In silico design and biological evaluation of a dual specificity kinase inhibitor targeting cell cycle progression and angiogenesis

Methodology: We have utilized a rational in silico-based approach to demonstrate the design and study of a novel compound that acts as a dual inhibitor of vascular endothelial growth factor receptor 2 (VEGFR2) and cyclin-dependent kinase 1 (CDK1). This compound acts by simultaneously inhibiting pro-Angiogenic signal transduction and cell cycle progression in primary endothelial cells. JK-31 displays potent in vitro activity against recombinant VEGFR2 and CDK1/cyclin B proteins comparable to previously characterized inhibitors. Dual inhibition of the vascular endothelial growth factor A (VEGF-A)-mediated signaling response and CDK1-mediated mitotic entry elicits anti-Angiogenic activity both in an endothelial-fibroblast co-culture model and a murine ex vivo model of angiogenesis

Site-directed M2 proton channel inhibitors enable synergistic combination therapy for rimantadine-resistant pandemic influenza.

Pandemic influenza A virus (IAV) remains a significant threat to global health. Preparedness relies primarily upon a single class of neuraminidase (NA) targeted antivirals, against which resistance is steadily growing. The M2 proton channel is an alternative clinically proven antiviral target, yet a near-ubiquitous S31N polymorphism in M2 evokes resistance to licensed adamantane drugs. Hence, inhibitors capable of targeting N31 containing M2 (M2-N31) are highly desirable. Rational in silico design and in vitro screens delineated compounds favouring either lumenal or peripheral M2 binding, yielding effective M2-N31 inhibitors in both cases. Hits included adamantanes as well as novel compounds, with some showing low micromolar potency versus pandemic "swine" H1N1 influenza (Eng195) in culture. Interestingly, a published adamantane-based M2-N31 inhibitor rapidly selected a resistant V27A polymorphism (M2-A27/N31), whereas this was not the case for non-adamantane compounds. Nevertheless, combinations of adamantanes and novel compounds achieved synergistic antiviral effects, and the latter synergised with the neuraminidase inhibitor (NAi), Zanamivir. Thus, site-directed drug combinations show potential to rejuvenate M2 as an antiviral target whilst reducing the risk of drug resistance

Design, synthesis and biological evaluation of inhibitors of FGFR, VEGFR-2 and Ras proteins

Deregulation of kinase activity has emerged as a major mechanism by which cancer cells evade normal physiological functions such as growth and survival. The identification of mutations in FGFR-3 within non invasive tumours of bladder cancer and over expression of this receptor in invasive tumours and superficial tumours makes FGFR-3 a promising target in developing a therapy for the treatment of urothelial bladder cancer. It was found that inhibiting VEGFR-2 together with FGFR provides an attractive strategy for the development of new anti-angiogenic agents as a potential anti -cancer therapy. In this project, the three dimensional structure of both the FGFR and VEGFR-2 was used in conjunction with the de novo methods such as SPROUT to generate oxindole- based novel inhibitors of FGFR and VEGFR-2 with ICso values in the range of 1-10 flM (Compound 4.7 inhibits FGFR and VEGFR-2 with ICso values of 3.9 uM and 1.6 flM for respectively (Chapter 4). These compounds show encouraging anti-angiogenic activity in the lower u M concentrations. In Chapter 5, intramolecular H-bonding concept was used to design pyrazole-based inhibitors of FGFR and VEGFR-2 with 29% and 63% inhibition at 10 uM respectively. Benzo-fused designs based on pyrazole scaffolds yielded indazole based compounds with inhibitory effects of 45%, 41 % and 74% for FGFR-1, FGFR-3 and VEGFR-2 respectively. Shape similarity was used to find novel hinge binders replacing the indazole core with hydroxy-quinolinone moiety which showed an encouraging level of inhibition with 30% and 64% for FGFR and VEGFR-2 respectively. Purine based inhibitors were designed to exploit the smaller gatekeeper residue in both the FGFR and VEGFR-2, but all the compounds show weaker or no inhibition at 10 u M (Chapter 5). In Chapter 6, structure guided method were used to design oxadiazole-based novel VEGFR-2 inhibitor with an ICso value of 1.6 j.1M. All these approaches are novel used in complementary to HTS. In Chapter 3, SPROUT was used to design novel inhibitors of Ras protein and the preliminary analysis of these inhibitors show preferential binding of these inhibitors to the active form compared to the inactive form. These molecules were first amongst the de novo/designed inhibitors of Ras which could serve as a starting point f-or the future potential inhibitors of protein-protein interaction.EThOS - Electronic Theses Online ServiceGBUnited Kingdo

