| 1. |
- Ayoola-Gustafsson, Kristin, et al.
(författare)
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Enrichment of pathogenic ASXL1 variants among patients with primary refractory chronic myeloid leukemia
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Annan publikation (övrigt vetenskapligt/konstnärligt)abstract
- In chronic myeloid leukemia in chronic phase (CP-CML), tyrosine kinase inhibitor (TKI) therapy is standard, yielding excellent long-term results. However, 5-10% of patients fail multiple TKIs, often resorting to allogeneic stem cell transplantation (Allo-SCT) with considerable risks. BCR::ABL1 kinase domain (KD) mutations and additional cytogenetic aberrations (ACA) represent common resistance mechanisms, but often the cause of resistance remains unknown. This study sequenced 54 myeloid neoplasm-related genes in 20 CP-CML patients refractory to TKI and without KD mutations or ACA, revealing pathogenic variants in 50% (n=10). ASXL1 mutations were most common (30%), followed by DNMT3A, IKZF1, GATA2, TP53 and NRAS. No pathogenic variant could be detected in a control cohort of 10 patients who were considered as optimal responders.
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| 2. |
- Gupta, Manu, et al.
(författare)
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Expression of BCR-ABL1 oncogene relative to ABL1 gene changes overtime in chronic myeloid leukemia
- 2008
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Ingår i: Biochemical and Biophysical Research Communications - BBRC. - : Elsevier BV. - 0006-291X .- 1090-2104. ; 366:3, s. 848-851
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Tidskriftsartikel (populärvet., debatt m.m.)abstract
- Using a quantitative single nucleotide polymorphism (SNP) assay we have investigated the changes in the expression of the BCR-ABL1 oncogene relative to the wild-type ABL1 and BCR alleles in cells from chronic myeloid leukemia (CML) patients not responding to therapy. The results show a progressive increase in the BCR-ABL1 oncogene expression at the expense of decreased expression of the ABL1 allele, not involved in the fusion. No relative changes in the expression of the two BCR alleles were found. These results demonstrate that allele-specific charities in gene expression, with selective, progressive silencing of the wild-type A BL1 allele in favor of the oncogenic BCR-ABL1 allele occur in CML patients with therapy-resistant disease.
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| 3. |
- Lampa, Erik, et al.
(författare)
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An investigation of the co-variation in circulating levels of a large number of environmental contaminants
- 2012
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Ingår i: Journal of Exposure Science and Environmental Epidemiology. - : Nature Publishing Group. - 1559-0631 .- 1559-064X. ; 22:5, s. 476-482
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Tidskriftsartikel (refereegranskat)abstract
- We are daily exposed to many different environmental contaminants. Mixtures of these contaminants could act together to induce more pronounced effects than the sum of the individual contaminants. To evaluate the effects of such mixtures, it is of importance to assess the co-variance amongst the contaminants. Thirty-seven environmental contaminants representing different classes were measured in blood samples from 1016 individuals aged 70 years. Hierarchical cluster analysis and principal component analysis were used to assess the co-variation among the contaminants. Within each identified cluster, possible marker contaminants were sought for. We validated our findings using data from the National Health and Nutrition Examination Survey (NHANES) 2003--2004 study. Two large clusters could be identified, one representing low/medium chlorinated polychlorinated biphenyls (PCBs) (<= 6 chlorine atoms), as well as two pesticides and one representing medium/high chlorinated PCBs (>= 6 chlorine atoms). PCBs 118 and 153 could be used as markers for the low/medium chlorinated cluster and PCBs 170 and 209 could be used as markers for the medium/high chlorinated cluster. This pattern was similar to data from the NHANES study. Apart from the PCBs, little co-variation was seen among the contaminants. Thus, a large number of chemicals have to be measured to adequately identify mixtures of environmental contaminants.
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| 4. |
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| 5. |
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| 6. |
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| 7. |
- Lampa, Erik, et al.
