Ahmed S. Mehanna

Liquid chromatographic determination of hippuric acid for the evaluation of ethacrynic acid as angiotensin converting enzyme inhibitor.

A rapid, simple and interference-free method is described to evaluate the inhibitory effects of organic compounds on the activity of angiotensin converting enzyme irrespective of their acid-base properties. The assay is based on the high performance liquid chromatographic separation of the synthetic substrate hippuryl-L-histidyl-L-leucine, the hydrolysis product hippuric acid and the test compound. Using the new method, the diuretic drug ethacrynic acid was found to act as an inhibitor for the enzyme in a non competitive mode.

Hydrophobized triphenyl phosphonium derivatives for the preparation of mitochondriotropic liposomes: choice of hydrophobic anchor influences cytotoxicity but not mitochondriotropic effect

Abstract Context: Nanocarrier-based strategies to achieve delivery of bioactives specifically to the mitochondria are being increasingly explored due to the importance of mitochondria in critical cellular processes. Objective: To test the ability of liposomes modified with newly synthesized triphenylphosphonium (TPP)–phospholipid conjugates and to test their use in overcoming the cytotoxicity of stearyl triphenylphosphonium (STPP)-modified liposomes when used for delivery of therapeutic molecules to the mitochondria. Methods: TPP–phospholipid conjugates with the dioleoyl, dimyristoyl or dipalmitoyl lipid moieties were synthesized and liposomes were prepared with these conjugates in a 1 mol% ratio. The subcellular distribution of the liposomes was tested by confocal microscopy. Furthermore, the liposomes were tested for their effect on cell viability using a MTS assay, on cell membrane integrity using a lactate dehydrogenase assay and on mitochondrial membrane integrity using a modified JC-1 assay. Results: The liposomes modified with the new TPP–phospholipid conjugates exhibited similar mitochondriotropism as STPP-liposomes but they were more biocompatible as compared to the STPP liposomes. While the STPP-liposomes had a destabilizing effect on cell and mitochondrial membranes, the liposomes modified with the TPP–phospholipid conjugates did not demonstrate any such effect on biomembranes. Conclusions: Using phospholipid anchors in the synthesis of TPP–lipid conjugates can provide liposomes that exhibit the same mitochondrial targeting ability as STPP but with much higher biocompatibility.

Design, synthesis and biological evaluation of a series of thioamides as non-nucleoside reverse transcriptase inhibitors.

A series of thioamides were designed as bio-isosteres to the non-nucleoside reverse transcriptase inhibitor trovirdine by replacement of the thiourea NH groups with methylene groups. Eight thioamides were synthesized and in vitro tested for inhibitory effects on the activity of HIV-1 reverse transcriptase wild and mutant types. Three of the 8-thioamides exhibited enzyme inhibitory activities with IC(50) values below 100 microM. While compound (2) exhibited activity against the mutant strain L100I with IC(50) of 70.1 microM, compound (4) showed activity against the mutant strain K103N with IC(50) of 92.7 microM, and compound (8) with activity against the wild type enzyme with IC(50) of 8.9 microM. Each of the three thioamides could serve as a lead compound for further activity optimization.

Synthesis of Triphenylphosphonium Phospholipid Conjugates for the Preparation of Mitochondriotropic Liposomes

Surface modification of liposomes with a ligand is facilitated by the conjugation of the ligand to a hydrophobic molecule that serves to anchor the ligand to the liposomal bilayer. We describe here a simple protocol to conjugate a triphenylphosphonium group to several commercially available functionalized phospholipids. The resulting triphenylphosphonium conjugated lipids can be used to prepare liposomes that preferentially associate with mitochondria when exposed to live mammalian cells in culture.

