Search

A strategy is introduced for enhancing the cellular selectivity of Amphotericin B (AmB) and other classes of membrane-disrupting agents. This strategy involves attaching the agent to a molecular umbrella to minimize the disruptive power of aggregated forms. Based on this approach, AmB has been coupled to a molecular umbrella derived from one spermidine and two cholic acid molecules and found to have antifungal activities approaching that of the native drug. However, in sharp contrast to AmB, the hemolytic activity and the cytotoxcity of this conjugate toward HEK293 T cells have been dramatically reduced.

A tetrawalled and an octawalled molecular umbrella conjugate of amphotericin B (AmB) have been synthesized. Both conjugates exhibit high water solubility, a low tendency to aggregate, negligible hemolytic activity at 100 μM, and an ability to release a derivative of AmB under reducing conditions that exhibits high antifungal activity. Whereas the larger, octawalled conjugate shows slight adsorption to liposomal membranes and an ability to cross them by passive transport, the tetrawalled analogue shows significant adsorption and much lower bilayer transport activity. The potential of molecular umbrella–AmB conjugates as therapeutic agents is briefly discussed., Please provide a synopsis

The synthesis, computer modeling, and biological activity of an octawalled molecular umbrella short interfacing RNA (siRNA) conjugate is described. This molecular umbrella-siRNA conjugate exhibited mRNA knockdown activity in vitro in the absence of a transfection reagent. Evaluation of this molecular umbrella conjugate in vivo, using the rat eye via intravitreal injection, resulted in sequence specific mRNA knockdown in the retina with no obvious signs of toxicity, as judged by ophthalmic examination.

A homologous series of calix[4]arene-, calix[5]arene-, and calix[6]arene-based surfactants, containing pendant trimethylammonium and n-hexadecyl groups, have been compared with respect to their ability (i) to undergo ionic cross-linking at the air/water interface, (ii) to incorporate poly(4-styrenesulfonate) (PSS) in Langmuir-Blodgett (LB) bilayers, and (iii) to act as barriers toward He, N(2), and CO(2) when assembled into cross-linked LB bilayers. As these calix[n]arenes increase in size, their ability to undergo ionic cross-linking has been found to increase, the thickness of corresponding glued LB bilayers has been found to decrease, and their barrier properties and permeation selectivities have been found to increase. The likely origin for these effects and the probable mechanism by which He, N(2), and CO(2) cross these ultrathin films are discussed.

A persulfated molecular umbrella derived from one spermine, four lysine, and eight deoxycholic acid molecules was found to exhibit ionophoric activity, as shown by pH discharge and Na(+) and Cl(-) transport experiments. In sharp contrast, a moderately more hydrophilic analogue derived from cholic acid showed no such ionophoric activity. Both molecular umbrellas crossed liposomal membranes by passive transport with experimental rates that were similar. These findings show how the interactions between such amphomorphic molecules and phospholipid bilayers are a sensitive function of the umbrella's hydrophilic/lipophilic balance (HLB). They also raise the possibility of exploiting molecular umbrellas in fundamentally new ways.

A fluorescently labeled, persulfated molecular umbrella ( 1) has been synthesized from cholic acid, lysine, spermine, and Coumarin 343 and found capable of entering live HeLa cells. The distributions of 1 throughout the cytoplasm and the nucleus were diffuse and punctate, respectively. This finding, together with its ability to cross liposomal membranes by passive diffusion, suggests that passive diffusion plays a significant role in the ability of 1 to enter cells. The fact that 1 is concentrated at the nucleus raises the possibility that molecular umbrellas of this type could be used for the nuclear targeting of drugs.

One of the most challenging questions that relates to the structure and function of biological membranes is whether the two halves of the bilayer "talk" to each other. In this letter, we show how the perturbation of the lateral organization of one leaflet of a fluid phospholipid bilayer by an external agent also alters the lateral organization of the adjoining leaflet. In addition, we show that the energy involved in such "cross talk" corresponds to ca. 100 cal/mol of phospholipid. These findings provide a basis for expecting similar cross talk to exist in cell membranes.

