Germline heterozygous SH2B3‐mutations and (idiopathic) erythrocytosis: Detection of a previously undescribed mutation

Abstract Erythrocytosis or polycythemia refers to a true or apparent increase in hemoglobin or hematocrit. When no etiology of erythrocytosis is identified, people are diagnosed with “idiopathic erythrocytosis” (IE). The identification of new contributing genes has recently improved the diagnostic workup of IE. As such mutations within the SH2B3 gene, which codes for the LNK protein and negatively regulates the JAK‐STAT pathway, have been identified in cases diagnosed as IE. This reports describes the presence of a previously undescribed germline SH2B3 variant p.(Thr335ArgfsTer4) within IE and emphasizes the advantages of gene panel sequencing as second step in the diagnostic work‐up.

by hypoxia, erythropoietin (EPO)-producing neoplasms or medication such as testosterone.Most primary cases are acquired, by example polycythemia vera (PV) in which approximately 95% of cases carry the JAK2 p.Val617Phe mutation.The Janus kinase-signal transducer and activator of transcription signaling pathway (JAK-STAT) is important in erythroid proliferation in response to EPO signaling.Inherited mutations may cause erythrocytosis by impacting hypoxia sensing, JAK-STAT signaling or epigenetic mechanisms [1,3].Identification of new causal/contributing genes improves the diagnostic workup of "idiopathic erythrocytosis" (IE).We report a new germline SH2B3 Somatic mutations of SH2B3 occur in 5%-7% of MPN patients and are associated with leukemic transformation [5,10].Most SH2B3 mutations occur in exon 2 (residues Glu208-Asp234), which codes for the PH-domain, residue 208 is considered as a mutational hotspot (p.Glu208Gln) [7,11].This is the first report describing the germline SH2B3 mutation p.(Thr335ArgfsTer4) in IE.Residue 335 is coded by exon 5.The Varsome database classifies this variant as "likely pathogenic," it is not found in the COSMIC and Human Gene Mutation Database.As in our case most SH2B3 mutations appear to be heterozygous, whether these contribute through haploinsufficiency or a dominant-negative effect is unknown [5].The lack of in vitro proof of concept of this variant could be considered as a limitation of this report.It is not explained why the mother has no signs of erythrocytosis, a synergistic effect of the patient's active smoking or other genes cannot be excluded.The patient carries mutated JAK3 and RELN with similar VAF.JAK3 belongs to the JAK-family of proteins and is mainly restricted to the hematopoietic lineage.LNK interacts with JAK3 and may serve as a scaffold enabling JAK3 autophosphorylation in the absence of a cytokine receptor.LNK mutations as Glu208Gln result in augmented phosphorylation [12].Mutated RELN, which encodes for proteins functioning in cellular interaction, is reported as megakaryocyte-unique somatic mutation in patients with MPN and may predispose to megakaryocytic clustering.
No clustering was present in the bone marrow of this patient.Mutated RELN is reported in autism spectrum disorder, as in our case, and in epilepsy [13,14].The potential role of these mutations in IE is unclear.
By following treatment guidelines we initiated low dose acetylsalicylic acid and performed venesection.An arbitrary hematocrit target of ≤55% is used; ≤45% is considered in patients with history of erythrocytosis-related thrombosis.Smoking cessation was advised [15].Hematocrit declined to ≤55% after four venesections performed within 7 weeks, thrombocyte count normalized (Supplemental Figure 1).
Although no specific phenotype is associated with mutated SH2B3 there is growing interest in its role in myeloproliferative disorders.This report supports gene panel sequencing as a second step in the diagnostic work-up of erythrocytosis as it is able to open new diagnostic opportunities of potential germline associated conditions [7].Identifying the genomic landscape in erythrocytosis will hopefully result in better risk-stratification and treatment optimization.

F I G U R E 1
Graphical representation of SH2B3 and the interaction with JAK-STAT.(A) The SH2B adaptor protein 3 gene, located on chromosome 12, codes for the lymphocyte adaptor protein (LNK), also known as SH2B3.This adaptor family also contains SH2B1 and SH2B3.The Src homology 2 (SH2) domain-containing adaptor family shows the same protein architecture, existing out of an N-terminal dimerization domain (DD), pleckstrin homology domain (PH) and a C-terminal SH2 domain.(B) LNK may directly inhibit the Janus Kinase-signal transducer and activator of transcription (JAK-STAT) signaling pathway.Other interacting proteins (semi-transparent color) such as phosphatases and suppressor of cytokine signaling (SOCS) are not discussed in this article.(Figure extracted from: Morris R, Butler L, Perkins A, Kershaw NJ, Babon JJ.The role of lnk (Sh2b3) in the regulation of jak-stat signaling in haematopoiesis.Pharmaceuticals 2022;15.https://doi.org/10.3390/ph15010024).

Blood tests at presentation Blood test (general) Blood test (specific) Value Reference range
Blood results at time of presentation.
eJHaem.2023;4:1143-1147.wileyonlinelibrary.com/journal/jha21143 TA B L E 1 Abbreviations: ALT, alanine aminotransferase; aPTT, activated partial thromboplastin time; AST, aspartate aminotransferase; DHEAS, dehydroepiandrosterone sulfate; GGT, gamma-glutamyl transferase; HBV, hepatitis B virus; HIV, human immunodeficiency virus; IgA, immunoglobulin A; IgG, immunoglobulin G; IgM, immunoglobulin M; INR, international normalized ratio.variant(p.Thr335ArgfsTer4) for the first time in erythrocytosis.The SH2B adaptor protein 3 gene (SH2B3) codes for the lymphocyte adaptor protein (LNK), which negatively regulates the JAK-STAT pathway [4-7].A 31-years old Caucasian male was referred because of polycythemia and mild thrombocytopenia (pseudothrombopenia excluded).He experienced no complaints of vasomotor symptoms, pruritus or B-symptoms.Blood oxygen levels were normal.He was an active smoker; there was no alcohol abuse or use of medication.There were hematological disorders known within family history.His medical history included autism spectrum disorder; there were no thromboembolic events.Clinical investigation was unremarkable, body-mass index was 24.6 kg/m 2 .Biochemical analysis (Table1) showed a hemoglobin and hematocrit value of respectively 242 g/L (reference range (RR):135-175 g/L) and 70.9% (RR:41.0-53.0%).Thrombocytes were 72 × 10ˆ9/L (RR:150-450 × 10ˆ9/L).Levels of testosterone and EPO resulted normal.Venous p50 was 25.1 mmHg.Thoraco-abdominal computed tomography and karyotyping (46,XY [10]) resulted normal.Bone marrow biopsy showed no signs of myeloproliferative neoplasms (MPN) according to current WHO-criteria.JAK2 p.Val617Phe, exon12 and other variants were absent.No mutations in the EPOR, VHL, erozygous carriership of mutated SH2B3.Paternal genetic analysis was not performed as contact was lost since several years.The SH2B3 gene codes for the LNK protein (figure 1) and is predominantly expressed in hematopoietic cells.LNK is member of the SH2-domain-containing adaptor family of proteins which exist out of a N-terminal dimerization domain, a central pleck-SH2B3 is described in MPN and nonmalignant diseases as IE.IE compromises a heterogenous group of disorders without evidence for MPN.MPNs are subdivided as BCR::ABL-positive or -negative, respectively indicating the presence/absence of a reciprocal translocation between chromosome 9 and 22.Typical chronic myeloid leukemia is BCR::ABL-positive.BCR::ABL-negative MPN are generally classified as PV, essential thrombocythemia and primary myelofibrosis.