Identification of PANoptosis-related signature reveals immune infiltration characteristics and immunotherapy responses for renal cell carcinoma.

PANoptosis is a specific type of inflammatory programmed cell death (PCD) modality that can be involved in three key modes of cellular programmed cell death-pyroptosis, apoptosis and necroptosis. We analyzed PANoptosis activity in three common renal cell carcinoma subtypes (Clear cell renal cell carcinoma, Papillary renal cell carcinoma, and Chromophobe renal cell carcinoma) separately and constructed a new PANoptosis immunity index (PANII). In three renal cell carcinomas, we found that PANII was an effective predictor of immunotherapy efficacy in KIRC, KIRP and KICH, and the high PANII group was characterized by high immune infiltration and sensitivity to immunotherapy, while the low PANII group was prone to immune escape and immunotherapy resistance. We performed molecular docking prediction of each core protein comprising PANII and identified natural small molecule compounds with the highest affinity to target proteins. In addition, we found that down-regulation of PYCARD inhibited the proliferation and migration of renal clear cell carcinoma cells by in vitro functional assays, suggesting that PYCARD could be a novel target for renal clear cell carcinoma therapy. Our findings that the PANoptosis characterization-based index (PANII) helps to elucidate the tumor microenvironmental features of three common renal cell carcinoma subtypes and identify patient populations that will benefit from immunotherapy, providing a new tool for the clinical diagnosis and treatment of patients with intermediate- and advanced-stage renal cell carcinoma.

A novel 17 apoptosis-related genes signature could predict overall survival for bladder cancer and its associations with immune infiltration.

Background: Apoptosis-related genes (ARGs) were used to develop a novel signature for forecasting overall survival (OS) and examining their relationships with immune infiltrates in bladder cancer (BC). Methods: Gene expression matrices as well as related clinical data were acquired for BC samples from online datasets. According to differentially expressed ARGs acquired from normal bladder tissues and cancer samples, functional enrichment analyses were conducted. With the assistance of LASSO and Cox regression analysis, a novel model was successfully established and evaluated by external and internal validations. Results: Eventually, 17 ARGs (SLC5A6, GULP1, TAP1, MMP9, P4HB, FOXL2, CIDEC, EN2, NES, EPHA7, SUSD2, TMPRSS3, HOXB7, SATB1, MEST, PCDHGC3, ASPM) were utilized to construct the signature. Our constructed signature significantly distinguished high-risk from low-risk BC patients of OS by internal and external validations and was also proven to be able to serve as an independent prognostic biomarker (all P < 0.05). Furthermore, a prognostic nomogram was also constructed based on TCGA dataset to predict OS prognosis in BC suffers. Besides, this ARG based model was markedly associated with clinical characteristics like tumor stage (P = 3.98e-06), race (P = 8.255e-06), N stage (P = 0.002), T stage (P = 3.679e-05) and M stage (P = 0.002). As for immune infiltration, our established model was significantly associated with seven tumor-infiltrating immune cells. Conclusions: A prognostic signature was successfully developed by us according to 17 ARGs in BC using external and internal verifications, enabling clinicians to predict BC suffers' OS and promote specific individualization of patient care.

A novel signature constructed by ferroptosis-associated genes (FAGs) for the prediction of prognosis in bladder urothelial carcinoma (BLCA) and associated with immune infiltration.

BACKGROUND: Ferroptosis, a novel form of regulated cell death, has been implicated in the pathogenesis of cancers. Nevertheless, the potential function and prognostic values of ferroptosis in bladder urothelial carcinoma (BLCA) are complex and remain to be clarified. Therefore, we proposed to systematically examine the roles of ferroptosis-associated genes (FAGs) in BLCA. METHODS: According to The Cancer Genome Atlas (TCGA) database, differently expressed FAGs (DEFAGs) and differently expressed transcription factors (DETFs) were identified in BLCA. Next, the network between DEFAGs and DETFs, GO annotations and KEGG pathway analyses were performed. Then, through univariate, LASSO and multivariate regression analyses, a novel signature based on FAGs was constructed. Moreover, survival analysis, PCA analysis, t-SNE analysis, ROC analysis, independent prognostic analysis, clinicopathological and immune correlation analysis, and experimental validation were utilized to evaluate the signature. RESULTS: Twenty-eight DEFAGs were identified, and four FAGs (CRYAB, TFRC, SQLE and G6PD) were finally utilized to establish the FAGs based signature in the TCGA cohort, which was subsequently validated in the GEO database. Moreover, we found that immune cell infiltration, immunotherapy-related biomarkers and immune-related pathways were significantly different between two risk groups. Besides, nine molecule drugs with the potential to treat bladder cancer were identified by the connectivity map database analysis. Finally, the expression levels of crucial FAGs were verified by the experiment, which were consistent with our bioinformatics analysis, and knockdown of TFRC could inhibit cell proliferation and colony formation in BLCA cell lines in vitro. CONCLUSIONS: Our study identified prognostic ferroptosis-associated genes and established a novel FAGs signature, which could accurately predict prognosis in BLCA patients.

