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LY353381.HCl: a novel raloxifene analog with improved SERM potency and efficacy in vivo.

Sato M, Turner CH, Wang T, Adrian MD, Rowley E, Bryant HU.

Department of Endocrine Research, Lilly Corporate Center, Indianapolis, Indiana, USA. Sato_Masahiko Lily.com

Body weight, uteri, serum cholesterol and bones were shown previously in vivo to be sensitive to circulating levels of estrogen, as well as to synthetic, nonsteroidal ligands termed selective estrogen receptor modulators (SERM). In this study, we examined the in vivo effects of a new potent SERM on these tissues in 6-month-old, ovariectomized rats that were orally dosed with 0.0001-10 mg/kg/day LY353381.HCl for 5 weeks. LY353381.HCl prevented the ovariectomy-induced increase in body weight and serum cholesterol levels of treated rats and lowered them to below sham levels in a dose dependent manner, with maximum efficacy similar to estrogen or raloxifene. However, LY353381.HCl was consistently more potent than raloxifene, with a half maximal efficacious dose of 0.001 mg/kg for the reduction of body weight and cholesterol. In the uterus, LY353381.HCl had marginal effects on uterine weight compared to ovariectomized controls (OVX) like raloxifene, but unlike estrogen. Histological examination of uterine epithelial cell height showed little to no stimulatory effect of LY353381.HCl on the endometrium. Quantitative computed tomographic analyses (pQCT) of tibiae showed that LY353381.HCl prevented loss of bone due to ovariectomy with an ED50 of about 0.01 mg/kg with maximal efficacy observed at 0.1-1 mg/kg/day. Maximally attainable bone mineral density and content with LY353381.HCl were not significantly different from Sham or ovariectomized rats treated with estrogen or raloxifene. Interestingly, assessment of bone quality by biomechanical analyses showed that LY353381.HCl preserved the strength of the femora neck and midshaft, while improving the Young's modulus of cortical bone to beyond estrogen, raloxifene or sham levels. In uteri of immature rats treated with estrogen, LY353381.HCl antagonized the estrogen-induced elevation in uterine weight down to vehicle-dosed control levels with ED50 of 0.03 mg/kg/day. Therefore, LY353381.HCl was 30-100 times more potent than raloxifene in preventing ovariectomy effects on body weight, serum cholesterol and bone, while maintaining estrogen antagonist effects on the uterus. These animal data suggest that LY353381.HCl may have advantages over estrogen or raloxifene in the prevention of bone loss and treatment of other tissues in postmenopausal women.

Online source: www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&list_uids=9765314&dopt=Abstract raloxifene Evista



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LY353381 x HCl: an improved benzothiophene analog with bone efficacy complementary to parathyroid hormone-(1-34).

Sato M, Zeng GQ, Rowley E, Turner CH.

Lilly Research Laboratories, Eli Lilly & Co., Lilly Corporate Center, Indianapolis, Indiana 46285, USA. sato_masahiko lilly.com

