Min-Kyung Yeo

MicroRNA expression profiles in placenta with severe preeclampsia using a PNA-based microarray

INTRODUCTION Preeclampsia (PE) is a leading cause of maternal and neonatal mortality and morbidity worldwide. However, the pathophysiology of this disease is not yet fully understood. MiRNA plays an important role in post-transcriptional gene regulation. Recent studies have suggested that dysregulation of miRNAs in placental tissue is involved in the pathogenesis of PE. Therefore, we investigated miRNA profiles in PE placenta to understand the miRNA function in PE pathogenesis. METHODS MiRNA profiling was performed in 20 formalin-fixed and paraffin-embedded samples (10 placentas from severe PE and 10 from a control group). We used a hybridization-based microarray with a PNA-probe comprised of 158 miRNAs. RESULTS Thirteen miRNAs (miR-92b, miR-197, miR-342-3p, miR-296-5p, miR-26b, miR-25, miR-296-3p, miR-26a, miR-198, miR-202, miR-191, miR-95, and miR-204) were significantly overexpressed and two miRNAs (miR-21 and miR-223) were underexpressed in PE compared with the control group. Among 15 differentially expressed miRNAs, miR-26b, miR-296-5p, and miR-223 were found to be consistent with results from previous studies. We identified 893 genes that were predicted by at least three of four computational algorithms. Target genes participated in several signaling pathways, adherens junction, focal adhesion, and regulation of the actin cytoskeleton. CONCLUSIONS Several miRNAs are found to be dysregulated in placentas of PE patients and they seem to be closely associated with the early pathogenesis of PE. Further study is necessary to develop tools for early detection and management.

Pyrosequencing cut‐off value identifying BRAFV600E mutation in fine needle aspiration samples of thyroid nodules

Context  Recently, tremendous efforts have been made towards the development of sensitive techniques to detect the BRAFV600E mutation in fine needle aspiration biopsy (FNAB) samples. However, newly developed quantitative and semi‐quantitative methods, such as dual‐priming oligonucleotide (DPO)‐based multiplex polymerase chain reaction (PCR), have the potential to generate false‐positive (FP) results.

Hippo pathway effector YAP inhibition restores the sensitivity of EGFR-TKI in lung adenocarcinoma having primary or acquired EGFR-TKI resistance.

The efficacy of EGFR-tyrosine kinase inhibitors (TKIs) is significantly limited by various resistance mechanisms to those drugs. The resistance to EGFR-TKI is largely divided by two classes; acquired resistance after EGFR-TKI treatment, and primary resistance marked by cancer cells dependence on other oncogene, such as KRAS. YAP has emerged as critical oncogene in conferring drug resistance against targeted therapy. In this study, we evaluated the role of YAP in primary and acquired EGFR-TKI resistance using gefitinib-resistant A549 and PC9 cells and their parental cell lines. Our study revealed that EGFR-TKI resistance is associated with enhanced YAP activity. Notably, YAP activation was independent of the Hippo pathway. We confirmed that AXL is a downstream target of YAP that confers EGFR-TKI resistance. And our results showed that YAP can induce ERK activation in lung adenocarcinoma. The combination of YAP inhibition with EGFR-TKI overcomes primary and acquired EGFR-TKI resistance. We also found increased YAP expression in human lung cancer after acquiring EGFR-TKI resistance. Collectively, we suggest a novel EGFR-TKI resistance mechanism involving YAP activation and suggest targeting YAP and EGFR simultaneously may be a breakthrough treatment of primary and acquired EGFR-TKI resistant lung cancer.

Pattern of expression of cell cycle-related proteins in malignant transformation of sinonasal inverted papilloma.

Background It has been suggested that sinonasal inverted papilloma (IP) can progress to squamous cell carcinoma (SCC); however, mechanisms of malignant transformation are not fully understood. This study investigated alterations in the expression of cell cycle–related proteins in a multistep process of malignant transformation of IPs. Methods The expression of cell cycle–related proteins, including p53, p21, p16, and p63, was evaluated by immunohistochemistry in 21, 56, 7, and 18 cases of nasal polyps, IPs, IPs with dysplasia, and IPs with SCC, respectively. Furthermore, we determined the possible correlation between the expression of these proteins and clinicopathological variables in patients with IPs with SCC. Results Expression of p53 was found only in 8 of 18 IPs with SCC (44.4%). The frequency of p21 positivity was significantly higher in IPs with dysplasia (71.4%) and IPs with SCC (77.8%) compared with nasal polyps (0%) and IPs (12.5%). A p16 positivity was observed in 82.1% of IPs and 57.1% of IPs with dysplasia, whereas 83.3% of IPs with SCC showed an apparent loss of p16 protein expression. A p63 positivity was found in all specimens, but the percentage of positive cells was significantly increased in IPs with dysplasia and IPs with SCC compared with nasal polyps and IPs. There was no significant correlation between the expression of these proteins and clinicopathological variables, such as tumor stage, histological differentiation, and the proportion of malignant areas in patients with IPs with SCC. Conclusion Alteration of cell cycle–related proteins may contribute importantly to the malignant transformation from IP to SCC.

