Abstract Background Deficiency of DNA mismatch repair (MMR) induces microsatellite instability (MSI). Pembrolizumab, an antibody targeting PD‐1 (an immune checkpoint inhibitor), is more effective against MMR‐deficient tumours than against MMR‐proficient tumours. The status of MMR is a useful biomarker for predicting the effectiveness of pembrolizumab administration. Although the status of MMR has attracted attention in skin tumours, there are few reports on MSI in extramammary Paget's disease (EMPD). Objectives To evaluate the status of MMR in patients with EMPD. Materials & Methods One hundred one patients with EMPD were included. MMR status of the genomic DNA of each subject was analysed using Promega panel (approved as a companion diagnostic agent for the administration of pembrolizumab). Results MSI testing showed the occurrence rates of MSI‐high (more than two markers are unstable), MSI‐low (one marker is unstable) and MSS (all markers are stable) tumour tissues were 0% (0/101), 1.0% (1/101) and 99.0% (100/101), respectively. Conclusion The status of MMR may not be useful for the potential therapeutic application of pembrolizumab.

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)-β2 and TGF-β3, inducers of versican synthesis. TGF-β3 expression was upregulated in both DM and SLE, while TGF-β2 expression was increased only in the DM epidermis. ΔDiHS-diS1, a component of heparan sulfate, was significantly increased only in DM. TGF-β2 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-β2-ΔDiHS-diS1 pathway may be responsible for the specific molecular change in DM skin.

Journal Article The miR‐146a rs2910164 C/G polymorphism is associated with telangiectasia in systemic sclerosis Get access A. Sakoguchi, A. Sakoguchi Department of Dermatology and Plastic Surgery, Faculty of Life Sciences, Kumamoto University, Kumamoto, Japan E‐mail: mjin@kumamoto-u.ac.jp Search for other works by this author on: Oxford Academic Google Scholar M. Jinnin, M. Jinnin Department of Dermatology and Plastic Surgery, Faculty of Life Sciences, Kumamoto University, Kumamoto, Japan E‐mail: mjin@kumamoto-u.ac.jp Search for other works by this author on: Oxford Academic Google Scholar T. Makino, T. Makino Department of Dermatology and Plastic Surgery, Faculty of Life Sciences, Kumamoto University, Kumamoto, Japan E‐mail: mjin@kumamoto-u.ac.jp Search for other works by this author on: Oxford Academic Google Scholar I. Kajihara, I. Kajihara Department of Dermatology and Plastic Surgery, Faculty of Life Sciences, Kumamoto University, Kumamoto, Japan E‐mail: mjin@kumamoto-u.ac.jp Search for other works by this author on: Oxford Academic Google Scholar K. Makino, K. Makino Department of Dermatology and Plastic Surgery, Faculty of Life Sciences, Kumamoto University, Kumamoto, Japan E‐mail: mjin@kumamoto-u.ac.jp Search for other works by this author on: Oxford Academic Google Scholar N. Honda, N. Honda Department of Dermatology and Plastic Surgery, Faculty of Life Sciences, Kumamoto University, Kumamoto, Japan E‐mail: mjin@kumamoto-u.ac.jp Search for other works by this author on: Oxford Academic Google Scholar W. Nakayama, W. Nakayama Department of Dermatology and Plastic Surgery, Faculty of Life Sciences, Kumamoto University, Kumamoto, Japan E‐mail: mjin@kumamoto-u.ac.jp Search for other works by this author on: Oxford Academic Google Scholar K. Inoue, K. Inoue Department of Dermatology and Plastic Surgery, Faculty of Life Sciences, Kumamoto University, Kumamoto, Japan E‐mail: mjin@kumamoto-u.ac.jp Search for other works by this author on: Oxford Academic Google Scholar S. Fukushima, S. Fukushima Department of Dermatology and Plastic Surgery, Faculty of Life Sciences, Kumamoto University, Kumamoto, Japan E‐mail: mjin@kumamoto-u.ac.jp Search for other works by this author on: Oxford Academic Google Scholar H. Ihn H. Ihn Department of Dermatology and Plastic Surgery, Faculty of Life Sciences, Kumamoto University, Kumamoto, Japan E‐mail: mjin@kumamoto-u.ac.jp Search for other works by this author on: Oxford Academic Google Scholar Clinical and Experimental Dermatology, Volume 38, Issue 1, 1 January 2013, Pages 99–100, https://doi.org/10.1111/j.1365-2230.2012.04453.x Published: 01 January 2013

