CURRENT CONCEPTS REVIEW GENES AND ORTHOPEDICS : FROM GENE TO CLINIC AND VICE VERSA Ph. DEBEER1,2,3, L. DE SMET1, W. J. M. VAN DE VEN3, G. FABRY1, J.-P. FRYNS2 Recent advances in molecular biology have greatly identify novel genes. Recent technical advances in helped in understanding the mechanisms involved in molecular biology and the completion of the normal skeletal morphogenesis. Multiple genes Human Genome Project (38, 61), which involved involved in normal skeletal development have been sequencing the three million base pairs of the identified, but several others still await discovery. human genome, have significantly facilitated the Mutations in these genes are often responsible for the possibility to locate and identify genes responsible congenital skeletal malformations that we see in the orthopedic clinics. In this overview we would like to for normal skeletal development. Further characte- emphasize the importance of the interaction between rization of these genes and elucidation of the func- orthopaedic surgeons, molecular biologists and tion of the corresponding proteins will undoubted- geneticists. ly provide us with more information regarding their role in normal limb- and skeletal development. Keywords : skeletal development ; congenital orthope- In this review, we present a brief overview of the dic malformations. current knowledge of the molecular aspects of Mots-clés : développement du squelette ; malformations orthopédiques. skeletal disease and illustrate how the recent advances in genetics may have practical implica- tions for patients with orthopaedic problems. INTRODUCTION Etiology of congenital malformations The embryonic development of the skeleton and Over 6000 human disorders exhibit simple gene limbs is an intriguing and complex process, which unifactorial or Mendelian inheritance. A disorder involves a cascade of molecular interactions. More determined by a gene on an autosome is said to specifically, differentiation, proliferation and pro- show autosomal inheritance, whereas a disorder grammed cell death play an important role. Much determined by a gene on one of the sex chromo- of what we know about the general mechanism underlying skeletal morphogenesis is the result of animal studies. The consequences of spontaneous ———————— 1 or induced mutations in the fruitfly (Drosophila Department of Orthopedics, University Hospital Pellenberg, Weligerveld 1, B-3212 Lubbeek (Pellenberg), melanogaster), worm (Caenorhabditis elegans), Belgium. zebra fish (Danio rerio), frog (Xenopus laevis), 2 Centre for Human Genetics, Herestraat 49, chicken and mouse have lead to the identification B-3000 Leuven, Belgium. of a multitude of genes involved in normal skeletal 3 Laboratory for Molecular Oncology, Herestraat 49, development (39, 41, 44, 45). Congenital deforma- B-3000 Leuven, Belgium. tions in humans, isolated or as part of a syndrome, Correspondence and reprints : Ph. Debeer, Department of Orthopaedic Surgery, U.Z. Pellenberg, Weligerveld 1, also provide us with the unique opportunity to B-3212 Lubbeek (Pellenberg), Belgium. Acta Orthopædica Belgica, Vol. 68 - 3 - 2002 204 PH. DEBEER, L. DE SMET, W. J. M. VA N D E V E N , G. FABRY, J.-P. FRYNS somes is said to show sex-linked inheritance. A tal malformation occasionally reveal a chromo- dominant trait manifests itself in a heterozygote, a somal aberration. These include abnormalities in person possessing both the abnormal or mutant chromosome number (aneuploidies, e.g. trisomy allele and the normal allele. The clinical features in 13 and trisomy 21, which give rise to polydacty- dominant traits can vary from person to person. ly and brachydactyly respectively) or abnormali- This is known as variable expressivity. In some ties in chromosome structure (for example the individuals of families with autosomal dominant ring chromosome 13 syndrome which is accom- traits, the presence of the mutation can go unde- panied by absence of the thumb). tected (‘skip a generation’), which is known as 4. The great majority of congenital malformations reduced penetrance. In autosomal dominant disor- however, are caused by a combination of genetic ders an affected person usually has an affected and environmental factors. In these cases parent. Sometimes a disorder can appear in an indi- patients have a genetic liability to develop a vidual whose parents (and all the previous genera- disease, but an environmental factor is also tions) are unaffected. This sudden appearance of a necessary. condition due to a mistake occurring in the trans- We will further focus our attention on congenital mission of a gene is called a new mutation (de novo disorders with an underlying genetic defect. mutation). Recessive disorders are only manifest when the mutant allele is present