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出境医 / 临床实验 / Lipa Gene Mutation in PED-LIPIGEN (Pediatric FH Subjects)

Lipa Gene Mutation in PED-LIPIGEN (Pediatric FH Subjects)

Study Description
Brief Summary:

Familial Hypercholesterolemia (FH) is a monogenic autosomal dominant disease also known as Autosomal Dominant Hypercholesterolemia - ADH) that leads to dramatically increased levels of Low Density Lipoprotein (LDL) and total cholesterol associated to tendon xanthomas, xanthelasma, corneal arcus, premature atherosclerosis and to an increased risk of coronary artery disease (CAD) and myocardial infarction.

FH is mainly caused by mutations in genes encoding for proteins affecting hepatic LDL cholesterol uptake including the LDL receptor (LDLR) gene or the gene encoding the only apolipoprotein of LDL, the apolipoprotein B (APOB), or the gene encoding a protease regulating LDLR levels on the cell membrane Lysosomal Acid Lipase A (LIPA) gene encode for Lysosomal acid lipase (LAL) enzyme responsible for hydrolyzing cholesterol esters and triglycerides that are delivered to lysosomes. Mutations in LIPA that completely inactivate LAL are the molecular cause of Wolman disease, a rapidly lethal disease of infancy while mutations in LIPA that result in residual enzymatic activity of LAL are responsible of a disorder characterized by a less severe phenotype known as cholesterol ester storage disease (CESD). Patients with CESD usually show a phenotype characterized by hepatic disease and mixed hyperlipidemia with elevated levels of LDL-C and triglycerides (TG) and decreased HDL-C levels.

A broader phenotypic presentation for loss of function mutations in LIPA suggests that LIPA mutations may be considered in patients with apparently monogenic FH in whom mutations in the known candidate genes are not detectable.

The project is aimed to evaluate the prevalence and the mutation rate of LIPA gene in subjects with a clinical diagnosis of FH and already genetically characterized in whom pathogenic mutations in the known candidate genes have not been identified. The analysis will be performed in about 250 FH pediatric subjects and putative causal mutations will be also tested for co-segregation in available families in affected and unaffected members.


Condition or disease Intervention/treatment
Lysosomal Acid Lipase Deficiency Other: Observational study

Detailed Description:

Lysosomal acid lipase (LAL) is encoded by LIPA gene located on chromosome 10q23.3-q23 and consists of 10 exons. LIPA mRNA (messenger RiboNucleic Acid) (GenBank accession number NM_000235) is 2782 bp long and encodes a mature protein of 375 residues (GenBank accession number NP_000226). The sequencing of all 10 exons of LIPA gene will consist of 10 PCR (Polymerase Chain Reaction) amplification reactions (for the 10 exons and the proximal promoter) followed by 20 sequence reactions (forward and reverse sequencing) with appropriate primers designed to include the intron-exon boundaries. This analysis will be performed in about 250 FH pediatric subjects as specified in project description.

The sequencing work will be performed taking advantage of 2 automated 8 capillaries automated DNA Sequencer (3500 Genetic Analyzer, Thermo Fisher Scientific, Monza, Italy) currently available in the laboratory of the Units involved in the project.

In case of identification of unreported sequence variants, the presence of these mutations will be assessed in a sample of at least 100 normolipidemic subjects of the population, in order to define whether the nucleotide changes are rare sequence variations (with a putative functional effect) or represent common polymorphisms. In case of finding of rare variants in the coding regions, an in silico analysis will be performed by using two different softwares (Polyphen, http://genetics.bwh.harvard.edu/pph/ and Panther, http://www.pantherdb.org/) to predict the putative damaging role of the mutations on the protein. In case of intronic variants, the specifically designed software Automated Splice Site Analysis will be applied (https://www.splice.uwo.ca/).

Putative causal mutations will be also tested for co-segregation in available families in affected and unaffected members.

In order to test the effect of variants on enzyme activity LAL-activity will be assayed with dried blood spot (DBS) technique using the inhibitors Lalistat 2 in carriers and non carriers of these mutations belonging to available kindred.

