CLINICAL REPORT
Copy Number Variants Including RAS Pathway Genes—How Much RASopathy is in the Phenotype? Christina Lissewski,1 Sarina G. Kant,2 Zornitza Stark,3 Ina Schanze,1 and Martin Zenker1* 1
Institute of Human Genetics, University Hospital Magdeburg, Magdeburg, Germany
2
Department of Clinical Genetics, Leiden University Medical Center, Leiden, The Netherlands
3
Victorian Clinical Genetics Service, MCRI, Royal Children’s Hospital, Melbourne, Australia
Manuscript Received: 12 March 2015; Manuscript Accepted: 27 April 2015
The RASopathies comprise a group of clinically overlapping developmental syndromes the common pathogenetic basis of which is dysregulated signal flow through the RAS-MAPK pathway. Mutations in several components or modifiers of the pathway have been identified in Noonan syndrome and related disorders. Over the past years copy number variants (CNVs) encompassing RAS pathway genes (PTPN11, RAF1, MEK2, or SHOC2) have been reported in children with developmental syndromes. These observations raised speculations that the associated phenotypes represent RASopathies, implying that the increased or reduced expression of the respective RAS pathway component and a consecutive dysregulation of RAS pathway signalling is responsible for the clinical picture. Herein, we present two individuals and three of their relatives harboring duplications of either 3p25.2 including the RAF1 locus or 19p13.3 including the MEK2 locus. Duplication carriers exhibited variable clinical phenotypes including non-specific facial dysmorphism, short stature, and learning difficulties. A careful review of the literature supported the impression that phenotypes associated with CNVs including RAS pathway genes commonly share nonspecific symptoms with RASopathies, while the characteristic “gestalt” is lacking. Considering the known molecular pathogenesis of RASopathies, it is questionable that a modest increase in the expression of a functionally normal signaling component can mimic the effects of a qualitatively abnormal (hyperactive) mutant protein. We thus argue that current empirical and biological evidence is still insufficient to allow the conclusion that an altered copy number of a RAS pathway component is indeed the mechanism that is critical for the phenotype associated with CNVs including RASopathy genes. © 2015 Wiley Periodicals, Inc.
Key words: RASopathy; Noonan syndrome; Noonan syndrome with multiple lentigines; cardio-facio-cutaneous syndrome; copy number variations
INTRODUCTION The RASopathies constitute a group of developmental disorders characterized by craniofacial anomalies, cardiac defects, short
© 2015 Wiley Periodicals, Inc.
How to Cite this Article: Lissewski C, Kant SG, Stark Z, Schanze I, Zenker M. 2015. Copy number variants including RAS pathway genes—How much RASopathy is in the phenotype? Am J Med Genet Part A 167A:2685–2690.
stature, and variable developmental delay [Rauen, 2013]. Noonan syndrome (NS; OMIM 163950) is the most common condition within this group. Less frequent RASopathies defined later share many key features of NS and have overlapping craniofacial anomalies, but differ in the level of cognitive impairment, ectodermal features, tumor risk, and other signs [Rauen, 2013]. They include cardio-facio-cutaneous syndrome (CFCS; OMIM 115150) and Costello syndrome (CS; OMIM 218040), as well as distinct variant types of NS such as Noonan syndrome with multiple lentigines (NSML), formerly known as LEOPARD syndrome (OMIM 151100), and Noonan-like syndrome with loose anagen hair (NSLH; OMIM 607721). The craniofacial appearance of individuals affected by RASopathies is recognizable especially in children and includes a wide forehead, hypertelorism, ptosis, a flat nasal bridge, low-set ears, and a broad neck. These features may become less obvious in adults with NS. The significance of the craniofacial phenotype in RASopathies is highlighted by established diagnostic criteria for NS [van der Burgt et al., 1994; van der Burgt, 2007] according to which the recognition of a typical or at least a suggestive craniofacial phenotype is required for the clinical diagnosis. The characteristic heart defects in RASopathies are pulmonary valve stenosis, hypertrophic cardiomyopathy or a Grant sponsor: German Research Foundation (DFG); Grant number: ZE 524/10-1. Correspondence to: Martin Zenker, Institute of Human Genetics, University Hospital Magdeburg, Leipziger Str. 44, 39129 Magdeburg, Germany. E-mail: [email protected] Article first published online in Wiley Online Library (wileyonlinelibrary.com): 14 May 2015 DOI 10.1002/ajmg.a.37155
2685
AMERICAN JOURNAL OF MEDICAL GENETICS PART A
