New Thoughts on Pediatric Genetic Obesity: Pathogenesis, Clinical Characteristics and Treatment Approach

Historically, some genetic syndromes and monogenic forms of obesity have been identified by clinical features and by sequencing candidate genes in patients with severe obesity. The phenotypic expression of genetic factors involved in obesity is variable, thereby allowing to distinguish several clinical pictures of obesity. Monogenic obesity is described as rare and severe early‐onset obesity with abnormal feeding behavior and endocrine disorders. Many of the findings emerged from studying families who displayed a classical Mendelian pattern of inheritance. On the contrary, patients with syndromic obesity show a various degree of intellectual disability, different dysmorphic features, and organ‐specific abnormalities. But to date, not all involved genes have been identified so far. New diagnostic tools, such as genome‐wide studies, array CGH, and whole‐exome sequencing, have highlighted more complex models of inheritance, and even more candidate genes were identified. This increase of knowledge may provide insights into the mechanisms involved in the regulation of body weight and finally lead to specific treatments. In these patients, hyperphagia is often a primary phenotypic component. Substantial gaps in understanding the molecular basis of inherited hyperphagia syndromes are present today with a lack of mechanistic targets that can serve as a basis for pharmacologic and behavioral treatments. We have evaluated retrospectively the literature data on weight, body mass index (BMI), clinical features, treatments, and treatment response in pediatric patients with forms of genetic obesity. However, this chapter provides an updated picture of emerging knowledge outlined by the more comprehensive genetic approaches, trying to outline more candidate genes for these forms of genetic obesity. Relevant papers will be identified through systematic searches of the PubMed, EMBASE and Cochrane databases. All published studies in the English language concerning these disorders will be evaluated.

In 1997, two severely obese cousins (an 8-year-old female child with a weight of 86 kg and a 5 2-year-old male child with a weight of 29 kg) were reported from a highly consanguineous 6 family of Pakistani origin [20]. Despite their severe obesity, both children had undetectable 7 levels of serum leptin and a mutation in the gene encoding leptin mapped at 7q32.1. The disease 8 is caused by mutations in the LEP gene (OMIM *164160) typically leading to defects in protein 9 synthesis or secretion, and therefore to the absence or very low blood levels of this hormone  However, recently the first cases of functional leptin deficiency have been described [23,24].
1 This entity is characterized by detectable immunoreactive levels of circulating leptin, but 2 bioinactivity of the hormone due to defective receptor binding [23,24].
3 So, serum leptin may be a useful marker in patients with severe early-onset obesity as an 4 undetectable serum leptin is highly suggestive of a diagnosis of congenital leptin deficiency 5 due to homozygous loss of function mutations in the LEP gene [12]. Leptin-deficient subjects 6 are born of normal birth weight but exhibit rapid weight gain in the first few months of life 7 resulting in severe obesity [25].

8
Leptin deficiency causes the loss of appetite control, so it is associated with hyperphagia, 9 increased energy intake and aggressive behavior when food is denied. Other phenotypic 14 Currently, the prevalence of mutations in leptin is about 1% [12].

21
In 1998 (1 year after the discovery of the congenital leptin deficiency), patients with similar 22 phenotypic characteristic of leptin deficiency, but with a high blood level of leptin, were 23 reported [28]. In these patients, a mutation in the leptin receptor (LEPR, OMIM *601007), 24 mapped at 1p31.3, has been described [28].

25
One subsequent study has demonstrated that 3% of a group of patients with severe, early-26 onset obesity had a pathogenic LEPR mutation, but blood levels of leptin were not very high, 27 suggesting that blood leptin levels cannot be used as a marker for leptin-receptor deficiency 28 [29].

33
The clinical phenotypes associated with congenital leptin-receptor deficiency are similar to 34 those of leptin deficiency, with severe obesity from the first few months of the life, hypothala-35 mic hypothyroidism and hypogonadotropic hypogonadism [12,26].

36
On the contrary, in these patients, because of a non-functional LEPR, leptin treatment is 37 ineffective. Other factors could possibly bypass normal leptin delivery systems, but these are 38 not yet currently available for the treatment of these patients [32].

