Pharmacogenetics of sodium-glucose co-transporter-2 inhibitors: Validation of a sex-agnostic pharmacodynamic biomarker.

AIM: To validate pharmacodynamic responses to sodium-glucose co-transporter-2 (SGLT2) inhibitors and test for association with genetic variants in SLC5A4, SLC5A9, and SLC2A9. METHODS: Canagliflozin (300 mg), a SGLT2 inhibitor, was administered to 30 healthy volunteers. Several endpoints were measured to assess clinically relevant responses, including drug-induced increases in urinary excretion of glucose, sodium and uric acid. RESULTS: This pilot study confirmed that canagliflozin (300 mg) triggered acute changes in mean levels of several biomarkers: fasting plasma glucose (-4.1 mg/dL; P = 6 × 10-5 ), serum creatinine (+0.05 mg/dL; P = 8 × 10-4 ) and serum uric acid (-0.90 mg/dL; P = 5 × 10-10 ). The effects of sex on glucosuria depended upon how data were normalized. Whereas males' responses were ~60% greater when data were normalized to body surface area, males and females exhibited similar responses when glucosuria was expressed as grams of urinary glucose per gram-creatinine. The magnitude of glucosuria was not significantly correlated with fasting plasma glucose, estimated glomerular filtration rate or age in those healthy individuals without diabetes with an estimated glomerular filtration rate of more than 60 mL/min/1.73m2 . CONCLUSIONS: Normalizing data relative to creatinine excretion will facilitate including data from males and females in a single analysis. Furthermore, because our ongoing pharmacogenomic study (NCT02891954) is conducted in healthy individuals, this will facilitate detection of genetic associations with limited confounding by other factors such as HbA1c and renal function.

Acute pharmacodynamic responses to exenatide: Drug-induced increases in insulin secretion and glucose effectiveness.

AIM: Glucagon-like peptide-1 receptor agonists provide multiple benefits to patients with type 2 diabetes, including improved glycaemic control, weight loss and decreased risk of major adverse cardiovascular events. Because drug responses vary among individuals, we initiated investigations to identify genetic variants associated with the magnitude of drug responses. METHODS: Exenatide (5 µg, subcutaneously) or saline (0.2 ml, subcutaneously) was administered to 62 healthy volunteers. Frequently sampled intravenous glucose tolerance tests were conducted to assess the impact of exenatide on insulin secretion and insulin action. This pilot study was a crossover design in which participants received exenatide and saline in random order. RESULTS: Exenatide increased first phase insulin secretion 1.9-fold (p = 1.9 × 10-9 ) and accelerated the rate of glucose disappearance 2.4-fold (p = 2 × 10-10 ). Minimal model analysis showed that exenatide increased glucose effectiveness (Sg ) by 32% (p = .0008) but did not significantly affect insulin sensitivity (Si ). The exenatide-induced increase in insulin secretion made the largest contribution to interindividual variation in exenatide-induced acceleration of glucose disappearance while interindividual variation in the drug effect on Sg contributed to a lesser extent (ß = 0.58 or 0.27, respectively). CONCLUSIONS: This pilot study provides validation for the value of a frequently sampled intravenous glucose tolerance test (including minimal model analysis) to provide primary data for our ongoing pharmacogenomic study of pharmacodynamic effects of semaglutide (NCT05071898). Three endpoints provide quantitative assessments of the effects of glucagon-like peptide-1 receptor agonists on glucose metabolism: first phase insulin secretion, glucose disappearance rates and glucose effectiveness.

Revisiting the Case of Sarah Newbury's Death from Mollities Ossium.

Multiple myeloma and its precursor and variant types represent some of the most common hematologic malignancies in adults. These plasma cell dyscrasias are well-known in modern medicine. There are well-established clinical, laboratory, and pathologic criteria for diagnosis and staging. There is debate about the diagnosis of some of the earliest cases of myeloma described in the literature. We present a critical review of one such case.

Impact of parental relatedness on reproductive outcomes among the Old Order Amish of Lancaster County.

