This supplemental application to Dr. Eckenhoff's PPG is for additional support for aims that were not funded following the previous review in 2006.
The aims now proposed are different from the original Aim 1.2, and have been expanded to include 3 parts:
Aim 1. 2 is to use computational methods to create in silico binding pocket models for the anesthetic binding sites in apoferritin and the L38M 4-Helix bundle protein (projects 2 &3).
Aim 1. 3 will use virtual screening of multi-conformer drug libraries to identify novel anesthetic candidates that are likely to bind to apoferritin and the 4HB.
Aim 1. 4 will test identified compounds for binding affinity to apoferritin and 4HB, confirm their location in the anesthetic sites using NMR and crystallography, and apply QSAR analysis to define an optimized pharmacophore. Another divergence from the original aims is that these studies now focus on alkyl phenol drugs (derivatives of propofol) and less emphasis is placed on volatile anesthetics. Strengths: Dr. Eckenhoff is an outstanding researcher and leader in the field of anesthetic mechanisms, and has established a productive research enterprise investigating anesthetic interactions with proteins. The environment, including collaborative colleagues, access to state-of-the-art methods, and institutional support at University of Pennsylvania, is outstanding. The redirection toward alkyl phenol drugs is justified by concerns about neurotoxicity associated with volatile ether anesthetics. This new direction for Dr. Eckenhoff's PPG project is also quite innovative, and represents a significant change from simply studying anesthetic binding to proteins-- the project now aims to use what has been learned to try and identify new anesthetic drugs. The computational tools that will be applied to achieve the aims also represent a novel approach to identifying candidate anesthetics. The application presents a strong logical, albeit largely conceptual argument for why the model 4HB proteins (apoferritin and L38M) may be surrogates for anesthetic binding sites in LGICs. The application presents a good deal of new preliminary data on the apoferritin pocket showing sufficient progress to raise confidence that the computational aims can be achieved. Weaknesses: The switch in focus from volatile anesthetics to alkyl phenol drugs weakens the links between Project 1 and other PPG projects, although Project 2 now adds propofol binding and co-crystallization studies with L38M. All of the preliminary data is also from the apoferritin site, which further reduces linkage to and interdependence with other projects studying model 4HBs that are similar to L38M. Given the hypothesis about the relationship between the model proteins and anesthetic target sites in LGICs, the computational approach to identifying novel anesthetic candidates seems rather round-about. It is not clear that alternative approaches have been considered. The application criticizes early studies for their narrow focus on a single class of anesthetics, but this project does the same thing by focusing almost entirely on alkyl phenol drugs. Given that the L38M 4HB structure has been solved both by NMR and x-ray diffraction (and an anesthetic-bound structure is solved with NMR), more analysis based on this model would have been desirable. Despite the significant amount of preliminary data presented, no anesthetic candidates are identified or tested for Aim 1.4. Thus the overall feasibility of this strategy remains very speculative. Budget: The consortium costs in Project 1 seem very high for in silico work. Dr. Liang's year 1 costs include $9420 for tuition and $3135 for supplies seems large for work that will be entirely computational Other Considerations: Protection of Human Subjects from Research Risks: NA Vertebrate Animals: NA Biohazards: No Concerns PROJECT 2: STRUCTURAL STUDIES ON ANESTHETIC COMPLEXES WITH SYNTHETIC FOUR-a-HELIX BUNDLE (Dr. Jonas Johansson, M.D., Ph.D.) PRIORITY SCORE: 206
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