Effects of novel raloxifene analogs alone or in combination with mechanical loading in the Col1a2G610c/+ murine model of osteogenesis imperfecta.

Osteogenesis imperfecta (OI) is a hereditary bone disease in which gene mutations affect collagen formation, leading to a weak, brittle bone phenotype that can cause severe skeletal deformity and increased fracture risk. OI interventions typically repurpose osteoporosis medications to increase bone mass, but this approach does not address compromised tissue-level material properties. Raloxifene (RAL) is a mild anti-resorptive used to treat osteoporosis that has also been shown to increase bone strength by a-cellularly increasing bone bound water content, but RAL cannot be administered to children due to its hormonal activity. The goal of this study was to test a RAL analog with no estrogen receptor (ER) signaling but maintained ability to reduce fracture risk. The best performing analog from a previous analog characterization project, named RAL-ADM, was tested in an in vivo study. Female wildtype (WT) and Col1a2G610C/+ (G610C) mice were randomly assigned to treated or untreated groups, for a total of 4 groups (n = 15). Starting at 10 weeks of age, all mice underwent compressive tibial loading 3×/week to induce an anabolic bone formation response in conjunction with RAL-ADM treatment (0.5 mg/kg; 5×/week) for 6 weeks. Tibiae were scanned via microcomputed tomography then tested to failure in four-point bending. RAL-ADM had reduced ER affinity, and increased post-yield properties, but did not improve bone strength in OI animals, suggesting some properties can be improved by RAL analogs but further development is needed to create an analog with decidedly positive impacts to OI bone.

The potential renoprotective effect of Raloxifene in renal ischemia-reperfusion injury in a male rat model.

Renal ischemia-reperfusion injury is caused by a temporary reduction in oxygen-carrying blood flow to the kidney, followed by reperfusion. During ischemia, kidney tissue damage induces overproduction of reactive oxygen species, which produces oxidative stress. The blood flow restoration during the reperfusion period causes further production of reactive oxygen species that ends with apoptosis and cell death. This study aimed to investigate the potential renoprotective effects of Raloxifene on bilateral renal ischemia-reperfusion injury in rats by looking into kidney function biomarkers, urea and creatinine, inflammatory cytokines, such as tumor necrosis factor-alpha (TNF-α) and interleukin-1 beta (IL-1ß). Additionally, antioxidant markers such as total antioxidant capacity (TAC) and the pro-apoptotic marker caspase-3 were assessed. Histopathological scores were also employed for evaluation. Our experimental design involved 20 rats divided into four groups: the sham group underwent median laparotomy without ischemia induction, the control group experienced bilateral renal ischemia for 30 minutes followed by 2 hours of reperfusion, the vehicle group received pretreatment with a mixture of corn oil and dimethyl sulfoxide (DMSO) before ischemia induction, and the Raloxifene-treated group was administered Raloxifene at a dose of 10 mg/kg before ischemia induction, followed by ischemia-reperfusion. Urea and creatinine, TNF-α, IL-1ß, and caspase-3 in the Raloxifene group were significantly lower compared to the control and vehicle groups. On the other hand, TAC levels in the Raloxifene group were significantly higher than in the control and vehicle groups. This study concluded that Raloxifene had a renoprotective impact via multiple actions as an anti-inflammatory, anti-apoptotic, and antioxidant agent.

The Interplay of Raloxifene and Sonochemical Bio-Oss in Early Maxillary Sinus Bone Regeneration: A Histological and Immunohistochemical Analysis in Rabbits.

The study aimed to assess the efficacy of using Raloxifene with ultrasonic processing to enhance Bio-Oss®, a bone graft substitute, for maxillary sinus bone height reconstruction. A total of 24 rabbit maxillary sinuses were distributed into three groups, each receiving different treatments: Bio-Oss® only, sonicated Bio-Oss, and sonicated Bio-Oss® with Raloxifene. Surgical procedures and subsequent histomorphometric and immunohistochemistry analyses were conducted to evaluate the bone formation, connective tissue, and remaining biomaterial, as well as the osteoblastic differentiation and maturation of collagen fibers. Results indicated that the sonicated Bio-Oss® and Bio-Oss® groups showed similar histological behavior and bone formation, but the Raloxifene group displayed inflammatory infiltrate, low bone formation, and disorganized connective tissue. The statistical analysis confirmed significant differences between the groups in terms of bone formation, connective tissue, and remaining biomaterial. In conclusion, the study found that while sonicated Bio-Oss® performed comparably to Bio-Oss® alone, the addition of Raloxifene led to an unexpected delay in bone repair. The findings stress the importance of histological evaluation for accurate bone repair assessment and the necessity for further investigation into the local application of Raloxifene. Future research may focus on optimizing bone substitutes with growth factors to improve bone repair.

