Ramnaraine Margaret, Pan Weihong, Clohisy Denis R
Department of Orthopedic Surgery, University of Minnesota, 420 Delaware Street SE, MMC 806, Minneapolis, MN 55455, USA.
Bone. 2006 Jan;38(1):4-12. doi: 10.1016/j.bone.2005.07.016. Epub 2005 Sep 1.
Cytosine deaminase (CD) catalyzes the deamination of 5-fluorocytosine (5FC) to produce the highly toxic chemotherapeutic agent 5-fluorouracil (5FU). A unique feature of the CD/5FC enzyme/prodrug system is its ability to kill adjacent cells via bystander killing. Bystander killing of cancer cells can be mediated by non-cancerous accessory cells transduced with the CD gene; one type of non-cancerous accessory cell found in primary bone cancer and breast cancer metastases to bone is the osteoclast. This manuscript determines if osteoclast precursor cells, transduced with the CD gene, can function as a gene delivery system capable of killing cancer cells. An osteoclast precursor cell line (RAW 264.7, RAW) and authentic bone marrow-derived osteoclast precursor cells were transduced with a retroviral vector containing the cytosine deaminase fusion gene (NCD) composed of the human nerve growth factor receptor and CD genes. RAW cells and bone marrow-derived osteoclast precursor cells transduced with NCD expressed NCD protein and converted 5FC to 5FU. Treatment of NCD-transduced osteoclast precursor cells with the 5FC prodrug resulted in significant killing in vitro. NCD-transduced osteoclasts were co-cultured with either DsRed2-labeled sarcoma cells (2472-DSR) or green fluorescent protein (GFP)-labeled breast cancer cells (GFP-4T1). Treatment of the NCD osteoclast/tumor cell co-cultures with 5FC resulted in bystander killing of 2472-DSR cells (P < 0.006) and GFP-4T1 cells (P < 0.004). These findings demonstrate that NCD-transduced osteoclasts can promote killing of cancer cells and introduce the exciting possibility for developing osteoclast-mediated, CD-based treatment of primary bone cancers and breast cancer metastases to bone.
胞嘧啶脱氨酶（CD）催化5-氟胞嘧啶（5FC）脱氨生成剧毒化疗药物5-氟尿嘧啶（5FU）。CD/5FC酶/前药系统的一个独特特征是其通过旁观者杀伤作用杀死邻近细胞的能力。癌细胞的旁观者杀伤可由转导了CD基因的非癌细胞辅助细胞介导；在原发性骨癌和乳腺癌骨转移灶中发现的一种非癌细胞辅助细胞是破骨细胞。本研究确定转导了CD基因的破骨细胞前体细胞是否能作为一种能够杀死癌细胞的基因传递系统。用含有由人神经生长因子受体和CD基因组成的胞嘧啶脱氨酶融合基因（NCD）的逆转录病毒载体转导破骨细胞前体细胞系（RAW 264.7，RAW）和真正的骨髓来源破骨细胞前体细胞。用NCD转导的RAW细胞和骨髓来源破骨细胞前体细胞表达NCD蛋白并将5FC转化为5FU。用5FC前药处理转导了NCD的破骨细胞前体细胞在体外导致显著杀伤。将转导了NCD的破骨细胞与DsRed2标记的肉瘤细胞（2472-DSR）或绿色荧光蛋白（GFP）标记的乳腺癌细胞（GFP-4T1）共培养。用5FC处理NCD破骨细胞/肿瘤细胞共培养物导致2472-DSR细胞（P<0.006）和GFP-4T1细胞（P<0.004）的旁观者杀伤。这些发现表明，转导了NCD的破骨细胞可促进癌细胞的杀伤，并为开发破骨细胞介导的、基于CD的原发性骨癌和乳腺癌骨转移治疗带来了令人兴奋的可能性。
