Protein Expression In Cancer Tissues Research Articles

Identification and Quantification of Radiotherapy-related Protein Expression in Cancer Tissues Using the Qupath Software and Prediction of Treatment Response.

Automated measurement of immunostained samples can enable more convenient and objective prediction of treatment outcome from radiotherapy. We aimed to validate the performance of the QuPath image analysis software in immune cell markers detection by comparing QuPath cell counting results with those of physician manual cell counting. CD8- and FoxP3-stained cervical, CD8-stained oropharyngeal, and Ku70-stained prostate cancer tumor sections were analyzed in 104 cervical, 92 oropharyngeal, and 58 prostate cancer patients undergoing radiotherapy at our Institution. QuPath and manual counts were highly correlated. When divided into two groups using ROC curves, the agreement between QuPath and manual counts was 89.4% for CD8 and 88.5% for FoxP3 in cervical cancer, 87.0% for CD8 in oropharyngeal cancer and 80.7% for Ku70 in prostate cancer. In cervical cancer, the high CD8 group based on QuPath counts had a better prognosis and the low CD8 group had a significantly worse prognosis [p=0.0003; 5-year overall survival (OS), 65.9% vs. 34.7%]. QuPath counts were more predictive than manual counts. Similar results were observed for FoxP3 in cervical cancer (p=0.002; 5-year OS, 62.1% vs. 33.6%) and CD8 in oropharyngeal cancer (p=0.013; 5-year OS, 80.2% vs. 47.2%). In prostate cancer, high Ku70 group had worse and low group significantly better outcome [p=0.007; 10-year progression-free survival (PFS), 56.0% vs. 93.8%]. QuPath showed a strong correlation with manual counting, confirming its utility and accuracy and potential applicability in clinical practice.

Effects of high-risk human papillomavirus infection on P53, pRb, and survivin in lung adenocarcinoma-a retrospective study.

To observe the effects of high-risk human papillomavirus (HR-HPV) infection on P53, pRb, and survivin in lung adenocarcinoma (LUAD). The cancerous and cancer-adjacent tissues of 102 patients with LUAD from January 2020 to April 2022 were selected for the study. HR-HPV infection was detected by flow fluorescence method, and P53, pRb, and survivin protein expression was detected by immunohistochemical staining method. Statistical analysis was performed to determine the differences in the HR-HPV infection and the expression of P53, pRb, and survivin proteins between LUAD tissues and cancer-adjacent tissues; the correlation between HR-HPV infection and P53, pRb, and survivin protein expression in cancer tissues; and the correlation between HR-HPV infection and clinicopathological features of LUAD. The infection rate of HR-HPV was higher in the LUAD tissues (28.43%) than in cancer-adjacent tissues (7.84%), and the difference was statistically significant (P<0.05). The positive rates of P53 and survivin protein were higher in the LUAD group (33.33% and 67.16%, respectively) than in the cancer-adjacent group (3.92% and 11.73%, respectively), and the difference was statistically significant (P<0.05). The positive rate of pRb protein was lower in the LUAD group (58.82%) than in the cancer-adjacent group (92.14%), and the difference was statistically significant (P<0.05). The positive rates of P53 and survivin proteins were significantly higher in the HR-HPV LUAD group (58.62% and 86.21%, respectively) than in the non-HR-HPV LUAD group (41.38% and 67.12%, respectively), and the difference was statistically significant (P<0.05). The expression rate of pRb protein was significantly lower in the HR-HPV LUAD group (37.93%) than in the non-HR-HPV LUAD group (67.12%), and the difference was statistically significant (P<0.05). The expression of p53 and survivin protein was positively correlated with HR-HPV infection (r=0.338 and 0.444, P<0.05), whereas the expression of pRb protein was negatively correlated with HR-HPV infection (r=-0.268, P<0.05). HR-HPV infection was not associated with gender, age, and smoking in patients with LUAD (P>0.05). HR-HPV infection was associated with lymph node metastasis and clinical stage of LUAD (P<0.05). HR-HPV infection was associated with lymph node metastasis and clinical stage of LUAD, which may be achieved by up-regulating p53 and survivin protein expression and down-regulating pRb protein expression.

Characterization of TIM-3 Expression and Its Correlation with TNF-α and IFN-γ in Patients with Surgically Resected Lung Adenocarcinoma.

