WO2010005303A2 - Nouveaux indicateurs de longévité humaine et du taux de vieillissement biologique - Google Patents

Nouveaux indicateurs de longévité humaine et du taux de vieillissement biologique Download PDF

Info

Publication number
WO2010005303A2
WO2010005303A2 PCT/NL2009/050409 NL2009050409W WO2010005303A2 WO 2010005303 A2 WO2010005303 A2 WO 2010005303A2 NL 2009050409 W NL2009050409 W NL 2009050409W WO 2010005303 A2 WO2010005303 A2 WO 2010005303A2
Authority
WO
WIPO (PCT)
Prior art keywords
longevity
nucleotide sequence
seq
substance
snps
Prior art date
Application number
PCT/NL2009/050409
Other languages
English (en)
Other versions
WO2010005303A3 (fr
Inventor
Pieternella Slagboom
Rudolf Gerardus Johannes Westendorp
Jeanne Jacobine Houwing - Duistermaat
Bastiaantheodoor Heijmans
Marian Beekman
Original Assignee
Publiekrechtelijke Rechtspersoon Academisch Ziekenhuis Leiden H.O.D.N. Leids Universitair Medisch Centrum
Priority date (The priority date is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the date listed.)
Filing date
Publication date
Application filed by Publiekrechtelijke Rechtspersoon Academisch Ziekenhuis Leiden H.O.D.N. Leids Universitair Medisch Centrum filed Critical Publiekrechtelijke Rechtspersoon Academisch Ziekenhuis Leiden H.O.D.N. Leids Universitair Medisch Centrum
Publication of WO2010005303A2 publication Critical patent/WO2010005303A2/fr
Publication of WO2010005303A3 publication Critical patent/WO2010005303A3/fr

Links

Classifications

    • 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
    • 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
    • C12Q1/6881—Nucleic acid products used in the analysis of nucleic acids, e.g. primers or probes for tissue or cell typing, e.g. human leukocyte antigen [HLA] probes
    • 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
    • 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
    • C12Q2600/118—Prognosis of disease development
    • 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
    • C12Q2600/136—Screening for pharmacological compounds
    • 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
    • C12Q2600/158—Expression markers

