Kazuyasu Fujii

Clinical characteristics associated with BRAF, NRAS and KIT mutations in Japanese melanoma patients

BACKGROUNDnThe importance of the genetic background of melanoma cells to the individual susceptibility to treatment has become apparent. In Caucasians, BRAF mutations are frequently detected in lesions on the skin of younger patients compared to NRAS and KIT mutations. However, clinical and pathological characteristics associated with BRAF, NRAS and KIT mutations have not been fully evaluated in East Asians.nnnOBJECTIVEnTo clarify clinical and pathological characteristics associated with BRAF, NRAS and KIT mutations in Japanese melanoma patients.nnnMETHODSnClinical data were retrospectively collected from 11 hospitals in Japan. BRAF, NRAS and KIT mutations were evaluated with polymerase chain reaction and Sanger sequencing. The relationships between these gene mutations and pathological and clinical findings were analyzed.nnnRESULTSnThe number of cases examined was 171 (primary: 135, metastases: 11, paired: 25), and all were Japanese patients. The detection rates of BRAF, NRAS and KIT mutations were 30.4%, 12.3% and 12.9%, respectively. Compared with the wild type, the presence of BRAF mutations was significantly associated with younger age (median, 50.0 years vs. 70.0 years, p<0.001). BRAF mutation was frequently detected in the lesions of the scalp (80%; 4/5), trunk (72.0%; 18/25), extremities (56.7%; 17/30) and neck (44.4%; 4/9), and the least prevalent were the face (22.2%; 2/9), nail (12.5%; 3/24), palm or sole (8.9%; 4/45) and mucosa (0%). NRAS mutations were prevalent in the face (33.3%) and palm or sole (20.0%), and the median age of these patients was 70.5 years. A KIT mutation was observed in the nail apparatus (25%), palm or sole (15.6%) and mucosa (18.2%). The median age of the patients with a KIT mutation was 63.0 years. Heterogeneity of mutations between primary and metastatic lesions was detected in six of 25 cases (24%). Solar elastosis was identified in 12 of 71 cases (15.3%), among which four cases harbored BRAF(V600E) (2 cases), BRAF(V600K), NRAS(Q61K) or NRAS(Q61L), respectively.nnnCONCLUSIONnSome clinical characteristics associated with BRAF, NRAS and KIT mutations were observed in Japanese patients, and we observed both similarities to and differences from those of Caucasians. Our findings could provide useful information in efforts to clarify the tumor genesis of malignant melanomas.

Romidepsin and azacitidine synergize in their epigenetic modulatory effects to induce apoptosis in CTCL

Purpose: Cutaneous T-cell lymphomas (CTCL) are a heterogeneous group of malignancies that despite available therapies commonly relapse. The emergence of combination epigenetic therapies in other hematologic malignancies have made investigation of such combinations in CTCL a priority. Here, we explore the synergistic antiproliferative effects of romidepsin, an HDAC inhibitor, and azacitidine, a demethylating agent, combination in CTCL. Experimental Design: The growth inhibition under combination treatment and single agent was explored by the MTT cell viability assay and the Annexin V/propidium iodide (PI) apoptosis assay in different CTCL cell lines and tumor cells derived from Sézary syndrome patients. Quantitative analysis of a dose–effect relationship of romidepsin and azacitidine was done by the CompuSyn software. Investigation of mechanism of action was performed by flow cytometry, immunoblotting, qRT-PCR arrays, and chromatin immunoprecipitation. Global CpG methylation sequencing was utilized to study genome methylation alteration under the treatment modalities. Results: The combination of romidepsin and azacitidine exerts synergistic antiproliferative effects and induction of apoptosis involving activation of the caspase cascade in CTCL cell lines and tumor cells derived from Sézary syndrome patients. We identified genes that were selectively induced by the combination treatment, such as the tumor suppressor gene RhoB that is linked to enhanced histone acetylation at its promoter region in parallel with pronounced expression of p21. Global CpG methylation sequencing in a CTCL cell line and tumor cells demonstrated a subset of genes with a unique change in methylation profile in the combination treatment. Conclusions: The synergistic antiproliferative effects of romidepsin and azacitidine combination treatment justify further exploration in clinical trials for advanced CTCL. Clin Cancer Res; 22(8); 2020–31. ©2015 AACR.

