New Roles for Vitamin D Superagonists: From COVID to Cancer.

Vitamin D is a potent steroid hormone that induces widespread changes in gene expression and controls key biological pathways. Here we review pathophysiology of vitamin D with particular reference to COVID-19 and pancreatic cancer. Utility as a therapeutic agent is limited by hypercalcemic effects and attempts to circumvent this problem have used vitamin D superagonists, with increased efficacy and reduced calcemic effect. A further caveat is that vitamin D mediates multiple diverse effects. Some of these (anti-fibrosis) are likely beneficial in patients with COVID-19 and pancreatic cancer, whereas others (reduced immunity), may be beneficial through attenuation of the cytokine storm in patients with advanced COVID-19, but detrimental in pancreatic cancer. Vitamin D superagonists represent an untapped resource for development of effective therapeutic agents. However, to be successful this approach will require agonists with high cell-tissue specificity.

Investigating the effect of testosterone by itself and in combination with letrozole on 1,25-dihydroxy vitamin D and FGF23 in male rats.

BACKGROUND: Testosterone deficiency might be associated with vitamin D levels in hypogonadal men, but it is not clear whether testosterone can affect vitamin D and fibroblast growth factor-23 (FGF23), either directly or indirectly via aromatization to estradiol. We aimed to investigate the role of testosterone on vitamin D metabolism and serum FGF23 in male rats. METHODS: A total of 48 male rats were divided into 4 equal groups: sham; O, orchiectomy; O + T, orchiectomized rats treated with testosterone; and O + T + L, orchiectomized rats treated with combination of testosterone and letrozole. We compare the vitamin D metabolism biochemical parameters in these four groups, before and after the study. RESULTS: We detected a significant reduction in 25-hydroxyvitamin D (25(OH)D), vitamin D binding protein (DBP), FGF23, and 1,25-dihydroxyvitamin D (1,25(OH)2D) serum level in O group compared to sham group (p = 0.004, p = 0.009, p < 0.001 and p < 0.001, respectively), and a significant increase in serum phosphorus, parathyroid hormone (PTH), and alkaline phosphatase (ALP) levels in orchiectomized rats in comparison to sham group (p < 0.001, p = 0.022, and p = 0.006, respectively). However, these changes were corrected by testosterone replacement in O + T and O + T + L groups. In addition, we found that DBP and 1,25(OH)2D serum levels were significantly higher in O + T group in comparison to O + T + L group (p = 0.030 and p = 0.026, respectively). CONCLUSIONS: Testosterone plays a significant role on regulating 25(OH)D, DBP, FGF23, phosphate (Phos), PTH, and 1,25(OH)2D serum levels in male rats. Also, testosterone has a potent effect on 1,25(OH)2D and DBP by its conversion to estradiol.

Role of vitamin D and vitamin D receptor (VDR) in oral cancer.

Oral cancer is known as one of the most common cancers, with a poor prognosis, related to delayed clinical diagnosis, either due to the lack of particular biomarkers related to the disease or costly therapeutic alternatives. Vitamin D executes its functions by interacting with the vitamin D receptor (VDR), both in healthy and diseased individuals, including oral cancer. This review discusses the role of vitamin D and VDR on tumorigenesis, emphasizing on oral cancer. Furthermore, regulation of VDR expression, mechanisms of anticancer effects of calcitriol, oral cancer chemoresistance and its relation with VDR and polymorphisms of VDR gene will be discussed. The manuscript is prepared mainly using the information collected from PubMed and MEDLINE.

Randomized trial of vitamin D supplementation for winter-related atopic dermatitis in children.