Genetically isolated populations that arise due to recent bottleneck events have reduced genetic variation reflecting the common set of founders. Increased genetic relatedness among members of isolated populations puts them at increased risk for some recessive disorders that are rare in outbred populations. To assess the burden on reproductive health, we compared frequencies of adverse reproductive outcomes between Amish couples who were both heterozygous carriers of a highly penetrant pathogenic or likely pathogenic variant and noncarrier couples from the same Amish community. In addition, we evaluated whether overall genetic relatedness of parents was associated with reproductive outcomes. Of the 1824 couples included in our study, 11.1% were at risk of producing a child with an autosomal recessive disorder. Carrier couples reported a lower number of miscarriages compared to noncarrier couples (p = 0.02), although the number of stillbirths (p = 0.3), live births (p = 0.9), and number of pregnancies (p = 0.9) did not differ significantly between groups. In contrast, higher overall relatedness between spouses was positively correlated with number of live births (p < 0.0001), pregnancies (p < 0.0001), and stillbirths (p = 0.03), although not with the number of miscarriages (p = 0.4). These results highlight a complex association between relatedness of parents and reproductive health outcomes in this community.

Heterozygosity for a Pathogenic Variant in SLC12A3 That Causes Autosomal Recessive Gitelman Syndrome Is Associated with Lower Serum Potassium.

BACKGROUND: Potassium levels regulate multiple physiologic processes. The heritability of serum potassium level is moderate, with published estimates varying from 17% to 60%, suggesting genetic influences. However, the genetic determinants of potassium levels are not generally known. METHODS: A whole-exome sequencing association study of serum potassium levels in 5812 subjects of the Old Order Amish was performed. A dietary salt intervention in 533 Amish subjects estimated interaction between p.R642G and sodium intake. RESULTS: A cluster of variants, spanning approximately 537 kb on chromosome 16q13, was significantly associated with serum potassium levels. Among the associated variants, a known pathogenic variant of autosomal recessive Gitelman syndrome (p.R642G SLC12A3) was most likely causal; there were no homozygotes in our sample. Heterozygosity for p.R642G was also associated with lower chloride levels, but not with sodium levels. Notably, p.R642G showed a novel association with lower serum BUN levels. Heterozygotes for p.R642G had a two-fold higher rate of self-reported bone fractures and had higher resting heart rates on a low-salt diet compared with noncarriers. CONCLUSIONS: This study provides evidence that heterozygosity for a pathogenic variant in SLC12A3 causing Gitelman syndrome, a canonically recessive disorder, contributes to serum potassium concentration. The findings provide insights into SLC12A3 biology and the effects of heterozygosity on electrolyte homeostasis and related subclinical phenotypes that may have implications for personalized medicine and nutrition.

The burden of pathogenic variants in clinically actionable genes in a founder population.

Founder populations may be enriched with certain genetic variants of high clinical impact compared to nonfounder populations due to bottleneck events and genetic drift. Using exome sequencing (ES), we quantified the load of pathogenic variants that may be clinically actionable in 6136 apparently healthy adults living in the Lancaster, PA Old Order Amish settlement. We focused on variants in 78 genes deemed clinically actionable by the American College of Medical Genetics and Genomics (ACMG) or Geisinger's MyCode Health Initiative. ES revealed 3191 total variants among these genes including 480 nonsynonymous variants. After quality control and filtering, we applied the ACMG/AMP guidelines for variant interpretation and classified seven variants, across seven genes, as either pathogenic or likely pathogenic. Through genetic drift, all seven variants, are highly enriched in the Amish compared to nonfounder populations. In total, 14.7% of Lancaster Amish individuals carry at least one of these variants, largely explained by the 13% who harbor a copy of a single variant in APOB. Other studies report combined frequencies of pathogenic/likely pathogenic (P/LP) variants in actionable genes between 2.0% and 6.2% in outbred populations. The Amish population harbors fewer actionable variants compared to similarly characterized nonfounder populations but have a higher frequency of each variant identified, offering opportunities for efficient and cost-effective targeted precision medicine.

Surveillance results and bone effects in the Gulf War depleted uranium-exposed cohort.

