ACUTE LEUKEMIAS

Acute leukemias (AL)

A heterogeneous group of diseases
characterized by proliferation of
immature (blastic) cells in bone marrow
and extramedullary tissues.
Normally, blasts are absent in peripheral blood
(PB) and are <5% in the bone marrow (BM)
The cytological criterium for AL diagnosis:
>20% blasts in BM

Blastic proliferation results in

suppression of normal clones and bone
marrow failure:
Anemia
Infection

Multipotential hematopoietic stem cell

(Hemocytoblast)

Hematopoiesis in Humans
Common
myeloid
progenitor

Megakaryoblast

Common
lymphoid
progenitor

Proerythroblast
(Pronormblast)

Myeloblast

Basophilic erythroblast

B. promyelocyte

N. promyelocyte

Lymphoblast

E. promyelocyte

Monoblast

Promegakaryoblast

Prolymphocy
te

Polychromatic erythroblast

B. myelocyte

N. myelocyte

E. myelocyte

Orthochromatic
erythroblast
(Normoblast)

B. metamyelocyte

N. metamyelocyte

E. metamyelocyte

Promonocyte

Megakaryocyte
Small lymphocyte
Polychromatic
erythrocyte
(Reticulocyte)

B. band

N. band

Natural killer cell

E. band

T lymphocyteB lymphocyte

Thrombocytes

Erythrocyte

Basophil

Mast cell

Neutrophil

Eosinophil

Monocyte

Myeloid

Lymphoid

 AL ~ 10% of hematological malignancies

The first cause of death by cancer in
subjects <35 years old.
According to the origin of blastic cells, AL
are subdivided in 2 large groups:
A. Acute lymphoblastic leukemia (ALL),
resulting from the transformation of the
multipotent lymphoid stem cell.
B. Acute myeoid leukemia (AML) resulting
from the transformation of the multipotent
myeloid stem cell

 Epidemiology
1. ALL
Incidence: 0,5-1,5/100.000/year.
ALL affects mostly children and young
adults
It is the most frequent malignancy <15
years
Afterwards, ALL incidence decreases
gradually: in the elderly, ALL is very
rare

2. AML
Incidence: 5-8/100.00/y increases with
age
2/100,000/year < 40 years

 Etiology.
Unknown
Genetic factors:
AL is more frequent in children with congenital
diseases such as Down syndrome, Fanconi
anemia, Li-Fraumeni syndrome, Klinefelter
syndrome.
Environmental factors
ALL the exaggerate hygiene hypothesis
AML
Exposure to radiation, solvents
Previous chemotherapy/radiotherapy for
other cancers
Secondary AML

 Pathogenesis

Genomic abnormalities appearing in a

single stem cell (leukemic stem cell).

Loss of differentiation capacity (maturation

arrest)
Increased proliferative capacity

Emergence of a growth advantage for

the leukemic clone compared to normal
clones: gradual replacement of
hematopoiesis by leukemic cells
AL requires several sequential genomic
abnormalities:

Example: the most frequent genetic

abnormality in ALL, the t(12;21) transocation,
results in the formation of a hybrid gene (TELAML1). This abnormality alone is not
leukemogenic but leads to genetic instability
with the appearance of new myutations that

AML pathogenesis The 2-hit theory (2002)

Normal hematopoiesis

1st hit

2nd hit

Leukemic clone

Class II
mutations
CEBPA, RARA,
RUNX1, MLL,
WT1
MATURATION ARREST

Class II
mutations
FLT3, RAS, KIT
PROLIFERATION

AML Pathogenesis 2014 Complex picture

Class I
mutations
(proliferation)

Class II
mutations
(maturation arrest)

FLT3

RUNX1

RAS

PML-RARA

Kit

CBF

JAK2

CEBPA

Class III mutations

(epigenetic changes)

IDH 1,2

DNMT

TET2

MLL

Class V mutations
Class IV
mutations
(Intercellular
adhesion/interactions)

(DNA repair tumor

suppressor genes)

NPM1
TP53

 Clinical picture
Usually sudden onset - main symptoms:
Anemia
Bleeding: petechiae, echymoses, mucosal bleeding
Infections

Other frequently encountered symptoms:

Bone, joint pain
Lymphadenopathy, hepatosplenomegaly - ALL
Rarely cutaneous infiltration - purple nodules ALL,
AML (monocytic)
Neurologic symptoms due to CNS leukemic
infiltration - ALL
Testicular infiltration more frequent in children,
ALL
Gingival hypertrophy more common in AML
(monocytic)

ALL Clinical aspects

AML Clinical
aspects

 Laboratory data
Blood counts:

Anemia
Thrombocitopenia
WBC: high, normal or low.

