title: Novel LEPR mutations in obese Pakistani children identified by PCR-based enrichment and next generation sequencing.
abstract: OBJECTIVE
  Mutations in leptin receptor gene (LEPR) result in early onset extreme adiposity. However, their prevalence in different populations is not known. Indeed, LEPR screening by gold standard Sanger sequencing has been limited by its large size and the cost. One-step PCR-based targeted enrichment could be an option for rapid and cost effective molecular diagnosis of monogenic forms of obesity.
  
  
  METHODS
  The study is based on 39 unrelated severely obese Pakistani children, previously shown to be negative for leptin (LEP) and melanocortin 4 receptor (MC4R) gene mutations, from an initial cohort of 62 probands. Patient samples were analyzed by microdroplet PCR-enrichment (RainDance technologies) targeting coding exons of 26 obesity-associated genes combined with next generation sequencing. Hormone levels were analyzed by ELISA.
  
  
  RESULTS
  The analysis revealed two novel homozygous LEPR mutations, an essential splice site mutation in exon 15 (c.2396-1 G>T), and a nonsense mutation in exon 10 (c.1675 G>A). Both probands had high leptin levels and were phenotypically indistinguishable from age-matched leptin-deficient subjects from the same population.
  
  
  CONCLUSIONS
  The two subjects carrying homozygous LEPR mutations, reported here for the first time in the Pakistani population, constitute 3% of the whole cohort of severely obese children (compared to 17% for LEP and 3% for MC4R).
authors: Department of Genomics of Common Disease, Imperial College London, London, UK.
authors: Sadia
authors: S
authors: Saeed
authors: Amélie
authors: A
authors: Bonnefond
authors: Jaida
authors: J
authors: Manzoor
authors: Julien
authors: J
authors: Philippe
authors: Emmanuelle
authors: E
authors: Durand
authors: Mohsin
authors: M
authors: Arshad
authors: Olivier
authors: O
authors: Sand
authors: Taeed A
authors: TA
authors: Butt
authors: Mario
authors: M
authors: Falchi
authors: Muhammad
authors: M
authors: Arslan
authors: Philippe
authors: P
authors: Froguel
mesh_terms: D016022:Case-Control Studies; D015331:Cohort Studies; D003241:Consanguinity; D005091:Exons / Q000235:genetics; D005260:Female; D006720:Homozygote; D006801:Humans; D007223:Infant; D008297:Male; D009154:Mutation / Q000235:genetics*; D009765:Obesity / Q000208:ethnology* / Q000235:genetics*; D010154:Pakistan; D010375:Pedigree; D010641:Phenotype; D016133:Polymerase Chain Reaction; D044105:Receptor, Melanocortin, Type 4 / Q000235:genetics; D054411:Receptors, Leptin / Q000235:genetics*; D017422:Sequence Analysis, DNA; D012720:Severity of Illness Index
chemical_list: C515448:LEPR protein, human; C470072:MC4R protein, human; D044105:Receptor, Melanocortin, Type 4; D054411:Receptors, Leptin