Definitions

  • the present invention relates to the field of molecular human genetics and epidemiology.
  • the invention relates to genetic and biochemical markers of longevity.
  • the GWAS was followed by replication analysis in additional cohorts using cross-sectional and prospective data.
  • One locus influenced the probability to survive into old age by affecting both the risk of cancer and cardiovascular mortality and was related to transcription of nearby genes.
  • hybridisation refers to the binding of two single stranded nucleic acids via complementary base pairing.
  • hybridizing specifically to refers to the binding, duplexing, or hybridizing of a nucleic acid molecule preferentially to a particular nucleotide sequence under stringent conditions.
  • stringent conditions refers to conditions under which a probe will hybridize preferentially to its target subsequence, and to a lesser extent to, or not at all to, other sequences in a mixed population (e.g., a cell lysate or DNA preparation from a tissue biopy)
  • a "stringent hybridization” and “stringent hybridization wash conditions” in the context of nucleic acid hybridization are sequence dependent, and are different under different environmental parameters.
  • Very stringent conditions are selected to be equal to the T m for a particular probe.
  • An example of stringent hybridization conditions for hybridization of complementary nucleic acids which have more than 100 complementary residues on an array or on a filter in a Southern or northern blot is 42 0 C using standard hybridization solutions (see, e.g., Sambrook and Russell (2001) Molecular Cloning: A Laboratory Manual (3rd ed.) Vol. 1-3, Cold Spring Harbor Laboratory, Cold Spring Harbor Press, NY, and detailed discussion, below), with the hybridization being carried out overnight.
  • An example of highly stringent wash conditions is 0.15 M NaCl at 72 0 C for about 15 minutes.
  • An example of stringent wash conditions is a 0.2 x SSC wash at 65 0 C for 15 minutes (see, e.g., Sambrook supra, for a description of SSC buffer). Often, a high stringency wash is preceded by a low stringency wash to remove background probe signal.
  • An example medium stringency wash for a duplex of, e.g., more than 100 nucleotides, is 1 x SSC at 45 0 C for 15 minutes.
  • An example of a low stringency wash for a duplex of, e.g., more than 100 nucleotides is 4 x to 6 x SSC at 40 0 C for 15 minutes.
  • nucleic acid or “nucleic acid molecule” as used herein refers to a deoxyribonucleotide or ribonucleotide in either single- or double-stranded form.
  • the term encompasses nucleic acids containing known analogues of natural nucleotides which have similar or improved binding properties, for the purposes desired, as the reference nucleic acid.
  • the term also includes nucleic acids which are metabolized in a manner similar to naturally occurring nucleotides or at rates that are improved for the purposes desired.
  • nucleic-acid-like structures with synthetic backbones are examples of synthetic backbones.
  • DNA backbone analogues provided by the invention include phosphodiester, phosphorothioate, phosphorodithioate, methylphosphonate, phosphoramidate, alkyl phosphotriester, sulfamate, 3'-thioacetal, methylene(methylimino), 3'-N-carbamate, morpholino carbamate, and peptide nucleic acids (PNAs); see Oligonucleotides and Analogues, a Practical Approach, edited by F. Eckstein, IRL Press at Oxford University Press (1991); Antisense Strategies, Annals of the New York Academy of Sciences, Volume 600, Eds. Baserga and Denhardt (NYAS 1992); Milligan (1993) J. Med. Chem.
  • PNAs contain non-ionic backbones, such as N-(2- aminoethyl) glycine units. Phosphorothioate linkages are described in WO 97/03211; WO 96/39154; Mata (1997) Toxicol. Appl. Pharmacol. 144: 189-197. Other synthetic backbones encompassed by the term include methyl-phosphonate linkages or alternating methylphosphonate and phosphodiester linkages (Strauss-Soukup (1997) Biochemistry 36: 8692-8698), and benzylphosphonate linkages (Samstag (1996) Antisense Nucleic Acid Drug Dev 6: 153-156).
  • nucleic acid molecule or sequence of the invention When a nucleic acid molecule or sequence of the invention is in single stranded form, the opposite, i.e. complementary strand of the nucleic acid molecule or sequence is expressly included within the scope of the invention.
  • An isolated nucleic acid means an object species of the invention that is the predominant species present (i.e., on a molar basis it is more abundant than any other individual species in the composition).
  • an isolated nucleic acid comprises at least about 50, 80 or 90 percent (on a molar basis) of all macro molecular species present.
  • the object species is purified to essential homogeneity (contaminant species cannot be detected in the composition by conventional detection methods).
  • array refers to an arrangement, on a substrate surface, of multiple nucleic acid molecules of predetermined identity, of which preferably the sequences are known. Each nucleic acid molecule is immobilized to a "discrete spot” (i.e., a defined location or assigned position) on the substrate surface.
  • micro-array more specifically refers to an array that is miniaturized so as to require microscopic examination for visual evaluation.
  • the arrays used in the methods of the invention are preferably microarrays.
  • the nucleic acid array as used herein is a plurality of target elements, each target element comprising one or more nucleic acid molecules (probes) immobilized on one or more solid surfaces to which sample nucleic acids can be hybridized.
  • the nucleic acids of a probe can contain sequence(s) from specific genes or clones, e.g. from specific genomic regions described in Tables herein. Other probes may contain, for instance, reference sequences.
  • the probes of the arrays may be arranged on the solid surface at different densities. The probe densities will depend upon a number of factors, such as the nature of the label, the solid support, and the like. One of skill will recognize that each probe may comprise a mixture of nucleic acids of different lengths and sequences.
  • a probe may contain more than one copy of a cloned piece of DNA or RNA, and each copy may be broken into fragments of different lengths.
  • the length and complexity of the nucleic acid fixed onto the target element is not critical to the invention. One of skill can adjust these factors to provide optimum hybridization and signal production for a given hybridization procedure, and to provide the required resolution among different genes or genomic locations.
  • probe or "nucleic acid probe”, as used herein, is defined to be one or more nucleic acid fragments whose specific hybridization to a sample can be detected.
  • the probe may be unlabelled or labelled as described below so that its binding to the target or sample can be detected.
  • the probe is produced from a source of nucleic acids from one or more particular (preselected) portions of a chromosome, e.g., one or more clones, an isolated whole chromosome or chromosome fragment, or a collection of polymerase chain reaction (PCR) amplification products.
  • the probes of the present invention are produced from nucleic acids found in the regions described herein.
  • the probe may also be isolated nucleic acids immobilized on a solid surface (e.g., nitrocellulose, glass, quartz, fused silica slides), as in an array.
  • the probe may be a member of an array of nucleic acids as described, for instance, in WO 96/17958.
  • Techniques capable of producing high density arrays can also be used for this purpose (see, e.g., Fodor (1991) Science 767-773; Johnston (1998) Curr. Biol. 8: R171-R174; Schummer (1997) Biotechniques 23: 1087-1092; Kern (1997) Biotechniques 23: 120-124; U.S. Pat. No. 5,143,854).
  • primer refers to a single-stranded oligonucleotide capable of acting as a point of initiation of template-directed DNA synthesis under appropriate conditions (i.e., in the presence of four different nucleoside triphosphates and an agent for polymerization, such as, DNA or RNA polymerase or reverse transcriptase) in an appropriate buffer and at a suitable temperature.
  • the appropriate length of a primer depends on the intended use of the primer but typically ranges from 15 to 30 nucleotides. Short primer molecules generally require cooler temperatures to form sufficiently stable hybrid complexes with the template.
  • a primer need not reflect the exact sequence of the template but must be sufficiently complementary to hybridize with a template.
  • primer site refers to the area of the target DNA to which a primer hybridizes.
  • primer pair means a set of primers including a 5' upstream primer that hybridizes with the 5' end of the DNA sequence to be amplified and a 3' downstream primer that hybridizes with the complement of the 3' end of the sequence to be amplified.
  • nucleic acid molecules and nucleotide sequences of the present invention that are to be used for the detection and/or for quantification of polymorphisms, genetic markers, (differentially) expressed sequences, such as e.g. probes and primers, are chosen such that they are specific for sequence to be detected in the context of the human genome and/or human transcriptome.
  • nucleic acid molecules and nucleotide sequences will comprise a unique sequence (in the context of the human genome and/or human transcriptome) consisting of sufficient length to be specific (i.e. at least 10, 11, 12, 13, 14, 15, 16, 17. 18.
  • sample as used herein relates to a material or mixture of materials, containing one or more components of interest. Samples include, but are not limited to, samples obtained from an organism and may be directly obtained from a source (e.g., such as a blood sample, a biopsy or from a tumor) or indirectly obtained e.g., after culturing and/or one or more processing steps.
  • a source e.g., such as a blood sample, a biopsy or from a tumor
  • indirectly obtained e.g., after culturing and/or one or more processing steps.
  • genomic refers to all nucleic acid sequences (coding and non-coding) and elements present in each cell type, preferably each somatic cell type, of a subject.
  • genome also applies to any naturally occurring or induced variation of these sequences that may be present in a mutant or disease variant of any cell type, including tumour cells.
  • genomic DNA and “genomic nucleic acid” are used herein interchangeably. They refer to nucleic acid isolated from a nucleus of one or more cells, and include nucleic acid derived from (i.e., isolated from, amplified from, cloned from as well as synthetic versions of) genomic DNA.
  • the human genome consists of approximately 3.0 x 10 9 base pairs of DNA organised into distinct chromosomes.
  • the genome of a normal diploid somatic human cell consists of 22 pairs of autosomes (chromosomes 1 to 22) and either chromosomes X and Y (males) or a pair of chromosome Xs (female) for a total of 46 chromosomes.
  • a genome of a cancer cell may contain variable numbers of each chromosome in addition to deletions, rearrangements and amplification of any subchromosomal region or DNA sequence.
  • each nucleic acid probe immobilised to a discrete spot on an array has a sequence that is specific to (or characteristic of) a particular genomic region.
  • the ratio of intensity of two differentially labelled test and reference samples at a given spot on the array reflects the genome copy number ratio of the two samples at a particular genomic region.
  • Linkage describes the tendency of genes, alleles, loci or genetic markers to be inherited together as a result of their location on the same chromosome.
  • Linkage disequilibrium or allelic association means the preferential association of a particular allele or genetic marker with a specific allele or genetic marker at a nearby chromosomal location more frequently than expected by chance for any particular allele frequency in the population. For example, if locus X has alleles a and b, which occur equally frequently, and linked locus Y has alleles c and d, which occur equally frequently, one would expect the combination ac to occur with a frequency of 0.25. If ac occurs more frequently, then alleles a and c are in linkage disequilibrium.
  • Linkage disequilibrium may result from natural selection of certain combination of alleles or because an allele has been introduced into a population too recently to have reached equilibrium with linked alleles.
  • a marker in linkage disequilibrium can be particularly useful in detecting susceptibility to disease (or other phenotype) notwithstanding that the marker does not cause the disease.
  • a marker (X) that is not itself a causative element of a disease, but which is in linkage disequilibrium with a gene (including regulatory sequences) (Y) that is a causative element of a phenotype can be used detected to indicate susceptibility to the disease in circumstances in which the gene Y may not have been identified or may not be readily detectable.
  • Polymorphism refers to the occurrence of two or more genetically determined alternative sequences or alleles in a population.
  • a polymorphic marker or site is the locus at which divergence occurs. Preferred markers have at least two alleles, each occurring at frequency of greater than 1%, and more preferably greater than 10% or 20% of a selected population.