Peripheral blood

Presence of immature cells (blasts)

Usually low neutrophil count (<1500/l)

Bone marrow

Blasts >20%
Reduced normal precursors

Lymphoblast vs. mature

lymphocyte

Lymphoid blasts in a case of

common ALL

Vacuolated blasts in a
case of Burkitt type
ALL

Myeloblast vs.
neutrophil

Monoblast vs.
monocyte

Blast with Auer body

AML2

Promyelocyte with
multiple Auer bodies
AML3
Abnormal erythroblast
AML6

Monoblasts
AML5

 Immunophenotyping by flow
cytometry: esential for diagnosis
Lymphoid markers:

B lymphoid markers (CD10, CD19, CD20)

T lymphoid markers (CD3, CD7)
Early lymphoid markers (HLA-DR, TdT)

Mieloid markers

Granulocytic CD13, CD33

Monocytic CD14, CD68
Megacaryocytic CD41, CD61
Erythrocytic glycophorin 1 (CD235)

Stem cell marker - CD34

Flow citometry in in a case of precursor B ALL: CD10, CD19, TDT, HLA-DR poz, CD20, CD7,

Flow-cytometry in AML: CD13, CD33, CD117

poz, HLA-DR, CD10, CD7 neg

 Cytogenetic aspects (karyotype).

ALL

Hyperdiploidia (>47 chromosomes) in children

no impact on prognosis.
Translocations:
t(12;21) - good prognosis
t(9;22), (Ph1) - 15% of ALL in adults like in
CML leads to the BCR-ABL fusion gene
good response to TKI but most cases
relapse bad prognosis
t(4;11) bad prognosis
Normal karyotype good prognosis

 AML Karyotype defines 3 prognostic groups:

Good prognosis: t(8;21), t(15;17), inv16, t(16;16): 10-15% of

cases
Bad prognosis: abnormalities of chromosomes 3, 5, 7,
multiple abnormalities (complex karyotype): 25-30% din
cazuri
Intermediate prognosis: normal karyotype, other
abnormalities: 40-50%

 AML Molecular genetics (PCR,

sequencing): detects genomic abnormalities
that are not visible by karyotyping useful
especially in normal cytogenetic cases
(NC-AML)
FLT3 (FMS-like tyrosine kinase-3)
mutations
20-30% af all cases
25-40% of NC-AML
Bad prognosis.

NPM1 (nucleophosmin 1) mutations

20-30% of all cases
40-55% of NC-AML
Good prognosis (when not associated with FLT3
mutations)

Many other mutations..

AML - t(8;21)
translocation
with the
formation of the
AML1-ETO
fusion gene
(RUNX1RUNX1T) on chr
21

AL Classification
ALL Immunophenotipic criteria
B-cell ALL ~ 80% of cases
Precursor B type (pB)

proB - (5% of children, 10% of adults) HLA-DR+,

TdT+, CD19+, CD34+
common - (65% of children, 50% adults) HLADR+, TdT+, CD19+, CD10+
preB (15% children, 10% adults) HLA-DR+,
TdT+, CD19+, CD10+/-, cytoplasmatic IgM (cIgM) +

Matur B type (3% of children, 5% of adults)

HLA-DR+, CD19+, CD20+/-, CD10+/-, sIgM + ;
cytologically Burkitt-like

T-cell ALL ~ 20% of cases

Precursor T (1% of children, 7% of adults)

TdT+, CD3+, CD7+
Mature T (11% of children, 17% of adults)
TdT+, CD3+, CD1a+, CD5+

 AML 2 classifications are used

1. FAB (French-American-British) classification
(the 80).
M1 = myeloblastic undifferentiated
M2 = mieloblastic with differentiation (>10%
maturation)
M3 = promyelocytic (APL)
M4 = myelo-monocytic: monocytic component
(monoblasts - monocytes) >20% in BM
M5 = monoblastic: monocytic component >80%
M6 = erythroleukemia: Ebl represent >50% of BM
cells
M7 = megakaryoblastic