Langmuir-Blodgett (LB) monolayers of 5,11,17,23,29,35-hexaformyl-37,38,39,40,41,42-hexakis(1-n-octyloxy)-calix[6]arene (2), deposited onto silylated silicon wafers, were cross-linked (i.e., "covalently glued") via Schiff base formation with poly(allylamine). Direct evidence for imine formation was obtained from X-ray photoelectron spectroscopy and from attenuated total reflection IR spectroscopy. These modified surfaces could be removed from the aqueous subphase into air with retention of the assembly and its orientation relative to the surface, as evidenced by atomic force microscopy, water contact angle measurements, and film thickness determinations by ellipsometry. Similar assemblies were also synthesized via a postgluing procedure, in which the substrate containing the LB monolayer was removed from the subphase and rapidly immersed into an aqueous solution containing poly(allylamine). The potential of combining postgluing methods with continuous LB film deposition as a surface modification technique is briefly discussed.

The introduction of poly(acrylamide) (PAM, Mw 10 000) to an aqueous subphase resulted in increased intermolecular association in the monolayer formed from 5,11,17,23,29,35-hexaamideoxime-37,38,39,40,41,42-hexakis(1-hexadecyloxy) calix[6]arene (2) at the air−water interface, as evidenced by surface viscosity measurements. Langmuir−Blodgett (LB) bilayers of 2, which were deposited onto silylated silicon wafers, showed the presence of PAM, as determined by X-ray photoelectron spectroscopy (XPS). On the basis of XPS analysis at two different takeoff angles (15 and 75°), both the amidoxime groups of the calix[6]arene and the amide groups of PAM, appear to be buried within the LB bilayer. Deposition of single LB bilayers of 2 containing PAM onto poly[1-(trimethylsilyl)-1-propyne] (PTMSP) supports and measurement of their their permeability with respect to He, N2, and CO2 showed a substantial improvement in their permeation selectivity properties, compared with analogous membranes that were devoid of PAM. Taken ...

A persulfated molecular umbrella, derived from cholic acid and spermidine, has been covalently attached to H-Tyr-D-Ala-Gly-Phe-D-Leu-OH (DADLE) by use of an o-dithiobenzyl carbamate linkage. Treatment of the resulting conjugate (1) with glutathione in solution resulted in the liberation of the free form of the peptide. Addition of 1 to glutathione-entrapped liposomes, prepared from 1-palmitoyl-2-oleyol-sn-glycero-3-phosphocholine (POPC), 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphatidylglycerol (POPG), and cholesterol [POPC/POPG/cholesterol, 72/4/24 (mol/mol/mol)], resulted in the delivery of DADLE into their aqueous interior.

Two molecular umbrella-nucleoside conjugates (1a and 1b) have been synthesized via thiolate-disulfide displacement by adenosine 5'-O-(3-thiomonophosphate) and adenosine 5'-O-(3-thiotriphosphate) on an activated dimer derived from cholic acid, spermidine, and 5,5'-dithiobis-(2-nitrobenzoic acid). Both conjugates readily enter the aqueous compartment of liposomes made from 1-palmitoyl-2-oleyol-sn-glycero-3-phosphocholine (POPC) and release the free nucleoside upon reaction with entrapped glutathione. Approximately 50% of the thiolated form of AMP is released within 20 min at 23 degrees C; 120 min is required for a similar release of the thiolated form of ATP. The facile cleavage of these conjugates by glutathione, together with the fact that mammalian cells contain millimolar concentrations of this tripeptide in their cytoplasm, suggest that such chemistry may be extended to the practical development of prodrugs, e.g., antisense oligonucleotides that can be delivered into cells.

Single Langmuir-Blodgett (LB) bilayers derived from calix[n]arene-based surfactants (n = 4, 5 and 6) and poly(acrylic acid) (PAA) exhibit exceptional H(2)/CO(2) permeation selectivities. Evidence for the importance of ionic crosslinking and self-healing processes is presented.