Endocrine Therapy for Primary and Secondary Prevention After Diagnosis of High-Risk Breast Lesions or Preinvasive Breast Cancer.

The Oncology Grand Rounds series is designed to place original reports published in the Journal into clinical context. A case presentation is followed by a description of diagnostic and management challenges, a review of the relevant literature, and a summary of the authors' suggested management approaches. The goal of this series is to help readers better understand how to apply the results of key studies, including those published in Journal of Clinical Oncology, to patients seen in their own clinical practice.Patients with high-risk breast lesions (HRLs) or preinvasive breast cancers face an elevated risk of future breast cancer diagnoses. Endocrine therapy in this setting reduces the risk of a future diagnosis but does not confer improved survival, thus the side effects of primary/secondary prevention must be considered relative to the benefits. Here, we discuss the available chemoprevention regimens for patients with HRLs and considerations for selecting a regimen, as well as the decision making surrounding use of adjuvant endocrine therapy for patients with ductal carcinoma in situ (DCIS). For patients with HRLs, available chemoprevention regimens differ by menopausal status, including tamoxifen 20 mg once daily for 5 years and more recently tamoxifen 5 mg once daily for 3 years in both premenopausal and postmenopausal women as well as raloxifene or aromatase inhibitors for postmenopausal women. We recommend a shared decision-making approach with attention to patient preferences related to risk tolerance and side-effect profiles. Low-dose tamoxifen appears to be a particularly favorable choice that is well tolerated, without risk of serious adverse events and offers comparable risk reduction to other regimens. For DCIS, the benefit of endocrine therapy in addition to radiation is small, and appears to be driven mainly by a reduction in contralateral breast diagnoses or new breast cancers. A strategy that reduces the side-effect profile of chemoprevention such as low-dose tamoxifen may be especially appealing in the setting of secondary prevention.

The Clinical Effectiveness of Denosumab (Prolia®) for the Treatment of Osteoporosis in Postmenopausal Women, Compared to Bisphosphonates, Selective Estrogen Receptor Modulators (SERM), and Placebo: A Systematic Review and Network Meta-Analysis.

To assess the effectiveness and safety of denosumab (Prolia®) compared to bisphosphonates (alendronate, ibandronate, risedronate, zoledronate), selective estrogen receptor modulators (SERMs; bazedoxifene, raloxifene) or placebo, for the treatment of osteoporosis in postmenopausal women (PMW). Systematic searches were run in PubMed, Embase & Cochrane Library on 27-April-2022. Randomized controlled trials (RCTs) that included osteoporotic PMW allocated to denosumab, SERMs, bisphosphonates, or placebo were eligible for inclusion. RCTs were appraised using Cochrane Risk of Bias 2.0. Bayesian network and/or pairwise meta-analyses were conducted on predetermined outcomes (i.e. vertebral/nonvertebral fractures, bone mineral density [BMD], mortality, adverse events [AEs], serious AEs (SAEs), withdrawals due to AEs, AEs caused by denosumab discontinuation). A total of 12 RCTs (k = 22 publications; n = 25,879 participants) were included in the analyses. Denosumab, reported a statistically significant increase in lumbar spine (LS) and total hip (TH) BMD, compared to placebo. Similarly, denosumab also resulted in a statistically significant increase in TH BMD compared to the raloxifene and bazedoxifene. However, relative to denosumab, alendronate, ibandronate and risedronate resulted in significant improvements in both femoral neck (FN) and LS BMD. With regards to vertebral fractures and all safety outcomes, there were no statistically significant differences between denosumab and any of the comparator. Relative to placebo, denosumab was associated with significant benefits in both LS and TH BMD. Additionally, denosumab (compared to placebo) was not associated with reductions in vertebral and nonvertebral fractures. Finally, denosumab was not associated with improvement in safety outcomes, compared to placebo. These findings should be interpreted with caution as some analyses suffered from statistical imprecision.

Vitamin D Receptor Gene Polymorphisms Affect Osteoporosis-Related Traits and Response to Antiresorptive Therapy.

