ABSTRACT
The lack of selectivity and low solubility of many chemotherapeutics impels the development of different biocompatible nanosized drug carriers. Amphiphilic block copolymers, composed of a hydrophilic and hydrophobic domain, show great potential because of their small size, large solubilizing power and loading capacity. In this paper, we introduce a new class of degradable temperature-responsive block copolymers based on the modification of N-(2-hydroxypropyl)methacrylamide (HPMA) with an ethyl group via a hydrolytically sensitive carbonate ester, polymerized by radical polymerization using a PEG-based macroinitiatior. The micellization and temperature-responsive behavior of the PEG-poly(HPMA-EC) block copolymer were investigated by dynamic light scattering (DLS). We observed that the polymer exhibits lower critical solution temperature (LCST) behavior and that above the cloud point (cp) of 17 °C the block copolymer self-assembles in micelles with a diameter of 40 nm. Flow cytometry analysis and confocal microscopy show a dose-dependent cellular uptake of the micelles loaded with a hydrophobic dye. The block copolymer nanoparticles were capable of delivering the hydrophobic payload into cancer cells in both 2D and 3D in vitro cultures. The block copolymer has excellent cytocompatibility, whereas loading the particles with the hydrophobic anticancer drug paclitaxel results in a dose-dependent decrease in cell viability.
Subject(s)
Acrylamides/chemistry , Polymers/chemistry , Animals , Antineoplastic Agents/chemistry , Antineoplastic Agents/pharmacology , Cell Line, Tumor , Cell Survival/drug effects , Drug Carriers/chemistry , Drug Delivery Systems/methods , Humans , Hydrolysis , Hydrophobic and Hydrophilic Interactions , MCF-7 Cells , Mice , Micelles , Nanoparticles/chemistry , Paclitaxel/chemistry , Paclitaxel/pharmacology , Particle Size , Polyethylene Glycols/chemistry , Solubility , TemperatureABSTRACT
We report the design and synthesis of a series of non-nucleoside MtbTMPK inhibitors (1-14) based on the gram-positive bacterial TMPK inhibitor hit compound 1. A practical synthesis was developed to access these analogues. Several compounds show promising MtbTMPK inhibitory potency and allow the establishment of a structure-activity relationship, which is helpful for further optimization.
Subject(s)
Antitubercular Agents/pharmacology , Mycobacterium tuberculosis/drug effects , Nucleoside-Phosphate Kinase/antagonists & inhibitors , Piperidines/pharmacology , Protein Kinase Inhibitors/pharmacology , Antitubercular Agents/chemical synthesis , Antitubercular Agents/chemistry , Dose-Response Relationship, Drug , Humans , Microbial Sensitivity Tests , Models, Molecular , Molecular Structure , Mycobacterium tuberculosis/enzymology , Nucleoside-Phosphate Kinase/metabolism , Piperidines/chemical synthesis , Piperidines/chemistry , Protein Kinase Inhibitors/chemical synthesis , Protein Kinase Inhibitors/chemistry , Structure-Activity RelationshipABSTRACT
We report on the design of a polymeric prodrug of the anticancer agent paclitaxel (PTX) by a grafting-from-drug approach. A chain transfer agent for reversible addition fragmentation chain transfer (RAFT) polymerization was efficiently and regioselectively linked to the C2'â position of paclitaxel, which is crucial for its bioactivity. Subsequent RAFT polymerization of a hydrophilic monomer yielded well-defined paclitaxel-polymer conjugates with high drug loading, water solubility, and stability. The versatility of this approach was further demonstrated by ω-end post-functionalization with a fluorescent tracer. Inâ vitro experiments showed that these conjugates are readily taken up into endosomes where native PTX is efficiently cleaved off and then reaches its subcellular target. This was confirmed by the cytotoxicity profile of the conjugate, which matches those of commercial PTX formulations based on mere physical encapsulation.
Subject(s)
Paclitaxel/analogs & derivatives , Paclitaxel/pharmacology , Polymers/chemistry , Polymers/pharmacology , Prodrugs/chemistry , Prodrugs/pharmacology , Cell Line, Tumor , Cell Survival/drug effects , Humans , Hydrophobic and Hydrophilic Interactions , Neoplasms/drug therapy , Paclitaxel/chemical synthesis , Polymerization , Polymers/chemical synthesis , Prodrugs/chemical synthesis , SolubilityABSTRACT
We report the evaluation of two alternative chemical dimerizer approaches aimed at increasing the sensitivity of MASPIT, a three-hybrid system that enables small-molecule target protein profiling in intact human cells. To circumvent the potential limitations related to the binding of methotrexate (MTX) to endogenous human dihydrofolate reductase (DHFR), we explored trimethoprim (TMP) as an alternative prokaryote-specific DHFR ligand. MASPIT evaluation of TMP fusion compounds with tamoxifen, reversine, and simvastatin as model baits, resulted in dose-response curves shifted towards lower EC50 values than those of their MTX congeners. Furthermore, a scalable azido-TMP reagent was synthesized that displayed a similar improvement in sensitivity, possibly owing to increased membrane permeability relative to the MTX anchor. Applying the SNAP-tag approach to introduce a covalent bond into the system, on the other hand, produced an inferior readout than in the MTX- or TMP-tag based assay.
