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WO2003062465A1 - Composes pour le diagnostic de la predisposition a l'osteoporose - Google Patents

Composes pour le diagnostic de la predisposition a l'osteoporose Download PDF

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WO2003062465A1
WO2003062465A1 PCT/ES2003/000034 ES0300034W WO03062465A1 WO 2003062465 A1 WO2003062465 A1 WO 2003062465A1 ES 0300034 W ES0300034 W ES 0300034W WO 03062465 A1 WO03062465 A1 WO 03062465A1
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polymorphism
regulatory region
osteoporosis
predisposition
seq
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PCT/ES2003/000034
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English (en)
Spanish (es)
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Natalia GARCÍ GIRALT
Francesc Xavier NOGUÉS SOLAN
Leonardo Gabriel Mellibovsky Saidler
Susana Balcells Comas
Adolfo DÍEZ PÉREZ
Daniel Raúl GRINBERG VAISMAN
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Universidad De Barcelona
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Publication of WO2003062465A1 publication Critical patent/WO2003062465A1/fr

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    • 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
    • C12Q2600/156—Polymorphic or mutational markers

Definitions

  • This invention relates to diagnostic methods and kits based on polymorphisms in a collagen gene. Specifically, it refers to methods and kits for the diagnosis of predisposition to certain pathological conditions, analyzing the presence of these polymorphisms. More specifically it refers to the diagnosis of predisposition to osteoporosis.
  • the invention also relates to the collagen gene that contains the polymorphisms.
  • Osteoporosis is a common condition characterized by a reduced bone mass, a deterioration of the microarchitecture of bone tissue and an increased risk of fracture.
  • the risk of osteoporotic fracture is related to bone mineral density (BMD).
  • BMD bone mineral density
  • Several inherited and environmental factors are involved in the pathogenesis of osteoporosis. Studies using families and pairs of twins indicate that osteoporosis has a substantial genetic component. The contribution of genetic factors to the variance of bone mineral density is between 65% and 92% in twin studies.
  • Type I collagen is the most abundant protein in the bone matrix. Mutations in the coding regions of the genes encoding the two chains of type I collagen (COL1A1 and COL1A2) result in a severe autosomal dominant pediatric condition called osteogenesis imperfecta. Unlike the severe effects produced by mutations located in the coding regions of type I collagen genes, changes in their regulatory regions may be responsible for subtle differences in gene expression in response to extracellular signals. This, in turn, can result in differences in BMD and fracture risk.
  • GT polymorphism has been described in intron 1 of the COL1A1 gene, which is within a recognition site for Sp1 transcription factor, and has been shown to be associated with BMD, both in the spine lumbar as in the neck of the femur, or with fractures (cf. US 5,922,542; SF Grant et al., Nat. Genet. 1996, vol. 14, pp. 203-5).
  • some studies have not been able to reproduce these results (cf. A. Heegaard et al., Calcif. Tissue Int. 2000, vol. 66, pp. 409-13; M. Liden, Calcif. Tissue Int. 1998, vol. 63, pp. 293-5; P. Garnero et al., J. Bone Miner Res. 1998, vol. 13, pp. 813-7).
  • the treatments currently available for osteoporosis include mainly hormone replacement therapy, bisphosphonates such as alendronate, selective estrogen receptor modulators, calcium, vitamin D metabolites or parathyroid hormone. These treatments generally manage to stop the progressive decrease in BMD characteristic of women suffering from this disease, but, in general, they are not able to reverse osteoporosis; that is, they are not able to induce an increase in BMD in patients. Thus, it is especially interesting to be able to identify those individuals with a predisposition or greater susceptibility to osteoporosis. In such a case, appropriate therapy could be applied before the effects of osteoporosis appear.
  • the present invention is the result of the search for new polymorphisms in the promoter of the COL1 A1 gene, in a region between nucleotide positions -2300 to -1500, which is relevant for its expression in osteoblasts in vivo.
  • Two new nucleotide exchange polymorphisms (“single nucleotide polymorphisms", SNPs), one of which shows association with BMD of the lumbar spine, have been found in a cohort of 256 Spanish postmenopausal women and the other is in a strong linkage disequilibrium (“linkage disequilibrium”) with the previously described GT polymorphism that shows to be associated with BMD, both in the lumbar spine and in the femoral neck (cf. US 5,922,542).
