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.

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

Aim: 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. Methods: 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). Results: 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 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. Conclusions: T30:T60 ratios for insulin and glucose during an OGTT provide useful indices to assess pharmacodynamic responses to DPP4 inhibitors.

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.

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 in response to VitD3-supplementation. 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. Suppression of 24-hydroxylase activity was triggered by mild-moderate VitD-deficiency, which sustains physiological levels of 1,25(OH)2D by suppressing metabolic clearance of 1,25(OH)2D. VitD metabolite ratios [e.g., 1,25(OH)2D/24,25(OH)2D] provide useful indices of VitD status. 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, a high 1,25(OH)2D/24,25(OH)2D ratio signifies triggering of homeostatic regulation in response to mild-moderate VitD-deficiency. Conclusions: Suppression of 24-hydroxylase activity 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 can provide a useful index of clinical VitD status.

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.

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) 2 D] and increasing PTH. Objective. To investigate whether baseline vitamin D (VitD) deficiency renders individuals vulnerable to this adverse effect and whether VitD3 supplementation is protective. Design. This study had a paired design comparing individual participants before and after VitD3 supplementation. Setting. Community-based outpatient. Patients. 11 VitD deficient (25-hydroxyvitamin D [25(OH)D] ≤ 20 ng/mL) individuals recruited from the Amish population in Lancaster PA. Interventions. Participants underwent two canagliflozin challenge protocols (300 mg daily for five 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 BMI for 4-6 weeks) to achieve 25(OH)D ≥ 30 ng/mL. Main Outcome Measures. Two co-primary endpoints were identified: effects of VitD3 supplementation on canagliflozin-induced changes in 1,25(OH) 2 D and PTH. Secondary endpoints 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=0.0006) and 24,25-dihydroxyvitamin D [24,25(OH) 2 D] from 1.0±0.1 to 4.3±0.6 ng/mL (p=0.0002). Mean 1,25(OH) 2 D and PTH were unchanged. VitD3 supplementation decreased the magnitude of canagliflozin-induced changes in 1,25(OH) 2 D (from -31.3%±4.7% to -9.3%±8.3%; p=0.04) and PTH (from +36.2%±6.2% to +9.7%±3.7%; p=0.005). Conclusions. 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) 2 D and PTH.

Pharmacogenetics of SGLT2 Inhibitors: Validation of a sex-agnostic pharmacodynamic biomarker.

Aim: SGLT2 inhibitors provide multiple benefits to patients with type 2 diabetes - including improved glycemic control and decreased risks of cardiorenal disease. Because drug responses vary among individuals, we initiated investigations to identify genetic variants associated with the magnitude of drug responses. Methods: Canagliflozin (300 mg) 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=6x10), 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 GFR, or age in these healthy non-diabetic individuals with estimated GFR>60 mL/min/1.73m 2 . 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 age and renal function. Registration: NCT02462421 ( clinicaltrials.gov ). Funding: Research grants from the National Institute of Diabetes and Digestive and Kidney Diseases: R21DK105401, R01DK108942, T32DK098107, and P30DK072488.

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

Background: GLP1R agonists provide multiple benefits to patients with type 2 diabetes - including improved glycemic 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, sc) or saline (0.2 mL, sc) 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 designed as a crossover study 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 demonstrated that exenatide increased glucose effectiveness (S g ) by 32% (p=0.0008) but did not significantly affect insulin sensitivity (S i ). The exenatide-induced increase in insulin secretion made the largest contribution to inter-individual variation in exenatide-induced acceleration of glucose disappearance while inter-individual variation in the drug effect on S g contributed to a lesser extent (ß=0.58 or 0.27, respectively). Conclusions: This pilot study provides validation for the value of an FSIGT (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 GLP1R agonists' effects on glucose metabolism: first phase insulin secretion, glucose disappearance rates, and glucose effectiveness. Registration: NCT02462421 (clinicaltrials.gov). Funding: American Diabetes Association (1-16-ICTS-112); National Institute of Diabetes and Digestive and Kidney Disease (R01DK130238, T32DK098107, P30DK072488).