The condensing action of cholesterol has been compared with that of a structural isomer having its hydroxyl group located at the C-25 position (i.e., 25-OH'), that is, an isomer favoring an "upside down" orientation in lipid membranes. Surface pressure-area isotherms of mixed monolayers made from 1,2-dimyristoyl-sn-glycero-3-phosphocholine (DMPC)/cholesterol and DMPC/25-OH' have established that 25-OH' has a weaker condensing effect than cholesterol. Nearest-neighbor recognition measurements in liposomes made from 1,2-dipalmitoyl-sn-glycero-3-phosphocholine (DPPC) have also shown that 25-OH' has a weaker condensing effect in the physiologically relevant fluid bilayer state. These findings provide support for surface occupancy playing a role in the condensing action of cholesterol.

This paper describes the conversion of poly(ethylene oxide) into a polyelectrolyte by use of an ionic linker strategy, and its subsequent use in fabricating polyelectrolyte multilayers (PEMs) with poly(allylamine hydrochloride) (PAH). The resulting thin films were found to exhibit reverse size-selectivity, in which the permeation of a relatively large CO2 molecule was favored over He. The potential of this new approach for modifying the bulk properties of PEMs is briefly discussed.

The use of partially quaternized, chloromethylated polystyrene as a covalent scavenger of cholate ion in aqueous media has been demonstrated. The ability of such polymers to scavenge organic anions by covalent as well as by ionic means has important implications in the areas of medicinal and environmental chemistry, which are briefly discussed.

There has been great interest recently in therapeutic use of nucleic acids including genes, ribozymes and antisense oligonucleotides. Despite recent improvements in delivering antisense oligonucleotides to cells in culture, nucleic acid-based therapy is still often limited by the poor penetration of the nucleic acid into the cytoplasm and nucleus of cells. In this report we describe nucleic acid delivery to cells using a series of novel cationic amphiphiles containing cholic acid moieties linked via alkylamino side chains. We term these agents 'molecular umbrellas' since the cationic alkylamino chains provide a 'handle' for binding of nucleic acids, while the cholic acid moieties are likely to interact with the lipid bilayer allowing the highly charged nucleic acid backbone to traverse across the cell membrane. Optimal gene and oligonucleotide delivery to cells was afforded by a derivative (amphiphile 5) containing four cholic acid moieties. With this amphiphile used as a constituent in cationic liposomes, a 4-5 log increase in reporter gene delivery was measured. This amphiphile used alone provided a 250-fold enhancement of oligo-nucleotide association with cells as observed by flow cytometry. A substantial fraction of cells exposed to complexes of amphiphile 5 and fluorescent oligo-nucleotide showed nuclear accumulation of the fluorophore. Enhanced pharmacological effectiveness of antisense oligonucleotides complexed with amphiphile 5 was observed using an antisense splicing correction assay that activates a Luciferase reporter. Intracellular delivery, nuclear localization and pharmacological effectiveness of oligonucleotides using amphiphile 5 were similar to those afforded by commercial cytofectins. However, in contrast to most commercial cytofectins, the umbrella amphiphile showed substantial delivery activity even in the presence of high concentrations of serum.

Chemical evidence has been obtained for transbilayer movement of a di-walled and a tetra-walled molecular umbrella in large unilamellar vesicles (200 nm) derived from 1-palmitoyl-2-oleyol-sn-glycero-3-phoshatidylglycerol (POPG). A di-walled molecular umbrella (1), bearing 2-mercaptopyridine (2-MP) as a “reactive tag”, was synthesized by reaction of N1,N3-spermidinebis[cholic acid amide] (3) with [N-1,2,3-benzotriazin-4(3H)one-yl]-3-(2-pyridyldithio)propionate [BPDP]. An analogous tetra-walled umbrella (2) was also prepared by condensing Fmoc-protected iminodiacetic acid with two molecules of 3, deprotecting the secondary amino group, and coupling the resulting intermediate (4) with BPDP. Reaction of vesicle-bound 1 with external glutathione (GSH) resulted in a rapid and quantitative release of 2-MP. A similar thiolate−disulfide interchange reaction that was carried out between membrane-bound 1 and GSH, which was captured within the aqueous interior of the vesicles, also resulted in rapid and complete rele...