39
New Thoughts on Pediatric Genetic Obesity: Pathogenesis, Clinical Characteristics and Treatment Approach 7

5
In 2010, it was described that the 220-kb 16p11.2 deletion (28.73-28.95 Mb) seen in three 6 patients co-segregated with severe early-onset obesity alone [14]. This deletion includes a 7 small number of genes, one of which was SH2B1, known to be involved in leptin and insulin 8 signaling [12]. However, several mutations in the SH2B1 gene have also been reported in 9 association with early-onset obesity, severe insulin resistance and behavioral abnormalities 10 in some patients [34].

11
The phenotype of the children with SH2B1-containing deletions is characterized by extreme 12 hyperphagia and fasting insulin levels disproportionately elevated compared to age and 13 obesity-matched controls [15]. As expected, obese SH2B1 KO mice develop hyperglycemia, 14 hyperinsulinemia, glucose intolerance, and insulin resistance and NIDDM [35]. Interestingly, 15 central and peripheral SH2B1 seem to regulate insulin sensitivity and glucose metabolism 16 independently of its action on body weight in man and mice [36].

17
In these patients, there is no specific treatment, but care must be taken in starting a specific 18 follow-up on the hyperphagia, obesity and alteration of gluco-insulinemic metabolism.

20
In 1997, a role of central melanocortin signaling in the control of energy homeostasis was 21 known [37]. Proopiomelanocortin (POMC) acts on anorectic targets of leptin in the brain [38].

28
Since POMC is the precursor of adrenocorticotropic hormone (ACTH) and melanocyte-

33
Two important POMC mutations have been described in literature: the first is the rare mutation 34 R236G that disrupts a di-basic cleavage site between β-MSH and β-endorphin, resulting in a 35 β-MSH/β-endorphin fusion protein that binds to MC4R but has reduced ability to activate the analog RM-493 [43,44], also known as setmelanotide, was awarded orphan drug status for 5 POMC deficiency and Prader-Willi syndrome [37]. Among all forms of monogenic obesity, the most common is caused by MC4-R deficiency.

8
Heterozygous mutations have been reported in many ethnic groups of obese patients and 9 prevalence varies from 0.5 to 1.0% in obese adults, up to 6% in individuals with severe infantile 14 Mutations of this gene are codominant with variable penetrance and expressivity in hetero-15 zygous carriers [48]. Both heterozygous and homozygous mutations in MC4R have been 16 implicated in obesity, but extreme obesity is incompletely penetrant in heterozygous patients 17 [3]. Also, in these patients, genetic and environmental factors influence the severity of obesity 18 associated with mutations of MC4-R.

19
The main clinical features include hyperphagia in early appearance (but not as severe as that 20 seen in leptin deficiency) and an increase in fat mass, lean mass and bone mineral density [45].

21
These patients also have an accelerated growth that seems to be a consequence of hyperinsu-22 linemia which such patients present from the earliest periods of life. It is apparently not related 23 to a dysfunction of the GH axis [3,49]. Despite this early hyperinsulinemia, obese adult subjects 24 who are heterozygous for mutations in the MC4R gene are not at increased risk of developing 25 glucose intolerance and NIDDM compared to controls of similar age and adiposity [12,45].

26
Currently, there are no specific therapies for the MC4-R deficiency, but these individuals may 27 benefit from surgical therapies, which could be taken into consideration in adults [12].

35
mapped at 5q15, present small bowel enteropathy, early-onset obesity and complex neuroen-36 docrine effects due to a failure to process the pro-hormones such as diabetes insipidus, 37 glucocorticoid deficiency, hypogonadism, and altered glucose homeostasis [51,52].

38
New Thoughts on Pediatric Genetic Obesity: Pathogenesis, Clinical Characteristics and Treatment Approach 9 A typical characteristic of these patients is a history of severe intestinal malabsorption in the 1 neonatal period, probably due to altered cleavage of intestinal peptides in the enteroendocrine 2 cells [51].

