Genotype-first in a cohort of 95 fetuses with multiple congenital abnormalities: when exome sequencing reveals unexpected fetal phenotype-genotype correlations
Authors: Anne-Sophie Lebre
Christel Thauvin-Robinet
Dominique Gaillard
Sophie Blesson
Alice Goldenberg
Sylvie Odent
Marion Aubert-Lenoir
Anne-Marie Beaufrere
Bernard Foliguet
Jean-Pierre Mazutti
Christine Francannet
Salima El Chehadeh
Marie-Hélène Saint-Frison
Antonio Vitobello
James Lespinasse
Mathilde Lefebvre
Christophe Philippe
Paul Kuentz
Sophie Nambot
Emilie Tisserant
Sophie Patrier-Sallebert
Nada Houcinat
Marie-José Perez
Mirna Assoum
Marjolaine Willems
Nicole Laurent
Sébastien Moutton
Elise Schaefer
Celine Poirisier
Elisabeth Alanio
Daphne Lehalle
Sophie Collardeau-Frachon
Anne-Marie Guerrot
Nicole Bigi
Nicolas Bourgon
Ange-Line Bruel
Jean-Louis Mandel
Valérie Kremer
Laetita Lambert
Renaud Touraine
Nathalie Marle
Frederic Tran Mau-Them
Nolwenn Jean-Marçais
Yline Capri
Anne-Claire Brehin
Maria Cristina Antal
Chloé Quélin
Melanie Fradin
Tania Attie-Bitach
Yannis Duffourd
Francine Arbez-Gindre
Philippe Jonveaux
Julien Thevenon
Françoise Girard-Lemaitre
Philippe Loget
Laurence Faivre
Journal: Journal of Medical Genetics (2021)
Discipline: CLINICAL MEDICINE
Keywords: complex traits; genetics; molecular genetics; reproductive medicine.
Affiliations: UFR Des Sciences de Santé, INSERM-Université de Bourgogne UMR1231 GAD « Génétique des Anomalies du Développement », FHU-TRANSLAD, Dijon, France., Laboratoire d'Anatomo-Pathologie, Plateforme de Biologie Hospitalo-Universitaire, CHU de Dijon Bourgogne, Dijon, France., Unité Fonctionnelle d'Innovation diagnostique des maladies rares, FHU-TRANSLAD, CHU Dijon Bourgogne, Dijon, France., Département d'Anatomo-pathologie, Hospices Civils de Lyon, Lyon, France., Laboratoire d'Embryologie et de Génétique des Malformations Congénitales, Hopital Necker, APHP, Paris Cedex 15, France., Service de Génétique Médicale, CHU de Strasbourg, Hôpital de Hautepierre, Strasbourg, France., Service de Fœtopathologie, CHU de Strasbourg, Hôpital de Hautepierre, Strasbourg, France., Laboratoire de Cytogénétique constitutionnelle et prénatale, CHU de Strasbourg, Strasbourg, France., Département Médecine translationnelle et neurogénétique, Institut de génétique et de biologie moléculaire et cellulaire, Strasbourg, France., Centre de Référence Maladies Rares « Anomalies du Développement et Syndrome Malformatifs » de L'Est, Hôpital D'Enfants, FHU-TRANSLAD, CHU Dijon Bourgogne, Dijon, France., Laboratoire de Génétique chromosomique et moléculaire, CHU de Dijon Bourgogne, Dijon, France., UF de Génétique médicale, Maternité régionale, CHU de Nancy, Nancy, France., Laboratoire de Génétique médicale, CHU de Nancy, Nancy, France., Laboratoire de Biologie de la Reproduction et du Développement Maternité de Nancy, CHU de Nancy, Nancy, France., Service de Foetopathologie, CHU de Reims, Reims, France., Laboratoire de Cytogénétique, CHU de Reims, Reims, France., Service de Génétique et Biologie de la Reproduction, CHU de Reims, Reims, France., Service