Praveen P. N. Rao

Amyloid cascade in Alzheimer's disease: Recent advances in medicinal chemistry.

Alzheimers disease is of major concern all over the world due to a number of factors including (i) an aging population (ii) increasing life span and (iii) lack of effective pharmacotherapy options. The past decade has seen intense research in discovering disease-modifying multitargeting small molecules as therapeutic options. The pathophysiology of Alzheimers disease is attributed to a number of factors such as the cholinergic dysfunction, amyloid/tau toxicity and oxidative stress/mitochondrial dysfunction. In recent years, targeting the amyloid cascade has emerged as an attractive strategy to discover novel neurotherapeutics. Formation of beta-amyloid species, with different degrees of solubility and neurotoxicity is associated with the gradual decline in cognition leading to dementia. The two commonly used approaches to prevent beta-amyloid accumulation in the brain include (i) development of beta-secretase inhibitors and (ii) designing direct inhibitors of beta-amyloid (self-induced) aggregation. This review highlights the amyloid cascade hypothesis and the key chemical features required to design small molecules that inhibit lower and higher order beta-amyloid aggregates. Several recent examples of small synthetic molecules with disease-modifying properties were considered and their molecular docking studies were conducted using either a dimer or steric-zipper assembly of beta-amyloid. These investigations provide a mechanistic understanding on the structural requirements needed to design novel small molecules with anti-amyloid aggregation properties. Significantly, this work also demonstrates that the structural requirements to prevent aggregation of various amyloid species differs considerably, which explains the fact that many small molecules do not exhibit similar inhibition profile toward diverse amyloid species such as dimers, trimers, tetramers, oligomers, protofibrils and fibrils.

Curcumin Binding to Beta Amyloid: A Computational Study

Curcumin, a chemical constituent present in the spice turmeric, is known to prevent the aggregation of amyloid peptide implicated in the pathophysiology of Alzheimers disease. While curcumin is known to bind directly to various amyloid aggregates, no systematic investigations have been carried out to understand its ability to bind to the amyloid aggregates including oligomers and fibrils. In this study, we constructed computational models of (i) Aβ hexapeptide 16KLVFFA21 octamer steric‐zipper β‐sheet assembly and (ii) full‐length Aβ fibril β‐sheet assembly. Curcumin binding in these models was evaluated by molecular docking and molecular dynamics (MD) simulation studies. In both the models, curcumin was oriented in a linear extended conformation parallel to fiber axis and exhibited better stability in the Aβ hexapeptide 16KLVFFA21 octamer steric‐zipper model (Ebinding = −10.05 kcal/mol) compared to full‐length Aβ fibril model (Ebinding = −3.47 kcal/mol). Analysis of MD trajectories of curcumin bound to full‐length Aβ fibril shows good stability with minimum Cα‐atom RMSD shifts. Interestingly, curcumin binding led to marked fluctuations in the 14HQKLVFFA21 region that constitute the fibril spine with RMSF values ranging from 1.4 to 3.6 Å. These results show that curcumin binding to Aβ shifts the equilibrium in the aggregation pathway by promoting the formation of non‐toxic aggregates.

Nonsteroidal Anti-Inflammatory Drugs (NSAIDs): Progress in Small Molecule Drug Development.

Ever since the discovery of aspirin, small molecule therapeutics have been widely prescribed to treat inflammation and pain. Aspirin and several small molecule NSAIDs are known to inhibit the enzymes cyclooxygenase-1 (COX-1) and -2 (COX-2). Despite the success of NSAIDs to treat inflammatory disorders, the development of a clinically useful small molecule NSAIDs with decreased side effect profiles is an ongoing effort. The recent discovery and development of selective COX-2 inhibitors was a step toward this direction. Emerging trends are represented by the progress in the development of hybrid agents such as nitric oxide donor-NSAIDs (NO-NSAIDs) and dual COX/lipoxygenase (LOX) inhibitors. This review focuses on the recent advances in the rational design of small molecule NSAIDs in therapy.

Design, synthesis and structure–activity relationship (SAR) studies of 2,4-disubstituted pyrimidine derivatives: Dual activity as cholinesterase and Aβ-aggregation inhibitors

A novel class of 2,4-disubstituted pyrimidines (7a-u, 8a-f, 9a-e) that possess substituents with varying steric and electronic properties at the C-2 and C-4 positions, were designed, synthesized and evaluated as dual cholinesterase and amyloid-β (Aβ)-aggregation inhibitors. In vitro screening identified N-(naphth-1-ylmethyl)-2-(pyrrolidin-1-yl)pyrimidin-4-amine (9a) as the most potent AChE inhibitor (IC(50)=5.5 μM). Among this class of compounds, 2-(4-methylpiperidin-1-yl)-N-(naphth-1-ylmethyl)pyrimidin-4-amine (9e) was identified as the most potent and selective BuChE inhibitor (IC(50)=2.2 μM, selectivity index=11.7) and was about 5.7-fold more potent compared to the commercial, approved reference drug galanthamine (BuChE IC(50)=12.6 μM). In addition, the selective AChE inhibitor N-benzyl-2-(4-methylpiperazin-1-yl)pyrimidin-4-amine (7d), exhibited good inhibition of hAChE-induced aggregation of Aβ(1-40) fibrils (59% inhibition). Furthermore, molecular modeling studies indicate that a central pyrimidine ring serves as a suitable template to develop dual inhibitors of cholinesterase and AChE-induced Aβ aggregation thereby targeting multiple pathological routes in AD.

