Pediatric autoimmune myelofibrosis: Experience from a large pediatric tertiary care center.

Autoimmune myelofibrosis (AIMF) is a rare disorder characterized by cytopenias and autoimmunity, with characteristic bone marrow findings that include lymphocytic infiltration and fibrosis. AIMF is described predominantly in adult populations who have systemic lupus erythematosis (SLE), with scant pediatric cases described mainly in older adolescents with SLE. Here, we described the largest single-center pediatric experience of pediatric autoimmune myelofibrosis (PAIMF) series, demonstrating both similarities and distinctions from the adult experience. Patients overall respond well to steroid therapy, but these patients were significantly younger, infrequently carried a diagnosis of SLE, and causative genetic lesions were identified in many cases.

Expansion of germline RPS20 mutation phenotype to include Diamond-Blackfan anemia.

Diamond-Blackfan anemia (DBA) is a ribosomopathy of variable expressivity and penetrance characterized by red cell aplasia, congenital anomalies, and predisposition to certain cancers, including early-onset colorectal cancer (CRC). DBA is primarily caused by a dominant mutation of a ribosomal protein (RP) gene, although approximately 20% of patients remain genetically uncharacterized despite exome sequencing and copy number analysis. Although somatic loss-of-function mutations in RP genes have been reported in sporadic cancers, with the exceptions of 5q-myelodysplastic syndrome (RPS14) and microsatellite unstable CRC (RPL22), these cancers are not enriched in DBA. Conversely, pathogenic variants in RPS20 were previously implicated in familial CRC; however, none of the reported individuals had classical DBA features. We describe two unrelated children with DBA lacking variants in known DBA genes who were found by exome sequencing to have de novo novel missense variants in RPS20. The variants affect the same amino acid but result in different substitutions and reduce the RPS20 protein level. Yeast models with mutation of the cognate residue resulted in defects in growth, ribosome biogenesis, and polysome formation. These findings expand the phenotypic spectrum of RPS20 mutation beyond familial CRC to include DBA, which itself is associated with increased risk of CRC.

Pediatric myeloid sarcoma: a single institution clinicopathologic and molecular analysis.

Myeloid sarcoma (MS) is a neoplastic condition composed of immature myeloid cells involving an extramedullary site. We investigated underlying chromosomal and molecular alterations to assess potential molecular markers of prognosis and outcome in this rare pediatric disease. We conducted a retrospective review of clinicopathologic and cytogenetic data from 33 pediatric patients with MS (ages 1 month-18 years) at our institution over a 32 year period (1984-2016). Tissue-based cancer microarray and targeted next-generation sequencing analysis were performed on six cases. The median age at diagnosis was 2.8 years with a male-to-female ratio of 2.6:1. MS is commonly presented with concomitant marrow involvement (n = 12, 36.4%) or as a recurrence of acute myeloid leukemia (AML; n = 14, 42.4%). The skin (n = 18, 54.5%) and soft tissue (n = 9, 27.3%) were the most common sites of involvement. Twenty-one of 25 samples (84.0%) harbored chromosomal aberrations; KMT2A alterations (n = 10, 40.0%) or complex cytogenetics (n = 7, 28.0%) were most frequent. Mutations in RAS, tyrosine kinase, cell signaling, and chromatin remodeling genes were detected. When compared to pediatric patients with AML without extramedullary involvement (EMI), inferior overall survival (OS) was observed (18.8 months vs. 89.3 months, p = .008). Pediatric patients with MS with non-favorable cytogenetics [abnormalities other than t(8;21), inv(16)/t(16;16), or t(15;17)] had a significantly lower OS compared to patients with AML with non-favorable cytogenetics and no extramedullary involvement (8.0 months vs. 28.1 months, p < .001). Pediatric MS is a rare disease with diverse clinical presentations. Non-favorable cytogenetics may be a poor prognostic marker for pediatric patients with MS and molecular diagnostics can assist with risk stratification and identify potentially actionable targets.

A clinicopathologic study of the spectrum of systemic forms of EBV-associated T-cell lymphoproliferative disorders of childhood: A single tertiary care pediatric institution experience in North America.

