1.1 Myeloid/lymphoid neoplasms with eosinophilia and tyrosine kinase gene fusions

The disease category was created in the 2008 (4th edition) WHO classification with the original name of “myeloid/lymphoid neoplasm with eosinophilia and abnormalities of PDGFRA, PDGFRB or FGFR1”, to include the cases characterized by constitutive tyrosine kinase signaling as a result of the gene fusion, frequent association with eosinophilia, and except for FGFR1, often excellent responses to the first generation tyrosine kinase inhibitors (TKI). These neoplasms originate from mutated pluripotent bone marrow (BM) stem cells and may differentiate along myeloid and/or lymphoid lines, often leading to complex and heterogeneous clinical manifestations.

The category name has been changed to M/LN-eo-TK in the ICC^{6, 7} to emphasize the molecular genetic changes underlying these hematopoietic neoplasms that lead to constitutive TK signaling, which may be amenable to targeted therapy.^{3, 8} M/LN-eo-TK applies to hematopoietic neoplasms bearing the respective genetic aberration at initial presentation, not cases with such abnormalities acquired in the course of a previously diagnosed hematopoietic neoplasm. Additional important changes made in this disease category are the inclusion of M/LN with t(9;12)(q34;p13)/ETV6::ABL1 and FLT3-rearrangements as new members; and promoting PCM1::JAK2 and its genetic variants such as ETV6::JAK2 and BCR::JAK2 to formal entities from their provisional state in the revised 4th edition WHO classification. Other potential candidates for M/LN-eo-TK include rearrangements involving LYN,⁹ FGFR2,¹⁰ and KIT. At this point, these cases are limited to individual case studies, and more data are needed to understand their clinicopathological features and response to TKI therapy.

The updates also provide a clear guidance on how to distinguish M/LN-eo-TK presenting as B- or T- lymphoblastic leukemia (ALL) from BCR::ABL1-like B-ALL or de novo T-ALL involving one of the respective TK genes; as well as M/LN-eo-TK presenting with a mast cell (MC) proliferation from systemic mastocytosis (SM). For example, the most common genetic variant of PDGFRB, t(5;12)(q32;p13.2)/ETV6::PDGFRB, often manifests with multilineage involvement, characteristic of M/LN-eo-TK; however, some PDGFRB rearrangements, especially those with alternative partners such as EBF1, SSBP2,¹¹ TNIP1, ZEB2, and ATF7IP, often present as de novo B-ALL without involvement of the myeloid lineage. The latter cases are best classified as BCR::ABL1-like B-ALL.¹² Similarly, JAK2 fusions with certain partner genes, such as t(5;9)(q14.1; p24.1)/STRN3::JAK2, and PAX5::JAK2 are usually seen in BCR::ABL1-like B-ALL, which are, by definition, not M/LN-eo-TK. To complicate the issue, some fusion partners can be observed in either entity. For example, ETV6::JAK2 is recognized as a genetic variant of PCM1::JAK2 in M/LN-eo-TK; however, more than half of the reported cases of ETV6::JAK2 are de novo B-ALL or de novo T-ALL.^{13, 14} Similarly, FLT3-rearrangement can occur in de novo B-ALL and T-ALL without a myeloid lineage involvement, and these cases should be classified as BCR::ABL1-like B-ALL or de novo T-ALL rather than M/LN-eo-TK.^{12, 15}

These decisions are driven by the general rule that, for cases of M/LN-eo-TK that initially present as B- or T-ALL, the TK gene fusion should involve the myeloid lineage in addition to lymphoblasts. In such instances, the chronic myeloid neoplasm (CMN) in M/LN-eo-TK may manifest either prior to or concomitantly or may emerge after therapy for the ALL. This concept is reminiscent of the distinction between CML presenting in B-lymphoblastic crisis and de novo BCR::ABL1+ B-ALL. For practical purposes, FISH-studies on granulocytes for detection of respective gene fusions in a clearly myeloid population may be very helpful in distinguishing M/LN-eo-TK from ALL.

An abnormal MC proliferation can be observed in M/LN-eo-TK (see below description). However, these cases consistently lack the KIT D816V mutation and show similar responses to TKI as other M/LN-eo-TK.¹⁶ In the current ICC, such cases are classified under M/LN-eo-TK if one of the TK gene fusions is detected.⁶ As such, it is highly recommended to test cases of putative SM for TK gene fusions, especially in cases associated with eosinophilia and lacking a detectable KIT mutation by high sensitivity molecular methods. In rare cases, a TK gene fusion and a KIT D816V mutation may coexist, and these may represent co-occurrence of two diseases with separate clones or a subclone with an acquired activating KIT mutation. These rare cases would fall within the spectrum of SM with an associated myeloid neoplasm (SM-AMN).

