USP6 rearrangement in pediatric nodular fasciitis.

Nodular fasciitis (NF) is a myofibroblastic proliferation that is uncommonly present in pediatric patients. These benign neoplasms can masquerade as more insidious sarcomatous proliferations on both clinical exam and initial histopathologic review, often prompting undue concern in patients, parents, and providers. While immunohistochemical analysis of NF can be variable, adding to the diagnostic uncertainty, molecular analysis documenting ubiquitin-specific protease 6 (USP6) gene rearrangement can help confirm the diagnosis as an association between NF and USP6 overexpression was first identified 10 years ago in an analysis that found rearrangements of the involved locus in over 90% of studied samples. In this report, we review one case of NF located on the chin of a nine-year-old girl in which molecular testing was essential to secure the correct diagnosis, and provide a summary of documented cases of USP6 overexpression in transient pediatric neoplasms.

Local immune cell infiltration in cutaneous acute graft versus host disease.

BACKGROUND: Hematopoietic stem cell transplant is a crucial intervention to definitively treat many hematopoietic malignancies, but it carries great risks of morbidity and mortality often associated with graft-versus-host disease (GVHD). Acute and chronic GVHD are distinct entities, defined by a combination of historical, clinical, and pathologic data, but both are generally thought to stem from self-propagating aberrantly activated immune cells inflicting end organ damage, with the potential to cause significant illness or even death. Event-free survival rates after hematopoietic stem cell transplant continue to improve each year, but GVHD remains a major hurdle in improving the efficacy and safety of transplant. OBJECTIVE: Recent studies demonstrating tissue-specific immune effector phenotypes underscore the need for a deeper understanding of the cellular and molecular pathways driving the destruction of target tissues in patients with acute GVHD. METHODS: Samples were collected from lesional and unaffected skin in five patients with acute cutaneous GHVD. Fresh tissue was processed for fluorescence-activated cell sorting and analysis of macrophages and lymphocytes. RESULTS: The percentage of lymphocytes and macrophages as a representation of total cells varied among patients and was not always consistent between lesional and unaffected sites. The heterogeneity in immune cell profiling observed in patients in this study could reflect the diverse demographics, conditioning, and transplant conditions of each individual. CONCLUSION: This study provides initial insight into the underlying molecular mechanisms of cutaneous GVHD progression and paves the way for additional studies to examine the cellular and molecular landscape in greater detail.

An Unusual Case of Morphea in the Setting of Aplastic Anemia.

Cutaneous sclerosis occurs in association with a variety of systemic diseases, including hematologic malignancy, plasma cell dyscrasias, solid organ tumors, and other systemic autoimmune conditions. Herein, we present a unique case of morphea/lichen sclerosus overlap arising in association with aplastic anemia. To expand upon this rare case, we also review the literature surrounding paraneoplastic sclerosing skin disorders. A 53-year-old man presented with a 13-month history of progressive and generalized skin changes. Exam revealed irregular, hypopigmented indurated plaques with focal areas of scale on the bilateral axillae and hips, as well as hyperpigmented brown papules and plaques on the back. Laboratory evaluation revealed pancytopenia and positive anti-nuclear antibody (1:160). Bone marrow biopsy demonstrated hypocellular marrow consistent with aplastic anemia. Furthermore, skin biopsies revealed lichen sclerosus overlying superficial morphea, consistent with a paraneoplastic sclerodermoid-like eruption. While preparations for hematologic-directed therapies were made, skin-directed therapy with a combination topical steroids and topical calcineurin inhibitors was initiated. Eosinophilic fasciitis and scleroderma have been linked to aplastic anemia, and herein, we expand upon this phenomenon by presenting our case of generalized plaque morphea/lichen sclerosus overlap arising in the setting of aplastic anemia. Dermatologists must be aware of this rare association in order to identify precocious hematologic disease.

SOX2 is a cancer-specific regulator of tumour initiating potential in cutaneous squamous cell carcinoma.

Although the principles that balance stem cell self-renewal and differentiation in normal tissue homeostasis are beginning to emerge, it is still unclear whether cancer cells with tumour initiating potential are similarly governed, or whether they have acquired distinct mechanisms to sustain self-renewal and long-term tumour growth. Here we show that the transcription factor Sox2, which is not expressed in normal skin epithelium and is dispensable for epidermal homeostasis, marks tumour initiating cells (TICs) in cutaneous squamous cell carcinomas (SCCs). We demonstrate that Sox2 is required for SCC growth in mouse and human, where it enhances Nrp1/Vegf signalling to promote the expansion of TICs along the tumour-stroma interface. Our findings suggest that distinct transcriptional programmes govern self-renewal and long-term growth of TICs and normal skin epithelial stem and progenitor cells. These programmes present promising diagnostic markers and targets for cancer-specific therapies.

Adult stem cell niches: cellular and molecular components.