T cell immunoglobulin and mucin-containing protein-3 (TIM-3) is an important immune checkpoint, but its role in lung cancer is still not clear. In this study, we investigated TIM-3 protein expression and its correlation with TNF-α and IFN-γ by examining the tissues of patients with lung adenocarcinoma. We detected the mRNA quantity of TIM-3, TNF-α, and IFN-γ in 40 surgically resected specimens from patients with lung adenocarcinoma by real-time quantitative polymerase chain reaction (qRT-PCR). The protein expression of TIM-3, TNF-α, and IFN-γ was assessed in normal tissues, paracarcinoma tissues, and tumor tissues by western blotting, respectively. The relevance between the expression and clinicopathological information of the patients was analyzed. The results showed that the expression level of TIM-3 was higher in tumor tissues than normal tissues and paracancerous tissues (P < 0.05). On the contrary, the expression of TNF-α and IFN-γ in tumor tissues was lower than normal tissues and paracarcinoma tissues (P < 0.05). However, the expression levels of IFN-γ mRNA were not observed to be significantly different between cancerous tissues and adjacent tissues. While TIM-3 protein expression in cancer tissues of patients with lymph node metastasis was higher than in patients without metastasis, the expression of TNF-α and IFN-γ was lower (P < 0.05). Importantly, the expression of TIM-3 was negatively correlated with the expression of TNF-α and IFN-γ, and the expression of TNF-α was found to be positively correlated with IFN-γ in the patient. The high expression of TIM-3, the low expression of TNF-α and IFN-γ, and the synergistic effect of TNF-α and IFN-γ in patients with lung adenocarcinoma were closely related to poor clinicopathological characteristics. Overexpression of TIM-3 may play an important role in the relationship between TNF-α and IFN-γ secretion and poor clinicopathological characteristics.

Analysis of the Clinical Value of MAGE-A9 Expressions in Cervical Cancer Tissues and PBMC.

Objective The aim of this study is to explore the expressions and clinical significance of melanoma-associated antigen-A9 (MAGE-A9) in cervical cancer tissues and peripheral blood mononuclear cells (PBMC). Methods 108 patients who were scheduled to undergo cervical conization or extensive hysterectomy between March 2019 and January 2021 due to cervical lesions were selected by convenient sampling. According to postoperative pathological results, the patients were divided into a cervical cancer group (n = 64) and cervical intraepithelial neoplasia (CIN) group (n = 44). The expression levels of MAGE-A9 mRNA in cervical lesion tissues and PBMC were detected by real-time fluorescence quantitative PCR, and the expression of MAGE-A9 protein in lesion tissues was detected by immunohistochemistry. The correlation between MAGE-A9 mRNA expressions in cancer tissues and PBMC and serum tumor markers in patients with cervical cancer and the relationship between MAGE-A9 protein expression in cancer tissues and clinicopathological characteristics were analyzed, and a receiver operating characteristic curve (ROC curve) was drawn to explore the diagnostic value of MAGE-A9 mRNA expressions in cancer tissues and PBMC on cervical cancer. Results The expression levels of MAGE-A9 mRNA in cervical lesion tissues and PBMC in the cervical cancer group were significantly higher than those in the CIN group (P < 0.05), and the levels of serum SCC-Ag, CA-125, and CEA were significantly higher than those in the CIN group (P < 0.05). The positive rate of the MAGE-A9 protein expression in cervical lesion tissues in the cervical cancer group was significantly higher than that in the CIN group (P < 0.05). The expression levels of MAGE-A9 mRNA in cancer tissues and PBMC of patients with cervical cancer were positively correlated with serum SCC-Ag, CA-125, and CEA (P < 0.05). The positive rate of the MAGE-A9 protein expression in cervical cancer tissues was related to FIGO stage, tumor diameter, degree of differentiation, lymph node metastasis, and high-risk HPV infection (P < 0.05) and was not correlated with age and pathological type (P > 0.05). The areas under the ROC curves of MAGE-A9 mRNA in lesion tissue and MAGE-A9 mRNA in PBMC were 0.925 and 0.900 in the diagnosis of cervical cancer (P < 0.05). Conclusion The expressions of MAGE-A9 in cancer tissues and PBMC of patients with cervical cancer are upregulated, which is related to the levels of serum tumor markers and the progression of disease. MAGE-A9 is expected to become an important marker for the diagnosis of early cervical cancer.

Single-Cell Transcriptome Analysis Reveals RGS1 as a New Marker and Promoting Factor for T-Cell Exhaustion in Multiple Cancers.

