Relaxin expression from tumor-targeting adenoviruses and its intratumoral spread, apoptosis induction, and efficacy
Relaxin expression from tumor-targeting adenoviruses and its intratumoral spread, apoptosis induction, and efficacy
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DOI：
10.1093/jnci/djj397
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发表时间：
2006-10-18
影响因子：
10.3
通讯作者：
Yun, Chae-Ok
中科院分区：
文献类型：
--
作者：
Kim, Joo-Hang;Lee, Young-Sook;Yun, Chae-Ok
溶瘤腺病毒在肿瘤中的渗透和分布不均匀，限制了其作为肿瘤基因治疗的应用。我们研究了腺病毒表达细胞基质降解蛋白松弛蛋白是否可以改善腺病毒在肿瘤中的分布和渗透。方法：将松弛蛋白基因插入E3腺病毒区域，构建松弛蛋白表达、复制能力强(dl-LacZ-RLX)和活性(Ad-Delta E1B-RLX)腺病毒。对照组为亲代腺病毒(dl-LacZ和Ad-Delta E1B)和磷酸盐缓冲液(PBS)(载体)。不能复制的病毒不能溶解细胞，用来评估转导效率。病毒在肿瘤球体中的传播是通过将肿瘤组织解剖成均匀的片段，通过报告基因(即LacZ)的表达来评估的。通过将腺病毒注射到裸鼠(n=8或9)的移植瘤中来评价对肿瘤生长的抑制作用。采用Kaplan-Meier方法评估患者的总体生存率。用Masson‘s三色染色检测细胞外基质。通过评估B16BL6黑色素瘤小鼠模型的自发性肺转移和原位移植肝癌(n=4-6)的生长抑制来评价治疗效果。所有的统计检验都是双面的。结果：在肿瘤球体和体内已建立的实体瘤中，dl-LacZ-RLX的转导效率高于亲本病毒和载体剂，病毒在肿瘤中的传播速度更快。与亲本病毒相比，携带Ad-Delta E1B-RLX的C33A荷瘤小鼠的存活率提高(例如，用Ad-Delta E1B-RLX治疗的C33A荷瘤小鼠的存活率为100%，95%可信区间[CI]=63.1%~100%，而用Ad-Delta E1B治疗的C33A荷瘤小鼠的存活率为50%，95%CI=15.7%~84.3%)。与未感染组织相比，Ad-Delta E1B-RLX感染可显著降低肿瘤组织的胶原含量，但不影响邻近正常组织的胶原含量。瘤内注射Ad-Delta E1B-RLX可抑制小鼠肺转移瘤的形成(PBS=268 mg/只，Ad-Delta E1B-RLX=10 mg；差异=258 mg，95%CI=94~426；P=0.003，Mann-Whitney检验)。全身应用Ad-Delta E1B-RLX可完全抑制Hep1肝癌的生长(PBS=20.2 mg/只，Ad-Delta E1B-RLX=0 mg；差异=20.2 mg，95%CI=3.7~36.7；P=0.004，Mann-Whitney检验)。结论：腺病毒表达的松弛蛋白降解细胞外基质增加了病毒的分布和肿瘤的穿透性，抑制了肿瘤的生长和转移，提高了小鼠的生存时间。
The use of oncolytic adenoviruses as cancer gene therapy is limited by their uneven penetration and distribution in tumors. We investigated whether the expression of the cell matrix-degradative protein relaxin by adenovirus could improve adenovirus distribution and penetration in tumors. Methods: We generated relaxin-expressing, replication-ineompetent (dl-lacZ-RLX) and -competent (Ad-Delta E1B-RLX) adenoviruses by inserting a relaxin gene into the E3 adenoviral region. Controls were parental adenoviruses (dl-lacZ and Ad-Delta E1B) and phosphate-buffered saline (PBS) (vehicle). Replication-incompetent viruses, which do not lyse cells, were used to assess transduction efficiency. Viral spread in tumor spheroids, made by dissecting tumor tissue into homogeneous fragments, was assessed by reporter gene (i.e., lacZ) expression. Tumor growth inhibition was assessed by injecting adenoviruses into xenograft tumors in athymic mice (n=8 or 9). Overall survival was assessed by the Kaplan-Meier method. Extracellular matrix was examined with Masson's trichrome staining. Therapeutic efficacy was evaluated by assessing spontaneous pulmonary metastasis in the B16BL6 melanoma mouse model and growth inhibition of orthotopically implanted hepatoma (n=4-6). All statistical tests were two-sided. Results: In tumor spheroids and established solid tumors in vivo, transduction with dl-lacZ-RLX, compared with parental virus or vehicle, elicited higher transduction efficiency and viral spread throughout the tumor mass. Infection with Ad-Delta E1B-RLX, compared with parental virus, elicited greater viral persistence and spread, leading to increased survival (e.g., 100%, 95% confidence interval [CI]=63.1% to 100%, for C33A tumor-bearing mice treated with Ad-Delta E1B-RLX, and 50%, 95% CI=15.7% to 84.3%, for C33A tumor-bearing mice treated with Ad-Delta E1B). Infection with Ad-Delta E1B-RLX substantially decreased the collagen content of tumor tissue but not of adjacent normal tissue, compared with noninfected tissues. Intratumoral injection of Ad-Delta E1B-RLX inhibited the formation of lung metastases in mice (PBS=268 mg of metastatic tumor per mouse and Ad-Delta E1B-RLX = 10 mg; difference = 258 mg, 95% Cl = 94 to 426; P=.003, Mann-Whitney test). Systemic treatment with Ad-Delta E1B-RLX completely inhibited the growth of Hep1 hepatocellular carcinomas (PBS=20.2 mg of tumor per mouse and Ad-Delta E1B-RLX=0 mg; difference=20.2 mg, 95% CI=3.7 to 36.7; P=.004, Mann-Whitney test). Conclusion: Extracellular matrix degradation by relaxin expressed by adenoviruses increased viral distribution and tumor penetration, inhibited tumor growth and metastasis, and increased survival of mice.