Gene Expression In Tumor Tissue Research Articles

Abstract 665: Cannabinoid receptor 2 tumor tissue gene expression positively affects lung cancer survival following radical surgery

Abstract Background: Non-small cell lung cancer (NSCLC) is a leading cause of cancer-related death with a 5-year survival rate of 21%. New treatment options are thus required, and the endogenous cannabinoid system is a potential new therapeutic target. We therefore analysed the expression of cannabinoid 1 and 2 receptor (CB1 and CB2) genes in NSCLC tissue and its influence on survival following radical surgery. Method: CB1 and CB2 gene expression in tumor tissue samples collected from 100 NSCLC patients was analysed using the quantitative real-time polymerase chain reaction. Since gene expression may not correlate with protein production, the presence of the CB2 protein was validated by immunohistochemistry. CB2 gene expression was correlated with disease stage, the presence of lymph node and distant metastases, tumor size, tumor grade, and survival. Result: Patients with tumors expressing CB2 gene had significantly longer overall survival (OS) (p<0.001), cancer-specific survival (CSS) (p=0.002), and disease-free survival (DFS) (p<0.001). They also had less advanced disease stage (p=0.047) and fewer lymph node metastases (p=0.011) at the time of surgery. A multivariate analysis showed that CB2 tumor tissue gene expression was a positive prognostic factor for OS (Hazard ratio (HR) = 0.402, p=0.013), CSS (HR=0.344, p=0.025), and DFS (HR=0.274, p=0.003). CB1 gene expression had no effect on survival. CB1 and CB2 gene expression was detected in 50% and 79.6% of the tumor tissue samples, respectively, and immunohistochemistry confirmed a correlation between CB2 gene expression and the presence of the CB2 protein in tumor tissues. Conclusion: In NSCLC patients undergoing radical surgery, CB2 gene expression in tumor tissue is associated with less advanced disease stage, fewer lymph node metastases, and longer survival (OS, CSS, DFS). Acknowledgement: This study was supported by Ministry of Health of the Czech Republic (NV18-03-00470), Ministry of Education, Youth and Sport of the Czech Republic (LM2018132), Palacky University Olomouc (LF 2020_007), Technological Agency of the Czech Republic (TN01000013), European Regional Development Fund (ENOCH CZ.02.1.01/0.0/0.0/16_019/0000868) and Cancer Research Czech Republic. Citation Format: Josef Srovnal, Monika Vidlarova, Emil Berta, Petr Prasil, Pavla Kourilova, Josef Chudacek, Marian Hajduch. Cannabinoid receptor 2 tumor tissue gene expression positively affects lung cancer survival following radical surgery [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 665.

Abstract 2601: Using chaperonin containing TCP1 as a marker to track clinically relevant circulating tumor cells

Abstract Current methods for assessing circulating tumor cells (CTC) in blood focus on capture and enumeration with epithelial markers that provide little information about the aggressive and invasive potential of CTC. As a result, valuable knowledge that improves monitoring of cancer recurrence and metastasis is lacking. To provide actionable information on CTC, we developed a CTC identification method based on detection of the second subunit of Chaperonin Containing TCP1 (CCT2). CCT is a macromolecular protein folding complex composed of eight subunits (CCT1-8) that drives expression of many tumorigenic factors. Bioinformatic analysis in multiple cancers, such as breast cancer, revealed increased gene expression in tumor tissue compared to normal tissue for CCT1-8 (p< 0.0001), with the largest differences observed in CCT2 and CCT3. Genomic alteration of CCT2 inversely correlated with cancer patient survival, and depletion of CCT2 prevented growth of murine breast tumors. In a correlative study of blood and tumor tissues from metastatic breast cancer patients, high CTC counts positively correlated with detectable CCT2 in tumor tissues. These results support CCT2 as a possible marker for CTC. To determine this, antibodies to CCT2 were validated for use in the CellSearch System (CSS), currently the only FDA approved CTC enrichment platform, and tested with breast cancer cells spiked into healthy human blood. Recovery using CCT2 for identification, led to increased collection up to 6% of spiked breast cancer cells that would have been lost by standard CTC classification using cytokeratin (CK). With prognostic thresholds around 3-5 CTC/tube of blood, increased detection of even 1-2 more CTC can be significant for assessing patients' risks. Analysis in an open software program called ACCEPT (Automated CTC Classification, Enumeration and PhenoTyping) confirmed that the majority of recovered spiked cancer cells expressed CCT2. Flow cytometry analysis of spiked cancer cells confirmed that CCT2+ enriched cells expressed cancer markers CD44 or estrogen receptor (ER). These results show that the addition of CCT2 staining to a CTC detection protocol has positive implications for enumeration of CTC from blood, such as increased detection of CTC with low expression of epithelial markers. This experiment was repeated using small cell lung cancer cell lines with similar results. The combined bioinformatics, histologic and experimental results indicate that identification of CCT2-expressing CTC can be incorporated into CTC detection platforms to monitor those patients with a heightened risk for cancer recurrence or metastasis. Citation Format: Amanda Cox, Ana Martini, Eunkyung Lee, Rebecca Moroose, Amr Khaled, Annette R. Khaled. Using chaperonin containing TCP1 as a marker to track clinically relevant circulating tumor cells [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 2601.

