Vitamin D receptor agonist VS-105 improves cardiac function in the presence of enalapril in 5/6 nephrectomized rats.

Vitamin D receptor (VDR) agonists (VDRAs) are commonly used to manage hyperparathyroidism secondary to chronic kidney disease (CKD). Patients with CKD experience extremely high risks of cardiovascular morbidity and mortality. Clinical observations show that VDRA therapy may be associated with cardio-renal protective and survival benefits in patients with CKD. The 5/6 nephrectomized (NX) Sprague-Dawley rat with established uremia exhibits elevated serum parathyroid hormone (PTH), hypertension, and abnormal cardiac function. Treatment of 5/6 NX rats with VS-105, a novel VDRA (0.05 and 0.5 µg/kg po by gavage), once daily for 8 wk in the presence or absence of enalapril (30 mg/kg po via drinking water) effectively suppressed serum PTH without raising serum calcium. VS-105 alone reduced systolic blood pressure (from 174 ± 6 to 145 ± 9 mmHg, P < 0.05) as effectively as enalapril (from 174 ± 6 to 144 ± 7 mmHg, P < 0.05). VS-105 improved cardiac functional parameters such as E/A ratio, ejection fraction, and fractional shortening with or without enalapril. Enalapril or VS-105 alone significantly reduced left ventricular hypertrophy (LVH); VS-105 plus enalapril did not further reduce LVH. VS-105 significantly reduced both cardiac and renal fibrosis. The lack of hypercalcemic toxicity of VS-105 is due to its lack of effects on stimulating intestinal calcium transport and inducing the expression of intestinal calcium transporter genes such as Calb3 and TRPV6. These studies demonstrate that VS-105 is a novel VDRA that may provide cardiovascular benefits via VDR activation. Clinical studies are required to confirm the cardiovascular benefits of VS-105 in CKD.

Two novel vitamin D receptor modulators with similar structures exhibit different hypercalcemic effects in 5/6 nephrectomized uremic rats.

BACKGROUND/AIMS: Vitamin D receptor modulators (VDRMs) are indicated for secondary hyperparathyroidism in chronic kidney disease (CKD). Clinical observations demonstrate that VDRM therapy provides cardiovascular (CV) benefit in CKD. Current on-market VDRMs have a narrow therapeutic index at 1- to 4-fold [hypercalcemic toxicity vs. parathyroid hormone (PTH)-suppressing efficacy]. Hypercalcemia leads to the need for frequent drug dose titration and serum calcium (Ca) monitoring. A VDRM with a wider therapeutic index and beneficial CV effects will be clinically useful. METHODS: Two structurally similar VDRMs were tested in the 5/6 nephrectomized (NX) rats with elevated PTH, endothelial dysfunction and left ventricular hypertrophy. RESULTS: VS-110 and VS-411 at 0.01-1 µg/kg (i.p. 3 times/week for 2 weeks) suppressed serum PTH effectively. VS-411 raised serum Ca with an 11% increase at 0.01 µg/kg (therapeutic index = ~1-fold), while VS-110 did not raise serum Ca even at 1 µg/kg (therapeutic index >50-fold). VS-110 improved endothelium-dependent aortic relaxation in a dose-dependent manner and significantly reduced left ventricular fibrosis without affecting serum Ca. VS-411 also exhibited effects on the CV parameters, but was less potent at the high doses with severe hypercalcemia. VS-110 and VS-411 specifically activated the reporter gene via a chimeric receptor containing the VDR ligand binding domain with EC(50) <0.1 nM. CONCLUSIONS: Structurally similar VDRMs can exhibit distinctly different hypercalcemic effects in 5/6 NX uremic rats. While differences exist for the Ca and CV effects of VS-110 and VS-411, the clinical implications are unclear. VS-110's results are promising but clinical outcome studies need to be performed.

Synthesis of VS-105: A novel and potent vitamin D receptor agonist with reduced hypercalcemic effects.

