Adjacent Control Tissues Research Articles

HMG20A was identified as a key enhancer driver associated with DNA damage repair in oral squamous cell carcinomas

BackgroundOral squamous cell carcinoma (OSCC) is the main type of oral cancer. Disturbing DNA repair is an invaluable way to improve the effectiveness of tumor treatment. Here, we aimed to explore the key enhancer drivers associated with DNA damage repair in OSCC cells.MethodsGene Set Enrichment Analysis (GSEA), Gene Set Variation Analysis (GSVA) and Kaplan-Meier analysis were applied to explore the relationship among DNA repair-related genes expression and clinical phenotypes based on The Cancer Genome Atlas (TCGA) database. HOMER software and Integrative Genomics Viewer were applied to identify and visualize enhancers using GSE120634. Toolkit for Cistrome Data Browser was applied to predict transcription factors. Human Protein Atlas Database was used to analyze the protein levels of transcription factors in OSCC and control tissues. Seventy-two OSCC patients were included in this study. qRT-PCR was used to detect transcription factor expression in OSCC and adjacent control tissues collected in this study. qRT-PCR and ChIP-qPCR were used to verify the binding of transcription factors to enhancers, and regulation of target genes transcription. Transcription factor knockdown and control cells were treated with cisplatin. CCK8 was used to detect cell viability and proliferation. Western blotting was implemented to detect the levels of DNA repair-related proteins. Transwell assay was used to detect cell invasion.ResultsDNA repair was positively associated with the OSCC metastatic phenotype. Patients in the cluster with high expression of DNA repair-related genes had a worse prognosis and a higher proportion of advanced stage, low-differentiation, alcohol consumption and smoking compared to the cluster with low DNA repair-related gene expression. Seventeen metastasis-specific enhancer-controlled upregulated DNA repair-related genes, with the top two upregulated genes being ADRM1 26 S proteasome ubiquitin receptor (ADRM1) and solute carrier family 12 member 7 (SLC12A7) were screened. High mobility group 20 A (HMG20A) was the key prognostic enhancer driver regulating metastasis-specific DNA repair-related genes, with higher expression in OSCC tissues than normal control tissues, and higher expression in metastatic OSCC tissues than non-metastatic OSCC tissues. HMG20A bound to the metastasis-specific enhancers of ADRM1 and SLC12A7, thereby promoting ADRM1 and SLC12A7 expression. Knockdown of HMG20A enhanced cisplatin sensitivity of cells, and inhibited OSCC cells from repairing DNA damage caused by cisplatin, as well as proliferation and invasion of OSCC cells.ConclusionHMG20A was identified as the key prognostic enhancer driver regulating DNA repair in OSCC cells, providing a new therapeutic target for OSCC.

Identify the immune characteristics and immunotherapy value of CD93 in the pan-cancer based on the public data sets.

CD93 is a transmembrane receptor that is mainly expressed on endothelial cells. A recent study found that upregulated CD93 in tumor vessels is essential for tumor angiogenesis in several cancers. However, the underlying mechanisms are largely unexplored. Our present research systematically analyzed the characteristics of CD93 in tumor immunotherapy among 33 cancers. CD93 levels and co-expression of CD93 on cancer and stromal cells were detected using public databases and multiple immunofluorescence staining. The Kaplan-Meier (KM) analysis identified the predictive role of CD93 in these cancer types. The survival differences between CD93 mutants and WT, CNV groups, and methylation were also investigated. The immune landscape of CD93 in the tumor microenvironment was analyzed using the SangerBox, TIMER 2.0, and single-cell sequencing. The immunotherapy value of CD93 was predicted through public databases. CD93 mRNA and protein levels differed significantly between cancer samples and adjacent control tissues in multiply cancer types. CD93 mRNA expression associated with patient prognosis in many cancers. The correlation of CD93 levels with mutational status of other gene in these cancers was also analyzed. CD93 levels significantly positively related to three scores (immune, stromal, and extimate), immune infiltrates, immune checkpoints, and neoantigen expression.. Additionally, single-cell sequencing revealed that CD93 is predominantly co-expressed on tumor and stromal cells, such as endothelial cells, cancer-associated fibroblasts (CAFs), neutrophils, T cells, macrophages, M1 and M2 macrophages. Several immune-related signaling pathways were enriched based on CD93 expression, including immune cells activation and migration, focal adhesion, leukocyte transendothelial migration, oxidative phosphorylation, and complement. Multiple immunofluorescence staining displayed the relationship between CD93 expression and CD8, CD68, and CD163 in these cancers. Finally, the treatment response of CD93 in many immunotherapy cohorts and sensitive small molecules was predicted from the public datasets. CD93 expression is closely associated with clinical prognosis and immune infiltrates in a variety of tumors. Targeting CD93-related signaling pathways in the tumor microenvironment may be a novel therapeutic strategy for tumor immunotherapy.

