Modeling mTOR network dynamics to achieve optimal drug rescue of congenital mTORopathies

PROJECT SUMMARY This R01 proposal focuses on unraveling the complexities of the mechanistic target of rapamycin (mTOR) signaling in neurons. mTOR is central to brain function and is often disrupted in a range of neurological disorders including epilepsy, autism, and neurodegenerative diseases. While much of the existing research has approached mTOR signaling through a linear model, targeting mTOR directly with drugs like Rapamycin, these strategies have yielded limited success in the clinic. We propose a paradigm shift: rather than viewing mTOR signaling as a linear cascade, we will investigate it as a dynamic, context-dependent protein interaction network. Our hypothesis is that understanding and targeting specific dysfunctional nodes within this network— upstream or downstream of mTOR—will lead to more effective therapeutic interventions for mTOR-related disorders. Using our innovative Quantitative Multiplex Co-Immunoprecipitation approach, we will map the entire mTOR protein interaction network (PIN) in neurons, revealing how these interactions reconfigure in response to cellular stimuli, genetic mutations, and pharmacological interventions. Preliminary data from our lab shows that mTOR-modifying drugs, such as PI3K or ERK inhibitors, induce non-linear effects on the network, depending on the specific node targeted. This insight points to a key flaw in the conventional approach of targeting mTOR directly: it fails to account for the complexity of the surrounding network and its role in determining cellular outcomes—particularly in neurons, where we show that network architecture differs substantially from that in proliferating cells. Our project is organized around three aims. Aim 1 will define the structure of the neuronal mTOR PIN under normal conditions, mapping how it responds to synaptic activity and or mTOR activation using tools like multiplex co-IP, optogenetic PI3K activation and electrophysiology. Aim 2 will assess how acute and chronic treatment with PI3K, AKT, mTOR, and ERK inhibitors affect the mTOR network, providing critical insights into drug effects on network dynamics. Aim 3 will use hyperactivating PI3K and PTEN mutations to model how disease-causing mutations distort mTOR signaling and whether these disruptions can be restored to normal function by targeting specific network nodes rather than mTOR itself. By adopting a network-based approach and focusing on protein-protein interactions within mTOR signaling, this proposal offers a new lens for understanding the molecular mechanisms underlying mTORopathies. The multidisciplinary team assembled for this project, with expertise in proteomics, electrophysiology, optogenetics, and behavioral neuroscience, is uniquely equipped to tackle the challenges of decoding these complex signaling networks. We anticipate that this work will lead to the identification of new, more precisely targeted therapeutic strategies for a broad range of neurological disorders linked to mTOR dysfunction.