KR101542275B1 - 폭스바이러스성 온콜리틱 벡터 - Google Patents
폭스바이러스성 온콜리틱 벡터 Download PDFInfo
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- KR101542275B1 KR101542275B1 KR1020107010821A KR20107010821A KR101542275B1 KR 101542275 B1 KR101542275 B1 KR 101542275B1 KR 1020107010821 A KR1020107010821 A KR 1020107010821A KR 20107010821 A KR20107010821 A KR 20107010821A KR 101542275 B1 KR101542275 B1 KR 101542275B1
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- C12N2710/24111—Orthopoxvirus, e.g. vaccinia virus, variola
- C12N2710/24141—Use of virus, viral particle or viral elements as a vector
- C12N2710/24143—Use of virus, viral particle or viral elements as a vector viral genome or elements thereof as genetic vector
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Abstract
Description
자살 유전자 | 예비약물(predrug) |
티미딘 키나제 | 간시클로버(Ganciclovir); 간시클로버 엘라이딘산 에스테르; 펜시클로버(Penciclovir); 아시클로버(Acyclovir); 발라시클로버(Valacyclovir); (E)-5-(2-브로모비닐)-2'-데옥시우리딘; 지도부딘(zidovudine); 2'-엑소(Exo)-메타노카르바티미딘 |
시토신 데아미나제 | 5-플루오로시토신 |
퓨린 뉴클레오시드 포스포릴라아제 | 6-메틸퓨린 데옥시리보시드; 플루다라빈(Fludarabine) |
우라실 포스포리보실 트랜스퍼라제 | 5-플루오로시토신; 5-플루오로우라실 |
티미딜레이트 키나제 | 아지도티미딘 |
도 2. 감염 후 5일째, 표시된 바이러스로 0.0001의 MOI에서 감염된 LoVo에서 바이러스의 시험관내 복제 효율. 값들은 세개의 개별 측정의 평균 ± SD로 나타나있다.
도 3. 바이러스의 정맥내 주사 후 스위스 무모 마우스에서 s.c LoVo의 평균 종양 체적 ± SEM. 종양 (촉진성 종양)의 접종으로부터 7일 후, 마우스를 107 pfu의 완충액 + 염수 (◇), 완충액 + 5-FC (◆), VVTK-F2L-/FCU1 + 염수 (□) 또는 VVTK-F2L-/FCU1 + 5-FC (■)으로 치료하였다. 동물들을 3주 동안 바이러스 주사로부터 7일 후, 경구 가바즈로 1일 2회 100 mg/kg/j에서 염수 또는 5-FC로 치료하였다. 종양 체적을 주 2회 측정하였다.
도 4. 바이러스의 정맥내 주사 후 스위스 무모 마우스에서 s.c HepG2 종양의 평균 체적 ± SEM. 종양 (촉진성 종양)의 접종으로부터 14일 후, 마우스를 완충액 + 물 (◇), 또는 완충액 + 5-FC (◆), 또는 106 pfu의 VVTK-/FCU1 + 물 (○), 또는 106 pfu의 VVTK-/FCU1 + 5-FC (●) (A); 또는 완충액 + 물 (◇), 또는 완충액 + 5-FC (◆) 또는 106 pfu의 VVTK-F2L-/FCU1 + 물 (◇), 또는 106 pfu의 VVTK-F2L-/FCU1 + 5-FC (◆) (B)으로 치료하였다. 동물들을 3주 동안 및 바이러스 주사로부터 7일 후, 경구 가바즈로 1일 2회 100 mg/kg에서 5-FC로 치료하였다. 종양 체적을 주 2회 측정하였다.
도 5. 분할 세포내 대 전면 세포내의 바이러스 수율 비율. PANC1 (췌장 인간 종양), H1299 (폐 인간 종양) 또는 U118MG (신경교종 인간 종양) 세포를 100 pfu의 (■) VVTK-/FCU1 또는 (□) VVTK-F2L-/FCU1로 감염시켰다. 감염 48시간 후, 바이러스 역가를 구하였다. 값들은 분할 세포내 대 전면 세포내의 바이러스의 수율간의 비율이다.
