br Mechanisms of action The focus here

Mechanisms of action The focus here is on compounds that act on bone solely via the ER and that have been proven to prevent bone loss. It is well known that estrogens prevent bone loss, but the precise mechanism is still not completely understood. Estrogens may act via ERα and/or ERβ and possess different affinities for these receptors. It has been proposed that ERβ is a type of co-repressor of ERα [18], although evidence for this is lacking in humans. Estradiol binds equally well to both receptors, as does estrone albeit with a weaker affinity [19], whilst ethinyl estradiol shows a two-fold higher affinity for ERα [20]. Pure agonists for the ER may all give the same maximal response in the various systems, but the potency may differ. Evidence exists that estrogens induce their own receptor, but when administered at high unphysiological doses, the ER can be down-regulated thus leading to a lower biological response. Both osteoblasts and osteoclasts contain ERs [4], [5]. Studies with intact ERKO and BERKO mice have shown that both receptors have a role in bone physiology, but it is important to note that endogenous hormone levels undergo drastic changes in intact KO-animals [21], [22] and it remains to be proven whether this is true under physiological circumstances. Further proof is required as to whether ER membrane receptors play a role in the maintenance of bone mass. It is also possible that other non-genomic pathways may be involved. Nevertheless, whatever the precise mechanism of action of estrogens may be, the imbalance between bone formation and resorption in postmenopausal women is restored by calpain inhibitor through inhibition of osteoclastogenesis resulting in a lower resorption rate and restoration of the balance between bone formation and bone loss. In mice, supraphysiological doses of estrogens may lead to bone formation, but this effect appears to be species-dependent. The high endogenous hormone concentrations in (B)ERKO mice [21], [22] may explain the lack of effect on bone after deletion of one of the ER genes. SERMs or partial agonists also bind to ERs, but functional assays have shown that partial transactivation occurs and it is possible that selectivity for one of the receptors may exist. Tamoxifen, a triphenylethylene derivative and the first generation SERM (although the term was only introduced later), has shown surprisingly positive effects on bone [23], [24]. The metabolite 4-OH-tamoxifen is the active metabolite and has a two–three-fold higher binding affinity for ERβ (Ki=0.04nM) [19] than for ERα. Raloxifene, which has a benzothiophene chemophore, is structurally different from tamoxifen and does not need to be metabolized in order to bind to the ER. The conformation of SERM–ER complexes is different from that of complexes with natural estradiol [25]. It is probable that the conformation with each estrogen is different and therefore each estrogen may have unique properties [26]. In the raloxifene–ER complex, helix 12 does not lie over the binding pocket, as seen with the estradiol–ER complex, and this may prevent coactivators from binding to the ligand binding domain of the ER. Transactivation experiments have shown that raloxifene and 4-OH-tamoxifen have ERα agonistic and antagonistic activity, whereas they possess full antagonistic activity at the ERβ [7]. In bone, the effects of raloxifene and tamoxifen show similarities to pure ER agonists, whilst in other tissues no activation is found [24], [27]. As with estrogens, the final effect on bone is an inhibition of resorption, although the effects are less strong. The main distinctions from estradiol are differences in the binding to ERα and ERβ and the fact that ER responsive genes are less strongly induced by SERMs.