Hyperthermia-triggered intracellular delivery of anticancer agent to HER2(+) cells by HER2-specific affibody (ZHER2-GS-Cys)-conjugated thermosensitive liposomes (HER2(+) affisomes).
Hyperthermia-triggered intracellular delivery of anticancer agent to HER2(+) cells by HER2-specific affibody (ZHER2-GS-Cys)-conjugated thermosensitive liposomes (HER2(+) affisomes).
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DOI：
10.1016/j.jconrel.2011.04.005
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发表时间：
2011-07-30
期刊：
影响因子：
--
通讯作者：
Puri A
中科院分区：
文献类型：
--
作者：
Smith B;Lyakhov I;Loomis K;Needle D;Baxa U;Yavlovich A;Capala J;Blumenthal R;Puri A
我们先前报道了HER 2（人表皮生长因子受体2）特异性亲和体（ZHER 2：342-Cys）缀合的热敏脂质体（HER 2 + Affisomes）的制剂和物理性质。在这里，我们研究了本地化的交付潜力，这些Affisomes通过监测细胞相互作用，细胞内摄取，高血压诱导的药物输送的影响。我们通过在C-末端半胱氨酸之前引入甘氨酸-丝氨酸间隔区（称为ZHER 2-GS-Cys）来修饰ZHER 2：342-Cys，以实现对细胞表面表达的HER 2的可接近性。这种修饰不影响HER 2特异性结合，并且ZHER 2-GS-Cys保留其与含有二棕榈酰磷脂酰胆碱：DSPE-PEG 2000-马来酰亚胺，96：04摩尔比（HER 2+亲和体）的脂质体缀合的能力。HER 2+亲和体（i）用罗丹明-PE和钙黄绿素荧光标记或（ii）负载抗癌药物阿霉素（DOX）。在37°C下，与HER 2 −细胞（MDA-MB-468）相比，荧光标记的HER 2+亲和体显示与HER 2+细胞（SK-BR-3）的结合至少增加10倍。使用游离ZHER 2-GS-Cys的竞争实验阻断了HER 2 + Affisomes-SK-BR-3细胞缔合。共聚焦显微镜成像显示，HER 2 + Affisomes在37°C下积聚在SK-BR-3细胞的胞质溶胶中。高血压诱导的细胞内释放实验显示，用45°C（±1°C）预平衡缓冲液处理HER 2+亲和体/SK-BR-3细胞复合物导致钙黄绿素的胞质递送。在45°C下在20分钟内观察到大量钙黄绿素释放，在这些条件下对细胞活力没有影响。类似地，与对照脂质体相比，负载DOX的HER 2+亲和体显示出在SK-BR-3细胞中至少2-3倍更高的DOX积累。DOX介导的细胞毒性在SK-BR-3细胞中更明显，特别是在较低剂量的HER 2+亲和体下。在用于细胞杀伤测定的孵育开始之前，脂质体-细胞复合物在45°C下的短暂暴露导致Affisomes和对照脂质体的细胞毒性增强。然而，Doxil（市售脂质体制剂）在相同条件下显示出显著更低的毒性。因此，我们的数据表明，HER 2 + Affisomes包括靶向和触发潜力，因此可以证明是可行的纳米药物递送载体用于乳腺癌治疗。
We previously reported the formulation and physical properties of HER2 (Human Epidermal Growth Factor Receptor 2)-specific Affibody (ZHER2:342-Cys) conjugated thermosensitive liposomes (HER2+ Affisomes). Here we examined localized delivery potential of these Affisomes by monitoring cellular interactions, intracellular uptake, and hyperthermia-induced effects on drug delivery. We modified ZHER2:342-Cys by introducing a glycine-serine spacer before the C-terminus cysteine (called ZHER2-GS-Cys) to achieve accessibility to cell-surface expressed HER2. This modification did not affect HER2-specific binding and ZHER2-GS-Cys retained its ability to conjugate to the liposomes containing dipalmitoyl phosphatidyl choline: DSPE-PEG2000-Malemide, 96:04 mole ratios (HER2+ Affisomes). HER2+ Affisomes were either (i) fluorescently labeled with rhodamine-PE and calcein or (ii) loaded with an anticancer drug Doxorubicin (DOX). Fluorescently labeled HER2+ Affisomes showed at least 10 fold increase in binding to HER2+ cells (SK-BR-3) when compared to HER2− cells (MDA-MB-468) at 37°C. A competition experiment using free ZHER2-GS-Cys blocked HER2+ Affisomes-SK-BR-3 cell associations. Imaging with confocal microscopy showed that HER2+ Affisomes accumulated in the cytosol of SK-BR-3 cells at 37°C. Hyperthermia-induced intracellular release experiments showed that the treatment of HER2+ Affisome/SK-BR-3 cell complexes with a 45°C (±1°C) pre-equilibrated buffer resulted in cytosolic delivery of calcein. Substantial calcein release was observed within 20 minutes at 45°C, with no effect on cell viability under these conditions. Similarly, DOX-loaded HER2+ Affisomes showed at least 2–3 fold higher accumulation of DOX in SK-BR-3 cells as compared to control liposomes. DOX-mediated cytotoxicity was more pronounced in SK-BR-3 cells especially at lower doses of HER2+ Affisomes. Brief exposure of liposome-cell complexes at 45°C prior to the onset of incubations for cell killing assays resulted in enhanced cytotoxicity for Affisomes and control liposomes. However, Doxil (a commercially available liposome formulation) showed significantly lower toxicity under identical conditions. Therefore, our data demonstrate that HER2+ Affisomes encompass both targeting and triggering potential and hence may prove to be viable nano drug delivery carriers for breast cancer treatment.
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DOI：
10.1615/critrevtherdrugcarriersyst.v26.i6.10
发表时间：
2009
影响因子：
2.7
作者：
Puri A;Loomis K;Smith B;Lee JH;Yavlovich A;Heldman E;Blumenthal R
通讯作者：
Blumenthal R
影响因子：
3.4
作者：
O'Donnell, Robert T.;Martin, Shiloh M.;Tuscano, Joseph M.
通讯作者：
Tuscano, Joseph M.
影响因子：
4.7
作者：
Beuttler, Julia;Rothdiener, Miriam;Kontermann, Roland E.
通讯作者：
Kontermann, Roland E.
影响因子：
56.9
作者：
YATVIN, MB;WEINSTEIN, JN;BLUMENTHAL, R
通讯作者：
BLUMENTHAL, R
影响因子：
10.8
作者：
Negussie, Ayele H.;Miller, Jenna L.;Reddy, Goutham;Drake, Steven K.;Wood, Bradford J.;Dreher, Matthew R.
通讯作者：
Dreher, Matthew R.