  • This invention provides diagnostic methods, compounds and kits for detecting individuals who have a predisposition or susceptibility to certain disease states, in particular, osteoporosis or low BMD.
  • the invention allows to identify human individuals who have such predisposition or susceptibility, identifying those individuals with an altered collagen 1A1 gene.
  • the invention also provides a therapy for those individuals who have a predisposition or susceptibility to conditions. Pathological mentioned above.
  • a first aspect of the invention provides a method of diagnosing the predisposition of a human individual to osteoporosis or a high probability of having low bone mineral density (BMD), which comprises detecting - in a sample of fluid or tissue obtained from said individual- the presence of a polymorphism selected from the group formed by an indel T polymorphism in the -1663 position and a polymorphism of change from G to T in the -1997 position in the regulatory region of the COL1A1 gene.
  • BMD bone mineral density
  • the diagnostic method involves examining an individual at risk of suffering a condition or disease by correlating it with the presence of one or both polymorphisms (that is, the presence of the less frequent allele of each SNP).
  • a diagnosis can use the polymerase chain reaction (PCR) or the single strand conformational polymorphism test, or both, and can determine whether an individual possesses a wild collagen gene or a polymorphism.
  • PCR polymerase chain reaction
  • each individual can be homozygous for the wild allele, heterozygous for the wild allele and polymorphism, or homozygous for the polymorphism.
  • Different genotypes correlate with a different predisposition to osteoporosis or a high probability of having low BMD.
  • the diagnostic method also includes the use of indicator means that react in the presence of one of the polymorphisms.
  • the indicating means typically induces that a detectable signal is produced in the presence of the polymorphism, and can induce a color change or a coagulation or a restriction site, detectable by additional analytical steps.
  • the indicator means also, can comprise an antibody that has binding affinity that distinguishes between the wild sequence and a polymorphism.
  • the diagnostic method comprises detecting the presence of a G to T polymorphism in the -1997 position in a regulatory region of the COL1A1 gene. In a preferred embodiment, this detection is carried out using PCR primers that amplify a segment of DNA that contains the nucleotide at the -1997 position in the regulatory region of the COL1A1 gene. In a more preferred embodiment a pair of PCR primers with the sequences SEQ ID NO: 5 and SEQ ID NO: 6 is used.
  • the diagnostic method comprises detecting the presence of an indel T polymorphism at position -1663 in the regulatory region of the COL1A1 gene.
  • this detection is carried out using PCR primers that amplify a segment of DNA containing the nucleotide at the -1663 position in the regulatory region of the COL1A1 gene.
  • a pair of PCR primers with the sequences SEQ ID NO: 2 and SEQ ID NO: 3 is used.
  • a second aspect of the invention provides diagnostic methods adapted to determine the presence of a polymorphism in a collagen gene.
  • the diagnostic method comprises a pair of PCR primers that amplify a segment of DNA from the regulatory region of the COL1 A1 gene, where the amplified DNA segment is up to 800 base pairs in length - preferably up to 500, more preferably up to 250- and the nucleotide position -1997 of the regulatory region of the COL1A1 gene is substantially towards the middle of the DNA segment.
  • the PCR primers are selected from the group consisting of SEQ ID NO: 5 and SEQ ID NO: 6.
  • the diagnostic method comprises a pair of PCR primers that amplify a segment of DNA from the regulatory region of the COL1A1 gene, where the amplified DNA segment is up to 800 base pairs in length - preferably up to 500, more preferably up to 500 250- and the nucleotide position -1663 of the regulatory region of the COL1A1 gene is substantially towards the middle of the DNA segment.
  • the PCR primers are selected from the group consisting of SEQ ID NO: 2 and SEQ ID NO: 3.
  • the invention also provides a diagnostic kit for the predisposition of a human individual to osteoporosis or a high probability of having low bone mineral density (BMD), comprising diagnostic means according to the second aspect of the invention, optionally within of a container
  • the diagnostic kit comprises a transport means such as a compartmentalized box that accommodates the diagnostic means according to the invention, optionally within a container such as a vial, a tube, a vial or something similar.
