Elucidating gastric cancer metastatic determinants through single-cell CRISPR screen (ELMET-scCRISPR)

• Applicants: David Koppstein, Ph.D.; Jovan Mircetic, Ph.D.; Professor Daniel Stange, Ph.D.
• Subject Area: Gastroenterology ; General Genetics and Functional Genome Biology
• Term: since 2024
• Project identifier: Deutsche Forschungsgemeinschaft (DFG) - Project number 546973095

Metastasis is the primary cause of cancer-related deaths, thus understanding its determinants is crucial. However, no specific genetic changes have been identified that drive metastasis. The concept of phenotypic plasticity is being increasingly used to explain how cancer cells adapt to different requirements during a metastatic cascade and develop treatment resistance. This plasticity is thought to be achieved through transcriptional programs controlled on the level of chromatin landscape and epigenetics. Gastric cancer is the third leading cause of cancer deaths worldwide, but the lack of sophisticated animal models has hindered research on tumor initiation and metastasis. New stomach-specific cancer mouse models, developed by the applicants, recapitulate human molecular subtypes, providing a new opportunity for metastasis research. Intriguingly, while some mouse models readily metastasized, the others formed tumors without apparent metastases. Thus, the models represent an ideal platform to probe for causative agents of metastatic capacity. CRISPR screens allow the assigning of quantifiable phenotypes to a range of genetic perturbations. Recent technical developments have integrated CRISPR technology with single-cell sequencing, with two important advancements. First, barcodes have been used to follow single cells and their progeny during metastasis, revealing the evolution of metastatic capacities. Second, new guide RNA (gRNA) designs allow simultaneous capturing of single-cell transcriptomes and gRNAs. However, to date, no screens have achieved both of these features together. Here, we made critical design adjustments to make such an analysis possible. By performing in vivo single-cell CRISPR screen with clonal tracking, the goals of the project are threefold: 1.) To investigate epigenetic cell-intrinsic factors that induce metastasis and identify transcriptional programs under the regulation of those epigenetic modulators. 2.) To directly compare transcriptomes of metastatic cells and their clonal progenitors in the primary tumor. Such comparison allows us to identify transcriptional programs induced by specific tumor microenvironments, assessing the role of cell-extrinsic factors. 3.) To compare single-cell transcriptomes of intrametastatic and intermetastatic cell populations to determine if and to what degree metastatic transcriptomes converge. Together, the experiments will identify critical pathways that guide metastatic competence, some of which may be druggable, thus providing novel treatment opportunities.

• DFG Programme: Research Grants