도 6. 1x106 PFU의 VVTK-/FCU1 (■) 또는 VVTK-F2L-/FCU1 (□)으로 피하 인간 종양을 보유하는 스위스 무모 마우스내로 정맥내 감염 후 6일째 및 21일째 기관 또는 종양에서의 바이러스 역가 (pfu/mg의 조직).
도 7. 정맥내 주사로 1x108 pfu의 VVTK-/FCU1 (■) 또는 VVTK-F2L-/FCU1 (◇)로의 치료 후 스위스 무모 마우스의 생존.
도 8. 정맥내 주사로 1x107 pfu (A) 또는 1x108 pfu (B)의 VVTK-/FCU1 (■) 또는 VVTK-F2L-/FCU1 (◇)로의 치료 후 면역적격 B6D2 마우스의 생존.
도 9. 감염 후 13일째 및 감염 후 34일째 스위스 무모 마우스내 1x106 pfu VVTK-/FCU1 또는 VVTK-F2L-/FCU1의 정맥내 주사 후 꼬리상 두진의 평균량.
도 10. 감염 후 15일째 및 감염 후 31일째 스위스 무모 마우스내 1x107 pfu VVTK-/FCU1 또는 VVTK-F2L-/FCU1의 정맥내 주사 후 꼬리상 두진의 평균량.
종양 | 폐 | 비장 | 신장 | 심장 | 림프절 | |
VVTK-/FCU1 | (0,2-3,3)x105 | 0.1-2 | 0-2.2 | 0-1.8 | 0-0.3 | 0-61 |
VVTK-F2L/FCU1 | 0-8.1x104 | 0-5.7 | 0-9.3 | 0.2-3 | 0,2 | 0-83 |
난소 | 피부 | 꼬리 | 골수 | 뇌 | 근육 | 심장 | |
VVTK-/FCU1 | 2.2-74 | 0.1-24 | 13.5-7.104 | 0-800 | 0-1.8 | 0-22 | 0-0.3 |
VVTK-F2L/FCU1 | 0-108 | 0-88 | 0.8-16.3 | 측정되지 않음(n.d) | 0-0.2 | 0-1.8 | 0,2 |
Claims (33)
- 결손 F2L 유전자,
결손 J2R 유전자, 및
자살 유전자를 포함하는 관심 핵산
을 포함하는 폭스바이러스. - 삭제
- 제1항에 있어서, 코르도폭스비리내 아과에 속하는 폭스바이러스.
- 제3항에 있어서, 백시니아 바이러스 종에 속하는 폭스바이러스.
- 제4항에 있어서, 백시니아 바이러스 균주 WR인 폭스바이러스.
- 제4항에 있어서, 백시니아 바이러스 균주 코펜하겐인 폭스바이러스.
- 제1항에 있어서, 상기 자살 유전자가, 적어도 시토신 데아미나제 활성을 가지는 단백질을 코딩하는 것인 폭스바이러스.
- 제7항에 있어서, 상기 자살 유전자가 FCY1, FCA1 또는 CodA인 폭스바이러스.
- 제7항에 있어서, 적어도 시토신 데아미나제 활성을 가지는 상기 단백질이 서열 확인번호 2에 나타난 FCU1-8 폴리펩티드인 폭스바이러스.
- 제1항에 있어서, 상기 자살 유전자가, 시토신 데아미나제 활성, 우라실 포스포리보실 트랜스퍼라제 활성, 또는 두 활성 모두를 가지는 단백질을 코딩하는 것인 폭스바이러스.
- 제10항에 있어서, 상기 자살 유전자가, 서열 확인번호 3 (coda::upp), 서열 확인번호 1 (FCU1) 또는 FCY1::FUR1의 아미노산 서열에 나타난 바와 같은 아미노산 서열을 포함하는 폴리펩티드를 코딩하는 것인 폭스바이러스.
- 제1항에 있어서, 투과효소를 코딩하는 유전자를 포함하는 핵산 서열을 추가적으로 포함하는 폭스바이러스.