Subject(s)
Indicators and Reagents/metabolism , Methotrexate/metabolism , Tetrahydrofolate Dehydrogenase/metabolism , Trimethoprim/chemistry , Trimethoprim/metabolism , Binding Sites , HEK293 Cells , Humans , Indicators and Reagents/chemical synthesis , Indicators and Reagents/chemistry , Ligands , Methotrexate/chemistry , Molecular Structure , Tetrahydrofolate Dehydrogenase/chemistry , Trimethoprim/chemical synthesisABSTRACT
In this study we report the synthesis of C5/C6-fused uridine phosphonates that are structurally related to earlier reported allosteric P2Y2 receptor ligands. A silyl-Hilbert-Johnson reaction of six quinazoline-2,4-(1H,3H)-dione-like base moieties with a suitable ribofuranosephosphonate afforded the desired analogues after full deprotection. In contrast to the parent 5-(4-fluoropheny)uridine phosphonate, the present extended-base uridine phosphonates essentially failed to modulate the P2Y2 receptor.
Subject(s)
Organophosphonates/chemical synthesis , Purinergic P2Y Receptor Agonists/chemical synthesis , Receptors, Purinergic P2Y2/metabolism , Uridine/chemical synthesis , Allosteric Regulation , Astrocytes/cytology , Astrocytes/drug effects , Astrocytes/metabolism , Cell Line, Tumor , Humans , Ligands , Organophosphonates/pharmacology , Purinergic P2Y Receptor Agonists/pharmacology , Quinazolines/chemistry , Uridine/analogs & derivatives , Uridine/pharmacologyABSTRACT
Two focused libraries based on two types of compounds, that is, thiazolidinediones and dioxazaborocanes were designed. Structural resemblances can be found between thiazolidinediones and well-known furanone type quorum sensing (QS) inhibitors such as N-acylaminofuranones, and/or acyl-homoserine lactone signaling molecules, while dioxazaborocanes structurally resemble previously reported oxazaborolidine derivatives which antagonized autoinducer 2 (AI-2) binding to its receptor. Because of this, we hypothesized that these compounds could affect AI-2 QS in Vibrio harveyi. Although all compounds blocked QS, the thiazolidinediones were the most active AI-2 QS inhibitors, with EC(50) values in the low micromolar range. Their mechanism of inhibition was elucidated by measuring the effect on bioluminescence in a series of V. harveyi QS mutants and by DNA-binding assays with purified LuxR protein. The active compounds neither affected bioluminescence as such nor the production of AI-2. Instead, our results indicate that the thiazolidinediones blocked AI-2 QS in V. harveyi by decreasing the DNA-binding ability of LuxR. In addition, several dioxazaborocanes were found to block AI-2 QS by targeting LuxPQ.
Subject(s)
Anti-Bacterial Agents/pharmacology , Boron Compounds/pharmacology , Quorum Sensing/drug effects , Thiazolidinediones/pharmacology , Vibrio/drug effects , Anti-Bacterial Agents/chemical synthesis , Anti-Bacterial Agents/chemistry , Boron Compounds/chemical synthesis , Boron Compounds/chemistry , Dose-Response Relationship, Drug , Microbial Sensitivity Tests , Molecular Structure , Quorum Sensing/genetics , Structure-Activity Relationship , Thiazolidinediones/chemical synthesis , Thiazolidinediones/chemistry , Vibrio/growth & developmentABSTRACT
Leishmaniasis causes high mortality and morbidity in tropical and subtropical regions of Africa, Asia, the Americas and southern Europe, and is characterized by diverse clinical manifestations. As a neglected tropical disease, limited resources are allocated for antileishmanial drug discovery. The Leishmania parasite is deficient in de novo purine synthesis, and therefore acquires purines from the host and processes these using a purine salvage pathway. By making use of purine transport systems and interfering with this salvage pathway, purine (nucleoside) analogues might exert a selective detrimental impact on its growth and survival. In vitro screening of an in-house purine nucleoside library and analogue synthesis afforded the 6-methyl-7-(2-pyridyl)-7-deazapurine ribonucleoside analogue 18 as a promising hit. Optimization of the 7-substituent afforded 31 and 32 which displayed potent activity against wild-type and resistant L. infantum, intracellular amastigote and extracellular promastigote forms, and favorable selectivity versus primary mouse macrophages (Mφ) and MRC-5 cells. Encouraged by the favorable in vitro metabolic stability of 32, an in vivo study was performed using an early curative L. infantum hamster model. When orally administrated at 50 mg/kg once daily (s.i.d) for 10 days, 32 was devoid of side effects, however, it only poorly reduced amastigote burdens in the major target organs.
