Ascl1 amplicon sequencing of RPM sgAscl1 targeted organoid clones.
We describe a rapid in vivo platform for identifying molecular determinants of neuroendocrine lineage transformation in prostate cancer utilizing engineered mouse prostatic organoids. Transplanted Rb1-/-; Trp53-/-; cMyc+ organoids progress to ASCL1+ neuroendcrine prostate cancer (NEPC), a highly aggressive, androgen receptor signaling inhibitor (ARSI)-resistant tumor. To understand the consequence of Ascl1 loss prior in vivo transdifferentiation, we targeted RPM organoids with sgRNAs against Ascl1. Individual targeted organoid clones were expanded and subjected to targeted Ascl1 amplicon sequencing to identify Ascl1 knockout clones used for transplantation in vivo assays.