T-cell exhaustion is one of the main reasons of tumor immune escape. Using single-cell transcriptome data of CD8+ T cells in multiple cancers, we identified different cell types, in which Pre_exhaust and exhausted T cells participated in negative regulation of immune system process. By analyzing the coexpression network patterns and differentially expressed genes of Pre_exhaust, exhausted, and effector T cells, we identified 35 genes related to T-cell exhaustion, whose high GSVA scores were associated with significantly poor prognosis in various cancers. In the differentially expressed genes, RGS1 showed the greatest fold change in Pre_exhaust and exhausted cells of three cancers compared with effector T cells, and high expression of RGS1 was also associated with poor prognosis in various cancers. Additionally, RGS1 protein was upregulated significantly in tumor tissues in the immunohistochemistry verification. Furthermore, RGS1 displayed positive correlation with the 35 genes, especially highly correlated with PDCD1, CTLA4, HAVCR2, and TNFRSF9 in CD8+ T cells and cancer tissues, indicating the important roles of RGS1 in CD8+ T-cell exhaustion. Considering the GTP-hydrolysis activity of RGS1 and significantly high mRNA and protein expression in cancer tissues, we speculated that RGS1 potentially mediate the T-cell retention to lead to the persistent antigen stimulation, resulting in T-cell exhaustion. In conclusion, our findings suggest that RGS1 is a new marker and promoting factor for CD8+ T-cell exhaustion and provide theoretical basis for research and immunotherapy of exhausted cells.

SNHG17 promotes colorectal tumorigenesis and metastasis via regulating Trim23-PES1 axis and miR-339-5p-FOSL2-SNHG17 positive feedback loop

BackgroundSmall nucleolar RNA host gene (SNHG) long noncoding RNAs (lncRNAs) are frequently dysregulated in human cancers and involved in tumorigenesis and progression. SNHG17 has been reported as a candidate oncogene in several cancer types, however, its regulatory role in colorectal cancer (CRC) is unclear.MethodsSNHG17 expression in multiple CRC cohorts was assessed by RT-qPCR or bioinformatic analyses. Cell viability was evaluated using Cell Counting Kit-8 (CCK-8) and colony formation assays. Cell mobility and invasiveness were assessed by Transwell assays. Tumor xenograft and metastasis models were applied to confirm the effects of SNHG17 on CRC tumorigenesis and metastasis in vivo. Immunohistochemistry staining was used to measure protein expression in cancer tissues. RNA pull-down, RNA immunoprecipitation, chromatin immunoprecipitation, and dual luciferase assays were used to investigate the molecular mechanism of SNHG17 in CRC.ResultsUsing multiple cohorts, we confirmed that SNHG17 is aberrantly upregulated in CRC and correlated with poor survival. In vitro and in vivo functional assays indicated that SNHG17 facilitates CRC proliferation and metastasis. SNHG17 impedes PES1 degradation by inhibiting Trim23-mediated ubiquitination of PES1. SNHG17 upregulates FOSL2 by sponging miR-339-5p, and FOSL2 transcription activates SNHG17 expression, uncovering a SNHG17-miR-339-5p-FOSL2-SNHG17 positive feedback loop.ConclusionsWe identified SNHG17 as an oncogenic lncRNA in CRC and identified abnormal upregulation of SNHG17 as a prognostic risk factor for CRC. Our mechanistic investigations demonstrated, for the first time, that SNHG17 promotes tumor growth and metastasis through two different regulatory mechanisms, SNHG17-Trim23-PES1 axis and SNHG17-miR-339-5p-FOSL2-SNHG17 positive feedback loop, which may be exploited for CRC therapy.

Effect of NLK on the proliferation and invasion of laryngeal carcinoma cells by regulating CDCP1.

The morbidity and mortality of laryngeal cancer are increasing rapidly, seriously threatening human health. There are several causes of laryngeal cancer, but the exact molecular mechanism is unclear. Finding the molecular targets of laryngeal cancer has become an emerging hot spot. Nemo-like kinase (NLK) is abnormally expressed in tumors, but its role in laryngeal cancer has not been reported. Real Time PCR and Western blot were used to detect NLK mRNA and protein expression in cancer tissues and adjacent tissues of laryngeal cancer patients. The laryngeal carcinoma cell line Hep-2 cells were cultured in vitro and randomly divided into three groups: control group, NC group, and NLK siRNA group followed by the analysis of cell proliferation by MTT assay, Caspase3 activity, and cell invasion by the transwell chamber. MMP-9 and CDCP1 expression was measured by Western blot. NLK mRNA and protein expression was significantly increased in laryngeal carcinoma tissues compared with those in adjacent tissues (p<0.05). NLK siRNA transfection into Hep-2 cells significantly down-regulated NLK expression, inhibited Hep-2 cell proliferation and invasion, increased Caspase-3 activity with statistical differences compared to control group (p<0.05). Down-regulation of NLK expression in Hep-2 cells inhibited MMP-9 expression and decreased CDCP1 expression. NLK is expressed in tumor tissues of patients with laryngeal cancer. The down-regulation of NLK expression may play a role in the proliferation, apoptosis, and invasion of laryngeal carcinoma cells and it is possible by regulating MMP-9 and CDCP1 expression.

Use of aspirin in the prevention of colorectal cancer through TIGIT-CD155 pathway.