We have synthesized a novel vitamin D receptor agonist VS-105 ((1R,3R)-5-((E)-2-((3αS,7αS)-1-((R)-1-((S)-3-hydroxy-2,3-dimethylbutoxy)ethyl)-7α-methyldihydro-1H-inden-4(2H,5H,6H,7H,7αH)-ylidene)ethylidene)-2-methylenecyclohexane-1,3-diol). Preparation of a-ring phenylphosphine oxide 11, followed by Wittig-Horner coupling of 11 with the protected 25-hydroxy Grundmann's ketone 22 generated the precursor 12. Deprotection of the TBDMS groups of 12 produced the target compound VS-105. The biological profiles of VS-105 were evaluated using in vitro assays (VDR receptor binding, VDR reporter gene and HL-60 differentiation) in comparison to calcitriol (the endogenous hormone) or paricalcitol. Furthermore, the PTH suppressing potency and hypercalcemic side effects of VS-105 were evaluated in the 5/6 nephrectomized uremic rats in comparison to paricalcitol. Combining various changes at 20-epi, 22-oxa, 24-methyl, and 2-methylene yielded VS-105 that not only is highly potent in inducing functional responses in vitro, but also effectively suppresses PTH in a dose range that does not affect serum calcium in the 5/6 nephrectomized uremic rats.

Differential effects of vitamin d receptor agonists on gene expression in neonatal rat cardiomyocytes.

PURPOSE: Vitamin D receptor (VDR) activation is associated with cardiovascular benefits in chronic kidney disease patients, but whether VDR's hormone and prehormone exhibit similar effects requires more studies. METHODS: Neonatal rat cardiomyocytes were treated with VDR agonists (calcitriol and/or paricalcitol) and the prehormone calcidiol in the presence of aldo (1 µM). The expression of VDR target genes were determined by real-time PCR and Western blotting. The expression and activity of CYP27B1 (the enzyme responsible for converting calcidiol to calcitriol) was measured. RESULTS: Treating cells with aldo (1 µM) for 24 h significantly reduced the VDR mRNA (29%) and protein levels (>90%). Calcitriol and calcidiol induced VDR expression in the presence of aldo with EC(50) at 0.3 and 7,952 nM, respectively. Calcitriol, paricalcitol and calcidiol stimulated CYP24A1 (EC(50) at 6.4, 4.5 and 992 nM, respectively) and suppressed NPPB expression (IC(50) at 1.9, 0.1 and 210 nM, respectively) in the presence of 1 µM aldo. Neonatal rat cardiomyocytes expressed CYP27B1 and converted calcidiol to calcitriol at a low rate (~10% in 24 h). CONCLUSIONS: VDR hormone calcitriol and its analog paricalcitol exhibit more potent effects than the prehormone calcidiol in cardiomyocytes.

A Novel Vitamin D Receptor Agonist, VS-105, Improves Bone Mineral Density without Affecting Serum Calcium in a Postmenopausal Osteoporosis Rat Model.

BACKGROUND AND OBJECTIVES: VS-105, a novel vitamin D receptor agonist with significantly less hypercalcemic side effects than calcitriol, is a useful tool to investigate whether or not a vitamin D receptor agonist at non-hypercalcemic doses could improve bone mineral density (BMD). METHODS: VS-105 and calcitriol were evaluated in an ovariectomized (OVX) osteoporosis rat model and in calvariae bone organ culture. RESULTS: Treatment of OVX rats by VS-105 (0.1, 0.2 or 0.5 µg/kg, intraperitoneal, 3×/week, for 90 days) significantly improved BMD in the L3 lumbar vertebra in a dose-dependent manner (sham vs. OVX/vehicle: 324 ± 14 vs. 279 ± 10 mg/cm2; VS-105 at 0.1, 0.2 and 0.5 µg/kg: 306 ± 9, 329 ± 12, and 327 ± 10 mg/cm2, respectively) without affecting serum calcium (Ca). Calcitriol at 0.1 µg/kg significantly increased BMD but it also increased serum Ca. VS-105 and calcitriol at the test doses significantly suppressed serum parathyroid hormone and promoted tibia bone growth. With respect to biomarkers of bone remodeling, calcitriol and VS-105 both significantly elevated serum osteocalcin. In the calvariae bone organ culture, net Ca release was significantly less in VS-105-treated groups (vs. calcitriol). CONCLUSIONS: VS-105 is efficacious in improving BMD in a dose range that does not affect serum Ca in OVX rats; the improvement in BMD by VS-105 is attributable to increased osteoblastic activity and reduced osteoclastic bone resorption.

