AML is a heterogeneous blood disorder in terms of cytogenetic, molecular, epigenetic, phenotypic and morphological characteristics. The diagnosis is established by the presence of 10% or more blasts of myeloid origin in the bone marrow or peripheral blood for AML with recurrent genetic abnormalities, or with 20% or more blasts of myeloid origin in case of mutated TP53, AML with myelodysplasia-related gene mutations or cytogenetic abnormalities, or for AML not otherwise specified. AML can be classified according to the international consensus classification as published in 2022.
AML is characterized by the inhibition of differentiation and the subsequent accumulation of cells at various stages of incomplete maturation, and by the reduced production of mature hematopoietic cells. Cytopenia’s cause clinical manifestations, with symptoms and signs of anemia (such as fatigue and dyspnea), neutropenia (such as infections), and thrombocytopenia (such as hemorrhage), which are usually present at the time of diagnosis and predominate during treatment.
AML is a complex and dynamic disease characterized by numerous driver mutations, i.e. acquired somatic mutations and competitive clones that contribute to the development of the disease over time.
Conventional cytogenetic analysis is mandatory if a diagnosis of AML is suspected.
Tests/procedures at diagnosis for a patient with AML
|Tests to establish the diagnosis|
|Complete blood count and differential counta|
|Bone marrow aspirateb|
|Bone marrow trephine biopsyc|
|Immunophenotyping by flow cytometry|
|Results preferably available within|
|Screening for gene mutations required for establishing the diagnosis and to identify actionable therapeutic targetse|
Screening for gene rearrangementsh
|PML::RARA, CBFB::MYH11, RUNX1::RUNX1T1, KMT2A rearrangements, BCR::ABL1, other fusion genes (if available)|
Additional genes recommended to test at diagnosisi
• ANKRD26, BCORL1, BRAF, CBL, CSF3R, DNMT3A, ETV6, GATA2, JAK2, KIT, KRAS, NRAS, NF1, PHF6, PPM1D, PTPN11, RAD21, SETBP1, TET2, WT1
Demographics and medical historyj
Detailed family historyk
Patient bleeding historyl
Analysis of comorbidities
Additional tests and procedures
Complete physical examinationm
Performance status (ECOG/WHO score)
Geriatric assessment (optional)
Biochemistry, coagulation testso
Hepatitis A, B, C; HIV-1 testing; CMV, EBV, HSV, VZV
Serum pregnancy testp
Eligibility assessment for allogenetic HCT (incl. HLA-typing)q
Chest x-ray, 12-lead electrocardiogram, echocardiography or MUGA (on indication)
Computed tomography of the chest (on indication)r
Lumbar puncture (on indication)s
Information on oocyte and sperm cryopreservationt
CMV, cytomegalovirus; EBV, Epstein-Barr virus; ECOG, Eastern Cooperative Oncology Group; HCT, hematopoietic cell transplantation; HSV, herpes simplex virus; MUGA, multigated acquisition; VZV, varicellazoster virus; WHO, World Health Organization.
a 200 nucleated cells on blood smears should be counted.
b 500 nucleated cells on bone marrow smears should be counted. Myeloblasts, monoblasts, and megakaryoblasts are included in the blast count. Monoblasts and promonocytes, but not abnormal monocytes, are counted as blast equivalents in AML with monocytic or myelomonocytic differentiation.
c In patients with a dry tap (punctio sicca); touch preparations from the core biopsy should be performed if a dry tap is suspected.
d At least 20 bone marrow metaphases are needed to define a normal karyotype, and recommended to describe an abnormal karyotype. Normal and abnormal karyotypes may be diagnosed from blood specimens with circulating blasts. In case of no analyzable metaphases, fluorescence in-situ hybridization is an alternative method to detect genetic abnormalities like RUNX1::RUNX1T1, CBFB::MYH11, KMT2A, and MECOM gene fusions, or myelodysplasia-related chromosome abnormalities, eg, loss of chromosome 5q, 7q, or 17p material.
e Screening for gene mutations is an evolving field of research; screening for single genes is increasingly replaced by gene panel diagnostics.
f FLT3: mutational screening should include the analysis of internal tandem duplications (ITD) and of tyrosine kinase domain (TKD) mutations. Longer FLT3-ITDs may be missed by next-generation sequencing, therefore, we recommend continuing to use capillary electrophoresis.
g The report should specify type of mutation: only in-frame mutations affecting the basic leucine zipper (bZIP) region of CEBPA, irrespective whether they occur as monoallelic or biallelic mutations, have been associated with favorable outcome.
h Screening for gene rearrangements should be performed if rapid information is needed for recommendation of suitable therapy, if chromosome morphology is of poor quality, or if there is typical morphology but the suspected cytogenetic abnormality is not present.
