Laboratorij za medicinsku genetiku / Laboratory for medical genetics

Voditelj:
Prof.dr.sc. Nina Canki-Klain
Pedijatar-Genetičar
Katedra za medicinsku Biologiju
Hrvatski institut za istraživanje mozga
Šalata 12, 10 000, Zagreb
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Suradnici:
Prof.dr.sc. Floriana Bulić-Jakuš
Prof.dr.sc. Sven Seiwerth
Dr.sc. Astrid Milić

Istraživački interes laboratorija predstavljaju bazična istraživanja nastanka te molekularni aspekti najčešćih monogenskih mišićnih i neuroloških poremećaja s naglaskom na epidemiološko stanje u Hrvatskoj. Ti podaci su neophodni. kako bi se pronašle usavršene mjere za sprječavanje , poboljšanje ili liječenje tih bolesti. Diferencijalna dijagnoza spomenutih poremećaja zahtijeva pomnu primjenu širokog opsega znanja: kliničku procjenu, istraživanje rodoslovlja, imunoblot analizu s upotrebom panela protutijela te opsežne molekularne genetske analize. Rad laboratorija je započeo sa širokim rasponom pristupa 1996. Bolesnici su se do 2009.odabirali u Neurogenetskoj ambulanti (Voditelj:Prof.dr. Nina Canki-Klain), Neurološka klinika, Klinički bolnički centar Zagreb. Danas se rad nastavlja u Neurogenetskoj ambulanti Hrvatskog Instituta za istraživanje mozga u okviru Medicinskog fakulteta Sveučilišta u Zagrebu.
Laboratorij je otkrio spektar mutacija nekih od najčešćih mišićnih bolesti u Hrvatskoj. Isto tako, radi se o laboratoriju koji jedini u zemlji obavlja slijedeće analize:

• Analiza genetskog povezivanja:
• Autosomnih recesivnih pojasnih mišićnih distrofija (LGMD2):
• Tip 2A – CAPN3 gen (Erbov juvenilni oblik mišićne distrofije ili kalpainopatija (LGMD2A) (Slika 1.)
• Tip 2B – DYSFgen (disferlinopatija / Miyoshyeva miopatija (LGMD2B) (Slika 2.)
• Tip2C – SGCG (gamasarkoglikanopatija) (LGMD2C)
• Tip 2D - SGCA gen (alfasarkoglikanopatija) (LGMD2D)
• Tip 2I – FKRPgen (pojasna mišićna distrofija LGMD2I (Slika 3a. i kongenitalna miš. distrofija MDC1D)
• Okulofaringealnemišićnedistrofije(PABP2gen) (OPMD) (Slika 4. i 5.)
• Direktna analiza
• Analiza šestmutacijaCAPN3: 550delA idelFWSAL (egzon 4); P82LiR49H (egzon 1); R541WiY537 (egzon 13)kojepokrivaju 96% CAPN3 kromosomaunašojpopulaciji. (Tablica 1.)
• Analiza C826 -najčešće mutacije FKRP gena koja uzrokuje LGMD2I: (Slika 3b.
• Analiza PABP2 gena uzročnika okulofaringealne mišićne distrofije (OPMD) (Slika 4.)
• Western blot analiza iz uzorka mišića:
• DMD gen (distrofin)
• CAPN3 (kalpain 3)
• SGCG (γ-sarkoglikan)
• SGCA (α-sarkoglikan)
• DYSF (disferlin).
• Neinvazivna Western blot analiza disferlina (DYSF)iz uzorka monocita periferne krvi (Slika 6.)

Projekt u tijeku:

• Genetika, priroda i epidemiologija značajnijih živčanih i mišićnih bolesti (ŽMB), Projetk br. 010800000003435 /2008 MZOS RHrvatske. Leader: Nina Canki-Klain

Završeni projekti napravljeni u laboratoriju

• Nacionalnienetic and epidemiological study of muscular dystrophy in Croatia. Grant No 0108052/2002 from the Ministry of Science, Education, and Sports Republic of Croatia
• Genetic and epidemiological study of muscular dystrophy in Croatia. Grant No 108041/1988 MZOŠ of Croatia

Bilateralni francusko-hrvatski

• COGITO 2004/5 :L’étude clinico-génétique des dystrophies musculaires des ceintures (LGMD) en Croatie: analyse comparative des différents types. INSERM U 567, Paris, France, Pr. Jamel Chelly, direktor/ Pr.Nina Canki-Klain, haed, Zagreb University Medical School
• COGITO 2003/4: Correlation phénotype-génotype dans les dystrophies musculaires de l'enfant et de l'adulte. INSERM, U 582, Paris, France, Dr.Pascale GUICHENEY,direktor/ Pr.Nina Canki-Klain, haed, Zagreb University Medical School

Međunarodni

• ECO-NET 2004/5 Approche multiparamètrique pour la connaissance de l'histoire naturelle des calpainopathies visant à déterminer l'efficacité de thérapies potentielles, Généthon, UMR8115 ,CNRS , Evry, France Dr.Isabelle Richard
• AFM, MNM1 1999-Funding-groupe 4,No 7233 Canki-Klain N, Milić A, Zurak N., Šoštarko M., Pažanin L., Bonne G., Urtizberea J.A., Kaplan J.-C, Leturcq F., Recan D. Clinical and genetic study of limb-Girdle muscular dystrophies in Croatia.

