Apomine, a novel 1,1-bisphosphonate ester, has been shown to lower plasma cholesterol concentration in several species. Here we show that Apomine reduced the levels of 3-hydroxy-3-methylglutaryl-coenzyme A reductase (HMGR), the rate-limiting enzyme in the mevalonate pathway, both in rat liver and in cultured cells. Apomine resembles sterols such as 25-hydroxycholesterol in its ability to potently accelerate the rate of HMGR degradation by the ubiquitin-proteasome pathway, a process that depends on the transmembrane domain of the enzyme. The similarity between Apomine and sterols in promoting rapid HMGR degradation extends to its acute requirements for ongoing protein synthesis and mevalonate-derived non-sterol product(s) as a co-regulator. Yet, at suboptimal concentrations, sterols potentiated the effect of Apomine in stimulating HMGR degradation, indicating that these agents act via distinct modes. Furthermore, unlike sterols, Apomine inhibited the activity of acyl-CoA:cholesterol acyltransferase in intact cells but not in cell-free extracts. Apomine stimulated the cleavage of the precursor of sterolregulatory element-binding protein-2 and increased the activity of low density lipoprotein receptor pathway. This Apomine-enhanced activation of sterol-regulatory element-binding protein-2 was prevented by sterols or mevalonate. Taken together, our results provide a molecular mechanism for the hypocholesterolemic activity of Apomine.In mammalian cells, cholesterol homeostasis is maintained by balancing cholesterol uptake and production. Cholesterol uptake is regulated through modulating the levels of the cell surface low density lipoprotein (LDL) 1 receptor (LDLR), and cholesterol synthesis is regulated primarily by changes in levels and activity of the enzyme 3-hydroxy-3-methylglutaryl-coenzyme A reductase (HMGR) (1, 2). HMGR is an endoplasmic reticulum (ER) enzyme that catalyzes the conversion of 3-hydroxy-3-methylglutaryl-CoA into mevalonate (MVA), the first committed step in the MVA pathway that leads to the synthesis of cholesterol as well as of essential non-sterol isoprenoid compounds (1, 2). When cells are starved for cholesterol and/or MVA, levels of HMGR and LDLR are elevated, thus increasing the rate of endogenous sterol synthesis and of LDL uptake (3, 4). Conversely, in cholesterol-replete cells, the levels of HMGR and LDLR decline, thereby lowering sterol production and LDL internalization.HMGR and LDLR are regulated at the transcriptional level by sterol-regulatory elements (SREs) in the promoter of their genes (1, 5). Specific transcription factors, designated SREbinding proteins (SREBPs), bind to these elements and activate transcription (5, 6). The SREBP family comprises three members that control different sets of genes. SREBP-2 regulates the transcription of genes mainly involved in sterol synthesis, whereas SREBP-1a and -1c regulate principally fatty acid biosynthetic genes (7). The nuclear, transcriptionally active SREBPs are derived from the NH 2 -terminal domain of large ER membrane-bound prec...

Apomine, a novel 1,1 bisphosphonate ester, increases the rate of degradation of HMG-CoA reductase, inhibiting the mevalonate pathway and thereby blocking cholesterol biosynthesis. We have investigated whether Apomine can induce myeloma cell apoptosis in vitro and modulate myeloma disease in vivo. Apomine induced a dose-dependent increase in apoptosis in NCI H929, RPMI 8226 and JJN-3 human myeloma cells. Apomine, unlike the bisphosphonate, alendronate, had no measurable effect on osteoclastic bone resorption in vitro. To investigate the effect of Apomine in vivo, 5T2MM murine myeloma cells were injected into C57BL/KaLwRij mice. After 8 weeks all animals had a serum paraprotein and were treated with Apomine (200 mg/kg), or vehicle, for 4 weeks. Animals injected with 5T2MM cells and treated with vehicle developed osteolytic bone lesions, reduced cancellous bone area, decreased bone mineral density (BMD) and increased osteoclast number. Apomine caused a decrease in serum paraprotein and a decrease in tumor burden. Apomine inhibited the development of osteolytic lesions and prevented the tumor-induced decreases in BMD. Apomine had no effect on osteoclast number in contrast to what had been seen previously with the bisphosphonate, zoledronic acid, suggesting that these are direct effects of Apomine on myeloma cells. This demonstrates that Apomine is able to promote myeloma cell apoptosis in vitro and inhibit the development of multiple myeloma and lytic bone disease in vivo. The use of bisphosphonate esters such as Apomine represents a novel therapeutic approach in the treatment of myeloma and, indirectly, the associated bone disease. ' 2007 Wiley-Liss, Inc.

No abstract

Tetra-iso-propyl 2-(3,5-di-tert-butyl-4-hydroxyphenyl)ethyl-1,1-diphosphonate (CAS 126411-13-0, SR-9223i) is a member of a new class of compounds with multiple antiatherosclerotic activities. This report not only describes the cholesterol-lowering properties in four species of animals fed normocholesterolaemic diets but also reductions in lipid deposition in the arteries of cholesterol-fed New Zealand white rabbits following the administration of SR-9223i. Plasma cholesterol concentrations were reduced in mice by 27% (200 mg/kg/day administered in the diet for 10 days), in hamsters by 33% (200 mg/kg/day administered in the diet for 8 days), in dogs by 16% (25 mg/kg/day p.o. for 28 days) and 23% (75 mg/kg/day p.o. for 28 days) and in monkeys by 22% (25 mg/kg/day p.o. for 28 days). Further, the deposition of cholesterol, especially in the esterified form, in the aortae of cholesterol-fed New Zealand white rabbits was inhibited by SR-9223i (50 and 100 mg/kg/day p.o.). At the higher dose, the cholesteryl ester content of the aorta was half that of control animals. SR-9223i, at both doses, also inhibited the accumulation of cholesterol in the liver. SR-9223i has been shown to suppress HMG CoA reductase activity, inhibit ACAT activity and prevent lipid oxidation. These activities, demonstrated in vitro, have now been shown to translate into lipid lowering and antiatherosclerotic activities in vivo.

No abstract