The present study analyzed the effect of vitamin D receptor (VDR) gene polymorphisms (ApaI, TaqI, BsmI, FokI, and Cdx2) on bone mineral density (BMD), biochemical parameters and bone turnover markers, fracture prevalence, and response to three types of antiresorptive therapy (estrogen-progesterone, raloxifene, and ibandronate) in 356 postmenopausal women from Slovakia. Association analysis revealed a significant effect of BsmI polymorphism on lumbar spine BMD, serum osteocalcin (OC), and ß-CrossLaps levels. While ApaI and Cdx2 polymorphisms were associated with OC and alkaline phosphatase, TaqI polymorphism affected all turnover markers. ApaI, TaqI, and BsmI genotypes increased the risk of spinal, radial, or total fractures with odds ratios ranging from 2.03 to 3.17. Each of therapy types evaluated had a beneficial effect on all osteoporosis-related traits; however, the VDR gene affected only ibandronate and raloxifene treatment. ApaI/aa, TaqI/TT, and BsmI/bb genotypes showed a weaker or no response to ibandronate therapy in femoral and spinal BMD. The impact of aforementioned polymorphisms on turnover markers was also genotype dependent. On the contrary, only TaqI and BsmI polymorphisms influenced raloxifene therapy, even only in lumbar spine BMD. These results point to the potential of using the VDR gene in personalized pharmacotherapy of osteoporosis.

Effect of Bone Resorption Inhibitors on Serum Cholesterol Level and Fracture Risk in Osteoporosis: Randomized Comparative Study Between Minodronic Acid and Raloxifene.

The positive link between osteoporosis and hypercholesterolemia has been documented, and bone resorption inhibitors, such as nitrogen-containing bisphosphonates (N-BP) and selective estrogen receptor modulators (SERMs), are known to reduce serum cholesterol levels. However, the relationship between the baseline cholesterol level and incident fracture rate under the treatment using the bone resorption inhibitors has not been documented. We investigated the relation between vertebral fracture incident and the baseline cholesterol levels and cholesterol-lowering effect of N-BP and SERM in osteoporosis through a prospective randomized open-label study design. Patients with osteoporosis (n = 3986) were allocated into two groups based on the drug used for treatment: minodronic acid (MIN) (n = 1624) as an N-BP and raloxifene (RLX) as an SERM (n = 1623). Serum levels of cholesterol and incidence of vertebral fracture were monitored for 2 years. The vertebral fracture rates between the two groups were compared using the pre-specified stratification factors. The patients receiving MIN with baseline low-density lipoprotein (LDL)-cholesterol level of ≥ 140 mg/dL, high-density lipoprotein cholesterol level < 40 mg/dL, age group of ≥ 75 years, and T score of BMD ≥ -3 SD had significantly lower vertebral fracture rates than those receiving RLX (incidence rate ratios (IRR) 0.45 [95% confidence interval (CI) 0.30 0.75, p = 0.001], 0.25 [95% CI 0.09 0.65, p = 0.005], 0.71 [95% CI 0.56 0.91, p = 0.006], 0.47 [95% CI 0.30 0.75, p = 0.0012], respectively). The cholesterol-lowering effect was stronger in the RLX group than in the MIN group, regardless of prior statin use. These results indicated that MIN treatment was more effective in reducing fracture risk in patients with higher LDL cholesterol levels, although its cholesterol-lowering ability was lesser than the RLX treatment.Trial registration University Hospital Medical Information Network-Clinical Trials Registry (UMIN-CTR), No. UMIN000005433; date: April 13, 2011.

Efficacy of Raloxifene as Add-on Therapy on Disease Activity of Postmenopausal Women with Rheumatoid Arthritis: A Double-blind, Randomized, Placebo-controlled Clinical Trial.

OBJECTIVE: The current study aimed to evaluate the effect of raloxifene on the disease activity of postmenopausal patients with rheumatoid arthritis (RA) and the prevention of glucocorticoid- induced osteoporosis. METHODS: This double-blind, randomized clinical trial was conducted at the Rheumatic Diseases Research Center affiliated with Mashhad University of Medical Sciences from 2015 to 2016. Postmenopausal women with RA were randomly treated with raloxifene or placebo after discontinuation of alendronate. Disease activity was evaluated using DAS28ESR, HAQ, and VAS before and every two months after the intervention. In addition, bone mineral densitometry was performed for patients before and 14 months after the intervention. The disease activity and densitometric criteria were compared between the two groups at a significant level of p <0.05. RESULTS: A total of 17 patients were allocated to each group. The two groups were similar at baseline in underlying disease, age, duration of RA, duration of alendronate use, laboratory findings, and rheumatoid arthritis drugs. Moreover, the mean scores of DAS28ESR, HAQ, and VAS during visits were not significantly different between the intervention and control groups (p >0.05). CONCLUSION: The current study results could not prove any clinical benefits of adding raloxifene to standard therapies for patients with rheumatoid arthritis in improving their disease activity compared to placebo. CLINICAL TRIAL REGISTRATION NUMBER: Trial registration number is NCT02982083.

Estradiol and raloxifene as adjunctive treatment for women with schizophrenia: A meta-analysis of randomized, double-blind, placebo-controlled trials.