Identification of novel genetic and epigenetic regulators of different tissue types of cervical cancer.

The study aimed to find differential gene mutations, DNA methylation, and expression profiles among different categories of cervical cancer samples. The study was based on freely available gene mutations, promoter methylation, and gene expression status of The Cancer Genome Atlas (TCGA) cervical cancer samples and adjacent normal tissues in the Genomic Data Commons (GDC) portal. The association of CpG island methylation with gene expression was determined through negative correlation analysis. We identified that the ErbB signaling pathway and proteoglycans pathway was significantly associated with adenocarcinoma cervical cancers patients. In these pathways, missense mutation especially S310F in the ERBB2 gene as well as G12D and A146T in the KRAS gene were significantly associated with adenocarcinoma cases. Furthermore, a comparison of SCC cases with adjacent control tissues revealed differential hypermethylation of two CpG positions of the KAAG1 gene and differential downregulation of NPY1R and NPY5R genes in cervical squamous cell carcinoma compared to cervical adenocarcinoma cases and adjacent normal tissues. Specifically, the hypermethylation of the promoter region of the KAAG1 gene might be responsible for the carcinogenesis of cervical squamous cells exclusively and methylation marks can be reversible by the widely used drug, azacytidine. In contrast, adenocarcinoma cervical cancer cases may be treated with floxuridine which is successfully utilized for other tissue-specific adenocarcinoma cases. These results provide valuable insight into the differential molecular markers among the categories of cervical cancer, which helps our ability to classify these cancers and for targeted therapy.

The pattern of expression and prognostic value of key regulators for m7G RNA methylation in hepatocellular carcinoma.

N7-methylguanosine (m7G) modification on internal RNA positions plays a vital role in several biological processes. Recent research shows m7G modification is associated with multiple cancers. However, in hepatocellular carcinoma (HCC), its implications remain to be determined. In this place, we need to interrogate the mRNA patterns for 29 key regulators of m7G RNA modification and assess their prognostic value in HCC. Initial, the details from The Cancer Genome Atlas (TCGA) database concerning transcribed gene data and clinical information of HCC patients were inspected systematically. Second, according to the mRNA profiles of 29 m7G RNA methylation regulators, two clusters (named 1 and 2, respectively) were identified by consensus clustering. Furthermore, robust risk signature for seven m7G RNA modification regulators was constructed. Last, we used the Gene Expression Omnibus (GEO) dataset to validate the prognostic associations of the seven-gene risk signature. We figured out that 24/29 key regulators of m7G RNA modification varied remarkably in their grades of expression between the HCC and the adjacent tumor control tissues. Cluster one compared with cluster two had a substandard prognosis and was also positively correlated with T classification (T), pathological stage, and vital status (fustat) significantly. Consensus clustering results suggested the expression pattern of m7G RNA modification regulators was correlated with the malignancy of HCC strongly. In addition, cluster one was extensively enriched in metabolic-related pathways. Seven optimal genes (METTL1, WDR4, NSUN2, EIF4E, EIF4E2, NCBP1, and NCBP2) were selected to establish the risk model for HCC. Indicating by further analyses and validation, the prognostic model has fine anticipating command and this probability signature might be a self supporting presage factor for HCC. Finally, a new prognostic nomogram based on age, gender, pathological stage, histological grade, and prospects were established to forecast the prognosis of HCC patients accurately. In essence, we detected association of HCC severity and expression levels of m7G RNA modification regulators, and developed a risk score model for predicting prognosis of HCC patients’ progression.

