Recent advances in the exploration of oxazolidinone scaffolds from compound development to antibacterial agents and other bioactivities.

Bacterial infections cause a variety of life-threatening diseases, and the continuous evolution of drug-resistant bacteria poses an increasing threat to current antimicrobial regimens. Gram-positive bacteria (GPB) have a wide range of genetic capabilities that allow them to adapt to and develop resistance to practically all existing antibiotics. Oxazolidinones, a class of potent bacterial protein synthesis inhibitors with a unique mechanism of action involving inhibition of bacterial ribosomal translation, has emerged as the antibiotics of choice for the treatment of drug-resistant GPB infections. In this review, we discussed the oxazolidinone antibiotics that are currently on the market and in clinical development, as well as an updated synopsis of current advances on their analogues, with an emphasis on innovative strategies for structural optimization of linezolid, structure-activity relationship (SAR), and safety properties. We also discussed recent efforts aimed at extending the activity of oxazolidinones to gram-negative bacteria (GNB), antitumor, and coagulation factor Xa. Oxazolidinone antibiotics can accumulate in GNB by a conjugation to siderophore-mediated ß-lactamase-triggered release, making them effective against GNB.

The N-Alkylation of Agelastatin A Modulates Its Chemical Reactivity.

Agelastatin A is a marine alkaloid with potent biological activity. To date, at least 17 different strategies have achieved its total synthesis, along with many analogues. The present study focuses on the acidity stability of some N-methyl derivatives of agelastatin A. The study made use of chemical reactions and spectroscopic acquisitions. The chemical structure of some derivatives can undergo a profound rearrangement. The results could shed light on the mechanism of action of agelastatin A and suggest the preparation of analogues with improved pharmacological efficacy.

Synthesis of the oxazolidinone fragment of thelepamide.

Thelepamide, an unique ketide-amino acid isolated from a marine annelid worm Thelepus crispus, has a unique oxazolidinone ring derived from cysteine, glycine and valine. Rareness in nature as well as promising bioactive possibility make the oxazolidinone ring an attractive synthetic target. The hydroxy oxazolidinone fragment of thelepamide was prepared by acid-catalysed N,O-acetal formation between a ketoamide and formaldehyde. Lactone-carbonyl selective isopropyl addition to an oxazilidine-dione under Grignard conditions also forms the target compound.

Simultaneous determination of a novel oxazolidinone anti-tuberculosis OTB-658 and its metabolites in monkey blood by LC-MS/MS.

OTB-658, a novel oxazolidinone anti-tuberculosis agent, has potent antibacterial activity against Mycobacterium tuberculosis, especially multi-drug resistant tuberculosis (MDR-TB) in vitro and in vivo. In this study, after metabolite identification of parent drug OTB-658, a specific and sensitive liquid chromatography-tandem mass spectrometry (LC-MS/MS) method was established and validated to quantify OTB-658 and its metabolites OTB-665 and OTB-698 in monkey blood. HHY-1442, an analogue compound of OTB-658, was used as the internal standard. Blood samples were prepared by direct protein precipitation. Separation was performed on a Zorbax SB C18 column (50 mm × 2.1 mm, 3.5 µm) with a gradient mobile phase of methanol/water at a flow rate of 0.3 mL/min. The detection was conducted by a positive electrospray ionization in multiple-reaction monitoring mode on a triple quadrupole MS. The monitored transitions were m/z 382.2 â†’ 221.1 for OTB-658, m/z 398.2 â†’ 308.1 for OTB-665, m/z 414.1 â†’ 372.3 for OTB-698 and m/z 418.2 â†’ 311.3 for HHY-1442, respectively. Good linearity was observed over the range of 10-2000 ng/mL for OTB-658 and OTB-665, and 5-1000 ng/mL for OTB-698. All the intra-day and inter-day precision for the three analytes was below 8.4%, and the accuracy ranged from 96.0% to 106.0%. All analytes were stable during storage, preparation, and analytical procedures. The validated method was successfully applied to pharmacokinetic and bioavailability studies of OTB-658 in cynomolgus monkeys and the absolute bioavailability of OTB-658 was 25.0% at an oral dose of 10 mg/kg.

Oxazolidinone-containing Hybrids with Antibacterial Activity against Methicillin-resistant Staphylococcus aureus (MRSA): A Mini-review.