OBJECTIVES: We conducted a comprehensive meta-analysis of all available trials to evaluate the efficacy and safety of estrogen and selective estrogen receptor modulators as adjunctive treatment for women with schizophrenia. METHODS: Multiple databases were searched from the inception until March 2022. Only randomized, double-blind, placebo-controlled studies (randomized controlled trials) were included. Mean differences (MDs) and their 95% confidence intervals (CIs) were calculated using random effects models. RESULTS: The meta-analysis included six estradiol versus placebo studies (n = 724) and seven raloxifene versus placebo studies (n = 419), covering a total of 1143 patients. Adjunctive estradiol outperformed the placebo in terms of the Positive and Negative Syndrome Scale (PANSS) total score (MD = -7.29; 95% CI = -10.67 to -3.91; I2  = 59.1%; p < 0.001; k = 9; N = 858), positive symptom score (MD = -1.54; 95% CI = -3.04 to -0.72; I2  = 45.8%; p < 0.001; k = 7; N = 624), negative symptom score (MD = -1.9; 95% CI = -1.77 to -0.34; I2  = 37.6%; p < 0.05; k = 14; N = 1042), and general psychopathology score (MD = -4.27; 95% CI = -7.14 to -1.41; I2  = 76.3%; p < 0.005; k = 7; N = 624). Adjunctive raloxifene outperformed the placebo in terms of the PANSS total score (MD = -6.83; 95% CI = -11.69 to -1.97; I2  = 67.8%; p = 0.006; k = 8; N = 432) and general psychopathology score (MD = -3.82; 95% CI = -6.36 to -1.28; I2  = 65.3%; p < 0.005; k = 8; N = 432). CONCLUSIONS: Our meta-analysis showed that estradiol and raloxifene are effective and safe adjunctive treatments that improve schizophrenia symptoms in women. Moreover, the effects of estradiol and raloxifene differed in terms of timing and dosage. Both are promising adjunctive treatments that merit further study.

Combined Thermoneutral Housing and Raloxifene Treatment Improves Trabecular Bone Microarchitecture and Strength in Growing Female Mice.

Thermoneutral housing and Raloxifene (RAL) treatment both have potential for improving mechanical and architectural properties of bone. Housing mice within a 30 to 32 °C range improves bone quality by reducing the consequences of cold stress, such as shivering and metabolic energy consumption (Chevalier et al. in Cell Metab 32(4):575-590.e7, 2020; Martin et al. in Endocr Connect 8(11):1455-1467, 2019; Hankenson et al. in Comp Med 68(6):425-438, 2018). Previous work suggests that Raloxifene can enhance bone strength and geometry (Ettinger et al. in Jama 282(7):637-645, 1999; Powell et al. in Bone Rep 12:100246, 2020). An earlier study in our lab utilized long bones to examine the effect of thermoneutral housing and Raloxifene treatment in mice, but no significant interactive effects were found. The lack of an impact is hypothesized to be connected to the short 6-week duration of the study and the type of bone analyzed. This study will examine the same question within the axial skeleton, which has a higher proportion of trabecular bone. After 6 weeks of treatment with RAL, vertebrae from female C57BL/6 J mice underwent microcomputed tomography (µCT), architectural analysis, and compression testing. Most of the tested geometric properties (bone volume/tissue volume percent, trabecular thickness, trabecular number, trabecular spacing) improved with both the housing and RAL treatment. The effect sizes suggested an additive effect when treating mice housed under thermoneutral conditions. While ultimate force was enhanced with the treatment and housing, force normalized by bone volume fraction was not significantly different between groups. For longer pre-clinical trials, it may be important to consider the impacts of temperature on mice to improve the accuracy of these models.

Identifying actionable druggable targets for breast cancer: Mendelian randomization and population-based analyses.

BACKGROUND: Drug repurposing provides a cost-effective approach to address the need for breast cancer prevention and therapeutics. We aimed to identify actionable druggable targets using Mendelian randomization (MR) and then validate the candidate drugs using population-based analyses. METHODS: We identified genetic instruments for 1406 actionable targets of approved non-oncological drugs based on gene expression, DNA methylation, and protein expression quantitative trait loci (eQTL, mQTL, and pQTL, respectively). Genome-wide association study (GWAS) summary statistics were obtained from the Breast Cancer Association Consortium (122,977 cases, 105,974 controls). We further conducted a nested case-control study using data retrieved from Swedish registers to validate the candidate drugs that were identified from MR analyses. FINDINGS: We identified six significant MR associations with gene expression levels (TUBB, MDM2, CSK, ULK3, MC1R and KCNN4) and two significant associations with gene methylation levels across 21 CpG islands (RPS23 and MAPT). Results from the nested case-control study showed that the use of raloxifene (targeting MAPT) was associated with 35% reduced breast cancer risk (odds ratio, OR, 0.65; 95% confidence interval, CI, 0.51-0.83). However, usage of estradiol, tolterodine, and nitrofurantoin (also targeting MAPT) was associated with increased breast cancer risk, with adjusted ORs and 95% CI of 1.10 (1.07-1.13), 1.16 (1.09-1.24), and 1.09 (1.05-1.13), respectively. The effect of raloxifene and nitrofurantoin lost significance in further validation analyses using active-comparator and new-user design. INTERPRETATION: This large-scale MR analysis, combined with population-based validation, identified eight druggable target genes for breast cancer and suggested that raloxifene is an effective chemoprevention against breast cancer. FUNDING: Swedish Research Council, Cancerfonden, Crafoordska Stiftelsen, Allmänna Sjukhusets i Malmö Stiftelsen för bekämpande av cancer, 111 Project and MAS cancer.