As stem cells (SCs) in adult organs continue to be identified and characterized, it becomes clear that their survival, quiescence, and activation depend on specific signals in their microenvironment, or niche. Although adult SCs of diverse tissues differ by their developmental origin, cycling activity, and regenerative capacity, there appear to be conserved similarities regarding the cellular and molecular components of the SC niche. Interestingly, many organs house both slow-cycling and fast-cycling SC populations, which rely on the coexistence of quiescent and inductive niches for proper regulation. In this review we present a general definition of adult SC niches in the most studied mammalian systems. We further focus on dissecting their cellular organization and on highlighting recently identified key molecular regulators. Finally, we detail the potential involvement of the SC niche in tissue degeneration, with a particular emphasis on aging and cancer.

Tbx18 targets dermal condensates for labeling, isolation, and gene ablation during embryonic hair follicle formation.

How cell fate decisions of stem and progenitor cells are regulated by their microenvironment or niche is a central question in stem cell and regenerative biology. Although functional analysis of hair follicle epithelial stem cells by gene targeting is well established, the molecular and genetic characterization of the dermal counterpart during embryonic morphogenesis has been lacking because of the absence of cell type-specific drivers. Here, we report that T-box transcription factor Tbx18 specifically marks dermal papilla (DP) precursor cells during embryonic hair follicle morphogenesis. With Tbx18(LacZ), Tbx18(H2BGFP), and Tbx18(Cre) knock-in mouse models, we demonstrate LacZ and H2BGFP (nuclear green fluorescent protein) expression and Cre activity in dermal condensates of nascent first-wave hair follicles at E14.5. As Tbx18 expression becomes more widespread throughout the dermis at later developmental stages, we use tamoxifen-inducible Cre-expressing mice, Tbx18(MerCreMer), to exclusively target DP precursor cells and their progeny. Finally, we ablate Tbx18 in full knockout mice, but find no perturbations in hair follicle formation, suggesting that Tbx18 is dispensable for normal DP function. In summary, our study establishes Tbx18 as a genetic driver to target for the first time embryonic DP precursors for labeling, isolation, and gene ablation that will greatly enhance investigations into their molecular functions during hair follicle morphogenesis.

RNAi screen of the protein kinome identifies checkpoint kinase 1 (CHK1) as a therapeutic target in neuroblastoma.

Neuroblastoma is a childhood cancer that is often fatal despite intense multimodality therapy. In an effort to identify therapeutic targets for this disease, we performed a comprehensive loss-of-function screen of the protein kinome. Thirty kinases showed significant cellular cytotoxicity when depleted, with loss of the cell cycle checkpoint kinase 1 (CHK1/CHEK1) being the most potent. CHK1 mRNA expression was higher in MYC-Neuroblastoma-related (MYCN)-amplified (P < 0.0001) and high-risk (P = 0.03) tumors. Western blotting revealed that CHK1 was constitutively phosphorylated at the ataxia telangiectasia response kinase target site Ser345 and the autophosphorylation site Ser296 in neuroblastoma cell lines. This pattern was also seen in six of eight high-risk primary tumors but not in control nonneuroblastoma cell lines or in seven of eight low-risk primary tumors. Neuroblastoma cells were sensitive to the two CHK1 inhibitors SB21807 and TCS2312, with median IC(50) values of 564 nM and 548 nM, respectively. In contrast, the control lines had high micromolar IC(50) values, indicating a strong correlation between CHK1 phosphorylation and CHK1 inhibitor sensitivity (P = 0.0004). Furthermore, cell cycle analysis revealed that CHK1 inhibition in neuroblastoma cells caused apoptosis during S-phase, consistent with its role in replication fork progression. CHK1 inhibitor sensitivity correlated with total MYC(N) protein levels, and inducing MYCN in retinal pigmented epithelial cells resulted in CHK1 phosphorylation, which caused growth inhibition when inhibited. These data show the power of a functional RNAi screen to identify tractable therapeutical targets in neuroblastoma and support CHK1 inhibition strategies in this disease.

Identification of ALK as a major familial neuroblastoma predisposition gene.

Neuroblastoma is a childhood cancer that can be inherited, but the genetic aetiology is largely unknown. Here we show that germline mutations in the anaplastic lymphoma kinase (ALK) gene explain most hereditary neuroblastomas, and that activating mutations can also be somatically acquired. We first identified a significant linkage signal at chromosome bands 2p23-24 using a whole-genome scan in neuroblastoma pedigrees. Resequencing of regional candidate genes identified three separate germline missense mutations in the tyrosine kinase domain of ALK that segregated with the disease in eight separate families. Resequencing in 194 high-risk neuroblastoma samples showed somatically acquired mutations in the tyrosine kinase domain in 12.4% of samples. Nine of the ten mutations map to critical regions of the kinase domain and were predicted, with high probability, to be oncogenic drivers. Mutations resulted in constitutive phosphorylation, and targeted knockdown of ALK messenger RNA resulted in profound inhibition of growth in all cell lines harbouring mutant or amplified ALK, as well as in two out of six wild-type cell lines for ALK. Our results demonstrate that heritable mutations of ALK are the main cause of familial neuroblastoma, and that germline or acquired activation of this cell-surface kinase is a tractable therapeutic target for this lethal paediatric malignancy.