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Histiocytic disorders of childhood represent a wide spectrum of conditions that share the common histologic feature of activated or transformed “histiocytes.” Langerhans cell histiocytosis (LCH) is the most common, with an incidence of approximately 5 per million children. LCH may be difficult to distinguish from more ubiquitous causes of skin rashes, bone pain, or fever. Current chemotherapy fails to cure more than 50% of children with multifocal disease, and treatment failure is associated with increased risks of long-term sequelae. Somatic activating mitogen-activated protein kinase (MAPK) pathway–activating mutations (most often BRAFV600E) have been identified in hematopoietic precursors in patients with LCH. Opportunities to improve outcomes with targeted therapies are under investigation. Juvenile xanthogranuloma (JXG) and Rosai-Dorfman disease (RDD) are less common than LCH and are distinguished by specific histologic and clinical features. Recurrent MAPK pathway gene mutations are also identified in JXG and RDD. In many cases, these conditions spontaneously resolve, but disseminated disease can be fatal. Although there has been historic debate regarding the nature of these conditions as inflammatory versus neoplastic, LCH, JXG, and RDD are now considered myeloid neoplastic disorders. In contrast, hemophagocytic lymphohistiocytosis (HLH) is clearly a disorder of immune dysregulation. HLH is characterized by extreme immune activation driven by hyperactivated T cells. HLH arises in approximately 1 child per million and is nearly universally fatal without prompt recognition and immune suppression. Outcomes of treated children are poor, with approximately 60% survival. Emapalumab, which targets interferon-γ signaling, was recently approved for patients with recurrent or refractory HLH, and additional cytokine-directed therapies are under investigation.

Four male infants with cystic fibrosis and prolonged neonatal jaundice underwent Kasai procedure to relieve biliary obstruction due to apparent biliary atresia. The excised remnants had viscid mucus accumulation in hypoplastic gallbladders and distended peribiliary glands. Main hepatic ducts were narrow and/or malformed. Microscopic differences between the gallbladder and extrahepatic bile ducts in cystic fibrosis and sporadic biliary atresia were unequivocal, despite some histologic overlap; no erosive or fibro-obliterative lesions typical of biliary atresia were seen. Common in liver, biopsies were small duct cholangiopathy with intense focal cholangiolitis and massive accumulation of ceroid pigment within damaged cholangiocytes, and in portal macrophages, portal fibrosis, and unequivocal features of large duct obstruction were inconspicuous compared with biliary atresia. Plugs of bile in small ducts tended to be pale and strongly periodic acid-Schiff-reactive in cystic fibrosis. Distinguishing the liver lesion from that of biliary atresia is challenging but possible. Liver biopsies from 2 additional infants with cystic fibrosis and prolonged jaundice that spontaneously resolved showed a similar small duct cholangiopathy. Small gallbladders and extrahepatic ducts challenge surgical judgment as findings in liver biopsies challenge the pathologist. The decision to perform a Kasai procedure is reasonable when mimicry of biliary atresia is grossly complete. We hypothesize that a disorder of bile volume/flow during development and/or early infancy linked to the CFTR mutation alone or in combination with the stresses of neonatal intensive care causes destructive cholangiolitis and intrahepatic reduction of bile flow with secondary hypoplasia of extrahepatic biliary structures.

Most children with high-risk Langerhans cell histiocytosis (LCH) have BRAFV600E mutation. BRAFV600E alleles are detectable in myeloid mononuclear cells at diagnosis but it is not known if the cellular distribution of mutation evolves over time. Here, the profiles of 16 patients with high-risk disease were analyzed. Two received conventional salvage chemotherapy, 4 patients on inhibitors were tracked at intervals of 3 to 6 years, and 10 patients, also given inhibitors, were analyzed more than 2 years after diagnosis. In contrast to the patients responding to salvage chemotherapy who completely cleared BRAFV600E within 6 months, children who received inhibitors maintained high BRAFV600E alleles in their blood. At diagnosis, mutation was detected predominantly in monocytes and myeloid dendritic cells. With time, mutation switched to the T-cell compartment, which accounted for most of the mutational burden in peripheral blood mononuclear cells, more than 2 years from diagnosis (median, 85.4%; range, 44.5%-100%). The highest level of mutation occurred in naïve CD4+ T cells (median, 51.2%; range, 3.8%-93.5%). This study reveals an unexpected lineage switch of BRAFV600E mutation in high-risk LCH, which may influence monitoring strategies for the potential withdrawal of inhibitor treatment and has new implications for the pathogenesis of neurodegeneration, which occurred in 4 patients.

Langerhans cell histiocytosis (LCH) is a potentially fatal condition characterized by granulomatous lesions with characteristic clonal mononuclear phagocytes (MNPs) harboring activating somatic mutations in mitogen-activated protein kinase (MAPK) pathway genes, most notably BRAFV600E. We recently discovered that the BRAFV600E mutation can also affect multipotent hematopoietic progenitor cells (HPCs) in multisystem LCH disease. How the BRAFV600E mutation in HPCs leads to LCH is not known. Here we show that enforced expression of the BRAFV600E mutation in early mouse and human multipotent HPCs induced a senescence program that led to HPC growth arrest, apoptosis resistance and a senescence-associated secretory phenotype (SASP). SASP, in turn, promoted HPC skewing toward the MNP lineage, leading to the accumulation of senescent MNPs in tissue and the formation of LCH lesions. Accordingly, elimination of senescent cells using INK-ATTAC transgenic mice, as well as pharmacologic blockade of SASP, improved LCH disease in mice. These results identify senescent cells as a new target for the treatment of LCH.

The upcoming 5th edition of the World Health Organization (WHO) Classification of Haematolymphoid Tumours is part of an effort to hierarchically catalogue human cancers arising in various organ systems within a single relational database. This paper summarizes the new WHO classification scheme for myeloid and histiocytic/dendritic neoplasms and provides an overview of the principles and rationale underpinning changes from the prior edition. The definition and diagnosis of disease types continues to be based on multiple clinicopathologic parameters, but with refinement of diagnostic criteria and emphasis on therapeutically and/or prognostically actionable biomarkers. While a genetic basis for defining diseases is sought where possible, the classification strives to keep practical worldwide applicability in perspective. The result is an enhanced, contemporary, evidence-based classification of myeloid and histiocytic/dendritic neoplasms, rooted in molecular biology and an organizational structure that permits future scalability as new discoveries continue to inexorably inform future editions.

Becker nevus (BN) is a benign cutaneous smooth muscle hamartoma that presents with a hyperpigmented patch or plaque with or without hypertrichosis.1 BN may be associated with ipsilateral breast hypoplasia or other musculoskeletal abnormalities, an association which has been termed Becker nevus syndrome (BNS).

Key Points Juvenile xanthogranuloma (JXG) usually presents with lesions isolated to the skin; however, aggressive, disseminated forms also occur. Identification of a novel MRC1-PDGFRB fusion in a child with JXG guided targeted therapy with dasatinib, leading to a dramatic response.

