Yujiro Takae

Antigen-independent development of Foxp3+ regulatory T cells suppressing autoantibody production in experimental pemphigus vulgaris

The CD4(+)CD25(+)Foxp3(+) regulatory T cells (Tregs) play suppressive roles in various types of autoimmunity. It has been reported that Tregs develop in the thymus after high-affinity interaction of their TCR with self-peptide/MHC ligands mostly utilizing TCR-transgenic system. In this study, we examined whether the specific antigen is involved in the development of polyclonal Tregs in pemphigus vulgaris (PV), an autoimmune blistering disease caused by anti-desmoglein 3 (Dsg3) IgG antibodies, as a model system. Adoptive transfer of splenocytes of Dsg3(-)(/-) mice immunized with recombinant mouse Dsg3 to Rag2(-)(/-) recipient mice expressing Dsg3 resulted in the stable production of anti-Dsg3 IgG and the development of PV phenotypes. We show here that Tregs control anti-Dsg3 antibody production in PV model mice: the adoptive transfer of Tregs and the depletion of endogenous Tregs suppressed and augmented, respectively, the anti-Dsg3 antibody production. To examine whether the endogenous expression of Dsg3 is involved in the generation of these PV-relevant Tregs, we compared the potential of wild-type Tregs with that of Tregs from Dsg3(-)(/-) mice. Polyclonal Tregs from Dsg3(-)(/-) mice were more potent than that of wild-type mice, in both adoptive transfer and Treg-depletion experiments, while suppressive activities against IgG production against an irrelevant antigen were similar between Tregs from wild-type and Dsg3(-)(/-) mice. Our observation implies that Tregs capable of suppressing T(h) cells that drive autoantibody production can develop in the absence of the target antigen.

Pemphigus mouse model as a tool to evaluate various immunosuppressive therapies.

Abstract:  Pemphigus vulgaris (PV) is an autoimmune bullous disease caused by immunoglobulin G (IgG) autoantibodies against desmoglein 3 (Dsg3). We have generated an active disease mouse model for PV by adoptive transfer of Dsg3−/− lymphocytes. In this study, we investigated the benefits and limitations of this model as a tool to evaluate various immunosuppressive therapeutic strategies. We used the following three measurements to evaluate the effects of the drugs during the time course: Dsg3 enzyme‐linked immunosorbent assay scores that represent the level of production of anti‐Dsg3 IgG, body weight loss that reflects the severity of oral erosions and PV score that reflects the extent of skin lesions. We examined various immunosuppressive agents currently used to treat patients with PV model mice in preventive protocol. Cyclophosphamide almost completely suppressed the production of anti‐Dsg3 IgG, development of body weight loss and the appearance of the PV phenotype in contrast with the control group without the drug. Azathioprine, cyclosporin A and tacrolimus hydrate also showed suppressive effects to various degrees. However, methylprednisolone and dexamethasone failed to show significant effects in contrast to the findings reported in humans. Knowing the advantages and limitations of this model will provide an important foundation for the future evaluation and development of novel therapeutic strategies.

A case of atypical POEMS syndrome without polyneuropathy

POEMS (Polyneuropathy, Organomegaly, Endocrinopathy, M‐protein, Skin changes) syndrome is a rare hematological disease associated with overproduction of pro‐inflammatory cytokines. Under the current nomenclature and diagnostic criteria for POEMS syndrome, the presence of characteristic polyneuropathy is required for diagnosis. We report a 43‐year‐old Japanese woman with organomegaly, endocrinopathy, M‐protein, skin lesions, as well as typical renal lesions and sclerotic bone lesions. Of note, neurological examinations and peripheral nerve conduction tests were normal in this patient. In view of the overwhelming number of otherwise characteristic signs and symptoms, we made a provisional diagnosis of ‘atypical POEMS syndrome without polyneuropathy’. If further similar cases are reported in the future, reconsideration of the nomenclature and/or diagnostic criteria for POEMS syndrome may be required.

