# Oncogenic Activity and Sorafenib Sensitivity of ARAF p.S214C Mutation in Lung Cancer

## TL;DR

This study shows that the ARAF p.S214C mutation in lung cancer cells is oncogenic and highly responsive to sorafenib, suggesting it could be used to guide personalized treatment.

## Contribution

The study identifies ARAF p.S214C as a novel biomarker for sorafenib sensitivity in lung cancer.

## Key findings

- ARAF p.S214C-engineered cells showed enhanced oncogenic potential and activated Raf-MEK-ERK signaling.
- These cells exhibited heightened sensitivity to sorafenib in vitro and in vivo.
- Sorafenib sensitivity was linked to inhibition of cell cycle and DNA replication in ARAF p.S214C mutants.

## Abstract

Lung cancer is often associated with aberrated signaling pathways like the RAF pathway. While most studies have focused on BRAF and CRAF mutations, the role of ARAF is less understood. This study shows that ARAF p.S214C mutation is a rare but important mutation that exhibits oncogenic properties in lung cancer models. ARAF p.S214C-mutated lung cancer cells demonstrate remarkable sorafenib sensitivity in vitro and in vivo, which aligns with a previous clinical report of a lung cancer patient carrying this mutation being an exceptional responder to sorafenib. These findings provide strong evidence that ARAF p.S214C mutation may serve as a novel biomarker for predicting sorafenib efficacy in lung cancer. Investigating the gene–drug sensitivity pairs in clinically exceptional responders in non-precision-based clinical trials could inform personalized cancer treatment strategies, leading to more effective and faster approval of targeted therapies based on specific tumor mutations.

Background/Objectives: RAF pathway aberrations are one of the hallmarks of lung cancer. Sorafenib is a multi-kinase inhibitor targeting the RAF pathway and is FDA-approved for several cancers, yet its efficacy in lung cancer is controversial. Previous clinical research showed that a ARAF p.S214C mutation exhibited exceptional responsiveness to sorafenib in lung adenocarcinoma. Methods: Considering this promising clinical potential, the oncogenic potential and sorafenib response of the ARAF p.S214C mutation were investigated using lung cancer models. ARAF p.S214C mutant, ARAF wild-type (WT), and EGFP control genes were ectopically expressed in lung adenocarcinoma cell lines retroviral transduction. In vitro and in vivo sorafenib sensitivity studies were performed, followed by transcriptomics and proteomics analyses. Results: Compared to the ARAF-WT and EGFP-engineered cells, the ARAF p.S214C-engineered cells activated Raf-MEK-ERK signaling and exhibited enhanced oncogenic potential in terms of in vitro cell proliferation, colony and spheroid formation, migration, and invasion abilities, as well as in vivo tumorigenicity. The ARAF p.S214C-engineered cells also displayed heightened sensitivity to sorafenib in vitro and in vivo. RNA sequencing and reverse-phase protein array analyses demonstrated elevated expression of genes and proteins associated with tumor aggressiveness in the ARAF p.S214C mutants, and its sorafenib sensitivity was likely moderated through inhibition of the cell cycle and DNA replication. The ERK and PI3K signaling pathways were also significantly deregulated in the ARAF p.S214C mutants regardless of sorafenib treatment. Conclusions: This study demonstrates the oncogenicity and sorafenib sensitivity of the ARAF p.S214C mutation in lung cancer cells, which may serve as a biomarker for predicting the sorafenib response in lung cancer patients. Importantly, investigating the gene–drug sensitivity pairs in clinically exceptional responders may guide and accelerate personalized cancer therapies based on specific tumor mutations.