Erdheim-Chester disease (ECD) is a rare histiocytosis that was recently recognized as a neoplastic disorder owing to the discovery of recurrent activating MAPK (RAS-RAF-MEK-ERK) pathway mutations. Typical findings of ECD include central diabetes insipidus, restrictive pericarditis, perinephric fibrosis, and sclerotic bone lesions. The histopathologic diagnosis of ECD is often challenging due to nonspecific inflammatory and fibrotic findings on histopathologic review of tissue specimens. Additionally, the association of ECD with unusual tissue tropism and an insidious onset often results in diagnostic errors and delays. Most patients with ECD require treatment, except for a minority of patients with minimally symptomatic single-organ disease. The first ECD consensus guidelines were published in 2014 on behalf of the physicians and researchers within the Erdheim-Chester Disease Global Alliance. With the recent molecular discoveries and the approval of the first targeted therapy (vemurafenib) for BRAF-V600–mutant ECD, there is a need for updated clinical practice guidelines to optimize the diagnosis and treatment of this disease. This document presents consensus recommendations that resulted from the International Medical Symposia on ECD in 2017 and 2019. Herein, we include the guidelines for the clinical, laboratory, histologic, and radiographic evaluation of ECD patients along with treatment recommendations based on our clinical experience and review of literature in the molecular era.

Contains fulltext : 249882.pdf (Publisher’s version ) (Closed access) ALK-positive histiocytosis is a rare subtype of histiocytic neoplasm first described in 2008 in 3 infants with multisystemic disease involving the liver and hematopoietic system. This entity has subsequently been documented in case reports and series to occupy a wider clinicopathologic spectrum with recurrent KIF5B-ALK fusions. The full clinicopathologic and molecular spectra of ALK-positive histiocytosis remain, however, poorly characterized. Here, we describe the largest study of ALK-positive histiocytosis to date, with detailed clinicopathologic data of 39 cases, including 37 cases with confirmed ALK rearrangements. The clinical spectrum comprised distinct clinical phenotypic groups: infants with multisystemic disease with liver and hematopoietic involvement, as originally described (Group 1A: 6/39), other patients with multisystemic disease (Group 1B: 10/39), and patients with single-system disease (Group 2: 23/39). Nineteen patients of the entire cohort (49%) had neurologic involvement (7 and 12 from Groups 1B and 2, respectively). Histology included classic xanthogranuloma features in almost one-third of cases, whereas the majority displayed a more densely cellular, monomorphic appearance without lipidized histiocytes but sometimes more spindled or epithelioid morphology. Neoplastic histiocytes were positive for macrophage markers and often conferred strong expression of phosphorylated extracellular signal-regulated kinase, confirming MAPK pathway activation. KIF5B-ALK fusions were detected in 27 patients, whereas CLTC-ALK, TPM3-ALK, TFG-ALK, EML4-ALK, and DCTN1-ALK fusions were identified in single cases. Robust and durable responses were observed in 11/11 patients treated with ALK inhibition, 10 with neurologic involvement. This study presents the existing clinicopathologic and molecular landscape of ALK-positive histiocytosis and provides guidance for the clinical management of this emerging histiocytic entity.

The historic term ‘histiocytosis’ meaning ‘tissue cell’ is used as a unifying concept for diseases characterized by pathogenetic myeloid cells that share histologic features with macrophages or dendritic cells. These cells may arise from the embryonic yolk sac, fetal liver, or post-natal bone marrow. Prior classification schemes align disease designation with terminal phenotype: for example, Langerhans cell histiocytosis (LCH) shares CD207(+) antigen with physiologic epidermal Langerhans cells. LCH, Erdheim-Chester disease (ECD), juvenile xanthogranoluma (JXG), and Rosai-Dorfman disease (RDD) are all characterized by pathologic ERK activation driven by activating somatic mutations in MAPK pathway genes. The title of this Primer (‘Histiocytic Disorders’) was chosen to differentiate the above diseases from Langerhans cell sarcoma and malignant histiocytosis, which are hyper-proliferative lesions typical of cancer. By comparison LCH, ECD, RDD, and JXG share some features of malignant cells including activating MAPK pathway mutations, but are not hyper-proliferative. ‘Inflammatory myelo-proliferative neoplasm’ may be a more precise nomenclature. By contrast, hemohagocytic lymphohistiocytosis (HLH) is associated macrophage activation and extreme inflammation, representing a syndrome if immune dysregulation. These diseases affect children and adults in varying proportions depending on which of the entities is involved.

Malignant histiocytic neoplasm with histiocytic sarcoma phenotype is a rare malignant neoplasm, distinguished by malignant cells with phenotypic characteristics of mature tissue histiocytes. Histiocytic sarcoma typically presents as a primary malignancy, although can also present as a secondary malignancy, and is rarely seen in the pediatric population. Due to the rarity of this condition, diagnosis of histiocytic sarcoma is difficult and considered a diagnosis of exclusion. We describe a unique case of a chronic upper eyelid lesion with biopsy findings of a highly atypical histiocytic neoplasm initially concerning for histiocytic sarcoma; however, after integration of clinical findings, non-progressive and quiescent molecular profile, concluded to be an atypical juvenile xanthogranuloma in a child treated with excision and observation alone. This report highlights the importance of an integrated team approach to diagnosis of unusual histiocytic neoplasms.

The family of juvenile xanthogranuloma family neoplasms (JXG) with ERK-pathway mutations are now classified within the “L” (Langerhans) group, which includes Langerhans cell histiocytosis (LCH) and Erdheim Chester disease (ECD). Although theBRAFV600E mutation constitutes the majority of molecular alterations in ECD and LCH, only three reported JXG neoplasms, all in male pediatric patients with localized central nervous system (CNS) involvement, are known to harbor theBRAFmutation. This retrospective case series seeks to redefine the clinicopathologic spectrum of pediatric CNS-JXG family neoplasms in the post-BRAF era, with a revised diagnostic algorithm to include pediatric ECD. Twenty-two CNS-JXG family lesions were retrieved from consult files with 64% (n = 14) having informativeBRAFV600E mutational testing (molecular and/or VE1 immunohistochemistry). Of these, 71% (n = 10) were pediatric cases (≤18 years) and half (n = 5) harbored theBRAFV600E mutation. As compared to theBRAFwild-type cohort (WT), theBRAFV600E cohort had a similar mean age at diagnosis [BRAF V600E: 7 years (3–12 y), vs. WT: 7.6 years (1–18 y)] but demonstrated a stronger male/female ratio (BRAF V600E: 4 vs WT: 0.67), and had both more multifocal CNS disease ( BRAFV600E: 80% vs WT: 20%) and systemic disease (BRAF V600E: 40% vs WT: none). Radiographic features of CNS-JXG varied but typically included enhancing CNS mass lesion(s) with associated white matter changes in a subset ofBRAFV600E neoplasms. After clinical-radiographic correlation, pediatric ECD was diagnosed in theBRAFV600E cohort. Treatment options varied, including surgical resection, chemotherapy, and targeted therapy with BRAF-inhibitor dabrafenib in one mutated case.BRAFV600E CNS-JXG neoplasms appear associated with male gender and aggressive disease presentation including pediatric ECD. We propose a revised diagnostic algorithm for CNS-JXG that includes an initial morphologic diagnosis with a final integrated diagnosis after clinical-radiographic and molecular correlation, in order to identify cases of pediatric ECD. Future studies with long-term follow-up are required to determine if pediatricBRAFV600E positive CNS-JXG neoplasms are a distinct entity in the L-group histiocytosis category or represent an expanded pediatric spectrum of ECD.