Head:
Prof.dr.sc. Nina Canki-Klain
Pediatric geneticist
Department of Medical Biology
Croatian Institute for Brain Research
Šalata 12, 10 000, Zagreb
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Associates:
Prof.dr.sc. Floriana Bulić-Jakuš
Prof.dr.sc. Sven Seiwerth
Dr.sc. Astrid Milić

Laboratory research interests are basic research of formation and molecular aspects of the most common monogenic muscle and neurological disorders with an emphasis on the epidemiological situation in Croatia. These data are essential in order to find a sophisticated measures to prevent, improve or treat these diseases. The differential diagnosis of these disorders requires a careful application of broad-based knowledge: clinical assessment, genealogy research, immunoblot analysis with the use of wide range of antibodies and extensive molecular genetic analysis. The work of the laboratory began with a wide range of access in 1996. Until 2009. patients were selected in Neurogeneticist's office (Head: Prof.dr.sc. Nina Canki-Klain), Department of Neurology, University Hospital Center Zagreb. Today, work continues in Neurogeneticist's office of Croatian Institute for Brain Research at the Medical School, University of Zagreb.
Laboratory discovered spectrum of mutations of some of the most common muscle diseases in Croatia. Also, it is the only lab in the country that performs the following analysis:

• Genetic linkage analysis:
• Limb-girdle muscular dystrophy (LGMD2):
• Type 2A – CAPN3 gene (Erb's muscular dystrophy or calpainopathy (LGMD2A) (Figure 1.)
• Type 2B – DYSF gene (dysferlinopathy / Miyoshy myopathy (LGMD2B) (Figure 2.)
• Type2C – SGCG (gamma-sarcoglycanopathy) (LGMD2C)
• Type 2D - SGCA gene (alphasarcoglycanopathy) (LGMD2D)
• Type 2I – FKRP gene (limb-girdle muscular dystrophy)(LGMD2I) (Figure 3a.) and congenital muscular dystrophy (MDC1D)
• Oculopharyngeal muscular dystrophy (PABP2gen) (OPMD) (Figure 4. i 5.)
• Direct analysis:
• Analysis of six mutations of CAPN3: 550 delA and delFWSAL (exon 4); P82LiR49H (exon 1); R541WiY537 (exon 13) that make 96% CAPN3 chromosomes in our population (Table 1.)
• Analysis of C826 –most common mutation of FKRP gene that causes LGMD2I: (Figure 3b.)
• Analysis of PABP2 gene that causes oculopharyngeal muscular dystrophy (OPMD) (Figure 4.)
• Western blot analysis from muscle tissue samples:
• DMD gene (dystrophin)
• CAPN3 (calpain 3)
• SGCG (γ-sarcoglycan)
• SGCA (α-sarcoglycan)
• DYSF (dysferlin).
• Noninvasive Western blot dysferline analysis (DYSF) from peripheral blood monocytes (Figure 6.)

Current project:

• Genetics, nature and epidemiology of major nervous and muscular disorders Grant number 010800000003435 /2008 MZOS. Leader: Nina Canki-Klain

Completed research grants

• Nacionalnienetic and epidemiological study of muscular dystrophy in Croatia. Grant No 0108052/2002 from the Ministry of Science, Education, and Sports Republic of Croatia

Genetic and epidemiological study of muscular dystrophy in Croatia. Grant No 108041/1988 MZOŠ of Croatia

Bilateral french-croatian

• COGITO 2004/5 :L’étude clinico-génétique des dystrophies musculaires des ceintures (LGMD) en Croatie: analyse comparative des différents types. INSERM U 567, Paris, France, Pr. Jamel Chelly, direktor/ Pr.Nina Canki-Klain, haed, Zagreb University Medical School
• COGITO 2003/4: Correlation phénotype-génotype dans les dystrophies musculaires de l'enfant et de l'adulte. INSERM, U 582, Paris, France, Dr.Pascale GUICHENEY,direktor/ Pr.Nina Canki-Klain, haed, Zagreb University Medical School

International

• ECO-NET 2004/5 Approche multiparamètrique pour la connaissance de l'histoire naturelle des calpainopathies visant à déterminer l'efficacité de thérapies potentielles, Généthon, UMR8115 ,CNRS , Evry, France Dr.Isabelle Richard
• AFM, MNM1 1999-Funding-groupe 4,No 7233 Canki-Klain N, Milić A, Zurak N., Šoštarko M., Pažanin L., Bonne G., Urtizberea J.A., Kaplan J.-C, Leturcq F., Recan D. Clinical and genetic study of limb-Girdle muscular dystrophies in Croatia.