Infiltration of PD-1-positive cells in combination with tumor site PD-L1 expression is a positive prognostic factor in cutaneous angiosarcoma

ABSTRACT Cutaneous angiosarcoma (CAS) is a malignant sarcoma with poor prognosis. Programmed cell death-1 (PD-1)/programmed cell death-1 ligand-1 (PD-L1) expression reflects antitumor immunity, and is associated with patient prognosis in various cancers. The purpose of this study is to investigate the relationship between PD-1/PD-L1 expression and CAS prognosis. CAS cases (n = 106) were immunohistochemically studied for PD-L1 and PD-1 expression, and the correlation with patient prognosis was analyzed. PD-L1 expression was assessed by flow cytometry on three CAS cell lines with or without IFNγ stimulation. A total of 30.2% of patients samples were positive for PD-L1, and 17.9% showed a high infiltration of PD-1-positive cells. Univariate analysis showed a significant relationship between a high infiltration of PD-1-positive cells with tumor site PD-L1 expression and favorable survival in stage 1 patients (p = 0.014, log-rank test). Multivariable Cox-proportional hazard regression analysis also showed that patients with a high infiltration of PD-1-positive cells with tumor site PD-L1 expression were more likely to have favorable survival, after adjustment with possible confounders (hazard ratio (HR) = 0.38, p = 0.021, 95% confidence interval (CI) 0.16–0.86). Immunofluorescence staining of CAS samples revealed that PD-L1-positive cells were adjacent to PD-1-positive cells and/or tumor stroma with high IFNγ expression. In vitro stimulation with IFNγ increased PD-L1 expression in two out of three established CAS cell lines. Our results suggest that PD-1/PD-L1 expression is related to CAS progression, and the treatment with anti-PD-1 antibodies could be a new therapeutic option for CAS.

Clues to diagnosis for unusual mucosal pemphigus demonstrating undetectable anti‐desmoglein 3 serum antibodies by routine tests

Pemphigus is an autoimmune blistering disease caused by immunoglobulin (Ig)G autoantibodies against desmogleins (Dsg). In mucosal‐dominant pemphigus vulgaris (PV), anti‐Dsg3 antibodies play a critical role in acantholysis. We followed two mucosal‐dominant PV cases who suffered from refractory oral mucosal erosions. In these cases, anti‐Dsg3 serum antibodies were not detected by indirect immunofluorescence and enzyme‐linked immunosorbent assay (ELISA). However, direct immunofluorescence showed the intercellular IgG deposition in the epidermis and histopathological findings revealed suprabasal acantholysis. In order to analyze the pathomechanisms in these cases, we first examined the Dsg3 expression patterns in lesional sites and compared them with those of typical mucosal‐dominant PV cases. In typical PV cases, the alteration of Dsg3 distribution was observed in lesional sites by immunostaining. The aggregation of Dsg3, which is the characteristic change in PV mucosal lesions, was observed as the initial change prior to acantholysis. In our cases, a clustering of Dsg3 was observed at mucosal lesions, and the expression levels of Dsg3 in acantholytic lesions were decreased, as observed in typical mucosal‐dominant PV cases. Although anti‐Dsg3 serum antibodies could not be detected by routine tests, anti‐Dsg3 serum antibodies were detected by Dsg3 ELISA using 10‐times more concentrated sera (highly sensitive ELISA). Moreover, purified and concentrated PV IgG showed high pathogenicity when examined by dissociation assay. In conclusion, the detection of morphological changes in Dsg3 distribution and highly sensitive ELISA method could be useful for the early diagnosis of PV recurrence.

Lymphomatoid granulomatosis initially presenting as ulcerated subcutaneous and muscle lesions without pulmonary involvement

Dear Editor, Lymphomatoid granulomatosis (LYG) is an angiocentric and angiodestructive lymphoproliferative disorder. It is comprised of Epstein–Barr virus (EBV)-positive B cells admixed with reactive T cells that tend to predominate. While LYG most commonly affects the lung, it can involve multiple extranodal sites. We report a patient who developed LYG after methotrexate (MTX) therapy for rheumatoid arthritis (RA). A 79-year-old man presented with an ulcerated subcutaneous induration in the left lumbar region that had appeared 10 months earlier (Fig. 1a). His RA had been treated with MTX for 9 years.

Rare case of Langerhans cell sarcoma with cutaneous manifestation arising on the inguinal region.

carcinoma-like features in a nodule. The fact that nodules with solid carcinoma-like histopathological features had appeared from an erythematous plaque leads us to hypothesize that the tumor cells differentiated into two directions of either tubular structures in an erythematous plaque or solid carcinoma-like features in a nodule. In accordance with this hypothesis, immunohistochemical staining shared a positive reaction with EMA in tumor cells of both syringomatous carcinoma and solid carcinoma-like features. The size of this tumor (24 cm 9 15 cm) is quite large compared with previous cases, and the location is relatively rare since most cases of syringomatous carcinoma occur on the face, axillae and scalp. In conclusion, we report a case that supports the concept of syringomatous carcinomas having several different histological expression patterns.

Spontaneous regression of a primary cutaneous diffuse large B-cell lymphoma, leg type.

