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Abstract Background In the evaluation of PD-L1 expression to select patients for anti-PD-1/PD-L1 treatment, uniform guidelines that account for different immunohistochemistry assays, different cell types and different cutoff values across tumor types are lacking. Data on how different scoring methods compare in breast cancer are scant. Methods Using FDA-approved 22C3 diagnostic immunohistochemistry assay, we retrospectively evaluated PD-L1 expression in 496 primary invasive breast tumors that were not exposed to anti-PD-1/PD-L1 treatment and compared three scoring methods (TC: invasive tumor cells; IC: tumor-infiltrating immune cells; TCIC: a combination of tumor cells and immune cells) in expression frequency and association with clinicopathologic factors. Results In the entire cohort, positive PD-L1 expression was observed in 20% of patients by TCIC, 16% by IC, and 10% by TC, with a concordance of 87% between the three methods. In the triple-negative breast cancer patients, positive PD-L1 expression was observed in 35% by TCIC, 31% by IC, and 16% by TC, with a concordance of 76%. Associations between PD-L1 and clinicopathologic factors were investigated according to receptor groups and whether the patients had received neoadjuvant chemotherapy. The three scoring methods showed differences in their associations with clinicopathologic factors in all subgroups studied. Positive PD-L1 expression by IC was significantly associated with worse overall survival in patients with neoadjuvant chemotherapy and showed a trend for worse overall survival and distant metastasis-free survival in triple-negative patients with neoadjuvant chemotherapy. Positive PD-L1 expression by TCIC and TC also showed trends for worse survival in different subgroups. Conclusions Our findings indicate that the three scoring methods with a 1% cutoff are different in their sensitivity for PD-L1 expression and their associations with clinicopathologic factors. Scoring by TCIC is the most sensitive way to identify PD-L1-positive breast cancer by immunohistochemistry. As a prognostic marker, our study suggests that PD-L1 is associated with worse clinical outcome, most often shown by the IC score; however, the other scores may also have clinical implications in some subgroups. Large clinical trials are needed to test the similarities and differences of these scoring methods for their predictive values in anti-PD-1/PD-L1 therapy.

B.cell lymphoma 6 (BCL6), a proto.oncogene, is an evolutionarily conserved zinc finger protein that functions as a transcriptional repressor. BCL6 is the master regulator of B.lymphocyte development, and it has been reported that BCL6 may serve an important role in breast cancer progression. The aim of the present study was to investigate the expression of BCL6, zinc finger E.box.binding homeobox (ZEB)1 and ZEB2 and their associations in breast cancer. The mRNA and protein expression of BCL6, ZEB1 and ZEB2 was assessed using in situ hybridization and immunohistochemistry, respectively, in 228 patients with breast cancer and 80 patients with benign breast disease. In addition, the association between BCL6, ZEB1 and ZEB2 expression and the clinicopathological characteristics and survival of patients with breast cancer were analyzed. The mRNA and protein expression of BCL6, ZEB1 and ZEB2 were significantly higher in breast cancer tissues compared with benign breast disease tissues (P<0.05). The expression of BCL6, ZEB1 and ZEB2 were significantly positively correlated with tumor size, lymph node metastasis and a higher tumor stage (P<0.05). Furthermore, patients with BCL6, ZEB1 and ZEB2 protein.positive primary tumors had significantly lower overall survival (P=0.001, 0.002 and 0.001, respectively) and relapse.free survival (P=0.002, 0.001 and 0.003, respectively) rates. The mRNA expressions of ZEB1 (rs=0.326, P<0.001) and ZEB2 (rs=0.382, P<0.001) were significantly positively correlated with BCL6 mRNA expression, and the protein expressions of ZEB1 ((rs=0.449, P<0.001) and ZEB2 (rs=0.669, P<0.001) were significantly positively correlated with BCL6 protein expression. These results suggest that BCL6, ZEB1 and ZEB2 are potential biomarkers for the invasion, metastasis and prognosis of breast cancer, and that BCL6 may be a regulator of the ZEB family. [ABSTRACT FROM AUTHOR]