This paper reports the design and synthesis of 5,11,17,23,29,35-hexaacetyl-37,38,39,40,41,42-hexakis(1-hexadecyloxy)calix[6]arene (2) and 5,11,17,23-tetraamideoxime-25,26,27,28-tetrakis(1-hexadecyloxy)calix[4]arene (3), plus a detailed comparison of the membrane properties of corresponding Langmuir-Blodgett films with those assembled from 5,11,17,23,29,35-hexaamideoxime-37,38,39,40,41,42-hexakis(1-hexadecyloxy)calix[6]arene (1a). Surface pressure-area isotherms that were recorded for 1a and 2 over a pure water subphase at 25 C showed significant hysteresis that was eliminated after two compression-expansion cycles; the limiting area in both cases was ca. 162 {angstrom}{sup 2}/molecule. In sharp contrast, 3 showed no hysteresis at 25 C and a limiting area of 107 {angstrom}{sup 2}/molecule. Careful examination of the dependence of surface viscosity on temperature has provided compelling evidence for a solid-analogous to liquid-analogous phase transition occurring at 31 and 28 C for monolayers of 1a and 2, respectively. Permeation measurements that were made across Langmuir-Blodgett (LB) films [four monolayers supported on poly-[1-(trimethylsilyl)-1-propyne] with respect to helium and nitrogen] revealed high selectivity in the case of 1a and much lower selectivity with 2 and 3 at ambient (21--23 C) temperatures. The implications of these findings, in terms of the further development of LB films as membranes for gas separations, are briefly discussed.

A new concept in chemical sensing has been introduced that uses changes in density of a cross-linked polymer to physically signal changes in solution composition. Prototype systems that have now be...

A series of calix[6]arene-based surfactants have been synthesized, which contain amide oxime head groups on their upper rim and 5,5-dimethylhexyl (I), n-octyl (II), n-dodecyl (III), and n-hexadecyl (IV) groups on their lower rim. Composite membranes that were fabricated from Langmuir−Blodgett (LB) multilayers derived from each surfactant plus poly[1-(trimethylsilyl)-1-propyne] (PTMSP) cast film showed a significant increase in their selectivity toward helium and nitrogen, relative to bare PTMSP. In sharp contrast, analogous composites that were prepared with LB multilayers of conventional single chain surfactants [arachidic acid (AA), cadmium arachidate (AA/Cd2+), and stearoamideoxime (SA)] exhibited permeation properties that were identical with those of bare PTMSP. When a polymeric surfactant [poly(1-octadecene-co-maleic anhydride), POM] was used for LB film construction, a modest increase in selectivity was observed. These findings, together with an analysis of representative composites by X-ray photoe...

This paper describes the design and synthesis of a series of conjugates derived from cholic acid, spermidine, and 5-(dimethylamino)-1-naphthalenesulfonyl (dansyl), which effectively shield the dansyl moiety from water. Direct coupling of cholic acid to both terminal amino groups of spermidine, and attachment of the environmentally-sensitive dansyl moiety to the remaining secondary amine, yields a “molecular umbrella” (Ia) whose fluorescent properties (λmax and emission intensity) reflect a nonpolar microenvironment in water and one that is relatively polar in intermediate dimethoxyethane/water mixtures. Comparison of Ia with analogous “single-walled” (II) and “no-walled” (III) umbrellas further indicates that a minimum of two walls is necessary in order to have “umbrella-like” properties. Examination of the fluorescent properties of a related double-walled umbrella, bearing a flexible (2-hydroxyethyl)carbamate moiety at the C-3 position of the sterol (Ib), reveals that “umbrella-like” properties are prese...

Structurally unmatched hyperthin (14 nm) polyelectrolyte multi-layers (PEMs) formed on poly[1-(trimethylsilyl)-1-propyne] supports exhibit exceptionally high CO(2)/N(2) permeation selectivities, ranging from 100 to 150. Structurally matched PEMs show similar selectivities with significantly higher permeances.