Colorectal cancer (CRC) is one of the most widespread malignant cancers, with a high incidence and mortality all over the world. Aspirin (ASA) otherwise known as acetylsalicylic acid, is a non‐steroidal anti‐inflammatory drug that has shown promising results in the prevention of chronic diseases, including several cancers. In previous studies, aspirin has been shown to reduce the incidence of CRC. Immune checkpoint blockade of T cell Ig and ITIM domain receptor (TIGIT) alone or combined with other immune checkpoint blockades moleculars has gained impressive results in the treatment of the melanoma and glioblastoma. Here, we found that TIGIT and Poliovirus receptor (PVR, CD155) are expressed in tumour cells; the TIGIT and CD155 protein expression in cancer tissue has been found to be significantly higher than that in the precancerous tissue. T cell Ig and ITIM domain receptor and CD226 were expressed in the lymphocytes near the tumour tissue and the adjacent tissues. Aspirin has been found to inhibit cancer cell viability and promote CRC cell apoptosis.Similarly, aspirin has also been found to increase pro‐apoptotic protein Bax's expression. We found that the expression of TIGIT decreased with an increase in the concentration of aspirin and that the suppression of TIGIT can affect the effect of aspirin on cell proliferation. In this paper, we found that aspirin attenuates cancer cell proliferation and induces CRC cells apoptosis by down‐regulating the expression of TIGIT, which provides new evidence for the application of aspirin in cancer treatment.

Extensive screening of microRNA populations identifies hsa-miR-375 and hsa-miR-133a-3p as selective markers for human rectal and colon cancer.

MicroRNAs (miRNAs) are ∼22-nt molecules exerting control of protein expression in cancer tissues. The current study determined the full spectrum of miRNA dysregulation in freshly isolated human colon or rectal cancer biopsies as well as in controls of healthy adjacent tissue (total of n = 100) using an Illumina sequencing technology. In this work, we aimed to identify miRNAs that may serve as future marker to discern between these two subtypes. DESeq2 analysis revealed 53 significantly dysregulated miRNAs in colon cancer, 67 miRNAs in rectal cancer, and 97 miRNAs in both at a Padj value < 0.05 and ≥ 10 read counts. 65% of miRNAs were upregulated in colon as well as rectal cancer. Highest significant dysregulation (Padj < 0.00001) was detected for hsa-miR-21-5p, -215-5p and -378a in both colon and rectal cancer. Among the group of miRNAs with Padj < 0.05 and more than 2-fold expression differences, hsa-miR-375 was detected in rectal cancer only, and hsa-miR-133a-3p only in colon cancer. Receiver operating characteristic (ROC) analysis confirmed highly distinct sensitivities for hsa-miR-375 to detect rectal cancer (area under the curve (AUC): 0.9), while hsa-miR-133a-3p (AUC: 0.89) had the highest sensitivity for detecting colon cancer. We conclude that hsa-miR-375 and hsa-miR-133a-3p may serve as new markers of rectal or colon cancer and should be further investigated to search for different etiologies of colorectal cancer.

Membranous or Cytoplasmic HER2 Expression in Colorectal Carcinoma: Evaluation of Prognostic Value Using Both IHC & BDISH

ABSTRACTBackground: Human epidermal growth factor recptor-2 (HER2) was identified as a driver gene in several types of cancers with both prognostic and predictive value. However, the molecular association of HER2 gene mutation with HER2 gene amplification and/or protein expression in cancer tissues has not been clearly defined. Moreover, there is little information available on HER2 status role in tumor progression and metastasis in colorectal carcinoma (CRC) compared to other solid tumors. The aim of this study was to evaluate both HER2 amplification and protein expression profiles using immunohistochemistry (IHC) and bright-field dual in situ hybridization (BDISH) techniques, respectively. Patients and methods: Tissue microarray (TMA) was constructed to accommodate a total of 243 CRC formalin-fixed paraffin embedded (FFPE) samples of consent patients and stained by IHC and BDISH methods. The expression patterns of HER2 protein status were evaluated and correlated to HER2 gene amplification status and then assessed for its prognostic value. Results: The expression profile of 58% samples showed cytoplasmic expression patterns of different categories. Interestingly, only 1% showed strong (+3) membranous expression pattern of HER2 with perfect match with their corresponding gene amplification status (>2). However, the cytoplasmic HER2 protein status did not show significant correlation with most clinicopathological features and survival outcomes except with age (p = 0.04) and tumor size (p = 0.03). Conclusion: We demonstrated that the membranous HER2 gene/protein status is infrequent, while the main fraction of HER2 overexpression was cytoplasmic and lacking prognostic value. This cytoplasmic HER2 overexpression was induced through a gene-amplification independent pathway, making the HER2 gene status evaluation approach in those cases not worthy. Further investigations about the molecular pathways of the cytoplasmic HER2 protein in CRC and its associations with survival outcomes are required to allow either a breakthrough in CRC management; or to confirm the hypothesis of a marginal role in CRC onset and progression.