BACKGROUND: Epidemiologic and preclinical data, and a small randomized trial in Boston, suggest that vitamin D supplementation may improve winter-related atopic dermatitis (AD). OBJECTIVE: To determine the effect of vitamin D supplementation on winter-related AD. METHODS: We performed a randomized, double-blind, placebo-controlled trial of Mongolian children with winter-related AD (clinicaltrials.gov identifier: NCT00879424). Baseline eligibility included age 2 to 17 years, AD score 10 to 72 using the Eczema Area and Severity Index (EASI), and winter-related AD (eg, history of AD worsening during the fall-to-winter transition). Subjects were enrolled in Ulaanbaatar during winter and randomly assigned to oral cholecalciferol (1000 IU/day) versus placebo for 1 month. All children and parents received emollient and patient education about AD and basic skin care. The main outcomes were changes in EASI score and in Investigator's Global Assessment. RESULTS: The 107 enrolled children had a mean age of 9 years (SD 5), and 59% were male. Their median age of AD onset was 3 months (interquartile range 2 months to 1 year) and mean EASI score at baseline 21 (SD 9). One-month follow-up data were available for 104 (97%) children. Compared with placebo, vitamin D supplementation for 1 month produced a clinically and statistically significant improvement in EASI score (adjusted mean change: -6.5 vs -3.3, respectively; P = .04). Moreover, change in Investigator's Global Assessment favored vitamin D over placebo (P = .03). There were no adverse effects in either group. CONCLUSION: Vitamin D supplementation improved winter-related AD among Mongolian children, a population likely to have vitamin D deficiency in winter.

Vitamin D in chronic kidney disease.

Vitamin D deficiency is highly prevalent in patients with chronic kidney disease (CKD). The low vitamin D status is, to a large extent, caused by dysregulation of vitamin D metabolism as a result of renal insufficiency. Recent studies indicate that vitamin D-deficiency may promote or accelerate the progression of CKD, whereas treatment with low calcemic vitamin D analogs can reduce proteinuria and ameliorate renal damage in animal models of kidney disease and in patients with CKD. The renoprotective activity of vitamin D regulates multiple signaling pathways known to play important roles in renal injury. These findings underscore the importance of correcting vitamin D deficiency with vitamin D supplementation or with activated vitamin D analogs in the management of CKD.

Parathyroid function in chronic kidney disease: role of FGF23-Klotho axis.

The parathyroid gland plays a central role in the regulation of mineral metabolism. In patients with chronic kidney disease (CKD), circulating levels of parathyroid hormone (PTH) are progressively increased as kidney function declines, as a result of phosphate retention, hypocalcemia, decreased production of 1,25-dihydroxyvitamin D [1,25(OH)2D], endogenous changes within the parathyroid gland, and skeletal resistance to the actions of PTH. In addition, the identification of fibroblast growth factor 23 (FGF23) and its cofactor Klotho offers important implications for the deeper understanding of disordered mineral metabolism in CKD. In early CKD, increased FGF23 to maintain neutral phosphate balance results in suppression of renal 1,25(OH)2D production and thereby triggers the early development of secondary hyperparathyroidism. FGF23 also acts directly on the parathyroid to decrease PTH synthesis and secretion, but this effect is blunted in advanced stages of CKD, due to decreased expression of the Klotho-FGF receptor 1 complex and increased concentrations of C-terminal FGF23 that competes with full-length FGF23 for binding to the receptor complex. Recent clinical studies also reported that high levels of FGF23 are associated with morbidity and mortality as well as treatment resistance to active vitamin D, suggesting the potential of FGF23 as a novel biomarker to guide treatment of disordered phosphate metabolism in CKD. This review will discuss the pathogenesis of secondary hyperparathyroidism, particularly focusing on the emerging role of the FGF23-Klotho axis in patients with CKD.

Caring for chronic kidney disease patients: focus on mineral and bone disorders.

This case study focuses on a patient with chronic kidney disease (CKD) with mineral and bone disorders (MBD) and the relationship and management strategies used in treating CKD-MBD. The various risks and issues pertaining to the CKD stage 5 patient population are addressed, including CKD-MBD and renal osteodystrophy. Proper management of CKD-MBD with diet, dialysis, laboratory testing, and medications is discussed. An interdisciplinary team that includes the patient and family is crucial for effective management of MBD.

Vitamin D derivatives induce apoptosis and downregulate ICAM-1 levels in peripheral blood mononuclear cells of inflammatory bowel disease patients.