Although littoral cell angiomas (LCAs) are phenotypically well characterized, the antibodies used to support the diagnosis identify many other cells in the normal spleen, and some may be found in other angiomatous lesions. Based on a langerin/CD207+ LCA index case, langerin and other selected immunohistochemical staining was performed on 10 LCAs, 20 other splenic angiomatous lesions, and 7 reactive lymph nodes to further investigate the role of langerin as a diagnostic tool. Ninety percent (9/10) of LCAs were langerin positive, whereas only 1 (5%) of 20 other splenic vascular lesions was partially positive (P < .00001). All LCAs were CD1a-, CD68+, CD34-, and CD8-; 20% were S100+, 70% CD21+, and 90% cyclin D1+. Ultrastructural studies of one LCA did not show Birbeck-type granules in definite lining cells. Sinus lining cells in 7 of 7 reactive lymph nodes showed partial langerin positivity, and 4 of 4 showed partial cyclin D1 positivity. In conclusion, langerin staining is an easily interpreted and highly sensitive and specific (sensitivity [0.90], specificity [0.95]) ancillary study to help distinguish LCA from other vascular tumors of the spleen. Whether this represents cross-reactivity or true CD207 expression is uncertain, as other immunohistochemical and ultrastructural studies do not support a Langerhans cell origin. The cyclin D1 staining seen in most LCA would be consistent with their expression of other selected vascular and histiocytic markers. The similar staining pattern in some lymph node sinus lining cells suggests a possible similar cell of origin, although LCA of lymph nodes is not described.

BACKGROUND “Langerhans cell histiocytosis” (LCH) is a term that encompasses single-system or multisystem disorders traditionally characterized by a proliferation of clonal CD1a+/CD207+ myeloid-derived histiocytes. In most cases of LCH, mitogen-activated protein kinase (MAPK) pathway somatic mutations lead to near universal upregulation of phosphorylated extracellular signal-regulated kinase expression. The clinical manifestations of LCH are numerous, but bone involvement is common. Intracranial lesions, especially as isolated manifestations, are rare. OBSERVATIONS The authors presented the case of a long-term survivor of exclusive intracranial LCH that manifested with isolated craniofacial bone and intraparenchymal central nervous system recurrences, which were managed with 3 decades of multimodal therapy. The patient was initially diagnosed with LCH at age 2 years, and the authors documented the manifestations of disease and treatment for 36 years. Most of the patient’s treatment course occurred before the discovery of BRAF V600E. Treatments initially consisted of chemotherapy, radiosurgery, and open resections for granulomatous LCH lesions. Into young adulthood, the patient had a minimal disease burden but still required additional radiosurgical procedures and open resections. LESSONS Surgical treatments alleviated the patient’s immediate symptoms and allowed for tumor burden control. However, surgical interventions did not cure the underlying, aggressive disease. In the current era, access to systemic MAPK inhibitor therapy for histiocytic lesions may offer improved outcomes.

Background Accurate risk stratification of Langerhans cell histiocytosis (LCH) is essential as management can range from conservative in single system, low risk for central nervous system (CNS) involvement lesions to intensive chemotherapy for multisystem or high-risk disease. Additionally, being able to differentiate metabolically active from inactive lesions is essential for both prognostic reasons and to avoid potentially unnecessary treatment. Methods A retrospective review was performed on all patients with histopathology-confirmed LCH at Cincinnati Children's Hospital Medical Center (CCHMC) between 2009 and 2019. Results One hundred seven positron emission tomography (PET)/computerized tomography (CT) images were included in the review. A discrepancy between PET/CT and conventional imaging occurred on 53 occasions. On 13 occasions, increased uptake was observed on PET in an area with no identifiable lesion on conventional imaging. On 40 occasions, lesions were found on conventional imaging where no increased uptake was observed on PET. On eight skeletal surveys, three other radiographs, four diagnostic CTs, five localization CTs, and one bone scan, no lesion was identified in an area with increased fluorodeoxyglucose (FDG) uptake. This occurred exclusively in bone. On nine skeletal surveys, one other radiograph, four diagnostic CTs, six localization CTs, 19 magnetic resonance imaging (MRI) scans, and one bone scan, a lesion was identified in a location without increased FDG uptake. This occurred in bone, CNS, and lungs. Conclusion F-18-FDG PET/CT is vital in the evaluation of LCH lesions given its ability to detect LCH lesions not detectable on conventional imaging modalities, as well as its ability to distinguish metabolically active from inactive disease. MRI and diagnostic CT are still useful adjunctive tests for identification of CNS and lung lesions.

Pediatric histiocytic neoplasms are hematopoietic disorders frequently driven by the BRAF-V600E mutation. Here we identified two BRAF gene fusions (novel MTAP-BRAF and MS4A6A-BRAF) in two aggressive histiocytic neoplasms. In contrast to previously described BRAF fusions, MTAP-BRAF and MS4A6A-BRAF do not respond to the paradox breaker RAF inhibitor (RAFi) PLX8394 due to stable fusion dimerization mediated by the N-terminal fusion partners. This highlights a significant and clinically relevant shift from the current dogma that BRAF-fusions respond similarly to BRAF-inhibitors. As an alternative, we show suppression of fusion-driven oncogenic growth with the pan-RAFi LY3009120 and MEK inhibition.

Pediatric histiocytic neoplasms are hematopoietic disorders frequently driven by the BRAF-V600E mutation. Here, we identified two BRAF gene fusions (novel MTAP-BRAF and MS4A6A-BRAF) in two aggressive histiocytic neoplasms. In contrast to previously described BRAF fusions, MTAP-BRAF and MS4A6A-BRAF do not respond to the paradox breaker RAF inhibitor (RAFi) PLX8394 due to stable fusion dimerization mediated by the N-terminal fusion partners. This highlights a significant and clinically relevant shift from the current dogma that BRAF-fusions respond similarly to BRAF-inhibitors. As an alternative, we show suppression of fusion-driven oncogenic growth with the pan-RAFi LY3009120 and MEK inhibition.

Summary Lipoblastomas can occasionally require further molecular confirmation when occurring outside of the usual age groups or demonstrating unusual morphology. We reviewed 28 lipoblastomas with 16 controls. Lipoblastomas were subdivided into myxoid (n = 7), classic (n = 9), or lipoma-like (n = 12) subtypes. PLAG1 immunohistochemistry, PLAG1 fluorescence in situ hybridization (FISH), and targeted RNA sequencing were performed on formalin-fixed paraffin-embedded tissue. Karyotypes were available in a subset of lipoblastomas (n = 9). Gene rearrangements were identified in 17/25 (68%) lipoblastomas, including PLAG1 (15/25, 60%) and HMGA2 (2/25, 8%). Five novel fusion partners (DDX6, KLF10, and KANSL1L with PLAG1 and EP400 and FGD6 with HMGA2) were found. PLAG1 immunohistochemistry was positive (nuclear, moderate/strong) in myxoid and classic subtypes lipoblastomas with preferential expression in mesenchymal cells within myxoid stroma and fibrous septa and negative in all controls. When comparing PLAG1 immunohistochemistry with molecular testing (FISH and/or RNA sequencing and/or karyotype), concordant results were noted in 13/25 (52%) cases, increasing to 15/25 (60%) after slight adjustment of the PLAG1 FISH positive threshold. In myxoid and classic lipoblastomas, PLAG1 immunohistochemistry seems to be a better surrogate marker for PLAG1 rearrangement, as compared with lipoma-like subtypes. In lipoma-like subtypes, targeted RNA sequencing appears to detect PLAG1 fusions better than FISH and immunohistochemistry. The preferential expression of PLAG1 in the mesenchymal and fibroblast-like cells deserves further investigation as the putative cell of origin in lipoblastoma.

