THE GENETICS OF OBESITY. One of the most serious health problem of the XXI. century

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1 THE GENETICS OF OBESITY One of the most serious health problem of the XXI. century

2 Obesity is a condition associated with the accumulation of excessive body fat resulting from chronic imbalance of energy whereby the intake of energy exceeds expenditure. Worldwide, more than one billion adults are overweight and over 300 million are obese.

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4 ANOREXIA NERVOSA OREXIS: appetite OREXIGEN: INCREASING APPETITE/ FOOD INTAKE ANOREXIGEN: DECREASING APPETITE/ FOOD INTAKE

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6 Hypothalamus

7 BMI= mass (kg)/ height 2 (m)

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9 WHY IS IT BAD TO BE FAT?

10 WHY IS IT BAD TO BE FAT?

11 STROKE WHY IS IT BAD TO BE FAT? BREATHING PROBLEMS CARDIOVASCULAR DISEASES EARLY DEATH ARTHRITIS DIABETES PROBLEMS WITH REPRODUCTION CANCER Manuel Uribe Mexico kg

12 Prevalence: 59 million (30%) Americans are obese (BMI>= 30) Rates are increasing faster than ever (epidemic proportions)

13 FACTORS AFFECTING OBESITY Energy Intake Genetic Predisposition Energy Partitioning Favoring Fat Accretion Obesity Overweight Energy Expenditure Adipogenesis

14 Sedentary time Smoking Prenatal factors Pollutants Viruses PAL Lactation Energy Intake Genetic Predisposition Energy Expenditure Energy Partitioning Favoring Fat Accretion Adipogenesis Obesity Overweight Nutrition Physical environment Social environment

15 BMI PHENOTYPE = ENVIRONMENTAL EFFECTS + GENOTYPE OBESOGENIC ENVIRONMENT RESTRICTIVE ENVIRONMENT NOT PRONE TO BE FAT PRONE TO BE FAT

16 EVOLUTIONARY THEORIES OF OBESITY -THRIFTY GENOTYPE (J. Neel 1962) -GLUTTONOUS GENOTYPE -SLOTHFUL GENOTYPE

17 Familial risk FAMILIAL AGGREGATION OF OBESITY Proband BMI = Lee JH et al, IJO, BMI in Relatives

18 THE ROLE OF FATTY FAMILIES IN THE SPREADING OF OBESITY FATTIES OFTEN MARRY FATTIES THE KIDS OF FAT PARENTS ARE THE MOST OBESE

19 GENETIC DIVERSITY OF OBESITY genes environment Monogenic high penetrance Monogenic low penetrance Poligenic Rare cases Syndromes Population study Genetic epidemiology Tissue investigation in clinical trials omic studies Example: Adipose tissue analysis

20 POLIGENIC OBESITY

21 SEARCHING FOR OBESITY GENES

22 Prevalence SEARCHING FOR OBESITY GENES Body fat

23 SEARCHING FOR OBESITY GENES Candidate Gene Candidate Position Quantitive Trait Locus (QTL) RELATION TO FUNCTIONS

24 LOD score What is QTL? QTL 0 cm

25 QTL-s in the Genome associated with obesity 1p Creighton FS Dutch FS Pima FS Quebec FS 2p San Antonio FHS Paris-Lille FS HERITAGE FS 2q Creighton FS HERITAGE FS 3q AA FS HERITAGE FS TOPS FS 4p Quebec FS Utah FS 4q Creighton FS HERITAGE FS Quebec FS 5p AA FS TOPS FS Pima FS Paris-Lille FS 6p AA FS Pima FS 7q Amish FS NHLBI FHS Quebec FS

26 QTL-s in the Genome associated with obesity 8q San Antonio FHS HERITAGE FS 9q HERITAGE FS Quebec FS 10p Amish FS Dutch FS HERITAGE FS Paris-Lille FS Pima FS 13q NHLBI FHS Quebec FS 14q Amish FS HERITAGE FS TOPS FS 17q Quebec FS San Antonio FHS TOPS FS 18q Quebec FS Finnish FS Pima FS 20q Univ Penn FS Pima FS

27 SEARCHING FOR OBESITY GENES QTL Inventory of genes Candidate gene Wild type and allelic versions GENE Sequence variants Sequencing Population studies FUNCTIONAL STUDIES Clinical studies Transgenic animals In vitro cell studies