(författare)
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The identification of complex interactions in epidemiology and toxicology : a simulation study of Boosted Regression Trees
- 2014
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Ingår i: Environmental Health. - 1476-069X. ; 13, s. 57-
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Tidskriftsartikel (refereegranskat)abstract
- Background: There is a need to evaluate complex interaction effects on human health, such as those induced by mixtures of environmental contaminants. The usual approach is to formulate an additive statistical model and check for departures using product terms between the variables of interest. In this paper, we present an approach to search for interaction effects among several variables using boosted regression trees. Methods: We simulate a continuous outcome from real data on 27 environmental contaminants, some of which are correlated, and test the method's ability to uncover the simulated interactions. The simulated outcome contains one four-way interaction, one non-linear effect and one interaction between a continuous variable and a binary variable. Four scenarios reflecting different strengths of association are simulated. We illustrate the method using real data. Results: The method succeeded in identifying the true interactions in all scenarios except where the association was weakest. Some spurious interactions were also found, however. The method was also capable to identify interactions in the real data set. Conclusions: We conclude that boosted regression trees can be used to uncover complex interaction effects in epidemiological studies.
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| 8. |
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| 9. |
- Löf, Liza, et al.
(författare)
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Flow Cytometric Measurement of Blood Cells with BCR-ABL1 Fusion Protein in Chronic Myeloid Leukemia
- 2017
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Ingår i: Scientific Reports. - : Nature Publishing Group. - 2045-2322. ; 7:1
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Tidskriftsartikel (refereegranskat)abstract
- Chronic myeloid leukemia (CML) is characterized in the majority of cases by a t(9;22)(q34;q11) translocation, also called the Philadelphia chromosome, giving rise to the BCR-ABL1 fusion protein. Current treatment with tyrosine kinase inhibitors is directed against the constitutively active ABL1 domain of the fusion protein, and minimal residual disease (MRD) after therapy is monitored by real-time quantitative PCR (RQ-PCR) of the fusion transcript. Here, we describe a novel approach to detect and enumerate cells positive for the BCR-ABL1 fusion protein by combining the in situ proximity ligation assay with flow cytometry as readout (PLA-flow). By targeting of the BCR and ABL1 parts of the fusion protein with one antibody each, and creating strong fluorescent signals through rolling circle amplification, PLA-flow allowed sensitive detection of cells positive for the BCR-ABL1 fusion at frequencies as low as one in 10,000. Importantly, the flow cytometric results correlated strongly to those of RQ-PCR, both in diagnostic testing and for MRD measurements over time. In summary, we believe this flow cytometry-based method can serve as an attractive approach for routine measurement of cells harboring BCR-ABL1 fusions, also allowing simultaneously assessment of other cell surface markers as well as sensitive longitudinal follow-up.
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| 10. |
- Olsson-Strömberg, Ulla, et al.
(författare)
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Molecular monitoring and mutation analysis of patients with advanced phase CML and Ph plus ALL receiving dasatinib
- 2010
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Ingår i: European Journal of Haematology. - : Wiley. - 0902-4441 .- 1600-0609. ; 85:5, s. 399-404
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Tidskriftsartikel (refereegranskat)abstract
- As a result of the excellent responses achieved in chronic phase chronic myeloid leukemia since the introduction of imatinib, sensitive techniques such as reverse transcriptase real-time PCR are warranted to monitor patients receiving tyrosine kinase inhibitors (TKI). Our objective was to determine the value of molecular monitoring Ph-positive leukemias under dasatinib treatment. We used real-time PCR and ABL1 kinase domain sequencing on sequential samples from 11 patients with Philadelphia-positive leukemias who received dasatinib. We were able to detect pre-existing mutations in the kinase domain of BCR-ABL1 in four patients, particularly in patients with high BCR-ABL1 transcript levels. Most mutations disappeared with dasatinib, however, in five patients a clone with T315I appeared during dasatinib treatment. We conclude that sensitive molecular monitoring with real-time PCR for BCR-ABL1 transcripts and mutation screening of the ABL1 kinase domain of patients with Philadelphia-positive leukemias are valuable for patient management, however, mutation findings should be interpreted with caution, as mutant clones not always behave in vivo as predicted by in vitro assays.
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| 11. |
- Pandzic, Tatjana, et al.
