LKB1-mutated NSCLCs represent a highly aggressive subtype lacking targeted therapeutic options. Previous preclinical studies demonstrated that these tumors are particularly sensitive to ERK inhibitors (ERKi). However, co-occurring alterations in the PI3K/Akt/mTOR pathway or other signaling networks may reduce ERKi efficacy. The project aimed to identify the most effective ERKi-based drug combinations for LKB1-deficient NSCLCs using advanced preclinical models, including patient-derived xenografts (PDXs) and organoids (PDXOs).

PDXs and PDXOs were generated from fresh tumor specimens and characterized histologically and molecularly. Their growth patterns and genetic profiles were analyzed by STR and Whole exome sequencing (WES) to confirm fidelity to the original tumor. A comprehensive drug screening of FDA-approved compounds was performed to identify candidates that enhance ERKi sensitivity. Key molecular mechanisms, were investigated by western blot analyses and CRISPR/Cas9-mediated gene deletion to understand ERKi synthetic lethality in LKB1-mutated models. Drug efficacy was validated in 2D and 3D cultures by carrying out distinct cell viability and cell death assays, with selected combinations tested in vivo.

From the drug-library screening, the ALK, NTRK, ROS1 inhibitor Entrectinib emerged as the best promising ERKi partner, showing the most significant impact in LKB1-deficient cells. This combination induced apoptosis, and demonstrated efficacy both in 2D and 3D models. To further clarify the specific mechanisms involved in Entrectinib activity, a pharmacological approach was applied by using Entrectinib analogues targeting only one or two receptors (ALK, NTRK, ROS1) and combined them with the ERK inhibitors. This approach revealed that inhibiting ALK is necessary, but probably not sufficient, to sensitize LKB1-mutated cells to ERKi. In vivo, the ERKi-Entrectinib combination reduced tumor growth, and improved survival rates, supporting its clinical potential. Further experiments are ongoing on patients-derived models to corroborate these results.

These findings highlight the therapeutic potential of ERKi-based combinations for LKB1-mutated NSCLCs, particularly with Entrectinib. The study provides a strong rationale for further preclinical and clinical evaluations to refine combination strategies, enhance efficacy, and minimize toxicity. This offers a new avenue for treating LKB1-deficient NSCLCs, a subtype with limited targeted therapies.