The FAB classification is no longer used:

Does not take into consideration cytogenetic

abnormalities
Is not useful in taking decision (except APL - M3)

2. The 2008 WHO AML classification

AML with recurrent genetic abnormalities

AML with t(8;21) (q22;q22 [RUNX1-RUNX1T1]

AML with inv(16)(p13.1;q22) or t(16;16)(p13.1;q22) [CBF/MYH11]
Acute promyelocytic leukemia with t(15;17) (q12;q22)
[PML/RARA]
AML with t(9;11)(p22;q23) [MLLT3-MLL]
Provisional entities molecularly defined:
AML with NPM1 mutations
AML with CEBPA mutations

AML with myelodysplasia - related changes (MRC)

AML and MDS, therapy related
AML not otherwise categorized

AML minimally differentiated

AML without maturation
AML with maturation
Acute myelomonocytic leukemia
Acute erythroid leukemia
Acute megakaryocytic leukemia
Acute basophilic leukemia
Acute panmyelosis with myelofibrosis
Vardiman et al. 2008

 Acu leukemia prognosis

1. ALL
Very good prognosis in children: 70-80% cure
after chemotherapy.
In adults prognosis is much worse:
20-30% cure after chemotherapy
40-50% after allogeneic stem cell transplant (SCT)

Prognostic factors in ALL

Bad:
WBC > 40.000/l
T and B mature phenotype
CNS involvement
Good:
Common immunophenotype

2. AML bad prognosis median survival <1

year
Prognostic factors
Good cure in 30-50%:

Age <60 years

FAB M3 (APL) with t(15;17) 70-80% cure
t(8;21) and inv16 cure 40-60%
NPM1 mutations

Bad cure 5-10%:

Age >60 de ani

AML with myelodysplasia - related changes (MRC)
AML therapy related
Bad karyotype
FLT3 mutations

 AL treatment
Treatment is very complex: supportive measures,
chemotherapy, transplant
The first step towards cure: Complete Remission
(CR)
CR can be defined as apparent cure (1000X
reduction of number of tumor cells).

CR criteria:
No lymphadenopathy, no hepato/splenomegaly.
PB: neutrophils >1500/l, platelets
>100.000/l, no blasts
<5% blasts n BM
Normal cerebrospinal fluid (CSF).
Normal karyotype

 General supportive measures:

Isolation
Hygene
Hyperuricemia prophylaxis: allopurinol,
rasburicase, alcalinisation.
Antibiotics, antifungals, antivirals
Transfusions:
Red cells : Hgb<7g/dl
Platelets: Plt <10-20.000/l.
Growth factors.
In severe neutropenia (<500/l): GCSF

Chemotherapy
1. ALL standard chemotherapy
Several steps:
Remission induction
Combinations: antracyclines, vincristine,
L-asparaginase, corticoids
CNS prophylaxis
MTX, Ara-C i.t.
Consolidation
High dose MTX, Ara-C
Maintenance up to 3 years
Low dose 6MP, MTX

GMALL
protocol

1.

Induction 1.

Vincristine (iv) 2mg, days 1, 8,15,22

Doxorubicin (iv, 30min), 25 mg/m2, days 1, 8,15,22

L-Asparaginase (iv, 30min), 5000 IU/m2, days

15,17,19,21,23,25,27

Prednisone (po) 60 mg/m2, days 1-28

Methotrexate (i.t.) - 15 mg, day 1

2. Induction 2.

Cyclophosphamide (iv) - 650 mg/m2, days 29, 43,57

Ara-C (iv, 1h) - 75 mg/m2, days 31-34, 38-41,45-48, 52-55

6-Mercaptopurine (po) - 60 mg/m2, days 29-57

Methotrexat (i.t.) 15 mg, days 31, 38,45,52

3. Consolidation 1. (HDARAC+Mitox) + (HDMTX+Asp+6MP)

Ara-C (iv, 3h, at 12h) - 1000 mg/m2, days 1-4

Mitoxantrone (iv, 30min) 10 mg/m2, days 3-5, then, after

hematological recovery:

Methotrexate (iv, 24h) - 1500 mg/m2, days 1, 15 +

Leucovorin

L-Asparaginase (iv, 30min) - 10000 U/m2, days 2,16

6-Mercaptopurine (po) - 25mg/m2, days 1-5, 15-19

4. Reinduction 1.

Vincristin (iv) 2mg, days 1, 8,15,22

Doxorubicin (iv, 30min) - 25 mg/m2, days 1, 8,15,22

Prednison (po) 60 mg/m2, days 1-28

MTX 15 mg + Ara-C 50 mg + Dexamethasone 4 mg (i.t.) d

1
5. Reinduction 2.

