European Review for Medical and Pharmacological Sciences 2019; 23: 1357-1378 Mendelian obesity, molecular pathways and pharmacological therapies: a review

S. PAOLACCI1, A. BORRELLI2, L. STUPPIA3, F.C. CAMPANILE4, T. DALLAVILLA1, J. KRAJČOVIČ5, D. VESELENYIOVA5, T. BECCARI6, V. UNFER7, M. BERTELLI8; GENEOB PROJECT

1MAGI’S Lab, Genetic Testing Laboratory, Rovereto (TN), Italy 2Obesity Center, Pineta Grande Hospital, Castelvolturno (CE), Italy 3Department of Psychological Sciences, Health and Territory, University “G. d’Annunzio”, Chieti-Pescara, Italy 4Department of Surgery, San Giovanni Decollato Andosilla Hospital, ASL VT, Civita Castellana, Italy 5Department of Biology, Faculty of Natural Sciences, University of ss. Cyril and Methodius, Trnava, Slovakia 6Department of Pharmaceutical Sciences, University of Perugia, Perugia, Italy 7Department of Developmental and Social Psychology, Faculty of Medicine and Psychology, Sapienza, University of Rome, Rome, Italy 8MAGI Euregio, Nonprofit Genetic Testing Laboratory, Bolzano, Italy

Abstract. – OBJECTIVE: In this qualitative re- Introduction view we analyze the major pathways and mecha- nisms involved in the onset of genetically-deter- Adipocytes are the major constituents of whi- mined obesity (Mendelian obesity), identifying te adipose tissue. They control energy balance by possible pharmacological treatments and trials. MATERIALS AND METHODS: We searched storing and mobilizing triacylglycerol, and they PubMed with the keywords (obesity[Title/Ab- play roles in endocrine and paracrine regulation. stract]) AND mutation[Title/Abstract], and OMIM Adipose tissue controls glucose metabolism, ap- with the keyword “obesity”. In both cases, we petite, immunological and inflammatory respon- selected non-syndromic Mendelian obesity. We ses, angiogenesis, blood pressure, and reproducti- then searched ClinicalTrials.gov with the follow- ve function through many specific factors. When ing criteria: “recruitment status: active, not re- adipocytes accumulate, they form adipose tissue. cruiting and completed”; “study type: interven- tional (clinical trial)”; “study results: with re- The amount of adipose tissue in the body is mainly sults”; type of intervention: “drug or dietary regulated by leptin, a hormone produced by adi- supplement”. pocytes themselves. Adipose tissue expands when RESULTS: From the PubMed and OMIM search- the number and size of adipocytes increases1. An es we obtained a total of 15 genes associated excessive amount or size of adipocytes can gene- with monogenic Mendelian obesity. From Clini- rate a morbid condition called obesity. Obesity is calTrials.gov we retrieved 46 completed or active trials of pharmacological treatments. highly heritable and arises from the interplay of CONCLUSIONS: We summarized the molecu- many genes and environmental factors. It is de- lar bases of Mendelian obesity and searched for fined as resulting from a prolonged imbalance any clinical trials completed or underway for the between calorie intake and energy utilization. An treatment of severe forms of obesity. Most Men- understanding of how these variations influence delian obesities are linked to dysfunctions in susceptibility to become or remain obese will ho- the leptin/melanocortin signaling pathway, and pefully provide insights into how obesity occurs most of the possible drugs target this pathway 2 in order to improve energy expenditure and re- and how to prevent and treat it . It is important to duce food intake. note that about 5% of cases of obesity are mono- genic3. Most mutant genes in Mendelian obesity Key Words: are also involved in a predisposition for common Mendelian obesity, Leptin/melanocortin pathway, obesity through the action of common single nu- Adipogenesis, Clinical trials. cleotide polymorphisms (SNPs) (minor allele fre-

Corresponding Author: Stefano Paolacci, MD; email: [email protected] 1357 S. Paolacci, A. Borrelli, L. Stuppia, F.C. Campanile, T. Dallavilla, et al. quency >1%). In this qualitative review, we clarify The Leptin/Melanocortin Pathway and analyze the major pathways and mechanisms Leptin (LEP) is secreted by adipose tissue and involved in the onset of genetically-determined binds to leptin receptors (LEPR) in the hypotha- obesity (Mendelian obesity). We also review dru- lamus, where it controls food intake through the gs that may be or are already used to treat obesity. melanocortin pathway. Leptin acts in the arcuate nucleus where it activates neurons that express the anorexigenic peptides pro-opiomelanocortin Materials and Methods (POMC), as well as cocaine- and amphetami- ne-related transcript preprotein (CARTPT) deri- To identify genes involved in isolated mono- vatives that control food intake. Leptin also has genic obesity we searched articles published in inhibitory effects on the neurons that express the English up to October 2018 in PubMed with the orexigenic peptides neuropeptide Y (NPY) and following combination of terms: (obesity[Title/ Agouti-related protein (AGRP), which increa- Abstract]) AND mutation[Title/Abstract], where- se food intake. Both POMC/CARTPT and NPY/ as in OMIM we searched using the word: “obesi- AGRP subsets of neurons have many connections ty”. From the records retrieved, we only selected with other hypothalamic nuclei. The binding of publications dealing with Mendelian non-syn- leptin to its receptor initiates a cascade of signal dromic obesity. We only considered disorders transduction pathways, deficits of which may cau- for which the pathogenicity of the mutations was se leptin resistance. With the help of Src homolo- supported by segregation studies in more than gy 2B adapter protein 1 (SH2B1), LEPR recruits one multigenerational family and/or by functio- JAK2, which in turn phosphorylates the intracel- nal studies. To find drugs useful to treat obesity lular domain of LEPR. The phosphorylated in- we searched ClinicalTrials.gov with the following tracellular domain of LEPR binds and activates criteria: “recruitment status: active, not recrui- STAT3, which with the aid of Tubby bipartite tran- ting, and completed”; “study type: interventional scription factor (TUB), translocates to the nucleus (clinical trial)”; “study results: with results”; type and acts as a transcription factor, mediating the of intervention “drug or dietary supplement”. anorexigenic effects of leptin4. Besides the JAK/ From the list obtained, we selected drugs that STAT pathway, another important intracellular were used to treat obesity, excluding drugs used pathway for energy homeostasis is the PI3K pa- only to treat obesity-related complications. thway. Indeed, LEPR stimulation mediates tyrosi- ne phosphorylation of IRS proteins by JAK2. IRS in turn activates PI3K that generates phosphati-

Results dylinositol-3,4,5-trisphosphate (PIP3) from pho- sphatidylinositol-4,5-bisphosphate (PIP2). The From the PubMed and OMIM searches, we importance of this pathway in the regulation of obtained 1878 and 655 entries, respectively, from energy balance is highlighted by the fact that ad- which we extracted a total of 15 genes associated ministration of myo-inositol (the core constituent with monogenic Mendelian non-syndromic obe- of PIP) may improve endocrine and metabolic pa- sity. From ClinicalTrials.gov we retrieved a total rameters, and reduce BMI in overweight patients 5 of 433 entries and as described in “Methods”, we with polycystic ovary syndrome . PIP3 induces selected 46 completed or active trials of pharma- activation of PDK1 that triggers a cascade inclu- cological treatments. ding Akt and members of the PKC family. Akt activates mTOR and inhibits the transcription factor FOXO16. Note the close interaction betwe- Mendelian Monogenic Obesity en STAT3 and FOXO1, both of which bind AGRP and POMC promoters. While STAT3 inhibits Most genes known to be associated with mo- expression of AGRP and activates expression of nogenic obesity are related to the leptin/melano- POMC, FOXO1 has the opposite effect6. cortin pathway (Figure 1) and are expressed in the After its synthesis, POMC is cleaved by hypothalamus. They code for proteins involved in prohormone convertase 1 and 2 (PC1/2) and by food intake/energy expenditure balance. Another carboxypeptidase E (CPE), which are expressed pathway associated with obesity leads to the for- in the feeding centers of the ARC, to yield mela- mation of mature white adipose tissue and invol- nocortins α-, β- and γ-MSH. The actions of me- ves several growth and transcription factors. lanocortins are mediated by two G-protein cou-

