scholarly journals High Expression of TWIST-1 in the Chronic Myeloid Leukemia Patients with T315I Mutation

2021 ◽  
Vol 12 (3) ◽  
Author(s):  
Nazanin Heidari ◽  
Fatemeh Noroozi ◽  
Najmaldin Saki
Keyword(s):  
Gene Expression ◽  
Chronic Myeloid Leukemia ◽  
Myeloid Leukemia ◽  
Amplification Refractory Mutation System ◽  
Chain Reaction ◽  
Mutation System ◽  
T315i Mutation ◽  
Polymerase Chain ◽  
Twist 1 ◽  
Age Range

Background: Among the known ABL mutations in chronic myeloid leukemia (CML), T315I is of particular importance. The T315I mutation may develop resistant cells that increase disease progression. TWIST-1 expression is impaired in patients with increased drug resistance. Objectives: The current study aimed to measure the expression of TWIST-1 gene in CML patients to investigate its association with T315I mutation. Methods: Peripheral blood samples were taken from 40 CML patients. The expression of TWIST-1 and BCR-ABL1 genes was quantified by real-time polymerase chain reaction (PCR). The gene expression was evaluated by REST software. cDNA was used for amplification refractory mutation system (ARMS)-PCR reaction. Results: Of the 40 patients (age range: 19 - 72 years) participating in the study, 23 (57.7%) were female, and 17 (42.5%) were male. The expression of TWIST-1 gene was 43 ± 184.09-fold. The T315I mutation was detected in 3 (7.5%) patients. Conclusions: According to our results, the TWIST-1 gene expression in patients with T315I mutation was significantly higher than patients without that mutation.

scholarly journals Amplification Refractory Mutation System – Polymerase Chain Reaction for Rapid Detection of rpoB Gene Mutations in Mycobacterium tuberculosis

2017 ◽  
Vol 5 (1) ◽  
pp. 81-85 ◽  
Author(s):  
Hemanta Kumari Chaudhary ◽  
Mitesh Shrestha ◽  
Prakash Chaudhary ◽  
Bal Hari Poudel
Keyword(s):  
Mycobacterium Tuberculosis ◽  
Rpob Gene ◽  
Amplification Refractory Mutation System ◽  
Chain Reaction ◽  
Arms Pcr ◽  
Mutation System ◽  
Mdr Tb ◽  
Total Dna ◽  
Polymerase Chain ◽  
Rifampin Resistance

Multidrug-resistant tuberculosis (MDR-TB) has become a serious worldwide threat including in Nepal. MDR-TB refers to the pathological condition whereby Mycobacterium tuberculosis becomes resistant to the first line of drug treatment i.e. rifampin and isoniazid. Resistance to rifampin (RIF) is mainly caused by the mutations in the rpoB gene which codes for the β-subunit of RNA polymerase. In this study, Amplification Refractory Mutation System – Polymerase Chain Reaction (ARMS – PCR) technique has been used to detect mutations in the rpoB gene of Mycobacterium tuberculosis. Total DNA samples of 34 phenotypic MDR-TB were subjected to ARMS – PCR using three different codon specific primers (516, 526 and 531). These three codons occupy large portion of total mutation responsible for rifampin resistance. Out of the total DNA samples, all were bearing mutation in at least one of the three codons mentioned. Of those bearing mutation, the highest number had mutation in codon 531 (97.05 %) followed by codon 516 (17.64 %) and finally in codon 526 (11.76%) respectively. Hence, ARMS – PCR may be used as an alternative diagnostic technique for detection of rifampin resistance in Mycobacterium tuberculosis strains, especially for a developing country like Nepal.Int. J. Appl. Sci. Biotechnol. Vol 5(1): 81-85

scholarly journals Detection of the molecular abnormality in chronic myeloid leukemia by use of the polymerase chain reaction