1.2 Idiopathic hypereosinophilic syndromes

HES is defined as persistent PB HE (≥1.5 × 10⁹/L) and/or tissue HE with associated tissue/organ damage/injury that is directly attributable to eosinophil-released cytokines or enzymes. The majority of HES are reactive due to an underlying inflammatory condition,^81-83 or infection, autoimmune disorder, allergy/vaccination, or neoplasm, such as solid tumors, lymphomas, or lymphocyte variant HES (LV-HES). HES is considered as idiopathic (iHES) when an underlying primary cause (either reactive or neoplastic) cannot be identified.^{2, 82} HES is generally divided into¹: (1) secondary HES (eosinophils are reactive and non-clonal) including lymphocyte variant-HES; (2) primary HES (associated with a hematopoietic neoplasms); (3) idiopathic HES. It is noteworthy that primary HES should be classified by the hematopoietic neoplasm according to ICC. The term of “idiopathic HE” or “HE of unknown significance” (HEus) is used to describe persistent HE (≥6 months) without associated organ/tissue damage.^{84, 85}

It has been historically challenging to differentiate iHES/HEus from CEL, NOS (a subtype of MPN with hypereosinophilia), due to the difficulty in proving clonality in the latter. In the 2008 WHO definition, CEL, NOS was separated from iHES/HEus by the presence of increased PB or BM blasts and/or a clonal karyotypic abnormality. With the advances in NGS and its ubiquitous diagnostic application, somatic mutations associated with myeloid neoplasms can be detected in 25%–30% of patients who would have been previously classified as “iHES.”^86-89 Such mutations have been found mostly in genes involved in DNA methylation and chromatin modification, such as ASXL1, TET2, EZH2, and DNMT3A, but also in other genes such as SRSF2, TP53, and SETBP1.^{83, 86-88} More recently, STAT5B N642H⁸⁹ has been reported in some patients with a referral diagnosis of eosinophilia (1.6% of patients in one series), including patients who would be otherwise diagnosed with iHES/HEus. While the presence of a mutation provides evidence of clonality, such mutations have also been reported in aging individuals lacking evidence of a myeloid neoplasm.^{90, 91} Some cases with clinical features typical of iHES/HEus and with a relatively benign clinical course have also been found to carry somatic mutations.^{83, 92} On the other hand, some true CEL, NOS, even those with karyotypic abnormalities, may not show detectable mutations at least by myeloid neoplasm targeted NGS panels.^{2, 83, 92} Although the underlying cause for iHES/HEus remains to be discovered, it is acknowledged that iHES/HEus lacking convincing evidence of clonality or abnormal BM morphology of a CMN should not be considered a myeloid neoplasm and should be kept separate from CEL, NOS and other true myeloid neoplasms with HE.

1.2.1 Histopathology

Recent studies have shown that the BM of patients with iHES/HEus is usually unremarkable except for increased eosinophils^{17, 83, 92} (Figure 1). The cellularity is often appropriate for age, and megakaryocytes are present in adequate numbers and exhibit normal morphology. Eosinophils in PB and BM are mostly of normal forms with bilobated nuclei, though, in some cases, mild uneven cytoplasmic granulation and hypersegmentation may be observed, often in a subset of eosinophils.^{83, 86, 92} Mast cells (MCs) are not significantly increased. If MCs are spindly and or in clusters, further work-up to rule out SM associated with eosinophilia is warranted.

Like iHES, the BM of LV-HES is also often unremarkable except for increased eosinophilia.^{93, 94} However, in LV-HES, abnormal T-cells, often with a TH2 type immunophenotype (CD3-, CD4+, CD8−, CD7−/or partially lost, CD5 bright+), are identified by flow cytometry but do not form a BM infiltrate.⁹⁵ Of note, T-cell clonality demonstrated by TCR PCR only is not sufficient for a diagnosis of LV-HES due to its non-specific nature. A diagnosis of iHES requires the exclusion of LV-HES. LV-HES is often sensitive to corticosteroids but carries a 5%–20% risk of progression to T-cell lymphoma with long-term follow-up. Some iHES may only present with tissue HE with a lower PB eosinophil count. Tissue HE is defined by an eosinophilic infiltrate or deposition of eosinophilic granules, significantly exceeding the normal physiologic ranges. BM HE is defined as 20% or more eosinophils of all nucleated BM cells. In some tissue biopsies, there may be only fibrosis with eosinophilic granules observed.