Serum levels of cytoskeleton remodeling proteins and their mRNA expression in tumor tissue of metastatic laryngeal and hypopharyngeal cancers

Actin-binding proteins (ABPs) and various signaling systems are involved in the process of squamous cell carcinoma of the larynx and hypopharynx (SCCLH) metastasis. The clinical significance of these proteins has not yet been determined. We analyzed the relationship between the mRNA levels of cofilin 1 (CFL1), profilin 1 (PFN1), adenylyl cyclase-associated protein 1 (CAP1), SNAI1 and RND3 and SCCLH metastasis. The serum levels of the above ABPs were estimated and the relationship between them and their mRNA expressions was analyzed. The expression levels of ABP mRNAs were measured by real-time RT-PCR in paired tissue samples taken from 54 patients with SCCLH (T1-4N0-1M0). Expression analysis was performed using the 2-ΔΔCT method. The levels of ABPs in the blood serum were measured by ELISA. Statistical analysis was carried out using the SPSS Statistica 20.0 software package. No significant difference in the mRNA gene expression in tumor tissue of patients with T1-3N0M0 SCCLH and patients with T2-4N1-2M0 SCCLH was found. High expression of RND3 mRNA was accompanied by an increase in mRNA expression of all studied ABPs. In the blood serum of T2-4N1-2M0 patients, the level of PFN1 was lower by 21% and the level of CAP1 was higher by 75% than those observed in T1-4N0M0 patients. The data obtained showed that RND3 is involved in the regulation of molecular cascades of SCCLH metastasis. PFN1 and CAP1 serum levels can be good classifiers of metastases in patients with SCCLH.

Identification of molecular heterogeneity of hepatocellular carcinoma based on immune gene expression signatures.

Although various molecular subtypes of hepatocellular carcinoma (HCC) have been investigated, most of these studies identify HCC subtype based on genomic profiling. Few studies have investigated the classification based on immune signatures, and none have classified HCC based on Immune activation and immunosuppressive. We performed immune gene expression of tumor tissue in 374 HCC patients from The Cancer Genome Atlas (TCGA) database and used unsupervised consensus clustering to stratify tumors. We then used HCC patients from the International Cancer Genome Consortium (ICGC) and Gene Expression Omnibus (GEO) as replication datasets. Based on the expression of 782 immune-related genes, HCC was stratified into four distinct immune subtypes. Tumors in one cluster (high immune activation; high-IA) indicate a higher level of Immune activation, which was characterized by higher anti-tumor immunity, higher pro-tumor immune-suppressive cell types, higher fractions of CD8+ T cells and M0 Macrophages compared with other subtypes. The high-IA also presents higher cancer-related hallmark signatures, such as epithelial-mesenchymal transition (EMT), angiogenesis, and apoptosis. We also found subpopulations of regulatory and exhaustion T lymphocyte were characterized by an opposite trend in high-IA, though samples in high-IA response to immunotherapy with better survival. The comparison of the immune profile in tumor and normal tissue indicates the activation of immune responses which only occurred in high-IA patients, while we conducted comparison of cirrhosis and non-cirrhosis tumor immune signatures, immune response activation was almost occurred in high-IA, but some of immune responses occurred in low-IA (low immune activation).

Reduction of T-Box 15 gene expression in tumor tissue is a prognostic biomarker for patients with hepatocellular carcinoma.

Genome-wide analysis is widely applied to detect molecular alterations during oncogenesis and tumor progression. We analyzed DNA methylation profiles of hepatocellular carcinoma (HCC), and investigated the clinical role of most heypermethylated of tumor, encodes T-box 15 (TBX15), which was originally involved in mesodermal differentiation. We conducted a genome-wide analysis of DNA methylation of tumor and non-tumor tissue of 15 patients with HCC, and revealed TBX15 was the most hypermethylated gene of tumor (Beta-value in tumor tissue = 0.52 compared with non-tumor tissue). Another validation set, which comprised 58 HCC with radical resection, was analyzed to investigate the relationships between tumor phenotype and TBX15 mRNA expression. TBX15 mRNA levels in tumor tissues were significantly lower compared with those of nontumor tissues (p < 0.0001). When we assigned a cutoff value = 0.5-fold, the overall survival 5-year survival rates of the low-expression group (n = 17) were significantly shorter compared with those of the high-expression group (n = 41) (43.3% vs. 86.2%, p = 0.001). Multivariate analysis identified low TBX15 expression as an independent prognostic factor for overall and disease-free survival. Therefore, genome-wide DNA methylation profiling indicates that hypermethylation and reduced expression of TBX15 in tumor tissue represents a potential biomarker for predicting poor survival of patients with HCC.