i Results from these genes are not required for establishing the diagnosis or for the identification of actionable therapeutic targets, rather they may be used for subsequent monitoring of the disease by next-generation sequencing-based techniques (with the exception of mutations consistent with pre-malignant clonal hematopoiesis, eg, DNMT3A, TET2, ASXL1); although these techniques are still investigational, this is a rapidly evolving field.
j Including race or ethnicity, prior exposure to toxic agents, prior malignancy, therapy for prior malignancy, information on smoking.
k Thorough family history needed to identify potential myeloid neoplasms with germline predisposition.
l History of bleeding episodes may inform cases of myeloid neoplasms with germline predisposition and preexisting platelet disorders.
m Special attention for skin (bleeding symptoms, leukemia cutis, Sweet syndrome), gingival hyperplasia, lymphadenopathy, testis enlargement, signs of infection (eg, pulmonary, perianal, mouth/teeth); symptoms of central nervous system involvement; signs of abnormalities associated with germline predisposition syndromes (see Table 2).
n Tests for objectively measured physical and cognitive function are particularly useful in the context of trials.
o Biochemistry: glucose, sodium, potassium, calcium, creatinine, aspartate amino transferase (AST), alanine amino transferase (ALT), alkaline phosphatase, lactate dehydrogenase (LDH), bilirubin, urea, total protein, uric acid, total cholesterol, total triglycerides, creatinine phosphokinase (CPK). Special attention should be given to tumor lysis syndrome. Coagulation tests: prothrombin time (PTT), international normalized ratio (INR) where indicated, activated partial thromboplastin time (aPTT).
p In women with childbearing potential.
q HLA typing and CMV testing should be performed in those patients eligible for allogeneic HCT. In patients in whom allogeneic HCT is likely to be indicated, it is also important to commence a search for sibling or volunteer unrelated donor at diagnosis.
r If suspicion of pulmonary infection.
s Required in patients with clinical symptoms suspicious of central nervous system involvement; patient should be evaluated by imaging study for intracranial bleeding, leptomeningeal disease, and mass lesion; lumbar puncture considered optional in other settings (eg, high white blood cell count).
t Cryopreservation to be done in accordance with the wish of the patient.u Pretreatment leukemic bone marrow and blood sample; preferably also normal tissue (eg, skin biopsy, nail clippings).
The scenario of driver mutations in AML reveals the presence of different molecular subgroups, which reflect different pathways in the development of AML and make it possible to classify the disease and make a prognostic stratification at the same.
In 2022 the European Leukemia Network (ELN) updated the risk stratification in three risk categories.
2022 European LeukemiaNet (ELN) risk classification by genetics at initial diagnosisa
a Frequencies, response rates and outcome measures should be reported by risk category, and, if sufficient numbers are available, by specific genetic lesions indicated.
b Mainly based on results observed in intensively treated patients. Initial risk assignment may change during the treatment course based on the results from analyses of measurable residual disease.
c Concurrent of KIT and/or FLT3 gene mutation does not alter risk categorization.
d AML with NPM1 mutation and adverse-risk cytogenetic abnormalities are categorized as adverse-risk.
e Only in-frame mutations affecting the basic leucine zipper (bZIP) region of CEBPA, irrespective whether they occur as monoallelic or biallelic mutations, have been associated with favorable outcome.
f The presence of t(9;11)(p21.3;q23.3) takes precedence over rare, concurrent adverse-risk gene mutations.
g Excluding KMT2A partial tandem duplication (PTD).
h Complex karyotype: ≥3 unrelated chromosome abnormalities in the absence of other class-defining recurring genetic abnormalities; excludes hyperdiploid karyotypes with three or more trisomies (or polysomies) without structural abnormalities.
i Monosomal karyotype: presence of two or more distinct monosomies (excluding loss of X or Y), or one single autosomal monosomy in combination with at least one structural chromosome abnormality (excluding corebinding factor AML).
j For the time being, these markers should not be used as an adverse prognostic marker if they co-occur with favorable-risk AML subtypes.
k TP53 mutation at a variant allele fraction of at least 10%, irrespective of the TP53 allelic status (mono- or biallelic mutation); TP53 mutations are significantly associated with AML with complex and monosomal karyotype.
The incidence of TP53 mutations in AML is around 5-10%.
Accumulating evidence indicates that from both a clinical and molecular perspective, TP53- mutant AML represents a distinct disease entity. Most TP53-mutant cases have complex karyotypes (≥3 aberrations), and in about half, TP53 mutations occur in the absence of other AML-associated gene mutations. Clinically, these myeloid neoplasms are associated with a very poor prognosis. The presence of a pathogenic TP53 mutation (at a variant allele fraction of at least 10%, with or without loss of the wild-type TP53 allele) defines the new entity AML with mutated TP53.