Synthesis, characterization, and radiosynthesis of fluorine-18-AVT-011 as a Pgp chemoresistance imaging marker.

P-glycoprotein (Pgp) is the most studied ATP-binding cassette (ABC) efflux transporter and contributes to chemoresistance. A few tracers have been developed to detect the in-vivo status of chemoresistance using positron emission tomography (PET) imaging. In our study, we have synthesized labeled AVT-011 with fluorine-18 (18F) followed by in-vitro and in-vivo analysis. Tosylate AVT-011 precursor was synthesized and characterized by 1H-NMR and 13C-NMR. AVT-011 was labeled with 18F using the nucleophilic substitution method, and a standard set of quality control was performed. The specificity for Pgp was tested in U87MG cells with and without an inhibitor (tariquidar). The biodistribution and in-vivo stability were tested in the small animals (mice). The biodistribution data of [18F]-AVT-011 was extracted from the PET-CT imaging of breast cancer patients (n = 6). The precursor was synthesized with 36 ± 4% yield and 97 ± 2% purity. The labeling was more than 95% with a 42 ± 2% yield, as evaluated by Radio-HPLC. The cell-binding assay showed a specificity of the tracer for Pgp as the uptake increased by twice after blocking the Pgp receptors. The radiotracer showed a hepatorenal excretion pathway for clearance in an animal study. The uptake was higher in the liver, lungs, spleen, and heart at 15 min and decreased at 60 min. The patients' distribution showed similar uptake patterns as observed in the small animals. [18F]AVT-011 was characterized successfully with high radiochemical purity and yield. The in-vitro and in-vivo studies proved its specificity for Pgp and safe for patient use.

Synthesis and biological evaluation of coumarin-thiazole hybrids as selective carbonic anhydrase IX and XII inhibitors.

A series of coumarin-linked thiazoles (6a-p) was synthesized and the synthesized compounds were evaluated against human carbonic anhydrases (hCAs) IX and XII, which have been implicated in cancer. All the compounds exhibited selective inhibition of both isoforms. The designed compounds inhibited hCA IX in a moderate nanomolar to submicromolar range. The hCA XII was inhibited in a low to moderate nanomolar range. Compound 6o exhibited the best inhibition of hCA XII with a Ki value of 91.1 nM. The hydrolyzed form of compound 6o also exhibited favorable interactions as well as good docking scores with both the isoforms. Hence, this compound can be taken as a template for the design of selective and potent hCA XII inhibitors.

Automated Radiosynthesis and Molecular Docking Studies of Coumarin- Triazole Hybrid with fluorine-18: A feasibility study.

BACKGROUND: Fluorine-18 is one of the promising radiotracers that can report target specific information related to its physiology to understand the disease status through the PET modality. In the current study, the radiochemical synthesis, purification, and molecular docking studies of fluorine-18 (18F) radiolabeled coumarin-triazole hybrid have been performed. OBJECTIVE: To develop target specific fluorine-18 radiotracer for the diagnosis in oncology. METHODS: GE Tracer-lab FX2N module with few modifications in the line connections was used for the radiosynthesis and purification of target molecule [18F]SG-2, 4-((2,6-dimethylmorpholino) methyl)-7-((1-(4-(fluoro-18F) benzyl)-1H-1,2,3-triazol-4-yl) oxy)-2H-chromen-2-one, through the nucleophilic radiofluorination mechanism. The radiochemical purity was measured by HPLC, and TLC analytical methods. The kryptofix levels were also evaluated by using the TLC method. The residual solvents like DMF, ethanol were measured using GC. The Schrödinger drug discovery suite 2018 was used to study the protein and ligand interactions. RESULTS: The quality control parameters revealed the purity, chemical identity, and limits of residual solvents. The radiochemical purity was 95.5 ± 2.3%, and dimethylformamide solvent limit was 89 ± 3 ppm. The molecular docking results had suggested that the cold target molecule has made strong electronic interactions and showed the possible pharmacokinetic (ADME) properties with galectin-1 protein. Overall, these results showed that [[18F]SG-2 radiolabeling with 18F radionuclide was feasible, and support of molecular docking studies suggest possible interactions with Galectin- 1. CONCLUSION: we reported a feasibility study for labeling coumarin-triazole hybrid with fluorine-18 through aromatic nucleophilic fluorination reaction (SNAr).

