suppressor miR200c plays important roles in many malignant tumors. This study aims to show that miR200c is a posttranscriptional regulator of insulin receptor substrate 1 (IRS1) and over-expression of miR200c suppresses prostate cancer cell growth. METHODS: Bioinformatics analysis was used to show potential post-translational regulation of IRS1 by miR200c. Dual reporter gene assays were chosen to test the binding of miR200c to the potential seed sequences in IRS1 3'UTR. RT-PCR, Q-PCR and western blot were applied to determine the regulation effect of miR200c on IRS1. CCK8 assay, soft agar assay, trypan blue exclusion assay and flow cytometric analysis were used to measure the biological effects of miR200c on prostate cancer cell proliferation and apoptosis. RESULTS: The 449-455 nt, 3061-3067 nt, and 3096-3102 nt of the IRS1 3'-UTR were identified as three potential seed sequences for miR200c. MiR200c directly binds to IRS1 through the seed sequences in IRS1 3'-UTR. Artificial overexpression of miR200c significantly downregulated the mRNA and protein levels of IRS1, together with decreased cell proliferation and increased cell death of PC3 and DU145 cells. CONCLUSIONS: Our results suggest that miR200c plays crucial roles in prostate cancer by post-transcriptional regulation of IRS1. The mir200c/IRS1 pathway may be a potential therapeutic target to prevent prostate cancer cell growth.

Deep learning-based prediction of H3K27M alteration in diffuse midline gliomas based on whole-brain MRI.

BACKGROUND: H3K27M mutation status significantly affects the prognosis of patients with diffuse midline gliomas (DMGs), but this tumor presents a high risk of pathological acquisition. We aimed to construct a fully automated model for predicting the H3K27M alteration status of DMGs based on deep learning using whole-brain MRI. METHODS: DMG patients from West China Hospital of Sichuan University (WCHSU; n = 200) and Chengdu Shangjin Nanfu Hospital (CSNH; n = 35) who met the inclusion and exclusion criteria from February 2016 to April 2022 were enrolled as the training and external test sets, respectively. To adapt the model to the human head MRI scene, we use normal human head MR images to pretrain the model. The classification and tumor segmentation tasks are naturally related, so we conducted cotraining for the two tasks to enable information interaction between them and improve the accuracy of the classification task. RESULTS: The average classification accuracies of our model on the training and external test sets was 90.5% and 85.1%, respectively. Ablation experiments showed that pretraining and cotraining could improve the prediction accuracy and generalization performance of the model. In the training and external test sets, the average areas under the receiver operating characteristic curve (AUROCs) were 94.18% and 87.64%, and the average areas under the precision-recall curve (AUPRC) were 93.26% and 85.4%. CONCLUSIONS: The developed model achieved excellent performance in predicting the H3K27M alteration status in DMGs, and its good reproducibility and generalization were verified in the external dataset.

Spatial transcriptomics reveals niche-specific enrichment and vulnerabilities of radial glial stem-like cells in malignant gliomas.

Diffuse midline glioma-H3K27M mutant (DMG) and glioblastoma (GBM) are the most lethal brain tumors that primarily occur in pediatric and adult patients, respectively. Both tumors exhibit significant heterogeneity, shaped by distinct genetic/epigenetic drivers, transcriptional programs including RNA splicing, and microenvironmental cues in glioma niches. However, the spatial organization of cellular states and niche-specific regulatory programs remain to be investigated. Here, we perform a spatial profiling of DMG and GBM combining short- and long-read spatial transcriptomics, and single-cell transcriptomic datasets. We identify clinically relevant transcriptional programs, RNA isoform diversity, and multi-cellular ecosystems across different glioma niches. We find that while the tumor core enriches for oligodendrocyte precursor-like cells, radial glial stem-like (RG-like) cells are enriched in the neuron-rich invasive niche in both DMG and GBM. Further, we identify niche-specific regulatory programs for RG-like cells, and functionally confirm that FAM20C mediates invasive growth of RG-like cells in a neuron-rich microenvironment in a human neural stem cell derived orthotopic DMG model. Together, our results provide a blueprint for understanding the spatial architecture and niche-specific vulnerabilities of DMG and GBM.

