采用具有针对自分泌调节生长途径和bcl-2的结合位点的双特异性反义寡核苷酸进行联合化疗以治疗前列腺肿瘤。
Combination chemotherapy employing bispecific antisense oligonucleotides having binding sites directed against an autocrine regulated growth pathway and bcl-2 for the treatment of prostate tumors.
作者信息
Rubenstein Marvin, Tsui Paulus, Guinan Patrick
机构信息
Division of Cellular Biology, Hektoen Institute for Medical Research, 2100 W. Harrison Street, Chicago, IL 60612, USA.
出版信息
Med Oncol. 2007;24(4):372-8. doi: 10.1007/s12032-007-0023-y.
In previous studies we demonstrated that antisense oligonucleotides (oligos) against transforming growth factor-alpha (TGF-alpha [MR1]), its binding site the epidermal growth factor receptor (EGFR [MR2]), and the anti-apoptosis protein bcl-2 (MR4) are efficacious against prostate tumors. In recent reports we also describe how two of these mRNA directed binding sites can be synthesized sequentially within a single linear complementary strand and administered either in the presence or absence of additional therapeutic agents. In these continuing experiments "bispecific" oligo pairs were further evaluated in the presence or absence of Cytoxan, Taxol, or DES. One oligo pair recognized the binding sites for TGF-alpha and EGFR mRNA (TGF-alpha/EGFR [MR12] and EGFR/TGF-alpha [MR21]); another pair recognized binding sites for EGFR and bcl-2 (EGFR/bcl-2 [MR24] and bcl-2/EGFR [MR42]). Oligo pairs differ in their linear 5' to 3' binding site orientations, and were tested in vitro against PC-3 and LNCaP prostate tumor cell lines. Following cell attachment, incubations were for 2 days with the agents followed by 2 days in their absence. When tested against PC-3 cells and combined with LD50 Cytoxan, MR2, MR4, MR24, MR42 significantly inhibited 47.3, 45.7, 68.3, and 64.9%; with LD50 Taxol MR2, MR4, MR24, MR42 significantly inhibited 49.8, 45.8, 64.1, and 59.2%; and with LD50 DES MR2, MR4, MR24, MR42 significantly inhibited 66.6, 67.6, 64.3, and 67.2% respectively. Each agent significantly increased the inhibition produced by either oligo alone.LNCaP cells were also incubated with mono- and bispecific oligos in either the presence or absence of chemotherapeutics. MR2, MR4, MR24, MR42 produced significant inhibitions of 57.4, 58.4, 69.4, and 68.6% with LD50 Cytoxan; 70.4, 70.1, 73.6, and 74.0% with LD50 Taxol; and 49.8, 50.1, 59.6, and 53.9%, respectively with LD50 DES.A complete PC-3 experiment compared MR1, MR2, MR4, MR12, MR21, MR24 and MR42, in the presence of LD50 Cytoxan. Each oligo combined with Cytoxan significantly inhibited: MR1 by 51.0, MR2 by 55.0, MR4 by 58.0; MR12 by 56.0; MR21 by 61.1, MR24 by 65.5 and MR42 by 66.0%. Bispecifics directed against two different pathways, MR24, and MR42 were the most effective.A complete LNCaP experiment compared the same series of oligos also in the presence of LD50 Cytoxan. Each oligo combined with Cytoxan significantly inhibited: MR1 by 49.0, MR2 by 50.0, MR4 by 53.0; MR12 by 52.0; MR21 by 58.6, MR24 by 53.9 and MR42 by 58.0%.
在先前的研究中，我们证明了针对转化生长因子-α（TGF-α [MR1]）、其结合位点表皮生长因子受体（EGFR [MR2]）以及抗凋亡蛋白bcl-2（MR4）的反义寡核苷酸（oligos）对前列腺肿瘤有效。在最近的报告中，我们还描述了如何在单一线性互补链内顺序合成这些mRNA导向的两个结合位点，并在有或没有其他治疗剂的情况下给药。在这些持续的实验中，在有或没有环磷酰胺、紫杉醇或己烯雌酚的情况下，对“双特异性”寡核苷酸对进行了进一步评估。一对寡核苷酸识别TGF-α和EGFR mRNA的结合位点（TGF-α/EGFR [MR12]和EGFR/TGF-α [MR21]）；另一对识别EGFR和bcl-2的结合位点（EGFR/bcl-2 [MR24]和bcl-2/EGFR [MR42]）。寡核苷酸对在其线性5'至3'结合位点方向上有所不同，并在体外针对PC-3和LNCaP前列腺肿瘤细胞系进行了测试。细胞贴壁后，先用这些试剂孵育2天，然后在无试剂的情况下再孵育2天。当与半数致死剂量（LD50）的环磷酰胺联合测试时，MR2、MR4、MR24、MR42对PC-3细胞的抑制率分别显著为47.3%、45.7%、68.3%和64.9%；与LD50的紫杉醇联合时，MR2、MR4、MR24、MR42的抑制率分别显著为49.8%、45.8%、64.1%和59.2%；与LD50的己烯雌酚联合时，MR2、MR4、MR24、MR42的抑制率分别显著为66.6%、67.6%、64.3%和67.2%。每种试剂均显著增强了单独一种寡核苷酸所产生的抑制作用。LNCaP细胞也在有或没有化疗药物的情况下与单特异性和双特异性寡核苷酸一起孵育。与LD50的环磷酰胺联合时，MR2、MR4、MR24、MR42的抑制率分别显著为57.4%、58.4%、69.4%和68.6%；与LD50的紫杉醇联合时，抑制率分别显著为70.4%、70.1%、73.6%和74.0%；与LD50的己烯雌酚联合时，抑制率分别显著为49.8%、50.1%、59.6%和53.9%。一项完整的PC-3实验在存在LD50环磷酰胺的情况下比较了MR1、MR2、MR4、MR12、MR21、MR24和MR42。每种寡核苷酸与环磷酰胺联合时均显著抑制：MR1为51.0%，MR2为55.0%，MR4为58.0%；MR12为56.0%；MR21为61.1%，MR24为65.5%，MR42为66.0%。针对两条不同途径的双特异性寡核苷酸MR24和MR42最为有效。一项完整的LNCaP实验也在存在LD50环磷酰胺的情况下比较了同一系列的寡核苷酸。每种寡核苷酸与环磷酰胺联合时均显著抑制：MR1为49.0%，MR2为50.0%，MR4为53.0%；MR12为52.0%；MR21为58.6%，MR24为53.9%，MR42为58.0%。
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