- Email: [email protected]
Association between the low density lipoprotein receptor-related protein (LRP) and Alzheimer's disease
Neuroscience Letters 227 (1997) 68–70
Association between the low density lipoprotein receptor-related protein (LRP) and Alzheimer’s disease Fabienne Wavrant-DeVrie`ze a , b ,*, Jordi Pe´rez-Tur a, Jean-Charles Lambert b, Bernard Frigard c, Florence Pasquier d, Andre´ Delacourte e, Philippe Amouyel b, John Hardy a, Marie-Christine Chartier-Harlin b a
Mayo Clinic Jacksonville, 4500 San Pablo Road, Jacksonville, FL 32224, USA Service de Sante´ Publique et d’Epidemiologie, CJF INSERM 95-05, Institut Pasteur de Lille, 1 rue Prof. Calmette, 59019 Lille cedex, France c Centre de ge´riatrie de Wasquehal Molinel, rue Salvador Allende, B.P. 165, 59444 Wasquehal, France d CHR et U de Lille, Clinique Neurologique, Centre de la me´moire, Hoˆpital Salengro, 59037 Lille cedex, France e Unite´ Inserm 422, 1 Place de Verdun, 59045 Lille cedex, France
b
Received 2 April 1997; revised version received 18 April 1997; accepted 18 April 1997
Abstract Alzheimer’s disease (AD) is the most common neurodegenerative disorder affecting elderly people. It usually occurs after 65 years old (late-onset AD). The e4 allele of apolipoprotein E (APOE) gene is a risk factor which contributes about 50% of the genetic risk for this form of the disease. The low density lipoprotein receptor-related protein (LRP) is a major receptor for APOE which is found in the senile plaques of AD brains. This makes it a good candidate gene for the disease. There is a polymorphism in the region upstream of the LRP gene that has been associated with AD in an American population. We examined this polymorphism by restriction fragment length polymorphism analysis in a French population with sporadic late-onset AD. In the previous report, a significant increase of the 87 bp allele was found in the AD cases; however, in our population, we observed a significant decrease with this same allele of the LRP gene. The possible reasons for this discrepancy, linkage disequilibrium or statistical anomaly, are discussed. 1997 Elsevier Science Ireland Ltd. Keywords: Late-onset Alzheimer’s disease; Low density lipoprotein receptor-related protein; Apolipoprotein E receptor; Association studies; Risk factor
Alzheimer’s disease (AD) is the most common neurodegenerative disorder affecting elderly people. Several genes have been reported to influence the occurrence of the disease [4], but the only established risk factor for the typical late-onset form of the disease is genetic variability at the apolipoprotein E (APOE) gene. Genetic variability at this locus has been shown to have a strong association with the disease [14]. The APOE gene is located on the chromosome 19q and codes for a protein which plays a central role in lipid metabolism. This protein is found in senile plaques and neurofibrillary tangles in AD brains [10]. The APOE gene has been implicated not only in 40–50% of the patients with
* Corresponding author. Tel.: +1 904 9530152; fax: +1 904 9537370; e-mail: [email protected]
late-onset disease [14], but also in some early-onset AD populations [2,16]. Several other genes have been studied as possible risk factors for typical late-onset disease including a-1-antichymotrypsin, the very low density lipoprotein receptor, and presenilin-1. However, in all these cases, there have been both positive and negative reports, and no consensus has developed about their involvement in disease etiology [3,7,11,12,15,18]. Most recently, a genetic association between late-onset AD and a polymorphism close to the low density lipoprotein receptor-related protein (LRP) gene has been reported [9]. The LRP gene is an attractive candidate for involvement in late-onset disease because it is directly involved with APOE metabolism [1] and it is also involved in the neuronal metabolism of the amyloid-b precursor protein [8]. Furthermore,
0304-3940/97/$17.00 1997 Elsevier Science Ireland Ltd. All rights reserved PII S0304-3940 (97 )0 0304-2
69
