Synthesis by diastereomeric resolution, biochemical evaluation and molecular modelling of chiral 3-hydroxyl b-lactam microtubule-targeting agents for the treatment of triple negative breast and chemoresistant colorectal cancers.

The synthesis and biochemical activity of a series of chiral trans 3-hydroxyl ß-lactams targeting tubulin is described. Synthesis of the series of enantiopure ß-lactams was achieved using chiral derivatising reagent N-Boc-l-proline. The absolute configuration was determined as 3S,4S for (+) enantiomer 4EN1 and 3R,4R for (-) enantiomer 4EN2. Antiproliferative studies identified chiral 3S,4S b-lactams with subnanomolar IC50 values across a range of cancer cell lines, improving potency with respect to the corresponding racemates. Fluoro-substituted (+)-(3S,4S)-4-(3-fluoro-4-methoxyphenyl)-3-hydroxy-1-(3,4,5-trimethoxyphenyl)azetidin-2-one (27EN1) was determined as the lead eutomer with dual antiproliferative activity in triple negative breast cancer cells (TNBC), and combretastatin A-4 resistant HT-29 colorectal cancer cells. IC50 values were in the range of 0.26-0.7 nM across four cell lines. Tubulin polymerisation assays, confocal microscopy and molecular modelling studies indicated that 3S,4S eutomers are microtubule destabilisers, while 3R,4R distomers have lower potency as microtubule destabilisers. 27EN1 demonstrated anti-mitotic and pro-apoptotic activity in MDA-MB-231 and HT-29 cells in addition to selective toxicity toward MCF-7 breast cancer versus non-tumorigenic MCF-10-2A cells. The related 3S,4S ß-lactam eutomer 4EN1 downregulated expression of key cell survival anti-apoptotic proteins Bcl-2 and Mcl-1 in MDA-MB-231 cells while 27EN1 downregulated Mcl-1 in HT-29 cells. Chiral ß-lactam 27EN1 will be further developed for treatment of TNBC and CA-4 resistant colorectal cancers.

Steric Repulsion Induced Conformational Switch in Supramolecular Structures.

Inspired by the rigidified architecture of 'picket-fence' systems, we propose a strategy utilizing strain to impose intramolecular tension in already peripherally overcrowded structures leading to selective atropisomeric conversion. Employing this approach, tuneable shape-persistent porphyrin conformations were acquired exhibiting distinctive supramolecular nanostructures based on the orientation of the peripheral groups. The intrinsic assemblies driven by non-covalent bonding interactions form supramolecular polymers while encapsulating small molecules in parallel channels or solvent-accessible voids. The developed molecular strain engineering methodologies combined with synthetic approaches have allowed the introduction of the pivalate units creating a highly strained molecular skeleton. Changes in the absorption spectrum indicated the presence of severe steric repulsions between the peripheral groups which were confirmed by single crystal X-ray analysis. To release the steric strain introduced by the peripheral units, thermal equilibration strategies were used to selectively convert the most abundant atropisomer to the desirable minor one.

Elucidating Atropisomerism in Nonplanar Porphyrins with Tunable Supramolecular Complexes.

Atropisomerism is a fundamental feature of substituted biaryls resulting from rotation around the biaryl axis. Different stereoisomers are formed due to restricted rotation about the single bond, a situation often found in substituted porphyrins. Previously NMR determination of porphyrin atropisomers proved difficult, if not almost impossible to accomplish, due to low resolution or unresolvable resonance signals that predominantly overlapped. Here, we shed some light on this fundamental issue found in porphyrinoid stereochemistry. Using benzenesulfonic acid (BSA) for host-guest interactions and performing 1D, 2D NMR spectroscopic analyses, we were able to characterize all four rotamers of the nonplanar 5,10,15,20-tetrakis(2-aminophenyl)-2,3,7,8,12,13,17,18-octaethylporphyirin as restricted H-bonding complexes. Additionally, X-ray structural analysis was used to investigate aspects of the weak host-guest interactions. A detailed assignment of the chemical signals suggests charge-assisted complexation as a key to unravel the atropisomeric enigma. From a method development perspective, symmetry operations unique to porphyrin atropisomers offer an essential handle to accurately identify the rotamers using NMR techniques only.

Short-Chained Anthracene Strapped Porphyrins and their Endoperoxides.

