Dopamine-selective potentiometric responses by new ditopic sensory elements based on a hexahomotrioxacalix[3]arene

New ditopic sensory elements 2 and 3 for catecholamines based on a hexahomotrioxacalix[3]arene, with a boronic acid substituent appended, were designed and synthesized. As an interesting mode of molecular recognition at membrane surfaces, the host, when incorporated into poly(vinyl chloride) (PVC) liquid membranes, displayed excellent potentiometric selectivity for dopamine over other catecholamines (noradrenaline and adrenaline) and inorganic cations (Na+, K+, and NH4+).     

Calix[4]arene C-90 selectively inhibits Ca2+,Mg2+-ATPase of myometrium cell plasma membrane

The supramolecular compound calix[4]arene C-90 (5,11,17,23-tetra(trifluoro)methyl(phenylsulfonylimino)-methylamino-25,26,27,28-tetrapropoxycalix[4]arene) is shown to efficiently inhibit the ATP hydrolase activity of Ca²⁺,Mg²⁺-ATPase in the myometrium cell plasma membrane fraction and also in a preparation of the purified enzyme solubilized from this subcellular fraction. The inhibition coefficient I _0.5 values were 20.2 ± 0.5 and 58.5 ± 6.4 μM for the membrane fraction and the solubilized enzyme, respectively. The inhibitory effect of calix[4]arene C-90 was selective comparatively to other ATPases localized in the plasma membrane: calix[4]arene C-90 did not influence the activities of Na⁺,K⁺-ATPase and “basal” Mg²⁺-ATPase. The inhibitory effect of calix[4]arene C-90 on the Ca²⁺,Mg²⁺-ATPase activity was associated with the cooperative action of four trifluoromethylphenyl sulfonylimine (sulfonylamidine) groups oriented similarly on the upper rim of the calix[4]arene macrocycle (the calix[4]arene “bowl”). The experimental findings seem to be of importance for studies, using calix[4]arene C-90, of membrane mechanisms of regulation of calcium homeostasis in smooth muscle cells and also for investigation of the participation of the plasma membrane Ca²⁺-pump in control of electro- and pharmacomechanical coupling in myocytes.     

Conjugation, immunoreactivity, and immunogenicity of calix[4]arenes; model study to potential calix[4]arene-based Ac3+ chelators.

For the development of calix[4]arene-based radiotherapeutic agents, the conjugation to biomolecules and immunogenicity in mice of potential 225Ac3+-chelating calix[4]arenes were studied. A calix[4]arene triethyl ester isothiocyanate and a bis(calix[4]arene) hexacarboxylic acid, containing a masked thiol functionality, were used in conjugation experiments to a mouse monoclonal antibody and serum albumins. All characterization techniques indicate that only the calix[4]arene carboxylic acid is successfully conjugated to the biomolecules. The immunoreactivity of the conjugates is not impaired when up to 6 equiv of calixarene are bound to the monoclonal antibody. Animal tests indicated that the immunogenicity toward the calix[4]arene is strongly influenced by the nature of the carrier, the dosage, and the injection method. No immune response occurred when a homologous carrier was used or when a heterologous carrier was applied at a dosage of 10 microg per immunization via intravenous injection. Under all other conditions, the presence of antibodies directed against the calix[4]arene was demonstrated. Thus, for the application in radioimmunotherapy, the conjugation of a calix[4]arene to a humanized antibody will probably not lead to an immune response, and the immunoreactivity will not be disturbed.     

Extraction selectivities of lower rim substituted calix[4]arene hosts induced by variations in the upper rim substituents

The compound 25,26,27,28-tetra-(2-dimethyldithiocarbamoylethoxy)calix[4]arene has been prepared from 25,26,27,28-tetra-(2-bromoethoxy)calix[4]arene by reaction with sodium dimethyldithiocarbamate. As an extractant for heavy metal ions 25,26,27,28-tetra-(2-dimethyldithiocarbamoylethoxy)calix[4]arene is effective for Hg²⁺, Ag⁺, Pd²⁺ and Au³⁺, but much less effective than 5,11,17,23-tetra-tert-butyl-25,26,27,28-tetra-(2-N,N-dimethyldithiocarbamoylethoxy)calix[4]arene for both Hg₂²⁺ and MeHg⁺. Calixarene alcohols also show selectivity as hosts. The alcohol derivative 25,26,27,28-tetra-(2-hydroxyethoxy)calix[4]arene undergoes slow occlusion of iodine into the lower rim, whereas with the alcohol 5,11,17,23-tetra-tert-butyl-25,26,27,28-tetra-(2-hydroxyethoxy)calix[4]arene no interaction is observed.     