SGLT2i Improves Glycemic Control in Patients With Congenital Severe Insulin Resistance.

Insulin-resistant diabetes in Rabson-Mendenhall syndrome (RMS) is relatively unresponsive to first-line antidiabetic treatments, including metformin and insulin. We report 2 patients with RMS treated with 2 different sodium-glucose cotransporter inhibitors 2: empagliflozin in an 11-year-old boy and dapagliflozin in a 12-year-old girl. In the first patient, we began empagliflozin at 2.5 mg/day and increased the dose to 10 mg/day over 3 months. During treatment with empagliflozin, the amount of time during which the patient maintained serum glucose in the 70 to 180 mg/dL target range increased by 2 hours per day. Hemoglobin A1C dropped from >14% to 11.9%, urinary calcium increased almost twofold, and ß-hydroxybutyrate remained <2.5 mmol/L. Because glycemic control did not further improve with dose escalation, we reverted to the 2.5 mg/day dose. We initiated dapagliflozin in a second patient at 5 mg/day and witnessed a reduction of hemoglobin A1C from 8.5% to 6.2% after 6 months and a mild increase in urinary excretion of phosphorus but not calcium. Insulin levels fell by >50%. In 2 patients with RMS, empagliflozin and dapagliflozin were well tolerated and improved glycemic control without significantly increasing ketonemia. Renal calcium excretion should be carefully monitored.

Genetic and functional evidence links a missense variant in B4GALT1 to lower LDL and fibrinogen.

Increased blood levels of low-density lipoprotein cholesterol (LDL-C) and fibrinogen are independent risk factors for cardiovascular disease. We identified associations between an Amish-enriched missense variant (p.Asn352Ser) in a functional domain of beta-1,4-galactosyltransferase 1 (B4GALT1) and 13.9 milligrams per deciliter lower LDL-C (P = 4.1 × 10­19) and 29 milligrams per deciliter lower plasma fibrinogen (P = 1.3 × 10­5). B4GALT1 gene­based analysis in 544,955 subjects showed an association with decreased coronary artery disease (odds ratio = 0.64, P = 0.006). The mutant protein had 50% lower galactosyltransferase activity compared with the wild-type protein. N-linked glycan profiling of human serum found serine 352 allele to be associated with decreased galactosylation and sialylation of apolipoprotein B100, fibrinogen, immunoglobulin G, and transferrin. B4galt1 353Ser knock-in mice showed decreases in LDL-C and fibrinogen. Our findings suggest that targeted modulation of protein galactosylation may represent a therapeutic approach to decreasing cardiovascular disease.

Pharmacological treatment of hyperglycemia in type 2 diabetes.

Diabetes mellitus is a major public health problem, affecting about 10% of the population. Pharmacotherapy aims to protect against microvascular complications, including blindness, end-stage kidney disease, and amputations. Landmark clinical trials have demonstrated that intensive glycemic control slows progression of microvascular complications (retinopathy, nephropathy, and neuropathy). Long-term follow-up has demonstrated that intensive glycemic control also decreases risk of macrovascular disease, albeit rigorous evidence of macrovascular benefit did not emerge for over a decade. The US FDA's recent requirement for dedicated cardiovascular outcome trials ushered in a golden age for understanding the clinical profiles of new type 2 diabetes drugs. Some clinical trials with sodium-glucose cotransporter-2 (SGLT2) inhibitors and glucagon-like peptide 1 (GLP1) receptor agonists reported data demonstrating cardiovascular benefit (decreased risk of major adverse cardiovascular events and hospitalization for heart failure) and slower progression of diabetic kidney disease. This Review discusses current guidelines for use of the 12 classes of drugs approved to promote glycemic control in patients with type 2 diabetes. The Review also anticipates future developments with potential to improve the standard of care: availability of generic dipeptidylpeptidase-4 (DPP4) inhibitors and SGLT2 inhibitors; precision medicine to identify the best drugs for individual patients; and new therapies to protect against chronic complications of diabetes.