28 MONOGENIC OBESITY

29 GENES RESULTING IN MONOGENIC OBESITY IN THE HUMAN GENOME

30 RODENT MODELS OF MONOGENIC OBESITY Strain Inheritance Encoded Protein Defect Obese (ob) Mouse Recessive Leptin Stop codon/promoter defect in leptin Diabetes (db) Mouse Recessive Leptin receptor Defect lepr Agouti yellow (a y ) Mouse Dominant Agouti Ectopic expression of melanocortin receptor antagonist Tubby (tub) Mouse Recessive Phosphodiesterase Apoptosis in the brain? Fat Mouse Recessive Carboxypeptidase E Carboxypeptidase E activity abolished Zucker/fatty (fa) Rat Recessive Leptin receptor Defect lepr Koletsky (kol) Rat Recessive Leptin receptor Defect lepr Corpulent (cp) Rat Recessive Leptin receptor Defect lepr

31 Gene Obesity HUMAN MONOGENIC OBESITY Birth weight Endocrine abnormalities Hyperphagia Inheritance Chromosome LEP Severe Normal Low leptin Hypogonadism High thyroid-stimulating hormone High insulin + Recessive 7q31.1 High leptin 1p31 LEPR Severe? Pituitary dysfunction Hypogonadotrophic hypogonadism Sympathetic dysfunction High Insulin + Recessive POMC Severe Normal Red hair pigmentation ACTH deficiency hypocortisolism Low - MSH + Recessive 2p23.3 PC1 Severe? Hypogonadotrophic hypogonadism Hypocortisolism High proinsulin, low insulin Postprandial hypoglycemia? Recessive 5q High POMC MC4-R Severe Normal Not observed + Dominant 18q22 NROB2 Mild High Mild hyperinsulinemia - Dominant 1p36.1 LEP, leptin; LEPR, leptin receptor; POMC, pro-opiomelanocortin; PC1, prohormone convertase1; MC4-R, melanocortin-4 receptor; ACTH, adrenocorticotropic hormone; - MSH, -melanocyte-stimulating hormone.

32 THE REGULATION OF FOOD INTAKE

33 THE REGULATION OF FOOD INTAKE HYPOTHALAMUS orexigenic anorexigenic -MSH

34 anorexigenic orexigenic

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37 ANOREXIGENIC SIGNALS LEPTIN LEPTIN RECEPTOR -MSH PROHORMONE CONVERTASE 1 MELANOCORTIN 4 RECEPTOR

38 LEPTIN LEPTIN RECEPTOR

39 Fat cells Leptin receptor Anterior. pituitary Hypothalamus TRH, CRH, GnRH Skeletal muscle Liver Pancreas Angiogenesis Other tissues Puberty Fertility Food intake Metabolic rates Glucose metabolism Lipid metabolism Immune functions

40 LEPTIN MUTATIONS LEPTIN: DECREASES APPETITE & INCREASES FAT METABOLISM 7q31 cebp * kb 1.9kb Gene= 18kb * cdna= 3.5 kb AAAA Leptin Leptin* 167 aa * 133aa Nature, 387, pp June 26, 1997 THE PROTEIN IS NOT SECRETED

41 LEPTIN RECEPTOR SIGNALING class I cytokine receptor superfamily The leptin receptor: (Ob-R), the diabetes (db) gene alternate splicing: six leptin receptor (Ob-Ra-f) isoforms Leptin signaling is blocked by suppressors of cytokine signaling-3 (SOCS-3) and by dephosphorylation by protein tyrosine phosphatase 1B (PTP-1B). SOCS-3 and PTP-1B deficient mice exhibit increased sensitivity to leptin and resistance to obesity.

42 LEPTIN RECEPTOR MUTATIONS LOSS OF TRANSMEMBRANE/INTRACELLULAR DOMAIN

43 LEPTIN: DECREASES APPETITE & INCREASES FAT METABOLISM In muscle Leptin activates 5 - AMPactivated protein kinase (AMPK) In liver, leptin turns down the activity of the gene for stearoyl-coa desaturase-1 (SCD-1)

44 LEPTIN THERAPY: MICE & MAN Left: Ob mouse 6 weeks post leptin therapy Right: Ob mouse 6 weeks post saline injections A child with a mutation in the leptin gene before and after leptin therapy

45 BEFORE LEPTIN THERAPY Initial body weight, 125 kg on average Daily leptin injection for ten months

46 AFTER 3 MONTHS LEPTIN THERAPY

47 AFTER 10 MONTHS LEPTIN THERAPY THEY LOST HALF OF THEIR BODY WEIGHT!

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49 POMC HYPOTHALAMUS orexigenic anorexigenic POMC PC MSH PROOPIOMELANOCORTIN

50 anorexigenic orexigenic

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53 POST-TRANSLATIONAL CLEAVAGE OF POMC Modified by Coll et al., 2004 NH 2 COOH ANTERIOR PITUITARY HYPOTHALAMUS N-teminal JP ACTH Β-LPH γ 3 -MSH γ-lph α-msh CLIP γ 1 -MSH PC1 cleavage site products PC2 cleavage site products β-msh β-end 1-27 Hypothalamus Anterior Pituitary