In contrast to iHES/HEus, the BM of CEL, NOS is often markedly hypercellular with pronounced megakaryocytic dysplasia, an increased myeloid: erythroid ratio, occasional dysgranulopoiesis, dyserythropoiesis, and myelofibrosis in 20%–30% cases, showing morphologic abnormalities similar to those seen in the BM of MDS/MPN, MDS, or MPN. In addition, eosinophils in CEL, NOS frequently demonstrate significant abnormal features (abnormal granulation, nuclear hyper- or hypo-segmentation, large size or, an increased in immature forms) (Figure 1).^{83, 92} A key update in the ICC is that abnormal BM histopathology is now incorporated among the required diagnostic criteria for CEL, NOS, allowing a more reliable separation from iHES/HEus.⁶ (Table 2).

TABLE 2. Diagnostic criteria and comparison of idiopathic hypereosinophilic syndrome (iHES)^a versus chronic eosinophilic leukemia, not otherwise specified (CEL, NOS).

	iHES	CEL, NOS
Definition	Persistentb HE with tissue damage/dysfunction attributing to eosinophils, not caused by a known reactive or neoplastic cause	A myeloproliferative neoplasm with a prominent and persistentb proliferation of eosinophils. Blasts constitute <20% cells in peripheral blood and bone marrow, not meeting any other diagnostic criteria for AML
Peripheral blood	Persistent peripheral blood hypereosinophilia (eosinophil count ≥1.5 × 109/L and ≥ 10% eosinophils)	Persistent peripheral blood hypereosinophilia (eosinophil count ≥1.5 × 109/L and ≥ 10% eosinophils)
Organ damage and/or dysfunction	Present, attributable to tissue eosinophilic infiltrate	Not required for diagnosis, variably present in some patients
Etiology	No evidence of a reactive, well-defined autoimmune disease or neoplastic condition/disorder underlying HE	A myeloid neoplasm with clonal eosinophil proliferation
Exclusions required	Rule out neoplastic and reactive HE including lymphocyte variant hypereosinophilic syndromec	No tyrosine kinase gene fusion including BCR::ABL1, ETV6::ABL1, PDGFRA, PDGFRB, FGFR1, JAK2, FLT3 fusions not meeting any other diagnostic criteria for AML, CMML, SM
Bone marrow findings	Bone marrow morphologically within normal limits except for increased eosinophils	Increased cellularity with dysplastic megakaryocytes with or without dysplastic features in other lineages including eosinophils and often significant fibrosis, associated with an eosinophilic infiltrate OR There are increased blasts ≥5% in the bone marrow and/or ≥2% in the peripheral blood
Molecular genetics	No molecular genetic clonal abnormality, with the caveat of clonal hematopoiesis of indeterminate potential (CHIP)	Demonstration of a clonal cytogenetic abnormality and/or somatic mutation(s)d

• ^a Hypereosinophilia of uncertain significance (HEus) has no tissue damage, otherwise, should follow the same diagnostic criteria for iHES.
• ^b Preferably a minimal duration of 6 months; however, in patients presenting with organ damage/dysfunction, immediate treatment is required, an interval of 4 weeks or two occasions with a minimum time interval of 2 weeks may suffice. Hypereosinophilia in the diagnosis of CEL, NOS is preferable 3 months, in patients required treatment, a minimum of 1 month at least two occasions.
• ^c The abnormal T-cell population needs to be detected immunophenotypically with or without TCR clonality by PCR.
• ^d In the absence of a clonal cytogenetic abnormality and/or somatic mutation(s) or increased blasts, bone marrow findings supportive of the diagnosis will suffice in the presence of persistent eosinophilia, provided other causes of eosinophilia having been excluded.

1.2.4 Prognosis and treatment

The early case series of HES reported a dismal prognosis in some patients, likely due to the inclusion of true myeloid neoplasms (so-called “myeloid HES”). In recent studies where an overt hematopoietic neoplasm is excluded, iHES demonstrates a much more indolent clinical course with a long-term disease related mortality of 10%–15%.^{87, 92, 100, 101} Mortality is associated with age > 60 years, cardiac involvement, cytopenia (anemia or thrombocytopenia), low absolute lymphocyte count⁸⁷ increased neutrophil to lymphocyte ratio,¹⁰¹ and hepatosplenomegaly.