The syntheses of short-chained anthracene-strapped porphyrins and their Zn(II)complexes are reported. The key synthetic step is a [2+2] condensation between a dipyrromethane and an anthracene bisaldehyde, 2,2'-((anthracene-9,10-diylbis(methylene))bis(oxy))dibenzaldehyde. Following exposure to white light, self-sensitized singlet oxygen and the anthracene moieties underwent [4+2] cycloaddition reactions to yield the corresponding endoperoxides. 1H NMR studies demonstrate that the endoperoxide readily formed in [D]chloroform and decayed at 85 °C. X-ray crystallography and absorption spectroscopy were used to confirm macrocyclic distortion in the parent strapped porphyrins and endoperoxides. Additionally, X-ray crystallography indicated that endoperoxide formation occurred exclusively on the outside face of the anthracene moiety.

Fixing the Conformation of Calix[4]arenes: When Are Three Carbons Not Enough?

Calix[4]arenes are unique macrocycles that through judicious functionalisation at the lower rim can be either fixed in one of four conformations or remain conformationally flexible. Introduction of propynyl or propenyl groups unexpectedly provides a new possibility; a unidirectional conformational switch, with the 1,3-alternate and 1,2-alternate conformers switching to the partial cone conformation, whilst the cone conformation is unchanged, under standard experimental conditions. Using 1 H NMR kinetic studies, rates of switching have been shown to be dependent on the starting conformation, upper-rim substituent, where reduction in bulk enables faster switching, solvent and temperature with 1,2-alternate conformations switching fastest. Ab initio calculations (DFT) confirmed the relative stabilities of the conformations and point towards the partial cone conformer being the most stable of the four. The potential impact on synthesis through the "click" reaction has been investigated and found not to be significant.

Lanthanide directed self-assembly of highly luminescent supramolecular "peptide" bundles from α-amino acid functionalized 2,6-bis(1,2,3-triazol-4-yl)pyridine (btp) ligands.

Ligands containing the [2,6-bis(1,2,3-triazol-4-yl)pyridine] (btp) motif have recently shown promise in coordination chemistry. The motif is synthesized via the Cu(I)-catalyzed "click" reaction and can be conveniently functionalized when compared to other terdentate chelating motifs. Ligand 1 was synthesized and shown to sensitize Eu(III) and Tb(III) excited states effectively. The use of these ions to synthesize self-assembly structures in solution was investigated by carrying out both (1)H NMR and photophysical titrations. The latter were used to determine high binding constants from changes in the absorption, ligand emission (fluorescence), and lanthanide-centered emission. A small library of amino acid derivatives of 1, ligands 3, were prepared upon coupling reactions with Gly, Ala, Phe, and Trp methyl esters, with a view to introducing biologically relevant and chiral moieties into such ligands. All of these derivatives were shown to form stable, emissive Ln(III) self-assemblies, emitting in the millisecond time range, which were studied by means of probing their photophysical properties in organic solutions using lanthanide ion titrations. All the Tb(III) complexes, with the exception of Trp based derivatives, gave rise to highly luminescent and bright complexes, with quantum yields of Tb(III) emission of 46-70% in CH3CN solution. In contrast, the Eu(III) complexes gave rise to more modest quantum yields of 0.3-3%, reflecting better energy match for the Tb(III) complexes, and hence, more efficient sensitization, as demonstrated by using low temperature measurements to determine the triplet state of 1.

Electrostatic interactions play an essential role in the binding of oleic acid with α-lactalbumin in the HAMLET-like complex: a study using charge-specific chemical modifications.

Human α-lactalbumin made lethal to tumor cells (HAMLET) and its analogs are partially unfolded protein-oleic acid (OA) complexes that exhibit selective tumoricidal activity normally absent in the native protein itself. To understand the nature of the interaction between protein and OA moieties, charge-specific chemical modifications of lysine side chains involving citraconylation, acetylation, and guanidination were employed and the biophysical and biological properties were probed. Upon converting the original positively-charged lysine residues to negatively-charged citraconyl or neutral acetyl groups, the binding of OA to protein was eliminated, as were any cytotoxic activities towards osteosarcoma cells. Retention of the positive charges by converting lysine residues to homoarginine groups (guanidination); however, yielded unchanged binding of OA to protein and identical tumoricidal activity to that displayed by the wild-type α-lactalbumin-oleic acid complex. With the addition of OA, the wild-type and guanidinated α-lactalbumin proteins underwent substantial conformational changes, such as partial unfolding, loss of tertiary structure, but retention of secondary structure. In contrast, no significant conformational changes were observed in the citraconylated and acetylated α-lactalbumins, most likely because of the absence of OA binding. These results suggest that electrostatic interactions between the positively-charged basic groups on α-lactalbumin and the negatively-charged carboxylate groups on OA molecules play an essential role in the binding of OA to α-lactalbumin and that these interactions appear to be as important as hydrophobic interactions.