- 제12항에 있어서, 상기 투과효소가 S. 세레비시애의 퓨린 또는 시토신 투과효소인 폭스바이러스.
- 제13항에 있어서, 상기 투과효소가 FCY2 및 Fur4를 포함하는 군으로부터 선택된 것인 폭스바이러스.
- 제1항에 있어서, 관심 핵산의 발현에 필요한 요소를 추가적으로 포함하는 폭스바이러스.
- 제12항에 있어서, 투과효소를 코딩하는 유전자를 포함하는 핵산 서열의 발현에 필요한 요소를 추가적으로 포함하는 폭스바이러스.
- 결손 F2L 유전자 및 결손 J2R 유전자를 포함하는 암 치료용 폭스바이러스.
- 제17항에 있어서, 관심 핵산을 추가적으로 포함하는 폭스바이러스.
- 제18항에 있어서, 상기 관심 핵산이, 치료 분자를 코딩하는 하나 이상의 관심 서열을 함유하는 것인 폭스바이러스.
- (i) 제1항에 따른 또는 제17항 내지 제19항 중 어느 한 항에서 정의된 바와 같은 폭스바이러스를 시험관 내에서(in vitro) 세포내로 도입하는 단계;
(ii) 상기 폭스바이러스가 제조될 수 있게 하기에 적절한 조건하에서 상기 세포를 배양하는 단계; 및
(iii) 상기 폭스바이러스를 세포 배양물로부터 회수하는 단계
의, 제1항에 따른 또는 제17항 내지 제19항 중 어느 한 항에서 정의된 바와 같은 폭스바이러스의 제조 방법. - 제약학상 허용가능한 부형제와 함께 제1항에 따른 또는 제17항 내지 제19항 중 어느 한 항에서 정의된 바와 같은 폭스바이러스를 포함하는, 암, 종양, AIDS, 재발협착증(restenosis), 류마티스성 관절염 또는 골다공증을 치료하기 위한 조성물.
- 제21항에 있어서, 5-플루오로시토신의 세포독성 효과를 강화시키는 하나 이상의 물질과 조합된 조성물.
- 제22항에 있어서, 5-플루오로시토신의 세포독성 효과를 강화시키는 상기 물질이 피리미딘의 신생합성의 경로의 효소를 억제하는 약물인 조성물.
- 제22항에 있어서, 5-플루오로시토신의 세포독성 효과를 강화시키는 상기 물질이 메토트렉세이트인 조성물.
- 제21항에 있어서, 암 치료를 위한 조성물.
- 제1항에 따른 또는 제17항 내지 제19항 중 어느 한 항에서 정의된 바와 같은 폭스바이러스를 포함하며, 암, 종양, AIDS, 재발협착증(restenosis), 류마티스성 관절염 또는 골다공증 치료를 필요로 하는 숙주 유기체 또는 세포에 투여되는, 상기 치료를 위한 조성물.
- 제26항에 있어서, 전신 경로를 통하여 투여되는 조성물.
- 제26항에 있어서, 추가적으로 제약학상 허용가능한 양의 전구약물과 함께 숙주 유기체 또는 세포에 투여되는 조성물.
- 제28항에 있어서, 상기 전구약물의 투여가 상기 조성물의 투여로부터 3일 이상 후에 실시되는 것인 조성물.
- 제29항에 있어서, 상기 전구약물의 투여가 상기 조성물의 투여로부터 4일 이상 후에 실시되는 것인 조성물.
- 제29항에 있어서, 상기 전구약물의 투여가 상기 조성물의 투여로부터 5일 이상 후에 실시되는 것인 조성물.
- 제29항에 있어서, 상기 전구약물의 투여가 상기 조성물의 투여로부터 7일 후에 실시되는 것인 조성물.
- 제23항에 있어서, 피리미딘의 신생합성의 경로의 효소를 억제하는 약물이 PALA, 레플루노미드 및 A771726으로 구성된 군으로부터 선택된 것인 조성물.
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