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WO2008144940A1 - Marqueur biologique pour l'hypertriglycéridémie - Google Patents

Marqueur biologique pour l'hypertriglycéridémie Download PDF

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WO2008144940A1
WO2008144940A1 PCT/CA2008/001057 CA2008001057W WO2008144940A1 WO 2008144940 A1 WO2008144940 A1 WO 2008144940A1 CA 2008001057 W CA2008001057 W CA 2008001057W WO 2008144940 A1 WO2008144940 A1 WO 2008144940A1
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allele
apoa5
htg
risk
mammal
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Robert A. Hegele
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The University Of Western Ontario
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    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12Q—MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEIC ACIDS OR MICROORGANISMS; COMPOSITIONS OR TEST PAPERS THEREFOR; PROCESSES OF PREPARING SUCH COMPOSITIONS; CONDITION-RESPONSIVE CONTROL IN MICROBIOLOGICAL OR ENZYMOLOGICAL PROCESSES
    • C12Q1/00—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions
    • C12Q1/68—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions involving nucleic acids
    • C12Q1/6876—Nucleic acid products used in the analysis of nucleic acids, e.g. primers or probes
    • C12Q1/6883—Nucleic acid products used in the analysis of nucleic acids, e.g. primers or probes for diseases caused by alterations of genetic material
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12Q—MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEIC ACIDS OR MICROORGANISMS; COMPOSITIONS OR TEST PAPERS THEREFOR; PROCESSES OF PREPARING SUCH COMPOSITIONS; CONDITION-RESPONSIVE CONTROL IN MICROBIOLOGICAL OR ENZYMOLOGICAL PROCESSES
    • C12Q2600/00—Oligonucleotides characterized by their use
    • C12Q2600/112—Disease subtyping, staging or classification
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12Q—MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEIC ACIDS OR MICROORGANISMS; COMPOSITIONS OR TEST PAPERS THEREFOR; PROCESSES OF PREPARING SUCH COMPOSITIONS; CONDITION-RESPONSIVE CONTROL IN MICROBIOLOGICAL OR ENZYMOLOGICAL PROCESSES
    • C12Q2600/00—Oligonucleotides characterized by their use
    • C12Q2600/156—Polymorphic or mutational markers
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12Q—MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEIC ACIDS OR MICROORGANISMS; COMPOSITIONS OR TEST PAPERS THEREFOR; PROCESSES OF PREPARING SUCH COMPOSITIONS; CONDITION-RESPONSIVE CONTROL IN MICROBIOLOGICAL OR ENZYMOLOGICAL PROCESSES
    • C12Q2600/00—Oligonucleotides characterized by their use
    • C12Q2600/172—Haplotypes

Definitions

  • the present invention relates to biomarkers useful in the determination of risk of hypertrigylceridemia in a mammal.
  • Hypertriglyceridemia is a commonly encountered phenotype that is a defining component of metabolic syndrome and is associated with numerous comorbidities, including coronary heart disease (CHD) and diabetes.
  • CHD coronary heart disease
  • TG plasma triglyceride
  • Plasma TG concentration >10 mmol/L is seen in ⁇ 1 in 600 adult North Americans.
  • Moderate HTG may generally be associated with plasma TG concentrations equal to or greater than 5 mmol/L and less than 10 mmol/L and may generally be associated with Frederickson type 4 hyperlipoproteinemia.
  • Mild hypertriglyceridemia may generally be associated with plasma TG concentrations equal to or greater than 2 mmol/L and less than 5 mmol/L and may generally also be associated with Frederickson type 4 hyperlipoproteinemia.
  • Plasma TG concentrations ⁇ 2 mmol/L may qualify as representing the normal range of TG levels. Patients with TG levels falling within the normal, mild, or moderate range remain at risk of developing severe HTG.
  • a method of assessing the risk of HTG in a mammal comprising determining in a nucleic acid- containing sample from the mammal the presence of polymorphisms within the APOA5 gene, wherein the identification of one or more APOA5 polymorphisms is indicative of a risk of HTG.
  • a method of assessing the risk of HTG in a mammal comprising determining in a nucleic acid-containing sample from the mammal the presence of at least one of the APOA5 polymorphisms, APOA5 S19W and APOA5 -1 131T>C, in combination with one or more secondary polymorphisms selected from the group consisting of the GCKR rs780094 A allele, TRIBl rsl7321515 G allele, GALNT2 rs4846914 G allele, TBL2 rsl7145738 T allele and the APOE non-E3 allele, wherein the identification of at least one APOA5 polymorphism in combination with one or more secondary polymorphisms is indicative of risk of HTG.