The use of organic polymeric membranes to separate gaseous mixtures provides an attractive alternative to other methods such as selective adsorption and cryogenic distillation. The primary advantages of membrane-based separations are their relative energy efficiency and lower costs. Because the flux of a gas across a membrane is inversely proportional to the membrane's thickness, this method relies on fabricating membranes that are as thin as possible. However, as researchers have tried to produce "hyperthin" membranes (less than 100 nm), these membranes often form defects and lose their permeation selectivity. In this Account, we review some of the progress in our laboratories at Lehigh University to create hyperthin membranes with high permeation selectivities. We focus special attention on gaseous permeants that are relevant for the production of clean energy (H2 and CO2 formed from CH4) and the reduction of global warming (CO2 and N2, the major components of flue gas). Our studies make extensive use of Langmuir-Blodgett (LB) methods and porous surfactants derived from calix[6]arenes. We specially designed each surfactant to form cohesive monolayers and multilayers, and we introduced a "gluing" technique, where we cross-link porous surfactants containing quaternary ammonium groups ionically with polymeric counterions. Using ellipsometry, atomic force microscopy, X-ray photoelectron spectroscopy, monolayer isotherm, surface viscosity, and permeation measurements, we have characterized these hyperthin films. While molecular sieving appears to make a significant contribution to the permeation selectivity of some of these membranes, solution-diffusion pathways predominate. We also describe initial studies in which we formed hyperthin films from poly(ethylene glycol)-based polyelectrolytes using layer-by-layer deposition (LbL) methods. We have found remarkably high H2/CO2 and CO2/N2 permeation selectivities with these LB- and LbL-based hyperthin membranes. These results suggest that such materials may lead the way to materials that researchers can exploit to purify hydrogen produced from CH4 and to capture CO2 from flue gas.

Polymeric surfactants were prepared by quaternization of poly(4-chloromethylstyrene) with N,N-dimethyl-N-n-dodecylamine and N,N-dimethyl-N-n-octylamine to give 1 and 2, respectively. Each of these polymers formed stable monolayers at the air/water interface. Injection of poly(acrylic acid) (PAA) beneath the surface of these films led to a substantial increase in their cohesiveness (i.e., “gluing”), as evidenced by a dramatic increase in their surface viscosity. Examination of monolayers of 1 by atomic force microscopy, after being transferred to silicon wafers that were surface-modified with n-octadecyltrichlorosilane, showed that the presence of PAA leads to intact film. In contrast, transfer of unglued monolayers resulted in poor coverage. Comparison of the barrier properties of single glued and unglued LB bilayers formed in the presence and in the absence of PAA have shown that PAA minimizes defect formation within these ultrathin assemblies.

A series of diwalled and tetrawalled molecular umbrellas have been synthesized using cholic acid, spermidine, and lysine as starting materials. Coupling of these molecular umbrellas to an octaarginine peptide afforded agents that were capable of promoting the transport of small interfering RNA to HeLa cells, as judged by the knockdown of enhanced green fluorescent protein expression. The efficiency of this knockdown was found to increase with an increasing number of facially amphiphilic walls present, and also when a cleavable disulfide linker was replaced with a noncleavable, maleimido moiety; i.e., a group that is not susceptible to thiolate-disulfide interchange. The knockdown efficiency that was observed for one tetrawalled molecular umbrella-octaargine conjugate was comparable to that observed with a commercially available transfection agent, Lipofectamine 2000, but the conjugate showed less cytotoxicity.

N-[O-1,2,3-Benzotriazin-4(3H)one-yl]-3-(2-pyridyldithio)propionate [BPDP] has been synthesized and found to react with secondary amines to give high yields of acylated product. In sharp contrast, the widely used heterobifunctional coupling reagent, N-[O-succinimdy]-3-(2-pyridyldithio) propionate [SPDP] afforded low to negligible yields of product.

Molecular umbrellas are “amphomorphic” compounds that can produce a hydrophobic or a hydrophilic exterior when exposed to a hydrophobic or hydrophilic microenvironment, respectively. Such molecules are composed of two or more facial amphiphiles that are connected to a central scaffold. Molecular umbrellas that have been synthesized to date, using bile acids as umbrella “walls”, polyamines such as spermidine and spermine as scaffold material, and L-lysine as “branches”, have been found capable of transporting certain hydrophilic peptides, nucleotides, and oligonucleotides across liposomal membranes by passive diffusion. They have also have been shown to increase in the water solubility and hydrolytic stability of a hydrophobic drug, and to exhibit significant antiviral activity. The ability of a fluorescently-labeled molecular umbrella to readily enter live HeLa cells suggests that such conjugates could find use as drug carriers.