Dissection of Mouse Hippocampus with Its Dorsal, Intermediate and Ventral Subdivisions Combined with Molecular Validation

Many research methods applied in molecular neuroscience require the collection of hippocampal samples, but a still poorly recognized problem is contamination with the choroid plexus during brain dissection. Because of a distinct pattern of gene expression, its inclusion in brain samples can obscure or even confound conclusions drawn from molecular studies. Therefore, we tested our dissection method designed for removal of tissue contamination using expression of the transthyretin gene (Ttr) as a marker of the choroid plexus. Additionally, we also validated dissection of the entire hippocampus into its dorsal, intermediate and ventral subdivisions using the expression of Trhr and Lct genes as molecular markers of anatomical subdivisions. The PCR analysis showed that Ttr is expressed at a residual level in hippocampal samples that display an mRNA level several hundred lower than the adjacent control tissue colocalized with the choroid plexus. This indicates that the applied method for dissecting the hippocampus from a fresh brain allows for replicable removal of the majority of choroid plexus from hippocampal samples. In turn, differences in expression of Lct and Trhr confirmed the proper dissection of dorsal, intermediate and ventral subdivisions from fresh brain tissue. Therefore, a special emphasis on the removal of tissue contamination and avoidance of tissue distortions makes our protocol especially suitable for molecular experiments performed either on the entire hippocampus or its subdivisions.

Systematic Construction and Validation of a Novel Macrophage Differentiation-Associated Prognostic Model for Clear Cell Renal Cell Carcinoma.

Background: Clear cell renal cell carcinoma (ccRCC) is a malignant tumor of the human urinary system. Macrophage differentiation is associated with tumorigenesis. Therefore, exploring the prognostic value of macrophage differentiation–associated genes (MDGs) may contribute to better clinical management of ccRCC patients. Methods: The RNA sequence data of ccRCC were obtained from The Cancer Genome Atlas (TCGA) database. Differentially expressed MDGs were unveiled in ccRCC and normal samples. The prognostic model was established according to the univariate and multivariate Cox regression analyses. By combining clinico-pathological features and prognostic genes, a nomogram was established to predict individual survival probability. The Tumor Immune Estimation Resource (TIMER) database was utilized to analyze the correlation between prognostic genes and immune infiltrating cells. Eventually, the mRNA and protein expression levels of prognostic genes were verified. Results: A total of 52 differentially expressed prognosis-related MDGs were identified in ccRCC. Afterward, a six-gene prognostic model (ABCG1, KDF1, KITLG, TGFA, HAVCR2, and CD14) was constructed through the Cox analysis. The overall survival in the high-risk group was relatively poor. Moreover, the risk score was identified as an independent prognostic factor. We constructed a prognostic nomogram with a well-fitted calibration curve based on risk score and clinical data. Furthermore, the prognostic genes were significantly related to the level of immune cell infiltration including B cells, CD8+T cells, CD4+T cells, macrophages, neutrophils, and dendritic cells. Finally, the mRNA expression of prognostic genes in clinical ccRCC tissues showed that the ABCG1, HAVCR2, CD14, and TGFA mRNA in tumor samples were increased compared with the adjacent control tissue samples, while KDF1 and KITLG were decreased, which was consistent with the verification results in the GSE53757. Conclusion: In conclusion, this study identified and validated a macrophage differentiation–associated prognostic model for ccRCC that could be used to predict the outcomes of the ccRCC patients.

DNA methylation of miR-138 regulates cell proliferation and EMT in cervical cancer by targeting EZH2

BackgroundEmerging evidence has identified miR-138 as a tumor suppressor that can suppress the proliferation of various cancers. Meanwhile, the cause of abnormal miR-138 expression in cervical cancer remains uncertain. This study clarified the mechanism by which miR-138 regulates proliferation, invasion, metastasis, and EMT in cervical cancer cells.ResultsmiR-138 expression in human cervical cancer and adjacent normal tissue was measured using qPCR. SiHa and C33A cells were used to determine the function of miR-138 via miR-138 mimic or inhibitor transfection, followed by wound healing, Cell Counting Kit-8, flow cytometry, and Transwell assays. Epithelial and mesenchymal marker expression was analyzed using Western blotting. DNA methylation in the miR-138 promoter was examined using bisulfite sequencing PCR. The downstream target genes of miR-138 were identified via bioinformatics analysis and luciferase reporter assays. A tumor xenograft model was employed to validate DNA methylation-induced miR-138 downregulation and tumor growth inhibition in cervical cancer in vivo. miR-138 levels were significantly lower in cervical cancer tissues than in adjacent control tissues. Furthermore, lower miR-138 expression and higher CpG methylation in the miR-138 promoter were identified in lymph node-positive metastatic cervical cancer tumors versus that in non-metastatic tumor tissues. Upon miR-138 overexpression, cell proliferation, metastasis, invasion, and EMT were suppressed. miR-138 agomir transfection and demethylating drug treatment significantly inhibited cervical tumor growth and EMT in tumor xenograft models. DNA methylation inhibited miR-138 transcription, and enhancer of zeste homolog 2 (EZH2) downregulation mediated the tumor suppressor function of miR-138 in cervical cancer.ConclusionWe demonstrated that miR-138 suppresses tumor progression by targeting EZH2 in cervical cancer and uncovered the role of DNA methylation in the miR-138 promoter in its downregulation. These findings demonstrated the potential of miR-138 to predict disease metastasis and/or function as a therapeutic target in cervical cancer.