2686 combination of both. A variety of other cardiac anomalies may occur, for example, septal defects, aortic coarctation, and valve anomalies other than pulmonary stenosis, but they lack a particular specificity for RASopathies. Developmental delays are usually mild in NS, and some patients do not experience intellectual disabilities [Pierpont et al., 2009], while these are more severe in CS and CFCS. Short stature is proportionate and typically of postnatal onset. The head size often exceeds the 97th percentile relative to height. Other symptoms of RASopathies include lymphatic abnormalities (manifesting as fetal nuchal edema or postnatal lymphedema), feeding difficulties and failure to thrive in infancy, bleeding diathesis, chest wall deformities, ectodermal signs (hyperkeratosis, curly hair), strabismus, refractive errors, and an increased risk of malignancies [Zenker, 2009; Rauen, 2013]. Mutations in multiple genes encoding components or modulators of the RAS-MAPK pathway have been identified as causative for NS and the other RASopathies (BRAF, CBL, HRAS, KRAS, MEK1, MEK2, NRAS, PTPN11, RAF1, SHOC2, and SOS1) [Rauen, 2013]. More recently mutations in RIT1 [Aoki et al., 2013] and RRAS [Flex et al., 2014] have been identified to cause NS. Notably, the mutations are almost exclusively missense mutations and a few small in-frame deletions or indels [Zenker, 2009]. It has consistently been shown that the mode of action of the resulting mutant proteins is a dysregulation (mostly hyperactivation) of the RAS-MAPK pathway [Keilhack et al., 2005; Tartaglia et al., 2006]. Despite more than a decade of research with many gene discoveries, the genetic etiology remained unclear in a significant proportion of individuals with a clinical diagnosis of NS or another RASopathy (20–30%, depending on clinical selection). Few observations of duplications encompassing RAS pathway genes have been reported in the literature and raised the hypothesis that increased gene dosage might also account for a RASopathy phenotype [Shchelochkov et al., 2008; Graham et al., 2009; Luo et al., 2012; Chen et al., 2014]. Herein we describe five individuals harboring duplications of 3p25.2 including RAF1 and of 19p13.3 including MEK2, respectively, from two unrelated families. We review previous cases from the literature and discuss the significance of these observations in light of the known molecular pathophysiology of RASopathies.
CLINICAL REPORTS Patient 1 The index patient was a 15-year-old boy (Dutch/German) with mild intellectual disability requiring special education. He was diagnosed with attention deficit and hyperactivity disorder and treated with methylphenidate. He had mild craniofacial dysmorphism including a broad nasal bridge and “interrupted” eyebrows, but no other hair abnormalities, and no cardiac anomalies (Fig. 1a, b). His measurements at age 15 years were: height 155.3 cm (
Copy Number Variants Including RAS Pathway Genes—How Much RASopathy is in the Phenotype? Christina Lissewski,1 Sarina G. Kant,2 Zornitza Stark,3 Ina Schanze,1 and Martin Zenker1* 1
Institute of Human Genetics, University Hospital Magdeburg, Magdeburg, Germany
2
Department of Clinical Genetics, Leiden University Medical Center, Leiden, The Netherlands
3
Victorian Clinical Genetics Service, MCRI, Royal Children’s Hospital, Melbourne, Australia
Manuscript Received: 12 March 2015; Manuscript Accepted: 27 April 2015
The RASopathies comprise a group of clinically overlapping developmental syndromes the common pathogenetic basis of which is dysregulated signal flow through the RAS-MAPK pathway. Mutations in several components or modifiers of the pathway have been identified in Noonan syndrome and related disorders. Over the past years copy number variants (CNVs) encompassing RAS pathway genes (PTPN11, RAF1, MEK2, or SHOC2) have been reported in children with developmental syndromes. These observations raised speculations that the associated phenotypes represent RASopathies, implying that the increased or reduced expression of the respective RAS pathway component and a consecutive dysregulation of RAS pathway signalling is responsible for the clinical picture. Herein, we present two individuals and three of their relatives harboring duplications of either 3p25.2 including the RAF1 locus or 19p13.3 including the MEK2 locus. Duplication carriers exhibited variable clinical phenotypes including non-specific facial dysmorphism, short stature, and learning difficulties. A careful review of the literature supported the impression that phenotypes associated with CNVs including RAS pathway genes commonly share nonspecific symptoms with RASopathies, while the characteristic “gestalt” is lacking. Considering the known molecular pathogenesis of RASopathies, it is questionable that a modest increase in the expression of a functionally normal signaling component can mimic the effects of a qualitatively abnormal (hyperactive) mutant protein. We thus argue that current empirical and biological evidence is still insufficient to allow the conclusion that an altered copy number of a RAS pathway component is indeed the mechanism that is critical for the phenotype associated with CNVs including RASopathy genes. © 2015 Wiley Periodicals, Inc.
Key words: RASopathy; Noonan syndrome; Noonan syndrome with multiple lentigines; cardio-facio-cutaneous syndrome; copy number variations
INTRODUCTION The RASopathies constitute a group of developmental disorders characterized by craniofacial anomalies, cardiac defects, short
© 2015 Wiley Periodicals, Inc.
