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1998 Fiscal Year Final Research Report Summary

Basic spproaches for gene therapy of hemoglobinopathy

Research Project

Project/Area Number 08457629
Research Category

Grant-in-Aid for Scientific Research (B)

Allocation TypeSingle-year Grants
Section一般
Research Field Human genetics
Research InstitutionKYUSHU UNIVERSITY

Principal Investigator

FUKUMAKI Yasuyuki  Kyushu University, Institute of Genetic Information, Professor, 遺伝情報実験施設, 教授 (90128083)

Co-Investigator(Kenkyū-buntansha) IWAKI Akiko  Kyushu University, Institute of Genetic Information, Assistant Professor, 遺伝情報実験施設, 助手 (30253454)
Project Period (FY) 1996 – 1998
Keywordsgene therapy / globin gene / thalassemia / AAV / insulator / HbE / antisense / splicing
Research Abstract

(1) Position-independent expression of transgenes in target cells is one of essential subjects for successful gene therapies. We prepared recombinant adeno-associated virus (rAAV) containing the combination of the DNaseI hypersensitive site 2 (HS2), 3 (HS3), and 4 (HS4) core elements from the human beta-globin locus control region (LOR), the human beta-globin gene, and the herpes virus tymidine kinase promoter driven neomycin-resistant gene (neo^R) (rHS432, rHS43, rHS42, rHS32, and rHS2), and also rAAV containing two copies of the 250-bp core fragment of the chicken beta-globin insulator on both sides of the rHS2 (rlns/HS2/2Ins). After isolating neomycin resistant mouse erythroleukemia (MEL) cells infected with each rAAV, we analyzed structure of proviral genome of rAAV and determined expression level of the human beta-globin gene. The high variability of the expression level was observed among clones infected with recombinant virus lacking the insulator. In contrast, in the clones inf … More ected with rIns/HS2/lns, the range of expression of the human beta-globin gene was from 52.8% to 58.3% of the mouse beta-major globin gene. Thus, these results indicate that the insulator dramatically functioned to reduce the variability oftransgene expression due to the position effect. This insulator-rAAV vectorsystem is a promising way to provide the constant level of the transgene expression for gene therapy regardless of insertion sites in chromatin.
(2) Hemoglobin (Hb) E is the most common Hb variant among Southeast Asian populations. The mutation in codon 26 (GAG to AAG) of the beta-globin gene (beta^E) induces an alternative splicing, resulting in production of normally and aberrantly spliced beta-globin mRNA.Compound heterozygotes for beta-thalassemia and beta^E have a severe beta-thalassemia- disease. Repression of aberrant splicing due to the BE mutation could ameliorate severity of such patients. We showed that the aberrant splicing was partially repressed in the cells treated with antisense oligoribonucleotide targeted to the aberrant 5 splice site. Maximum effect of the antisense oligoribonucleotide was observed at concentration of 0.4 muM and 36 hrs after the treatment in our experiment. We also analyzed the effect of transient and stable expression of SF2/ASF on aberrant splicing in cells expressing the beta^E-globin gene. Partial repression of the aberrant splicing was also observed in both expression systems. These results imply that antisense oligoribonucleotide treatment and SF2/ASF expression are possible therapeutic applications for beta-thalassemia/HbE disease. Less

  • Research Products

    (49 results)

All Other

All Publications (49 results)

  • [Publications] Manabe,J.et al.: "Two novel mutations in the reduced nicotinamide adenine dinucleotide(NADH)-cytochrome b5 reductase gene of a patinet with generalized type,hereditary methemoglobinemia." Blood. 88. 3208-3215 (1996)

    • Description
      「研究成果報告書概要(和文)」より
  • [Publications] Sumimoto,H.et al.: "Assembly and activation of the phagocyte NADPH oxidase." J.Biol.Chem.271. 22152-22158 (1996)

    • Description
      「研究成果報告書概要(和文)」より
  • [Publications] Ogawa,T.,et al.: "Accelerated evolution of snake venom phospholipase A2 isozymes for aquisition of diverse physiological functions." Toxicon. 34. 1229-1236 (1996)