For patients with iHES, treatment decisions largely depend on the pattern and extent of organ infiltration/damage. iHES with multi-organ involvement needs similar treatment considerations like ANCA-negative eosinophilic granulomatosis with polyangiitis. In general, corticosteroids are effective in producing a rapid reduction in eosinophil count or tissue/organ dysfunction in most of the patients.^{85, 102} However, long-term treatment with steroids to suppress eosinophilia and organ damage can carry significant side effects, with recommendations to taper once symptom control and a reduction of the eosinophil count to <1.5 × 10⁹/L is achieved.³ Steroid resistance or intolerance occurs in about 20% of patients. Failure to control symptoms, organ damage, or persistently elevated PB eosinophil count with a prednisone dose >10 mg daily is an indication for the addition of other agents. The IL-5 antagonist mepolizumab¹⁰³ has been shown to significantly reduce the occurrence of symptom flares, with marked reductions in PB eosinophilia, and can be associated with a reduced need for oral corticosteroids and other cytoreductive therapy. Benralizumab is an anti-IL-5 receptor antibody that also demonstrates activity in PDGFRA-negative HES,¹⁰⁴ and is further being evaluated in a phase 3 randomized, double-blind, placebo-controlled study (ClinicalTrials.gov Identifier: NCT04191304).

Figure 2 is an algorithm to illustrate the ICC recommendation in the diagnosis and classification of hypereosinophilia.

2.1 Introduction

Mastocytosis is characterized by an accumulation of clonal MCs infiltrating one or more organs, including BM, skin, gastrointestinal tract, liver, and spleen.¹⁰⁵ Mastocytosis can be divided into cutaneous mastocytosis (CM), systemic mastocytosis (SM), and mast cell sarcoma (MCS). CM which can present in different forms,¹⁰⁶ is usually confined to the skin (pure CM) in children but almost always “systemic” in adults (i.e., represents a manifestation of SM).

SM is mostly a nonaggressive disease, commonly presenting as indolent SM (ISM) and more rarely as smoldering SM (SSM). Advanced SM (AdvSM) comprises aggressive SM (ASM), SM with an associated myeloid neoplasm (SM-AMN), and mast cell leukemia (MCL). MCS is a localized, destructive MC infiltration with an aggressive behavior, and around 20% of MCS are associated with underlying SM.¹⁰⁷ If the MCS develops in a patient with SM, it is more appropriate to call those cases MCS-like progression of the SM rather than MCS. It is known that MCS associated with SM usually carries a KIT D816V mutation, whereas most cases of primary MCS lack the KIT D816V mutation.¹⁰⁸

In the ICC 2022 update,^{6, 109} the diagnostic criteria for SM, including the major criteria and minor criteria, are mostly the same as those outlined in the revised 4th edition WHO, but some refinements have been introduced (Table 3). Some changes have also been made in the assessment of “disease burden” B findings (Figure 3). The diagnostic criteria for MCL have been modified to ensure that the entity of MCL represents a high-grade aggressive form of SM and to eliminate the term “chronic MCL”. These changes will be further discussed in the following sections.

2.2 Histopathology

In the ICC update, there are a number of changes involving the use of histopathology in defining mastocytosis. The major criterion for the diagnosis of SM remains the presence of multifocal dense MC infiltrates (≥15 MCs in aggregates) in BM sections and/or other extracutaneous organ(s), but it now requires demonstration of CD117 (often strong) and MC tryptase (often weaker and may be partial) expression to identify and confirm the MC infiltrate.¹¹⁰ With the support of immunohistochemistry, the major criterion on its own can be sufficient for diagnosing SM, when M/LN-eo-TK is carefully excluded. The use of immunohistochemistry is also important to better estimate the MC infiltrate burden and to assess MC morphology. Regarding the latter, the presence of ≥25% atypical MCs in tissue sections or aspirate smears is the first minor criterion for the diagnosis of SM. Atypical MCs include spindle-shaped MCs (often also hypogranular), promastocytes, metachromatic blasts, multinucleated or highly pleomorphic MCs, and recently recognized well-differentiated MCs. Well-differentiated MCs are heavily granulated and have round central nuclei and often abundant cytoplasm¹¹¹ (Figure 4). Well-differentiated MCs have been observed in various subcategories of SM, and therefore do not designate a specific subtype of SM. CD30 expression, either by flow cytometry¹¹² or immunohistochemistry,¹¹³ can be detected in up to 80%–90% SM. CD30 is now accepted as an additional marker to CD25 and CD2 in defining immunophenotypic aberrancy of MCs,¹¹² which is the second minor criterion in the diagnosis of SM. CD30 is particularly valuable in the assessment of SM with well-differentiated MCs that are often negative for CD25 and CD2, but frequently positive for CD30.¹¹¹ Like CD25 and CD2, CD30 may be occasionally expressed in a subset of normal/reactive MCs at a low level.¹¹⁴ However, expression of two or more aberrant markers or strong uniform expression of at least one of the three markers is strongly associated with SM. It is important to be aware that in some cases of SM,¹¹⁵ the major criterion of aggregates of ≥15 MC in the BM or tissue biopsy is not present, and the diagnosis relies on the presence of three or all four minor criteria; therefore, in this setting, documentation of MC morphology and identifying a MC-associated aberrant immunophenotype become essential.

