The term “acute myeloid leukemias” (AML) unites a group of acute leukemias arising from the precursor cell of myelopoiesis and differing in certain morphological, immunophenotypic and cytogenetic characteristics. About 10% of AML have an erythroid or megakaryocytic orientation, therefore the term “acute non-lymphoblastic leukemia” (ONLL) is also considered eligible. Thus, AML, or ONLL, is a group of dissimilar leukemias that require different therapeutic approaches.

Acute myeloid leukemia (AML) is diagnosed at any age, but the frequency of its occurrence increases in older age groups. The median age at which AML is diagnosed is 60–65 years, that is, it is a disease of the elderly. On average, AML affects 2 people per 100,000 population per year.

Unlike ALL, myeloid leukemia may be preceded by a pre-leukemic phase (pre-leukemia, myelodysplastic syndrome). Almost 30-40% of elderly patients with the so-called primary (de novo) ONLL detected myelodysplasia.

For acute myeloid leukemias, the connection with radiation, chemical factors (benzene), alkylating agents (mustargen), and epipodophyllotoxins (etoposide) has been proven more clearly than for ALL. The incidence of AML increases when interacting with substances such as totratrast (radiographic contrast agent), pesticides, dyes, rubber, etc.

The clinical manifestations of the disease are very non-specific. Weakness and malaise may precede the diagnosis many months before it is established. Pallor, dizziness can be manifestations of anemic syndrome. Fever and sweating in the debut of the disease are observed in 15-20% of patients, and they may not be associated with any infectious process, which is also often determined at the onset of the disease (tonsillitis, pneumonia, etc.). Frequent symptoms are those or other manifestations of hemorrhagic syndrome.

Petechial rash, ecchymosis is detected at the time of diagnosis in 50% of patients. Sometimes the only symptom of the disease can be bleeding: uterine or nasal, from the gastrointestinal tract, gums, kidneys, etc.

About half of patients complain of a slight weight loss. Ossalgia is noted in 20% of patients. Organomegaly is not a bright diagnostic sign of AML, but an increase in the size of the liver, spleen, lymph nodes is found in 50% of patients. In 10% of patients, specific skin infiltration is defined – leukemides, and most often in acute myeloblastic and monoblastic leukemias. For these same options, a characteristic feature is gum infiltration.

Diagnosis of acute leukemia is based on the assessment of morphological features of bone marrow cells and peripheral blood. The diagnosis is established only when so-called blast cells in the bone marrow or peripheral blood are characterized by a delicate net structure of nuclear chromatin. The number of blast cells in this case should be 20% or more.

If it is less in the bone marrow, but its content in the blood is 20% or more, then the diagnosis of acute leukemia is also established. Determining the belonging of tumor cells to the myeloid or lymphoid hematopoietic lines when using the usual Romanovsky – Giemsa staining is possible only in 70% of cases. For a more precise definition, other diagnostic approaches are needed: immunophenotyping, cytochemical, cytogenetic, molecular biological and cultural studies.

A new WHO classification can be called a list of diseases characterized by a specific set of features, since it lacks a single classification feature that existed, for example, in developing the FAB classification (morphology and cytochemistry of blast cells). As a result, some forms of OL can simultaneously be assigned to different categories.

Thus, acute leukemias with Hq23 translocations are assigned to the first classification category, but the 11q23 segment anomaly is often detected in patients with secondary acute leukemia. At the same time, secondary acute myeloblastic leukemias are separated into a separate category, so it is not clear how to classify AML, which arose after previous chemotherapy of a tumor and is characterized by anomalies of the 11q23 segment, as a disease of the first category or the third.

There is also a question regarding other consistently detectable chromosomal aberrations: for example, why in the first classification category such anomalies as t (6; 9) and anomalies of chromosome 3 that have clear morphological and clinical characteristics are not considered.

In those situations when it is not possible to analyze the karyotype of blast cells, the only principle by which acute leukemia, morphological, will be classified. Again, it is unclear to which classification category AML is classified, which, for example, according to the FAB classification, is characterized as myelomonoblastic, but arose after prior chemotherapy and in which signs of myelodysplasia are determined in the bone marrow.

