FAST GENERATION OF CONDITIONAL ROSA26-BASED MOUSE MODELS THAT RECAPITULATE GENOMIC EVENTS IN HUMAN T-ALL
(Abstract release date: 05/19/16)
EHA Library. Pieters T. 06/12/16; 135294; S800
Dr. Tim Pieters
Contributions
Contributions
Abstract
Abstract: S800
Type: Oral Presentation
Presentation during EHA21: On Sunday, June 12, 2016 from 08:30 - 08:45
Location: Hall C11
Background
T-cell acute lymphoblastic leukemias (T-ALLs) are aggressive hematologic tumors that result from the malignant transformation of T-cell progenitors. Due to intensified chemotherapy, the prognosis of T-ALL has gradually improved. Nevertheless, this clinical improvement is most pronounced in pediatric treatment protocols, whereas adult patients more often present with primary resistant or relapsed disease.
Aims
Our Aim is to develop new mouse models that mimic oncogenic lesions identified in human disease. These mouse models are critically required to further enhance our knowledge on the molecular mechanisms that drive T-cell leukemogenesis. Moreover, they serve as important pre-clinical models to evaluate new therapeutic strategies for the treatment of human T-ALL.
Methods
Here, we generated a fast method for embryonic stem cell (ESC) targeting and mouse chimera production. We constructed gateway-compatible vectors that allow tailor-made targeting vector design, including conditional expression of a transgene combined with an eGFP/luciferase reporter from the ROSA26(R26)-promoter or from (inducible) exogenous promoters. The final targeting vector is recombination-mediated cassette exchange (RMCE)-compatible and can be inserted in a genetically engineered R26-locus via RMCE. Correct integration of the incoming construct reactivates the NeoR gene and results in up to 100% ESC targeting efficiencies.
Results
Using our technology, we generated R26-based conditional knock-in mouse models for putative oncogenes that have previously been implicated in T-ALL disease biology. More specifically, the MYB leucine zipper transcription factor is aberrantly activated in a subset of T-ALL patients through T-cell receptor driven translocations (t(6;7)(q23;q34)) or genomic duplications of the MYB locus itself. Moreover, Myb was found to be crucial for the initiation of oncogenic superenhancers in T-ALL. In addition, another subset of T-ALLs show specific TCR-mediated translocations that drive aberrant expression of the cell cycle regulator Cyclin D2 (CCND2). To study the in vivo roles of cMyb and Ccnd2 in the pathogenesis of T-ALL, we used the above-mentioned genomic engineering technology to generate cMyb and Ccnd2 conditional knockin mouse models. These animals were crossed with Lck-Cre mice to obtain T-cell specific expression of the oncogenes of interest at moderate pathophysiological relevant levels. Currently, we are monitoring tumor development in T-cell specific homozygous knockin mice. In addition, Myb and Ccnd2 transgenic animals are crossed into a Pten null background to accelerate disease onset. Preliminary data shows that T-cell specific Ccnd2 overexpression cooperates with Pten loss to decrease T-ALL latency.
Conclusion
All together, we have used a fast genomic engineering pipeline to develop new and pathophysiological relevant conditional overexpression mouse models for human T-ALL.
Session topic: ALL Biology - Transcriptional dysregulation
Keyword(s): Acute lymphoblastic leukemia, Embryonic stem cells, Mouse model, Transgenic mice
Type: Oral Presentation
Presentation during EHA21: On Sunday, June 12, 2016 from 08:30 - 08:45
Location: Hall C11
Background
T-cell acute lymphoblastic leukemias (T-ALLs) are aggressive hematologic tumors that result from the malignant transformation of T-cell progenitors. Due to intensified chemotherapy, the prognosis of T-ALL has gradually improved. Nevertheless, this clinical improvement is most pronounced in pediatric treatment protocols, whereas adult patients more often present with primary resistant or relapsed disease.
Aims
Our Aim is to develop new mouse models that mimic oncogenic lesions identified in human disease. These mouse models are critically required to further enhance our knowledge on the molecular mechanisms that drive T-cell leukemogenesis. Moreover, they serve as important pre-clinical models to evaluate new therapeutic strategies for the treatment of human T-ALL.
