Sun A. Kim

Human plasma carboxylesterase 1, a novel serologic biomarker candidate for hepatocellular carcinoma

To identify and characterize a serologic glycoprotein biomarker for hepatocellular carcinoma (HCC), multi‐lectin affinity chromatography was used to isolate intracellular N‐linked glycoprotein fractions from five paired non‐tumor and tumor tissues. From the series of 2‐D DIGE targeted differentially expressed N‐linked glycoproteins, we identified human liver carboxylesterase 1 (hCE1), which was remarkably down‐regulated in tumor tissues, a finding confirmed by Western blot, a quantitative real‐time RT‐PCR, and immunohistochemical staining of non‐tumor and tumor tissues from total 58 HCC patients. To investigate whether hCE1 is also present in human plasma, we employed a magnetic bead‐based immunoprecipitation followed by nano‐LC‐MS/MS analysis, and we found for the first time that hCE1 is present in human plasma as opposed to that in liver tissues. That is, from normalization of hCE1 signal by the immunoprecipitation and Western blot analysis, hCE1 levels were increased in plasma specimens from HCC patients than in plasma from other disease patient groups (e.g. liver cirrhosis, chronic hepatitis, cholangiocarcinoma, stomach cancer, and pancreatic cancer). From the receiver operating characteristic analysis in HCC, both sensitivity and specificity were shown to be greater than 70.0 and 85.0%, respectively. Thus, the high‐resolution proteomic approach demonstrates that hCE1 is a good candidate for further validation as a serologic glycoprotein biomarker for HCC.

Identification of human complement factor B as a novel biomarker candidate for pancreatic ductal adenocarcinoma

Pancreatic cancer (PC; pancreatic ductal adenocarcinoma) is characterized by significant morbidity and mortality worldwide. Although carbohydrate antigen (CA) 19-9 has been known as a PC biomarker, it is not commonly used for general screening because of its low sensitivity and specificity. Therefore, there is an urgent need to develop a new biomarker for PC diagnosis in the earlier stage of cancer. To search for a novel serologic PC biomarker, we carried out an integrated proteomic analysis for a total of 185 pooled or individual plasma from healthy donors and patients with five disease groups including chronic pancreatitis (CP), PC, and other cancers (e.g., hepatocellular carcinoma, cholangiocarcinoma, and gastric cancer) and identified complement factor b (CFB) as a candidate serologic biomarker for PC diagnosis. Immunoblot analysis of CFB revealed more than two times higher expression in plasma samples from PC patients compared with plasma from individuals without PC. Immunoprecipitation coupled to mass spectrometry analysis confirmed both molecular identity and higher expression of CFB in PC samples. CFB showed distinctly higher specificity than CA 19-9 for PC against other types of digestive cancers and in discriminating PC patients from non-PC patients (p < 0.0001). In receiver operator characteristic curve analysis, CFB showed an area under curve of 0.958 (95% CI: 0.956 to 0.959) compared with 0.833 (95% CI: 0.829 to 0.837) for CA 19-9. Furthermore, the Y-index of CFB was much higher than that of CA 19-9 (71.0 vs 50.4), suggesting that CFB outperforms CA 19-9 in discriminating PC from CP and other gastrointestinal cancers. This was further supported by immunoprecipitation and qRT-PCR assays showing higher expression of CFB in PC cell lines than in normal cell lines. A combination of CFB and CA 19-9 showed markedly improved sensitivity (90.1 vs 73.1%) over that of CFB alone in the diagnosis of PC against non-PC, with similar specificity (97.2 vs 97.9%). Thus, our results identify CFB as a novel serologic PC biomarker candidate and warrant further investigation into a large-scale validation and its role in molecular mechanism of pancreatic carcinogenesis.

A novel HDAC inhibitor, CG200745, inhibits pancreatic cancer cell growth and overcomes gemcitabine resistance

Pancreatic cancer is predominantly lethal, and is primarily treated using gemcitabine, with increasing resistance. Therefore, novel agents that increase tumor sensitivity to gemcitabine are needed. Histone deacetylase (HDAC) inhibitors are emerging therapeutic agents, since HDAC plays an important role in cancer initiation and progression. We evaluated the antitumor effect of a novel HDAC inhibitor, CG200745, combined with gemcitabine/erlotinib on pancreatic cancer cells and gemcitabine-resistant pancreatic cancer cells. Three pancreatic cancer-cell lines were used to evaluate the antitumor effect of CG200745 combined with gemcitabine/erlotinib. CG200745 induced the expression of apoptotic proteins (PARP and caspase-3) and increased the levels of acetylated histone H3. CG200745 with gemcitabine/erlotinib showed significant growth inhibition and synergistic antitumor effects in vitro. In vivo, gemcitabine/erlotinib and CG200745 reduced tumor size up to 50%. CG200745 enhanced the sensitivity of gemcitabine-resistant pancreatic cancer cells to gemcitabine, and decreased the level of ATP-binding cassette-transporter genes, especially multidrug resistance protein 3 (MRP3) and MRP4. The novel HDAC inhibitor, CG200745, with gemcitabine/erlotinib had a synergistic anti-tumor effect on pancreatic cancer cells. CG200745 significantly improved pancreatic cancer sensitivity to gemcitabine, with a prominent antitumor effect on gemcitabine-resistant pancreatic cancer cells. Therefore, improved clinical outcome is expected in the future.