A small group of Gulf War I veterans wounded in depleted uranium (DU) friendly-fire incidents have been monitored in a clinical surveillance program since 1993. During the spring of 2017, 42 members of the cohort were evaluated with a protocol which includes exposure monitoring for total and isotopic uranium concentrations in urine and a comprehensive assessment of health outcomes including measures of bone metabolism, and for participants >50 years, bone mineral density (BMD) determination. Elevated urine U concentrations were observed in cohort members with retained DU shrapnel fragments. Only the mean serum estradiol concentration, a marker of bone metabolism, was found to be significantly different for lower-vs- higher urine U (uU) cohort sub-groups. For the first time, a significant deficit in BMD was observed in the over age 50, high uU sub-group. After more than 25 years since first exposure to DU, an aging cohort of military veterans continues to exhibit few U-related adverse health effects in known target organs of U toxicity. The new finding of reduced BMD in older cohort members, while biologically plausible, was not suggested by other measures of bone metabolism in the full (all ages) cohort, as these were predominantly within normal limits over time. Only estradiol was recently found to display a difference as a function of uU grouping. As BMD is further impacted by aging and the U-burden from fragment absorption accrues in this cohort, a U effect may be clarified in future surveillance visits.

Genetic Variants Associated with Circulating Parathyroid Hormone.

Parathyroid hormone (PTH) is a primary calcium regulatory hormone. Elevated serum PTH concentrations in primary and secondary hyperparathyroidism have been associated with bone disease, hypertension, and in some studies, cardiovascular mortality. Genetic causes of variation in circulating PTH concentrations are incompletely understood. We performed a genome-wide association study of serum PTH concentrations among 29,155 participants of European ancestry from 13 cohort studies (n=22,653 and n=6502 in discovery and replication analyses, respectively). We evaluated the association of single nucleotide polymorphisms (SNPs) with natural log-transformed PTH concentration adjusted for age, sex, season, study site, and principal components of ancestry. We discovered associations of SNPs from five independent regions with serum PTH concentration, including the strongest association with rs6127099 upstream of CYP24A1 (P=4.2 × 10-53), a gene that encodes the primary catabolic enzyme for 1,25-dihydroxyvitamin D and 25-dihydroxyvitamin D. Each additional copy of the minor allele at this SNP associated with 7% higher serum PTH concentration. The other SNPs associated with serum PTH concentration included rs4074995 within RGS14 (P=6.6 × 10-17), rs219779 adjacent to CLDN14 (P=3.5 × 10-16), rs4443100 near RTDR1 (P=8.7 × 10-9), and rs73186030 near CASR (P=4.8 × 10-8). Of these five SNPs, rs6127099, rs4074995, and rs219779 replicated. Thus, common genetic variants located near genes involved in vitamin D metabolism and calcium and renal phosphate transport associated with differences in circulating PTH concentrations. Future studies could identify the causal variants at these loci, and the clinical and functional relevance of these variants should be pursued.

WRN Mutation Update: Mutation Spectrum, Patient Registries, and Translational Prospects.

Werner syndrome (WS) is a rare autosomal recessive disorder characterized by a constellation of adult onset phenotypes consistent with an acceleration of intrinsic biological aging. It is caused by pathogenic variants in the WRN gene, which encodes a multifunctional nuclear protein with exonuclease and helicase activities. WRN protein is thought to be involved in optimization of various aspects of DNA metabolism, including DNA repair, recombination, replication, and transcription. In this update, we summarize a total of 83 different WRN mutations, including eight previously unpublished mutations identified by the International Registry of Werner Syndrome (Seattle, WA) and the Japanese Werner Consortium (Chiba, Japan), as well as 75 mutations already reported in the literature. The Seattle International Registry recruits patients from all over the world to investigate genetic causes of a wide variety of progeroid syndromes in order to contribute to the knowledge of basic mechanisms of human aging. Given the unusually high prevalence of WS patients and heterozygous carriers in Japan, the major goal of the Japanese Consortium is to develop effective therapies and to establish management guidelines for WS patients in Japan and elsewhere. This review will also discuss potential translational approaches to this disorder, including those currently under investigation.

Multistage genome-wide association meta-analyses identified two new loci for bone mineral density.