BACKGROUND: Systemic forms of EBV-associated T-cell lymphoproliferative disorders of childhood (S-EBV-T-LPD) comprise three major forms: EBV-positive hemophagocytic lymphohistiocytosis (EBV-HLH), systemic EBV-positive T-cell lymphoma (S-EBV-TCL), and systemic chronic active EBV infection (S-CAEBV). These disorders occur rarely in children in Western countries. Here, we described eight children of such entities. DESIGN: Eight cases (six clinical and two autopsy) with S-EBV-T-LPD of childhood were retrospectively identified from 1990 to 2015. Clinicopathologic parameters including histomorphology, immunophenotype, EBV studies, and T-cell receptor gene rearrangement studies were recorded. RESULTS: Patients include five females and three males of Hispanic, Asian, and Caucasian origins with an age range of 14 months to 9 years. Fever, hepatosplenomegaly, cytopenias, abnormal EBV serologies, and very high EBV viral loads were common findings. Histologic findings showed EBV+ T-cell infiltrates with variable degrees of architectural distortion and cytologic atypia ranging from no to mild cytologic atypia to overt lymphoma and tissue hemophagocytosis. All showed aberrant CD4+ or CD8+ T cells with dim to absent CD5, CD7, and CD3, and bright CD2 and CD45 by flow cytometry or loss of CD5 by immunohistochemistry. TCR gene rearrangement studies showed monoclonal rearrangements in all clinical cases (6/6). Outcomes were poor with treatment consisting of chemotherapy per the HLH-94 or HLH-2004 protocols with or without bone marrow transplant. CONCLUSION: In this large pediatric clinicopathologic study of S-EBV-T-LPD of childhood in the United States, EBV-HLH, S-EBV-TCL, and S-CAEBV show many overlapping features. Diagnosis is challenging, and overall outcome is poor using current HLH-directed therapies.

Steroid-responsive atypical marginal zone hyperplasia of the lip in a child.

Atypical marginal zone hyperplasia (AMZH) is a recently described disease entity seen mainly in children. AMZH most commonly affects tonsils and appendices. Cutaneous AMZH is rare. The authors report here a recurrent AMZH in the lip of a 9-year-old child who presented originally with a lip swelling for approximately 3 months. The lip lesion recurred after each incomplete excision for 4 times. Pathologically, the lesion demonstrated marginal zone B-cell hyperplasia with kappa monoclonality by flow cytometry and immunohistochemistry studies. Lymphoepithelial lesions were noted with involvement of minor salivary glands. Polymerase chain reaction for immunoglobulin heavy-chain gene rearrangement has been repeatedly negative. Polymerase chain reaction for Borrelia species DNA was negative on both paraffin-embedded tissue and plasma. Serum antibodies IgG and IgM for Helicobacter Pylori were positive. A diagnosis of AMZH was made. Two courses of anti H. Pylori therapy did not improve the lip lesion, which completely regressed after a course of prednisone therapy. With differential diagnosis of cutaneous marginal zone lymphoma, the case illustrated diagnostic challenges, especially with recurrent lesions. This is the first case of recurrent cutaneous AMZH that has uncharacteristic kappa light-chain restriction. AMZH should be considered in children with mucocutaneous lesions with features of marginal zone lymphoma.

Therapeutic implications of CD1d expression and tumor-infiltrating macrophages in pediatric medulloblastomas.

Immunobiology of medulloblastoma (MB), the most common malignant brain tumor in children, is poorly understood. Although tumor cells in some MBs were recently shown to express CD1d and be susceptible to Vα24-invariant natural killer T (NKT)-cell cytotoxicity, the clinical relevance of CD1d expression in MB patients remains unknown. We investigated the expression of CD1d in pediatric MBs and correlated with molecular and clinical characteristics. Specifically, we explored if NKT cell therapy can be targeted at a subset of pediatric MBs with poorer prognosis. Particularly, infantile MBs have a worse outcome because radiotherapy is delayed to avoid neurocognitive sequelae. Immunohistochemistry for CD1d was performed on a screening set of 38 primary pediatric MBs. Gene expression of the membrane form of M2 macrophage marker, CD163, was studied in an expanded cohort of 60 tumors. Outcome data was collected prospectively. Thirteen of 38 MBs (34.2 %) expressed CD1d on immunohistochemistry. CD1d was expressed mainly on MB tumor cells, and on some tumor-associated macrophages. Majority (18/22, 82 %) of non sonic-hedgehog/Wingless-activated MBs (group 3 and 4) were CD1d-negative (p = 0.05). A subset of infantile MBs (4/9, 44.4 %) expressed CD1d. Macrophages infiltrating MB expressed CD163 apart from CD1d. Molecular subtypes demonstrated statistical differences in CD163 expression, SHH-tumors were the most enriched (p = 0.006). Molecular and clinical subtypes of pediatric MB exhibit distinct differences in CD1d expression, which have important therapeutic implications. High CD1d expression in infantile MBs offers potential new immunotherapeutic treatment with NKT cell therapy in infants, where treatment is suboptimal due delayed radiotherapy.

