Real-time quantification of fusion transcripts with ligase chain reaction by direct ligation of adjacent DNA probes at fusion junction

The Analyst ◽  
2020 ◽  
Vol 145 (11) ◽  
pp. 3977-3982
Author(s):  
Fengxia Su ◽  
Jianing Ji ◽  
Pengbo Zhang ◽  
Fangfang Wang ◽  
Zhengping Li
Keyword(s):  
Real Time ◽  
Fusion Transcript ◽  
Dna Probes ◽  
Chain Reaction ◽  
Ligase Chain Reaction ◽  
Fusion Transcripts ◽  
Fusion Junction

A fusion transcript assay is developed based on direct ligation and ligase chain reaction.

Real-Time Detection of PML-RARA and RARA-PML Fusion Transcripts in High Risk Acute Promyelocytic Leukemia (APL) Treated with Arsenic Trioxide (ATO): Implications for Realization of Pre-Emptive Therapy for Molecular Relapse.

Blood ◽  
2006 ◽  
Vol 108 (11) ◽  
pp. 493-493 ◽  
Author(s):  
David Grimwade ◽  
J.V. Jovanovic ◽  
D. Diverio ◽  
E. Nugent ◽  
A.Y. Ho ◽  
...  
Keyword(s):  
Real Time ◽  
Residual Disease ◽  
Survival Rates ◽  
Fusion Transcript ◽  
Additional Treatment ◽  
Molecular Response ◽  
Sample Type ◽  
Assay Sensitivity ◽  
Fusion Transcripts ◽  
Kinetics Of

Abstract A key challenge in the management of APL is to improve upon survival rates now achieved with ATRA and anthracycline-based chemotherapy. One approach involves minimal residual disease (MRD) monitoring to identify patients otherwise destined to relapse, whose outcome may be improved by pre-emptive therapy. There is also scope for improvement through reduction in induction deaths and treatment-related mortality, with recent studies suggesting that molecularly targeted therapy involving ATO can achieve long term remissions with limited toxicity. However, such strategies are critically dependent upon optimized schedules for MRD detection to rapidly identify those patients needing additional treatment. Yet, there are limited data concerning assay sensitivity, optimal sample type or kinetics of relapse, that are fundamental to delivery of MRD-directed therapy. We determined expression levels of PML-RARA and reciprocal RARA-PML transcripts in diagnostic BM samples from 102 patients using real-time quantitative PCR (RQ-PCR). Normalized PML-RARA transcript levels varied by 3-logs, equating with marked inter-patient variability in maximal achievable sensitivities for MRD detection, ranging between 1 in 102 and 1 in 105 (med. 1 in 103.8). RARA-PML fusion transcripts were expressed in 68/102 (67%). Parallel detection of both transcripts increased sensitivity for MRD detection in 35/102 (34%); yet, the detection limit was still <1 in 104 in 49% due to the low level of fusion transcript expression. Serial MRD monitoring was undertaken in patients (n=32) treated with ATO for relapsed APL in UK and Eire since Nov 02: 22 in hematologic relapse (HR) and 10 treated pre-emptively in 1st molecular relapse (MR). RQ-PCR assessment prior to MR/HR showed a median increase in PML-RARA transcripts of 0.8 log/month (range 0.2–1.9). Analysis of paired BM and blood (PB) samples revealed that molecular conversion in BM invariably preceded that in PB; including a patient testing negative in PB despite a significant level of disease in BM (6% by FISH). Parallel use of the RARA-PML assay in evaluable patients increased rates of MRD detection, in some instances providing earlier warning of impending relapse. ATO was given using the conventional schedule (0.15mg/kg/d - Regimen A) or a regimen involving loading followed by intermittent dosing (0.3mg/kg/d x 5d, then 0.25mg/kg 2x/wk - Regimen B). No difference in kinetics of molecular response was observed; however amongst patients treated in HR, hyperleucocytosis was less common with Regimen B (peak WBC 5 vs 126 x 109/l, p=0.007), indicating that this schedule may favor apoptosis over differentiation and merits evaluation in newly diagnosed APL. OS from time of relapse was 60% at 3yrs; of 10 patients treated pre-emptively, 7 are in 2nd molecular CR at median of 23mo (7–40mo), 2 died in relapse, treatment is on-going in 1. This study highlights sample type, assay sensitivity (which demonstrates inter-patient and -sample variability) and kinetics of relapse as critical factors that need to be taken into account for the successful delivery of MRD-directed therapy, which is becoming increasingly pertinent to the management of other subsets of acute leukemia.

