LPD Nanoparticles in Anti-Cancer Therapy

描述（由申请人提供）：我们最近开发了一种氨酰胺靶向纳米颗粒制剂，称为LPD，用于在胸腺裸鼠模型中静脉给药siRNA到人肺癌细胞NCI-H460（表达sigma受体）。siRNA被设计用来沉默表皮生长因子受体（EGFR）。由于肿瘤的高水平摄取（每g组织注射剂量为70-80%），EGFR在整个肿瘤中完全沉默。尽管siRNA的高效递送和靶癌基因的沉默，但肿瘤细胞在体内仅表现出部分（~15%）凋亡和部分生长抑制。因此，该项目将开发几个独立的策略来增强LPD介导的治疗活性。在目标1中，我们将封装针对不同细胞癌基因的多组siRNA，以显示至少在肿瘤杀伤方面具有添加性（如果不是协同性）效果。我们还将在模型中测试Let-7 miRNA的抑瘤活性。化学修饰的siRNA，包括含有硼磷酸盐的siRNA，也将测试其长期沉默效果。我们发现阳离子脂质DOTAP是LPD制剂的主要成分，具有抗凋亡活性，可保护肿瘤细胞免于死亡。我们发现了另一种阳离子脂质，它本身就能杀死肿瘤细胞，可以用来代替siRNA制剂中的DOTAP。初步数据表明，茴香胺靶向LPD也可以将siRNA传递到小鼠黑色素瘤B16F10细胞的肺转移中。在目标2中，我们将传递不同的siRNA来诱导小鼠黑色素瘤细胞凋亡。我们还将测试一个剪接移位寡核苷酸，它将抗凋亡的Bcl-xL转化为凋亡的Bcl-xS，从而诱导肿瘤细胞死亡。由于该传递系统还可以与siRNA一起传递质粒DNA，因此可以使用不同的抗癌基因与siRNA进行联合治疗。在目标3中，我们将采用另一种纳米颗粒制剂，即LPH，它含有硫酸肝素来递送强效抗癌药物阿霉素（dox），既可以单独递送，也可以与EGFR siRNA一起递送。由于dox和EGFR沉默通过不同的机制诱导细胞凋亡，我们预计至少是一种加法效应，如果不是协同效应的话。Dox将化学偶联到硫酸肝素并在LPH中共配制。该项目的目标是为癌症治疗开发有效的抗肿瘤纳米颗粒药物。公共卫生相关性：该项目的目标是开发用于癌症治疗的siRNA的肿瘤特异性全身递送系统。一种自组装的纳米粒子制剂将被用作递送载体。该项目以小鼠肺癌和肺转移为疾病模型。

DESCRIPTION (provided by applicant): We have recently developed an anisamide-targeted nanoparticle formulation, called LPD, for intravenous delivery of siRNA to human lung cancer cells NCI-H460 (expressing the sigma receptor) in an athymic nude mouse model. The siRNA was designed to silence the epidermal growth factor receptor (EGFR). As the result of a high level of uptake by the tumor (70-80% injected dose per g of tissue), the EGFR was completely silenced through out the entire tumor. Despite the high efficiency delivery of the siRNA and silencing of the target oncogene, the tumor cells only showed a partial (~15%) apoptosis and partial growth inhibition in vivo. Thus, the project will develop several independent strategies to enhance the therapeutic activity mediated by LPD. In aim 1, we will encapsulate multiple sets of siRNA targeting to different cellular oncogenes to show at least an additive, if not synergistic, effect in tumor killing. Let-7 miRNA will also be tested for its tumor suppression activity in the model. Chemically modified siRNA including those containing boranophosphate will also be tested for prolonged silencing effect. We have discovered that cationic lipid DOTAP, which is a major component in the LPD formulation, shows an anti-apoptotic activity which protects the tumor cells from death. We have discovered another cationic lipid which by itself kills tumor cells and can be used to replace DOTAP in the siRNA formulation. Preliminary data indicate that the anisamide targeted LPD can also deliver siRNA to the pulmonary metastasis of mouse melanoma B16F10 cells. In aim 2, we will deliver different siRNA to induce apoptosis of the melanoma cells in mice. We will also test a splice shifting oligo which will convert the anti-apoptotic Bcl-xL to the apoptotic Bcl-xS to induce tumor cell death. Since the delivery system can also deliver a plasmid DNA along with siRNA, different anti-cancer genes can be used in a combination therapy with siRNA. In aim 3, we will employ another nanoparticle formulation, i.e., LPH, which contains heparin sulfate to deliver doxorubicin (dox), a potent anti-cancer drug, either by itself or together with EGFR siRNA. Since dox and EGFR silencing induce apoptosis by different mechanisms, we expect at least an additive, if not synergistic, effect. Dox will be chemically conjugated to heparin sulfate and co-formulated in LPH. The goal of the project is to develop efficient anti-tumor nanoparticle agents for cancer therapy. PUBLIC HEALTH RELEVANCE: The goal of the project is to develop a tumor specific, systemic delivery system for siRNA for cancer therapy. A self-assembled nanoparticle formulation will be used as a delivery vehicle. The project uses lung cancer and lung metastasis in mice as the disease model.