Khatri Aparajita, Zhang Bing, Doherty Eboney, Chapman Jane, Ow Kim, Pwint Hnin, Martiniello-Wilks Rosetta, Russell Pamela J
Oncology Research Centre, Prince of Wales Hospital Clinical School of Medicine, The University of New South Wales, Randwick, NSW 2031, Australia.
J Gene Med. 2006 Sep;8(9):1086-96. doi: 10.1002/jgm.944.
We aimed to evaluate the efficacy of gene-directed enzyme-prodrug therapy (GDEPT) using cytosine deaminase in combination with uracil phosphoribosyl transferase (CDUPRT) against intraprostatic mouse androgen-refractory prostate (RM1) tumors in immunocompetent mice. The product of the fusion gene, CDUPRT, converts the prodrug, 5-fluorocytosine (5FC), into 5-fluorouracil (5FU) and other cytotoxic metabolites that kill both CDUPRT-expressing and surrounding cells, via a 'bystander effect'.
Stably transformed andogen-independent mouse prostate cancer (PC) cells, RM1-CDUPRT, -GFP or GFP/LacZ cells were used. To assess the local bystander effects of CDUPRT-GDEPT, immunocompetent C57BL/6 mice implanted with cell mixtures of RM1-GFP/CDUPRT and RM1-GFP cells in different proportions intraprostatically were treated with 5FC. Pseudo-metastases in the lungs were established by a tail vein injection of untransfected RM1 cells. At necropsy, prostate weight/volume and lung colony counts were assessed. Tumors, lymph nodes, spleens and lungs were frozen or fixed for immunohistochemistry.
CDUPRT expression in RM1-GFP/CDUPRT cells or tumors was confirmed by enzymic conversion of 5FC into 5FU, using HPLC. Treatment of mice bearing intraprostatic RM1-GFP/CDUPRT tumors with 5FC resulted in complete regression of the tumors. A 'local bystander effect' was seen, even though only 20% of the cells expressed CDUPRT. More importantly a significant reduction in pseudo-metastases of RM1 cells in lungs indicated a 'distant bystander effect'. Immunohistochemical evaluation of the treated tumors showed increased necrosis and apoptosis, with decreased tumor vascularity. There was also a significant increase in tumour-infiltration by macrophages, CD4+ T and natural killer cells.
We conclude that CDUPRT-GDEPT significantly suppressed the aggressive growth of RM1 prostate tumors and lung pseudo-metastases via immune mechanisms involving necrosis and apoptosis.
我们旨在评估使用胞嘧啶脱氨酶联合尿嘧啶磷酸核糖基转移酶（CDUPRT）的基因导向酶-前药疗法（GDEPT）对免疫功能正常小鼠前列腺内雄激素抵抗性小鼠前列腺癌（RM1）肿瘤的疗效。融合基因CDUPRT的产物将前药5-氟胞嘧啶（5FC）转化为5-氟尿嘧啶（5FU）和其他细胞毒性代谢产物，这些产物通过“旁观者效应”杀死表达CDUPRT的细胞和周围细胞。
使用稳定转化的雄激素非依赖性小鼠前列腺癌细胞RM1-CDUPRT、-GFP或GFP/LacZ细胞。为了评估CDUPRT-GDEPT的局部旁观者效应，对前列腺内植入不同比例的RM1-GFP/CDUPRT和RM1-GFP细胞混合物的免疫功能正常的C57BL/6小鼠用5FC进行治疗。通过尾静脉注射未转染的RM1细胞在肺部建立假转移灶。尸检时，评估前列腺重量/体积和肺集落计数。将肿瘤、淋巴结、脾脏和肺冷冻或固定用于免疫组织化学分析。
使用高效液相色谱法通过将5FC酶促转化为5FU证实了RM1-GFP/CDUPRT细胞或肿瘤中CDUPRT的表达。用5FC治疗前列腺内携带RM1-GFP/CDUPRT肿瘤的小鼠导致肿瘤完全消退。即使只有20%的细胞表达CDUPRT，也观察到了“局部旁观者效应”。更重要的是，肺部RM1细胞假转移灶的显著减少表明存在“远处旁观者效应”。对治疗后的肿瘤进行免疫组织化学评估显示坏死和凋亡增加，肿瘤血管减少。巨噬细胞、CD4+T细胞和自然杀伤细胞对肿瘤的浸润也显著增加。
我们得出结论，CDUPRT-GDEPT通过涉及坏死和凋亡的免疫机制显著抑制了RM1前列腺肿瘤的侵袭性生长和肺假转移灶。
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