54 MSH: MELANOCYTE STIMULATING HORMONE MC4-R: MELANOCORTIN 4 RECEPTOR POMC ACTH MSH MSH ACTH MC1-R s MC2-R MC4-R s Eumelanin pigment synthesis skin Glycocorticoid Adrenal cortex Inhibiting food intake Hypothalamus

55 human mutant POMC proteins ARG236GLY mutation in the POMC gene leads to EARLY-ONSET OBESITY in children

56 prohormone convertase1 PC1

57 PC 1 MUTATION SEVERE OBESITY PROHORMON MATURATION PROBLEMS (Proinsulin, POMC) Fraction Number O Rahilly et al, NEJM, 1995 &Jackson et al, Nature Genet 1997, Jackson, Nat Genet 2003

58 Melanocortin 4 Receptor MC4R

59 THE REGULATION OF FOOD INTAKE HYPOTHALAMUS orexigenic anorexigenic -MSH

60 anorexigenic orexigenic

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62 305 E V F V T L G V I S L L E N I L V I V A I A N K N L S C I L L T I I I T E S G N S V S V L M D A V A F F Y D N V I D S V I C S S L L A S I C S L L S I A D R Y F F T I I T M F F T M L A L M A S L Y V H M F L M A R L H I K R I I G S V T C A A W I C S I I I G V R K V T L F F P A W C V V F V G I L I T L T I A G K M N A G Q R I L N Y A L F N L Y L I L I M C N S I I D P L I Y A H S P M S T T V N I A L Q Y H N M I I I S S Y D A S V I I C L V L P G T A G C P Q N P Y C V C F M S H Y P N S V F L E A S H L R Y S S Q F R S Q E L R K T F K E I I C C L P G L L G C D L S S R Y H L W N R G M H T S L V N S T H R M K G G L S E S C Y G G D S Y D S T F Q A D I L H S I Y F K V H S H L C F* M C S M L I D T P L T D I P W Q -- V T P S E I S S* Y S P H T W S T Q A V* S L K T S R K V L L C R N S S N R NH2 COOH Extracellular Intracellular more than 90 mutation in the MC4R gene 2-3% of obesity cases MC4R

63 MELANOCORTIN SIGNALING

64 IMC 70,00 LEPR mutation MC4R homozygote ( AG ) MC4R homozygote (I166V) MC4R heterozygote children (13) Obese children with wild type MC4R (40) Clement Nature 1998 ; Dubern, et al J Pediatr 2001 and ,00 66,00 64,00 62,00 60,00 58,00 56,00 54,00 52,00 50,00 48,00 46,00 44,00 42,00 40,00 38,00 36,00 34,00 32,00 30,00 28,00 26,00 24,00 22,00 20,00 18,00 16,00 14,00 12,00 10, Age (ans) 97P 50P LepR Mutation P97 LepR P95 Mutation P90 P75 P50 P25 P5 Homo (V166I) LepR 1 LepR 2 Non mutes Het MC4R

65 functional consequences of MC4R mutations AGRP N - + MSH MEMBRANE EXPRESSION Intracellular retention => 56% of MC4R mutations X C G Prot AC camp RECEPTOR ACTIVITY? Weak MSH response 80% of MC4R mutations Food Intake Energy homeostasis

66 MC4R Deficiency 9-year-old boy, homozygous for a mutation in MC4R 16-year-old brother with a normal genotype Severe early childhood obesity Dominant inheritance Farooqi et al, NEJM, 2003

67 OREXIGENIC SIGNALS Marco Ferreri: La Grande Bouffe

68 Neuropeptid Y (NPY)

69 THE REGULATION OF FOOD INTAKE HYPOTHALAMUS orexigenic anorexigenic -MSH

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71 NEUROPEPTIDE Y stimulation of food intake (orexigenic) receptor: Y1 & Y5 (GPCR) NPY/AgRP neuronos are inhibited by leptin & insulin NPY/AgRP neurons inhibit the POMC neurons (with GABA) NPY/AgRP neurons activate anabolic processes & inhibit catabolic processes

72 Ghrelin

73 THE REGULATION OF FOOD INTAKE HYPOTHALAMUS orexigenic anorexigenic -MSH

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76 Ghrelin: Endogenous Ligand for the GH Secretagogue Receptor stimulation of food intake (orexigenic) stomach & hypothalamus stimulate GH secretion Decreases fat oxidation Prader-Willi syndrome: chronic hunger, high plasma ghrelin levels Ghrelin Vaccination against obesity?

77 GHRELIN

78 GHRELIN 117AA ghrelin precursor (preproghrelin) Protease cleavage and acyl modification: 28AA mature ghrelin peptide The acyl modification, mainly n-octanoyl modification, is essential for the activity of ghrelin. S, signal peptide.