Methods
Here, we generated a fast method for embryonic stem cell (ESC) targeting and mouse chimera production. We constructed gateway-compatible vectors that allow tailor-made targeting vector design, including conditional expression of a transgene combined with an eGFP/luciferase reporter from the ROSA26(R26)-promoter or from (inducible) exogenous promoters. The final targeting vector is recombination-mediated cassette exchange (RMCE)-compatible and can be inserted in a genetically engineered R26-locus via RMCE. Correct integration of the incoming construct reactivates the NeoR gene and results in up to 100% ESC targeting efficiencies.
Results
Using our technology, we generated R26-based conditional knock-in mouse models for putative oncogenes that have previously been implicated in T-ALL disease biology. More specifically, the MYB leucine zipper transcription factor is aberrantly activated in a subset of T-ALL patients through T-cell receptor driven translocations (t(6;7)(q23;q34)) or genomic duplications of the MYB locus itself. Moreover, Myb was found to be crucial for the initiation of oncogenic superenhancers in T-ALL. In addition, another subset of T-ALLs show specific TCR-mediated translocations that drive aberrant expression of the cell cycle regulator Cyclin D2 (CCND2). To study the in vivo roles of cMyb and Ccnd2 in the pathogenesis of T-ALL, we used the above-mentioned genomic engineering technology to generate cMyb and Ccnd2 conditional knockin mouse models. These animals were crossed with Lck-Cre mice to obtain T-cell specific expression of the oncogenes of interest at moderate pathophysiological relevant levels. Currently, we are monitoring tumor development in T-cell specific homozygous knockin mice. In addition, Myb and Ccnd2 transgenic animals are crossed into a Pten null background to accelerate disease onset. Preliminary data shows that T-cell specific Ccnd2 overexpression cooperates with Pten loss to decrease T-ALL latency.
Conclusion
All together, we have used a fast genomic engineering pipeline to develop new and pathophysiological relevant conditional overexpression mouse models for human T-ALL.
Session topic: ALL Biology - Transcriptional dysregulation
Keyword(s): Acute lymphoblastic leukemia, Embryonic stem cells, Mouse model, Transgenic mice
Abstract: S800
Type: Oral Presentation
Presentation during EHA21: On Sunday, June 12, 2016 from 08:30 - 08:45
Location: Hall C11
Background
T-cell acute lymphoblastic leukemias (T-ALLs) are aggressive hematologic tumors that result from the malignant transformation of T-cell progenitors. Due to intensified chemotherapy, the prognosis of T-ALL has gradually improved. Nevertheless, this clinical improvement is most pronounced in pediatric treatment protocols, whereas adult patients more often present with primary resistant or relapsed disease.
Aims
Our Aim is to develop new mouse models that mimic oncogenic lesions identified in human disease. These mouse models are critically required to further enhance our knowledge on the molecular mechanisms that drive T-cell leukemogenesis. Moreover, they serve as important pre-clinical models to evaluate new therapeutic strategies for the treatment of human T-ALL.
Methods
Here, we generated a fast method for embryonic stem cell (ESC) targeting and mouse chimera production. We constructed gateway-compatible vectors that allow tailor-made targeting vector design, including conditional expression of a transgene combined with an eGFP/luciferase reporter from the ROSA26(R26)-promoter or from (inducible) exogenous promoters. The final targeting vector is recombination-mediated cassette exchange (RMCE)-compatible and can be inserted in a genetically engineered R26-locus via RMCE. Correct integration of the incoming construct reactivates the NeoR gene and results in up to 100% ESC targeting efficiencies.
