Nidhi Chadha

A New SiF–Dipropargyl Glycerol Scaffold as a Versatile Prosthetic Group to Design Dimeric Radioligands: Synthesis of the [18F]BMPPSiF Tracer to Image Serotonin Receptors

A novel SiX–dipropargyl glycerol scaffold (X: H, F, or 18F) was developed as a versatile prosthetic group that provides technical advantages for the preparation of dimeric radioligands based on silicon fluoride acceptor pre‐ or post‐labeling with fluorine‐18. Rapid conjugation with the prosthetic group takes place in microwave‐assisted click conjugation under mild conditions. Thus, a bivalent homodimeric SiX–dipropargyl glycerol derivatized radioligand, [18F]BMPPSiF, with enhanced affinity was developed by using click conjugation. High uptake of the radioligand was demonstrated in 5‐HT1A receptor‐rich regions in the brain with positron emission tomography. Molecular docking studies (rigid protein–flexible ligand) of BMPPSiF and known antagonists (WAY‐100635, MPPF, and MefWAY) with monomeric, dimeric, and multimeric 5‐HT1A receptor models were performed, with the highest G score obtained for docked BMPPSiF: −6.766 as compared with all three antagonists on the monomeric model. Multimeric induced‐fit docking was also performed to visualize the comparable mode of binding under in vivo conditions, and a notably improved G score of −8.455 was observed for BMPPSiF. These data directly correlate the high binding potential of BMPPSiF with the bivalent binding mode obtained in the biological studies. The present study warrants wide application of the SiX–dipropargyl glycerol prosthetic group in the development of ligands for imaging with enhanced affinity markers for specific targeting based on peptides, nucleosides, and lipids.

Synthesis and biological evaluation of newly designed phosphonate based bone-seeking agent.

A cyclic tetraaza based bifunctional triphosphonate ligand 10-(2-aminoethyl)-1,4,7,10-tetraazacyclododecane-1,4,7-tris(methylenephosphonic acid) (DO3MP-EA) was synthesized as bone-seeking theranostic agent. The compound was characterized by spectroscopic techniques and labelled with (99m)Tc with more than 97% purity. Blood clearance of 99mTc labelled compound a quick wash out from the circulation. The compound was excreted mainly via kidneys and accumulation of (99m)Tc-DO3MP-EA in bone was 9.53 ± 1.06% of injected dose per gram of bone at 1 h. The preliminary CADD analysis confirms the efficacy of DO3MP-EA (G Score -7.005) as better binding agent for osteocalcin (pdb 1Q8H) rather than other known clinical agents. Subsequently stability constant of chelate with Ga(III) was found to be 18.6 which confirms its efficacy as (68)Ga labelled PET radiopharmaceutical for bone.

Synthesis, Biological Evaluation and Molecular Docking Studies of High-Affinity Bone Targeting N,N'-Bis (alendronate) Diethylenetriamene-N,N'-Triacetic Acid: A Bifunctional Bone Scintigraphy Agent

A bisphosphonate derivative DTPA‐bis(alendronate) conjugate has been synthesized and evaluated as potential radiopharmaceutical for bone imaging. The compound was synthesized by the covalent coupling of DTPA‐bis(anhydride) with alendronate and was char‐acterized on the basis of IR, NMR and mass spectroscopy. It was labelled with 99mTc with 96% efficacy and was found stable for about 24 h under physiological conditions. Blood kinetic studies of 99mTc DTPA‐bis(alendronate) showed a biexponential pattern as well as quick washout from the blood circulation. The biological t1/2(F) and t1/2(S) were found to be 50 min ± 0.001 and 6 h 30 min ± 0.005, respectively. Imaging and biodistribution studies showed a significant accumulation of 99mTc DTPA‐bis(alendronate) conjugate at bone site. Bone‐to‐muscles ratios were 12.08 ± 0.001 at 1 h, 45.33 ± 0.001 at 4 h and 35.83 ± 0.001 at 24 h after post‐injection, respectively. The receptor binding of the 99mTc‐DTPA‐bis (alendronate) was established on human bone cell line (Soas‐2) revealed KD = 0.86 nm. The preliminary result of the 99mTc‐DTPA‐bis(alendronate) is encouraging to carrying out further in vivo experiment for targeted bone imaging because of good‐bone‐to‐normal‐organ contrast. Further docking analysis with molecular targets, farnesyl diphosphate synthase, geranylgeranyl pyrophosphate and osteocalcin revealed the high affinity of −17.419 and thus represents strong potential of bone‐imaging agent.