The increasing danger of methicillin-resistant Staphylococcus aureus (MRSA) and the limited therapeutic options for invasive MRSA infections make an urgent demand for the development of novel anti-MRSA agents. Oxazolidinone derivatives could inhibit protein synthesis by acting on the ribosomal 50S subunit of the bacteria and prevent the formation of a functional 70S initiation complex, so oxazolidinones are a novel class of antimicrobial agents with potential activity against a wide range of clinically significant multidrug-resistant Gram-positive pathogens. However, oxazolidinones such as linezolid are associated with significant adverse events, and myelosuppression represents the main unfavorable side effects. Moreover, MRSA isolates that are resistant to oxazolidinones have already emerged. Hybridization of oxazolidinone with other antibacterial pharmacophores has the potential to interact with multiple targets or to counterbalance the known side effects associated with each pharmacophore. Thus, oxazolidinone-containing hybrids are useful scaffolds for the development of novel anti-MRSA agents. This review covers the recent advances of oxazolidinonecontaining hybrids with anti-MRSA activity developed in the last decade to set up the direction for the design and development of oxazolidinone-containing hybrids with high efficiency and low toxicity.

Anti-progressive Effects of a Series of Glycinyl and Alaninyl Triazolyl-oxazolidinones on Kelly Neuroblastoma Cell Line.

BACKGROUND/AIM: Neuroblastoma (NB), the most common extracranial malignant childhood tumor accounts for about 15% of cancer-related deaths in children. Despite the intensive treatment of patients with high-risk scarification of NB, clinical outcomes indicate tumor recurrence greater than 50% and late severe adverse effects. Oxazolidinones are 5-membered heterocyclic compounds with antibacterial activity against resistant bacterial strains. Structural modifications around the oxazolidinone moiety have resulted in derivatives with anti-cancer properties against proliferation, motility, and invasion of breast cancer cells. This study aimed to examine the anti-cancer potential of novel oxazolidinones against a model of a neuroblastoma cell line. MATERIALS AND METHODS: Newly synthesized and characterized triazolyl-oxazolidinone derivatives were incubated with neuroblastoma Kelly cells. The anti-proliferation and anti-progression effects of the compounds were evaluated by MTT, and adhesion with migration assays. RESULTS: The 5-nitrofuroyl glycinyl-oxazolidinone containing 4-methyltriazolyl group demonstrated the most potent activity with an IC50=6.52 µM. Furthermore, the D-isomer of 5-nitrothiophenecarbonyl alaninyl containing derivative reduced the adhesion to fibronectin by 56.34%, while the D-isomer of 5-nitrofuroyl alaninyl derivative reduced the migration of Kelly cells by 29.14%. CONCLUSION: The presence of the 4-methyltriazolyl moiety seems to enhance the anti-proliferative property of triazolyl-oxazolidinone derivatives, as demonstrated by PH-145. There is little or no effect of the stereochemistry of the alanine side-chain on the antiproliferative effect, as demonstrated by the 5-nitrofuroyl D- and L-alaninyl containing derivatives with similar IC50 values. The observed differences in the inhibition of adhesion and migration by the oxazolidinones on Kelly cells provide a new therapeutic approach that needs further investigation.

Discovery of a Conformationally Constrained Oxazolidinone with Improved Safety and Efficacy Profiles for the Treatment of Multidrug-Resistant Tuberculosis.

Tuberculosis (TB) remains a serious public health challenge, and the research and development of new anti-TB drugs is an essential component of the global strategy to eradicate TB. In this work, we discovered a conformationally constrained oxazolidinone 19c with improved anti-TB activity and safety profile through a focused lead optimization effort. Compound 19c displayed superior in vivo efficacy in a mouse TB infection model compared to linezolid and sutezolid. The druggability of compound 19c was demonstrated in a panel of assays including microsomal stability, cytotoxicity, cytochrome P450 enzyme inhibition, and pharmacokinetics in animals. Compound 19c demonstrated an excellent safety profile in a battery of safety assays, including mitochondrial protein synthesis, hERG K+, hCav1.2, and Nav1.5 channels, monoamine oxidase, and genotoxicity. In a 4 week repeated dose toxicology study in rats, 19c appeared to have less bone marrow suppression than linezolid, which has been a major liability of the oxazolidinone class.