The MADS transcription factor GhFYF is involved in abiotic stress responses in upland cotton (Gossypium hirsutum L.).

Cotton is an important textile industry raw material crops, which plays a critical role in the development of society. MADS transcription factors (TFs) play a key role about the flowering time, flower development, and abiotic stress responses in plants, but little is known about their functions on abiotic stress in cotton. In this study, a MIKCC subfamily gene from cotton, GhFYF (FOREVER YOUNG FLOWER), was isolated and characterized. Our data showed that GhFYF localized to the nucleus. A ß-glucuronidase (GUS) activity assay revealed that the promoter of GhFYF was mainly expressed in the flower and seed of ProGhFYF::GUS transgenic A. thaliana plants. The GUS staining of flowers and seeds was deepened after drought, salt treatment, and the expression level of the GUS gene and corresponding stress genes AtERD10, AtAnnexin1 are up-regulated in the inflorescence. Overexpression GhFYF in A. thaliana could promote the seed germination and growth under different salt concentrations, and determin the proline content. Yeast two-hybrid (Y2H) assays showed that GhFYF interacted with the HAD-like protein GhGPP2, which has responds to abiotic stress. Our findings indicate that GhFYF is involved in abiotic stress responses, especially for salt stress. This work establishes a solid foundation for further functional analysis of the GhFYF gene in cotton.

A novel promoter-associated non-coding small RNA paGLI1 recruits FUS/P65 to transactivate GLI1 gene expression and promotes infiltrating glioma progression.

Glioma-associated oncogene homolog 1 (GLI1) is a core component of the Hedgehog (HH) signalling pathway and is a transcription activator of numerous oncogenes, such as SOX9, VEGFA, BCL2, and CDK2. The complex regulation of GLI1 involves numerous pathways and molecules, including HH-dependent and independent, epigenetic and post-transcriptional mechanisms. Here, we report the discovery, characterization and function of a novel sense promoter-associated ncRNA, paGLI1 that is overexpressed in infiltrating glioma. We show that paGLI1 promotes GLI1 gene transcription through binding to and recruitment of the transcription factor complex FUS/P65 by interacting with paGLI1 DNA sequence. This interaction facilitates FUS/P65 binding to the GLI1 promoter to activate GLI1 transcription and hence its downstream oncogenes, which results in enhancement of glioma cell proliferation and invasiveness. Importantly, over-expression of paGLI1 is a significant unfavorable prognosticator for both disease-specific and progression-free survival in glioma patients, with relative risks being 2.932 (95% confidence interval: 1.280 to 6.713) (P < 0.05) and 2.284 (95% confidence interval: 1.051 to 4.966) (P < 0.05), respectively. The novel paGLI1/FUS/P65 regulatory mechanisms play important roles in infiltrating glioma progression and may serve as potential targets for future therapeutics.

Novel Recurrent Altered Genes in Chinese Patients With Anaplastic Thyroid Cancer.

BACKGROUND: Anaplastic thyroid cancer (ATC) is a rare but lethal malignancy, and few systematic investigations on genomic profiles of ATC have been performed in Chinese patients. METHODS: Fifty-four ATC patients in West China Hospital between 2010 to 2020 were retrospectively analyzed, while 29 patients with available samples were sequenced by whole-exome sequencing (WES). The associations between genomic alterations and clinical characteristics were statistically evaluated. RESULTS: The median overall survival was 3.0 months in the entire cohort, which was impacted by multiple clinical features, including age, tumor size, and different treatment strategies. In the WES cohort, totally 797 nonsilent mutations were detected; the most frequently altered genes were TP53 (48%), BRAF (24%), PIK3CA (24%), and TERT promoter (21%). Although these mutations have been well-reported in previous studies, ethnic specificity was exhibited in terms of mutation frequency. Moreover, several novel significantly mutated genes were identified including RBM15 (17%), NOTCH2NL (14%), CTNNA3 (10%), and KATNAL2 (10%). WES-based copy number alteration analysis also revealed a high frequent gain of NOTCH2NL (41%), which induced its increased expression. Gene mutations and copy number alterations were enriched in phosphatidylinositol 3-kinase/AKT/mechanistic target of rapamycin (mTOR), NOTCH, and WNT pathways. CONCLUSIONS: This study reveals shared and ethnicity-specific genomic profiles of ATC in Chinese patients and suggests NOTCH2NL may act as a novel candidate driver gene for ATC tumorigenesis.