Pediatric histiocytic neoplasms are clonal hematopoietic disorders driven by mutations activating the mitogen-activated protein kinase (MAPK) pathway, such as BRAF-V600E. In non-BRAFV600E cases, we investigated alternative MAPK mutations and found two novel BRAF gene fusions. We investigated the distinct responsiveness of novel BRAF fusions to RAFi therapies and explored the mechanistic basis of such differential responses compared to other BRAF fusions. Two histiocytic patient tumors were analyzed using the CHOP Comprehensive Next-Gen Sequencing Solid Tumor Panel and a targeted RNA-seq panel for 106 fusion partner genes. In the two M- and L-type histiocytic neoplasms assessed, we found novel and rare BRAF gene fusions, MTAP-BRAF and MS4A6A-BRAF, respectively. Both BRAF fusions activated the MAPK/ PI3K pathways and showed homo- and hetero-dimerization with BRAF and the respective N-terminal fusion partner. In contrast to common BRAF fusions, MTAP-BRAF and MS4A6A-BRAF did not respond to PLX8394 due to a lack of disruption of active fusion homo- and hetero-dimers, which was in turn due to the untargeted, stable dimerization mediated by the N-terminal fusion partners. Conversely, we observed robust suppression with LY3009120 that bound fusion dimers and kept them in an inactivate confirmation. MEKi were found to successfully suppress fusion driven signaling and oncogenic phenotypes. Our finding that PLX8394 does not disrupt MTAP-BRAF or MS4A6A-BRAF dimerization due to contribution of N-terminal partners defines a novel paradigm for the distinct mechanisms sought by BRAF fusions in response to RAFi therapy. Overall, this study highlights the unique and differential biology hijacked by BRAF fusions in response to RAFi and further warrants detailed mechanistic classification of BRAF fusions based on their responsiveness to targeted agents.

Langerhans cell histiocytosis (LCH) is a myeloid neoplasia, driven by sporadic activating mutations in the MAPK pathway. The misguided myeloid dendritic cell (DC) model proposes that high-risk, multisystem, risk-organ–positive (MS-RO+) LCH results from driver mutation in a bone marrow (BM)-resident multipotent hematopoietic progenitor, while low-risk, MS-RO− and single-system LCH would result from driver mutation in a circulating or tissue-resident, DC-committed precursor. We have examined the CD34+c-Kit+Flt3+ myeloid progenitor population as potential mutation carrier in all LCH disease manifestations. This population contains oligopotent progenitors of monocytes (Mo’s)/macrophages (MΦs), osteoclasts (OCs), and DCs. CD34+c-Kit+Flt3+ cells from BM of MS-RO+ LCH patients produced Langerhans cell (LC)-like cells in vitro. Both LC-like and DC offspring from this progenitor carried the BRAF mutation, confirming their common origin. In both high- and low-risk LCH patients, CD34+c-Kit+Flt3+ progenitor frequency in blood was higher than in healthy donors. In one MS-RO+ LCH patient, CD34+c-Kit+Flt3+ cell frequency in blood and its BRAF-mutated offspring reported response to chemotherapy. CD34+c-Kit+Flt3+ progenitors from blood of both high- and low-risk LCH patients gave rise to DCs and LC-like cells in vitro, but the driver mutation was not easily detectable, likely due to low frequency of mutated progenitors. Mutant BRAF alleles were found in Mo’s /MΦs, DCs, LC-like cells, and/or OC-like cells in lesions and/or Mo and DCs in blood of multiple low-risk patients. We therefore hypothesize that in both high- and low-risk LCH, the driver mutation is present in a BM-resident myeloid progenitor that can be mobilized to the blood.

Capillary hemangioma is a rare benign lesion in the testicle, particularly in pediatrics. It can mimic malignancy, leading to radical orchiectomy. We present a case of a testicular hemangioma in a child, and review the literature on testicular hemangiomas in this age group. A hypervascular testicular lesion without elevated tumor markers may warrant intraoperative biopsy to direct surgical management, which may include testis-sparing surgery if amenable.

Hemophagocytic lymphohistiocytosis (HLH) and macrophage activation syndrome (MAS) are life-threatening hyperferritinemic systemic inflammatory disorders. Although profound cytotoxic impairment causes familial HLH (fHLH), the mechanisms driving non-fHLH and MAS are largely unknown. MAS occurs in patients with suspected rheumatic disease, but the mechanistic basis for its distinction is unclear. Recently, a syndrome of recurrent MAS with infantile enterocolitis caused by NLRC4 inflammasome hyperactivity highlighted the potential importance of interleukin-18 (IL-18). We tested this association in hyperferritinemic and autoinflammatory patients and found a dramatic correlation of MAS risk with chronic (sometimes lifelong) elevation of mature IL-18, particularly with IL-18 unbound by IL-18 binding protein, or free IL-18. In a mouse engineered to carry a disease-causing germ line NLRC4(T337S) mutation, we observed inflammasome-dependent, chronic IL-18 elevation. Surprisingly, this NLRC4(T337S)-induced systemic IL-18 elevation derived entirely from intestinal epithelia. NLRC4(T337S) intestines were histologically normal but showed increased epithelial turnover and upregulation of interferon-γ–induced genes. Assessing cellular and tissue expression, classical inflammasome components such as Il1b, Nlrp3, and Mefv predominated in neutrophils, whereas Nlrc4 and Il18 were distinctly epithelial. Demonstrating the importance of free IL-18, Il18 transgenic mice exhibited free IL-18 elevation and more severe experimental MAS. NLRC4(T337S) mice, whose free IL-18 levels were normal, did not. Thus, we describe a unique connection between MAS risk and chronic IL-18, identify epithelial inflammasome hyperactivity as a potential source, and demonstrate the pathogenicity of free IL-18. These data suggest an IL-18–driven pathway, complementary to the cytotoxic impairment of fHLH, with potential as a distinguishing biomarker and therapeutic target in MAS.

Juvenile granulosa cell tumors of the ovary are rare sex cord-stromal ovarian tumors that are typically diagnosed during the first 2 decades of life. Most patients present with precocious puberty in the early stages of disease. We present a rare case of asymptomatic uterine torsion from a 15-cm juvenile granulosa cell tumors in a 5-year-old girl with elevated inhibin B, breast development, vaginal bleeding, and a palpable right-sided abdominal mass.

Hemophagocytic lymphohistiocytosis (HLH) is an often-fatal disorder characterized by the overactivation of T cells and macrophages that excessively produce proinflammatory cytokines, including interferon-γ (IFN-γ). Previously, we reported that the JAK inhibitor ruxolitinib dampens T-cell activation and lessens inflammation in a model of HLH in which perforin-deficient (Prf1−/−) mice are infected with lymphocytic choriomeningitis virus (LCMV). Ruxolitinib inhibits signaling downstream of IFN-γ, as well as several other JAK-dependent cytokines. As a consequence, it remained unclear whether ruxolitinib was exerting its beneficial effects in HLH by inhibiting IFN-γ signaling or by targeting signaling initiated by other proinflammatory cytokines. To address this question, we compared the effects of ruxolitinib with those obtained using an IFN-γ–neutralizing antibody (αIFN-γ) in 2 murine HLH models. In both models, ruxolitinib and αIFN-γ reduced inflammation-associated anemia, indicating that ruxolitinib operates in an IFN-γ–dependent manner to reverse this HLH manifestation. In contrast, the number and activation status of T cells and neutrophils, as well as their infiltration into tissues, were significantly reduced following treatment with ruxolitinib, but they remained unchanged or were increased following treatment with αIFN-γ. Notably, despite discontinuation of ruxolitinib, LCMV-infected Prf1−/− mice exhibited enhanced survival compared with mice in which αIFN-γ was discontinued. This protective effect could be mimicked by transient treatment with αIFN-γ and a neutrophil-depleting antibody. Thus, ruxolitinib operates through IFN-γ–dependent and -independent mechanisms to dampen HLH by targeting the deleterious effects of T cells and neutrophils, with the latter representing an unappreciated and understudied cell type that contributes to HLH pathogenesis.