The presence of myelodysplasia is the basis for the selection of the second classification category. As noted earlier, morphological signs of dysplasia are detected in patients with different types of AML (myeloblastic, myelomonous and monoblastic, erythroblastic, rarely promyelocytic, etc.), with AML developed from MDS, secondary AML. This pattern was previously noted by other international experts, so it is not quite clear why a special category was selected.

Previously, it was believed that the presence of signs of myelodysplasia determines an unfavorable prognosis, and therefore, it seems that leukemias with these symptoms were considered as a separate category. However, it has now been established that the symptoms of myelodysplasia in the debut of the disease are observed quite often and do not affect the results of therapy.

In the new WHO classification, acute lymphoblastic leukemia is considered in the section of tumors arising from the precursor cells of T and B lymphocytes. In the section of lymphatic tumors from the early progenitor cells are presented:
1) lymphoblastic leukemia / lymphoma from B-lymphocyte progenitor cells (synonym: acute lymphoblastic leukemia from B-cell precursors);
2) lymphoblastic leukemia / lymphoma from T-lymphocyte progenitor cells (synonym: acute lymphoblastic leukemia from T-lymphocyte progenitor cells).

Perhaps the equivalent use of the presented definitions, the authors of the classification only believe that when the content of blast cells in the bone marrow is 25% and it is more expedient to speak of acute leukemia, less than 25% of lymphoblastic lymphoma. However, most often these terminological difficulties are speculative, since the therapy is the same in either case.

In accordance with the FAB classification, the described forms of acute lymphoblastic leukemia were defined as variants L1 and L2, however, at present, these names are practically not used. According to the FAB classification, the third form of ALL, in the modern classification, is assigned to a large section of tumors from mature (determined by immunophenotype) B-cells as Berkitt-like leukemia / lymphoma. Unfortunately, the new classification does not give clear immunophenotypic characteristics of specific sub-variants of B-and T-cell lymphoblastic leukemias, which often causes difficulties in interpreting the results of flow fluorocytometry.

Differential diagnosis of acute lymphoblastic leukemia

The differential diagnosis of acute lymphoblastic leukemia and non-Hodgkin’s lymphomas (NHL) in the blast-type leukemization stage necessitates a detailed immunological study. Those hemoblastosis in which the blasts have the phenotype of early progenitors are designated as ALL / NHL, and the rest – as NHL from peripheral cells. So, when expressing antigens of early B-precursors (CD19, CD20, CD22), TdT enzyme and in the absence of membrane immunoglobulin, B-ALL / NHL is diagnosed, and in the presence of T-precursor phenotype – T-ALL / NHL.

Similar difficulties are encountered in the differential diagnosis of acute non-lymphoblastic leukemia (ONL) and myeloid sarcomas. Currently, myeloid sarcomas in the WHO classification are considered in the section of acute leukemia and can be presented as the first manifestation of the disease with rapid leukemization by the type of acute non-lymphoblastic leukemia (ONLL).

The cellular composition of myelosarcoma can be represented by blasts of various types or by maturing elements of the granulocyte series. Morphocytochemical and immunophenotypic characteristics of blasts, cytogenetic abnormalities are identical to those observed in acute non-lymphoblastic leukemia (ONLL).

In patients with multiple myeloma, plasma cell leukemia may occur. It usually manifests itself as a terminal phase of myeloma, although there are descriptions of cases when it is regarded as an independent disease. The leukemic population in the blood and bone marrow is represented by plasmablasts, as well as protoplasmocytes and plasma cells. All elements of the plasma line are morphocytochemically and immunophenotypically characterized as B cells with clonal expression of immunoglobulins.