F. Wavrant-DeVrie`ze et al. / Neuroscience Letters 227 (1997) 68–70
LRP immunostaining has been demonstrated in the neuritic plaques of AD brains [13]. No mutations have yet been reported in this large and complex gene, although a polymorphism has been reported upstream of the gene. This polymorphism involves a tetranucleotide repeat (TTTC)n which can generate four different alleles [5,19]; the 91 and 87 bp alleles which are found most frequently, and the rarer 95 and 83 bp alleles. Lendon et al. [9] have reported a significant increase in the frequency of the 87 bp allele in late-onset AD studying an American population. We have sought to confirm this potential association in a French population with late-onset disease compared to relevant controls. The study was performed in a population consisting of 144 sporadic AD patients (100 women, 44 men) and 153 controls (106 women, 47 men) with a similar mean age (AD cases, 77.3 ± 8.9 years; controls, 79.3 ± 9.3 years). None of the subjects had a family history of AD consistent with Mendelian inheritance. All cases fulfilled NINCDS/ ADRDA criteria for probable AD. The mean age of onset for the AD patients was 73.4 ± 8.7 years. Genomic DNA was extracted from white blood cells, and the relevant part of the LRP gene, containing the TTTC repeat, was amplified by polymerase chain reaction (PCR) in order to genotype the subjects. PCR amplifications were carried out using primers previously described [19]. The amplified product was analyzed by electrophoresis on 20% acrylamide-bisacrylamide (19:1) gels stained with ethidium bromide. The APOE genotype was performed as described by Hixson and Vernier [6]. The genotype distributions were in Hardy–Weinberg equilibrium as assessed by a Chi-square test, and were similar in men and women. The Pearson Chi-square (x2) test of association was used to compare allele and genotype distributions. Where appropriate, the SPSS statistical package (version 7.5) was used to analyze the data with a multivariate logistic regression model giving odds ratios (OR) and 95% confidence intervals (95% CI). As expected the allelic frequencies of the APOE gene showed a strong association between AD cases and the e4 Table 1 Distribution of APOE genotypes and alleles APOE
Number of subjects Genotype (%)*
Allele frequencies (%)*
*P , 0.0001.
E2/E3 E2/E4 E3/E3 E3/E4 E4/E4 E2 E3 E4
AD cases
Controls
144 4.2 4.2 41.0 39.6 11.0 4.2 62.8 33.0
153 10.5 3.3 66.0 19.6 0.6 6.9 81.0 12.1
Table 2 Distribution of LRP genotypes and alleles LRP
Number of subjects Genotype (%)*
Allele frequencies (%)**
83/87 83/91 87/87 87/91 91/91 83 87 91
AD cases
Controls
144 0.7 5.5 13.9 39.6 40.3 3.1 34.0 62.9
153 2.0 0.7 17.6 47.7 32.0 1.3 42.5 56.2
*P = 0.041; **P = 0.046.
allele (P , 0.0001) (Table 1). The odds ratio to develop the disease in the presence of at least one e4 allele was 3.950 (95% CI = [2.402–6.495]). We did not see a statistically significant protective effect of the e2 allele in this population (OR = 0.571; 95% CI = [0.270–1.209]; P = 0.139). The genotype distributions of the LRP polymorphism and the allele frequencies are shown in Table 2. The 83 bp allele was rarely observed (0.013), and the 95 bp allele was absent in this population. In the control group, the LRP allele frequencies were similar to those previously described [5,19]. The genotype distributions showed a significant difference between AD cases and controls (P = 0.041). This difference in genotype distributions was reflected in the allele frequencies. Comparison of the distribution of the LRP alleles indicated a depletion of the 87 bp allele and an enrichment of the 91 bp allele in AD cases compared to controls (P = 0.046). This effect is exactly the opposite of the results reported by Lendon et al. [9]. The genotypes containing the 83 bp allele, which is rare, were combined either with the genotypes containing the 91 bp allele, or the 87 bp allele. The results showed a non-significant increase in the genotypes with the 91 bp allele in the AD cases; but a significant decrease of the genotypes with at least one 87 bp allele