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1. The first step in AML diagnosis to establish the primitive nature of the abnormal cells:

The primitive nature of blast cells in AML may be confirmed by a number of approaches:

The use of CD45 together with other properties (most often side scatter) to distinguish primitive cells

The use of markers associated with early differentiation, most often CD34 although others are often helpful

The identification of lineage markers typically acquired by more primitive myeloid cells

MARKERS USEFUL TO CONFIRM PRIMITIVE NATURE OF BLAST CELLS
CD45	A marker expressed by all leukocytes and their precursors. In AML expression is characteristically "weak" i.e. significantly less intense than normal lymphocytes or monocytes. In monocytic AML expression may be stronger.
CD34	Frequently expressed by AML blast cells (40-80% of cases) - most often in less differentiated forms of AML. Expression is also seein frequently (and often more strongly) in ALL
Other markers that may be associated with primitive nature in AML
In the context of a proven AML diagnosis a number of markers may be expressed that are considered to predominantly reflect their primitive nature. Some of these are more frequently associated with lymphoid disorders, but providing other criteria for AML are met their presence does not change lineage assignment. These include CD38, HLA-DR, TDT, CD7.

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MARKERS USEFUL TO CONFIRM MYELOID LINEAGE OR DIFFERENTIATION
Frequently expressed "general" markers of myeloid lineage These markers may be considered the most frequent set of markers expressed by AML. Having said that the expression of any single marker is not required for the diagnosis of AML and it is recognised that (excluding MPO), and each may be expressed by cells of ALL. General concepts: Expression of MPO is lineage defining and should indicate AML (or MPAL if lymphoid lineage also confirmed) In the absence of markers suggesting alternative lineage any of the remaining markers (CD117, CD33, CD13) may (alone) be used to support a diagnosis of AML Expression of one or more of CD117, CD33, CD13 markers on otherwise typical ALL is allowed and does not change lineage assignment
MPO	A lineage-defining marker in AML when expressed (around 80% of cases). More frequent in cases with granulocytic maturation. When detected by flow cytometry is diagnostic of myeloid differentiation (either AML or MPAL)
CD117	An early marker of myeloid lineage, seen in up to 80% of AML and vauable in recognising more primitive differentaiion forms (note that aberrant expression is seen in up to 20% of ALL cases)
CD33	A good marker for AML, particularly for those cases with granulocytic maturation, CD33 is often less strongly expressed in AML with monocytic dfferentiation and strongly expressed in APL.
CD13	A good lineage marker for AML that is acquired a little later in differentation than CD117 or CD33; expression of CD13 is often higher than CD33 in AML with monocytic differentiation.

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MARKERS USEFUL TO CONFIRM MYELOID LINEAGE OR DIFFERENTIATION
Confirmatory markers of AML or markers of subtype Some additional markers are regarded as indicating primitive nature when expressed by AML blasts; these markers may be more usually assocated with T-lineage or B-lineage ALL, but when expressed in AML they are considered to indicate less differentiated forms.
Granulocytic lineage markers
CD10	One of tne of the first lineage-markers acquired in AML with good relative specificity
CD11b	One of tne of the first lineage-markers acquired in AML with good relative specificity
Monocytic lineage markers
CD11c	One of tne of the first lineage-markers acquired in AML with good relative specificity
CD14	A good marker for AML that is often less strongly expressed in monocytic forms
CD64	A good lineage marker for AML aquired a little later, higher than CD13 in monocytic forms
Features associated with erythroid differentiation
Most often CD34, CD45 and HLA-DR are weak or negative, although CD117 and CD36 are generally expressed. There may be some expression of platelet markers in some cases.
CD71	Frequently expressed though not fully lineage specific
CD235	A good marker of erythroid differentiation but acquired late and therefore may not be expressed
Features associated with megakaryocytic differentiation
Most often CD34, CD45 and HLA-DR are weak or negative, although CD13 and CD33 may be expressed
CD41	Platelet glycoprotein IIbIIIa
CD61	Platelet glycoprotein IIIa
CD42b	Platelet gycoprotein Ib
CD36	Relatively non-specific (seen in erythroid and monocytic leukaemias) but often strongly expressed

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NON MYELOID MARKERS FREQUENTLY "ABERENTLY" EXPRESSED IN AML
Markers of lymphoid lineage indicating primitive dfferentiation in AML Some additional markers are regarded as indicating primitive nature when expressed by AML blasts; these markers may be more usually assocated with T-lineage or B-lineage ALL, but when expressed in AML they are considered to indicate less differentiated forms.
TDT	Expressed in some cases of AML (5-20%) particularly in less differentiated blast cells, often on a sub-population of cells. More typically associated with primitive ALL blast cells.
CD7	Predominantly seen as a T-cell antigen, CD7 tends may be expressed by AML blasts (20-40%) where it most often indicates a more primitive forms or MPAL. CD7 is most consistently a marker of T-lineage (including T-ALL and more mature froms).
Markers of lymphoid differentation abererantly expressed in AML Some additional markers are regarded as indicating primitive nature when expressed by AML blasts; these markers may be more usually assocated with T-lineage or B-lineage ALL, but when expressed in AML they are considered to indicate less differentiated forms.
TDT	Expressed in some cases of AML (5-20%) particularly in less differentiated blast cells, often on a sub-population of cells. More typically associated with primitive ALL blast cells.
CD7	Predominantly seen as a T-cell antigen, CD7 tends may be expressed by AML blasts (20-40%) where it most often indicates a more primitive forms or MPAL. CD7 is most consistently a marker of T-lineage (including T-ALL and more mature froms).

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IMPORTANT ADDITIONAL POINTS 1
AML subtypes with distinctive phenotype The first step in AML diagnosis to establish the primitive nature of the abnormal cells: (1) Typically AML blasts have low CD45 expression and cause low side scatter. This means that they form a relatively uniform and distinctive population that is clearly separate from that of lymphocytes on CD45/SSc plots (for further details see this section) also not all AML forms may fit this pattern - particularly APL and monocytic AML. (2) Additional markers often expressed on myeloid cells may help confirm the primitive nature of the cells: in AML this is most often CD34, although other markers may contribute.
Mixed Phenotype Acute leukaemia (MPAL)
Formerly known as leukaemia of ambiguous lineage
Early T-precursor acute lymphoblastic leukaemia (ETP-ALL)
A (relatively new entity)

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IMPORTANT ADDITIONAL POINTS 2
Acute leukaemias with ambiguous lineage features The first step in AML diagnosis to establish the primitive nature of the abnormal cells: (1) Typically AML blasts have low CD45 expression and cause low side scatter. This means that they form a relatively uniform and distinctive population that is clearly separate from that of lymphocytes on CD45/SSc plots (for further details see this section) also not all AML forms may fit this pattern - particularly APL and monocytic AML. (2) Additional markers often expressed on myeloid cells may help confirm the primitive nature of the cells: in AML this is most often CD34, although other markers may contribute.
Mixed Phenotype Acute leukaemia (MPAL)
Formerly known as leukaemia of ambiguous lineage: Myeloid MPO or monocytic defined by at least two markers (CD14, CD11c, CD64, NSE, lysozyme. T lineage CD3 (s or m), STRONG Cd19, with strong CD79a cCD22 or CD10 OR wk CD19 also with obove
Early T-precursor acute lymphoblastic leukaemia (ETP-ALL)
A (relatively new entity)
Undifferentiated acute leukaeemia
A (relatively new entity)
Non-haematopoietic tumours
need to know problems

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