1358 Mendelian obesity, molecular pathways and pharmacological therapies: a review pled receptors known as melanocortin 3 and 4 culating leptin (LEP) binds to the leptin receptor receptors (MC3R, MC4R) that play a central role (LEPR) in the hypothalamus, which activates in the control of body weight, decreasing food downstream signaling pathways that inhibit fee- intake and favoring weight loss6. Melanocortin ding and promote energy expenditure. Consistent- 2 receptor accessory protein 2 (MRAP2) can re- ly, loss of function mutations in this gene leads to duce the responsiveness of MC3R and MC4R to the onset of obesity. Obesity linked to mutations α- and β-MSH. On the other hand, single-minded in LEP has a prevalence of < 1/1,000,000 and 1 (SIM1) acts as a facilitator of MC4R activi- autosomal recessive inheritance. LEP was first ty. MC4R activity also stimulates the release of linked to human Mendelian obesity in 1997 by brain-derived neurotrophic factor (BDNF), which Montague and collaborators studying an inbred binds to the neurotrophin receptor (NTRK2) and Pakistani family. They found a homozygous de- influences food intake and energy expenditure7. letion of a G in LEP in codon 133 in two indivi- duals (c.398delG). The mutation disrupted the re- Adipogenesis ading frame, leading to a premature stop codon10. Mesenchymal stem cells (MSCs) are the precur- In 2002, a third child from an inbred Pakistani sors of adipocytes. They can differentiate into adi- family with the same single-base deletion was re- pocytes, chondrocytes, myocytes or osteocytes. ported11. A fourth case with severe hyperphagia Differentiation towards the adipocyte lineage is and obesity, reported by Gibson et al12 in 2004, induced by chronic excessive energy intake and had the recurrent homozygous deletion12. In three elevated glucose uptake. Lineage determination is obese individuals with very low serum concentra- regulated by a network of extracellular signaling tions of leptin, Strobel et al13 (1998) found the first factors8. Although many molecular details of adi- pathogenic missense mutation in LEP (c.313C > pogenesis are still unknown, numerous factors in- T; p.Arg105Trp). All three affected individuals volved in this process have been identified. Some were hyperphagic and one was hyperglycemic13. stimulators include peroxisome proliferator-acti- Obesity due to mutations in LEPR has autoso- vated receptor γ (PPARG), enhancer binding pro- mal recessive inheritance and unknown prevalen- teins α, β and δ (CEBPA, CEBPB, CEBPD), single ce. Clement et al14 (1998) were the first to report a transducers and activators of transcription (STA- mutation in the human leptin receptor gene, a G Ts), the transcriptional factor sterol-regulatory-e- > A transition at the donor splice site in intron 16 lement-binding protein-1 (SRBP1), insulin-like (c.2598 + 1G > A) that causes obesity and pituitary growth factor I (IGF1), macrophage colony sti- dysfunction14. The mutation was homozygous in mulating factor, fatty acids, prostaglandins, glu- an inbred family of Algerian origin in which 3 out cocorticoids, Kruppel-like factors (KLFs), win- of 9 siblings had morbid obesity with onset in ear- gless and INT-1 proteins (WNTs), various cell ly childhood. In addition to obesity, the affected cycle proteins, clock proteins (BMAL1, NR1D1), sibs had no pubertal development and reduced interferon regulatory factors (IRF3, IRF4), B-cell secretion of growth hormone and thyrotropin14. factor 1 (EBF1), GATA-binding proteins 2 and 3, To determine the prevalence of pathogenic LEPR bone morphogenetic proteins (BMPs), transfor- mutations in severely obese patients, Farooqi et ming growth factor β (TGF-β), fibroblast growth al15 (2007) sequenced LEPR in 300 patients with factors (FGFs), insulin-like growth factor (IGF), hyperphagia and severe early-onset obesity, fin- Notch-mediated signaling and pro-inflamma- ding nonsense or missense LEPR mutations in tory cytokines. Inhibitors include glycoproteins, 2.7% of them. All missense mutations resulted in TGF-β, inflammatory cytokines and growth hor- impaired receptor signaling. Affected individuals mone1. In particular, PPARG is necessary and were characterized by hyperphagia, severe obe- sufficient for adipocyte differentiation, and is sity, alterations in immune function and hypogo- therefore a master regulator of adipogenesis. It is nadotropic hypogonadism. Their clinical features not surprising that mutations in this gene cause a were less severe than those of patients with con- form of Mendelian obesity9. genital leptin deficiency15. In general, the clini- cal phenotypes associated with leptin and LEPR Obesity Due to mutations in the Leptin deficiencies are broadly similar10,15. Patients have Gene, LEP, and its Receptor, LEPR normal birth weight but show rapid weight gain in LEP encodes a protein that is secreted into cir- the first few months of life. They often accumu- culation by white adipocytes and plays a major late subcutaneous fat over the trunk and limbs. role in the regulation of energy homeostasis. Cir- The most notable feature is intense hyperphagia.

1359 S. Paolacci, A. Borrelli, L. Stuppia, F.C. Campanile, T. Dallavilla, et al.

Children with leptin deficiency also have striking and severe hyperinsulinemia, whereas homozy- abnormalities in T cell number and function11. gotes were more severely affected than heterozy- Leptin and LEPR deficiency are generally asso- gotes. MC4R mutations are therefore inherited in ciated with hypothalamic hypothyroidism and a co-dominant manner21. hypogonadotropic hypogonadism11,13. Obesity Due to Mutations in POMC Obesity Due to Mutations in TUB POMC encodes a preproprotein that undergoes Tubby bipartite transcription factor (TUB) is a extensive post-translational processing via clea- member of the tubby-like proteins. It is a mem- vage by prohormone convertases. There are eight brane-bound transcription regulator that translo- cleavage sites in the preproprotein and processing cates to the nucleus in response to phosphoino- may yield as many as ten biologically active pep- sitide hydrolysis16. TUB is a substrate for insulin tides, involved in various cell functions. Peptides and leptin receptors in the hypothalamus. It is with roles in energy homeostasis, such as al- translocated to the nucleus after binding to LEPR pha-melanocyte-stimulating hormone (α-MSH), via JAK2. Inhibition of TUB expression in the which has anorexigenic effects, are generated hypothalamus results in increased food intake, in the hypothalamus25. Obesity associated with fasting glucose levels and hepatic glucose output, POMC mutations has autosomal recessive inheri- and in reduced oxygen consumption and POMC tance and a prevalence of < 1/1,000,000 in the ge- sensitivity to leptin17. Only a single mutation in neral population. The first pathogenic mutations TUB has so far been reported. The subject was in POMC were discovered in 1998. One patient a boy with deteriorating vision, obesity and nor- had biallelic loss-of-function mutations in exon 3 mal glucose/cholesterol/triglycerides levels from of POMC (c.7013G > T; p.Glu79* and c.7133delC) an inbred Caucasian family18. The mutation was that interfered with the appropriate synthesis a homozygous frameshift variant in TUB (c.1194_ of adrenocorticotropic hormone (ACTH) and 1195delAG, p.Arg398Serfs*9) that results in a α-MSH. The other patient was homozygous for truncated form of TUB. a mutation (c.-11C > A) in exon 2 that abolished POMC translation. The POMC gene was chosen Obesity Due to Mutations in for investigation because studies in animal mo- Melanocortin Receptor MC4R dels elucidated a central role of α-MSH in the MC4R is a G-protein-coupled receptor mainly regulation of food intake by activation of brain expressed in the brain and involved in energy in- MC4R26. In addition to obesity, patients with take and expenditure19. MC4R mutations are the POMC mutations displayed hypocortisolism, hair most common cause of human monogenic obesi- and skin hypopigmentation, neonatal hypoglyce- ty, having a frequency of 1.5%4 and a prevalence mia, seizures, cholestasis, and voracious appe- of 1-5/10,000 in the general population. Besides tite26. Although most research has been focused obesity, MC4R-deficient children also display on α-MSH, a missense loss-of-function mutation hyperinsulinemia and increased linear growth20. in β-MSH (c.14G > A; p.Cys5Tyr) has also been Interestingly, patients experience an increase in associated with childhood obesity. The lack of adiposity as well as lean mass21. The first muta- function of β-MSH reduces the amount of MSH tions in MC4R, c.631_634delGAGA and c.732in- peptide in the POMC/MC4R pathway, resulting sCATT, were discovered in humans in 1998. They in obesity27. are heterozygous frameshifts and were found in a severely obese child and an adult, respectively. Obesity Due to Mutations in PCSK1 Neither had evidence of impaired adrenal fun- Another gene involved in the onset of Mende- ction, and sexual development and fertility were lian obesity is PCSK1 that translates for pro-pro- normal22,23. Several other cases were subsequent- tein convertase subtilisin/kexin type 1. Obesity ly reported in extremely obese individuals whose linked to PCSK1 mutations has a prevalence of < BMIs all exceeded the 99th percentile4,24. Farooqi 1/1,000,000 in the general population and autoso- et al21 (2003) defined the clinical spectrum, mode mal recessive inheritance. Complete prohormone of inheritance and genotype-phenotype corre- convertase 1 deficiency was reported for the first lations. They found patients with heterozygous time in a patient with a compound heterozygous and homozygous loss-of-function mutations: he- mutation in PCSK1 (c.1447G > A; p.Gly483Arg, terozygous carriers had severe obesity, increased c.285 + 4A > C) characterized by severe early-on- lean mass, increased linear growth, hyperphagia, set obesity, hyperphagia, hypoglycemia, hypo-