YIPF5 mutations cause neonatal diabetes and microcephaly: progress for precision medicine and mechanistic understanding.

Identifying genes that result in monogenic diabetes can provide insights that can build a scientific foundation for precision medicine. At present, nearly 20% of neonatal diabetes cases have unknown causes. In this issue of the JCI, De Franco and Lytrivi et al. sequenced the genome of two probands with a rare neonatal diabetes subtype that also associated with microcephaly and epilepsy. The authors revealed mutations in the YIPF5 gene. YIPF5 resides in the Golgi apparatus and is thought to play a critical role in vesicular trafficking. Notably, disrupting YIPF5 in ß cell-based models induced ER stress signaling and resulted in the accumulation of intracellular proinsulin. We believe that utilizing registries and biobanks to reveal other monogenic atypical forms of diabetes is an important approach to gaining insight and suggest that an insulin sensitizer may alleviate ER stress associated with YIPF5 disruption by decreasing the demand for insulin secretion.

The complicated clinical course in a case of atypical lipodystrophy after development of neutralizing antibody to metreleptin: treatment with setmelanotide.

SUMMARY: A patient with atypical partial lipodystrophy who had a transient initial response to metreleptin experienced acute worsening of her metabolic state when neutralizing antibodies against metreleptin appeared. Because her metabolic status continued to deteriorate, a therapeutic trial with melanocortin-4 receptor agonist setmelanotide, that is believed to function downstream from leptin receptor in the leptin signaling system, was undertaken in an effort to improve her metabolic status for the first time in a patient with lipodystrophy. To achieve this, a compassionate use (investigational new drug application; IND) was initiated (NCT03262610). Glucose control, body fat by dual-energy X-ray absorptiometry and MRI, and liver fat by proton density fat fraction were monitored. Daily hunger scores were assessed by patient filled questionnaires. Although there was a slight decrease in hunger scales and visceral fat, stimulating melanocortin-4 receptor by setmelanotide did not result in any other metabolic benefit such as improvement of hypertriglyceridemia or diabetes control as desired. Targeting melanocortin-4 receptor to regulate energy metabolism in this setting was not sufficient to obtain a significant metabolic benefit. However, complex features of our case make it difficult to generalize these observations to all cases of lipodystrophy. It is still possible that melanocortin-4 receptor agonistic action may offer some therapeutic benefits in leptin-deficient patients. LEARNING POINTS: A patient with atypical lipodystrophy with an initial benefit with metreleptin therapy developed neutralizing antibodies to metreleptin (Nab-leptin), which led to substantial worsening in metabolic control. The neutralizing activity in her serum persisted for longer than 3 years. Whether the worsening in her metabolic state was truly caused by the development of Nab-leptin cannot be fully ascertained, but there was a temporal relationship. The experience noted in our patient at least raises the possibility for concern for substantial metabolic worsening upon emergence and persistence of Nab-leptin. Further studies of cases where Nab-leptin is detected and better assay systems to detect and characterize Nab-leptin are needed. The use of setmelanotide, a selective MC4R agonist targeting specific neurons downstream from the leptin receptor activation, was not effective in restoring metabolic control in this complex patient with presumed diminished leptin action due to Nab-leptin. Although stimulating the MC4R pathway was not sufficient to obtain a significant metabolic benefit in lowering triglycerides and helping with her insulin resistance as was noted with metreleptin earlier, there was a mild reduction in reported food intake and appetite. Complex features of our case make it difficult to generalize our observation to all leptin-deficient patients. It is possible that some leptin-deficient patients (especially those who need primarily control of food intake) may still theoretically benefit from MC4R agonistic action, and further studies in carefully selected patients may help to tease out the differential pathways of metabolic regulation by the complex network of leptin signaling system.