Israyelyan Anna H, Melancon Jeffrey M, Lomax Larry G, Sehgal Inder, Leuschner Carola, Kearney Michael T, Chouljenko Vladimir N, Baghian Abolghasem, Kousoulas Konstantin G
Division of Biotechnology and Molecular Medicine and Department of Pathobiological Sciences, School of Veterinary Medicine, Louisiana State University, Baton Rouge, LA 70803, USA.
Hum Gene Ther. 2007 May;18(5):457-73. doi: 10.1089/hum.2006.145.
A new oncolytic and fusogenic herpes simplex virus type 1 (HSV-1) was constructed on the basis of the wildtype HSV-1(F) strain. To provide for safety and tumor selectivity, the virus carried a large deletion including one of the two alpha4, gamma(1)34.5, alpha0 genes and the latency-associated transcript region. The gamma(1)34.5 gene, a major neurovirulence factor, was replaced by a gene cassette constitutively expressing the red fluorescent protein gene. Homologous recombination was used to transfer the fusogenic gBsyn3 mutation to the viral genome to produce the OncSyn virus. OncSyn causes extensive virus-induced cell fusion (syncytia) and replicates to higher titers than the parental Onc and HSV-1(F) strains in breast cancer cells. Biochemical analysis revealed that the OncSyn virus retains a stable genome and expresses all major viral glycoproteins. A xenograft mouse model system using MDA-MB-435S-luc (MM4L) human breast cancer cells constitutively expressing the luciferase gene implanted within the interscapular region of animals was used to test the ability of the virus to inactivate breast tumor cells in vivo. Seventy-two mice bearing MM4L breast cancer xenografts were randomly divided into three groups and given two rounds of three consecutive intratumoral injections of OncSyn, inactivated OncSyn, or phosphate-buffered saline 3 days apart. A single round of virus injections resulted in a drastic reduction of tumor sizes (p <or= 0.0001) and diminution of chemiluminescence emitted by the cancer cells (p <or= 0.0002). This effect was enhanced by a second round of virus injections into the tumors 3 days after the first round (p <or= 0.0001). Systematic necropsy and pathological evaluation of the primary tumors revealed that the single round of injections resulted in extensive necrosis of tumor cells (p <or= 0.0001), which was enhanced by the second round of injections (p <or= 0.0002). Internal organs were not affected by virus inoculation. Mouse weights were not significantly impacted by any treatment during the course of the entire study (p = 0.46). These results show that the attenuated, fusogenic, and oncolytic HSV-1(F) virus strain OncSyn may effectively treat human breast tumors in vivo.
一种新型的溶瘤性和融合性单纯疱疹病毒1型（HSV-1）是在野生型HSV-1(F)毒株的基础上构建的。为确保安全性和肿瘤选择性，该病毒携带一个大的缺失区域，包括两个α4、γ(1)34.5、α0基因中的一个以及潜伏相关转录区。γ(1)34.5基因是一种主要的神经毒力因子，被一个组成型表达红色荧光蛋白基因的基因盒所取代。利用同源重组将融合性gBsyn3突变转移到病毒基因组中，从而产生OncSyn病毒。OncSyn在乳腺癌细胞中可引起广泛的病毒诱导的细胞融合（多核巨细胞），并且比亲代Onc和HSV-1(F)毒株复制到更高的滴度。生化分析表明，OncSyn病毒保留了稳定的基因组，并表达所有主要的病毒糖蛋白。使用在动物肩胛间区域植入的组成型表达荧光素酶基因的MDA-MB-435S-luc（MM4L）人乳腺癌细胞的异种移植小鼠模型系统，来测试该病毒在体内使乳腺肿瘤细胞失活的能力。72只携带MM4L乳腺癌异种移植瘤的小鼠被随机分为三组，每隔3天进行两轮连续三次瘤内注射OncSyn、灭活的OncSyn或磷酸盐缓冲盐水。单次病毒注射导致肿瘤大小急剧减小（p≤0.0001），并且癌细胞发出的化学发光减弱（p≤0.0002）。第一轮注射3天后对肿瘤进行第二轮病毒注射可增强这种效果（p≤0.0001）。对原发性肿瘤进行系统的尸检和病理评估发现，单次注射导致肿瘤细胞广泛坏死（p≤0.0001），第二轮注射可增强这种坏死（p≤0.0002）。病毒接种未影响内部器官。在整个研究过程中，任何治疗对小鼠体重均无显著影响（p = 0.46）。这些结果表明，减毒、融合性和溶瘤性的HSV-1(F)病毒株OncSyn可能在体内有效治疗人类乳腺肿瘤。