A Novel Set of Immune-associated Gene Signature predicts Biochemical Recurrence in Localized Prostate Cancer Patients after Radical Prostatectomy.

Background: Decision-making regarding biochemical recurrence (BCR) in localized prostate cancer (PCa) patients after radical prostatectomy (RP) mainly relies on clinicopathological parameters with a low predictive accuracy. Currently, accumulating evidence suggests that immune-associated genes (IAGs) play irreplaceable roles in tumorigenesis, progression and metastasis. Considering the critical role of immune in PCa, we therefore attempted to identify the novel IAGs signature and validate its prognostic value that can better forecast the risk for BCR and guide clinical treatment. Methods: RNA-sequencing and corresponding clinicopathological data were downloaded from the Gene Expression Omnibus (GEO) database and the Cancer Genome Atlas (TCGA) database. Weighted gene co-expression network analysis (WGCNA) was utilized to screen out the candidate module closely related to BCR, and univariate and LASSO Cox regression analyses were performed to build the gene signature. Kaplan-Meier (KM) survival analysis, time-dependent receiver operating curve (ROC), independent prognostic analysis and nomogram were also applied to evaluate the prognostic value of the signature. Besides, Gene ontology analysis (GO), Kyoto encyclopedia of genes and genomes (KEGG) and gene set enrichment analysis (GSEA) were used to explore potential biological pathways. Results: A total of six IAGs (SSTR1, NFATC3, NRP1, TUBB3, IL1R1, GDF15) were eventually identified and used to establish a novel IAGs signature. The Kaplan-Meier analysis revealed that patients with low-risk scores had longer recurrence-free survival (RFS) than those with high-risk scores in both GSE70769 and TCGA cohorts. Further, our signature was also proven to be a valuable independent prognostic factor for BCR. We also constructed a nomogram based on the gene signature and related clinicopathologic features, which excellently predict 1-year, 3-year and 5-year prognosis of localized PCa patients after RP. Moreover, functional enrichment analysis demonstrated the vital biological processes, and stratified GSEA revealed that a crucial immune-related pathway (T cell receptor signaling pathway) was notably enriched in the high-risk group. Conclusions: We successfully developed a novel robust IAGs signature that is powerful in BCR prediction in localized PCa patients after RP, and created a prognostic nomogram. In addition, the signature might help clinicians in selecting high-risk subpopulation, predicting survival status of patients and promoting more individualized therapies than traditional clinical factors.

Identification and validation of a prognostic immune-related lncRNA signature in bladder cancer.