Collapse of the keratin filament network through the expression of mutant keratin 6c observed in a case of focal plantar keratoderma

Focal palmoplantar keratoderma (PPK) with severe pain is a hallmark of pachyonychia congenita, a rare autosomal dominant disorder involving PPK and hypertrophic nail dystrophy. Some families present focal PPK with either minimal or no nail changes. Dominant‐negative mutations in any of the four identified keratin genes, KRT6A, KRT6B, KRT16 or KRT17, lead to pachyonychia congenita. However, the majority of families with focal PPK showing minimal or no nail changes do not harbor mutations in these genes. Recently, mutations of KRT6C were identified in families with focal PPK alone. Here, we report a 26‐year‐old Japanese man with focal plantar hyperkeratosis that developed at approximately 10 years of age with no palmar involvement and no nail alterations. We identified a missense KRT6C mutation c.1414G>A resulting in an p.Glu472Lys substitution, as reported in other Japanese patients. When the mutant keratin 6c protein is exogenously expressed in human HaCaT cells, a collapse of the keratin filament network is observed in a dose‐dependent manner, suggesting the mutation has a dominant‐negative effect on keratin filament network formation. The mutated residue is located at the helix termination motif of keratin 6c. The peptide sequence around this residue is highly conserved among type II, III and IV intermediate filament proteins. Glu to Lys mutations of the equivalent residue have been reported in a variety of inherited diseases, including neurodegenerative diseases, corneal dystrophy and skin disorders, suggesting that this residue is vital to keratin function.

Demodicidosis as a cause of facial eruption developing early after allogeneic hematopoietic stem cell transplantation

Graft-versus-host disease (GVHD) is the most common and clinically important cause of skin eruption after allogeneic hematopoietic stem cell transplantation (HSCT). Other causes include viral infection such as human herpesvirus 6, allergic reactions to drugs, and the toxicity of the conditioning regimens. Despite the rarity, infectious microorganisms may also cause this complication. We report our experience as regards two patients developing skin eruption limited to the face early after allogeneic HSCT, which was diagnosed as demodicidosis. A 53-year-old woman with acute lymphoblastic leukemia (Patient 1) and a 44-year-old woman with chronic myelogenous leukemia (Patient 2) underwent bone marrow transplantation from an unrelated donor. Tacrolimus and short-term methotrexate were used as a prophylaxis for GVHD. Their post-transplant courses were complicated with acute skin GVHD, which was successfully treated with a topical steroid. Around day 110 post-transplant, both patients developed an itchy eruption limited to the forehead, cheeks, and jaw (Fig. 1). The eruptions were characterized by multiple papules and pustules with erythema. Microscopic examination found numerous Demodex mites in the pustules (Fig. 2), and the diagnosis of demodicidosis was made. Histological examination was not performed. Eruptions promptly resolved with topical sulfur, and completely resolved within 3–4 weeks. No recurrence was observed after the cessation of the treatment in Patient 2. However, 2 weeks after the cessation of the treatment, the demodicidosis recurred in Patient 1, and the same therapy was initiated, which was again effective. An additional 1-month treatment resulted in the complete resolution of the skin eruption without further recurrence. Demodex mites inhabit human pilosebaceous ducts and are frequently found in healthy populations. Because of the distribution of pilosebaceous ducts, Demodex are exclusively found in the human face. Demodicidosis associated with immunocompromised hosts has been reported in patients with HIV infection and lymphoid malignancies, and in a recipient of allogeneic HSCT [1–7]. Thus, although still controversial, these reports strongly suggest that immunodeficiency may significantly contribute to its pathogenicity. The common feature of these reported immunocompromised settings is long-term impairment of cell-mediated immunity. Although there has only been one reported case of demodicidosis after allogeneic HSCT, long-term impairment of cell-mediated immunity could contribute to the susceptibility of recipients of HSCT and organ transplantation to demodicidosis.

