RESUMEN

BACKGROUND: Ovarian cancer (OC) is one of the most lethal cancers in women. The active form of vitamin D3, 1,25-dihydroxyvitamin D3 (1,25D3, calcitriol) has anticancer activity in several cancers, including ovarian cancer, but the required pharmacological doses may cause hypercalcemia. We hypothesized that newly developed, low calcemic, vitamin D analogs (an1,25Ds) may be used as anticancer agents instead of calcitriol in ovarian cancer cells. METHODS: We used two patient-derived high-grade serous ovarian cancer (HGSOC) cell lines with low (13781) and high (14433) mRNA expression levels of the gene encoding 1,25-dihydroxyvitamin D3 24-hydroxylase CYP24A1, one of the main target genes of calcitriol. We tested the effect of calcitriol and four structurally related series of an1,25Ds (PRI-1906, PRI-1907, PRI-5201, PRI-5202) on cell number, viability, the expression of CYP24A1, and the vitamin D receptor (VDR). RESULTS: CYP24A1 mRNA expression increased in a concentration-dependent manner after treatment with all compounds. In both cell lines, after 4 h, PRI-5202 was the most potent analog (in 13781 cells: EC50 = 2.98 ± 1.10 nmol/L, in 14433 cells: EC50 = 0.92 ± 0.20 nmol/L), while PRI-1907 was the least active one (in 13781 cells: EC50 = n/d, in 14433 cells: EC50 = n/d). This difference among the analogs disappeared after 5 days of treatment. The 13781 cells were more sensitive to the an1,25Ds compared with 14433 cells. The an1,25Ds increased nuclear VDR levels and reduced cell viability, but only in the 13781 cell line. CONCLUSIONS: The an1,25Ds had different potencies in the HGSOC cell lines and their efficacy in increasing CYP24A1 expression was cell line- and chemical structure-dependent. Therefore, choosing sensitive cancer cell lines and further optimization of the analogs' structure might lead to new treatment options against ovarian cancer.

Asunto(s)

Supervivencia Celular , Neoplasias Ováricas/tratamiento farmacológico , Receptores de Calcitriol/genética , Vitamina D3 24-Hidroxilasa/genética , Vitamina D/farmacología , Antineoplásicos/metabolismo , Antineoplásicos/farmacología , Línea Celular Tumoral , Células Cultivadas , Ergocalciferoles/metabolismo , Ergocalciferoles/farmacología , Femenino , Regulación Neoplásica de la Expresión Génica , Humanos , Neoplasias Ováricas/enzimología , Neoplasias Ováricas/metabolismo , Vitamina D/análogos & derivados

RESUMEN

BACKGROUND: Novel ways of determining cardiovascular risk are needed as a consequence of population ageing and the increased prevalence of chronic kidney disease (CKD), both of which favour vascular calcification. Since the formation of arterial calcium deposits has a genetic component, single nucleotide polymorphisms (SNPs) could predict cardiovascular events. METHODS: A selection of 1927 CKD patients and controls recruited by the NEFRONA study were genotyped for 60 SNPs from 22 candidate genes. A calcium score was calculated from the echogenicity of arterial atherosclerotic plaques and the presence of cardiovascular events during a 4-year period was recorded. Association of SNPs with the calcium score was identified by multiple linear regression models and their capacity to predict events was assessed by means of Cox proportional hazards regression and receiver operating characteristics curves. RESULTS: Two variants, rs2296241 of CYP24A1 and rs495392 of KL, were associated with the calcium score. Despite this, only heterozygotes for rs495392 had a lower risk of suffering an event compared with homozygotes for the major allele {hazard ratio (HR) 0.67 [95% confidence interval (CI) 0.48-0.93]}. Of note, the calcium score was associated with an increased risk of cardiovascular events [HR 1.71 (95% CI 1.35-2.17)]. The addition of the rs495392 genotype to classical cardiovascular risk factors did not increase the predictive power [area under the curve (AUC) 71.3 (95% CI 61.1-85.5) versus 71.4 (61.5-81.4)]. CONCLUSIONS: Polymorphisms of CYP24A1 and KL are associated with the extent of calcification but do not predict cardiovascular events. However, the echogenic determination of the extent of calcium deposits seems a promising non-irradiating method for the scoring of calcification in high-risk populations.