    • Description
      「研究成果報告書概要(和文)」より
  • [Publications] Nobuhisa,I.et al.: "Accelerated evolution of Trimeresurus okinavensis venom gland phopholipase A2 isozyme-encoding genes." Gene. 172. 267-272 (1996)

    • Description
      「研究成果報告書概要(和文)」より
  • [Publications] Deshimaru,M.,et al.: "Accelerated evolution of crotalinae snake venom gland serine proteases." FEBS Letters. 397. 83-88 (1996)

    • Description
      「研究成果報告書概要(和文)」より
  • [Publications] Sriroongrueng,W.et al.: "Analysis of β-thalassemia mutations and β-locus control region hypersensitive sites 2,3 and 4 in Southern Thailand." Asian J.Trop.Med.Pub.Health. 28. 120-127 (1997)

    • Description
      「研究成果報告書概要(和文)」より
  • [Publications] Fucharoen,S.et al.: "Molecular and hematological characterization of Hb Tak and Hb Pyrgos in Thailand." Southweast Asian J.Trop.Med.Pub.Health. 28. 110-114 (1997)

    • Description
      「研究成果報告書概要(和文)」より
  • [Publications] Iwaki,A.et.al.: "Identification and characterization of the gene encoding a new member of the α-crystallin/small hsp family, closely linked to the αB-crystallin gene in a head-to-head" Genomics. 45. 386-394 (1997)

    • Description
      「研究成果報告書概要(和文)」より
  • [Publications] Nobuhisa,I.et al.: "Characterization and evolution of a gene encoding a Trimeresurus flavoviridis serm protein that inhibits basic phosphlipase A2 jsozymes in the snake's venom." Eur.J.Biochem.249. 838-845 (1997)

    • Description
      「研究成果報告書概要(和文)」より
  • [Publications] Kamakura,S.,et al.: "Cloning and characterization of tha 5'-flanking region of the human dopamine D4 receptor gene." Biohem.Biophys.Res.Commun.235. 321-326 (1997)

    • Description
      「研究成果報告書概要(和文)」より
  • [Publications] Ohba,Y.et al.: "β-Thalassemia mutations in Japanese and Koreans." Hemoglobin. 21. 191-200 (1997)

    • Description
      「研究成果報告書概要(和文)」より
  • [Publications] Nobuhisa,I.et al.: "Structures of gene encoding phospholipase A2 inhibitors from the serum of Trimeresurus flaviridis snake." Gene. 191. 31-37 (1997)

    • Description
      「研究成果報告書概要(和文)」より
  • [Publications] Furuya,H.et al.: "Adult onset globoid cell leukodystrophy(Krabbe disease): analysis of galactosylceramidase cDNA from four Japanese patients." Hum.Gnenet.100. 450-456 (1997)

    • Description
      「研究成果報告書概要(和文)」より
  • [Publications] Furuumi,H.et al.: "Molecular basis of β-thalassemia in the Maldives." Hemoglobin. 22. 141-151 (1998)

    • Description
      「研究成果報告書概要(和文)」より
  • [Publications] Tanphaichitr,V.et al.: "Type I and II hereditary methemoglobinemia due to NADH-cytochrome b5 reductase deficiency in Thai patients." Internal Medicine. 13. 81-85 (1998)

    • Description
      「研究成果報告書概要(和文)」より
  • [Publications] Nobiuhisa,I.et al.: "Structural elements of Trimeresurusflaviridis serum inhibitors for recognition of its venome phospholipase A2 isozymes." FEBS Letters. 429. 385-389 (1998)

    • Description
      「研究成果報告書概要(和文)」より
  • [Publications] Yamada,M.et al.: "Thrombopoietin supports in vitro erythroid differentiation via its specific receptor c-MpI in a human leukemia cell line." Cell Growth & Differentiation. 9. 487-496 (1998)

    • Description
      「研究成果報告書概要(和文)」より
  • [Publications] Nobuhisa,I.et al.: "Retrotramsposable CR1-like elements in crotalinae sanke genomes." Toxicon. 36. 915-920 (1998)

