1995 Fiscal Year Annual Research Report

リポフェクション法を用いたLDL受容体遺伝子導入による難治性高脂血症治療法の開発

Research Project

Project/Area Number	06557059
Research Institution	Osaka University
Principal Investigator	松沢佑次大阪大学, 医学部, 教授 (70116101)
Co-Investigator(Kenkyū-buntansha)	金田安史大阪大学, 細胞生体工学センター, 助教授 (10177537) 野崎秀一大阪大学, 医学部, 助手 (30252646) 山下静也大阪大学, 医学部, 助手 (60243242) 竹村芳大阪大学, 医学部, 講師 (00161240) 河田純男大阪大学, 医学部, 助教授 (90183285)
Keywords	家族性高コレステロール血症 / HVJ-liposome法 / 遺伝子治療
Research Abstract	LDL受容体遺伝子異常を原因とする家族性高コレステロール血症(FH)に対する遺伝子治療を開発するため、昨年度に引き続き、効率の良いin vivo gene introduction法であるHVJ-liposome法を用いたLDL受容体遺伝子導入実験を行った。昨年度の結果から、当法によるLDL受容体遺伝子導入で、ある程度のコレステロール低下が見込めることが明らかになったが、その効果はなお不十分のため、今年度はvectorの改良により導入発現効率を上げることを目標とした。昨年度使用したvector(pMy3)では、liposomeに包み込むのに大きすぎることやtransferrin promoterが肝においてregulationを受ける可能性があったため、promotorをchicken β actin promotorとし、hLDL receptor cDNAを組み込んで、7.5kbに比較し、Northern blotおよび免疫組織学的検索により明らかに遺伝子導入効率が高いことが確認された。また、pCL1を用いた遺伝子導入によるLDLの取り込みの増加が標識LDLを用いて確認された。以上より、HVJ-liposome法を用いたLDL受容体遺伝子導入による難治性高脂血症治療法の開発は、その基礎段階が完了したと考えられる。一方、さらにWHHLウサギおよびノックアウトマウスなどモデル動物での遺伝子導入発現実験を行ったが、HVJ-liposome法による導入遺伝子発現期間が短く、発現量も不十分であり、さらに改良が必要と考えられた。今後、長期発現に向けてEBvirus vectorの開発やneonateへの投与、あるいはretrovirus法との組み合わせなどの工夫を行っていく予定である。

Research Products
(15 results)

All Other

All Publications (15 results)

[Publications] N.Sakai: "Decreased affinity of low density lipoprotein(LDL) particles for LDL receptors in patients with cholesteryl ester transfer protein deficiency" Eur.J.Clin.Invest.25. 332-339 (1995)
[Publications] T.Nakagawa: "Marked hypocholesterolemia in a case with adrenal adenoma: Enhanced catabolism of low density lipoprotein(LDL) via the LDL receptors of tumor cells." J Clin Endocrinol Metab. 80(1). 92-96 (1995)
[Publications] T.Funahashi: "A compound heterozygote for familial hypercholesterolemia with a homozygous mother." J.Intern.Med.239. 187-190 (1996)
[Publications] S.Nozaki: "The effects of pravastatin on plasma and urinary mevalonate concentrations in subjects with familial hypercholesterolemia:a comparison of morning and evening administration." Eur.J.Clin Pharmacol.(in press).
[Publications] T.Maruyama: "Common mutations in the low density lipoprotein leceptor gene causing familial hypercholesterolemia in the Japanese popuration." Arterioscl.Thoromb.Vasc.Biol.15. 1713-1718 (1995)
[Publications] T.Maruyama: "A single point mutation in the splice donor site of the low density lipoprotein receptor gene produces an intron read-through,an exon-skipped and a cryptic site utilized transcripts." Eur.J.Biochem.232. 700-705 (1995)
[Publications] N.Tomita: "A novel gene-transfer technique mediated by HVJ(Sendai virus),nuclear protein,and liposomes." Cancer-Detect-Prev.18(6). 485-491 (1994)
[Publications] R.Morishita: "A gene therapy strategy using a transcription factor decoy of the E2F bindinng site inhibits smooth muscle proliferation in vivo." Proc-Natl-Acad-Sci-U-S-A. 92(13). 5855-5859 (1995)
[Publications] R.Morishita: "Evidence for direct local effect of angiotensin in vascular hypertrophy.In vivo gene transfer of angiotensin converting enzyme." J-Clin-Invest.94(3). 978-984 (1994)
[Publications] R.Morishita: "Autocrine and paracrine effects of atrial natriuretic peptide gene transfer on vascular smooth muscle and endothelial cellular growth." J-Clin-Invest.94(2). 824-829 (1994)
[Publications] R.Morishita: "Intimal hyperplasia after vascular injury is inhibited by antisense cdk 2 kinase oligonucleotides." J-Clin-Invest.93(4). 1458-1464 (1994)
[Publications] N.Tomita: "Transient decrease in high blood pressure by in vivo transfer of antisense oligodeoxynucleotides against rat angiotensinogen." Hypertension.26(1). 131-136 (1995)
[Publications] M.J.Mann: "Genetic engineering of vein grafts resistant to atherosclerosis" Proc-Natl-Acad-Sci-U-S-A.92(10). 4502-4506 (1995)
[Publications] von-der-Leyen-HE: "Gene therapy inhibiting neointimal vascular lesion:in vivo transfer of endothelial sell nitric oxide synthase gene." Proc-Natl-Acad-Sci-U-S-A.92(4). 1137-1141 (1995)
[Publications] M.Arai: "In vivo transfection of genes for renin and angiotensinogen into the glomerular sells induced phenotypic change of the mesangial cells and glomerular sclerosis." Biochem-Biophys-Res-Commun.206(2). 525-532 (1995)