Application of 2D EXSY and qNMR Spectroscopy for Diastereomeric Excess Determination Following Chiral Resolution of ß-Lactams.

Trans-ß-lactam isomers have garnered much attention as anti-cancer microtubule targeting agents. Currently available synthetic methods are available for the preparation of enantiopure ß-lactams and favour isomeric cis/trans ß-lactam mixtures. Indirect chiral resolution offers the opportunity for isolation of exclusively enantiopure trans-ß-lactams. In this study, liquid chromatography chiral resolution of ß-lactams derivatized as diastereomer mixtures with a panel of N-protected amino acids is explored, where N-(Boc)-L-proline served as the optimal chiral derivatising reagent. High-performance liquid chromatography failed to adequately determine diastereomeric excess (de) of resolved diastereomers. Variable temperature, 1 H NMR and 2D EXSY spectroscopic analyses of proline-derivatised diastereomers were successfully employed to characterise equilibrating rotamers of resolved diastereomers and determine their de. Integration of resolved resonances corresponding to H3 and H4 of the ß-lactam ring served as a quantitative qNMR tool for the calculation of de following resolution.

The analysis of amphetamine-like cathinone derivatives using positive electrospray ionization with in-source collision-induced dissociation.

RATIONALE: Amphetamine-like cathinone derivatives have become popular as recreational drugs over the past several years but their identification for forensic purposes is made difficult as they undergo extensive fragmentation under commonly used electron ionization (EI) conditions to afford ambiguous mass spectra. To overcome this, the feasibility of using positive electrospray ionization (ESI) with in-source collision-induced dissociation (CID) to produce distinguishable product ion mass spectra was examined. METHODS: A set of six homologous cathinone derivatives was analyzed using an LTQ/Orbitrap™ high-resolution mass spectrometer to establish if there are any commonalities or uniqueness in their mass spectra. These compounds and a number of other cathinone derivatives were also analyzed on a single quadrupole mass spectrometer to establish the feasibility of using in-source CID for their identification in forensic drug samples. RESULTS: The ESI product ion mass spectra of the [M + H](+) ions of six model compounds were found to be readily interpretable and product ion formation pathways are presented. The use of such mass spectral data in the analysis of forensic drug samples facilitated the discrimination of closely related cathinone derivatives that were difficult to distinguish using conventional gas chromatography/electron ionization mass spectrometry. A product ion mass spectral library of 22 commonly encountered cathinone derivatives was also developed. CONCLUSIONS: It has been shown that the product ion ESI mass spectra of cathinone derivatives are readily interpretable and are useful for the identification of this drug group in forensic samples.

Importance of molecular symmetry for enantiomeric excess recognition by NMR.

Recently prochiral solvating agents (pro-CSA) came under the spotlight for the detection of enantiopurity by NMR. Chemical shift non-equivalency in achiral hosts introduced by the presence of chiral guests yields observable resonance signal splitting (Δδ) correlating to the enantiomeric excess (e.e.). In this work, symmetry is our lens to explain porphyrin-based supramolecular receptor activity in a chiral environment. Based on extensive NMR analyses of the atropisomeric receptors, the host symmetry is shown to be affected by porphyrin nonplanarity and further desymmetrized in the presence of a chiral guest. As such, the exposed porphyrin inner core (N-H), with its strong hydrogen bond abilities, for the first time, has been exploited in enantiomeric composition analysis. Our approach in e.e. detection by N-H signals appearing in a previously underutilized region of the spectrum (below 0 ppm) shows chemical shift splitting (Δδ) three times more sensitive to enantiomeric compositions than previously reported systems.

Photochemical [2 + 2] cycloaddition reactions of 6-alkenyl-3-phenylcyclohex-2-en-1-ones: using biradical conformation control to account for exceptions to the "rule of five".

A series of 6-alkenyl-3-phenylcyclohex-2-enones has been synthesised and the structures of the products obtained from them on irradiation have been determined. The 6-propenyl compounds afforded a tricyclic 'parallel' [2 + 2] cycloaddition product and a bicyclic enone resulting from hydrogen abstraction in the biradical intermediate. The 6-butenyl and 6-pentenyl analogues gave 'crossed' cycloaddition products only. Although the regiochemistry of these cycloaddition reactions cannot be explained in terms of the 'rule of five', it is compatible with the concept of 'biradical conformation control' which is based on a consideration of the energy and structure of the possible 1,4-biradical intermediates.

Synthesis and in vitro toxicity of 4-MTA, its characteristic clandestine synthesis byproducts and related sulfur substituted alpha-alkylthioamphetamines.