Expression of fascin_1 protein in cancer tissues of patients with nonsmall cell lung cancer and its relevance to patients' clinicopathologic features and prognosis.

The aim of this study is to explore the expression of Fascin_1 protein in cancer tissues of patients with nonsmall cell lung cancer (NSCLC) and its relevance to patients' clinicopathologic features and prognosis. Retrospective analysis was performed on 81 patients with NSCLC who were admitted to our hospital and received surgical treatment from February 2012 to March 2016. To analyze the correlation of Fascin_1 protein expression in cancer tissues with the clinicopathologic features and prognosis of the patients, the immuno-histochemical method was applied to examine Fascin_1 protein expression in cancer and normal paracarcinoma tissues of the included 81 NSCLC patients. Fascin_1 protein was diffused in cancer tissues, and cancer cells surrounding the carcinoma lesions were strongly stained. The protein was mainly located on the cytoplast or membrane of positive cells and manifested as a focal or diffuse brown granular distribution. The negative, weakly positive, and strongly positive expression rates in cancer tissues were 23.5% (19/81), 35.8% (29/81), and 40.7% (33/81), respectively. The negative, weakly positive, and strongly positive expression rates in paracarcinoma tissues were 71.6% (58/81), 12.3% (10/81), and 16% (13/81), respectively. The difference was statistically significant (P < 0.05), and the positive expression rate of Fascin_1 in cancer tissues was significantly higher than that of in paracarcinoma tissues (P < 0.05). The positive expression of Fascin_1 in cancer tissues was significantly correlated with the neoplasm stage (P < 0.05) and mediastinal lymph node metastases (P < 0.05); however, it was not relevant to the patients' age, gender, tumor size, pathological type, tumor grading, or pleural effusion (P > 0.05). The median survival times for patients with positive Fascin_1 and negative Fascin_1 expression in cancer tissues were 26 and 44 months, respectively, indicating that the median survival time of the Fascin_1-positive group was significantly lower than that of in the Fascin_1-negative group (heart rate = 2.74, 95% confidence interval: 1.32-6.87, P < 0.05). Fascin_1 protein expression in NSCLC tissues and normal paracarcinoma issues were different. Fascin_1 expression in cancer tissues was related to the poor prognosis of NSCLC patients.

Cancer-testis antigen GAGE-1 expression and serum immunoreactivity in hepatocellular carcinoma.

To explore the use of cancer-testis antigen G antigen 1 (GAGE-1) in the diagnosis and potential therapeutic targeting of hepatocellular carcinoma (HCC), we measured the expression of GAGE-1 protein levels in HCC tissues and its serum immunoreactivity in HCC patients. We detected the expression of GAGE-1 protein in HCC by immunohistochemistry (IHC). We then analyzed the clinical significance of GAGE-1 expression in HCC with respect to clinicopathological parameters. We observed positive anti-GAGE-1 antibody reactivity in HCC patient serum, liver cirrhosis patients (LC), hepatitis B patients (HB), and normal human individuals (NHS) by enzyme-linked immunosorbent assay. The IHC results showed that the positive rates of GAGE-1 protein expression in cancer tissues and adjacent tissues were 43.3% (26/60) and 5% (3/60), respectively. The expression level of GAGE-1 protein in HCC tissues was significantly higher than that in tumor-adjacent tissues (P < 0.05). Positive GAGE-1 protein expression was not correlated with clinicopathological parameters (P > 0.05). Positive serum anti-GAGE-1 antibody reactivity in HCC patients, LC, HB, and NHS was 23.33% (14/59), 13.1% (8/61), 3.3% (2/60), and 3.4% (2/59), respectively. The frequency of anti-GAGE-1 antibody-positive sera in HCC patients and LC was significantly different than that in HB and NHS (P < 0.01), but no significant differences were found between HCC patients and LC (P = 0.485) or between HB and NHS (P = 0.410). Positive anti-GAGE-1 antibody reactivity was not correlated with clinicopathological parameters (P > 0.05). These data illustrate that the GAGE-1 protein exhibits moderate cancer-restricted pattern of expression and immunogenicity, laying the foundation for the application of GAGE-1 in immunotherapy and for the diagnosis of HCC.

