Background: Bronchial carcinoids (BC) are rare neoplasm, still orphan of medical therapy, which arise from neuroendocrine cells. It has been previously demonstrated that the atypical BC human cell line NCI-H720 is sensitive to everolimus (E), an m-TOR inhibitor, in terms of cell viability reduction, with a G0 cell-cycle arrest and a Cyclin D1 protein reduction. On the contrary, the typical human BC cell line NCI-H727 is not sensitive to E, despite the Cyclin D1 reduction. The mechanisms underlying this phenomenon have not been fully clarified, yet.
Aim: Our aim is to further investigate cell cycle mechanisms that underlie resistance to mTOR inhibitors in BC cells, in order to identify new therapeutic approaches.
Materials and methods: Human BC cell line cultures (NCI-H720 and NCI-H727 cells) were investigated by Western blot, cell viability and caspase activation assays before and after a challenge with E.
Results: Protein profiling of the main complexes regulating G0/G1 >> S progression (CDK2/Cyclin E and CDK4/Cyclin D1) and CDK-inhibitors (p27 and phospho p27_SER10) was performed. At basal levels the two BC cell lines showed a different protein profile with the positive regulators of the cell cycle (Cyclin D/CDK4 and Cylin E/CDK2) more expressed in NCI-H727 cells (resistant to E) as compared to NCI-H720 cells (sensitive to E), suggesting that in the Resistant BC cell line p27kip1, despite being over-expressed, has an impaired function. We also found that E reduced Cyclin D/CDK4 protein complex as well and Cyclin E expression, but induced CDK2 expression, possibly explaining the escape of NCI-H727 from the antiproliferative effects of E. On the other hand, no modification of cell cycle control protein profile has been observed in NCI-H720 after treatment with E, indicating that the latter exerts its antiproliferative effects on this cell line by a cyclin-independent mechanism.
Conclusion: Our data indicate that resistance to mTOR inhibitors may be linked to a deranged cell cycle control protein profile. Therefore the characterization of these proteins may represent a putative marker of resistance to E, possibly contributing to a better patients selection for a therapeutic approach with mTOR inhibitors.