Effects of L-690,488, a prodrug of the bisphosphonate inositol monophosphatase inhibitor L-690,330, on phosphatidylinositol cycle markers.
In order to enhance the entry into cells of L-690,330, a bisphosphonate inhibitor of inositol monophosphatase (IMPase; a key, enzyme in the phosphatidylinositol (Pl) cell signaling pathway), the tetrapivaloyloxymethyl ester prodrug, L-690,488 [tetrapivaloyloxymethyl 1-(4-hydroxyphenoxy)ethane-1,1-bisphosphonate], was synthesized. The effects of L-690,488 were studied in cholinergically (carbachol)-stimulated rat cortical slices and Chinese hamster ovary cells stably transfected with the human muscarinic m1 receptor (m1 CHO cells). The accumulation of [3H]inositol monophosphates or [3H]cytidine monophosphorylphosphatidate ([3H]CMP-PA) after [3H]inositol or [3H]cytidine prelabeling, respectively, and inositol 1,4,5-trisphosphate and inositol 1,3,4,5-tetrakisphosphate mass were measured. In rat cortical slices and m1 CHO cells, the maximum response and time course of accumulation of [3H]inositol monophosphates for L-690,488 and lithium were similar. However, the concentrations of L-690,488 required to produce these effects (EC50 values of 3.7 +/- 0.9 and 1.0 +/- 0.2 microM in cortical slices and m1 CHO cells, respectively) were much lower than with lithium (0.3-1.5 mM). Likewise, the time course and maximum accumulation of [3H] CMP-PA in L-690,488-treated m1 CHO cells was similar to lithium but L-690,488 was again much more potent (EC50 values = 3.5 +/- 0.3 microM and 0.52 +/- 0.03 mM for L-690,488 and lithium, respectively). In addition, L-690,488 attenuated the carbachol-induced elevation of inositol 1,4,5-trisphosphate and inositol 1,3,4,5-tetrakisphosphate in m1 CHO cells, an effect reported previously with lithium. These results are all consistent with L-690,488 and lithium both depleting intracellular inositol as a consequence of inhibition of IMPase. That these effects of L-690,488 on the PI cycle are indeed due to inositol depletion is shown by the observation that the effects of L-690,488 on CMP-PA accumulation could be overcome by addition of exogenous myo-inositol (EC50 = 1.7 +/- 0.5 mM). These data show that inhibition of IMPase produces effects on the PI cycle comparable to lithium. As a corollary, the effects of lithium on the PI cycle are therefore consistent with its major mechanism of action being inhibition of IMPase.
Merck Sharp & Dohme Research Laboratories, Neuroscience Research Centre, Harlow, Essex, England.
J R
JR
Atack
A M
AM
Prior
S R
SR
Fletcher
K
K
Quirk
R
R
McKernan
C I
CI
Ragan
D000818:Animals; D016466:CHO Cells / Q000187:drug effects / Q000378:metabolism; D002217:Carbachol / Q000494:pharmacology; D002463:Cell Membrane Permeability / Q000187:drug effects; D002540:Cerebral Cortex / Q000187:drug effects / Q000378:metabolism; D006224:Cricetinae; D003562:Cytidine / Q000378:metabolism; D003568:Cytidine Monophosphate / Q000031:analogs & derivatives / Q000378:metabolism; D004164:Diphosphonates / Q000494:pharmacology*; D020404:Glycerophospholipids; D007294:Inositol / Q000378:metabolism; D015544:Inositol 1,4,5-Trisphosphate / Q000378:metabolism; D007295:Inositol Phosphates / Q000378:metabolism; D008094:Lithium / Q000494:pharmacology; D008297:Male; D010712:Phosphatidic Acids / Q000378:metabolism; D010716:Phosphatidylinositols / Q000378:metabolism*; D010744:Phosphoric Monoester Hydrolases / Q000037:antagonists & inhibitors*; D011355:Prodrugs / Q000494:pharmacology*; D051381:Rats; D017207:Rats, Sprague-Dawley; D015290:Second Messenger Systems / Q000187:drug effects / Q000502:physiology; D014316:Tritium
D004164:Diphosphonates; D020404:Glycerophospholipids; D007295:Inositol Phosphates; D010712:Phosphatidic Acids; D010716:Phosphatidylinositols; D011355:Prodrugs; C088379:cytidine monophosphorylphosphatidate; D014316:Tritium; C081655:inositol-1,3,4,5-tetrakisphosphate; C088378:L 690488; C079103:L 690330; D007294:Inositol; D003562:Cytidine; D015544:Inositol 1,4,5-Trisphosphate; D002217:Carbachol; D008094:Lithium; D010744:Phosphoric Monoester Hydrolases; C021822:myo-inositol-1 (or 4)-monophosphatase; D003568:Cytidine Monophosphate