Blood ◽  
1988 ◽  
Vol 72 (6) ◽  
pp. 2063-2065
Author(s):  
A Dobrovic ◽  
KJ Trainor ◽  
AA Morley
Keyword(s):  
Polymerase Chain Reaction ◽  
Chronic Myeloid Leukemia ◽  
Blood Cells ◽  
Myeloid Leukemia ◽  
Lymphoblastic Leukemia ◽  
Leukemic Cell ◽  
Great Sensitivity ◽  
Chain Reaction ◽  
Molecular Abnormality ◽  
Polymerase Chain

The bcr-abl translocation characteristic of chronic myeloid leukemia (CML) was detected by the polymerase chain reaction (PCR) modified to use mRNA as the starting material. Amplification of a sequence spanning the bcr-abl junction was obtained by using peripheral blood cells from all of 20 patients with classic CML, one patient with acute lymphoblastic leukemia probably secondary to CML, and two cell lines derived from patients with CML. The presence of bcr exon 3 in the mRNA was determined from the size of the amplified sequence; it was present in 14 and absent in seven patients. One leukemic cell per 1,000 nonleukemic cells could be readily detected, thus indicating the great sensitivity of the method. This technique is of routine value in CML both for diagnosis and for following the course of treatment.

Determination of a T/G polymorphism at nucleotide 3206 of the apolipoprotein C III gene by amplification refractory mutation system

1994 ◽  
Vol 40 (12) ◽  
pp. 2235-2239 ◽  
Author(s):  
M Y Tsai ◽  
N Q Hanson ◽  
K R Copeland ◽  
I Beheshti ◽  
U Garg
Keyword(s):  
Epidemiological Studies ◽  
Amplification Refractory Mutation System ◽  
Chain Reaction ◽  
Apolipoprotein C ◽  
Significant Difference ◽  
Mutation System ◽  
Polymerase Chain ◽  
Exon 4 ◽  
And Control

Abstract We used the amplification refractory mutation system (ARMS)--a polymerase-chain-reaction-based method--to determine the 3206 T-to-G polymorphism on exon 4 of the apolipoprotein (apo) C III gene. Apo C III is an inhibitor of the enzyme lipoprotein lipase (EC 3.1.1.34). Previous studies have demonstrated that a polymorphism at nucleotide 3175 on exon 4 of this gene is associated with hypertriglyceridemia. We studied 45 hypertriglyceridemic and 46 age-matched controls for the 3206 T-to-G polymorphism. The results showed a significant difference in the distribution of the genotypes with respect to this allele between the hypertriglyceridemic and control individuals. We also determined the presence of the SacI site at nucleotide 3175 in these same individuals and found no significant difference in SacI genotypes between the two groups. This study reaffirms the usefulness of ARMS as a simple, reliable method for detecting mutations and polymorphisms in clinical and epidemiological studies.

scholarly journals Detection of residual leukemia after bone marrow transplant for chronic myeloid leukemia: role of polymerase chain reaction in predicting relapse

Blood ◽  
1991 ◽  
Vol 77 (4) ◽  
pp. 874-878 ◽  
Author(s):  
TP Hughes ◽  
GJ Morgan ◽  
P Martiat ◽  
JM Goldman
Keyword(s):  
Bone Marrow ◽  
Polymerase Chain Reaction ◽  
Chronic Myeloid Leukemia ◽  
Bone Marrow Transplant ◽  
Blood Cells ◽  
Myeloid Leukemia ◽  
Marrow Transplant ◽  
Early Assessment ◽  
Chain Reaction ◽  
Polymerase Chain