Diagnosis of hemoblastosis

Diagnosis of hemoblastosis can be difficult with scanty punctate bone marrow. This can be observed in aplastic conditions, myelofibrosis, MDS with fibrosis or tumor metastases. In cases where a proliferative pathological clone has been identified (myeloblasts, megakaryoblasts), acute leukemia with myelofibrosis is diagnosed, otherwise acute or chronic idiopathic myelofibrosis is diagnosed. In acute panmielosis, in contrast to chronic idiopathic myelofibrosis, young forms of myelopoiesis, including mononuclear megakaryocytes, predominate in the bone marrow.

Acute leukemias also have to be differentiated from blast crises in the inventories and, in particular, CML. Special difficulties are presented when the CML manifests with a blast crisis. The presence of the Ph-chromosome in most cases helps to establish the diagnosis of CML. It should be noted that the Philadelphia chromosome is also detected in rare cases of myeloblastic and in 25% lymphoblastic leukemia, which makes diagnosis difficult. Imperious crises of CML can be myeloid and lymphoid, and differential diagnosis is carried out with the appropriate variant of acute leukemia. The picture of the bone marrow in the myeloid blast crisis of CML is much more variegated than in ONLL: eosinophils, basophils may be present in the granulocyte sprout, pathological microforms of megakaryocytes are found.

Cells in the blast population are more diverse in their morphoimmunological features compared with acute leukemia, myeloblasts, erythroblasts, megakaryoblasts can be detected at the same time, and lymphoblasts in some cases. The picture of lymphoid blast crisis is more monomorphic and similar to that in ALL. Ph-positive clone in CML and ALL is different in its functional features, which is associated with the structural features of BCR / ABL oncogenes in these two hemoblastosis. In ALL, the BCR / ABL gene encodes the abnormal p190 protein, and in CML, the p210 protein is produced. With the help of molecular research methods (PCR reaction) establish an accurate diagnosis.

Certain diagnostic difficulties may also occur in the differential diagnosis of ONLL and metastases of alveolar rhabdomyosarcoma and neuroblastoma in children.

Acute non-lymphoblastic leukemia differentiate

M5a acute nonlymphoblastic leukemia is differentiated from leukemias of the M0, Ml and M7 variants, as well as from acute lymphocytic leukemia. In these cases, the leading criteria are cytochemical and immunophenotypic parameters. A specific feature of monoblasts M5a and M5b is α-naphthyl acetate esterase, which is inhibited by sodium fluoride. Acute monoblastic leukemia with maturity is also differentiated from atypical promyelocytic leukemia without grain. Characteristic cytochemical markers (non-specific esterase and peroxidase) make it easy to distinguish them.

In the differential diagnosis of M6 and other variants of acute non-lymphoblastic leukemia, it is necessary to take into account the possibility of two subvariants of the disease: with the presence of an extended pathological red sprout (erythromyelosis) and with total bone marrow metaplasia with leukemic erythroblasts (erythroid leukemia).

In the first case, differentiated from RAIB MDS. If the number of erythroid progenitors is more than 50%, then the number of blasts should be recalculated to the non-erythroid fraction. In the event that the number of blasts exceeds 20%, M6 is diagnosed with acute non-lymphoblastic leukemia, in the opposite – RIBS MDS. If dysplasia is expressed in more than 50% of myelopoiesis cells, then, at the suggestion of the WHO classification, a variant of acute non-lymphoblastic leukemia with multilinear dysplasia is diagnosed.

In erythroleukemia, differential diagnosis is carried out with megakaryoblastic and lymphoblastic leukemias. The presence of erythroid erythroblasts (NAE3, NAE9, glycophorin A) and the absence of megakaryocytic and lymphoid antigens are taken into account.

Megakaryoblastic leukemia is distinguished from M0 and M6 in acute non-lymphoblastic leukemia based on the expression of specific antigens (CD41 and CD61). It should be noted that leukemic megakaryoblasts sometimes differ in some morphocytochemical features, which indirectly can suggest the need for advanced immunophenotypic studies. Thus, the irregular shape of the cells, the presence of outgrowths of the cytoplasm, its pronounced basophilia, the presence of megakaryocytes in the blood and their nuclei suggest a megakaryoblastic variant of acute non-lymphoblastic leukemia.