Anti-amyloid Aggregation Activity of Natural Compounds: Implications for Alzheimer’s Drug Discovery

Several plant-derived natural compounds are known to exhibit anti-amyloid aggregation activity which makes them attractive as potential therapies to treat Alzheimer’s disease. The mechanisms of their anti-amyloid activity are not well known. In this regard, many natural compounds are known to exhibit direct binding to various amyloid species including oligomers and fibrils, which in turn can lead to conformational change in the beta-sheet assembly to form nontoxic aggregates. This review discusses the mechanism of anti-amyloid activity of 16 natural compounds and gives structural details on their direct binding interactions with amyloid aggregates. Our computational investigations show that the physicochemical properties of natural products do fit Lipinski’s criteria and that catechol and catechol-type moieties present in natural compounds act as lysine site-specific inhibitors of amyloid aggregation. Based on these observations, we propose a structural template to design novel small molecules containing site-specific ring scaffolds, planar aromatic and nonaromatic linkers with suitably substituted hydrogen bond acceptors and donors. These studies will have significant implications in the design and development of novel amyloid aggregation inhibitors with superior metabolic stability and blood-brain barrier penetration as potential agents to treat Alzheimer’s disease.

Development and evaluation of multifunctional agents for potential treatment of Alzheimer's disease: application to a pyrimidine-2,4-diamine template.

We investigated a group of 2-benzylpiperidin-N-benzylpyrimidin-4-amines with various electron-withdrawing or electron-donating groups (EWGs or EDGs, respectively) as multi-targeted Alzheimers disease (AD) therapeutics. The synthesized derivatives were screened for anti-cholinesterase (AChE and BuChE), anti-Aβ-aggregation (AChE- and self-induced) and anti-β-secretase (BACE-1) activities in an effort to identify lead, multifunctional candidates as part of our multi-targeted approach to treat AD. Biological assessment revealed that the nature of the substituent on the C-4 benzylamine group (e.g., halogen vs methoxy-based) greatly affected the biological profile. In vitro screening identified N(2)-(1-benzylpiperidin-4-yl)-N(4)-(3,4-dimethoxybenzyl)pyrimidine-2,4-diamine (7h) as the lead candidate with a dual ChE (AChE IC(50)=9.9 μM; BuChE IC(50)=11.4 μM), Aβ-aggregation (AChE-induced=59.3%; self-induced=17.4% at 100 μM) and BACE-1 (34% inhibition at 10 μM) inhibitory profile along with good cell viability (% neuroblastoma cell viability at 40 μM=81.0%). Molecular modeling studies indicate that a central pyrimidine-2,4-diamine ring serves as a suitable template to develop novel small molecule candidates to target multiple pathological routes in AD.

Synthesis and biological evaluation of isoxazolo[4,5-d]pyridazin-4-(5H)-one analogues as potent anti-inflammatory agents.

In this study, eighteen new isoxazolo[4,5-d]pyridazin-4(5H)-one derivatives possessing either a 1,3,4-thiadiazole or a 1,2,4-triazole-5-thione moiety were synthesized and tested for anti-inflammatory activity in vitro (COX-1/COX-2, 5-LOX) and in vivo (rat paw edema assay). Compounds 15, 16, 25, 26 and 28-30 showed dual COX-2 (IC(50)s in the 2.1-10.9 μM range), and 5-LOX (IC(50)s in the 6.3-63.5 μM range) inhibitory activity. When administered orally to rats, dual COX-2/5-LOX inhibitors showed higher anti-inflammatory activity in vivo (30-45% reduction of the inflammatory response) than the reference drug ibuprofen (18%). Among dual COX-2/5-LOX inhibitors, the most potent compound (28) exhibited the best anti-inflammatory profile by inhibiting both COX-2 (IC(50)=2.1 μM) and 5-LOX (IC(50)=6.3 μM) enzymes. We investigated the binding interactions of compound 28 by an enzyme-ligand molecular modeling (docking) studies, which showed favorable binding interactions in both COX-2 and 5-LOX active sites. Furthermore, the dual acting COX-2/5-LOX compound 28 exhibited a superior gastrointestinal safety profile (ulcer index=0.25) compared to the reference drug ibuprofen (UI=7.0) when administered orally at the same molar dose. These observations suggest that isoxazolo[4,5-d]pyridazin-4(5H)-one analogs represent a new scaffold to design potent, effective, and safe anti-inflammatory agents possessing dual COX-2/5-LOX inhibitory activity.