LY353381 x HCl is a benzothiophene analog that is structurally related to raloxifene with potent selective estrogen receptor modulator activity in the ovariectomized rat model of postmenopausal osteoporosis. The effects of LY353381 x HCl on bones, body weight, and uterine weight were evaluated in 7-month-old rats with osteopenia that was induced by ovariectomizing animals for 1 month before initiation of treatment with several agents individually, in combination, or in sequence. LY353381 x HCl was administered daily by itself for 90 days, in combination with the amino-terminal fragment of PTH-(1-34) (PTH) for 90 days, or sequentially after PTH when PTH was discontinued after 45 days of treatment. Additionally, comparisons were made of animals treated with PTH alone, 17alpha-ethynyl estradiol alone, equine estrogens (Premarin) alone, raloxifene alone, or combinations of PTH and equine estrogens or raloxifene. Ovariectomy induced increases in the rate of bone turnover and body weight while decreasing bone mineral density, bone mineral content, bone strength, trabecular bone volume, trabecular thickness, trabecular number, and uterine weight. LY353381 x HCl at 0.01-1 mg/kg had marginal effects on body weight and no effect on uterine weight compared with those in ovariectomized controls, in contrast to 17alpha-ethynyl estradiol or equine estrogens. LY353381 x HCl prevented further bone loss due to ovariectomy in tibia, femora, and lumbar vertebra, like 17alpha-ethynyl estradiol but unlike equine estrogens. LY353381 x HCl prevented the resorption of trabecular bone spicules, like 17alpha-ethynyl estradiol, but inhibited bone formation activity to a lesser extent than 17alpha-ethynyl estradiol. In this model, 17alpha-ethynyl estradiol appeared to be more efficacious after 3 months of treatment than equine estrogens in the proximal tibia metaphysis, suggesting efficacy differences between metabolites of 17beta-estradiol in bone. PTH at 10 microg/kg had no effect on body weight or uterine weight, but significantly increased bone mass to beyond those in sham-operated controls, baseline controls, and groups receiving other individual treatments at both axial and appendicular sites. The combination of LY353381 x HCl and PTH increased bone mass at a faster rate and to a greater extent than PTH alone or the combinations of equine estrogens/PTH and raloxifene/PTH at trabecular bone sites. The LY353381 x HCl/PTH combination improved bone mass and quality beyond any agent alone in regions enriched for cancellous bone, but was not significantly better than PTH alone on cortical bone. Additionally, when PTH was discontinued at 45 days, LY353381 x HCl prevented the rapid loss of bone observed in controls. Therefore, LY353381 x HCl appears to be useful by itself, in combination, or in sequence with PTH to replace lost bone in postmenopausal women.

Online source: www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&list_uids=9794476&dopt=Abstract raloxifene Evista



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Effects of raloxifene in a guinea pig model for leiomyomas.

Porter KB, Tsibris JC, Porter GW, Fuchs-Young R, Nicosia SV, O'Brien WF, Spellacy WN.

Departments of Obstetrics and Gynecology, Biochemistry and Molecular Biology, and Pathology, University of South Florida College of Medicine, Tampa, Florida, USA.

OBJECTIVE: Chronic exposure of oophorectomized guinea pigs to 17beta-estradiol causes leiomyoma formation. Our aims were to determine whether these leiomyomas can become estradiol independent after exposure to estradiol and if raloxifene inhibits leiomyoma growth when given concomitantly with estradiol. STUDY DESIGN: To induce leiomyoma development, 6 oophorectomized animals received two estradiol implants for 140 days. Next, the estradiol implants were replaced with empty implants in 3 animals, whereas the other 3 received 2 new estradiol implants and raloxifene given per os 10 mg/kg per day for 60 days. Tumor size was monitored biweekly by ultrasonography. RESULTS: On estradiol removal, abdominal wall leiomyomas regressed within 15 to 30 days; when estradiol implants were reintroduced, leiomyomas redeveloped. Within 30 days on raloxifene, all abdominal leiomyomas (n = 9) regressed as determined by ultrasonography and verified at laparotomy. Serum raloxifene and estradiol levels were 432 +/- 46 pg/mL and 78 +/- 13 pg/mL (mean +/- SEM, n = 3), respectively, after 60 days of treatment. CONCLUSIONS: Leiomyomas did not become estradiol independent, even after long exposure to estradiol; ultrasonography allowed frequent, noninvasive assessment of leiomyoma size, and raloxifene rapidly regressed leiomyomas in this animal model.

Online source: www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&list_uids=9822517&dopt=Abstract raloxifene Evista



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Synthesis and binding affinities of fluoroalkylated raloxifenes.

Lee KC, Moon BS, Lee JH, Chung KH, Katzenellenbogen JA, Chi DY.

Department of Chemistry, Inha University, 253 Yonghyundong, Namgu, 402-751, Inchon, South Korea.

Three fluoroalkylated derivatives (1-3) of the selective estrogen receptor modulator (SERM), raloxifene, have been synthesized. The key step in the synthesis is the C-C bond formation of benzo[b]thiophene and a substituted phenyl group (ring C) using a Stille reaction. The in vitro binding affinities of the substituted raloxifenes 1-3 are 45, 60, 89%, respectively, relative to the affinity of estradiol, which is higher than the affinity of raloxifene itself (25%). When labeled with the positron-emitting radionuclide, these compounds might be useful as PET imaging agents for estrogen receptor-positive breast tumors.

Online source: www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&list_uids=12901910&dopt=Abstract raloxifene Evista



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Raloxifene induces neurite outgrowth in estrogen receptor positive PC12 cells.