Asunto(s)

Enfermedades Cardiovasculares , Proteínas Klotho/genética , Calcificación Vascular , Vitamina D3 24-Hidroxilasa/genética , Enfermedades Cardiovasculares/etiología , Enfermedades Cardiovasculares/genética , Humanos , Polimorfismo de Nucleótido Simple , Factores de Riesgo , Calcificación Vascular/diagnóstico por imagen , Calcificación Vascular/genética

RESUMEN

BACKGROUND/AIMS: The CYP24A1 gene encodes the vitamin D 24-hydroxylase enzyme, which hydroxylates active forms of vitamin D into inactive forms. Biallelic mutations in the CYP24A1 gene can lead to elevated levels of active vitamin D metabolites and, consequently, to hypercalcemia, hypercalciuria, nephrocalcinosis, and nephrolithiasis; however, monoallelic mutations have been associated only with milder phenotypes. In the present manuscript, we report the case of a young male patient who presented hypercalcemia and nephrolithiasis, suppressed parathormone, and elevated 1,25 dihydroxy vitamin D levels. METHODS: Biochemical analyses were performed on Cobas 8000, F. Hoffmann-La Roche AG, Basel, Switzerland. The proband was initially evaluated for occult malignancies by body imaging, serum electrophoresis, and tumor markers, which did not reveal any pathology. DNA samples of the proband and his sibling were then examined using Sanger sequencing. RESULTS: Genetic analysis revealed 2 compound heterozygous CYP24A1 mutations (p.L148P and p.R223*). The novel nonsense CYP24A1 mutation, p.R223*, was also found heterozygously in other family members with a medical history of nephrolithiasis. CONCLUSIONS: The identification of this gene mutation causing hypercalcemia, hypercalciuria, and renal stones allows the specific management of endogenous vitamin D production.

Asunto(s)

Cálculos Renales/genética , Mutación , Vitamina D3 24-Hidroxilasa/genética , Humanos , Hipercalcemia , Hipercalciuria , Masculino , Análisis de Secuencia de ADN , Hermanos , Vitamina D/sangre , Adulto Joven

RESUMEN

A homology model of human CYP27B1 was built using MOE and was further optimised by molecular dynamics simulations of the hCYP27B1 homology model and a hCYP27B1-SDZ-88357 complex. Docking results from the hCYP27B1-SDZ-88357 complex showed amino acids Arg107, Asn387 and Asp320 have an important role in binding interaction, with Asp320 part of the important acid-alcohol pair situated in the I-helix with the conserved sequence (A/G) GX (E/D) (T/S), which assumes an essential role in the binding of an oxygen molecule for catalysis. Additional docking experiments with selective hCYP27B1 or hCYP24A1 inhibitors using both the hCYP27B1 model and a triple mutant hCYP24A1 model provided further support for the importance of H-bonding interactions with the three identified active site amino acids. To confirm the role of Arg107, Asn387 and Asp320 in the active site of hCYP27B1 compounds were designed that would form H-bonding interactions, as determined from docking experiments with the hCYP27B1 model. Subsequent synthesis and CYP24A1 and CYP27B1 enzyme assays of the designed compounds 1a and 1b showed a∼5-fold selectivity for CYP27B1 confirming the importance of Asp320 in particular and also Asn387 and Arg107 as important amino acids for CYP27B1 inhibitory activity.