in a double dose How can we find novel genes involved in skeletal (homozygosity). Both parents of the affected person malformations ? are obligate heterozygous for the mutation. They are perfectly healthy carriers. Many disorders Several approaches can be used in order to detect demonstrate a type of inheritance that is not similar genes involved in human disease (11, 12). Some- to any recognised Mendelian pattern (for instance times information about the underlying biochemi- congenital dislocation of the hip). This pattern of cal defect is used. Using a functional cloning inheritance is referred to as multifactorial. Both approach, the identification of a gene is based on genes and environmental factors play a role in these pre-existing information about the underlying bio- disorders. chemical defect. No reference is made to any chro- When searching for the possible causes of con- mosomal map position or sequence information. genital malformations, four categories must be Positional cloning assumes no functional informa- considered. tion and must locate the responsible genes on the 1. Sometimes, a clear exogenous insult can be basis of map position. In a first step, linkage analy- identified as the main causative factor. Examples sis of multiple affected families can assist in map- include the limb anomalies caused by thalido- ping the disease gene on a specific chromosome. mide, congenital malformations caused by The identification of patients with chromosomal maternal illness (infections, hypertension, dia- aberrations (chromosomal deletions, inversions, betes, toxemia) and transverse limb deficiencies translocations, duplications,...) is often of great caused by early chorion villus sampling with help. After mapping the cytogenetic aberration to fetal vascular shock. Poland syndrome (unilater- its correct location on the chromosome, the candi- al symbrachydactyly with ipsilateral aplasia of date interval is narrowed down until the gene is the pectoralis major) is probably due to a prima- identified. The genetic tools used mainly consist of ry defect in the development of the subclavian previously constructed chromosomal maps and artery. sequence information. All these data can be found 2. In some Mendelian inherited conditions, a single at various sites on the World Wide Web. The com- major gene can be identified, for example the pletion of the Human Genome Project greatly faci- HOXD13 mutations in synpolydactyly (1, 47). litates the search for genes involved in human 3. Chromosome studies in patients with a congeni- disease. The candidate gene approach makes use of Acta Orthopædica Belgica, Vol. 68 - 3 - 2002 GENES AND ORTHOPEDICS 205 information of already known genes. Features of Foot-Genital-Syndrome (42, 43)). Figure 1 illus- the disease are compared with those of the gene trates how currently all the available data and tech- and, depending on the results of this comparison, a niques can be combined in searching for human specific gene is considered a good candidate or not. disease genes. There is a clear interplay between Mutation analysis of this gene in affected patients clinical work, computer analysis (“cyber cloning”) may prove that this gene is indeed responsible for and laboratory benchwork. It is obvious that the the observed phenotype. The positional candidate advances in the Human Genome Project greatly gene approach combines the two latter methods. speed up this approach. First the disease locus is mapped to the correct chromosomal region, followed by a search of this Molecular genetics of skeletal disease : an interval to see whether any functionally interesting overview candidate genes have been positioned there. In our search for genes involved in skeletal malforma- A detailed description of all the disorders given tions, information derived from other species is below can be found in the OMIM database (Online often of great help. Several mouse models with dis- Mendelian Inheritance in Man ; http://www.ncbi. tinct phenotypes have been described (39, 41). nlm.nih.gov/Omim/). Table 1 gives an overview of Sometimes a human limb deformity shows great some congenital malformations with orthopedic resemblance with that of a previously described implications. The underlying genetic defect, if mouse. If the genetic defect in the mouse is known, known, is indicated together with the mode of identification of its human counterpart is possible inheritance (AD = autosomal dominant ; AR= auto- (for example the identification of hoxa13 mutations somal recessive) and the OMIM number. Neuro- in the hypodactyly mouse and the subsequent iden- muscular and metabolic disorders were not in- tification of human HOXA13 mutations in Hand- cluded. Fig. 1. — This flowchart illustrates the important interplay between