Nilsen J, Mor G, Naftolin F.

Department of Obstetrics and Gynecology and Center for Reproductive Biology, Yale University School of Medicine, New Haven, Connecticut 06520-8063, USA.

OBJECTIVE: It is well established that gonadal steroids have direct in vivo and in vitro effects on neurons. To further study these effects, we used rat PC 12 cells to examine the effects of estrogen receptor (ER) ligands on neuronal morphology. DESIGN: PC 12 cells constitutively express ER beta, but only strongly express ER alpha after long-term priming with nerve growth factor (NGF). We therefore primed PC12 cells with NGF for 14 days before testing them for estradiol (10(-9)M)- and/or raloxifene (10(-7) M)-induced neurite growth. Neurite growth was assessed by quantitative light microscopy. As control, ER status of the PC12 cells was assessed by reverse transcription-polymerase chain reaction (RT-PCR). RESULTS: In this study, both estradiol and raloxifene induced the outgrowth of neurites in NGF-treated PC 12 cells (p < 0.05). The combination of estradiol- and raloxifene-induced neurite growth was statistically greater than the effects of either agent alone. RT-PCR confirms that NGF-treated PC 12 cells express both ERalpha and ERbeta. CONCLUSIONS: This report is the first on the neurotrophic effect of raloxifene. At 10(-7) M, raloxifene's effect equaled that of estradiol; moreover, raloxifene did not block the neurite growth of simultaneously estradiol-treated PC 12 cells, despite its functional antiestrogenic effects in vivo. We conclude that raloxifene is estrogen agonistic in this animal model and therefore studies are warranted to delineate the relationship between steroidal estrogen and raloxifene.

Online source: www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&list_uids=9872486&dopt=Abstract raloxifene Evista



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Both estrogen and raloxifene cause G1 arrest of vascular smooth muscle cells.

Takahashi K, Ohmichi M, Yoshida M, Hisamoto K, Mabuchi S, Arimoto-Ishida E, Mori A, Tsutsumi S, Tasaka K, Murata Y, Kurachi H.

Department of Obstetrics and Gynecology, Yamagata University School of Medicine, 2-2-2 Iidanishi, Yamagata, Yamagata 990-9585, Japan.

The proliferation of vascular smooth muscle cells (VSMC) is a crucial pathophysiological process in the development of atherosclerosis. Although estrogen is known to inhibit the proliferation of VSMC, the mechanism responsible for this effect remains to be elucidated. In addition, the effect of raloxifene on VSMC remains unknown. We have shown here that 17beta-estradiol (E(2)) and raloxifene significantly inhibited the platelet-derived growth factor (PDGF)-stimulated proliferation of cultured human VSMC. Flow cytometry demonstrated that PDGF-stimulated S-phase progression of the cell cycle in VSMC was also suppressed by E(2) or raloxifene. We found that PDGF-induced phosphorylation of retinoblastoma protein (pRb), whose hyperphosphorylation is a hallmark of the G1-S transition in the cell cycle, was significantly inhibited by E(2) and raloxifene. These effects were associated with a decrease in cyclin D1 expression, without a change in cyclin-dependent kinase 4 or cyclin-dependent kinase inhibitor, p27(kip1) expression. ICI 182,780 abolished the inhibitory effects of E(2) and raloxifene on PDGF-induced pRb phosphorylation. Next, we examined which estrogen receptor (ER) is necessary for these effects of E(2) and raloxifene. Since VSMC express both ERalpha and ERbeta, A10, a rat aortic smooth muscle cell line that expresses ERbeta but not ERalpha, was used. The dose-dependent stimulation of A10 cell proliferation by PDGF was not inhibited by E(2) or raloxifene in contrast to the results obtained in VSMC. Moreover, E(2) and raloxifene significantly inhibited the PDGF-induced cyclin D1 promoter activity in A10 cells transfected with cDNA for ERalpha but not in the parental cells. These results suggested that E(2) and raloxifene exert an antiproliferative effect in VSMC treated with PDGF, at least in part through inhibition of pRb phosphorylation, and that the inhibitory effects of E(2) and raloxifene may be mainly mediated by ERalpha.

Online source: www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&list_uids=12904179&dopt=Abstract raloxifene Evista