ALK-POSITIVE ANAPLASTIC LARGE CELL LYMPHOMA WITH THE VARIANT EEF1G-, RNF213- AND ATIC-ALK FUSIONS IS FEATURED BY COPY NUMBER GAIN OF THE REARRANGED ALK GENE
(Abstract release date: 05/19/16)
EHA Library. Wlodarska I. 06/09/16; 132936; E1387
Prof. Dr. Iwona Wlodarska
Contributions
Contributions
Abstract
Abstract: E1387
Type: Eposter Presentation
Background
Molecular pathogenesis of ALK-positive Anaplastic Large Cell Lymphoma (ALK+ ALCL) is not completely understood. Approximately 80% of ALK+ ALCL cases harbor the t(2;5)(p23;q35)-associated NPM1-ALK rearrangement, while variant 2p23/ALK translocations involving at least nine partner genes have been identified in the remaining cases. The 5’ALK partners play a key role in the constitutive activation of the chimeric protein by mediating its oligomerization and its subcellular localization. In addition, they impact a range of biological activities of ALK chimeras, including proliferation, transformation and metastatic capacities. Comparative analysis of biological properties of ALK oncoproteins, however, is hampered by the relative low frequency of different variant ALK fusions.
Aims
The aim of the present study was to identify novel ALK-related fusions in ALK+ ALCL and to gain more insight into the molecular pathogenesis of this tumor.
Methods
Six cases of ALK+ ALCL with a cytoplasmic expression of ALK recently diagnosed in our institution were selected. The ALK fusions were characterized using 5’RACE PCR, low coverage full genome sequencing (LCFGS), FISH, array CGH and QRT-PCR. Functional studies were performed on the IL3-dependent Ba/F3 cell line transformation model.
Results
The ALK partner genes identified in the present cases included EEF1G (Eukaryotic translation elongation factor 1 gamma), a novel ALK partner located at 11q12.3 (one case), and the already known genes, RNF213/ALO17 (17q25) (one case) and ATIC (2q35) (four cases). Notably, all six cases displayed a similar LSI ALK break-apart FISH pattern showing copy number gain of the rearranged ALK gene. The LSI 3’ALK/red signal was duplicated on der(11)t(2;11)(p23;q12.3) in the case with EEF1G-ALK and amplified in cases with the RNF213-ALK (5-7 extra red signals) and ATIC-ALK (2-4 extra red signals) rearrangements. FISH pattern in the ATIC-ALK cases suggests the presence of underlying inv(2)(p23q35) associated with one or two copies of the derivative i(2)(q10) carrying two additional ATIC-ALK loci each, as previously reported (PMID: 10706887). FISH findings were confirmed by array CGH in two available cases. These data provide a strong evidence that ALCL driven by at least three variant ALK fusions (EEF1G-, RFN213- and ATIC-ALK), but not by the classic NPM1-ALK, requires an increased gene dosage of the rearranged ALK. To assess whether this need is caused by the weaker promoter of EEF1G, RFN213 and ATIC, compared to the NPM1 promoter, we determined at first the relative mRNA expression level of the four ALK partner genes. The study revealed a significantly lower expression of EEF1G, RNF213 and ATIC in nonmalignant lymph nodes when compared to NPM1. In the next step, we compared oncogenic potential of all four fusions in the murine hematopoietic IL-3 dependent Ba/F3 cell line. We found that EEF1G-ALK, RNF213-ALK and ATIC-ALK are less potent to transform BaF3 cells than NPM1-ALK, which showed the highest oncogenic potential.
Conclusion
ALK+ ALCL driven by three variant ALK fusions, EEF1G-ALK (novel), RNF213-ALK and ATIC-ALK, are characterized by copy number gain of the rearranged ALK. These lymphomas, but not the NPM1-ALK-positive ALCL, likely require an increased gene dosage of the rearranged ALK to compensate the relatively low and insufficient expression of the chimeric gene driven by the partner genes. Occurrence of the ATIC-ALK rearrangement in four out of six cases analyzed confirms that ATIC-ALK is the most prevalent variant fusion in ALK+ ALCL.
Session topic: E-poster
Keyword(s): ALK/ALCL, Chromosomal translocation, Gene dosage
Type: Eposter Presentation
Background
Molecular pathogenesis of ALK-positive Anaplastic Large Cell Lymphoma (ALK+ ALCL) is not completely understood. Approximately 80% of ALK+ ALCL cases harbor the t(2;5)(p23;q35)-associated NPM1-ALK rearrangement, while variant 2p23/ALK translocations involving at least nine partner genes have been identified in the remaining cases. The 5’ALK partners play a key role in the constitutive activation of the chimeric protein by mediating its oligomerization and its subcellular localization. In addition, they impact a range of biological activities of ALK chimeras, including proliferation, transformation and metastatic capacities. Comparative analysis of biological properties of ALK oncoproteins, however, is hampered by the relative low frequency of different variant ALK fusions.
Aims
The aim of the present study was to identify novel ALK-related fusions in ALK+ ALCL and to gain more insight into the molecular pathogenesis of this tumor.
