Serum cell-free DNA levels are a useful marker for extramammary Paget disease.

BACKGROUND: Although carcinoembryonic antigen (CEA) and cytokeratin 19 fragment (CYFRA) are useful markers for extramammary Paget disease (EMPD), serum CEA and CYFRA levels are not elevated in most patients with EMPD without metastasis. Cell-free (cf)DNA has attracted attention as an indicator of clinical conditions in several cancers. OBJECTIVES: To identify further useful biomarkers for the detection of EMPD, including early lesions, and to study the clinical implications of cfDNA in EMPD. METHODS: cfDNA were isolated from serum of patients with EMPD with and without metastasis, and from healthy volunteers. Serum extracts were amplified using polymerase chain reaction. RESULTS: Serum cfDNA levels were significantly elevated in patients with EMPD with or without metastasis compared with those in healthy controls. Serum cfDNA was a better diagnostic marker for the presence of EMPD than serum CYFRA. Moreover, the postoperative serum cfDNA levels were significantly lower than those from the preoperative samples, and the change in serum cfDNA levels reflected the clinical courses of patients with EMPD treated with chemotherapy. CONCLUSIONS: Taking the evidence together, serum cfDNA levels may be a useful marker for diagnosis and disease progression in EMPD. What's already known about this topic? Serum levels of carcinoembryonic antigen (CEA) and cytokeratin 19 fragment (CYFRA) are not elevated in most patients with extramammary Paget disease (EMPD) without metastasis. Cell-free (cf)DNA has attracted attention as an indicator of clinical conditions in several cancers. There are few reports of the clinical implications of cfDNA in dermatology. What does this study add? Serum cfDNA levels were significantly elevated in patients with EMPD with or without metastasis compared with those in healthy controls. Postoperative serum cfDNA levels were significantly lower than those from the preoperative samples. Changes in serum cfDNA levels reflected the clinical courses of patients with EMPD treated with chemotherapy. What is the translational message? Serum cfDNA levels in patients with EMPD are a useful marker for the detection of EMPD, including localized EMPD. Changes in serum cfDNA levels in an individual patient may reflect the clinical course of EMPD.

Inhibition of heat shock protein 90 exerts an antitumour effect in angiosarcoma: involvement of the vascular endothelial growth factor signalling pathway.

BACKGROUND: Angiosarcoma is a rare malignant neoplasm derived from endothelial cells, and because advanced angiosarcoma is resistant to standard chemotherapy its prognosis is poor. Therefore, new therapies are urgently needed. Heat shock protein (HSP)90 has been identified as a molecular chaperone that regulates various cancer-related proteins. Numerous clinical trials are currently testing the effectiveness of HSP90 inhibitors in various types of malignancies. OBJECTIVES: To investigate the role of HSP90 in the pathogenesis of angiosarcoma and whether the inhibition of HSP90 may have antitumour activity. METHODS: The expression of HSP90 protein in angiosarcoma was examined using immunohistochemistry and immunoblotting. The effects of HSP90 inhibition were proven using proliferation, migration and invasion assay in angiosarcoma cells. The mechanism of antitumour effect by HSP90 inhibition was investigated by the transfection of small interfering RNA (siRNA). RESULTS: The levels of HSP90 protein expression in cultured angiosarcoma cell lines were markedly increased compared with those in normal tissue cell lines. Immunohistochemical analyses revealed that the expression of HSP90 protein was strongly detected in angiosarcoma tissues compared with that in normal dermal vessels or senile angioma tissues. Ganetespib, an HSP90 inhibitor, with or without taxanes, inhibited the proliferation of angiosarcoma cells via apoptosis in a dose-dependent manner. HSP90 siRNA suppressed the proliferation, migration and invasion of angiosarcoma cells. Knock-down of HSP90 did not suppress vascular endothelial growth factor receptor 2 directly, but selectively suppressed several downstream targets of vascular endothelial growth factor signalling in angiosarcoma cells. CONCLUSIONS: HSP90 could be a novel therapeutic target for angiosarcoma.

The role of PSMB9 upregulated by interferon signature in the pathophysiology of cutaneous lesions of dermatomyositis and systemic lupus erythematosus.