1360 Mendelian obesity, molecular pathways and pharmacological therapies: a review gonadotropic hypogonadism, hypocortisolism, A girl with loss of one functional copy of elevated plasma proinsulin and elevated POMC BDNF, due to a de novo chromosomal inversion but low insulin concentrations28. Partial loss-of- (inv(11)(p13p15.3)), presented with hyperphagia, function heterozygous mutations in PCSK1 seem severe obesity, cognitive impairment, and hype- associated with a non-fully penetrant intermedia- ractivity35. Since cases associated with mutations te obesity phenotype29. in NTRK2 and BDNF are rare, their prevalence in the general population is unknown. Obesity Due to Mutations in SH2B1 The protein translated by SH2B1 is an adap- Obesity Due to Mutations in SIM1 ter protein for several members of the tyrosine Single-minded homolog 1 (SIM1) is a mem- kinase receptor family involved in multiple si- ber of the helix-loop-helix PAS family of nuclear gnaling pathways mediated by JAK and receptor transcription factors. It is expressed in kidneys tyrosine kinases, such as the receptor of insulin and the central nervous system and plays an es- or the receptor of brain-derived neurotrophic sential role in the formation of the paraventri- factor (BDNF). In leptin signaling, SH2B1 is cular nucleus (PVN) of the hypothalamus37,38. a key regulator of leptin sensitivity, binding to This could be a mechanism in which SIM1 plays and potentiating activation of JAK2 by globally a role in energy homeostasis, as PVN neurons enhancing downstream pathways. SH2B1 binds express MC4R which inhibits food intake5. Seve- simultaneously to JAK2 and IRS1 or IRS2, thus re early-onset obesity was observed in a girl with mediating formation of a JAK2, SH2B1 and IRS1 haploinsufficiency of SIM1. The proband had ear- or IRS2 complex. The result of this interaction is ly-onset obesity and a de novo balanced translo- the activation of the PI3K pathway30. Obesity as- cation between chromosomes 1p22.1 and 6q16.2. sociated with mutations in this gene has autoso- Her weight gain was due to excessive food inta- mal dominant inheritance with a prevalence of < ke39. Further support for the involvement of SIM1 1/1,000,000 in the general population. In humans, in obesity came from studies in which patients loss-of-function mutations in SH2B1 result in se- displayed Prader-Willi-like phenotype due to he- vere early-onset obesity. Eight different mutations terozygous mutations in SIM1. In particular, three have been reported: seven missense and one fra- mutations showed strong loss-of-function effects meshift variant31,32. All patients exhibit hyperpha- (p.Thr46Arg, p.His323Tyr, and p.Thr714Ala) and gia, early onset obesity, insulin resistance, and were associated with high intra-family risk for short stature31. Note that recurrent deletions of obesity40. In another study, at least six severely the SH2B1-containing region on the short arm of damaging heterozygous variants in SIM1 (p.Se- chromosome 16 have been associated with beha- r71Arg, p.Arg171His, p.Leu238Arg, p.Pro497Arg, vioral disorders and obesity33. p.Arg550His, p.Thr712Ile) were associated with severe obesity and neurobehavioral phenotype. Obesity Due to Mutations in the It is noteworthy that SIM1-mutant patients share Receptor NTRK2 and its Ligand BDNF many features with melanocortin-deficient pa- Neurotrophic receptor tyrosine kinase 2 tients, but they do not share the accelerated linear (NTRK2) is a member of the neurotrophin fa- growth and increased final height seen in MC4R mily, known to be involved in the development deficiency41. and in maintenance and function of peripheral and central neurons; it is thought to play a role Obesity Due to Mutations in KSR2 in mediating neuronal plasticity in the hypotha- Kinase suppressor of Ras 2 (KSR2) is a mo- lamus34. NTRK2 and its ligand BDNF are also lecular scaffold that facilitates and regulates the known to be involved in the regulation of food intensity and duration of Raf/MEK/ERK signa- intake and body weight35. A de novo heterozy- ling42. KSR2 also promotes activation, by di- gous missense mutation (c.2165A > G; p.Tyr- rect interaction, of the primary regulator of cell 722Cys) in the neurotrophic tyrosine kinase re- energy homeostasis, 5′-adenosine monophospha- ceptor type 2 (NTRK2) gene was found in a boy te-activated protein kinase (AMPK)43,44. The in- with early onset obesity, hyperphagia, develop- teraction between KSR2 and AMPK is suggested mental delay, impaired short-term memory, and to be involved in obesity, high insulin levels and impaired nociception. Further analysis showed impaired glucose tolerance observed in Ksr2 KO an impairment in BDNF-stimulated protein ki- mice45. Under conditions of nutrient deprivation nase phosphorylation36. and cell stress, intracellular ATP levels fall and

1361 S. Paolacci, A. Borrelli, L. Stuppia, F.C. Campanile, T. Dallavilla, et al. levels of AMP rise, promoting AMPK activa- receptors. Furthermore, NR0B2 has the ability to tion that in turn promotes catabolic processes modulate the transcriptional activity of the nucle- and inhibits anabolic pathways46. Many of the ar receptor 4-alpha (HNF4A) involved in maturi- variants studied impaired signaling through the ty onset diabetes of the young (MODY). In 2001, Raf-MEK-ERK pathway, while some reduced the NR0B2 was screened in 173 Japanese patients interaction between KSR2 and AMPK, compared diagnosed with MODY. The authors reported to wild-type KSR247. In 2013, Pearce et al47 se- that seven heterozygous mutations (p.Arg57Trp, quenced the coding exons of KSR2 in 1770 Eu- p.Gly189Glu, p.His53Alafs, p.Leu98_Ala101de- ropeans with severe, early-onset obesity (age of linsPro, p.Arg34*, p.Ala195Ser, p.Arg213Cys), onset < 10 years) to clarify whether genetic va- found in 12 subjects who were mildly or mode- riants in KSR2 contribute to obesity. They also rately obese at the onset of diabetes, were asso- retrospectively analyzed data from whole-exo- ciated with obesity rather than with diabetes. All me sequencing of a further 331 unrelated indivi- were truncating or loss-of-function mutations48. duals belonging to a cohort patients with severe, Although 7% of Japanese obese and diabetic pa- early-onset obesity. They identified 27 different tients showed NR0B2 mutations, this association rare variants in 45 of the 2101 unrelated seve- seems absent in the UK and very rare in Danish rely obese individuals screened. Most variants subjects49,50. Studies in mice suggest that NR0B2 were found in heterozygous form, although one may also be linked to the hypothalamic-pituitary severely obese subject was homozygous for two axis, important in the regulation of appetite and variants (p.Arg253Trp and p.Asp323Glu). Twen- energy expenditure51. ty-three variants were only identified in severely obese individuals; many were predicted to be fun- Obesity Due to Mutations in PPARG ctionally deleterious and altered highly conserved PPARG encodes a member of the peroxisome residues47. Nine of them were truncating variants. proliferator-activated receptor (PPAR) subfamily of nuclear receptors. PPARs form heterodimers Obesity Due to Mutations in ADCY3 with retinoid X receptors (RXRs). Three subtypes ADCY3 encodes adenylyl cyclase 3 which is of PPARs are known: PPAR-alpha, PPAR-delta, a membrane-associated enzyme that catalyzes and PPAR-gamma. The protein encoded by this the formation of the secondary messenger cyclic gene is PPAR-gamma, a regulator of adipocyte adenosine monophosphate (cAMP). This protein differentiation. PPAR-gamma has been implica- is widely expressed in various human tissues, ted in the pathology of diseases such as obesity showing high levels in subcutaneous and visceral and diabetes. fat cells. The cAMP is an essential second mes- In 1998, Ristow et al53 studied 121 obese senger in intracellular signaling of key metabolic subjects and identified the p.Pro115Gln mutation factors such as glucagon-like peptide 1, ghrelin in exon 6 of PPARG in four of them. The four and α-melanocyte stimulating hormone. cAMP German patients with the mutant allele had seve- signaling has been linked to control of adipose re obesity, their body mass indexes ranging from tissue development and function, as well as insu- 37.9 to 47.352. Significantly, the mutation was in lin secretion in beta cells. Leptin resistance may the codon immediately adjacent a Ser114 pho- possibly occur through disrupted cAMP signaling sphorylation site that down-regulates the tran- in primary cilia in the hypothalamus, affecting scriptional activity of the protein53. Over-expres- downstream signaling and neuron morphology46. sion of the mutant gene in murine fibroblasts led Two papers published in 2018 link homozygous to the production of a protein with defective pho- loss-of-function mutations to onset of obesity and sphorylation of Ser114, accelerated differentiation type 2 diabetes46,47. The studies were performed into adipocytes and greater cell accumulation of in a small inbred population (Greenlandic) and in triglycerides than for the wild-type53. an inbred Pakistani family, respectively46,47. Obesity Due to Mutations in DYRK1B Obesity Due to Mutations in NR0B2 DYRK1B encodes a member of a family of nu- NR0B2 belongs to the nuclear hormone recep- clear-localized protein kinases and participates in tor family and contains a putative ligand-binding cell cycle regulation. DYRK1B inhibits the SHH domain, while lacking a conventional DNA-bin- and WNT1 pathways, thereby enhancing adipo- ding domain. The protein has been shown to inte- genesis. In affected members of three Iranian fa- ract with retinoid, thyroid hormone, and estrogen milies with metabolic syndrome and early-onset