  • a polymorphic locus may be as small as one base pair.
  • Polymorphic markers include restriction fragment length polymorphisms, variable number of tandem repeats (VNTR's), hypervariable regions, minisatellites, dinucleotide repeats, trinucleotide repeats, tetranucleotide repeats, simple sequence repeats, and insertion elements such as AIu.
  • allelic form occurring most frequently in a selected population is sometimes referred to as the wildtype form. Diploid organisms may be homozygous or heterozygous for allelic forms.
  • a diallelic polymorphism has two forms.
  • a triallelic polymorphism has three forms.
  • a single nucleotide polymorphism or SNP occurs at a polymorphic site occupied by a single nucleotide, which is the site of variation between allelic sequences. The site is usually preceded by and followed by highly conserved sequences of the allele (e.g., sequences that vary in less than 1/100 or 1/1000 members of the populations). SNPs are most frequently diallelic.
  • a single nucleotide polymorphism usually arises due to substitution of one nucleotide for another at the polymorphic site.
  • a transition is the replacement of one purine by another purine or one pyrimidine by another pyrimidine.
  • a transversion is the replacement of a purine by a pyrimidine or vice versa.
  • Single nucleotide polymorphisms can also arise from a deletion of a nucleotide or an insertion of a nucleotide relative to a reference allele.
  • To identify common genetic variation associated with longevity we have performed a genome wide association scan in the Leiden Longevity study and followed the results up for replication in other cohorts of elderly subjects.
  • the present invention therefore relates to genetic markers of longevity.
  • genetic markers of longevity are preferably markers for mammalian longevity, more preferably human longevity, of which markers for longevity in the Caucasian population are most preferred.
  • the genetic marker for longevity is a nucleic acid molecule comprising a nucleotide sequence that is in linkage disequilibrium with an SNP selected from the group consisting of the SNPs listed in Table 2.1.1.
  • a nucleotide sequence that is in linkage disequilibrium with an SNP preferably is a polymorphic nucleotide sequence, Preferably a polymorphic nucleotide sequence is in linkage disequilibrium with the minor allele of an SNP.
  • a preferred genetic marker for longevity is a nucleic acid molecule comprising a nucleotide sequence that is in linkage disequilibrium with an SNP selected from the group consisting of the SNPs listed in Table 2.1.3.
  • the genetic marker for longevity is a nucleic acid molecule comprising a nucleotide sequence that is in linkage disequilibrium with an SNP selected from the group consisting of the SNPs listed in Table 2.1.2. Still more preferably the genetic marker for longevity is a nucleic acid molecule comprising a nucleotide sequence that is in linkage disequilibrium with an SNP selected from the group consisting of the SNPs listed in Table 2.1.4. Most preferably the genetic marker for longevity is a nucleic acid molecule comprising a nucleotide sequence that is in linkage disequilibrium with an SNP selected from the group consisting of the SNPs listed in Table 2.1.5.
  • the genetic marker for longevity is a nucleic acid molecule comprising a nucleotide sequence that is in linkage disequilibrium with an SNP selected from the group consisting of the SNPs listed in Table 2.1.6.
  • the nucleotide sequence comprises a polymorphism in the human population that is associated with longevity as listed in Table 2.1.1.
  • the polymorphism is a SNP selected from the group consisting of the SNPs listed in Table 2.1.3. More preferably, the polymorphism is a SNP selected from the group consisting of the SNPs listed in Table 2.1.2.
  • the polymorphism is a SNP selected from the group consisting of the SNPs listed in Table 2.1.4. Most preferably the polymorphism is a SNP selected from the group consisting of the SNPs listed in Table 2.1.5. In an alternative preferred embodiment the polymorphism is a SNP selected from the group consisting of the SNPs listed in Table 2.1.6.
  • the nucleic acid molecule comprises at least 10, 12, 15, 20, 25 or 30 contiguous nucleotides from a nucleotide sequence that has at least 80, 85, 90, 95, 98, 99% sequence identity with a nucleotide sequence as defined in any one of SEQ ID NO.'s comprising an SNP listed in the above-mentioned Tables (see Table 3.1).
  • the nucleic acid molecule comprises at least 10, 12, 15, 20, 25 or 30 contiguous nucleotides from a nucleotide sequence that has at least 80, 85, 90, 95, 98, 99% sequence identity with a nucleotide sequence as defined in any one of SEQ ID NO.'s: 1 - 94, more preferably the nucleic acid molecule comprises at least 10, 12, 15, 20, 25 or 30 contiguous nucleotides from a nucleotide sequence that has at least 80, 85, 90, 95, 98, 99% sequence identity with a nucleotide sequence as defined in any one of SEQ ID NO.'s: 17, 18, 22, 23, 32, 39, 41, 44, 45, 56, 83 and 84.
  • a particularly preferred genetic marker for longevity is a nucleic acid molecule comprising a nucleotide sequence that is in linkage disequilibrium with an SNP selected from the group consisting of the SNPs rsl6905070, rs7814049 and rs7013830.
  • SNPs rsl6905070, rs7814049 and rs7013830 are a nucleotide sequence present at chromosome 8q24.22.
  • a nucleotide sequence that is in linkage disequilibrium with SNPs rsl6905070, rs7814049 and rs7013830 is a nucleotide sequence comprised in a chromosomal fragment extending from at least 500 kb upstream of SNPs rsl 6905070, rs7814049 and rs7013830, to at least 450 kb downstream of SNPs rsl6905070, rs7814049 and rs7013830.
  • a nucleotide sequence that is in linkage disequilibrium with SNPs rsl6905070, rs7814049 and rs7013830 is a nucleotide sequence comprised in a chromosomal fragment extending from ST3GAL1 to ZFAT.
  • Another particularly preferred genetic marker for longevity is a nucleic acid molecule comprising a nucleotide sequence that is in linkage disequilibrium with an SNP selected from the group consisting of the SNPs rs4513644 rs4700233, rs854050 and rs4700231.
  • a nucleotide sequence that is in linkage disequilibrium with SNPs rs4513644 rs4700233, rs854050 and rs4700231 is a nucleotide sequence present at chromosome 5ql 1.2.
  • a nucleotide sequence that is in linkage disequilibrium with SNPs rs4513644 rs4700233, rs854050 and rs4700231 is a nucleotide sequence comprised in a chromosomal fragment extending from at least 400 kb upstream of SNPs rs4513644 rs4700233, rs854050 and rs4700231, to at least 650 kb downstream of SNPs rs4513644 rs4700233, rs854050 and rs4700231.
  • a nucleotide sequence that is in linkage disequilibrium with SNPs rs4513644 rs4700233, rs854050 and rs4700231 is a nucleotide sequence comprised in a chromosomal fragment extending from ACTBL2 to PLK2. If the nucleic acid molecule does not comprise the polymorphism (SNP) itself, it is preferred that nucleic acid molecule comprises at least a nucleotide sequence in the proximity of the polymorphism (SNP), i.e.
  • nucleic acid molecule comprises at least a nucleotide sequence immediately adjacent to the polymorphism (SNP).
  • nucleotide sequence that is in linkage disequilibrium with an SNP as herein defined as a nucleotide sequence showing an r 2 within 0.4, 0.5, 0.6, 0.7, 0.8, 0.9, or 1.0 and/or D' within 0.6, 0.7, 0.8, 0.9, or 1.0 with a SNP identified in Table 9 and where r 2 and D' are measures indicating the extent of the LD between markers.
  • a nucleotide sequence that is in linkage disequilibrium with an SNP as herein defined is thus at least physically linked to the SNP and will preferably be present in the vicinity of the SNP, whereby the vicinity of the SNP is understood to mean within no more than 750, 500, 200, 100, 50, 20, 10 or 5 kb from the chromosomal location of the SNP.
  • a preferred nucleotide sequence that is in linkage disequilibrium with an SNP as herein defined and that is physically linked to the SNP is an expressed nucleotide sequence of a gene as listed in Table 2.1.2.
  • a more preferred nucleotide sequence that is in linkage disequilibrium with an SNP as herein defined and that is physically linked to the SNP is an expressed nucleotide sequence of a gene as listed in Table 2.1.5.
  • the genetic marker is a nucleic acid molecule comprising a nucleotide sequence that is differentially expressed between a population that expresses excess survival and a control population.
  • a population that expresses excess survival is herein defined as a population that has a lifespan of or above 85 years.
  • a nucleotide sequence that is differentially expressed between a population that expresses excess survival and a control population preferably is a nucleotide sequence that is differentially expressed between one or more of the cohorts of the Leiden Longevity Study and their corresponding control populations (samples) as described in the Examples herein.
  • a differentially expressed genetic marker for longevity may be useful in the methods of the present invention without knowledge of the polymorphism that underlies the difference in expression.
  • a preferred differentially expressed genetic marker for longevity is a nucleic acid molecule comprising at least 10, 12, 15, 20, 25 or 30 contiguous nucleotides from a transcript (or complement thereof) that specifically hybridises to a probe selected from the group consisting of GE488443 (KALRN), GE83396 (C3orf26), GE4871(NFIA), GE88135 (TCF4), GE 785523 (METT5D1), GE624013 and GE535567 (MARCHIII), GE749029 and GE57513 (SNRPN).
  • Genbank accession no.'s, Unigene No.'s of expressed sequences identified by these probes are given in Tables 2.4.2 and 2.4.3.
  • nucleic acid molecule comprising at least 10, 12, 15, 20, 25 or 30 contiguous nucleotides from a nucleotide sequence that has at least 80, 85, 90, 95, 98, 99% sequence identity with a nucleotide sequence with a Genbank accession no. as provide in Tables 2.4.2 and 2.4.3.
  • a particularly preferred differentially expressed genetic marker for longevity is a nucleic acid molecule comprising at least 10, 12, 15, 20, 25 or 30 contiguous nucleotides from a transcript (or complement thereof) having at least 80, 85, 90, 95, 98, 99% sequence identity with a nucleotide sequence selected from the group consisting of SEQ ID NO: 120 (ST3GAL1), SEQ ID NO: 121 (ZFAT), SEQ ID NO: 122 (ACTBL2) and SEQ ID NO: 123 (PLK2).
  • a most preferred differentially expressed genetic marker for longevity is a nucleic acid molecule comprising at least 10, 12, 15, 20, 25 or 30 contiguous nucleotides from a transcript (or complement thereof) having at least 80, 85, 90, 95, 98, 99% sequence identity with the nucleotide sequence of SEQ ID NO: 120 (ST3GAL1).
  • gene and SNP information as used herein and in the Tables is expressed by gene symbols, gene locations, polymorphism names and their identifications in public SNP database (dbSNP ID), and the genotypes using the standard expression used in molecular biology, and they are readily understood by one of ordinary skill in the art.
  • Gene symbol is the acronym or abbreviation corresponding to a given gene name. Genes and markers may have multiple symbols and names due to rediscovery or correlation to function following discovery.
  • a nucleotide sequence of the invention that is a genetic marker for longevity may be a nucleotide sequence of which a human allele is associated with genetic predisposition for deficiency in a health function, such as e.g.
  • risk factors for these diseases such as high serum cholesterol, triglycerides, blood pressure etc., ) or other parameters of impaired functions of brain, heart, endocrine systems involved in metabolism, skeleton, muscles .
  • a portfolio of markers for use in the invention may contain any number of two or more and any type of marker as disclosed herein.
  • the markers in the portfolio are genetic markers as disclosed herein.
  • One embodiment of the invention relates to a portfolio comprising at least 2, 3, 4, 5, 6, 8, 10, 12, 15, 20 or 30 (isolated) nucleotide sequences or their complements, wherein each nucleotide sequence is in linkage disequilibrium with an SNP selected from the group consisting of the SNPs listed in any one of Tables 3.1 and 2.1.1 to 2.1.4.
  • the portfolio comprises at least 2, 3, 4, 5, 6, 8, 10, 12, 15, 20 or 30 (isolated) nucleotide sequences or their complements, wherein each nucleotide sequence is in linkage disequilibrium with an SNP selected from the group consisting of the SNPs listed in Table 2.1.5 or 2.1.6.
  • the portfolio comprises at least two nucleotide sequences, which at least two nucleotide sequences are in linkage disequilibrium with at least two different SNPs listed in any one of Tables 3.1 and 2.1.1 to 2.1.4, whereby at least one nucleotide sequence is in linkage disequilibrium with a SNP selected from the group consisting of the SNPs rsl6905070, rs7814049, rs7013830, rs4513644 rs4700233, rs854050 and rs4700231.
  • the polymorphic nucleotide sequences are in linkage disequilibrium with the minor allele of the SNPs.