BACKGROUND: Dermatomyositis (DM) and systemic lupus erythematosus (SLE) have common skin features, including dermal mucin deposition and interferon signature, although their roles are unknown. OBJECTIVES: To identify common or specific molecular changes in DM and SLE skin. METHODS: Proteomic analysis was performed using DM and healthy skin. Glycosaminoglycans were analysed by high-performance liquid chromatography. RESULTS: The expression of 60 proteins was upregulated or downregulated in DM skin compared with healthy skin in the proteomic analysis. Among those proteins, PSMB9, an immunoproteasome subunit, was upregulated in the epidermis of DM and SLE, but not in other skin diseases. Furthermore, versican V1, a core protein for glycosaminoglycans, was upregulated, while type I collagen was downregulated in the dermis of DM and SLE skin. Interferon stimulated PSMB9 expression in cultured keratinocytes and reduced collagen expression in dermal fibroblasts, but did not affect versican expression. The PSMB9 knock-down in keratinocytes led to significant suppression of transforming growth factor (TGF)-ß2 and TGF-ß3, inducers of versican synthesis. TGF-ß3 expression was upregulated in both DM and SLE, while TGF-ß2 expression was increased only in the DM epidermis. ΔDiHS-diS1, a component of heparan sulfate, was significantly increased only in DM. TGF-ß2 expression significantly increased the ΔDiHS-diS1 expression in dermal fibroblasts in vitro. CONCLUSIONS: The interferon signature in DM and SLE skin reduces collagen in dermal fibroblasts, whereas overexpression of PSMB9 induced by interferon stimulates versican inducers in epidermal keratinocytes. In addition, the TGF-ß2-ΔDiHS-diS1 pathway may be responsible for the specific molecular change in DM skin.

Role of c-Jun N-terminal kinase isoforms in the cellular activity of melanoma cell lines.

BACKGROUND: The c-Jun N-terminal kinase (JNK) is thought to be involved in inflammation, proliferation and apoptosis. AIM: To examine the role of JNK isoforms in metastasis, proliferation, migration and invasion of the malignant melanoma (MM) cell lines SK-MEL-28, SK-MEL-3 and WM164, using a kinase-specific inhibitor or isoform-specific small interfering (si)RNAs. RESULTS: SK-MEL-3, a cell line established from metastatic MM, showed slightly increased phosphorylation of both JNK1 and JNK2, whereas WM164, a cell line derived from primary MM, showed significant phosphorylation of JNK1. A JNK inhibitor, SP600125, inhibited cell proliferation of SK-MEL-3 but not SK-MEL-28 or WM164. Transfection of JNK1-specific siRNA reduced the migratory activity of WM164 cells, while silencing of either JNK1 or JNK2 strongly suppressed the invasive activity of SK-MEL-3. CONCLUSIONS: Our study suggests that JNK isoforms have different roles in MM. Metastasis of MM may be regulated by JNK2, while invasion is regulated by both JNK1 and JNK2. JNK1 and JNK2 respectively mediate cell migration and cell proliferation. Further understanding of the specific roles of JNK isoforms in the pathogenesis of MM may lead to the development of therapies targeting specific isoforms.

Association of linear IgA bullous disease with ulcerative colitis: a case of successful treatment with infliximab.

Linear IgA bullous disease (LABD) has been reported in association with inflammatory bowel disease, in particular ulcerative colitis (UC). We reporting a 34-year-old female who developed LABD during a flare-up of UC. We administered infliximab, which has been approved for the treatment of UC; infliximab dramatically improved the cutaneous lesions and bowel symptoms. This is the first report showing a marked effect of infliximab on LABD. First, we hypothesize that infliximab works for UC and then calms down excessive production of inflammatory cytokines and autoantibodies, and so stricter control of UC by infliximab is beneficial against the skin condition of LABD. Second, we suggest that TNF-α production in the lesion of LABD is increased, so TNF-α plays an important role in developing cutaneous lesions. This case suggests that infliximab, a monoclonal antibody against TNF-α, is efficacious in the cutaneous symptoms of LABD.

Serum epidermal growth factor receptor levels in patients with malignant melanoma.