Key Points Demonstration of BRAF-V600E in Rosai-Dorfman-Destombes disease requires sensitive molecular assays and molecular-based tissue immunostain. BRAF-V600E blood testing is important for disease-monitoring BRAF-mutated histiocytosis and can guide inhibitor treatment plans.

Langerhans cell histiocytosis (LCH) is a potentially fatal condition characterized by granulomatous lesions with characteristic clonal mononuclear phagocytes (MNPs) harboring activating somatic mutations in mitogen-activated protein kinase (MAPK) pathway genes, most notably BRAF(V600E). We recently discovered that the BRAF(V600E) mutation can also affect multipotent hematopoietic progenitor cells (HPCs) in multisystem LCH disease. How the BRAF(V600E) mutation in HPCs leads to LCH is not known. Here we show that enforced expression of the BRAF(V600E) mutation in early mouse and human multipotent HPCs induced a senescence program that led to HPC growth arrest, apoptosis resistance and a senescence-associated secretory phenotype (SASP). SASP, in turn, promoted HPC skewing toward the MNP lineage, leading to the accumulation of senescent MNPs in tissue and the formation of LCH lesions. Accordingly, elimination of senescent cells using INK-ATTAC transgenic mice, as well as pharmacologic blockade of SASP, improved LCH disease in mice. These results identify senescent cells as a new target for the treatment of LCH.

Histiocytoses are clonal hematopoietic disorders frequently driven by mutations mapping to the BRAF and MEK1 and MEK2 kinases. Currently, however, the developmental origins of histiocytoses in patients are not well understood, and clinically meaningful therapeutic targets outside of BRAF and MEK are undefined. In this study, we uncovered activating mutations in CSF1R and rearrangements in RET and ALK that conferred dramatic responses to selective inhibition of RET (selpercatinib) and crizotinib, respectively, in patients with histiocytosis.

The histiocytoses are rare disorders characterized by the accumulation of macrophage, dendritic cell, or monocyte-derived cells in various tissues and organs of children and adults. More than 100 different subtypes have been described, with a wide range of clinical manifestations, presentations, and histologies. Since the first classification in 1987, a number of new findings regarding the cellular origins, molecular pathology, and clinical features of histiocytic disorders have been identified. We propose herein a revision of the classification of histiocytoses based on histology, phenotype, molecular alterations, and clinical and imaging characteristics. This revised classification system consists of 5 groups of diseases: (1) Langerhans-related, (2) cutaneous and mucocutaneous, and (3) malignant histiocytoses as well as (4) Rosai-Dorfman disease and (5) hemophagocytic lymphohistiocytosis and macrophage activation syndrome. Herein, we provide guidelines and recommendations for diagnoses of these disorders.

BACKGROUND Post-transplant lymphoproliferative disorders (PTLDs) are well characterized in tissue sections, but their evaluation in exfoliative cytology specimens is limited. This study reports a 25-year experience with PTLDs in exfoliative cytology specimens. METHODS All solid organ or allogeneic stem cell transplant recipients with PTLDs and exfoliative cytology specimens from 1987 to 2011 were identified. The cytomorphology, Epstein-Barr virus (EBV) status, flow cytometry, immunohistochemistry, and molecular studies were reviewed from all exfoliative cytology specimens previously diagnosed as atypical lymphoid proliferations or PTLDs. RESULTS A total of 55 patients (age range, 1-72 years) with PTLDs had 434 exfoliative cytology specimens. Thirty-six of the 55 patients (65%) had 54 specimens with abnormal lymphoid proliferations (12% of the specimens), and 26 of these patients had 37 specimens available for review (15 cerebrospinal fluid specimens, 12 peritoneal fluid specimens, 9 pleural fluid specimens, and 1 bronchoalveolar lavage fluid specimen). Thirty percent of the reviewed cytology specimens were diagnostic of PTLDs, including 8 cases of monomorphic post-transplant lymphoproliferative disorder (M-PTLD) with abnormal B/T-cell populations identified with flow cytometry/immunohistochemistry and 3 EBV-positive specimens with a differential diagnosis of polymorphic PTLD versus M-PTLD. All cases diagnostic of a PTLD had 1 to 3 ancillary studies performed. Forty percent of the cytology specimens (15 of 37) were suspicious for a PTLD, but ancillary studies were performed for only a third of them, and they did not support a definitive diagnosis of a PTLD. Thirty percent of the cytology specimens (11 of 37) appeared reactive, but they lacked sufficient ancillary studies to exclude a PTLD. CONCLUSIONS Atypical lymphoid proliferations are common in exfoliative cytology specimens from patients with PTLDs, and they require ancillary studies at least including immunophenotyping and EBV evaluations for a definitive diagnosis. Cancer (Cancer Cytopathol) 2016. © 2016 American Cancer Society.

Neonatal cholestasis is a potentially life-threatening condition requiring prompt diagnosis. Mutations in several different genes can cause progressive familial intrahepatic cholestasis, but known genes cannot account for all familial cases. Here we report four individuals from two unrelated families with neonatal cholestasis and mutations in NR1H4, which encodes the farnesoid X receptor (FXR), a bile acid-activated nuclear hormone receptor that regulates bile acid metabolism. Clinical features of severe, persistent NR1H4-related cholestasis include neonatal onset with rapid progression to end-stage liver disease, vitamin K-independent coagulopathy, low-to-normal serum gamma-glutamyl transferase activity, elevated serum alpha-fetoprotein and undetectable liver bile salt export pump (ABCB11) expression. Our findings demonstrate a pivotal function for FXR in bile acid homeostasis and liver protection., Neonatal cholestasis is a result of elevated bile acid levels, and is associated with mutations in genes regulating bile acid homeostasis. Here the authors identify mutations in the bile acid sensing farnesoid X receptor in four individuals with neonatal cholestasis from two unrelated families.

We report an unusual case of a patient with primary CNS-HS a very rare neoplasm of histiocytic lineage with usually poor prognosis. An 8 year old boy presented with a one month history of headaches, nausea and vomiting. Physical examination revealed nystagmus and dysmetria. Brain MRI revealed a localized 2.4 cm posterior fossa (cerebellar) mass with restricted diffusion. The patient underwent a gross total resection of the mass. Initial post-operative lumbar puncture was positive for rare malignant cells. Pathology showed a focally necrotic neoplasm, composed of nests and cords of large relatively uniform cells with abundant eosinophilic cytoplasm, moderately pleomorphic nuclei and numerous mitotic figures, consistent with CNS-HS with juvenile xanthogranuloma phenotype, as supported by positive IHC expression of CD163, CD68, CD14, fascin, and Factor XIIIa, while negative for CD1a, Lymphoid and Myeloid markers, and BRAFv600e mutation. He was treated with two cycles of clofarabine and cytarabine and triple intrathecal (IT) chemotherapy. He developed generalized seizures and MRI showed demyelination consistent with IT methotrexate toxicity; MTX was then discontinued. He was then given two additional cycles of cladribine and weekly intrathecal therapy prior to consolidation with an Allo-HCT using a 10/10 HLA allelic-matched unrelated donor. His conditioning regimen included total body irradiation and cyclophosphamide. He did well post-transplant with peripheral blood chimerism at 1 year showing > 95% donor cells. He remains disease-free with an excellent quality of life since August 2016. We report one of the few known survivors of this unusual and highly malignant entity.