  • a method of assessing the risk of HTG in a mammal comprising:
  • the invention provides a method of assessing the risk of disease associated with HTG in a mammal comprising the step of identifying the occurrence in said mammal of at least one APOA5 polymorphism, wherein the identification of one or more APOA5 polymorphisms is indicative of a risk of disease associated with HTG.
  • a kit useful to assess risk of HTG in a mammal comprising at least one reagent useful to identify the presence of at least one APOA5 polymorphism.
  • an array useful to assess the risk of HTG in a mammal comprising reagents useful to detect at least one APOA5 polymorphism, and at least one secondary polymorphism selected from the group consisting of the GCKR rs780094 A allele, TRIBl rsl7321515 G allele, GALNT2 rs4846914 G allele, TBL2 rsl7145738 T allele and the APOE non-E3 allele.
  • Figure 1 is a bar graph illustrating the plasma lipoprotein response in patients with severe HTG to fibrate monotherapy
  • Figure 2 illustrates he relationship between plasma TG quartile and APOA5 variant frequencies
  • Figure 3 is a line graph illustrating that the presence of APOA5 S19W, -
  • HLP Fredrickson hyperlipoproteinemia
  • Figure 4 illustrates the risk associated with select clinical and genetic variables for severe HTG.
  • a method of assessing the risk of HTG in a mammal comprising identifying the occurrence in the mammal of one or more polymorphisms within the APOA5 gene. Detection of an APOA5 polymorphism is indicative of a risk of severe HTG.
  • HTG refers to a TG concentration that may fall within the normal, mild, or moderate range, e.g. from ⁇ 2 mmol/L to > 10 mmol/L, while the term “severe HTG” refers to a fasting plasma triglyceride (TG) concentration greater than 10 mmol/L documented on at least 2 distinct occasions, wherein “fasting” refers to no intake of anything other than water for at least 12 hours.
  • TG fasting plasma triglyceride
  • mamal refers to human and non-human mammals such as domestic animals, livestock and wild animals.
  • polymorphism refers to one of two or more alternate forms
  • APOA5 refers to the apolipoprotein A-V gene having the nucleotide sequence depicted by accession no. rs662799.
  • nucleic acid-containing sample is used to refer to any biological sample that may be obtained from the mammal that contains nucleic acid.
  • Appropriate DNA- containing biological samples for use in the present method include, but are not limited to, saliva, urine, semen and other bodily secretions, as well as hair, epithelial cells and the like.
  • invasively-obtained DNA- containing biological samples may also be used in the present method, including for example, blood, serum, bone marrow, cerebrospinal fluid (CSF) and tissue biopsies such as lymph node samples. Techniques for the invasive process of obtaining such samples are known to those of skill in the art.
  • DNA extraction it may be necessary, or preferable, to extract the DNA from the biological sample prior to polymorphism determination.
  • Methods of DNA extraction are well-known to those of skill in the art and include chemical extraction techniques utilizing phenol- chloroform (Sambrook et al., 1989), guanidine-containing solutions, or CTAB-containing buffers.
  • commercial DNA extraction kits are also widely available from laboratory reagent supply companies, including for example, the QIAamp DNA Blood Minikit available from QIAGEN (Chatsworth, CA), or the Extract- N-Amp blood kit available from Sigma (St. Louis, MO).
  • genotyping assays such as TaqMan assays or restriction endonuclease analysis.
  • APOA5 polymorphisms in both the coding and non-coding regions of the gene have been identified as biomarkers associated with risk of HTG.
  • the polymorphisms, APOA5 S19W in the coding region of the gene, and APOA5 -1131T>C in the non-coding region of the gene are each distinct biomarkers associated with risk of HTG in a mammal.
  • distinct it is meant that each of these polymorphisms is itself a biomarker of HTG.
  • the identification of APOA5 polymorphisms in combination with one or more secondary polymorphisms may provide a more definitive risk assessment, i.e. a risk assessment having an odds ratio of at least about 2.00, preferably at least about 4.00, and more preferably at least about 10.
  • the greater the odds ratio the greater the degree of risk in a mammal of developing HTG.
  • secondary polymorphism is used herein to refer to a polymorphism which, when it occurs in conjunction with an APOA5 polymorphism indicative of severe HTG risk e.g. one or the other of , it will generally increase the odds ratio associated with HTG risk.