Asunto(s)

25-Hidroxivitamina D3 1-alfa-Hidroxilasa/química , 25-Hidroxivitamina D3 1-alfa-Hidroxilasa/metabolismo , Modelos Químicos , Simulación de Dinámica Molecular , Vitamina D3 24-Hidroxilasa/química , Vitamina D3 24-Hidroxilasa/metabolismo , Secuencia de Aminoácidos , Sitios de Unión , Dominio Catalítico , Inhibidores Enzimáticos/química , Inhibidores Enzimáticos/farmacología , Humanos , Ligandos , Estructura Molecular , Unión Proteica/efectos de los fármacos , Homología de Secuencia

RESUMEN

Cross-sectional studies depict an inverse relationship between vitamin D (VD) status reflected by plasma 25-hydroxy-vitamin D and obesity. Furthermore, recent studies in vitro and in animal models tend to demonstrate an impact of VD and VD receptor on adipose tissue and adipocyte biology, pointing to at least a part-causal role of VD insufficiency in obesity and associated physiopathological disorders such as adipose tissue inflammation and subsequent insulin resistance. However, clinical and genetic studies are far less convincing, with highly contrasted results ruling out solid conclusions for the moment. Nevertheless, prospective studies provide interesting data supporting the hypothesis of a preventive role of VD in onset of obesity. The aim of this review is to summarise the available data on relationships between VD, adipose tissue/adipocyte physiology, and obesity in order to reveal the next key points that need to be addressed before we can gain deeper insight into the controversial VD-obesity relationship.

RESUMEN

Eldecalcitol shows higher binding affinity for vitamin D-binding protein (DBP), tighter binding to vitamin D receptor (VDR), and resistance to metabolic degradation via 24-hydroxylation. In silico analysis of the mode of binding demonstrated that the 3-hydroxypropyloxy (3-HP) group of eldecalcitol offers additional hydrogen bond and CH-π interaction for the binding to DBP and VDR. However, the 3-HP group interferes with the binding of eldecalcitol to CYP24A1, causing poor metabolic clearance of eldecalcitol by this enzyme. These characteristics may contribute to the stronger effect of eldecalcitol than calcitriol. The present post-hoc analysis also demonstrate that the incidence of hypercalcemia and hypercalciuria is slightly higher in eldecalcitol than in alfacalcidol group especially in patients with CKD stage 3B, that both serum and urinary calcium return to the baseline levels shortly after cessation of the treatment in both treatment groups, that the incidence of urolithiasis is higher in patients with higher eGFR and is similar between alfacalcidol and eldecalcitol groups, and that eGFR is transiently reduced by both alfacalcidol and eldecalcitol treatment especially among patients with higher eGFR but recovers after the end of both treatment. Eldecalcitol can be used for the treatment of osteoporosis without Ca supplementation to reduce the incidence of hypercalcemia and hypercalciuria, and enough hydration is recommended in order to avoid hypercalcemia, urolithiasis and deterioration of renal function.

Asunto(s)

Osteoporosis/tratamiento farmacológico , Fracturas Osteoporóticas/prevención & control , Receptores de Calcitriol/metabolismo , Proteína de Unión a Vitamina D/metabolismo , Vitamina D3 24-Hidroxilasa/metabolismo , Vitamina D/análogos & derivados , Anciano , Densidad Ósea/efectos de los fármacos , Calcitriol/farmacología , Calcio/metabolismo , Ensayos Clínicos Fase III como Asunto , Cristalografía por Rayos X , Femenino , Tasa de Filtración Glomerular , Humanos , Masculino , Persona de Mediana Edad , Osteoporosis/metabolismo , Fracturas Osteoporóticas/metabolismo , Conformación Proteica/efectos de los fármacos , ARN Mensajero/genética , Reacción en Cadena en Tiempo Real de la Polimerasa , Receptores de Calcitriol/química , Receptores de Calcitriol/genética , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Vitamina D/química , Vitamina D/farmacología , Proteína de Unión a Vitamina D/química , Proteína de Unión a Vitamina D/genética , Vitamina D3 24-Hidroxilasa/química , Vitamina D3 24-Hidroxilasa/genética , Vitaminas/farmacología