Down-regulation of metabolic proteins in hepatocellular carcinoma with portal vein thrombosis

BackgroundHepatocellular carcinoma is an aggressive malignancy with poor prognosis and easy to recur even the tumor is totally removed by surgery. Portal vascular invasion is one of the major factors contributing to tumor recurrence and poor prognosis. However, why hepatocellular carcinoma is easy to grow into vessels is unclear.MethodsSurgical specimens from seven hepatocellular carcinoma patients with portal vein thrombosis and seven patients without vascular invasion were utilized to analyze protein expression by proteomic technique. The proteins in the tumors were separated by 2-dimensional electrophoresis. Protein patterns in the gels were recorded as digitalized images. The differences of expression in hepatocellular carcinoma with or without portal vein thrombosis were identified by mass spectrometry.ResultsClinically, the tumors with portal vein thrombosis were larger than those without portal vein thrombosis. The median survival time for the patients with portal vein thrombosis was much shorter [4 (ranged 2.5–47) vs. 53 (ranged 33–85) months, p = 0.002]. By analyzing the protein expression in cancer tissues with or without portal vein thrombosis, the differences of protein expression were mainly metabolic enzymes. Carbonic anhydrase I, betaine–homocysteine S-methyltransferase 1, fumarate hydratase, isovaleryl-CoA dehydrogenase, short-chain specific acyl-CoA dehydrogenase and arginase-1 were all down-regulated in the tumors with portal vein thrombosis.ConclusionMetabolic enzymes and cytosol carbonic anhydrases were downregulated in hepatocellular carcinoma with portal vein thrombus. The deficiency of metabolic enzymes and cytosol carbonic anhydrases may alter cellular metabolisms and acid–base balance in hepatocellular carcinoma, which may facilitate to invade portal vein.

Fibroblast activating protein-α expression in squamous cell carcinoma of the esophagus in primary and irradiated tumors: the use of archival FFPE material for molecular techniques.

There are numerous reports suggesting that fibroblast activating protein-α (FAP-α) plays an important role in invasion of various tumor types. We studied the expression pattern of FAP-α in esophageal squamous cell carcinoma (ESCC) patients who had not been treated primarily and those who had received neoadjuvant radiochemotherapy. Our goal was to establish whether readily available tissue specimens fixed in formalin and stored in paraffin blocks for years might still be a source of FAP-α RNA for PCR analysis. The study included 20 patients divided into two groups, 10 patients in each group. We evaluated the expression of FAP- α by PCR techniques in fresh frozen and in paraffin-embedded tissues, and compared it to the expression in non-cancer tissues. To detect the protein expression level of FAP-α in paraffin-embedded tissues we used chromogenic immunohistochemical (IHC) staining. Data were analyzed by t-test or the nonparametric Wilcoxon matched pair test using Statistica 12.5 software. We observed an increased level of the FAP-α gene and protein expression in cancer tissues when compared with their corresponding normal tissues. However, statistically significant differences were found only in the group of patients untreated before surgery. RNA extracted from paraffin-embedded tissue sections had very low quality, especially in the context of degradation. FAP-α remains a highly altered participant of a complex microenvironment in esophageal squamous cell carcinoma, and its role in cell signaling requires further study. In this paper, we conclude that the use of a regular RT-PCR method for diagnostic purposes, which we have presented in an earlier paper, can be as good as qRT-PCR. Also, immunohistochemistry proved to be very useful and the only reliable method that can be used on longterm stored formalin-fixed, paraffin-embedded tissues.

Oxidative stress balance is dysregulated and represents an additional target for treating cholangiocarcinoma

ABSTRACTBackground: Pancreatico-biliary malignancies exhibit similar characteristics, including obesity-related features and poor prognosis, and require new treatment strategies. Oxidative stress is known to induce DNA damage and carcinogenesis, and its reduction is viewed as being favorable. However, it also has anti-infection and anti-cancer functions that need to be maintained. To reveal the effect of oxidative stress on cancer progression, we evaluated oxidative stress and anti-oxidative balance in pancreatic cancer (PC) and cholangiocarcinoma (CC) patients, as well as the effect of add-on antioxidant treatment to chemotherapy in a mouse cholangiocarcinoma model.Methods: We recruited 84 CC and 80 PC patients who were admitted to our hospital. Serum levels of reactive oxygen metabolites (ROM) and the anti-oxidative OXY-adsorbent test were determined and the balance of these tests was defined as an oxidative index. A diabetic mouse-based cholangiocarcinoma model was utilized to evaluate the effects of add-on antioxidant therapy on cholangiocarcinoma chemotherapy.Results: Serum ROM was higher and anti-oxidant OXY was lower in CC patients with poor outcomes. These parameters were not significantly different in PC patients. In mice, vitamin E administration induced antioxidant hemeoxygenase (HO)-1 protein expression in cancer tissue, while the number of stem-like cells increased. l-carnitine administration improved intestinal microbiome and biliary acid balance, upregulated the hepatic mitochondrial membrane uptake related gene Cpt1 in non-cancerous tissue, and did not alter stem-like cell numbers.Conclusion: Oxidative stress balance was dysregulated in cholangiocarcinoma with poor outcome. The mitochondrial function-supporting agent l-carnitine is a good candidate to control oxidative stress conditions.