MicroRNA-1236-3p inhibits human osteosarcoma growth.

Osteosarcoma (OS) is a common bone tumor with high mortality worldwide. The long-term survival rate of patients with metastatic or recurrent disease is <20%. The present study explored the biological role of microRNA (miRNA/miR)-1236-3p in OS. miRNA and mRNA expression levels were measured via reverse transcription-quantitative PCR. Fluorescence in situ hybridization was performed to determine miR-1236-3p expression levels in clinical specimens. Protein expression was measured via western blotting. Immunohistochemical analysis was used to detect Wnt target gene expression in tumor tissues. The interaction between the Wnt3a 3′untranslated region and miR-1236-3p was assessed via dual-luciferase reporter assays. Cell cycle, Transwell, Cell Counting Kit-8 and wound healing assays were conducted to evaluate the function of the miR-1236-3p/Wnt3a axis. Human OS (HOS) cells stably transfected with vector or miR-1236-3p sponge were injected subcutaneously into nude mice to assess the role of miR-1236-3p in vivo. miR-1236-3p expression was downregulated in OS tissues compared with chondroma tissues, and miR-1236-3p overexpression inhibited OS cell migration and proliferation compared with the negative control group. Furthermore, in vivo xenograft assays displayed enhanced tumour growth rates in the miR-1236-3p sponge group compared with the vector control group. In the present study, the results indicated that miR-1236-3p inhibited OS progression and Wnt3a was identified as a target of miR-1236-3p.

Effects of Low Dose Radiation on Biological Processes in Peri-tumoral and Tumoral Areas n Squamous Cell Skin Cancer

Very low dose ionizing radiation (LDR) may have paradoxically beneficial effects via an adaptive response. This adaptive response can mitigate against the potential adverse effects of high-dose radiation therapy and chemotherapy. In this study we aimed to:1.identify the specific molecular details behind the adaptive response to LDR and2.integrate the LDR adaptive response into clinical practice. Our aim was to determine how low dose radiation modifies genetic expression in both healthy and tumor tissue. To determine this, a human clinical trial (n=8) was conducted in 2018 to ascertain the biological effects of integrating 0.1Gy of LDR into standard radiotherapy (2Gy fractions) over the first two weeks of radiation treatment for epidermoid skin cancers. RNA was obtained from skin biopsies and processed for genetic expression using Affymetrix Human gene chips Clarion-D (138K genes). 8 patients with squamous skin cancer, were studied according to the experimental design showed in Figure 1. Biopsy samples were collected and total RNA preparation (100 ng), labeled cRNA synthesis, hybridization, scanning, and image analysis were performed. Changes in biological response were measured using Affymetrix microarrays. Experiments 17F1 and 17F2 were performed in duplicate, and experiments 18F3 and 18F4 were performed in triplicate. Pathway analysis was performed using the Affymetrix Transcriptome Analysis Console software and the Wikipaths database. Comparisons were performed as noted in Table 1. The difference in gene expression in healthy tissues due to LDR exposure is quite low (only 325 genes, mostly downregulated) as noted in Figures 2A and 3A and Tables 2 and 3. The 20 most important biological processes enriched are shown in Figure 4 and include Senescence and Autophagy in malignant cells, and the Genotoxicity Pathway (both downregulated). The difference in gene expression in tumor tissue due to LDR exposure is significantly more pronounced (5,651 genes, mostly upregulated) as noted in Figures 2C and 3C. The 20 most important biological processes enriched are shown in Figures 5 and 6 and Tables 4-6, and include the Allograft rejection pathway in skin cancer (upregulated). Our results suggest that very low doses of radiation (0.1Gy) have a narrow effect on gene expression when applied in the healthy peritumoral area. Down regulated genes predominate, and DNA repair mechanisms are stimulated by the first low dose in healthy tissue.