Subject(s)
Antiprotozoal Agents , Leishmania , Leishmaniasis , Purines , Ribonucleosides , Animals , Antiprotozoal Agents/pharmacology , Antiprotozoal Agents/therapeutic use , Cricetinae , Leishmania/drug effects , Leishmania/metabolism , Leishmaniasis/drug therapy , Mice , Nucleosides/pharmacology , Nucleosides/therapeutic use , Purine Nucleosides/pharmacology , Purine Nucleosides/therapeutic use , Purines/pharmacology , Purines/therapeutic use , Ribonucleosides/pharmacology , Ribonucleosides/therapeutic useABSTRACT
Chagas disease and leishmaniasis are two poverty-related neglected tropical diseases that cause high mortality and morbidity. Current treatments suffer from severe limitations and novel, safer and more effective drugs are urgently needed. Both Trypanosoma cruzi and Leishmania are auxotrophic for purines and absolutely depend on uptake and assimilation of host purines. This led us to successfully explore purine nucleoside analogues as chemotherapeutic agents against these and other kinetoplastid infections. This study extensively explored the modification of the 6-amino group of tubercidin, a natural product with trypanocidal activity but unacceptable toxicity for clinical use. We found that mono-substitution of the amine with short alkyls elicits potent and selective antitrypanosomal and antileishmanial activity. The methyl analogue 15 displayed the best in vitro activity against both T. cruzi and L. infantum and high selectivity versus host cells. Oral administration for five consecutive days in an acute Chagas disease mouse model resulted in significantly reduced peak parasitemia levels (75, 89 and 96% with 12.5, 25 and 50 mg/kg/day, respectively). as well as increased animal survival rates with the lower doses (83 and 67% for 12.5 and 25 mg/kg/day, respectively).
Subject(s)
Chagas Disease , Leishmania , Trypanocidal Agents , Trypanosoma cruzi , Animals , Chagas Disease/drug therapy , Mice , Nucleosides/pharmacology , Purines/pharmacology , Purines/therapeutic use , Structure-Activity Relationship , Trypanocidal Agents/pharmacology , Trypanocidal Agents/therapeutic useABSTRACT
The kinetoplastid parasites Trypanosoma brucei, Trypanosoma cruzi and Leishmania spp. are the causative agents of neglected tropical diseases with a serious burden in several parts of the world. These parasites are incapable of synthesizing purines de novo, and therefore rely on ingenious purine salvage pathways to acquire and process purines from their host. Purine nucleoside analogs that may interfere with these pathways therefore constitute a privileged source of new antikinetoplastid agents. In this study, we synthetized a collection of C-nucleosides employing five different heterocyclic nucleobase surrogates. C-nucleosides are chemically and enzymatically stable and allow for extensive structural modification. Inspired by earlier 7-deazaadenosine nucleosides and known antileishmanial C-nucleosides, we introduced different modifications tailored towards antikinetoplastid activity. Both adenosine and inosine analogs were synthesized with the aim of discovering new antikinetoplastid hits and expanding knowledge of structure-activity relationships. Several promising hits with potent activity against Trypanosoma brucei, Trypanosoma cruzi and Leishmania infantum were discovered, and the nature of the nucleobase surrogate was found to have a profound influence on the selectivity profile of the compounds.