d'Imagerie médicale, CHU de Besançon, Besançon, France., Service de Fœtopathologie, CHU de Besançon, Besançon, France., Service de Génétique Clinique, Hôpital Sud, CLAD Ouest, CNRS UMR6290 Génétique et Pathologies du Développement, Université de Rennes, Rennes, France., Service de Fœtopathologie, CHU de Rennes, Rennes, France., Equipe Maladies Génétiques de l'Enfant et de l'Adulte, CHU de Montpellier, Montpellier, France., Service de Fœtopathologie, CHU de Montpellier, Montpellier, France., Service de Génétique, CHU de Tours, Tours, France., Service de Génétique médicale, CHU de Clermont-Ferrand, Clermont-Ferrand, France., Service de Fœtopathologie, CHU de Clermont-Ferrand, Clermont-Ferrand, France., Service de Fœtopathologie, CHU de Rouen, Rouen, France., Service de Génétique clinique, CHU de Rouen, Rouen, France., Service de Génétique clinique, CH de Chambéry, Chambéry, France., Service de Genetique Clinique, C.H.U. De Saint Etienne-Hopital Nord, Saint Etienne Cedex 2, France., Service de génétique clinique, Hôpital Robert Debré - APHP, Paris, France., Service de foetopathologie, Hôpital Robert Debré - APHP, Paris, France., Département de Génétique et Procréation, CHU Grenoble Alpes, Université Grenoble Alpes, Grenoble, France., UFR Des Sciences de Santé, INSERM-Université de Bourgogne UMR1231 GAD « Génétique des Anomalies du Développement », FHU-TRANSLAD, Dijon, France  ., Centre de Référence Déficiences Intellectuelles de Causes Rares, Hôpital D'Enfants, CHU Dijon Bourgogne, Dijon, France.

Abstract:  Purpose: Molecular diagnosis based on singleton exome sequencing (sES) is particularly challenging in fetuses with multiple congenital abnormalities (MCA). Indeed, some studies reveal a diagnostic yield of about 20%, far lower than in live birth individuals showing developmental abnormalities (30%), suggesting that standard analyses, based on the correlation between clinical hallmarks described in postnatal syndromic presentations and genotype, may underestimate the impact of the genetic variants identified in fetal analyses. Methods: We performed sES in 95 fetuses with MCA. Blind to phenotype, we applied a genotype-first approach consisting of combined analyses based on variants annotation and bioinformatics predictions followed by reverse phenotyping. Initially applied to OMIM-morbid genes, analyses were then extended to all genes. We complemented our approach by using reverse phenotyping, variant segregation analysis, bibliographic search and data sharing in order to establish the clinical significance of the prioritised variants. Results: sES rapidly identified causal variant in 24/95 fetuses (25%), variants of unknown significance in OMIM genes in 8/95 fetuses (8%) and six novel candidate genes in 6/95 fetuses (6%). Conclusions: This method, based on a genotype-first approach followed by reverse phenotyping, shed light on unexpected fetal phenotype-genotype correlations, emphasising the relevance of prenatal studies to reveal extreme clinical presentations associated with well-known Mendelian disorders. 

