Identification of autoantibody against cytokeratin 8 in HBx transgenic mouse and its application to diagnosis of human breast cancer

Ah Ruem Kim,Chang-Kyu Heo,Hai-min Hwang,Dae-Yeul Yu,Ju Yeon Lee,Jong Shin Yoo,Sejeong Oh,Jeong Heon Ko,Eun-Wie Cho 한국당과학회 2012 한국당과학회 학술대회 Vol.2012 No.1

In many cases of cancers, aberrantly expressed or modified tumor-associated antigens leads to the formation of tumor-associated autoantibodies. Recent studies have demonstrated that autoantibodies are appropriate to serve as biomarkers for the early detection of cancers, if autoantibodies and their target antigens are well characterized. In previous studies, we established a B cell hybridoma pool derived from hepatitis B virus X (HBx) transgenic mouse, a hepatocellular carcinoma model mouse, and characterized autoantibodies from B cell hybridoma and their specific target antigens using purified autoantibodies. In this study, XE278 autoantibody, one of them, was purified and characterized. Flow cytometric analysis revealed that target antigen of XE278 autoantibody is highly expressed in various cancer cell lines as compared to normal ones, especially in breast cancer cell lines. Its target antigen, with molecular weight of 55 kDa, was identified by immunoprecipitation and MS-based proteomic methods as cytokeratin 8 (CK8), an intermediate filament protein which is expressed in epithelial and carcinoma cells and correlated with increased invasiveness of tumor. Moreover, two specific mimotopes (XE278-p1 and XE278-p2) against XE278 autoantibody were screened from the cyclic random hepta-peptide phage libraries, which can be used as baits for capturing target autoantibodies. Using XE278-p2-display M13 phage as coating antigen in enzyme-linked immunosorbent assay (ELISA), we could distinguish breast cancer patients from normal subjects. Furthermore, the combination assay using XE278-p2 and K94-p1, another specific mimotope against K94 autoantibody which targets CK8/18 complex, improved the specificity and sensitivity of diagnosis to discriminate cancer patients from normal ones. These results indicate that anti-CK8 autoantibody is induced in breast cancer patients and the detection of anti-CK8 autoantibody can be used for the diagnosis of breast cancer.

Distinct roles of two ceramide synthases, CaLag1p and CaLac1p, in the morphogenesis of <i>Candida albicans</i>

Cheon, Seon Ah,Bal, Jyotiranjan,Song, Yunkyoung,Hwang, Hai‐,min,Kim, Ah Ruem,Kang, Woo Kyu,Kang, Hyun Ah,Hannibal‐,Bach, Hans K.,Knudsen, Jens,Ejsing, Christer S.,Kim, Jeong‐,Yoon Blackwell Publishing Ltd 2012 Molecular microbiology Vol.83 No.4

<P><B>Summary</B></P><P>Lag1p and Lac1p catalyse ceramide synthesis in <I>Saccharomyces cerevisiae.</I> This study shows that Lag1 family proteins are generally required for polarized growth in hemiascomycetous yeast. However, in contrast to <I>S. cerevisiae</I> where these proteins are functionally redundant, <I>C. albicans</I> Lag1p (CaLag1p) and Lac1p (CaLac1p) are functionally distinct. Lack of CaLag1p, but not CaLac1p, caused severe defects in the growth and hyphal morphogenesis of <I>C. albicans</I>. Deletion of <I>CaLAG1</I> decreased expression of the hypha‐specific <I>HWP1</I> and <I>ECE1</I> genes. Moreover, overexpression of <I>CaLAG1</I> induced pseudohyphal growth in this organism under non‐hypha‐inducing conditions, suggesting that CaLag1p is necessary for relaying signals to induce hypha‐specific gene expression. Analysis of ceramide and sphingolipid composition revealed that CaLag1p predominantly synthesizes ceramides with C24:0/C26:0 fatty acid moieties, which are involved in generating inositol‐containing sphingolipids, whereas CaLac1p produces ceramides with C18:0 fatty acid moieties, which are precursors for glucosylsphingolipids. Thus, our study demonstrates that CaLag1p and CaLac1p have distinct substrate specificities and physiological roles in <I>C. albicans</I>.</P>

경량 전기 항공기용 65kW급 추진전동기의 비행 프로파일에 따른 특성 해석

김래은(Rae-Eun Kim),노아름(Ah-Ruem Ro),이정훈(JeongHoon Lee),서정무(Jung-Moo Seo) 한국항공우주학회 2022 한국항공우주학회 학술발표회 논문집 Vol.2022 No.11

Identification of a mimotope for circulating anti-cytokeratin 8/18 antibody and its usage for the diagnosis of breast cancer

HEO, CHANG-KYU,HWANG, HAI-MIN,RUEM, AH,YU, DAE-YEUL,LEE, JU YEON,YOO, JONG SHIN,KIM, IN GYU,YOO, HYANG SOOK,OH, SEJEONG,KO, JEONG HEON,CHO, EUN-WIE D.A. Spandidos 2013 International journal of oncology Vol.42 No.1

