Mechanisms of ERK action in MEK inhibitor-insensitive lung cancers
Introduction
Small cell lung cancer (SCLC) and other neuroendocrine (NE) lung tumors represent 25% of all lung cancers and are known for their aggressive nature and poor prognosis. SCLC are considered recalcitrant tumors as the 5-year survival is lower than 7%. Besides, the few available treatments display low efficacy and relapse eventually occurs in the majority of the patients, due to acquired resistance. The lack of driver oncogenes involved in the regulation of kinase signaling makes these tumors hard to treat. Interestingly, unlike other tumor types, including non-small cell lung carcinoma (NSCLC), tumorigenic mutations in the RAS-ERK pathway are uncommon in SCLC. Previous studies have shown that ERK activation interferes with SCLC growth, but the exact mechanism remains unknown. The transcription factor ASCL1 plays a critical role in the survival of most SCLC and NE lung tumors by regulating DUSP6 transcription, leading to the suppression of ERK activity. Conversely, ERK activation affects cell cycle progression and survival by downregulating ASCL1.
Methods
Individual deletion of each ERK isoform and DUSP6 was performed by CRISPR Cas9 technologies. Viability and proliferation assays were used to compare the effects of DUSP6 KO and pharmacological inhibition with the DUSP6 inhibitor BCI in HCC1833 NSCLC-NE cells. ASCL1 expression was assessed upon treatment with several pharmacological inhibitors of different signaling pathways. ERK activation dynamics, histone acetylation and ASCL1 expression were tested by western blot analyses.
Results
BCI, DUSP6 inhibitor, has been used to sustain ERK activation by decreasing ASCL1 expression, but concerns about potential off-target effects of BCI exist. Both pharmacological inhibition of DUSP6 and genetic ablation of DUSP6 reduced cell viability and proliferation in ASCL1-high cells, no toxic BCI effect was detected in cells lacking the target. Inhibitors promoting histone acetylation provoked a negative impact in the expression of ASCL1 as well as ERK activation. Furthermore, complete knockout of ERK1/2 was not achievable in these tumors due to the essential role of some ERK activity for their viability, but individual deletion of each ERK isoform was possible. Notably, ERK2 KO cells underwent an adaptation process that resulted in higher proliferation compared to wild-type (WT) cells, likely attributed to constitutive Akt activation.
Conclusion
The findings suggest that one of the crucial roles of ASCL1 in the survival of SCLC and NE tumors is regulating ERK activity through DUSP6 to protect its transcription. BCI, an inhibitor of DUSP6, can sustain ERK activation by impeding DUSP6 activity in ASCL1-high cells without showing toxicity in cells lacking the DUSP6 target. However, some ERK activity is necessary for tumor viability and complete ERK1/2 knockout is not feasible. Besides, high chromatin acetylation levels correlated with low ASCL1 expression. Further research is needed to understand the implications of ERK activity in NE lung tumors and the potential mechanisms of resistance that may arise upon targeted therapies. A better understanding of SCLC biology and the identification of resistance mechanisms are crucial for developing effective treatment strategies for these challenging tumors.