Endothelial dysfunction and chronic kidney disease: treatment options.

Endothelial cells detect physical and chemical changes in the blood vessels, and release various factors to counter these changes to maintain homeostasis. Traditional cardiovascular disease risk factors, such as hypertension, dyslipidemia and diabetes, cause endothelial dysfunction characterized by off-balanced vasodilation/vasoconstriction, increased oxidative stress and inflammation, deregulation of thrombosis and fibrinolysis, abnormal smooth muscle cell proliferation, and a deficient repair mechanism. Patients with chronic kidney disease (CKD) have a much higher risk of cardiovascular disease and mortality than the general population. Endothelial dysfunction is commonly observed in CKD, likely preceding other cardiovascular complications. Lipid-lowering agents, such as statins, improve endothelial functions and are effective in reducing cardiovascular disease risk in the general population, but have not demonstrated comparable efficacy in the CKD patient population. Similarly, antidiabetic agents, such as thiazolidinediones, that improve endothelial function in the general population are less efficacious than expected in slowing disease progression and reducing cardiovascular disease risk in CKD patients. Interestingly, agents that activate the vitamin D receptor (VDR) for the treatment of hyperparathyroidism secondary to CKD are associated with a survival benefit in CKD patients that is likely mediated through the effects of the VDR on modulating key components involved in endothelial dysfunction. However, a randomized, clinical study is required to confirm the survival benefit of VDR activation therapy for CKD patients. Results from clinical studies suggest that managing hypertension alone may not be adequate in slowing CKD progression and its related cardiovascular complications. Angiotensin-converting enzyme inhibitors and angiotensin receptor blockers that target the renin-angiotensin system slow CKD progression, possibly due to their effects on improving endothelial function, independent of controlling blood pressure.

Vascular calcification in chronic kidney failure: role of vitamin D receptor.

Chronic kidney disease (CKD) patients encounter an increased risk of cardiovascular disease and mortality compared with healthy individuals, most likely due to the presence of severe atherosclerosis and accelerated vascular calcification. Vascular calcification is an active, regulated process resulting from an imbalance between losses in inhibitory factors and gains in inducing factors present in cells and the blood circulation. However, exactly which inhibitory and inducing factors are involved remains unknown. The vitamin D receptor (VDR) is a nuclear receptor present in over 30 different tissues. Several VDR activators (VDRAs), including paricalcitol and calcitriol, are currently available for the treatment of secondary hyperparathyroidism in patients with CKD. Recent clinical observations demonstrate that VDRA therapy provides survival benefits for CKD patients in the order of paricalcitol > calcitriol > no VDRA therapy, independent of serum parathyroid hormone, phosphorus and calcium levels. The survival benefit of VDRAs seems contradictory to the perception that VDRAs, due to their potential impact of increasing serum phosphorus and calcium, may cause calcification in vessels. A review of the current literature shows that inconsistent data exist regarding the role of VDRAs in vascular calcification. A possible explanation is that the VDR may be involved in regulating several different pathways as an endocrine, paracrine and/or autocrine factor, and different VDRAs may have differential effects on the endocrine versus the paracrine/autocrine aspect.

Differential effects of Vitamin D analogs on calcium transport.