Abstract 6450: Circulating hepsin as a novel serum biomarker in prostate cancer patients

Abstract Hepsin, a type II transmembrane serine protease, is commonly overexpressed in a variety of epithelial cancers, where its overexpression and concurrent proteolytic activity correlate with tumor progression. Recent studies indicate that Hepsin undergoes activation via autocatalysis and subsequent ectodomain shedding, thus highlighting its potential as a serum-based biomarker. Using transgenic mouse models of prostate adenocarcinoma, Hepsin overexpression has previously been implicated in disease progression and, notably, metastases that are neuroendocrine in nature. Collectively, these findings led us to explore the potential diagnostic utility of Hepsin expression in prostate cancer progression, inclusive of metastatic variants. Here, we reanalyzed pre-existing, publicly available gene expression datasets (2 datasets, 326 patient samples), which revealed a significant upregulation of Hepsin gene expression in tumor tissue versus benign tissue and further pronounced upregulation in androgen-dependent and -independent metastatic lesions. To further examine the potential correlation of Hepsin overexpression to androgen-independence, we reanalyzed in vitro data comparing LNCaP parental versus androgen-independent LNCaP subclones. This analysis revealed a stepwise increase in Hepsin expression over the course of androgen deprivation, which was accompanied by a concomitant decrease in PSA expression. Lastly, to evaluate circulating hepsin protein levels as they occur in patient serum, a total of 424 subjects suspected of prostate cancer were analyzed for circulating Hepsin levels from 2015 to 2018 at the time of biopsy and followed for recurrence or survival until the end of the study. Serum Hepsin levels, as determined by enzyme-linked immunosorbent assay, rendered a specificity of 89% (Gleason &amp;lt; 6 (3+3); Hepsin positive, &amp;gt;100 ng/mL). Within this cohort, 18 patients presented with biochemical recurrence, of which 44% were Hepsin positive and exhibited a 22-month accelerated clinical path to recurrence versus Hepsin negative patients. Collectively, these results highlight the longitudinal diagnostic potential of circulating Hepsin levels in the prostate cancer setting. Citation Format: John Beard, Sam Eddington, Noah Bowman, Adam Cole, Timothy O'Brien, Blake P. Johnson. Circulating hepsin as a novel serum biomarker in prostate cancer patients [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 6450.

Metformin is distributed to tumor tissue in breast cancer patients in vivo: A 11C-metformin PET/CT study.

Epidemiological studies and randomized clinical trials suggest that the antidiabetic drug, metformin, may have anti-neoplastic effects. The mechanism that mediates these beneficial effects has been suggested to involve direct action on cancer cells, but this will require distribution of metformin in tumor tissue. The present study was designed to investigate metformin distribution in vivo in breast and liver tissue in breast cancer patients. Seven patients recently diagnosed with ductal carcinoma were recruited. Using PET/CT, tissue distribution of metformin was determined in vivo for 90min after injection of a carbon-11-labeled metformin tracer. After surgery, tumor tissue was investigated for gene expression levels of metformin transporter proteins. Tumor tissue displayed a distinct uptake of metformin compared to normal breast tissue AUC0-90min (75.4 ± 5.5 vs 42.3 ± 6.3) g/ml*min (p = 0.01). Maximal concentration in tumor was at 1min where it reached approximately 30% of the activity in the liver. The metformin transporter protein with the highest gene expression in tumor tissue was multidrug and toxin extrusion 1 (MATE 1) followed by plasma membrane monoamine transporter (PMAT). This study confirms that metformin is transported into tumor tissue in women with breast cancer. This finding support that metformin may have direct anti-neoplastic effects on tumor cells in breast cancer patients. However, distribution of metformin in tumor tissue is markedly lower than in liver, an established metformin target tissue.

Experimental Study on the Effect of a Weifufang on Human Gastric Adenocarcinoma Cell Line BGC-823 Xenografts and PTEN Gene Expression in Nude Mice.

Background: This study aims at investigating the effect of the Weifufang, an effective prescription for the treatment of gastric cancer developed by the Traditional Chinese Medicine (TCM)/Combination of TCM and Western Medicine Department of the Hunan Cancer Hospital, on gastric cancer xenografts in nude mice and its effect on the PTEN gene; it also aims at exploring the possible tumor suppression mechanism. Methods: Nude mice with xenografts were treated with different concentrations of the Weifufang for 2 weeks, and changes in tumor volume were observed. The histopathology of the tumor was detected by hematoxylin and eosin staining; PTEN gene expression in tumor tissues was detected by immunohistochemistry (IHC) and western blot. Results: After 2 weeks of treatment, tumor inhibition rates in the 5-flourouracil (5-FU) group, and in the Weifufang low-, middle-, and high-dose groups were 30.67%, 19%, 49.52%, and 29.36%, respectively. The IOD of the PTEN gene was detected by IHC. The values in the water group, the 5-FU group, and the Weifufang low-, middle-, and high-dose groups were 0.013 ± 0.004, 0.085 ± 0.062, 0.041 ± 0.024, 0.128 ± 0.032, and 0.061 ± 0.052, respectively. Except for the 5-FU group, the differences between the gastric compound middle dose-group and the other groups were statistically significant (p < 0.05). Results of PTEN expression detection by western blot: The expression levels in the water group, 5-FU group, and the Weifufang low-, middle-, and high-dose groups were 0.2240 ± 0.0172, 0.4200 ± 0.0228, 0.2760 ± 0.0163, 0.3840 ± 0.0133, and 0.3040 ± 0.0211, respectively. Except for the 5-FU group, differences between the Weifufang middle-dose group and the other groups were statistically significant (p < 0.05). Conclusion: The Weifufang may inhibit the growth of gastric cancer xenografts by upregulating PTEN gene expression. The middle-dose group had the best effect.