BACKGROUND: Lymphocytes are crucial in the pathogenesis of inflammatory bowel disease (IBD) and are an important target for drug development. Our aim was to verify whether 2 vitamin D derivatives, 1,25-dihydroxyvitamin D3 [1,25(OH)2D3] and EB 1089, could induce cell apoptosis and affect cell-cell interaction by regulating adhesion molecule levels. METHODS: Peripheral blood mononuclear cell (PBMC) proliferation was studied by [3H]thymidine incorporation and apoptosis was determined using an enzyme-linked immunosorbent assay (ELISA) kit. (Poly(ADP-ribose)polymerase (PARP) cleavage, caspase-3, and ICAM-1 protein levels were determined by Western blot analysis. RESULTS: Our results indicate that 1,25(OH)2D3 or EB 1089 or anti-TNF-alpha (infliximab) induce apoptosis in PBMC obtained from healthy subjects. In IBD patients apoptosis is induced by vitamin D derivatives and by anti-TNF-alpha only in CD patients. Caspase-3 activation and PARP cleavage are registered when PBMC were treated with vitamin D derivatives. ICAM-1 levels remarkably increase when PBMC was incubated with lipopolysaccharide (LPS) or TNF-alpha. The treatment with the vitamin D derivatives, alone or in combination with LPS or TNF-alpha, significantly decreases ICAM-1 levels both in healthy subjects and IBD patients. In HUVEC cocultured with PBMC, previously incubated with LPS or TNF-alpha associated with 1,25(OH)2D3, ICAM-1 levels decrease both in healthy subjects and IBD patients. CONCLUSIONS: 1,25(OH)2D3 and EB 1089 inhibit PBMC proliferation, induce apoptosis in PBMC of healthy subjects and IBD patients, and affect ICAM-1 expression on PBMC and on HUVEC cocultured with PBMC, suggesting that the ICAM-1 downregulation could provide a new target for controlling the recruitment of leukocytes at the sites of inflammation in IBD.

Alteraciones en el metabolismo mineral óseo en pacientes con urolitiasis de repetición y polimorfismos del gen del receptor de la vitamina D. Resultados preliminares / Alterations in bone mineral metabolism in patients with calcium kidney stone disease and polymorphism of vitamin D receptor, preliminary results

Existe controversia sobre la afectación ósea en la litiasis renal cálcica. Por otro lado, algunos estudios genéticos han encontrado asociación entre los polimorfismos del receptor de la vitamina D (VDR) y la urolitiasis.Objetivo principal: Relacionar la nefrolitiasis cálcica de repetición con el metabolismo óseo y los polimorfismos del gen del VDR. Material y métodos: Estudio de casos y controles, estando el grupo de casos formado por 51 pacientes con litiasis renal de repetición, que subdividimos enno hipercalciúricos (NHC, n = 27), hipercalciúricos absortivos (HCA, n = 10) e hipercalciúricosrenales (HCR, n = 14); el grupo control, formado por 21 sujetos sin historia de litiasis renal ni hipercalciuria. Se les determinaron parámetros del metabolismo fosfo-cálcico, marcadores de remodelado óseo, densidad mineral ósea (DMO) en columna lumbar y en cuello femoral, y polimorfismos del gen del VDRpara los loci b, a y t. Resultados: Los pacientes litiásicos presentaron frente a los controles una DMO inferior tanto en L2-L4 como en cuello femoral (Z-score, p = 0,045 y 0,031), niveles superiores de 1,25 (OH)2 vitamina D (p = 0,002) e inferiores de PTH (p = 0,049), y una menor ingesta cálcica (p < 0,001). Los HCA mostraron una mayor DMO frente a los NHC (sólo significativo en cuello femoral). Los pacientes con LRC no mostraron diferencias en las frecuencias genotípicas estudiadas frente a los controles. Al reagrupar los alelos, sólo se apreció una menor frecuencia del genotipo BB respecto al Bb-bb, y del tt frente al TT-Tt, en los pacientes litiásicos (p =0,098 y p = 0,051, respectivamente). Conclusiones: La litiasis renal cálcica pareceinfluir en la DMO de cuello femoral. Los pacientes litiásicos mostraron niveles elevados de 1,25 (OH)2 vitamina D, posiblemente relacionado con la baja dieta cálcica. Los genotipos homocigóticos BB y tt parecen ser menos frecuentes entre los pacientes con litiasis renal cálcica

