Involvement of the JAK-STAT pathway in the molecular landscape of tyrosine kinase fusion-negative hypereosinophilic syndromes: A nationwide CEREO study.

We investigated using a custom NGS panel of 149 genes the mutational landscape of 64 consecutive adult patients with tyrosine kinase fusion-negative hypereosinophilia (HE)/hypereosinophilic syndrome (HES) harboring features suggestive of myeloid neoplasm. At least one mutation was reported in 50/64 (78%) patients (compared to 8/44 (18%) patients with idiopathic HE/HES/HEUS used as controls; p < .001). Thirty-five patients (54%) had at least one mutation involving the JAK-STAT pathway, including STAT5B (n = 18, among which the hotspot N642H, n = 13), JAK1 (indels in exon 13, n = 5; V658F/L, n = 2), and JAK2 (V617F, n = 6; indels in exon 13, n = 2). Other previously undescribed somatic mutations were also found in JAK2, JAK1, STAT5B, and STAT5A, including three patients who shared the same STAT5A V707fs mutation and features consistent with primary polycythemia. Nearly all JAK-STAT mutations were preceded by (or associated with) myelodysplasia-related gene mutations, especially in RNA-splicing genes or chromatin modifiers. In multivariate analysis, neurologic involvement (hazard ratio [HR] 4.95 [1.87-13.13]; p = .001), anemia (HR 5.50 [2.24-13.49]; p < .001), and the presence of a high-risk mutation (as per the molecular international prognosis scoring system: HR 6.87 [2.39-19.72]; p < .001) were independently associated with impaired overall survival. While corticosteroids were ineffective in all treated JAK-STAT-mutated patients, ruxolitinib showed positive hematological responses including in STAT5A-mutated patients. These findings emphasize the usefulness of NGS for the workup of tyrosine kinase fusion-negative HE/HES patients and support the use of JAK inhibitors in this setting. Updated classifications could consider patients with JAK-STAT mutations and eosinophilia as a new "gene mutated-entity" that could be differentiated from CEL, NOS, and idiopathic HES.

World Health Organization and International Consensus Classification of eosinophilic disorders: 2024 update on diagnosis, risk stratification, and management.

DISEASE OVERVIEW: The eosinophilias encompass a broad range of non-hematologic (secondary or reactive) and hematologic (primary or clonal) disorders with the potential for end-organ damage. DIAGNOSIS: Hypereosinophilia (HE) has generally been defined as a peripheral blood eosinophil count greater than 1.5 × 109/L, and may be associated with tissue damage. After the exclusion of secondary causes of eosinophilia, diagnostic evaluation of primary eosinophilias relies on a combination of various tests. They include morphologic review of the blood and marrow, standard cytogenetics, fluorescence in situ hybridization, molecular testing and flow immunophenotyping to detect histopathologic or clonal evidence for an acute or chronic hematolymphoid neoplasm. RISK STRATIFICATION: Disease prognosis relies on identifying the subtype of eosinophilia. After evaluation of secondary causes of eosinophilia, the 2022 World Health Organization and International Consensus Classification endorse a semi-molecular classification scheme of disease subtypes. This includes the major category "myeloid/lymphoid neoplasms with eosinophilia and tyrosine kinase gene fusions" (MLN-eo-TK), and the MPN subtype, "chronic eosinophilic leukemia" (CEL). Lymphocyte-variant HE is an aberrant T-cell clone-driven reactive eosinophila, and idiopathic hypereosinophilic syndrome (HES) is a diagnosis of exclusion. RISK-ADAPTED THERAPY: The goal of therapy is to mitigate eosinophil-mediated organ damage. For patients with milder forms of eosinophilia (e.g., <1.5 × 109/L) without symptoms or signs of organ involvement, a watch and wait approach with close follow-up may be undertaken. Identification of rearranged PDGFRA or PDGFRB is critical because of the exquisite responsiveness of these diseases to imatinib. Pemigatinib was recently approved for patients with relapsed or refractory FGFR1-rearranged neoplasms. Corticosteroids are first-line therapy for patients with lymphocyte-variant HE and HES. Hydroxyurea and interferon-α have demonstrated efficacy as initial treatment and in steroid-refractory cases of HES. Mepolizumab, an interleukin-5 (IL-5) antagonist monoclonal antibody, is approved by the U.S Food and Drug Administration for patients with idiopathic HES. Cytotoxic chemotherapy agents, and hematopoietic stem cell transplantation have been used for aggressive forms of HES and CEL, with outcomes reported for limited numbers of patients. Targeted therapies such as the IL-5 receptor antibody benralizumab, IL-5 monoclonal antibody depemokimab, and various tyrosine kinase inhibitors for MLN-eo-TK, are under active investigation.