Sensitive quantification of messenger RNA with a real-time ligase chain reaction by using a ribonucleotide-modified DNA probe

2014 ◽  
Vol 50 (86) ◽  
pp. 13093-13095 ◽  
Author(s):  
Yanlei Hu ◽  
Hongxia Jia ◽  
Yucong Wang ◽  
Yongqiang Cheng ◽  
Zhengping Li
Keyword(s):  
Real Time ◽  
Messenger Rna ◽  
Dna Probe ◽  
Chain Reaction ◽  
Ligase Chain Reaction ◽  
Modified Dna

scholarly journals Higher Incidence of Co-Expression of BCR-ABL Fusion Transcripts in an Eastern Indian Population

2021 ◽  
Author(s):  
Ajeet Kumar ◽  
Vatsal Mishra ◽  
Chandra Bhan Singh ◽  
Rashmi Patel ◽  
Siddharth Samrat ◽  
...  
Keyword(s):  
Indian Population ◽  
Fusion Transcript ◽  
Break Point ◽  
Cdna Synthesis ◽  
Chain Reaction ◽  
Hematopoietic Stem ◽  
Fusion Transcripts ◽  
Ph Chromosome ◽  
Polymerase Chain ◽  
Cell Disorder

Abstract Background Chronic myeloid leukaemia (CML) is a hematopoietic stem cell disorder, caused by a balanced reciprocal translocation (t(9;22) (q34;q11))that lead to the formation of BCR (Break point Cluster Region)-ABL (Abelson) fusion transcripts known as Philadelphia (Ph) chromosome. Prevalence of BCR-ABL fusion transcripts in Indian CML population is poorly understood and few studies have been reported from India. The aim of present study was to determine the frequencies as well as prognostic effects of the three fusion transcripts i.e. b2a2, b3a2 and e1a2 in an Indian population. Methods RNA was isolated from total 123 sample 27 bone marrow (BM) sample and 96 Peripheral blood (PB) sample of CML patient followed by cDNA synthesis. Real-time quantitative reverse-transcription polymerase chain reaction (qRT-PCR) was performed using TaqMan→ assay (ABI, CA, USA) to monitor BCR-ABL transcript. Results Ph' chromosome was observed in 103 patients whereas it was not detected in 20 cases. qRT-PCR revealed that the b3a2 fusion transcripts was the most common transcript in CML patients (63.41%) while b2a2 fusion transcript was present in 16.26% cases. Co-expression of b3a2+b2a2 fusion transcript was observed in 0.81% cases whereas co-expression of b3a2+e1a2 fusion transcript was found in 1.63% cases. There was no co-relation observed between b3a2 fusion transcript and platelet count. The fusion transcript b2a2 was observed in relatively younger patients compared to b3a2 fusion transcript. Although this correlation was not statistically significant. Conclusion The co-expression of BCR-ABL fusion transcripts was higher (63.41% aggregate of b3a2) in the present population in contrast to other populations reported. This finding was consistent with the frequency data reported from Sudan.

Real-Time Gap Ligase Chain Reaction

2004 ◽  
Vol 10 (7) ◽  
pp. 2379-2385 ◽  
Author(s):  
Susan V. Harden ◽  
David C. Thomas ◽  
Nicole Benoit ◽  
Khalid Minhas ◽  
William H. Westra ◽  
...  
Keyword(s):  
Real Time ◽  
Chain Reaction ◽  
Ligase Chain Reaction ◽  
Time Gap
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