Individuals in general populations contain two functional copies of Neurofibromatosis type 1 (NF1) gene. However, NF1 patients either inherit or acquire one defective NF1 copy during development. Consequently, every cell in NF1 patients only has one functional copy of the NF1 gene. In general, two mechanisms are responsible for NF1-associated lesions. One mechanism involves a genetic process known as loss of heterozygosity (LOH), in which affected cells lose the functional or normal NF1 copy, leading to a complete loss of NF1 function. This mechanism is involved in benign and malignant tumor formation in NF1 patients. Examples of such NF1-associated tumors include neurofibroma and malignant peripheral nerve sheath tumor (MPNST) in the peripheral nervous system. The second mechanism, termed haploinsufficiency -- loss of only one functional copy of NF1 -- leads to disease manifestations. Many of the non-tumor related symptoms including learning disabilities, bone defects, pain, and blood vessel abnormalities are believed to result from haploinsufficiency. In haploinsufficient lesions, it is thought that one functional copy of NF1 gene is not sufficient to perform normal cellular functions. While it is relatively easy to understand that a complete loss of NF1 function leads to disease manifestations, it may seem counterintuitive for the mechanisms underlying NF1 haploinsufficient lesions. Particularly, clinical studies showed that some NF1 patients can live a relatively normal life and that siblings with the same NF1 mutations can have dramatically different disease manifestations. These observations indicate that upon loss of one functional NF1 copy, some cells, but not others, cannot perform normal functions. In other words, NF1 haploinsufficient lesions are context-dependent, influenced by other factors. Thus, one fundamental question in NF1 is to understand the nature of context-dependent NF1 haploinsufficient lesions. Moreover, one potentially effective therapeutic strategy for NF1-haploinsufficient lesions is to increase NF1 protein availability in the affected cells, as they still have one functional copy. However, the mechanisms that control NF1 protein levels remain largely unknown.

In our preliminary study, we identified the signals and mechanisms that control degradation (or destruction) of NF1 proteins in cultured cells. Moreover, we identified one specific context in which cells with one functional NF1 copy receive higher growth signals triggering a greater level of degradation of NF1 proteins, which further lowers NF1 protein levels leading to haploinsufficient lesions. In this application, we propose the experiments to extend these important findings from cultured cells to mouse models (going from in vitro to in vivo settings). Furthermore, we will use these findings to attempt to increase NF1 protein levels in cells with one functional NF1 copy and to perform preclinical trials to reverse learning disabilities in Nf1 mouse models. We will use both genetic and pharmacological approaches to increase NF1 protein levels in tissues of the Nf1 mouse models to reverse learning disabilities. The pharmacological agent proposed in the application is currently being tested in clinical trials for treating cancers. Thus, the studies proposed in this application have high potential for clinical applications to treat NF1 haploinsufficient lesions.

In summary, from a basic science point of view, our studies will open a new research area to study what signals regulate NF1 protein degradation and how NF1 proteins are degraded. More importantly, these studies are potentially highly clinically relevant, if they provide insights on the nature of context-dependent NF1 haploinsufficient lesions and the design of novel therapies to treat them. Finally, NF1 haploinsufficient lesions contribute to much of the morbidity and mortality associated with NF1 patients. For example, learning disabilities are found in over 50% of NF1 children, which is the most common complication affecting quality of life in these children. Furthermore, these deficits are stable both across childhood and into adulthood. Moreover, bone dysplasia, pain, and cardiovascular diseases also affect a significant number of NF1 patients. In addition to non-tumor related symptoms, recent studies suggest that NF1 haploinsufficiency also contributes to tumor formation such as neurofibroma and optic glioma. Together, our studies have the potential to benefit a large population of NF1 patients.