Subject(s)
Antiprotozoal Agents/pharmacology , Leishmania infantum/drug effects , Nucleosides/pharmacology , Trypanosoma brucei brucei/drug effects , Trypanosoma cruzi/drug effects , Antiprotozoal Agents/chemical synthesis , Antiprotozoal Agents/chemistry , Dose-Response Relationship, Drug , Molecular Structure , Nucleosides/chemical synthesis , Nucleosides/chemistry , Parasitic Sensitivity Tests , Structure-Activity RelationshipABSTRACT
Kinetoplastid parasites are the causative agents of Chagas disease (CD), leishmaniasis and human African trypanosomiasis (HAT). Despite a sustained decrease in the number of HAT cases, more efforts are needed to discover safe and effective therapies against CD and leishmaniasis. Kinetoplastid parasites lack the capability to biosynthesize purines de novo and thus critically depend on uptake and processing of purines from host cells. As such, modified purine nucleoside analogues may act as broad-spectrum antikinetoplastid agents. This study assessed the in vitro activity profile of 7-modified 6-methyl tubercidin derivatives against Trypanosoma cruzi, Leishmania infantum, Trypanosoma brucei brucei and T. b. rhodesiense, and led to the identification of analogues that display activity against all these species, such as 7-ethyl (13) and 7-chloro (7) analogues. These selected analogues also proved sufficiently stable in liver microsomes to warrant in vivo follow-up evaluation.
Subject(s)
Antiprotozoal Agents , Trypanosoma brucei brucei , Trypanosoma cruzi , Antiprotozoal Agents/pharmacology , Humans , Nucleosides/pharmacology , Purines , Structure-Activity RelationshipABSTRACT
Chagas disease is a tropical infectious disease resulting in progressive organ-damage and currently lacks efficient treatment and vaccine options. The causative pathogen, Trypanosoma cruzi, requires uptake and processing of preformed purines from the host because it cannot synthesize these de novo, instigating the evaluation of modified purine nucleosides as potential trypanocides. By modifying the pyrimidine part of a previously identified 7-aryl-7-deazapurine nucleoside, we found that substitution of a 6-methyl for a 6-amino group allows retaining T. cruzi amastigote growth inhibitory activity but confers improved selectivity towards mammalian cells. By keeping the 6-methyl group unaltered, and introducing different 7-aryl groups, we identified several analogues with sub-micromolar antitrypanosomal activity. The 7-(4-chlorophenyl) analogue 14, which was stable in microsomes, was evaluated in an acute mouse model. Oral administration of 25â mg/kg b.i.d. suppressed peak parasitemia and protected mice from infection-related mortality, gave similar reductions as the reference drug of blood parasite loads determined by qPCR, but as benznidazole failed to induce sterile cure in the short time period of drug exposure (5â days).
Subject(s)
Nucleosides/pharmacology , Purines/pharmacology , Trypanocidal Agents/pharmacology , Trypanosoma cruzi/drug effects , Dose-Response Relationship, Drug , Molecular Structure , Nucleosides/chemical synthesis , Nucleosides/chemistry , Parasitic Sensitivity Tests , Purines/chemical synthesis , Purines/chemistry , Structure-Activity Relationship , Trypanocidal Agents/chemical synthesis , Trypanocidal Agents/chemistryABSTRACT
Chagas disease and visceral leishmaniasis are two neglected tropical diseases responsible for numerous deaths around the world. For both, current treatments are largely inadequate, resulting in a continued need for new drug discovery. As both kinetoplastid parasites are incapable of de novo purine synthesis, they depend on purine salvage pathways that allow them to acquire and process purines from the host to meet their demands. Purine nucleoside analogues therefore constitute a logical source of potential antiparasitic agents. Earlier optimization efforts of the natural product tubercidin (7-deazaadenosine) involving modifications to the nucleobase 7-position and the ribofuranose 3'-position led to analogues with potent anti-Trypanosoma brucei and anti-Trypanosoma cruzi activities. In this work, we report the design and synthesis of pyrazolo[3,4-d]pyrimidine nucleosides with 3'- and 7-modifications and assess their potential as anti-Trypanosoma cruzi and antileishmanial agents. One compound was selected for in vivo evaluation in an acute Chagas disease mouse model.
Subject(s)
Nucleosides/therapeutic use , Pyrazoles/therapeutic use , Pyrimidines/therapeutic use , Trypanocidal Agents/therapeutic use , Trypanosoma cruzi/drug effects , Animals , Chagas Disease/drug therapy , Drug Design , Drug Stability , Humans , Leishmania infantum/drug effects , Male , Mice , Microsomes, Liver/metabolism , Molecular Structure , Nucleosides/chemical synthesis , Nucleosides/pharmacology , Pyrazoles/chemical synthesis , Pyrazoles/pharmacology , Pyrimidines/chemical synthesis , Pyrimidines/pharmacology , Structure-Activity Relationship , Trypanocidal Agents/chemical synthesis , Trypanocidal Agents/pharmacologyABSTRACT
An efficient protocol for the construction of C-6-(hetero)aryl-substituted uridine phosphonate analogues utilizing an aerobic, ligand-free Suzuki-Miyaura cross-coupling reaction of a 6-iodo-precursor in aqueous media has been established. The method presents a modular approach toward the target compounds as demonstrated by the synthesis of a small library comprising 14 novel nucleoside phosphonates.