Human liver carboxylesterase 1 outperforms alpha‐fetoprotein as biomarker to discriminate hepatocellular carcinoma from other liver diseases in Korean patients

Although alpha‐fetoprotein (AFP) is currently the major serologic biomarker for hepatocellular carcinoma (HCC), it cannot efficiently distinguish this cancer from other forms of liver disease in early diagnosis due to its low sensitivity. The aim of this study is to compare sensitivity and specificity of human carboxylesterase 1 (hCE1) and AFP biomarker. Antibody‐based assays for hCE1 and AFP were used to test both biomarkers with respect to diagnostic efficiency, Youdens index and the area under the curve (AUC) through receiver operating characteristic (ROC) analysis in plasma from 208 patients with HCC (n=57), liver cirrhosis (n=27), chronic hepatitis (n=37), cholangiocarcinoma (n=22), gastric cancer (n=31) and pancreatic cancer (n=34), along with 52 healthy donors (HDs). The levels of hCE1 were significantly higher in patients with HCC than HDs and the other diseases (p<0.005), further verified by AUC values and Youdens index. In the set of HCC versus liver cirrhosis the AUC values were 0.744 (AFP), 0.918 (hCE1) and 0.938 (combination of AFP and hCE1), respectively. These results indicate that hCE1 is not only a more potent and specific marker in distinguishing cancer from liver diseases, in particular cirrhosis, but the combination of hCE1 and AFP shows also synergistic potential for greater sensitivity and specificity in early diagnosis. Therefore the antibody‐based hCE1 assay appears to have high diagnostic efficiency for discriminating HCC from other forms of liver disease. It is now feasible to further validate this novel plasma‐based biomarker in the large cohort we assembled.

Combined use of CEMIP and CA 19-9 enhances diagnostic accuracy for pancreatic cancer

Carbohydrate antigen (CA) 19-9 is the only diagnostic marker used in pancreatic cancer despite its limitations. Here, we aimed to identify the diagnostic role of CEMIP (also called KIAA1199) combined with CA 19-9 in patients with pancreatic cancer. A retrospective analysis of prospectively collected patient samples was performed to determine the benefit of diagnostic markers in the diagnosis of pancreatic cancer. We investigated CEMIP and CA 19-9 levels in 324 patients with pancreatic cancer and 49 normal controls using serum enzyme-linked immunosorbent assay. Median CA 19-9 and CEMIP levels were 410.5 U/ml (40.8–3342.5) and 0.67 ng/ml (0.40–1.08), respectively, in patients with pancreatic cancer. The AUROC for CA 19-9 and CEMIP were 0.847 (95% confidence interval [CI]: 0.806–0.888) and 0.760 (95% CI: 0.689–0.831), respectively. Combination of CA 19-9 with CEMIP showed markedly improved AUROC over CA 19-9 alone in pancreatic cancer diagnosis (0.94 vs. 0.89; P < 0.0001). CEMIP showed a diagnostic yield of 86.1% (68/79) in CA 19-9 negative pancreatic cancer. Combined use with CEMIP showed significantly improved diagnostic value compared with CA 19-9 alone in pancreatic cancer. Especially, CEMIP may be a complementary marker in pancreatic cancer patients with normal CA 19-9 levels.

Abstract 259: Novel biliary tract cancer stem cell biomarker TAGLN2 regulates migration via c-met signaling cascades in biliary tract cancer cells.

Proceedings: AACR 104th Annual Meeting 2013; Apr 6-10, 2013; Washington, DC Background/Aims : Biliary tract cancer is one of the lethal malignancy. Most biliary tract cancer is diagnosed at advanced stages and have poor prognosis. Currently, there are no standard biomarkers and chemotherapeutic agents for biliary tract cancers. In order to find the biomarkers and therapeutic targets for biliary tract cancer, we have focused on the cancer stem cells (CSCs) with tumor initiating/self renewing/metastatic capacity and chemo/radio-resistance. In this study, we investigated the function of biliary tract CSC related biomarker TALGN2 discovered from comparision of cDNA microarray between sphere and adherent cell populations of biliary tract cancer cell lines. Methods : Analysis using cDNA microarray between sphere and adherent cell populations and selection of CSC specific biomarkers were performed. Among them, we selected TAGLN2 as a biliary CSC related gene. TAGLN2 expression was confirmed by RT-PCR, western blot and immunohistochemistry. To evaluate the function of TAGLN2 in biliary carcinogenesis, we performed targeted knockdown via siRNA in SNU1196 biliary tract cancer cell line. Results : The expression of stemness and c-met signaling cascades were up-regulated in sphere cells compared with adherent cells. TAGLN2 was also up-regulated in sphere cells. TAGLN2 was highly expressed in most of biliary tract cancer cell lines and biliary tract cancer tissues. TAGLN2 knockdown cells showed decreased cell migration and increased chemosentitivity compared with mock cells. In addition, we found that c-met signaling cascades including PI3K and MAPK were inactivated in TAGLN2 knockdown cells. Conclusions : These results suggest that biliary tract CSC biomarker TAGLN2 is implicated in biliary tract cancer progression and TAGLN2 may be a potential biomarker and therapeutic target for biliary tract cancer. Citation Format: Soo Been Park, Sun A Kim, Si Young Song. Novel biliary tract cancer stem cell biomarker TAGLN2 regulates migration via c-met signaling cascades in biliary tract cancer cells. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 259. doi:10.1158/1538-7445.AM2013-259

Role of overexpression of factors regulating pancreatic embryogenesis in acinar to ductal transdifferentiation in vitro and pancreatic carcinogenesis and establishment of two normal rat pancreatic ductal cell lines under the concept of acinar-ductal transidifferentiation

of the Na-K ATPase in the e/f ~-spectrin mutant phenotype resembles the Drosophila ~3spectrin mutant that is in turn reminiscent of the Drosophila labial phenotype, particularly in the gut. Control of the homeotic gene labial is dependent upon extmceilular gradients of wingless and decapentapleo~c (the Drosophila homologue of TGF-[3 ) during embryogenesis, suggesting that TGF-~/elf interactions are important for gut epithelial cell formation and are conserved through evolution.