in AD cases (OR = 0.574; 95% CI = [0.358–0.919]; P = 0.020). This association persisted when the analysis was performed with a multivariate logistic regression on the whole population adjusted on gender and age (OR = 0.567; 95% CI = [0.350–0.919]; P = 0.021), or when adjusted on gender, age and APOE (OR = 0.537; 95% CI = [0.321–0.897]; P = 0.018). The decrease in the 87 bp allele in the AD group results in a concomitant increase in the frequencies of both the 83 and the 91 bp alleles. Our results, however, are in conflict with those of Lendon et al. We show an association between the LRP gene and disease; however, this association is opposite to that reported previously. In this report, the association is with a decrease in the frequency of the 87 bp allele; in the previous report it was an increase of this allele. There are two possibilities to explain these observations. The first is that both reports are type I statistical errors (false positives),
70
F. Wavrant-DeVrie`ze et al. / Neuroscience Letters 227 (1997) 68–70
as the weak significance of the associations would suggest (Lendon’s P = 0.041; this study P = 0.046); the second is that alleles at this locus are in linkage disequilibrium with biologically relevant variability elsewhere in the gene, and that different alleles are in disequilibrium in different populations. This latter suggestion could be due to the different genetic backgrounds between the populations used for these studies (American in Lendon et al.; French in this report). This possibility is further supported by the fact that both reports have borderline P values for the analysis of the LRP genotypes. However, they did not find any effect of the 87 bp allele after correction by logistic regression (P = 0.27) whereas we still had a significant result after this correction (P = 0.021). Sequence analysis of the LRP gene is a formidable undertaking as it has 89 coding exons [17], but will probably be needed to resolve this issue. A possible clue as to which part of the gene may be involved could be given by exploring the interaction between the LRP and APOE genes. We have found that the frequency of the 87 bp allele of the LRP gene is decreased in AD cases who are e4 carriers; however, the lack of significance may be due to the small size of the population and the low frequency of the e4 allele in the control group. This hypothesis needs to be investigated in a larger population before targeting the APOE binding domains for sequence analysis. Finally, it is of interest that there have been two positive linkage reports suggesting a locus on chromosome 12 may be involved in late-onset disease (Tanzi, personal communication; Pericak-Vance, personal communication). It remains possible that these positive linkage reports relate to the LRP locus. This work was supported by an NIH project grant: LRP and Alzheimer’s disease (J.H.), the Mayo Foundation, and also supported by INSERM and the Conseil Re´gional du Nord-Pas de Calais ‘axe re´gional de recherche sur les maladies neurode´ge´ne´ratives et le vieillissement ce´re´bral’ (P.A., M.C.C.H., F.P.). [1] Beisiegel, U., Weber, W., Ihrke, G., Herz, J. and Stanley, K.K., The LDL-receptor-related protein, LRP, is an apolipoprotein E-binding protein, Nature, 341 (1989) 162–164. [2] Chartier-Harlin, M.-C., Parfitt, M., Legrain, S., Pe´rez-Tur, J., Brousseau, T., Evans, E., Berr, C., Vidal, O., Roques, P., Gourlet, V., Fruchart, J.-C., Delacourte, A., Rossor, M. and Amouyel, P., Apolipoprotein E, e4 allele as a major risk factor for sporadic early and late-onset forms of Alzheimer’s disease: analysis of the 19q13.2 chromosomal region, Hum. Mol. Genet., 3 (1994) 569–574. [3] Chung, H., Roberts, C.T., Greenberg, S., Rebeck, G.W., Christie, R., Wallace, R., Jacob, H.J. and Hyman, B.T., Lack of association of trinucleotide repeat polymorphisms in the very-low-density lipoprotein receptor gene with Alzheimer’s disease, Ann. Neurol., 6 (1996) 800–803. [4] Hardy, J., Molecular genetics of Alzheimer’s disease (review), Acta Neurol. Scand. (Suppl.), 165 (1996) 13–17.