In a substantial subset of SM, there may be an associated hematological neoplasm (SM-AHN). The most common association is CMML, followed by MDS, MPN including CEL, NOS, other MDS/MPN and AML. In SM associated with hypereosinophilia, if the BM fulfills the diagnostic criteria of CEL, NOS, the diagnosis would be SM-CEL; otherwise, it will be SM with eosinophilia. A shared clonal origin has been demonstrated between SM and the associated myeloid neoplasms either by cytogenetic abnormality¹¹⁶ and/or KIT mutations. In rare cases, SM may present with a concurrent lymphoid neoplasm or plasma cell neoplasm, and in such cases these two components are instead considered clonally unrelated, co-incident diseases.^116-118 In the ICC 2022 update, SM-AHN is now limited to the association with a myeloid neoplasm (SM-AMN). A high degree of suspicion for an underlying myeloid neoplasm should be raised by the presence of significant cytopenia(s) or cytosis (leukocytosis, monocytosis, eosinophilia, thrombocytosis), splenomegaly, elevated LDH, high KIT D816V variant allele frequency, as well as the detection of additional somatic mutations in genes associated with myeloid malignancies. Morphologic assessment of the BM should include BM cellularity, megakaryocyte morphology, dyserythropoiesis, dysgranulopoiesis, monocytosis, eosinophilia, and blasts. The AMN should be subclassified according to established criteria. On the other hand, if a KIT mutation is detected in a myeloid neoplasm, performing CD117 and MC tryptase stains is strongly recommended to identify an occult associated accumulation of neoplastic MCs,¹¹⁹ especially in CMML and AML with t(8;21).

In SM, disease burden and aggressiveness are measured by B- and C-findings. B findings refer to organ enlargement without organ failure, while C-findings refer to MC-related organ damage. In the ICC update, the vague morphological description in the second criterion of the B finding in the revised 4th edition of the WHO “signs of dysplasia or myeloproliferation in non–MC lineage(s), but criteria are not met for a definitive diagnosis of an associated hematological neoplasm” is now removed to avoid ambiguity in disease definition. As a result, the first part of the B finding is now simplified to “cytopenia (not meeting criteria for C findings) or -cytosis. Reactive causes are excluded, and criteria for other myeloid neoplasms are not met” (Figure 3).

Another important change is in the definition of MCL. In the past classifications including the revised 4th edition of the WHO, MCL was simply defined as SM with ≥20% MCs on BM aspirate smears, regardless of MC morphology or C findings. In most cases of SM, MCs are bland-appearing spindled cells associated with significant reticulin fibrosis, yielding very few MCs in BM aspirates even in cases with a high MC burden. The cases with ≥20% MCs on BM aspirate smears are those without significant fibrosis and mostly round-shaped MCs with high grade cytology (i.e., atypical immature MCs, see below) or with well-differentiated morphology. The latter have been described in a proportion of cases of the so-called chronic MCL.¹²⁰ Due to the same reason of BM aspirate for MCs, cases of ASM with 5%–19% MCs on BM aspirates have been found to be similarly aggressive and are referred to as “ASM in transformation.”¹⁰⁸ Of note, MCL is the only disease in the hematolymphoid neoplasm classification which exclusively relies on BM aspirate smear count independently of the biopsy. In the ICC 2022 update, MCL is redefined as a subset of SM with atypical immature morphology in 20% or more of the BM cells. Atypical immature MCs include metachromatic blasts, promastocytes, multinucleated, anaplastic, or highly pleomorphic MCs.^{121, 122} Metachromatic blasts¹²³ are blast-like (immature chromatin, with or without nucleoli) with metachromatic granules; while promastocytes are immature MCs with bilobated nuclei and cytoplasms packed with metachromatic MC granules (Figure 5). Spindled MCs and MC with well-granulated morphology (well-differentiated MC), though different from normal MC, are not considered as atypical immature MCs. The ICC does not recognize the still poorly characterized entity termed chronic MCL.¹²⁰ Following the ICC approach, if the aspirate is suboptimal or a dry tap, a BM biopsy showing a diffuse MC infiltration with at least 20% of atypical immature MCs by CD117, and/or tryptase immunohistochemistry stains is sufficient to support a diagnosis of MCL. In addition, the ICC has further removed “aleukemic” or “leukemic” from the subclassification of MCL. Of note, the majority of MCL are aleukemic in that ≥10% circulating MCs are very infrequent.¹²⁴ Recently, by flow cytometry, circulating MCs were detected in nearly half of the patients with ISM, almost all patients with AdvSM and SSM.¹²⁵

