Abstract
Cytotoxic chemotherapy agents (e.g., cisplatin) are the first-line drugs to treat non-small cell lung cancer (NSCLC) but NSCLC develops resistance to the agent, limiting therapeutic efficacy. Despite many approaches to identifying the underlying mechanism for cisplatin resistance, there remains a lack of effective targets in the population that resist cisplatin treatment. In this study, we sought to investigate the role of cytoplasmic RAP1, a previously identified positive regulator of NF-κB signaling, in the development of cisplatin resistance in NSCLC cells. We found that the expression of cytoplasmic RAP1 was significantly higher in high-grade NSCLC tissues than in low-grade NSCLC; compared with a normal pulmonary epithelial cell line, the A549 NSCLC cells exhibited more cytoplasmic RAP1 expression as well as increased NF-κB activity; cisplatin treatment resulted in a further increase of cytoplasmic RAP1 in A549 cells; overexpression of RAP1 desensitized the A549 cells to cisplatin, and conversely, RAP1 depletion in the NSCLC cells reduced their proliferation and increased their sensitivity to cisplatin, indicating that RAP1 is required for cell growth and has a key mediating role in the development of cisplatin resistance in NSCLC cells. The RAP1-mediated cisplatin resistance was associated with the activation of NF-κB signaling and the upregulation of the antiapoptosis factor BCL-2. Intriguingly, in the small portion of RAP1-depleted cells that survived cisplatin treatment, no induction of NF-κB activity and BCL-2 expression was observed. Furthermore, in established cisplatin-resistant A549 cells, RAP1 depletion caused BCL2 depletion, caspase activation and dramatic lethality to the cells. Hence, our results demonstrate that the cytoplasmic RAP1–NF-κB–BCL2 axis represents a key pathway to cisplatin resistance in NSCLC cells, identifying RAP1 as a marker and a potential therapeutic target for cisplatin resistance of NSCLC.
Highlights
NSCLC, among which are the apoptosis-induction chemotherapy agents such as cisplatin (CP).[1,2] But considerably high rate of relapse occurs following CP treatment despite the approaches aiming to overcome resistance.[3,4,5] It is necessary to develop strategies that act on the CP-resistant subpopulation of NSCLC and reverse the resistance in these tumor cells
To test the hypothesis that cytoplasmic RAP1 is a biomarker for higher-grade NSCLC, we immunostained for RAP1 and quantified its cytoplasmic and nuclear expression in 93 lung adenocarcinoma and 75 lung squamous cell carcinoma tissues
These analyses identify cytoplasmic RAP1 as an indicator of high-grade NSCLC, suggesting that it may have a critical role in cancer progression
Summary
NSCLC, among which are the apoptosis-induction chemotherapy agents such as cisplatin (CP).[1,2] But considerably high rate of relapse occurs following CP treatment despite the approaches aiming to overcome resistance.[3,4,5] It is necessary to develop strategies that act on the CP-resistant subpopulation of NSCLC and reverse the resistance in these tumor cells. RAP1 was identified to bind with extratelomeric sites through a consensus motif, helping with gene transcriptional regulation.[15,16] cytoplasmic RAP1 has been revealed for its essential role in modulating NF-κB signaling.[17] the RAP1–NF-κB axis was demonstrated to promote invasion of breast cancer cells.[17] Given the broad activation of NF-κB signaling in human cancer, one would infer that RAP1 might contribute to the progression of other types of tumors via a similar mechanism. Our results demonstrate an extratelomeric function for RAP1 in tumor progression and suggest that RAP1 is a potential therapeutic target for CP-resistant NSCLCs
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