Blocking the transfer of damaged mitochondria from cancer cells to RGS5+ MYL9+ CAFs reduces persistent tolerance to TKIs in EGFR-mutant lung cancer

Eliminating drug-tolerant persister (DTP) cells remain a significant challenge in overcoming resistance to tyrosine kinase inhibitors (TKIs) in EGFR-mutant lung adenocarcinoma. Utilizing single-cell RNA sequencing, we identify a novel RGS5+MYL9+ cancer associated fibroblasts (CAFs) subset. This subset is linked to TKI resistance and correlates with poorer prognosis. The RGS5+MYL9+ CAFs are spatially positioned close to DTP cells, a proximity facilitated by CCL11 recruitment from DTP cells. TKIs induced mitochondria ROS activates Rho GTPase 1 (Miro1) and RhoA of tumor cells, subsequently promoting the formation of tunneling nanotube connections with neighboring RGS5+MYL9+ CAFs. These CAFs play a protective role for DTP cells, aiding in the transfer of damaged mitochondria from tumor cells through these nanotubes. By employing the Rho kinase inhibitor Fasudil to block this transfer of damaged mitochondria, we observed a significant reduction in persistent tolerance to Osimertinib in a xenograft mouse model. Our study highlights the critical role of RGS5+MYL9+ CAFs in mediating resistance to EGFR-TKIs and suggests potential therapeutic strategies for overcoming this challenge. Overall design: Single-cell RNA sequencing was performed on tumor tissues from patients with EGFR-mutant lung adenocarcinoma, and bulk RNA sequencing was conducted on parental 827 cells, TKI-induced DTP 827 cells, and DTP 827 cells co-cultured with distinct CAF populations.