It is well known that the efficiency of intestinal active calcium transport is regulated by the Vitamin D receptor pathway and Vitamin D analogs seem to exhibit differential effects on intestinal active calcium transport. To investigate the molecular basis for the difference among Vitamin D analogs, we tested three Vitamin D analogs: 1,25-dihydroxyvitamin D(3), 19-nor-1,25-dihydroxyvitamin D(2), and 1alpha-hydroxyvitamin D(2) ex vivo and in vitro. In 5/6 nephrectomized rat intestinal active calcium transport, 19-nor-1,25-dihydroxyvitamin D(2) did not show a significant effects on intestinal active calcium transport at all the concentrations tested, while 1alpha-hydroxyvitamin D(2) at 0.33 and 0.67 microg/kg and 1,25-dihydroxyvitamin D(3) at 1microg/kg significantly stimulated calcium transport. In Caco-2 cells, 19-nor-1,25-dihydroxyvitamin D(2) did not show a significant effect on calcium transport, while 1,25-dihydroxyvitamin D(3) and 1,25-dihydroxyvitamin D(2) (the active form of 1alpha-hydroxyvitamin D(2)) stimulated calcium transport by 934 and 501% at 0.1microM, respectively. 1,25-Dihydroxyvitamin D(2) potently induced the expression of CALB3 and TRPV6 mRNA with an EC(50) of 0.3 and 1.0nM, whereas 19-nor-1,25-dihydroxyvitamin D(2) was 10-fold less potent than 1,25-dihydroxyvitamin D(2) in inducing CALB3 and TRPV6 mRNA. The three Vitamin D analogs had no significant effect on the expression of PMCA1 mRNA. These Vitamin D analogs did not change the expression of Vitamin D receptor (VDR) up to 10nM, but stimulated CYP24A1 expression in a dose-dependent manner with the potency in the order of 1,25-dihydroxyvitamin D(3)>1,25-dihydroxyvitamin D(2)=19-nor-1,25-dihydroxyvitamin D(2). These results suggest that the differential effect of Vitamin D analogs on stimulating intestinal and Caco-2 calcium transport may be in part due to its different effect on stimulating CALB3 and TRPV6 mRNA expression.

Differential inhibition of renin mRNA expression by paricalcitol and calcitriol in C57/BL6 mice.

BACKGROUND/AIMS: Vitamin D receptor activators (VDRAs) may suppress renin expression and VDR-mediated renin inhibitors may offer a novel mechanism to control the RAS. METHODS: We delineated the effects of paricalcitol and calcitriol on PTH, renin, and iCa(2+) in C57/BL6 mice administered vehicle, paricalcitol, or calcitriol (0.01, 0.03, 0.10, 0.33, 1.0 microg/kg s.c.) 3 days/week for 9 days. RESULTS: Paricalcitol produced PTH suppression from 0.03 to 1.0 microg/kg (values between 9.7 +/- 3.3 and 20.7 +/- 4.7 pg/ml; vehicle = 88.0 +/- 16.9) and elicited dose-dependent reductions in renin/GAPDH expression at 0.33 and 1.0 microg/kg (0.037 +/- 0.002, 0.027 +/- 0.003; vehicle = 0.054 +/- 0.003) but produced no increases iCa(2+) at any dose tested. Calcitrol produced PTH suppression at all doses tested (between 6.4 +/- 1.2 and 29.5 +/- 17.2 pg/ml) and renin suppression at 0.10, 0.33, and 1.0 microg/kg (0.029 +/- 0.002, 0.031 +/- 0.003, and 0.038 +/- 0.02). However, at 0.33 and 1.0 mg/kg, calcitriol produced increases iCa(2+) (1.31 +/- 0.03 and 1.48 +/- 0.02 mmol/l; vehicle = 1.23 +/- 0.02 mmol/l). CONCLUSIONS: Paricalcitol produces significant, dose-dependent suppression of renin expression in the absence of hypercalcemia at doses 10-fold above those necessary for PTH suppression. Calcitriol also produced suppression of renin at doses at least 10-fold above those required for PTH suppression, but increases in iCa(2+) were observed at doses only 3-fold above those necessary to elicit renin suppression.

Vitamin D receptor agonist VS-105 directly modulates parathyroid hormone expression in human parathyroid cells and in 5/6 nephrectomized rats.