• Milić A, Malnar M, Canki-Klain N. Non-invasive protein analysis in the first Translational Neuroscience • 2(3) • 2011 • 241-245
• Alfirević-Ungarov T. Canki-Klain N,. Handžić J,. Jokić-Begić N, Kovač I,. Kovač V,. Richard J.L.F. LEMMERS, OZRETIĆ D, Sinanović O, Stiglmayer N, Štifter S, Vranješ D,. Vukojević N Novosti u kliničkoj genetici:Molekularna dijagnostika, terapija i prevencija facioskapulohumeralne mišićne distrofije (FSHD) Canki-Klain , Nina (ur.). Best copy, Zagreb, 2011, 116 str.
• Alfirević-Ungarov, Taida; Bilić, Erivna; Canki-Klain, Nina; Guicheney, Pascale; Kosi, Damir; Kovač, Biserka; Kovač, Branislav; Milić, Astrid; Mitrović, Zoran; Sinanović, Osman; Šoštarko, Marija; Vranješ, Davorka. Novosti u kliničkoj genetici: genetske mišićne bolesti u svakodnevnoj praksi / Canki-Klain , Nina (ur.). Zagreb : Kolding, 2004., 161 str.
• Canki-Klain, Nina; Fardeau, Michel; Jurčić, Dragan; Jurić, Gordana; Kovač, Ida; Ljubanović, Danica; Milić, Astrid; Sinanović, Osman; Stiglmayer, Neda; Tocilj-Šimunković, Gorana; Večerina, Santa; Vranješ, Davorka; Žagar, Marija. Novosti u kliničkoj genetici: multidisciplinarni pristup dijagnostici, terapiji i prevenciji okulofarinegealne mišićne distrofije (OPMD) / Canki-Klain , Nina (ur.). Zagreb : Kolding, 2004. 99str.
• Bushby, Kate; Canki-Klain, Nina; Stiglmayer, Neda; Fučić, Aleksandra; Jurić, Gordana; Mitrović, Zoran; Milić, Astrid. Novosti u kliničkoj genetici: dijagnostički postupci u bolesnika s kongenitalnim miopatijama i mišićnim distrofijama / Canki-Klain , Nina (ur.).Zagreb : Kolding, 2003., 74 str.
• Milić A, Canki-Klain N. Calpainopathy (LGMD2A) in Croatia: Molecular and Haplotype Analysis. Croat Med J 2005; 46/4: 657-663.
• Piluso G, Politano L, Aurino S, Fanin M, Ricci E, Ventriglia VM, Belsito A, Totaro A, Saccone V, Topaloglu H, Nascimbeni AC, Fulizio L,Brocetolini A, Canki-Klain N, Comi Li, Nigro G, Angelini C, Nigro V.Extensive scanning of the calpain-3 gene broadens the spectrum of LGMD2A phenotypes. J Med Genet 2005; 42:686-613.
• Nina Canki-Klain, Astrid Milic, Biserka Kovac, Anuska Trlaja, Damir Grgicevic, Niko Zurak, Michel Fardeau, France Leturcq,Jean-Claude Kaplan, J Andoni Urtizberea, Luisa Politano , and Josue Feingold.Prevalence of the 550delA mutation in calpainopathy (LGMD 2A) in Croatia. Am J Med Genet 2004; 125A: 152-156.
• Hecimovic S, Klepac N, Vlasic J, Vojta A, Janko D, Skarpa-Prpic I I, Canki-Klain N, Markovic D, Bozikov J, Relja M, Pavelic K.Genetic background of Huntington disease in Croatia: Molecular analysis of CAG, CCG, and Delta2642 (E2642del) polymorphisms. Hum Mutat 2002; 20/3:233-40.
• N.Canki-Klain, D.Recan, D.Milicic, S.Llense ,F.Leturcq, N.Deburgrave, J.C.Kaplan, M.Debevec, N.Zurak: Clinical Variability and Molecular Diagnosis in a Four-generation Family with X-linked Emery-Dreifuss Muscular Dystrophy. Croat Med J 2000; 41: 389-395.
• Canki N, Tivadar I, Župančič N, Debevec M. Citogenetska študija sedmih bolnic z ataksijo-teleangiektazijo. Zdrav Vestn 1983; 52: 567-70.
• Canki N, Dutrillaux B, Tivadar I. Dystrophie musculaire de Duchenne chez une petite fille porteuse d'une translocation t(X;3)(p21;q13) de novo. Ann Génét 1979; 22: 33-9.