Hippo effector YAP directly regulates the expression of PD-L1 transcripts in EGFR-TKI-resistant lung adenocarcinoma

Developments of EGFR-TKI and immunotherapy targeting the PD1/PD-L1 pathway are considered most important medical breakthroughs in lung cancer treatment. Nowadays, 3rd generation EGFR TKI is widely used for T790M positive 1st and 2nd EGFR-TKI resistant lung cancer patients. Immunotherapy is powerful option for lung cancer patients without drug targets and chemotherapy resistant patients. It also has changed the concept of conventional anti-cancer therapy in the point of regulating tumor microenvironment. There are many studies linking these two important pathways. Recent studies demonstrated that PD-L1 expression is significantly correlated to the mutation status of EGFR, and activation of EGFR signaling can also induce the expression of PD-L1. However, the real linker between PD-L1 and EGFR signaling remains to be revealed. Our previous study revealed that the Hippo pathway effector YAP confers EGFR-TKI resistance in lung adenocarcinoma, and inhibition of YAP restores sensitivity to EGFR-TKIs. Thus, we examined whether PD-L1 is relevant, in terms of conferring EGFR-TKI resistance and whether YAP directly regulates the expression of PD-L1 in this context. First, we compared the expression levels of PD-L1 and YAP between EGFR-TKI-resistant PC9 cells and the parental PC9 adenocarcinoma cells. The expression levels of both YAP and PD-L1 were markedly higher in the EGFR-TKI-resistant cells compared to the parental cells, suggesting differential expression pattern between two cell types. YAP knockdown significantly decreased the expression of PD-L1 in the EGFR-TKI-resistant cells, while YAP overexpression increased the expression of PD-L1 in the parental PC9 cells. Then, our results revealed that YAP regulates the transcription of PD-L1, and the YAP/TEAD complex binds to the PD-L1 promoter. Surprisingly, knockdown of PD-L1 was sufficient to decrease cell proliferation and wound healing in the EGFR-TKI-resistant PC9 cells. These data suggest a PD1-independent oncogenic function of PD-L1. The Hippo effector YAP plays a crucial role in linking the PD-L1 and EGFR-TKI resistance by directly regulating the expression of PD-L1 in lung cancer. Targeting PD-L1 directly or via YAP could provide an effective therapeutic strategy for EGFR-TKI-resistant lung adenocarcinoma.

Pattern of expression of cyclooxygenase-2 in malignant transformation of sinonasal inverted papilloma

OBJECTIVE Cyclooxygenases (COXs) are enzymes that catalyze the conversion of arachidonic acid to prostaglandins. Many studies have suggested that COX-2, the inducible form of COX, is important in carcinogenesis. However, little is known about the pattern of expression of COX-2 in a multistep process of malignant transformation of sinonasal inverted papilloma (IP). In this study, we investigated COX-2 expression in IPs, IPs with dysplasia, IPs with squamous cell carcinoma (SCC), and primary SCCs of sinonasal tract. STUDY DESIGN A retrospective study was conducted. SETTING The setting was a tertiary care referral center. SUBJECTS AND METHODS The expression of COX-2 was evaluated by immunohistochemistry in 56, 7, 18, and 17 cases of IPs, IPs with dysplasia, IPs with SCC, and primary SCCs, respectively. Furthermore, we investigated the possible correlation between the expression of COX-2 and clinicopathologic variables in patients with IPs with SCC and primary SCC patients. RESULTS Positive immunoreactivity for COX-2 was observed in 3 (5.4%) of 56 IPs, 7 (38.9%) of 18 IPs with SCC, and 7 (41.2%) of 17 primary SCCs, whereas it was not observed in IPs with dysplasia. The percentage of tumors with COX-2-positive immunostaining was significantly higher in IPs with SCC and primary SCCs compared with benign IPs. There was no significant correlation between the expression of COX-2 and clinicopathologic variables, such as tumor stage, histologic differentiation, and the proportion of malignant areas in patients with IPs with SCC. CONCLUSION Cyclooxygenase-2 may play an important role in the process of malignant transformation from IP to SCC.

Clinicopathological roles of S100A8 and S100A9 in cutaneous squamous cell carcinoma in vivo and in vitro.