Synthesis and biological evaluation of novel 4,7-disubstituted coumarins as selective tumor-associated carbonic anhydrase IX and XII inhibitors.

With an aim to develop novel potential anti-cancer agents we have designed a series of novel 4,7-disubstituted coumarin hybrids synthesis and evaluated for their inhibitory activity against the human carbonic anhydrase isoforms namely CA I, CA II, CA IX and CA XII. The results of CA inhibition clearly showed that the novel 4, 7-disubstituted coumarin hybrids (7a-i & 8a-j) exhibited selective inhibition towards tumor associated isoforms, CA IX and CA XII without inhibiting CA I and CA II isoforms. Among all the compound 8b showed a significant inhibition against hCA IX with a Ki of 0.58 µM whereas, the compound 7c showed a significant inhibition against hCA XII with a Ki of 0.36 µM respectively. All other compounds have shown a good inhibition against hCA IX and hCA XII over hCA I and hCA II within the range of 0.46 to 9.35 µM. Therefore, compound 8b and 7c would be the potential leads for developing selective cytotoxic agents targeting hCA IX and XII.

Human Galectin-1 in Multiple Cancers: A Privileged Molecular Target in Oncology.

Galectin-1 (Gal-1), a 14kDa carbohydrate-binding protein of the galectin family found in humans, affects intracellular signaling pathways upon interaction with ß-galactosides on cell-surface, cytosol, and nucleus. The structural information reveals that it consists of a monovalent dimer composed of subunits with one Carbohydrate Recognition Domain (CRD), which is the main active site to interact with various glycoproteins, and carbohydrates in the body to regulate cellular functions. Gal-1 contributes towards different events associated with cancer biology, including tumor transformation, cell cycle regulation, apoptosis, cell adhesion, migration, and inflammation. The extracellular existence and function of Gal-1 have been well-established, and it is known to express in many tumor types, including astrocytoma, melanoma, prostate, colon, bladder, and ovarian carcinomas, etc. Several studies suggested the upregulation of Gal-1 levels in multiple cancer cells. Thus, Gal-1 is a promising molecular target for the development of new therapeutic tools. The present review focuses on the functions of Gal-1 in tumor progression in multiple cancers and its structural insights.

Carbon-11: Radiochemistry and Target-Based PET Molecular Imaging Applications in Oncology, Cardiology, and Neurology.

The positron emission tomography (PET) molecular imaging technique has gained its universal value as a remarkable tool for medical diagnosis and biomedical research. Carbon-11 is one of the promising radiotracers that can report target-specific information related to its pharmacology and physiology to understand the disease status. Currently, many of the available carbon-11 (t1/2 = 20.4 min) PET radiotracers are heterocyclic derivatives that have been synthesized using carbon-11 inserted different functional groups obtained from primary and secondary carbon-11 precursors. A spectrum of carbon-11 PET radiotracers has been developed against many of the upregulated and emerging targets for the diagnosis, prognosis, prediction, and therapy in the fields of oncology, cardiology, and neurology. This review focuses on the carbon-11 radiochemistry and various target-specific PET molecular imaging agents used in tumor, heart, brain, and neuroinflammatory disease imaging along with its associated pathology.

Radiosynthesis challenges of 11C and 18F-labeled radiotracers in the FX2C/N tracerlab and their validation through PET-MR imaging.