Synthesis and Structure-Activity Relationships of N-(4-Benzamidino)-Oxazolidinones: Potent and Selective Inhibitors of Kallikrein-Related Peptidase 6.

Kallikrein-related peptidase 6 (KLK6) is a secreted serine protease that belongs to the family of tissue kallikreins. Aberrant expression of KLK6 has been found in different cancers and neurodegenerative diseases, and KLK6 is currently studied as a potential target in these pathologies. We report a novel series of KLK6 inhibitors discovered in a high-throughput screen within the European Lead Factory program. Structure-guided design based on docking studies enabled rapid progression of a hit cluster to inhibitors with improved potency, selectivity and pharmacokinetic properties. In particular, inhibitors 32 ((5R)-3-(4-carbamimidoylphenyl)-N-((S)-1-(naphthalen-1-yl)propyl)-2-oxooxazolidine-5-carboxamide) and 34 ((5R)-3-(6-carbamimidoylpyridin-3-yl)-N-((1S)-1-(naphthalen-1-yl)propyl)-2-oxooxazolidine-5-carboxamide) have single-digit nanomolar potency against KLK6, with over 25-fold and 100-fold selectivities against the closely related enzyme trypsin, respectively. The most potent compound, 32, effectively reduces KLK6-dependent invasion of HCT116 cells. The high potency in combination with good solubility and low clearance of 32 make it a good chemical probe for KLK6 target validation in vitro and potentially in vivo.

In vitro anti-glioblastoma activity of L-valine derived boroxazolidones.

In the present study, a series of L-valine derived boroxazolidones, previously synthesized and reported to have residual activity in a human epithelial cell line, have been evaluated in vitro for their anti-glioblastoma activity. A boroxazolidone derivative containing 2,4-difluorophenyl moieties (6) was found to have higher cytotoxicity than the standard drug, Temozolomide (TMZ). Compound 6 was found to exhibit dose-dependent growth inhibitory effects with an IC50 of 49 µM and 53 µM for LN229 and SNB19 cells, respectively. Additionally, 6 was assessed for its role in apoptosis, caspase 3/7 activation and oxidative stress in SNB19 and LN229 cells. SNB19 cells treated with 6 showed 45.3% apoptosis in the population, while TMZ had 24.7%. In LN229 cells, the percentage of apoptotic cells treated with compound 6 and TMZ were the same. Both 6 and TMZ induced apoptosis through the activation of caspase 3/7 in SNB19 and LN229 cells. Interestingly, 6 exhibited a higher effectivity in promoting reactive oxygen species production in LN229, while it was 6-fold less in SNB19. Boroxazolidone-treated GBM cell lines increased reactive oxygen species production, suggesting that such species may be interlinked with the observed programmed cell death. Additionally, the treatment of both GBM cell lines with 6 led to G2/M phase arrest. The magnitude of anti-GBM effect of 6 is significantly higher than the known chemotherapeutic agent TMZ. This work further demonstrates the anticancer properties of L-valine derived boroxazolidones, adding another potential derivative to the collection of promising chemotherapeutic agents for GBM treatment.

Probing the gas-phase structure of charge-tagged intermediates of a proline catalyzed aldol reaction - vibrational spectroscopy distinguishes oxazolidinone from enamine species.

An l-proline based catalyst with a charged phenyl-pyridium substituent (1) was used to analyze intermediates of an organocatalyzed aldol reaction by infrared multi-photon dissociation (IRMPD) mass spectrometry after transfer into the gas phase via electrospray ionization (ESI). IRMPD spectra were interpreted with the aid of density functional theory (DFT) computations. A structurally restricted enamine species was used as a reference molecule for the calculated vibrational frequencies. A close correlation between theory and experiment was found for the energetically most favoured oxazolidinone structures.

Synthesis and Biological Evaluation of Novel Dehydroabietic Acid-Oxazolidinone Hybrids for Antitumor Properties.