A tetrawalled and an octawalled molecular umbrella conjugate of amphotericin B (AmB) have been synthesized. Both conjugates exhibit high water solubility, a low tendency to aggregate, negligible hemolytic activity at 100 μM, and an ability to release a derivative of AmB under reducing conditions that exhibits high antifungal activity. Whereas the larger, octawalled conjugate shows slight adsorption to liposomal membranes and an ability to cross them by passive transport, the tetrawalled analogue shows significant adsorption and much lower bilayer transport activity. The potential of molecular umbrella-AmB conjugates as therapeutic agents is briefly discussed.

A series of molecular umbrella conjugates, derived from cholic acid, deoxycholic acid, spermidine, lysine, and 5-mercapto-2-nitrobenzoic acid, have been synthesized and found capable of transporting an attached 16-mer oligonucleotide (S-dT16) across liposomal membranes made from 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine (POPC), 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphatidyldglycerol (POPG), and cholesterol [POPC/POPG/cholesterol (65/5/30; mol/mol/mol, v/v/v)] at 37 degrees C. Those molecular umbrellas containing four choloyl (or deoxycholoyl) groups resulted in significantly faster rates of transport as compared to those containing only two such moieties. A model that accounts for these membrane transport processes is proposed.

Single Langmuir-Blodgett bilayers derived from 5,11,17,23,29,35-hexakis[(N,N,N-trimethylammonium)-N-methyl-37,38,39,40,41,42-hexakis-n-hexamedecyloxy-calix[6]arene hexachloride (1), which have been ionically cross-linked (i.e., "glued together") with poly(acrylic acid), have been found to exhibit He/CO2 permeation selectivities as high as 150. This degree of selectivity, for a membrane that is less than 6 nm in thickness, is without precedent. In principle, materials of this type could lead the way to improved membranes for hydrogen purification.

A series of persulfated molecular umbrellas have been synthesized from putrescine, spermidine, spermine, lysine, and cholic acid (1a, 2a, 3a, 4a, and 5a) and their anti-HIV and anti-HSV activities determined. Despite it size, the most active of these conjugates (5a) was able to cross phospholipid bilayers made from 1-palmitoyl-2-oleyol-sn-glycero-3-phosphocholine (POPC) and 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphatidylglycerol (POPG). The unique combination of anti-viral activity, an ability to cross hydrophobic barriers, a lack of cytotoxicity, and a simple three-step synthesis from biogenic starting material suggests that 5a and related conjugates may be exploitable as a novel class of anti-viral agents for systemic and topical applications.

This paper describes the synthesis and monolayer properties of two porous surfactants derived from the calix[5]arene framework that exist as stable cone conformers [i.e., 5,11,17,29,29-pentaformyl-91,92,99,94,95-pentakis(n-octyloxy)calix[5]arene (1a) and an N,N-dimethylthioamide derivative 2a] plus calix[4]arene and calix[6]arene analogs. The introduction of 1a, in and of itself, represents a significant advance in the calixarene area because it provides access to a wide-cavity calixarene that is amenable for further modification at its wider rim

A series of cross-linked polymers have been synthesized by reaction of 2% cross-linked, chloromethylated polystyrene with a tertiary amine derivative (7) of cholic acid (prepared by reaction of cholic acid methyl ester with N,N-dimethyl-ethylenediamine), followed by exhaustive quaternization with trimethylamine. Increased loadings of 7 result in enhanced binding of taurocholate ion and a decrease in its rate of release upon exposure to 50 mM aqueous NaCl. Examination of particle-size effects indicates that film diffusion and particle diffusion both contribute to the observed kinetics. Interruption tests that were carried out confirmed that particle diffusion is kinetically important. The relevance of this bioconjugate approach to the design of bile acid sequestrants for the treatment of hypercholesterolemia is briefly discussed.