Study Design
Layout table for study information
Study Type : Observational
Estimated Enrollment : 1000 participants
Observational Model: Other
Time Perspective: Retrospective
Official Title: Prevalence and Mutation Rate of Lipa Gene in LIPIGEN Subjects With Clinical Diagnosis of FH
Actual Study Start Date : September 1, 2017
Estimated Primary Completion Date : July 1, 2021
Estimated Study Completion Date : July 1, 2021
Arms and Interventions
Group/Cohort Intervention/treatment
FH pediatric patients
1000 clinically diagnosed FH pediatric patients (age <18 years) included in the LIPIGEN (Lipid TransPort Disorders italian Genetic Network) database
 Other: Observational study
Observational study: There is no intervention.
Outcome Measures
Primary Outcome Measures :
  1. Prevalence of patients with mutations of LIPA gene among clinically diagnosed FH subjects [ Time Frame: 2 years from start of the study ]
    Percentage of patients with at least one mutation of LIPA gene among clinically diagnosed FH subjects according to a "Dutch Lipid Clinic Network" score of 6 or above


Secondary Outcome Measures :
  1. Frequency of specific mutations of LIPA gene among clinically diagnosed FH subjects [ Time Frame: 2 years from start of the study ]
    Numbers of patients carrying specific mutations of LIPA gene among clinically diagnosed FH subjects for each mutation identified by sequencing of all 10 exons of LIPA gene.


Biospecimen Retention:   Samples With DNA

Venous blood samples were taken after 12 hours of fasting. Serum total cholesterol, triglycerides and HDL-cholesterol levels were measured in a centralized laboratory using enzymatic methods.

DNA samples, serum, plasma and whole blood were aliquoted and preserved at -80°C.


Eligibility Criteria
Layout table for eligibility information
Ages Eligible for Study:   up to 18 Years   (Child, Adult)
Sexes Eligible for Study:   All
Accepts Healthy Volunteers:   No
Sampling Method:   Non-Probability Sample
Study Population
Clinically diagnosed FH pediatric patients (age <18 years) included in the Lipid TransPort Disorders italian Genetic Network (LIPIGEN) database, already genetically characterized.
Criteria

Inclusion Criteria:

  • Pediatric subjects (<18 years old) with a clinical diagnosis of FH and without identified pathogenic mutations in the known candidate genes.

Exclusion Criteria:

  • Subjects with a clinical diagnosis of FH with identified pathogenic mutations in the known candidate genes.
Contacts and Locations

Contacts
Layout table for location contacts
Contact: Alberico L Catapano alberico.catapano@unimi.it
Contact: Alessia Tincani +39026173276 catapano.centroatero@gmail.com

Locations
Layout table for location information
Italy
CENTRO PER LO STUDIO DELL'ATEROSCLEROSI - Ospedale Bassini Recruiting
Cinisello Balsamo, Mi, Italy
Contact: Alberico L Catapano       alberico.catapano@unimi.it   
Laboratorio di biochimica delle lipoproteine - DIPARTIMENTO DI SCIENZE BIOMEDICHE Recruiting
Modena, Italy
Contact: Patrizia M. Tarugi       patriziamaria.tarugi@unimore.it   
Centro Di Riferimento Regionale Per La Prevenzione, Diagnosi E Cura Delle Malattie Rare Del Metabolismo Recruiting
Palermo, Italy
Contact: Maurizio Averna       maurizio.averna@unipa.it   
Centro Per L'Arteriosclerosi Dipartimento Di Medicina Interna E Specialità Mediche Recruiting
Roma, Italy
Contact: Marcello Arca       marcello.arca@uniroma1.it   
Sponsors and Collaborators
Fondazione SISA (Societa Italiana per lo Studio della Arteriosclerosi)
Investigators
Layout table for investigator information
Study Director: Maurizio Averna Fondazione SISA
Tracking Information
First Submitted Date May 21, 2019
First Posted Date June 12, 2019
Last Update Posted Date July 23, 2020
Actual Study Start Date September 1, 2017
Estimated Primary Completion Date July 1, 2021   (Final data collection date for primary outcome measure)
Current Primary Outcome Measures
 (submitted: June 11, 2019) 		Prevalence of patients with mutations of LIPA gene among clinically diagnosed FH subjects [ Time Frame: 2 years from start of the study ]
Percentage of patients with at least one mutation of LIPA gene among clinically diagnosed FH subjects according to a "Dutch Lipid Clinic Network" score of 6 or above
Original Primary Outcome Measures Same as current
Change History
Current Secondary Outcome Measures
 (submitted: June 11, 2019) 		Frequency of specific mutations of LIPA gene among clinically diagnosed FH subjects [ Time Frame: 2 years from start of the study ]
Numbers of patients carrying specific mutations of LIPA gene among clinically diagnosed FH subjects for each mutation identified by sequencing of all 10 exons of LIPA gene.
Original Secondary Outcome Measures Same as current
Current Other Pre-specified Outcome Measures Not Provided
Original Other Pre-specified Outcome Measures Not Provided
 