2.3 Cytogenetics

An abnormal karyotype is reported in 10%–15% SM in two large series.^126-128 The majority of the clonal abnormalities are identified in patients with SM-AMN (25%–30%), followed by MCL (15%–20%) and aggressive SM (<10%). The karyotypic abnormalities, which are not unique to SM, are likely related to the underlying AMN (CMML, MDS, MDS/MPN, or MPN). These include −7, +8, or complex karyotypes, and less frequently, del(5q) or del7q. In ISM, clonal cytogenetic abnormalities are extremely rare. Whenever present, they are low risk abnormalities such as del(20)q, +Y, del(12)p, or inv(2). An abnormal karyotype is more frequently associated with male gender, older age, and cytopenia(s).¹²⁷ While an abnormal karyotype did not identify it as an independent prognostic factor in patients with SM,¹²⁷ a study of MCL patients found that it was associated with inferior outcomes.¹²⁶ Monosomy 7 and complex karyotypes in advanced SM (including SM-AMN) have been reported to be associated with disease progression to MCL¹²⁴ and secondary AML.¹²⁸

2.4 Molecular genetics

KIT D816 mutation is detected in >90% of SM, mostly in the indolent forms of SM, but >50 other rare KIT mutations have also been described in SM.¹²⁹ Over 75% of ISM and SSM have an isolated KIT mutation without other co-mutations.^{130, 131} Due to a low yield of MCs in BM aspirates, the use of a high sensitivity PCR assay for KIT D816, such as quantitative allele-specific polymerase chain reaction (PCR) or digital droplet PCR (ddPCR) assays with a sensitivity down to 0.01%–0.1%,¹³² is highly recommended in suspected SM whenever NGS fails to reveal a KIT mutation. While KIT D816 mutation is uncommon or absent in MCS or SM with well-differentiated morphology as well as in some AdvSM, other activating KIT mutations are not infrequently encountered.^{107, 111, 129, 133} As these other activating KIT mutations can render the MCs responsive to TKI, complete KIT gene sequencing should be strongly considered.^134-136 If a KIT mutation is absent, particularly in cases with eosinophilia, TK gene fusions associated M/LN-Eo-TK must be excluded. As previously discussed, M/LN-eo-TK may show morphologic findings similar to those seen in true SM.

KIT D816V mutation in non-MC lineages can be found in ISM, but is much more frequent in AdvSM, particularly SM-AMN. The KIT D816V VAF in PB and BM correlates with the subtype of SM, multilineage involvement, serum tryptase level, organ involvement, disease progression and survival.^{117, 125, 137} In SM-AMN, especially SM-CMML and SM-MDS, a KIT D816V mutation has been clearly demonstrated not only in SM, but also in the myeloid cells.¹¹⁷ The measurement of KIT D816V VAF has been incorporated as a benchmark for treatment response in studies of non-advanced and advanced SM.^{138, 139} Additionally, a 10% or higher KIT D816V allele burden has been proposed as a novel additional B-finding¹⁰⁸ and is included in the 5th edition WHO Classification.⁹⁷ While a correlation between KIT D816V VAF and disease burden may be observed, it was felt by the ICC that more evidence is needed before a precise threshold can be determined and clinical utility assessed. In fact, a recent study¹⁴⁰ showed that the presence of ≥3.5% (not ≥10%) KIT D816V-mutated cells as well as a dynamically unstable KIT D816V mutation burden (increase or decrease over time) in blood and/or BM are associated with a significantly shortened progression-free survival (PFS). In addition, a high mutational VAF can be due to a clonally related concomitant myeloid neoplasm, which may or may not be readily apparent at the time of diagnosis of SM, or may be unmasked when SM-directed therapy is administered.