Role of chloride on carrier-mediated transport of p-aminohippurate in rat renal basolateral membrane vesicles

Effect of inorganic anions on p-amino[3H]hippurate transport in renal basolateral membranes has been studied using the vesicles preloaded with unlabeled p-aminohippurate (countertransport condition). The uptake of p-amino[3H]hippurate was stimulated by the outward gradient of unlabeled p-aminohippurate and the labeled substrate was accumulated into the vesicles against its concentration gradient in the presence of Cl-. The substitution of SCN- and SO4(2-) for Cl- in both sides of the vesicles depressed the initial rate and the overshoot magnitude of p-amino[3H]hippurate uptake. These results suggest that Cl- may play an important role for the carrier-mediated transport system of organic anion in renal basolateral membranes.     

Chiral calix[4]arenes bearing amino alcohol functionality as membrane carriers for transport of chiral amino acid methylesters and mandelic acid

Novel chiral calix[4]arene derivatives bearing amino alcohol moieties at the lower rim have been synthesized from the reaction of p-tert-butylcalix[4]arene diester with various amino alcohols. The transport of amino acid esters (phenylglycine, phenylalanine, and tryptophan methyl esters hydrochloride) and mandelic acid were studied through chloroform bulk liquid membrane system using chiral calix[4]arenes 15-20. All these receptors have been found to act as carriers for transport of aromatic amino acid methylesters and mandelic acid from the aqueous source phase to the aqueous receiving phase. The influence of calixarene and guest structures upon transport through liquid membrane is discussed.     

Pyrene-p-tert-butylcalixarenes inclusion complexes formation: a surface photochemistry study.

Diffuse reflectance and luminescence techniques were used to study the photophysics and photochemistry of pyrene within p-tert-butylcalix[n]arenes with n = 4, 6, and 8, and to study their ability to form inclusion complexes in heterogeneous media. Evidences for inclusion complex formation were found for the three hosts under study. Ground state diffuse reflectance results have shown the formation of ground state dimers of pyrene inside the cavity of calix[6]arene and calix[8]arene, with this feature much more evident for calix[6]arene. For calix[4]arene, only a monomer fits inside the cavity and the presence of pyrene microcrystals outside the cavity was detected. A luminescence lifetime distribution analysis was performed, revealing the presence of prompt emissions from the pyrene microcrystals outside the cavity in the case of calix[4]arene and from the constrained dimers inside the cavities of calix[6]arene and calix[8]arene. Transient absorption results have shown the presence of pyrene radical cation and also of trapped electrons for the three hosts under study. The formation of the phenoxyl radical of the calixarene following the laser pulsed excitation of pyrene at 355 nm is increased for calix[6]arene and calix[8]arene. This feature is particularly relevant for calix[6]arene, suggesting a very favourable situation for the hydrogen atom abstraction to occur. The analysis of the degradation products revealed the presence of hydroxypyrene as a major photodegradation product for the three hosts. Dihydro-hydroxypyrene was also formed in the case of calix[6]arene and calix[8]arene. The formation of the calixarene's phenoxyl radical and subsequent hydrogen abstraction is consistent with the formation of dihydro-dihydroxypyrene.     

Calix[8]arene-mediated self-assembly of tetrapeptide H-Leu-Leu-Ile-Leu-OMe

Conformational analysis of peptide 1, H-Leu-Leu-Ile-Leu-OMe on complexing with macro cycle calix[8]arene has been carried out using (1)H-NMR and FTIR spectroscopic techniques. Stoichiometry of the complex formed in the 1:8 ratio was evidenced by a Job plot. NMR studies of the above peptide show a marked downfield shift and an increase in (3)J values for NH resonances on complexing with calix[8]arene. The characteristic NOE connectivity between N(i+1)H and C(ialpha)H confirm beta-sheet conformation in the complexed state. Both (1)H-NMR and FTIR results indicate that the alpha-amino group of Leu I is proximal to the macrocycle and is involved in hydrogen bond formation with phenolic hydrogen atom of the calix[8]arene. This suggests that calix[8]arene provides a suitable platform for peptide 1 to self-assemble in a parallel beta-sheet conformation. The nature of calix[8]arene interaction with peptide 1 has been studied using dynamic NMR studies, which concludes that a bifurcated hydrogen bonding interaction exists in the molecular interfaces of the assembly.     