BACKGROUND: Recently, researches have implied that immune-related lncRNAs (IR-lncRNAs) have a vital role in tumor occurrence and development. However, the study in bladder cancer (Bca) is still unclear. New biomarkers and reliable prognostic models for Bca are still limited. Here, we investigated the potential application value of immune-related lncRNAs in the prognostic evaluation of Bca patients. METHODS: We obtained clinical information and corresponding sequencing data from the The Cancer Genome Atlas (TCGA) database, and the cohort of Bca patients was divided into training and validation cohorts (ratio 7:3) randomly. An immune-related lncRNA co-expression network was constructed to identify immune-related lncRNAs. The candidate module intimately associated with overall survival (OS) was identified by using weighted gene co-expression network analysis (WGCNA). Univariate, multivariate Cox regression and LASSO analysis were performed to build the immune-related lncRNA signature. A prognostic model was further developed and its prognostic value was evaluated by Kaplan-Meier (KM) analysis. GSEA, KEGG analysis and GO annotation were used for functional annotation in this study. RESULTS: Totally, we identified 1,249 differentially expressed IR-lncRNAs were and six of which (AC005674.2, AC090948.1, TFAP2A-AS1, AL354919.2, AC011468.1 and AC018809.2) were finally selected in the gene signature. According to survival analysis, patients with high-risk scores were significantly related to poor survival outcomes. Furthermore, we established a novel gene signature demonstrated high prognostic value, and can be utilized as an independent risk factor (validation cohort: P<0.001, HR =3.832; training cohort: P<0.001, HR =2.843). Additionally, we built a nomogram on account of the clinicopathologic characteristics and gene signature to predict the survival probability of 1-, 3- and 5-year in Bca patients. The value of AUC curve of 1-, 3- and 5-year survival probability was 0.724, 0.777 and 0.77, severally. Furthermore, some enrichment pathways were identified by KEGG and GO analysis, and might be useful to display the potential mechanism for Bca. CONCLUSIONS: Our results indicated that the signature of six immune-related lncRNAs had an underlying value in the prognosis of Bca patients and might be helpful for the immunotherapy of Bca.

Identification and validation of a five apoptosis-related genes signature for prediction of disease-free survival for testicular germ cell tumors.

BACKGROUND: More and more studies have paid attention to the role of apoptosis in tumorigenesis. A variety of apoptosis-related genes (ARGs) are related to tumor progression and resistance to chemotherapy drugs. Therefore, this study aims to establish a prognostic marker for ARG-based testicular germ cell tumors (TCGT). METHODS: TCGT sequencing data and clinical information were downloaded from The Cancer Genome Atlas (TCGA) database and GEO database. The sequencing data of normal tissues came from the GTEx database. Through univariate COX, LASSO, and multiple COX regression analyses, we screened out key ARGs related to prognosis and constructed a risk signature and a prognostic nomogram. Finally, we performed internal and external verification to verify the signature we have established. RESULTS: Five ARGs, including CHGA, LPCAT1, PPP1CA, PSMB5, UBR2 were selected out and utilized to establish a novel signature. Based on this signature, TCGT patients were divided into high-risk groups and low-risk groups. The results showed that the disease-free survival (DFS) of patients in the high-risk group was lower than that in the low-risk group (P=0.02268). The subsequent univariate and multivariate Cox regression analysis further proved that the features we established are valuable independent prognostic factors (P<0.05). Also, a prognostic nomogram was created to visualize the relationship between various prognostic-related factors and the 1-, 3-, and 5-year DFS of TCGT in the TCGA cohort. CONCLUSIONS: We constructed a new nomogram based on ARGs to predict the risk of testicular tumor recurrence. It can help clinicians better and more intuitively predict the survival of patients.

A novel 10 glycolysis-related genes signature could predict overall survival for clear cell renal cell carcinoma.

BACKGROUND: The role of glycolysis in tumorigenesis has received increasing attention and multiple glycolysis-related genes (GRGs) have been proven to be associated with tumor metastasis. Hence, we aimed to construct a prognostic signature based on GRGs for clear cell renal cell carcinoma (ccRCC) and to explore its relationships with immune infiltration. METHODS: Clinical information and RNA-sequencing data of ccRCC were obtained from The Cancer Genome Atlas (TCGA) and ArrayExpress datasets. Key GRGs were finally selected through univariate COX, LASSO and multivariate COX regression analyses. External and internal verifications were further carried out to verify our established signature. RESULTS: Finally, 10 GRGs including ANKZF1, CD44, CHST6, HS6ST2, IDUA, KIF20A, NDST3, PLOD2, VCAN, FBP1 were selected out and utilized to establish a novel signature. Compared with the low-risk group, ccRCC patients in high-risk groups showed a lower overall survival (OS) rate (P = 5.548Ee-13) and its AUCs based on our established signature were all above 0.70. Univariate/multivariate Cox regression analyses further proved that this signature could serve as an independent prognostic factor (all P < 0.05). Moreover, prognostic nomograms were also created to find out the associations between the established signature, clinical factors and OS for ccRCC in both the TCGA and ArrayExpress cohorts. All results remained consistent after external and internal verification. Besides, nine out of 21 tumor-infiltrating immune cells (TIICs) were highly related to high- and low- risk ccRCC patients stratified by our established signature. CONCLUSIONS: A novel signature based on 10 prognostic GRGs was successfully established and verified externally and internally for predicting OS of ccRCC, helping clinicians better and more intuitively predict patients' survival.