In addition, megacaryoblastic leukemia is differentiated from acute panmielosis with myelofibrosis. The presence of more than 50% of blasts expressing specific antigens, and the absence of trilinear myeloid cell dysplasia, make it possible to diagnose acute megakaryoblastic leukemia, even with fibrosis.

Differential diagnostics

Differential diagnosis of M2 for acute non-lymphoblastic leukemia is performed with myeloblastic without maturation, myelomonoblastic and basophilic-cellular variants. In the M2 variant, in contrast to M1, the number of granulocyte maturation cells is more than 10%. In contrast to variant M4, when M2 there are no monoblasts containing nonspecific esterase. In basophilic cell leukemia, unlike myeloid blast, metachromasia is determined in blasts and peroxidase is absent, the serum of patients with elevated heparin and serotonin levels.

Of particular practical importance is the identification of promyelocytic leukemia among patients with acute non-lymphoblastic leukemia, since the treatment for this variant, including ATRA, is fundamentally different from the treatment regimens used for the remaining myeloid leukemia. The leading diagnostic feature of the M3 blasts is the characteristic dvulobolny form of the nuclei. However, the number of such cells in the blast population may be less than 25%, which significantly complicates the differential diagnosis of variants of MH and M2. Only a careful morphological analysis allows to identify characteristic cells. The lack of grit and Auer rods in the M3v variant makes leukemic promyelocytes similar to maturation monoblasts.

The differential diagnosis of M3v and M5b acute non-imphobular leukemia is based on the high activity of granulocyte enzymes in the first case and non-specific esterase in the second.

Diagnosing a rare variant of APL with t (11; 17) (PLZF / RARa), resistant to ATRA, with powerful cells similar to myeloblasts, presents great difficulties for morphologists. Practically, the diagnosis of this variant is based on data from a cytogenetic study. In the diagnosis of myelomonoblastic acute non-lymphoblastic leukemia, the definition of biclonal (most frequent) and biphenotypic variants is necessary. The visual difference between them is determined by carrying out successive reactions on a single smear of the bone marrow, first with ASD chloroacetate and then with α-naphthylacetate.

In the biclonal variant, 2 types of positive cells are detected: ASD-chloroacetate and α-naphthylacetate. In the bifenotypical variant, granules of both enzymes, colored in different (blue and red) colors, are found in all blasts. An increase in the number of atypical eosinophils suggests that the 16th chromosome is abnormal.

Differentiate M4 from options M2 and M5 according to cytochemical studies and the determination of the content of monocytoid cells.

Modern differential diagnosis of acute leukemia variants

The polymorphism of leukemic elements in acute leukemia is pronounced significantly. The presented schematic description of the signs of cells of different types does not include the whole variety of forms, therefore the differential diagnosis of disease variants can cause certain difficulties. Differences in the diagnosis of three morphologists in the study of the same observations can be 5-10%.

One of the most difficult diagnostic options for acute non-lymphoblastic leukemia is myeloblastic with minimal differentiation. It was singled out by the authors of the FAB classification among the latter. This variant of leukemia has to be differentiated from acute lymphoblastic, megakaryoblastic, bi-phenotypic, and true erythroid leukemias.

The difference between M0 and acute lymphoblastic leukemia and large cell lymphoma in the leukemization stage lies in the expression of myeloid (CD33, CD13) and the absence of lymphoid antigens. PAS-positive substance is more often defined in diffuse, but not in granular form.

In the differential diagnosis of MO variants from M6 and M7, the criterion is the absence of specific immunological markers of the erythroid (GlyA) and megakaryocytic (CD41, CD61) series. In contrast to biphenotypic leukemia, only myeloid antigens are detected on blasts and lymphoid antigens are absent.