3
Over the past few years, two meta-analysis about PCSK1 mutations have been published: the 4 first in 2014 confirmed the association of PCSK1 SNPs with obesity and provides the first 5 evidence that the association between PCSK1 rs6232 and obesity is stronger for childhood 6 obesity than for adult obesity; the second meta-analysis tried to study the association of PCSK1 7 variants rs6232 and rs6234/rs6235 with quantitative BMI variation and common obesity risk in 8 subjects from diverse ethnic groups. In this study, cohort age-group significantly modulated 9 the association between rs6232, rs6234/rs6235 and obesity with the effect sizes for both SNPs 10 being stronger in children/adolescents than in adults.

11
It is thought also that the most common PCSK1 variants predispose to obesity especially in an 12 "obesogenic" environment with free access to high-caloric food [53].

7
In mice, hyperphagia associated with SIM1 deficit can be improved by the administration of 8 oxytocin, a neurotransmitter involved in the modulation of emotion (impaired oxytocinergic 9 signaling is also one possible mechanism implicated in the obesity) [58]. Mutations of the BDNF (brain-derived neurotrophic factor, OMIM *113505, mapped at 11p14.1) 12 and its receptor TrKb (tyrosin kinase B receptor, OMIM *600456, mapped at 9q21. 33   tion between rs6265 and BMI [60-62]. In addition, the minor C allele of intronic rs12291063 28 SNP was associated with lower BDNF expression and higher BMI [63].

29
NTRK2 (TrkB) mutation (which interferes with receptor autophosphorylation) causes the same 30 symptoms of BDNF deficiency such as hyperphagia, obesity, impaired nociception, and 31 intellectual disability [64,65]. Recently, a de novo mutations in TrkB was found in a boy with 32 severe obesity and impairment in learning, memory and nociception, and in a girl with 33 hyperphagia and severe obesity [66].

34
Another cause of non-syndromic monogenic obesity is due to a gene mutation of CART

35
(cocaine-and amphetamine-regulated transcript, OMIM *602606), mapped at 5q13.2. CART is an   In these syndromes, obesity can be caused by hyperphagia because are involved genes related 12 to central nervous system appetite control centers.

13
Recently, the genetic bases for some of these syndromes have been elucidated and are begin-14 ning to provide insights into the pathogenesis of the derangements of energy homeostasis.

20
The estimated prevalence for ALMS is one to nine cases per 1,000,000 individuals with nearly

12
The diagnosis is based on the phenotype of the patient, and it is confirmed when two muta-

13
tions in ALMS1 gene are identifies through molecular analysis.
14 However, it is difficult to diagnose early ALMS first of all because symptoms arise gradually 15 and secondly because the phenotypes overlap, in particular with BBS in the case of ALMS [98].

16
In recent times, thanks to the discovery of new genetic tools, in particular next-generation

24
The WES is a rapid and easier technique because it analyzes all coding regions in the genome

25
[100]. Thanks to it, in fact, mutations in ALMS1 gene have been identified in individuals, whose 26 phenotype did not seem to be typical of ALMS; therefore, it is fundamental to identifying

34
Moreover, these mutations have been shown also in consanguineous Leber congenital 35 amaurosis families through homozygosity mapping followed by WES [103].

36
As evidenced by these studies, the simultaneous use of different genetic techniques is funda-

Obesity
For management of the disease and to identify an accurate treatment, it is important for both 1 the present of typical clinical features that an appropriate genetic diagnosis, which may be 2 carried out by NGS techniques, thanks to its low cost compared with traditional polymerase 3 chain reaction and direct Sanger sequencing [103].

21
The cardinal features of PWS include infantile hypotonia, feeding difficulties due to a poor

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Initially, two nutritional phases have been described in children with PWS:

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• phase 1: the individual often presents FTT; he exhibits hypotonia with difficult feeding;

35
To date, instead, seven different nutritional phases (five main phases and sub-phases in phases 36 1 and 2) have been identified.