Aiming to identify novel genetic variants and to confirm previously identified genetic variants associated with bone mineral density (BMD), we conducted a three-stage genome-wide association (GWA) meta-analysis in 27 061 study subjects. Stage 1 meta-analyzed seven GWA samples and 11 140 subjects for BMDs at the lumbar spine, hip and femoral neck, followed by a Stage 2 in silico replication of 33 SNPs in 9258 subjects, and by a Stage 3 de novo validation of three SNPs in 6663 subjects. Combining evidence from all the stages, we have identified two novel loci that have not been reported previously at the genome-wide significance (GWS; 5.0 × 10(-8)) level: 14q24.2 (rs227425, P-value 3.98 × 10(-13), SMOC1) in the combined sample of males and females and 21q22.13 (rs170183, P-value 4.15 × 10(-9), CLDN14) in the female-specific sample. The two newly identified SNPs were also significant in the GEnetic Factors for OSteoporosis consortium (GEFOS, n = 32 960) summary results. We have also independently confirmed 13 previously reported loci at the GWS level: 1p36.12 (ZBTB40), 1p31.3 (GPR177), 4p16.3 (FGFRL1), 4q22.1 (MEPE), 5q14.3 (MEF2C), 6q25.1 (C6orf97, ESR1), 7q21.3 (FLJ42280, SHFM1), 7q31.31 (FAM3C, WNT16), 8q24.12 (TNFRSF11B), 11p15.3 (SOX6), 11q13.4 (LRP5), 13q14.11 (AKAP11) and 16q24 (FOXL1). Gene expression analysis in osteogenic cells implied potential functional association of the two candidate genes (SMOC1 and CLDN14) in bone metabolism. Our findings independently confirm previously identified biological pathways underlying bone metabolism and contribute to the discovery of novel pathways, thus providing valuable insights into the intervention and treatment of osteoporosis.

Biologic monitoring and surveillance results for the department of veterans affairs' depleted uranium cohort: Lessons learned from sustained exposure over two decades.

BACKGROUND: A small group of Gulf War I veterans wounded in depleted uranium (DU) friendly fire incidents have been monitored in a clinical surveillance program at the Veterans Affairs Medical Center, Baltimore since 1994. METHODS: An in-patient clinical surveillance protocol was performed on 35 members of the cohort, including exposure monitoring for total and isotopic uranium concentrations in urine and a comprehensive assessment of health outcomes. RESULTS: Although urine U concentrations continue to be elevated in this group, illustrating on-going in situ mobilization of U from embedded fragments, no consistent U-related health effects have been observed. CONCLUSIONS: Now more than 20 years since first exposure to DU, an aging cohort of military veterans continues to show no U-related health effects in known target organs of U toxicity. As tissue concentrations continue to accrue with exposure duration, critical tissue-specific U concentration thresholds may be reached, thus recommending on-going surveillance of this veteran cohort.

WNT16 influences bone mineral density, cortical bone thickness, bone strength, and osteoporotic fracture risk.

We aimed to identify genetic variants associated with cortical bone thickness (CBT) and bone mineral density (BMD) by performing two separate genome-wide association study (GWAS) meta-analyses for CBT in 3 cohorts comprising 5,878 European subjects and for BMD in 5 cohorts comprising 5,672 individuals. We then assessed selected single-nucleotide polymorphisms (SNPs) for osteoporotic fracture in 2,023 cases and 3,740 controls. Association with CBT and forearm BMD was tested for ∼2.5 million SNPs in each cohort separately, and results were meta-analyzed using fixed effect meta-analysis. We identified a missense SNP (Thr>Ile; rs2707466) located in the WNT16 gene (7q31), associated with CBT (effect size of -0.11 standard deviations [SD] per C allele, P = 6.2 × 10(-9)). This SNP, as well as another nonsynonymous SNP rs2908004 (Gly>Arg), also had genome-wide significant association with forearm BMD (-0.14 SD per C allele, P = 2.3 × 10(-12), and -0.16 SD per G allele, P = 1.2 × 10(-15), respectively). Four genome-wide significant SNPs arising from BMD meta-analysis were tested for association with forearm fracture. SNP rs7776725 in FAM3C, a gene adjacent to WNT16, was associated with a genome-wide significant increased risk of forearm fracture (OR = 1.33, P = 7.3 × 10(-9)), with genome-wide suggestive signals from the two missense variants in WNT16 (rs2908004: OR = 1.22, P = 4.9 × 10(-6) and rs2707466: OR = 1.22, P = 7.2 × 10(-6)). We next generated a homozygous mouse with targeted disruption of Wnt16. Female Wnt16(-/-) mice had 27% (P<0.001) thinner cortical bones at the femur midshaft, and bone strength measures were reduced between 43%-61% (6.5 × 10(-13)

Evidence for threshold effects of 25-hydroxyvitamin D on glucose tolerance and insulin resistance in black and white obese postmenopausal women.