1362 Mendelian obesity, molecular pathways and pharmacological therapies: a review coronary artery disease, Keramati et al54 identi- discussed in the following sub-section, whereas fied a heterozygous missense mutation in DYR- other drugs will only be enlisted in Table I. K1B (p.Arg102Cys) that segregated with disease Liraglutide is a glucagon-like peptide 1 (GLP1) in all three families. Functional characterization agonist. It modulates AMPK, a protein expressed showed that the p.Arg102Cys allele has a gain-of- in the ventromedial hypothalamus and involved function effect. To further confirm the involve- in activation of thermogenic activity in beige adi- ment of DYRK1B in obesity, analysis of the gene pocytes. While liraglutide increases AMPK pho- in 300 morbidly obese Caucasian individuals sphorylation in various tissues (endothelium, he- with coronary artery disease and multiple meta- art, liver, muscle and white fat), it reduces AMPK bolic phenotypes identified another heterozygous phosphorylation in pancreatic beta cells and the missense mutation (p.His90Pro) in five unrelated hypothalamus (Figure 1). The main effect of li- patients54,55. raglutide is a decrease in inflammation, impro- vement of insulin sensitivity, induction of β-cell proliferation and an increase in beige adipocyte Discussion thermogenic capacity58. Since 2009, liraglutide has been used to treat type 2 diabetes, and sub- Obesity is generally a multifactor disorder as- sequently obesity. Its mechanism of action is ba- sociated with the Western lifestyle and linked to sed on glucose-dependent stimulation of insulin genetic and environmental factors. However, in a secretion by pancreatic β-cells, inhibition of glu- small percentage of cases (~5%)3, obesity is clear- cagon secretion by α-cells under normoglycemic ly associated with specific mutations in specific conditions, slowing of gastric emptying, and ap- genes. 5% might seem a rather small percentage petite suppression. Liraglutide results in weight but if one considers that million people are obese, loss in more than 5% of patients with obesity. In the absolute number might be quite high. These obese non-diabetics, the therapeutic effects are forms of obesity are defined as Mendelian obesity significant compared to placebo (5.9% weight re- or monogenic obesity. Many studies have already duction in obese diabetics and 8.0% in obese and elucidated the pathways deregulated in patients overweight non-diabetics compared to about 2% with monogenic obesity. In most cases, it is the in placebo-treated patients). Liraglutide also redu- leptin/melanocortin hypothalamic pathway that ces the probability of developing type 2 diabetes regulates food intake and energy expenditure. Pa- and has a positive effect on blood pressure and thways that lead to adipocyte differentiation have lipid profile when used in the treatment of obesity also recently been found deregulated in Mende- (Table I)59. lian obesity. Considering the role played by ge- Orlistat is a gastric and pancreatic lipase inhi- netics in obesity, we developed genetic tests for bitor that reduces dietary fat absorption from the the diagnosis of Mendelian obesity and for the gastrointestinal tract by around 30%60. Approved identification of genes encoding elements of the- by the FDA in 1999, it is indicated for obesity se pathways. We also analyze genes involved in management, including weight loss and weight candidate pathways, studied in mice, or for whi- maintenance when used in conjunction with a ch mutations have been identified in single cases low-calorie diet, and to reduce the risk of wei- without additional information about pathogenici- ght regain after weight loss. Patients on orlistat ty56,57. As the understanding of the genetic, beha- are prescribed a balanced, low-calorie diet with vioral and biochemical basis of obesity increases, approximately 30% of calories from fat. They more and more drugs are being tested to treat this should also take a multivitamin dietary supple- disorder. We summarized the molecular bases of ment during orlistat therapy, as the drug may de- Mendelian obesity and searched for clinical trials, crease absorption of fat-soluble vitamins (A, D, completed or underway, of drugs to treat severe E, K). In a double-blind prospective study that forms of obesity. Some of these drugs, such as li- randomized 3305 patients with a BMI ≥ 30 kg/m2 raglutide and topiramate activate the thermogenic with orlistat or placebo, mean weight loss was si- activity of beige adipocytes, inducing the catabo- gnificantly greater with orlistat (5.8 kg) than with lism of the stored fat; others reduce fat absorption placebo (3 kg) after 4 years; 53% of the patients such as the lipase inhibitor orlistat; other drugs assigned orlistat lost ≥ 5%, and 26.2% lost ≥10% target hypothalamic neurons in order to redu- of their initial body weight. Besides promoting ce food consumption or increase energy expen- weight loss, orlistat lowers serum levels of glu- diture. Some of these drugs will be extensively cose and improves insulin sensitivity. However,

1363 S. Paolacci, A. Borrelli, L. Stuppia, F.C. Campanile, T. Dallavilla, et al. orlistat is not commonly used for obesity manage- the management of obesity for patients with an ment because of side effects such as oily stool and initial body mass index of ≥ 30 kg/m2 or ≥ 27 kg/ fecal incontinence (Table I)61. m2 and other risk factors (hypertension, diabetes, Lorcaserin is a 5-hydroxytryptamine (5-HT; hyperlipidemia). It generally appears to be rela- serotonin) 2C receptor agonist. Its precise mecha- tively well-tolerated65. The exact mechanism of nism of action is not known but it is believed to phentermine is not known but it is known to sti- reduce food consumption and promote satiety by mulate neurons to release or maintain high levels selectively activating 5-HT2C receptors located of catecholamines, such as dopamine and nore- on anorexigenic pro-opiomelanocortin neurons in pinephrine. High levels of these catecholamines the hypothalamus. Available data suggests it does tend to suppress hunger signals and appetite. The not alter energy expenditure. The 5-hydroxytryp- drug seems to inhibit reuptake of noradrenaline, tamine (serotonin) system is a fundamental com- dopamine, and serotonin through inhibition of ponent of brain control of energy homeostasis. reuptake transporters, and to inhibit monoamine The neurotransmitter serotonin has a well-defined oxidase enzymes leaving more neurotransmitter role in eating behavior and is associated with de- available at synapses. Through catecholamine creased food intake, increased satiety, and appe- elevation, phentermine may also indirectly raise tite suppression. Pharmacological agents that se- leptin levels in the brain (Figure 1). Increased le- lectively target the 5-HT2C receptor subtypes that vels of catecholamines are theoretically respon- mediate the anorectic effects of 5-HT offer a novel sible for halting neuropeptide Y, that initiates ea- approach to improving weight-loss intervention ting, decreases energy expenditure, and increases programs. 5-HT2C receptors are located in the fat storage66. choroid plexus, limbic structures, extrapyrami- Topiramate was approved for seizure treat- dal pathways, thalamus, and hypothalamus. The ment in 1996 and migraine prophylaxis in 2004. key mechanism of 5HT2C modulation of body Its weight-loss properties were initially repor- weight occurs in the brain melanocortin circuit. ted in patients with seizure, mood, binge eating, About 40% of hypothalamic pro-opiomelanocor- and borderline personality disorders. Topirama- tin neurons (POMC) express 5HT2C receptors. te treatment leads to an average weight loss of POMC activation leads to the release of α-MSH, 5.3 kg67. It blocks voltage-gated sodium chan- which binds to melanocortin 4 receptors leading nels, reduces L-type calcium currents, increases to a reduction in appetite and increased energy potassium conductance, antagonizes α-amin- expenditure (Figure 1). In addition, pharmaco- o-3-hydroxy-5-methyl-4-isoxazolepropionic acid logical activation of 5-HT2C receptors improves (AMPA) receptor/kainate glutamate receptors glycemic control, whereas their inactivation leads and enhances γ-aminobutyric-acid-mediated to weight gain and insulin resistance. Lorcaserin chloride channels68. Studies in mice have provi- is indicated as an adjunct to diet and exercise for ded insights into its mechanisms of action in the chronic weight management, including weight hypothalamus. The administration of topiramate loss and maintenance, in patients with obesity to mice fed a high-fat diet reduced food intake and [BMI ≥ 30 kg/m2], or in patients with overweight increased energy expenditure, leading to dimini- (BMI ≥ 27 kg/m2) and at least one weight-related shed adiposity and increased insulin and leptin comorbid condition (hypertension, dyslipidemia signaling in the hypothalamus. The signaling of or type 2 diabetes). The safety and tolerability insulin and leptin in the hypothalamus is a key profile of lorcaserin has been documented in three to inducing anorexia and weight loss. Treatment Phase III studies (NCT00603902, NCT00395135, with topiramate also reduces hypothalamic levels NCT00603291) (Table I)62,63. of negative modulators of insulin signaling, such Phentermine, initially approved for obesity tre- as the phosphatases LAR, PTP1B, and PHLPP1, atment in 1959, is a derivative of amphetamine suggesting that their reduction may have a role in but has less abuse potential than the latter. It is improving the action of insulin. Topiramate-trea- estimated that phentermine can lead to an average ted obese mice also showed activation of the lep- weight loss of 3.6 kg in obese patients64. Its an- tin-induced JAK2/STAT3 pathway, important for tiobesity effect could be due to its ability to rele- food intake reduction and weight loss (Figure 1). ase norepinephrine in the brain. Phentermine is This pathway is necessary to enhance the down- currently approved as a short-term adjunct in a stream melanocortin pathway, which contributes regimen of weight reduction based on exercise, to hypophagia and increased energy expenditure. behavioral modification, and calorie restriction in The MAPK/ERK pathway is regulated by leptin