Descriptive Information
Brief Title Lipa Gene Mutation in PED-LIPIGEN (Pediatric FH Subjects)
Official Title Prevalence and Mutation Rate of Lipa Gene in LIPIGEN Subjects With Clinical Diagnosis of FH
Brief Summary

Familial Hypercholesterolemia (FH) is a monogenic autosomal dominant disease also known as Autosomal Dominant Hypercholesterolemia - ADH) that leads to dramatically increased levels of Low Density Lipoprotein (LDL) and total cholesterol associated to tendon xanthomas, xanthelasma, corneal arcus, premature atherosclerosis and to an increased risk of coronary artery disease (CAD) and myocardial infarction.

FH is mainly caused by mutations in genes encoding for proteins affecting hepatic LDL cholesterol uptake including the LDL receptor (LDLR) gene or the gene encoding the only apolipoprotein of LDL, the apolipoprotein B (APOB), or the gene encoding a protease regulating LDLR levels on the cell membrane Lysosomal Acid Lipase A (LIPA) gene encode for Lysosomal acid lipase (LAL) enzyme responsible for hydrolyzing cholesterol esters and triglycerides that are delivered to lysosomes. Mutations in LIPA that completely inactivate LAL are the molecular cause of Wolman disease, a rapidly lethal disease of infancy while mutations in LIPA that result in residual enzymatic activity of LAL are responsible of a disorder characterized by a less severe phenotype known as cholesterol ester storage disease (CESD). Patients with CESD usually show a phenotype characterized by hepatic disease and mixed hyperlipidemia with elevated levels of LDL-C and triglycerides (TG) and decreased HDL-C levels.

A broader phenotypic presentation for loss of function mutations in LIPA suggests that LIPA mutations may be considered in patients with apparently monogenic FH in whom mutations in the known candidate genes are not detectable.

The project is aimed to evaluate the prevalence and the mutation rate of LIPA gene in subjects with a clinical diagnosis of FH and already genetically characterized in whom pathogenic mutations in the known candidate genes have not been identified. The analysis will be performed in about 250 FH pediatric subjects and putative causal mutations will be also tested for co-segregation in available families in affected and unaffected members.
Detailed Description

Lysosomal acid lipase (LAL) is encoded by LIPA gene located on chromosome 10q23.3-q23 and consists of 10 exons. LIPA mRNA (messenger RiboNucleic Acid) (GenBank accession number NM_000235) is 2782 bp long and encodes a mature protein of 375 residues (GenBank accession number NP_000226). The sequencing of all 10 exons of LIPA gene will consist of 10 PCR (Polymerase Chain Reaction) amplification reactions (for the 10 exons and the proximal promoter) followed by 20 sequence reactions (forward and reverse sequencing) with appropriate primers designed to include the intron-exon boundaries. This analysis will be performed in about 250 FH pediatric subjects as specified in project description.

The sequencing work will be performed taking advantage of 2 automated 8 capillaries automated DNA Sequencer (3500 Genetic Analyzer, Thermo Fisher Scientific, Monza, Italy) currently available in the laboratory of the Units involved in the project.

In case of identification of unreported sequence variants, the presence of these mutations will be assessed in a sample of at least 100 normolipidemic subjects of the population, in order to define whether the nucleotide changes are rare sequence variations (with a putative functional effect) or represent common polymorphisms. In case of finding of rare variants in the coding regions, an in silico analysis will be performed by using two different softwares (Polyphen, http://genetics.bwh.harvard.edu/pph/ and Panther, http://www.pantherdb.org/) to predict the putative damaging role of the mutations on the protein. In case of intronic variants, the specifically designed software Automated Splice Site Analysis will be applied (https://www.splice.uwo.ca/).

Putative causal mutations will be also tested for co-segregation in available families in affected and unaffected members.

