1. Cytogenetics available
COMPARATIVE ANALYSIS OF MOLECULAR GENETIC AND CONVENTIONAL CYTOGENETIC DETECTION OF DIAGNOSTICALLY IMPORTANT TRANSLOCATIONS
Biswajoy Pal, Dr. Deepak Mishra, Dr. Mayur Parihar,  Poonam Santra, Sourav Sarma Chowdhury, Dr. Saurabh Bhave, Dr. Anupam Chakrapani, Dr Arpita Bhattacharya, Dr Shekhar krishnan, Dr. Reena Nair, Dr Vaskar Saha, Dr. Mammen Chandy,
Dr. Neeraj Arora.
Department of Molecular Genetics & Cytogenetics, Tata Medical Center, Kolkata.
2014
INTRODUCTION
RESULTS
Fig-3: concordance and discordance results of karyotyping, FISH and RT-PCR
The contemporary diagnosis of leukaemia relies on a multifaceted approach.
Techniques like cytogenetics, RT-PCR and FISH are the commonly used methods in the diagnostic laboratory.
Though many translocations can be detected by cytogenetics alone, RT-PCR and FISH are increasingly used for detection of diagnostically significant translocations in a clinical laboratory.
The aim of the study was to test the usefulness of these three techniques for detection of common recurrent chromosomal translocations in leukemia so that they can be used in a cost effective manner.
486 specimens from 239 unique patients were tested by RT-PCR for seven common translocations: t(8;21), t(15;17), inv(16), t(9;22), t(12;21), t(4;11), and t(1;19), with a detection rate of 18.9% (92/486) in 2 years.
Of 92 positive cases, 30 having previous treatment history were excluded.
Of remaining 62, FISH analysis was performed on 44 (71%) and karyotyping in 30 (48.4%) cases only.
Discordance rate of RT-PCR/FISH and RT-PCR/karyotyping was 4.5% (2/44) and 6.6% (2/30). Both these cases were diagnosed as APML.
Total RTPCR positive
n=62
FISH available
n=44
Concordant
42 (95.4%)
Discordant
2 (4.5%)
Cytogenetics available
n=30
28 (93.3%)
2 (6.6%)
CONCLUSION
This analysis shows 8.3%(2/24) PML-RARA [(t15;17)] discordance between both karyotyping/RT-PCR and FISH/RT-PCR due to cryptic translocation in ALL cases which was not detected by karyotyping and/or FISH.
Both Conventional Cytogenetics and Molecular Genetics based assays are powerful diagnostic tools for detection of chromosomal aberrations in leukemias.
Although there is sporadic discordance between these two techniques and each method has its own limitations, the judicious use of these techniques will deliver the best cost effective treatment in leukaemias.
ENMZ(MALT)
Fig-1 :LIST OF RT-PCR POSITIVE CASES
AT DIAGNOSIS
MATERIALS AND METHODS
Total at diagnosis
RT-PCR positive
n=62
CML positive n = 17
ALL positive n = 21
BCR-ABL positive n= 12
E2A-PBX positive n = 4
MLL-AF4 positive n = 1
TEL-AML1 positive n = 4
APML positive n = 24
This is a retrospective study carried out in the Department of Molecular Genetics and Cytogenetics, Tata Medical Center, Kolkata.
All newly diagnosed cases of acute leukemia from August 2012 to August 2014 referred for RT-PCR analysis were included. Patients were diagnosed and sub classified according to the WHO 2008 classification.
All RT-PCR positive cases were compared to the karyotyping and/or FISH wherever available. Cytogenetic and FISH analyses were carried out using standard protocols.
All patients with leukemia at diagnosis who were positive for any of the fusion transcripts by RT-PCR, were included in this study.
Table-1 : concordance and discordance of karyotyping, FISH and RT-PCR in various tested molecular aberrations
Particulars
Diagnosis
CML
APML
Ph+ALL
t{12;21}
t{1;19}
Total cases 17 24 10 4
Concordance RT-PCR/ Karyotyping (%)
100
(17/17)
91.7
(22/24)
(6/6)
(4/4)
(2/2)
Discordance RT-PCR/ Karyotyping (%)
(0/17)
8.3
(2/24)
(0/6)
(0/4)
(0/2)
Concordance RT-PCR/FISH (%) NA
Discordance RT-PCR/FISH (%)
REFERENCES
• WHO classification of tumuors,2008.
• A comparative analysis of FISH, RT-PCR, and
cytogenetics for the diagnosis of bcr-abl-positive
leukemias. MC Cox et al, Am J Clin Pathol. 1998
Jan;109(1):24-31.