1364 Mendelian obesity, molecular pathways and pharmacological therapies: a review and insulin in the hypothalamus. Activation of the dopaminergic tone in the hypothalamus is as- SHP2/ERK pathway, induced by leptin, mediates sociated with obesity. The hypothalamic mela- thermogenesis and energy expenditure: topira- nocortin system is, therefore, a potential site of mate may interfere with energy use efficiency, action of bupropion. Indeed, bupropion stimulates enhancing thermogenesis. Topiramate treatment the activity of POMC cells in vitro and increases also increases UCP-1 and PGC-1α levels in brown α-MSH secretion. It reduces short-term food in- adipocytes of obese mice. In addition, it can lead take and increases energy expenditure by incre- to increased phosphorylation of AMPK in skele- asing heat production, although the overall effect tal muscle, heart, adipose tissue and liver, causing of bupropion on body weight in mice is modest. enhanced glucose uptake and fatty acid oxida- In humans, weight loss is a common side effect of tion69. bupropion use for the treatment of depression. In Although phentermine and topiramate, taken overweight and obese adults, bupropion treatment singly, have body weight reduction effects, their leads to limited weight loss72. combination is more effective. Comparison of the Combining naltrexone and bupropion was pro- effects of single and combined use is the subject posed on the basis of in vitro studies in the mouse of a study that showed that combined phentermi- hypothalamus, and was linked to melanocortin ne/topiramate led to a ~11% reduction in baseline and reward pathways. The effect of bupropion in weight, compared with ~2% for placebo, ~5% for increasing POMC activity is limited by µ-opioid phentermine and ~6% for topiramate. Patients receptors, which mediate autoinhibition of POMC who received high doses of phentermine/topira- cells by β-endorphin, resulting in the modest wei- mate achieved an average weight loss of ~9%, ght loss and reduction of calorie intake of bupro- compared with ~1.5% with placebo, ~6% with pion monotherapy. On the other hand, blockade phentermine, ~6% with topiramate. This demon- of the µ-opioid receptor with naltrexone alone strates additive weight loss when the two drugs gradually increases POMC activity. The simulta- are combined (Table I)70,71. neous administration of bupropion and naltrexone Naltrexone is an opioid antagonist with high produces a large increase in POMC activity (Fi- affinity for the µ-opioid receptor, implicated in gure 1): the combination stimulates POMC cells eating behavior. Its activity may influence food (bupropion) and removes β-endorphin inhibition intake and body weight via the hypothalamic me- on POMC cells (naltrexone). At the same time, lanocortin and reward systems containing opioid injection of bupropion or naltrexone alone direct- neurons. Indeed, studies in animals indicate that ly into the reward system is sufficient to reduce acute naltrexone administration influences the food intake in hungry mice, while direct injection activity of the reward system and eating beha- of naltrexone and bupropion produces a synergi- vior. Systemic naltrexone prevents the increase stic reduction in food intake. Initial Phase 2 cli- in dopamine in the nucleus accumbens caused nical studies comparing the naltrexone/bupropion by food ingestion and reduces food intake, food (NB) combination with naltrexone or bupropion seeking, binge-like eating and preference for hi- monotherapy for weight loss in obese subjects gh-calorie foods. Human studies demonstrate that demonstrated that NB produces synergistic wei- naltrexone reduces the subjective pleasantness of ght loss. In Phase 3 studies, NB-treated subjects palatable foods, in line with the role of opioids in showed more than twice the weight loss of tho- the rewarding aspects of eating. Despite the in- se in the bupropion monotherapy group and NB dications of these preliminary studies, naltrexone therapy was associated with corresponding re- monotherapy-mediated blockade of opioid neu- ductions in visceral and total body fat (Table I)72. rotransmission is insufficient to produce reliable Exenatide is a GLP-1 agonist and was appro- decreases in food intake in humans72. ved for the treatment of type 2 diabetes mellitus Bupropion is an antidepressant that inhibi- (T2DM) in 2005. It is a synthetic exendin-4 (a ts reuptake of catecholamines, such as dopami- hormone found in the saliva of the lizard Helo- ne and norepinephrine. In mice, acute treatment derma suspectum) that is resistant to peptidase with bupropion produces changes in extracellular degradation, but has a short half-life due to rapid dopamine and norepinephrine concentrations in clearance by glomerular filtration. The short- the brain and alters the activity of dopamine- and acting compounds suppress postprandial gluco- norepinephrine-releasing neurons. The activi- se elevation by inhibiting gastric emptying73. In ty of the melanocortin system is influenced by studies of patients with T2DM, exenatide treat- both dopamine and norepinephrine, and reduced ment was usually associated with reductions in

1365 S. Paolacci, A. Borrelli, L. Stuppia, F.C. Campanile, T. Dallavilla, et al.

Figure 1. Graphical representation of the leptin/melanocortin pathway in the hypothalamus, with the main molecular mem- bers of this signaling pathway. Drugs that may be beneficial for the treatment of obesity are indicated.

body weight of about 2-3 kg. This was also seen 3T3-L1 adipocytes (a cell line derived from mu- in obese non-diabetics with normal or impaired rine 3T3 cells that is used in research on adipose glucose tolerance or impaired fasting glucose tissue) treated with gymnemic acids conjugated and BMI 39.6 ± 7 kg/m2, who showed a progres- with gold nanoparticles (GA-AuNPs) showed sive reduction in body weight over 24 weeks and enhancement in glucose uptake efficiency by improved glucose tolerance. In a small, dou- translocating GLUT4 glucose transport vesicles ble-blind, placebo-controlled crossover study, 41 from the cytosol to the plasma membrane. These obese (BMI 33.1 ± 4.1 kg/m2) female non-diabe- findings suggest that GA-AuNPs may act as sup- tics, treated with exenatide for 16 weeks, showed pressors of mitochondrial ATP synthase in the a significant weight loss of 2.49 ± 0.66 kg. Other presence of insulin or an inducer of AMPK (an clinical trials are testing the effect of exenatide activator of fatty acid oxidation) in the absence of on obese patients74. Exenatide was also tested in insulin78,79. Although clinical approval and scien- patients with type 1 diabetes: a crossover study tific validation are necessary before they can be with high doses of exenatide was performed for approved for the treatment of diabetes, several 6 months in 13 adults who showed a significant studies have reported anti-diabetic effects, sugar average weight loss of 4.2 kg (Table I)73. inactivation properties and anti-obesity action of Gymnema sylvestre (GS) is a tropical plant gymnemic acids in pre-clinical studies (cell and of the Asclepiadaceae family. It is well known animal models) and clinical studies75,77,80-85. A for its antihypercholesterolemic, anti-inflamma- recent interventional, randomized, double-blind tory, antimicrobial, antidiabetic and anti-obesity clinical study (NCT02370121) reported a statisti- effects. The leaves are known to contain gym- cally significant reduction in body weight, redu- nemic acids, stigmasterol, quercitol and ami- ced BMI and lower values for very-low-density no acid derivatives of betaine, ethylamine, and lipoprotein (VLDL) (Table I)86, thereby justi- choline. Major active compounds are gymnemic fying the interest of the scientific community in acids found in all parts of the plant75. Gymnemic this promising molecule. acids are molecules similar to that of glucose that bind the taste buds receptors of the tongue, temporarily destroying the taste of sweetness76. Conclusions They also suppress absorption of glucose from the intestine into the blood by binding Na+-glu- We found that most Mendelian forms of obesi- cose symporters in the intestine77. Furthermore, ty are linked to dysfunctions in the leptin/mela-

1366 Mendelian obesity, molecular pathways and pharmacological therapies: a review Table continued Table Sponsor/Collaborators

Madison University Wisconsin, of Development, LLC Janssen Research & Development, L.L.C. Pharmaceutical Research & Johnson & Johnson Yale University Yale University Hospital Chonbuk National University Hospital Chonbuk National Diabetes Association Hospital, American Massachusetts General