In order to test the effect of variants on enzyme activity LAL-activity will be assayed with dried blood spot (DBS) technique using the inhibitors Lalistat 2 in carriers and non carriers of these mutations belonging to available kindred.
Study Type Observational
Study Design Observational Model: Other
Time Perspective: Retrospective
Target Follow-Up Duration Not Provided
Biospecimen Retention:   Samples With DNA
Description:

Venous blood samples were taken after 12 hours of fasting. Serum total cholesterol, triglycerides and HDL-cholesterol levels were measured in a centralized laboratory using enzymatic methods.

DNA samples, serum, plasma and whole blood were aliquoted and preserved at -80°C.
Sampling Method Non-Probability Sample
Study Population Clinically diagnosed FH pediatric patients (age <18 years) included in the Lipid TransPort Disorders italian Genetic Network (LIPIGEN) database, already genetically characterized.
Condition Lysosomal Acid Lipase Deficiency
Intervention Other: Observational study
Observational study: There is no intervention.
Study Groups/Cohorts FH pediatric patients
1000 clinically diagnosed FH pediatric patients (age <18 years) included in the LIPIGEN (Lipid TransPort Disorders italian Genetic Network) database
Intervention: Other: Observational study
Publications *
  • ABRAMOV A, SCHORR S, WOLMAN M. Generalized xanthomatosis with calcified adrenals. AMA J Dis Child. 1956 Mar;91(3):282-6.
  • Bernstein DL, Hülkova H, Bialer MG, Desnick RJ. Cholesteryl ester storage disease: review of the findings in 135 reported patients with an underdiagnosed disease. J Hepatol. 2013 Jun;58(6):1230-43. doi: 10.1016/j.jhep.2013.02.014. Epub 2013 Feb 26. Review.
  • Bertolini S, Pisciotta L, Rabacchi C, Cefalù AB, Noto D, Fasano T, Signori A, Fresa R, Averna M, Calandra S. Spectrum of mutations and phenotypic expression in patients with autosomal dominant hypercholesterolemia identified in Italy. Atherosclerosis. 2013 Apr;227(2):342-8. doi: 10.1016/j.atherosclerosis.2013.01.007. Epub 2013 Jan 19.
  • Burke JA, Schubert WK. Deficient activity of hepatic acid lipase in cholesterol ester storage disease. Science. 1972 Apr 21;176(4032):309-10.
  • Futema M, Plagnol V, Li K, Whittall RA, Neil HA, Seed M; Simon Broome Consortium, Bertolini S, Calandra S, Descamps OS, Graham CA, Hegele RA, Karpe F, Durst R, Leitersdorf E, Lench N, Nair DR, Soran H, Van Bockxmeer FM; UK10K Consortium, Humphries SE. Whole exome sequencing of familial hypercholesterolaemia patients negative for LDLR/APOB/PCSK9 mutations. J Med Genet. 2014 Aug;51(8):537-44. doi: 10.1136/jmedgenet-2014-102405. Epub 2014 Jul 1.
  • Hamilton J, Jones I, Srivastava R, Galloway P. A new method for the measurement of lysosomal acid lipase in dried blood spots using the inhibitor Lalistat 2. Clin Chim Acta. 2012 Aug 16;413(15-16):1207-10. doi: 10.1016/j.cca.2012.03.019. Epub 2012 Mar 29.
  • Hopkins PN, Toth PP, Ballantyne CM, Rader DJ; National Lipid Association Expert Panel on Familial Hypercholesterolemia. Familial hypercholesterolemias: prevalence, genetics, diagnosis and screening recommendations from the National Lipid Association Expert Panel on Familial Hypercholesterolemia. J Clin Lipidol. 2011 Jun;5(3 Suppl):S9-17. doi: 10.1016/j.jacl.2011.03.452. Epub 2011 Apr 3.
  • Nordestgaard BG, Chapman MJ, Humphries SE, Ginsberg HN, Masana L, Descamps OS, Wiklund O, Hegele RA, Raal FJ, Defesche JC, Wiegman A, Santos RD, Watts GF, Parhofer KG, Hovingh GK, Kovanen PT, Boileau C, Averna M, Borén J, Bruckert E, Catapano AL, Kuivenhoven JA, Pajukanta P, Ray K, Stalenhoef AF, Stroes E, Taskinen MR, Tybjærg-Hansen A; European Atherosclerosis Society Consensus Panel. Familial hypercholesterolaemia is underdiagnosed and undertreated in the general population: guidance for clinicians to prevent coronary heart disease: consensus statement of the European Atherosclerosis Society. Eur Heart J. 2013 Dec;34(45):3478-90a. doi: 10.1093/eurheartj/eht273. Epub 2013 Aug 15.
  • Patrick AD, Lake BD. Deficiency of an acid lipase in Wolman's disease. Nature. 1969 Jun 14;222(5198):1067-8.
  • Risk of fatal coronary heart disease in familial hypercholesterolaemia. Scientific Steering Committee on behalf of the Simon Broome Register Group. BMJ. 1991 Oct 12;303(6807):893-6.
  • Stitziel NO, Fouchier SW, Sjouke B, Peloso GM, Moscoso AM, Auer PL, Goel A, Gigante B, Barnes TA, Melander O, Orho-Melander M, Duga S, Sivapalaratnam S, Nikpay M, Martinelli N, Girelli D, Jackson RD, Kooperberg C, Lange LA, Ardissino D, McPherson R, Farrall M, Watkins H, Reilly MP, Rader DJ, de Faire U, Schunkert H, Erdmann J, Samani NJ, Charnas L, Altshuler D, Gabriel S, Kastelein JJ, Defesche JC, Nederveen AJ, Kathiresan S, Hovingh GK; National Heart, Lung, and Blood Institute GO Exome Sequencing Project. Exome sequencing and directed clinical phenotyping diagnose cholesterol ester storage disease presenting as autosomal recessive hypercholesterolemia. Arterioscler Thromb Vasc Biol. 2013 Dec;33(12):2909-14. doi: 10.1161/ATVBAHA.113.302426. Epub 2013 Sep 26.
  • Talmud PJ, Futema M, Humphries SE. The genetic architecture of the familial hyperlipidaemia syndromes: rare mutations and common variants in multiple genes. Curr Opin Lipidol. 2014 Aug;25(4):274-81. doi: 10.1097/MOL.0000000000000090.
  • Williams RR, Hunt SC, Schumacher MC, Hegele RA, Leppert MF, Ludwig EH, Hopkins PN. Diagnosing heterozygous familial hypercholesterolemia using new practical criteria validated by molecular genetics. Am J Cardiol. 1993 Jul 15;72(2):171-6.