Transcriptional Activity of Some Genes Involved in Apoptosis in Patients with Vulvar Lichen Sclerosus.

Lichen sclerosus of the vulva is a common, but poorly studied disease. We assessed the level of transcriptional activity of APAF1, BAX, BCL2, BIRC5, CCND1, DAPK1, MCL1, and MYC genes encoding products that control apoptosis in the samples of tissues affected by vulvar lichen sclerosus and adjacent control tissues (n=24). Analysis of transcriptional activity was performed by real-time PCR using specific primers and SYBR Green intercalating dye. After the total group was divided by the presence of the concomitant gynecological diseases, a significant increase in the transcriptional activity of the CCND1 gene was revealed in patients with concomitant uterine fibroids. This may indicate the possible role of the activation of mitosis during tumor initiation.

Elevated Expression of SATB1 Predicts Unfavorable Clinical Outcomes in Colon Adenocarcinoma.

Special AT-rich sequence-binding protein 1 (SATB1) belongs to the chromatin-remodeling protein which regulates different genes expression. High expression of SATB1 was found to be associated with the development of certain carcinomas. However, the functions of SATB1 in colon adenocarcinoma (CAC) remains unclear yet. Our study aims to investigate the potential role of SATB1 in CAC and whether it is associated with the unfavorable symptoms of CAC patients. The expression pattern of SATB1 was measured in CAC samples and adjacent noncancerous samples through quantitative real-time polymerase chain reaction and immunohistochemistry staining. We performed univariate and multivariate analyses to evaluate the clinical role of SATB1 in enrolled patients. The Kaplan-Meier analyses and log-rank tests were carried out to assess the clinicopathologic characteristics. The effect of SATB1 in human colon cancer cells was examined through cellular experiments. The expression level of SATB1 in CAC tissues was significantly elevated compared with adjacent control tissues. High expression of SATB1 in tumor tissue was found to be associated with lymph node metastasis and advanced TNM stage. Higher SATB1 level in CAC patients indicated a worse 5-year survival time. Moreover, high SATB1 was defined as an independent poor prognostic factor. Cellular experiments showed that inhibition of the SATB1 protein level in human colon cells could suppress the migration and invasion capabilities. Our findings revealed that high expression of SATB1 was significantly correlated with the poor clinical features and prognosis of CAC patients. It indicated that SATB1 might serve as a potential prognostic predictor and novel drug target for CAC treatment.

The Novel Methylation Biomarker NPY5R Sensitizes Breast Cancer Cells to Chemotherapy.

Breast cancer (BC) is the most common tumor in women, and the molecular mechanism underlying its pathogenesis remains unclear. In this study, we aimed to investigate gene modules related to the phenotypes of BC, and identify representative candidate biomarkers for clinical prognosis of BC patients. Using weighted gene co-expression network analysis, we here identified NPY5R as a hub gene in BC. We further found that NPY5R was frequently downregulated in BC tissues compared with adjacent tumor-matched control tissues, due to its aberrant promoter CpG methylation which was confirmed by methylation analysis and treatment with demethylation agent. Higher expression of NPY5R was closely associated with better prognosis for BC patients. Gene set enrichment analysis showed that transcriptome signatures concerning apoptosis and cell cycle were critically enriched in specimens with elevated NPY5R. Ectopic expression of NPY5R significantly curbed breast tumor cell growth, induced cell apoptosis and G2/M arrest. Moreover, NPY5R also promoted the sensitivity of BC cells to doxorubicin. Mechanistically, we found that NPY5R restricted STAT3 signaling pathway activation through interacting with IL6, which may be responsible for the antitumor activity of NPY5R. Collectively, our findings indicate that NPY5R functions as a tumor suppressor but was frequently downregulated in BC.