  • Such secondary polymorphisms include, but are not necessarily limited to, GCKR rs780094 A allele, TRIBl rsl 7321515 G allele, GALNT2 rs4846914 G allele, TBL2 rsl7145738 T allele and APOE non-E3 allele, e.g. E2 (C112(rs429358) + C158(rs7412)) and E4 (R112(s429358) + R158(rs7412)) alleles.
  • GNKR glucokinase regulatory protein
  • TBL2 transducin-(beta)-like 2 gene
  • APOE apolipoprotein E gene
  • APOA5 and secondary polymorphisms as set out above, in combination with one or more clinical factors may also provide a more definitive risk assessment, i.e. a risk assessment having an odds ratio of at least about 2.00, and preferably at least about 4.00.
  • clinical factors is used herein to refer to factors such as diabetes, obesity (defined as having a BMI>33kg/m 2 ), age, sex, diet, alcohol intake, ethnicity (e.g. asian, aboriginal), total fat content and psychological state (e.g. stress) of the candidate mammal to be assessed.
  • a method of assessing the risk of disease associated with HTG in a mammal is provided.
  • the method comprises the step of identifying the occurrence in said mammal of at least one APOA5 polymorphism. Identification of one or more APOA5 polymorphisms is indicative of a risk of disease associated with HTG, including but not limited to metabolic syndrome, disorders in which there is an increased risk of developing cardiovascular disease, diabetes, hypertension, polycystic ovarian syndrome, non-alcoholic fatty liver disease, as well as pancreatitis.
  • Assessment of risk may be calculated manually, or automatically using computer technology.
  • the relevant genetic and clinical factors (or characteristics) of a mammal may be input into a computer adapted to calculate the risk based on these factors, e.g. a system comprising the appropriate software to conduct the risk assessment, and the computer will provide an output of the risk for the mammal of developing severe HTG, or a disease associated with severe HTG.
  • kits that is useful in the determination of risk in a mammal of developing HTG.
  • the kit comprises one or more reagents useful to determine the presence of an APOA5 polymorphism in a nucleic acid- containing sample from the mammal.
  • the kit may optionally include reagents useful to identify the presence of one or more secondary polymorphisms such as the GCKR rs780094 A allele, TRIBl rsl7321515 G allele, GALNT2 rs4846914 G allele, TBL2 rsl 7145738 T allele and the APOE non-E3 allele.
  • the reagent useful to identify the presence of an APOA5 polymorphism, or a secondary polymorphism may comprise one or more probes, primers or other nucleic acid capable of specific binding to a selected polymorphism, or any other component useful to detect a selected polymorphism.
  • the reagents may be detectably labeled and may optionally be fixed to a solid support.
  • the kit may include controls, buffers, and instructions for use.
  • an array is provided that is useful to assess the risk of HTG in a mammal.
  • the array will include a support to which a series of reagents, as set out above, is bound that is useful to identify the APOA5 and secondary polymorphisms identified herein.
  • the reagents typically polynucleotide probes obtained by, e.g., polymerase chain reaction (PCR) amplification of specific gene segments that target the polymorphism of interest, are affixed to the array support using methods well established in the art.
  • Detection of target polymorphisms within a nucleic acid-containing sample from a mammal is conducted using the array in conjunction with a labeling technique, as one of skill in the art will appreciate, which enables detection of binding of the polymorphism-containing nucleic acid.
  • SHARE Study of Health Assessment and Risk in Ethnic groups
  • Table 1 Baseline attributes of study subjects severe HTG controls P-value number 167 277 percent female 32.9% 53.7% O.0001 age (years) 50.9 ⁇ 13.0 50.1 ⁇ 14.8 NS treatment for diabetes mellitus 35.3% 1.4% ⁇ 0.0001 body mass index (kg/m 2 ) 30.3 ⁇ 4.8 27.0 ⁇ 4.6 O.0001 body mass index >33 kg/m 2 28.0% 8.6% O.0001 total cholesterol (mmol/L) 11.9 ⁇ 5.9 5.0 ⁇ 0.9 ⁇ 0.0001 triglycerides (mmol/L) 31.3 ⁇ 25.0 1.15 ⁇ 0.41 O.0001 HDL-cholesterol (mmol/L) 0.77 ⁇ 0.36 1.25 ⁇ 0.36 ⁇ 0.0001 [0036] Severe HTG cases and controls were matched for age.
  • Body mass index (BMI) was significantly higher in severe HTG patients.
  • severe HTG patients had markedly higher plasma TG and total cholesterol and significantly lower plasma HDL cholesterol.
