Sekvenciranje celotnega genoma bolnikov s hipogonadotropnim hipogonadizmom

Breznik, Nika (author), Trebušak Podkrajšek, Katarina (mentor), Avbelj Stefanija, Magdalena (co-mentor)

Hipogonadotropni hipogonadizem (HH) je heterogena bolezen z variabilno klinično sliko, ki nastane zaradi nepravilnega razvoja olfaktorne plakode, nepravilnosti v migraciji nevronov GnRH in motenj v uravnavanju izločanja ali delovanja GnRH. Prirojeni HH delimo glede na zmožnost zaznave vonja na Kallmannov sindrom (KS) in normozmični idiopatski HH (nIHH). Opredelitev genetskega vzroka HH je pogosto težavna in v 50 % ostane nepojasnjena. V magistrsko nalogo smo vključili 20 preiskovancev iz 18 družin, pri katerih do sedaj uporabljeni genetski pristopi pri prepoznavanju genetskega vzroka bolezni niso bili uspešni. Klinična diagnoza KS je bila postavljena pri 15 preiskovancih, pri 5 preiskovancih je bila postavljena diagnoza nIHH. Dodatne klinične značilnosti so bile prisotne pri 13 preiskovancih, 12 preiskovancev pa je imelo pozitivno družinsko anamnezo. Pri vseh preiskovancih smo izvedli sekvenciranje celotnega genoma in nato analizirali spremembe posameznega nukelotida, manjše delecije, insercije in spremembe v številu kopij izbranih 366 genov ter tako poskušali opredeliti genetski vzrok bolezni. Pri preiskovancih, pri katerih smo imeli podatke celotnega genoma tudi za družinske člane, smo analizirali intronska področja osnovnega nabora 34 genov, ki so znano povezani s HH. Kot potrditveno metodo za opredelitev družinske segregacije sprememb smo uporabili sekvenciranje po Sangerju. Genetsko etiologijo smo dokončno pojasnili pri 30 % (6/20) preiskovancev in delno pri dodatnih 45 % (9/20) preiskovancev. Pri preiskovancih smo opredelili devet novih genetskih sprememb v genih ANOS1, CCDC141, FGFR1, IGSF10, PROK2, SEMA3A, SEMA3G in SPRY4, ki so znani vzročni geni za HH. Identificirali smo tudi spremembo v kandidatnem genu LGR4, ki do sedaj še ni bil povezan s HH. Pri samo 2 preiskovancih nismo prepoznali sprememb, ki bi lahko pojasnile nastanek klinične slike HH. Pri 3 preiskovancih iz iste družine smo v genu FGFR1 identificirali globoko intronsko spremembo, ki bi lahko bila vzrok za nastanek klinične slike pri tej družini, a glede na obstoječe podatke to težko opredelimo. Globoke intronske spremembe do sedaj še niso bile prepoznane pri bolnikih s HH. Genetska diagnostika HH je zahtevna zaradi velike heterogenosti, oligogenosti in nepopolne penetrance patoloških sprememb, kar je bilo ugotovljeno tudi pri naši skupini bolnikov.

• Language: Slovenian
• Keywords: hipogonadotropni hipogonadizem, Kallmannov sindrom, normozmični idiopatski hipogonadotropni hipogonadizem, sekvenciranje celotnega genoma
• Work type: Master's thesis/paper
• Organization: FFA - Faculty of Pharmacy
• Year: 2023
• PID: 20.500.12556/RUL-149360
• Publication date in RUL: 07.09.2023

Secondary language

• Language: English
• Title: Whole genome sequencing in patients with hypogonadotropic hypogonadism

Abstract

Hypogonadotropic hypogonadism (HH) is a heterogeneous disorder with a variable clinical presentation due to abnormal development of the olfactory placode, abnormalities in GnRH neurons migration and dysregulation of GnRH secretion or function. Congenital HH is divided into Kallmann syndrome (KS) and normosmic idiopathic HH (nIHH) according to the ability to detect smell. Identifying the genetic cause of HH is often challenging, as approximately 50 % of cases remain unexplained. We included 20 subjects from 18 families in whom genetic approaches used so far have not been successful in identification of genetic etiology. A clinical diagnosis of KS was made in 15 subjects and a diagnosis of nIHH was made in 5. Additional clinical features were present in 13 subjects and 12 subjects had a positive family history. Whole genome sequencing was performed in all subjects, followed by analysis of single nucleotide variants, small deletions, insertions, and copy number variations in 366 genes with the aim of explaining the genetic etiology. In subjects with available genome-wide data from family members, we also examined intronic regions of 34 genes known to be associated with HH. Sanger sequencing was used to confirm segregation of the identified genetic variants within families. Genetic etiology was explained in 30 % (6/20) of subjects and partially in another 45 % (9/20) of subjects. We identified nine novel genetic variants in the genes ANOS1, CCDC141, FGFR1, IGSF10, PROK2, SEMA3A, SEMA3G and SPRY4, all known HH-associated genes. We also identified a genetic variant in the candidate gene LGR4 which has not previously been associated with HH. In only two subjects, we were not able to identify genetic variants that may have contributed to the development of HH. In three subjects from the same family, we detected a deep intronic variant in the FGFR1 gene that may have contributed to their clinical presentation. However, identification of deep intronic variants is challenging based on the available data. To the best of our knowledge, deep intronic variants have not yet been identified in patients with HH. Genetic diagnosis in HH is challenging due to high genetic heterogeneity, oligogenity and incomplete penetrance of pathogenic variants all of which was also observed in our cohort.

• Keywords: hypogonadotropic hypogonadism, Kallmann syndrome, normosmic idiopathic hypogonadotropic hypogonadism, whole genome sequencing