Contributions
Abstract: EP392
Type: E-Poster Presentation
Session title: Acute myeloid leukemia - Biology & Translational Research
Background
Acute myeloid leukemia (AML) with translocation 8;21 (AML8;21) is a core-binding factor AML characterized by the oncogenic fusion protein RUNX1-RUNXT1. AML8;21 is one of the most common types of AML and is associated with a favourable prognosis with an estimated 5-year overall survival of 60-70%. However, 30% of patients relapse following first line treatment, leaving these patients with very limited treatment options. Patients older than 65 years represent a particular challenge as many are not candidates for treatment with cytarabine and daunorubicin. The RUNX1-RUNX1T1 fusion protein impairs differentiation and maturation of hematopoietic cells and is unique to the malignant clone, thus constituting a potential target for treatment. In recent years, CRISPR-Cas9 technology has shown great promise in various gene therapy applications. In cancers with known oncogenic drivers, such as AML8;21, there is further potential for novel treatment strategy development utilizing CRISPR-Cas9.
Aims
In this study, we aimed to utilize a novel CRISPR-based treatment strategy to target the RUNX1-RUNX1T1 fusion gene.
Methods
The oncogenic fusion gene RUNX1-RUNX1T1, was targeted with CRISPR-Cas9 in the human AML cell line, Kasumi-1, and in archival cells obtained from RUNX1-RUNX1T1-positive AML patients treated at the Department of Hematology at Aarhus University Hospital, Denmark. CRISPR-Cas9-based disruption of the RUNX1-RUNX1T1 fusion gene was validated using targeted polymerase chain reaction (PCR) and Sanger sequencing. Kasumi-1 cell population growth and cell population proliferation was investigated following CRISPR-CAS9-based disruption of RUNX1-RUNX1T1. Six nude mice were injected with CRISPR-Cas9 RUNX1-RUNX1T1 disrupted Kasumi-1 cells or non-treated Kasumi-1 cells in the right and left flank, respectively. Animals were sacrificed after four weeks and tumor volume was evaluated.
Results
We show CRISPR-Cas9-based disruption of RUNX1-RUNX1T1 in Kasumi-1 cell line and AML-patient-derived cells. In vitro cell growth rate and proliferation were decreased by 58 % and 74 %, respectively, in CRISPR-Cas9 RUNX1-RUNX1T1 disrupted Kasumi-1 cells compared to controls. Four out of 6 mice had tumor growth following injection with RUNX1-RUNX1T1 disrupted or control Kasumi-1 cells an thus allowed for analysis of RUNX1-RUNX1T1 disruption effect on tumor volume. Two mice developed a tumor in only the control flank while the two other mice developed tumors in both flanks. In the two mice with tumor development in both flanks, tumor volumes were decreased by 69 % and 91 % in the CRISPR-Cas9 RUNX1-RUNX1T1 disrupted flank, as compared to the control flank.
Conclusion
In this proof of principle study, we demonstrate a novel CRISPR-based technology that effectively inhibits proliferation and decreases tumor volume in vitro and in vivo. Our results suggest that targeting the RUNX1-RUNX1T1 fusion gene can potentially be a future treatment strategy for patients with AML8;21.
Keyword(s): Acute myeloid leukemia, Gene therapy
Abstract: EP392
Type: E-Poster Presentation
Session title: Acute myeloid leukemia - Biology & Translational Research
Background
Acute myeloid leukemia (AML) with translocation 8;21 (AML8;21) is a core-binding factor AML characterized by the oncogenic fusion protein RUNX1-RUNXT1. AML8;21 is one of the most common types of AML and is associated with a favourable prognosis with an estimated 5-year overall survival of 60-70%. However, 30% of patients relapse following first line treatment, leaving these patients with very limited treatment options. Patients older than 65 years represent a particular challenge as many are not candidates for treatment with cytarabine and daunorubicin. The RUNX1-RUNX1T1 fusion protein impairs differentiation and maturation of hematopoietic cells and is unique to the malignant clone, thus constituting a potential target for treatment. In recent years, CRISPR-Cas9 technology has shown great promise in various gene therapy applications. In cancers with known oncogenic drivers, such as AML8;21, there is further potential for novel treatment strategy development utilizing CRISPR-Cas9.
Aims
In this study, we aimed to utilize a novel CRISPR-based treatment strategy to target the RUNX1-RUNX1T1 fusion gene.
Methods
The oncogenic fusion gene RUNX1-RUNX1T1, was targeted with CRISPR-Cas9 in the human AML cell line, Kasumi-1, and in archival cells obtained from RUNX1-RUNX1T1-positive AML patients treated at the Department of Hematology at Aarhus University Hospital, Denmark. CRISPR-Cas9-based disruption of the RUNX1-RUNX1T1 fusion gene was validated using targeted polymerase chain reaction (PCR) and Sanger sequencing. Kasumi-1 cell population growth and cell population proliferation was investigated following CRISPR-CAS9-based disruption of RUNX1-RUNX1T1. Six nude mice were injected with CRISPR-Cas9 RUNX1-RUNX1T1 disrupted Kasumi-1 cells or non-treated Kasumi-1 cells in the right and left flank, respectively. Animals were sacrificed after four weeks and tumor volume was evaluated.
Results
We show CRISPR-Cas9-based disruption of RUNX1-RUNX1T1 in Kasumi-1 cell line and AML-patient-derived cells. In vitro cell growth rate and proliferation were decreased by 58 % and 74 %, respectively, in CRISPR-Cas9 RUNX1-RUNX1T1 disrupted Kasumi-1 cells compared to controls. Four out of 6 mice had tumor growth following injection with RUNX1-RUNX1T1 disrupted or control Kasumi-1 cells an thus allowed for analysis of RUNX1-RUNX1T1 disruption effect on tumor volume. Two mice developed a tumor in only the control flank while the two other mice developed tumors in both flanks. In the two mice with tumor development in both flanks, tumor volumes were decreased by 69 % and 91 % in the CRISPR-Cas9 RUNX1-RUNX1T1 disrupted flank, as compared to the control flank.
Conclusion
In this proof of principle study, we demonstrate a novel CRISPR-based technology that effectively inhibits proliferation and decreases tumor volume in vitro and in vivo. Our results suggest that targeting the RUNX1-RUNX1T1 fusion gene can potentially be a future treatment strategy for patients with AML8;21.
Keyword(s): Acute myeloid leukemia, Gene therapy