  • Plasma TG concentration in severe HTG patients ranged from 10.1 to 180 mmol/L.
  • 47/167 severe HTG patients (28.1%) had been hospitalized on >1 occasion with pancreatitis.
  • Corresponding data were not available for the control group.
  • Coding regions and intron-exon boundaries of APOA5 (4 exons) were amplified, purified and then directly sequenced in 5'- and 3'- directions in an ABI 3730 DNA Analyzer using reagents shown in Table 2.
  • exon primer sequence Tanneal ( 0 C) size (bp)
  • Genomic DNA sequences were analyzed using Sequence Navigator software. Sequence variants were confirmed on an independent sample on a second day. Screening of normolipidemic controls for sequence variants found in severe HTG patients was performed using allele-specific methods such as restriction endonuclease analysis or a method called SNaPshot, as summarized in Table 3.
  • RE restriction enzyme
  • ASO allele-specific oligonucleotide method (SNaPshot)Blinded between-day replicated genotypes of a random 5% of samples showed >99.9% concordance.
  • the PANTHER database was used to impute dysfunction of sequence variants.
  • the output of PANTHER is the subPSEC score, which represents the negative logarithm of the probability ratio of dysfunction of the wild-type and mutant amino acids at a particular position of the gene product; scores range between 0 (neutral) and -10 (most likely to be deleterious).
  • PANTHER also calculates a probability of the mutation having a deleterious effect between 0 (neutral) and 1 (certainly deleterious). Predictions of dysfunction are very highly correlated with in vitro functional assessment.
  • P.S19W known SNP -4.02 0.73 associated with 0.162 /0.049f dyslipidemia (ref 31-34); 50% reduced secretion from HepG2 cells
  • Plasma lipoprotein profiles were determined as described for lipid clinic patients (Hegele RA et al. (2003) Arterioscler Thromb Vase Biol 23: 111-6, the relevant contents of which are incorporated herein by reference) and for normal controls according to Anand et al. 2000, the relevant contents of which are incorporated herein by reference.
  • Subjects were classified as having familial hypercholesterolemia (FH; HLP type 2A) based on the presence of definite diagnostic criteria as set out in Yuan G et al. (2006) CMAJ 174: 1124-9, the relevant contents of which are incorporated herein by reference, which in all cases included demonstration of heterozygosity for a disease-causing mutation (as described in Wang J et al (2005) .
  • DBL dysbetalipoproteinemia
  • HLP type 3 dysbetalipoproteinemia
  • HCG hypertriglyceridemia
  • HLP type 4 based on TG concentrations exceeding age- and sex-specific 90th percentile values, but not exceeding 10 mmol/L, with no documented chylomicronemia and absence of other lipoprotein phenotypes. Subjects were classified has having severe HTG, sometimes also called 'mixed hyperlipidemia' (MHL; HLP type 5) based on fasting plasma TG >10 mmol/L documented on >2 occasions with documented chylomicronemia. Children with fasting plasma TG >10 mmol/L with documented chylomicronemia and homozygous or compound heterozygous mutations in LPL were excluded.
  • MHL 'mixed hyperlipidemia'
  • APOA5 S19W (dbSNP rs3135506) was genotyped using a validated TaqMan genotyping assay (Assay ID C 25638153 10; TaqMan® SNP Genotyping Assays, Applied Biosystems, Foster City, CA).
  • APOA5 T[- 1131C] (dbSNP rsl 729411) was genotyped using a custom designed TaqMan genotyping assay (TaqMan® SNP Custom Genotyping Assays, Applied Biosystems, Foster City, CA).
  • a 600 nucleotide sequence (300 upstream and 300 downstream) from NT 033899.7 was submitted to RepeatMasker (www.repeatmasker.org) to detect repetitive sequences and then the sequence was submitted to BLASTN2.2.17 to confirm unique alignment to the human Build 36 genome database. After passing these criteria, the 600 nucleotide sequence was edited to place an "N" where any other SNPs or indels were present to allow for Applied Biosystems to design the custom probe.
  • the custom probe uses primers as follows: 5'- CCC TGC GAG TGG AGT TCA -3' and 5'- CTC TGA GCC CCA GGA ACT G.
  • SNP genotyping was performed using an allelic discrimination assay using the 7900HT Fast Real-Time PCR System and genotypes were read using automated software (SDS 2.3, Applied Biosystems, Foster City, CA). Reactions were run in 5 ⁇ L volumes using an amplification protocol of 95 0 C for 10 minutes, followed by 50 cycles of 95 0 C for 15 seconds, then 6O 0 C for 1.5 minutes.