Abstract We used the polymerase chain reaction (PCR) to detect residual leukemia- specific mRNA in blood and marrow from 37 patients in complete hematologic and cytogenetic remission after allogeneic bone marrow transplant (BMT) for chronic myeloid leukemia (CML). Our two-step PCR method involved the use of “nested primers” in the second step and could detect one K562 cell diluted into 10(5) normal cells. Elaborate measures were taken to exclude false-positive and false-negative results. In nine patients whose blood and marrow were studied simultaneously the results were concordant (two positive and seven negative). Twenty-three patients transplanted in chronic phase (CP) with unmanipulated donor marrow were studied. Blood cells from nine of these patients were studied 3 to 6 months post-BMT and six were PCR positive; three were negative on subsequent studies. Blood cells from 18 patients studied between 8 months and 8 years post-BMT were all PCR negative. Nine patients transplanted in CP with T-cell-depleted marrow cells were studied. Blood from five was positive 3 to 24 months post- BMT; blood from five was negative 3 to 6 years post-BMT. Four patients no longer in first CP were studied after BMT with unmanipulated donor marrow. Blood from all four was positive 5 to 19 months post-BMT. Based on the known clinical results of transplant in these three cohorts we conclude that PCR may be positive within 6 months of BMT in patients who can expect long-lasting remission, whereas PCR positivity later after BMT may indicate that the probability of cure is reduced. Thus, the technique may prove useful for early assessment of new transplant protocols that might inadvertently increase the risk of relapse.

scholarly journals A new method of "in-cell reverse transcriptase-polymerase chain reaction" for the detection of BCR/ABL transcript in chronic myeloid leukemia patients

Blood ◽  
1996 ◽  
Vol 87 (9) ◽  
pp. 3822-3827 ◽  
Author(s):  
N Testoni ◽  
G Martinelli ◽  
P Farabegoli ◽  
A Zaccaria ◽  
M Amabile ◽  
...  
Keyword(s):  
Polymerase Chain Reaction ◽  
Chronic Myeloid Leukemia ◽  
Messenger Rna ◽  
Myeloid Leukemia ◽  
Residual Disease ◽  
Single Cells ◽  
Fluorescent Microscopy ◽  
Rt Pcr ◽  
Chain Reaction ◽  
Polymerase Chain

Methods of detecting minimal residual disease (MRD) in chronic myeloid leukemia (CML) include chromosome analysis, Southern blotting, polymerase chain reaction (PCR) and fluorescent in situ hybridization (FISH) techniques. We report a novel method to detect intracellular messenger RNA (mRNA) by combining the techniques of reverse transcription (RT) and PCR performed directly inside the cells, without extraction of the nucleic acid. We applied this method, which we call “in-cell RT-PCR”, to detect hybrid BCR/ABL transcript within single cells. After cellular permeabilization and fixation of single cells in suspension, the neoplastic mRNA was reverse transcribed into cDNA, and the cDNA was amplified by PCR with fluorescent primers, specific for bcr/abl. Flow cytometry was used to detect cells positive for the amplified DNA within the cell cytoplasm. After transferring the amplified cells onto slides by cytospin, the positive cells for BCR/ABL cDNA were observed by fluorescent microscopy. The technique was capable of detecting low abundancy signals and distinguishing different levels of gene expression. The amplification products were found in the cells and supernatants. The distribution was critically affected by the protease digestion condition. The specificity of amplification was confirmed by a nested RT-PCR of BCR/ABL performed on extracted mRNA from the same sample, and by reamplification of supernatants. We have used the technique to study 10 Ph+ CML patients and three normal subjects as controls. Four patients were 100% Ph+ at diagnosis time and RT-PCR+ at cytogenetic and molecular analysis, respectively. In-cell RT- PCR showed that the residual non-neoplastic cells could be observed in all cases. In two patients undergoing interferon-alpha (IFN-alpha) therapy and in four bone-marrow transplanted patients, the in-cell RT- PCR was used to compare the level of Ph+ positivity detected by cytogenetic analysis with the number of cells expressing BCR/ABL transcript. In this manner, we could estimate the MRD. Our preliminary application of the technique suggests that it is capable of accurately identifying cells transcribing bcr/abl, and that it may have significant clinical applications in the detection of MRD.

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