79 GHRELIN MUTATIONS G274A Arg51Gln Leu72Met Exon 1 Exon 2 Exon 3 Exon 4 Mature ghrelin product

80 THE CONSEQUENCES OF GHRELIN MUTATIONS Cleavage site of a signal peptide * 1 MPSPGTVCSLLLLGMLWLDLAMAGSSFLSP 30 Mature Ghrelin Product Arg51Gln mutation 31 EHQRVQQRKESKKPPAKLQPRALAGWLRPE 60 C-terminal-processing site 61 DGGQAEGAEDELEVRFNAPFDVGIKLSGVQ 90 Leu72Met polymorphism 91 YQQHSQALGKFLQDILWEEAKEAPADK 117 * n-octanoyl modification site

81 OREXIN, MCH, ENDOCANNABINOIDS

82 OREXIN stimulation of food intake (orexigenic) Orexin-A, Orexin-B, prepro-orexin, 130 AA receptor: OX1R & OX2R (GPCR) NARCOLEPSY & OBESITY

83 MELANIN-CONCENTRATING HORMONE stimulation of food intake (orexigenic) MCH, 19AA Prohormon: Pmch receptor: MCHR1 & MCHR2 (GPCR) MCH OVEREXPRESSION: FAT MICE MCHR1- KO: LEAN MICE

84 ENDOCANNABINOIDS anandamide & 2-arachidonyl glycerol receptor: CB1 cannabinoid receptor (GPCR) stimulation of food intake (orexigenic) CB1 antagonists are potential medicines against obesity

85 Short term regulation of food ingestion buffers the swings in nutrient availability. For example, following a meal blood glucose is high at which time the pancreatic hormone, insulin, is secreted to lower blood glucose by producing liver glycogen. Between meals, blood glucose falls causing secretion of another pancreatic hormone, glucagon, which coverts glycogen back to glucose. The liver monitors glucose and fatty acid titers to elicit feeding behavior via the vagus nerve. You feel sated after a meal due to distention of the stomach and release of hormones (CCK) into the blood. Both inhibit the satiety center in the medulla. Long term regulation of food ingestion involves neuropeptides which stimulate feeding (e.g., Orexin, Melaninconcentrating hormone, and Neuropeptide Y) or inhibit feeding (CART and alpha-msh). These neuropeptides reside in the Arcuate Nucleus and other hypothalamic nuclei. Leptin is a peptide hormone that is secreted when adequate white fat stores are present. Leptin inhibits feeding behavior to regulate body weight, and it is critical for reproductive competence.

86 Monogenic syndromes with obesity Syndrome Name (reference) Clinical heterogenity Transmission Loci / Genes Prader-Willi Muscular Hypotony Mental retardation Hyperphagia Hypogonadism Short stature Autosomic dominant imprinting 15q11 SRNPN Micro deletion Maternal Disomy Bardet-Biedel Mykytyn Nature Genet 2002 Hypogonadism Pigmentary retinopathy Polydactyly Mental retardation Autosomic recessive BBS (1-12) chaperonin Protein MKKS (Chr 20) Ciliary cells proteins Alström Hearn Nature Genet 2002 Collin Nature Genet 2002 Myocardiopathy Sensory deficit (retinopathy, deafness) Dyslipidemia, diabetes Autosomique recessive 2p14 ALMS1 Börjson-Forssman- Lehman Lower Nature genet 2002 Morbid obesity, epilepsy Hypogonadism, facial dysmorphy Xq26.3 / Plant homeodomain like finger gene

87 BARDET-BIEDL SYNDROME (BBS)

88 BBS: CILIOPATHY Name Locus Gene Protein BBS1 11q13 BBS1 Ciliary protein (M390R mutation 80%) BBS2 16q21 BBS2 Ciliary Protein BBS3 3p13 BBS3 G-ADP ribosylation BBS4 15q22 BBS4 PCM1 recruitment (pericentriolar material protein) BBS5 2q13 BBS5 synthesis cilia flagella BBS6 20p12 MKKS McKusick-Kaufman (chaperonin) BBS7 4q27 BBS7 Ciliary protein (close to BBS 1 & 2) BBS8 14q32 BBS8(TTC8) Cell Motility (primary cilia) BBS9 7p14 PTHB1 regulated by Parahormone BBS10 12q BBS10 Chaperonin Protein (new)* BBS11 9q33.1 BBS11? Unconfirmed BBS12 4q27 BBS12 Chaperonin Protein (new)*

89 Most of our cells have motile or primary cilium

90 Dysfunction of movement regulation (BBS7,8)

91 Evolution in the Status of the Human Obesity Gene Map Single-gene a) Mendelian disorders Animal QTLs Human QTLs Candidate genes a) Number of genes, not the number of mutations Compiled from the Obesity Research Series

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