MADR: Rapid generation of somatic mosaics with locus-specific, stably integrated transgenic elements for generation of "personalized" mouse models and human organoid tumor models.

In situ transgenesis methods such as virus and electroporation can create somatic transgenic mice quickly, but they lack the exquisite control over copy number, zygosity, and locus specificity. We have recently established mosaic analysis by dual recombinase-mediated cassette exchange (MADR), which permits stable labeling of mutant cells expressing transgenic elements from precisely defined chromosomal loci. MADR provides a toolkit of elements for combinatorial labeling, inducible/reversible transgene manipulation, VCre recombinase expression, and genetic manipulation of human cells. Further, we have demonstrated the versatility of MADR by creating glioma models with mixed, reporter-identified zygosity or with “personalized” driver mutations from pediatric glioma. For example, introducing H3f3a mutation variants with MADR regulates the spatiotemporal profile of glioma, and single-cell RNA and ATAC sequencing analysis demonstrates a recapitulation of developmental hierarchy seen in K27M mutant human glioma. Moreover, we have generated novel models of supratentorial ependymoma using patient-derived oncofusion transgenes. These models display a high degree of fidelity, and we now compare these models on a single-cell level with our previous models and human tumor cell transcriptomes. In addition, we now demonstrate the ability to generalize the MADR technology to other non-CNS tissues using local plasmid delivery. Finally, we have engineered human cells to allow for MADR transgenesis and somatic transgenic organoids. These combined approaches will enable researchers to discover disease mechanisms and test therapeutics in more physiologically relevant cancer models. MADR is extensible to thousands of existing mouse lines and can be adapted to human cells, providing a flexible platform to democratize the generation of somatic transgenic disease models. This abstract is also being presented as Poster B46. Citation Format: Gi Bum Kim, David Rincon Fernandez Pacheco, David Saxon, Amy Yang, Sara Sabet, Marina Dutra-Clarke, Rachelle Levy, Ashley Watkins, Hannah Park, Aslam Abbasi Akhtar, Paul W. Linesch, Naomi Kobritz, Swasty S. Chandra, Katie Grausam, Alberto Ayala- Sarmiento, Jessica Molina, Kristyna Sedivakova, Daniel S. Gareau, Mariella G. Filbin, Serguei Bannykh, Jie Tang, Mario Suva, Bin Chen, Moise Danielpour, Joshua J. Breunig. MADR: Rapid generation of somatic mosaics with locus-specific, stably integrated transgenic elements for generation of “personalized” mouse models and human organoid tumor models [abstract]. In: Proceedings of the AACR Special Conference on the Evolving Landscape of Cancer Modeling; 2020 Mar 2-5; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2020;80(11 Suppl):Abstract nr PR11.