Malignant Histiocytosis Comprises a Phenotypic Spectrum That Parallels the Lineage Differentiation of Monocytes, Macrophages, Dendritic Cells, and Langerhans Cells.

Malignant histiocytoses (MHs), or the 'M group' of the Histiocyte Society classification, are characterized by neoplastic histiocytes with large pleomorphic nuclei. MH encompasses the diagnoses of histiocytic sarcoma, interdigitating dendritic cell sarcoma, and Langerhans cell sarcoma. We aimed to define the phenotypic spectrum of MH and examine the genotypic features across this spectrum. Using immunohistochemistry, we arranged the 22 cases into 4 subtypes that correspond to the lines of differentiation from monocytic and dendritic cell precursors as follows: (1) macrophage (n = 5): CD68+, CD163+, CD14+, and Factor 13a+; (2) monocyte-macrophage (n = 5): CD68+, CD163+, CD14+, S100+, and OCT2+; (3) dendritic cell (n = 6): CD68+, CD11c+, S100+, lysozyme+, ZBTB46+, and CD1a/langerin < 5%; and (4) Langerhans cell (n = 6): CD68+, CD11c+, S100+, ZBTB46+, CD1a+, and langerin+. The phenotypic subtypes align with those seen in low-grade histiocytic neoplasms as follows: MH-macrophage type correlates with Erdheim-Chester disease phenotype; MH-monocyte-macrophage type with Rosai-Dorfman disease phenotype, and MH-Langerhans cell type with Langerhans cell histiocytosis. Activating mutations in MAPK-pathway genes were identified in 80% of MH cases; 29% had mutations in the PI3k-AKT-mTOR pathway and 59% had mutations in epigenetic modulating genes. Strong expression of cyclin D1 was present in all cases, whereas p-ERK and p-AKT were not uniformly expressed. Eight of 22 (36%) MH cases were proven to be clonally related to a prior B-cell lymphoma. Defining the phenotypic spectrum of MH provides a guide to diagnosis and allows further exploration into the potential biological and clinical significance.

Clinical Validation of a Targeted Next-Generation Sequencing Panel for Lymphoid Malignancies.

Lymphoid malignancies are a heterogeneous group of hematological disorders characterized by a diverse range of morphologic, immunophenotypic, and clinical features. Next-generation sequencing (NGS) is increasingly being applied to delineate the complex nature of these malignancies and identify high-value biomarkers with diagnostic, prognostic, or therapeutic benefit. However, there are various challenges in using NGS routinely to characterize lymphoid malignancies, including pre-analytic issues, such as sequencing DNA from formalin-fixed, paraffin-embedded tissue, and optimizing the bioinformatic workflow for accurate variant calling and filtering. This study reports the clinical validation of a custom capture-based NGS panel to test for molecular markers in a range of lymphoproliferative diseases and histiocytic neoplasms. The fully validated clinical assay represents an accurate and sensitive tool for detection of single-nucleotide variants and small insertion/deletion events to facilitate the characterization and management of patients with hematologic cancers specifically of lymphoid origin.

MICon Contamination Detection Workflow for Next-Generation Sequencing Laboratories Using Microhaplotype Loci and Supervised Learning.

Innovation in sequencing instrumentation is increasing the per-batch data volumes and decreasing the per-base costs. Multiplexed chemistry protocols after the addition of index tags have further contributed to efficient and cost-effective sequencer utilization. With these pooled processing strategies, however, comes an increased risk of sample contamination. Sample contamination poses a risk of missing critical variants in a patient sample or wrongly reporting variants derived from the contaminant, which are particularly relevant issues in oncology specimen testing in which low variant allele frequencies have clinical relevance. Small custom-targeted next-generation sequencing (NGS) panels yield limited variants and pose challenges in delineating true somatic variants versus contamination calls. A number of popular contamination identification tools have the ability to perform well in whole-genome/exome sequencing data; however, in smaller gene panels, there are fewer variant candidates for the tools to perform accurately. To prevent clinical reporting of potentially contaminated samples in small next-generation sequencing panels, we have developed MICon (Microhaplotype Contamination detection), a novel contamination detection model that uses microhaplotype site variant allele frequencies. In a heterogeneous hold-out test cohort of 210 samples, the model displayed state-of-the-art performance with an area under the receiver-operating characteristic curve of 0.995.

Follicular lymphoma and diffuse large B-cell lymphoma with BCL2 and IRF4 rearrangements in adult patients.

Follicular lymphoma (FL) and diffuse large B-cell lymphoma (DLBCL) with concurrent BCL2 and IRF4 rearrangements are rare. It is unclear whether such cases should be classified as large B- cell lymphoma with IRF4 rearrangement or FL/DLBCL-not otherwise specified. We identified 5 adult patients (FL, N = 3 and FL/DLBCL, N = 2) with concurrent BCL2 and IRF4 rearrangements. The median age at presentation was 77 years, and three patients presented with advanced stage disease. Both nodal and extranodal sites were involved and involvement was not limited to head and neck region. With a median follow-up of 18 months, 1 patient died and 4 patients were alive, including 3 who received chemotherapy and 1 who was observed. The neoplasms were histologically heterogeneous, including grade 2 and 3 FL and DLBCL. Four cases coexpressed CD10, BCL6, BCL2 and MUM1/IRF4. The Ki67 labelling index ranged from 20% to 95%. In 4 patients, the percentage of cells with BCL2 rearrangement was equal to or slightly greater than the cells harboring IRF4 rearrangement. Two cases underwent next generation sequencing tailored for lymphoid neoplasms. Both lacked mutations involving IRF4 and NF-kB pathway genes that are frequently detected in large B-cell lymphoma with IRF4 rearrangement, and one case showed DLBCL-EZH2 type mutations, including KMT2D and BCL2 mutations, similar to 2 previously reported DLBCL with BCL2 and IRF4 rearrangements. Adults with FL and FL/DLBCL with BCL2 and IRF4 rearrangements display clinicopathologic and mutational features more akin to FL and DLBCL and should not be characterized as large B-cell lymphoma with IRF4 rearrangement.