Bioinformatic identification of renal cell carcinoma microenvironment-associated biomarkers with therapeutic and prognostic value

Renal cell carcinoma (RCC) is the ninth most prevalent form of malignancy worldwide. The tumor microenvironment significantly affects gene expression in tumor tissues, which subsequently impacts the prognosis of RCC patients. Available datasets such as The Cancer Genome Atlas (TCGA) can be utilized to improve diagnostic methods and search for novel tumor therapeutic targets and prognostic biomarkers. The current study used the ESTIMATE algorithm to explore the immune and stromal components in RCC. Differentially expressed genes (DEGs) were identified by comparing the gene expression patterns in groups with high and low immune/stromal scores. Functional enrichment analysis was conducted and Kaplan-Meier survival curves were plotted to explore the functions of the DEGs in the tumorigenesis, progression, and prognosis of RCC. Our results revealed that immune and stromal scores are associated with specific clinicopathologic variables in RCC. These variables include gender, tumor grade, tumor stage, tumor size, distant metastasis and prognosis. A total of 48 upregulated and 47 downregulated genes were obtained. Functional enrichment analysis demonstrated a correlation between DEGs and the tumor microenvironment, tumor immune response and RCC tumorigenesis. Kaplan-Meier survival curves showed that 43 out of the 48 identified tumor microenvironment related genes are involved in the prognosis of RCC. Three genes, IL10, IGLL5 and POU2AF1, were selected as the hub genes, and their kinase targets were identified as MAPK1 and PPKCA. A positive correlation was obtained between the expression of IL/POU2AF1 and the abundance of six immune cells. Our study provides potential biomarkers for the therapy and prognosis of RCC.

383TiP - Bintrafusp alfa (M7824) and eribulin mesylate in treating patients with metastatic triple negative breast cancer (TNBC)(NCT03579472)

BackgroundTNBC is an aggressive subtype of metastatic breast cancer. Eribulin has been shown to have activity in metastatic TNBC and has enhanced efficacy if TGF-β is downregulated. Additionally, down-regulating TGF-β in the tumor microenvironment may decrease tumor growth and increase responsiveness to checkpoint blockade. Bintrafusp alfa* (M7824) is an innovative first-in-class bifunctional fusion protein composed of the extracellular domain of the TGF-βRII receptor (a TGF-β “trap”) fused to a human IgG1 mAb blocking PD-L1. Therefore the combination of eribulin with bintrafusp alfa is being tested in this phase I trial to evaluate safety and efficacy for patients with metastatic TNBC. Trial designThis is an open-label, single arm phase Ib study in patients with metastatic TNBC. The primary objective is to evaluate the recommended phase-2 dosing of eribulin and bintrafusp. Patients undergo core tissue biopsy and blood work at enrollment. Patients will then receive study drug (bintrafusp alfa) and eribulin with tumor assessments at 6 and 12 weeks. Patients will be treated in a cohort size of 4 with an expansion of up to 20 patients. Bayesian optimal interval (BOIN) design will be employed to identify the RP2D of eribulin by conducting dose de-escalations based on dose-limiting toxicity. Patients undergo repeat core tissue biopsy after 12 weeks of therapy and will continue on study drug until progression of disease or a sustained CR for one year. Eligibility: Inclusion: metastatic TNBC, adequate organ, bone marrow and cardiac parameters. Exclusion: prior immunotherapy, IBC, history of autoimmune disease, HIV, Hep-B, Hep-C, active tuberculosis, pregnant. PD-L1 positivity is not an eligibility requirement. CORRELATIVE SCIENCE: Biopsies and blood are being obtained at baseline and after 2 months of therapy as well as requested at the time of progression of disease. Peripheral blood mononuclear cells (PBMCs will be evaluated for immunophenotyping, T cell activation and function as well as T cell repertoire analysis. PD-L1 staining will be done on tumor and infiltrating immune cells. From the tumor samples, TIL will be measured and characterized as well as tumor-tissue gene expression. Clinical trial identificationNCT03579472. Legal entity responsible for the studyThe University of Texas MD Anderson Cancer Center. FundingMerck/EMD-Serono. DisclosureJ.K. Litton: Research grant / Funding (institution): astra zeneca; Advisory / Consultancy, uncompensated: astra zeneca; Advisory / Consultancy, uncompensated: Pfizer; Research grant / Funding (institution): Pfizer; Research grant / Funding (institution): EMD Serono; Research grant / Funding (institution): Novartis; Advisory / Consultancy, uncompensated: Genentech; Research grant / Funding (institution): Genentech. S. Damodaran: Research grant / Funding (institution): emd serono. I.I. Wistuba: Research grant / Funding (institution): emd serono. L. Ojalvo: Full / Part-time employment: emd serono. I. Dussault: Full / Part-time employment: emd serono. C. Helwig: Full / Part-time employment: Merck. All other authors have declared no conflicts of interest.