Introduction: Pediatric histiocytic neoplasms are clonal hematopoietic disorders driven by mutations activating the mitogen-activated protein kinase (MAPK) pathway, such as BRAF- V600E. In non-BRAFV600E cases, we investigated alternative MAPK mutations and found two novel BRAF gene fusions. We have previously shown effective targeting of BRAF fusions (KIAA1549-BRAF) in pediatric low-grade gliomas (PLGGs) with paradox breaker RAF inhibitor (RAFi), PLX8394 and MEK inhibitors (MEKi). Here, we investigate the distinct responsiveness of novel BRAF fusions to RAFi therapies and explore the mechanistic basis of such differential responses compared to other BRAF fusions. Method: Two histiocytic patient tumors were analyzed using the CHOP Comprehensive Next- Gen Sequencing Solid Tumor Panel and a targeted RNA-seq panel for 106 fusion partner genes. Novel BRAF fusions identified were sub-cloned to create heterologous cell models. Immunoblotting of serum starved cells measured MAPK and PI3K pathway activation and soft agar assay tested for oncogenic phenotypes driven by BRAF fusions, along with response to PLX8394, LY3009120 (pan-RAF dimer inhibitor), and MEKi. Co-immunoprecipitation assays using Myc- and Flag-tagged BRAF fusion constructs assessed dimerization profiles, with or without inhibitors. Result: In the two M- and L-type histiocytic neoplasms assessed, we found novel and rare BRAF gene fusions, MTAP-BRAF and MS4A6A-BRAF, respectively. Both BRAF fusions activated the MAPK/ PI3K pathways and showed homo- and hetero-dimerization with BRAF and the respective N-terminal fusion partner. Compared to common BRAF fusions, MTAP-BRAF and MS4A6A- BRAF did not respond to PLX8394 due to no disruption of active fusion homo- and hetero-dimers, which was in turn was due to the untargeted, stable dimerization mediated by the N-terminal fusion partners. Conversely, we observed robust suppression with LY3009120 that bound fusion dimers and kept them in an inactivate confirmation. MEKi selumetinib and trametinib also suppressed fusion driven signaling and oncogenic phenotypes. Conclusion: We show that novel MTAP-BRAF and MS4A6A-BRAF fusions, found in histiocytic tumors, do not respond to RAFi PLX8394 but are targeted by LY3009120 or MEKi. Our finding that PLX8394 does not disrupt MTAP-BRAF or MS4A6A-BRAF dimerization due to contribution of N-terminal partners defines a novel paradigm for the distinct mechanisms sought by BRAF fusions in response to RAFi therapy. We show potent suppression of these mechanistically distinct BRAF fusions with MEKi or LY3009120 which function independent of vulnerability to fusion dimerization. Overall, this study highlights the unique and differential biology hijacked by BRAF fusions in response to RAFi and further warrants detailed mechanistic classification of BRAF fusions based on their responsiveness to targeted agents. Citation Format: Payal Jain, Lea F. Surrey, Joshua Straka, Pierre Russo, Richard Womer, Marilyn Li, Phillip B. Storm, Angela J. Waanders, Michael Hogarty, Jennifer Picarsic, Adam C. Resnick. Novel BRAF gene fusions in pediatric histiocytic neoplasms respond differentially to RAF targeted therapies based on dimerization profiles [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 632.

Introduction: In children, approximately 7% of the non-CNS solid tumors include extremely rare histologies such as angiosarcomas and histiocytic neoplasms. While most recurrent histologies can be identified with a commonly occurring alteration or associated with a known biologic mechanism, rare solid tumors remain poorly characterized given their low frequency of occurrence and intrapatient tumor heterogeneity. Our group and others have demonstrated gene fusions (GFs) as unique oncogenic alterations in pediatric gliomas to guide prognosis, diagnosis, and therapeutic interventions. Here, we investigated the role of unique GFs as individualized therapeutic targets in 3 patients with rare non-CNS malignancies and explored GF-directed precision medicine approaches. Methods: We selected a patient with the rare diagnosis of pediatric angiosarcoma (pAS) and two with pediatric histiocytic neoplasms spanning the juvenille xanthogranuloma (JXG) family and malignant histiocytoses groups. Neoplasms were analyzed using the CHOP Comprehensive Next-Generation Sequencing Solid Tumor Panel as well as a targeted RNA-seq panel for 106 fusion partner genes. For novel gene fusions identified, we performed molecular and therapeutic characterization, including subcloning to create heterologous cell models. Cellular assays were used to assess oncogenic transformation, test mechanistic activation of related downstream signaling pathways via cellular assays, and examine targeting of GFs with specific small-molecule inhibitors. Results and Discussion: We identified a novel NOTCH1-ROS1 gene fusion in pAs, and rare BRAF-fusions, MTAP-BRAF and MS4A6A-BRAF, in malignant and JXG family histiocytic neoplasms, respectively. We also identified a germline TP53 p.T123Wfs*12 pathogenic variant, and the cancer predisposition team confirmed Li Fraumeni syndrome in the angiosarcoma patient. Our studies demonstrate that the NOTCH1-ROS1 fusion and both BRAF-fusions are potent oncogenes capable of inducing neoplastic transformation in cells and tumor formation in a murine allograft model. The kinase domains of ROS1 and BRAF are retained and activate the MAPK/PI3K/mTOR and JAK-STAT pathways in respective GF- expressing models, driven by potent dimerization of the GFs. Upon testing ROS1-targeted tyrosine kinase inhibitors (TKI) against NOTCH1-ROS1, we observe dose-dependent suppression of Notch1-Ros1 driven cellular activity and concurrent inhibition of tumor growth as well as prolonged survival after oral monotherapy. Interestingly, BRAF-fusions do not respond to second-generation BRAF inhibitors but can be suppressed by RAF dimer-inhibitor LY3009120 and MEK inhibitors. Overall, these data suggest that ongoing genomic profiling of rare pediatric tumors may reveal actionable drivers, and molecular testing of putative oncogenic fusions is imperative for improvement of patient outcomes through precision therapy. Citation Format: Payal Jain, Lea F. Surrey, Joshua Straka, Tiffany Smith, Sudarshan Iyer, Elizabeth Fox, Monika Davare, Jennifer Picarsic, Marilyn Li, Angela J. Waanders, Adam C. Resnick. Unique gene fusions inform targeted therapeutic strategies across extremely rare, non-CNS pediatric solid tumors [abstract]. In: Proceedings of the AACR Special Conference on the Advances in Pediatric Cancer Research; 2019 Sep 17-20; Montreal, QC, Canada. Philadelphia (PA): AACR; Cancer Res 2020;80(14 Suppl):Abstract nr A37.

Rosai-Dorfman-Destombes disease (RDD) is a rare non–Langerhans cell histiocytosis characterized by accumulation of activated histiocytes within affected tissues. RDD, which now belongs to the R group of the 2016 revised histiocytosis classification, is a widely heterogeneous entity with a range of clinical phenotypes occurring in isolation or in association with autoimmune or malignant diseases. Recent studies have found NRAS, KRAS, MAP2K1, and ARAF mutations in lesional tissues, raising the possibility of a clonal origin in some forms of RDD. More than 1000 reports have been published in the English literature; however, there is a lack of consensus regarding approach for the clinical management of RDD. Although in most cases RDD can be observed or treated with local therapies, some patients with refractory or multifocal disease experience morbidity and mortality. Here we provide the first consensus multidisciplinary recommendations for the diagnosis and management of RDD. These recommendations were discussed at the 32nd Histiocyte Society Meeting by an international group of academic clinicians and pathologists with expertise in RDD. We include guidelines for clinical, laboratory, pathologic, and radiographic evaluation of patients with RDD together with treatment recommendations based on clinical experience and review of the literature.

