Project Summary
Neurodegenerative disorders (such as Alzheimer’s, Parkinson’s, Huntington’s diseases, and others) result in
severe neurological, cognitive, and motor deficits. Neuronal nitric oxide synthase (nNOS) inhibition is emerging
as a promising strategy to treat neurodegenerative disorders, but the development of nNOS inhibitors is often
hindered by poor pharmacokinetic (PK) properties. In the past few years, we have developed several classes of
highly potent and selective inhibitors of nNOS, which exhibit improved membrane permeability, elevated
metabolic stability, or promising blood-brain barrier penetration in animals. However, we have yet to identify
compounds that encompass all of these desired properties, and further optimization of these nNOS inhibitors
will be needed. Our current R35 award allows us to identify promising nNOS inhibitors and characterize drug-
like properties in a limited capacity. We have expertise and capability to design and synthesize 40-50 novel
nNOS inhibitors every year. However, only 5-6 compounds can be evaluated in comprehensive in vitro studies
of absorption, distribution, metabolism, excretion and toxicity (ADMET), and only 2-3 compounds are advanced
in PK in vivo studies using external resources. To facilitate our discovery of nNOS inhibitors with desired CNS
drug properties, we have tried to carry out some of these ADMET/PK studies using commercially available
materials and internal resources, while the following quantitative analyses were impeded by the lack of reliable
and continuous LC-MS/MS support. Therefore, through this proposal, we aim to describe how the requested LC-
MS/MS system will be valuable and supportive of our current R35 award to improve the druglike properties of
novel nNOS inhibitors with higher efficiency and productivity. The plan of research encompasses the following
aims:
Specific Aims
Aim 1 Installation of a new triple-quadrupole mass spectrometer coupled with high-performance liquid
chromatography to be completed within three months. A routine training/maintenance policy will be established,
assisted by technicians from the vendor.
Aim 2 To establish an ADMET/PK routine screening platform within three months using internal resources.
Cellular and animal experiments will be carried out at the core facilities of Northwestern University with
appropriate training and approvals. Final quantitative analyses will be carried out using the new LC-MS/MS
system.
Aim 3 To evaluate 20 new analogues through different in vitro ADMET studies every year to identify nNOS
inhibitors with improved druglike parameters for CNS applications. Three to five inhibitors that are chosen from
in vitro assessment will then be advanced to brain/plasma PK studies.