[5] Harris, M., Sanghera, D.K. and Kamboh, M.I., Two new alleles in the tetranucleotide repeat polymorphism in the LDL-receptor-related protein (LRP) gene, Clin. Genet., 50 (1996) 54–55. [6] Hixson, J.E. and Vernier, D.T., Restriction isotyping of human apolipoprotein E by gene amplification and cleavage with HhaI, J. Lipid Res., 31 (1990) 545–548. [7] Kamboh, M.I., Sanghera, D.K., Ferrell, R.E. and DeKosky, S.T., APOE*4-associated Alzheimer’s disease risk is modified by a1antichymotrypsin polymorphism, Nature Genet., 10 (1995) 486– 488. [8] Kounnas, M., Moir, R., Rebeck, W., Bush, A., Argraves, S., Tanzi, R., Hyman, B. and Strickland, D., LDL receptor-related protein, a multifunctional ApoE receptor, binds secreted b-amyloid precursor protein and mediates its degradation, Cell, 82 (1995) 331–340. [9] Lendon, C., Talbot, C., Craddock, N., woo Han, S., Wragg, M., Morris, J. and Goate, A., Genetic association studies between dementia of the Alzheimer’s type and three receptors for apolipoprotein E in a Caucasian population, Neurosci. Lett., 222 (1997) 187–190. [10] Namba, Y., Tomonaga, M., Kawasaki, H., Otomo, E. and Ikeda, K., Apolipoprotein E immunoreactivity in cerebral amyloid deposits and neurofibrillary tangles in Alzheimer’s disease and kuru plaque amyloid in Creutzfeldt–Jakob disease, Brain Res., 541 (1991) 163– 166. [11] Okuizumi, K., Onodera, O., Namba, Y., Ikeda, K., Yamamoto, T., Seki, K., Ueki, A., Nanko, S., Tanaka, H., Takahashi, H., Oyanagi, K., Mizusawa, H., Kanazawa, I. and Tsuji, S., Genetic association of the very low density lipoprotein (VLDL) receptor gene with sporadic Alzheimer’s disease, Nature Genet., 11 (1995) 207–209. [12] Pe´rez-Tur, J., Wavrant-De Vrie`ze, F. and Lambert, J.-C., ChartierHarlin, M.-C. and the Alzheimer’s Study Group, Presenilin-1 polymorphism and Alzheimer’s disease, Lancet, 347 (1996) 1560–1561. [13] Rebeck, G.W., Harr, S.D., Strickland, D.K. and Hyman, B.T., Multiple, diverse senile plaque-associated proteins are ligands of an apolipoprotein receptor, the alpha 2-macroglobulin receptor/low densitylipoprotein receptor-related protein, Ann. Neurol., 37 (1995) 211– 217. [14] Strittmatter, W.J., Saunders, A.M., Schmechel, D., Pericak-Vance, M., Enghild, J., Salvesen, G.S. and Roses, A.D., Apolipoprotein E: high-avidity binding to beta-amyloid and increased frequency of type 4 allele in late-onset familial Alzheimer’s disease, Proc. Natl. Acad. Sci. USA, 90 (1993) 1977–1981. [15] Talbot, C., Houlden, H., Craddock, N., Crook, R., Hutton, M., Lendon, C., Prihar, G., Morris, J.C., Hardy, J. and Goate, A., Polymorphism in AACT gene may lower age of onset of Alzheimer’s disease, NeuroReport, 7 (1996) 534–536. [16] van Duijn, C.M., de Knijff, P., Cruts, M., Wehnert, A., Havekes, L.M., Hofman, A. and Van Broeckhoven, C., Apolipoprotein E4 allele in a population-based study of early-onset Alzheimer’s disease, Nature Genet., 7 (1994) 74–78. [17] Van Leuven, F., Stas, L., Hilliker, C., Lorent, K., Umans, L., Serneels, L., Overbergh, L., Torrekens, S., Moechars, D., De Strooper, B. and Van den Berghe, H., Structure of the gene (LRP1) coding for the human a2-macroglobulin receptor lipoprotein receptor-related protein, Genomics, 24 (1994) 78–89. [18] Wragg, M. and Hutton, M., Talbot, C. and the Alzheimer’s Disease Collaborative Group, Genetic association between intronic polymorphism in presenilin-1 gene and late-onset Alzheimer’s disease, Lancet, 347 (1996) 509–512. [19] Zuliani, G. and Hobbs, H.H., Tetranucleotide length polymorphism 5′ of the a2-macroglobulin receptor (A2MR)/LDL receptor-related protein (LRP) gene, Hum. Mol. Genet., 3 (1994) 215.