  • HLP2A HLP2B
  • HLP3 HLP4
  • patients controls number 88 92 48 38 151 678 373 percent female 454% 46 7% 396% 10 5% 31 8% 43 1%
  • age (years) 57 7+13 6 56 6 ⁇ 11 7 51 6 ⁇ 12 0 59 5 ⁇ 13 4 50 8+12 7 54 7+14 7 472+15 2 body mass index (kg/m 2 ) 24 1+3 4 29 1+4 3 28 9+3 1 31 2+8 1 30 5+4 8 28 7 ⁇ 4 7 27 2 ⁇ 42 plasma cholesterol (mmol/L)
  • APOAS S19W or -1131T>C neither 71 5% (63 63 1% (58) 54 2% (26) 55 3%(21) 42 3%(64) 64 8% (439) 82 6% (308) either 28 5% (25) 369% (34) *** 45 8% (22) *** 44 7 (17) *** 57 6% (87) *** 35 2% (239) *** 174 (65)
  • Estimates of pairwise linkage disequilibrium between APOA5 S19W and -1 131T>C in HLP subgroups was similarly non-significant, with r values ranging from 0.004 to 0.165 and P-values ranging from 0.23 to 0.98.
  • the alleles of the two SNPs were not significantly associated with each other and SNP genotypes could be considered as being independent of each other.
  • APOA5 S19W and -1131T>C genotype frequencies did not deviate significantly from expectations of Hardy- Weinberg equilibrium.
  • Table 2 shows that APOA5 S19W was found at significantly higher allele frequency compared to controls (9.4% vs. 4.2%, respectively, PO.0001) and at significantly higher carrier frequency compared to controls (18.1% vs. 7.2%, respectively, P ⁇ 0.0001).
  • APOA5 -1 131T>C was found at significantly higher allele frequency compared to controls (10.5% vs. 5.4%, respectively, PO.0001) and at significantly higher carrier frequency compared to controls (19.8% vs. 10.4%, respectively, PO.0001).
  • Carriers of either APOA5 S19W or -1131T>C were significantly more frequent among lipid clinic patients compared to controls (carrier frequency of 35.2% vs. 17.4%, respectively, PO.0001).
  • the overall odds ratio for carriers of APOA5 S19W, -1131T>C or either one among lipid clinic patients was 2.98 (95% confidence interval [CI] 1.93 to 4.60), 2.01 (95% CI 1.38 to 2.95) and 2.58 (95% CI 1.89 to 3.52), respectively.
  • a stepwise relationship between APOA5 S19W carrier frequency and TG quartile: 11.8%, 13.4%, 20.0% and 30.5% in quartiles 1, 2, 3 and 4, respectively (P for trendO.OOOl) was similarly observed.
  • a significant increasing trend of APOA5 -1131T>C allele frequency across TG quartiles: 6.5%, 7.0%, 11.5% and 15.9% in quartiles 1, 2, 3 and 4, respectively (P for trend ⁇ 0.0001) was observed, as well as a stepwise relationship between APOA5 -1131T>C carrier frequency and TG quartile: 12.4%, 14.0%, 21.2% and 28.8% in quartiles 1, 2, 3 and 4, respectively (P for trendO.OOOl).
  • HLP types 2A, 2B, 3, 4 and 5 were unequivocally classified with FH, CHL, DBL, HTG and MHL (HLP types 2A, 2B, 3, 4 and 5, respectively).
  • HLP type 1 defined as children or adolescents with LPL deficiency due to absent post-heparin LPL activity and/or mutated LPL or APOC2 alleles.
  • Clinical, biochemical and genetic features of study subjects are shown in Table 2. For all HLP phenotypes, APOA5 S19W and -1131T>C genotype frequencies did not deviate significantly from expectations of the Hardy- Weinberg equilibrium.
  • APOA5 allele and carrier frequencies were significantly higher than in control subjects for all HLP phenotypes, except for FH as illustrated in Figure 3.
  • AP O AS S19W carrier odds ratios for HLP types 2B, 3, 4 and 5 were 3.11 (95% confidence interval [CI] 1.63 to 5.95), 4.76 (95% CI 2.25 to 10.1), 2.89 (95% CI 1.17 to 7.18) and 6.16 (95% CI 3.66 to 10.3), respectively.
  • APOA5 -1 131T>C carrier odds ratios for HLP types 2B, 3, 4 and 5 were 2.23 (1.21 to 4.08), 3.18 (95% CI 1.55 to 6.52), 3.95 (95% CI 1.85 to 8.45) and 4.24 (95% CI 2.64 to 6.81), respectively.