Myelodysplastic neoplasms evolving from inherited bone marrow failure syndromes / germline predisposition syndromes: Back under the microscope.

Inherited bone marrow failure syndromes and germline predisposition syndromes (IBMFS/GPS) are associated with increased risk for hematologic malignancies, particularly myeloid neoplasms, such as myelodysplastic neoplasms (MDS) and acute myeloid leukemia (AML). The diagnosis of MDS in these syndromes poses difficulty due to frequent bone marrow hypocellularity and the presence of some degree of dysplastic features related to the underlying germline defect causing abnormal maturation of one or more cell lines. Yet, the diagnosis of MDS is usually associated with a worse outcome in several IBMFS/GPS. Criteria for the diagnosis of MDS in IBMFS/GPS have not been standardized with some authors suggesting a mixture of morphologic, cytogenetic, and genetic criteria. This review highlights these challenges and suggests a more standardized approach to nomenclature and diagnostic criteria.

The 5th edition of the World Health Organization Classification of Haematolymphoid Tumours: Myeloid and Histiocytic/Dendritic Neoplasms.

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.

Utility of Flow Cytometry of Bronchoalveolar Lavage in the Diagnosis of T and NK Cell Malignancies in Critically Ill Patients.

OBJECTIVE: The value of bronchoalveolar lavage (BAL) in the diagnosis of pulmonary diseases of diverse etiologies is widely accepted. Cytospin and cell-block preparations for cytomorphological (CM) evaluation and for immunohistochemical studies are the standard method to evaluate BAL, though it may be time-consuming. Flow cytometric (FC) evaluation, on the other hand, has a short turnaround time, and is a useful methodology to differentiate reactive processes from hematological neoplasms, or detect a small aberrant population in an inflammatory background. BAL specimens provide an excellent source for FC studies. CASE REPORTS: We describe two cases of critically ill patients with no history of hematolymphoid neoplasms, who presented with non-specific symptoms. Abnormal pulmonary imaging studies prompted bronchoscopic evaluation and collection of BAL during the initial evaluation. FC analysis of the BAL fluid aided to the early diagnosis of aggressive NK cell leukemia and adult T cell leukemia/lymphoma, respectively. CONCLUSIONS: Flow cytometric immunophenotyping in addition to the CM assessment increases the diagnostic value and provides timely diagnosis from BAL specimens, which is especially important for critically ill patients.

Inherited Bone Marrow Failure Syndromes: Biology and Diagnostic Clues.

Inherited bone marrow failure syndromes are a group of genetic disorders associated with bone marrow production defects resulting in single or multiple cytopenias. Many of these disorders predispose the patient to hematologic and nonhematologic malignancies, requiring life-long follow-up. A positive family history of hematologic disorders or malignancies is frequent, as these disorders commonly run in families, and selection of family members as potential bone marrow donors should be performed with caution to avoid transplanting potentially defective stem cells. This review highlights the most common genetic disorders associated with bone marrow failure.

Hemoglobin fractionation by Sebia Capillarys 2 Flex Piercing System as primary method for evaluation of hemoglobinopathies.

BACKGROUND: Hemoglobin fractionation by capillary zone electrophoresis (CE) is becoming a popular method for the identification of hemoglobin variants that can cause hemoglobinopathies. The goal of this study was to compare the performance of capillary electrophoresis using Sebia Capillarys 2 Flex Piercing system (CE-S) with high-pressure liquid chromatography (HPLC) using Primus Ultra2 Resolution Variants System (HPLC-P) as a primary method in hemoglobinopathy work-up. METHODS: A total of 306 blood specimens submitted for evaluation of hemoglobinopathies were studied using HPLC-P and CE-S. RESULTS: The reference ranges for Hb A, A2 and F agreed well between methods. All common variants containing Hb S and Hb C were detected by both methods. Quantification of Hb A2 with HPLC-P required a correction in the presence of Hb S, while quantification of Hb A2 was slightly overestimated by CE-S in the presence of Hb C. Of 41 samples containing other variants, 2 were not identified by HPLC-P and 3 were not identified by CE-S. CONCLUSION: CE-S provides comparable information to that obtained by HPLC-P and it is a reliable primary method for the evaluation of hemoglobin variants.

Maximizing the potential of aggressive mouse tumor models in preclinical drug testing.