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New Thoughts on Pediatric Genetic Obesity: Pathogenesis, Clinical Characteristics and Treatment Approach 17 As following, focusing on nutrition, although in the early phases, the child has poor appetite, 1 the latter increases in phase 2b and leads progressively to hyperphagia, evident in phase 3 2 ( 4 Table 3. Clinical characteristics of the nutritional phases seen in Prader-Willi syndrome.

5
Analyzing the seven phases, we highlight the following:

6
• phase 0: the infant has growth restriction and decreased fetal movements;

7
• sub-phase 1a: the infant is hypotonic with difficulty feeding and with or without FTT;

8
• sub-phase 1b: the infant grows normally, and he improves appetite, also if weight gain is 9 normal;

10
• sub-phase 2a: the child has a weight gain although there is not an increased appetite or 11 caloric intake;

12
• sub-phase 2b: in addition to weight gain, there is an increased appetite;

13
• phase 3: the individual is hyperphagic; he seeks foods and presents the loss of sense of 14 satiety;

15
• phase 4: it is typical of adults, who have an insatiable appetite and are able to feel full [107].

16
As said previously, individuals with PWS present an appetite that gradually increases and 17 leads to obesity. In recent years, some studies have been conducted to understand the mech-18 anisms controlling appetitive behavior, energy expenditure and body composition.

19
The central nervous system, in particular the hypothalamus that determines changes in energy 20 balance, is involved in these processes.

Obesity
One of the determining factors for the development of obesity in these patients is ghrelin, a 28 1 amino acid peptide produced in the stomach, that transmit satiety signal and whose level in 2 obese PWS individuals is high [116,117]. Circulating ghrelin levels are elevated in young 3 children with PWS long before the onset of hyperphagia, especially during the early phase of 4 poor appetite and feeding [118].

5
The literature reports that about 25% of the adults with PWS presents NIDDM (non-insulin-

9
This syndrome, as mentioned, represents an human disorder related to genomic imprinting.

10
Although the DNA sequence of the imprinted maternally and paternally inherited alleles is

29
Children with CS tend to manifest failure to thrive in infancy and early childhood but

9
The obesogenic changes of our environment in recent decades, especially the unlimited supply 10 of cheap food with high palatability and high energy density, associated with genetic suscept-

11
ibility are the causes of the current obesity epidemic [132].

12
The recent rapid rise in prevalence of childhood obesity suggests that, probably, environmental

24
New Thoughts on Pediatric Genetic Obesity: Pathogenesis, Clinical Characteristics and Treatment Approach 21 Therefore, unlike monogenic obesity, many genes and chromosome regions contribute to 1 common obesity phenotype.   folate, methionine and vitamin B12, which affect methylation, become very important [147].

34
One study showed that prenatal exposure to malnutrition can determine abnormal DNA

13
The pathogenesis of NAFLD appears to be multifactorial. The principal risk factor for fatty 14 liver in childhood is obesity, but several other factors contribute to NAFLD development,

39
Another gene that acts together with PNPLA3 in determining hepatic steatosis is the glucoki-

19
In conclusion, obesity and fatty liver disease often go hand in hand even in the pediatric 20 population, and both are pathologies related to genetic and environmental factors.

32
The genetic contribution to common obesity has been established initially through family, twin

27
The rare variant-common disease hypothesis-suggests that rare variants contribute  report abdominal discomfort, which improves when the drug is taken with food. There is also 7 a risk of vitamin B12 deficiency; therefore, a multivitamin is recommended. The risk of lactic 8 acidosis has been observed in adults but not seen in pediatric patients [147].

9
Octreotide, a somatostatin analogue, has been investigated as a treatment for hypothalamic 10 obesity. It binds receptors on the beta cells of the pancreas and inhibits insulin release [147]. A

33
Contraindications for GH therapy in PWS patients are severe obesity, uncontrolled diabetes 34 mellitus, untreated severe OSA, active cancer and psychosis [108].

35
A number of the PWS features, such as hyperphagia, obesity and behavioral anomalies, may  foods, slow satiety mechanisms and high metabolic efficiency [198].

27
The control of food intake and energy expenditure consists of a complex network of neural 28 and hormonal systems that involving many genes [199]: in particular, the informations are 29 collected at the peripheral level (intestine, stomach, adipose tissue); then, they are processed 30 at the hypothalamic level and, finally, generate behavioral, endocrine and autonomic output 31 [198].