A synthetic ionophore (1) has been prepared via condensation of 23,24-bisnor-5-cholenic acid with both amino groups of 1,17-diamino-3,6,9,12,15-pentaoxaheptadecane and used to probe the mixing behavior of a series of unsaturated phosphocholines (PCs) in the fluid bilayer state. Specifically, the ability of 1 to promote the transport of Na+ across membranes made from equimolar mixtures of “long” and “short” PCs has been compared with that in analogous membranes derived from single-component PCs of intermediate length. Relative ionophoric activities have been measured by use of 23Na+ NMR spectroscopy. The results of this study provide compelling evidence that a hydrophobic mismatch of four methylene groups can lead to nonideal mixing of phosphocholines in fluid bilayers.

The ability of a series of molecular umbrellas, derived from cholic acid, L-lysine, spermidine, and Cascade Blue, to cross fluid liposomal membranes made from 1-palmitoyl-2-oleyol-sn-glycero-3-phosphocholine (POPC)/1-palmitoyl-2-oleoyl-sn-glycero-3-phosphatidylglycerol (POPG) (95/5, mol/mol) has been determined. In sharp contrast to the classic "size/lipophilicity" rule of membrane transport, those molecular umbrellas that were larger in size and less lipophilic crossed these liposomal membranes more readily. The likely origin for this unusual behavior is briefly discussed.

Double- and tetrawalled molecular umbrella-spermine conjugates (I and II) have been synthesized, and their binding to calf thymus DNA (CT-DNA), poly[d(AT)], and poly[d(GC)] compared with that of a single-walled analogue (III). At moderate salt concentrations (8 mM NaCl), I and II show significantly greater affinity toward each DNA, relative to III; at high salt concentrations (150 mM NaCl), strong binding of I and II (but not III) was maintained toward poly[d(GC)]. Examination of the influence of I-III on the melting behavior of poly[d(AT)] has provided strong evidence that the binding of I and II reflects highly cooperative interactions among DNA-bound conjugates and that the DNA duplex serves as a nucleation site for umbrella aggregation. The implications of these findings for the rational design of novel drug conjugates that operate at the nuclear level, and also novel transfection agents, are briefly discussed.

This paper describes a new approach for modifying hydrophobic surfaces that is based on the use of ionically cross-linked (i.e., "glued") Langmuir-Blodgett monolayers. Specifically, this work shows how monolayers of 5,11,17,23,29,35-hexa(trimethylammonium methyl)-37,38,39,40,41,42-hexakis-n-hexadecyloxy-calix[6]arene (1a), which have been ionically cross-linked with poly(4-styrene-sulfonate) (PSS, MW 30,000-50,000), can be deposited onto silylated silicon wafers with their polar headgroups extending outward into air. This work also demonstrates the feasibility of using glued monolayers of 1a as "anchors" for attaching alternating layers of poly(diallyldimethylammonium) [PDADMA] and PSS ions onto these same hydrocarbon surfaces.

A di-walled molecular umbrella, composed of two choloyl groups, one spermidine moiety, and a 5-thiol(2-nitrobenzoyl) "handle", was covalently attached to a 16-mer oligonucleotide (S-dT16) through a disulfide bond. Incubation of this conjugate (1) with vesicles made from 1-palmitoyl-2-oleyol-sn-glycero-3-phosphocholine and 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphatidylglycerol (95/5, mol/mol) (200 nm diameter, extrusion) that contained entrapped glutathione (GSH) at 37 degrees C resulted in the liberation of the oligonucleotide and the umbrella-bound 5-mercapto(2-nitrobenzoyl) moiety (USH) via thiolate-disulfide interchange. The appearance of USH, together with the absence of leakage of entrapped GSH and a vesicular capture of the oligonucleotide that matches the extent of USH formation, provides compelling evidence for delivery of S-dT16 into the aqueous compartment of these vesicles. In a sense, the molecular umbrella functions like a "needle" in providing a pathway for the oligonucleotide (the "thread") to cross the membrane.