Retracted: No

PubPeer comments:
 I really don t understand the strategy used for this family. ![file] ![file] ![file] The fetus is stillborn and three members of the family are affected. I guess that clinical examination of this family leads to the correct diagnosis. In this case, only one gene is known accounting for this disease. So, why did the authors choose to perform such a complicated approach? , 
 I m surprised by the case associating craniosynostosis and vertebral segmentation defects: ![file] First, I found craniosynostosis badly described. What are the sutures that are involved? Second, facial dysmorphism is also badly described. At last, haemivertebrae means nothing. The correct term is hemivertebrae. If you cross craniosynostosis and hemivertebrae on Pubmed, you obtain: Anderson et al. Cervical spine in Pfeiffer s syndrome. J Craniofac Surg 1996;7:275-279. Consequently, analysis of FGFR1 and FGFR2 should be performed in this case. I think that molecular diagnosis of this case could have been obtained by a simple analysis of the phenotype. The authors stated that the diagnosis is surprising because: ![file] This is the normal finding in Pfeiffer syndrome type III. This case shows that a correct analysis of the phenotype could lead to the genetic diagnosis without needing exome sequencing. , 
 About the case with *HRAS* mutation: - What is  partal vermian hypoplasia ? I don t know this malformation. It is important since it is listed as phenotype particularity. - What is  interventricular communication ? I suppose that the correct term is ventricular septal defect. Am I right? , 
 The case presenting *B3GLCT* mutation (Peters-plus syndrome) is supposed to present phenotypic particularities: posterior corpus callosum hypoplasia. Corpus callosum could be absent in this syndrome (see Camera G. *et al*. Clin Dysmorphol 1993;2:317-321; Schoner K. *et al*. Prenat Diagn 2013;33:75-80). Furthermore, the case published by Khatri D. *et al*. (J Pediatr Neurosci 2019;14:148-153) showed posterior corpus callosum hypoplasia. This means that such a finding is not a phenotypic particularity. In the case presented by the authors, ocular malformations (that seems very badly described for me since we know nothing about the anterior chamber) should point to Peters anomaly. The association with hydrocephaly, short stature and labial palatal clefts should point to Peters-plus. I think it would be very simple to search for *B3GLCT* mutation since it is the only gene involved. In this case also, I think exome sequencing is not necessary since the diagnosis should be proposed by experts in fetal pathology. , 
 For another case, the authors described the association between distal arthrogryposis and unilateral preaxial hexadactyly ![file] Hexadactyly was not precisely described (left or right?, upper or lower limb?). Exome sequencing in this case leads to evidencing a variant in *MYH3 *gene: ![file] The authors indicated that hexadactyly is a  phenotypic particularity . Finding genes is important but asking biological questions is much more important in my opinion. *MYH3* is only expressed by muscular cells. Consequently, a variant of this gene cannot explain an abnormal development of the digits. In my opinion, hexadactyly in this case is not linked with the phenotype. , 
 In the case of Kabuki syndrome linked with *KMT2D* variant, the authors claimed that hypoplastic left heart is a  phenotypic particularity . But see Digilio MC *et al*. Am J Med Genet 2017;173A:2912-2922; and Yoon JK *et al*. Korean J Pediatr 2015;58:256-262. Such an association has already been reported. , 
 For the case with *TUBA1A* variant: ![file] Termination of pregnancy took place at the 17th gestational week (15th week after fertilization). I found neuropathology very badly described. What is gyration defect at this age? Could the authors describe the aspect of the telencephalon? How is the cortical plate? How is the ventricular subventricular layer? Could the authors precise if the cerebellum is globally hypoplastic or not? How is the pons? Of course  hypocampal hypoplasia should be written  hippocampal hypoplasia . For a better description of the case, the authors could be helped by the paper published by Fallet-Bianco *et al*. (Acta Neuropathol Comm 2014). Could the authors illustrate what they consider as labial and palate medial cleft? Is it premaxillary agenesis? It is important to present a very good description of the phenotype in this sort of papers. , 
 For the case with *ZNF3* mutations, I have some problems with the phenotype. ![file] - What is partial maxillary agenesis? - What is partial thalamic fusion? I just want to mention that adhesion between the two thalami is a normal finding in neuroanatomy. Consequently, it should be interesting to know what is abnormal with this finding. Furthermore, expression pattern of *ZNF3* has never been published. It seems very premature to be certain that this gene is responsible for this phenotype. , 
 Another  strange case : Cornelia de Lange ![file] - Facial dysmorphism is not described. Is it compatible with Cornelia de Lange? - Complex cardiopathy is not described. The authors stated: ![file] ![file] But diaphragmatic hernia is associated with this syndrome: ![file] Furthermore: ![file] According to Gupta et al. J Pediatr Surg 2021 In conclusion, the analysis of this case is far from being precise. ,