Raloxifene potentiates the effect of gefitinib in triple-negative breast cancer cell lines.

Triple-negative breast cancers (TNBCs) are characterized by a lack of approved targeted therapies and remain a challenge in the clinic. Several overexpressed proteins, including epidermal growth factor receptor (EGFR), have been associated with TNBCs and are considered potential therapeutic targets. However, EGFR inhibitors alone failed to demonstrate a cutting-edge advantage for treating TNBCs over conventional chemotherapies. Studies have shown that selective estrogen receptor modulators (SERMs) tamoxifen and raloxifene also affect TNBC cell viability. The combination of gefitinib and raloxifene was assessed against TNBC cell lines in vitro. Two TNBC cell lines, MDA-MB-231 and MDA-MB-468, were used to investigate the combination of gefitinib and raloxifene on cell viability, DNA synthesis, and apoptosis. The combination was assessed on intracellular signaling pathways, colony formation, migration, and angiogenesis. In the present study, raloxifene, in combination with gefitinib, decreased cell viability. The combination potentiates apoptosis and affects the expression and phosphorylation pattern of proteins involved in cell proliferation, such as NFκB, ß-catenin, and EGFR. Furthermore, evidence of apoptosis activation was also observed, along with a decreased cell migration and tumorigenicity of TNBC cells. Moreover, the combined treatment decreased the ability of neovascularization as assessed by tube formation of endothelial cells. These results suggested the potential of the combination of raloxifene and gefitinib for the prevention of TNBC growth and the appearance of metastatic events. Our findings provide the basis for future studies on the mechanism involved in raloxifene-gefitinib inhibition of ER-negative tumor growth.

Patterns of Osteoporosis Medications Selection After Drug Holiday or Continued Therapy: A Real-World Experience.

OBJECTIVE: Published literature on physicians' preferences and sequential treatment patterns of osteoporosis therapy is scarce. METHODS: A retrospective cohort study of patients who received bisphosphonates, denosumab, and/or raloxifene for at least 3 consecutive years or teriparatide for at least 18 months for osteoporosis. Data collection spanned 10 years, from October 2007 to September 2016, at a tertiary care center in the United States. RESULTS: In total, 12 885 patients were identified on the basis of receiving at least 1 treatment at any point in time; 1814 patients were randomly reviewed, and 274 patients met the inclusion criteria. The mean age was 68.8 ± 10.7 years, and women represented 90.9% of all the cases. Primary care physicians and rheumatologists constituted 65.7% and 22.6% of the prescribers, respectively. Before instituting a drug holiday, alendronate was the most common initial treatment (percentage, mean duration ± standard deviation in years: 69%, 5.4 ± 2.4 years) followed by ibandronate (9.5%, 4.9 ± 2.1 years) and raloxifene (9.1%, 5.2 ± 1.6 years). Denosumab was the most common second course of treatment, accounting for 29.3% of 82 patients who were subsequently prescribed another therapy, followed by alendronate (24.4%) and zoledronate (20.7%). Among patients who were placed on a drug holiday and eventually restarted on osteoporosis therapy, denosumab was the most common treatment instituted (n = 21), accounting for 40% of the total patients, followed by alendronate (32%) and zoledronate (16%). There was a progressive decline in osteoporosis therapy over the duration of the study. CONCLUSION: Alendronate was the most common initial therapy. Denosumab was the most common second course of treatment prescribed.

Treatment patterns of long-dose-interval medication for persistent management of osteoporosis in Taiwan.

