The PI3K-mTOR Pathway in Mammals: From Therapeutics to Fundamentals
Cells from multicellular organisms are under the control of extracellular signals to ensure the activation of certain cellular processes only in specific contexts. These extracellular signals act through intracellular signaling components that share some features with unicellular organisms such as yeast, but with additional layers of complexity and regulation. In metazoans, activation of the PI3K-mTOR signaling network is a shared response to engagement of diverse types of extracellular signals. Depending on the cell type and stimulus, activation of this pathway can promote different cellular fates including cell growth, proliferation, survival, migration and differentiation. Dysregulation of these processes is highly implicated in a variety of diseases such as cancer, diabetes, and immune disorders. Indeed, either decreased or hyperactivation of the PI3K-mTOR signaling pathway is observed in many human diseases making this network a crucial target for therapy. In this dissertation, I present a series of preclinical studies showing that indeed, pharmacological or genetic perturbation of the PI3K-mTOR pathway has an impact in the context of cancer and immunity.
Since the identification of phosphoinositide 3-kinase (PI3K) enzyme activity in transformed cells nearly three decades ago, there has been significant advancement in the field leading to the development of a variety of novel chemical tools for therapeutic intervention. In chapter 2, I focus on the published promising finding that cancer cells with mutations in one of the most frequently mutated PI3K isoforms, p110α (encoded by PIK3CA), can be selectively targeted with a novel p110α-selective chemical inhibitor while preserving adaptive immune function. As it is becoming increasingly clear that an intact adaptive immune system is critical for tumor regression, these findings raise confidence that selective p110α inhibitors in cancer therapy will not be as immunosuppressive as global PI3K inhibitors currently in clinical trials.
Chapter 3 focuses on the surprising immunomodulatory effects of targeting the mammalian target of rapamycin (mTOR) downstream of PI3K with second-generation ATP-competitive mTOR kinase inhibitors (TOR-KIs). Unlike the long-known immunosuppressant rapamycin that targets mTOR through an allosteric mechanism, we observed immunoenhancing effects of TOR-KIs specifically in the context of B cell antibody class-switch recombination (CSR). I will present published work where I genetically validated that the mechanism of action of TOR-KIs on CSR is through inhibition of the mTOR complex 2 (mTORC2) signaling axis. These findings have strong implications for utilizing currently available inhibitors of the PI3K-mTOR pathway beyond cancer therapy and in modulating immunity.
Chapter 4 returns to the evolutionarily conserved fundamental role of the PI3K-mTOR pathway, which is to regulate cellular growth (mass increase). A working concept has been proposed and experimentally supported in that cells from multicellular organisms can uncouple cell growth from other cellular processes such as cell proliferation through engagement of distinct mTORC1 effectors. However, given the unique characteristics of primary lymphocytes in that they require a very long phase of growth prior to rapid divisions to produce a large number of identical cells, I hypothesized that a common intracellular effector may coordinate both cell growth and proliferation in this cell type. This work has led to the identification of the eukaryotic translation initiation factor 4E-binding proteins (4E-BPs) downstream of mTORC1 as the critical effector in coordinating the two processes during lymphocyte activation. These results are surprising as the 4E-BPs specifically regulate only cell proliferation in other mammalian cell types. The findings highlight the amazing specificity of this pathway in coordinating cell growth to cell proliferation in a cell-type specific manner.
Despite the fundamental aspect of this research, this work has provided some major advances in the clinical context as well. The FDA-approved immunosuppressant rapamycin has long been known to have selective potency against lymphocytes although its target mTOR is ubiquitously expressed. In work presented in Chapter 4, I show that rapamycin acts upon the 4E-BPs specifically in primary lymphocytes but not other cell types, providing an explanation for its selective effects on these cells. Implications of this work beyond immunity will be discussed in Chapter 5.
In terms of basic cell biology, a fascinating aspect of this research is that during metazoan evolution, distinct cell types have adopted specialized mechanisms to utilize the PI3K-mTOR pathway for fundamental cellular processes such as growth and proliferation, in order to serve each of their roles in a multicellular system. These fundamental studies are critical as loss of proper PI3K-mTOR regulation in each cell type can ultimately lead to diseases such as cancer where cells have lost the properties to adapt to multicellularity.
Lastly, I believe that my findings support and provide more evidence towards a paradigm-shifting concept that protein synthesis can no longer be regarded as a housekeeping function that automatically pumps out proteins at the end of gene regulation. Rather, the findings strongly argue that protein synthesis itself can be regulated and perhaps is the most important step of gene regulation given that proteins are the most important effectors that mediate cellular functions. This paradigm will be discussed and extended in the context of understanding adaptive immunity. Recent progress in understanding lymphocyte activation has mainly focused on concepts of transcriptional regulation and metabolic reprogramming. Protein synthesis in lymphocyte activation has been studied vigorously in the late 70s. The finding that cap-dependent translation is a critical step in lymphocyte activation not only rejuvenates the idea that regulation of protein synthesis plays a critical role in immunity and suggests the possibility of regulation of select transcripts that may have therapeutic potential.