The international consensus classification of eosinophilic disorders and systemic mastocytosis.

Based on new data and increased understanding of disease molecular genetics, the international consensus classification (ICC) has made several changes in the diagnosis and classification of eosinophilic disorders and systemic mastocytosis. Myeloid/lymphoid neoplasms with eosinophilia (M/LN-eo) and gene rearrangements have been renamed as M/LN-eo with tyrosine kinase gene fusions (M/LN-eo-TK). The category has been expanded to include ETV6::ABL1 and FLT3 fusions, and to accept PCM1::JAK2 and its genetic variants as formal members. The overlaps and differences between M/LN-eo-TK and BCR::ABL1-like B-lymphoblastic leukemia (ALL)/de novo T-ALL sharing the same genetic lesions are addressed. Besides genetics, ICC for the first time has introduced bone marrow morphologic criteria in distinguishing idiopathic hypereosinophilia/hypereosinophilic syndrome from chronic eosinophilic leukemia, not otherwise specified. The major diagnostic criteria for systemic mastocytosis (SM) in the ICC remain largely based on morphology, but several minor modifications/refinements have been made in criteria related to diagnosis, subclassification, and assessment of disease burden (B- and C-findings). This review is to focus on the ICC updates related to these disease entities, illustrated through changes related to morphology, molecular genetics, clinical features, prognosis, and treatment. Two practical algorithms are provided in navigating through the diagnosis and classification systems of hypereosinophilia and SM.

Lymphomatoid papulosis associated with myeloid neoplasm with eosinophilia and FIP1L1::PDGFRA rearrangement: Successful imatinib treatment in two cases.

Lymphomatoid papulosis (LyP) is a benign condition, listed among primary cutaneous CD30+ lymphoproliferative disorders. Its typical picture consists of relapsing-remitting papular lesions and it can be encountered in the course of a hematologic disease, at times representing its first manifestation. Hypereosinophilic syndromes are a heterogeneous group of disorders characterized by persistent peripheral blood hypereosinophilia that may lead to life-threatening organ damage. Among eosinophilic disorders, the subtype identified as myeloid/lymphoid neoplasm with eosinophilia and tyrosine kinase gene fusions has aroused particular interest due to its excellent response to tyrosine kinase inhibitors, including imatinib. Here, we described the case of two 33-year-old men presenting with LyP and myeloid neoplasm with eosinophilia and FIP1L1::PDGFRA rearrangement who achieved complete clinical and molecular remission of both conditions a few months after starting imatinib.

Detection of FIP1L1-PDGFRA fusion gene-positive cells in the skin lesion of a patient with hypereosinophilic syndrome.

Hypereosinophilic syndrome (HES) is a heterogeneous group of diseases, characterized by persistent hypereosinophilia and end-organ damage. The FIP1L1-PDGFRA (F/P) fusion gene is found in 3-25% of patients with HES and is an oncogenic driver of myeloid neoplasms with clonal eosinophilia. Although cutaneous symptoms are the most common type of symptom in patients who have F/P fusion gene-positive HES (F/P HES), histological reports are limited. We herein present the case of a 78-year-old man with erythematous macules and severe pruritus on his trunk and extremities. Laboratory investigations revealed marked eosinophilia and elevated serum vitamin B12. A histological examination showed massive infiltration of eosinophils and mast cells around the vessels in the upper dermis. Fluorescence in situ hybridization revealed F/P fusion genes in nuclei in the peripheral blood and the skin lesion. The patient was diagnosed with F/P HES, and showed an excellent clinical and haematological response to imatinib.

Diagnostic spectrum of hypereosinophilia based on bone marrow pathology: 10 years' experience at a tertiary care hospital.

