الفهرس 
INTRODUCTIONOnly 14 pages are availabe for public view
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Abstract
Acute myeloid leukemia (AML) is a clonal proliferation of myeloid progenitorcells in the bone marrow and often also in the peripheral blood. These cells are still capable
of selfrenewal but have generally lost the ability to undergo terminal differentiation. In such cases, normal hematopoiesis in the bone marrow is diminished due to the excessive
proliferation of the leukemic cells and may result in anemia, granulocytopenia and thrombocytopenia. AML may emerge de novo, following other hematopoietic
malignancies or after cytotoxic therapy for other disorders. According to the FAB classification, AML can be subdivided into various
subtypes depending on the percentages and cytological features of the bone marrow blasts, erythroblasts and the presence of leukemic promyelocytes into M0, M1, M2, M3, M4,M4eo, M5, M6 and M7. The FAB classification system is useful and is still commonlyused to group AML into subtypes. However, it doesn’t take into account many of the
factors that are known to impact prognosis of patient. The World Health Organization (WHO) has published a system that includes some of these factors to try to help better classify cases of AML, which included, in addition to the morphologic and
immunophenotypic results, clinical features, the cytogenetic and molecular genetic findings. The WHO classification will be more difficult to apply than the FAB classification. Nevertheless, The WHO classification will prove even more useful when therapies become available for the different types of diseases. Non-random clonal chromosome aberrations (i.e., balanced translocations,inversions, deletions, monosomies, and trisomies) are detectable in the leukemic blasts of
approximately 55% of adults with AML and patients can be stratified in groups reflecting
good, intermediate or poor prognosis. Numerous molecular markers have been recently
discovered as potential prognostic factors in acute myeloid leukemia (AML). Internal tandem duplications in the hematopoietic growth factor receptor Flt3 have been shown to
separate a subset of high risk patients from intermediate or low risk cases. Because of its prognostic importance, the mutational status of the FLT3 gene should always be
ascertained in AML patients according to the WHO recommendations. NG2 is a novel cellsurface
antigen expressed on blast cells that is not expressed during normal hematopoitic
development and is implicated as a marker with poor prognosis. Identification of poor
prognosis markers will aid AML patients to receive the most appropriate therapeutic
regimen and involvement in the BMT program.
In an effort to decipher the phenotype, genotype as well as risk stratification of adult patients with AML, our aim was to stratify AML patients into low risk, intermediate or high risk groups according to cytogenetic results to be able to provide the most appropriate
treatment to prevent hazards of unnecessary chemotherapy. Detection of the bad prognostic factors including NG2 expression and FLT3/ITD mutation was performed to allow early enrolment
in bone marrow transplantation programs.
The study involved 50 adult patients diagnosed as AML aged below or over 60 years
who didn’t receive induction chemotherapy. The patients were subjected to the following
tests:
1. History taking: including age and sex
2. Thorough clinical examination
3. Hematological assessment:
A. Complete blood count (CBC) and morphological examination on PB smears
stained by Leishman stain to detect hematological abnormalities.
B. M aspiration:
I. For morphological examination: done by preparation, staining and
examination of bone marrow smears to classify acute myeloid leukemia into
FAB groups.
II. For flow cytometry immunophenotyping to:
• Screen for myeloid markers expressed by AML cells
• Identify aberrantly expressed lymphoid marker in AML
These were held using acute panel of monoclonal antibodies directed against the
following cluster of differentiation (CD) antigens:
1) Lymphoid lineage markers: CD2, CD3, CD5 and CD7 for T-cell lineage; and
CD10 and CD19 for B-cell lineage.
2) Myelomonocytic lineage markers: CD11b, CD13, CD14, CD33, CD64, CD56.
3) Megakaryocytic lineage markers: CD41 and CD61
4) Erythroblasts markers: CD235a (Glycophorin A [GPA]) and CD71
5) Non-lineage specific markers: CD34, CD45, HLA-DR
• Detect NG2 expression.
4. Genetic assessment was done to stratify patients at the appropriate risk group and
included:
A. chromosome examination using conventional cytogenetic technique.
B. Molecular detection of FLT3/ITD mutations using conventional RT-PCR
technique, where RNA was extracted using SV Total RNA Isolation System
(Promega Corporation, Madison, USA) according to the techniqual manual protocol
followed by one step conventional RT-PCR using Maxime RT-PCR PreMix Kit
(Introne Biotechnology, USA) according to the manufacturer protocol.