RESUMEN

BACKGROUND: Loss-of-function mutations of vitamin D-24 hydroxylase have recently been recognized as a cause of hypercalcaemia and nephrocalcinosis/nephrolithiasis in infants and adults. True prevalence and natural history of this condition are still to be defined. METHODS: We describe two adult patients with homozygous mutations and six related heterozygous carriers. Mineral and hormonal data in these patients were compared with that in 27 patients with stage 2-3 chronic kidney disease and 39 healthy adult kidney donors. RESULTS: Probands had recurrent nephrolithiasis, chronic hypercalcaemia with depressed parathyroid hormone (PTH) and increased 1,25(OH)(2)D levels; carriers had nephrolithiasis (two of six), hypercalciuria (two of six) and high or normal-high 1,25(OH)(2)D (four of four). Corticosteroids did not reduce plasma and urine calcium levels, but ketoconazole did, indicating that 1,25(OH)(2)D production is not maximally depressed despite coexisting hypercalcaemia, high 1,25(OH)(2)D and depressed PTH, and that 1,25(OH)(2)D degradation through vitamin D-24 hydroxylase is a regulator of plasma 1,25(OH)(2)D levels. Both probands had vascular calcifications and high bone mineral content. One developed stage 3b renal failure: in this patient 1,25(OH)(2)D decreased within normal limits as glomerular filtration rate (GFR) fell and PTH rose to high-normal values, yet hypercalcaemia persisted and the ratio of 1,25(OH)(2)D to GFR remained higher than normal for any degree of GFR. CONCLUSIONS: This natural model indicates that vitamin D-24 hydroxylase is a key physiologic regulator of calcitriol and plasma calcium levels, and that balanced reduction of 1,25(OH)(2)D and GFR is instrumental for the maintenance of physiologic calcium levels and balance in chronic kidney diseases.

Asunto(s)

Hipercalcemia/genética , Fallo Renal Crónico/genética , Vitamina D3 24-Hidroxilasa/genética , Estudios de Casos y Controles , Estudios de Asociación Genética , Predisposición Genética a la Enfermedad , Humanos , Hipercalcemia/complicaciones , Hipercalcemia/enzimología , Fallo Renal Crónico/enzimología , Masculino , Persona de Mediana Edad , Mutación , Hormona Paratiroidea/sangre , Linaje

RESUMEN

PURPOSE: Hypercalciuria is the most common cause of kidney stone disease and genetic factors have an important role in nearly half of these cases. Recently loss-of-function mutations of CYP24A1, the gene encoding vitamin D 24-hydroxylase, were identified in idiopathic infantile hypercalcemia. We describe the clinical and molecular basis of severe long-standing kidney stone disease in adults caused by CYP24A1 mutations. MATERIALS AND METHODS: Three subjects from 2 Israeli families with nephrolithiasis and nephrocalcinosis were clinically characterized. Genomic DNA was isolated from peripheral blood and sequencing of CYP24A1 was performed. RESULTS: All subjects presented with severe kidney stone disease, the cause of which was not discovered for decades despite extensive evaluation. They all had hypercalciuria, nephrocalcinosis and intermittent hypercalcemia, and chronic kidney insufficiency developed in the oldest subject. All patients had a typical pattern of test results, including normal-high serum calcium, low parathyroid hormone levels, high vitamin D 25-(OH)D3 and 1,25-(OH)2D3, and low 24,25-(OH)2D3. Overall 3 CYP24A1 loss-of-function mutations were identified, including a homozygous deletion (delE143) in consanguinous family 1, and compound heterozygous mutations L409S and the novel W268-stop in family 2. CONCLUSIONS: Loss-of-function mutations of CYP24A1 gene, encoding for 1,25-dihydroxyvitamin D3 24-hydroxylase, cause severe hypercalciuric nephrolithiasis and nephrocalcinosis. The mutations may present in adults and may lead to chronic renal insufficiency. Our results support a recessive mode of inheritance. CYP24A1 mutations should be considered in the differential diagnosis of hypercalciuric nephrolithiasis, especially as many adults are now prescribed supplemental oral vitamin D.

Asunto(s)

Hipercalciuria/genética , Mutación , Nefrocalcinosis/genética , Nefrolitiasis/genética , Esteroide Hidroxilasas/genética , Adulto , Consanguinidad , Humanos , Israel , Masculino , Persona de Mediana Edad , Linaje , Fenotipo , Reacción en Cadena de la Polimerasa , Vitamina D3 24-Hidroxilasa