Treatment persistence was higher among the patients who initially received an anti-osteoporosis medication (AOM) with a long-dose-interval. PURPOSE: With long-dose-interval anti-osteoporosis medications (AOMs) available for osteoporosis management, it is important to evaluate persistence of any AOM as long as it is continuously used. The purpose of this study was to investigate the treatment pattern and persistence of AOMs, allowing for medication switch. METHODS: This study was an observational retrospective cohort study using Taiwan's National Health Insurance claims data. We selected patients who first initiated an AOM between January 1, 2013, and June 30, 2016. AOM therapy included alendronate, raloxifene, teriparatide, denosumab, zoledronate, and ibandronate; the latter three were categorized as long-dose-interval medications. Persistence was defined as continual prescription of any AOM at a given time point with a grace period of 45 days within which to obtain prescription refill. The competing risk model was used to examine the factors affecting patients switching their initial AOM. RESULTS: During the study period, 126,539 patients with mean age of 75 years met the inclusion criteria; 85% were female. For initial AOM, 43.3%, 25.6%, 14.6%, 9.3%, 5.3%, and 1.9% of the patients received alendronate, denosumab, raloxifene, zoledronate, ibandronate, and teriparatide, respectively. During a mean 36-month follow-up, 29.6% of the patients who received at least two AOM pharmacy claims throughout the study period have ever switched their initial medication. Long-dose-interval medications, mainly denosumab and zoledronate, were the preferred choice for medication switch. Treatment persistence was higher in patients who initiated with long-dose-interval AOMs. CONCLUSION: The real-world data reveal long-dose-interval therapy as an initial treatment or at the first switch stage may improve management of persistent AOM treatment.

Bazedoxifene, a Postmenopausal Drug, Acts as an Antimalarial and Inhibits Hemozoin Formation.

Despite a remarkable improvement in health care and continued drug discovery efforts, malaria control efforts are continuously challenged by the emergence of drug-resistant parasite strains. Given a long and risky development path of new drugs, repurposing existing drugs for the treatment of malaria is an attractive and shorter path. Tamoxifen, a selective estrogen receptor modulator (SERM) for the treatment and prevention of estrogen receptor-positive breast cancer, possesses antibacterial, antifungal, and antiparasitic activities. Hence, we assessed tamoxifen, raloxifene, and bazedoxifene, which represent the first-, second-, and third-generation SERMs, respectively, for antimalarial activity. Raloxifene and bazedoxifene inhibited the erythrocytic development of Plasmodium falciparum with submicromolar 50% inhibitory concentration (IC50) values. Among the three, bazedoxifene was the most potent and also decreased P. berghei infection in female mice but not in male mice. However, bazedoxifene similarly inhibited P. falciparum growth in erythrocytes of male and female origin, which highlights the importance of sex-specific host physiology in drug efficacy. Bazedoxifene was most potent on early ring-stage parasites, and about 35% of the treated parasites did not contain hemozoin in the food vacuole. Bazedoxifene-treated parasites had almost 34% less hemozoin content than the control parasites. However, both control and bazedoxifene-treated parasites had similar hemoglobin levels, suggesting that bazedoxifene inhibits hemozoin formation and that toxicity due to accumulation of free heme could be a mechanism of its antimalarial activity. Because bazedoxifene is in clinical use and bazedoxifene-chloroquine combination shows an additive antiparasitic effect, bazedoxifene could be an adjunctive partner of currently used antimalarial regimens. IMPORTANCE The emergence and spread of drug-resistant strains of the human malaria parasite Plasmodium falciparum has necessitated new drugs. Selective estrogen receptor modulators are in clinical use for the prevention and treatment of breast cancer and postmenopausal osteoporosis. We demonstrate that bazedoxifene, a third-generation selective estrogen receptor modulator, has potent inhibitory activity against both susceptible and drug-resistant strains of Plasmodium falciparum. It also blocked the development of Plasmodium berghei in mice. The inhibitory effect was strongest on the ring stage and resulted in the inhibition of hemozoin formation, which could be the major mechanism of bazedoxifene action. Hemozoin is a nontoxic polymer of heme, which is a by-product of hemoglobin degradation by the malaria parasite during its development within the erythrocyte. Because bazedoxifene is already in clinical use for the treatment of postmenopausal osteoporosis, our findings support repurposing of bazedoxifene as an antimalarial.

Characterization of raloxifene as a potential pharmacological agent against SARS-CoV-2 and its variants.