  • the presence of either allele was similarly strongly associated with HLP types: the overall odds ratio for the presence of either allele in lipid clinic patients was 2.58 (95% CI 1.89 to 3.52).
  • the ORs for the TG-containing HLP phenotypes are shown in
  • APOA5 variants S19W and -1131T>C are strongly and specifically associated with HTG in lipid clinic patients and with several HLP phenotypes defined by elevated plasma TG concentration.
  • the findings confirm the importance of these APOA5 variants in the study of patient pathogenesis and response to intervention, and also for diagnosis of HTG and TG-containing HLP phenotypes.
  • HLP type 2B, 3, 4 and 5 phenotypes Between 30 and 60% of subjects with these four classical HLP phenotypes were carriers of either the APOA5 S 19W or - 1131T>C alleles, with significant odds ratios between 2 and 7. Thus, the results indicate that both variants individually have comparably strong associations with HTG and HLP 2B, 3, 4 and 5. Furthermore, given the absence of significant linkage disequilibrium between APOA5 S19W and -1131T>C, the genotypes function effectively as independent determinants of HTG and the essentially independent information that each provides can be combined.
  • APOA5 S19W and -1131T>C are herein shown to be clinical genetic markers for HTG.
  • APOA5 S19W and -1131T>C are consistent and important genetic determinants of complex traits defined by elevated TG, specifically classical HLP phenotypes, as well as general HTG.
  • markers that were replicably associated with plasma TG and showed relatively strong association were selected.
  • the selected genes and dbSNP identification numbers were: GALNT2 rs4846914, TBL2 rsl7145738, TRIBl rsl7321515, ANGPTL3 rsl2130333, GCKR rs780094, APOA5 rs3135506 (S 19W) and LPL rs328 (S447X).
  • These genes were genotyped using validated genotyping assays (TaqMan® SNP Genotyping Assays, Applied Biosystems, Foster City, CA).
  • APOA5 -1131T>C was genotyped using a custom designed genotyping assay (TaqMan® SNP Custom Genotyping Assays, Applied Biosystems, Foster City, CA).
  • the custom probe uses primers as follows: 5'- CCC TGC GAG TGG AGT TCA -3' and 5'- CTC TGA GCC CCA GGA ACT G.
  • SNP genotyping was performed using an allelic discrimination assay using the 7900HT Fast Real-Time PCR System (Applied Biosystems, Foster City, CA) and genotypes were read using automated software (SDS 2.3, Applied Biosystems, Foster City, CA).
  • Reactions were run in 5 ⁇ L volumes using an amplification protocol of 95 0 C for 10 minutes, followed by 42 cycles of 95 0 C for 15 seconds, then 6O 0 C for 1.5 minutes.
  • An established method was used to genotype APOE isoforms (described in Hixson et al. (1990). J Lipid Res 31, 545-548. the relevant contents of which are incorporated herein by reference).
  • LPL, APOC2 or APOA5 132 patients or cases with severe HTG remained for analysis. These were each matched with up to 4 normolipidemic controls based on age within 5 years and sex. By definition, severe HTG patients had markedly higher plasma TG and total cholesterol and significantly lower HDL cholesterol (Table 7). Plasma TG concentration in severe HTG patients ranged from 10.1 to 180 mmol/L. In addition, 37/132 severe HTG patients (28.0%) had been hospitalized on >1 occasion with pancreatitis.
  • APOA5 gene encoding apolipoprotein A-V, GCKR, gene encoding glucokinase receptor, TRIBl, gene encoding homologue ofDrosophila Tnbbles 1, GALNT2, gene encoding UDP-N-acetyl-alpha-D-galactosamine polypeptide N-acetylgalactosaminyltransferase, TBL2 gene encoding transducin-beta-like-2, ANGPTL3 gene encoding angiopoietin-hke 3, APOE, gene encoding apolipoprotein E, LPL, gene encoding lipoprotein lipase [0069] Minor allele frequencies (MAFs) for each genotype in severe HTG cases and controls are shown in Table 9.