Results

in fat mass Significant reduction

canagliflozin phentermine plus phentermine and body weight with Significant reduction in of theof drug correlated with dose reductions are positively body mass index. Both in body weight and Significant reduction No significantNo results

No significantNo results weight, body mass index total visceral fat, body in body fat mass, Significant reduction cholesterol lipoprotein (LDL) in density low Significant reduction

Aim oxidation total of fat consumption increases determine CLA whether compared to control; to results in loss body of fat of conjugated linoleic acid (CLA) (CLA) conjugated acid linoleic of consumption a purified of form To determineTo whether obese non-diabeticsobese of overweightof and placebo in the treatment phentermine with canagliflozin and administration of To test the effectsTo co- of non-diabetics overweight and obese promoting weight loss in (JNJ-28431754) in (JNJ-28431754) effectiveness canagliflozin of To assess theTo safety and women with binge eating disorder bupropion on overweight hypercholesterolemic subjects test the efficacyTo of cholesterol in mildly serum and LDL total reduce visceral fat, β-glucan supplements To investigateTo whether barley

Korean subjects of actiponinof in obese To test the efficacyTo and safety order to reduce free fatty acids can improve mitochondria in To examineTo whether acipimox of Number participants

24 (drug); 24 (placebo)

phentermine mg) 15 84 (canagliflozin300 mg/ 84 (canagliflozin300 mg), 85 (phentermine85 mg), 15 82 (placebo),82

89 (placebo)89 96 (canagliflozin300 mg), 93 (canagliflozin93 100 mg), 98 (canagliflozin98 50 mg),

(drug); (placebo) 30 31

40 (drug); 40 (placebo)

40 (drug); 40 (placebo) 20 (drug); 19 (placebo)20 (drug); 19

Condition (age of treated patients in years)

Obesity (18-44)

Obesity (18-65)

Obesity (18-65) obesity (18-65) disorder and Binge eating

(19-70) hyperlipidemia Overweight and

Obesity (19-65) hypertriglyceridemia insulin resistance and Abdominal obesity with ID NCT (phase)

(2) NCT00204932

(2) NCT02243202

(2) NCT00650806

and(2 3) NCT00414167

and(2 3) NCT01402128

(2) NCT01667224

(2) NCT01488409

List of active (not recruiting) active List of (not and completed interventional clinical trials using drugs or dietary supplements the for treatment obesity of (retrieved from ClinicalTrials.gov). metabolism)

(fat oxidation) (fat conjugated Linoleic acid, pathway) dopamine/leptin lumen and glucose from tubule (re-absorption of Phentermine Canagliflozin +

tubule lumen) glucose from renal (re-absorption of Canagliflozin pathway) (melanocortin Bupropion

supplement dietary Barley β-glucan,

(AMPK pathway) supplement Actiponin, dietary (18-55)

activity) Drug (target pathway/ (target Acipimox (lipase 1367 Table I. Table S. Paolacci, A. Borrelli, L. Stuppia, F.C. Campanile, T. Dallavilla, et al. 1368 Table continued Table Sponsor/Collaborators University Guadalajara of (NHLBI) Lung, and Blood Institute Hospital, National Heart, Massachusetts General Hanyang University Company AstraZeneca, Eli Lilly and Digestive and Kidney Diseases (NIDDK) Institute Diabetes of and Mayo Clinic,Mayo National Minnesota Hospitals and Clinics of Science Institute, Children’s Clinical and Translational University Minnesota of - Science Institute Clinical and Translational University Minnesota of - Clinical Center (CC) National Institute Health, of Kidney Diseases (NIDDK), Diabetes and Digestive and National Institute of

Results

analysis provided significantNo results tolerance; no statistical and increase in glucose fatty tissue, triglycerides Reduction in visceral No significantNo results body weight, BMI Significant decrease in sample size too small emptying time but Reduction in gastric

sample size too small Decrease in BMI but

sample size too small Decrease in BMI but expenditure increase in energy body weight, and in energy intake, Significant decrease

Aim Gymnema sylvestre on metabolic syndrome and insulin resistance To evaluate theTo effect of in obesity cardiovascular function abdominal fat and improving synthetic GHRH in decreasing safety Glycin of max (L.) Merr. peel extract on decrement body of fat evaluate theTo effect of in non-diabetics obese evaluate theTo efficacy and exenatide on body weight To test the effectTo of satiation and weight loss in obese participants gastric emptying, satiety, exenatide vs placebo on To evaluate theTo effect of

extreme obesity exenatide in pediatric weight loss test the effectTo of glucose tolerance, on glucose disposal, GLP-1 agonist GLP-1 (exenatide) To test the effectTo a of

in obese non-diabetics leads to weight loss exenatide treatment To evaluate whetherTo of Number participants

12 (drug); 12 (placebo) (drug); 12 12

(drug); (placebo) 29 31 40 (drug); 40 (placebo)

80 (drug); (placebo) 83

10 (drug); 10 (placebo) (drug); 10 10

(placebo) (drug); 13 13

11 (drug); 11 (placebo) (drug); 11 11

41 (drug); 39 (placebo) (drug); 39 41 Condition (age of treated (30-60) Metabolic syndrome

deficiency(18-55) and growth hormone Abdominal obesity Obesity (19-65)

Obesity 18) (>

Obesity (18-70)

Obesity (12-19)

Obesity (8-19)

Obesity (18-55) patients in years) ID NCT (phase) (2) NCT02370121

(2) NCT00675506 and(2 3) NCT02108691 (2) NCT00500370

(4) NCT02160990

(2) NCT01237197

(2) NCT00886626

(3) NCT00856609 List of active (not recruiting) active List of (not and completed interventional clinical trials using drugs or dietary supplements the for treatment obesity of (retrieved from ClinicalTrials.gov). .

metabolism) (glucose metabolism, insulin pathway) Gymnema sylvestre

pathway) (growth hormone releasing hormone dietary supplement Growth hormone Merr. peel extract, maxGlycin (L.)

metabolism) Drug (target pathway/ (target Exenatide (glucose Table I (Continued) Table Mendelian obesity, molecular pathways and pharmacological therapies: a review Table continued Table Novo NordiskNovo A/S Clinic,Mayo National Institute Diabetes of and Digestive and Kidney In Qpharm Group NordiskNovo A/S Novo NordiskNovo A/S Sponsor/Collaborators

Results Significant reduction in body weight, glycosylated hemoglobin, waist circumference Significant reduction in body weight and increase in gastric Reduction in body weight, body fat, waist circumference, but no statistical analysis provided Significant reduction in body weight and number patientsof with type 2 diabetes Diseases (NIDDK), Novo Nordisk A/S Decrease in body weight only with highest dose liraglutide;of

Aim

emptying time but without statistical analysis no statistical analysis

To test the effectTo liraglutide of on body weight in overweight/ on weight loss and gastric unctions in obesity obese subjects with type 2 diabetes test the effectTo liraglutide of To evaluate theTo safety and in efficacyIQP-VV-102 of on body weight in reducing body weight in overweight and obese subjects test the effectTo liraglutide of obese/overweight non-diabetics with

on body weight in obese To test the effectTo liraglutide of subjects without diabetes co-morbidities of Number participants 757 (placebo) 757 423 (liraglutide423 3 mg), (liraglutide mg), 211 1.8 212 (placebo) 212 (placebo) (drug); 21 19 60 (drug); 60 (placebo) no pre-diabetes); (liraglutide959 3 mg, (placebo);487 (liraglutide1528 3 mg, pre-diabetes);

2.4 mg/liraglutide 98 (placebo/liraglutide98 (liraglutide 95 3 mg), mg/liraglutide1.2 3 mg), 90 (liraglutide mg/ 1.8 liraglutide 93 3 mg), (liraglutide 2.4 mg/ liraglutide 3 mg), (liraglutide93 3 mg), (orlistat) 95 Condition (age of treated patients in years)

Obesity 18) (> Obesity (18-65) Obesity or overweight (18-60) Obesity 18) (>

Obesity (18-65) ID NCT (phase) List of active (not recruiting) active List of (not and completed interventional clinical trials using drugs or dietary supplements the for treatment obesity of (retrieved from .