Atypical teratoid rhabdoid tumor (ATRT) is an aggressive embryonal brain tumor among infants and young children. Two challenges exist for preclinical testing in ATRT. First, genetically quiet, ATRT is a difficult tumor to target molecularly. Tumor cells need to divide to propagate tumor growth-intercepting the common crossroads in cell cycle progression is a feasible strategy. KIF11 is needed for bipolar spindle formation in metaphase. We identified KIF11 as a universal target of all ATRT-molecular-subtypes. Ispinesib, a KIF11-inhibitor, effectively inhibited tumor proliferation in all seven cell lines. A second challenge-a major challenge in preclinical drug testing in-vivo among aggressive tumor models, is the narrow therapeutic window to administer drugs within the limited murine lifespan. Our most aggressive ATRT tumor model was lethal in all mice within ~ 1 month of tumor implantation. Such short-surviving mouse models are difficult to employ for preclinical drug testing due to the narrow time window to administer drugs. To overcome this time restriction, we developed a clinical staging system which allowed physically-fit mice to continue treatment, in contrast to the conventional method of fixed drug-dose-duration regimen in preclinical testing which will not be feasible in such short-surviving mouse models. We validated this approach in a second embryonal brain tumor, medulloblastoma. This is a clinically relevant, cost-efficient approach in preclinical testing for cancer and non-cancer disease phenotypes. Widely used preclinical mouse models are not the most accurate and lack the aggressive tumor spectrum found within a single tumor type. Mice bearing the most aggressive tumor spectrum progress rapidly in the limited murine life-span, resulting in a narrow therapeutic window to administer drugs, and are thus difficult to employ in preclinical testing. Our approach overcomes this challenge. We discovered ispinesib is efficacious against two embryonal brain tumor types.

The emerging spectrum of cardiopulmonary pathology of the coronavirus disease 2019 (COVID-19): Report of 3 autopsies from Houston, Texas, and review of autopsy findings from other United States cities.

This paper collates the pathological findings from initial published autopsy reports on 23 patients with coronavirus disease 2019 (COVID-19) from 5 centers in the United States of America, including 3 cases from Houston, Texas. Findings confirm that COVID-19 is a systemic disease with major involvement of the lungs and heart. Acute COVID-19 pneumonia has features of a distinctive acute interstitial pneumonia with a diffuse alveolar damage component, coupled with microvascular involvement with intra- and extravascular fibrin deposition and intravascular trapping of neutrophils, and, frequently, with formation of microthombi in arterioles. Major pulmonary thromboemboli with pulmonary infarcts and/or hemorrhage occurred in 5 of the 23 patients. Two of the Houston cases had interstitial pneumonia with diffuse alveolar damage pattern. One of the Houston cases had multiple bilateral segmental pulmonary thromboemboli with infarcts and hemorrhages coupled with, in nonhemorrhagic areas, a distinctive interstitial lymphocytic pneumonitis with intra-alveolar fibrin deposits and no hyaline membranes, possibly representing a transition form to acute fibrinous and organizing pneumonia. Multifocal acute injury of cardiac myocytes was frequently observed. Lymphocytic myocarditis was reported in 1 case. In addition to major pulmonary pathology, the 3 Houston cases had evidence of lymphocytic pericarditis, multifocal acute injury of cardiomyocytes without inflammatory cellular infiltrates, depletion of splenic white pulp, focal hepatocellular degeneration and rare glomerular capillary thrombosis. Each had evidence of chronic cardiac disease: hypertensive left ventricular hypertrophy (420 g heart), dilated cardiomyopathy (1070 g heart), and hypertrophic cardiomyopathy (670 g heart). All 3 subjects were obese (BMIs of 33.8, 51.65, and 35.2 Kg/m2). Overall, the autopsy findings support the concept that the pathogenesis of severe COVID-19 disease involves direct viral-induced injury of multiple organs, including heart and lungs, coupled with the consequences of a procoagulant state with coagulopathy.

Clinical features and outcomes of patients with Shwachman-Diamond syndrome and myelodysplastic syndrome or acute myeloid leukaemia: a multicentre, retrospective, cohort study.

