Identification of CCDC62‐2 as a novel cancer/testis antigen and its immunogenicity

Abstract

Cancer/testis (CT) antigens are expressed in normal germ line tissues and various cancers. They are considered promising target molecules for immunotherapy for patients with various cancers. To identify CT antigens, we performed serological identification of antigens by recombinant expression cloning. The humoral immune response of cancer patients against a newly defined antigen was analyzed. A testicular cDNA library was immunoscreened with serum obtained from a gastric adenocarcinoma patient whose primary cancer had regressed once and most liver metastasies had disappeared transiently. We isolated 55 positive cDNA clones comprising 23 different genes. They included 4 genes with testis-specific expression profiles in the Unigene database, including coiled-coil domain containing 62 (CCDC62). RT-PCR analysis showed that the expression of 2 splice variants of CCDC62 was restricted to the testis in normal adult tissues. In malignant tissues, CCDC62 variant 2 (CCDC62-2) was aberrantly expressed in a variety of cancers, including stomach cancer. A serological survey of 191 cancer patients with a range of different cancers by ELISA revealed antibodies to CCDC62-2 in 13 patients, including stomach cancer. None of the 41 healthy donor serum samples were reactive in the same test. The serum reaction against CCDC62-2 was confirmed by western blot. CCDC62-2 is a CT antigen that is immunogenic in cancer patients.

Mass spectrometry of peptides and proteins from human blood

“…Early indications were that parent ion intensities from MALDI analysis were in agreement with those of LC-ESI-MS (Griffin et al, 2001) and that parent and fragment ion intensities between separate runs of normal and diseased samples were consistent with Western blots or ELISA analysis of the same sample fractions (Marshall et al, 2003;Reynolds et al, 2003;Zhang et al, 2004;Malik et al, 2005;Yokoi et al, 2005;Yang et al, 2006;Haqqani et al, 2007;Morgan et al, 2007;Wilson et al, 2007;Barba de la Rosa et al, 2008;Gramolini et al, 2008;Hammerer-Lercher et al, 2008;Hao et al, 2008;Kolialexi et al, 2008;Kulasingam et al, 2008;Luque-Garcia et al, 2008;Marchi et al, 2008;Nicol et al, 2008;Plavina et al, 2008;Reichel, 2008;Xing et al, 2008;Wu et al, 2008a;Wang et al, 2008b;Zhao et al, 2008b;Domae et al, 2009;Fortin et al, 2009). Hence, where there is doubt regarding the best fit of the data, a second method such as accurate parent mass, targeted MS/MS, MRM, synthetic chromatography and spectrometry standards or immunological confirmation may be employed (Marshall et al, 2003;Zhang et al, 2004;Malik et al, 2005;Yokoi et al, 2005;Gao et al, 2005a;Haqqani et al, 2007;Plavina et al, 2007;Sardana,...…”

Cancer/testis antigens: A prospective reagent as diagnostic and immunotherapeutic targets for squamous cell carcinoma of the head and neck

“…SSX2, SYCP-1, and NY-ESO-1 were isolated using cDNA libraries from cancer or normal testis tissues [26][27][28]. Additional CT antigens, such as XAGE-1, CCDC62-2, GKAP1, and TEKT5, were identified in our SEREX analysis [29][30][31][32].…”

“…Western Blot analysis revealed reactions against recombinant CCDC62-2 molecules. CCDC62-2 was identified as a novel CT antigen that is immunogenic in cancer patients [30].…”

“…It was isolated by a yeast two-hybrid system using a mouse embryo library and reported as the male germ cell-specific 42-kDa protein, GKAP42 [63]. We successfully cloned human GKAP1 cDNA and demonstrated that the expression of GKAP1 mRNA was the highest in the testis of normal adult tissues and in various cancers [30,31]. GKAP1 functions as an anchoring protein for cGK-I␣ and appears to be similar to AKAP (A kinase anchoring protein), which binds to cAK via its regulatory subunits.…”

Isolation and characterization of human lung cancer antigens by serological screening with autologous antibodies

“…The screening procedure was described previously [18,20]. In brief, serum samples that had been diluted 1:10 were preabsorbed with lysate from Escherichia coli Y1090/Y1089 and coupled to Sepharose 4B (BioDynamics Lab Inc., Tokyo, Japan).…”