GPR56 CONTRIBUTES TO THE DEVELOPMENT OF ACUTE MYELOID LEUKEMIA IN MICE
(Abstract release date: 05/19/16)
EHA Library. Kirsten N. 06/12/16; 135300; S806
Mrs. Nicole Kirsten
Contributions
Contributions
Abstract
Abstract: S806
Type: Oral Presentation
Presentation during EHA21: On Sunday, June 12, 2016 from 08:45 - 09:00
Location: Hall A3
Background
One of the major goals in AML research is to identify key characteristics of leukemic stem cells (LSCs) in patients with AML. Gene expression profiling in functionally validated primary AML LSCs has established a gene signature, which consists of 42 genes associated with LSC properties and inferior treatment outcome in patients with AML (Eppert et al., Nature Medicine 2011). Among these 42 genes the G-protein coupled receptor 56 (GPR56) was highly expressed in the LSC enriched fraction compared to the CD34-negative leukemic bulk.
Aims
The aim of this study was to determine the functional role of Gpr56 in AML and to test its accessibility for antibody – mediated targeting of human AML LSCs.
Methods
For this retrovirally engineered overexpression of Gpr56 and the murine bone marrow transplantation model was used. For analyses in the human system NSG xenografting was employed.
Results
43 clinically and molecularly annotated AML samples of different genotypes were analysed by RQ-PCR, confirming high expression in NPM1c mutated AML and lower expression in the CBF INV16 AML subtype with a 4.7fold difference between the two genotypes. Of note, expression of GPR56 was highest in normal CD34+ hematopoietic stem and progenitor cells, indicating that there is substantial, but not aberrantly high GPR56 expression in human AML cells. To test whether expression levels of GPR56 correlated with treatment outcome, microarray based GPR56 expression of 423 clinically and molecularly annotated newly diagnosed patients treated in two independent prospective clinical trials was correlated with event free and overall survival: when the median expression level of GPR56 was taken as cut-off, high GPR56 expression was associated with inferior event free and overall survival in the total cohort of patients (n=423) as well as in the patients with normal karyotype (n = 184) Functional relevance of GPR56 expression was validated in mice, in which co-expression of Gpr56 and Hoxa9 induced leukemia in transplanted mice after a median time of 148 days post transplantation (range 93 – 264) in contrast to the Hoxa9 and Gpr56 controls, which did not develop any signs of disease in the observation period of up to 380 days post transplant. Vice versa, the onset of leukemia was significantly delayed by shRNA-mediated knockdown of GPR56 in Hoxa9/Meis1 transduced cells when transplanted into mice. Overexpression of Gpr56 grossly changed the molecular phenotype of Hoxa9 transduced cells affecting pathways involved in G – coupled protein receptors and associated intracellular signaling. As GPR56 is part of the LSC signature in human AML, associated with inferior prognosis and as we had shown that GPR56 in collaboration with the oncogene Hoxa9 contributes to myeloid leukemogenesis, it was tested whether antibody-mediated blockage of the surface receptor GPR56 impairs leukemic engraftment in the NSG mouse model. Blockage of GPR56 with a monoclonal naked antibody significantly impaired engraftment into NSG mice. Importantly, also FLT3-ITD3+ NPM1c- primary patient samples showed an up to 68 % decrease in BM engraftment at 12 weeks (p< 0.04).
Conclusion
Taken together, these data demonstrate that high expression of GPR56 is able to contribute to AML development and characterize the GPR56 as a potential novel target for antibody – mediated anti-leukemic strategies.
Session topic: AML Biology - Novel targeted therapies
Keyword(s): Acute myeloid leukemia, G-protein-coupled receptors, Leukemic stem cell
Type: Oral Presentation
Presentation during EHA21: On Sunday, June 12, 2016 from 08:45 - 09:00
Location: Hall A3
Background
One of the major goals in AML research is to identify key characteristics of leukemic stem cells (LSCs) in patients with AML. Gene expression profiling in functionally validated primary AML LSCs has established a gene signature, which consists of 42 genes associated with LSC properties and inferior treatment outcome in patients with AML (Eppert et al., Nature Medicine 2011). Among these 42 genes the G-protein coupled receptor 56 (GPR56) was highly expressed in the LSC enriched fraction compared to the CD34-negative leukemic bulk.
Aims
The aim of this study was to determine the functional role of Gpr56 in AML and to test its accessibility for antibody – mediated targeting of human AML LSCs.
Methods
For this retrovirally engineered overexpression of Gpr56 and the murine bone marrow transplantation model was used. For analyses in the human system NSG xenografting was employed.