  • a third aspect of the invention provides a collagen gene, or a fragment thereof, in which, or a guanosine nucleotide at the -1997 position is replaced by a thymidine nucleotide, or a series of eight thymidines in the range of -1663 to -1670 is replaced by a series of seven thymidines in the range of -1663 to -1669, or both.
  • the gene or the gene fragment is an isolated DNA molecule, free of other naturally occurring DNA molecules naturally associated with it.
  • the isolated DNA molecule comprises a regulatory region of the collagen gene in which a guanosine in the -1997 position is replaced by a thymidine; in a preferred case this isolated DNA molecule comprises the oligonucleotide of SEQ ID NO: 4.
  • the isolated DNA molecule comprises a collagen gene regulatory region in which a series of eight thymidines in the range of -1663 a -1670 is replaced by a series of seven thymidines in the range of -1663 to -1669; In a preferred case this isolated DNA molecule comprises the oligonucleotide of SEQ ID NO: 1.
  • a fourth aspect of the invention provides a method of osteoporosis therapy comprising: the examination of a human individual for the genetic predisposition to osteoporosis; and, if said predisposition is identified, the treatment of said individual to prevent or reduce osteoporosis or delay the onset of osteoporosis, where the predisposition to osteoporosis correlates with a polymorphism of a COL1A1 gene of said individual, selected from the group formed by a polymorphism of G to T in the -1997 position of the regulatory region of the COL1A1 gene, and an indel T polymorphism in the -1663 position of the regulatory region of the COL1A1 gene.
  • the predisposition to osteoporosis correlates with a G to T polymorphism at the -1997 position of the regulatory region of the COL1A1 gene. In other embodiment, such predisposition correlates with an indel T polymorphism at position -1663 of the regulatory region of the COL1A1 gene.
  • the treatment of the individual is performed by hormone replacement therapy; In another preferred case for each of these two embodiments, the treatment of the individual is performed by an antiosteoporotic agent selected from the group consisting of: bisphosphonates, selective estrogen receptor modulators, calcium, vitamin D metabolites and parathyroid hormone.
  • an individual who is heterozygous for both polymorphisms is classified as having the lowest risk (as illustrated in group D of FIG. 2).
  • An individual who is either G / G + I / I, or G / G + D / D, or G / T + l / l is Classify as having a moderate risk.
  • an individual who is either G / G + l / D is classified in the highest risk category.
  • the risk is assessed in reference to both the presence of one of the two polymorphisms of the collagen gene, as well as other known indications (genetic, physiological, nutritional, etc.).
  • the invention provides additional information on which the risk of an individual can be based.
  • FIG. 1 shows the strategy followed for the search and genotyping of the new polymorphisms in the promoter of the COL1A1 gene.
  • A Position of the three amplimers used in the search. The +1 position corresponds to the transcription start site. The relevant region for osteoblastic expression in vivo is indicated.
  • B Genotyping of polymorphism -1663indelT by an improved SSCP method. The three genotypes are distinguished at the single chain level (ss). In addition, heterozygous (ID) individuals are easily distinguished from homozygous by the formation of heteroduplex molecules at the level of double chains (ds).
  • C Genotyping of PCOL2 polymorphism by restriction analysis.
  • the genotypes ee (homozygous for the Eco31 l site), EE (homozygous without the site) and Ee (heterozygous) correspond to GG, TT and GT, respectively.
  • the first lane corresponds to the molecular weight marker. The size, in base pairs, is shown on the right.
  • FIG. 2 represents the analysis of compound genotypes and haplotypes.
  • A Mean BMD values for the five groups (called A through E) of composite genotypes that have frequencies greater than 5%.
  • FIG. 3 illustrates the electrophoretic mobility shift test ("Electrophoretic Mobility Shift Assays", EMSA) using the PCOL1-I probe and a nuclear extract of primary osteoblasts.
  • A Scheme of the three different probes used: double chain, direct single chain (“forward"), and single reverse chain (“reverse”).
  • B A double chain PCOL1-I probe was incubated in the presence (+) or absence (-) of the extract. The competitors are indicated above.