BACKGROUND: Hypereosinophilia (HE) is defined as peripheral blood (PB) eosinophil count exceeding 1.5 × 109 /L. As the causes of HE can be diverse, the work-up of patients was complicated. In this study, we aimed to categorize the underlying diseases associated with HE and demonstrate minimum diagnostic approach. METHODS: Cases presenting with HE within 7 days of bone marrow (BM) examination conducted between 2008 and 2019 were selected. Cases were classified by the revised 2022 WHO and ICC classification. We also assessed morphologic features of unclassified persisting HE (>4 weeks) patients according to the morphologic criteria suggested a previous study by Wang et al. RESULTS: A total of 364 patients were included. The work-up confirmed primary HE in 38.7%, secondary HE in 48.9%, HE patients with insufficient evaluation in 13.7%. When conducted a slide review of HE patients with sustained HE more than 4 weeks among HE patients with insufficient evaluation, the morphological features showed abnormal eosinophils in PB/BM (69.0%/81.0%), hypercellularity (26.2%), myelofibrosis (7.1%), increased M:E ratio (5.3%), and dysmegakaryopoiesis (4.8%). Of these patients, 14 patients who met all morphologic criteria were suspected of CEL. CONCLUSIONS: This study demonstrates that HE is associated with variable conditions. BM morphological assessment based on a robust criterion can help to confirm a MN irrespective of the presence of clonal markers. The work-up of patients in whom ruled out the common secondary causes of HE requires a systematic but sufficient approach including at a minimum BM karyotyping, PDGFRA testing, lymphocyte immunophenotyping and TCR gene rearrangement.

Updates on eosinophilic disorders.

This review addresses changes and updates in eosinophilic disorders under the International Consensus Classification (ICC). The previous category of myeloid/lymphoid neoplasm with eosinophilia (M/LN-eo) and a specific gene rearrangement is changed to M/LN-eo with tyrosine kinase gene fusions to reflect the underlying genetic lesions. Two new members, M/LN-eo with ETV6::ABL1 fusion and M/LN-eo with various FLT3 fusions, have been added to the category; and M/LN-eo with PCM1::JAK2 and its genetic variants ETV6::JAK2 and BCR::JAK2 are recognized as a formal entity from their former provisional status. The updated understanding of the clinical and molecular genetic features of PDGFRA, PDGFRB and FGFR1 neoplasms is summarized. Clear guidance as to how to distinguish these fusion gene-associated disorders from the overlapping entities of Ph-like B-acute lymphoblastic leukemia (ALL), de novo T-ALL, and systemic mastocytosis is provided. Bone marrow morphology now constitutes one of the diagnostic criteria of chronic eosinophilic leukemia, NOS (CEL, NOS), and idiopathic hypereosinophilia/hypereosinophilic syndrome (HE/HES), facilitating the separation of a true myeloid neoplasm with characteristic eosinophilic proliferation from those of unknown etiology and not attributable to a myeloid neoplasm.

Two pediatric oncologic cases of hypereosinophilic syndrome and review of the literature.

BACKGROUND: Persistent peripheral blood hypereosinophilia may cause tissue damage, leading to hypereosinophilic syndrome (HES) with end-organ dysfunction. Here we discuss two unique pediatric cases of primary hypereosinophilic syndrome with oncologic etiologies to highlight the importance of early recognition, workup and treatment of HES. CASE 1: A previously healthy 7-year-old male presented with acute myocardial infarction and transient ischemic attack and found to have significant hyperleukocytosis with a total white blood count of 131 000 and hypereosinophilia with an absolute eosinophil count of 99 560. He was ultimately diagnosed with precursor B-cell acute lymphoblastic leukemia with immunoglobulin heavy chain gene rearrangement. He completed standard treatment without significant complications and remains in remission at about 2 years off therapy. He is in overall good health and has normal cardiac function. CASE 2: A 13-year-old female was referred for iron deficiency and reported a history of severe anxiety, shortness of breath and anorexia. She had experienced fatigue and dizziness associated with frequent panic attacks and shortness of breath with strenuous activity since the age of five. Serial laboratory investigations revealed persistent hypereosinophilia (AEC 4000-6000/µl). Additional workup revealed elevated vitamin B12 (>2000 pg/ml; normal range: 243-894) and tryptase (16.4 ng/ml; normal range: ≤10.9). The FIP1L1-PDGFRA gene fusion was detected by fluorescence in situ hybridization (FISH) on peripheral blood, diagnostic for myeloid/lymphoid neoplasm with eosinophilia. Evaluation for end-organ damage associated with persistent hypereosinophilia included an echocardiogram which revealed severe restrictive cardiomyopathy with pulmonary hypertension. Monotherapy with imatinib was initiated, after which she achieved a rapid hematologic response and remains in molecular remission, though she continues to have persistent asymptomatic severe pulmonary hypertension in the setting of severe diastolic dysfunction. CONCLUSION: Persistent hyperosinophilia can be a silent cause of significant and often irreversible tissue damage and should therefore always prompt workup for both primary and secondary causes.