    • Description
      「研究成果報告書概要(和文)」より
  • [Publications] Mizuki,K.et al.: "Functional modules and expression of mouse p40^<phox> and p65^<phox>, SH3-domain-cotaining proteins involved in the phagocyte NADPH oxidase complex." Eur.J.Biochem.251. 573-582 (1998)

    • Description
      「研究成果報告書概要(和文)」より
  • [Publications] Mamiya,K.et al.: "Association of polymorphisms in the cytochrome P450(CYP)2C19 and 2C18 genes in Japanese epileptic patients." Pharmacogenetics. 8. 87-90 (1998)

    • Description
      「研究成果報告書概要(和文)」より
  • [Publications] Matsumoto,M.et al.: "High pressure induces G2 arrest in murine erythroleukemia cells." J.Biochem.123. 87-93 (1998)

    • Description
      「研究成果報告書概要(和文)」より
  • [Publications] Higasa,K.et al.: "Molecular basis of hereditary methemoglbinemia types,I and II." Brt.J.Haematol.103. 922-930 (1998)

    • Description
      「研究成果報告書概要(和文)」より
  • [Publications] Inoue,T.et al.: "Position-independent human β-globin gene expression mediated by a recombinant adeno-associated virus vector carrying the chicken β-globin insulator." J.Hum.Genet.印刷中. (1999)

    • Description
      「研究成果報告書概要(和文)」より
  • [Publications] Hamamura,M.et al.: "Differential dicreases in c-fos and aldolase C mRNA expression in the rat cerebellum after repeated methamphetamine administration." Mol,Brain Res.印刷中. (1999)

    • Description
      「研究成果報告書概要(和文)」より
  • [Publications] 服巻保幸: "グロビン遺伝子 「赤血球」" 藤井寿一編 医学書院, 114-127 (1998)

    • Description
      「研究成果報告書概要(和文)」より
  • [Publications] Manabe, J., Roopen, A., Sumimoto, H., Yubisui, T., Bellingham, A.J., Layton, D.M.and Fukumaki, Y: "Two novel mutations in the reduced nicotinamide adenine dinucleotide (NADH)-cytochromeb5 reductase gene of a patinet with generalized type, hereditary methemoglobinemia." Blood. 88. 3208-3215 (1996)

    • Description
      「研究成果報告書概要(欧文)」より
  • [Publications] Sumimoto, H., Hata, K., Mizuki, K., Ito, T., Kage, Y., Sakaki, Y., Fukumaki, Y., Nakamura, M.and Takeshige, K.: "Assembly and activation of the phagocyte NADPH oxidase." J.Biol.Chem.271. 22152-22158 (1996)

    • Description
      「研究成果報告書概要(欧文)」より
  • [Publications] Ogawa, T., Nakashima, K., Nobuhisa, I., Deshimaru, M., Shimohigashi, Y., Fukumaki, Y., Sakaki, Y., Hattori, S.and Ohno, M.: "Accelerated evolution of snake venom phospholipase A2 isozymes for aquisition of diverse physiological functions." Toxicon. 34. 1229-1236 (1996)

    • Description
      「研究成果報告書概要(欧文)」より
  • [Publications] Nobuhisa, I., Nakashima, K., Deshimaru, M., Ogawa, T., Shimohigashi, Y., Fukumaki, Y., Sakaki, Y., Hattori, S., Kihara, H.and Ohno, M.: "Accelerated evolution of Trimeresurus okinavensis venom gland phopholipase A2 isozyme-encoding genes." Gene. 172. 267-272 (1996)

    • Description
      「研究成果報告書概要(欧文)」より
  • [Publications] Deshimaru, M., Ogawa, T., Nakashima, K., Nobuhisa, I., Chijiwa, T., Shimohigashi, Y., Fukumaki, Y., Niwa, M., Yamashina, I., Hattori, S.and Ohno, M.: "Accelerated evolution of crotalinae snake venom gland serine proteases." FEBS Letters. 397. 83-88 (1996)