Integrative analysis of ceRNA network and DNA methylation associated with gene expression in malignant pheochromocytomas: a study based on The Cancer Genome Atlas.

BACKGROUND: Competitive endogenous RNAs (ceRNAs) have revealed a new mechanism of interaction between RNAs. Epigenetic regulation in the gene expression dynamics has become increasingly important in malignant pheochromocytomas (PCCs). We performed an integrative analysis of ceRNA networks and DNA methylation to identify key biomarkers and contribute to the understanding of the molecular biological mechanisms of malignant PCCs. METHODS: Differentially expressed genes in malignant PCCs and controls were identified from The Cancer Genome Atlas database by using the Limma package in R (v3.4.4). An abnormal lncRNA-miRNA-mRNA ceRNA network was constructed for malignant PCCs, and function enrichment analysis was performed using the Database for Annotation, Visualization, and Integrated Discovery. For DNA methylation datasets, the methylation analysis package was used in identifying differential methylation genes, and potential prognostic genes were identified by Kaplan-Meier survival analysis. RESULTS: A total of 447 lncRNAs, 26 miRNAs, and 1,607 mRNAs were found to be differentially expressed in malignant PCCs as compared with those in normal samples. We then constructed an abnormal lncRNA-miRNA-mRNA ceRNA network for malignant PCCs. The network consisted of 12 lncRNAs, 6 miRNAs, and 220 mRNAs. Functional enrichment analysis showed that differentially expressed mRNAs were particularly enriched in the biological process, cellular component, and molecular function. Furthermore, four differentially expressed mRNAs from ceRNAs were identified through the cross-analysis of gene expression and DNA methylation profiles. LncRNA C9orf147 and 6 out of 220 mRNAs were indicated as prognostic biomarkers for patients with malignant PCCs (P<0.05). CONCLUSIONS: Our research increases the understanding of the pathogenesis of malignant PCCs and offers potential genes as underlying therapeutic targets or prognostic biomarkers.

Cell fate regulation by reticulon-4 in human prostate cancers.

Reticulon-4 (RTN4), a reticulon family protein localized in the endoplasmic reticulum, is reported to be involved in multiple physiological processes like neuroendocrine secretion and membrane trafficking in neuroendocrine cells. Previous studies have presented a great potential of RTN4 for the treatment of autoimmune-mediated demyelinating diseases and spinal cord injury regeneration. While interaction with Bcl-2 and Bcl-2-like family in apoptosis modulation implicated its possible role in various human cancers. However, the investigation of this gene in prostate cancer is mainly ignored. Here in our current study, we focused on its role in prostate cancer and found that RTN4 DNA copy numbers were higher in prostate cancer than normal prostate gland while its RNA and protein expressions were relatively lower. Chromosomal neighbor gene EML6 had similar expression patterns with RTN4 in prostate cancer tissues and cell lines, and further research found that they could be both targeted by miR-148a-3p. Lentivirus-mediated RTN4 overexpression potently inhibited DU145 and LNCaP cells proliferation. Cell cycle was blocked in G2/M phase and significant cell senescence was observed in RTN4 overexpressed prostate cancer cells. Finally, interaction networks in the normal prostate gland and cancer tissues further revealed that RTN4 maybe phosphorylated by MAPKAPK2 and FYN at tyrosine 591 and serine 107, respectively. All these results implied that RTN4 might somehow participate in prostate tumor progression, and this elicits possibility to develop or identify selective agents targeting RTN4 for prostate cancer therapy.

Loss of scinderin decreased expression of epidermal growth factor receptor and promoted apoptosis of castration-resistant prostate cancer cells.

Most patients with prostate cancer will eventually develop the castration-resistant form characterised by metastasis. Cytoskeleton constituents, including F-actin, play important roles in maintaining epithelial integrity and their disruption is a major cause of cancer progression. We previously showed that scinderin (SCIN), an important regulator of F-actin organisation, is highly expressed in poorly differentiated cancer tissues. This study aimed to explore the mechanism of its regulation of cell proliferation. We discovered that SCIN knockdown significantly downregulated epidermal growth factor receptor (EGFR) protein expression, and inhibited epidermal growth factor (EGF)-mediated cell proliferation and activation of the downstream mitogen-activated protein kinase kinase (MEK)/extracellular signal-regulated kinase (ERK) signalling pathway. Silencing of SCIN promoted apoptosis in two cell lines (PC-3 and DU145), inhibited B-cell lymphoma-extra-large (Bcl-xl) expression and activated caspase signalling. Furthermore, in vivo studies showed that SCIN deletion slowed tumour growth and decreased EGFR expression. Thus, we conclude that SCIN promotes prostate cancer cell survival by stabilising EGFR and MEK/ERK signalling.