Non-covalent (iso)guanosine-based ionophores for alkali(ne earth) cations

Different (iso)guanosine-based self-assembled ionophores give distinctly different results in extraction experiments with alkali(ne earth) cations. A lipophilic guanosine derivative gives good extraction results for K⁺, Rb⁺, Ca²⁺, Sr²⁺, and Ba²⁺ and in competition experiments it clearly favors the divalent Sr²⁺ (and Ba²⁺) cations. 1,3-Alternate calix[4]arene tetraguanosine hardly shows any improvement in the extraction percentages compared to its reference compound 1,3-alternate calix[4]arene tetraamide. This indicates that one G-quartet does not provide efficient cation complexation under these conditions. In the case of the lipophilic isoguanosine derivative there is a cation size dependent affinity for the monovalent cations (Cs⁺ ? Rb⁺ ? K⁺), but not for the divalent cations (Ca²⁺ > Ba²⁺ > Sr²⁺ > Mg²⁺). In competition experiments the isoguanosine derivative, unlike guanosine, does not discriminate between monovalent and divalent cations, giving an almost equal extraction of Cs⁺ and Ba²⁺.     

Spectroscopic investigation of the interaction of water-soluble azocalix[4]arenes with bovine serum albumin

In this work, three hydrosoluble azocalix[4]arene derivatives, 5-(o-methylphenylazo)-25,26,27-tris(carboxymethoxy)-28-hydroxycalix[4]arene (o-MAC-Calix), 5-(m-methylphenylazo)-25,26,27-tris(carboxymethoxy)-28-hydroxycalix[4]arene (m-MAC-Calix) and 5-(p-methylphenylazo)-25,26,27-tris(carboxymethoxy)-28-hydroxycalix[4]arene (p-MAC-Calix) were synthesized. Their structures were characterized by infrared spectrum (IR), nuclear magnetic resonance spectrum (¹H NMR and ¹³C NMR) and mass spectrum (MS). The interactions between these compounds and bovine serum albumin (BSA) were studied by fluorescence spectroscopy, UV–vis spectrophotometry and circular dichroic spectroscopy. According to experimental results, three azocalix[4]arene derivatives can efficiently bind to BSA molecules and the o-MAC-Calix displays more efficient interactions with BSA molecules than m-MAC-Calix and p-MAC-Calix. Molecular docking showed that the o-MAC-Calix was embedded in the hydrophobic cavity of helical structure of BSA molecular and the tryptophan (Trp) residue of BSA molecular had strong interaction with o-MAC-Calix. The fluorescence quenching of BSA caused by azocalix[4]arene derivatives is attributed to the static quenching process. In addition, the synchronous fluorescence spectroscopy indicates that these azocalix[4]arene derivatives are more accessible to Trp residues of BSA molecules than the tyrosine (Tyr) residues. The circular dichroic spectroscopy further verified the binding of azocalix[4]arene derivatives and BSA.     

Calixarene-based phosphinic acids as inhibitors of protein tyrosine phosphatases

In the present work, the derivatives of calix[4]arene, thiacalix[4]arene, and sulfonylcalix[4]arene bearing four methylene(phenyl)phosphinic acid groups on the upper rim of the macrocycle were synthesized and studied as inhibitors of human protein tyrosine phosphatases. The inhibitory capacities of the three compounds towards PTP1B were higher than those for protein tyrosine phosphatases TC–PTP, MEG1, MEG2, and SHP2. The most potent sulfonylcalix[4]arene phosphinic acid displayed K_i value of 32?nM. The thiacalix[4]arene phosphinic acid was found to be a low micromolar inhibitor of PTP1B with selectivity over the other PTPs. The kinetic experiments showed that the inhibitors compete with the substrate for the active site of the enzyme. Molecular docking was performed to explain possible binding modes of the calixarene-based phosphinic inhibitors of PTP1B.     

Structural and functional bases of the intermolecular interaction of calix[4]arene C-97 with myosin subfragment-1 of myometrium

Calix[4]arene C-97 (code is shown) is the macrocyclic compound which has lipophilic intramolecular higly-structured cavity formed by four aromatic cycles, one of which on the upper rim is modified by methylene bisphosphonic group. It was shown that calix[4]arene C-97 (100 microM) efficiently inhibits ATPase activity of myosin subfragment-1 from pig myometrium, the inhibition coefficient I(0.5) being 83 +/- 7 microM. At the same time, this compound at 100 microM concentration significantly increases the effective hydrodynamic diameter of myosin subfragment-1, that may be indicative of intermolecular complexation between the calix[4]arene and myosin head. Computer simulation methods (docking, molecular dynamics, involving the Grid) have been used to clarify structural basis of the intermolecular interaction of calix[4]arene C-97 with myosin subfragment-1 of the myometrium; participation of hydrophobic, electrostatic and pi-pi (stacking) interactions between calix[4]arene C-97 and amino acid residues of myosin subfragment-1, some of them being located near the active site of the ATPase has been found out.     