Isoquinoline-1,3-diones as Selective Inhibitors of Tyrosyl DNA Phosphodiesterase II (TDP2)

Tyrosyl DNA phosphodiesterase II (TDP2) is a recently discovered enzyme that specifically repairs DNA damages induced by topoisomerase II (Top2) poisons and causes resistance to these drugs. Inhibiting TDP2 is expected to enhance the efficacy of clinically important Top2-targeting anticancer drugs. However, TDP2 as a therapeutic target remains poorly understood. We report herein the discovery of isoquinoline-1,3-dione as a viable chemotype for selectively inhibiting TDP2. The initial hit compound <b>43</b> was identified by screening our in-house collection of synthetic compounds. Further structure–activity relationship (SAR) studies identified numerous analogues inhibiting TDP2 in low micromolar range without appreciable inhibition against the homologous TDP1 at the highest testing concentration (111 μM). The best compound <b>64</b> inhibited recombinant TDP2 with an IC₅₀ of 1.9 μM. The discovery of this chemotype may provide a platform toward understanding TDP2 as a drug target

Receptor tyrosine kinase structure and function in health and disease

Receptor tyrosine kinases (RTKs) are membrane proteins that control the flow of information through signal transduction pathways, impacting on different aspects of cell function. RTKs are characterized by a ligand-binding ectodomain, a single transmembrane α-helix, a cytosolic region comprising juxtamembrane and kinase domains followed by a flexible C-terminal tail. Somatic and germline RTK mutations can induce aberrant signal transduction to give rise to cardiovascular, developmental and oncogenic abnormalities. RTK overexpression occurs in certain cancers, correlating signal strength and disease incidence. Diverse RTK activation and signal transduction mechanisms are employed by cells during commitment to health or disease. Small molecule inhibitors are one means to target RTK function in disease initiation and progression. This review considers RTK structure, activation, and signal transduction and evaluates biological relevance to therapeutics and clinical outcomes

Design, Synthesis, and Biological Evaluations of Hydroxypyridone­carboxylic Acids as Inhibitors of HIV Reverse Transcriptase Associated RNase H

Targeting the clinically unvalidated reverse transcriptase (RT) associated ribonuclease H (RNase H) for human immunodeficiency virus (HIV) drug discovery generally entails chemotypes capable of chelating two divalent metal ions in the RNase H active site. The hydroxypyridone­carboxylic acid scaffold has been implicated in inhibiting homologous HIV integrase (IN) and influenza endonuclease via metal chelation. We report herein the design, synthesis, and biological evaluations of a novel variant of the hydroxypyridone­carboxylic acid scaffold featuring a crucial <i>N</i>-1 benzyl or biarylmethyl moiety. Biochemical studies show that most analogues consistently inhibited HIV RT-associated RNase H in the low micromolar range in the absence of significant inhibition of RT polymerase or IN. One compound showed reasonable cell-based antiviral activity (EC₅₀ = 10 μM). Docking and crystallographic studies corroborate favorable binding to the active site of HIV RNase H, providing a basis for the design of more potent analogues

Design, Synthesis, and Biological Evaluations of Hydroxypyridone­carboxylic Acids as Inhibitors of HIV Reverse Transcriptase Associated RNase H

Targeting the clinically unvalidated reverse transcriptase (RT) associated ribonuclease H (RNase H) for human immunodeficiency virus (HIV) drug discovery generally entails chemotypes capable of chelating two divalent metal ions in the RNase H active site. The hydroxypyridone­carboxylic acid scaffold has been implicated in inhibiting homologous HIV integrase (IN) and influenza endonuclease via metal chelation. We report herein the design, synthesis, and biological evaluations of a novel variant of the hydroxypyridone­carboxylic acid scaffold featuring a crucial <i>N</i>-1 benzyl or biarylmethyl moiety. Biochemical studies show that most analogues consistently inhibited HIV RT-associated RNase H in the low micromolar range in the absence of significant inhibition of RT polymerase or IN. One compound showed reasonable cell-based antiviral activity (EC₅₀ = 10 μM). Docking and crystallographic studies corroborate favorable binding to the active site of HIV RNase H, providing a basis for the design of more potent analogues

Inhibition of VEGFR2 and CDK1/cyclin B kinase activity by JK-31.

<p>IC₅₀ values were derived from curves (<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0110997#pone-0110997-g001" target="_blank">Figures 1E and 1F</a>) generated using an <i>in vitro</i> kinase assay (see <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0110997#s2" target="_blank">Methods</a>).</p><p>Inhibition of VEGFR2 and CDK1/cyclin B kinase activity by JK-31.</p