20.500.12556/RUL-182445 Bioinformatična analiza rezultatov sekvenciranja naslednje generacije in analiza segregacije sprememb v genih CDON, CEP290, CFTR, CNTNAP2, NOTCH2 in TRPS1 v družinah z orofacialno shizo Bioinformatic analysis of next-generation sequencing results and segregation analysis of variants in the CDON, CEP290, CFTR, CNTNAP2, NOTCH2 and TRPS1 genes in families with orofacial clefts Orofacialne shize so najpogostejše prirojene nepravilnosti glave in vratu. Pojavijo se zgodaj v embrionalnem razvoju in se običajno delijo v dve večji skupini: razcep ustnice z ali brez razcepa neba in izoliran razcep neba. Posamezniki z orofacialnimi shizami se poleg estetskih težav pogosto soočajo s funkcionalnimi težavami, kot so težave pri hranjenju in govoru. Orofacialne shize so kompleksne prirojene nepravilnosti in čeprav se lahko pojavijo kot posledica monogenskih sprememb, so pogosteje multifaktorskega izvora, pri čemer na njihov razvoj vpliva kombinacija genetskih in okoljskih dejavnikov. V magistrski nalogi smo poskušali poiskati genetske vzroke za orofacialne shize v 25 družinah z družinsko anamnezo orofacialnih shiz neznanega vzroka. Pridobili smo podatke sekvenciranja celotnega eksoma za 28 indeksnih bolnikov z orofacialno shizo. Najprej smo z bioinformatično analizo rezultatov sekvenciranja celotnega eksoma v 40 kandidatnih genih preverjali genetske spremembe, ki bi lahko vplivale na pojav orofacialnih shiz. Skupno smo preverili 400 sprememb, pri čemer smo patološki potencial najdenih sprememb opredelili z orodjem Franklin (Genoox) glede na kriterije Ameriškega kolegija za medicinsko genetiko in genomiko. Med pregledanimi genetskimi polimorfizmi smo identificirali 55 sprememb nejasnega pomena, 4 so bile verjetno patogene, 2 spremembi pa sta bili patogeni. Prisotnost verjetno patogenih in patogenih sprememb pri indeksnih bolnikih in njihovih prizadetih in neprizadetih sorodnikih smo nato potrjevali z genotipizacijo s hidrolizirajočimi sondami ali s sekvenciranjem po Sangerju. Dodatno smo potrjevali še prisotnost dveh sprememb v genih NOTCH2 in TRPS1, ki so jih identificirali predhodno v sklopu magistrske naloge. Za spremembo rs376493409 v genu CEP290 smo dokazali, da bi lahko segregirala z bolezenskim fenotipom. Za spremembo rs73715573 v genu CFTR segregacije z bolezenskim fenotipom nismo mogli ne potrditi in ne ovreči. Za spremembi rs113993960 v CFTR in rs374564199 v CDON smo potrdili, da v pripadajočih družinah ne segregirata z bolezenskim fenotipom. Prisotnosti delecij v genih CEP290, CNTNAP2, NOTCH2 in TRPS1 pri indeksnih bolnikih in njihovih sorodnikih nismo potrdili. V genih CEP290, CFTR in NOTCH2 smo neodvisno od sekvenciranja celotnega eksoma odkrili še tri spremembe, ki pa naj bi bile benigne oz. verjetno benigne. Orofacial clefts are the most common congenital anomalies of the head and neck. They arise early in embryonic development and are typically classified into two major groups: cleft lip with or without cleft palate, and cleft palate only. In addition to aesthetic problems, individuals with orofacial clefts often face functional difficulties, such as problems with feeding and speech. Orofacial clefts are complex congenital anomalies and, although they may occur as a result of monogenic genetic variants, they are more often of multifactorial origin, influenced by a combination of genetic and environmental factors. In this master's thesis, we aimed to identify genetic causes of orofacial clefts in 25 families with a family history of orofacial clefts of unknown origin. We obtained whole exome sequencing data for 28 index patients with orofacial cleft. First, we performed a bioinformatic analysis of whole exome sequencing data, where we screened 40 candidate genes to identify genetic variants that could influence the development of orofacial clefts. In total, 400 variants were evaluated, and their clinical significance was assessed using the Franklin (Genoox) platform according to the criteria of the American College of Medical Genetics and Genomics. Among the identified genetic variants, 55 were classified as variants of uncertain significance, 4 as likely pathogenic variants, and 2 as pathogenic variants. The presence of likely pathogenic and pathogenic variants in index patients and their affected and unaffected relatives was then confirmed using genotyping with hydrolysis probes or Sanger sequencing. Additionally, we verified the presence of two variants in the NOTCH2 and TRPS1 genes, which had been identified previously in another master's thesis. For the variant rs376493409 in gene CEP290, we demonstrated the possibility that it could segregate with the disease phenotype. For the variant rs73715573 in gene CFTR, we were unable to confirm or exclude segregation with the disease phenotype. For the variants rs113993960 in CFTR and rs374564199 in CDON, we confirmed that the variants do not segregate with the disease phenotype in the respective families. We did not confirm the presence of deletions in the CEP290, CNTNAP2, NOTCH2, and TRPS1 genes in the index patients and their relatives. Independently of whole exome sequencing results, we identified three additional variants in the CEP290, CFTR, and NOTCH2 genes. However these variants are predicted to be benign or likely benign. orofacialne shize genetske spremembe bioinformatična analiza genotipizacija študija družin orofacial clefts genetic variants bioinformatic analysis genotyping family study true false false [Z. Janža] Slovenski jezik Angleški jezik Magistrsko delo/naloga 2026-05-12 08:45:24 2026-05-12 08:45:35 2026-05-14 08:14:49 0000-00-00 00:00:00 2026 0 0 0000-00-00 NiDoloceno NiDoloceno NiDoloceno 0000-00-00 0000-00-00 0000-00-00 1970-01-01 616.315‑007.254(043.2) 125314 277983747 32116.pdf 32116.pdf 1 F057BE92E5A02CC1290B9C1B1B2B653E 5cf0e4bf7dd9dc59e8757e4c1f629bee479a96864d53a1406ea0c1b691bd9bef e0384ffd-4dcd-11f1-b0ab-0050569b8976 https://repozitorij.uni-lj.si/Dokument.php?lang=slv&id=233708 Fakulteta za farmacijo 0 0 0