MicroRNA-146a (miR-146a) is one of the microRNAs (miRNAs) implicated in the pathogenesis of various cancers. Recently, single nucleotide polymorphisms (SNPs) located in miRNAs themselves, so-called MIRSNPs, have attracted attention because of their possible involvement in the pathogenesis of various diseases. Such MIRSNPs may play functional roles because of the alteration of the miRNA. In this study, we investigated whether MIRSNP rs2910164 in miR-146a is involved in the pathogenesis of malignant melanoma (MM). DNA samples were collected from 50 patients with MM and 107 controls and genotyped by a PCR-restriction fragment. In patients with MM, the genotype distributions were 15 CC (30.0%), 35 CG (70.0%), and 0 GG (0.0%). The CG genotype was significantly increased in the patients compared with the controls (P = 0.02). The minimum free energy between miR-146a and its complementary strand with the G allele was determined to be -26.8 kcal/mol, whereas that of the C allele was -24.0 kcal/mol, indicating that the change from C to G may increase the stability of the miR-146a. However, there was no significant difference between the CC and the CG genotypes in terms of the relative expression levels of miR-146a. Human melanoma cell lines with the G allele showed significantly higher proliferation, migration, and invasion than those with the C allele. In conclusion, miR-146a may be involved in the pathogenesis of MM, and individuals with the CG genotype have an increased risk of developing MM. Melanoma Res 23: 13-20 (C) 2013 Wolters Kluwer Health vertical bar Lippincott Williams & Wilkins. Melanoma Research 2013, 23:13-20

Overexpression of integrins in dermal fibroblasts is thought to play a key role in the pathogenesis of systemic sclerosis (SSc), but the mechanism is unknown. We evaluated the possibility that microRNAs (miRNAs) are involved in the regulation of integrin β3 in these cells. The miRNA expression profile was determined by miRNA PCR array and real-time PCR. Protein expression of integrin β3 was determined by immunoblotting. In vivo detection of miRNA in paraffin section was performed by in situ hybridization. miR-150 expression was decreased in SSc fibroblasts both in vivo and in vitro. The transfection of miR-150 inhibitor into normal fibroblasts induced expression of integrin β3, phosphorylated Smad3, and type I collagen, whereas forced overexpression of the miRNA resulted in their down-regulation in SSc fibroblasts. Treatment of SSc fibroblasts with 5-AdC revealed that miR-150 down-regulation in these cells is caused by DNA methylation. In addition, we found that miR-150 is detectable and quantitative in serum. Serum miR-150 levels were decreased in SSc patients, and the SSc patients with lower serum miR-150 levels tended to have more severe clinical manifestations. miR-150 may play an important role in the pathogenesis of SSc via overexpression of integrin β3. Investigation of the regulatory mechanisms of tissue fibrosis by miR-150 could lead to development of new diagnostic tools and new treatments using miRNA.

It has been shown recently that immunotherapy for advanced melanoma is effective. However, in order to improve the efficacy of immunotherapy, the identification of more specific melanoma-associated antigens is urgently needed. Kinesin family member 20A (KIF20A) has been reported to be a promising immunotherapeutic target for pancreatic cancer. To investigate the expression of KIF20A in melanoma, we performed quantitative reverse transcript (RT)-PCR and western blotting analyses of melanoma cell lines. We also investigated primary melanomas and naevus tissues with immunohistochemistry and real-time RT-PCR. KIF20A expression was detected in 59% of melanomas and 12% of naevi by immunohisto-chemistry, and 64% of melanomas and 60% of naevi by real-time RT-PCR. The primary melanomas that were positive for KIF20A showed a significantly greater thickness than those that were negative, and patients with KIF20A+ melanoma tended to develop recurrence earlier. These results suggest that immunotherapy with KIF20A may be a novel treatment option for advanced melanoma.

Background. Recently, increased evidence has shown that serum micro (mi)RNA levels are a useful biomarker for the diagnosis, prognosis and therapeutic value of various diseases. However, serum miRNA has not been investigated in patients with systemic sclerosis (SSc), to our knowledge. Aim. To investigate the possibility that serum levels of Homo sapiens miR-142 stemloop (hsa-miR-142-3p), one of the miRNAs regulating the expression of integrin aV, could be a specific disease marker for SSc. Methods. Serum samples were obtained from 61 patients with SSc and 20 healthy controls. Patients with systemic lupus erythematosus (SLE), dermatomyositis (DM) and scleroderma spectrum disorder (SSD), who did not fulfil American College of Rheumatology criteria for SSc but might develop SSc in the future, were included as disease controls in this study. miRNAs were purified from serum, and miR-142-3p levels were measured with a quantitative real-time PCR assay. Results. Serum miR-142-3p levels in patients with SSc were significantly higher than in patients with SSD, SLE or DM, and healthy control groups. Patients with increased miR-142-3p levels tended to have a short sublingual frenulum. Conclusions. Our data indicate that serum levels of miR-142-3p may be elevated specifically in patients with SSc, correlating with the severity of this disease, and may be useful diagnostic markers for the presence of SSc and for the differentiation of SSc from SSD.