In myeloblastic leukemia without maturation, two types of blasts can occur. In blasts of type I, a large nucleol is determined in the nucleus, primary granules are absent, the nuclear-cytoplasmic ratio is high. For blasts of type II, small nucleols are characteristic, the presence in the cytoplasm of several primary granules, a moderate nuclear-cytoplasmic ratio. Acute myeloid leukemia with type I blasts with a low peroxidase content (less than 10%) is differentiated from acute lymphoblaqueosis. In these cases, the data of immunophenotypic research, namely the expression of myeloid antigens CD33 and CD13, are very significant.

In type II myeloblasts, differential diagnosis is performed with the M2 variant. The presence in the bone marrow of less than 10% of ripening granulocytes gives grounds to establish the variant Ml, and not M2 of acute non-lymphoblastic leukemia.

Myeloblastic leukemia with maturation is heterogeneous in its characteristics. Blasts vary in size, number of forms with granularity, degree of disgranulopoiesis, cytogenetic parameters. Comparison of M2 patients with acute non-lymphoblastic leukemia with t (8; 21) and expression of the AML / ETO gene and the absence of this anomaly showed that in the former, compared to the latter in the bone marrow, Auer’s rods were more common, the peroxidase activity was higher, trophilia was determined by pink staining, the number of eosinophils was slightly increased (2–6%).

It should be noted that, on the basis of morphological parameters without data from the karyotype study, it is difficult to distinguish these groups of patients. The clinical course of leukemia with this translocation proceeded in patients more favorably than in its absence.

Acute lymphoblastic leukemia (ALL) – diagnosis

Acute lymphoblastic leukemia (ALL) includes leukemias, the tumor substrate of which is represented by progenitor cells of the lymphoid series. There are two variants of leukemia, depending on the direction of differentiation of blast cells: acute B-lymphoblastic leukemia / lymphoma from progenitor cells and acute T-lymphoblastic leukemia / lymphoma from progenitor cells.

Most of the observations are regarded as acute leukemia, however, cases with a pronounced extramedullary lesion and a number of blasts in the bone marrow of less than 25%, according to the WHO classification, belong to lymphomas.

The presence of individual cytogenetic abnormalities in acute lymphoblastic leukemia (ALL) has significant prognostic significance, therefore, in B-acute lymphoblastic leukemia (ALL), a number of variants are distinguished depending on the characteristics of the karyotype.
1. B-lymphoblastic leukemia / precursor lymphoma: t (9; 22) (q34; qll); BCR / ABL; t (v; llq23); MLL restructuring; t (l; 19) (q23; pl3) PBX / E2A; t (12; 21) (pl2; q22) TEL / AMN; ALL hypodiploid; ALL hyperdiploid (> 50).
2. T-lymphoblastic leukemia / precursor lymphoma.

The allocation of morphological types of lymphoblasts according to the FAB classification depending on their size [prevalence of microform L1 or mesoform P2] did not play a significant role in determining treatment tactics or in the prognosis of the disease. The type of blasts with basophilia and vacuolation of LZ cytoplasm is more characteristic of Burkitt’s lymphoma and is not specific for progenitor cells. The morphocytochemical signs of B- and T-lymphoblasts have some differences: • B-lymphoblasts are characterized by significant morphological diversity. Cells can be regular or elongated, sometimes they resemble the shape of a hand mirror. The size of the blasts varies from micro to mesoform. The nuclei are round, oval or folded, with dense chromatin and rare nucleoli. The cytoplasm is scanty, slightly basophilic, sometimes contains a slight granularity. The nuclear cytoplasmic ratio is high or moderate.

• B-lymphoblasts do not contain peroxidases, lipids and ASD-chloroacetate esterase. In half of the cases, small granules of PAS-positive substance and acid phosphatase in a fine granular form are found in them.

• T-lymphoblasts are usually monomorphic, medium-sized, rounded, with irregularly folded nuclei. The cytoplasm is scanty, moderately basophilic. The nuclear cytoplasmic ratio is high.

• T-lymphoblasts do not contain peroxidase, ASD-chloroacetate esterase and lipids. PAS-positive substance is rarely found in the form of a single granule. A characteristic feature is the presence of large granules of enzymes: acid phosphatase, nonspecific esterase, and acid nonspecific esterase.