Comprehensive Analysis of Differentially Expressed Profiles of mRNA N6-Methyladenosine in Colorectal Cancer.

Aim: To comprehensively profile the landscape of the mRNA N6-methyladenosine (m6A) modification in human colorectal cancer (CRC). Methods: Methylated RNA immunoprecipitation sequencing (MeRIP-seq) was explored to compare the difference in mRNA N6-methyladenosine (m6A) methylation between CRC tissues and adjacent normal control (NC) tissue. RNA-sequencing (RNA-seq) was performed to transcribe differentially expressed mRNAs. Conjoint analysis of MeRIP-seq and RNA-seq data was conducted to predict RNA-binding proteins (RBPs). Results: MeRIP-seq identified 1110 differentially m6A methylated sites (DMMSs) and 980 differentially m6A methylated genes (DMMGs) in CRC, with 50.13% of all modified genes showing unique m6A-modified peaks in CRC. RNA-seq showed 915 upregulated genes and 1463 downregulated genes in CRC. QRT-PCR verified the RNA-seq results by detecting the expression of some mRNAs. Conjoint analysis of MeRIP-seq and RNA-seq identified 400 differentially m6A methylated and expressed genes (DEGs), and pathway analysis detected that DMMGs and DEGs were closely related to cancer. After analyzing these DMMGs and DEGs through the GEPIA database, we found that the expression of B3GNT6, DKC1, SRPK1, and RIMKLB were associated with prognosis, and the expression of B3GNT6 and RIMKLB were associated with clinical stage. 17 RBPs were identified based on the DMMGs and DEGs, among which FXR1, FXR2, FMR1, IGF2BP2, IGF2BP3, and SRSF1 were obviously highly expressed in CRC, and FMR1, IGF2BP2, and IGF2BP3 were closely related to methylation, and might be involved in the development of CRC. Conclusion: This study comprehensively profiled m6A modification of mRNAs in CRC, which revealed possible mechanisms of m6A-mediated gene expression regulation.

MiR-145-5p Inhibits the Invasion of Prostate Cancer and Induces Apoptosis by Inhibiting WIP1.

Prostate cancer (PCa) is a common malignant tumor of the male genitourinary system that seriously affects the quality of life of patients. Studying the pathogenesis and therapeutic targets of PCa is important. In this study, we investigated the role of miR-145-5p in PCa and its potential molecular mechanisms. The expression levels of miR-145-5p in PCa tissues and adjacent control tissues were detected by real-time quantitative polymerase chain reaction. The effects of miR-145-5p overexpression on PCa were studied using cell proliferation, migration, and invasion experiments. Furthermore, WIP1 was the target gene of miR-145-5p through the bioinformatics website and dual-luciferase reporter gene experiment. Further studies found that WIP1 downregulation could inhibit the proliferation, invasion, and cloning of PCa cells. Overexpression of WIP1 reversed the anticancer effects of miR-145. The anticancer effect of miR-145 was achieved by inhibiting the PI3K/AKT signaling pathway and upregulating ChK2 and p-p38MAPK. Taken together, these results confirmed that miR-145-5p inhibited the growth and metastasis of PCa cells by inhibiting the expression of proto-oncogene WIP1, thereby playing a role in tumor suppression in PCa and may become a potential therapeutic target for the treatment of PCa.

KIFC1 promotes aerobic glycolysis in endometrial cancer cells by regulating the c-myc pathway.