Results
43 clinically and molecularly annotated AML samples of different genotypes were analysed by RQ-PCR, confirming high expression in NPM1c mutated AML and lower expression in the CBF INV16 AML subtype with a 4.7fold difference between the two genotypes. Of note, expression of GPR56 was highest in normal CD34+ hematopoietic stem and progenitor cells, indicating that there is substantial, but not aberrantly high GPR56 expression in human AML cells. To test whether expression levels of GPR56 correlated with treatment outcome, microarray based GPR56 expression of 423 clinically and molecularly annotated newly diagnosed patients treated in two independent prospective clinical trials was correlated with event free and overall survival: when the median expression level of GPR56 was taken as cut-off, high GPR56 expression was associated with inferior event free and overall survival in the total cohort of patients (n=423) as well as in the patients with normal karyotype (n = 184) Functional relevance of GPR56 expression was validated in mice, in which co-expression of Gpr56 and Hoxa9 induced leukemia in transplanted mice after a median time of 148 days post transplantation (range 93 – 264) in contrast to the Hoxa9 and Gpr56 controls, which did not develop any signs of disease in the observation period of up to 380 days post transplant. Vice versa, the onset of leukemia was significantly delayed by shRNA-mediated knockdown of GPR56 in Hoxa9/Meis1 transduced cells when transplanted into mice. Overexpression of Gpr56 grossly changed the molecular phenotype of Hoxa9 transduced cells affecting pathways involved in G – coupled protein receptors and associated intracellular signaling. As GPR56 is part of the LSC signature in human AML, associated with inferior prognosis and as we had shown that GPR56 in collaboration with the oncogene Hoxa9 contributes to myeloid leukemogenesis, it was tested whether antibody-mediated blockage of the surface receptor GPR56 impairs leukemic engraftment in the NSG mouse model. Blockage of GPR56 with a monoclonal naked antibody significantly impaired engraftment into NSG mice. Importantly, also FLT3-ITD3+ NPM1c- primary patient samples showed an up to 68 % decrease in BM engraftment at 12 weeks (p< 0.04).
Conclusion
Taken together, these data demonstrate that high expression of GPR56 is able to contribute to AML development and characterize the GPR56 as a potential novel target for antibody – mediated anti-leukemic strategies.
Session topic: AML Biology - Novel targeted therapies
Keyword(s): Acute myeloid leukemia, G-protein-coupled receptors, Leukemic stem cell
Abstract: S806
Type: Oral Presentation
Presentation during EHA21: On Sunday, June 12, 2016 from 08:45 - 09:00
Location: Hall A3
Background
One of the major goals in AML research is to identify key characteristics of leukemic stem cells (LSCs) in patients with AML. Gene expression profiling in functionally validated primary AML LSCs has established a gene signature, which consists of 42 genes associated with LSC properties and inferior treatment outcome in patients with AML (Eppert et al., Nature Medicine 2011). Among these 42 genes the G-protein coupled receptor 56 (GPR56) was highly expressed in the LSC enriched fraction compared to the CD34-negative leukemic bulk.
Aims
The aim of this study was to determine the functional role of Gpr56 in AML and to test its accessibility for antibody – mediated targeting of human AML LSCs.
Methods
For this retrovirally engineered overexpression of Gpr56 and the murine bone marrow transplantation model was used. For analyses in the human system NSG xenografting was employed.
Results
43 clinically and molecularly annotated AML samples of different genotypes were analysed by RQ-PCR, confirming high expression in NPM1c mutated AML and lower expression in the CBF INV16 AML subtype with a 4.7fold difference between the two genotypes. Of note, expression of GPR56 was highest in normal CD34+ hematopoietic stem and progenitor cells, indicating that there is substantial, but not aberrantly high GPR56 expression in human AML cells. To test whether expression levels of GPR56 correlated with treatment outcome, microarray based GPR56 expression of 423 clinically and molecularly annotated newly diagnosed patients treated in two independent prospective clinical trials was correlated with event free and overall survival: when the median expression level of GPR56 was taken as cut-off, high GPR56 expression was associated with inferior event free and overall survival in the total cohort of patients (n=423) as well as in the patients with normal karyotype (n = 184) Functional relevance of GPR56 expression was validated in mice, in which co-expression of Gpr56 and Hoxa9 induced leukemia in transplanted mice after a median time of 148 days post transplantation (range 93 – 264) in contrast to the Hoxa9 and Gpr56 controls, which did not develop any signs of disease in the observation period of up to 380 days post transplant. Vice versa, the onset of leukemia was significantly delayed by shRNA-mediated knockdown of GPR56 in Hoxa9/Meis1 transduced cells when transplanted into mice. Overexpression of Gpr56 grossly changed the molecular phenotype of Hoxa9 transduced cells affecting pathways involved in G – coupled protein receptors and associated intracellular signaling. As GPR56 is part of the LSC signature in human AML, associated with inferior prognosis and as we had shown that GPR56 in collaboration with the oncogene Hoxa9 contributes to myeloid leukemogenesis, it was tested whether antibody-mediated blockage of the surface receptor GPR56 impairs leukemic engraftment in the NSG mouse model. Blockage of GPR56 with a monoclonal naked antibody significantly impaired engraftment into NSG mice. Importantly, also FLT3-ITD3+ NPM1c- primary patient samples showed an up to 68 % decrease in BM engraftment at 12 weeks (p< 0.04).