Bone health, within calcium kidney stone disease is a matter of controversy. On the other hand, some genetic studies have shown an association between some Vitamin Dreceptor polymorphisms and calcium kidney stone disease. Main objective: To study the possible association between calcium kidney stone disease with bone metabolismand some Vitamin D receptor polymorphisms. Patients and methods: This is a casecontrol study, with seventy-two subjects of both genders divided into two groups: Group I: cases, composed by 51 patients suffering from calcium kidney stone disease. Twenty-four of them had no hypercalciuria, 16 had absortive hypercalciuria and 11 had renal hypercalciuria. Group II: controls, composed by 21 people, without either urolithiasis or hypercalciuria. We performed a complete study including biochemical markers of bone mineral remodelling, bone mineral density (BMD) was estimatedboth in the lumbar spine (L2-L4) and femoral neck, and also VDR polymorphism for the loci b, a and t. Results: Patients with urolithiasis had lower values of BMD both in the lumbar spine and femoral neck, compared to controls. Z-score were lower in the lumbar spine and femoral neck (p = 0.045 y 0.031, respectively). Those patients with absorptive hypercalciuria had higher BMD in the femoral neck than those with renal hypercalciuria and non-hypercalciuria. Because they had more weight and height all the statistical study was performed alter adjusting by these two variables and statisticalsignificance was then only stated between patients with hypercalciuria and without it. Patients with urolithiasis had higher values of 1.25 (OH)2 vitamin D (p = 0.002), and lower of PTH (p = 0.049), without any relationship to hypercalciuria and its subtypes.Seventy six percent of the patients had a daily calcium intake lower than 800 mg/day. The distribution of VDR alleles in patients with urolithiasis was similar to controls, although after grouping genotypes, a lower distribution of BB and tt polymorphisms wereobserved in patients suffering from urolithiasis. Conclusions: Calcium kidney stone diseaseby itself produces a decrease in BMD, more intense in femoral neck, independently the presence or absence of hypercalciuria. Patients suffering from urolitihiasishave higher values of 1.25 (OH)2 vitamin D than non-hypercalciuric patients and lower values of PTH probably due to a low dietary calcium intake. In our populationstudied there is no relationship between VDR polymorphisms and the presence of calcium kidney stone disease. Because the reduced number of patients of our study,more studies are needed to obtain definitely conclusions

Ligands with a 3,3-diphenylpentane skeleton for nuclear vitamin D and androgen receptors: Dual activities and metabolic activation.

Ligands possessing dual vitamin D(3) (VD(3))-agonistic and androgen-antagonistic activities with various activity spectra were prepared based on a substituted 3,3-diphenylpentane (DPP) skeleton. Among the compounds, (R,S)-DPP-1023 [(R,S)-7b] and (S,S)-DPP-0123 [(S,S)-7c] showed the most potent vitamin D(3)-agonistic activity [with potency comparable to that of 1alpha,25-dihydroxyvitamin D(3) (1,25-VD(3))] and nuclear androgen receptor (AR)-binding activity (with higher affinity than that of hydroxyflutamide), respectively. Metabolic activation (reduction of the carbonyl group) of pivaloyl analogs [DPP-1113 (3a), DPP-1013 (3b), DPP-0113 (3c), and DPP-0013 (3d)] in HL-60 cells was found to be necessary for binding to nuclear vitamin D(3) receptor (VDR).

Non-genomic effects of the androgen receptor and vitamin D agonist are involved in suppressing invasive phenotype of prostate cancer cells.