BACKGROUND: Epidermal growth factor receptor (EGFR) is known to be abnormally expressed in many human carcinomas, suggesting that there may be an increase in serum EGFR levels in patients with malignant melanoma (MM) and that this might be a possible new tumour marker. AIM: To assess whether serum EGFR levels might be a marker of MM. METHODS: Serum samples were obtained from 66 patients with MM and 12 healthy controls, and EGFR levels were measured by double-determinant ELISA. RESULTS: Patients with in situ or stage I MM had significantly higher serum EGFR levels compared with healthy controls. Interestingly, serum EGFR levels decreased gradually with the stage of the tumour, being highest at stage I and lowest at stage IV. There was also a trend towards a reverse correlation between tumour thickness and serum EGFR levels. Moreover, a longitudinal study identified a trend for serum EGFR levels in patients with preoperative MM to decrease compared with patients with recurrent MM. CONCLUSIONS: To our knowledge, this is the first report investigating the serum EGFR levels of patients with MM, and gives new insight into the relationship between EGFR and MM. We found that serum EGFR levels were significantly increased in patients with early-stage MM such as in situ and stage I tumours. Measurements of serum EGFR levels might be of clinical value in the detection of early-stage MM.

Overexpression of hepatocyte growth factor receptor in scleroderma dermal fibroblasts is caused by autocrine transforming growth factor ß signaling.

Cutaneous fibrosis seen in systemic sclerosis (SSc) is caused by fibroblast activation and abnormal collagen accumulation due to 'autocrine transforming growth factor (TGF)-ß/Smad signaling'. Hepatocyte growth factor (HGF) may have therapeutic value against SSc, because of its inducible effect on the expression of matrix metalloproteinase (MMP)-1. Previous studies indicated SSc dermal fibroblasts overexpress HGF receptor c-met, which suggest specific and effective induction of MMP-1 in SSc fibroblasts caused by HGF treatment. However, the exact mechanism of c-met overexpression in SSc cells was hardly investigated. We hypothesized that such c-met overexpression is also caused by autocrine TGF-ß/Smad signaling. Expression of c-met protein in cultured SSc dermal fibroblasts was significantly up-regulated compared with that in normal fibroblasts. Ectopic TGF-ß stimulation induced c-met synthesis in normal fibroblasts, while a TGF-ß knockdown normalized the up-regulated c-met levels in SSc fibroblasts. Furthermore, we found the c-met promoter contains a putative binding site for Smads, and the binding activity of Smad2/3 to the c-met promoter was constitutively up-regulated in SSc fibroblasts as well as in normal fibroblasts treated with exogenous TGF-ß1. Taken together, c-met may be overexpressed due to autocrine TGF-ß/Smad signaling in SSc. Considering that HGF has an antifibrotic effect, such c-met overexpression in SSc fibroblasts may be a negative feedback against cutaneous fibrosis. Clarifying the mechanisms of c-met overexpression and controlling the HGF/c-met pathway may lead to a new therapeutic approach for this disease.

The expression of HER-2 in extramammary Paget's disease.

Extramammary Paget's disease (EMPD) is a rare intraepidermal adenocarcinoma. The common sites of EMPD involvement are the vulva, perineal, perianal, scrotal and penile skin. Several studies have shown that HER-2/neu, also known as c-erbB-2, is amplified and overexpressed in many cancers. In this study, we investigated the expression and clinical significance of HER-2 in Japanese patients with EMPD. Keratinocytes in epidermis were slightly positive for HER-2. As for EMPD, 19 of 31 EMPD were positive for HER-2 (61%). There is significant correlation between the presence of invasion and strong positivity (3+) for HER-2 (p < 0.02). Furthermore, there is significant correlation between the presence of lymph node metastasis and strong positivity (3+) for HER-2 (p < 0.02). These results suggest that patients with EMPD strongly positive for HER-2 may have high risk for lymph node metastasis and should be followed up carefully. The observed overexpression of HER-2 in EMPD presents a potential therapeutic target for adjuvant treatment of this disease. Treatment with trastuzumab is well established in breast cancer with HER-2 overexpression and is recommended by several consensus statements. The results of the present study indicate that targeting therapies for HER-2, such as trastuzumab, may be used for EMPD particularly in patients with invasive and/or metastatic EMPD.

microRNA-mediated keratinocyte hyperproliferation in psoriasis vulgaris.