We identified normal vs. abnormal 25-hydroxyvitamin D [25(OH)D] concentrations by examining the relation of 25(OH)D to non-bone-related measures (plasma glucose, insulin resistance, lipids, blood pressure, fitness, obesity, and regional adiposity) and asking whether there is a 25(OH)D concentration above and below which the relation between 25(OH)D and outcome changes. We examined the relation between 25(OH)D and outcome by race to see whether race-specific normal ranges are needed, and we examined the role of insulin-like growth factor-1 (IGF-1) in modulating the relation between 25(OH)D and outcome. In a cross-sectional study of 239 overweight and obese, sedentary postmenopausal women without diabetes (83 black, 156 white), outcome measures included plasma lipids, glucose, insulin, homeostasis model assessment of insulin resistance (HOMA-IR), IGF-1, parathyroid hormone (PTH), aerobic fitness, body composition, subcutaneous abdominal and visceral fat, and blood pressure. We identified threshold effects in the association between 25(OH)D and these variables using piecewise linear regressions. We found that 25(OH)D was inversely related to fasting glucose, fasting and 2-h insulin, HOMA-IR, visceral abdominal fat, percentage fat, PTH, and triglycerides. Evidence for a threshold effect of 25(OH)D was found for 2-h glucose, 2-h insulin, fasting insulin, and HOMA-IR. There was no evidence suggesting the need for race-specific normal 25(OH)D concentrations. IGF-1 modulated the relation between 25(OH)D and outcome but only below, and not above, a threshold 25(OH)D concentration. Our findings suggest a threshold effect of 25(OH)D on glucose-insulin metabolism such that 25(OH)D ≥ ∼26 µg/L (65.0 pmol/L) supports normal glucose homeostasis and that the same cut point defining normal 25(OH)D concentration can be used in black and white women. This study was registered at clinicaltrials.gov as NCT01798030.

Meta-analysis of genome-wide studies identifies MEF2C SNPs associated with bone mineral density at forearm.

BACKGROUND: Forearm fractures affect 1.7 million individuals worldwide each year and most occur earlier in life than hip fractures. While the heritability of forearm bone mineral density (BMD) and fracture is high, their genetic determinants are largely unknown. AIM: To identify genetic variants associated with forearm BMD and forearm fractures. METHODS: BMD at distal radius, measured by dual-energy x-ray absorptiometry, was tested for association with common genetic variants. We conducted a meta-analysis of genome-wide association studies for BMD in 5866 subjects of European descent and then selected the variants for replication in 715 Mexican American samples. Gene-based association was carried out to supplement the single-nucleotide polymorphism (SNP) association test. We then tested the BMD-associated SNPs for association with forearm fracture in 2023 cases and 3740 controls. RESULTS: We found that five SNPs in the introns of MEF2C were associated with forearm BMD at a genome-wide significance level (p<5×10(-8)) in meta-analysis (lead SNP, rs11951031[T] -0.20 SDs per allele, p=9.01×10(-9)). The gene-based association test suggested an association between MEF2C and forearm BMD (p=0.003). The association between MEF2C variants and risk of fracture did not achieve statistical significance (SNP rs12521522[A]: OR=1.14 (95% CI 0.92 to 1.35), p=0.14). Meta-analysis also revealed two genome-wide suggestive loci at CTNNA2 and 6q23.2. CONCLUSIONS: These findings demonstrate that variants at MEF2C were associated with forearm BMD, implicating this gene in the determination of BMD at forearm.

Critical Role for 24-Hydroxylation in Homeostatic Regulation of Vitamin D Metabolism.