Suppression of invasive phenotype is essential in developing new therapeutic tools to treat prostate cancer (PC). Evidence indicates that androgen-dependent (AD) prostate cancer cells are characterized by a lower malignant phenotype. We have demonstrated that transfection with an androgen receptor (AR) expression vector of the androgen-independent (AI) prostate cancer cell line PC3 decreases invasion of these cells through modulation of alpha6beta4 integrin expression, indicating a genotropic effect of androgens in inhibiting invasion ability of AD PC cells. Later on, we have shown that also a non-genotropic mechanism is involved in such an effect. By using immunoconfocal fluorescent microscopy, we demonstrated that AR in PC3-AR cells co-localizes with the EGFR receptors (EGFR) in PC3-AR cells. Co-immunoprecipitation studies both in PC3-AR cells and in the AD cell line LNCaP that physiologically express both receptors, confirm the occurrence of an interaction between of the two proteins. In PC3-AR cells, we demonstrated a disruption of EGFR signalling properties (reduced EGF-induced EGFR autotransphosphorylation, reduced EGF-stimulated PI3K activity as well as EGFR-PI3K interaction) contributing to the lower invasive phenotype of these cells. In another study, we investigated the effects of a new Vitamin D analogue, BXL628, on invasion in response to KGF in the androgen-independent PC cell line DU145. We found that the compound was able to reduce proliferation and invasion of the cells in response to the growth factor. In addition, we found that KGF-induced autotransphosphorylation of KGF receptor (KGFR) and PI3K activation were suppressed after short-term (5min) pre-treatment with the analogue before addition of KGF. Collectively, these studies demonstrate that a non-genotropic effect due to a direct interaction of the androgen receptor with EGFR and to a rapid effect of a Vitamin D agonist on KGFR may disrupt signalling of GF leading to decreased tumorigenicity and a less malignant phenotype of PC cells in vitro.

NMR assignments of tryptophan residue in apo and holo LBD-rVDR.

Binding sites in the full-length, ligand-binding domain of rat vitamin D receptor (LBD-rVDR) for an active hormone derived from vitamin D (1alpha,25-dihydroxyvitamin D(3)) and three of its C-2 substituted analogs were compared by nuclear magnetic resonance (NMR) spectroscopy. Specific residue labeled with [UL]-(15)N(2) Trp allowed assignment of the side-chain H(epsilon1) and N(epsilon1) resonances of the single tryptophan residue at position 282 in LBD-rVDR. Comparison of (1)H[(15)N] Heteronuclear Single Quantum Correlation (HSQC) spectra of apo and holo LBD-rVDR revealed that the position of the Trp282 H(epsilon1) and N(epsilon1) signals are sensitive to the presence of the ligand in the receptor cavity. Binding of the ligands to LBD-rVDR results in a shift of both Trp H(epsilon1) and N(epsilon1) resonances to lower frequencies. The results indicate that the interaction between the ligands and Trp282 is not responsible for differences in calcemic activity observed in vitamin D analogs.

Altered Vitamin D receptor-coactivator interactions reflect superagonism of Vitamin D analogs.

The active form of Vitamin D, 1alpha,25-dihydroxyvitamin D(3) [1,25-(OH)(2)D(3)], has potent antiproliferative actions on various normal and malignant cells. Calcemic effects, however, hamper therapeutic application of 1,25-(OH)(2)D(3) in hyperproliferative diseases. Two 14-epi-analogs of 1,25-(OH)(2)D(3) namely 19-nor-14-epi-23-yne-1,25-(OH)(2)D(3) (TX522) and 19-nor-14,20-bisepi-23-yne-1,25-(OH)(2)D(3) (TX527), display reduced calcemic effects coupled to an (at least 10-fold) increased antiproliferative potency when compared with 1,25-(OH)(2)D(3). Altered cofactor recruitment by the Vitamin D receptor (VDR) might underlie the superagonism of these 14-epi-analogs. Therefore, this study aims to evaluate their effects at the level of VDR-coactivator interactions. Mammalian two-hybrid assays with VDR and the coactivators TIF2 and DRIP205 showed the 14-epi-analogs to be more potent inducers of VDR-coactivator interactions than 1,25-(OH)(2)D(3). TX522 and TX527 require 30- and 40-fold lower doses to obtain the VDR-DRIP205 interaction induced by 1,25-(OH)(2)D(3) at 10(-8)M. Evaluation of additional 1,25-(OH)(2)D(3)-analogs and their impact on VDR-coactivator interactions revealed a strong correlation between the antiproliferative potency of an analog and its ability to induce VDR-coactivator interactions. In conclusion, these data show that altered coactivator binding by the VDR is one possible explanation for the superagonistic action of the two 14-epi-analogs TX522 and TX527.