Preliminary investigation into construction of sensitivity model of adjuvant radiotherapy in cervical cancer based on gene expression data in vivo

Objective To construct a radiosensitivity model based on gene expression in tumor tissues, and to explore the feasibility of using this model to select the patients benefiting from adjuvant radiotherapy in cervical cancer. Methods Patients underwent radical hysterectomy alone or radical hysterectomy plus adjuvant radiotherapy were selected from The Cancer Genome Atlas (TCGA) cervical cancer dataset. Candidate radiosensitive genes were identified through multivariate Cox regression analysis with recurrence free survival (RFS) as end-point events. The sum of regression coefficients and products of expression levels of candidate genes was calculated as radiosensitivity index. Time-dependent receiver operating characteristic (ROC) curve analysis was used to analyze the predictive accuracy of this model for RFS. Kaplan-Meier plot coupled with Log-rank test was used to evaluate difference in RFS between subgroups with different radiosensitivity. Results A total of 155 patients underwent radical hysterectomy was retrieved from the phenotype dataset among which 18 patients was completely treated with planned adjuvant radiotherapy. Expression values of 17 156 genes and 480 microRNAs after probes filtration were included into analysis. Twenty-three genes were identified consistent with statistical criterion. Hierarchical cluster analysis revealed 7 subgroups existed across these 23 genes. The MAT2A, PIGY, SUSD1, ZNF341, TMEM85, HIF1AN and CLEC18A genes were finally selected to calculate radiosensitivity index. The median value and range of RI was 0.7258 (-5.501 - 9.046). In subset consist of patients underwent radical hysterectomy alone, adjusted for intravascular tumor thrombus, radiosensitivity index was independent prognostic factor (HR = 1.456, 95% CI 1.192-1.780, P < 0.01). Whereas, radiosensitivity index was remained significantly associated with RFS after adjustment for the number of positive nodes in subset comprised of patients underwent radical hysterectomy plus adjuvant radiotherapy (HR = 0.572, 95% CI 0.328-0.998, P = 0.049). Moreover, radiosensitivity index exhibited more accuracy in prediction for 3 or 5 years RFS compared with clinical prognostic factors N stage and intravascular tumor thrombus. Conclusion The radiosensitivity model based on gene expression data in tumor tissues has potential value to select the cervical cancer patients benefiting from adjuvant radiotherapy. Key words: Uterine cervical neoplasms; Gene expression; Neoadjuvant therapy; Radiation tolerance; Radiotherapy

Prolonged survival of renal cancer patients is concomitant with a higher regucalcin gene expression in tumor tissues: Overexpression of regucalcin suppresses the growth of human renal cell carcinoma cells in vitro.

Renal cell carcinoma (RCC), which is a type of cancer found in the kidney tubule, is among the 10most frequently occurring human cancers. Regucalcin plays a potential role as a regulator of transcriptional activity, and its downregulated expression or activity may contribute to the promotion of human cancers. In this study, we investigated the involvement of regucalcin in human RCC. Regucalcin expression was compared in 23normal and 29tumor samples of kidney cortex tissues of patients with clear cell RCC obtained through the Gene Expression Omnibus (GEO) database (GSE36895). Regucalcin expression was downregulated in the tumor tissues. The prolonged survival of patients with clear cell RCC was demonstrated to be associated with a higher regucalcin gene expression in the TCGA dataset. The overexpression of regucalcin suppressed the colony formation, proliferation and the death of human clear cell RCC A498 cells invitro. Mechanistically, the overexpression of regucalcin induced the G1andG2/M phase cell cycle arrest of A498 cells through the suppression of multiple signaling components, including Ras, PI3 kinase, Akt and mitogen‑activated protein(MAP) kinase. Importantly, the overexpression of regucalcin led to an elevation in the levels of the tumor suppressors, p53, Rb and the cell cycle inhibitor, p21. The levels of the transcription factors, c‑fos, c‑jun, nuclear factor‑κB p65, β‑catenin and signal transducer and activator of transcription3, were suppressed by regucalcin overexpression. On the whole, the findings of this study suggest that regucalcin plays a suppressive role in the promotion of human RCC. The overexpression of regucalcin by gene delivery systems may thus prove to be a novel therapeutic strategy for RCC.