Partial tandem duplication of KMT2A gene in patient afflicted with hypereosinophilic syndrome: A case report.

Eosinophilia is defined as a condition with increased eosinophil cell counts in blood more than the standard value. In this regard, when extensive evaluation fails to reveal the underlying causes of this disease, hypereosinophilic syndrome (HES) diagnosis should be considered. Moreover, it is possible that the mutation of the tyrosine kinase genes, as the most common type of cryptic mutations, is associated with HES syndrome. We report a case of a 47-year-old man who was initially diagnosed with HES by Microscopic examination of bone marrow aspiration and peripheral blood cell analysis. After diagnosis, the patient was administered with cortisone acetate, leading to an initial remission. One month after the initial remission, the disease relapsed, and the patient eventually died. This case report provides the first report of HES, in which a novel variant of partial tandem duplication (PTD) was detected in the KMT2A gene.

A retrospective cohort study of patients with eosinophilia referred to a tertiary centre.

INTRODUCTION: Patients with eosinophilia (an increased number of eosinophilic granulocytes > 0.5 × 108/l in the blood) are encountered in all medical specialties and frequently need thorough workup to identify the eliciting causes and decide whether treatment is indicated. In Denmark, highly specialised centres for eosinophilic diseases or conditions have been established to provide a foundation for the management of complicated cases. Here, we present experiences from such a multidisciplinary centre. METHODS: This was a retrospective study of all patients seen in our tertiary centre for eosinophilia in the 2016-2019 period. RESULTS: Referrals mainly derived from specialised secondary care and to a lesser degree from primary care physicians. Patients were either asymptomatic or exhibited symptoms from up to three organ systems and presented a median eosinophil count of 1.7 × 108/l. Up to eight new clonality analyses or imaging studies per patient were performed after referral. One of these, T-cell receptor analysis, was performed frequently but provided limited information, whereas, e.g., flow cytometry proved more clinically applicable owing to its broader diagnostic range. In total, 51 patients were evaluated and classified as secondary (59%), myeloid neoplasm with PDGFRA rearrangement (2%), idiopathic hypereosinophilic syndrome (31%) and idiopathic hypereosinophilia (8%). CONCLUSION: The value of a multidisciplinary and versatile approach in a highly specialised centre has a positive impact on diagnostic processes as well as on the evaluation of treatment need. FUNDING: none. TRIAL REGISTRATION: not relevant.

Comprehensive characterization of central BCL-2 family members in aberrant eosinophils and their impact on therapeutic strategies.

PURPOSE: Hypereosinophilia represents a heterogenous group of severe medical conditions characterized by elevated numbers of eosinophil granulocytes in peripheral blood, bone marrow or tissue. Treatment options for hypereosinophilia remain limited despite recent approaches including IL-5-targeted monoclonal antibodies and tyrosine kinase inhibitors. METHODS: To understand aberrant survival patterns and options for pharmacologic intervention, we characterized BCL-2-regulated apoptosis signaling by testing for BCL-2 family expression levels as well as pharmacologic inhibition using primary patient samples from diverse subtypes of hypereosinophilia (hypereosinophilic syndrome n = 18, chronic eosinophilic leukemia not otherwise specified n = 9, lymphocyte-variant hypereosinophilia n = 2, myeloproliferative neoplasm with eosinophilia n = 2, eosinophilic granulomatosis with polyangiitis n = 11, reactive eosinophilia n = 3). RESULTS: Contrary to published literature, we found no difference in the levels of the lncRNA Morrbid and its target BIM. Yet, we identified a near complete loss of expression of pro-apoptotic PUMA as well as a reduction in anti-apoptotic BCL-2. Accordingly, BCL-2 inhibition using venetoclax failed to achieve cell death induction in eosinophil granulocytes and bone marrow mononuclear cells from patients with hypereosinophilia. In contrast, MCL1 inhibition using S63845 specifically decreased the viability of bone marrow progenitor cells in patients with hypereosinophilia. In patients diagnosed with Chronic Eosinophilic Leukemia (CEL-NOS) or Myeloid and Lymphatic Neoplasia with hypereosinophilia (MLN-Eo) repression of survival was specifically powerful. CONCLUSION: Our study shows that MCL1 inhibition might be a promising therapeutic option for hypereosinophilia patients specifically for CEL-NOS and MLN-Eo.