Tau-derived-hexapeptide 306VQIVYK311 aggregation inhibitors: nitrocatechol moiety as a pharmacophore in drug design.

The nitrocatechol derivatives tolcapone (1) and entacapone (2), used as adjunctive therapy in the treatment of Parkinsons disease, were investigated for their potential to inhibit the tau-derived-hexapeptide 306VQIVYK311. They were compared to small molecules that contain similar pharmacophores including the catechol derivatives (dopamine 3 and epinephrine 4), nitroderivatives (nifedipine 5 and chloramphenicol 6), nitrocatechol isomers (7 and 8), and a tolcapone derivative (13) lacking the nitrocatechol moiety. The aggregation kinetics by thioflavin S fluorescence assay indicates that both tolcapone (1) and entacapone (2) exhibit antiaggregation properties. These findings were supported by transmission electron microscopy (TEM) and circular dichroism (CD) spectroscopy measurements which suggest that the nitrocatechol (3,4-dihydroxy-5-nitrophenyl) moiety is a suitable pharmacophore in the design of new tau-aggregation inhibitors. Furthermore, tolcapone (1) was identified as most active compound with antiaggregation activity (46% inhibition of fluorescence intensity at 50 μM), which was supported by TEM data. The in silico steric zipper model of the tau-derived-hexapeptide 306VQIVYK311 indicates that the 3,4-dihydroxy-substituent present in tolcapone (1) and entacapone (2) underwent polar contacts with lysine side chains (VQIVYK), whereas the charged 5-nitrosubstituent was in close contact with lysine side chain present in the steric zipper region suggesting the critical role of a nitrocatechol (3,4-dihydroxy-5-nitrophenyl) pharmacophore present in tolcapone (1) and entacapone (2) in tau-hexapeptide binding and prevention of β-sheet assembly. Our results have significant implications in the design and development of tau-aggregation inhibitors.

Development of 2-substituted-N-(naphth-1-ylmethyl) and N-benzhydrylpyrimidin-4-amines as dual cholinesterase and Aβ-aggregation inhibitors: Synthesis and biological evaluation

A group of 2-substituted N-(naphth-1-ylmethyl)pyrimidin-4-amines (6a-k) and N-benzhydrylpyrimidin-4-amines (7a-k) in conjunction with varying steric and electronic properties at the C-2 position were designed, synthesized and evaluated as dual cholinesterase and amyloid-β (Aβ)-aggregation inhibitors. The naphth-1-ylmethyl compound 6f (2-(4-cyclohexylpiperazin-1-yl)-N-(naphth-1-ylmethyl)pyrimidin-4-amine) exhibited optimum dual ChE (AChE IC(50)=8.0 μM, BuChE IC(50)=3.9 μM) and hAChE-promoted Aβ-aggregation inhibition (30.8% at 100 μM), whereas in the N-benzhydryl series, compound 7f (N-benzhydryl-2-(4-cyclohexylpiperazin-1-yl)pyrimidin-4-amine) exhibited optimum combination of dual ChE (AChE IC(50)=10.0 μM, BuChE IC(50)=7.6μM) and hAChE-promoted Aβ-aggregation inhibition (32% at 100 μM). These results demonstrate that a 2,4-disubstituted pyrimidine ring serves as a suitable template to target multiple pathological routes in AD, with a C-2 cyclohexylpiperazine substituent providing dual ChE inhibition and potency whereas a C-4 diphenylmethane substituent provides Aβ-aggregation inhibition.

Design, synthesis and evaluation of 2,4-disubstituted pyrimidines as cholinesterase inhibitors

A group of 2,4-disubstituted pyrimidine derivatives (7a-e, 8a-e and 9a-d) that possess a variety of C-2 aliphatic five- and six-membered heterocycloalkyl ring in conjunction with a C-4 arylalkylamino substituent were designed, synthesized and evaluated as cholinesterase (ChE) inhibitors. The steric and electronic properties at C-2 and C-4 positions of the pyrimidine ring were varied to investigate their effect on ChE inhibitory potency and selectivity. The structure-activity relationship (SAR) studies identified N-benzyl-2-thiomorpholinopyrimidin-4-amine (7c) as the most potent cholinesterase inhibitor (ChEI) with an IC(50)=0.33 microM (acetylcholinesterase, AChE) and 2.30muM (butyrylcholinesterase, BuChE). The molecular modeling studies indicate that within the AChE active site, the C-2 thiomorpholine substituent was oriented toward the cationic active site region (Trp84 and Phe330) whereas within the BuChE active site, it was oriented toward a hydrophobic region closer to the active site gorge entrance (Ala277). Accordingly, steric and electronic properties at the C-2 position of the pyrimidine ring play a critical role in ChE inhibition.