    • Description
      「研究成果報告書概要(欧文)」より
  • [Publications] Sriroongrueng, W., Schleiemacher, E., Panich, V., Nopparatana, C., Saechan, V., Laosombat, V., Pornpatkul, M.and Fukumaki, Y.: "Analysis of beta-thalassemia mutations and beta-locus control region hypersensitive sites 2,3 and 4 in Southern Thailand." Southeast Asian J.Trop.Med.Pub.Health. 28. 120-127 (1997)

    • Description
      「研究成果報告書概要(欧文)」より
  • [Publications] Fucharoen, S., Fucharoen, G., Sae-ung, N., Sanchaisuriya, K.and Fukumaki, Y.: "Molecular and hematological characterization of Hb Tak and Hb Pyrgos in Thailand." Southweast Asian J.Trop.Med.Pub.Health. 28. 110-114 (1997)

    • Description
      「研究成果報告書概要(欧文)」より
  • [Publications] Iwaki, A., Nagano, T., Nakagawa, M., Iwaki, T.and Fukumaki, Y.: "Identification and characterization of the gene encoding a new member of the alpha-crystallin/small hsp family, closely linked to the alphaB-crystallin gene in a head-to-head manner." Genomics. 45. 386-394 (1997)

    • Description
      「研究成果報告書概要(欧文)」より
  • [Publications] Nobuhisa, I., Inamasu, S., Nakai, M., Tatsui, A., Mimori, T., Ogawa, T., Shimohigashi, Y., Fukumaki, Y., Hattori, S.Kihara, H.and Ohno, M.: "Characterization and evolution of a gene encoding a Trimeresurus flavoviridisserm protein that inhibits basic phosphlipase A2 isozymes in the snake's venom." Eur.J.Biochem.249. 838-845 (1997)

    • Description
      「研究成果報告書概要(欧文)」より
  • [Publications] Kamakura, S., Iwaki, A., Matsumoto, M.and Fukumaki, Y.: "Cloning and characteriztion of the 5'-flanking region of the human dopamine D4 receptor gene." Biochem.Biophys.Res.Commun.235. 321-326 (1997)

    • Description
      「研究成果報告書概要(欧文)」より
  • [Publications] Ohba, Y.Hattori, Y., Harano, T., Harano, K., Fukumaki, Y., Ideguchi, H., Cho, H.I.and Park, S.S.: "beta-Thalassemia mutations in Japanese and Koreans." Hemoglobin. 21. 191-200 (1997)

    • Description
      「研究成果報告書概要(欧文)」より
  • [Publications] Nobuhisa, I.and Deshimaru, M., Chijiwa, T., Nakashima, K., Ogawa, T., Shimohigashi, Y., Fukumaki, Y., Hattori, S.Kihra, H.and Ohno, M.: "Structures of genes encoding phospholipase A2 inhibitors from the serum of Trimeresurus flaviridis snake." Gene. 191. 31-37 (1997)

    • Description
      「研究成果報告書概要(欧文)」より
  • [Publications] Furuya, H., Kukita, Y., Nagano, S., Sakai, Y., Yamashita, Y., Fukuyama, H., Inatomi, Y., Saito, Y., Koike, R., Tsuji, S., Fukumaki, Y., Hayashi, K.and Kobayashi, T.: "Adult onset globoid cell leukodystrophy (Krabbe disease) : analysis of galactosylceramidase cDNA from four Japanese patients." Hum.Gnenet. 100. 450-456 (1997)

    • Description
      「研究成果報告書概要(欧文)」より
  • [Publications] Furuumi, H., Firdous, N., Inoue, T., Ohta, H., Winichagoon, P., Fucharoen, S.and Fukumaki, Y.: "Molecular basis of beta-thalassemia in the Maldives." Hemoglobin. 22. 141-151 (1998)