(författare)
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Five Percent Variant Allele Frequency Is a Reliable Reporting Threshold for TP53 Variants Detected by Next Generation Sequencing in Chronic Lymphocytic Leukemia in the Clinical Setting
- 2022
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Ingår i: HemaSphere. - : Lippincott Williams & Wilkins. - 2572-9241. ; 6:8
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Tidskriftsartikel (refereegranskat)abstract
- The clinical significance of small TP53 clones detected with next generation sequencing (NGS) in chronic lymphocytic leukemia is an issue of active debate. According to the official guidelines, treatment decisions should be guided only by variants with variant allele frequency (VAF) >= 10%. We present data on 325 consecutive patients with chronic lymphocytic leukemia analyzed with NGS. In total 47 pathogenic/likely pathogenic (P/LP), TP53 variants were detected in 26 patients (8%). Eleven of these (23%) were in the 5% to 10% VAF range and reported according to our institutional policy. All TP53 variants in the 5% to 10% VAF range were confirmed (100% concordance) with a second NGS panel. Our results where further validated with the performance of Sanger sequencing and digital droplet PCR (ddPCR). In 12 patients with available fluorescence in situ hybridization data and TP53 mutations within 5% to 10% VAF, deletion of chromosome 17p (del(17p)) was detectable in only 1 patient. We propose a robust diagnostic algorithm, which allows the safe detection and reporting of TP53 variants with VAF down to 5% in the clinical setting. Our study provides evidence that NGS is equally potent to detect variants with VAF 5% to 10% compared to those with VAF 10% to 15%, highlighting the urgent need for harmonization of NGS methodologies across diagnostic laboratories.
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| 12. |
- Paul, Esbjörn, et al.
(författare)
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Low p14ARF expression in de novo acute myeloid leukemia with normal karyotype is associated with poor survival
- 2009
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Ingår i: Leukemia and Lymphoma. - : Informa UK Limited. - 1042-8194 .- 1029-2403. ; 50:9, s. 1512-1518
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Tidskriftsartikel (refereegranskat)abstract
- The p14ARF protein activates the p53 tumor suppressor by binding to and inhibiting its negative regulator HDM-2. We have studied the prognostic impact of p14ARF in acute myeloid leukemia (AML). Leukemic cells from 57 adult patients with normal karyotype de novo AML were analyzed for p14ARF mRNA expression level using real-time polymerase chain reaction (RT-PCR). We also tested the effect of conventional anti-leukemic drugs and the mutant p53-targeting small molecule PRIMA-1 in vitro. Patients whose cells expressed more p14ARF mRNA than the 75th percentile (0.26) had significantly better survival compared with those expressing lower levels, 61 vs. 30% 3-year survival (p = 0.046). The difference remained significant also when NPM1/FLT3 status was considered. The mean effects of all the tested conventional anti-leukemic drugs were greater in leukemic cell samples expressing p14ARF mRNA >or= 0.26, but the differences were not statistically significant. In contrast, PRIMA-1 had a significantly greater effect on leukemic cell samples with low levels of p14ARF mRNA. We conclude that low levels of p14ARF mRNA in leukemic cells from patients with normal karyotype AML is associated with poor prognosis. Treatment with drugs targeting p53 may be a future possibility to improve outcome for these patients.
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| 13. |
- Pfeifer, H., et al.
(författare)
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Standardisation and consensus guidelines for minimal residual disease assessment in Philadelphia-positive acute lymphoblastic leukemia (Ph plus ALL) by real-time quantitative reverse transcriptase PCR of e1a2 BCR-ABL1
- 2019
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Ingår i: Leukemia. - : NATURE PUBLISHING GROUP. - 0887-6924 .- 1476-5551. ; 33:8, s. 1910-1922
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Tidskriftsartikel (refereegranskat)abstract
- Minimal residual disease (MRD) is a powerful prognostic factor in acute lymphoblastic leukemia (ALL) and is used for patient stratification and treatment decisions, but its precise role in Philadelphia chromosome positive ALL is less clear. This uncertainty results largely from methodological differences relating to the use of real-time quantitative PCR (qRT-PCR) to measure BCR-ABL1 transcript levels for MRD analysis. We here describe the first results by the EURO-MRD consortium on standardization of qRT-PCR for the e1a2 BCR-ABL1 transcript in Ph + ALL, designed to overcome the lack of standardisation of laboratory procedures and data interpretation. Standardised use of EAC primer/probe sets and of centrally prepared plasmid standards had the greatest impact on reducing interlaboratory variability. In QC1 the proportion of analyses with BCR-ABL1/ABL1 ratios within half a log difference were 40/67 (60%) and 52/67 (78%) at 10(-3) and 36/67 (53%) and 53/67 (79%) at 10(-4)BCR-ABL1/ABL1. Standardized RNA extraction, cDNA synthesis and cycler platforms did not improve results further, whereas stringent application of technical criteria for assay quality and uniform criteria for data interpretation and reporting were essential. We provide detailed laboratory recommendations for the standardized MRD analysis in routine diagnostic settings and in multicenter clinical trials for Ph + ALL.
