Sweet Vector for Gene Delivery: the Sugar Decoration of Polyplexes Reduces Cytotoxicity with a Balanced Effect on Gene Expression.

The use of sugar-functionalized polyplexes as a nonviral gene delivery vector with lower cytotoxicity than the well-known polymeric carrier branched polyethyleneimine (BPEI) is investigated. The substitution of primary amine groups in the BPEI chains with lactose residues leads to larger polyplexes, presumably due to the higher amount of polymer required to complete DNA condensation. Nevertheless, the sugar functionalization substantially reduces the cytotoxicity of the assemblies. The nanocomplexes are taken up by the cells to a greater extent, whereas the levels of gene expression are maintained compared to those obtained using BPEI, which is known for its excellent transfection efficiency. Accordingly, the preparation of lower-cytotoxicity polyplexes while maintaining gene expression, which is highly relevant to the field, is demonstrated.

Measurement of cystic fibrosis transmembrane conductance regulator activity using fluorescence spectrophotometry.

Cystic fibrosis (CF) is a frequent autosomal recessive disease caused by mutations that impair the CF transmembrane conductance regulator (CFTR) protein function. CFTR is a chloride channel activated by cyclic AMP (cAMP) via protein kinase A (PKA) and ATP hydrolysis. We describe here a method to measure CFTR activity in a monolayer of cultured cells using a fluorescence spectrophotometer and the chloride-sensitive probe 6-methoxy-N-(3-sulfopropyl)quinolinium (SPQ). Modifying a slice holder, the spectrophotometer quartz cuvette was converted in a perfusion chamber, allowing measurement of CFTR activity in real time, in a monolayer of T84 colon carcinoma cells. The SPQ Stern-Volmer constant (K(Cl(-))) for chloride in water solution was 115.0 ± 2.8M(-1), whereas the intracellular (K(Cl(-))) was 17.8 ± 0.8 M(-1), for T84 cells. A functional analysis was performed by measuring CFTR activity in T84 cells. The CFTR transport inhibitors CFTR(inh)-172 (5 µM) and glibenclamide (100 µM) showed a significant reduction (P<0.05) in CFTR activity. This simple method allows measuring CFTR activity in a very simple, reproducible, and sensitive way.

Solvent and temperature effects on the conformational equilibria of a dihydrobenzo[h]quinolinium fluoroborate.

Spectra of the N-phenyl-5,6-dihydro-2,4-diphenylbenzo[h]quinolinium tetrafluoroborate (1) and of the N-phenyl-5,6,8,9-tetrahydro-7-phenyldibenzo[c,h]acridinium tetrafluoroborate (2) were recorded in various solvents and temperatures. The analysis of the (1)H-NMR spectra of the tetrafluoroborate salt 1, recorded in acetone, acetonitrile, 1,1,2,2-tetrachloroethane and chloroform, revealed the existence of an equilibrium between two conformers in solution. Tight ion-pairing in chloroform led to a smaller barrier for interconversion between the two conformers. In more polar solvents, where the dihydrobenzoquinolinium exists as a free cation, theoretical calculations predicted larger barriers. The spectra of 1 in 1,1,2,2-tetrachloroethane also varied with temperature, resembling at higher temperatures the spectrum in CDCl(3) and at 300K spectra in more polar media. Spectra of 2 did not vary with the solvent or the temperature, in an indication of a much higher barrier to conformational interconversion, because of a greater steric hindrance between the N-phenyl substituent and the dihydrobenzo rings.

Synthesis and antimalarial activity of E-2-quinolinylbenzocycloalcanones.

A series of E-2-quinolinylbenzocycloalcanones 5-21 were prepared and evaluated for their activity to inhibit beta-hematin formation and the hydrolysis of hemoglobin in vitro. Positive compounds for both assays were also tested for their efficacy in rodent Plasmodium berghei. Compounds 6, 16, 19, and 20, were the most promising. Inhibition of beta-hematin formation was minimal when a hydrogen or methoxy groups were present on the position 8 of the quinoline and position 4' of the indanone ring as it appeared for compounds 5, 7-15, 17, 18, and 21, and greatest with compounds (52%) and (90%) with a substitution of methoxy on position 6 and 7 or methyl on position 8 of the quinoline nucleus and methoxy or methyl groups on position 4' of the indanone. The most active compound to emerge from this study is 2-chloro-8-methyl-3-[(4'-methoxy-1'-indanoyl)-2'-methyliden]-quinoline 20 effective as antimalarial that target beta-hematin formation and the inhibition of the hydrolysis of hemoglobin in vitro together with a good survival in a murine malaria model, which should help delay the rapid onset of resistance to drugs acting at only a single site. Results with these assays suggest that quinolinylbenzocycloalcanones exert their antimalarial activity via multiple mechanisms.