BACKGROUND: Data to inform surveillance and treatment for leukaemia predisposition syndromes are scarce and recommendations are largely based on expert opinion. This study aimed to investigate the clinical features and outcomes of patients with myelodysplastic syndrome or acute myeloid leukaemia and Shwachman-Diamond syndrome, an inherited bone marrow failure disorder with high risk of developing myeloid malignancies. METHODS: We did a multicentre, retrospective, cohort study in collaboration with the North American Shwachman-Diamond Syndrome Registry. We reviewed patient medical records from 17 centres in the USA and Canada. Patients with a genetic (biallelic mutations in the SBDS gene) or clinical diagnosis (cytopenias and pancreatic dysfunction) of Shwachman-Diamond syndrome who developed myelodysplastic syndrome or acute myeloid leukaemia were eligible without additional restriction. Medical records were reviewed between March 1, 2001, and Oct 5, 2017. Masked central review of bone marrow pathology was done if available to confirm leukaemia or myelodysplastic syndrome diagnosis. We describe the clinical features and overall survival of these patients. FINDINGS: We initially identified 37 patients with Shwachman-Diamond syndrome and myelodysplastic syndrome or acute myeloid leukaemia. 27 patients had samples available for central pathology review and were reclassified accordingly (central diagnosis concurred with local in 15 [56%] cases), 10 had no samples available and were classified based on the local review data, and 1 patient was excluded at this stage as not eligible. 36 patients were included in the analysis, of whom 10 (28%) initially presented with acute myeloid leukaemia and 26 (72%) initially presented with myelodysplastic syndrome. With a median follow-up of 4·9 years (IQR 3·9-8·4), median overall survival for patients with myelodysplastic syndrome was 7·7 years (95% CI 0·8-not reached) and 0·99 years (95% CI 0·2-2·4) for patients with acute myeloid leukaemia. Overall survival at 3 years was 11% (95% CI 1-39) for patients with leukaemia and 51% (29-68) for patients with myelodysplastic syndrome. Management and surveillance were variable. 18 (69%) of 26 patients with myelodysplastic syndrome received upfront therapy (14 haematopoietic stem cell transplantation and 4 chemotherapy), 4 (15%) patients received no treatment, 2 (8%) had unavailable data, and 2 (8%) progressed to acute myeloid leukaemia before receiving treatment. 12 patients received treatment for acute myeloid leukaemia-including the two patients initially diagnosed with myelodysplastic who progressed- two (16%) received HSCT as initial therapy and ten (83%) received chemotherapy with intent to proceed with HSCT. 33 (92%) of 36 patients (eight of ten with leukaemia and 25 of 26 with myelodysplastic syndrome) were known to have Shwachman-Diamond syndrome before development of a myeloid malignancy and could have been monitored with bone marrow surveillance. Bone marrow surveillance before myeloid malignancy diagnosis was done in three (33%) of nine patients with leukaemia for whom surveillance status was confirmed and 11 (46%) of 24 patients with myelodysplastic syndrome. Patients monitored had a 3-year overall survival of 62% (95% CI 32-82; n=14) compared with 28% (95% CI 10-50; n=19; p=0·13) without surveillance. Six (40%) of 15 patients with available longitudinal data developed myelodysplastic syndrome in the setting of stable blood counts. INTERPRETATION: Our results suggest that prognosis is poor for patients with Shwachman-Diamond syndrome and myelodysplastic syndrome or acute myeloid leukaemia owing to both therapy-resistant disease and treatment-related toxicities. Improved surveillance algorithms and risk stratification tools, studies of clonal evolution, and prospective trials are needed to inform effective prevention and treatment strategies for leukaemia predisposition in patients with Shwachman-Diamond syndrome. FUNDING: National Institute of Health.

Marginal zone variant of mantle cell lymphoma: CD5-negative cyclin D1-positive variant posing a diagnostic dilemma.

Described herein is an unusual case of mantle cell lymphoma (MCL) histologically mimicking marginal zone lymphoma (MZL). An 83-year-old man presented with multiple adenopathies and a hilar mass encroaching on the right lung. A transbronchial biopsy showed small blue cells suspicious for small cell carcinoma. On further analysis the cells were predominantly small cleaved and CD20 positive, suggesting follicular lymphoma, grade 2. An axillary lymph node biopsy showed germinal centers surrounded by monocytoid B cells. Flow cytometry was negative for CD5 and CD23 and the diagnosis of MZL was considered. Because of the aggressive clinical behavior, including extensive necrosis on imaging studies, immunohistochemistry for cyclin D-1 was performed and was positive. Bone marrow was extensively involved and it showed t(11;14), in addition to other complex cytogenetic abnormalities. Differentiating MCL from MZL has prognostic and therapeutic implications, particularly when considering the potential role of targeted therapy and cell cycle modulators.

Bone marrow cell cycle markers in inherited bone marrow failure syndromes.

Patients with inherited bone marrow failure syndromes (IBMFS) are at increased risk of acute myeloid leukemia (AML) and myelodysplastic syndrome (MDS), possibly related to cell cycle dysregulation. In a cross-sectional analysis of bone marrow from 77 IBMFS, 71 sporadic conditions (AML, MDS, acquired aplastic anemia) and 22 normal controls we found overexpression of p53 in IBMFS, AML, and MDS; of Ki-67 in IBMFS and AML; and of survivin in IBMFS compared with all other groups. The patterns of expression of cell cycle markers in IBMFS are thus distinct. Longitudinal studies will determine the diagnostic and prognostic significance of these findings.