How to Cite this Article: Lissewski C, Kant SG, Stark Z, Schanze I, Zenker M. 2015. Copy number variants including RAS pathway genes—How much RASopathy is in the phenotype? Am J Med Genet Part A 167A:2685–2690.
stature, and variable developmental delay [Rauen, 2013]. Noonan syndrome (NS; OMIM 163950) is the most common condition within this group. Less frequent RASopathies defined later share many key features of NS and have overlapping craniofacial anomalies, but differ in the level of cognitive impairment, ectodermal features, tumor risk, and other signs [Rauen, 2013]. They include cardio-facio-cutaneous syndrome (CFCS; OMIM 115150) and Costello syndrome (CS; OMIM 218040), as well as distinct variant types of NS such as Noonan syndrome with multiple lentigines (NSML), formerly known as LEOPARD syndrome (OMIM 151100), and Noonan-like syndrome with loose anagen hair (NSLH; OMIM 607721). The craniofacial appearance of individuals affected by RASopathies is recognizable especially in children and includes a wide forehead, hypertelorism, ptosis, a flat nasal bridge, low-set ears, and a broad neck. These features may become less obvious in adults with NS. The significance of the craniofacial phenotype in RASopathies is highlighted by established diagnostic criteria for NS [van der Burgt et al., 1994; van der Burgt, 2007] according to which the recognition of a typical or at least a suggestive craniofacial phenotype is required for the clinical diagnosis. The characteristic heart defects in RASopathies are pulmonary valve stenosis, hypertrophic cardiomyopathy or a Grant sponsor: German Research Foundation (DFG); Grant number: ZE 524/10-1. Correspondence to: Martin Zenker, Institute of Human Genetics, University Hospital Magdeburg, Leipziger Str. 44, 39129 Magdeburg, Germany. E-mail: [email protected] Article first published online in Wiley Online Library (wileyonlinelibrary.com): 14 May 2015 DOI 10.1002/ajmg.a.37155
2685
AMERICAN JOURNAL OF MEDICAL GENETICS PART A
2686 combination of both. A variety of other cardiac anomalies may occur, for example, septal defects, aortic coarctation, and valve anomalies other than pulmonary stenosis, but they lack a particular specificity for RASopathies. Developmental delays are usually mild in NS, and some patients do not experience intellectual disabilities [Pierpont et al., 2009], while these are more severe in CS and CFCS. Short stature is proportionate and typically of postnatal onset. The head size often exceeds the 97th percentile relative to height. Other symptoms of RASopathies include lymphatic abnormalities (manifesting as fetal nuchal edema or postnatal lymphedema), feeding difficulties and failure to thrive in infancy, bleeding diathesis, chest wall deformities, ectodermal signs (hyperkeratosis, curly hair), strabismus, refractive errors, and an increased risk of malignancies [Zenker, 2009; Rauen, 2013]. Mutations in multiple genes encoding components or modulators of the RAS-MAPK pathway have been identified as causative for NS and the other RASopathies (BRAF, CBL, HRAS, KRAS, MEK1, MEK2, NRAS, PTPN11, RAF1, SHOC2, and SOS1) [Rauen, 2013]. More recently mutations in RIT1 [Aoki et al., 2013] and RRAS [Flex et al., 2014] have been identified to cause NS. Notably, the mutations are almost exclusively missense mutations and a few small in-frame deletions or indels [Zenker, 2009]. It has consistently been shown that the mode of action of the resulting mutant proteins is a dysregulation (mostly hyperactivation) of the RAS-MAPK pathway [Keilhack et al., 2005; Tartaglia et al., 2006]. Despite more than a decade of research with many gene discoveries, the genetic etiology remained unclear in a significant proportion of individuals with a clinical diagnosis of NS or another RASopathy (20–30%, depending on clinical selection). Few observations of duplications encompassing RAS pathway genes have been reported in the literature and raised the hypothesis that increased gene dosage might also account for a RASopathy phenotype [Shchelochkov et al., 2008; Graham et al., 2009; Luo et al., 2012; Chen et al., 2014]. Herein we describe five individuals harboring duplications of 3p25.2 including RAF1 and of 19p13.3 including MEK2, respectively, from two unrelated families. We review previous cases from the literature and discuss the significance of these observations in light of the known molecular pathophysiology of RASopathies.
CLINICAL REPORTS Patient 1 The index patient was a 15-year-old boy (Dutch/German) with mild intellectual disability requiring special education. He was diagnosed with attention deficit and hyperactivity disorder and treated with methylphenidate. He had mild craniofacial dysmorphism including a broad nasal bridge and “interrupted” eyebrows, but no other hair abnormalities, and no cardiac anomalies (Fig. 1a, b). His measurements at age 15 years were: height 155.3 cm (