Ethnopharmacological relevance Vasculogenic mimicry (VM) has been suggested to be present in various malignant tumors and associated with tumor nutrition supply and metastasis, leading to poor prognosis of patients. Notch1 has been demonstrated to contribute to VM formation in hepathocellular carcinoma (HCC). Celastrus orbiculatus extract (COE), a mixture of 11 terpenoids isolated from the Chinese Herb Celastrus orbiculatus Vine, has been suggested to be effective in cancer treatment. Aim of the study In the current study, experiments were carried out to examine the effect of COE on VM formation and HCC tumor growth both in vitro and in vivo . Materials and methods CCK-8 assay and Nikon live-work station were used to observe the viability of malignant cells treated with COE. Cell invasion was examined using Transwell. Matrigel was used to establish a 3-D culture condition for VM formation. Changes of mRNA and protein expression were examined by RT-PCR and Western Blot respectively. Tumor growth in vivo was monitored using in vivo fluorescence imaging device. PAS-CD34 dual staining and electron microscopy were used to observe VM formation. Immunohistochemical staining (IHC) was used to examine Notch1 and Hes1 expression in tumor tissues. Results Results showed that COE can inhibit HCC cells proliferation and invasion in a concentration-dependent manner. VM formation induced by TGF-β1 was blocked by COE. In mouse xenograft model, COE inhibited tumor growth and VM formation. Both in vitro and in vivo studies showed that COE can downregulate expression of Notch1 and Hes1. Conclusion The current results indicate that COE can inhibit VM formation and HCC tumor growth by downregulating Notch1 signaling. This study demonstrates that COE is superior to other anti-angiogenesis agents and can be considered as a promising candidate in HCC treatment. [ABSTRACT FROM AUTHOR]

Guanine nucleotide-binding protein subunit alpha-s (Gnas) is a small subunit of the G protein-couple signaling pathway, which is involved in the formation of coat color. The expression level and distribution of Gnas were detected by quantitative real-timepolymerase chain reaction (qPCR), western blot, and immunohistochemistry to investigate the underlying mechanisms of coat color in white and black skin tissues of mice. qPCR and western blot results suggested that Gnas was expressed at significantly higher levels in black mice compared with that of white mice, and transcripts and protein possessed the same expression in both colors. Immunohistochemistry demonstrated Gnas staining in the root sheath and dermal papilla in hair follicle of mice skins. The results indicated that the Gnas gene was expressed in both white and black skin tissues, and the expression level of Gnas in the two types of color was different. Therefore, Gnas may be involved in the coat color formation in mice. [ABSTRACT FROM AUTHOR]

Wild-type p53-induced phosphatase 1 (Wip1), also termed PPM1D, is a member of the protein phosphatase 2C family, which is characterized by distinctive oncogenic properties. Overexpression of Wip1 is observed in certain types of human tumors that are associated with significantly poor prognosis. The present study aimed to detect the expression of Wip1 in non-small cell lung cancer (NSCLC) and to analyze its prognostic value in such patients. The protein expression level of Wip1 was compared between NSCLC and normal lung tissue specimens using by immunohistochemistry, and it was found that Wip1 was highly expressed in NSCLCs but was absent or weakly expressed in normal lung tissues. Detailed clinical and demographic information of patients were retrospectively collected pre- and postoperatively, and Kaplan-Meier survival and Cox's regression analyses were performed to evaluate the prognosis of patients. Survival analysis revealed that the overall survival rate for patients in the Wip1-positive expression group was significantly lower than that of the Wip1-negative group, and Cox multivariate analysis indicated that positive Wip1 expression, pN classification and pathological stage were significant prognostic predictors. The results of the current study suggest that Wip1 may be associated with pathological diagnosis and prognostic evaluation of NSCLC. [ABSTRACT FROM AUTHOR]

Metabotropic glutamate receptor 4 (mGluR4) has been associated with the pathogenesis of osteosarcoma. The aim of this study was to investigate mGluR4 expression and its clinical significance in osteosarcoma patients. mGluR4 expression was investigated using immunohistochemistry (IHC) in 58 osteosarcomas and 32 giant-cell tumors of bone. The correlations between mGluR4 expression and clinicopathological characteristics were analyzed with the Chi-squared test and survival curves were generated using the Kaplan-Meier method. The IHC results demonstrated that 20.69% (12/58) of the osteosarcomas and 43.75% (14/32) of the giant-cell tumors were mGluR4-positive. The statistical analysis revealed that mGluR4 expression was correlated with gender, age, Enneking stage and tumor volume in osteosarcomas (P<0.05). In the multivariate stepwise Cox regression analysis, Enneking stage was found t o b e statistically significantly associated with survival (P<0.05) and the survival analysis demonstrated that the survival probability was significantly higher in patients with higher mGluR4 expression compared with those with lower expression (P<0.05). Therefore, mGluR4 expression may be used to estimate the prognosis of osteosarcoma patients. [ABSTRACT FROM AUTHOR]