<P>A novel circulating tumor-associated autoantibody, K94, obtained from a hepatocellular carcinoma (HCC) mouse model was characterized. The target antigen of K94 autoanti-body was expressed in various tumor cell lines including liver cancer, and its secretion was detectable using MCF-7 breast carcinoma cells. Proteomic analysis revealed that the protein bands reactive to K94 included cytokeratin (CK) 8 and 18, which are known to be related to tumorigenesis and form a heterotypic complex with each other. However, K94 showed no activity toward CK8 or CK18 separately. The epitope of the K94 antibody was only presented by a complex between CK8 and CK18, which was confirmed by analysis using recombinant CK8 and CK18 proteins. To formulate an assay for anti-CK8/18 complex autoantibody, a mimotope peptide reactive to K94 was selected from loop-constrained heptapeptide (-CX<SUB>7</SUB>C-) display phage library, of which sequence was CISPDAHSC (K94p1). A mimotope enzyme-linked immunosorbent assay (ELISA) using phage-displayed K94p1 peptide as a coating antigen was able to discriminate breast cancer (n=30) patients from normal subjects (n=30) with a sensitivity of 50% and a specificity of 82.61%. CA15.3 was detected at very low levels in the same breast cancer subjects and did not discriminate breast cancer patients from normal subjects, although it is a conventional biomarker of breast cancer. These results suggest that a mimotope ELISA composed of K94p1 peptide may be useful for the diagnosis of breast cancer.</P>

Autoantibody responses to AFP in liver diseases and its usage for the early diagnosis of Hepatocellular Carcinoma (HCC)

Hai-Min Hwang,Chang-Kyu Heo,Ah Ruem Kim,Chulhun Chang,Jeong Heon Ko,Eun-Wie Cho 한국당과학회 2012 한국당과학회 학술대회 Vol.2012 No.1

Hepatocellular carcinoma (HCC) is one of the most common cancers worldwide, particularly prevalent in Asia, with very poor prognosis. Although α-fetoprotein (AFP) is the most effective biomarker available to detect HCC, it has a low clinical sensitivity and specificity as biomarker. Therefore, there is a need for the development of more sensitive and specific assays that can supplement AFP for the early detection of liver cancer. Recently, autoantibodies, which are generated by immune system recognizing the presence of the abnormal tumor-associated antigens, are suggested as promising biomarkers for the early detection of tumors. In this study, to improve the early detection and diagnosis of HCC, determination of autoantibody responses to AFP in HCC patients was performed and analyzed its correlation with the progression of disease. The serum AFP levels were measured using sandwich ELSA commercially available and anti-AFP autoantibody levels were determined by ELISA using purified AFP from HepG2 cells as coating antigen. Assays were performed for 50 healthy controls, 50 patients with HCC, 36 patients with liver cirrhosis, and 35 patients with chronic hepatitis. Anti-AFP autoantibodies were detected in patients’ sera showing elevated AFP levels, which revealed that AFP secreted from tumor cells elicit immune response which induce tumor-associated autoantibodies. Even when the AFP level was very low anti-AFP responses were detectable, impling that autoantibody response to AFP can be used as an early detection biomarker. Several patient sera showing highly elevated AFP, however, did not show anti-AFP antibody response, which might be related to the intrinsic property of the AFP molecule as an immune suppressor. In addition, simultaneous detection of AFP and anti-AFP response in patients’ sera improved the diagnosis of HCC and we discuss about its usage as HCC diagnosis biomarker.

Identification of a Tumor-Associated Autoantibody against Aminoimidazole Carboxamide Ribonucleotide Transformylase/Inosine Monophosphate Cyclohydrolase (ATIC) in Hepatocellular Carcinoma (HCC) Mouse Model and Its Application to Diagnosis of HCC

Chang-Kyu Heo,Mi-Kyung Woo,Hai-Min Hwang,Ah Ruem Kim,Dae-Yeul Yu,Ju Yeon Lee,Jong Shin Yoo,Chulhun Chang,Jeong Heon Ko,Eun-Wie Cho 한국당과학회 2012 한국당과학회 학술대회 Vol.2012 No.1

Autoantibodies, which are generated by immune system recognizing the presence of the abnormal tumor-associated antigens, are promising biomarkers for the early detection of tumors. Recently, we established a B cell hybridoma pool derived from HBx transgenic mouse, as a source of tumor-associated autoantibodies without using any extracellular antigens, and have characterized the specific target antigens against them. XC154 autoantibody, one of them, has been investigated in this study and its target antigen was identified by mass spectrometric analysis as aminoimidazole carboxamide ribonucleotide transformylase / inosine monophosphate cyclohydrolase (ATIC) that catalyzes the last two steps of de novo purine biosynthesis. A specific mimotope against XC154 autoantibody was screened from the cyclic random hepta-peptide phage library and, using mimotope display M13 phage as a coating antigen for ELISA, we could distinguish patients with hepatocellular carcinoma (HCC) vs. normal subjects with 86.96% sensitivity and 88.24% specificity. These results imply that anti-ATIC autoantibody is induced in patients with HCC and detection of anti-ATIC autoantibody can be used for the diagnosis of HCC. Now, several attempts have been performed to establish a more reliable and convenient assay to detect tumor-assocoated autoantibodies using recombinant proteins displaying cycling peptide instead of mimotope display M13 phage and the usefulness of improved assay will be discussed.