Association between the low density lipoprotein receptor-related protein (LRP) and Alzheimer’s disease Fabienne Wavrant-DeVrie`ze a , b ,*, Jordi Pe´rez-Tur a, Jean-Charles Lambert b, Bernard Frigard c, Florence Pasquier d, Andre´ Delacourte e, Philippe Amouyel b, John Hardy a, Marie-Christine Chartier-Harlin b a
Mayo Clinic Jacksonville, 4500 San Pablo Road, Jacksonville, FL 32224, USA Service de Sante´ Publique et d’Epidemiologie, CJF INSERM 95-05, Institut Pasteur de Lille, 1 rue Prof. Calmette, 59019 Lille cedex, France c Centre de ge´riatrie de Wasquehal Molinel, rue Salvador Allende, B.P. 165, 59444 Wasquehal, France d CHR et U de Lille, Clinique Neurologique, Centre de la me´moire, Hoˆpital Salengro, 59037 Lille cedex, France e Unite´ Inserm 422, 1 Place de Verdun, 59045 Lille cedex, France
b
Received 2 April 1997; revised version received 18 April 1997; accepted 18 April 1997
Abstract Alzheimer’s disease (AD) is the most common neurodegenerative disorder affecting elderly people. It usually occurs after 65 years old (late-onset AD). The e4 allele of apolipoprotein E (APOE) gene is a risk factor which contributes about 50% of the genetic risk for this form of the disease. The low density lipoprotein receptor-related protein (LRP) is a major receptor for APOE which is found in the senile plaques of AD brains. This makes it a good candidate gene for the disease. There is a polymorphism in the region upstream of the LRP gene that has been associated with AD in an American population. We examined this polymorphism by restriction fragment length polymorphism analysis in a French population with sporadic late-onset AD. In the previous report, a significant increase of the 87 bp allele was found in the AD cases; however, in our population, we observed a significant decrease with this same allele of the LRP gene. The possible reasons for this discrepancy, linkage disequilibrium or statistical anomaly, are discussed. 1997 Elsevier Science Ireland Ltd. Keywords: Late-onset Alzheimer’s disease; Low density lipoprotein receptor-related protein; Apolipoprotein E receptor; Association studies; Risk factor
Alzheimer’s disease (AD) is the most common neurodegenerative disorder affecting elderly people. Several genes have been reported to influence the occurrence of the disease [4], but the only established risk factor for the typical late-onset form of the disease is genetic variability at the apolipoprotein E (APOE) gene. Genetic variability at this locus has been shown to have a strong association with the disease [14]. The APOE gene is located on the chromosome 19q and codes for a protein which plays a central role in lipid metabolism. This protein is found in senile plaques and neurofibrillary tangles in AD brains [10]. The APOE gene has been implicated not only in 40–50% of the patients with
* Corresponding author. Tel.: +1 904 9530152; fax: +1 904 9537370; e-mail: [email protected]
late-onset disease [14], but also in some early-onset AD populations [2,16]. Several other genes have been studied as possible risk factors for typical late-onset disease including a-1-antichymotrypsin, the very low density lipoprotein receptor, and presenilin-1. However, in all these cases, there have been both positive and negative reports, and no consensus has developed about their involvement in disease etiology [3,7,11,12,15,18]. Most recently, a genetic association between late-onset AD and a polymorphism close to the low density lipoprotein receptor-related protein (LRP) gene has been reported [9]. The LRP gene is an attractive candidate for involvement in late-onset disease because it is directly involved with APOE metabolism [1] and it is also involved in the neuronal metabolism of the amyloid-b precursor protein [8]. Furthermore,
0304-3940/97/$17.00 1997 Elsevier Science Ireland Ltd. All rights reserved PII S0304-3940 (97 )0 0304-2
69
F. Wavrant-DeVrie`ze et al. / Neuroscience Letters 227 (1997) 68–70