BACKGROUND: Psoriasis is a chronic inflammatory skin disease characterized by intense proliferation and abnormal differentiation of keratinocytes, although the pathogenesis is still not completely clarified. OBJECTIVES: We investigated the mechanism of keratinocyte proliferation seen in psoriasis, focusing on microRNA (miRNA). MATERIALS AND METHODS: miRNAs were extracted from tissues and sera of psoriasis, atopic dermatitis and healthy control. To determine pathogenic miRNAs, we performed miRNA polymerase chain reaction (PCR) array analysis. The results were confirmed with quantitative real-time PCR, in situ hybridization, immunohistochemistry, transient transfection of siRNA and inhibitor in cultured keratinocytes and Western blotting. RESULTS: PCR array analysis using tissue miRNA demonstrated miR-424 level was markedly decreased in psoriasis skin in vivo. Protein expression of mitogen-activated protein kinase kinase 1 (MEK1) or cyclin E1, predicted target genes of miR-424, was increased in psoriatic skin, although their mRNA levels were not. The transfection of specific inhibitor of miR-424 in normal human keratinocytes led to upregulation of MEK1 or cyclin E1 protein, and resulted in increased cell proliferation. On the other hand, cell number was significantly decreased when cells were transfected with siRNA for MEK1 or cyclin E1. Furthermore, we first investigated serum miRNA levels in psoriasis. Although not significant, serum miR-424 concentration tended to be decreased in patients with psoriasis compared with healthy controls. CONCLUSIONS: Decreased miR-424 expression and subsequently increased MEK1 or cyclin E1 may play a key role in the pathogenesis of psoriasis. Investigation of the regulatory mechanisms of keratinocyte proliferation by miRNA may lead to new treatments and a disease activity marker.

Clinical and laboratory features of Japanese patients with scleroderma and telangiectasia.

BACKGROUND: Systemic sclerosis (SSc) is a connective tissue disease characterized by sclerotic changes of the skin and internal organs. Telangiectasia is a frequent complication of patients with SSc. OBJECTIVE: To examine the prevalance of telangiectasia in patients with SSc and investigate the clinical and laboratory features of patients with SSc and telangiectasia. METHODS: In total, 211 patients with SSc who fulfilled the diagnostic criteria for SSc of the American College of Rheumatology were examined by laboratory and clinical methods. The average of disease duration time was 7.4 years. RESULTS: Telangiectasia was found in 119 of the 211 patients (56%) with SSc. The prevalence of oesophageal involvement, decreased diffusing capacity for carbon monoxide (DLCO), heart involvement, calcinosis, shortening of the sublingual frenulum, or pitting scars was significantly greater in patients with telangiectasia than in those without telangiectasia. CONCLUSION: Our study suggests that the presence of telangiectasia may be a marker of oesophageal involvement, decreased DLCO, and heart involvement.

Elevated circulating soluble CD40 ligand in patients with mixed connective tissue disease.

Abnormalities of CD40 ligand have been demonstrated in various kinds of diseases. Our objective was to determine serum soluble CD40 ligand (sCD40L) levels in patients with mixed connective tissue disease (MCTD). Serum sCD40L levels of 23 patients with MCTD and 21 healthy individuals were measured with specific enzyme-linked immunosorbent assays. Serum levels of sCD40L were significantly higher in patients with MCTD than in healthy individuals (median 0.179 ng/ml vs 0.99 ng/ml, 25th-75th percentile; 0.117-0.296 ng/ml vs 0.82-1.70 ng/ml, P<0.005), whereas there were no significant correlation between elevated serum sCD40L levels and clinical or serological features in patients with MCTD. These results suggest that sCD40L plays a role in the pathogenesis of MCTD. Further studies are needed to clarify this.

A case of linear scleroderma with muscle calcification.

We report a 21-year-old man with linear scleroderma with muscle calcification revealed by a computed tomographic (CT) scan. Levels of serum calcium and phosphorus were normal. This is the first report of localized scleroderma with muscle calcification. Muscle involvement may cause contracture or functional abnormalities of the extremities. Therefore, CT scans may be useful in evaluating muscle involvement in patients with localized scleroderma.