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Frequency of FANCA and FANCG G enes Mutation in Sudanese Fanconi Anemia Patients . By Ream Elzain Abdelgadir Supervisor: Imad Fadl-Elmula. Historical background. Guido Fanconi (1927) . Incidence (Worldwide). Uroepithelial cancer. Uroepithelial cancer. - PowerPoint PPT Presentation
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FREQUENCY OF FANCA AND FANCG GENES MUTATION IN SUDANESE
FANCONI ANEMIA PATIENTS
By Ream Elzain AbdelgadirSupervisor: Imad Fadl-Elmula
Historical background
Guido Fanconi (1927)
Uroepithelial cancerUroepithelial cancer
E54,000 new casesE12,500 deathsEThree times more frequently in €than in �
E7th most common cancer in�4th most common cancer in€
E In areas where Schistosoma haematobiumis endemicEThe most common cancer (25%) in€
ERarely occur before the age of 40 years
25% up to 30% of pediatric aplastic patients
Incidence (Worldwide)
Fanconi genes
Clinical Presentation
Chromosomal breakage test
Fibroblasts cultures Cell cycle
arrestDetermination of complementation
groups
Treatment
Supportive treatment
Bone marrow transplantation
Present StudyDate of mission 2011
To identify the prevalence of Fanconi anemia among patients in aplastic anemia Sudanese patients To characterize FA genetic profile in Sudanese To assess the influence of the WBCs in
chromosomal breakage test
Specific Objective
1- To detect the frequency of Fanconi anemia patients among aplastic anemia using chromosomal breakage test.
2- To determine the prevalence of FANCA gene among Sudanese patients with FA.
3- To identify the molecular genetics profile of FANCA gene mutation (exon 10, 11, 12 and31) in Sudanese patients with Fanconi anemia.
4- To detect FANCG 637-643 deletion mutation among Sudanese AA patients.
5- To determine the relationship between total white blood cell count and successful of cytogenetic cell culture among aplastic anemia patients.
6- To correlate between the TWBC count and amount of blood to obtain a successful cytogenetic cell culture among aplastic anemia patients.
working planningFrom 2011-2014
Year Tasks Year 12 11 10 9 8 7 6 5 4 3 2 1
Work Planning
2011
Sampling and practical analysis
2011
Sampling and practical analysis
2012
Practical analysis
2013
Statistical analysis
2013
Thesis correction
2013
Exam
2014
Material and methods
Sample collection69 child with aplastic
anemia
Gaffer Ibnaof Hospital Hematology unit
During 2011-2012
5 ml Venous blood
Clinical Presentation
Café au lait Short stature
Abnormal thumbs Microcephaly
Complete HomogramUsing Sysmex
Chromosomal breakage test
Culture of blood in RPM1 media after adding MM-C. Incubate 37C° for 72 hours.
Chromosomal breakage test
Harvesting steps: Adding Colcemid, hypotonic solution, Fixative.
Chromosomal breakage test
Slide preparation: Metaphase dropping, Ageing of cells, SSC treatment, staining.
Analysis of chromosomal breakage test
Analysis of 5-25 metaphase.
PCR Technique
Sequencing in Korea company
Salting out method
PCR mixture
DNA Amplification
Data AnalysisFrequency, percentage and correlation
carried out by SPSS 13 software program.
Demographic and hematological result
69study patients
Af:72%Nilo:16%Nig:12%
Age : 4-8 (66%)
GR:57%
Hb89.9%RBCs87%
TWBC88%Plt94%
E.BMF39%Ad.BMF50%NoBMF11%
M:65%F: 35%
Cons +ve 77%
Cytogenetic Results Prevalence of FA among AA patients
Negative 65%
Positive35%
Analysis of chromosomal breakage test
Breakage positive: Breakage, gaps, redial, ring.
Transformation to Leukemia4 patients transform to AML
AML M4
AML M5
AML M6
Female 5 (23.8%)
Male 16 ( 76.2%)
18 (85.7%)
18(85.7%)
Bone marrow Failure19
(90.5%)
Clinical presentation
Café au lait
Growth re
tardation
Short sta
ture
Microce
phaly
No abnormaliti
es
Kidneys abnorm
alities
Thumbs abnorm
alities
Eye abnormaliti
es
Mental retardation 0
102030405060708090
%
BMF in Fanconi anemiaChart Title
AdvanceBMFEarly BMFNo BMF
Molecular results #16/ 21 was +ve for FANCA gene Prevalence
of FANCA gene was 76.1%.
#A total of 47 mutations were detected of which 34 were intronic mutation whereas 13 were exonic mutation. Only one was reported.
Frame shift mutation 3 Nonsense mutation 9 Missense mutation 18 Silent mutation 17
Molecular results
Correlation between WBC count and the success of cell culture
A total of 60/69 (87%) showed an adequate number of metaphase for chromosomal breakage analysis. Of the 60 cases 8 (11.6%) cases showed excellent metaphase appearance, 25 (36.2%) showed good metaphase appearances, and 27 (39.2%) revealed bad metaphase appearances. The remaining 9 (13 %) showed failure in culture.
Correlation between WBC count and the success of cell culture
Direct correlation between :WBC count Metaphase NumberWBC count Metaphase qualityWBC count amount of needed blood
Correlation between WBC count and the success of cell culture
Conclusion and recommendation
*In conclusion , the prevalence of FA among Sudanese patients are higher than the other countries although the disease is consider as a rare syndrome. This make the importance of screening FA among aplastic anemia patients to use the appropriate treatment and prevent the appearance of the disease complication.
Conclusion and recommendation
* The equation developed by the present study improved the out come of chromosomal breakage test and thus should be used to calculate the needed amount of blood to perform successful chromosomal breakage test.
Acknowledgment Our acknowledge to the patients
and their family who agreed to participate in this study. Our thanks extend to the staff of department of Pediatrics, Hematology unit, Gaffer Ibnaof hospital for children and Elite for Health & Medical Services center for genetic study at Khartoum-Sudan.
Acknowledgment
ليت البيان مطاوعا لمرادى .... حتي أصوغ تحيه لعماد
عقل تفجر فى مرابع قطرنا .... أزرى عاله بشامخ االطواد
علم الوراثه كان رائد بحثه .... فاعجب بني بسابق الرواد
ليت القريض يمدنى ببيانه .... حتى أريك روائع االنشاد
فأنهض الى العلياء دون تردد .... ال تستجيب لمقاله الحساد
الله يحفظكم بحسن عناية ... يكفيك شر قوائل االحقاد
ريم الزين
ريم