Cyclophosphamida (iv) - 650 mg/m2, day 29

Ara-C (iv, 1h) - 75 mg/m2, days 31-34, 38-41

6-Thioguanina (po) - 60 mg/m2, days 29-57

MTX 15 mg + Ara-C 50 mg + Dexametazona 4 mg (i.t.)

day 29
6. Consolidation 2.

Etoposide (iv, 1h) - 100 mg/m2, days 1-5

Ara-C (iv, 1h) - 150 mg/m2, days 1-5, then, after

hematological recovery:

Cyclophosphamide (iv) - 1000 mg/m2, day 1

Ara-C (iv, 24h) - 500 mg/m2, day 1

MTX 15 mg + Ara-C 50 mg + Dexametasone 4 mg (i.t.)

day 1
7. Maintenance 2 years.

6-Mercaptopurine (po) 60mg/m2, weekly, days 1-5

Methotrexate (po) 12,5mg/m2, weekly, day 6

HyperCVAD protocol

1. Cycle A.

Cyclophosphamide (iv, 3h, at 12 h) 300mg/m2,

days 1,2,3

Methotrexate (i.t.) 15 mg, day 2

Doxorubicin (iv, 30min) 50mg/m2, day 4

Vincristine (iv) 2mg, days 4,11

Dexamethasone (iv or po) 40mg days 1-4 and

11-14

Cytarabine (i.t.) - 70mg, day 7

2. Cycle B

Methotrexate (iv, 24h) 1000mg/m2, day 1

Leucovorin (iv, 24 h after MTX) 6 doses every 6

h

Ara-C (iv, 2 f, at 12 h) 3000mg/m2, days 2,3

A total of 8 cycles (4A + 4B)
3. Maintenance POMP 2 years

Vincristine (iv) - 2mg, day 1

Prednisone (po) 60mg/m2, days 1-5

6-Mercaptopurine (po) 60mg/m2, weekly, days

1-5

Methotrexate (po) 12,5mg/m2, weekly, day 6

Treatment of certain ALL

subtypes:
Ph1-ALL [t(9;22)]: Tyrosine kinase
inhibitors - imatinib 600-800mg/day
or dasatinib 140mg/day are added,
a la longue.
B-ALL(Burkitt), CD20+: monoclonal
anti-CD20 antibodies (rituximab) are
added

2. AML Standard chemotherapy

(except M3):
Remission induction: 12 cycles 3/7 regimen
Idarubicin 12mg/m2 days 1,3,5
Cytarabine (Ara-C) 100-200mg/m2, days 1-7
Remission consolidation: High-dose Ara-C 3-4 cycles
Ara-C (iv 3h, every 12 ore) 3g/m2 days 1,3,5

M3 (Acute promylocytic leukemia, APL)

Induction:

All-trans retinoic acid (ATRA) 45mg/m2, p.o, day 1 till CR

(usually 14-30 days) induces leukemic cell differentiation

Idarubicin 12mg/m2, days 2, 4, 6, 8

Consolidation:

Idarubicin 5mg/m2days 1-4, repeated at 28 days, 3 cycles

Maintenance up to 2 years from onset

- 6MP 90mg/m2/day, p.o. Days 1-5 of every week

 Stem cell transplant

Allogeneic stem cell transplant (allo-SCT)
Indicated as soon as possible, in 1st CR:
ALL
ALL Ph+
In adults with bad prognostic factors (<60
years)
AML
All intermediate/bad risk patients <60 years
In relapse/refractory disease all patients <60 ani

Autologous stem cell transplant (auto-SCT)

When no donor is found for allo-SCT
It is not clear if there is an advantage over
chemotherapy alone