The Journal of Clinical Endocrinology & Metabolism, 2022, 107, e1797–e1806 https://doi.org/10.1210/clinem/dgac064 Advance access publication 3 February 2022 Clinical Research Article Received: 15 November 2021. Editorial Decision: 28 January 2022. Corrected and Typeset: 24 February 2022 © The Author(s) 2022. Published by Oxford University Press on behalf of the Endocrine Society. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com Contribution of Clinical and Genetic Approaches for Diagnosing 209 Index Cases With 46,XY Differences of Sex Development Nathalia Lisboa Gomes, 1,2, Rafael Loch Batista, 1, Mirian Y. Nishi, 1 Antônio Marcondes Lerário, 3 Thatiana E. Silva, 1 Amanda de Moraes Narcizo, 4 Anna Flávia Figueredo Benedetti, 4 Mariana Ferreira de Assis Funari, 1 José Antônio Faria Junior, 1 Daniela Rodrigues Moraes, 1 Lia Mesquita Lousada Quintão, 1 Luciana Ribeiro Montenegro, 1 Maria Teresa Martins Ferrari, 1 Alexander A. Jorge, 1,5 Ivo J. P. Arnhold, 1 Elaine Maria Frade Costa, 1 Sorahia Domenice, 1 and Berenice Bilharinho Mendonca 1, 1 Unidade de Endocrinologia do Desenvolvimento/ LIM42, Hospital das Clínicas da Faculdade de Medicina da Universidade de São Paulo, São Paulo, Brazil 2 Unidade de Adrenal, Serviço de Endocrinologia, Santa Casa de Belo Horizonte, Belo Horizonte, Brazil 3 Division of Metabolism, Department of Internal Medicine, Endocrinology and Diabetes, University of Michigan, Ann Arbor, MI, USA 4 Laboratório de Sequenciamento em Larga Escala (SELA), Faculdade de Medicina da Universidade de São Paulo FMUSP, São Paulo, Brazil 5 Unidade de Endocrinologia Genética, Laboratório de Endocrinologia Celular e Molecular LIM25, Disciplina de Endocrinologia da Faculdade de Medicina da Universidade de São Paulo, São Paulo, Brazil Correspondence: Berenice B. Mendonca, Hospital das Clínicas, Faculdade de Medicina da Universidade de São Paulo, Disciplina de Endocrinologia e Metabologia., Av. Dr. Enéas de Carvalho Aguiar, 155, 2° andar, bloco 6, CEP: 05403-900, São Paulo, Brazil. Email: beremen@usp.br/; Nathalia Gomes, PhD, Hospital das Clínicas, Faculdade de Medicina da Universidade de São Paulo, Disciplina de Endocrinologia e Metabologia., Av. Dr. Enéas de Carvalho Aguiar, 155, 2° andar, bloco 6, CEP: 05403-900, São Paulo, Brazil. Email: nathalialisboa.endocrino@gmail.com. Abstract Context: Massively parallel sequencing (MPS) technologies have emerged as a first-tier approach for diagnosing several pediatric genetic syn- dromes. However, MPS has not been systematically integrated into the diagnostic workflow along with clinical/biochemical data for diagnosing 46,XY differences of sex development (DSD). Objective: To analyze the contribution of phenotypic classification either alone or in association with genetic evaluations, mainly MPS, for diagnosing a large cohort of 46,XY DSD patients. Design/patients: 209 nonsyndromic 46,XY DSD index cases from a Brazilian DSD center were included. Patients were initially classified into 3 subgroups according to clinical and biochemical data: gonadal dysgenesis (GD), disorders of androgen secretion/action, and DSD of unknown etiology. Molecular genetic studies were performed by Sanger sequencing and/or MPS. Results: Clinical/biochemical classification into either GD or disorders of hormone secretion/action was obtained in 68.4% of the index cases. Among these, a molecular diagnosis was obtained in 36% and 96.5%, respectively. For the remainder 31.6% classified as DSD of clinically un- known etiology, a molecular diagnosis was achieved in 31.8%. Overall, the molecular diagnosis was achieved in 59.3% of the cohort. The com- bination of clinical/biochemical and molecular approaches diagnosed 78.9% of the patients. Clinical/biochemical classification matched with the genetic diagnosis in all except 1 case. DHX37 and NR5A1 variants were the most frequent genetic causes among patients with GD and DSD of clinical unknown etiology, respectively. Conclusions: The combination of clinical/biochemical with genetic approaches significantly improved the diagnosis of 46,XY DSD. MPS poten- tially decreases the complexity of the diagnostic workup as a first-line approach for diagnosing 46,XY DSD. Key Words: differences in sex development, 46,XY gonadal dysgenesis, massively parallel sequencing, DHX37 Diagnosing patients with 46,XY differences of sex develop- ment (DSD) is quite challenging due to the complexity of the process of sex development. As a result, 46,XY DSD encom- passes a myriad of conditions with distinct molecular/genetic causes but highly overlapping phenotypic features that re- quire an intensive diagnostic workup to be distinguished from each other (1). Until the 1990s, the etiological diagnosis of patients with 46,XY DSD was based on the results of exten- sive clinical, hormonal, and imaging studies (2). The incorp- oration of genetic tests to the clinical practice increased the diagnostic accuracy of 46,XY DSD (1). Initially, clinical/bio- chemical diagnostic workflows were used to guide sequencing of a presumptive candidate gene by Sanger method. This ap- proach usually confirms most of the diagnosis of patients with Downloaded from https://academic.oup.com/jcem/article/107/5/e1797/6521524 by guest on 02 August 2023