Posted: Thursday, May 19, 2022
Brian D. Brown, PhD, of the Precision Immunology Institute, Icahn School of Medicine at Mount Sinai, New York, and colleagues developed an approach for spatial functional genomics called Perturb-map. This creation is a protein barcode system that uses triplet combinations of linear epitopes to create a higher order set of barcodes that are able to mark cells expressing CRISPR guide RNAs. Published in the journal Cell, these findings show that this technique has the potential to determine how genes influence the tumor microenvironment in lung cancer.
“If specific gene mutations are keeping T cells out of a subclonal region, this can serve as a pocket of resistance to immunotherapies like [pembrolizumab],” stated Dr. Brown in an institutional press release. “The local and distal effects of many other genes on tumor composition are still not known, but the Perturb-map platform will now give scientists a powerful means to tackle the problem.”
Mouse KRAS G12D p53 lung cancer cells and 4T1 breast cancer cells were transduced with a pool of lentiviral vectors encoding 84 or 120 different Pro-Codes to knock out dozens of genes in parallel. How each knockout influenced immune composition, tumor growth, and histopathology was assessed simultaneously. Additionally, Perturb-map was paired with spatial transcriptomics for an impartial analysis of tumors that were edited with CRISPR.
It was discovered that in Tgfbr2 knockout tumors, the tumor microenvironment was changed into a fibromucinous state. In addition, these tumors were excluded in T cells, and this process occurred simultaneously with upregulated TGF-β and TGF-β–mediated fibroblast activation. This was a clear indication that the loss of TGF-β receptors effectively increased its bioavailability, as well as its immunosuppressive effects on the tumor microenvironment.
Disclosure: For full disclosures of the study authors, visit cell.com.