Richard J. Brown

The Intracellular Target for the Antiresorptive Aminobisphosphonate Drugs in Dictyostelium discoideum Is the Enzyme Farnesyl Diphosphate Synthase

Aminobisphosphonate (aBP) drugs inhibit osteoclast‐mediated bone resorption and also growth of amoebas of Dictyostelium discoideum apparently by interaction with the same intracellular target. Identification of the target in Dictyostelium therefore could also identify the target in osteoclasts. The aBPs (100 μM alendronate and 30 μM YM‐175) inhibited conversion of [14C]mevalonate into sterols by cultures of Dictyostelium amoebas. One of three enzymes (isopentenyl diphosphate [IDP] isomerase, farnesyl diphosphate [FDP] synthase, and squalene synthase) appeared to be the target for this inhibition because conversion of [14C]IDP into squalene, the immediate precursor for sterol biosynthesis, was inhibited in extracts of wild‐type amoebas by alendronate (IC50 = 75 nM) or risedronate (IC50 = 30 nM) whereas, when the extract had been prepared from amoebas of strains selected for having partial resistance to the growth‐inhibitory effects of alendronate (strain MR102) or risedronate (strain RB101), the values of IC50 were increased to 700 nM for alendronate (MR102 extract) or 130 nM for risedronate (RB101 extract). Neither IDP isomerase nor squalene synthase was inhibited significantly by alendronate or risedronate but both of these aBP drugs, and all others tested, inhibited FDP synthase. Determination of the nucleotide sequences of complementary DNAs (cDNAs) encoding FDP synthase in the wild‐type and aBP‐resistant strains of Dictyostelium indicated that there had been no changes in the amino acid sequence of the enzyme in the mutant strains. However, both mutant strains overproduce FDP synthase. It is concluded that FDP synthase is the intracellular target for the aBP drugs. (J Bone Miner Res 2000;15:971–981)

Differential Effects of Aminosubstituted Analogs of Hydroxy Bisphosphonates on the Growth of Dictyostelium discoideum

Replacing the hydroxyl group in the bone‐binding site of three clinically useful bisphosphonates (etidronate, pamidronate, and olpadronate) by an amino group resulted in great differences in their antiresorptive potencies in vitro. In the present study, this is also shown in vivo in mice treated with the six bisphosphonates at doses of up to 16 μM/kg/day for 12 days. Because binding to bone mineral is nearly the same for all tested bisphosphonates, these findings suggest that the aminosubstitution affects the cellular action of the bisphosphonates. This was tested in the cellular slime mould Dictyostelium discoideum in which cellular effects of bisphosphonates can be examined independently of binding to bone mineral. Etidronate and its aminosubstituted analog were equipotent in inhibiting amebal growth, while pamidronate was somewhat more potent than its analog. Whereas olpadronate was a potent inhibitor of axenic growth of Dictyostelium amebae, the aminosubstitution reduced its potency drastically (IC50 12 μM and 700 μM, respectively). The similarities between the inhibitory effects of the bisphosphonates tested on bone resorption in vitro and in vivo and on the growth of Dictyostelium amebae confirm that the differences in antiresorptive potencies found reflect differences in cellular effects and suggest that bisphosphonates may bind to more than one intracellular target.

Discovery of Narrow Spectrum Kinase Inhibitors: New Therapeutic Agents for the Treatment of COPD and Steroid-Resistant Asthma

The discovery of a novel series of therapeutic agents that has been designed and optimized for treating chronic obstructive pulmonary disease is reported. The pharmacological strategy was based on the identification of compounds that inhibit a defined subset of kinase enzymes modulating inflammatory processes that would be effective against steroid refractory disease and exhibit a sustained duration of action after inhaled delivery.

The Synthesis and Evaluation of Quaternary Pyridinium Bisphosphonates as Potent Anti-Resorptives

Abstract Several novel quaternary pyridinium bisphosphonates have been synthesised and their efficacy as potential anti-resorptive bone agents have been tested in Dictyostelium discoideum. This assay has been shown to accurately reflect the potency of a bisphosphonic acid as an anti-resorptive compound. All the quaternary bisphosphonates are very potent growth inhibitors but results indicate that the more potent compounds are those containing hydrophobic, bulky groups.