Serum Anti-BPAG1 Auto-Antibody Is a Novel Marker for Human Melanoma

Malignant melanoma is one of the most aggressive types of tumor. Because malignant melanoma is difficult to treat once it has metastasized, early detection and treatment are essential. The search for reliable biomarkers of early-stage melanoma, therefore, has received much attention. By using a novel method of screening tumor antigens and their auto-antibodies, we identified bullous pemphigoid antigen 1 (BPAG1) as a melanoma antigen recognized by its auto-antibody. BPAG1 is an auto-antigen in the skin disease bullous pemphigoid (BP) and anti-BPAG1 auto-antibodies are detectable in sera from BP patients and are used for BP diagnosis. However, BPAG1 has been viewed as predominantly a keratinocyte-associated protein and a relationship between BPAG1 expression and melanoma has not been previously reported. In the present study, we show that bpag1 is expressed in the mouse F10 melanoma cell line in vitro and F10 melanoma tumors in vivo and that BPAG1 is expressed in human melanoma cell lines (A375 and G361) and normal human melanocytes. Moreover, the levels of anti-BPAG1 auto-antibodies in the sera of melanoma patients were significantly higher than in the sera of healthy volunteers (p<0.01). Furthermore, anti-BPAG1 auto-antibodies were detected in melanoma patients at both early and advanced stages of disease. Here, we report anti-BPAG1 auto-antibodies as a promising marker for the diagnosis of melanoma, and we discuss the significance of the detection of such auto-antibodies in cancer biology and patients

Keratinocyte Responsive Element 3: Analysis of a Keratinocyte-specific Regulatory Sequence in the 230-kDa Bullous Pemphigoid Antigen Gene Promoter

The 230-kDa bullous pemphigoid antigen gene is expressed primarily, if not exclusively, in basal keratinocytes of the epidermis. Keratinocyte responsive element 3, a cis-element at position –216 to –197 of the human 230-kDa bullous pemphigoid antigen gene promoter, confers tissue-specific expression to this gene (Tamai et al: J Biol Chem 270:7609–7614, 1995). In this study, we investigated the functional characteristics of keratinocyte responsive element 3 on the 230-kDa bullous pemphigoid antigen gene core promoter by transient transfections of cultured normal human keratinocytes and normal human fibroblasts, as well as of lung carcinoma (A549), osteosarcoma (OST), and gastric adenocarcinoma (GT3TKB) cell lines. A 230-kDa bullous pemphigoid antigen gene core promoter/luciferase reporter gene plasmid construct, pBPL, was modified to develop a series of constructs (pKBPL–p4KBPL), which have insertions of one, two, three, or four tandem repeats of keratinocyte responsive element 3, and these plasmids were used in transient transfections of the cultured cells. The promoter activities of pKBPL–p4KBPL constructs, relative to pBPL, in normal human keratinocytes were 7.6-, 15.5-, 4.6-, and 2.7-fold higher, respectively, whereas no upregulatory effect by keratinocyte responsive element 3 insertion was observed in other cell lines tested. prKBPL, a plasmid constructed with keratinocyte responsive element 3 in reverse orientation, showed essentially no activity in normal human keratinocytes. Insertion of a random 20 bp sequence between keratinocyte responsive element 3 and the 230-kDa bullous pemphigoid antigen gene core promoter resulted in about 40% reduction of luciferase activity in normal human keratinocytes. These data suggest that keratinocyte responsive element 3 functions as a position-, copy number-, and orientation-dependent cis-element contributing to tissue-specific regulation of the 230-kDa bullous pemphigoid antigen gene

Integration-free T cell-derived human induced pluripotent stem cells (iPSCs) from a patient with recessive dystrophic epidermolysis bullosa (RDEB) carrying two compound heterozygous mutations in the COL7A1 gene

Expanded human T cells from a Japanese female with recessive dystrophic epidermolysis bullosa (RDBE) were used to generate integration-free induced pluripotent stem cells (iPSCs) by exogenous expression of four reprogramming factors, OCT3/4, SOX2, cMYC, KLF4, using Sendai virus vector (SeVdp). The authenticity of established iPSC line, RDEB-iPSC26, was confirmed by the expressions of stem cell markers and the differentiation capability into three germ layer. RDEB-iPSC26 may be a useful cell resource for the establishment of in vitro RDEB modeling and the study for developing gene and cell therapy

Hallopeau-Siemens型劣性栄養障害型表皮水疱症における新規COL7A1ミスセンス変異

Dystrophic epidermolysis bullosa (DEB) is caused by mutations in the COL7A] gene encoding type VII collagen. DEB is clinically characterized by mucocutaneous blistering in response to minor trauma. followed by scarring and nail dystrophy. DEB is inherited in either an autosomal dominant (DDEB) or recessive (RDEB) fashion. DDEB basically results from a glycine substitution mutation within the collagenous domain on one COL7A1 allele. while a combination of mutations such as premature termination codon. missense. splice-site mutations on both alleles causes RDEB. In this study, we examined a Japanese EB boy with generalized blistering and scar formation. and made a diagnosis of the Hallopeau-Siemens type RDEB (HS-RDEB). the most severe form of RDEB. Mutational analysis of the COL7A1 gene revealed a novel missense mutation A80P and a novel nonsense mutation Q1211X. In general, HS-RDEB is caused by combination of premature termination codon mutations. but 3 HS-RDEB cases have been reported to have combination of premature termination codon and missense mutations one of which was S48P. This study suggests that even missense mutation, which leads to substitution for proline in amino terminal end of type VII collagen. can cause HS-RDEB