Endometrial cancer (EC) is a common gynecological malignant tumor worldwide. It is imperative to study pathogenesis and therapeutic targets for improving the prognosis of EC. The present study aimed to explore the function and mechanism of kinesin family member C1 (KIFC1) in EC. EC tumor and adjacent normal tissues were collected from 68 pairs of patients. The expression of KIFC1 in tissues and EC cells was analyzed by immunohistochemistry, qRT-PCR or western blot. MTT assay was used to test the cell viability. Flow cytometry was used to determine apoptosis and the cell cycle. Glucose uptake, lactate production, ATP contents and lactate dehydrogenase (LDH) activity were evaluated by a glucose metabolism kit. The expression of HMGA1, c-myc and glycolytic genes was assessed using western blot or qRT-PCR. A mouse xenograft model was established in BALB/c mice to detect tumor growth in vivo. KIFC1 was significantly upregulated in EC tumor tissues compared to adjacent normal control tissues. The upregulated expression of KIFC1 was correlated with poor prognosis in patients. Lentiviral-mediated overexpression of KIFC1 observably enhanced cell viability and reduced the apoptotic rate of Ishikawa and HEC-1B cells. Cell cycle progression was also expedited in the KIFC1 vector group. Moreover, overexpression of KIFC1 elevated glucose uptake, lactate production, ATP contents and LDH activity. However, knockdown of KIFC1 by short hairpin RNA (shRNA) showed the reverse effect on cellular functions. In addition, the expression of c-myc, GLUT1, LDHA and HK2 was increased by the KIFC1 vector. Moreover, HMGA1 regulated the expression of c-myc and glycolytic genes. Upregulated HMGA1 could rescue the effect of KIFC1 knockdown on cellular functions and the expression of glycolytic genes. Finally, KIFC1 knockdown inhibits tumor growth in vivo. The upregulation of KIFC1 was correlated with poor prognosis in EC. KIFC1 promoted aerobic glycolysis in endometrial cancer cells by regulating the HMGA1/c-myc pathway. KIFC1 may be a potential target for the diagnosis and therapy of EC.

Construction and Validation of an Immune-Related Gene Prognostic Index for Esophageal Squamous Cell Carcinoma.

Immune checkpoint inhibitor (ICI) therapy may benefit patients with advanced esophageal squamous cell carcinoma (ESCC); however, novel biomarkers are needed to help predict the response of patients to treatment. Differentially expressed immune-related genes within The Cancer Genome Atlas ESCC dataset were selected using the weighted gene coexpression network and lasso Cox regression analyses. Based on these data, an immune-related gene prognostic index (IRGPI) was constructed. The molecular characteristics of the different IRGPI subgroups were assessed using mutation information and gene set enrichment analysis. Differences in immune cell infiltration and the response to ICI therapy and other drugs were also analyzed. Additionally, tumor and adjacent control tissues were collected from six patients with ESCC and the expression of these genes was verified using real-time quantitative polymerase chain reaction. IRGPI was designed based on CLDN1, HCAR3, FNBP1L, and BRCA2, the expression of which was confirmed in ESCC samples. The prognosis of patients in the high-IRGPI group was poor, as verified using publicly available expression data. KMT2D mutations were more common in the high-IRGPI group. Enrichment analysis revealed an active immune response, and immune infiltration assessment showed that the high-IRGPI group had an increased infiltration degree of CD8 T cells, which contributed to the improved response to ICI treatment. Collectively, these data demonstrate that IRGPI is a robust biomarker for predicting the prognosis and response to therapy of patients with ESCC.

Molecular and Clinical Implications of Somatostatin Receptor Profile and Somatostatin Analogues Treatment in Oral Cavity Squamous Cell Carcinoma.

Simple SummaryThe treatment of oral squamous cell carcinoma (OSCC) represents a significant problem worldwide. Among cancers with the highest incidence, OSCC renders one of the worst prognoses. Therefore, novel prognostic biomarkers and therapeutic tools to tackle OSCC are urgently needed. Somatostatin-analogues (SSA) are an invaluable therapeutic option in the treatment of several cancers. We aimed to determine the expression levels of all somatostatin-receptors (SSTs) in OSCC, compared to adjacent healthy control tissues, to analyze the relationship of SSTs expression with key clinical and histopathological data, and to explore the direct in vitro effect of different SSAs on OSCC cancer cells. Our findings highlight a potential role of SST2 as a good prognostic biomarker for recurrence and metastasis in OSCC and unveil that SSA exerts antitumoral effects on OSCC cells, providing a relevant clinical conclusion, which should be soon tested for their use in humans.Oral squamous cell carcinoma (OSCC) incidence has increased by 50% over the last decade. Unfortunately, surgery and adjuvant radiotherapy and chemotherapy are still the mainstream modality of treatment, underscoring the need for alternative therapies. Somatostatin-analogues (SSA) are efficacious and safe treatments for a variety of tumors, but the presence of somatostatin-receptors (SSTs) and pharmacological effects of SSA on OSCC are poorly known. In this study, we demonstrated that SST2 and SST3 levels were significantly higher in OSCC, compared to adjacent healthy control tissues. SST2 expression was associated with less regional metastasis and a lower recurrence rate. Moreover, SST2 was elevated in OSCC and associated with histopathological good prognosis factors, such as high peritumoral inflammation, smaller depth of invasion, and expansive vs. infiltrative front of tumor invasion. Importantly, treatment with different SSA (octreotide, lanreotide, and pasireotide) significantly reduced cell-proliferation in OSCC primary cell cultures. Altogether, this study demonstrated that SST2 is overexpressed in OSCC vs. healthy tissues and could represent a novel prognostic biomarker, since its expression is associated with tumors that show better prognostic factors and less recurrent rate. Moreover, our data unveil clear antitumoral effects of SSAs on OSCC, opening new avenues to explore their potential as targeting therapy to OSCC.