Inhibition of Yersinia protein tyrosine phosphatase by phosphonate derivatives of calixarenes

Inhibition of Yersinia protein tyrosine phosphatase by calix[4]arene mono-, bis-, and tetrakis(methylenebisphosphonic) acids as well as calix[4]arene and thiacalix[4]arene tetrakis(methylphosphonic) acids have been investigated. The kinetic studies revealed that some compounds in this class are potent competitive inhibitors of Yersinia PTP with inhibition constants in the low micromolar range. The binding modes of macrocyclic phosphonate derivatives in the enzyme active center have been explained using computational docking approach. The results obtained indicate that calix[4]arenes are promising scaffolds for the development of inhibitors of Yersinia PTP.     

The synthesis and structural characterisation of two polynuclear hafnium(IV) complexes

Attempts to form extended supramolecular structures incorporating hafnium(IV) complexes of p-sulfonatocalix[6]arene resulted in the serendipitous formation of two polynuclear-hafnium clusters. Subsequent reaction between hafnium triflate and 1 M sulfuric or p-toluenesulfonic acid solutions also led to the formation of the same clusters, identified as cations based on Hf₁₇ and Hf₄, respectively.     

Calix[4]arenes C-136 and C-137 hyperpolarize myometrium mitochondria membranes

Calixarenes are supramolecular compounds interacting with bioactive molecules and ions, causing changes in biochemical and biophysical processes. The aim of this work was to study the effects of calix[4]arenes C-136, C-137, and C-138 at the level of polarization of the rat myometrium mitochondria membrane. The structure of synthesized calix[4]arene molecules was confirmed by the methods of ¹H NMR and infrared spectroscopy. Calix[4]arenes C-136 and C-137 each possess two chalcone amide moieties at the lower rim, while calix[4]arene C-138, only one. Calix[4]arenes C-136 and C-137 differ by the presence of ether or hydroxyl groups, respectively, at the lower rim of calix[4]arene skeleton, as well as the length of alkyl spacer between chalcone amide group and the macrocycle. It was shown that calix[4]arenes C-136, C-137, and C-138 form micelles in aqueous medium and in dimethylformamide (DMF). Irradiation of micelles with an argon laser on the flow cytometer results in the rise of autofluorescence. In an aqueous medium, calix[4]arene micelles interact with a positively charged voltage-sensitive fluorescent probe TMRM, which can testify to the presence of negative charge in these structures. However, calix[4]arene micelles do not interact with TMRM in DMF solution. The mitochondrial membrane potential was measured using fluorescent dyes MTG and TMRM with confocal microscopy and fluorescent dye TMRM with flow cytometry. Experiments were conducted on myometrium cells in culture and on suspension of digitonin-permeabilized uterus myocytes. It was shown that the fluorescent signal was stable during the time of experiment. Calix[4]arenes C-136 and C-137 (10 μM) hyperpolarize mitochondria membranes. At maximum, the effect was 173% relative to the control. At the same time, calix[4]arene C-138 did not influence the mitochondria membrane potential. The relationship between the structural organization of investigated calix[4]arene molecules and their effect on polarization of the mitochondria membrane is discussed.     

Calix[6]arene bypasses human pancreatic cancer aggressiveness: Downregulation of receptor tyrosine kinases and induction of cell death by reticulum stress and autophagy