LRP immunostaining has been demonstrated in the neuritic plaques of AD brains [13]. No mutations have yet been reported in this large and complex gene, although a polymorphism has been reported upstream of the gene. This polymorphism involves a tetranucleotide repeat (TTTC)n which can generate four different alleles [5,19]; the 91 and 87 bp alleles which are found most frequently, and the rarer 95 and 83 bp alleles. Lendon et al. [9] have reported a significant increase in the frequency of the 87 bp allele in late-onset AD studying an American population. We have sought to confirm this potential association in a French population with late-onset disease compared to relevant controls. The study was performed in a population consisting of 144 sporadic AD patients (100 women, 44 men) and 153 controls (106 women, 47 men) with a similar mean age (AD cases, 77.3 ± 8.9 years; controls, 79.3 ± 9.3 years). None of the subjects had a family history of AD consistent with Mendelian inheritance. All cases fulfilled NINCDS/ ADRDA criteria for probable AD. The mean age of onset for the AD patients was 73.4 ± 8.7 years. Genomic DNA was extracted from white blood cells, and the relevant part of the LRP gene, containing the TTTC repeat, was amplified by polymerase chain reaction (PCR) in order to genotype the subjects. PCR amplifications were carried out using primers previously described [19]. The amplified product was analyzed by electrophoresis on 20% acrylamide-bisacrylamide (19:1) gels stained with ethidium bromide. The APOE genotype was performed as described by Hixson and Vernier [6]. The genotype distributions were in Hardy–Weinberg equilibrium as assessed by a Chi-square test, and were similar in men and women. The Pearson Chi-square (x2) test of association was used to compare allele and genotype distributions. Where appropriate, the SPSS statistical package (version 7.5) was used to analyze the data with a multivariate logistic regression model giving odds ratios (OR) and 95% confidence intervals (95% CI). As expected the allelic frequencies of the APOE gene showed a strong association between AD cases and the e4 Table 1 Distribution of APOE genotypes and alleles APOE
Number of subjects Genotype (%)*
Allele frequencies (%)*
*P , 0.0001.
E2/E3 E2/E4 E3/E3 E3/E4 E4/E4 E2 E3 E4
AD cases
Controls
144 4.2 4.2 41.0 39.6 11.0 4.2 62.8 33.0
153 10.5 3.3 66.0 19.6 0.6 6.9 81.0 12.1
Table 2 Distribution of LRP genotypes and alleles LRP
Number of subjects Genotype (%)*
Allele frequencies (%)**
83/87 83/91 87/87 87/91 91/91 83 87 91
AD cases
Controls
144 0.7 5.5 13.9 39.6 40.3 3.1 34.0 62.9
153 2.0 0.7 17.6 47.7 32.0 1.3 42.5 56.2
*P = 0.041; **P = 0.046.
allele (P , 0.0001) (Table 1). The odds ratio to develop the disease in the presence of at least one e4 allele was 3.950 (95% CI = [2.402–6.495]). We did not see a statistically significant protective effect of the e2 allele in this population (OR = 0.571; 95% CI = [0.270–1.209]; P = 0.139). The genotype distributions of the LRP polymorphism and the allele frequencies are shown in Table 2. The 83 bp allele was rarely observed (0.013), and the 95 bp allele was absent in this population. In the control group, the LRP allele frequencies were similar to those previously described [5,19]. The genotype distributions showed a significant difference between AD cases and controls (P = 0.041). This difference in genotype distributions was reflected in the allele frequencies. Comparison of the distribution of the LRP alleles indicated a depletion of the 87 bp allele and an enrichment of the 91 bp allele in AD cases compared to controls (P = 0.046). This effect is exactly the opposite of the results reported by Lendon et al. [9]. The genotypes containing the 83 bp allele, which is rare, were combined either with the genotypes containing the 91 bp allele, or the 87 bp allele. The results showed a non-significant increase in the genotypes with the 91 bp allele in the AD cases; but a significant decrease of the genotypes with at least one 87 bp allele in AD cases (OR = 0.574; 95% CI = [0.358–0.919]; P = 