Conclusion
Taken together, these data demonstrate that high expression of GPR56 is able to contribute to AML development and characterize the GPR56 as a potential novel target for antibody – mediated anti-leukemic strategies.
Session topic: AML Biology - Novel targeted therapies
Keyword(s): Acute myeloid leukemia, G-protein-coupled receptors, Leukemic stem cell
Type: Oral Presentation
Presentation during EHA21: On Sunday, June 12, 2016 from 08:45 - 09:00
Location: Hall A3
Background
One of the major goals in AML research is to identify key characteristics of leukemic stem cells (LSCs) in patients with AML. Gene expression profiling in functionally validated primary AML LSCs has established a gene signature, which consists of 42 genes associated with LSC properties and inferior treatment outcome in patients with AML (Eppert et al., Nature Medicine 2011). Among these 42 genes the G-protein coupled receptor 56 (GPR56) was highly expressed in the LSC enriched fraction compared to the CD34-negative leukemic bulk.
Aims
The aim of this study was to determine the functional role of Gpr56 in AML and to test its accessibility for antibody – mediated targeting of human AML LSCs.
Methods
For this retrovirally engineered overexpression of Gpr56 and the murine bone marrow transplantation model was used. For analyses in the human system NSG xenografting was employed.
Results
43 clinically and molecularly annotated AML samples of different genotypes were analysed by RQ-PCR, confirming high expression in NPM1c mutated AML and lower expression in the CBF INV16 AML subtype with a 4.7fold difference between the two genotypes. Of note, expression of GPR56 was highest in normal CD34+ hematopoietic stem and progenitor cells, indicating that there is substantial, but not aberrantly high GPR56 expression in human AML cells. To test whether expression levels of GPR56 correlated with treatment outcome, microarray based GPR56 expression of 423 clinically and molecularly annotated newly diagnosed patients treated in two independent prospective clinical trials was correlated with event free and overall survival: when the median expression level of GPR56 was taken as cut-off, high GPR56 expression was associated with inferior event free and overall survival in the total cohort of patients (n=423) as well as in the patients with normal karyotype (n = 184) Functional relevance of GPR56 expression was validated in mice, in which co-expression of Gpr56 and Hoxa9 induced leukemia in transplanted mice after a median time of 148 days post transplantation (range 93 – 264) in contrast to the Hoxa9 and Gpr56 controls, which did not develop any signs of disease in the observation period of up to 380 days post transplant. Vice versa, the onset of leukemia was significantly delayed by shRNA-mediated knockdown of GPR56 in Hoxa9/Meis1 transduced cells when transplanted into mice. Overexpression of Gpr56 grossly changed the molecular phenotype of Hoxa9 transduced cells affecting pathways involved in G – coupled protein receptors and associated intracellular signaling. As GPR56 is part of the LSC signature in human AML, associated with inferior prognosis and as we had shown that GPR56 in collaboration with the oncogene Hoxa9 contributes to myeloid leukemogenesis, it was tested whether antibody-mediated blockage of the surface receptor GPR56 impairs leukemic engraftment in the NSG mouse model. Blockage of GPR56 with a monoclonal naked antibody significantly impaired engraftment into NSG mice. Importantly, also FLT3-ITD3+ NPM1c- primary patient samples showed an up to 68 % decrease in BM engraftment at 12 weeks (p< 0.04).
Conclusion
Taken together, these data demonstrate that high expression of GPR56 is able to contribute to AML development and characterize the GPR56 as a potential novel target for antibody – mediated anti-leukemic strategies.
Session topic: AML Biology - Novel targeted therapies
Keyword(s): Acute myeloid leukemia, G-protein-coupled receptors, Leukemic stem cell
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