Results
Using our technology, we generated R26-based conditional knock-in mouse models for putative oncogenes that have previously been implicated in T-ALL disease biology. More specifically, the MYB leucine zipper transcription factor is aberrantly activated in a subset of T-ALL patients through T-cell receptor driven translocations (t(6;7)(q23;q34)) or genomic duplications of the MYB locus itself. Moreover, Myb was found to be crucial for the initiation of oncogenic superenhancers in T-ALL. In addition, another subset of T-ALLs show specific TCR-mediated translocations that drive aberrant expression of the cell cycle regulator Cyclin D2 (CCND2). To study the in vivo roles of cMyb and Ccnd2 in the pathogenesis of T-ALL, we used the above-mentioned genomic engineering technology to generate cMyb and Ccnd2 conditional knockin mouse models. These animals were crossed with Lck-Cre mice to obtain T-cell specific expression of the oncogenes of interest at moderate pathophysiological relevant levels. Currently, we are monitoring tumor development in T-cell specific homozygous knockin mice. In addition, Myb and Ccnd2 transgenic animals are crossed into a Pten null background to accelerate disease onset. Preliminary data shows that T-cell specific Ccnd2 overexpression cooperates with Pten loss to decrease T-ALL latency.
Conclusion
All together, we have used a fast genomic engineering pipeline to develop new and pathophysiological relevant conditional overexpression mouse models for human T-ALL.
Session topic: ALL Biology - Transcriptional dysregulation
Keyword(s): Acute lymphoblastic leukemia, Embryonic stem cells, Mouse model, Transgenic mice
Type: Oral Presentation
Presentation during EHA21: On Sunday, June 12, 2016 from 08:30 - 08:45
Location: Hall C11
Background
T-cell acute lymphoblastic leukemias (T-ALLs) are aggressive hematologic tumors that result from the malignant transformation of T-cell progenitors. Due to intensified chemotherapy, the prognosis of T-ALL has gradually improved. Nevertheless, this clinical improvement is most pronounced in pediatric treatment protocols, whereas adult patients more often present with primary resistant or relapsed disease.
Aims
Our Aim is to develop new mouse models that mimic oncogenic lesions identified in human disease. These mouse models are critically required to further enhance our knowledge on the molecular mechanisms that drive T-cell leukemogenesis. Moreover, they serve as important pre-clinical models to evaluate new therapeutic strategies for the treatment of human T-ALL.
Methods
Here, we generated a fast method for embryonic stem cell (ESC) targeting and mouse chimera production. We constructed gateway-compatible vectors that allow tailor-made targeting vector design, including conditional expression of a transgene combined with an eGFP/luciferase reporter from the ROSA26(R26)-promoter or from (inducible) exogenous promoters. The final targeting vector is recombination-mediated cassette exchange (RMCE)-compatible and can be inserted in a genetically engineered R26-locus via RMCE. Correct integration of the incoming construct reactivates the NeoR gene and results in up to 100% ESC targeting efficiencies.
Results
Using our technology, we generated R26-based conditional knock-in mouse models for putative oncogenes that have previously been implicated in T-ALL disease biology. More specifically, the MYB leucine zipper transcription factor is aberrantly activated in a subset of T-ALL patients through T-cell receptor driven translocations (t(6;7)(q23;q34)) or genomic duplications of the MYB locus itself. Moreover, Myb was found to be crucial for the initiation of oncogenic superenhancers in T-ALL. In addition, another subset of T-ALLs show specific TCR-mediated translocations that drive aberrant expression of the cell cycle regulator Cyclin D2 (CCND2). To study the in vivo roles of cMyb and Ccnd2 in the pathogenesis of T-ALL, we used the above-mentioned genomic engineering technology to generate cMyb and Ccnd2 conditional knockin mouse models. These animals were crossed with Lck-Cre mice to obtain T-cell specific expression of the oncogenes of interest at moderate pathophysiological relevant levels. Currently, we are monitoring tumor development in T-cell specific homozygous knockin mice. In addition, Myb and Ccnd2 transgenic animals are crossed into a Pten null background to accelerate disease onset. Preliminary data shows that T-cell specific Ccnd2 overexpression cooperates with Pten loss to decrease T-ALL latency.
Conclusion
All together, we have used a fast genomic engineering pipeline to develop new and pathophysiological relevant conditional overexpression mouse models for human T-ALL.
Session topic: ALL Biology - Transcriptional dysregulation
Keyword(s): Acute lymphoblastic leukemia, Embryonic stem cells, Mouse model, Transgenic mice
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