Identifying cellular drivers of DICER1 predisposition in pediatric rhabdomyosarcoma

MicroRNAs are small, single-stranded noncoding RNA molecules that regulate gene expression. Dysregulated gene expression can lead to cancer, but the exact mechanisms and relationships are still not fully understood. One factor involved in producing functional microRNAs is DICER1. When DICER1 is mutated, microRNA-mediated regulation can be disrupted, which may play a role in cancer development.

Individuals with DICER1 cancer predisposition have lost a germline copy of the DICER1 gene. Cancer develops when affected individuals acquire a mutation in the remaining copy of DICER1, disrupting microRNA function, but it was unknown which specific genes or cells were responsible for this. Published in Developmental Cell, corresponding author Mark Hatley, MD, PhD, Division of Molecular Oncology director and Department of Oncology associate member, and co-first author Jason Hanna, PhD, former St. Jude postdoctoral fellow, examined DICER1’s impact using mouse models of rhabdomyosarcoma.

“When we specifically mutated DICER1 in cancer cells, there was no difference in tumor growth; however, when we mutated DICER1 in all cells, those mutations did promote cancer growth,” explained Hatley. “We discovered that the tumor promotion activity is not driven by the cancer cells, but rather by cells in the tumor microenvironment.”

When the scientists compared tumors developing in the context of DICER1 mutations to those without the mutations, they identified a common type of white blood cell called neutrophils as key contributors.

Co-first author Randy Larsen, PhD, St. Jude Graduate School of Biomedical Sciences, performed single-cell RNA sequencing of the tumor stroma, confirming the enrichment of neutrophils and enabling a precise identification of the cells driving cancer progression.

Larsen identified a diverse population ranging from immature to fully differentiated neutrophil cells. He then showed that removing neutrophils in mice with germline DICER1 mutations eliminated the mutations’ cancer-promoting effects.

Larsen and Hatley sought to understand how neutrophils promote tumor growth. “We went back to the sequencing data and saw an increased expression of genes for NETosis,” Hatley said. “NETosis is a neutrophil process in which neutrophil extracellular traps (NETs) made of DNA and enzymes are released into the tumor microenvironment.

“Randy also showed that one of the molecules released into the tumor microenvironment during NETosis is a peptide called cathelicidin antimicrobial peptide, or CAMP,” Hatley continued. “CAMP activates growth-promoting signaling pathways in rhabdomyosarcoma cells.”

The researchers determined that NET formation releases and creates proliferative signals near the tumor, explaining how DICER1 mutations promote rhabdomyosarcoma. They then used disulfiram, a drug that blocks NET formation and prevents neutrophils from rupturing and performing NETosis. When given to their mouse models, disulfiram blocked the tumor-promoting effect of DICER1 germline mutations.

To confirm the findings in humans, Larsen analyzed patient tumor data from multiple registries and databases. He found that tumors with DICER1 mutations were enriched for neutrophil-related genes. The findings provide a mechanistic link between DICER1 mutations and rhabdomyosarcoma that suggests a new therapeutic opportunity.