In contrast to ISM and SSM, the genetic profile of AdvSM is more complex, with 60%–70% of cases showing mutations in addition to KIT.^{130, 141-145} These additional somatic mutations are strikingly similar to those encountered in various myeloid neoplasms such as SRSF2, ASXL1, RUNX1, TET2, DNMT3A, JAK2, CBL, and NRAS. The genetic profile and the VAF of individual genes may substantially affect clinical phenotype, response to treatment, and prognosis. Mutations in one or more of the SRSF2, ASXL1, and/or RUNX1 (so-called S/A/R gene panel) genes are associated with a poorer prognosis in AdvSM, including MCL.^{141, 143} Therefore, it is recommended to test other pathogenic mutations in genes with known prognostic impact in SM patients for further identification of patients at higher risk. It is noteworthy that all of the above molecular markers should be used in combination with other disease features for accurate risk stratification of patients with SM.

2.5 Clinical features

The clinical presentations vary among different subtypes of SM that are closely related to B- and C- findings. In the ICC 2022, the “burden of disease” B-findings are largely unchanged but simplified and refined (Figure 3). In addition to removing the description of BM morphology as discussed above, the third criterion has been modified to specify “splenomegaly” as “palpable splenomegaly without features of hypersplenism including thrombocytopenia” and “lymphadenopathy” as “lymph node size of >1 cm” (a threshold of >2 cm is used by the ECNM-AIM consensus criteria¹⁰⁸). No changes have been made to the “C”-findings (Figure 3).

ISM is the most common form of SM,¹⁴⁶ defined by less than 2 B-findings and absence of C-findings. ISM has a median onset age of 45–57 years.^146-148 Its presenting symptoms include maculopapular skin lesions, itching, flushing, abdominal pain, acid reflux, nausea, diarrhea, brain fogginess, and musculoskeletal pain. These symptoms significantly affect the quality of life in about two thirds of patients. Secondary osteopenia and osteoporosis are reported in up to one third of patients. These may be the presenting symptom of the disease.^{149, 150} Isolated BM mastocytosis (BMM) is a clinicopathologic variant of ISM (ICC), which account for 25%–60% of all ISM.^{150, 151} BMM is characterized by absence of skin/cutaneous lesions, isolated BM involvement with a lower MC burden, tryptase level < 125 ng/mL, no B finding, older age, and male predominance.^{149, 150, 152} The incidence of hymenoptera allergic/anaphylaxis reactions to the sting by a bee, wasp or other hymenoptera insects due to an IgE-mediated allergy to its venom,^{149, 150} is significantly higher in BMM than other ISM (62% vs. 16%).¹⁵⁰ Overall, the prognosis of ISM is excellent, with reported disease progression to AdvSM in <3% patients.¹⁴⁸

SSM is characterized by a high MC burden, defined by the presence of ≥2 B-findings but no C findings. SSM has a higher rate (9%–15%) of progression to AdvSM (up to 7%) than ISM.^146-148 In contrast, ASM has ≥1 C-finding(s) due to SM-induced organ damage. ASM can occur de novo or as progression from ISM or SSM.^{146, 147, 153, 154} Skin lesions of ASM are less common than ISM (present in ~50%), but constitutional symptoms (60%), hepatosplenomegaly (50%), and lymphadenopathy (30%) are frequent.¹⁵⁵ While patients with ISM may have osteoporosis, patients with ASM often present with osteosclerosis and may present with pathologic fracture(s) due to lytic bone lesions. SM-AHM should be excluded before making a diagnosis of SSM and ASM.

In SM-AMN, the SM and the underlying myeloid neoplasm may be diagnosed at the same time (67%), or at a variable interval (3–370 months) apart.¹⁵⁶ In patients with SM-AMN, skin lesions are present in 60%–70% of cases¹⁵⁶ and cytopenia, cytosis, hepatosplenomegaly, increased LDH are common, depending on the type of AMN. It can be extremely challenging to attribute potential C findings, including clinical symptoms, signs of organ involvement, and abnormal laboratory findings, to SM versus the concomitant AMN. BM biopsy to assess MC burden as well as the AMN component, especially blast count, may provide important information for clinical management and therapeutic decision making, as the prognosis of SM-AMN tends to depend largely on the type and stage of the AMN.

