Establishing A Genetic Diagnosis System For 46, XY DSD And Analyzing Gene Mutation For 46, XY Female /AIS/5α-RD2 Deficiency

Background:Disorders of sex development (DSD) refer to "congenital conditions in which development of chromosomal, gonadal, or anatomical sex is atypical" and consist of a wide spectrum of complex diseases which is difficult to identify or diagnosis, mostly caused by changes in genetic material. According to different karyotypes, DSD is classified into sex chromosome DSD,46,XX DSD and 46,XY DSD. The genetic background of 46,XY DSD is the most complex one among them, and 46,XY DSD involves more genes than 46,XX DSD. Over 20 pathogenic genes have been cloned for 46,XY DSD. Among 46,XY DSD, tumour incidence is highest in 46,XY female, caused by X, Y chromosomal or autosomal gene mutation.46,XY females generally do not have offspring from their own gametes, so there is no risk of future generations. But it is well know that tumors occur with increased frequency (20% to 50%) in 46,XY female. The most common one among 46,XY DSD is androgen insensitivity syndrome (AIS), X-linked recessive inheritance, and 5a-reductase type 2 deficiency (5a-RD2 deficiency), autosomal recessive inheritance. Patients with 46,XY DSD are desperate for diagnosis and treatment because of their physical and mental pains. Molecular genetic diagnosis can not only confirm the diagnosis, more importantly, but also guide treatment and achieve prenatal diagnosis. Objective:Establishing the standardized genetic diagnosis system according to features of 46,XY DSD and sequencing genes for 46,XY female, AIS and 5a-RD2 deficiency, providing genetic diagnosis, possibility of prenatal diagnosis and informative genetic counseling, involving to prevent tumour, for these patients.Methods:46, XY DSD patients are admitted in our genetic clinics to learn more about various aspects of them. Establishing the standardized genetic diagnosis system according to clinical features, medical examination, karyotype analysis, hormone testing, B-ultrasound and imaging, even endoscopy or biopsy results, and sex chromosome DSD and 46, XX DSD are excluded. Then screening all exons of sex-determining region of the Y chromosome (SRY), desert hedgehog (DHH) and nuclear receptor subfamily 5, group A, member 1 (NR5A1)/steroidogenic factor-1 (SF1) for six 46,XY female patients from six unrelated families; screening all exons of androgen receptor (AR) for 10 patients (from five different families) with complete androgen insensitivity syndrome and the sporadic 66 patients with 46,XY DSD, maybe androgen insensitivity syndrome or otherwise, and identifying female member carriers in the five families having pathogenic mutations; screening the whole all exons of steroid 5 alpha reductase 2 isoenzyme (SRD5A2) for 19 patients with hypospadias and those who could not be excluded from hypospadias; screening the gene for 100 normal men as control according to the de novo mutation.Results:(1) Among 46,XY females, two de novo mutations of SRY (c.254T>C, C.391C>T), one de novo mutation of NR5A1 (c.740-787del (48) were detected in patient 1, patient 2, patient 3 respectively. No mutations of DHH were detected in all six patients. (2) Among the patients with complete androgen insensitivity syndrome, one de novo mutation of AR (c.2609T>C), three known pathogenic mutations of AR (c.2547insA, c.2522G>A, c.2248A>G) were detected. And four female members from two different families with AR pathogenic mutation were confirmed as carriers. Among sporadic 66 patients with 46,XY DSD, two de novo mutations of AR (c.2465T>A, c.623G>A), two known pathogenic mutations of AR (c.1847G>A, (CAG)39) and one samesense mutation of AR (c.2673C>T) were detected. The polymorphism of (CAG)n of AR in 75 patients (excluding patient 38 with pathogenic repeat) showed n=(16-30) with 9 kinds of alleles, and n=22 to the most common, accounting for 57.33%(43/75). Polyglycine (GGC)n polymorphism of AR gene in 76 patients, n= 17 in 74 patients and n= 16 in only 2. The polymorphism of AR (c.639G>A) was not detected. (3) Among 19 patients with hypospadias or likely hypospadias, two known pathogenic mutation of SRD5A2 (c.578A>G, c.680G>A) were detected. Conclusions:(1) Establishing the standardized genetic diagnosis system for 46,XY DSD. (2) Testing five genes for 46,XY female, AIS and 5a-RD2 deficiency, providing clinical genetic counseling information and prenatal diagnosis information for them. Then analyzing genotype and phenotype for 46,XY female, AIS and 5a-RD2 deficiency. And AR polymorphism provides different information, may benefit to the further research of different ethnic groups and relationship of AR gene variation and disease. (3) Detecting six novel mutations of three genes in six pedigrees.