  • MAFs Minor allele frequencies
  • Variables entered into model are defined as follows: APOA5 Wl 9 dominant had the test genotypes SW and WW and the reference genotype SS; APOA5 -1 131C dominant had the test genotypes CC and TC and the reference genotype TT; GALNT2 G recessive had the test genotype GG and the reference genotypes AA and AG; GCKR A recessive had the test genotype AA and the reference genotypes GA and GG; TBL2 C recessive had the test genotype CC and the reference genotypes CT and TT; APOE non-E3 allele had the test genotypes 2/2, 4/2, 4/4 and the reference genotypes 3/2, 3/3 and 4/3; TRIBl A recessive had the test genotype AA and the reference genotypes AG and GG; LPL S447 recessive had the test genotype SS and the reference genotypes SX (there were no XX individuals); ANGPTL3 C recess
  • the multivariate ORs for severe HTG were calculated using the WaId statistic in multivariate logistic regression analysis with stepwise addition of variables and P ⁇ 0.05 for each step (Table 1 1).
  • the first model which included two clinical variables in addition to nine genetic variables, found that diabetes, obesity, two APOA5 markers, APOE non-E3 genotype and GCKR, TRIBl and TBL2 genotypes were significantly associated with severe HTG.
  • the C-statistic which corresponds to the area under the receiver-operator curve for a diagnostic test, was 0.869 for this particular combination of clinical and genetic markers (Table 11).
  • the model used backward elimination and had a nominal P-value of 0 05 for each variable [0076]
  • the proportion of contribution of specific variables to severe HTG was calculated using partial i ⁇ -values in multivariate linear regression analysis with stepwise addition of variables and P ⁇ 0.05 for each step (Table 11).
  • the first model which included two clinical variables in addition to nine genetic variables, found that diabetes, APOA5 markers, obesity, TBL2 genotype, APOE genotype, TRIBl genotype and GCKR genotype were significantly associated with severe HTG.
  • the model explained -43% of total variation in case versus control status, and of the explained variation, the total contribution of the genetic variables was -40% (range -1 to 25%).
  • the second model assessed only genetic variables: the same genotypes from the first model remained significantly associated in the second model with one additional significantly associated genotype - namely GALNT2.
  • the model accounted for -25% of total variation in case versus control status.
  • genetic markers accounted for at least -1% each as shown in Figure 4.

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Abstract

Cette invention concerne un procédé permettant d'évaluer le risque d'une hypertriglycéridémie sévère chez un mammifère, lequel procédé consiste à déterminer dans un échantillon contenant des acides nucléiques prélevé sur le mammifère la présence d'au moins un polymorphisme dans le gène APOA5. L'identification d'un polymorphisme APOA5 indiquant un risque d'hypertriglycéridémie sévère.
PCT/CA2008/001057 2007-05-31 2008-05-30 Marqueur biologique pour l'hypertriglycéridémie WO2008144940A1 (fr)

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Cited By (7)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
ES2338856A1 (es) * 2009-12-29 2010-05-12 Universidad De Malaga Conjunto de cebadores, sondas, procedimiento y kit para el genotipado del polimorfismo genetico - 1131t/c del gen apo a5.
WO2014193247A1 (fr) * 2013-05-31 2014-12-04 Livestock Improvement Corporation Limited Marqueurs génétiques du nanisme et leur utilisation
CN110358839A (zh) * 2019-06-06 2019-10-22 佛山科学技术学院 与猪饲料转化率相关的gckr基因的snp分子遗传标记
WO2019214591A1 (fr) * 2018-05-07 2019-11-14 创观（苏州）生物科技有限公司 Produit sanguin dérivé d'un porc knock-out de gène et son utilisation
CN111662977A (zh) * 2020-06-30 2020-09-15 西安市精神卫生中心 一种用于抗精神药物的代谢影响检测的基因探针、芯片、试剂盒及其应用
CN111909996A (zh) * 2020-07-08 2020-11-10 广西医大睿谷医学检验有限公司 一种个体化用药相关基因多态性检测试剂盒
