David Grierson

Professor

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Graduate Student Supervision

Doctoral Student Supervision (Jan 2008 - May 2019)
Benzoisoxazolone-based scaffolds for development of diversity driven small molecule libraries : an effective tool to identify novel molecules targeting key therapeutic applications (2017)

The screening of compound libraries has repeatedly demonstrated its effectiveness as a research tool in drug discovery. Parallel synthesis is a powerful approach for the assembly of compound libraries possessing a wide range of structural and functional group diversity for evaluation against both new and established biological targets.This thesis project was focused on the development of four amino and/or carboxy-substituted benzoisoxazolone (B to E) as privileged scaffolds for the parallel synthesis of diversity driven libraries to identify isoform selective inhibitors of Rho-kinase (ROCK) for the treatment of diabetic cardiovascular disease and different cancers, as well as potential anti-HIV agents that block HIV pre-mRNA alternative splicing, and inhibitors of protein arginine methyltransferase 4 (PRMT4) for the treatment of cancer. In the context of developing a synthesis to the C5-amino substituted benzoisoxazolone scaffold B, the general methodology to construct the benzoisoxazolone ring system found in all four scaffolds is described. This involved reaction of a requisite salicylic acid derivative with hydroxylamine followed by dehydrative cyclization to assemble the isoxazolone ring. Also described in Chapter 2 was the synthesis of the C4-amino substituted scaffold C. The crucial issue encountered was control of chemoselectivity during dehydrative ring closure. The intent was that scaffold B and C-based libraries would contain type 2 ROCK inhibitors. The concept described in Chapter 3 was the “formal” use of the C5-carboxy substituted benzoisoxazolone scaffold D to generate novel molecules that mimic the tetracyclic indole anti-HIV agent IDC16, through conservation of the central (B and C-rings) in its structure. Based on the results of virtual screening/docking experiments on PRMT4, chapter 4 describes the synthesis of the 5-carboxy-7-amino substituted benzoisoxazolone scaffold E and its functionalization such that the benzyl substituents introduced at N-2 and the 5-benzamide motif interact with hydrophobic residues along the PRMT4 substrate binding site, and that the appending side chain at position 7, incorporating a urea sub-element, binds strongly in the arginine binding region in PRMT4. Overall, the synthesis of scaffolds B to E on a multigram scale has been achieved, opening the way for diversity driven parallel synthesis of novel compound libraries.

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Parallel synthesis of small molecule compound libraries : isoform selective Rho-Kinase inhibitors (2015)

The objective of this thesis was to use parallel synthesis to build small molecule libraries of novel compounds with potential as isoform selective type II rho-kinase (ROCK1/2) inhibitors. Considerable effort is ongoing to identify isoform selective ROCK inhibitors, as hyperactivity of the ROCK is implicated in cardiovascular diseases, cancer, diabetes and many other diseases that affect millions of Canadians, and billions of people worldwide. In the absence the extensive structural details of the type II kinase inhibitor binding mode, we have used an empirical approach to the design of type II ROCK inhibitors, based on the conception that such molecules correspond to elongated structures with H-bonding functional elements in their central portion, a motif at one end that mimics the adenine ring in ATP, and a hydrophobic moiety at the other end of the molecule that will interact with an allosteric pocket in the ATP binding region of the kinase. Based on these very general structural requirements, eleven different representative libraries of novel compounds (not described in CAS) were designed and synthesized. In the initial series of compounds, a 2-pyridinone motif was employed as the hinge binding element, and the central portion corresponded to carboxamide substituted oxazoline, oxadiazole, or 2-aminothiazole system, joined through the amide bond to a set of structurally diverse aromatic, heteroaromatic and benzylamine subunits, corresponding to the diversity elements. The preliminary assay results demonstrated that, overall, these compounds were weak and non-selective ROCK1/2 inhibitors when compared to H-1152 as a positive control at 10 μM concentration. However, further structural modification revealed the interest in using an indazole motif as the hinge binder in conjunction with the 2-aminothiazole carboxamide linker. Indeed, more potent activities were observed in the single point assay for a significant portion of the libraries of molecules built around these structural components. Further evaluation of 18 active compounds in a 10-point assay, by Invitrogen, to determine IC50’s revealed that indazole-based inhibitors are active at low micromolar concentrations (1-10 µM), but do not display any significant isoform selectivity.

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