The term histiocyte has come to encompass two cell lines, the monocyte-macrophages and the specialized antigen-presenting dendritic cells (DCs). The nomenclature, cellular origin, and overlapping functional capabilities of the mononuclear phagocytic system (MPS) are under continual refinement. The understanding of these cells in steady state, inflammatory states, and various disease states is discussed, with special emphasis on the histiocytic disorders and neoplasms in relationship to their revised classifications and molecular characterization. These include Langerhans cell histiocytosis (LCH), Erdheim-Chester disease (ECD), juvenile xanthogranuloma family (JXG) lesions, Rosai-Dorfman disease (RDD), and histiocytic malignancies (histiocytic sarcoma, Langerhans cell sarcoma, indeterminate and interdigitating cell sarcoma). Updates in the field of hemophagocytic lymphohistiocytosis, both primary and secondary forms, are also reviewed.

Langerhans cell histocytosis (LCH) and Erdheim-Chester disease are two rare histiocytic disorders. Their occurrence in the same patient is more infrequent, but has been described. We report a case of a 38-year-old woman who presented with a diagnosis of single system cutaneous LCH. Subsequently, she developed multiple papules on her extremities consistent with a non-LCH xanthogranuloma type lesion. BRAF(V600E) mutation immunostain, VE1 was positive in the skin lesion, which was confirmed by molecular polymerase chain reaction (PCR) studies, initiating a complete systemic workup for Erdheim-Chester disease. Systemic involvement was confirmed with bilateral sclerotic bone lesions and retroperitoneal and pelvic fibrosing disease. She was also found to have a BRAF(V600E) mutation positive papillary thyroid carcinoma. New suspicious cutaneous lesions presenting in patients with a history of LCH need to be biopsied. A BRAF(V600E) mutation in a non-LCH histiocytic lesion with a xanthogranuloma phenotype (CD163/CD68/CD14/fascin/Factor 13a) should prompt an Erdheim-Chester disease workup. This is a unique case of a woman with BRAF(V600E) mutation positive Erdheim-Chester disease and cutaneous LCH, while also being, to our knowledge, the first reported case in the English literature of it occurring in a patient with a BRAF(V600E) mutation positive papillary thyroid carcinoma.

Papillary thyroid carcinoma (PTC) is rare in children, although it is a known secondary malignancy after treatment for neuroblastoma (NB). The interval between NB treatment completion and PTC is usually more than 5 years. A 4-year-old, female patient with a high risk adrenal NB was found to have a 2.9-cm, right thyroid nodule on surveillance chest computed tomography (CT) 6 months after completion of her NB treatment (induction chemotherapy, tumor resection, autologous stem cell transplantation, external beam radiation to the abdominal tumor site, immunotherapy, and retinoic acid). Posttreatment surveillance included iodine-123-metaiodobenzylguanidine scans and CT scans. Fine-needle aspiration of the thyroid nodule diagnosed a follicular neoplasm, which was negative for BRAF, NRAS, KRAS, HRAS, PAX8/PPARg, and RET/PTC mutations, without evidence of metastatic NB. Nodule histology demonstrated an encapsulated follicular variant of PTC (FVPTC). Next-generation sequence analysis for a 46 cancer-gene profile was performed on both tumors with subsequent peripheral blood DNA testing. A heterozygous missense mutation in STK11 (F354L) was identified in both the NB and FVPTC. This mutation was also detected in peripheral blood mononuclear cells. Two additional heterozygous somatic missense mutations of uncertain significance were identified: KDR/VEGF receptor 2 (Q472H) on chromosome 4 and MET (N375S) on chromosome 7. To our knowledge, this is the shortest reported duration from completion of NB treatment to detection of thyroid cancer. The association of the STK11 gene with Peutz-Jeghers syndrome, lung adenocarcinomas, and medullary thyroid cancer leads to a possible association between this genetic variant and our patient's tumors.

Thyroid nodules are less common in children than adults, but the risk of malignancy in thyroid nodules is much higher in children. The ability to characterize pediatric thyroid nodules has improved with the use of ultrasound-guided fine-needle aspiration, the Bethesda System for Reporting Thyroid Cytopathology (TBSRTC) classification system, and expanded molecular testing. Nevertheless, stratification criteria to predict thyroid malignancy in children are poorly defined. Our objective was to determine if clinical presentation and molecular genetics could predict malignancy in pediatric thyroid nodules.Retrospective chart review of patients ≤18 years of age at the Children's Hospital of Pittsburgh of the University of Pittsburgh Medical Center with the diagnosis of a thyroid nodule from January 2007 to January 2012 was conducted. Eighty-nine subjects fulfilled the inclusion criteria: 1) thyroid nodule ≥0.8 cm and biopsy (n=76), or 2) thyroid nodule ≥0.8 cm, no biopsy, and ultrasound follow-up for at least 2 years (n=13).Twenty-four (27%) of 89 patients were diagnosed with thyroid cancer (50% papillary thyroid carcinoma [PTC], 50% follicular variant of papillary thyroid carcinoma [FVPTC]). Features associated with malignancy included larger nodule size, palpable nodule, or palpable lymphadenopathy. There were no differences in presenting features between patients with PTC and those with FVPTC. Thyroid malignancy was diagnosed in all nine patients with a molecular abnormality (BRAF, RAS, RET/PTC, PAX8/PPARγ).Clinical features, FNA cytology, and molecular genetics are valuable tools to discriminate benign from malignant nodules in pediatric patients. This information is important to direct subsequent clinical management.

Thymic involvement by Langerhans cell histiocytosis (LCH) has been described mainly in isolated case reports. A description of the histopathologic patterns of LCH proliferations in the thymus, together with therapeutic implications, has not, to our knowledge, been previously addressed. The pathology consultation files at Children's Hospital of Pittsburgh of the University of Pennsylvania Medical Center were reviewed for cases of thymic involvement by LCH. Relevant cases in the literature were also reviewed, and the histopathology and clinical course of those cases were collected. Nine consultation cases of thymic involvement were reviewed, together with 23 cases in the literature, which provided adequate pathologic description and ancillary confirmation (n = 32), revealing 4 distinct pathologic groups. Group 1 showed microscopic collection of hyperplastic LCH-like cells in incidental thymectomies of patients without LCH disease, requiring no further treatment (n = 7; 22%). Group 2 showed solitary and/or cystic LCH of the thymus with gland disruption, and at least 3 cases resolved without systemic therapy (n = 10; 31%). Group 3 showed more variable thymic involvement in multisystemic LCH disease, with either a medullary restricted pattern or more diffuse gland involvement, requiring adjuvant therapy and having a higher mortality rate (n = 13; 41%). Group 4 showed a mixed histiocytic lesion with a concurrent LCH and juvenile xanthogranuloma-like proliferation (n = 2; 6%). Thymic involvement in LCH is quite rare. Based on our cases and those in the literature, we propose 4 distinct pathologic groups of thymic involvement in Langerhans cell proliferations with relevance for diagnosis and treatment.