Figure 3 provides a practical algorithm for the diagnosis and classification of SM following the ICC criteria.

2.6 Prognosis and treatment

The prognosis and survival in ISM are favorable. Progression of ISM to SSM occurs in around 2% of patients and to AdvSM in 1%–3%.^{147, 148, 154, 157} Progression of SSM to AdvSM occurs in up to 7% of patients.¹⁵⁸ For patients with ISM and SSM, the treatment is to manage SM mediator symptoms and avoid triggers that may exacerbate symptoms. Antihistamine medications (anti H1 and/or H2 receptor) are recommended depending on the constellation of mediator symptoms. ISM patients (as well as patients with all mastocytosis variants) are encouraged to carry epinephrine autoinjectors to mitigate potentially life-threatening episodes of anaphylaxis. For patients who are prone to anaphylaxis due to bee/wasp venom allergies, venom immunotherapy to induce immune tolerance has shown to provide protection.¹⁵⁹ Anti-IgE antibody treatment (e.g., omalizumab) has shown benefit in patients to prevent life-threatening reactions.¹⁶⁰ Other treatment for ISM/SSM includes leukotriene inhibitors, cromolyn sodium, and ketotifen. Bisphosphonates and RANK ligand inhibitors (e.g., denosumab) may be considered for osteoporosis which can be more frequent and severe in patients with mastocytosis. Historically, PEG-interferon-α, corticosteroids, and cytoreduction with cladribine have been reserved for patients with severe, refractory symptomatic ISM; however, both short- and longer-term toxicities, need to be considered carefully before using such agents in these patients who otherwise exhibit a (near) normal life expectancy. Highly selective KIT D816V inhibitors (e.g., avapritinib and bezuclastinib) are currently being assessed in clinical trials of ISM patients with refractory symptoms despite best supportive care. Validated mastocytosis-specific patient-reported outcome measures should be used for treatment response assessment.¹⁶¹ In trials of KIT inhibitors in ISM, BM MC burden, serum tryptase level, and KIT D816V VAF are being evaluated as biomarkers of response but do not necessarily correlate with the severity of mediator symptoms and quality of life.

AdvSM carries a poor prognosis, typically with a median survival of less than 4 years.^{146, 147, 162} Midostaurin is a multikinase/KIT inhibitor that targets not only D816V-mutated KIT but also wild-type (WT) KIT, PDGFRα/β, VEGFR2, and FLT3.¹⁶³ Avapritinib is a potent and highly selective oral type 1 multi-kinase inhibitor with activity against KIT D816V.^164-167 Based on trials of AdvSM patients, midostaurin¹⁶³ and avapritinib,^{164, 165, 167, 168} were approved by the FDA for frontline treatment of AdvSM, in 2017 and 2021, respectively.¹⁶⁹ Bezuclastinib is an oral highly selective TKI with minimal brain penetration which is currently being evaluated in AdvSM.¹⁷⁰ In patients with pure MCL and ASM without an AMN component, treatment with midostaurin or avapritinib generally result in better responses compared to patients with SM-AMN. However, treatment decisions for SM-AMN should be individually tailored, prioritizing the treatment for the component felt to be mostly contributing to the patient's clinical symptoms and organopathy. This should include consideration of the patient's mediator symptoms, complete blood count, BM findings including MC burden and blast count/stage of AMN, tryptase level, presence and severity of organ damage (e.g., cytopenias, liver function abnormalities, hypoalbuminemia with weight loss), and molecular profile including KIT D816V VAF and presence of other myeloid molecular abnormalities, especially the poor-risk SRSF2/ASXL1/RUNX1 mutations. The SM component often responds to KIT targeting agents, with molecular remissions of KIT D816V achievable with highly selective KIT D816V inhibitors such as avapritinib. However, progression of disease is usually related to the AMN component. In this regard, how best to combine KIT inhibitors with AMN-targeted therapy is a major focus of future clinical trial development in AdvSM. Allo-SCT may be considered for patients with SM-AMN, especially in younger patients with good performance status who have achieved a pre-SCT complete remission or significant reduction of the MC component. In the KIT inhibitor era, the ability of new potent TKIs (e.g., avapritinib) to elicit pathologic complete remissions and deep molecular responses is increasingly attainable, which should make more patients eligible for transplant if they are felt to be appropriate candidates.¹⁷¹ The role of KIT inhibitors in the posttransplant setting to reduce relapse also merits investigation.