Pancreatic cancer ranks fourth among cancer-related causes of death in North America. Minimal progress has been made in the diagnosis and treatment of patients with late-stage tumors. Moreover, pancreatic cancer aggressiveness is closely related to high levels of pro-survival mediators, which can ultimately lead to rapid disease progression, resistance and metastasis. The main goal of this study was to define the mechanisms by which calix[6]arene, but not other calixarenes, efficiently decreases the aggressiveness of a drug resistant human pancreas carcinoma cell line (Panc-1). Calix[6]arene was more potent in reducing Panc-1 cell viability than gemcitabine and 5-fluorouracil. In relation to the underlying mechanisms of cytotoxic effects, it led to cell cycle arrest in the G0/G1 phase through downregulation of PIM1, CDK2, CDK4 and retinoblastoma proteins. Importantly, calix[6]arene abolished signal transduction of Mer and AXL tyrosine kinase receptors, both of which are usually overexpressed in pancreatic cancer. Accordingly, inhibition of PI3K and mTOR was also observed, and these proteins are positively modulated by Mer and AXL. Despite decreasing the phosphorylation of AKT at Thr308, calix[6]arene caused an increase in phosphorylation at Ser473. These findings in conjunction with increased BiP and IRE1-α provide a molecular basis explaining the capacity of calix[6]arene to trigger endoplasmic reticulum stress and autophagic cell death. Our findings highlight calix[6]arene as a potential candidate for overcoming pancreatic cancer aggressiveness. Importantly, we provide evidence that calix[6]arene affects a broad array of key targets that are usually dysfunctional in pancreatic cancer, a highly desirable characteristic for chemotherapeutics.     

A molecular level study of selective cation capture by a host–guest mechanism for 25,26,27,28-tetramethoxycalix[4]arene in MClO4 solution (M = Na,K)

25,26,27,28-Tetramethoxycalix[4] arene selectively captures cations by changing the conformation. In this study, a hybrid approach of ab initio molecular orbital calculation and statistical mechanics for molecular liquid was utilised to understand the capture mechanism in electrolyte solution phase at molecular level. The association free energy and solvation structure were evaluated on the basis of statistical mechanics for molecular liquids. The selectivity is correctly reproduced by the computation, namely cone conformer captures Na⁺ while K⁺ is recognised by the partial-cone conformer.     

Proton-coupled organic cation transport in renal brush-border membrane vesicles

We had previously proposed that organic cations are transported across the brush-border membrane in the canine kidney by a H+ exchange (or antiport) system (Holohan, P.D. and Ross, C.R. (1981) J. Pharmacol. Exp. Ther. 216, 294-298). In the present report, we demonstrate that in brush-border membrane vesicles the transport of organic cations is chemically coupled to the countertransport of protons, by showing that the uphill or concentrative transport of a prototypic organic cation, N1-methylnicotinamide (NMN), is chemically coupled to the flow of protons down their chemical gradient. In a reciprocal manner, the concentrative transport of protons is coupled to the counterflow of organic cations down their concentration gradient. The transport of organic cations is monitored by measuring [3H]NMN while the transport of protons is monitored by measuring changes in acridine orange absorbance. The functional significance of the coupling is that a proton gradient lowers the Km and increases the Vmax for NMN transport.     

Design,synthesis and application of chiral tetraoxacalix[2]arene[2]triazine‐based organocatalysts in asymmetric Michael addition reactions

A novel type of oxacalix[2]arene[2]triazine‐based organocatalysts for asymmetric Michael reactions are reported for the first time. Chiral subunits were attached to the heteroatom‐bridged calixaromatic platform by a reaction of (R)‐ and (S)‐1‐aminotetraline with tetraoxacalix[2]arene[2]triazine in both enantiomeric forms. To evaluate the catalytic efficiency of the novel organocatalysts, isobutyraldehyde reacted with various substituted and unsubstituted aromatic trans‐β‐nitrostyrenes in tetrahydrofuran (THF), leading to Michael adducts in excellent yields and enantioselectivites (up to 97% yield and 99% ee).     

Chiral mono and diamide derivatives of calix[4]arene for enantiomeric recognition of chiral amines

Novel chiral mono and diamide derivatives of calix[4]arene have been prepared from the aminolysis reaction of 5,11,17,23-tetra-tert-butyl-25,27-diethoxycarbonyl-methoxy-26,28-dihydroxycalix[4]arene 1 and 25,27-diethoxycarbonyl-methoxy-26,28-dihydroxycalix[4]arene 2 with chiral (S)-(-)-1-phenylethylamine (PEA) and (1S,2S)-(+)-2-amino-1-(4-nitrophenyl)-1,3-propanediol, respectively. Spectrophotometric titrations have been performed in CHCl(3) at 20-30 degrees C in order to obtain the binding constants (K) and the thermodynamic quantities (DeltaH and DeltaS) for the stoichiometric 1:1 inclusion complexation of various chiral amines with these new host compounds. Preliminary experiments were undertaken to confirm the complexation properties of receptors 9 and 13 with PEA by (1)H NMR in CDCl(3) at room temperature. The molecular recognition abilities and enantioselectivities for guests (R and S)-alpha-PEA and (R and S)-cyclohexylethylamine (CHEA) are discussed from a thermodynamic point of view.