Vitamin D receptor (VDR) agonists (VDRAs) are commonly used to treat secondary hyperparathyroidism (SHPT) associated with chronic kidney disease (CKD). Current VDRA therapy often causes hypercalcemia, which is a critical risk for vascular calcification. Previously we have shown that a novel VDRA, VS-105, effectively suppresses serum parathyroid hormone (PTH) without affecting serum calcium levels in 5/6 nephrectomized (NX) uremic rats. However, it is not known whether VS-105 directly regulates PTH gene expression. To study the direct effect of VS-105 on modulating PTH, we tested VS-105 and paricalcitol in the spheroid culture of parathyroid cells from human SHPT patients, and examined the time-dependent effect of the compounds on regulating serum PTH in 5/6 NX uremic rats (i.p. 3x/week for 14days). In human parathyroid cells, VS-105 (100nM) down-regulated PTH mRNA expression (to 3.6% of control) and reduced secreted PTH (to 43.9% of control); paricalcitol was less effective. VS-105 effectively up-regulated the expression of VDR (1.9-fold of control) and CaSR (1.8-fold of control) in spheroids; paricalcitol was also less effective. In 5/6 NX rats, one single dose of 0.05-0.2µg/kg of VS-105 or 0.02-0.04µg/kg of paricalcitol effectively reduced serum PTH by >40% on Day 2. Serum PTH remained suppressed during the dosing period, but tended to rebound in the paricalcitol groups. These data indicate that VS-105 exerts a rapid effect on suppressing serum PTH, directly down-regulates the PTH gene, and modulates PTH, VDR and CaSR gene expression more effectively than paricalcitol.

Effects of Vitamin D analogs on gene expression profiling in human coronary artery smooth muscle cells.

Vitamin D analogs provide survival benefit for chronic kidney disease patients with cardiovascular complications. Activation of smooth muscle cells plays a role in cardiovascular diseases. It is not known how Vitamin D analogs modulate gene expression in smooth muscle cells. In this study, DNA microarray technology was used to assess the gene expression profile in human coronary artery smooth muscle cells treated with 0.1microM 1alpha,25-dihydroxyvitamin D3 (calcitriol) or paricalcitol (an analog of calcitriol) for 30 h. The effects of calcitriol and paricalcitol were similar. A total of 176 target genes were identified with 115 up-regulated and 61 down-regulated genes in the paricalcitol group. Target genes fall into various categories including cell differentiation/proliferation. Real-time RT-PCR analysis demonstrated that paricalcitol dose- and time-dependently regulated the expression of IGF1, WT1 and TGFbeta3, three genes known to modulate cell proliferation. Paricalcitol also down-regulated the expression of natriuretic peptide precursor B and thrombospondin 1. Both drugs inhibited cell proliferation in a dose-dependent manner. This study identified genes not previously known to be regulated by VDR, providing insight into understanding the role of VDR on regulating smooth muscle cell growth, thrombogenicity, fibrinolysis and endothelial regeneration.

Cardiovascular disease in chronic kidney failure: the role of VDR activators.

Vitamin D3 is modified by vitamin D3 25-hydroxylase in the liver, and by 25-hydroxyvitamin D3 1alpha-hydroxylase (CYP27B1) in the kidney, to form the active metabolite 1alpha,25-dihydroxyvitamin D3. Several vitamin D receptor (VDR) activators, including paricalcitol and calcitriol, are currently available for the treatment of hyperparathyroidism secondary to chronic kidney disease (CKD). CKD patients encounter a much higher risk of cardiovascular disease than do members of the general public, and recent clinical observations have shown that VDR activator therapy provides survival benefit for CKD patients in the rank order of paricalcitol > calcitriol > no VDR activator therapy, independent of parathyroid hormone, phosphorus and calcium. One possible explanation for this observation is that VDR activators exert a positive impact on cardiovascular functions. Studies in animals with disrupted genes involved in the vitamin D signaling pathway have provided some interesting data. For example, in mice lacking VDR or CYP27B1, it was found that in addition to the expected phenotype (hypocalcemia, secondary hyperparathyroidism and osteomalacia), expression of renin or atrial natriuretic peptide was elevated. The mice also developed hypertension and cardiac hypertrophy. Gene expression profiling studies have revealed that VDR may play a role in regulating smooth-muscle-cell (SMC) proliferation, thrombosis, fibrinolysis and vessel relaxation. Paricalcitol and calcitriol are equally potent at suppressing plasminogen activator inhibitor-1 synthesis and inhibiting cellular proliferation in human coronary artery SMCs. The effect of VDR activators on the modulation of renin expression and vascular functions may be factors that contribute to reduced mortality and morbidity risk in VDR-activator-treated CKD patients. In this review, we discuss recent preclinical and clinical data regarding the role of VDR and its ligands in the cardiovascular system.