Summary and Conclusions Hind SA Yagoub, 2014
125
The results of the present study were as follows:
• The median age of all patients at diagnosis was 49 years old (range 17-77 years
old).
• Patients aged <60 years were 78% with a median age of 45 years (range 17-59),
whereas patients aged >60 years 22% with a median age of 66 years old (range 63-
77).
• Fifty four percent were male patients and 46% were females with a male to female
ratio of 1.17
• According to the type of AML, 84% were de novo cases, whereas 16% were
patients with secondary AML.
• De novo AML type is classified by the FAB subclassification system and included:
16% M1 patients, 28% M2, 6% M3 subtypes, 10% M4, 4% M4eo, 16% M5 and
4% M7
• Secondary AML included 10% of cases ontop of CML, 2% ontop of each of MF,
ontop of CMML as well as t-AML (therapy induced).
• Clinical data recorded for patients at presentation included anemia (86%), infection
(48%) and hemorrhagic manifestations (38%) as findings of BM infiltration,
whereas clinical findings for EM infiltration included splenomegaly (54%),
hepatomegaly (38%), lymphadenopathy (26%), gum hypertrophy (12%) and
chloroma (4%).
• Hepatomegaly and splenomegaly significantly more frequent in secondary AML
versus de novo AML patients, whereas infection and bleeding manifestations were
more frequently present in de novo cases.
• As regard to hematological findings at diagnosis, the WBC profile revealed that the
mean WBC count was 29.97X109/L, the mean monocyte count was 6.27%, the
mean PB and BM blasts were 48.86% and 61.72%, respectively; the mean RBC
count was 2.83X109/L and Hb concentration was 8.14%; and the mean platelet
count was 66.64 X109/L.
• The anomalous CD56 marker inferring poor prognosis was expressed in 16% and
the aberrant lymphoid markers CD7 and CD19 were expressed in 16% and 6%,
respectively.
• No patients had expressed the poor prognosis marker NG2.
• This study revealed that EMI were significantly associated with expression of
CD56 and CD7
• As regards to the cytogenetic results and risk stratification
chromosomal abnormalities was detected in 69% of cases, whereas 31% were
cases with normal chromosome complement
Summary and Conclusions Hind SA Yagoub, 2014
126
Cytogenetic risk stratification included 34.6% patients in the intermediate risk
group, while 65.3% were patients in the unfavorable risk group; and no patient
was stratified in the favorable risk group
The median age of patients was 52 years (35-65) in the intermediate risk groups
and 49 years in the unfavorable risk group.
Patients aged <60 years were 33.3% and 66.7% in the intermediate and
unfavorable risk groups, respectively; whereas patients aged >60 years were
37.5% and 62.5% in the intermediate and adverse risk groups, respectively.
The intermediate risk group findings included 31% CN-AML and 3.84%
trizomy 21.
The unfavorable risk group aberrations were 15.3% noncomplex MK, 38.5%
monosomal positive complex karyotype as well as 3.8 each of CK, del(11q23)
and near-tetraploidy.
• As regard to the correlation of the cytogenetic findings with patient characteristics
Females were significantly more frequent in the intermediate risk group,
whereas males were significantly predominant in the poor risk stratification
group
WBC and platelet counts were significantly higher in the intermediate rather
than the adverse risk group
CD56, CD7 and CD19 marker expression were significantly more encountered
in the unfavorable risk group.
Patients in the intermediate risk group significantly achieved CR more than
patients with adverse risk had achieved.
• Concerning the molecular genetic results for the FLT3/ITD aberration detection
Twenty percent of patients under study harbored the mutation with 18% and 2%
in heterozygous and homozygous forms, respectively.
The median age of patients harboring the mutation was 46 years (18.6-70),
where 20% and 18.2 % of the patients were below and above 60 years,
respectively.
12% were males versus 8% females.
Half of the patients with mutation belonged to M1 FAB subtype.
The mutation was significantly associated with higher WBCS count, PB and
BM blasts and the aberrant poor prognostic marker CD7, adding to the adverse
outcome of such patients.
FLT3/ITD mutation was significantly associated with splenomegaly and
hepatomegaly.
• When characteristics of patients with and without EMI were compared, EMI were
significantly associated with males with 1.77:1 male to female ratio, higher WBC
count, M4 FAB subtype and FLT3/ITD mutation.