  • the portfolio comprises at least two nucleotide sequences selected the group consisting of: i) nucleotide sequences comprising at least 10 contiguous nucleotides from a transcript or complement thereof that specifically hybridises to a probe selected from the group consisting of probes having a nucleotide sequences of SEQ ID NO: 95 - 103; and, ii) nucleotide sequences that specifically hybridise to a transcript having at least 80% sequence identity with at least one nucleotide sequence selected from the group consisting of SEQ ID NO: 120, SEQ ID NO: 121, SEQ ID NO: 122, SEQ ID NO: 123 and their complements; whereby the portfolio comprises at least one nucleotide sequence selected from ii).
  • each nucleotide sequence in the portfolio is in linkage disequilibrium with a different SNP in the group.
  • one or more of the nucleotide sequences themselves comprise a polymorphism in the human population. This polymorphism may comprise an SNP as defined above.
  • the nucleotide sequences comprise at least 10, 12, 15, 20, 25 or 30 contiguous nucleotides from a nucleotide sequence that has at least 80, 85, 90, 95, 98, 99% sequence identity with a nucleotide sequence as defined in any one of SEQ ID NO.'s comprising an SNP listed in the above-mentioned Tables (see Table 3.1).
  • the nucleotide sequences comprises at least 10, 12, 15, 20, 25 or 30 contiguous nucleotides from a nucleotide sequence that has at least 80, 85, 90, 95, 98, 99% sequence identity with a nucleotide sequence as defined in any one of SEQ ID NO.'s: 1 - 94, more preferably the nucleotide sequences comprise at least 10, 12, 15, 20, 25 or 30 contiguous nucleotides from a nucleotide sequence that has at least 80, 85, 90, 95, 98, 99% sequence identity with a nucleotide sequence as defined in any one of SEQ ID NO.'s: 17, 18, 22, 23, 32, 39, 41, 44, 45, 56, 83 and 84.
  • the nucleotide sequences in the portfolio are expressed nucleotide sequences.
  • the invention relates to the use of any of the markers for longevity as defined herein in any of the methods of the invention.
  • the invention relates to the use of the longevity markers of the invention in methods for determining genetic predisposition for longevity, in methods of screening for a substance that modulates the biological aging rate (and promotes/inhibits a healthy aging process), or a substance that is capable of modulation of longevity and/or life expectancy, and/or in methods for assessing the physiological age of (a sample from) a subject.
  • the invention in a fourth aspect relates to a method for determining a genetic predisposition for longevity. It is herein understood that methods for determining a genetic predisposition for longevity more generally include methods for determining a genetic predisposition for a certain life expectancy, including both a genetic predisposition against longevity as well as a genetic predisposition for longevity and all intermediate phenotypes.
  • the methods for determining genetic predisposition for longevity of a person (or a subject) preferably is an ex vivo method, e.g. performed in vitro on a sample obtained from the subject.
  • the method preferably comprises determining the genotype of the subject with respect to one or more genetic markers of longevity as herein defined above.
  • the method comprises detecting the presence of a polymorphism that is in linkage disequilibrium with an SNP selected from the group consisting of the SNPs listed in any one of Tables 2.1.1 to 2.1.4, more preferably a polymorphism that is in linkage disequilibrium with an SNP selected from the group consisting of the SNPs listed in Table 2.1.5 or 2.1.6, wherein the presence of the polymorphism is indicative of longevity.
  • the method comprises detecting the presence of a polymorphism selected from the group consisting of the SNPs listed in any one of Tables 2.1.1 to 2.1.4., more preferably a SNP selected from the group consisting of the SNPs listed in Table 2.1.5 or 2.1.6, wherein the presence of the polymorphism is indicative of longevity.
  • a genotype that is indicative of longevity is a carrier of an allele that is indicative of longevity i.e. a carrier of a protective allele.
  • This may be deduced from the odds ratios and the indications of major and minor alleles for each SNP as presented in Table 2.1.1.
  • an odds ratios (OR) greater than one indicates that a given minor allele from a polymorphism in Table 2.1.1 is the allele that is indicative of longevity
  • odds ratios less than one indicate that a given minor allele is the allele that is indicative of mortality.
  • the allele that is indicative of longevity may also be referred to as the protective allele. For a given allele that is indicative of longevity (i.e.
  • the other alternative allele at the SNP position may be considered a mortality and/or disease risk allele.
  • the other alternative allele at the SNP position may be considered an allele that is indicative of longevity (i.e. the allele protective against mortality and disease).
  • the genotype "a carrier of the protective allele of a given SNP" is understood to mean a genotype that is homozygous or heterozygous for the protective allele of that SNP.
  • the present methods may be performed using any known biological or biochemical method in which genetic polymorphisms, such as SNPs, can be detected or visualized.
  • genetic polymorphisms such as SNPs
  • Such methods include, but are not limited to, DNA sequencing, allele specific PCR, PCR amplification followed by an allele/mutant specific restriction digestion, oligonucleotide ligation assays, primer hybridization and primer extension assays, optionally combined with or facilitated by microarray analysis.
  • Alternative methods for determining allelic variants and gene polymorphisms are readily available to the skilled person in the art of molecular diagnostics.
  • oligonucleotides capable of hybridizing to sequences in or flanking genes (e.g., polymorphic regions) involved in adenosine metabolism, and the use of these oligonucleotides for performing these methods.
  • Primers may be designed to amplify (e.g., by PCR) at least a fragment of a gene encoding an adenosine metabolism- associated enzyme.
  • a polymorphism may be present within the amplified sequence and may be detected by, for example, a restriction enzyme digestion or hybridization assay.
  • the polymorphism may also be located at the 3' end of the primer or oligonucleotide, thus providing means for an allele or polymorphism specific amplification, primer extension or oligonucleotide ligation reaction, optionally with a labelled nucleotide or oligonucleotide.
  • the label may be an enzyme (e.g., alkaline phosphatase, horseradish peroxidase), radiolabel ( 32 P, 33 P, 3 H, 125 1, 35 S etc.), a fluorescent label (Cy3, Cy5, GFP, EGFP, FITC, TRITC and the like) or a hapten/ligand (e.g., digoxigenin, biotin, HA, etc.).
  • the detection is carried out using oligonucleotides physically linked to a solid support, and may be performed in a microarray format.
  • a kit comprising one or more oligonucleotides capable of hybridizing to, or adjacent to, any of the polymorphic sites in any genetic markers as defined hereinabove.
  • the oligonucleotide(s) may be provided in solid form, in solution or attached on a solid carrier such as a DNA microarray.
  • the kit may provide detection means, containers comprising solutions and/or enzymes and a manual with instructions for use.
  • the method for determining genetic predisposition for longevity of a subject comprises determining the expression level of a nucleotide sequence that is in linkage disequilibrium with a SNP selected from the group consisting of the SNPs listed in Table 2.1.1.
  • Preferred examples of expressed nucleotide sequences that are in linkage disequilibrium with the SNPs listed in Table 2.1.1 are provide in Tables 2.1.2, 2.1.5, 2.1.7, 2.4.2 and 2.4.3.
  • the expression level is determined of a nucleotide sequence that is in linkage disequilibrium with an SNP selected from the group consisting of the SNPs listed in Table 2.1.1 to 2.1.4, or yet more preferably in Table 2.1.5 or 2.1.6.
  • the expression level is determined of a transcript (or complement thereof) that specifically hybridises to a probe selected from the group consisting of GE488443 (KALRN), GE83396 (C3orf26), GE4871(NFIA), GE88135 (TCF4), GE 785523 (METT5D1), GE624013 and GE535567 (MARCHIII), GE749029 and GE57513 (SNRPN).
  • Genbank accession no.'s, Unigene no.'s of expressed sequences identified by these probes are given in Tables 2.4.2 and 2.4.3.
  • the expression level can be determined of a transcript that has at least 80, 85, 90, 95, 98, 99% sequence identity with a nucleotide sequence with a Genbank accession no. as provide in Tables 2.4.2 and 2.4.3.
  • the expression level is determined of a transcript (or complement thereof) having at least 80, 85, 90, 95, 98, 99% sequence identity with a nucleotide sequence selected from the group consisting of SEQ ID NO: 120 (ST3GAL1), SEQ ID NO: 121 (ZFAT), SEQ ID NO: 122 (ACTBL2) and SEQ ID NO: 123 (PLK2).
  • the expression level is determined of a transcript (or complement thereof) having at least 80, 85, 90, 95, 98, 99% sequence identity with the nucleotide sequence of SEQ ID NO: 120 (ST3GAL1).
  • Expressed nucleotide sequence (or nucleotide sequences) that are indicative of longevity are preferably those nucleotide sequences that are differentially expressed as up regulated or down regulated (in cells) in a sample from a population that expresses excess survival as compared to corresponding (cells) in a sample from a control population.
  • a modulated nucleotide sequence is (part of) a gene that is differentially expressed between one or more of the cohorts of the Leiden Longevity Study and their corresponding control populations (samples) as described in the Examples herein.
  • Up regulation and down regulation are relative terms meaning that a detectable difference (beyond the contribution of noise in the system used to measure it) is found in the amount of expression of the nucleotide sequences relative to a baseline.
  • a baseline preferably comes from a pool of subjects that do not express excess survival.
  • a pool of these subjects preferably contains 1, 3, 5, 10, 20, 30, 100, 400, 500, 600 or more subjects.
  • the expression level of a differentially expressed nucleotide sequence that is indicative of longevity is then considered either up regulated or down regulated relative to a baseline level using the same measurement method.
  • the assessment of the expression level of a nucleotide sequence in order to assess whether a gene/nucleotide sequence is modulated is preferably performed using classical molecular biology techniques to detect mRNA levels, such as (real time) reverse transcriptase PCR (whether quantitative or semi-quantitative), mRNA (micro)array analysis or Northern blot analysis, or other methods to detect RNA.
  • the expression level of a nucleotide sequence is determined indirectly by quantifying the amount of the polypeptide encoded by the gene/nucleotide sequence. Quantifying an amount of polypeptide may be carried out by any known technique. Preferably, an amount of polypeptide is quantified by Western blotting.
  • the quantification of a substrate of the corresponding polypeptides or of any compound known to be associated with the function of the corresponding polypeptides or the quantification of the function or activity of the corresponding polypeptide using a specific assay is encompassed within the scope of the prognosticating method of the invention.
  • the assessment of the expression level of a nucleotide sequence is carried out using (micro)arrays as defined herein.
  • the expression levels of a nucleotide sequence and/or amounts of a corresponding polypeptide are preferably measured in a sample from a subject.
  • the expression level (of a nucleotide sequence or polypeptide) is determined ex vivo in a sample obtained from a subject.
  • a sample may be liquid, semi- liquid, semi-solid or solid.
  • a preferred sample comprises 100 or more cells and/or a tissue from a subject to be tested taken in a biopsy.
  • a sample may comprises blood of a subject.
  • the skilled person knows how to isolate and optionally purify RNA and/or protein present in such a sample. In case of RNA, the skilled person may further amplify it using known techniques.
  • An increase (or upregulation) (which is synonymous with a higher expression level) or decrease (or downregulation) (which is synonymous with a lower expression level) of the expression level of a nucleotide sequence (or steady state level of the encoded polypeptide) is preferably defined as being a detectable change of the expression level of the nucleotide sequence (or steady state level of the encoded polypeptide or any detectable change in the biological activity of the polypeptide) using a method as defined earlier on as compared to the expression level of the corresponding nucleotide sequence (or steady state level of the corresponding encoded polypeptide) in a baseline.
  • an increase or decrease of a polypeptide activity is quantified using a specific assay for the polypeptide activity.
  • an increase of the expression level of a nucleotide sequence means an increase of at least 2, 3, 4, 5, 6, 7, 8, or 9% of the expression level of the nucleotide sequence using arrays. More preferably, an increase of the expression level of a nucleotide sequence means an increase of at least 10 or 11%, even more preferably at least 20%, at least 30%, at least 40%, at least 50%, at least 70%, at least 90%, at least 150% or more.