Oxime-dipeptides as anticholinesterase, reactivator of phosphonylated-serine of AChE catalytic triad: probing the mechanistic insight by MM-GBSA, dynamics simulations and DFT analysis

Neuropathological cascades leading to reduced cholinergic transmission in Alzheimer’s disease led to development of AChE-inhibitors. Although lethal dose of some inhibitors cause interruption with AChE mediated mechanism but reversible AChE inhibitors can assist in protection from inhibition of AChE and hence in an aim to probe potential molecules as anticholinesterase and as reactivators, computationally structure-based approach has been exploited in this work for designing new 2-amino-3-pyridoixime-dipeptides conjugates. We have combined MD simulations with flexible ligand docking approach to determine binding specificity of 2-amino-3-pyridoixime dipeptides towards AChE (PDB 2WHP). PAS residues are found to be responsible for oxime-dipeptides binding along with π–π interactions with Trp86 and Tyr286, hydrogen bonding with side chains of Asp74 and Tyr341 (Gscore –10.801 and MM-GBSA free energy –34.89 kcal/mol). The docking results depicted complementary multivalent interactions along with good binding affinity as predicted from MM-GBSA analysis. The 2-amino-3-pyridoxime-(Arg-Asn) AChE systems subjected to MD simulations under explicit solvent systems with NPT and NVT ensemble. MD simulations uncovered dynamic behavior of 2-amino-3-pyridoxime-(Arg-Asn) and exposed its mobile nature and competence to form strong long range-order contacts towards active site residues to approach inhibited serine residue and facilitated via large contribution from hydrogen bonding and water bridges along with slow and large movements of adjacent important residues. In an effort to evaluate the complete potential surface profile, 2-amino-3-pyridoxime induced reactivation pathway of sarin–serine adduct has been investigated by the DFT approach at the vacuum MO6/6–311G (d, p) level along with the Poisson-Boltzmann solvation model and found to be of relatively low energy barrier. The pKa evaluation has revealed the major deprotonated 2-amino-3-pyridoixime species having pKa of 6.47 and hence making 2-amino-3-pyridoxime-(Arg-Asn) potential anticholinesterase and reactivator for AChE under the physiological pH.

Synthesis, preclinical evaluation and molecular modelling of macrocyclic appended 1-(2-methoxyphenyl)piperazine for 5-HT1A neuroreceptor imaging

5-HT1A receptors are known to be implicit in a number of neuropsychiatric fluctuations related to mood and anxiety. Their visualization in the human brain using PET, SPECT or MRI is of great importance in the management and treatment of neurological disorders. The present work focuses on the metal complexes (Gd3+, Eu3+ and Ga3+) of DO3A-butyl-MPP to be used as brain (cerebral cortex, hippocampus, and amygdala) imaging agents using different modalities. Synthesis of 2,2′,2′′-(10-(2-(4-(4-(2-methoxyphenyl)piperazin-1-yl)butylamino)-2-oxoethyl)-1,4,7,10-tetraazacyclododecane-1,4,7-triyl)triacetic acid (DO3A-butyl-MPP) was achieved by conjugating the chloroacetylated derivative of 1-(2-methoxyphenyl)piperazine-butylamine with trisubstituted cyclen with subsequent cleavage with trifluoroacetic acid (TFA). The resulting compound was then labeled with GdCl3 and 68GaCl3 to perform MRI (relaxivity studies) and PET respectively. The longitudinal relaxivity (r1), and transverse relaxivity (r2), were determined to be 6.66 and 11.486 mM−1 s−1 respectively on 7 T at 21 °C. The SD (Sprague Dawley) rat brain uptake was 3.91% ID per g (percentage of the injected dose per gram) at 30 min post injection. Homology modeling and docking studies at the shallow antagonist binding pocket of 5-HT1A show a high G score of −12.132 that confers high binding of the ligand at the target receptor.