Apolipoprotein C1 promotes prostate cancer cell proliferation in vitro.

Here, we aimed to investigate the carcinogenic effects of apolipoprotein C1 (APOC1) in prostate cancer (PCa). APOC1 expression was evaluated in PCa and normal prostate specimens, and lentivirus-mediated RNA interference was used to knockdown APOC1 in DU145 cells. The effects of APOC1 silencing on cell proliferation, cell cycle arrest, and apoptosis were assessed. APOC1 expression was much higher in PCa tissues than in normal tissues. Moreover, APOC1 silencing inhibited cell proliferation and colony formation, arrested cell cycle progression, and enhanced apoptosis in DU145 cells. Additionally, APOC1 silencing decreased survivin, phospho-Rb, and p21 levels and increased cleaved caspase-3 expression. These data supported the procarcinogenic effects of APOC1 in the pathogenesis of PCa and suggested that targeting APOC1 may have applications in the treatment of PCa.

Knockdown of PYCR1 inhibits cell proliferation and colony formation via cell cycle arrest and apoptosis in prostate cancer.

Pyrroline-5-carboxylate reductase 1 (PYCR1) is an enzyme involved in cell metabolism, which has been shown to be up-regulated in cancers. However, the functions of PYCR1 in prostate cancers (PCa) are still largely unknown. In the present study, we found that PYCR1 was highly expressed in prostate cancer tissues and then knocked down PYCR1 in PCa cell lines (DU145, PC-3 and LNCap) via lentivirus-mediated gene delivery and analyzed its biological function. Both qRT-PCR and western blotting indicated that PYCR1 was suppressed efficiently after sh-PYCR1 infection. Further analysis indicated knockdown of PYCR1 significantly inhibited PCa cell growth and colony formation ability. The inhibition effects on growth were likely due to G2/M-phase arrest and enhanced cell apoptosis, as determined by flow cytometer analysis. At last, we verified the expression levels of cell cycle regulatory proteins, including CDK1, CDK2, CDK4 and Cyclin B1 were all downregulated and cell apoptotic-related proteins, including cleaved caspase 3 and cleaved PARP were increased in PCa cells after PYCR1 knockdown. Furthermore, PYCR1 has been shown not to be directly regulated by androgen receptor (AR) levels. These results show the functions of PYCR1 in PCa tumorigenesis for the first time and suggest that PYCR1 might be a good potential therapy approach for treating PCa.

MYO6 knockdown inhibits the growth and induces the apoptosis of prostate cancer cells by decreasing the phosphorylation of ERK1/2 and PRAS40.

Prostate cancer is the second most frequently diagnosed cancer among males around the world. Myosin VI (MYO6), as a motor protein, has been reported to be implicated in cancer-related cell migration and cellular functions. To investigate the role of MYO6 in prostate cancer, immunohistochemical analysis was firstly applied to prostate cancer tissues and revealed that MYO6 was closely related with the Gleason score in prostate cancer. Then we used specific short hairpin RNA (shRNA) to downregulate MYO6 expression in DU145 and PC-3 cells and found that decreased MYO6 expression significantly suppressed cell proliferation, as determined by MTT and colony formation assays. Flow cytometry confirmed that the suppression of MYO6 promoted cell cycle arrest at the G2/M and sub-G1 phase in the DU145 cells. Furthermore, PathScan intracellular signaling array analysis demonstrated that the phosphorylation of ERK1/2 and PRAS40 was downregulated in the DU145 cells following MYO6 knockdown. Knockdown of MYO6 downregulated the expression of AKT3 and upregulated the expression of PARP, as confirmed by western blot analysis. These results suggest that MYO6 plays an essential role in the progression of prostate cancer and silencing of MYO6 may be a promising therapeutic approach for prostate cancer.