  • a decrease of the expression level of a nucleotide sequence means a decrease of at least 2, 3, 4, 5, 6, 7, 8, or 9% of the expression level of the nucleotide sequence using arrays. More preferably, a decrease of the expression level of a nucleotide sequence means an decrease of at least 10%, even more preferably at least 20%., at least 30%, at least 40%, at least 50%, at least 70%, at least 90%, at least 150% or more.
  • an increase of the expression level of a polypeptide means an increase of at least 2, 3, 4, 5, 6, 7, 8, or 9% of the expression level of the polypeptide using western blotting.
  • an increase of the expression level of a polypeptide means an increase of at least 10%, even more preferably at least 20%, at least 30%, at least 40%, at least 50%, at least 70%, at least 90%, at least 150% or more.
  • a decrease of the expression level of a polypeptide means a decrease of at least 2, 3, 4, 5, 6, 7, 8, or 9% of the expression level of the polypeptide using western blotting.
  • a decrease of the expression level of a polypeptide means a decrease of at least 10%, even more preferably at least 20%, at least 30%, at least 40%, at least 50%, at least 70%, at least 90%, at least 150% or more.
  • an increase of the polypeptide activity means an increase of at least 2,
  • an increase of the polypeptide activity means an increase of at least 10%, even more preferably at least 20%, at least 30%, at least 40%, at least 50%, at least 70%, at least 90%, at least 150% or more.
  • a decrease of the polypeptide activity means a decrease of at least 2, 3,
  • a decrease of the polypeptide activity means a decrease of at least 10%, even more preferably at least 20%, at least 30%, at least 40%, at least 50%, at least 70%, at least 90%, at least 150% or more.
  • a transcript or complement thereof
  • a probe selected from the group consisting of GE488443 (KALRN), GE83396 (C3orf26), GE4871 (NFIA), GE88135 (TCF4), GE 785523 (METT5D1), GE624013 and GE535567 (MARCHIII), GE749029 and GE57513 (SNRPN) as a function of the age and sex of a subject is indicative of longevity.
  • up- or down-regulation can be determined of a transcript that has at least 80, 85, 90, 95, 98, 99% sequence identity with a nucleotide sequence with a Genbank accession no. as provide in Table 2.4.2 and 2.4.3.
  • the up- or down-regulation of a transcript having at least 80, 85, 90, 95, 98, 99% sequence identity with a nucleotide sequence selected from the group consisting of SEQ ID NO: 120 (ST3GAL1), SEQ ID NO: 121 (ZFAT), SEQ ID NO: 122 (ACTBL2) and SEQ ID NO: 123 (PLK2), as a function of the age and sex of a subject is indicative of longevity.
  • the up- or down-regulation of a transcript (or complement thereof) having at least 80, 85, 90, 95, 98, 99% sequence identity with the nucleotide sequence of SEQ ID NO: 120 (ST3GAL1), as a function of the age and sex of a subject is indicative of longevity.
  • the invention in a fifth aspect relates to a method for assessing physiological age of a subject, preferably a human subject.
  • the rate of aging is very species specific, where a human may be aged at about 50 years; and a rodent at about 2 years.
  • a natural progressive decline in body systems starts in early adulthood, but in humans it becomes most evident several decades later.
  • One arbitrary way to define old age more precisely in humans is to say that it begins at conventional retirement age, around about 60, around about 65 years of age.
  • Another definition sets parameters for aging coincident with the loss of reproductive ability, which is around about age 45, more usually around about 50 in humans, but will, however, vary with the individual. It has been found that individuals age at different rates, even within a species.
  • the method for assessing physiological age of a subject preferably is performed or measured in a sample from a subject, Thus, preferably the physiological age is assessed ex vivo in a sample obtained from a subject.
  • the method preferably comprises: a) determining expression information in a sample obtained from the subject, of one or more differentially expressed nucleotide sequences as herein defined above, preferably expression information of a portfolio of expressed nucleotide sequences as herein defined above; b) using the expression information to generate an age signature for the sample; and, c) comparing the age signature obtained in b) with a control age signature; wherein a statistically significant match with a positive control or a statistically significant difference from a negative control is indicative of age in the sample.
  • Method for generating an age signature for a sample, for determining control age signatures and for determining statistically significant matches of differences with controls are described in detail in US20070161022, which is incorporated by reference herein.
  • the invention in a sixth aspect relates to a method for identification of a substance that modulates (or is capable of modulating) the biological aging rate in a subject, and/or that modulates (or is capable of modulating) longevity and/or life expectancy.
  • the method preferably comprises the steps of a) contacting the substance to a test cell or administering the substance to a test organism; b) determining in the test cell or in (a sample from) the test organism the expression level of one or more differentially expressed nucleotide sequences as herein defined above, preferably determining the expression level of a portfolio of expressed nucleotide sequences as herein defined above; c) comparing the expression level of the nucleotide sequence(s) with the expression level of the corresponding nucleotide sequence(s) in a test cell that is not contacted with the substance or in (a sample from) the test organism that is not contacted with the substance; and, d) identifying a substance that produces a difference in expression level of at least one of
  • a substance is identified as a substance the promotes longevity, extends lifespan and/or improves health, when the substance upregulates a nucleotide sequence that is upregulated in a population that expresses excess survival or when the substance downregulates a nucleotide sequence that is downregulated in a population that expresses excess survival.
  • the up- or down-regulation respectively in a population that expresses excess survival means that a nucleotide sequence is up- or down-regulated respectively in a population of cells or organisms that expresses excess survival as compared to a corresponding control population (e.g. one or more of the cohorts of the Leiden Longevity Study and their corresponding control populations as described in the Examples herein).
  • the expression level of the nucleotide acid sequence may be determined indirectly by quantifying the amount of polypeptide encoded by the nucleotide acid sequence.
  • the cells in the test cell population are preferably mammalian cells, preferably human cells.
  • the test cell population that is contacted with the substance and the test cell population that is not contacted with the substance are derived from one cell population, preferably from one cell line, more preferably from one cell.
  • the test organism may is a non-human mammal or a human volunteer.
  • the invention in a seventh aspect relates to a method of improving health of a subject.
  • the method preferably comprises: a) determining a genetic predisposition for longevity using a method as defined herein above; and, b) if the subject does not have the genetic predisposition for longevity, 1) providing, to the subject, a substance that modulates (or is capable of modulating) the biological aging rate identified in a method as described above; or 2) providing to the subject other medication or other dosages of medication on the basis of the biological age established by the methods of the invention method as compared the medication or dosage thereof normally provided to the subject.
  • the determination of the genetic predisposition for longevity in a) comprises determining in the test cell or in (a sample from) the test organism the expression level of one or more differentially expressed nucleotide sequences as herein defined above, more preferably determining the expression level of a portfolio of expressed nucleotide sequences as herein defined above; whereby in b) a substance is provided to the subject that modulates expression or activity of the (differentially) expressed nucleotide sequence(s) or its gene product(s).
  • the substance upregulates expression or activity of those (differentially) expressed nucleotide sequence(s) or its gene product(s) that are upregulated in a population that expresses excess survival or that the substance downregulates expression or activity of those (differentially) expressed nucleotide sequence(s) or its gene product(s) a nucleotide sequence that are downregulated in a population that expresses excess survival.
  • LLC 18 Longevity Study (LLS 18 ), Leiden 85plus Study 20 ' 21 , Danish 1905 cohort 22 and PROSPER 23 were published previously and is provided together with numbers and description of mortality data below. Numbers of cases and controls are summarized in
  • Inclusion criteria called for men and women between the ages of 70 and 82 years with a total plasma cholesterol of 155- 350 mg/dL (4-9/mmol/L) and triglyceride levels ⁇ 200 mg/dL (6 mmol/L). Patients were excluded if they showed signs of cognitive decline, indicated by a score of 23 or less on the Mini Mental State Examination and a series of psychometric tests. The study population was distributed evenly with respect to existing vascular disease and qualifying risk factors. Patients were followed every 3 months for an average of 3.2 years. The primary composite endpoint, definite or suspected death from coronary heart disease (CHD), nonfatal myocardial infarction (MI), or fatal/nonfatal stroke, was measured at 3-year follow-up. A total of 604 subjects died during follow up. In statistical analysis adjustments for study cohort as well as use of Pravastatin were made. 1.1.5 German cohort
  • the unrelated German study participants were drawn from population-based collections and comprised 1447 long-lived individuals of exceptional age (810 nonagenarians, 637 centenarians; age range: 95 - 110 years; mean age: 99 years) and 1104 younger control subjects (age range: 60-75 years; mean age: 67 years) (Nebel, A. et al, 2005, Proc. Natl. Acad. Sci. U. S. A 102, 7906-7909).
  • the gender ratio was about 75% females vs. 25% males.
  • the controls match the long-lived individuals in terms of ancestry, gender and geographical origin within the country and genetic differences between the case-control samples are considered to be very low (Wilicox, B. J. et al., 2008, Proc. Natl. Acad. Sci. U. S. A 105, 13987-13992).
  • the survival benefit of these families is marked by a 30 % excess survival observed in the proband, the parental and the offspring generation (Schoenmaker et al., 2006, supra).
  • the male offspring of the long-lived subjects have a lower prevalence of diabetes and cardiovascular disease as compared to their partners.
  • the offspring has lower serum glucose than their partners and beneficial lipoprotein profiles in the sense that on average offspring has significantly larger LDL particle sizes than partners, a feature even stronger represented in the nonagenarian siblings (Heijmans et al., 2006, PLoS Med. 2006 Dec; 3 ⁇ 12):e495).
  • 1.2 Genotyping A flow chart of the approach used to identify SNPs associated to survival is provided in Figure 1 and a description of population is provided in Table 1.1.
  • Genotyping for stage 1 of the GWAS was performed at Perlegen Sciences by applying the first generation genome-wide SNA array Affymetrix Gene Chip Human Mapping 500K Array set comprising two arrays (262K + 238K) and for stage 2 by using an in-house Perlegen platform.
  • a MAF > 0.02 a successful call rate > 80%
  • P HWE ⁇ 10 "4 In total 357,162 SNPs were used for GWAS analysis with a mean genotype call rate of 95%. Genotype data were used to confirm sex and family relationships.
  • stage 1 genotypes due to population substructure was assessed by clustering analysis of IBS matrix using Plink. One cluster was identified indicating that the Leiden Longevity Study is a homogenous population. The quantile-quantile plot showed that the p-value distribution of stage 1 conformed to a null distribution at all but the extreme tail (data not shown) The genomic inflation factor ( ⁇ ), which measures over-dispersion of test statistics from association tests indicating population stratification was 1.019.
  • Stage 2 SNPs were selected for analyses when the mean call rate > 95%, MAF > 0.02 and P HWE > 0.01. Genotyping quality control was performed using duplicate DNA samples within the LLS and SNP assays of the Sequenom MassARRAY platform to confirm genotyping accuracy of SNPs genotyped in stage 1 and 2 combined for which 99.67% concordant results were obtained. Genotyping for replication studies was performed with MassARRAY Iplex (Sequenom, San Diego). Four i-plexes containing 96 out of 104 prioritised SNPs could be designed for replication studies 81 of which were successfully typed in 100% of the samples at a genotype call rate of .
  • stage 1 of the GWAS comparing genotype frequencies between cases and controls we applied the Cochrane- Armitage test for additive effects and the Fishers exact test for recessive effects.
