PROJECT SUMMARY The proposed Cancer Systems Therapeutics (CaST) Center focuses on understanding and targeting molecularly distinct yet co-existing cell states and transitions that establish and maintain tumors and tumor microenvironments at a systems level. These goals demand analytical methods with single-cell resolution. The Advanced Single Cell Technology Innovation Core (ASCeTIC) will provide state-of-the-art technology for single-cell genomic analysis, multiplexed single-cell molecular perturbation, and in situ validation to CaST investigators. We will offer a suite of tools including multi-omic methods for joint single-cell RNA/ATAC-seq, single-cell imaging/RNA-seq (SCOPE-seq), and single-cell CRISPRi/RNA-seq (CROP-seq). Recognizing the importance of spatial profiling for both in situ validation and studies of cell-cell interactions in the tumor microenvironment, we will also offer immunohistochemical analysis, RNA-FISH, and cyclic immunofluorescence (cycIF), which can simultaneously quantify tens of protein epitopes in individual cells in a single tissue section.