Identifikacija genetskih vzrokov orofacialne shize v družinah s pozitivno družinsko anamnezo z bioinformatično analizo rezultatov sekvenciranja naslednje generacije

Košir, Katja (avtor), Karas Kuželički, Nataša (mentor)

Orofacialne shize (OFC) so najpomembnejše prirojene napake glave in vratu. Delimo jih na razcep ustnice, čeljustnega grebena in neba. Poznamo sindromske OFC, ki imajo monogenski vzrok nastanka in nesindromske OFC, ki imajo lahko večfaktorsko etiologijo. Po kliničnem pomenu jih delimo na heiloshize, gnatoshize, palatoshize in najtežjo obliko, ki združuje vse zgoraj omenjene, heilognatopalatoshize. Iz predhodne študije družin z večgeneracijsko OFC v sklopu doktorata z naslovom Genetske spremembe, povezane z orofacialnimi shizami v slovenski populaciji, je ostalo 25 družin, kjer po pregledu 75 kandidatnih genov niso ugotovili vzroka za OFC. Namen magistrske naloge je bil analizirati podatke sekvenciranja naslednje generacije 28 indeksnih bolnikov iz 25 družin z družinsko anamnezo OFC neznanega vzroka. Genetski vzrok smo poskušali identificirati s pregledom 43 kandidatnih genov: ABCA3, ABCC6, COG1, COL11A1, COL1A2, COL6A2, CTNND1, ECEL1, ERCC2, FBN1, GDF11, GJB6, GRIN2A, IDUA, IFT122, IFT140, IFT172, IKBKG, LRP6, KMT2A, KMT2D, MCPH1, NKX3-2, NOTCH2, PALB2, PCGF2, PGM1, PKLR, PROKR2, RPGRIP1L, SEC23A, SF3B4, SH3PXD2B, SOX9, SRCAP, SZT2, TBX1, TFR2, TNNT3, TP63, TRPS1, TUBB2B in UBE3B. Kandidatne gene, povezane z nesindromskimi OFC, smo izbrali na osnovi pregleda dveh znanstvenih člankov. Posledice odkritih sprememb v kandidatnih genih smo pregledali s pomočjo platforme Franklin (Genoox). Vsako genetsko spremembo smo ovrednotili s kriteriji ACMG in s programom Progeny narisali družinska drevesa preučevanih družin. Upoštevali smo spremembe, ki izpolnjujejo pogoj kvalitete genotipizacije, so v splošni populaciji redkejše od 1 % in so z večino bioinformatičnih napovednih modelov ugotovljene kot spremembe nejasnega pomena, verjetno patogene ali patogene. Po analizi sekvenciranja naslednje generacije rezultatov indeksnih bolnikov smo odkrili 34 redkih genetskih sprememb, ki se nahajajo v 22 različnih genih. Izmed 34 odkritih sprememb so bile po smernicah ACMG tri klasificirane kot verjetno patogene, ostale kot spremembe nejasnega pomena. Tri preučevane družine so imele po dva indeksna bolnika. Samo v eni družini (OFC_182) pa smo odkrili spremembo, ki se je pojavila pri obeh indeksnih bolnikih in segregira z bolezenskim fenotipom. Gre za intronsko spremembo v genu IDUA. Dve družini (OFC_185 in OFC_186) sta v tesnem sorodstvu. Pri njih smo odkrili drugačnosmiselno spremembo v genu ERCC2, ki segregira z bolezenskim fenotipom v obeh družinah.

• Jezik: Slovenski jezik
• Ključne besede: bioinformatična analiza, orofacialne shize, genetske spremembe, segregacija, študija družin
• Vrsta gradiva: Magistrsko delo/naloga
• Organizacija: FFA - Fakulteta za farmacijo
• Leto izida: 2024
• PID: 20.500.12556/RUL-165910
• Datum objave v RUL: 13.12.2024

Sekundarni jezik

• Jezik: Angleški jezik
• Naslov: Identification of genetic causes of orofacial clefts in families with a positive family history using bioinformatic analysis of next-generation sequencing results

Izvleček

Orofacial clefts (OFC) are the most common congenital defects of the head and neck. They are divided into syndromic OFCs, with a monogenic cause, and non-syndromic OFCs, which can have a multifactorial etiology. Clinically, they are classified as cleft lip, cleft alveolus, cleft palate, and the most severe form, which combines all of the above, cleft lip, alveolus, and palate. A previous study Genetic Variants Associated with Orofacial Clefts in the Slovenian Population, was unable to identify the genetic cause of OFC in 25 families. The purpose of this master's thesis is to analyze next-generation sequencing data from 28 index patients from 25 families with a family history of OFC of unknown origin. We tried to identify the genetic cause by examining 43 candidate genes: ABCA3, ABCC6, COG1, COL11A1, COL1A2, COL6A2, CTNND1, ECEL1, ERCC2, FBN1, GDF11, GJB6, GRIN2A, IDUA, IFT122, IFT140, IFT172, IKBKG, LRP6, KMT2A, KMT2D, MCPH1, NKX3-2, NOTCH2, PALB2, PCGF2, PGM1, PKLR, PROKR2, RPGRIP1L, SEC23A, SF3B4, SH3PXD2B, SOX9, SRCAP, SZT2, TBX1, TFR2, TNNT3, TP63, TRPS1, TUBB2B, and UBE3B. Candidate genes associated with non-syndromic OFCs were selected based on a review of two scientific articles. The functional consequences of the discovered changes in candidate genes were examined using the platform Franklin (Genoox). Each genetic variant was evaluated using ACMG criteria, and family trees of the investigated families were drawn using the Progeny program. We considered variants that met the criteria for genotype quality, were rarer than 1% in the general population, and were identified by most predictive models as a variant of uncertain significance, likely pathogenic, or pathogenic. After analyzing the results of the next-generation sequencing of index patients, we discovered 34 rare genetic variants in 22 genes. Among the 34 discovered variants, three were classified as likely pathogenic according to ACMG guidelines, while the rest were classified as variants of uncertain significance. Three of the investigated families had two index patients but only in one such family (OFC_182) a candidate variant was present in both index patients, meaning it segregates with OFC within the family. This was an intronic mutation in the IDUA gene. Two families (OFC_185 and OFC_186) were closely related, and we discovered a missense mutation in the ERCC2 gene that segregated with the disease phenotype in both families.

• Ključne besede: bioinformatic analysis, orofacial clefts, genetic variants, segregation, family study