  • stage 1 of the GWAS comparing genotype frequencies between cases and controls
  • Table 1.1 we applied the Cochrane- Armitage test for additive effects and the Fishers exact test for recessive effects.
  • SNPs with P-value below 0.01 for the trend were selected for the second stage.
  • Restricted space on the chip for stage 2 allowed us to select a limited number of additional SNPs for which reason SNPs associating in the recessive model at a p-value below 0.005 for Fishers exact test were selected for genotyping in the second stage.
  • stage 1 and 2 For the joint analysis of stage 1 and 2 a variance-modified version of these score- tests was used for additive and recessive effects. The relatedness between the highly aged sibling cases was taken into account when computing the variances of the scores 25 . Unless otherwise stated, subsequent association analyses of the SNPs in additional cohorts were restricted to only the genetic model corresponding to the most significant result in the GWAS. Odds ratios were estimated and corresponding 95% confidence intervals were computed based on empirical standard errors. For meta-analyses a fixed effect approach was used. Scores and their variances were computed within each study and combined across the three studies to obtain a single meta-statistic. P-values below 5 x 10 ⁇ 8 were considered as genome wide significant 26 . Heterogeneity between studies was assessed by estimating the between study variance using random effects meta analysis.
  • RNA expression analysis For testing association to survival of disease susceptibility alleles, the same methods as for the GWAS and meta-analysis were used. A linear regression model with the number of disease risk alleles as outcome and longevity status as covariate was performed. For testing within the Leiden Longevity Study, empirical standard errors were used. To search for a set of single SNPs predicting survival, a cross-validation based model selection algorithm was applied 27 . In addition to main effects the method also considers all pairwise interactions. 1.4 RNA expression analysis
  • RNA yield was evaluated on the 2100 Bioanalyzer (Agilent Technologies) and the concentration was measured using a NanoDrop spectrophotometer (NanoDrop Technologies). Quality criteria included a 28S/18S ratio as measured by the Bioanalyzer of at least 1.2, and a total RNA yield of at least 3 ⁇ g.
  • the samples were hybridized on 54k CodeLink Human Whole Genome Bioarrays (GE Healthcare, currently of Applied Micro arrays). Images were quantified with CodeLink Expression software (version 4.0). For the analysis of expression levels, 11 Codelink Bioarray probes were selected on their correspondence to the ST3GAL1 (7 probes) or ZFAT locus (4 probes).
  • ST3GAL1 and one ZFAT probe were present in an exon of these genes, while the other probes corresponded to one or more UniGene Cluster or Expressed Sequence Tags (ESTs) within intronic regions 1.5 Selection for a set of disease susceptibility alleles
  • a GWAS was performed in two stages in subjects of the Leiden Longevity Study (LLS) and controls.
  • LLC Leiden Longevity Study
  • genotype data of 357,162 SNPs that passed quality control were analysed in a comparison of 417 unrelated probands from a nonagenarian sibling pair (94 years on average) and 470 controls (60 years on average, i.e. partners of offspring).
  • a flow chart of the consecutive analysis steps is depicted in Figure 1 and a description of the population samples investigated in the GWAS and subsequent replication studies is given in Table 1.1.
  • the association analysis for survival into old age is based only on the age difference between cases and controls, not on health status.
  • the nearest genes of the SNPs associating with p ⁇ 10 "4 are depicted in Table 2.1.2.
  • a multiplex genotyping assay could be designed and successful typing was obtained in the replication cohorts. Association analysis was performed comparing 1236 cases of a mean age of 87 years from the Leiden 85 Plus Study and 1644 cases of a mean age of 93 years from the Danish 1905 Cohort and appropriate populations of younger controls (Table 1.1). Both are population based cohort studies from a genetic background similar to the LLS cohort 30 .
  • the survival-associated SNPs at 8q24.22 cover a chromosome region of 72 kb that contains no known protein coding gene located within a 100 kb distance of the SNPs. Since long range cis-acting regulating variation has been documented 31 we tested whether the expression of the nearest genes to the chromosome 8q24.22 SNPs, including ZFAT, 391kb upstream of rsl6905070 and ST 3 GALl at 515 kb downstream of the top SNP, correlated to the variation. Gene expression micro-array data were available from 50 unrelated long-lived siblings from LLS families, 50 of their offspring and 50 controls 32 .
  • Seven probes on the microarray corresponded to the ST3GAL1 genome region and four probes to the ZFA T region.
  • the expression levels of these probes were compared between 60 carriers (nonagenarians, offspring and partners combined) and 90 non-carriers of any of the minor alleles of the three 8q24.22 SNPs.
  • the effect size and direction was similar in siblings, offspring and partner groups separately and comparable but non-significant increases were observed for the other five probes targeting this locus (Table 2.1.7) of which one is present in a known exon.
  • RNA expression analysis 150 RNA samples belonging to 50 trios (50 unrelated nonagenarian siblings, 50 of their offspring, 50 partners of this offspring) were analysed using CodeLink microarrays. The characteristics of these samples are summarized in Table 2.4.1. A T- test analysis showed that none of the variables in Table 2.4.1 differ significantly between offspring and partners (which was one of the criteria for selecting the 50 trios), but that several parameters differ between sibs and offspring or partners (on which item the trios were not selected).
  • the gene expression data files were read into R (from Bioconductor) by using the 'Codelink' package. Normalisation was performed using the Cyclic Lowess method, resulting in a Spearman correlation between all samples of 0.97 and 0.98-0.99 between the technical replicates. In total 87.7% of the probes on the CodeLink array (53.423 in total) was expressed in at least 10% of the samples and 66% of the probes were expressed in at least 90% of the samples indicating that a major part of the genes on the array are expressed in blood. We tested which transcripts were differentially expressed with age, comparing the nonagenarian sibs to partners of offspring. Corrected for gender 3127 transcripts were found.
  • the SNP is located on 8q24.22 and was identified in a meta-analysis of GWAS candidate SNPs in the Leiden Longevity Study (LLS) and two population-based cohort studies, consisting in total of 3700 highly aged and 4153 middle-aged controls.
  • LLS Leiden Longevity Study
  • P 6.63 x 10 "9 ).
  • the minor allele associated to a 33% decreased probability of becoming long-lived.
  • Prospective analysis of survival data in an additional large cohort revealed that this allele indeed associated from seventy years onwards with an increased mortality risk due to cancer and cardiovascular causes.
  • Pleiotropic effects of genetic variation have previously been reported such as for variation at HFNlB influencing the risk of prostate cancer and diabetes 33 .
  • the offspring of the nonagenarian siblings in the LLS have a decreased prevalence of myocardial infarction and hypertension 19 as compared to partners, whereas the somewhat older offspring of centenarians have a decreased prevalence of both cancer and cardiovascular disease 5 .
  • the proximal chromosome band 8q24.21 has frequently been implicated in the risk of various types of cancer (prostate, ovary, breast, bladder 34 ), however, none of the 8q24.22 SNPs in our study are in LD with the cancer associated SNPs on 8q24.21, 6Mb centromeric of the 8q24.22 SNP. Consistent asssociations of genetic variation to survival in the general population have not frequently been described 9 . This was further illustrated by applying the same cross-sectional study design to test the effect on survival into old age for human longevity of 22 GWAS-identified susceptibility loci for coronary artery disease, cancer and type-2 diabetes.
  • GWAS identified SNPs have a relatively low predictive value for disease risk and explain only a small fraction of the heritability of the traits involved 35 . It would be expected that such alleles only marginally affect population-wide survival.
  • the subjects surviving into old age may carry protective genetic variation or be subject to environmental features counteracting the disease promoting effect of disease susceptibility alleles.
  • Carriers of the minor alleles of any of the three 8q24.22 SNPs were found to have an 1.11-fold increased expression of mRNA probes covering the ST3GAL1 gene located 515 kb from rsl6905070. Small long-range effects were also observed for variation in the 5pl3 gene desert that correlated with expression of the PTGER4 gene at a 270 kb distance 31 .
  • the STSGALl gene encodes ST3 beta-galactoside alpha-2,3-sialyltransferase 1, a member of glycosyltransferase family 29.
  • This type II membrane protein catalyzes the transfer of sialic acid from CMP-sialic acid to galactose-containing substrates and sialic acid modulates immune interactions.
  • ST3Gal I modulates surface sialylated structures during the generation of dendritic cells from monocytes 36 .
  • Dendritic cells are antigen-presenting cells with high endocytic capacity that play a central role in immune regulation. The regulation of glycosylation has been implicated in immune responsiveness, multiple diseases and aging 37 .
  • Kenyon C The plasticity of aging: insights from long-lived mutants. Cell 2005; 120(4):449-460.
  • apolipoprotein E gene is a "frailty gene," not a "longevity gene”. Genet Epidemiol 2000; 19(3):202-210.
  • TCF7L2 Transcription factor 7-like 2
  • Raskin,L. et al. FGFR2 is a breast cancer susceptibility gene in Jewish and Arab Israeli populations. Cancer Epidemiol. Biomarkers Prev. 17, 1060-1065 (2008).
  • CAD coronary artery disease
  • T2D Type-2 diabetes
  • HF heart failure
  • BC breast cancer
  • PC prostate cancer
  • CC colon carcinoma
  • LC lung cancer
  • CVD cardiovascular disease.
  • SNPs selected for replication analysis associating at p ⁇ 6.43X10 " 4 to longevity in the GWAS analysis of LLS stage 1 and 2 combined. Chromosome position according to build 36. Position according to dbSNPbuild 129. MAF indicates minimal allele frequency in all 953 Dutch controls. Major/minor refers to the allele with the highest or lowest frequency in controls. Ptrend, Precessive refers to the p value obtained in either the additive or recessive model. OR indicates Odds Ratio of the most significant model. OR's above 1 indicate the increased probability to become long-lived based on the minor allele being overrepresented in the elderly as compared to young controls. OR's below 1 indicate the opposite. Supplementary Table 3.1 present the sequences of the SNP identifiers of the prioritized SNPs.
  • CodeLink Bioarray probes were selected on basis of their correspondence to the ST3GAL1 or ZFAT locus.
  • One ST3GAL1 (GE57639) and one ZFAT (GE86200) probe were present in an exon of these genes, while the other probes corresponded to one (or more) Expressed Sequence Tags (ESTs) within intronic regions. Expression differences within these probes were analyzed using Linear regression in Stata/SE 8.0 between 60 carriers and 90 non-carriers of one or more of the 8q24.22 SNPs.
  • FC Fold Change
  • a M indicates that the risk allele contributes to metabolic disease (CAD or T2D); C indicates that the risk allele contributes to cancer.
  • the major allele (A) has been associated with risk for cancer, while the minor allele G) has been associated with T2D.
  • c The major allele is indicated in bold.
  • Z) Logistic regression with long-lived/control status as outcome, the study as covariate and the SNP genotypes as independent variable (Stata/SE 8.0). Analyses were performed with robust standard errors to take into account family dependency in the Leiden Longevity Study.
  • Table 2.4.1 Medians of parameters measured in donors of the 150 RNA samples that were used in the microarray comparison study. *: p ⁇ 0.05 between long-lived sibs and offspring or partners. None of the parameters is significantly different between partners and offspring.
  • Lymphocytes E-9 cells/L* 2.01 2.03 1.19
  • Eosinophils E- 10 cells/L 1.63 1.84 1.56
  • RNA from 150 subjects from the Leiden Longevity study results from the microarray comparison of RNA from 150 subjects from the Leiden Longevity study. Probes that are 1) differentially expressed between offspring of long-lived siblings and their partners and 2) reside in transcripts in which SNPs were associated with longevity in the Leiden Longevity Study with p-value ⁇ 0.05 (Combination expression/GWA) .
  • GE Probe ID which is a unique identifier for the probe sequence in the CodeLink WEBB database. This is an internal GE Healthcare relational database that held all gene associated annotations and linked them to the specific codelink probe ID.