Methods
Six cases of ALK+ ALCL with a cytoplasmic expression of ALK recently diagnosed in our institution were selected. The ALK fusions were characterized using 5’RACE PCR, low coverage full genome sequencing (LCFGS), FISH, array CGH and QRT-PCR. Functional studies were performed on the IL3-dependent Ba/F3 cell line transformation model.
Results
The ALK partner genes identified in the present cases included EEF1G (Eukaryotic translation elongation factor 1 gamma), a novel ALK partner located at 11q12.3 (one case), and the already known genes, RNF213/ALO17 (17q25) (one case) and ATIC (2q35) (four cases). Notably, all six cases displayed a similar LSI ALK break-apart FISH pattern showing copy number gain of the rearranged ALK gene. The LSI 3’ALK/red signal was duplicated on der(11)t(2;11)(p23;q12.3) in the case with EEF1G-ALK and amplified in cases with the RNF213-ALK (5-7 extra red signals) and ATIC-ALK (2-4 extra red signals) rearrangements. FISH pattern in the ATIC-ALK cases suggests the presence of underlying inv(2)(p23q35) associated with one or two copies of the derivative i(2)(q10) carrying two additional ATIC-ALK loci each, as previously reported (PMID: 10706887). FISH findings were confirmed by array CGH in two available cases. These data provide a strong evidence that ALCL driven by at least three variant ALK fusions (EEF1G-, RFN213- and ATIC-ALK), but not by the classic NPM1-ALK, requires an increased gene dosage of the rearranged ALK. To assess whether this need is caused by the weaker promoter of EEF1G, RFN213 and ATIC, compared to the NPM1 promoter, we determined at first the relative mRNA expression level of the four ALK partner genes. The study revealed a significantly lower expression of EEF1G, RNF213 and ATIC in nonmalignant lymph nodes when compared to NPM1. In the next step, we compared oncogenic potential of all four fusions in the murine hematopoietic IL-3 dependent Ba/F3 cell line. We found that EEF1G-ALK, RNF213-ALK and ATIC-ALK are less potent to transform BaF3 cells than NPM1-ALK, which showed the highest oncogenic potential.
Conclusion
ALK+ ALCL driven by three variant ALK fusions, EEF1G-ALK (novel), RNF213-ALK and ATIC-ALK, are characterized by copy number gain of the rearranged ALK. These lymphomas, but not the NPM1-ALK-positive ALCL, likely require an increased gene dosage of the rearranged ALK to compensate the relatively low and insufficient expression of the chimeric gene driven by the partner genes. Occurrence of the ATIC-ALK rearrangement in four out of six cases analyzed confirms that ATIC-ALK is the most prevalent variant fusion in ALK+ ALCL.
Session topic: E-poster
Keyword(s): ALK/ALCL, Chromosomal translocation, Gene dosage
Abstract: E1387
Type: Eposter Presentation
Background
Molecular pathogenesis of ALK-positive Anaplastic Large Cell Lymphoma (ALK+ ALCL) is not completely understood. Approximately 80% of ALK+ ALCL cases harbor the t(2;5)(p23;q35)-associated NPM1-ALK rearrangement, while variant 2p23/ALK translocations involving at least nine partner genes have been identified in the remaining cases. The 5’ALK partners play a key role in the constitutive activation of the chimeric protein by mediating its oligomerization and its subcellular localization. In addition, they impact a range of biological activities of ALK chimeras, including proliferation, transformation and metastatic capacities. Comparative analysis of biological properties of ALK oncoproteins, however, is hampered by the relative low frequency of different variant ALK fusions.
Aims
The aim of the present study was to identify novel ALK-related fusions in ALK+ ALCL and to gain more insight into the molecular pathogenesis of this tumor.
Methods
Six cases of ALK+ ALCL with a cytoplasmic expression of ALK recently diagnosed in our institution were selected. The ALK fusions were characterized using 5’RACE PCR, low coverage full genome sequencing (LCFGS), FISH, array CGH and QRT-PCR. Functional studies were performed on the IL3-dependent Ba/F3 cell line transformation model.
Results
The ALK partner genes identified in the present cases included EEF1G (Eukaryotic translation elongation factor 1 gamma), a novel ALK partner located at 11q12.3 (one case), and the already known genes, RNF213/ALO17 (17q25) (one case) and ATIC (2q35) (four cases). Notably, all six cases displayed a similar LSI ALK break-apart FISH pattern showing copy number gain of the rearranged ALK gene. The LSI 3’ALK/red signal was duplicated on der(11)t(2;11)(p23;q12.3) in the case with EEF1G-ALK and amplified in cases with the RNF213-ALK (5-7 extra red signals) and ATIC-ALK (2-4 extra red signals) rearrangements. FISH pattern in the ATIC-ALK cases suggests the presence of underlying inv(2)(p23q35) associated with one or two copies of the derivative i(2)(q10) carrying two additional ATIC-ALK loci each, as previously reported (PMID: 10706887). FISH findings were confirmed by array CGH in two available cases. These data provide a strong evidence that ALCL driven by at least three variant ALK fusions (EEF1G-, RFN213- and ATIC-ALK), but not by the classic NPM1-ALK, requires an increased gene dosage of the rearranged ALK. To assess whether this need is caused by the weaker promoter of EEF1G, RFN213 and ATIC, compared to the NPM1 promoter, we determined at first the relative mRNA expression level of the four ALK partner genes. The study revealed a significantly lower expression of EEF1G, RNF213 and ATIC in nonmalignant lymph nodes when compared to NPM1. In the next step, we compared oncogenic potential of all four fusions in the murine hematopoietic IL-3 dependent Ba/F3 cell line. We found that EEF1G-ALK, RNF213-ALK and ATIC-ALK are less potent to transform BaF3 cells than NPM1-ALK, which showed the highest oncogenic potential.