Rapidly developed BRAF inhibitor-induced verrucous keratosis

Dear Editor, BRAF inhibitors (BRAFi) are utilized for the treatment of various malignancies, including melanoma. These agents are associated with the development of squamoproliferative lesions such as squamous cell carcinomas (SCC), keratoacanthomas and BRAFi-induced verrucous keratosis (BIVK). Compared with Caucasians, these lesions rarely occur in Asian subjects. Here, we report the first case of BIVK in an Asian subject, which developed rapidly with unique initial lesion, HRAS mutation and cyclin-dependent kinase inhibitor 2A (CDKN2A) amplification. A 74-year-old Japanese woman presented with melanoma that had originated in the sacral region skin, with BRAF V600E mutation and multiple in-transit metastases; her disease progressed through treatment with a clinical trial of combination BRAFi and mitogen-activated protein kinase kinase inhibitor (MEKi). Six months after the clinical trial started, a new in-transit metastasis occurred and she was withdrawn. The lesion was surgically removed and 1 month after discontinuation of the drug, vemurafenib monotherapy was started. After 4 days, she developed a 25 mm 9 10 mm pearly reddish plaque on the outside of the lower lip (Fig. 1a) with skin rash and milia. Prednisolone 10 mg/day was administrated for rash, the plaque did not shrink but changed like a verruca (Fig. 1b). Histopathologically, the lesion showed acanthosis with papillomatosis and hypergranulosis without koilocytes or keratinocyte atypia (Fig. 1c,d). The immunohistochemical (IHC) staining pattern of p53 showed so-called wild-type pattern, interestingly activated Notch 1 (Fig. 1e) and Ki-67 (Fig. 1f) positive from the epidermal basal cell layer. A negative finding of p16 IHC staining and human papillomavirus (HPV) in situ hybridization denied the involvement of HPV infection. By using an original gene panel, “NCC oncopanel v3”, the targeted gene sequencing analysis revealed that our case had HRAS mutation (G12C) and CDKN2A amplification. As for the development of squamoproliferative lesions associated with BRAF inhibitors, paradoxical mitogenactivated protein kinase activation was thought to be mediated. Among them, HRAS mutations were reported as a most predominant in BIVK (90% of lesions). In our case, BIVK occurred only after use of BRAFi monotherapy, but not after MEKi combination. The process supported the hypothesis. On the other hand, CDKN2A gene amplification has not been reported in BIVK but has nevertheless been reported in SCC. In our case, the amplification of CDKN2A could be associated with rapid occurrence and unique initial appearance. Physicians need to pay attention for the development of BIVK even when the patients are Asian.

Development of varicella after allogeneic cord blood transplantation in a varicella zoster virus seropositive patient

Dear Sir Cord blood (CB) has been established as 1 of the stem cell sources for both adult and pediatric patients requiring allogeneic hematopoietic stem cell transplantation (HSCT) [1,2]. The most common life-threatening complication after CB transplantation (CBT) is infection, mostly bacterial infection. The high incidence of bacterial infection after CBT can be explained by the delayed neutrophil engraftment particularly in adult patients [3,4]. Furthermore, several investigators and we have reported a notably high incidence of viral infection after CBT as well as cytomegalovirus and human herpes virus type-6 [5,6]. A high incidence of viral infection is considered attributable to the absence of antigen-specific T-lymphocytes in cord blood [5 7]. Because of the exclusive component of naive Tlymphocytes in cord blood, a primary infection of varicella zoster virus (VZV) could occur more frequently after CBT than HSCT using other stem cell sources. However, most of the reported infection of VZV is its reactivation causing herpes zoster, as in other types of HSCT [8]. These observations could be explained by the recipient’s residual humoral immunity or the transferred immunity from the donor. Specific IgG is generally used as a useful parameter of the host’s defense against primary VZV infection, varicella, even in HSCT recipients [9]. Therefore, the recipients of HSCT at risk for developing varicella are identified according to their VZV serostatus [9]. In contrast to this general practice, we have experienced a patient who developed varicella after CBT despite the fact that the patient possessed a high titer of anti-VZV IgG. A 33-y-old male with acute myelogenous leukemia (AML) underwent CBT from a human leukocyte antigen (HLA)-mismatched unrelated donor. The conditioning regimen consisted of total body irradiation (12 Gy), cytarabine (8 g/m), and cyclophosphamide (120 mg/kg). Before transplantation, he was positive for anti-VZV IgG, with an index of 43.0 by enzyme-linked immunosorbent assay (ELISA; positive index ]2.0). Cyclosporine A (CsA) and short-term methotrexate were used as a prophylaxis for graft-versus-host disease (GVHD). His posttransplant course was complicated with acute GVHD involving the skin and gastrointestinal tract, which was successfully treated by adding prednisolone to CsA. He did not develop chronic GVHD, and CsA was discontinued 7 months after CBT. One y after CBT, a relapse of AML was observed, and 2 courses of high-dose cytarabine were given. 28 d after the second course of high-dose cytarabine, multiple vesicular skin eruptions appeared over the upper and lower extremities, and over the trunk and head without pain or paresthesia. A Tzanck smear from the vesicle showed multinucleated giant cells, the sign of herpes virus infection. Based on these findings, varicella was diagnosed and intravenous acyclovir was given for 14 d in combination with intravenous immunoglobulin. All the eruptions had dried out by the completion of this treatment. At the