Novel representatives of the important group of biologically-active, dehydroabietic acid-bearing oxazolidinone moiety were synthesized to explore more efficacious and less toxic antitumor agents. Structures of all the newly target molecules were confirmed by IR, ¹H-NMR, 13C-NMR, and HR-MS. The inhibitory activities of these compounds against different human cancer cell lines (MGC-803, CNE-2, SK-OV-3, NCI-H460) and human normal liver cell line LO2 were evaluated and compared with the commercial anticancer drug cisplatin, using standard MTT (methyl thiazolytetrazolium) assay in vitro. The pharmacological screening results revealed that most of the hybrids showed significantly improved antiproliferative activities over dehydroabietic acid and that some displayed better inhibitory activities compared to cisplatin. In particular, compound 4j exhibited promising cytotoxicity with IC50 values ranging from 3.82 to 17.76 µM against all the test cell lines and displayed very weak cytotoxicity (IC50 > 100 µM) on normal cells, showing good selectivity between normal and malignant cells. Furthermore, the action mechanism of the representative compound 4j was preliminarily investigated by Annexin-V/PI dual staining, Hoechst 33258 staining, which indicated that the compound can induce cell apoptosis in MGC-803 cells in a dose-dependent manner and arrest the cell cycle in G1 phase. Therefore, 4j may be further exploited as a novel pharmacophore model for the development of anticancer agents.

Synthesis and antibacterial activity evaluation of novel biaryloxazolidinone analogues containing a hydrazone moiety as promising antibacterial agents.

A series of linezolid analogues containing a hydrazone moiety were designed, synthesized and evaluated for their antibacterial activity. Most compounds exhibited more potent antibacterial activity against S.aureus, MRSA, MSSA, LREF and VRE pathogens as compared with linezolid and radezolid. Compounds 9a, 9c, 9f, 9g, 10m and 10t were more potent against tested clinical isolates of MRSA, MSSA, VRE and LREF as compared to linezolid. Compound 9a exhibited comparable activity with linezolid against human MAO-A for safety evaluation and showed moderate metabolism in human liver microsome. The most promising compound 9a showed remarkable antibacterial activity against S.aureus, MRSA, MSSA, LREF and VRE pathogens with MIC value of 0.0675 mg/mL, respectively, which was 15- to 30-fold more potent than linezolid.

Oxazolidinone-Mediated Sequence Determination of One-Bead One-Compound Cyclic Peptide Libraries.

A novel one-bead one-compound (OBOC) dual ring-opening/cleavage approach for cyclic peptide sequencing was developed. The method selectively modifies serine, cysteine, threonine, and/or glutamic acid to an oxazolidinone-derived moiety, thereby increasing the susceptibility of the modified peptide backbone toward hydrolysis. The resulting linear peptide was then sequenced in 1 min by tandem mass spectrometry on a quadrupole time-of-flight instrument incorporating two-dimensional liquid chromatography and ion mobility spectrometry separation. To evaluate this approach, a library of cyclic peptides was successfully sequenced with 98% overall accuracy, demonstrating its robustness and broad substrate scope.

Mechanistic insights of the controlled release properties of amide adhesive and hydroxyl adhesive.

Although interactions between drugs and acrylate pressure sensitive adhesives (PSAs) containing amide groups were reported in the previous studies, detailed studies elucidating their mechanism of action are still lacking. In the present study, an amide PSA (AACONH2) and a hydroxyl PSA (AAOH, as the control) were synthesized, and their molecular mechanism of controlled drug release was described. Using zolmitriptan (ZOL) and etodolac (ETO) as model drugs, in vitro drug release and skin permeation experiments were performed. Intermolecular interactions between drugs and PSAs were determined by Flory-Huggins model, FT-IR spectroscopic analysis and molecular modeling. In addition, PSA mobility was evaluated using differential scanning calorimetry and rheology study. Release percent of ZOL and ETO from AACONH2 were 43.9 ±â€¯0.3% and 50.0 ±â€¯2.0% respectively, while from AAOH, the release percent of ZOL and ETO were 61.4 ±â€¯1.2% and 81.0 ±â€¯1.2% separately. As a consequence of controlled drug release, skin permeation of both drugs was significantly controlled by AACONH2. It was demonstrated that AACONH2 markedly interacted with drugs, especially with ETO, through hydrogen bonding and weak intermolecular forces (e.g. dipole-dipole and van der waals). PSA mobility of AACONH2 was significantly increased due to drug-PSA interactions. In conclusion, AACONH2 had stronger controlled release properties compared with AAOH, which was mainly caused by the stronger interactions between amide groups and drugs. The amide PSA synthesized in the present study was a potential sustained-release excipient for transdermal drug delivery system.