  • the triangle encompasses the three lanes in which increasing amounts of PCOL1-l-ds cold probe (molar excesses of 10 3 , 10 4 and 10 5 of cold probe with respect to labeled probe) were added.
  • the triangle encompasses the two lanes in which increasing amounts of PCOL1-l-ss cold probe (10 3 and 10 4 molar excesses of cold probe with respect to the marked one) were added.
  • Each of the non-specific competitors (PCOL1-l-ss direct -8A- and GRE-ss inverse) is at a molar excess of 10 4 .
  • the arrow indicates a band that probably corresponds to the double chain generated by the hybridization of the two complementary PCOL1-l-ss oligonucleotides.
  • FIG. 4 shows an EMSA using PCOL2-G probes and an osteoblastic nuclear extract.
  • A Scheme of the three different probes used: double chain, direct single chain ("forward"), and single reverse chain (“reverse”).
  • B A double-chain PCOL2-G probe was incubated in the absence (-) or presence (+) of the extract. The competitors are indicated above.
  • the triangle encompasses the four lanes in which increasing amounts of double-stranded PCOL2 cold oligonucleotide were added. From left to right, the alleles and molar excess are the following: G-10 3 , G-10 4 , T-10 4 and G-2x10 4 .
  • each of the non-specific competitors is in a molar excess of 10 4 .
  • C Comparison of the binding capabilities of the double-chain and single-chain PCOL2-G probes. The labeled double-stranded PCOL2-G oligonucleotides (probe 1), single straight chain (probe 2) and reverse single strand (probe 3) in the absence (-) or presence (+) of the extract were incubated.
  • D A PCOL2-G reverse single chain probe was incubated in the absence (-) or presence (+) of the extract. The competitors are indicated above.
  • the triangle encompasses the two lanes in which increasing amounts of PCOL2-G-ss cold oligonucleotide are added (molar excesses of 10 3 and 10 4 ).
  • Each of the non-specific competitors (Sp1-ds and reverse GRE-ss) is in a molar excess of 10 4 .
  • FIG. 5 shows a comparison of the binding capacities of the two alleles (G and T) of the -1997 position.
  • a reverse PCOL2-G-SS (C * ) probe was incubated in the absence (-) or presence (+) of the osteoblastic nuclear extract.
  • Lanes 3 through 8 contain increasing amounts (molar excesses of 10 2 , 10 3 and 10 4 ) of the inverse PCOL2-G-ss probe (C) (lanes 3 to 5) or of the inverse PCOL2-T-ss probe (A) (lanes 6 to 8).
  • Bone mineral density (BMD) of the lumbar spine (L2-L4) and non-dominant femur neck was measured using dual x-ray absorptiometry (Hologic QDR 4500 SL ® , Hologig, Waltham, MA, USA). BMD was expressed in g / cm 2 .
  • Genomic DNA was obtained from blood leukocytes using the Kit Wizard (Promega, Madison, Wl, U.S.A.) for genomic DNA purification.
  • the promoter region of COL1A1 comprising positions -2292 to -1499 in three fragments of about 300 bp overlapping each other was amplified by PCR (FIG. 1), using the primers described in TABLE 1.
  • the amplification reactions contained 100 ng of genomic DNA, 20 pmol of each primer, 10 nmol of each dNTP, 1.25 U of Taq polymerase (Promega, Madison, Wl, USA) and MgCI 2 at 1.5 mM (fragment 1) or at 2 mM (fragments 2 and 3) in one volume final 50 ⁇ l.
  • the single chain conformation polymorphisms (SSCP) assay was performed as follows: 4-5 ⁇ l of each PCR reaction was mixed with 7 ⁇ l of STOP buffer (95% formamide, 10 mM EDTA, 0 xylene cyanol , 05%, bromophenol blue 0.05%), heat denaturation at 80 ° C for 3 min and cool on ice before electrophoresis at 150-200 V in 8% polyacrylamide gels (acrylamide: bisacrylamide 29: one).
  • the optimal SSCP conditions for polymorphisms -1663indelT (in fragment 1) and -1997 G / T (in fragment 2) were at room temperature and 4 ° C, respectively. For the visualization of the bands, the gels were dyed with silver.