The new coronavirus SARS-CoV-2 is the causative agent of the COVID-19 pandemic, which so far has caused over 6 million deaths in 2 years, despite new vaccines and antiviral medications. Drug repurposing, an approach for the potential application of existing pharmaceutical products to new therapeutic indications, could be an effective strategy to obtain quick answers to medical emergencies. Following a virtual screening campaign on the most relevant viral proteins, we identified the drug raloxifene, a known Selective Estrogen Receptor Modulator (SERM), as a new potential agent to treat mild-to-moderate COVID-19 patients. In this paper we report a comprehensive pharmacological characterization of raloxifene in relevant in vitro models of COVID-19, specifically in Vero E6 and Calu-3 cell lines infected with SARS-CoV-2. A large panel of the most common SARS-CoV-2 variants isolated in Europe, United Kingdom, Brazil, South Africa and India was tested to demonstrate the drug's ability in contrasting the viral cytopathic effect (CPE). Literature data support a beneficial effect by raloxifene against the viral infection due to its ability to interact with viral proteins and activate protective estrogen receptor-mediated mechanisms in the host cells. Mechanistic studies here reported confirm the significant affinity of raloxifene for the Spike protein, as predicted by in silico studies, and show that the drug treatment does not directly affect Spike/ACE2 interaction or viral internalization in infected cell lines. Interestingly, raloxifene can counteract Spike-mediated ADAM17 activation in human pulmonary cells, thus providing new insights on its mechanism of action. A clinical study in mild to moderate COVID-19 patients (NCT05172050) has been recently completed. Our contribution to evaluate raloxifene results on SARS-CoV-2 variants, and the interpretation of the mechanisms of action will be key elements to better understand the trial results, and to design new clinical studies aiming to evaluate the potential development of raloxifene in this indication.

Implications and Efficacy of Aromatase Inhibitors in Combination and Monotherapy for the Treatment of Lung Cancer.

BACKGROUND: Lung tumors express high levels of aromatase enzyme compared to surrounding normal tissue. Inhibition of aromatase has emerged as a recent therapeutic approach for the treatment of breast cancer. However, the role of aromatase inhibition in lung cancer treatment requires further investigation. METHODS: The anti-proliferative effects of aromatase inhibitors were evaluated by MTT assay. Cell migration was assessed using a wound healing assay. The mechanism of cell death was determined using the annexin VFITC/ propidium iodide staining flow cytometry method. The soft agar colony formation assay evaluated cells' capability to form colonies. RESULT: Exemestane and curcumin significantly inhibited the growth of lung cancer cell lines in a dose- and timedependent manner. The IC50 values after 48 hours of treatment with exemestane were 176, 180, and 120 µM in A549, H661, and H1299, respectively. Curcumin IC50 values after 48 hours were 80, 43, and 68 µM in A549, H661, and H1299, respectively. The combined treatment of exemestane or curcumin with cisplatin, raloxifene, and celecoxib resulted in a synergistic effect in the A549 lung cell line with a combination index of less than 1, suggesting synergism. Exemestane resulted in approximately 96% inhibition of wound closure at 100 µM, while curcumin resulted in approximately 63% inhibition of wound closure at 50 µM. Exemestane and curcumin inhibited the formation of cell colonies by reducing the number and size of formed colonies of A549, H661, and H1299 cell lines in a concentration dependent manner. Exemestane and curcumin had significantly induced apoptosis in A549 cells compared to control of untreated cells. CONCLUSION: Aromatase inhibition by exemestane or curcumin had significantly inhibited the growth of lung cancer cell lines, synergized with cisplatin, raloxifene, and celecoxib, suppressed lung cancer cell migratory potential, induced apoptosis, and reduced colony formation of lung cancer cells.

Raloxifene Use After Denosumab Discontinuation Partially Attenuates Bone Loss in the Lumbar Spine in Postmenopausal Osteoporosis.

Discontinuation of denosumab (DMab) is associated with decline in bone density. Whether raloxifene can be effective to attenuate bone loss after DMab discontinuation in certain conditions when other antiresorptives cannot be used remains unclear. Data on postmenopausal women with osteoporosis who discontinued DMab treatment after short-term use (1-to-4 doses) at Severance Hospital, Seoul, Korea, between 2017 and 2021 were reviewed. Changes in bone mineral density (BMD) at 12 months after DMab discontinuation was compared between sequential raloxifene users (DR) and those without any sequential antiresorptive (DD) after 1:1 propensity score matching. In matched cohort (66 patients; DR n = 33 vs. DD n = 33), mean age (69.3 ± 8.2 years) and T-score (lumbar spine - 2.2 ± 0.7; total hip - 1.6 ± 0.6) did not differ between two groups at the time of DMab discontinuation. Sequential treatment to raloxifene in DR group attenuated the bone loss in lumbar spine after DMab discontinuation compared to DD group (DR vs. DD; - 2.8% vs. - 5.8%, p = 0.013). The effect of raloxifene on lumbar spine BMD changes remained robust (adjusted ß + 2.92 vs. DD, p = 0.009) after adjustment for covariates. BMD loss at femoral neck (- 1.70% vs. - 2.77%, p = 0.673) and total hip (- 1.42% vs. - 1.44%, p = 0.992) did not differ between two groups. Compared to BMD at DMab initiation, DR partially retained BMD gain by DMab treatment in lumbar spine (+ 3.7%, p = 0.003) and femoral neck (+ 2.8%, p = 0.010), whereas DD did not. Raloxifene use after DMab treatment attenuated lumbar spine BMD loss in postmenopausal women with short exposures (< 2 years) to DMab.