*   Includes publications given by the data provider as well as publications identified by ClinicalTrials.gov Identifier (NCT Number) in Medline.
 
Recruitment Information
Recruitment Status Recruiting
Estimated Enrollment
 (submitted: June 11, 2019) 		1000
Original Estimated Enrollment Same as current
Estimated Study Completion Date July 1, 2021
Estimated Primary Completion Date July 1, 2021   (Final data collection date for primary outcome measure)
Eligibility Criteria

Inclusion Criteria:

  • Pediatric subjects (<18 years old) with a clinical diagnosis of FH and without identified pathogenic mutations in the known candidate genes.

Exclusion Criteria:

  • Subjects with a clinical diagnosis of FH with identified pathogenic mutations in the known candidate genes.
Sex/Gender
Sexes Eligible for Study: All
Ages up to 18 Years   (Child, Adult)
Accepts Healthy Volunteers No
Contacts
Contact: Alberico L Catapano alberico.catapano@unimi.it
Contact: Alessia Tincani +39026173276 catapano.centroatero@gmail.com
Listed Location Countries Italy
Removed Location Countries  
 
Administrative Information
NCT Number NCT03984149
Other Study ID Numbers LIPIGEN-002
Has Data Monitoring Committee Yes
U.S. FDA-regulated Product
Studies a U.S. FDA-regulated Drug Product: No
Studies a U.S. FDA-regulated Device Product: No
IPD Sharing Statement
Plan to Share IPD: Yes
Plan Description: Upon preventive request for a scientific collaboration
Supporting Materials: Study Protocol
Supporting Materials: Statistical Analysis Plan (SAP)
Supporting Materials: Informed Consent Form (ICF)
Supporting Materials: Clinical Study Report (CSR)
Supporting Materials: Analytic Code
Time Frame: For three years from the end of the study
Access Criteria: Upon preventive request for a scientific collaboration
Responsible Party Fondazione SISA (Societa Italiana per lo Studio della Arteriosclerosi)
Study Sponsor Fondazione SISA (Societa Italiana per lo Studio della Arteriosclerosi)
Collaborators Not Provided
Investigators
Study Director: Maurizio Averna Fondazione SISA
PRS Account Fondazione SISA (Societa Italiana per lo Studio della Arteriosclerosi)
Verification Date July 2020

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