In silico thermodynamics stability change analysis involved in BH4 responsive mutations in phenylalanine hydroxylase: QM/MM and MD simulations analysis

The mammalian tetrahydrobiopterin (BH4)-dependent phenylalanine hydroxylases (PAH), involved in important metabolic pathways of phenylalanine, belong to non-heme iron-containing aromatic acid hydroxylases’ enzyme (AAH) family. AAHs utilize BH4 as protein co-factor and thus promote hydroxylation reactions of their substrates. Any alterations in BH4 -mediated AAH’s pathway or mutations in these enzymes are responsible for various disorders, and thus highlights the importance of mutational analysis to assess the effect on their biosynthetic pathways. Our present studies are aimed at single-site mutations in PAH that lead to thermodynamic stability change upon folding and further validation of designed non-reduced BH2 designed co-factors. We have presented single-site mutational analysis of PAH where single-site mutations have been identified from known literature. Further, in silico studies with the PAH, in silico mutant PAH, and crystallized known mutant A313T forms, involved QM/MM and Molecular Dynamics (MD) simulations analysis. The modified co-factor A showed high affinity with PAH and all mutant PAH with high G-score of −14.851. The best pose high affinity co-factor A subjected to QM/MM optimization which leads to square-pyramidal coordination of non-heme active site. The structural and energetic information obtained from the production phase of 20 ns MD simulation of co-factor-metalloprotein complex results helped to understand the binding mode and involvement of three molecules throughout the reaction pathways’ catalysis of PAH. The free energies of binding (dG) of A were found to be −68.181 kcal/mol and −72.249 for 1DMW and 1TDW for A313T mutant. Binding of Co-factor A do not perturb the coordination environment of iron at the active site which resides in 2-Histdine and 1-Glutamate triad, and may enhance the percentage response towards co-factor-mediated therapy.

Engineered Nanoparticles Associated Metabolomics

AbstractNanotechnology is of great economic importance due to rapid growth in industrial and household applications and their continuous release into environmental matrices has increased the concerns regarding its potential impact on human and environment. To evaluate the potential toxicity of nanoparticles (NPs), the impact on humans and ecosystems needs to be understood from mechanistic insight by metabolomics. The analytical methods and omics technologies are particularly well-suited to evaluate these two dimensions in environment and both in vitro and in vivo systems, respectively. In this paper, the currently most effective methods for sampling and detection are presented together for future standardization of protocols providing profiles of NPs and associated metabolomics. All these approaches are designed without compromising the reliability and quality of the analytical results for their application in understanding the metabolomics and chemistry of NPs in the environmental matrices. Among samplin...

Preclinical Evaluation of a Potential GSH Ester Based PET/SPECT Imaging Probe DT(GSHMe)2 to Detect Gamma Glutamyl Transferase Over Expressing Tumors