Glucose is the renowned source of the energy for the cancer growth, that's the reason for [18F]FDG success and make it widely used radiotracer. Though [18F]FDG has its own inherent limitations therefore many tracers have been developed to target specific receptors, and other metabolic routes. We have used FX2C and FX2N Tracerlab modules for the synthesis of the [11C]methionine, [18F]choline and [18F]fluorodopa via nucleophilic pathway in FX2C/N module. [11C]methionine was standardized in FX2C module using two different precursors, and purified using C18 cartridge based technique. [18F]methylcholine was synthesized using dimethylaminoethanol precursor and purified using cartridge-based method. [18F]fluorodopa was synthesized using nucleophilic precursor and purified using in-built preparative HPLC on FX2N module. All radioactive intermediates and chemical impurities were evaluated by analytical HPLC. The radiochemical purity of D and L-[11C]methionine were 4.6 ± 3.2% and 95.4 ± 3.6% while other chemical impurities were less than prescribed limits with yield of 20 ± 5%. [18F]fluoromethylcholine was prepared with high radiochemical purity of 97.3 ± 2.6% with yield of 8 ± 3%. [18F]fluorodopa was synthesized with high radiochemical purity of 95.8 ± 1.4% with 15 ± 3% yield. The adaptation of [18F]fluorodopa synthesis to FX2N module via designing synthesis sequence and purified through on-line HPLC has provided high radiochemical purity. PET-MR imaging was done using these tracers which have validated the synthesis and their availability for future clinical applications.

Recent Developments of Target Based Coumarin Derivatives as Potential Anticancer Agents.

Cancer is the second life-threatening disease worldwide, and it resulted in around 9.6 million deaths globally in 2018. The multidrug-resistant cancers and non-selectivity create an urge for the development of novel anticancer drugs with a diverse mechanism of action. Coumarin is one of the most widely used scaffolds for the development of highly effective anticancer agents. It has versatile anticancer profiles with diverse mechanisms to inhibit tumor progression. The facile synthetic strategies are also favoured for target-based coumarin derivatives. Structural Activity Relationship (SAR) studies determine anticancer potentials with minimal side effects through the substitution pattern of coumarin. A number of coumarin derivatives have been developed against the Galectin-1 (Gal-1), Carbonic anhydrases (CAs), Tubulin protein, and other essential targets for the treatment in cancer therapy. Therefore, in this review, we have mainly focused on different target based coumarin derivatives, and synthetic strategies.

Synthesis, 18F-radiolabeling and apoptosis inducing studies of novel 4, 7-disubstituted coumarins.

In present study, a new series of 4, 7-disubstituted coumarin derivatives (7a-y) have been synthesized as galectin-1 targeting apoptosis inducing agents and evaluated for their in vitro cytotoxic potentials against a panel of selected human cancer cell lines namely, Brest (MCF7), Ovarian (SKOV3), Prostate (PC-3 & DU145) and normal embryonic kidney (HEK293T) cells, using MTT assay. Most of the compounds exhibited potent growth inhibitory action against the treated cancer cell lines with an IC50 range of 10-30 µM. Compound 7q exhibited a significant growth inhibition against prostate cancer (PC-3 & DU145) cell lines with an IC50 value of 7.45 ± 0.03 µM, 8.95 ± 0.17 µM respectively. Further, the target compound 7q was radiolabeled with fluorine-18 [18F] to be used as a novel PET radiotracer for imaging of tumors via targeting galectin-1, using appropriate reaction conditions in the GE Tracer-lab FX2N synthesis module. The purification of the [18F] radiolabeled compound [18F]-7q was successfully achieved with 60% ethanol. The radiochemical purity was>85% and residual solvent limits of DMF was 65 ± 3 ppm as analysed by HPLC, TLC & GC analytical methods. The apoptosis studies confirm the inhibition of cell proliferation with morphological changes like cell shrinkage, blebbing and cell wall deformation, increasing the ROS levels, and loss of mitochondrial membrane potential by Acridine orange/Ethidium bromide staining, Hoechst-33342 staining, H2DCFDA staining, annexin V-FITC/PI, and JC-1 staining methods. In flow cytometric analysis, 7q selectively arrested the sub-G1 phase of the cell cycle in a dose-dependent manner. In Gal-1 ELISA studies, compound 7q efficiently reduced the levels of Gal-1 protein in dose-dependent manner with an IC50 value of 100 µM. The binding constant (Ka) of 7q with Gal-1 was observed as 1.3 × 104 M-1 by fluorescence spectroscopy. The molecular docking studies clearly showed possible interactions and the pharmacokinetic (ADMET) properties of compound 7q with Gal-1. Hence, the novel 4, 7-disubstituted coumarins could be a potential cytotoxic and PET imaging agents via Gal-1.