Hydrogen Bonding: Between Strengthening the Crystal Packing and Improving Solubility of Three Haloperidol Derivatives

The purpose of this study is to confirm the impact of polar functional groups on inter and intra-molecular hydrogen bonding in haloperidol (HP) and droperidol (DP) and, hence, their effects on dissolution using a new approach. To confirm our theory, a new molecule: deshydroxy-haloperidol (DHP) was designed and its synthesis was requested from a contract laboratory. The molecule was then studied and compared to DP and HP. Unlike DHP, both the HP and DP molecules have hydrogen donor groups, therefore, DHP was used to confirm the relative effects of the hydrogen donor group on solubility and crystal packing. The solid dispersions of the three structurally related molecules: HP, DP, and DHP were prepared using PVPK30, and characterized using XRPD and IR. A comparative dissolution study was carried out in aqueous medium. The absence of a hydrogen bonding donor group in DHP resulted in an unexpected increase in its aqueous solubility and dissolution rate from solid dispersion, which is attributed to weaker crystal pack. The increased dissolution rate of HP and DP from solid dispersions is attributed to drug-polymer hydrogen bonding that interferes with the drug-drug intermolecular hydrogen bonding and provides thermodynamic stability of the dispersed drug molecules. The drug-drug intermolecular hydrogen bond is the driving force for precipitation and crystal packing.

High Throughput Kinetic Assay for Screening Potential Inhibitors of SickleHemoglobin Polymerization

Previously we reported the capacity of the bio-inspired Fe(III) complexes, Fe2PO and Fe2PC, to mimic the activity of superoxide dismutase and peroxidase enzymes, as well as their capacity to reduce the cytotoxicity generated by superoxide radicals in human peripheral blood mononuclear cells, under stress conditions. Based on those findings, we decided to evaluate the cytotoxicity, antioxidant and redox state modulation capacity of Fe2PO and Fe2PC complexes, pondering them as drug candidates against free radicals unbalance disorders. Fe2PO and Fe2PC deactivated the ABTS synthetic radical with an IC50 value of 38.4 ± 0.9 μM and 28.9 ± 0.2 μM, respectively, while against the DPPH radical, the IC50 value was over the higher concentration tested (>200 μM) for both complexes. As desirable for any drug candidate, none of the metallic complexes (at 25, 50 and 100 μM) induced cytotoxicity on M12.C3.F6 cells (a murine B-cell lymphoma model), but differences in the redox state modulation were observed on the basis of fluorescence detection of a 2′,7′-dichlorofluorescin probe by flow cytometry. Cells under normal conditions and preincubated with Fe2PO and Fe2PC complexes slightly augmented the reactive oxygen species concentration, meanwhile, cells under stress condition preincubated with H2O2 and metallic complexes, showed a higher augmentation in the reactive oxygen species concentration, in comparison to the controls. Finally, a cellular internalisation assay was performed, showing that Fe2PO and Fe2PC exert those effects from the outside of the cells. All these results suggest the ability of Fe2PO and Fe2PC complexes to selectively increase the reactive oxygen species concentration in cells with a free radical unbalance, without inducing mortality in cells under normal conditions.

Design, Synthesis and Calcium Channel Blocking Activity of Diltiazem- Verapamil Hybrid Molecules

The current manuscript describes the design, synthesis, and in vitro testing of four thioacetanilides with diltiazemverapamil hybrid structural features as potential calcium channel blockers. The current hybrid strategy of drug design aimed to generate compounds that could span, with a single compound, the trans-membrane locales where the two drugs bind, with the ultimate goal of increasing the blocking activity. The latter, was assessed by measuring the inhibitory effects, expressed as IC50, on calcium-induced contractions of potassium depolarized isolated rat aorta strips. The assessment of the binding locales was determined by incubating the test compound with aortic strips for two different periods, 10-minutes and 2-hours, before adding the contractile calcium ions to the assay medium. Diltiazem IC50 values were 0.26 and 0.14 μM, after 10-minutes and 2-hours, respectively, reflecting less than two fold increase in activity and confirming previous reports that its locale of binding in mostly on the exterior side of the membrane. On the other hand, verapamil IC50 values were 0.47 and 0.14 μM after 10-minutes and 2-hour incubation respectively, reflecting approximately a 3-4 fold increase in activity and confirming previous reports that it binds mainly to the interior domains of the membrane. The four designed hybrid compounds showed, after 10-minute incubation, an IC50 value range of 3.7-12.0 μM, and after 2-hour incubation an IC50 range of 0.78-2.12 μM, reflecting approximately a 5-fold increase in activity suggesting more similarity to the verapamil binding profile. The data indicate that the designed compounds are with moderate activities, but generally less active as calcium channel blockers than either of the two parent drugs.