TP53 mutant cell lines selected for resistance to MDM2 inhibitors retain growth inhibition by MAPK pathway inhibitors but a reduced apoptotic response

Chiao-En Wu,Tsin Shue Koay,Penny Lovat,Yi-Hsuan Ho,John Lunec

doi:10.1186/s12935-019-0768-3

Chiao-En Wu, Tsin Shue Koay + Show 3 more

Open Access

https://doi.org/10.1186/s12935-019-0768-3

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Abstract

BackgroundEmergence of resistance to molecular targeted therapy constitutes a limitation to clinical benefits in cancer treatment. Cross-resistance commonly happens with chemotherapeutic agents but might not with targeted agents.MethodsIn the current study, TP53 wild-type cell lines with druggable MAPK pathway mutations [BRAFV600E (WM35) or NRASQ61K (SJSA-1)] were compared with their TP53 mutant sublines (WM35-R, SN40R2) derived by selection for resistance to MDM2/p53 binding antagonists.ResultsThe continued presence of the druggable MAPK pathway targets in the TP53 mutant (TP53MUT) WM35-R and SN40R2 cells was confirmed. Trametinib and vemurafenib were tested on the paired WM35/WM35-R and SJSA-1/SN40R2 cells and similar growth inhibitory effects on the paired cell lines was observed. However, apoptotic responses to trametinib and vemurafenib were greater in WM35 than WM35-R, evidenced by FACS analysis and caspase 3/7 activity, indicating that these MAPK inhibitors acted on the cells partially through p53-regulated pathways. SiRNA mediated p53 knockdown in WM35 replicated the same pattern of response to trametinib and vemurafenib as seen in WM35-R, confirming that p53 plays a role in trametinib and vemurafenib induced apoptosis. In contrast, these differences in apoptotic response between WM35 and WM35-R were not seen with the SJSA-1/SN40R2 cell line pair. This is likely due to p53 suppression by overexpressed MDM2 in SJSA-1.ConclusionThe TP53MUT cells selected by resistance to MDM2 inhibitors nevertheless retained growth inhibitory but not apoptotic response to MAPK pathway inhibitors.

Highlights

Emergence of resistance to molecular targeted therapy constitutes a limitation to clinical benefits in cancer treatment
Selection for resistant cells SN40R2 and WM35-R cell lines with resistance to MDM2/p53 binding antagonists were generated by continuously exposing SJSA-1 osteosarcoma cells [20] and WM35 cutaneous melanoma cells to either Nutlin-3 or RG7388 respectively
WM35‐R cells are resistant to other MDM2 inhibitors WM35-R cells were selected from a parental WM35 culture by exposure to a final concentration of 5 μM RG7388

Summary

Introduction

Emergence of resistance to molecular targeted therapy constitutes a limitation to clinical benefits in cancer treatment. SiRNA mediated p53 knockdown in WM35 replicated the same pattern of response to trametinib and vemurafenib as seen in WM35-R, confirming that p53 plays a role in trametinib and vemurafenib induced apoptosis These differences in apoptotic response between WM35 and WM35-R were not seen with the SJSA-1/SN40R2 cell line pair. RAS and RAF oncogenes are frequently mutated in human cancer leading to a constitutively activated MAPK pathway which is critical for oncogenesis, tumour proliferation and survival [1, 2]. These genetic alterations provide important targets druggable by low molecular weight compounds which have been evaluated in clinical. RG7388 [15] and HDM201 [16] are new generations of MDM2 inhibitors which are more potent and specific than Nutlin-3 and clinical trials are ongoing to investigate their efficacy in clinical settings

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