NCT01272232 (3) (2) NCT02647944 NCT01681069 (3) (3) NCT01272219

(2) NCT00422058

metabolism)

Drug (target pathway/ (target IQP-VV-102, dietary supplement pathway) Liraglutide (AMPK

orlistat (AMPK Liraglutide and pathway and lipase activity) 1369 Table I (Continued) Table ClinicalTrials.gov). S. Paolacci, A. Borrelli, L. Stuppia, F.C. Campanile, T. Dallavilla, et al. 1370 Table continued Table Columbia University Arena Pharmaceuticals Arena Pharmaceuticals Arena Pharmaceuticals University Colombo of Sponsor/Collaborators

Results No significantNo results

Significant reduction in body weight Significant reduction in body weight Significant reduction in body weight Reduction in body weight, BMI, insulin resistance; no statistical analysis provided

Aim To determineTo whether oral leucine supplementation in overweight/obese individuals increases basal metabolic rate, reduces body weight, improves glucose tolerance and/or insulin sensitivity, and/or reduces circulatingreduces LDL-cholesterol levels To test the effectsTo lorcaserin of hydrochloride on overweight efficacylorcaserinof or obese volunteers assess theTo safety and hydrochloride in obese patients effect lorcaserin of in To assess theTo weight loss overweight and obese patients with II Type diabetes mellitus treated with metformin, sulfonylurea or both with other oral hypoglycemic agents. To evaluate theTo effect of metformin on changes in insulin resistance, fatty liver status, body fat content, BMI and other metabolic markers of Number participants 8 (drug); 8 (control)

802 (lorcaserin mg); 10 (lorcaserin1603 mg); 10 lorcaserin mg); 10 1603 (placebo) 1603 (year 1, 1595 (year placebo); 1, 1587 573 (year573 2, lorcaserin 256 once a day); 10 mg (year10 1)/lorcaserin mg (year 283 10 2) (year 2, lorcaserin m 10 (year (year 1)/placebo 2); (year697 2, placebo) (lorcaserin95 mg, 10 (lorcaserin mg, twice 10 a day); 253 (placebo) 253 a day);

(control) (drug); 173 166

Condition (age of treated patients in years) Overweight or obesity (18-65)

Obesity (18-65) Obesity (18-65)

Obesity (18-65)

Obesity (8-16)

ID NCT (phase) List of active (not recruiting) active List of (not and completed interventional clinical trials using drugs or dietary supplements the for treatment obesity of (retrieved from . NCT00683826 (NA)

NCT00603902 (3) (3) NCT00395135 (3) NCT00603291

NCT02274948 (4)

metabolism) (target pathway/ (target L-leucine, dietary supplement Drug

Lorcaserin (serotonin/ melanocortin pathway)

Metformin (AMPK pathway)

Table I (Continued) Table ClinicalTrials.gov). Mendelian obesity, molecular pathways and pharmacological therapies: a review Table continued Table Sponsor/Collaborators Orexigen Therapeutics Inc. Orexigen Therapeutics Inc. Orexigen Therapeutics Inc. Yale University Yale Orexigen Therapeutics Inc. GlaxoSmithKline

Results Significant decrease in body weight, waist circumference, triglycerides and fasting glucose, increase in HDL No significantNo results Significant decrease in body weight, waist circumference and triglycerides, increase in HDL

No significantNo results Significant reduction in body weight, waist circumference, triglycerides, glucose, triglycerides, fasting insulin, insulin resistance and binge eating, increase in HDL visceral fat mass, Significant reduction in body weight, total fat mass

Aim To test the safetyTo and efficacy of theof combination of naltrexone and SR bupropion compared SR to placebo in obese subjects and in overweight/obese subjects with hypertension and/or dyslipidemia assess theTo effects of naltrexone / bupropion SR SR on overweight or obese subjects To test the safetyTo and efficacy of theof combination of naltrexone and SR bupropion SR compared to placebo in obese subjects and in overweight/obese subjects with hypertension hypertension with subjects and/or dyslipidemia To test the effectTo the of combination naltrexone of and bupropion relative to placebo on binge eating in persons with obesity and binge eating disorder To assess theTo effects of combination therapy with naltrexone SR/bupropion SR SR SR/bupropion naltrexone (NB) and comprehensive comprehensive and (NB) lifestyle intervention (CLI) on body weight and cardiovascular risk factors compared to the effects usual of care in overweight or obese dyslipidemic subjects with or without controlled hypertension orlistat affects visceral adipose To determineTo if weight loss with tissue compared to placebo of Number participants 578 (naltrexone 16 mg/ (naltrexone 16 578 bupropion mg 360 /day); 583 (naltrexone mg/583 32 bupropion 360 mg /day); bupropion mg 360 /day); 581 (placebo) 581

(drug);23 (placebo) 23

1001 (drug); 495 (placebo) (drug); 495 1001

12 (drug); 10 (placebo) (drug); 10 12

(drug); (control) 89 153

65 (drug); 6665 (placebo)

(age of treated patients in years) Obesity or overweight (18-65)

Obesity (18-45)

Obesity or overweight (18-65)

Binge-eating disorder (21-65)

(18-60) Obesity or overweight

Obesity or overweight

ID NCT Condition (phase) List of active (not recruiting) active List of (not and completed interventional clinical trials using drugs or dietary supplements the for treatment obesity of (retrieved from . NCT00532779 (3)

(2) NCT00711477

(3) NCT00567255

NCT02317744 (NA)

(3) NCT01764386

(4) NCT00752726

(18-60)

Drug metabolism) (target pathway/ (target Naltrexone +

Bupropion (opioid Bupropion (opioid and melanocortin pathway)

Orlistat

(lipase activity) 1371 Table I (Continued) Table ClinicalTrials.gov). S. Paolacci, A. Borrelli, L. Stuppia, F.C. Campanile, T. Dallavilla, et al. 1372 Table continued Table VIVUS Inc., Medpace Inc . VIVUS Inc., Medpace Inc. VIVUS Inc., Medpace Inc. VIVUS Inc., Medpace Inc. Mayo Clinic,Mayo National Institute Diabetes of and Digestive and Kidney Diseases (NIDDK) New York State York New Institute, AstraZeneca Sponsor/Collaborators

Results Significant reduction in body weight Significant reduction in body weight Significant reduction in body weight Significant reduction in body weight significantNo results due to the number low of participants

No significantNo results

Aim To evaluate theTo efficacy and safety compared VI-0521 of to placebo in the treatment of obesity (BMI ≥35) To evaluate theTo long-term safety and efficacy of compared VI-0521 to placebo in the treatment of adult overweight and obesity evaluate theTo safety and efficacy of compared VI-0521 to placebo in the treatment of overweight and obese adults compare theTo effect VI-0521 of (topiramate+phetermine) or topiramate or phentermine alone with placebo treatment obesity of in adults determineTo the effect of phentermine and topiramate ER on gastric emptying, gastric accommodation, satiety and satiation in obesity To determineTo whether phentermine changes food intake compared to placebo administration of Number participants 241 (phentermine241 + topiramate, dose); low 512 (phentermine+512 topiramate, high dose); (placebo) 514 154 (phentermine154 + topiramate, dose); low 295 (phentermine 295 + topiramate, high dose); (placebo)227 (phentermine154 + topiramate, dose); low 295 (phentermine295 + topiramate, high dose); (placebo)227 mg phentermine); (7.5 108 107 (46 mg topiramate); (46 107 108 (7.5 mg/46 mg (7.5 108 phentermine/topiramate); mg phentermine); (15 107 mg topiramate (92 108 mg/9215 mg phentermine/ topiramate); (placebo) 109 (phentermine+12 topiramate); (placebo) 12

7 (drug); 6 (placebo)

(age of treated patients in years) Obesity (18-70)

Obesity and type 2 diabetes (18-70)

Obesity (18-70)

Obesity (18-70)

Obesity (18-70)

Obesity (18-60)

ID NCT Condition (phase) List of active (not recruiting) active List of (not and completed interventional clinical trials using drugs or dietary supplements the for treatment obesity of (retrieved from . (3) NCT00554216

(3) NCT00553787

(3) NCT00796367

(3) NCT00563368

(4) NCT01834404

(4) NCT01886937

Drug metabolism) Phentermine+ Topiramate (dopamine/leptin, insulin/leptin signaling)

(target pathway/ (target Phentermine Psychiatric (dopamine/leptin pathway)

Table I (Continued) Table ClinicalTrials.gov). Mendelian obesity, molecular pathways and pharmacological therapies: a review Table continued Table AstraZeneca AstraZeneca Massachusetts General Hospital, Genentech Inc. Göteborg University, Sahlgrenska University Hospital, Sweden Merck Sharp & Dohme Corp. University Minnesota, of Clinical and Translational Science Institute Sponsor/Collaborators

Results Significant reduction in body weight Significant reduction in body weight significantNo results due to the number low of participants statisticalNo significance provided No significantNo results No statisticalNo significance provided

Aim

To examineTo the safety, tolerability and effect on body weight of metreleptin and pramlintide in obese and overweight subjects To examineTo the long-term safety, tolerability and effect on body pramlintideweight of combination in administered with metreleptin test the effectTo recombinant of human growth hormone (rhGH) on reduction abdominal of fat and cardiovascular risk in obese girls investigateTo whether treatment with rosuvastatin reduces visceral fat in obese men To evaluate theTo effect SCH of on weight in497079 obese and overweight participants To evaluate theTo safety and efficacyof topiramate therapy with meal replacement therapy in adolescents with severe obesity of