Depending on the stage of lymphoblasts differentiation, subvariants of B-and T-linear acute lymphoblastic leukemia (ALL) are distinguished. The diagnosis is based on cell immunophenotyping data. The frequency of detection of these subvariants is different. Different chromosomal abnormalities are characteristic of different subvariants.

Leukemia from B-linear progenitor cells occurs in approximately 75% of adult patients with acute lymphoblastic leukemia (ALL). In all cases, blast cells express CD19 and / or CD79a and / or cytoplasmic CD22 molecules. As differentiation to these antigens are added, new, specific to the subsequent stages of maturation. Among B-linear leukemias, the following immunosupportants are distinguished: pro-B, pre-pre-B, pre-B, and B-ALL. The presence of t (9; 22), t (4; ll) and t (l; 19), observed in pro-B and pre-pre-B variants, is an independent indicator of an unfavorable prognosis.

Translocation (9; 22) occurs in adult patients with acute lymphoblastic leukemia (ALL) quite often (about 25%). Ph-positive and Ph-negative lymphoblasts do not have morphocytochemical and immunophenotypic differences, therefore the data of cytogenetic studies are necessary when choosing the tactics of treatment in patients with acute lymphoblastic leukemia (ALL). The prognostic value for subvariants of B-linear acute lymphoblastic leukemia (ALL) also has a change in the number of chromosomes in the blasts: hyperploidy is favorable, and hypoploidy is an unfavorable sign.

T-linear variants occur in approximately 25% of cases of acute lymphoblastic leukemia (ALL). Two immunosuppressants are distinguished: early pre-T and mature T-ALL. Both of them are characterized by the expression of the CD7 antigen. Pre-T-acute lymphoblastic leukemia (ALL), in turn, is subdivided according to the presence of the “common” antigen into CD10-positive and CD10-negative subvariants. T-linear leukemias are characterized by rearrangements of T-cell receptor genes located on the 14th and 7th chromosomes, in particular t (10; 14), t (ll; 14), t (l; 14) and inv14.

Acute leukemia of unclear differentiation line – diagnosis

A special group is made up of acute leukemias, in which the cells simultaneously exhibit a myeloid and lymphoid orientation of differentiation. In this case, two subclones of cells can be determined, each of which expresses markers characteristic of a single line. These leukemias are called biclonal. In other cases, the blasts coexpress both myeloid and lymphoid or B-or T-lymphoid antigens. This variant of the disease is called biphenotypic acute leukemia (BAL).

Myeloid differentiation is determined by the presence of peroxidase, esterase and / or expression of myeloid (CD13, CD33 and CD65) antigens in blasts. B-lymphoid differentiation is detected by expression of the antigens CD79a, cytoplasmic CD22, cytoplasmic IgM, CD19, CD10 and CD20. T-lymphoblasts are characterized by reactions with MCA to CD3, TCR, CD2, CD5, CD8.

In order to make a diagnosis of biphenotypic leukemia, it is necessary that a positive result be observed with at least two ICA of the same line. It should be noted that some antigens (T-linear CD7, CDla and B-linear CD24) are often present on myeloid cells and cannot serve as markers of the bifenotype. Due to the difficulties in interpreting the results, it became necessary to unify the diagnosis of hybrid leukemia. According to the BAL system, acute leukemia is considered biphenotypic if the sum of cells with myeloid and lymphoid markers exceeds 120. The presence of mixed differentiation is sometimes found only at the molecular-biological level.

It has been shown that during ONLL, rearrangements of the genes of immunoglobulin chains or T-cell receptor genes can be detected.

Biphenotypic leukemias are characterized by a high frequency of cytogenetic abnormalities. In a third of cases of B-lymphoid / myeloid proliferation, the Philadelphia chromosome is determined by t (9; 22), in separate cases — 1 (4; 11) (q21; q23) and anomalies llq23. In patients with T-lymphoid / myeloid biphenotypic leukemia, complex abnormalities of the karyotype are often found.