Expression of Autophagy and Mitophagy Markers in Breast Cancer Tissues.

Mitochondria play important roles in regulating cell bioenergetics status and reactive oxygen species (ROS) generation. ROS-induced mitochondrial damage is among the main intracellular signal inducers of autophagy. Autophagy is a cellular catabolic process that regulates protein and organelle turnover, while a selective form of autophagy, mitophagy, specifically targets dysfunctional mitochondrial degradation. This study aims to measure the levels of autophagy, mitophagy, oxidative stress, and apoptosis in invasive breast carcinoma tissues using immunohistochemistry (IHC). Tissue microarrays of 76 patients with breast cancer were stained with six IHC markers (MnSOD, Beclin-1, LC3, BNIP3, Parkin, and cleaved caspase 3). The expression intensity was determined for each tumor tissue and the adjacent tumor-matched control tissues. Intermediate and strong staining scores of MnSOD, Beclin-1, LC-3, BNIP-3, and Parkin were significantly higher in tumor tissues compared to the adjacent matched control. The scoring intensity was further classified into tissues with negative staining and positive staining, which showed that positive scores of Beclin-1 and Parkin were significantly high in tumor tissues compared to other markers. Positive association was also noted between BNIP-3 and Beclin-1 as well as LC-3 and cleaved caspase-3 immunostaining. To our knowledge, this is one of the first studies that measure both mitophagy and autophagy in the same breast cancer tissues and the adjacent matched control. The findings from this study will be of great potential in identifying new cancer biomarkers and inspire significant interest in applying anti-autophagy therapies as a possible treatment for breast cancer.

MicroRNA Prognostic Signature for Postoperative Success of Metastatic Orthopedic Cancers: Implications for Precision Microsurgery.

The importance of miRNA prognostic signature in cancer, particular cancer metastasis is increasingly being realized. Bone metastasis from several primary human cancers can be managed in clinics by surgical intervention but the prognostic impact of miRNA signature on post-surgery outcome of patients is unknown. This study evaluated a miRNA signature for post-operative outcome of patients with bone metastatic disease. First, the miRNAs, miR-135, miR-203, miR-10b, miR-194, miR-886, and miR-124 were evaluated in bone metastatic tissues, relative to adjacent control tissue. The cohorts of samples (n = 44) consisted of bone metastatic cancer patients with primary lung (n = 18) or breast cancer (n = 26). miR-203 was significantly down-regulated while miR-10b was significantly up-regulated in bone metastasis. Additionally, miR-135 was significantly differentially expressed in the primary lung cancer patients while miR-194 in primary breast cancer patients. The low miR-203- high miR-10b expression was designated high risk group and, compared to the low risk group (high miR-203-low miR-10b expression). Patients with the signature high risk fared significantly better with surgical intervention, in terms of survival at 12 months time point (40% survival with surgery vs. 10% survival without surgery), as revealed by retrospective analysis of patient data. This work reveals potential utilization of miRNA expression levels in not only the general prognosis of cancer metastasis but also the prognosis of surgical intervention with implication for better stratification of patients.

MiR-34a inhibits melanoma growth by targeting ZEB1.

Melanoma is a highly lethal cutaneous cancer with the tendency for early invasion and metastasis. Integrated miRNA transcriptome sequence analysis of human melanoma tumors and adjacent control tissues identified 17 miRNAs differentially expressed in melanoma tissues: let-7a-5p, let-7b-5p, let-7c, miR-374a-3p, miR-100-5p, miR-7, miR-195, miR-1908, miR-214, miR-221, miR-199a-5p, miR-21, miR-18, miR-34a, miR-199a-3p, miR-92a and miR-106b. Among these, miR-34a was most significantly down-regulated in melanoma tissues, and its expression correlated with TNM melanoma stage. miR-34a overexpression inhibited expression and activity of the transcription factor ZEB1, resulting in decreased proliferation and migration of melanoma cells. Moreover, miR-34a overexpression inhibited ZEB1 expression and melanoma tumor growth in vivo, in a melanoma nude mouse model. Together, these findings demonstrate that miR-34a inhibits melanoma growth by targeting the proto-oncogene ZEB1 and suggest the miR-34a -ZEB1 axis may serve as a novel target for melanoma treatment.