Landscapes

  • Chemical & Material Sciences (AREA)
  • Life Sciences & Earth Sciences (AREA)
  • Proteomics, Peptides & Aminoacids (AREA)
  • Health & Medical Sciences (AREA)
  • Organic Chemistry (AREA)
  • Wood Science & Technology (AREA)
  • Analytical Chemistry (AREA)
  • Zoology (AREA)
  • Engineering & Computer Science (AREA)
  • Immunology (AREA)
  • Genetics & Genomics (AREA)
  • Bioinformatics & Cheminformatics (AREA)
  • Microbiology (AREA)
  • Molecular Biology (AREA)
  • Biotechnology (AREA)
  • Biophysics (AREA)
  • Biochemistry (AREA)
  • Physics & Mathematics (AREA)
  • General Engineering & Computer Science (AREA)
  • General Health & Medical Sciences (AREA)
  • Pathology (AREA)
  • Cell Biology (AREA)
  • Measuring Or Testing Involving Enzymes Or Micro-Organisms (AREA)

Abstract

La présente invention concerne des marqueurs génétiques et biochimiques de longévité, qui ont été identifiés comme variation génétique commune associée à la longévité dans des cohortes de sujets âgés. Les marqueurs génétiques de longévité de l’invention concernent en particulier une collection de molécules d’acide nucléique comprenant une séquence de nucléotides qui est en déséquilibre de liaison avec un SNP associé à la longévité dans des cohortes de sujets âgés. Dans un autre mode de réalisation de l’invention, les marqueurs génétiques de longévité sont des molécules d’acide nucléique qui comprennent des séquences de nucléotides qui sont exprimées différemment entre une population qui exprime une survie en excès et une population témoin. L’invention concerne, dans un autre aspect, un portefeuille comprenant des sous-ensembles des marqueurs génétiques de longévité. Les marqueurs génétiques de longévité de l’invention peuvent être utilisés dans des procédés de détermination de la prédisposition génétique à la longévité, des procédés de criblage d’une substance qui module le taux de vieillissement biologique ou une substance qui est capable de modulation de la longévité et/ou de l’espérance de vie, et/ou des procédés d’évaluation de l’âge physiologique.
PCT/NL2009/050409 2008-07-07 2009-07-07 Nouveaux indicateurs de longévité humaine et du taux de vieillissement biologique WO2010005303A2 (fr)