Lentinan inhibits tumor angiogenesis via interferon \u03b3 and in a T cell independent manner

BackgroundAntiangiogenic agents are commonly used in lung and colon cancer treatments, however, rapid development of drug resistance limits their efficacy.MethodsLentinan (LNT) is a biologically active compound extracted from Lentinus edodes. The effects of LNT on tumor angiogenesis were evaluated by immunohistochemistry in murine LAP0297 lung and CT26 colorectal tumor models. The impacts of LNT on immune cells and gene expression in tumor tissues were determined by flow cytometry, qPCR, and ELISA. Nude mice and IFNγ blockade were used to investigate the mechanism of LNT affecting on tumor angiogenesis. The data sets were compared using two-tailed student’s t tests or ANOVA.ResultsWe found that LNT inhibited tumor angiogenesis and the growth of lung and colon cancers. LNT treatments elevated the expression of angiostatic factors such as IFNγ and also increased tumor infiltration of IFNγ-expressing T cells and myeloid cells. Interestingly, IFNγ blockade, but not T cell deficiency, reversed the effects of LNT treatments on tumor blood vessels. Moreover, long-lasting LNT administration persistently suppressed tumor angiogenesis and inhibited tumor growth.ConclusionsLNT inhibits tumor angiogenesis by increasing IFNγ production and in a T cell-independent manner. Our findings suggest that LNT could be developed as a new antiangiogenic agent for long-term treatment of lung and colon cancers.

Prolonged survival of patients with colorectal cancer is associated with a higher regucalcin gene expression: Overexpression of regucalcin suppresses the growth of human colorectal carcinoma cells invitro.

Regucalcin plays a crucial role as a regulator of transcriptional signaling activity, and its decreased expression or activity may contribute to the promotion of human carcinogenesis. A higher regucalcin expression in the tumor tissues has been demonstrated to prolong the survival of patients with various types of cancer, including pancreatic cancer, breast cancer, liver cancer and lung adenocarcinoma. The involvement of regucalcin in human colorectal cancer was investigated in the current study. Regucalcin gene expression and the survival data of 62patients with colorectal cancer were obtained though the Gene Expression Omnibus(GEO) database (GSE12945) for outcome analysis. The data of gene expression revealed that the prolonged survival of patients with colorectal cancer was associated with a higher regucalcin gene expression in tumor tissues. The overexpression of regucalcin suppressed colony formation and proliferation, and induced the death of human colorectal carcinoma RKO cells cultured in a medium containing fetal bovine serum invitro. Mechanistically, the overexpression of regucalcin induced the G1 and G2/M phase cell cycle arrest of the RKO cells through the suppression of multiple signaling pathways, including Ras, Akt, mitogen-activated protein (MAP) kinase and SAPK/JNK. Of note, the overexpression of regucalcin induced an increase in the levels of the tumor suppressors, p53 and Rb, and the cell cycle inhibitor, p21. Moreover, the levels of the transcription factors, c‑fos, c‑jun, nuclear factor (NF)‑κB p65, β-catenin and signal transducer and activator of transcription3 (Stat3), were suppressed by the overexpression of regucalcin. On the whole, the findings of this study suggest that regucalcin plays a crucial role as a suppressor in human colorectal cancer, and that the suppressed expression of the regucalcin gene may predispose patients to the promotion of colorectal cancer. The overexpression of regucalcin by gene delivery may thus prove to be a novel therapeutic strategy for colorectal cancer.

Development of a computational promoter with highly efficient expression in tumors

BackgroundGene therapy is a potent method to increase the therapeutic efficacy against cancer. However, a gene that is specifically expressed in the tumor area has not been identified. In addition, nonspecific expression of therapeutic genes in normal tissues may cause side effects that can harm the patients’ health. Certain promoters have been reported to drive therapeutic gene expression specifically in cancer cells; however, low expression levels of the target gene are a problem for providing good therapeutic efficacy. Therefore, a specific and highly expressive promoter is needed for cancer gene therapy.MethodsBioinformatics approaches were utilized to analyze transcription factors (TFs) from high-throughput data. Reverse transcription polymerase chain reaction, western blotting and cell transfection were applied for the measurement of mRNA, protein expression and activity. C57BL/6JNarl mice were injected with pD5-hrGFP to evaluate the expression of TFs.ResultsWe analyzed bioinformatics data and identified three TFs, nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB), cyclic AMP response element binding protein (CREB), and hypoxia-inducible factor-1α (HIF-1α), that are highly active in tumor cells. Here, we constructed a novel mini-promoter, D5, that is composed of the binding sites of the three TFs. The results show that the D5 promoter specifically drives therapeutic gene expression in tumor tissues and that the strength of the D5 promoter is directly proportional to tumor size.ConclusionsOur results show that bioinformatics may be a good tool for the selection of appropriate TFs and for the design of specific mini-promoters to improve cancer gene therapy.