Conclusion
ALK+ ALCL driven by three variant ALK fusions, EEF1G-ALK (novel), RNF213-ALK and ATIC-ALK, are characterized by copy number gain of the rearranged ALK. These lymphomas, but not the NPM1-ALK-positive ALCL, likely require an increased gene dosage of the rearranged ALK to compensate the relatively low and insufficient expression of the chimeric gene driven by the partner genes. Occurrence of the ATIC-ALK rearrangement in four out of six cases analyzed confirms that ATIC-ALK is the most prevalent variant fusion in ALK+ ALCL.
Session topic: E-poster
Keyword(s): ALK/ALCL, Chromosomal translocation, Gene dosage
Type: Eposter Presentation
Background
Molecular pathogenesis of ALK-positive Anaplastic Large Cell Lymphoma (ALK+ ALCL) is not completely understood. Approximately 80% of ALK+ ALCL cases harbor the t(2;5)(p23;q35)-associated NPM1-ALK rearrangement, while variant 2p23/ALK translocations involving at least nine partner genes have been identified in the remaining cases. The 5’ALK partners play a key role in the constitutive activation of the chimeric protein by mediating its oligomerization and its subcellular localization. In addition, they impact a range of biological activities of ALK chimeras, including proliferation, transformation and metastatic capacities. Comparative analysis of biological properties of ALK oncoproteins, however, is hampered by the relative low frequency of different variant ALK fusions.
Aims
The aim of the present study was to identify novel ALK-related fusions in ALK+ ALCL and to gain more insight into the molecular pathogenesis of this tumor.
Methods
Six cases of ALK+ ALCL with a cytoplasmic expression of ALK recently diagnosed in our institution were selected. The ALK fusions were characterized using 5’RACE PCR, low coverage full genome sequencing (LCFGS), FISH, array CGH and QRT-PCR. Functional studies were performed on the IL3-dependent Ba/F3 cell line transformation model.
Results
The ALK partner genes identified in the present cases included EEF1G (Eukaryotic translation elongation factor 1 gamma), a novel ALK partner located at 11q12.3 (one case), and the already known genes, RNF213/ALO17 (17q25) (one case) and ATIC (2q35) (four cases). Notably, all six cases displayed a similar LSI ALK break-apart FISH pattern showing copy number gain of the rearranged ALK gene. The LSI 3’ALK/red signal was duplicated on der(11)t(2;11)(p23;q12.3) in the case with EEF1G-ALK and amplified in cases with the RNF213-ALK (5-7 extra red signals) and ATIC-ALK (2-4 extra red signals) rearrangements. FISH pattern in the ATIC-ALK cases suggests the presence of underlying inv(2)(p23q35) associated with one or two copies of the derivative i(2)(q10) carrying two additional ATIC-ALK loci each, as previously reported (PMID: 10706887). FISH findings were confirmed by array CGH in two available cases. These data provide a strong evidence that ALCL driven by at least three variant ALK fusions (EEF1G-, RFN213- and ATIC-ALK), but not by the classic NPM1-ALK, requires an increased gene dosage of the rearranged ALK. To assess whether this need is caused by the weaker promoter of EEF1G, RFN213 and ATIC, compared to the NPM1 promoter, we determined at first the relative mRNA expression level of the four ALK partner genes. The study revealed a significantly lower expression of EEF1G, RNF213 and ATIC in nonmalignant lymph nodes when compared to NPM1. In the next step, we compared oncogenic potential of all four fusions in the murine hematopoietic IL-3 dependent Ba/F3 cell line. We found that EEF1G-ALK, RNF213-ALK and ATIC-ALK are less potent to transform BaF3 cells than NPM1-ALK, which showed the highest oncogenic potential.
Conclusion
ALK+ ALCL driven by three variant ALK fusions, EEF1G-ALK (novel), RNF213-ALK and ATIC-ALK, are characterized by copy number gain of the rearranged ALK. These lymphomas, but not the NPM1-ALK-positive ALCL, likely require an increased gene dosage of the rearranged ALK to compensate the relatively low and insufficient expression of the chimeric gene driven by the partner genes. Occurrence of the ATIC-ALK rearrangement in four out of six cases analyzed confirms that ATIC-ALK is the most prevalent variant fusion in ALK+ ALCL.
Session topic: E-poster
Keyword(s): ALK/ALCL, Chromosomal translocation, Gene dosage
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