Blood Res 2013; 48(2):
Published online June 25, 2013
https://doi.org/10.5045/br.2013.48.2.128
© The Korean Society of Hematology
Department of Laboratory Medicine, University of Ulsan College of Medicine and Asan Medical Center, Seoul, Korea.
Correspondence to : Correspondence to Hyun-Sook Chi, M.D., Ph.D. Department of Laboratory Medicine, University of Ulsan College of Medicine and Asan Medical Center, 88 Olympic-ro 43-gil, Songpa-gu, Seoul 138-736, Korea. Tel: +82-2-3010-4502, Fax: +82-2-478-0884, hschi@amc.seoul.kr
This is an Open Access article distributed under the terms of the Creative Commons Attribution Non-Commercial License (http://creativecommons.org/licenses/by-nc/3.0/) which permits unrestricted non-commercial use, distribution, and reproduction in any medium, provided the original work is properly cited.
We aimed to evaluate the feasibility of using the allele burden of Janus kinase 2 (
We collected 70 peripheral blood (PB) and 81 bone marrow (BM) samples from patients diagnosed with Ph-MPN. Real-time quantitative PCR (RQ-PCR) and Amplification Refractory Mutation System (ARMS) assays were performed for each sample. We compared the allele burden of
The
The allele burden of
Keywords Allele, Discrimination, Janus Kinase 2, Mutation, Myeloproliferative disorders, Real-time polymerase chain reaction
Polycythemia vera (PV), essential thrombocythemia (ET), and primary myelofibrosis (PMF) are subtypes of Philadelphia chromosome-negative myeloproliferative neoplasm (Ph-MPN) [1]. The Janus kinase 2 (
According to the revised World Health Organization (WHO) classification of Ph-MPN, the 3 different disease subtypes show considerable overlap in terms of their clinical presentation, disease course, and associated bone marrow histopathology. Bone marrow histopathology is recommended for discriminating among the subtypes of Ph-MPN [5]. Although the
Recently, real-time quantitative PCR (RQ-PCR) methods that determine the allele burden of
In this study, we performed RQ-PCR to detect the allele burden of
We enrolled 151 patients diagnosed with Ph-MPN at the Asan Medical Center between January 2008 and July 2011. We obtained 70 peripheral blood (PB) and 81 bone marrow (BM) aspiration samples from these patients at initial diagnosis. The diagnosis of Ph-MPN was based on BM histopathology performed by an expert in this area. The numbers of patients diagnosed with PV, ET, PMF, and other subtypes of Ph-MPN were 41, 54, 24, and 32, respectively. Patients diagnosed as secondary Ph-MPN, such as post-ET or PV-PMF, were excluded from the study population to minimize the effect of a primary disorder on the secondary disorder.
Mutation analysis of
For quantitative analysis of the allele burden of the
The Kruskal-Wallis test was performed to evaluate whether the allele burden of
Of the 151 samples analyzed, 38 (25.2%) samples showed wild types by both RQ-PCR and ARMS. Analysis by using ARMS revealed that 97 (64.2%) were heterozygous and 2 (1.3%) were homozygous for the
Among the 151 patients, no gender-related pattern was observed with respect to disease subtypes (
The
For the diagnosis of Ph-MPN, the revised WHO classification emphasizes both BM histopathologic findings and presence of
Until recently, most of the studies that focused on the role of
Our data shows that the results of RQ-PCR correspond well with those of allele-specific PCR, with concordance rates of 90.7% and a kappa value of 0.780. The present study showed a low frequency of homozygous positivity in ARMS (only 1 case each of PV and PMF: 1.3%), which may have been caused by the ARMS interpretation method applied at the authors' institution. That is, because the presence of 2 bands was always considered as heterozygous positivity, irrespective of the signal strength; moreover, in this study, the frequency of heterozygous positivity may have been overestimated and that of homozygous positivity may have been underestimated.
Among the cases where the 2 tests showed discrepant results, 12 showed positive RQ-PCR results and negative ARMS results. The majority of these cases were 9 patients with other diseases and a relatively low allele burden of
Notably, our results also revealed that the allele burden of
In addition, our results showed that the hemogram results and allele burden of
In conclusion, the allele burden of
Comparison of the allele burden of the
Table 1 Comparison of the test results of quantitative real-time PCR with those of allele-specific PCR for the detection of the
Abbreviations: RQ-PCR, quantitative real-time polymerase chain reaction; ARMS, amplification refractory mutation system.
Table 2 Demographic and laboratory test results for the 151 patients diagnosed with Philadelphia-negative myeloproliferative neoplasm with respect to disease subtypes.
Abbreviations: PV, polycythemia vera; ET, essential thrombocythemia; PMF, primary myelofibrosis; WBC, white blood cell; ARMS, amplification refractory mutation system; RQ-PCR, real time quantitative polymerase chain reaction.
Table 3 Comparison of
Abbreviations: PV, polycythemia vera; ET, essential thrombocythemia; PMF, primary myelofibrosis.
Blood Res 2013; 48(2): 128-132
Published online June 25, 2013 https://doi.org/10.5045/br.2013.48.2.128
Copyright © The Korean Society of Hematology.
