Abstract B10: Inhibiting eIF4F-mediated adaptive translatome reprogramming is a salvage strategy for targted melanoma therapy

Abstract

Abstract BRAFV600E mutation is observed in ~50% of melanomas. Once melanomas have progressed with acquired resistance to BRAF-targeted therapy, mutational heterogeneity presents a major challenge. While enormous effort has gone into understanding the molecular events in mutational acquired resistance, not much attention has been attributed to what happens during therapy phase when patients still respond to BRAF-targeted treatment. It has been hypothesized that initally drug-tolerant cells without bona fide mutational resistance may survive by cellular adaptations and undergo evolution over time towards acquired genetic mutational resistance. We defined the nature of eIF4F-mediated translatome reprogramming and its role during the early phase of targeted melanoma therapy before acquired mutational resistance developed. To explore the possibility that melanoma cells can develop resistance via drug-tolerant evolution (versus pre-existing mutation), we cultured over 1000 small pools (5000 cells each) of parental A375 cells (BRAFV600E) in the presence of vemurafenib and cobimetinib and monitored the emergence of resistant clones. 99.7% of the wells contained a small number of surviving, drug-tolerant cells after 3 weeks of drug exposure. Strikingly, to further demonstrate adaptive drug-tolerant cells exist as a mechanism of resistance evolution, we established A375 subclones derived from single cells to eliminate possible pre-existing mutational resistant cells. Indeed, in five independent single-cell subclones, no early resistant colonies emerged after 3 weeks of drug treatment. These drug-tolerant cells are transiently resistant to re-exposure of anti-BRAF or anti-MEK treatment, and the cells drifted back to the similar sensitivity compared to parental cells after ~9 days of drug-free culture. These drug-tolerant cells are instead more sensitive to eIF4F inhibition compared to parental cells, indicating that eIF4F plays a central role in reprogramming adaptive translatome of drug-tolerant cells. To study the eIF4F activity at single cell level, we developed a single cell image analysis pipeline by using proximity ligation assay and automatic image analysis. By cell image machine learning, we identified five types of eIF4F localization in a large population of melanoma cells. With the classification model, we classified each single drug-tolerant cell into one of the five localization type. Rather than modifying global translation rate in drug-tolerant cells, we revealed that cell-to-cell heterogeneity of the subcellular localization of eIF4F complex discriminates the parental and drug-tolerant cells. This indicates that differential localized eIF4F might initiate the translation of different subsets of genes in drug-tolerant cells. Particularly, the localization pattern of eIF4F is also dynamic. Consistent with the sensitivity of the drug-tolerant cells, the localization pattern of eIF4F drifted back to the similar distribution of parental cells after ~9 days of drug-free culture. Thus, we delineated a novel level of adaptive regulation of gene expression at translational level during the course of targeted melanoma therapy, which might provide a general mechanism of network rewiring during tumor evolution. This translational controlled network reprogramming is likely to be missed if one looks merely at the genetic alone, we thus stress the need to break the current “glass ceiling” of relying solely on genomic dataset analysis to advance cancer therapies. Note: This abstract was not presented at the conference. Citation Format: Shensi Shen, Caroline Robert, Stephan Vagner. Inhibiting eIF4F-mediated adaptive translatome reprogramming is a salvage strategy for targted melanoma therapy. [abstract]. In: Proceedings of the AACR Special Conference on Translational Control of Cancer: A New Frontier in Cancer Biology and Therapy; 2016 Oct 27-30; San Francisco, CA. Philadelphia (PA): AACR; Cancer Res 2017;77(6 Suppl):Abstract nr B10.