Number participants 77 (pramlintide77 mcg); 360 72 (metreleptin72 5 mg); 75 (pramlintide75 mcg 180 + metreleptin 78 2.5 mg); (pramlintide mcg + 180 metreleptin 78 5 mg); (pramlintide mcg 360 + metreleptin mg); 1.25 (pramlintide78 mcg 360 + metreleptin 2.5 mg); (pramlintide75 mcg 360 + metreleptin 75 5 mg); (placebo) 242 (drug); 31 (placebo) (drug);242 31

(placebo) (drug); 11 11

(drug); (placebo)30 29

267 (drug); 134 (placebo) (drug); 134 267

16 (drug); 14 (placebo) (drug); 14 16

(age of treated patients in years) Overweight or obesity

Obesity (18-65)

Obesity (13-21)

Abdominal obesity (40-65)

Obesity or overweight (>18)

Obesity (12-17)

ID NCT Condition (phase) List of active (not recruiting) active List of (not and completed interventional clinical trials using drugs or dietary supplements the for treatment obesity of (retrieved from . (2) NCT00673387

(2) NCT00819234

(2) NCT01169103 (3) NCT01068626

NCT00642993 (2) (2) NCT01859013

Drug metabolism) (target pathway/ (target Pramlintide and metreleptin (calcitonin receptor pathway and leptin/ melanocortin pathway)

Recombinant human growth hormone (growth hormone pathway) Rosuvastatin (lipid synthesis)

SCH 497079 (Histamine H3 receptor pathway) Topiramate (insulin/ leptin signaling)

1373 Table I (Continued) Table ClinicalTrials.gov). S. Paolacci, A. Borrelli, L. Stuppia, F.C. Campanile, T. Dallavilla, et al. 1374 Heart and Diabetes Center North-Rhine Westfalia Duke University, National Institute Diabetes of and Digestive and Kidney Diseases (NIDDK) Aaron Cypess, National Institutes of Health (NIH), Health Institutes of National Institute of Diabetes and Digestive and Kidney Diseases (NIDDK), Beth Israel Deaconess Medical Center Sponsor/Collaborators

Results No statisticalNo significance provided

No statisticalNo significance provided

Significant increase BAT in activity

Aim To investigateTo the influence of vitaminof D supplement on weight loss and body composition in overweight patients To test the effectTo zonisamide of on weight reduction in obese adults

To test the hypothesisTo that human brown adipose tissue can be activated using a β3-adrenergic receptor agonist. The efficacy of β3-ARof agonist will be compared with exposure cold and a placebo control of Number participants 100 (drug); 100 (placebo) (drug); 100 100

76 (zonisamide76 200 mg; 75

12 (drug); 12 (placebo) (drug); 12 12

(age of treated patients in years) Overweight or obesity (18-70)

Obesity (18-65)

Obesity (18-65)

ID NCT Condition (phase) List of active (not recruiting) active List of (not and completed interventional clinical trials using drugs or dietary supplements the for treatment obesity of (retrieved from . NCT00493012 (NA)

NCT00275834 (NA)

(2) NCT01783470

Drug metabolism) (target pathway/ (target Vitamin dietary D, supplement

Zonisamide (GABAergic synaptic activity)

β3-adrenergic receptor agonist (β-adrenergic pathway)

Table I (Continued) Table ClinicalTrials.gov). Mendelian obesity, molecular pathways and pharmacological therapies: a review nocortin hypothalamic signaling pathway. Most 7) Morton GJ, Cummings DE, Baskin DG, Barsh GS, of the drugs being evaluated target this pathway Schwartz MW. Central nervous system control of with the aim of improving energy expenditure food intake and body weight. Nature 2006; 443: and/or reducing food intake. The results of the- 289-295. Tang QQ, Lane MD. se trials are promising in many cases, though in 8) Adipogenesis: from stem cell to adipocyte. Annu Rev Biochem 2012; 81: 715- others the sample has been too small to obtain 736. significant results. Of course, the possibility of 9) Lefterova M, Haakonsson A, Lazar M, Mandrup severe side effects should be weighed by clini- S. PPARγ and the global map of adipogenesis cians when prescribing these medications. and beyond. Trends Endocrinol Metab 2014; 25: 293-302. 10) Montague CT, Farooqi IS, Whitehead JP, Soos MA, Rau H, Wareham NJ, Sewter CP, Digby JE, Mohammed SN, Hurst JA, Cheetham CH, Earley AR, Barnett AH, Conflict of Interest Prins JB, O’Rahilly S. Congenital leptin deficiency The Authors declare that they have no conflict of interests. is associated with severe early-onset obesity in humans. Nature 1997; 387: 903-908. 11) Farooqi IS, Depaoli AM, Rahilly SO, Farooqi IS, Mata- Acknowledgements rese G, Lord GM, Keogh JM, Lawrence E, Agwu C, San- We would like to thank Helen Ampt for the English language na V, Jebb SA, Perna F, Fontana S, Lechler RI, Depaoli editing and all the laboratory staff of the MAGI group. AM, O’Rahilly S. Beneficial effects of leptin on obesi- ty, T cell hyporesponsiveness, and neuroendocrine/ metabolic dysfunction of human congenital leptin Contributorship Statement deficiency. J Clin Invest 2002; 110: 1093-1103. MB conceived the study; SP, TD, DV and MB collected in- 12) Gibson WT, Farooqi IS, Moreau M, DePaoli AM, formation; SP wrote the manuscript; SP, AB, LS, FCC, TD, Lawrence E, O’Rahilly S, Trussell RA. Congeni- JK, DV, TB, VU, MB reviewed and edited the text; MB su- tal leptin deficiency due to homozygosity for the pervised the work. Δ133G mutation: report of another case and eval- uation of response to four years of leptin therapy. J Clin Endocrinol Metab 2004; 89: 4821-4826. Funding 13) Strobel A, Issad T, Camoin L, Ozata M, Strosberg A. The authors did not receive any funding for this study. A leptin missense mutation associated with hypo- gonadism and morbid obesity. Nat Genet 1998; 18: 213-215. 14) Clément K, Vaisse C, Lahlou N, Cabrol S, Pelloux References V, Cassuto D, Gourmelen M, Dina C, Chambaz J, Lacorte JM, Basdevant A, Bougnères P, Lebouc Y, 1) Ali AT, Hochfeld WE, Myburgh R, Pepper MS. Adipo- Froguel P, Guy-Grand B. A mutation in the human cyte and adipogenesis. Eur J Cell Biol 2013; 92: leptin receptor gene causes obesity and pituitary 229-236. dysfunction. Nature 1998; 392: 398-401. 2) Rao KR, Lal N, Giridharan NV. Genetic and epi- 15) Farooqi IS, Wangensteen T, Collins S, Kimber W, genetic approach to human obesity. Indian J Med Matarese G, Keogh JM, Lank E, Bottomley B, Lo- Res 2014; 140: 589-603. pez-Fernandez J, Ferraz-Amaro I, Dattani MT, Ercan 3) Kleinendorst L, Massink MPG, Cooiman MI, Savas M, O, Myhre AG, Retterstol L, Stanhope R, Edge JA, van der Baan-Slootweg OH, Roelants RJ, Janssen McKenzie S, Lessan N, Ghodsi M, De Rosa V, Perna F, ICM, Meijers-Heijboer HJ, Knoers NVAM, Ploos van Fontana S, Barroso I, Undlien DE, O’Rahilly S. Clin- Amstel HK, van Rossum EFC, van den Akker ELT, van ical and molecular genetic spectrum of congeni- Haaften G, van der Zwaag B, van Haelst MM. Ge- tal deficiency of the leptin receptor. N Engl J Med netic obesity: next-generation sequencing results 2007; 356: 237-247. of 1230 patients with obesity. J Med Genet 2018; 16) Carroll K, Gomez C, Shapiro L. Tubby proteins: the 55: 578-586. plot thickens. Nat Rev Mol Cell Biol 2004; 5: 55-63. Yazdi FT, Clee SM, Meyre D 4) . Obesity genetics in 17) Kim J, Kim H, Kim S, Park J. Insulin phosphorylates mouse and human: back and forth, and back tyrosine residue 464 of Tub and translocates Tub- again. PeerJ 2015; 3: e856. by into the nucleus in HIRcB cells. Endocrinol Me- 5) Saleem F, Rizvi SW. New therapeutic approaches in tab (Seoul) 2014; 29: 163-168. obesity and metabolic syndrome associated with 18) Borman AD, Pearce LR, Mackay DS, Nagel-Wolfrum polycystic ovary syndrome. Cureus 2017; 9: 1-11 K, Davidson AE, Henderson R, Garg S, Waseem NH, 6) Oswal A, Yeo GSH. The leptin melanocortin path- Webster AR, Plagnol V, Wolfrum U, Farooqi IS, way and the control of body weight: lessons from Moore AT. A homozygous mutation in the TUB human and murine genetics. Obes Rev 2007; 8: gene associated with retinal dystrophy and obe- 293-306. sity. Hum Mutat 2014; 35: 289-293.

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