0.020). This association persisted when the analysis was performed with a multivariate logistic regression on the whole population adjusted on gender and age (OR = 0.567; 95% CI = [0.350–0.919]; P = 0.021), or when adjusted on gender, age and APOE (OR = 0.537; 95% CI = [0.321–0.897]; P = 0.018). The decrease in the 87 bp allele in the AD group results in a concomitant increase in the frequencies of both the 83 and the 91 bp alleles. Our results, however, are in conflict with those of Lendon et al. We show an association between the LRP gene and disease; however, this association is opposite to that reported previously. In this report, the association is with a decrease in the frequency of the 87 bp allele; in the previous report it was an increase of this allele. There are two possibilities to explain these observations. The first is that both reports are type I statistical errors (false positives),
70
F. Wavrant-DeVrie`ze et al. / Neuroscience Letters 227 (1997) 68–70
as the weak significance of the associations would suggest (Lendon’s P = 0.041; this study P = 0.046); the second is that alleles at this locus are in linkage disequilibrium with biologically relevant variability elsewhere in the gene, and that different alleles are in disequilibrium in different populations. This latter suggestion could be due to the different genetic backgrounds between the populations used for these studies (American in Lendon et al.; French in this report). This possibility is further supported by the fact that both reports have borderline P values for the analysis of the LRP genotypes. However, they did not find any effect of the 87 bp allele after correction by logistic regression (P = 0.27) whereas we still had a significant result after this correction (P = 0.021). Sequence analysis of the LRP gene is a formidable undertaking as it has 89 coding exons [17], but will probably be needed to resolve this issue. A possible clue as to which part of the gene may be involved could be given by exploring the interaction between the LRP and APOE genes. We have found that the frequency of the 87 bp allele of the LRP gene is decreased in AD cases who are e4 carriers; however, the lack of significance may be due to the small size of the population and the low frequency of the e4 allele in the control group. This hypothesis needs to be investigated in a larger population before targeting the APOE binding domains for sequence analysis. Finally, it is of interest that there have been two positive linkage reports suggesting a locus on chromosome 12 may be involved in late-onset disease (Tanzi, personal communication; Pericak-Vance, personal communication). It remains possible that these positive linkage reports relate to the LRP locus. This work was supported by an NIH project grant: LRP and Alzheimer’s disease (J.H.), the Mayo Foundation, and also supported by INSERM and the Conseil Re´gional du Nord-Pas de Calais ‘axe re´gional de recherche sur les maladies neurode´ge´ne´ratives et le vieillissement ce´re´bral’ (P.A., M.C.C.H., F.P.). [1] Beisiegel, U., Weber, W., Ihrke, G., Herz, J. and Stanley, K.K., The LDL-receptor-related protein, LRP, is an apolipoprotein E-binding protein, Nature, 341 (1989) 162–164. [2] Chartier-Harlin, M.-C., Parfitt, M., Legrain, S., Pe´rez-Tur, J., Brousseau, T., Evans, E., Berr, C., Vidal, O., Roques, P., Gourlet, V., Fruchart, J.-C., Delacourte, A., Rossor, M. and Amouyel, P., Apolipoprotein E, e4 allele as a major risk factor for sporadic early and late-onset forms of Alzheimer’s disease: analysis of the 19q13.2 chromosomal region, Hum. Mol. Genet., 3 (1994) 569–574. [3] Chung, H., Roberts, C.T., Greenberg, S., Rebeck, G.W., Christie, R., Wallace, R., Jacob, H.J. and Hyman, B.T., Lack of association of trinucleotide repeat polymorphisms in the very-low-density lipoprotein receptor gene with Alzheimer’s disease, Ann. Neurol., 6 (1996) 800–803. [4] Hardy, J., Molecular genetics of Alzheimer’s disease (review), Acta Neurol. Scand. (Suppl.), 165 (1996) 13–17.