Suppression of α-methylacyl-coenzyme A racemase by miR200c inhibits prostate adenocarcinoma cell proliferation and migration.

Overexpression of α-methylacyl-coenzyme A racemase (AMACR/P504S) is a major abnormality that has been observed in prostate cancer, whereas microRNA (miRNA/miR) 200c, is downregulated. The aim of the present study was to explore whether miR200c was able to exert any regulatory effects on AMACR. To meet this aim, bioinformatics analysis was performed to identify potential binding sites for miR200c in the 3'-untranslated region (3'-UTR) of AMACR. Recombinant adenoviral and dual reporter gene assays were designed to examine the binding of miR200c to the potential seed sequences in the AMACR 3'-UTR. Conventional reverse transcription (RT)-PCR, RT-quantitative (q)PCR and western blotting were also used to examine the regulatory effects of miR200c on AMACR at the mRNA and protein levels. Furthermore, Cell Counting Kit-8, wound healing and Transwell assays were performed to investigate the biological effects of miR200c-AMACR deregulation on prostate cancer cell proliferation, migration and invasion. It was revealed that miR200c post-transcriptionally suppressed AMACR expression by interacting with the 90-97 nucleotide sequence of the AMACR mRNA 3'-UTR. Artificial overexpression of miR200c significantly downregulated the mRNA and protein levels of AMACR in DU145 and PC-3 prostate cancer cells. Knockdown of AMACR by RNA interference, or overexpression of miR200c by recombinant adenoviral Ad-miR200c, inhibited prostate cancer cell proliferation, migration and invasiveness. Taken together, the results of the present study revealed that miR200c may suppress the AMACR expression level post-transcriptionally. The results also indicate that perturbation of the miR200c-AMACR regulatory mechanism may be involved in prostate carcinogenesis and that this may be exploited in future therapeutic approaches to prostate cancer.

Identification, Expression, and Functional Analysis of the Group IId WRKY Subfamily in Upland Cotton (Gossypium hirsutum L.).

WRKY transcription factors have diverse functions in regulating stress response, leaf senescence, and plant growth and development. However, knowledge of the group IId WRKY subfamily in cotton is largely absent. This study identified 34 group IId WRKY genes in the Gossypium hirsutum genome, and their genomic loci were investigated. Members clustered together in the phylogenetic tree had similar motif compositions and gene structural features, revealing similarity and conservation within group IId WRKY genes. During the evolutionary process, 14 duplicated genes appeared to undergo purification selection. Public RNA-seq data were used to examine the expression patterns of group IId WRKY genes in various tissues and under drought and salt stress conditions. Ten highly expressed genes were identified, and the ten candidate genes revealed distinct expression patterns under drought and salt treatments by qRT-PCR analysis. Among them, Gh_A11G1801 was used for functional characterization. GUS activity was differentially induced by various stresses in Gh_A11G1801p::GUS transgenic Arabidopsis plants. The virus-induced gene silencing (VIGS) of Gh_A11G1801 resulted in drought sensitivity in cotton plants, which was accompanied by elevated malondialdehyde (MDA) content and reduced catalase (CAT) content. Taken together, these findings obtained in this study provide valuable resources for further studying group IId WRKY genes in cotton. Our results also enrich the gene resources for the genetic improvements of cotton varieties that are suitable for growth in stressful conditions.

Downregulation of miR-199a-5p promotes prostate adeno-carcinoma progression through loss of its inhibition of HIF-1α.

Hypoxia-inducible factor-1 alpha (HIF-1α) plays key roles in cell survival under both hypoxia and normoxia conditions. Regulation of HIF-1α is complex and involves numerous molecules and pathways, including post-transcriptional regulation by microRNAs (miRNAs). Although upregulation of HIF-1α has been shown to promote prostate adenocarcinoma (PCa) progression, the mechanism by which miRNAs modulate HIF-1α in prostate cancer has not been clarified. Here, we show that miR-199a-5p is underexpressed in prostate adenocarcinoma. Artificial overexpression of miR-199a-5p decreased cell proliferation, motility, and tumor angiogenesis and increased apoptosis in PCa cell liness PC-3 and DU145 by directly targeting the 3'-untranslated region (UTR) of HIF-1α mRNA, which reduced HIF-1α levels as well as downstream genes transactivated by HIF-1α (such as VEGF, CXCR4, BNIP3 and BCL-xL). Abnormalities of miR-199a-HIF regulation may contribute significantly to PCa pathogenesis and progression.