Background: This study aimed to investigate the expression of CD147 and MMP-11 in human colorectal cancer (CRC) and to evaluate their clinical significance.Methods: Real-time polymerase chain reaction was used to evaluate CD147 and MMP-11 mRNA level in 56 pairs of fresh CRC samples matched with adjacent normal mucosa. The protein expression of CD147 and MMP-11 in CRC specimens and corresponding normal colorectal mucosa were evaluated by immunohistochemistry on CRC tissue microarrays. Expression and co-localization of these two proteins in human colorectal cancer tissue were also evaluated by laser scanning confocal microscopy. Furthermore, their correlations with clinicopathological factors and overall survival after surgery were evaluated.Results: Both CD147 and MMP-11 were demonstrated to be over-expressed at mRNA level (P < 0.001, both) and protein level (P < 0.001, both) in CRC tissue than paired normal mucosa. Spearman rank test showed a positive correlation between these two proteins (P = 0.025). Immunofluorescence double staining confirmed the co-localization of CD147 and MMP-11 in paraffin-embedded tissues of CRC patients. Expression of CD147 and MMP-11 were both correlated with CRC lymph node metastasis (P = 0.021 and P = 0.031, respectively), distant metastasis (P < 0.001 and P = 0.013, respectively) and TNM stage (P = 0.006 and P = 0.049, respectively). Univariate survival analysis showed that both CD147 and MMP11 expression was significantly associated with shorter survival time (P = 0.001 and P = 0.009, respectively). Additionally, in multivariate analysis, both CD147 and MMP-11 were proved to be independent prognostic factors (P = 0.009, 0.028, respectively).Conclusions: These results indicated that both CD147 and MMP-11 may be involved in the progression of colorectal cancer, and they are potential prognostic factors and might become new therapeutic targets for CRC patients. [ABSTRACT FROM AUTHOR]

As the main active constituent of Andrographis paniculata that was applied in treatment of many diseases including inflammation in ancient China, andrographolide (ANDRO) was found to facilitate reduction of edema and analgesia in arthritis. This suggested that ANDRO may be promising anti-inflammatory agent to relieve destruction and degeneration of cartilage after inflammation. In this study, the effect of ANDRO on rabbit articular chondrocytes in vitro was investigated. Results showed that not more than 8f(M ANDRO did no harm to chondrocytes (P < 0.05). DNA content and glycosaminoglycan (GAG) /DNA were, respectively, improved in ANDRO groups comparing to the control (P < 0.05). ANDRO could promote expression of aggrecan, collagen II, and Sox9 genes while downregulating expression of collagen I gene (P < 0.05). Furthermore, hypertrophy that may result in chondrocyte ossification could not be detected in all groups (P > 0.05). The viability assay, hematoxylin-eosin, safranin O, and immunohistochemical staining also showed better performances in ANDRO groups. As to the doses, 3 µM ANDRO showed the best performance. The results indicate that ANDRO can accelerate proliferation of rabbit articular chondrocytes in vitro and meanwhile maintain the phenotype, which may provide valuable references for further exploration on arthritis. [ABSTRACT FROM AUTHOR]

Background We aimed to detect CD40 mutant expression and evaluate its clinical significance in gastric cancer. Methods CD40 mutant expression in 78 cases of gastric cancer tissues, 10 cases of normal gastric tissues, and 10 cases of gastric adenoma tissues by immunohistochemical test. Survival analyses were also performed. Results The positive CD40 mutant rate in gastric cancer was 55.1% (43/78). No positive CD40 mutant staining was observed in the normal gastric tissue or the gastric adenoma. CD40 mutants expression was significantly correlated with invasive depth, lymph metastasis, and TNM stage (P <0.05). Cases with negative CD40 mutant expression had a significantly longer median survival time than those with positive CD40 mutant expression (40 vs. 14 months, P <0.05). A lower death risk in negative CD40 mutant cases was observed comparing with positive CD40 mutant cases. Conclusions Positive CD40 mutant expression suggests a poorer prognosis of gastric cancer cases. [ABSTRACT FROM AUTHOR]

Aims: This study attempts to characterize the feature of immunologically competent cells (ICCs) and evaluate its clinical implication in patients with acute-on-chronic liver failure (ACLF) in relation to chronic hepatitis B virus (HBV) infection. Methods: Circulating ICCs were examined in ACLF patients ( n = 75), as well as in patients with hepatitis B (CHB, n = 31), CHB-related liver cirrhosis (LC, n = 36), and normal controls (NC, n = 30). Intrahepatic ICCs in some patients were further analyzed via immunohistochemical and flow cytometric assays. Results: Total lymphocytes, CD4+ T cells, CD8+ T cells, and NK cells in circulation were numerically lower in the ACLF and LC groups compared to the CHB and NC groups. Importantly, the number of these cells was significantly lower in non-surviving ACLF patients compared with surviving ACLF patients. In comparison to NC, ACLF patients displayed a significantly higher ratio of liver-infiltrating CD4+ T-cell frequency than its circulating counterpart, suggesting that the possiblility of the ICCs compartmentalization from the peripheral blood into the liver in ACLF. Immunohistochemical analysis showed that intrahepatic CD4+ cells, CD8+ cells, and CD56+ cells were significantly higher in the ACLF group compared with the other three groups, suggesting a stronger cellular immune response-mediated inflammation in ACLF group than other patient groups. Conclusions: The abnormal prevalence of circulating and intrahepatic ICCs possibly acts as an important factor that may drive the progression of HBV-related ACLF. [ABSTRACT FROM AUTHOR]