Abstract P2-07-06: Periostin is associated with breast tumor progression and epithelial periostin expression is correlated with poor survival in patients with invasive breast carcinoma

Abstract Background: Invasion and metastasis are the direct causes of mortality in patients with breast cancer and require reciprocal interactions between cancer cells and the extracellular matrix (ECM). Periostin, a fasciclin-containing adhesive ECM glycoprotein, is frequently overexpressed in various types of human cancer, and its overexpression in cancer-associated stroma and/or cancer cells is usually associated with poor clinical outcomes. However, the expression of periostin in the successive steps of breast tumorigenesis and its association with outcome variables have not been well established in breast carcinoma. The purposes of the present study were to assess the role of periostin alteration in breast tumorigenesis and evaluate the putative prognostic value of periostin as a function of its compartmentalization. Materials and Methods: Immunohistochemical staining with anti-periostin antibody was performed in a total of 300 patients (26 patients with normal breasts, 76 patients with ductal carcinoma in situ [DCIS], and 198 patients with invasive breast carcinoma [IBC]) using tissue microarray. Periostin immunoreactivity was assessed in both epithelial tissue and the surrounding stromal compartment. In addition, the mRNA and protein expression of periostin was analyzed in 10 paired normal/invasive cancer frozen specimens by quantitative real time-polymerase chain reaction and western blot analysis, respectively. Results: Periostin mRNA and protein expression in cancer tissues was increased compared with that in adjacent normal tissues. Both epithelial and stromal periostin staining scores were significantly increased with disease progression in a stepwise manner to DCIS and IBC compared with those in normal breast tissues (P = 0.000 and 0.000, respectively). High epithelial and stromal periostin expression was observed in 109 of 189 (57.7%) and 158 of 189 (83.6%) cases of IBC, respectively. High epithelial periostin expression was more frequently observed in the distant metastatic relapse-positive group than in the distant metastatic relapse-negative group (41 [80.4%] of 51 cases versus 68 [49.3%] of 138 cases [P = 0.000]). Furthermore, high epithelial periostin expression was associated with reduced disease-free and overall survival on univariate and multivariate analyses. Conclusion: Periostin might play an important role in the progression of breast tumor, and epithelial periostin expression may serve as a new parameter for prognostic prediction in patients with IBC. Further study of periostin expression and its potential as a target of therapy for IBC appears warranted. Citation Format: Lee JS, Kim G-E, Park MH, Yoon JH. Periostin is associated with breast tumor progression and epithelial periostin expression is correlated with poor survival in patients with invasive breast carcinoma. [abstract]. In: Proceedings of the Thirty-Eighth Annual CTRC-AACR San Antonio Breast Cancer Symposium: 2015 Dec 8-12; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2016;76(4 Suppl):Abstract nr P2-07-06.

Effect of Liuweidihuang pill and Jinkuishenqi pill on inhibition of spontaneous breast carcinoma growth in mice

ObjectiveTo investigate the preventing and treating action of Liuweidihuang pill (LP) and Jinkuishenqi pill (JP) on spontaneous breast carcinoma in mice. MethodsA model of spontaneous breast carcinoma was derived from 11.5-month-old female Kunming breeding mice following the delivery of several litters. The mice were randomly divided into five groups: model control group (C), Liuweidihuang pill high-dose group (LH; 4.6 g • kg−1 • d−1), Liuweidihuang pill low-dose group (LL; 2.3 g • kg−1 • d−1), Jinkuishenqi pill high-dose group (JH; 4.6 g • kg−1 • d−1) and Jinkuishenqi pill low-dose group (JL; 2.3 g • kg−1 • d−1). Cancer tissue volume was measured by water immersion. Histopathology was analyzed by hematoxylin and eosin staining. Vascular endothelial growth factor (VEGF), extracellular signal-regulated kinase (ERK) and cyclin D1 protein expression in cancer tissue was assayed by western blotting. ResultsCompared with the control group, cancer tissue volume and weight were lower in the LP and JP groups, and survival time was longer. The expression of VEGF, ERK and Cyclin D1 were inhibited in the LP and JP groups (P < 0.05), and cell differentiation was increased. Tumor weights and volumes and VEGF, ERK and Cyclin D1 expression in LL or LH were significantly lower than in JL and JH (P < 0.01). ConclusionBoth LP and JP could restrain cancer growth and promote cancer cell differentiation; moreover, LP was more effective than JP The likely mechanism of action was via inhibition of VEGF, ERK and cyclin D1.