4-Methylthioamphetamine (4-MTA) is recognised as a 3,4-methylenedioxymethamphetamine (MDMA)-like drug of abuse. Such amphetamine-type drugs often contain byproducts of uncontrolled, illegal clandestine synthetic processes. We report the isolation and structural identification of a number of novel pyridines, dihydropyridone and N,N-di(1-aryl-2-propyl) amines as route-specific byproducts associated with clandestine synthesis of 4-MTA and related amphetamines. We report the in vitro cytotoxicity of 4-MTA, its synthesis byproducts together with some structurally related sulfur substituted alpha-alkyl phenethylamines in cell lines overexpressing human monoamine transporters as well as in a primary neuronal cell line model and a dopaminergic neuroblastoma cell line. 4-MTA along with a number of other structurally related amphetamine derivatives and synthetic impurities were found to be cytotoxic to these cells within pharmacologically defined concentrations implying that 4-MTA is a cytotoxic agent in vitro and therefore might have the potential to be a neurotoxic agent in vivo.

Design, Synthesis and Biochemical Evaluation of Novel Ethanoanthracenes and Related Compounds to Target Burkitt's Lymphoma.

Lymphomas (cancers of the lymphatic system) account for 12% of malignant diseases worldwide. Burkitt's lymphoma (BL) is a rare form of non-Hodgkin's lymphoma in which the cancer starts in the immune B-cells. We report the synthesis and preliminary studies on the antiproliferative activity of a library of 9,10-dihydro-9,10-ethanoanthracene based compounds structurally related to the antidepressant drug maprotiline against BL cell lines MUTU-1 and DG-75. Structural modifications were achieved by Diels-Alder reaction of the core 9-(2-nitrovinyl)anthracene with number of dienophiles including maleic anhydride, maleimides, acrylonitrile and benzyne. The antiproliferative activity of these compounds was evaluated in BL cell lines EBV- MUTU-1 and EBV+ DG-75 (chemoresistant). The most potent compounds 13j, 15, 16a, 16b, 16c, 16d and 19a displayed IC50 values in the range 0.17-0.38 µM against the BL cell line EBV- MUTU-1 and IC50 values in the range 0.45-0.78 µM against the chemoresistant BL cell line EBV+ DG-75. Compounds 15, 16b and 16c demonstrated potent ROS dependent apoptotic effects on the BL cell lines which were superior to the control drug taxol and showed minimal cytotoxicity to peripheral blood mononuclear cells (PBMCs). The results suggest that this class of compounds merits further investigation as antiproliferative agents for BL.

Metal-directed synthesis of enantiomerially pure dimetallic lanthanide luminescent triple-stranded helicates.

The synthesis and photophysical evaluation of two enatiomerially pure dimetallic lanthanide luminescent triple-stranded helicates is described. The two systems, formed from the chiral (R,R) ligand 1 and (S,S) ligand 2, were produced as single species in solution, where the excitation of either the naphthalene antennae or the pyridiyl units gave rise to Eu(III) emission in a variety of solvents. Excitation of the antennae also gave rise to circularly polarized Eu(III) luminescence emissions for Eu(2):1(3) and Eu(2):2(3) that were of equal intensity and opposite sign, confirming their enantiomeric nature in solution providing a basis upon which we were able to assign the absolute configurations of Eu(2):1(3) and Eu(2):2(3).

Light-harvesting, 3rd generation RuII/CoII MOF with a large, tubular channel aperture.

A photoactive, hetero-metallic CoII/RuII-based metal-organic framework (MOF) with a large channel aperture, ca. 21 Å, is reported. The photophysical properties of the MOF are derived from the RuII nodes giving rise to emission centred at ca. 620 nm and relatively long triplet 3MLCT lifetimes. In addition to the optical attributes, the 1H-imidazo [4,5-f][1,10]-phenanthroline ligand imparts structural functionality to the MOF which is composed of alternating CoII- and RuII-based nodes of Δ and Λ helicity. The framework maintains its integrity upon activation and shows gas sorption behaviour that is characteristic of mesoporous materials promoting high CO2 sorption capacities and selectivities over N2.

Colorimetric sensing of anions in aqueous solution using a charge neutral, cleft-like, amidothiourea receptor: tilting the balance between hydrogen bonding and deprotonation in anion recognition.

The design, synthesis and physical evaluation of 1, a visible colorimetric 'naked eye' pyridyl based bis-amidothiourea sensor for anions, is described. This charge neutral sensor gives rise to significant changes in the absorption spectra upon interactions with several important biological anions such as AMP and ADP in 4:1 DMSO-H(2)O solution, while ATP was not detected. These colorimetric changes are due to the formation, or the combination of hydrogen bonding complexes and/or deprotonation between these anions and 1.

