Contributions
Abstract: PB1712
Type: Publication Only
Background
Acute myeloid leukemia (AML) is a complex and dynamic disorder characterized by a wide range of recurrent driver genetic defects. The introduction of next generation sequencing (NGS) in AML genome investigation led to a better understanding of the mutational spectrum intricacy. Characterization of the patterns of mutation co-occurrence has become equally significant, as most AML patients bear more than one driver mutation. The functional consequences and clinical relevance of mutations and patterns of co-mutation are not fully understood, thus further studies are needed in order to inform clinical practice.
Aims
We report on the results of targeted NGS and fragment analysis for characterization of the mutational landscape in a group of 12 Romanian AML patients with normal cytogenetics (NC-AML).
Methods
Targeted NGS testing with Ion AmpliSeq™ AML Research Panel (ThermoFisher Scientific) was performed on genomic DNA extracted from diagnostic bone marrow samples according to manufacturer’s recommendations. Nineteen genes were targeted for entire coding regions (CEPBA, DNMT3A, GATA2, TET2, TP53) or mutational hotspots (ASXL1, BRAF, CBL, FLT3, IDH1/2, JAK2, KIT, KRAS, NRAS, NPM1, PTPN11, RUNX1, WT1). Ion PGM System (ThermoFisher Scientific) and NextGENe v.2.4.2.1 (SoftGenetics) were used for sequencing and data analysis. Internal tandem duplication of FLT3 gene (FLT3-ITD) were assessed by PCR amplification of exons 13 and 14 and the spanning intronic region of the FLT3 gene. The amplified products were analysed by automated electrophoresis on the 2100 Bioanalyzer system (Agilent Technologies).
Results
A total of 38 mutations, with a median of 3 mutations per patient (ranging from 2 to 5 mutations/patient), were identified in our patient group. Among these, 31 were driver mutations recurrent in myeloid malignancies, while 7 mutations, with in silico predicted deleterious effects, were not previously reported in cancer databases (COSMIC, ClinVar). The highest mutation frequency was observed for NPM1, as previously reported in NC-AML, followed by DNMT3A gene. The mutation co-occurrence patterns in our patient group followed the previously reported ones, however some uncommon association were observed, such as between NPM1 and NRASQ61. None of the investigated patients showed FLT3-ITD. However, two patients presented FLT3-TKD point mutations in association with NPM1 mutations.
Conclusion
Targeted NGS combined with fragment analysis proved a successful approach for molecular profiling of NC-AML highlighting the complexity of the molecular profiles in this AML subgroup. The study represent one of the first NGS analyses of Romanian patients with NC-AML. The mutational data obtained in our patients are consistent with previously published data, while also revealing new insights, such as previously unreported variants or new mutational associations. The data contribute to a better understanding of the mutational landscape of AML and, ultimately, to an improved patient care.
Session topic: 3. Acute myeloid leukemia - Biology & Translational Research
Keyword(s): Acute Myeloid Leukemia, mutation analysis
Abstract: PB1712
Type: Publication Only
Background
Acute myeloid leukemia (AML) is a complex and dynamic disorder characterized by a wide range of recurrent driver genetic defects. The introduction of next generation sequencing (NGS) in AML genome investigation led to a better understanding of the mutational spectrum intricacy. Characterization of the patterns of mutation co-occurrence has become equally significant, as most AML patients bear more than one driver mutation. The functional consequences and clinical relevance of mutations and patterns of co-mutation are not fully understood, thus further studies are needed in order to inform clinical practice.
Aims
We report on the results of targeted NGS and fragment analysis for characterization of the mutational landscape in a group of 12 Romanian AML patients with normal cytogenetics (NC-AML).
Methods
Targeted NGS testing with Ion AmpliSeq™ AML Research Panel (ThermoFisher Scientific) was performed on genomic DNA extracted from diagnostic bone marrow samples according to manufacturer’s recommendations. Nineteen genes were targeted for entire coding regions (CEPBA, DNMT3A, GATA2, TET2, TP53) or mutational hotspots (ASXL1, BRAF, CBL, FLT3, IDH1/2, JAK2, KIT, KRAS, NRAS, NPM1, PTPN11, RUNX1, WT1). Ion PGM System (ThermoFisher Scientific) and NextGENe v.2.4.2.1 (SoftGenetics) were used for sequencing and data analysis. Internal tandem duplication of FLT3 gene (FLT3-ITD) were assessed by PCR amplification of exons 13 and 14 and the spanning intronic region of the FLT3 gene. The amplified products were analysed by automated electrophoresis on the 2100 Bioanalyzer system (Agilent Technologies).
Results
A total of 38 mutations, with a median of 3 mutations per patient (ranging from 2 to 5 mutations/patient), were identified in our patient group. Among these, 31 were driver mutations recurrent in myeloid malignancies, while 7 mutations, with in silico predicted deleterious effects, were not previously reported in cancer databases (COSMIC, ClinVar). The highest mutation frequency was observed for NPM1, as previously reported in NC-AML, followed by DNMT3A gene. The mutation co-occurrence patterns in our patient group followed the previously reported ones, however some uncommon association were observed, such as between NPM1 and NRASQ61. None of the investigated patients showed FLT3-ITD. However, two patients presented FLT3-TKD point mutations in association with NPM1 mutations.
Conclusion
Targeted NGS combined with fragment analysis proved a successful approach for molecular profiling of NC-AML highlighting the complexity of the molecular profiles in this AML subgroup. The study represent one of the first NGS analyses of Romanian patients with NC-AML. The mutational data obtained in our patients are consistent with previously published data, while also revealing new insights, such as previously unreported variants or new mutational associations. The data contribute to a better understanding of the mutational landscape of AML and, ultimately, to an improved patient care.
Session topic: 3. Acute myeloid leukemia - Biology & Translational Research
Keyword(s): Acute Myeloid Leukemia, mutation analysis