Differential effects of Vitamin D analogs on bone formation and resorption.

Deficiency in Vitamin D and its metabolites leads to a failure in bone formation primarily caused by dysfunctional mineralization, suggesting that Vitamin D analogs might stimulate osteoblastic bone formation and mineralization. In this study, we compare the effect of selected Vitamin D analogs and active metabolite, 1alpha,25-dihydroxyvitamin D(3), 19-nor-1alpha, 25-dihydroxyvitamin D(2), and 1alpha-hydroxyvitamin D(2) or 1alpha,25-dihydroxyvitamin D(2) on bone formation and resorption. In a mouse calvariae bone primary organ culture system, all Vitamin D analogs and metabolite tested-stimulated collagen synthesis in a dose-dependent manner and 19-nor-1alpha, 25-dihydroxyvitamin D(2) was the most efficacious among three. 19-nor-1alpha, 25-dihydroxyvitamin D(2) and 1alpha,25-dihydroxyvitamin D(2) showed similar potencies and 1alpha,25-dihydroxyvitamin D(3) was less potent than others. Osteocalcin was also up-regulated in a dose-dependent manner, suggesting that the three Vitamin D analogs have the equal potencies on bone formation. 25-Hydroxyvitamin D-24-hydroxylase expression was induced in a dose-dependent manner and 19-nor-1alpha, 25-dihydroxyvitamin D(2) was less potent than other two compounds. In a mouse calvariae organ culture, all induced a net calcium release from calvariae in a dose-dependent manner, but the potency is in the order of 1alpha,25-dihydroxyvitamin D(2) congruent with1alpha,25-dihydroxyvitamin D(3)>19-nor-1alpha, 25-dihydroxyvitamin D(2). In a Vitamin D/calcium-restricted rat model, all caused an elevation in serum calcium in a dose-dependent manner. There is no significant difference between 1alpha,25-dihydroxyvitamin D(3) and 1alpha-hydroxyvitamin D(2) in potencies, but 19-nor-1alpha, 25-dihydroxyvitamin D(2) is at least 10-fold less potent than the other two compounds. Our results suggest that Vitamin D analogs have direct effects on bone resorption and formation, and 19-nor-1alpha, 25-dihydroxyvitamin D(2) may be more effective than 1alpha,25-dihydroxyvitamin D(3) and 1alpha-hydroxyvitamin D(2) on stimulating anabolic bone formation.

Expression of VDR and CYP24A1 mRNA in human tumors.

1,25-dihydroxyvitamin D3 (1,25(OH)2D3) and its analogues have been shown to inhibit proliferation of human cancer cells mediated by vitamin D receptor (VDR). The over-expression of 25-hydroxyvitamin D-24-hydroxylase (CYP24A1), an enzyme involved in the metabolism of 1,25(OH)2D3 and its analogues, is associated with poor prognosis of some human cancers. In this study, we employed real-time reverse transcription PCR to examine the expression of VDR and CYP24A1 mRNA in a cohort of human breast, lung, colon and ovary tumor samples. We found that CYP24A1 mRNA was significantly up-regulated in colon, ovary and lung tumors, but down-regulated in breast tumor relative to the analogous normal tissues. As a comparison, VDR mRNA was modestly down-regulated in colon, breast and lung tumors, but highly up-regulated in ovarian tumors. Treatment of two breast cancer cell lines, SW-620 and MCF-7, and one colon cancer cell line, HT-29, by 1,25(OH)2D3 for 48 h profoundly stimulated CYP24A1 mRNA expression (EC50=0.6, 0.8 and 29.5 nM in SW-620, HT-29 and MCF-7, respectively), but did not significantly affect VDR mRNA expression. Growth as assessed by DNA synthesis was modestly arrested by 1,25(OH)2D3 after 72 h of incubation, but was not altered after a 5-day incubation period. These data suggest that the VDR signaling pathway may be compromised via the modulation of CYP24A1 and VDR in human tumors.

Vitamin D receptor (VDR) localization in human promyelocytic leukemia cells.