Fluorine-18: A radionuclide with diverse range of radiochemistry and synthesis strategies for target based PET diagnosis.

Fluorine-18 is one of the most widely used radionuclides for the production of radiopharmaceuticals for positron emission tomography (PET). The radiolabeling methods like nucleophilic, electrophilic, Cu mediated mechanisms or prosthetic groups are widely using to achieve high regioselective radiochemical yields. It acts as a powerful tool to identify new drug targets through cellular uptake, pharmacokinetic (ADME) and pharmacodynamic parameters of the 18F labeled tracer or drug. These PET tracers have been developed based on the receptors expressed in a disease condition. A number of 18F radiotracers have been developed against the Tropomyosin Receptor Kinases (Trks), Carbonic anhydrases (CAs), Epidermal Growth Factor Receptors (EGFR), Poly ADP ribose polymerase (PARP) etc. for the diagnosis in cancer therapy. The current research also focused on the development of novel 18F radiotracers for neurological conditions for deciphering underlying physiology in diseases like Alzheimer, and Parkinson. Therefore, in this review we have focused on 18F labeling methods, and radiotracers developed against common cancers and neurological conditions.

Synthesis of 1-benzyl-1H-benzimidazoles as galectin-1 mediated anticancer agents.

In our pursuit to develop novel non-carbohydrate small molecule Galectin-1 Inhibitors, we have designed a series of 1-benzyl-1H-benzimidazole derivatives and demonstrated their anticancer activity. The compound 6g, 4-(1-benzyl-5-chloro-1H-benzo[d]imidazol-2-yl)-N-(4-hydroxyphenyl) benzamide was found to be most potent with an IC50 of 7.01 ±â€¯0.20 µM and arresting MCF-7 cell growth at G2/M phase and S phase. Induction of apoptosis was confirmed by morphological changes like cell shrinkage, blebbing and cell wall deformation, dose dependent increase in the mitochondrial membrane potential (ΔΨm) and ROS levels. Further, dose dependent decrease in Gal-1 protein levels proves Gal-1 mediated apoptosis by 6g. Molecular docking studies were performed to understand the Gal-1 interaction with compound 6g. In addition, RP-HPLC studies showed 85.44% of 6g binding to Gal-1. Binding affinity studies by fluorescence spectroscopy and Surface Plasmon Resonance (SPR) showed that 6g binds to Gal-1 with binding constant (Ka) of 1.2 × 104 M-1 and equilibrium constant KD value of 5.76 × 10-4 M respectively.

Synthesis and biological evaluation of morpholines linked coumarin-triazole hybrids as anticancer agents.

A series of novel morpholines linked coumarin-triazole hybrids (6a-6v) has been synthesized and evaluated for their anti-proliferative potential on a panel of five human cancer cell lines, namely bone (MG-63), lung (A549), breast (MDA-MB-231), colon (HCT-15) and liver (HepG2), using MTT assay. Among all, the compound 6n {7-((1-(2,4-dichlorobenzyl)-1H-1,2,3-triazol-4-yl) methoxy)-4-((2,6-dimethylmorpholino) methyl)-2H-chromen-2-one} showed significant growth inhibition against MG-63 cells with an IC50 value of 0.80 ± 0.22 µM. Further, induction of apoptosis by 6n of MG-63 cells confirmed as a result of morphological changes, the sub-G1 phase arrest, increased percentage of apoptotic cells, and decrease in mitochondrial membrane potential and increase in reactive oxygen species levels. The in vitro Gal-1 expression in cell culture supernatant of MG-63 cells treated with compound 6n showed dose-dependent reduction. The binding constant (Ka ) of 6n with Gal-1 was calculated from the intercept value which was observed as 3.0 × 105  M-1 by fluorescence spectroscopy. Surface plasmon resonance showed that 6n binds to Gal-1 with binding constant (Ka ) of 1.29E+04 1/Ms and equilibrium constant KD value of 7.54E-07 M, respectively. Molecular docking studies revealed the binding interactions of 6n with Gal-1.