32
In particular, much larger portions of the nervous system of animals and humans, including and lifestyle with the human body [198]. By focusing on the neural reward systems and the 37 interaction between reward and homeostatic functions, it is possible to infer that the disturb-

38
ance of this relationship determines obesity (Figure 6). 8 The alteration of reward functions may be a cause (i.e., excessive caloric intake modulated by 9 hedonic value of food (1)) and/or a consequence (induced by obese state (3)) of obesity [198].

10
As schematically depicted in Figure 6, several potential interactions exist between food reward 11 and obesity.

12
In particular, there are three fundamental mechanisms involved in the development of obesity, functions induced by obesity [198].

17
It is also important to realize that hyperphagia is not always necessary for obesity to develop,

18
as the macronutrient composition of food can independently favor fat deposition.

19
In this regard, there is the "gluttony hypothesis" emerged from several studies in animals: in 20 particular, although reward functions are not altered unlimited access to palatable food and
2 However, it is important to underline that not all individuals exposed to environment of plenty 3 show an increased food intake and weight gain; this means that there are genetic and epigenetic 4 pre-existing alterations that make some individuals more vulnerable to the increased availa-5 bility of palatable food and food cues [198].

6
One of the key questions is how the motivation to get a reward will translate into action. In 7 most cases, the motivation for something comes from the pleasure that this has generated in 8 the past, or in other words, to obtain what has been helpful. The dopamine signal seems to be 9 a critical component in this process [198].

10
The limited information available suggests that repeated sucrose access can upregulate  addiction [198].

18
An issue on which to focus is that excessive caloric intake, as part of a disease, can gradually

35
In particular, obesity is associated with dysregulated signaling systems, such as leptin and

36
insulin resistance, as well as increased signaling through proinflammatory cytokines and 37 pathways activated by oxidative and endoplasmatic reticulum stress [204] (Figure 7).

AQ1
New Thoughts on Pediatric Genetic Obesity: Pathogenesis, Clinical Characteristics and Treatment Approach 33 1 Figure 7. Secondary effects of obesity on reward circuitry and hypothalamic energy balance regulation. Adapted with-2 permission from Berthoud et al. [198].

3
As schematically depicted in Figure 7, obesity and, in turn, neurodegenerative diseases may 4 be caused by leptin resistance, central insulin and altered regulation of energy balance, 5 controlled by hypothalamus. About the latter, the literature shows that mitochondrial and 6 oxidative stress increase due to high-fat diets leading to neural/glial dysfunction and, conse-7 quently, cytotoxic effects [198].

8
However, these toxic effects do not stop at the level of the hypothalamus but can also affect 9 brain areas involved in reward processing [198].

17
The family, especially the parents, should be actively involved in the therapeutic program and 18 become protagonists. The targeted intervention with "individual" programs only for the child,

19
on the contrary, is often unsuccessful and frustrating for the child himself [205].

15
In particular, nutrition offered to obese children must also ensure the maintenance of adequate 16 rhythms of growth and promote the maintenance of lean body mass (in particular of muscle 17 mass), which represents the metabolically active compartment, and it is the large part of the 18 total energy expenditure. Therefore, it must necessarily guarantee the macro-and micro-

19
nutrients intake in relation to their age [205].

20
In overweight and obese children and young people, it is important a multidisciplinary 21 intervention that includes dietary recommendations appropriate for age and complies with 22 the principles for a healthy nutrition (in these patients, total energy intake should be below 23 their energy expenditure) [206].

24
Dietary changes should be tailored to food preferences and allow for a flexible and individual 25 approach to reducing calorie intake; it is important not to use unduly restrictive and nutri-26 tionally unbalanced diets because they are ineffective in the long term and can be harmful [206].

27
In these patients, it is also necessary that an intervention about physical activity is important 28 not only for lose weight, but also for other health benefits, such as reduction risk of type 2 29 diabetes or heart diseases [206].