Gamma Glutamyl Transferase (GGT) is an important biomarker in malignant cancers. The redox processes ensuing from GGT-mediated metabolism of extracellular GSH are implicated in critical aspects of tumor cell biology. Reportedly, Glutathione monoethyl ester (GSHMe) is a substrate of GGT, which has been used for its rapid transport over glutathione. Exploring GGT to be an important target, a homobivalent peptide system, DT(GSHMe)2 was designed to target GGT-over expressing tumors for diagnostic purposes. DT(GSHMe)2 was synthesized, characterized and preclinically evaluated in vitro using toxicity, cell binding assays and time dependent experiments. Stable and defined radiochemistry with 99mTc and 68Ga was optimized for high radiochemical yield. In vivo biodistribution studies were conducted for different time points along with scintigraphic studies of radiolabeled DT(GSHMe)2 on xenografted tumor models. For further validation, in silico docking studies were performed on GGT (hGGT1, P19440). Preclinical in vitro evaluations on cell lines suggested minimal toxicity of DT(GSHMe)2 at 100 μM concentration. Kinetic analysis revealed transport of 99mTc-DT(GSHMe)2 occurs via a saturable high-affinity carrier with Michaelis constant (Km) of 2.25 μM and maximal transport rate velocity (Vmax) of 0.478 μM/min. Quantitative estimation of GGT expression from western blot experiments showed substantial expression with 41.6 ± 7.07 % IDV for tumor. Small animal micro PET (Positron Emission Tomography)/CT(Computed Tomography) coregistered images depicted significantly high uptake of DT(GSHMe)2 at the BMG-1 tumor site. ROI analysis showed high tumor to contra lateral muscle ratio of 9.33 in PET imaging studies. Avid accumulation of radiotracer was observed at tumor versus inflammation site at 2 h post i.v. injection in an Ehrlich Ascites tumor (EAT) mice model, showing evident specificity for tumor. We propose DT(GSHMe)2 to be an excellent candidate for prognostication and tumor imaging using PET/SPECT.

Investigation for the interaction of tyramine-based anthraquinone analogue with human serum albumin by optical spectroscopic technique.

A newly synthesized anthraquinone derivative ‘N‐(2‐methylanthraquinone)‐4‐(2‐aminoethyl) phenol’ (Tyan) were characterized as a fluorophore from photophysical analysis by measuring the UV‐Vis absorptive (λex = 325 nm) and fluorescence emitive (λem = 660 nm) values. Density functional theory additionally supported the spectroscopic data by modulation of highest occupied molecular orbital rather than lowest unoccupied molecular orbital due to the affect of tyramine moiety present in Tyan. The pharmacological importance of Tyan was evaluated by molecular docking with human serum albumin. The molecular docking of the Tyan was performed with the crystal structure of human serum albumin (PDB entry 1E78), which shows binding in all the three domains of human serum albumin corresponding to −7.74 as the GScore. Moreover, the interactions of human serum albumin with Tyan were assessed employing fluorescence spectroscopy under simulative physiological conditions, and the binding constant for the interaction at 25 °C was found to be 0.6 × 103/m.

Picolinic acid based acyclic bifunctional chelating agent and its methionine conjugate as potential SPECT imaging agents: syntheses and preclinical evaluation

Bifunctional chelate, 6,6′-(2-aminoethylazanediyl)bis(methylene)dipicolinic acid (H2pentapa-en-NH2), has been synthesized and labeled with 99mTc with a specific activity of 135–140 MBq μmol−1 in >95% yield. The in vitro stability of the labeled chelate in both PBS and human serum shows only 85% yield and labeled with 99mTc in 96.2% radiochemical yield with a specific activity of 110–125 MBq μmol−1. The conjugate probe exhibited high serum stability (>94% at 24 h). The in vivo blood kinetic studies of radiocomplexes of H2pentapa-en-NH2 and its methionine conjugated derivative exhibited fast clearance with t1/2(F) = 32 ± 0.14 min, t1/2(S) = 4 h 20 min ± 0.21 min and t1/2(F) = 27 ± 0.3 min, t1/2(S) = 4 h 01 min ± 0.11 min, respectively. In vivo scintigraphy and ex vivo biodistribution studies in EAT tumor bearing mice demonstrated a high retention of H2pentapa-en-met2 at the site of the tumor with tumor to muscle ratio of 6.52 at 1 h, indicating the high specificity of 99mTc-pentapa-en-met2 toward tumors.