Context: The body has evolved homeostatic mechanisms to maintain free levels of Ca+2 and 1,25-dihydroxyvitamin D [1,25(OH)2D] within narrow physiological ranges. Clinical guidelines emphasize important contributions of PTH in maintaining this homeostasis. Objective: To investigate mechanisms of homeostatic regulation of vitamin D (VitD) metabolism and to apply mechanistic insights to improve clinical assessment of VitD status. Design: Crossover clinical trial studying participants before and after VitD3-supplementation. Setting: Community. Participants: 11 otherwise healthy individuals with VitD-deficiency (25-hydroxyvitamin D [25(OH)D] ≤20 ng/mL). Interventions: VitD3-supplements (50,000 IU once or twice a week depending on BMI, for 4-6 weeks) were administered to achieve 25(OH)D≥30 ng/mL. Results: VitD3-supplementation significantly increased mean 25(OH)D by 2.7-fold and 24,25-dihydroxyvitamin D [24,25(OH)2D] by 4.3-fold. In contrast, mean levels of PTH, FGF23, and 1,25(OH)2D did not change. Mathematical modeling suggested that 24-hydroxylase activity was maximal for 25(OH)D≥50 ng/mL and achieved a minimum (~90% suppression) with 25(OH)D<10-20 ng/mL. The 1,25(OH)2D/24,25(OH)2D ratio better predicted modeled 24-hydroxylase activity (h) (ρ=-0.85; p=0.001) compared to total plasma 25(OH)D (ρ=0.51; p=0.01) and the 24,25(OH)2D/25(OH)D ratio (ρ=0.37; p=0.3). Conclusions: Suppression of 24-hydroxylase provides a first line of defense against symptomatic VitD-deficiency by decreasing metabolic clearance of 1,25(OH)2D. The 1,25(OH)2D/24,25(OH)2D ratio provides a useful index of VitD status since it incorporates 24,25(OH)2D levels and therefore, provides insight into 24-hydroxylase activity. When VitD availability is limited, this suppresses 24-hydroxylase activity - thereby decreasing the level of 24,25(OH)2D and increasing the 1,25(OH)2D/24,25(OH)2D ratio. Thus, an increased 1,25(OH)2D/24,25(OH)2D ratio signifies triggering of homeostatic regulation, which occurs at early stages of VitD-deficiency.

Vitamin D Deficiency Increases Vulnerability to Canagliflozin-induced Adverse Effects on 1,25-Dihydroxyvitamin D and PTH.

CONTEXT: Canagliflozin has been reported to increase the risk of bone fracture-possibly mediated by decreasing 1,25-dihydroxyvitamin D (1,25(OH)2D) and increasing parathyroid hormone (PTH). OBJECTIVE: This work investigated whether baseline vitamin D (VitD) deficiency renders individuals vulnerable to this adverse effect and whether VitD3 supplementation is protective. METHODS: This community-based, outpatient study had a paired design comparing individual participants before and after VitD3 supplementation. Eleven VitD-deficient (25-hydroxyvitamin D [25(OH)D] ≤ 20 ng/mL) individuals were recruited from the Amish population in Lancaster, Pennsylvania. Participants underwent 2 canagliflozin challenge protocols (300 mg daily for 5 days): the first before and the second after VitD3 supplementation. In the VitD3 supplementation protocol, participants received VitD3 supplementation (50 000 IU once or twice a week depending on body mass index for 4-6 weeks) to achieve 25(OH)D of 30 ng/mL or greater. Two coprimary end points were identified: effects of VitD3 supplementation on canagliflozin-induced changes in 1,25(OH)2D and PTH. Secondary end points included effects of VitD3 supplementation on baseline levels of VitD metabolites and PTH. RESULTS: VitD3 supplementation increased mean 25(OH)D from 16.5 ± 1.6 to 44.3 ± 5.5 ng/mL (P = .0006) and 24,25-dihydroxyvitamin D (24,25(OH)2D) from 1.0 ± 0.1 to 4.3 ± 0.6 ng/mL (P = .0002). Mean 1,25(OH)2D and PTH were unchanged. VitD3 supplementation decreased the magnitude of canagliflozin-induced changes in 1,25(OH)2D (from -31.3%±4.7% to -9.3%±8.3%; P = .04) and PTH (from +36.2%±6.2% to +9.7%±3.7%; P = .005). CONCLUSION: VitD deficiency rendered individuals more vulnerable to adverse effects of canagliflozin on biomarkers associated with bone health. VitD3 supplementation was protective against canagliflozin's short-term adverse effects on 1,25(OH)2D and PTH.