Raloxifene, a cannabinoid type-2 receptor inverse agonist, mitigates visual deficits and pathology and modulates microglia after ocular blast.

Visual deficits after ocular blast injury (OBI) are common, but pharmacological approaches to improve long-term outcomes have not been identified. Blast forces frequently damage the retina and optic nerves, and work on experimental animals has shown the pro-inflammatory actions of microglia can further exacerbate such injuries. Cannabinoid type-2 receptor (CB2) inverse agonists specifically target activated microglia, biasing them away from the harmful pro-inflammatory M1 state toward the helpful reparative M2 state. We previously found that treating mice with CB2 inverse agonists after traumatic brain injury, produced by either focal cranial air blast or dorsal cranial impact, greatly attenuated the visual deficits and pathology that otherwise resulted. Here we examined the consequences of single and repeat OBI and the benefit provided by raloxifene, an FDA-approved estrogen receptor drug that possesses noteworthy CB2 inverse agonism. After single OBI, although the amplitudes of the A- and B-waves of the electroretinogram and pupil light response appeared to be normal, the mice showed hints of deficits in contrast sensitivity and visual acuity, a trend toward optic nerve axon loss, and significantly increased light aversion, which were reversed by 2 weeks of daily treatment with raloxifene. Mice subjected to repeat OBI (5 blasts spaced 1 min apart), exhibited more severe visual deficits, including decreases in contrast sensitivity, visual acuity, the amplitudes of the A- and B-waves of the electroretinogram, light aversion, and resting pupil diameter (i.e. hyperconstriction), accompanied by the loss of photoreceptor cells and optic nerve axons, nearly all of which were mitigated by raloxifene. Interestingly, optic nerve axon abundance was strongly correlated with contrast sensitivity and visual acuity across all groups of experimental mice in the repeat OBI study, suggesting optic nerve axon loss with repeat OBI and its attenuation with raloxifene are associated with the extent of these two deficits while photoreceptor abundance was highly correlated with A-wave amplitude and resting pupil size, suggesting a prominent role for photoreceptors in these two deficits. Quantitative PCR (qPCR) showed levels of M1-type microglial markers (e.g. iNOS, IL1ß, TNFα, and CD32) in retina, optic nerve, and thalamus were increased 3 days after repeat OBI. With raloxifene treatment, the overall expression of M1 markers was more similar to that in sham mice. Raloxifene treatment was also associated with the elevation of IL10 transcripts in all three tissues compared to repeat OBI alone, but the results for the three other M2 microglial markers we examined were more varied. Taken together, the qPCR results suggest that raloxifene benefit for visual function and pathology was associated with a lessening of the pro-inflammatory actions of microglia. The benefit we find for raloxifene following OBI provides a strong basis for phase-2 efficacy testing in human clinical trials for treating ocular injury.

The effects of raloxifene on endothelial function and Inflammation in Postmenopausal women: A Meta-analysis of randomized controlled trials.

BACKGROUND AND AIM: Raloxifene treatment has been reported to be associated with cardiovascular benefits if prescribed to women during the postmenopausal period. However, a final conclusion regarding this hypothesis has not yet been achieved. We conducted a systematic review and meta-analysis to evaluate the effect of raloxifene on the endothelial function and inflammation in postmenopausal women. METHODS: We systematically searched the following 4 databases from inception to 23 January 2021 without any language restrictions: Web of Science, PubMed/Medline, Embase and Scopus. The eligible randomized controlled trials (RCTs) reporting the effects of raloxifene on the flow-mediated dilatation (FMD), C-reactive protein (CRP), carotid intima-media thickness (CIMT), intercellular adhesion molecule 1 (ICAM-1), vascular cell adhesion molecule 1 (VCAM-1) and E-selectin levels, were included in the final meta-analysis. RESULTS: A total of 16 RCTs were included in the final analysis. Raloxifene administration had no significant effect on ICAM-1 and E-selectin levels. However, we observed a decrease of the CIMT (WMD: -0.071 mm, 95% CI: -0.09 to -0.04, P = 0.000), CRP (WMD: -0.342 mg/L, 95% CI: -0.591, -0.094, p = 0.007), and VCAM-1 (WMD: -197.90 mg/L, 95% CI: -269.58 to -126.23, P = 0.000) levels in the intervention versus control groups following the prescription of this pharmacological agent. Moreover, raloxifene treatment resulted in a significant elevation of the FMD (WMD: 1.64%, 95% CI: 0.46 to 2.81, P = 0.006), particularly if the intervention was equal to or exceeded 12 weeks. CONCLUSION: Raloxifene might emerge as a potential therapeutic option in the management of endothelial dysfunction and inflammation in postmenopausal women.