    • Description
      「研究成果報告書概要(欧文)」より
  • [Publications] Tanphaichitr, V., Mahasandana, C., Pung-amritt, P., Veerakul, G., Kankirawatana, S., Suvatte, V., Tritilanunt, S.and Fukumaki, Y.: "Type I and II hereditary methemoglobinemia due to HADH-cytochrome b5 reductase deficiency in Thai patients." Internal Medicine. 13. 81-85 (1998)

    • Description
      「研究成果報告書概要(欧文)」より
  • [Publications] Nobiuhisa, I., Chiwata, T., Fukumaki, Y., Hattori, S., Shimohigashi, Y.and Ohno, M.: "Structural elements of Trimeresurus flaviridis serum inhibitors for recognition of its venome phospholipase A2 isozymes." FEBS Lettrs. 429. 385-389 (1998)

    • Description
      「研究成果報告書概要(欧文)」より
  • [Publications] Yamada, M., Komatsu, N., Kirito, K., Kashii, Y., Tomizuka, H., Okada, K., Endo, T., Fukumaki, Y., Shinjo, K., Abe, K.and Miura, Y.: "Thrombopoietin supports in vitro erythroid differentiation via its specific receptor c-MpI in a human leukemia cell line." Cell Growth & Differentiation. 9. 487-496 (1998)

    • Description
      「研究成果報告書概要(欧文)」より
  • [Publications] Nobuhisa, I., Ogawa, T., Deshimaru, M., Chijiwa, T., Nakashima, K., Chuman, Y., Shimohigashi, Y., Fukumaki, Y., Hattori, and Ohno, M.: "Retrotramsposable CR1-like elemnts in crotalinae sanke genomes." Toxicon. 36. 915-920 (1998)

    • Description
      「研究成果報告書概要(欧文)」より
  • [Publications] Mizuki, K., Kadomatsu, K., Hata, K., ito, T., Fan, Q., Kage, Y., Fukumaki, Y., Sakaki, Y., Takeshige, K.and Sumimoto, H.: "Functional modules and expression of mouse p40^<phox> and p65^<phox> SH3-domain-containing proteins involved in the phagocyte NADPH oxidase complex." Eur.J.Biochem.251. 573-582 (1998)

    • Description
      「研究成果報告書概要(欧文)」より
  • [Publications] Mamiya, K., Ieiri, I., Miyahara, S., Imai, J., Furuumi, H., Fukumaki, Y., Ninomiya, H., Tashiro, N., Yamnada, H.and Higuchi, S.: "Association of polymorphisms in the cytochrome P450 (CYP) 2C19 and 2C18 genes inn Japanese epileptic patients." Pharmacogenetics. 8. 87-90 (1998)

    • Description
      「研究成果報告書概要(欧文)」より
  • [Publications] Matsumoto, M., Yamaguchi, T., Fukumaki, Y., Yasunaga, R.and Terada, S.: "High pressure induces G2 arrest in murine erythroleukemia cells." J.Biochem.123. 87-93 (1998)

    • Description
      「研究成果報告書概要(欧文)」より
  • [Publications] Higasa, K., Manabe, J., Yubisui, T., Sumimoto, H., Amritt, P., Tanpahichitr, V., Fukumaki, Y.: "Molecular basis of hereditary methemoglbinemia types, I and II." Brt.J.Haematol. 103. 922-930 (1998)

    • Description
      「研究成果報告書概要(欧文)」より
  • [Publications] Inoue, T., Yamaza, H., Sakai, Y., Mizuno, S., Ohno, M., Hamasaki, N., Fukumaki, Y.: "Position-independent human beta-globin gene expression mediated by a recombinant adeno-associated virus vector carrying the chicken beta-globin insulator." J.Hum.Genet.(in press).

    • Description
      「研究成果報告書概要(欧文)」より
  • [Publications] Hamamura, M., Ozawa, H., Kimuro, Y., Higasa, K., Iwaki, A., Fukumaki, Y.: "Differential dicreases in c-fos and aldolase C mRNA expression in the rat cerebellum after repeated methamphetamine administration." Mol.Brain Res.(in press).

    • Description
      「研究成果報告書概要(欧文)」より

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Published: 1999-12-08  

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