Critical Role for 24-Hydroxylation in Homeostatic Regulation of Vitamin D Metabolism.

CONTEXT: The body has evolved homeostatic mechanisms to maintain free levels of Ca+2 and 1,25-dihydroxyvitamin D [1,25(OH)2D] within narrow physiological ranges. Clinical guidelines emphasize important contributions of PTH in maintaining this homeostasis. OBJECTIVE: To investigate mechanisms of homeostatic regulation of vitamin D (VitD) metabolism and to apply mechanistic insights to improve clinical assessment of VitD status. DESIGN: Crossover clinical trial studying participants before and after VitD3-supplementation. SETTING: Community. PARTICIPANTS: 11 otherwise healthy individuals with VitD-deficiency (25-hydroxyvitamin D [25(OH)D] ≤20 ng/mL). INTERVENTIONS: VitD3-supplements (50,000 IU once or twice a week depending on BMI, for 4-6 weeks) were administered to achieve 25(OH)D≥30 ng/mL. RESULTS: VitD3-supplementation significantly increased mean 25(OH)D by 2.7-fold and 24,25-dihydroxyvitamin D [24,25(OH)2D] by 4.3-fold. In contrast, mean levels of PTH, FGF23, and 1,25(OH)2D did not change. Mathematical modeling suggested that 24-hydroxylase activity was maximal for 25(OH)D≥50 ng/mL and achieved a minimum (∼90% suppression) with 25(OH)D<10-20 ng/mL. The 1,25(OH)2D/24,25(OH)2D ratio better predicted modeled 24-hydroxylase activity (h) (ρ=-0.85; p=0.001) compared to total plasma 25(OH)D (ρ=0.51; p=0.01) and the 24,25(OH)2D/25(OH)D ratio (ρ=0.37; p=0.3). CONCLUSIONS: Suppression of 24-hydroxylase provides a first line of defense against symptomatic VitD-deficiency by decreasing metabolic clearance of 1,25(OH)2D. The 1,25(OH)2D/24,25(OH)2D ratio provides a useful index of VitD status since it incorporates 24,25(OH)2D levels and therefore, provides insight into 24-hydroxylase activity. When VitD availability is limited, this suppresses 24-hydroxylase activity - thereby decreasing the level of 24,25(OH)2D and increasing the 1,25(OH)2D/24,25(OH)2D ratio. Thus, an increased 1,25(OH)2D/24,25(OH)2D ratio signifies triggering of homeostatic regulation, which occurs at early stages of VitD-deficiency.

Acute pharmacodynamic responses to sitagliptin: Drug-induced increase in early insulin secretion in oral glucose tolerance test.

DPP4 inhibitors are widely prescribed as treatments for type 2 diabetes. Because drug responses vary among individuals, we initiated investigations to identify genetic variants associated with the magnitude of drug responses. Sitagliptin (100 mg) was administered to 47 healthy volunteers. Several endpoints were measured to assess clinically relevant responses - including the effect of sitagliptin on glucose and insulin levels during an oral glucose tolerance test (OGTT). This pilot study confirmed that sitagliptin (100 mg) decreased the area under the curve for glucose during an OGTT (p = 0.0003). Furthermore, sitagliptin promoted insulin secretion during the early portion of the OGTT as reflected by an increase in the ratio of plasma insulin at 30 min divided by plasma insulin at 60 min (T30:T60) from mean ± SEM 0.87 ± 0.05 to 1.62 ± 0.36 mU/L (p = 0.04). The magnitude of sitagliptin's effect on insulin secretion (as judged by the increase in the T30:T60 ratio for insulin) was correlated with the magnitude of sitagliptin-induced increase in the area under the curve for intact plasma GLP1 levels during the first hour of the OGTT. This study confirmed previously reported sex differences in glucose and insulin levels during an OGTT. Specifically, females exhibited higher levels of glucose and insulin at the 90-180 min time points. However, we did not detect significant sex-associated differences in the magnitude of sitagliptin-induced changes in T30:T60 ratios for either glucose or insulin. In conclusion, T30:T60 ratios for insulin and glucose during an OGTT provide useful indices to assess pharmacodynamic responses to DPP4 inhibitors.