1995 Fiscal Year Annual Research Report

リポフェクション法を用いたLDL受容体遺伝子導入による難治性高脂血症治療法の開発

Principal Investigator

松沢 佑次 大阪大学, 医学部, 教授 (70116101)

Research Products

[Publications] N.Sakai: "Decreased affinity of low density lipoprotein(LDL) particles for LDL receptors in patients with cholesteryl ester transfer protein deficiency" Eur.J.Clin.Invest.25. 332-339 (1995)

[Publications] T.Nakagawa: "Marked hypocholesterolemia in a case with adrenal adenoma: Enhanced catabolism of low density lipoprotein(LDL) via the LDL receptors of tumor cells." J Clin Endocrinol Metab. 80(1). 92-96 (1995)

[Publications] T.Funahashi: "A compound heterozygote for familial hypercholesterolemia with a homozygous mother." J.Intern.Med.239. 187-190 (1996)

[Publications] S.Nozaki: "The effects of pravastatin on plasma and urinary mevalonate concentrations in subjects with familial hypercholesterolemia:a comparison of morning and evening administration." Eur.J.Clin Pharmacol.(in press).

[Publications] T.Maruyama: "Common mutations in the low density lipoprotein leceptor gene causing familial hypercholesterolemia in the Japanese popuration." Arterioscl.Thoromb.Vasc.Biol.15. 1713-1718 (1995)

[Publications] T.Maruyama: "A single point mutation in the splice donor site of the low density lipoprotein receptor gene produces an intron read-through,an exon-skipped and a cryptic site utilized transcripts." Eur.J.Biochem.232. 700-705 (1995)

[Publications] N.Tomita: "A novel gene-transfer technique mediated by HVJ(Sendai virus),nuclear protein,and liposomes." Cancer-Detect-Prev.18(6). 485-491 (1994)

[Publications] R.Morishita: "A gene therapy strategy using a transcription factor decoy of the E2F bindinng site inhibits smooth muscle proliferation in vivo." Proc-Natl-Acad-Sci-U-S-A. 92(13). 5855-5859 (1995)

[Publications] R.Morishita: "Evidence for direct local effect of angiotensin in vascular hypertrophy.In vivo gene transfer of angiotensin converting enzyme." J-Clin-Invest.94(3). 978-984 (1994)

[Publications] R.Morishita: "Autocrine and paracrine effects of atrial natriuretic peptide gene transfer on vascular smooth muscle and endothelial cellular growth." J-Clin-Invest.94(2). 824-829 (1994)

[Publications] R.Morishita: "Intimal hyperplasia after vascular injury is inhibited by antisense cdk 2 kinase oligonucleotides." J-Clin-Invest.93(4). 1458-1464 (1994)

[Publications] N.Tomita: "Transient decrease in high blood pressure by in vivo transfer of antisense oligodeoxynucleotides against rat angiotensinogen." Hypertension.26(1). 131-136 (1995)

[Publications] M.J.Mann: "Genetic engineering of vein grafts resistant to atherosclerosis" Proc-Natl-Acad-Sci-U-S-A.92(10). 4502-4506 (1995)

[Publications] von-der-Leyen-HE: "Gene therapy inhibiting neointimal vascular lesion:in vivo transfer of endothelial sell nitric oxide synthase gene." Proc-Natl-Acad-Sci-U-S-A.92(4). 1137-1141 (1995)

[Publications] M.Arai: "In vivo transfection of genes for renin and angiotensinogen into the glomerular sells induced phenotypic change of the mesangial cells and glomerular sclerosis." Biochem-Biophys-Res-Commun.206(2). 525-532 (1995)

松沢佑次大阪大学, 医学部, 教授 (70116101)