Subject(s)
Carbon/chemistry , Models, Chemical , Organophosphonates/chemical synthesis , Oxygen/chemistry , Uridine/chemical synthesis , Alkenes/chemistry , Boronic Acids/chemistry , Hydrogenation , Indicators and Reagents , Ligands , Magnetic Resonance Spectroscopy , Organophosphonates/chemistry , Oxidation-Reduction , Temperature , Uridine/chemistryABSTRACT
The aqueous methanolic leaf extract of Fargesia robusta var. Pingwu was evaluated in vitro for its antioxidant capacity using the TEAC and ORAC assays. C-Glycosyl flavones, farobin A (1) and farobin B (2), together with three known compounds, tricin-5-O-glucopyranoside (3), 2''-O-α-rhamnosyl-6-C-(6-deoxy-ribo-hexos-3-ulosyl)luteolin (4), and luteolin-6-C-glucopyranoside (homoorientin) (5), were isolated from the hydroalcoholic extract of the leaves of F. robusta. The structures of the compounds were determined by spectroscopic analyses including UV, 1D and 2D NMR, and MS. Compounds 1, 4, and 5 exhibited potent antioxidant activity in the TEAC assay, while compounds 1, 3, and 5 showed the highest antioxidant capacity in the ORAC assay.
Subject(s)
Antioxidants/isolation & purification , Antioxidants/pharmacology , Flavones/isolation & purification , Flavones/pharmacology , Glycosides/isolation & purification , Glycosides/pharmacology , Sasa/chemistry , Antioxidants/chemistry , Belgium , Flavones/chemistry , Glycosides/chemistry , Molecular Structure , Nuclear Magnetic Resonance, Biomolecular , Plant Leaves/chemistryABSTRACT
Acid-sensitive paclitaxel (PTX)-polymer conjugates were designed by applying a grafting-from-drug RAFT approach. PTX was linked through either a cyclic or a linear, acid-sensitive acetal moiety. Relative to direct esterification of PTX, which occurred regioselectively at the C2' OH-group, direct acetalization was observed at either the C2' or the C7 OH-group of PTX. This yielded two regioisomers of acetal-based PTX-functionalized RAFT chain transfer agents (CTAs). Subsequent polymerization with N,N-dimethylacrylamide (DMA) resulted in amphiphilic highly defined, acetal-based PTX-polymer conjugates with nearly identical features in terms of polymer definition and micellar self-assembly behavior, but with distinct PTX release kinetics and absence of burst release. This was further reflected by their in vitro biological performance, giving insights into the difference of the release mechanism between ester- and acetal-based PTX-polymer conjugates.
ABSTRACT
Chloroform extracts of the underground parts of two Balkan endemic Laserpitium species, Laserpitium zernyi Hayek and Laserpitium ochridanum Micevski, were chemically investigated. Five unknown guaianolides from the class of slovanolides, of which four were additionally 2ß-esterified, as well as two lactones, previously identified in other Laserpitium species, were isolated from the L. ochridanum extract. From the L. zernyi extract one slovanolide derivative was isolated for the first time in the genus Laserpitium. In addition, the phenylpropanoid latifolone and six known sesquiterpene lactones, characterised as derivatives of slovanolide and silerolide, were isolated from the extracts of both species. The cytotoxic activities of the total extracts and the isolated compounds were tested using MTT and SRB assays on the two human breast cancer cell lines, MCF 7/6 and MCF 7/AZ. The extracts exerted cytotoxic activities with the IC(50) values ranging 65.21-348.25 µg/mL. The L. ochridanum extract was most potent in the MTT test with IC(50) values of 65.21 and 66.09 µg/mL in the MCF 7/AZ and MCF 7/6 cell lines, respectively. The highest cytotoxic activity exerted 2ß,8α-di-angeloyloxy-10ß-hydroxy-6αH-guaian-3,(7-11)-dien-12,6-olide, a slovanolide derivative with an additional double bond in lactone ring, on highly invasive MCF 7/6 cell line, with IC(50) value 0.7 µM in both assays tested. Generally, guaianolides with a higher number of ester moieties at the positions 2ß, 8α, 10ß or 11α exhibited IC(50) values in the micromolar range, while eudesmanolides and guaianolides with a lower number of esters did not induce significant cytotoxicity.