Although vitamin D analogs are known to induce the differentiation of the HL-60 promyelocytic leukemia cells, the effect of vitamin D analogs on the distribution of vitamin D receptor (VDR) in these cells is not well studied. This report showed, by confocal microscopy, that VDR mainly resided in the cytoplasm in the absence of VDR ligands. When cells were treated with 19-nor-1alpha,25-(OH)(2)D(2) or 1,25(OH)(2)D(3), VDR moved from the cytoplasm into the nucleus in a time-dependent manner. VDR could be observed in the nucleus as early as 6 h after drug treatment and was still observed in the nucleus 3 days after one single addition of 100 nM 19-nor-1alpha,25-(OH)(2)D(2) or 1,25(OH)(2)D(3). The VDR protein level was significantly increased by 19-nor-1alpha,25-(OH)(2)D(2) or 1,25(OH)(2)D(3) in a dose-dependent manner, while the VDR mRNA level was not affected by either compound. These results suggest that binding of vitamin D analogs to VDR induced receptor translocation into the nucleus, which stabilizes the receptor, resulting in an accumulation of the VDR protein.

Effects of vitamin D analogs on the expression of plasminogen activator inhibitor-1 in human vascular cells.

INTRODUCTION: Vitamin D analogs such as paricalcitol and calcitriol have been shown to provide survival benefit for Stage 5 chronic kidney disease (CKD) patients, possibly due to their positive impact on the cardiovascular system. Plasminogen activator inhibitor-1 (PAI-1) is one of the risk markers for coronary artery disease. MATERIALS AND METHODS: Human coronary artery smooth muscle cells (SMC) and endothelial cells (CAEC) were treated with vitamin D analogs to assess the effects of the drugs on the expression of PAI-1 mRNA and protein. RESULTS: In SMC, both paricalcitol and calcitriol down-regulated the expression of PAI-1 mRNA and protein in a dose-dependent manner. The EC(50) values of paricalcitol and calcitriol on suppressing PAI-1 mRNA were 3.0 and 2.8 nM, respectively. Interestingly, these two drugs had no significant effect on the expression of PAI-1 protein or mRNA in CAEC. Further analysis showed that CAEC did not express functional vitamin D receptor (VDR) and paricalcitol failed to induce the expression of 25-hydroxyvitamin D-24-hydroxylase (CYP24A1) mRNA, a gene known to be regulated by VDR. As a comparison, SMC expressed VDR and paricalcitol induced CYP24A1 mRNA in SMC (>150-fold at 10 nM) dose-dependently. The effect of paricalcitol on suppressing PAI-1 in SMC was blocked by cycloheximide, suggesting that protein synthesis was involved. CONCLUSION: These results demonstrate that vitamin D analogs suppress PAI-1 in SMC, but not in CAEC. Suppression of PAI-1 in SMC may be one of the factors contributing to the survival benefits of vitamin D analog therapy in CKD patients.

Preclinical studies of VS-505: a non-absorbable highly effective phosphate binder.

BACKGROUND AND PURPOSE: Phosphate imbalance is often present in chronic kidney disease (CKD), and it contributes to a higher cardiovascular mortality rate. A phosphate binder is typically part of a treatment strategy for controlling phosphate imbalance. However, safety concerns and low compliance are two well-recognized disadvantages of on-market phosphate binders. This report describes the preclinical studies of VS-505, a non-absorbable, calcium- and aluminum-free, plant-derived polymer currently being evaluated in haemodialysis patients in Australia. EXPERIMENTAL APPROACH: Normal Sprague Dawley (SD) rats or uraemic SD rats induced by 5/6 nephrectomy fed a high-phosphate diet were treated with VS-505 or sevelamer (0.05-10% in food) for 5 and 28 days respectively. KEY RESULTS: Urinary and serum phosphate levels were significantly elevated in untreated rats, and were decreased by VS-505 and sevelamer. VS-505 increased faecal phosphate levels in a dose-dependent manner. High-phosphate diet also caused an increase in serum FGF-23 and parathyroid hormone in nephrectomized (NX) rats, effects prevented by VS-505 or sevelamer. Significant aortic calcification was observed in NX rats treated with 5% sevelamer, whereas VS-505 at all doses tested did not show effects. VS-505 had no effects on small intestine histomorphology and intestinal sodium-dependent phosphate cotransporter gene expression. In vitro characterizations showed that VS-505 has a relatively high density and low expansion volume when exposed to simulated gastric fluid. CONCLUSIONS AND IMPLICATIONS: VS-505 is a safe and effective phosphate binder and may offer the advantage of having a reduced pill burden and minimal GI side effects for CKD patients.