30
Therefore, obese and overweight children must be encouraged to become more active and to 31 reduce inactive behaviors, such as sitting and watching television, using a computer or playing 32 video games and to do at least 60 min of moderate or greater intensity physical activity each 33 day. The activity can be in 1 session or several sessions lasting 10 min or more [206].

34
It is important to make the choice of activity with the child and ensure that it is appropriate to 35 the child's ability and confidence, giving children the opportunity and support to do more 36 exercise in their daily lives (e.g., walking, cycling, using the stairs and active play) or to do 37 more regular, structured physical activity (e.g., football, swimming or dancing) [206].

38
Children affected by genetic obesity (e.g., PWS) often eat more than necessary for anxiety, 39 sadness, boredom: in this case, it is important not only to reduce the amount of foods but also parents must learn to celebrate each small goal, large or small, and to appreciate the acquisition 10 of any new skill [208].

11
In these children, there are behavior changes that become more apparent and severe with age:

13
These behaviors are caused specially by their insatiable appetite that causes physical, emo-

15
For these reasons, it is important to intervene to reduce stress not only for children, but also 16 for the whole family.

17
However, to control the anxious behavior in children with PWS, the following information 18 may be useful:

19
• having a regular daily routine, following appropriate food program;

20
• giving your child transitional warnings-that is, "after you finish that puzzle, it is time for 21 bath" [209];

22
• preparing your child ahead of time if there is going to be a change in routine;

23
• re-directing your child to another activity;

25
• speaking to your child in a calm, yet firm matter-of-fact tone [209].

26
In children with PWS, it is essential management food, based also on control food access, to 27 ensure adequate nutrition, weight regulation and appropriate eating behaviors.

28
Crucial in this regard is the role of parents, who must support their children in these changes 29 by adopting appropriate strategies.

30
However, each family will find the best way for them and for the specific need of their child.

31
First of all, it is important to follow an adequate food program that helps parents to monitor 32 their food intake and reassures the child that the food will always be available: therefore, it 33 represents the beginning for him to acquire the habit of eating healthy so that food can be 34 controlled and could become a part of his daily routine [209].
This program is based on three main meals (breakfast, lunch, and dinner) and two or three 1 snacks (mid-morning snack, afternoon snack, and perhaps evening snack) [209]. It is funda-2 mental to respect scheduled times (food must be given every 2-3 h), avoiding giving food 3 outside mealtimes. Whenever possible, all family members should eat at the same time and 4 others should not eat in front of the child when it is not their scheduled meal/snack time [209].

5
Portion control is another adequate strategy: it must not be excessive, but appropriate for the 6 child's age to ensure adequate growth [209].

7
However, food must be healthy considering that in children with PWS, calorie needs are lower 8 due to reduced metabolism. Food must be given only by parents/caregivers and served on the 9 plate prior to being eaten, avoiding other platters/bowls of food visible on the table and to 10 share or offer them other food [209].

11
At the end of the meal, it is important to remove the empty plate from the table and encourage 12 the child to play away from the table or from the kitchenette until all food has been taken away.

13
It is important to keep food out of sight and reach of children, keeping it under lock and key 14 if necessary (Figure 8). predisposition to develop an obese phenotype. This chapter may also help to understand better 1 the genetic diversity that could be associated in subjects with genetic forms of obesity.
2 However, this chapter may help to understand this complex problem and the different 3 approaches to treatment. In these forms of genetic obesity, the team approach to therapy (nurse 4 educators, nutritionists, exercise physiologists, and counsellors) is the basis for treatment.

5
Dramatic reductions in BMI are difficult to achieve and sustain, so counselling and therapy 6 should start with realistic goals that emphasize gradual reductions of body fat and BMI and 7 maintenance of weight loss. Finally, this chapter may provide news on the need for new 8 therapeutic approaches in the field of childhood obesity as the basis of the hyperphagia 9 treatment, a typical feature of these syndromes.

10
Acknowledgements 11 Conflicting interests: The authors declare that they have no conflicting interests.

12
Financial conflicting interests: The authors do not have any financial and non-financial 13 conflicting interests in relation to this manuscript.