S100A8 and S100A9 are members of the S100 protein family and exist in neutrophils, monocytes, and macrophages. Recent studies have shown that S100A8 and S100A9 are associated with various neoplastic disorders; however, their roles in cutaneous squamous cell carcinoma (SCC) are not well defined. To investigate the expression and function of S100A8 and S100A9 in skin tumors, we examined the expression levels of S100A8 and S100A9 between premalignant and malignant skin tumors and investigated the functional roles of S100A8 and S100A9 in vitro and in vivo using recombinant adenovirus expressing S100A8 or S100A9. The immunopositive staining rates and intensities of S100A8 and S100A9 were higher in SCC than in premalignant skin tumors. When S100A8 and/or S100A9 were overexpressed in SCC12 cells using a recombinant adenovirus, cell growth and motility were increased. Similarly, when mouse skin was intradermally injected with SCC12 cells overexpressing S100A8 and/or S100A9, there were remarkable increases in tumor growth and volume. Both S100A8 and S100A9 are highly expressed in cutaneous SCC and play important roles in tumorigenesis. We suggest that S100A8 and S100A9 may be potential therapeutic targets for the prevention or treatment of SCC in skin.

The usefulness of a novel fully automated PCR-based Idylla test for detection of the BRAF V600E mutation in thyroid tissue: comparison with PNA-clamping PCR, real-time PCR and pyrosequencing

Introduction The BRAF V600E mutation is the most common genetic event in papillary thyroid carcinoma (PTC). The BRAF V600E mutational status has a significant diagnostic and prognostic role in PTC since it can be detected in 32%–87% of PTC by various molecular methods. Aim(s) A novel, fully automated real-time PCR-based Idylla test is assessed to detect the BRAF mutation in formalin-fixed paraffin-embedded (FFPE) thyroid samples. Methods 99 PTC and 11 nodular hyperplasia FFPE thyroid tissues are evaluated for the BRAF V600E mutation by the Idylla tests and compared with peptide nucleic acid-clamping PCR, real-time PCR and pyrosequencing. Results The sensitivity and specificity of the Idylla test to detect BRAF V600E are 98.8% and 100%, which is superior to real-time PCR and pyrosequencing. The concordance between Idylla and true positive is highest at 0.974. Conclusions This study validates that the Idylla test is a sensitive and specific method to detect BRAF V600E in FFPE thyroid tissues. A simple, quick and easy to handle Idylla test is a useful and reliable molecular technique to evaluate BRAF mutations.

Clinical usefulness of the free web-based image analysis application ImmunoRatio for assessment of Ki-67 labelling index in breast cancer

Aims Ki-67 is a prognostic marker in breast cancer; however, the use of the Ki-67 labelling index (LI) in clinical practice requires a consistent and easily accessible scoring method. The present study evaluated the use of the free internet-based image analysis program ImmunoRatio to score Ki-67 LI in breast cancer in comparison with manual counting. Methods Ki-67 immunohistochemical detection was performed in 577 breast cancer cases, and the Ki-67 LI was determined by ImmunoRatio and manual counting. Results The Ki-67 LI determined by ImmunoRatio correlated well with that obtained by manual counting. The concordance rate between ImmunoRatio and manual counting was excellent (κ coefficient of 0.881) at a Ki-67 LI cut-off value of 20%. Cases with high Ki-67 LI by ImmunoRatio were associated with poor overall survival, in particular in the hormone receptor positive group. Conclusions The web-based automated image analysis program ImmunoRatio is an attractive alternative to manual counting to determine the Ki-67 LI in breast cancer.

Association of PD-L1 expression and PD-L1 gene polymorphism with poor prognosis in lung adenocarcinoma and squamous cell carcinoma

Programmed cell death 1 receptor (PD-1)/programmed death-1 ligand-1 (PD-L1) interaction has been linked to tumor immune evasion. PD-L1 expression has been indicated in identifying non-small cell lung carcinoma (NSCLC) patients for treatment with anti-PD-1 or anti-PD-L1 therapy. The goal of this study was to evaluate the clinicopathologic values of PD-L1 expression and single-nucleotide polymorphisms (SNPs) in the PD-L1 gene in lung adenocarcinoma (ADC) and squamous cell carcinoma (SqCC). The 147 NSCLC tissues consisted of 84 samples of ADC and 63 samples of SqCC. All tissue microarray paraffin blocks were used for PD-L1 immunohistochemical assays with 22C3, SP263, and SP142 clones. Three SNPs in the PD-L1 gene, rs4143815, rs822336, and rs822337, were genotyped using SNP pyrosequencing. The PD-L1 expression was significantly higher in SqCC than in ADC. Among ADCs, PD-L1 expression was significantly higher in papillary and solid types than in lepidic and acinar types. Statistical associations of the PD-L1 expression with a shorter disease-free survival outcome and lymph node metastasis in the ADCs were found but no associations in SqCCs. Among the three SNPs, the rs4143815 genotype CC was statistically associated with positive 22C3 PD-L1 labeling in NSCLCs. The rs4143815 genotype GG instead showed a trend of shorter survival outcomes but did not reach statistical significance in the ADCs. Our results showed a significantly higher prevalence of positive PD-L1 expression in lung SqCC than in ADC. However, the PD-L1 expression and rs4143815 genotype GG might be useful for the prediction of poor prognosis in lung ADC cases.