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| 14. |
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| 15. |
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| 16. |
- Schaal, Wesley, PhD, et al.
(författare)
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Migrating to Long-Read Sequencing for Clinical Routine BCR-ABL1 TKI Resistance Mutation Screening
- 2022
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Ingår i: Cancer Informatics. - : Sage Publications. - 1176-9351. ; 21, s. 1-8
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Tidskriftsartikel (refereegranskat)abstract
- OBJECTIVE: The aim of this project was to implement long-read sequencing for BCR-ABL1 TKI resistance mutation screening in a clinical setting for patients undergoing treatment for chronic myeloid leukemia.MATERIALS AND METHODS: Processes were established for registering and transferring samples from the clinic to an academic sequencing facility for long-read sequencing. An automated analysis pipeline for detecting mutations was established, and an information system was implemented comprising features for data management, analysis and visualization. Clinical validation was performed by identifying BCR-ABL1 TKI resistance mutations by Sanger and long-read sequencing in parallel. The developed software is available as open source via GitHub at https://github.com/pharmbio/clampRESULTS: The information system enabled traceable transfer of samples from the clinic to the sequencing facility, robust and automated analysis of the long-read sequence data, and communication of results from sequence analysis in a reporting format that could be easily interpreted and acted upon by clinical experts. In a validation study, all 17 resistance mutations found by Sanger sequencing were also detected by long-read sequencing. An additional 16 mutations were found only by long-read sequencing, all of them with frequencies below the limit of detection for Sanger sequencing. The clonal distributions of co-existing mutations were automatically resolved through the long- read data analysis. After the implementation and validation, the clinical laboratory switched their routine protocol from using Sanger to long-read sequencing for this application.CONCLUSIONS: Long-read sequencing delivers results with higher sensitivity compared to Sanger sequencing and enables earlier detection of emerging TKI resistance mutations. The developed processes, analysis workflow, and software components lower barriers for adoption and could be extended to other applications.KEYWORDS: Long-read sequencing, SMRT sequencing, drug resistance, chronic myeloid leukemia, BCR-ABL1, CML, mutation screening
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| 17. |
- Thörn, Ingrid, 1957-, et al.
(författare)
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Monitoring minimal residual disease with flow cytometry, antigen-receptor gene rearrangements and fusion transcript quantification in Philadelphia-positive childhood acute lymphoblastic leukemia
- 2009
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Ingår i: Leukemia Research. - : Elsevier BV. - 0145-2126 .- 1873-5835. ; 33:8, s. 1047-1054
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Tidskriftsartikel (refereegranskat)abstract
- In this study, we followed minimal residual disease (MRD) in eight children with Philadelphia-positive acute lymphoblastic leukemia (Ph+ ALL) using (i) flow cytometry (FCM), (ii) real-time quantitative PCR of IG/TCR gene rearrangements and (iii) RT-PCR detecting fusion gene transcripts. In six of the eight cases the kinetics of MRD clearance was comparable. One of the two discordant cases could be explained by presence of an alternative fusion transcript. The other discordant case showed high BCR-ABL1 RNA level while the other methods did not detect any MRD. In our limited material quantitative RT-PCR of fusion gene transcripts seemed particularly useful to measure MRD in Ph+ ALL. However, BCR-ABL1 expression may not reflect the percentage of leukemic cells as FCM and IG/TCR rearrangement quantification do, and these methods are thus complementary.
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| 18. |
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