Sanford Burnham Prebys Medical Discovery Institute 6 articles published in JoVE Developmental Biology Generation of 3D Whole Lung Organoids from Induced Pluripotent Stem Cells for Modeling Lung Developmental Biology and Disease Sandra L. Leibel1,2,3, Rachael N. McVicar2,3, Alicia M. Winquist2,3, Evan Y. Snyder1,2,3 1Department of Pediatrics, University of California, San Diego School of Medicine, 2Sanford Consortium for Regenerative Medicine, 3Sanford Burnham Prebys Medical Discovery Institute The article describes step wise directed differentiation of induced pluripotent stem cells to three-dimensional whole lung organoids containing both proximal and distal epithelial lung cells along with mesenchyme. Cancer Research Assessing Cellular Target Engagement by SHP2 (PTPN11) Phosphatase Inhibitors Lester J. Lambert1, Celeste Romero1, Douglas J. Sheffler1, Maria Celeridad1, Nicholas D. P. Cosford1, Lutz Tautz1 1Cancer Metabolism & Signaling Networks Program, NCI-Designated Cancer Center, Sanford Burnham Prebys Medical Discovery Institute The ability to assess target engagement by candidate inhibitors in intact cells is crucial for drug discovery. This protocol describes a 384 well format cellular thermal shift assay that reliably detects cellular target engagement of inhibitors targeting either wild-type SHP2 or its oncogenic variants. Genetics Tumorsphere Derivation and Treatment from Primary Tumor Cells Isolated from Mouse Rhabdomyosarcomas Francesca Boscolo Sesillo1,2,3, Alessandra Sacco2 1Graduate School of Biomedical Sciences, Sanford Burnham Prebys Medical Discovery Institute, 2Development, Aging and Regeneration Program, Sanford Burnham Prebys Medical Discovery Institute, 3Department of Obstetrics, Gynecology, and Reproductive Sciences, Division of Female Pelvic Medicine and Reconstructive Surgery, University California San Diego This protocol describes a reproducible method for isolation of mouse rhabdomyosarcoma primary cells, tumorsphere formation and treatment, and allograft transplantation starting from tumorspheres cultures. Cancer Research Investigation of Genetic Dependencies Using CRISPR-Cas9-based Competition Assays Anagha Deshpande1, Bo Rui Chen1, Luyi Zhao1, Kelsey Saddoris1, Mayuri Kerr1, Nan Zhu2, Prashant Mali3, Aniruddha J. Deshpande1 1Tumor Initiation and Maintenance Program, Sanford Burnham Prebys Medical Discovery Institute, 2Blood Research Institute, Blood Center of Wisconsin, 3Department of Bioengineering, University of California, San Diego This manuscript describes a Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR) CRISPR-Cas9-based method for simple and expeditious investigation of the role of multiple candidate genes in Acute Myeloid Leukemia (AML) cell proliferation in parallel. This technique is scalable and can be applied in other cancer cell lines as well. Developmental Biology Generation of First Heart Field-like Cardiac Progenitors and Ventricular-like Cardiomyocytes from Human Pluripotent Stem Cells Michael S. Yu*1,2, Sean Spiering*1, Alexandre R. Colas1 1Sanford Burnham Prebys Medical Discovery Institute, 2Department of Bioengineering, University of California at San Diego Here we describe a scalable method, using a simple combination of Activin A and lentivirus-mediated Id1-overexpression, to generate first heart field-like cardiac progenitors and ventricular-like cardiomyocytes from human pluripotent stem cells. Genetics High Fat Diet Feeding and High Throughput Triacylglyceride Assay in Drosophila Melanogaster Soda Balla Diop1, Ryan T. Birse1, Rolf Bodmer1 1Development, Aging and Regeneration Program, Sanford Burnham Prebys Medical Discovery Institute This is a high fat diet feeding protocol to induce obesity in Drosophila, a model for understanding fundamental molecular mechanisms implicated in lipotoxicity. It also provides a high throughput triacylglyceride assay for measuring fat accumulation in Drosophila and potentially other (insect) models under various dietary, environmental, genetic or physiological conditions.