The effect of the linker size in C2-symmetrical chiral ligands on the self-assembly formation of luminescent triple-stranded di-metallic Eu(iii) helicates in solution.

Chiral lanthanide-based supramolecular structures have gained significant importance in view of their application in imaging, sensing and other functional purposes. We have designed chiral C2-symmetrical ligands (L) based on the use of two 2,6-pyridine-dicarboxylic-amide moieties (pda), that differ from one another by the nature of the diamine spacer groups (from 1,3-phenylenedimethanamine (1(S,S), 2(R,R)) and benzene-1,3-diamine (3(S,S), 4(R,R)) to much bulkier 4,4'-(cyclohexane-1,1-diyl)bis(2,6-dimethylaniline) (5(S,S), 6(R,R))) between these two pda units. The self-assembly between L and Eu(iii) ions were investigated in CH3CN solution at low concentration whereby the changes in the absorbance, fluorescence and Eu(iii)-centred emission spectra allowed us to model the binding equilibria occurring in the solution to the presence of [Eu:L2], [Eu2:L2], [Eu2:L3] assemblies and reveal their high binding constant values. The self-assembly in solution were also studied at higher concentration by following the changes in the 1H NMR spectra of the ligands upon Eu(iii) addition, as well as by using MALDI-MS of the isolated solid state complexes. The chiroptical properties of the ligands were used in order to study the structural changes upon self-assembly between the ligands and Eu(iii) ions using circular dichroism (CD) and circularly polarised luminescence (CPL) spectroscopies. The photophysical properties of [Eu2:L3] complexes were evaluated in solution and showed a decrease of luminescence quantum yield when going from the ligand with smaller (1(S,S)) to bulkier (5(S,S)) linker from ∼5.8% to ∼2.6%. While mass-spectrometry revealed the possible formation of trinucler assemblies such as [Eu3:L3] and [Eu3:L2].

Identification of pyrolysis products of the new psychoactive substance 2-amino-1-(4-bromo-2,5-dimethoxyphenyl)ethanone hydrochloride (bk-2C-B) and its iodo analogue bk-2C-I.

2-Amino-1-(4-bromo-2,5-dimethoxyphenyl)ethanone hydrochloride (bk-2C-B) has recently emerged as a new psychoactive substance (NPS). It is most commonly consumed orally, although there are indications that it might also be ingested by inhalation or 'smoking'. Information about the stability of bk-2C-B when exposed to heat is unavailable and the potential for pyrolytic degradation and formation of unknown substances available for inhalation prompted an investigation using a simulated 'meth pipe' scenario. Twelve products following pyrolysis of bk-2C-B were detected and verified by organic synthesis of the corresponding standards. In addition, 2-amino-1-(4-iodo-2,5-dimethoxyphenyl)ethanone hydrochloride (bk-2C-I) was characterized for the first time and subjected to pyrolysis as well. Similar products were formed, which indicated that the replacement of the bromo with the iodo substituent did not affect the pyrolysis pattern under the conditions used. Two additional products were detected in the bk-2C-I pyrolates, namely 1-(2,5-dimethoxyphenyl)-ethanone and 1-iodo-4-ethenyl-5-methoxyphenol. The potential ingestion of pyrolysis products with unknown toxicity adds an element of concern. Copyright © 2017 John Wiley & Sons, Ltd.

A folded [2 × 2] metallo-supramolecular grid from a bis-tridentate (1,2,3-triazol-4-yl)-picolinamide (tzpa) ligand.

A flexible ditopic ligand 1 containing two N,N,O-tridentate (1,2,3-triazol-4-yl)-picolinamide chelating pockets is reported and the formation of multimetallic architectures is explored in the solid and the solution phase. The self-assembled ZnII complex [Zn4(1)4](ClO4)8 exhibited a folded [2 × 2] square grid supramolecular architecture that selectively assembled in MeCN solution as shown using various spectroscopic techniques. The closely related FeII complex shows equivalent behaviour in the solid state, while a discrete dinuclear species [Cu2(NO3)41]·5MeCN was the sole product observed in the solid state from the reaction between 1 and CuII under similar conditions.

Reduction of dipyrido-[3,2-a:2',3'-c]-phenazine (dppz) by photolysis in ethanol solution.

Photolysis of dipyrido-[3,2-a:2',3'-c]-phenazine (dppz) (1) in ethanol solution leads to the formation of 9,14-dihydrodipyridophenazine (2), which has been characterised by detailed NMR analysis, UV/VIS absorption spectroscopy, and theoretical calculations which reveal that its red colour is due to a low-lying intramolecular charge transfer state.