CN118256611A (zh) * 2023-05-16 2024-06-28 山东大学齐鲁医院 筛查高脂血症和/或急性胰腺炎患者或评估高脂血症和/或急性胰腺炎易感性的系统

Citations (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US20030150003A1 (en) * 2001-08-27 2003-08-07 Edward Rubin Novel apolipoprotein gene involved in lipid metabolism

Patent Citations (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US20030150003A1 (en) * 2001-08-27 2003-08-07 Edward Rubin Novel apolipoprotein gene involved in lipid metabolism

Non-Patent Citations (10)

* Cited by examiner, † Cited by third party
Title
DE BEER F. ET AL.: "Apolipoprotein E2 (Lys146 Gln) causes hypertriglyceridemia due to an apolipoprotein E variant-specific inhibition of lipolysis of very low density lipoproteins-triglycerides", ARTERIOSCLER. THROMB. VASC. BIOL., vol. 20, no. 7, July 2000 (2000-07-01), pages 1800 - 1806 *
KATHIRESAN S. ET AL.: "Six new loci associated with blood low-density lipoprotein cholesterol, high-density lipoprotein cholesterol or triglycerides in humans", NAT. GENET., vol. 40, no. 2, February 2008 (2008-02-01), pages 189 - 197, XP002516887, DOI: doi:10.1038/NG.75 *
PENNACCHIO L.A. AND RUBIN E.: "Apolipoprotein A5, a newly identified gene that affects plasma triglyceride levels in humans and mice", ARTERIOSCLER. THROMB. VASC. BIOL., vol. 23, no. 4, April 2003 (2003-04-01), pages 529 - 534 *
PENNACCHIO L.A. ET AL.: "An apolipoprotein influencing triglycerides in humans and mice revealed by comparative sequencing", SCIENCE, vol. 294, no. 5540, 5 October 2001 (2001-10-05), pages 169 - 173 *
PRIORE OLIVA C. ET AL.: "Inherited apolipoprotein A-V deficiency in severe hypertriglyceridemia", ARTERIOSCLER. THROMB. VASC. BIOL., vol. 25, no. 2, February 2005 (2005-02-01), pages 411 - 417 *
SAXENA R. ET AL.: "Genome-wide association analysis identifies loci for type 2 diabetes and triglyceride levels", SCIENCE, vol. 316, no. 5829, 1 June 2007 (2007-06-01), pages 1331 - 1336 *
SPARSO T. ET AL.: "The GCKR rs780094 polymorphism is associated with elevated fasting serum triacyclglycerol, reduced fasting and OGTT-related insulinaemia, and reduced risk of type 2 diabetes", DIABETOLOGIA, vol. 51, no. 1, January 2008 (2008-01-01), pages 70 - 75, XP019558703 *
TALMUD P.J. ET AL.: "Determination of the functionality of common APOA5 polymorphisms", J. BIOL. CHEM., vol. 280, no. 31, 5 August 2005 (2005-08-05), pages 28215 - 28220 *
WANG J. ET AL.: "Resequencing genomic DNA of patients with severe hypertriglyceridemia (MIM 144650)", ARTERIOSCLER. THROMB. VASC. BIOL., vol. 27, no. 11, November 2007 (2007-11-01), pages 2450 - 2455 *
WILLER C.J. ET AL.: "Newly identified loci that influence lipid concentrations and risk of coronary artery disease", NAT. GENET., vol. 40, no. 2, February 2008 (2008-02-01), pages 161 - 169, XP002516888, DOI: doi:10.1038/ng.76 *

Cited By (10)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
ES2338856A1 (es) * 2009-12-29 2010-05-12 Universidad De Malaga Conjunto de cebadores, sondas, procedimiento y kit para el genotipado del polimorfismo genetico - 1131t/c del gen apo a5.
ES2338856B2 (es) * 2009-12-29 2011-01-27 Universidad De Malaga Conjunto de cebadores, sondas, procedimiento y kit para el genotipadodel polimorfismo genetico - 1131t/c del gen apo a5.
WO2014193247A1 (fr) * 2013-05-31 2014-12-04 Livestock Improvement Corporation Limited Marqueurs génétiques du nanisme et leur utilisation
WO2019214591A1 (fr) * 2018-05-07 2019-11-14 创观（苏州）生物科技有限公司 Produit sanguin dérivé d'un porc knock-out de gène et son utilisation
CN112105369A (zh) * 2018-05-07 2020-12-18 创观（苏州）生物科技有限公司 源自基因敲除猪的血液产品及其用途
CN112105369B (zh) * 2018-05-07 2024-05-14 创观（苏州）生物科技有限公司 源自基因敲除猪的血液产品及其用途
CN110358839A (zh) * 2019-06-06 2019-10-22 佛山科学技术学院 与猪饲料转化率相关的gckr基因的snp分子遗传标记
CN111662977A (zh) * 2020-06-30 2020-09-15 西安市精神卫生中心 一种用于抗精神药物的代谢影响检测的基因探针、芯片、试剂盒及其应用
CN111909996A (zh) * 2020-07-08 2020-11-10 广西医大睿谷医学检验有限公司 一种个体化用药相关基因多态性检测试剂盒
CN118256611A (zh) * 2023-05-16 2024-06-28 山东大学齐鲁医院 筛查高脂血症和/或急性胰腺炎患者或评估高脂血症和/或急性胰腺炎易感性的系统

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