1 Harding JJ, Pulitzer M, Chapman PB. Vemurafenib sensitivity skin reaction after ipilimumab. N Engl J Med 2012; 366: 866–868. 2 Imafuku K, Yoshino K, Ishiwata K et al. Severe rash associated with vemurafenib administration following nivolumab therapy. J Eur Acad Dermatol Venereol 2016; 30: e84–e86. 3 Minor DR, Rodvien R, Kashani-Sabet M. Successful desensitization in a case of Stevens-Johnson syndrome due to vemurafenib. Melanoma Res 2012; 22: 410–411. 4 Johnson DB, Wallender EK, Cohen DN et al. Severe cutaneous and neurologic toxicity in melanoma patients during vemurafenib administration following anti-PD-1 therapy. Cancer Immunol Res 2013; 1: 373–377.

Long-lasting Localized Pemphigus Vulgaris without Detectable Serum Autoantibodies Against Desmoglein 3 and Desmoglein 1.

Pemphigus vulgaris (PV) is an autoimmune blistering disease elicited by anti-desmoglein (DsG) 3 antibody. Although skin lesions tend to be distributed over the entire body, in some patients, they are confined to a restricted area. We report two patients who presented with long-lasting localized PV without detectable anti-DsG antibodies after suffering antibody-positive systemic PV. Initial treatment with prednisolone (PSL) was successful in both patients, but a local relapse occurred on the cheek or lower lip after a reduction in the PSL dose. Biopsy of the localized lesions showed suprabasal acantholysis; no serum DsG antibodies were found. Local immunosuppression therapy was effective in both patients. Based on our findings, we suggest that localized PV without detectable antibodies can develop after systemic PV.

Ethosuximide-induced Stevens-Johnson syndrome: Beneficial effect of early intervention with high-dose corticosteroid therapy

We report two rare cases of childhood epilepsy patients who developed ethosuximide‐induced Stevens–Johnson syndrome (SJS). Unlike typical SJS, the initial eruption of both patients presented well‐demarcated, infiltrating firm papules mainly on the cheeks and the extensor aspects of the arms (case 1), and multiple vesicles on the soles and oral aphthosis (case 2), which closely mimicked viral exanthema. We diagnosed both patients with ethosuximide‐induced SJS, based on the dosing period and the positive results of drug‐induced lymphocyte stimulation test. Systemic corticosteroids are usually selected as a standard therapy for SJS, despite controversial results regarding their effectiveness. In case 1, an i.v. pulse therapy of methylprednisolone (30 mg/kg, 3 days consecutively) was initiated on day 7 from the onset of illness, and an i.v. immunoglobulin (400 mg/kg, 5 days consecutively) was added the following day. In case 2, an i.v. prednisone treatment (1 mg/kg, for 1 week) was initiated on day 4 from the onset. Eventually, the early therapeutic interventions resulted in good outcomes in both patients.

Hypohidrotic ectodermal dysplasia with strabismus

Dear Editor, We encountered a patient with hypohidrotic ectodermal dysplasia (HED) with strabismus. A 26-year-old man with a history of bronchial asthma and atopic dermatitis was admitted for the evaluation of congenital heat retention. His brother manifested the same symptoms and died at the age of 4 months; his two sisters had no such history. His scalp hair and eyebrows were sparse (Fig. 1a), and axillary and pubic hair were absent. Frontal bossing, saddlenose, thick lips, strabismus, hypodontia, dry skin, but no nail deformities were observed (Figs. 1a–c). Abdominal skin biopsy showed sweat gland hypoplasia. A 15-min sweat test (41°C, 22% humidity) elicited no sweating except on his palms; his body temperature rose by 1°C. Topical injection of 5% acetylcholine chloride at the anhidrotic area elicited no sweating. Genetic analysis using peripheral blood cells revealed the deletion of 36 bases in exon 4 of the ectodysplasin A (EDA) gene (c.646_681del), resulting in the loss of 12 amino acids from the 216th proline to the 227th glycine (Fig. 1d). Because the mutation is an in-frame deletion, it can be predicted that the mutant transcript is stably expressed in the patient’s skin. Ectodermal dysplasias (ED), characterized by disorders of various organs including the skin, is attributable to the abnormal development of the primordial external germ layer. It is roughly divided into HED and hidrotic ED depending on the presence or absence of sweat gland hypoplasia. Typical clinical manifestations of HED include hypotrichosis with fine and slow-growing scalp and body hair, sparse eyebrows, hypohidrosis, nail anomalies, hypodontia, frontal bossing, saddle-nose and thick lips. In some patients, the body temperature is as high as 42°C. Atopic dermatitis and bronchial asthma are frequent complications of HED. Besides typical findings, our patient manifested strabismus which is often seen in hidrotic ED. We found no HED patient with strabismus in the published work. Approximately 200 ED subtypes have been reported; the implicated gene has been studied in 30. X-linked HED, its most common form, accounted for approximately 70% of HED. It is due to mutations in the ED1 gene encoding EDA. EDA plays