Early diagnostic role of PSA combined miR-155 detection in prostate cancer.

As a kind of malignant tumor in the male genitourinary system, prostate cancer exhibits significantly increased occurrence. Prostate-specific antigen (PSA) expression can be seen in the prostate cancer, prostatitis, and other diseases, therefore, lack of diagnostic specificity. The miR-155 expression is abnormally increased in the tumors. Therefore, this study aims to explore the clinical significance of PSA combined miR-155 detection in the early diagnosis of prostate cancer. A total of 86 patients diagnosed with prostate cancer were enrolled in this study. PSA and miR-155 gene expression in tumor tissue were detected by using Real-time PCR. The serum levels of PSA were measured by using enzyme-linked immunosorbent assay (ELISA). The correlation of PSA and miR-155 expression with age, body mass index (BMI), tumor volume, tumor-node-metastasis (TNM) stage, lymph node metastasis (LNM), and other clinicopathological features were analyzed, respectively. Serum PSA expression and PSA gene in tumor tissue were significantly higher compared to that in adjacent tissues (p<0.05). PSA gene and protein increased significantly with the clinical stage of TNM and decreased following the increase of grade (p<0.05). The miR-155 level was significantly elevated in the tumor tissue compared with para-carcinoma tissue (p<0.05). PSA and miR-155 expressions were positively correlated with TNM stage, tumor volume, and LNM, and negatively correlated with grade (p<0.05). PSA and miR-155 were closely related to the clinicopathological features of prostate cancer. Combined detection is helpful for the early diagnosis of prostate cancer.

Reduction of T-Box 15 gene expression in tumor tissue as a prognostic biomarker for patients with hepatocellular carcinoma.

341 Background: Genome-wide analysis is widely applied to detect molecular alterations in tumor and non-tumor tissue during oncogenesis and tumor progression. We analyzed DNA methylation profiles of hepatocellular carcinoma (HCC), which was is the third-highest cause of cancer-related deaths worldwide, and investigated the clinical role of most heyper-methylated gene, encodes T-box 15 (TBX15), which was originally involved in mesodermal differentiation. Methods: We used an Illumina Infinium HumanMethylation450 BeadChip Kit to conduct a genome-wide analysis of DNA methylation of tumor and non-tumor tissue of 15 patients with HCC. Methylation scores for CpG sites were assigned a Beta-value. Another validation set, which comprised 58 patients with HCC who underwent radical resection, was analyzed to investigate the relationships between tumor phenotype, prognosis, and TBX15 mRNA expression. Results: TBX15 was the most hyper-methylated gene (Beta-value in tumor tissue = 0.52 compared with non-tumor tissue), and TBX15 mRNA levels in tumor tissues were significantly lower compared with those of non-tumor tissues (p &lt; 0.0001). When we assigned a cutoff value = 0.5-fold to define differential expression of TBX15 mRNA, low TBX15 expression significantly correlated with higher serum DCP levels, and the overall survival 5-year survival rates of the low-expression group (n = 17) were significantly shorter compared with those of the high-expression group (n = 41) (43.3% vs. 86.2%, p = 0.001). Multivariate analysis identified low TBX15 expression as an independent prognostic factor for overall and disease-free survival. Conclusions: Genome-wide DNA methylation profiling indicates that hypermethylation and reduced expression of TBX15 in tumor tissue represents a potential biomarker for predicting tumor progression and poor survival of patients with HCC.

Design of the First-in-Class, Highly Potent Irreversible Inhibitor Targeting the Menin-MLL Protein-Protein Interaction.

The structure-based design of M-525 as the first-in-class, highly potent, irreversible small-molecule inhibitor of the menin-MLL interaction is presented. M-525 targets cellular menin protein at sub-nanomolar concentrations and achieves low nanomolar potencies in cell growth inhibition and in the suppression of MLL-regulated gene expression in MLL leukemia cells. M-525 demonstrates high cellular specificity over non-MLL leukemia cells and is more than 30 times more potent than its corresponding reversible inhibitors. Mass spectrometric analysis and co-crystal structure of M-525 in complex with menin firmly establish its mode of action. A single administration of M-525 effectively suppresses MLL-regulated gene expression in tumor tissue. An efficient procedure was developed to synthesize M-525. This study demonstrates that irreversible inhibition of menin may be a promising therapeutic strategy for MLL leukemia.