Introduction Ectopic thymic tissue can arise as an asymptomatic neck mass, which may be detected on imaging studies. The aim of this study was to determine the incidence of ectopic thymic tissue in paediatric FNAs and to the correlate clinical, radiological and cytomorphological findings. Methods FNAs in children with neck and mediastinal lesions performed between January 2012 and July 2016 were reviewed for cases of ectopic thymus. These were then evaluated and correlated with the cytology findings. Results Of 739 FNAs, 13 (1.8%) cases from 11 patients showed ectopic thymic tissue. The targeted lesions were in the thyroid (n = 7), submandibular region (n = 1), superior mediastinum (n = 1) and paratracheal region (n = 1). The most common indication was for microcalcifications concerning for papillary thyroid carcinoma on ultrasound (n = 6). Imaging findings included fusiform lesions with linear and punctuate bright echoes. The cytology evaluation showed small lymphocytes with discohesive epithelioid cells in most cases, and proteinaceous fluid in the cystic case. There were rare macrophages and Hassall's corpuscles. Flow cytometry and/or immunostains were performed in all cases, supporting thymic origin. Conclusion Ectopic thymic tissue is rarely present as a neck mass or thyroid nodule on FNA biopsy. The ultrasound imaging findings reveal a well-defined fusiform lesion with punctate bright echoes that could be misinterpreted as papillary thyroid carcinoma. The aspirates show a small lymphoid population, immunophenotypically compatible with thymic T-cells, in addition to scattered epithelial cells. Therefore, knowledge of the typical ultrasonographic and cytopathological features can help make a definitive diagnosis and avoid more invasive procedures in paediatric patients.

The pathology of histiocytic disorders and neoplasms of the macrophage-dendritic cell lineages is heterogeneous and may have overlapping features, all the while made more difficult by their rarity. The morphologic features together with immunophenotype and pattern of involvement should be taken together with the clinical and radiographic findings to make a unifying diagnosis. The “molecular microscope” of these lesions is further unraveling their biologic potential, which in some cases has helped to solidify their standing as inflammatory myeloid neoplasms. The histopathologic clues for the diagnosis of Langerhans cell histiocytosis (LCH), Erdheim-Chester Disease (ECD), the “juvenile xanthogranuloma” (JXG) family of lesions, Rosai-Dorfman disease (RDD), malignant histiocytoses including histiocytic sarcoma and Langerhans cell sarcoma, and other less common histiocytic-related disorders, including hemophagocytic lymphohistiocytosis (HLH), are reviewed in this chapter.

Background Histopathologic features of late graft dysfunction (LGD) in endomyocardial biopsies (EMBs) after pediatric heart transplantation (HT) have been incompletely described and rarely quantified. We employed automated, morphometric analysis of whole-slide EMB images to objectively quantify fibrosis and microvasculopathy after pediatric HT. Methods Nine recipients with clinical LGD were matched with controls on age, listing diagnosis, crossmatch and time since HT. Fibrosis was quantified as percent tissue area with fibrosis and capillary density as capillaries per unit area, number of capillary “neighbors” within 30 μm of each myocyte and myocyte-to-nearest-capillary diffusion distance. Clinical data, including all EMB reports, were also reviewed. Results The groups were well matched for age at HT (median 4.0 vs 3.1 years), listing diagnosis (50% congenital heart disease for each), positive crossmatch (11% each) and days post-HT (2,628 vs 2,894, p = 0.69). Despite a similar number of previous EMBs (median 23 each, p = 0.43), areas occupied by fibrosis were greater in LGD cases (44.5% vs 23.2%, p = 0.012). Capillary number/area data were not statistically different between LGD cases and controls (378/mm 2 vs 559/mm 2 , p = 0.57), but LGD cases more commonly had zero capillary neighbors (35% vs 20%, p = 0.02) and greater myocyte-to-nearest-capillary distances (27.1 μm vs 18.7 μm, p = 0.005). Cumulative rejection history correlated with fibrosis ( r = 0.49, p = 0.039) and myocyte-to-nearest-capillary distance ( r = 0.5, p = 0.036). Conclusions LGD after pediatric HT is associated with previous rejection and characterized histologically by fibrosis and microvasculopathy, which are not readily appreciated by traditional semi-quantitative EMB analysis. Software-assisted EMB analysis may enable greater pathophysiologic understanding of LGD and identification of targets for future study and intervention.

Rosai–Dorfman disease (RDD) is a benign non-Langerhans cell histiocytic disorder that usually presents with massive lymphadenopathy and sinus histiocytosis, although at least 40% of patients can have extranodal involvement. Recent studies found NRAS, KRAS, and ARAF mutations in a subset of patients with RDD, but no BRAF-V600E mutations have been reported yet. RDD is often associated with autoimmune or malignant disease, and rare cases of hereditary RDD have been reported as well. In most cases, RDD is self-limited and observation is the standard approach. However, 7–12% of patients die from the disease. Treatment is restricted to patients with life-threatening, multiple-relapsing, or autoimmune-associated disease.

Langerhans cell histiocytosis (LCH) can affect virtually any organ of the human body, including the central nervous system (CNS). Based on correlation of clinical, imaging, and pathology findings, two patterns of CNS-LCH have been identified: granulomatous (“tumorous”) and non-granulomatous “neurodegenerative” (ND)-CNS-LCH. Granulomatous CNS-LCH can manifest with polyuria and polydipsia, seizures, increased intracranial pressure, or other signs, depending on location. The typical location on MRI is extra-axial (hypothalamic-pituitary region, meninges, pineal gland, choroid plexus), and pathology in a proliferative phase reveals CD1a/langerin+ histiocytes. ND-CNS-LCH manifests insidiously with cerebellar and bulbar symptoms. MRI typically reveals parenchymal signal alterations in the cerebellum, pons, basal ganglia, and supratentorial white matter consistent with neuronal loss and demyelination. Biopsies of such lesions are rare. In the few published cases biopsied, variable pathologies are described with perivascular inflammatory changes, variable neuronal loss, demyelination, and gliosis, but are nondiagnostic for CD1a+/CD207+ LCH cells. Granulomatous LCH can be responsive to chemotherapy agents such as vinblastine, cytarabine, or cladribine, while the optimal treatment of ND-CNS-LCH is yet to be defined.

Autoinflammatory diseases are caused by pathologic activation of the innate immune system. Primary hemophagocytic lymphohistiocytosis (HLH) is an aggressive syndrome of excessive immune activation caused by monogenic mutations resulting in cytotoxic cell defects and subsequent failure to eliminate activated macrophages. Secondary HLH is often diagnosed in cases without a known Mendelian inheritance. However, some cases of “secondary” HLH have been shown to harbor mutations with partial dysfunction of the cytotoxic system. Recently, macrophage intrinsic abnormalities caused by NLRC4 inflammasome mutations have been linked to autoinflammation and recurrent macrophage activation syndromes resembling a primary HLH. We report a case of a former 28-week preterm infant with congenital anemia, ascites, and a heavy edematous placenta with fetal thrombotic vasculopathy, who developed hepatosplenomegaly and unexplained systemic inflammation with laboratory features of HLH in the early postnatal course and died at 2 months of age. Postmortem examination confirmed the hepatosplenomegaly with marked sinusoidal hemophagocytosis, along with striking hemophagocytosis in the bone marrow and lymph nodes. There was extensive acute and chronic ischemic bowel disease with matted bowel loops, fibrous adhesions, and patchy necrotizing enterocolitis features. Whole exome sequencing analysis demonstrated a novel mosaic heterozygous NLRC4 512 C> T (p.Ser171Phe) de novo mutation predicated to cause a dominant, gain-of-function mutation resulting in a constitutively active protein. The assembly of NLRC4-containing inflammasomes via an induced self-propagation mechanism likely enables a perpetuating process of systemic macrophage activation, presumed to be initiated in utero in this patient.