Foundation-OneR-Heme Next Generation Sequencing of Hairy Cell Leukemia Variant Lymphocytes
Authors: Pawloski John R. , Randolph Brion , Bajic Paulie 
Published: October 2019

Abstract
Hairy Cell Leukemia variant (HCL-V) is a rare disorder of small, mature, B-cell lymphocytes, and is characterized clinically by splenomegaly, an elevated white blood cell (WBC) count, and a hypercellular bone marrow. It accounts for 0.4% of chronic lymphoid and 10% of all cases of hairy cell leukemia (HCL) cases, respectively, and is now considered by the World Health Organization (WHO) to be a distinct provisional entity, biologically unrelated to classical HCL (HCL-C). The median survival is 9 years, with 42% of patients dying of unrelated causes. Transformation to large cell lymphoma is seen in 6% of patients. Whereas HCL-C patients are typically middle-aged males, who manifest lymphocytosis, splenomegaly, and cytopenias, with malignant lymphocytes that express CD103 and CD25, the median age of HCL-V patients is 71 years with no gender preference, and whose cells are characterized by expression of CD103, but lacking CD25. HCL-V also has a more aggressive clinical course, with far lower response rates to purine nucleoside analogs, but better responses to anti-CD20 and anti-CD22 immunotherapies. The BRAF-V600E mutation is considered the molecular hallmark of HCL-C, while 16% have recurrent inactivating mutations of the cell cycle inhibitor CDKN1B, making it the second most common mutation in HCL-C. In contrast, while del17p13 and mutations of the TP53 and MAP2K1 genes are frequent in HCL-V, there are no defining chromosomal or genetic abnormalities, and little is known about the genetic underpinnings of this rare condition.
We evaluated a 50 year-old African-American male who was referred to us for an incidental asymptomatic lymphocytosis of 11,990/uL. Flow cytometry showed these monoclonal B lymphocytes to express CD103, CD11c (bright), CD20, CD22, and CD52, but they did not express CD5, CD10, or CD25, with a chronic lymphocytic leukemia (CLL) immunophenotypic (Matutes) score of 0, consistent with HCL-V. A CCND/IgVH translocation was seen in 5% of these cells, and the IgVH gene was mutated at a rate of 7.2%. The patient had a normal hemoglobin and platelet count, and a computed tomography (CT) scan with contrast showed no adenopathy or splenomegaly. Foundation One' Heme Next Generation Sequencing (NGS) was performed on this patient's peripheral blood lymphocytes, and revealed several variants of unknown significance (VUS) including ALK P1599H, BRD4 Q969_L9, CARD11 D764G, CBL L43_S44, CHEK2 D438Y, FANCC Q465R, FANCL S82R, FOX03 P138L, GNAS T398M, GSK3B R396Q, HDAC7 T161M, MET V919I, PIM1 E142D, SOX2 T232A, SYK Y348C, TRAF2 R372H, and YY1AP1 S47P. Notably, no abnormalities in the BRAF, TP53, MAP2K1, and CCND family of genes were observed.
A search of multiple literature and genetic databases for the gene sequence variants listed above was performed to assess for their known correlations to lymphoproliferative malignancies, especially HCL-V and HCL-C. These databases included PubMed (http://www.ncbi.nlm.nih.gov/pubmed), Genetics Home Reference (https://ghr.nlm.nih.gov/) Cancer Genetics Web (http://www.cancer-genetics.org/), and Gene Cards (http://www.genecards.org/). Mutations or translocations in the genes ALK, CARD11, HDAC7, MET, leukemia, Burkitt's lymphoma and Hodgkin's disease leukemia, Burkitt's lymphoma and Hodgkin's disease leukemia, Burkitt's lymphoma and Hodgkin's diseasePIM1, SOX2, and SYK, were found to have clear associations with various lymphoid malignancies, including large B cell lymphoma (CARD11, PIM1), Burkitt lymphoma (HDAC7, MET), anaplastic large cell lymphoma (ALK, SOX2), Hodgkin lymphoma (MET), mantle cell lymphoma (SOX2), pro-B cell ALL (HDAC7), and rare peripheral T cell lymphomas (SYK). Alterations in CARD11 have also been associated with congenital B cell lymphocytosis. However, none of the specific variants/mutations identified by NGS of our patient's HCL-V cells have previously been described to be associated with either HCL-V or HCL-C. These results may thus help to elucidate the molecular pathogenesis of this rare hematologic malignancy, and emphasize the need to apply this technology to a greater number of HCL-V cases to assess for recurring genetic variations that may distinguish this unique disorder.

Disclosures
No relevant conflicts of interest to declare.