  • Polymorphism -1663NdelT is type with an improved PCR-SSCP protocol, as follows: a 180 bp fragment (which becomes 179 bp in the polymorphism) was amplified using the PCOL1 F primer of SEQ ID NO: 2 (5 'TAGCCCCTGCAGTCTCCCTC 3') and primer PCOL1 R of SEQ ID NO: 3 (5 'AAGATTCCATTGCCTCCCCC 3') under the following conditions: 10 pmol of each primer, 5 nmol of each dNTP, 0.7 U of Taq polymerase (Promega, Madison, Wl, USA), MgCI 2 to 1, 5 mM and 100 ng of genomic DNA in a final volume of 25 ⁇ l.
  • the program consisted of an initial denaturation step of 1 min at 94 ° C, 35 cycles of 40 s at 94 ° C and 30 s at 62 ° C, followed by a final step of 5 min at 72 ° C.
  • the PCR products were prepared for loading on SSCP gels as described above and were subjected to electrophoresis in 12% polyacrylamide gels containing 5% glycerol at room temperature and 140 V. The bands were revealed by silver staining .
  • the -1997 G / T polymorphism is type using a PCR-RFLP (restriction fragment length polymorphism) strategy, as follows: DNA samples were subjected to PCR amplification of fragment 2 with primers F2 and R2 which are shown in TABLE 1, and were subsequently digested with the restriction enzyme Eco31 l. The digestion products were resolved in 8% polyacrylamide gels. The most frequent allele appears as two bands of 212 and 81 bp, while the less frequent remains undigested (293 bp).
  • PCR-RFLP restriction fragment length polymorphism
  • Human bone cells were obtained from surgical samples of healthy subjects undergoing surgery for acute trauma and free of bone remodeling disease.
  • a protocol based on a described method was used (PJ Marie et al., In Vitro Cell. Dev. Biol. 1989, vol. 25, pp. 373-80) with some modifications (M. Nacher et al., Bone Miner. 1994, vol. 26, pp. 231-43; C. Garcia-Moreno et al., Bone 1998, vol. 22, pp. 233-9).
  • a culture of primary osteoblasts was established from a pool of cells from four donors.
  • DMEM Dulbecco-modified Eagle medium
  • FCS fetal bovine serum
  • Nuclear extracts were prepared from confluent cells according to a published protocol (E. Schreiber et al., Nucleic Acids Res. 1989, vol. 17, p. 6419), using a modified C buffer (10% glycerol , MgCl 2 to 1.5 mM). Protein concentrations were determined by the method Bradford, and the nuclear extracts were stored at -80 ° C until use. Testing of electrophoretic mobility change (EMSA)
  • underlined bases correspond to the polymorphic sites.
  • the corresponding complementary probes were also synthesized.
  • the double chain probe was obtained by hybridization of the complementary oligonucleotides. All probes were labeled at the end with [ ⁇ - 32 P] ATP using the T4 polynucleotide kinase (Promega, Madison, Wl, USA).
  • Binding reactions normally contain 10 ⁇ g of nuclear extract, 2 ⁇ g of double stranded poly (dl-dC) (Amersham Pharmacia Biotech, Freiburg, Germany), 6 ⁇ g of acetylated BSA and 50,000 cpm of radioactively labeled probe.
  • the mixture is incubated for 15-20 min at 37 ° C in a buffer containing 20 mM HEPES and pH 7.9, 60 mM KCI, 1 mM EDTA, 1 mM DTT and 10% glycerol in one volume total of 20 ⁇ l.
  • the DNA-protein complexes were resolved from the free probes in non-denaturing gels of 5% polyacrylamide (29: 1) containing 2.5% glycerol.
  • Electrophoresis was performed at 4 ° C in TBExl buffer at 15-18 mA for approximately 2 h. The gels were dried under vacuum and exposed to X-ray films with intensifying screens at -70 ° C for 24-48 hours. In competition assays, binding reactions were made in the presence of an excess of unlabeled competing oligonucleotide, as indicated in each case.
