Noninvasive detection of lentiviral-mediated choline kinase targeting in a human breast cancer xenograft.
Noninvasive detection of lentiviral-mediated choline kinase targeting in a human breast cancer xenograft.
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DOI：
10.1158/0008-5472.can-08-4120
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发表时间：
2009-04-15
期刊：
影响因子：
11.2
通讯作者：
Bhujwalla ZM
中科院分区：
文献类型：
--
作者：
Krishnamachary B;Glunde K;Wildes F;Mori N;Takagi T;Raman V;Bhujwalla ZM
高磷胆碱（PC）和总胆碱代谢物是包括乳腺癌在内的大多数癌细胞广泛确定的特征。胆碱激酶（chk）是一种将胆碱转化为磷酸胆碱的酶，有效的沉默与肿瘤生长的减少有关。使用RNAi的功能重要性和chk的下调已被先前确定。在这里，我们报告了慢病毒载体介导的使用短发夹RNA （shRNA）在人乳腺癌细胞衍生的肿瘤中下调胆碱激酶（chk）的临床前评估。将表达shRNA的浓缩慢病毒静脉注射于MDA-MB-231型荷瘤雌性严重联合免疫缺陷（SCID）小鼠的尾静脉。通过增强的绿色荧光蛋白（EGFP）表达以及chk mRNA和蛋白水平来评估细胞和肿瘤体内的转导效率。在细胞中几乎100%的转导效率后，chk mRNA和蛋白质减少了80%。chk-shRNA转导后，细胞和肿瘤提取物的1H磁共振谱（MRS）显示，与对照组相比，PC和总胆碱水平降低（P分别< 0.01和0.05）。采用肿瘤组织中31P MRS、细胞和肿瘤组织提取物中1H MRS无创监测PC水平。体内chk-shRNA转导肿瘤的非侵入性31P MR谱显示，较低的PC和磷酸单酯（PME）水平与肿瘤生长和增殖减少有关。本研究展示了慢病毒载体在人乳腺癌异种移植物中靶向chk的使用，以及这种靶向的无创MRS检测。
Elevated phosphocholine (PC) and total choline metabolites are widely established characteristics of most cancer cells including breast cancer. Effective silencing of choline kinase (chk), the enzyme that converts choline to phosphocholine, is associated with reduced tumor growth. The functional importance and down regulation of chk using RNAi has been previously established. Here, we report on the preclinical evaluation of lentiviral vector-mediated downregulation of choline kinase (chk) using short hairpin RNA (shRNA), in established tumors derived from human breast cancer cells. Concentrated lentivirus expressing shRNA against chk was injected intravenously in the tail vein of MDA-MB-231 tumor bearing female severe combined immunodeficient (SCID) mice. Transduction efficiency in cells and tumors in vivo, was assessed optically by enhanced green fluorescent protein (EGFP) expression and additionally from chk mRNA and protein levels. An 80% reduction in chk mRNA and protein was achieved following almost 100% transduction efficiency in cells. Post-transduction with chk-shRNA, 1H magnetic resonance spectroscopy (MRS) of cell and tumor extracts showed decreases in PC and total choline levels (P < 0.01 and 0.05 respectively) in comparison to controls. PC levels were monitored non-invasively by 31P MRS in tumors, and by 1H MRS in cell and tumor tissue extracts. Noninvasive 31P MR spectra of chk-shRNA transduced tumors in vivo showed lower PC and phosphomonoester (PME) levels that were associated with reduced tumor growth and proliferation. This study demonstrates the use of lentiviral vectors to target chk in a human breast cancer xenograft, and noninvasive MRS detection of this targeting.