Applications Claiming Priority (2)

Application Number Priority Date Filing Date Title
EP08159827.8 2008-07-07
EP08159827 2008-07-07

Publications (2)

Publication Number Publication Date
WO2010005303A2 true WO2010005303A2 (fr) 2010-01-14
WO2010005303A3 WO2010005303A3 (fr) 2010-04-29

Family

ID=39938277

Family Applications (1)

Application Number Title Priority Date Filing Date
PCT/NL2009/050409 WO2010005303A2 (fr) 2008-07-07 2009-07-07 Nouveaux indicateurs de longévité humaine et du taux de vieillissement biologique

Country Status (1)

Country Link
WO (1) WO2010005303A2 (fr)

Citations (7)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO1998048785A2 (fr) * 1997-04-29 1998-11-05 Kenneth Blum, Inc. Diagnostic d'un syndrome d'insatisfaction a l'aide de polygene allelique et traitement associe
WO2002014552A2 (fr) * 2000-08-11 2002-02-21 Children's Medical Center Corporation Loci genetiques indicateurs de la propension a la longevite et procedes d'identification de la propension a la resistance aux maladies liees a l'age
WO2004050898A2 (fr) * 2002-12-04 2004-06-17 Elixir Pharmaceuticals, Inc. Composants de la voie ampk
US20040259104A1 (en) * 2002-10-22 2004-12-23 Lamberts Steven Willem Jan Test
US20060068432A1 (en) * 2003-03-20 2006-03-30 Nir Barzilai Biological markers for longevity and diseases and uses thereof
US20070161022A1 (en) * 2005-11-30 2007-07-12 Kim Stuart K Signatures for human aging
WO2007131345A1 (fr) * 2006-05-12 2007-11-22 The Hospital For Sick Children Facteur de risque génétique dans les gènes sod1 et sfrs15 dans la néphropathie, la cataracte diabétique, la maladie cardiovasculaire et la longévité

Patent Citations (7)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO1998048785A2 (fr) * 1997-04-29 1998-11-05 Kenneth Blum, Inc. Diagnostic d'un syndrome d'insatisfaction a l'aide de polygene allelique et traitement associe
WO2002014552A2 (fr) * 2000-08-11 2002-02-21 Children's Medical Center Corporation Loci genetiques indicateurs de la propension a la longevite et procedes d'identification de la propension a la resistance aux maladies liees a l'age
US20040259104A1 (en) * 2002-10-22 2004-12-23 Lamberts Steven Willem Jan Test
WO2004050898A2 (fr) * 2002-12-04 2004-06-17 Elixir Pharmaceuticals, Inc. Composants de la voie ampk
US20060068432A1 (en) * 2003-03-20 2006-03-30 Nir Barzilai Biological markers for longevity and diseases and uses thereof
US20070161022A1 (en) * 2005-11-30 2007-07-12 Kim Stuart K Signatures for human aging
WO2007131345A1 (fr) * 2006-05-12 2007-11-22 The Hospital For Sick Children Facteur de risque génétique dans les gènes sod1 et sfrs15 dans la néphropathie, la cataracte diabétique, la maladie cardiovasculaire et la longévité

Non-Patent Citations (6)

* Cited by examiner, † Cited by third party
Title
AKATSUKI KOKAZE ET AL: "Longevity-associated mitochondrial DNA 5178 C/A polymorphism and its interaction with cigarette consumption are associated with pulmonary function in middle-aged Japanese men" JOURNAL OF HUMAN GENETICS, SPRINGER-VERLAG, TO, vol. 52, no. 8, 17 July 2007 (2007-07-17), pages 680-685, XP019520274 ISSN: 1435-232X *
CAPRI ET AL: "Human longevity within an evolutionary perspective: The peculiar paradigm of a post-reproductive genetics" EXPERIMENTAL GERONTOLOGY, ELSEVIER SCIENCE, OXFORD, GB, vol. 43, no. 2, 17 January 2008 (2008-01-17), pages 53-60, XP022424984 ISSN: 0531-5565 *
DATABASE EMBL [Online] 16 May 2001 (2001-05-16), "602723676T1 NIH_MGC_113 Homo sapiens cDNA clone IMAGE:4849987 3', mRNA sequence." XP002554551 retrieved from EBI accession no. EMBL:BG746166 Database accession no. BG746166 *
DATABASE EMBL [Online] 29 December 1999 (1999-12-29), "xr04c06.x1 NCI_CGAP_Brn53 Homo sapiens cDNA clone IMAGE:2759146 3', mRNA sequence." XP002554550 retrieved from EBI accession no. EMBL:AW264502 Database accession no. AW264502 *
IMUSCE FRANCESCHI ET AL: "The extreme longevity: The state of the art in Italy" EXPERIMENTAL GERONTOLOGY, ELSEVIER SCIENCE, OXFORD, GB, vol. 43, no. 2, 17 January 2008 (2008-01-17), pages 45-52, XP022424983 ISSN: 0531-5565 *
REINER ET AL: "Common promoter polymorphisms of inflammation and thrombosis genes and longevity in older adults: The cardiovascular health study" ATHEROSCLEROSIS, ELSEVIER IRELAND LTD, IE, vol. 181, no. 1, 1 July 2005 (2005-07-01), pages 175-183, XP005270342 ISSN: 0021-9150 *

Also Published As

Publication number Publication date
WO2010005303A3 (fr) 2010-04-29

Similar Documents

Publication Publication Date Title
KR101546058B1 (ko) 대사증후군 예측용 ｓｎｐ 마커 및 이의 용도
Saarela et al. PRKCA and multiple sclerosis: association in two independent populations
KR101536213B1 (ko) 복부비만 예측용 ｓｎｐ 마커 및 이의 용도
KR101532308B1 (ko) 복부비만 예측용 ｓｎｐ 마커 및 이의 용도
KR102543907B1 (ko) 치주질환 위험도 평가용 유전자 마커
US20030032099A1 (en) Methods for predicting susceptibility to obesity and obesity-associated health problems
US10731219B1 (en) Method for preventing progression to metabolic syndrome
US20090092987A1 (en) Polymorphic Nucleic Acids Associated With Colorectal Cancer And Uses Thereof
KR101546069B1 (ko) 대사증후군 예측용 ｓｎｐ 마커 및 이의 용도
WO2009055596A2 (fr) Procédés d'utilisation de variants génétiques permettant de diagnostiquer et de prédire un syndrome métabolique et des traits associés
WO2010005303A2 (fr) Nouveaux indicateurs de longévité humaine et du taux de vieillissement biologique
JP2006296270A (ja) Ｐｒｋａａ２遺伝子多型による２型糖尿病発症素因の検出方法
JP2007517511A (ja) ヒトチオプリンｓ−メチルトランスフェラーゼ欠損に関連するハプロタイプおよび多型
KR101304535B1 (ko) Ｋｌｏｔｈｏ 유전자의 단일염기다형을 이용한 심혈관계 질환 예측 방법
KR101092580B1 (ko) 위암 감수성 예측용 ｖｃａｎ 다형성 마커 및 이를 이용한위암 감수성 예측 방법
KR101543774B1 (ko) 복부비만 예측용 ｓｎｐ 마커 및 이의 용도
KR101546070B1 (ko) 대사증후군 예측용 ｓｎｐ 마커 및 이의 용도
KR20110011306A (ko) 텔로미어 유지 유전자를 이용한 폐암 감수성 진단용 마커 및 폐암 감수성 예측 및 판단 방법
WO2009053513A1 (fr) Procédé et kit de détermination d'une prédisposition, d'un risque de développement ou du diagnostic du psoriasis
US20040076975A1 (en) Methods for assessing the risk of non-insulin-dependent diabetes mellitus based on allelic variations in the 5'-flanking region of the insulin gene and body fat
KR20240057772A (ko) 우울증 진단용 단일염기다형성 및 이의 용도
KR20150115700A (ko) 한국인의 고혈압 예측용 ｓｎｐ 마커
KR20150092937A (ko) 한국인의 고혈압 예측용 ｓｎｐ 마커
US20100184839A1 (en) Allelic polymorphism associated with diabetes
KR20220113305A (ko) 단일염기다형성을 이용한 안지오텐신 전환효소억제제 이상반응 진단용 조성물 및 이를 이용한 방법

Legal Events

Date Code Title Description
121 Ep: the epo has been informed by wipo that ep was designated in this application

Ref document number: 09788216

Country of ref document: EP

Kind code of ref document: A2

NENP Non-entry into the national phase in:

Ref country code: DE

122 Ep: pct application non-entry in european phase

Ref document number: 09788216

Country of ref document: EP

Kind code of ref document: A2