  • the oligonucleotides containing the binding sites of known transcription factors (Sp1 and the glucocorticoid receptor) were: Sp1 5 'ATTCGATCGGGGCGGGGCGAGC 3' (SEQ ID NO: 13)
  • a new polymorphism was detected in fragment 1.
  • Four of the seven control individuals showed a different SSCP pattern, detected mostly in the heteroduplex region of the gel. Sequencing revealed a deletion of a T nucleotide in a series of eight T residues, located at positions -1663 to -1670. The allele that carried 8 Ts was called I (by the insertion of a residue T) and the one carrying 7 Ts was called D (by the deletion of a T), and the polymorphism was called -1663indelT.
  • I by the insertion of a residue T
  • D by the deletion of a T
  • the polymorphism was called -1663indelT.
  • To establish genotypic frequencies an improved SSCP strategy was followed (see above), using a smaller PCR fragment (FIG. 1B). Allelic and genotypic frequencies for this polymorphism have been established in 256 women (TABLE 2).
  • Sp1 polymorphism in intron 1 of COL1A1 is type.
  • TABLE 2 shows the genotypic and allelic frequencies.
  • genotypes and BMD of the lumbar spine in 256 postmenopausal women were analyzed.
  • the mean BMD value, the standard deviation (SD) and the range in our sample were 0.894 g / cm 2 , 0.135 and 0.567-1, 297.
  • the mean for each of the genotypes of -1663indelT, -1997 G / T and Sp1 is shown in TABLE 3.
  • Genotypes II 155 GG 194 SS 161 (0.60) (0.76) (0.63)
  • BMD means ⁇ SD of the individuals carrying the different genotypes -1663indelT, -1997 G / T and Sp1.
  • the BMD means of the lumbar spine of compound genotypes that include all three polymorphic sites were also analyzed. Only genotypes that had frequencies greater than 5% (5 of the 27 possible combinations) were included.
  • FIG. 2 shows the means of BMD for these five compound genotypes, called A to E.
  • the highest average BMD (0.982 ⁇ 0.186) corresponded to group D (triple heterozygous), while the lowest value (0.853 ⁇ 0.137) corresponded to group C
  • a competition test was performed (FIG. 4D). The addition of a 10 4 molar excess of both the single chain GRE oligonucleotide and the double chain Sp1 oligonucleotide showed no competition with respect to the delayed band.
  • FIG. 5 shows an EMSA in which the labeled probe was the single chain oligonucleotide carrying the C (PCOL2-G, reverse chain), and the competitors were the single cold chain carrying the C (molar excesses of 10 3 , 10 4 and 10 5 ), or the same amounts of the simple cold chain carrying the A (reverse chain of the T allele).
  • a competition difference was detected, which is best observed when comparing lanes 4 and 7 (molar excesses of 10 4 ).
  • 1663indelT and Sp1 suggests that -1663indelT contributes to the effect of Sp1 that has been detected in some populations.

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Abstract

Deux nouveaux polymorphismes de changement d'un nucleótide (SNPs) ont été mis en évidence dans la zone régulatrice du gène COL1A1. Des études réalisées chez des femmes postménopausiques ont révélé que le polymorphisme de changement d'une guanine G à une thymine T en position -1997 est associé à la densité minérale osseuse (DMO) de la colonne lombaire et du col du fémur. Le polymorphisme -1663indelT, qui correspond a une délétion d'une T d'une série de huit T (-1670a -1663), est en fort déséquilibre de liaison avec ledit polymorphisme Sp1 de l'intron 1 qui s'est avéré être associé à la DMO. Des interactions significatives ont été établies entre -1997 G/T et -1663indelT, et entre -1997 G/T et Sp1. L'invention est utile dans le diagnostic et dans le traitement de l'ostéoporose, particulièrement en tant que kit de diagnostic comprenant de nouveaux amorceurs de PCR.
PCT/ES2003/000034 2002-01-25 2003-01-23 Composes pour le diagnostic de la predisposition a l'osteoporose WO2003062465A1 (fr)

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* Cited by examiner, † Cited by third party
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EP2521793A4 (fr) * 2010-01-06 2013-07-03 Univ Florida Procédés et trousses pour détecter des facteurs de risque pour le développement d'une ostéonécrose des mâchoires, et procédés pour leur traitement

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