Cardiovascular disease in chronic kidney failure: is there a role for vitamin D analogs?

Vitamin D3 is modified by vitamin D3-25-hydroxylase in the liver, and 25-hydroxyvitamin D3-1alpha-hydroxylase in the kidney, to form the active metabolite, 1,25-dihydroxyvitamin D3. Chronic kidney disease (CKD) is characterized by reduced synthesis of 1,25-dibydroxyvitamin D3, inadequate renal phosphate clearance and calcium imbalance, secondary hyperparathyroidism (SHPT) and bone disease. CKD patients encounter a much higher risk of cardiovascular disease (CVD) than the general public. The cardiovascular risk factors for CKD patients include conventional factors such as age, gender, hypertension, diabetes, dyslipidemia and smoking, and non-conventional factors, such as anemia, uremia, reduced vascular compliance, inflammation and various hormonal factors. Several vitamin D analogs are currently available for the treatment of SHPT, and recent clinical data show that these analogs provide survival benefit for CKD patients in the order of paricalcitol > calcitriol > no vitamin D analog, independent of parathyroid hormone and calcium. Moreover, the survival benefit seems to be associated with cardiovascular causes. The observations made from these clinical studies raised intriguing questions about the involvement of the vitamin D receptor locus (VDR) in the cardiovascular system. This review discusses recent data regarding the role of vitamin D and its analogs in the CVD associated with CKD.

Different vitamin D receptor agonists exhibit differential effects on endothelial function and aortic gene expression in 5/6 nephrectomized rats.

Endothelial dysfunction, common in chronic kidney disease (CKD), significantly increases cardiovascular disease risk in CKD patients. This study investigates whether different vitamin D receptor agonists exhibit different effects on endothelial function and on aortic gene expression in an animal CKD model. The 5/6 nephrectomized (NX) rat was treated with or without alfacalcidol (0.02, 0.04 and 0.08µg/kg), paricalcitol (0.04 and 0.08µg/kg), or VS-105 (0.004, 0.01 and 0.16µg/kg). All three compounds at the test doses suppressed serum parathyroid hormone effectively. Alfacalcidol at 0.08µg/kg raised serum calcium significantly. Endothelial function was assessed by pre-contracting thoracic aortic rings with phenylephrine, followed by treatment with acetylcholine or sodium nitroprusside. Uremia significantly affected endothelial-dependent aortic relaxation, which was improved by all three compounds in a dose-dependent manner with alfacalcidol and paricalcitol exhibiting a lesser effect. DNA microarray analysis of aorta samples revealed that uremia impacted the expression of numerous aortic genes, many of which were normalized by the vitamin D analogs. Real-time RT-PCR analysis confirmed that selected genes such as Abra, Apoa4, Fabp2, Hsd17b2, and Hspa1b affected by uremia were normalized by the vitamin D analogs with alfacalcidol exhibiting less of an effect. These results demonstrate that different vitamin D analogs exhibit different effects on endothelial function and aortic gene expression in 5/6 NX rats. This article is part of a Special Issue entitled '17th Vitamin D Workshop'.

Endothelin receptor antagonists as therapeutic agents for cancer.

Endothelins (ETs) are 21-amino acid peptides involved in many different pathological conditions. ETs, mediated by the ET(A) receptor, are mitogenic for many types of cancer cells. ETs seem to both promote and inhibit apoptosis, dependent on the cell and tissue types. The anti-apoptotic effect of ETs is mediated predominantly by the ET(A) receptor. The ET system also plays a role in metastases. Indeed, animal studies and human clinical trials with ET(A) receptor antagonists suggest that they may be useful therapeutic agents for treating cancer. This paper reviews recent studies on the role of the ET system in tumorigenesis and provides an update on the recent development of ET receptor antagonists, in clinical studies for cancer.