KDF1, a Novel Tumor Suppressor in Clear Cell Renal Cell Carcinoma.

KDF1 has been identified as a key regulator of epidermal proliferation and differentiation, but it is unknown whether KDF1 is involved in the pathogenesis of malignancy. No study has reported the expression and function of KDF1 in renal cancer. To explore the pathologic significance of KDF1 in clear cell renal cell carcinoma (ccRCC), the expression level of KDF1 protein in the tumor tissue of ccRCC patients was examined by immunohistochemistry and Western blot while the expression level of KDF1 mRNA was analyzed by using the data from TCGA database. In vitro cell experiments and allogeneic tumor transplantation tests were performed to determine the effects of altered KDF1 expression on the phenotype of ccRCC cells. Both the KDF1 mRNA and protein were found to be decreasingly expressed in the tumor tissue of ccRCC patients when compared with the adjacent non-tumor control tissue. The expression level of KDF1 in the tumor tissue was found to correlate negatively with the tumor grade. Patients with higher KDF1 in the tumor tissue were found to have longer overall survival and disease-specific survival time. KDF1 was shown to be an independent factor influencing the disease-specific survival of the ccRCC patients. Overexpression of KDF1 was found to inhibit the proliferation, migration and invasion of ccRCC cells, which could be reversed by decreasing the expression of KDF1 again. ccRCC cells with KDF1 overexpression were found to produce smaller transgrafted tumors. These results support the idea that KDF1 is involved in ccRCC and may function as a tumor suppressor.

Proteome and phospho-proteome profiling for deeper phenotype characterization of colorectal cancer heterogeneity.

e15536 Background: The rise of precision oncology therapeutics requires deep understanding of the molecular mechanisms implicated in cancer biology. Colorectal cancer (CRC) is one of the first solid tumors to be molecularly characterized by defined genes and pathways. Advances in tumor profiling have revealed a profound molecular heterogeneity in CRC leading to the definition of several consensus molecular subtypes (CMS). However, this molecular heterogeneity is still largely defined on the genomic and transcriptomics level. To complement the understanding of genetically defined molecular subgroups, we performed large-scale deep proteomic and phospho-proteomic profiling of CRC patient biopsies and adjacent healthy control tissue, which has enabled to explore the phenotype and obtain more functional insights in cancer biology. Methods: Sample processing from 5-10 mg of tissue per sample was performed using a liquid handling robot. Phospho-peptide enrichment was carried out with a Kingfisher Flex device and MagReSyn Ti-IMAC magnetic beads. Data-Independent Acquisition (DIA) LC-MS/MS was performed on multiple platforms consisting of a Thermo Scientific Q Exactive HF-X mass spectrometer coupled to a Waters M-Class LC. Chromatography was operating at 5 µL/min, and separation was achieved using 45 min (whole proteome) and 60 min (phospho-proteome) gradients. Results: Indivumed has built IndivuType, the world’s first multi-omics database for individualized cancer therapy, analyzing the highest quality cancer biospecimens to generate the most comprehensive dataset, including genomics, transcriptomics, proteomics, and clinical outcome information. Enabled by the DIA technology, a mass spectrometric method developed by Biognosys that obtains peptide fragmentation data in a highly parallelized way with high sensitivity, more than 7,000 proteins in the whole proteome and 20,000 phospho-peptides in the phospho-proteome workflow were profiled across more than 900 resected tissue samples of various CMS of CRC. The resulting proteome and phospho-proteome data were integrated into the IndivuType database and cross-analyzed with genomic and transcriptomic markers. Through this combined analysis, novel insights in clinically relevant signaling pathways in CRC subtypes were revealed. Conclusions: The deep phenotypic profiling of cancer samples, using next generation proteomics and phospho-proteomics, has enabled us to go beyond the genomic level in the characterization of tumor molecular heterogeneity. This multi-omics approach provides a solid foundation to advance the understanding of cancer biology, unravel key molecular events, and support the identification of novel therapeutic targets for precision medicine in CRC.