IDH1/2 gene hotspot mutations in central nervous system tumours: analysis of 922 Chinese patients.

Mutations of isocitrate dehydrogenase 1 (IDH1) or 2 (IDH2) genes have been identified as early molecular events in the development of astrocytomas and oligodendrogliomas. Data regarding the status and prevalence of IDH1/2 mutations in Chinese patients are limited. Herein we report our data from West China Hospital, a major Chinese medical centre. IDH1(R132H) mutation was analysed by immunohistochemistry with the mutation-specific IDH1(R132H) antibody in 1011 patients, including 922 central nervous system (CNS) tumours and 89 non-neoplastic CNS lesions, and PCR-based direct sequencing of IDH1/2 gene mutation in 570 of these samples. Correlation with clinicopathological features and immunohistochemical expression of p53, EGFR, PTEN and Ki-67 was examined. Our data showed that IDH1/2 mutation was present in oligodendrogliomas, anaplastic oligodendrogliomas, diffuse or anaplastic astrocytomas, and glioblastomas, with decreasing frequency, but not in other types of CNS tumours or non-neoplastic lesions examined. IDH1(R132) mutation was most frequent in oligodendrogliomas (57/62, 91.9%), with IDH1(R132H) mutation as the most frequent mutation form. Only one case for each of the rare mutations (R132C, R132G, R132L, and R132S) was identified in the 570 samples analysed by sequencing. Younger age, low expression of p53 and low Ki-67 index were significantly correlated with IDH1 mutation status (p=0.000). All tumours with IDH1(R132) mutations were supratentorial, with frontal lobe as the most frequent site for IDH-mutated gliomas. Only three IDH2(R172) mutation cases were detected in this series. Univariate survival analysis in 459 glioma patients with diffusely infiltrating gliomas showed that IDH1 mutations as well as the more classical prognosticators (age, WHO grade, p53 and Ki-67 index) were of prognostic significance. Multivariate analysis by Cox proportional hazard regression model demonstrated that lack of IDH1 mutation was an independent prognostic factor for both progression-free survival [relative risk (RR)=2.450, 95% confidence interval (CI)=1.351-4.444] and disease-specific survival (RR=2.489, 95%CI=1.155-5.363).

SOX9 was involved in TKIs resistance in renal cell carcinoma via Raf/MEK/ERK signaling pathway.

Renal cell carcinoma (RCC) is common genitourinary malignancy in human, 30-40% of patients with RCC would be diagnosed with metastatic RCC (mRCC). Even in the era of targeted therapy, patients with mRCC would inevitably progress due to drug resistance. Herein, exploration of the mechanisms of resistance is noteworthy to study. In the present study, we firstly reported the expression profile of SOX9 in renal carcinoma cells and tissues, and found that its expression was significantly associated with Fuhrman grading. Dual luciferase analysis confirmed that Raf/MEK/ERK pathway could directly be regulated by SOX9, and sequential experiments demonstrated that, renal carcinoma cells could sensitize to Sorafenib/Sunitinib through Raf/MEK/ERK signaling pathway inhibition regulated by SOX9 down-regulation. In a small cases with mRCC treated with Sorafenib/Sunitinib (n=38), comparative analysis showed that patients with SOX9 (-) had much better therapeutic response to TKIs than those with SOX9 (+) (PD: 9.1% vs. 56.2%, P=0.002, DCR: 90.9% vs. 43.8%, P=0.002). Based on these findings, we concluded that, SOX9 was firstly described to be highly expressed in renal cell carcinoma, and its expression was involved in TKIs drug resistance through activation of Raf/MEK/ERK pathway. In vitro, patients with SOX9 (-) was related to better response to TKIs treatment than those with SOX9 (+). SOX9 could be expected to be a promising biomarker predicting TKIs response and even expected to be another novel target in the treatment of mRCC.