Sang Hyuk Park, Hyun-Sook Chi*, Young-Uk Cho, Seongsoo Jang, and Chan-Jeoung Park
Department of Laboratory Medicine, University of Ulsan College of Medicine and Asan Medical Center, Seoul, Korea.
Correspondence to: Correspondence to Hyun-Sook Chi, M.D., Ph.D. Department of Laboratory Medicine, University of Ulsan College of Medicine and Asan Medical Center, 88 Olympic-ro 43-gil, Songpa-gu, Seoul 138-736, Korea. Tel: +82-2-3010-4502, Fax: +82-2-478-0884, hschi@amc.seoul.kr
This is an Open Access article distributed under the terms of the Creative Commons Attribution Non-Commercial License (http://creativecommons.org/licenses/by-nc/3.0/) which permits unrestricted non-commercial use, distribution, and reproduction in any medium, provided the original work is properly cited.
We aimed to evaluate the feasibility of using the allele burden of Janus kinase 2 (
We collected 70 peripheral blood (PB) and 81 bone marrow (BM) samples from patients diagnosed with Ph-MPN. Real-time quantitative PCR (RQ-PCR) and Amplification Refractory Mutation System (ARMS) assays were performed for each sample. We compared the allele burden of
The
The allele burden of
Keywords: Allele, Discrimination, Janus Kinase 2, Mutation, Myeloproliferative disorders, Real-time polymerase chain reaction
Polycythemia vera (PV), essential thrombocythemia (ET), and primary myelofibrosis (PMF) are subtypes of Philadelphia chromosome-negative myeloproliferative neoplasm (Ph-MPN) [1]. The Janus kinase 2 (
According to the revised World Health Organization (WHO) classification of Ph-MPN, the 3 different disease subtypes show considerable overlap in terms of their clinical presentation, disease course, and associated bone marrow histopathology. Bone marrow histopathology is recommended for discriminating among the subtypes of Ph-MPN [5]. Although the
Recently, real-time quantitative PCR (RQ-PCR) methods that determine the allele burden of
In this study, we performed RQ-PCR to detect the allele burden of
We enrolled 151 patients diagnosed with Ph-MPN at the Asan Medical Center between January 2008 and July 2011. We obtained 70 peripheral blood (PB) and 81 bone marrow (BM) aspiration samples from these patients at initial diagnosis. The diagnosis of Ph-MPN was based on BM histopathology performed by an expert in this area. The numbers of patients diagnosed with PV, ET, PMF, and other subtypes of Ph-MPN were 41, 54, 24, and 32, respectively. Patients diagnosed as secondary Ph-MPN, such as post-ET or PV-PMF, were excluded from the study population to minimize the effect of a primary disorder on the secondary disorder.
Mutation analysis of
For quantitative analysis of the allele burden of the
The Kruskal-Wallis test was performed to evaluate whether the allele burden of
Of the 151 samples analyzed, 38 (25.2%) samples showed wild types by both RQ-PCR and ARMS. Analysis by using ARMS revealed that 97 (64.2%) were heterozygous and 2 (1.3%) were homozygous for the
Among the 151 patients, no gender-related pattern was observed with respect to disease subtypes (
The
For the diagnosis of Ph-MPN, the revised WHO classification emphasizes both BM histopathologic findings and presence of
Until recently, most of the studies that focused on the role of
Our data shows that the results of RQ-PCR correspond well with those of allele-specific PCR, with concordance rates of 90.7% and a kappa value of 0.780. The present study showed a low frequency of homozygous positivity in ARMS (only 1 case each of PV and PMF: 1.3%), which may have been caused by the ARMS interpretation method applied at the authors' institution. That is, because the presence of 2 bands was always considered as heterozygous positivity, irrespective of the signal strength; moreover, in this study, the frequency of heterozygous positivity may have been overestimated and that of homozygous positivity may have been underestimated.
Among the cases where the 2 tests showed discrepant results, 12 showed positive RQ-PCR results and negative ARMS results. The majority of these cases were 9 patients with other diseases and a relatively low allele burden of
Notably, our results also revealed that the allele burden of
In addition, our results showed that the hemogram results and allele burden of
In conclusion, the allele burden of
Comparison of the allele burden of the
Table 1 . Comparison of the test results of quantitative real-time PCR with those of allele-specific PCR for the detection of the
Abbreviations: RQ-PCR, quantitative real-time polymerase chain reaction; ARMS, amplification refractory mutation system..
Table 2 . Demographic and laboratory test results for the 151 patients diagnosed with Philadelphia-negative myeloproliferative neoplasm with respect to disease subtypes..
Abbreviations: PV, polycythemia vera; ET, essential thrombocythemia; PMF, primary myelofibrosis; WBC, white blood cell; ARMS, amplification refractory mutation system; RQ-PCR, real time quantitative polymerase chain reaction..
Table 3 . Comparison of
Abbreviations: PV, polycythemia vera; ET, essential thrombocythemia; PMF, primary myelofibrosis..
Yoon Hwan Chang
Blood Res 2021; 56(S1): S51-S64Hee Suk Choung, Hee Jin Kim, Chul Won Jung, Sun Hee Kim
Korean J Hematol 2008; 43(2): 118-121Kyung Soon Song, Hyun Kyung Kim, Young Sook Park
Korean J Hematol 2003; 38(4): 274-278
Comparison of the allele burden of the