COX-2, MMP-7 expression in oral lichen planus and oral squamous cell carcinoma

ObjectiveTo observe cyclooxygenase (COX)-2 expression in normal oral mucosa (NOM), oral lichen planus (OLP) and oral squamous cell carcinoma (OSCC) and explore its significance in the incidence of oral cancer. MethodsThe immunohistochemical method and RT-PCR method were applied to detect the expression of COX-2 and MMP-7 in 10 cases with NOM, 33 cases of with OLP and 38 cases with OSCC. ResultsThe expression of COX-2 mRNA in OSCC tissues (68.4%, 26/38) was significantly higher than in the OLP (24.2%, 8/33) and NOM (0.0%, 0/10) (P<0.01). The expression of MMP-7 mRNA in OSCC tissues (65.8%, 25/38) was significantly higher than in the OLP (30.3%, 10/33) and NOM (0.0%, 0/10) (P<0.01). The expression of MMP-7 in OLP was significantly higher than in the NOM (P<0.05). There was no significant expression of COX-2 protein in NOM, and the positive rate was 42.4% (14/33) and 89.5% (34/38) in OLP and OSCC group, respectively. The COX-2 expression in cancer tissues was significantly higher than in NOM and OLP (P<0.05). The MMP-7 protein expression in cancer tissues (84.2%, 32/38) was significantly higher than in NOM (10.0%, 1/10) and in OLP (42.4%, 14/33), and the positive rate in OLP was significantly higher than in NOM (P< 0.01). The COX-2 expression was associated with clinical stage (P<0.05), the MMP-7 expression was associated with clinical stage and lymph node metastasis (P<0.05). The expressions of COX-2 and MMP-7 mRNA were positively correlated with OSCC. ConclusionsThe abnormal expressions of COX-2 and MMP-7 are closely related to the biological behavior of OSCC, the MMP-7 may be induced by COX-2, and further lead to the invasion and metastasis of OSCC.

P73 G4C14-to-A4T14 polymorphisms are positively correlated with triple-negative breast cancer in southwestern China

p73 gene shares structural and functional similarities to p53 and plays an important role in modulating cell cycle arrest and apoptosis. A common non-coding polymorphism of p73 G4C14-to-A4T14 (rs2273953 and rs1801173) at exon 2 may affect gene expression, thus, it may lead to functional significance. The correlation of this polymorphism with clinicopathologic variables of patients with breast cancer has not been investigated. In this study, single-nucleotide polymorphisms (SNPs) of p73 G4C14-to-A4T14 were genotyped by Sequenom MassArray-iPLEX GOLD System in 170 patients with breast cancer. Data were analyzed via t test, chi-square test, and logistic regression analysis. There was no significant correlation between p73 G4C14-to-A4T14 polymorphisms and the patient characteristics, such as clinical TNM stage, menopausal status, axillary lymph node metastasis, pathological type, estrogen receptor (ER), progesterone receptor (PR), human epidermal growth factor receptor 2 (HER2). No significant relationship was observed between the p73 G4C14-to-A4T14 polymorphism and p73 protein expression in cancer tissues. The frequency of GC/GC genotype in patients with triple-negative breast cancer (TNBC) was 78.9 %, that of patients with others was 57.6 %, and the difference had statistical significance (χ ( 2 ) = 5.74, P = 0.02). p73 G4C14-to-A4T14 polymorphisms were negatively correlated with chemosensitivity for anthracycline-based chemotherapy in breast cancer (P > 0.05). p73 G4C14-to-A4T14 polymorphisms are positively correlated with TNBC, and p73 gene may play a critical role in a novel therapeutic strategy to TNBC. Additional larger studies are required to test these hypotheses.

PEBP4 gene expression and its significance in invasion and metastasis of non-small cell lung cancer

The goal of this study was to investigate the function of phosphatidylethanolamine-binding protein 4 (PEBP4) in invasion and metastasis of non-small cell lung cancer (NSCLC). PEBP4 mRNA and protein expression in 56 cases of NSCLC tissues were detected using RT-PCR and Western blot, and the relationship between PEBP4 expression and invasion and metastasis of NSCLC was analyzed. The change in the invasive ability of human NSCLC cell line HCC827 was observed after knocking down PEBP4 expression using RNA interference. PEBP4 mRNA and protein expression in cancer tissues of patients with lymph node metastasis were significantly higher than those in patients without lymph node metastasis (p < 0.05). PEBP4 expression significantly decreased in HCC827 cells after transfection with PEBP4 siRNA (p < 0.01), and the number of HCC827 cells that migrated through Transwell chambers was significantly lower than that of non-transfected control and transfected control cells (p < 0.01). PEBP4 over-expression may promote the invasion and metastasis of NSCLC.