Gene therapy in a murine model of chronic eosinophilic leukemia-not otherwise specified (CEL-NOS).

Chronic eosinophilic leukemia-not otherwise specified (CEL-NOS) is a rare, aggressive, fatal disease characterized by blood eosinophilia and dysfunction of organs infiltrated with eosinophils. Clinically, the disease manifests with weight loss, cough, weakness, diarrhea, and multi-organ dysfunction that is unresponsive to therapy. We developed a one-time gene therapy for CEL-NOS using an adeno-associated virus (AAV) expressing an anti-eosinophil monoclonal antibody (AAVrh.10mAnti-Eos) to provide sustained suppression of eosinophil numbers in blood, thus reducing eosinophil tissue invasion and organ dysfunction. A novel CEL-NOS model was developed in NOD-scid IL2rγnull (NSG) mice by administration of AAV expressing the cytokine IL5 (AAVrh.10mIL5), resulting in marked peripheral and tissue eosinophilia of the heart, lung, liver, and spleen, and eventually death. Mice were administered AAVrh.10mAnti-Eos (1011 genome copies) 4 wk after administration of AAVrh.10mIL5 and evaluated for anti-eosinophil antibody expression, blood eosinophil counts, organ eosinophil invasion, and survival. AAVrh.10mAnti-Eos expressed persistent levels of the anti-eosinophil antibody for >24 wk. Strikingly, CEL-NOS treated mice had markedly lower blood eosinophil levels and reduced mortality when compared with control treated mice. These results suggest that a single treatment with AAVrh.10mAnti-Eos has the potential to provide substantial therapeutic benefit to patients with CEL-NOS, a fatal malignant disorder.

Chronic Eosinophilic Leukaemia Associated with JAK2 Exon 13 Insertion/Deletion Mutations.

Chronic eosinophilic leukaemia, not otherwise specified (CEL, NOS), is a diagnosis of exclusion made in cases in which there is clonal eosinophilia but an absence of genetic aberrations that define other disease subtypes. There is a need for further characterization of these cases in order to inform risk stratification and management. The importance of JAK2 mutations in myeloproliferative neoplasms (MPN) as a whole is well established, although their role specifically in eosinophilic disorders is less clear, with only a minority of cases demonstrating JAK2 abnormalities. Here, we report 2 cases with an exon 13 insertion-deletion (indel) mutation in JAK2: one with CEL-NOS and the second with an unspecified eosinophilic disorder. JAK2 indels were not detected in a screen of suspected MPN cases (n = 592) without eosinophilia that tested negative for common MPN driver mutations. Our findings thus provide further evidence for a specific association between this rare mutation and clonal eosinophilic disorders.

A novel activating JAK1 mutation in chronic eosinophilic leukemia.

Hypereosinophilia (HE) has been defined as persistent eosinophilia >1.5 × 109/L; it is broadly divided into primary HE (clonal or neoplastic; HEN), secondary/reactive HE (HER), or HE of undetermined significance (HEUS) when no cause is identified. The use of myeloid next-generation sequencing (NGS) panels has led to the detection of several mutations in patients previously diagnosed with HEUS, reassigning some patients to the category of HEN, specifically the World Health Organization category of chronic eosinophilic leukemia, not otherwise specified (CEL, NOS). Here, we describe a novel somatic JAK1 pseudokinase domain mutation (R629_S632delinsSA) in a patient with HE that had initially been characterized as a variant of uncertain significance. We performed functional studies that demonstrated that this mutation results in growth factor independence of Ba/F3 cells in vitro and activation of the JAK-STAT pathway. These effects were abrogated by the JAK1/JAK2 inhibitor ruxolitinib. R629_S632delinsSA is the first known somatic mutation in JAK1 linked to a clonal eosinophilic neoplasm, and highlights the importance of the JAK-STAT pathway in eosinophil survival.