A coupled bimodal SPECT-CT imaging and brain kinetics studies of zolmitriptan-encapsulated nanostructured polymeric carriers.

The present investigation deals with preparation and characterization of anti-migraine zolmitriptan (ZMT) nanostructured polymeric carriers for nose to brain drug targeting. The drug-loaded colloidal nanocarriers of ZMT were prepared by modified ionic gelation of cationic chitosan with anionic sodium tripolyphosphate and characterized for particle size, zeta potential, and entrapment efficiency. Further, in order to investigate nose to brain drug targeting, biodistribution, and brain kinetics studies were performed using 99mtechnetium radiolabeled nanocarriers (99mTc-ZMTNP) in Swiss albino mice. The results were compared with intranasal pure drug solution (99mTc-ZMT) and intravenous nanocarriers (99mTc-ZMTNP). A single photon emission computerized tomography (SPECT) radioimaging studies were also carried out to visualize and confirm brain uptake of nanocarriers. The optimized nanocarriers showed particle size of 161 nm, entrapment efficiency of 80.6%, and zeta potential of + 23.7 mV. The pharmacokinetic parameters, Cmax, and AUC0-∞ values for ZMT concentration in the brain expressed as percent radioactivity per gram of brain in intranasal and intravenous route of administration were calculated. The brain Cmax and AUC0-∞ values found in three groups, intranasal 99mTc-ZMTNP, intranasal 99mTc-ZMT, and intravenous 99mTc-ZMTNP were (0.427 and 1.889), (0.272 and 0.7157), and (0.204 and 0.9333), respectively. The higher Cmax values of intranasal 99mTc-ZMTNP suggests better brain uptake as compared to other routes of administration. The significant higher values of nose to brain targeting parameters namely, drug targeting index (5.57), drug targeting efficiency (557.08%), and nose to brain drug direct transport (82.05%) confirmed drug targeting to brain via nasal route. The coupled bimodal SPECT-CT scintigrams confirm the brain uptake of intranasal 99mTc-ZMTNP demonstrating major radioactivity accumulation in brain. This study conclusively demonstrated the greater uptake of ZMT-loaded nanocarriers by nose to brain drug targeting, which proves promising drug delivery system.

Structure-guided design, synthesis and evaluation of oxazolidinone-based inhibitors of norovirus 3CL protease.

Acute nonbacterial gastroenteritis caused by noroviruses constitutes a global public health concern and a significant economic burden. There are currently no small molecule therapeutics or vaccines for the treatment of norovirus infections. A structure-guided approach was utilized in the design of a series of inhibitors of norovirus 3CL protease that embody an oxazolidinone ring as a novel design element for attaining optimal binding interactions. Low micromolar cell-permeable inhibitors that display anti-norovirus activity have been identified. The mechanism of action, mode of binding, and structural rearrangements associated with the interaction of the inhibitors and the enzyme were elucidated using X-ray crystallography.

Design, synthesis and biological activity of 3-pyrazine-2-yl-oxazolidin-2-ones as novel, potent and selective inhibitors of mutant isocitrate dehydrogenase 1.

Isocitrate dehydrogenases (IDHs) catalyze the oxidative decarboxylation of isocitrate to alpha-ketoglutarate (α-KG) generating carbon dioxide and NADPH/NADH. Evidence suggests that the specific mutations in IDH1 are critical to the growth and reproduction of some tumor cells such as gliomas and acute myeloid leukemia, emerging as an attractive antitumor target. In order to discovery potent new mutant IDH1 inhibitors, we designed, synthesized and evaluated a series of allosteric mIDH1 inhibitors harboring the scaffold of 3-pyrazine-2-yl-oxazolidin-2-ones. All tested compounds effectively suppress the D-2-hydroxyglutarate (D-2-HG) production in cells transfected with IDH1-R132H and IDH1-R132C mutations at 10 µM and 50 µM. Importantly, compound 3g owns the similar inhibitory activity to the positive agent NI-1 and shows no significant toxicity at the two concentrations. The parallel artificial membrane permeation assay of the blood-brain barrier (PAMPA-BBB) identified 3g with a good ability to penetrate the blood-brain barrier (BBB). These findings indicate that 3g deserves further optimization as a lead compound for the treatment of patients with IDH1 mutated brain cancers.

