Parker L P, Wolf J K, Price J E
The University of Texas M. D. Anderson Cancer Center, Houston, USA.
Ann Clin Lab Sci. 2000 Oct;30(4):395-405.
Our objective was to determine the efficacy of adenoviral-mediated gene therapy with wild-type p53 or p21 in human breast cancer cells and investigate interactions with radiation and chemotherapy. Two human breast cancer cell lines, MDA-MB-231 and MDA-MB-435, both with p53 mutations, were transduced with adenoviral vectors containing wild-type p53 (Ad5CMV-p53) or p21/WAF1/Cip1 (Ad5CMV-p21), and the effects on growth were determined. Infection was combined with low-dose (1.4 - 3.7 Gy) irradiation to see if this would improve transduction efficiency and enhance numbers of cells killed. Transduction with either vector resulted in expression of p21WAF1/cip1 and growth inhibition, although Ad5CMV-p53 transduction produced greater growth inhibition than did Ad5CMV-p21. The cell lines differed in sensitivity to the vectors. The Ad5CMV-p53 vector in a multiplicity of infection (MOI) of 125 resulted in 50% to 80% inhibition of MDA-MB-231, while MOI 250 of the same vector resulted in 27% inhibition of MDA-MB-435. Infection with Ad5CMV-p21 produced modest growth inhibition in both cell lines (< or = 40% at MOI 200), although protein expression was detected at lower viral doses. Low dose gamma-irradiation (1.4 to 3.7 Gy) was used to try and improve the rate of gene transfer. Modest increases in transduction efficiency and duration of expression of a vector containing beta-galactosidase occurred in irradiated breast cancer cells. Radiation 24 hr before transduction with Ad5CMV-p53 increased the proportions of apoptotic MDA-MB-231 cells. The cells transduced with Ad5CMV-p21 were arrested in G1, yet when they were irradiated before adenoviral transduction, the overexpression of p21 protected the cells from the cytotoxic effects of the radiation. Clonogenic assays showed that Ad5CMV-p21 reduced the sensitivity of MDA-MB-231 to VP-16 and paclitaxel. Combining these drugs with Ad5CMV-p53 did not consistently or significantly decrease clonogenic survival.
我们的目标是确定腺病毒介导的野生型p53或p21基因治疗对人乳腺癌细胞的疗效，并研究其与放疗和化疗的相互作用。将含有野生型p53（Ad5CMV-p53）或p21/WAF1/Cip1（Ad5CMV-p21）的腺病毒载体转导至两种均存在p53突变的人乳腺癌细胞系MDA-MB-231和MDA-MB-435中，并测定其对细胞生长的影响。将感染与低剂量（1.4 - 3.7 Gy）照射相结合，以观察这是否会提高转导效率并增加杀伤细胞数量。尽管Ad5CMV-p53转导产生的生长抑制作用比Ad5CMV-p21更强，但两种载体转导均导致p21WAF1/cip1表达及生长抑制。这两种细胞系对载体的敏感性不同。感染复数（MOI）为125的Ad5CMV-p53载体导致MDA-MB-231细胞生长抑制50%至80%，而相同载体MOI为250时导致MDA-MB-435细胞生长抑制27%。Ad5CMV-p21感染在两种细胞系中均产生适度的生长抑制（MOI为200时≤40%），尽管在较低病毒剂量时即可检测到蛋白表达。使用低剂量γ射线照射（1.4至3.7 Gy）来尝试提高基因转移率。照射后的乳腺癌细胞中，含有β-半乳糖苷酶的载体的转导效率和表达持续时间有适度增加。在用Ad5CMV-p53转导前24小时进行照射可增加MDA-MB-231凋亡细胞的比例。用Ad5CMV-p21转导的细胞停滞在G1期，然而，当它们在腺病毒转导前接受照射时，p21的过表达保护细胞免受辐射的细胞毒性作用。克隆形成试验表明，Ad5CMV-p21降低了MDA-MB-231对VP-16和紫杉醇的敏感性。将这些药物与Ad5CMV-p53联合使用并未持续或显著降低克隆形成存活率。
