الفهرس 
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Abstract
Acute lymphoblastic leukemia (ALL) is a malignant disorder of lymphoid progenitor cells that proliferate and replace the normal hematopoietic cells of the bone marrow. These lymphoblasts replace the normal bone marrow elements that result in a marked decrease in the production of normal blood cells.
There are many prognostic factors in ALL such as age, sex, leukemic burden, laboratory criteria (initial TLC, hemoglobin level and platelet count), immunophenotyping, cytogenetic profile, duration of induction of remission, drug resistance profiles, and minimal residual disease. Assessment of these factors is mandatory for therapeutic assignment.
Cytogenetic abnormalities are independent prognostic variables that predict the outcome of adult ALL patients. Recent genomic studies have provided a refined genetic map of ALL and increased the number of potential prognostic markers.
The Philadelphia chromosome (ph1) results from a translocation involving the break-point cluster region of the BCR gene on chromosome 22 and the ABL gene on chromosome 9. Ph1 positive ALL represent a high-risk cytogenetic subset, accounts for 25-30% of adult ALL cases and this chromosome is known to be associated with the worst prognosis among patients with ALL.
The use of ES-FISH probes in interphase nuclei of a large series of BCR/ABL+ve leukemias is associated with the observation of a variable number of different interphase FISH patterns. The most frequently detected patterns corresponded to typical BCR/ABL gene rearrangements involving the MBCR and the mBCR breakpoints. Discrimination between these two breakpoint regions could not be achieved with the single fusion or double fusion D-FISH probe. Interestingly, additional chromosomal abnormalities (eg; supernumerary Ph, gain or loss of chromosomes 9 and 22, as well as deletions of 9q and 22q) can occur in BCR/ABL+ve ALL patients.
The present study aimed to detect BCR/ABL genes fusion, amplification, deletion and/or other aberrations in acute lymphoblastic leukemic patients, using extra signal fluorescence in situ hybridization (ES-FISH), and to assess their relation with other standard prognostic factors and therapeutic response.
The current study was carried out on 39 newly diagnosed adults ALL patients. Informed consent was obtained from patients to use their samples in this study. Patients were evaluated at day 14 of therapy to assess therapeutic response. All patients were subjected to the following:
A. History and clinical examination laying stress on the presence of hepatosplenomegaly, lymphadenopathy and CNS infelteration.
B. Laboratory investigations, which included:
1. Complete blood count using Sysmex XN-1000 & SA-01.
2. Examination of Leishman stained PB smears laying stress on differential leucocytic count , assessment of blast cell number and morphology.
3. Bone marrow aspiration and examination of Leishman stained smears.
4. Immunophenotyping on BM or PB samples, using acute leukemia panel performed on EPICS XL Coulter Flow Cytometer, USA.
5. Fluorescence in situ hybridization using the following probes:
- LSI dual color single fusion and double fusion BCR/ABL probes for detection of t(9;22)(q34; q11).
- LSI dual color extrasignal BCR/ABL probe for detection of t(9;22) with other aberrations as; amplification, deletion or duplication.
- LSI dual color double fusion TCF2/PBX1 for detection of t(1;19)(q23;p13.3).
- LSI dual color breakapart rearrangement MLL probe for detection of 11q23 rearrangement.
In this work, BCR/ABL fusion observed with a percentage of 28.2% (11/39 cases) in adult ALL patients. The ph1 associated with other aberrations were presented in 8 of 11 patients with a frequency of (72.7%) and represented (20.5%) of total 39 cases of ALL adults patients. The most common two atypical FISH signal patterns were amplifications and deletions. ABL amplifications were observed in three cases with a frequency of 7.7% in ALL patients while derivative chromosome 9q34 deletion was observed in three patients with a frequency of 7.7% in ALL patients the same as amplifications. Duplication was observed in one patient with a frequency of 2.56% in ALL patients and one patient show combination of amplification and deletion with a frequency of 2.56%.
Follow up was done at day 14 of chemotherapy. Out of the 39 newly diagnosed patients; 16 (41%) patients achieved complete remission while 23 (59%) patients showed incomplete remission.
In the present study there was high significant, negative relation between outcome and positive philidelphia chromosome t(9;22). Among 11 cases with ph1 +ve, nine patients had IR and only two patients had CR.
Statistical analysis of patients’outcome with prognostic markers revealed significant association (p<0.05) of CR with age <35 years and TLC <50x109/L. On the other hand, gender, hepatosplenomegaly, lymphadenopathy, CNS infilteration, Hb, platelet count and immunophenotyping showed non significant statistical difference between the patients who achieved complete remission and those with incomplete remission (p>0.05).
Analyzing the relationship of t(9;22) with various studied standard prognostic factors, revealed significant association (p<0.05) of ph1 +ve patients with age >35 years, hepatosplenomegaly, absence of lymphadenopathy, TLC ≥50X109/L and absolute-PB blasts ≥4.4X109/L, immunophenotyping and other aberrations. On the other hand, gender, CNS infilteration, Hb and platelet count showed non significant statistical difference (p>0.05).
Regarding the presence of other genetic aberrations associated ph1 in relation to different prognostic factors. The present study showed a significant positive association between their presence and age ≥35 years and also with absolute-PB blasts ≥4.4X109/L with (p<0.05). With no significance to other prognostic factors.
In conclusion, BCR/ABL fusion gene analysis by ES-FISH may serve as a powerful prognostic marker in adulthood ALL. The age, TLC and t(9;22) represent the significant standard prognostic factors in relation to patients’ outcome. Moreover, philidelphia chromosome with additional chromosomal abnormalities and gene amplification affecting BCR/ABL are efficiently detected by ES-FISH and show significant association with patients’ outcome that may be used as prognostic indicators for therapeutic response.