The convergent synthesis and anticancer activity of broussonetinines related analogues.

The convergent synthesis of broussonetinines related congeners 3 and 4 with the simple C13 alkyl side chain and differently configured pyrrolidine skeleton has been achieved. Our approach relied on the [3,3]-sigmatropic rearrangements of chiral allylic substrates derived from d-xylose. Cross metathesis of the common oxazolidinone intermediates 7 and 8 with tridec-1-ene followed by alkylative cyclization completed the construction of both C-alkyl iminosugars. The targeted compounds 3 and 4 were screened for antiproliferative/cytotoxic activities against multiple cancer cell lines by MTT assay. Compound 3 exhibited very good in vitro potency on Caco-2 and Jurkat cell lines with IC50 value of 5.1 µM and 5.8 µM, respectively.

Synthesis and Evaluation of Agelastatin Derivatives as Potent Modulators for Cancer Invasion and Metastasis.

The synthesis of new agelastatin alkaloid derivatives and their anticancer evaluation in the context of the breast cancer microenvironment is described. A variety of N1-alkyl and C5-ether agelastatin derivatives were accessed via application of our strategy for convergent imidazolone synthesis from a common thioester along with appropriately substituted urea and alcohol components. These agelastatin derivatives were evaluated in our three-dimensional coculture assay for the effects of mammary fibroblasts on associated breast cancer cells. We have discovered that agelastatin alkaloids are potent modulators for cancer invasion and metastasis at noncytotoxic doses. Herein, we discuss the increased potency of (-)-agelastatin E as compared to (-)-agelastatin A in this capacity, in addition to identification of new agelastatin derivatives with activity that is statistically equivalent to (-)-agelastatin E. The chemistry described in this report provides a platform for the rapid synthesis of agelastatin derivatives with excellent potency (50-100 nM) as modulators for cancer invasion and metastasis.

Hepatoprotective Effect of Aqueous Extract from the Seeds of Orychophragmus violaceus against Liver Injury in Mice and HepG2 Cells.

Orychophragmus violaceus (O. violaceus) is a kind of edible wild herb in north China and its seeds have medical potential, however, the effect of O. violaceus seeds on liver injury and the mechanism of action remains poorly understood. Thus, the purpose of the present study is to investigate the effect of O. violaceus seeds on liver injury and further explore the molecular mechanism of the beneficial effects using aqueous extract from the seeds of O. violaceus (AEOV). Mice were orally administrated with saline, AEOV, and biphenyldicarboxylate for 4 days, and were then injected subcutaneously with 0.1% carbon tetrachloride (CCl4) dissolved in corn oil. Sixteen hours later, mice were sacrificed and blood samples were collected. Then, the serum was separated and used for biochemical assay. Livers were excised and were routinely processed for histological examinations. Enzyme activities and protein levels in liver homogenates were detected using commercial kits or by western blot analysis. Additionally, the hepatoprotective effect of AEOV in vitro was evaluated using epigoitrin, the major alkaloid compound isolated from AEOV. We found that AEOV attenuated liver injury induced by CCl4 as evidenced by decreased levels of alanine aminotransferase (ALT) and aminotransferase (AST) in serum, improvement of liver histopathological changes, and substantial attenuation of oxidative stress and inflammation via regulation of nuclear factor-erythroid 2-related factor-2 (Nrf2) and nuclear factor κB (NFκB) pathways. These effects of AEOV were comparable to that of biphenyldicarboxylate which was commonly used as a hepatoprotective reference. Moreover, pretreatment of HepG2 cells with epigoitrin improved cell viability, decreased lactate dehydrogenase (LDH) and malondialdehyde (MDA) levels, increased superoxide dismutase (SOD) and glutathione peroxidase (GSH-Px) activity, attenuated the NFκB pathway, and elevated the Nrf2 pathway after exposure to H2O2. These results suggest that AEOV could effectively prevent CCl4-induced liver injury in mice via regulating the Nrf2 and NFκB pathways, and reveal the cytoprotective effects of epigoitrin against H2O2-induced oxidative stress in HepG2 cells.