Abstract 4436: Mechanistic insights into CXCR7 activation and resistance pathways in EGFR-Mutant NSCLC with mesenchymal phenotype
Authors: Laura C. Gunder, Inés Pulido, Jeffery H. Becker, Frida Pantoja, Jennifer Graeber, Malek G. Massad, Takeshi Shimamura
Journal: Cancer Research (2025)

EGFR tyrosine kinase inhibitors (TKIs) have significantly improved outcomes for NSCLC patients with EGFR mutations. However, resistance inevitably develops, with the T790M mutation identified in over 50% of resistant tumors, alongside mechanisms like MET amplification, SCLC transformation, and Epithelial Mesenchymal Transition (EMT). Osimertinib, a third-generation EGFR TKI approved for first-line treatment, specifically targets the T790M mutation and mitigates T790M-related resistance. Despite this, non-T790M resistance mechanisms, including mesenchymal transformation, are increasingly prevalent. Our group previously reported that the atypical GPCR CXCR7 is critical for EGFR TKI resistance associated with a mesenchymal phenotype. CXCR7 activates the MAPK and PI3K pathways when mutant EGFR receptors are inhibited, offering alternative survival pathways. Although CXCR7 plays a pivotal role in drug resistance with a mesenchymal phenotype, the precise mechanisms underlying its activation remain unclear. Profiling cytokine-chemokine secretion from gefitinib-resistant HCC4006GeR cells identified elevated levels of CXCL12, MIF, and chitinase-3 like-protein-1 (CHI3L1). Exposure of HEK293 cells ectopically overexpressing CXCR7 to canonical CXCR7 ligands, CXCL12 and MIF, activated the MAPK pathway within 30 minutes. We found that CRISPR depletion of CXCR4 in HEK293 cells abrogated MAPK activation following exposure to CXCL12 and MIF. In contrast, HCC4006-CXCR7 cells lacking CXCR4 required eight hours for CXCL12 and MIF to activate MAPK in the presence of osimertinib. Notably, exposure of HCC4006-CXCR7 cells to CXCL12 and MIF in the absence of EGFR TKI failed to augment the MAPK signal. We discovered that CXCR7 co-immunoprecipitated with constitutively active mutant EGFR in NSCLC cells, and this association ceased upon EGFR inhibition. Thus, dissociation of CXCR7 from mutant EGFR appears to be a prerequisite for MAPK activation. Additionally, we identified CH3L1 as a previously unrecognized chemokine capable of activating MAPK in HCC4006-CXCR7 more effectively than CXCL12 or MIF. CXCR7 activation increased the expression of TWIST, ZEB1, and ZEB2, essential for maintaining the mesenchymal phenotype. Depletion of CXCR7 in HCC4006Ge-R cells with a mesenchymal phenotype led to the attenuation of these transcription factors, a mesenchymal-to-epithelial transition, and restoration of EGFR TKI sensitivity. Collectively, these findings reveal a unique ligand-induced mechanism of CXCR7 activation and downstream MAPK pathway modulation by putative ligands. This complex CXCR7 activation mechanism in EGFR-mutant NSCLC suggests that preventing receptor-ligand binding may not be a viable strategy for overcoming EGFR TKI resistance. Citation Format: Laura C. Gunder, Inés Pulido, Jeffery H. Becker, Frida Pantoja, Jennifer Graeber, Malek G. Massad, Takeshi Shimamura. Mechanistic insights into CXCR7 activation and resistance pathways in EGFR-Mutant NSCLC with mesenchymal phenotype [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2025; Part 1 (Regular Abstracts); 2025 Apr 25-30; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2025;85(8_Suppl_1):Abstract nr 4436.