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Abstract :
[en] ABSTRACT
[en] Introduction
[en] PI3K/mTORC1- and Ras/MAPK-signalling pathways are aberrantly regulated in most cancers. Specific resistances to drugs targeting these pathways can emerge during tumour evolution. Preexisting, innate resistance mechanisms, such as stemming from feedback-loops, should ideally be known already during drug-target nomination. However, as the example of B-Raf-inhibitors that paradoxically activate MAPK-signalling has shown, feedback mechanisms may only become apparent at very late drug development stages.
[en] Material and methods
[en] HEK cells expressing FRET-pairs of Ras proteins were used to study specific effects on Ras isoforms (nanoclustering). Breast cancer cells were grown in 2D for Western blotting of Ras and mTORC1-pathway proteins, or as spheres to analyse stemness traits.
[en] Results and discussions
[en] Here, we describe two broad feedback loops from the mTor-pathway back to the nanoscopic membrane signalling complexes (nanocluster) of H-ras and K-ras4B (hereafter K-ras). Increased nanoclustering typically correlates with increased Ras output. The first, upstream loop leads to an inadvertent rapalog induced promotion of stemness traits and tumorigenicity in Ras transformed cells. This is due to an induction of the H-ras nanocluster scaffold galectin-1, when FKBP12 levels are low. Surprisingly we find that rapalogs do not only bind to but induce a loss of FKBP12 protein. Thus, rapalog treatment induces galectin-1, which stimulates H-ras signal output and stemness traits. Secondly, modulation of the activity in the mTORC1 pathway downstream of the major lipidome regulator SREBP1, oppositely regulates H-ras and K-ras nanoclustering. Thus, ablation of SREBP1 increases K-ras, but decreases H-ras nanoclustering and signal output. We show that altered levels of phosphatidic acid downstream of SREBP1 are sufficient for the opposite regulation of the two Ras isoforms.
[en] Conclusion
[en] The described feedback loops may only become apparent in certain tumour settings. For example, tumour promotion during rapalog-treatment may only be relevant in H-ras mutant cancers, which make up a small portion of human cancers. In those cases, rapalog efficacy may be improved in combination with novel anti-galectin-1 drugs. Targeting the mTORC1 pathway downstream of SREBP1, may have opposite effects in H-ras and K-ras mutant cancers. Thus, care may have to be taken when targeting the mTORC1-pathway in a mutant Ras setting.