Value of Vitamin D Metabolite Ratios in 3 Patients as Diagnostic Criteria to Assess Vitamin D Status.

Although clinical guidelines recommend measuring total plasma 25-hydroxyvitamin D (25[OH]D) to assess vitamin D (VitD) status, this index does not account for 3-fold inter-individual variation in VitD binding protein (VDBP) level. We present 3 individuals with total plasma 25(OH)D levels of 10.8 to 12.3 ng/mL (27-30.7 nmol/L). Because Endocrine Society guidelines define VitD deficiency as 25(OH)D ≤ 20 ng/mL (50 nmol/L), all 3 would be judged to be VitD deficient. VitD3 supplementation increased 25(OH)D to the range of 31.7 to 33.8 ng/mL (79.1-84.4 nmol/L). Patient #1 exhibited secondary hyperparathyroidism; VitD3 supplementation decreased parathyroid hormone (PTH) by 34% without a clinically significant change in PTH levels in the other 2 individuals. Thus, 25(OH)D level did not distinguish between the 1 patient who had secondary hyperparathyroidism and the 2 who did not. We therefore inquired whether VitD metabolite ratios (which are VDBP-independent) might distinguish among these 3 individuals. Of all the assessed ratios, the 1,25(OH)2D/24,25(OH)2D ratio was the most informative, which had a value of 102 pg/ng in the individual with secondary hyperparathyroidism but lower values (41 and 20 pg/ng) in the other 2 individuals. These cases illustrate the value of the 1,25(OH)2D/24,25(OH)2D ratio to provide clinically relevant information about VitD status.

Causal relationship between obesity and vitamin D status: bi-directional Mendelian randomization analysis of multiple cohorts.

BACKGROUND: Obesity is associated with vitamin D deficiency, and both are areas of active public health concern. We explored the causality and direction of the relationship between body mass index (BMI) and 25-hydroxyvitamin D [25(OH)D] using genetic markers as instrumental variables (IVs) in bi-directional Mendelian randomization (MR) analysis. METHODS AND FINDINGS: We used information from 21 adult cohorts (up to 42,024 participants) with 12 BMI-related SNPs (combined in an allelic score) to produce an instrument for BMI and four SNPs associated with 25(OH)D (combined in two allelic scores, separately for genes encoding its synthesis or metabolism) as an instrument for vitamin D. Regression estimates for the IVs (allele scores) were generated within-study and pooled by meta-analysis to generate summary effects. Associations between vitamin D scores and BMI were confirmed in the Genetic Investigation of Anthropometric Traits (GIANT) consortium (n = 123,864). Each 1 kg/m(2) higher BMI was associated with 1.15% lower 25(OH)D (p = 6.52×10⁻²7). The BMI allele score was associated both with BMI (p = 6.30×10⁻6²) and 25(OH)D (-0.06% [95% CI -0.10 to -0.02], p = 0.004) in the cohorts that underwent meta-analysis. The two vitamin D allele scores were strongly associated with 25(OH)D (p≤8.07×10⁻57 for both scores) but not with BMI (synthesis score, p = 0.88; metabolism score, p = 0.08) in the meta-analysis. A 10% higher genetically instrumented BMI was associated with 4.2% lower 25(OH)D concentrations (IV ratio: -4.2 [95% CI -7.1 to -1.3], p = 0.005). No association was seen for genetically instrumented 25(OH)D with BMI, a finding that was confirmed using data from the GIANT consortium (p≥0.57 for both vitamin D scores). CONCLUSIONS: On the basis of a bi-directional genetic approach that limits confounding, our study suggests that a higher BMI leads to lower 25(OH)D, while any effects of lower 25(OH)D increasing BMI are likely to be small. Population level interventions to reduce BMI are expected to decrease the prevalence of vitamin D deficiency.