[5] Harris, M., Sanghera, D.K. and Kamboh, M.I., Two new alleles in the tetranucleotide repeat polymorphism in the LDL-receptor-related protein (LRP) gene, Clin. Genet., 50 (1996) 54–55. [6] Hixson, J.E. and Vernier, D.T., Restriction isotyping of human apolipoprotein E by gene amplification and cleavage with HhaI, J. Lipid Res., 31 (1990) 545–548. [7] Kamboh, M.I., Sanghera, D.K., Ferrell, R.E. and DeKosky, S.T., APOE*4-associated Alzheimer’s disease risk is modified by a1antichymotrypsin polymorphism, Nature Genet., 10 (1995) 486– 488. [8] Kounnas, M., Moir, R., Rebeck, W., Bush, A., Argraves, S., Tanzi, R., Hyman, B. and Strickland, D., LDL receptor-related protein, a multifunctional ApoE receptor, binds secreted b-amyloid precursor protein and mediates its degradation, Cell, 82 (1995) 331–340. [9] Lendon, C., Talbot, C., Craddock, N., woo Han, S., Wragg, M., Morris, J. and Goate, A., Genetic association studies between dementia of the Alzheimer’s type and three receptors for apolipoprotein E in a Caucasian population, Neurosci. Lett., 222 (1997) 187–190. [10] Namba, Y., Tomonaga, M., Kawasaki, H., Otomo, E. and Ikeda, K., Apolipoprotein E immunoreactivity in cerebral amyloid deposits and neurofibrillary tangles in Alzheimer’s disease and kuru plaque amyloid in Creutzfeldt–Jakob disease, Brain Res., 541 (1991) 163– 166. [11] Okuizumi, K., Onodera, O., Namba, Y., Ikeda, K., Yamamoto, T., Seki, K., Ueki, A., Nanko, S., Tanaka, H., Takahashi, H., Oyanagi, K., Mizusawa, H., Kanazawa, I. and Tsuji, S., Genetic association of the very low density lipoprotein (VLDL) receptor gene with sporadic Alzheimer’s disease, Nature Genet., 11 (1995) 207–209. [12] Pe´rez-Tur, J., Wavrant-De Vrie`ze, F. and Lambert, J.-C., ChartierHarlin, M.-C. and the Alzheimer’s Study Group, Presenilin-1 polymorphism and Alzheimer’s disease, Lancet, 347 (1996) 1560–1561. [13] Rebeck, G.W., Harr, S.D., Strickland, D.K. and Hyman, B.T., Multiple, diverse senile plaque-associated proteins are ligands of an apolipoprotein receptor, the alpha 2-macroglobulin receptor/low densitylipoprotein receptor-related protein, Ann. Neurol., 37 (1995) 211– 217. [14] Strittmatter, W.J., Saunders, A.M., Schmechel, D., Pericak-Vance, M., Enghild, J., Salvesen, G.S. and Roses, A.D., Apolipoprotein E: high-avidity binding to beta-amyloid and increased frequency of type 4 allele in late-onset familial Alzheimer’s disease, Proc. Natl. Acad. Sci. USA, 90 (1993) 1977–1981. [15] Talbot, C., Houlden, H., Craddock, N., Crook, R., Hutton, M., Lendon, C., Prihar, G., Morris, J.C., Hardy, J. and Goate, A., Polymorphism in AACT gene may lower age of onset of Alzheimer’s disease, NeuroReport, 7 (1996) 534–536. [16] van Duijn, C.M., de Knijff, P., Cruts, M., Wehnert, A., Havekes, L.M., Hofman, A. and Van Broeckhoven, C., Apolipoprotein E4 allele in a population-based study of early-onset Alzheimer’s disease, Nature Genet., 7 (1994) 74–78. [17] Van Leuven, F., Stas, L., Hilliker, C., Lorent, K., Umans, L., Serneels, L., Overbergh, L., Torrekens, S., Moechars, D., De Strooper, B. and Van den Berghe, H., Structure of the gene (LRP1) coding for the human a2-macroglobulin receptor lipoprotein receptor-related protein, Genomics, 24 (1994) 78–89. [18] Wragg, M. and Hutton, M., Talbot, C. and the Alzheimer’s Disease Collaborative Group, Genetic association between intronic polymorphism in presenilin-1 gene and late-onset Alzheimer’s disease, Lancet, 347 (1996) 509–512. [19] Zuliani, G. and Hobbs, H.H., Tetranucleotide length polymorphism 5′ of the a2-macroglobulin receptor (A2MR)/LDL receptor-related protein (LRP) gene, Hum. Mol. Genet., 3 (1994) 215.