Management of renal osteodystrophy in children.

Prevention and treatment of renal osteodystrophy (ROD) are great challenges for pediatric nephrologists. The strategies for prevention and treatment of ROD in children with chronic renal failure (CRF) should be created on an individual basis. The following factors should be considered: age, type of primary disease, rate of progression of CRF, nutrition, acidosis, type of dialysis, and drugs (corticosteroids, growth hormone, etc). The treatment should start very early in the course of renal insufficiency with close monitoring of serum calcium, phosphate, alkaline phosphatase and parathormone (PTH) levels. Maintenance of serum phosphate within age- appropriate limits is essential for prevention of secondary hyperparathyroidism. PTH levels should be kept within normal limits in predialysis children and 2-3 times over upper normal limit in those on dialysis. Aggressive treatment with calcium-based phosphate binders and vitamin D derivates should be avoided to prevent PTH oversuppression and development of adynamic bone disease. The advantage in this respect is the development of calcium- and aluminum-free phosphate binders, of which there is limited pediatric experience with sevelamer hydrochloride. Paricalcitol is a non-hypercalcemic vitamin D analogue, and preliminary favorable experience has been reported in children. Calcimimetics like cinacalcet hydrochloride, which directly stimulate calcium sensing receptor and potently suppress PTH secretion without increasing plasma calcium in adults, are very promising agents, but pediatric experience is lacking.

Molecular mechanism of the vitamin D antagonistic actions of (23S)-25-dehydro-1alpha-hydroxyvitamin D3-26,23-lactone depends on the primary structure of the carboxyl-terminal region of the vitamin d receptor.

We reported that (23S)-25-dehydro-1alpha-hydroxyvitamin D(3)-26,23-lactone (TEI-9647) antagonizes vitamin D receptor (VDR)-mediated genomic actions of 1alpha,25-dihydroxyvitamin D(3) [1alpha,25(OH)(2)D(3)] in human cells but is agonistic in rodent cells. Human and rat VDR ligand-binding domains are similar, but differences in the C-terminal region are important for ligand binding and transactivation and might determine the agonistic/antagonistic effects of TEI-9647. We tested TEI-9647 on 1alpha,25(OH)(2)D(3) transactivation using SaOS-2 cells (human osteosarcoma) or ROS 24/1 cells (rat osteosarcoma) cotransfected with human or rodent VDR and a reporter. In both cell lines, TEI-9647 was antagonistic with wild-type human (h)VDR, but agonistic with overexpressed wild-type rat (r)VDR. VDR chimeras substituting the hVDR C-terminal region (activation function 2 domain) with corresponding rVDR residues diminished antagonism and increased agonism of TEI-9647. However, substitution of 25 C-terminal rVDR residues with corresponding hVDR residues diminished agonism and increased antagonism of TEI-9647. hVDR mutants (C403S, C410N) demonstrated that Cys403 and/or 410 was necessary for TEI-9647 antagonism of 1alpha,25(OH)(2)D(3) transactivation. These results suggest that species specificity of VDR, especially in the C-terminal region, determines the agonistic/antagonistic effects of TEI-9647 that determine, in part, VDR interactions with coactivators and emphasize the critical interaction between TEI-9647 and the two C-terminal hVDR Cys residues to mediate the antagonistic effect of TEI-9647.