Background. Recently, increased evidence has shown that serum micro (mi)RNA levels are a useful biomarker for the diagnosis, prognosis and therapeutic value of various diseases. However, serum miRNA has not been investigated in patients with systemic sclerosis (SSc), to our knowledge.Aim. To investigate the possibility that serum levels of Homo sapiens miR-142 stemloop (hsa-miR-142-3p), one of the miRNAs regulating the expression of integrin aV, could be a specific disease marker for SSc.Methods. Serum samples were obtained from 61 patients with SSc and 20 healthy controls. Patients with systemic lupus erythematosus (SLE), dermatomyositis (DM) and scleroderma spectrum disorder (SSD), who did not fulfil American College of Rheumatology criteria for SSc but might develop SSc in the future, were included as disease controls in this study. miRNAs were purified from serum, and miR-142-3p levels were measured with a quantitative real-time PCR assay.Results. Serum miR-142-3p levels in patients with SSc were significantly higher than in patients with SSD, SLE or DM, and healthy control groups. Patients with increased miR-142-3p levels tended to have a short sublingual frenulum.Conclusions. Our data indicate that serum levels of miR-142-3p may be elevated specifically in patients with SSc, correlating with the severity of this disease, and may be useful diagnostic markers for the presence of SSc and for the differentiation of SSc from SSD.

BACKGROUND:Vascular abnormalities are one of the primary pathological components of systemic sclerosis (SSc). However, it has not been determined if there are also abnormalities in the formation of lymphatic vessels in SSc.OBJECTIVE:To evaluate lymphangiogenic activity in SSc skin.METHODS:The numbers of D2-40-positive lymphatic vessels in skin specimens from healthy control subjects and patients with SSc were counted and compared. Quantitative real-time polymerase chain reaction (PCR) was performed to determine mRNA levels of vascular endothelial growth factor (VEGF)-D and Flt-4 (fms-related tyrosine kinase 4, VEGFR-3, one of the receptors for VEGF-D) in the skin. Serum VEGF-D levels were measured with specific enzyme-linked immunosorbent assays. RESULTSZ: The number of lymphatic vessels in patients with SSc was significantly decreased compared with healthy control subjects. Mean relative transcript levels of FIGF (VEGF-D) and FLT4 (Flt-4) in skin tissue from patients with SSc were significantly increased compared with healthy control subjects. By the analysis of the association between serum VEGF-D levels and the clinical or laboratory features, we found that patients with SSc with higher serum VEGF-D levels more frequently have skin ulcers than those with normal VEGF-D levels.CONCLUSIONS:A systemic increase of VEGF-D, as well as local overexpression of FIGF and FLT4, may be the cause of disturbed lymphangiogenesis in SSc skin and play a role in the pathogenesis of SSc. We showed the possibility that regulation of VEGF-D/Flt-4 signalling could lead to new treatment of skin ulcers in SSc by controlling the formation of lymphatic vessels.

Background Although vascular endothelial growth factor (VEGF)-A/VEGF receptor 2 (KDR) signalling may play a major role in the microangiopathy of systemic sclerosis (SSc), serum levels of soluble KDR (sKDR) in this disease have not yet been determined. Objectives To evaluate the possibility that serum sKDR levels can be a specific disease marker of SSc. Methods Serum sKDR levels of 42 patients with SSc, 10 patients with Raynaud’s phenomenon (RP) and 22 healthy controls were measured with specific enzyme-linked immunosorbent assays. Quantitative real-time polymerase chain reaction (PCR) was performed to determine KDR mRNA levels. Results In females, the serum sKDR levels were significantly higher in patients with SSc, especially limited cutaneous SSc, than in patients with RP or healthy controls. Quantitative real-time PCR with RNA from skin sections revealed that KDR mRNA levels were also increased in the skin of patients with SSc with elevated serum sKDR levels. A significantly lower prevalence of pulmonary fibrosis, higher percentage vital capacity, and a higher incidence of telangiectasia were seen in female patients with SSc with elevated serum sKDR levels than those with normal levels. Conclusions These results suggest that the skin can be one of the sources of elevated serum sKDR levels, and that serum sKDR levels are useful for diagnosis and may be a marker of microangiopathy in patients with SSc, especially females. The VEGF-A/KDR signalling system may be involved in the pathogenesis of the disease.