Translational Development of a Targeted and Stroma-breaking Nanoparticle Drug for Pancreatic Cancer Therapy

项目摘要 耐药是胰腺癌治疗面临的主要临床挑战。药物输送效率低 和侵袭性的肿瘤生物学是治疗反应差的主要原因。众所周知， 致密的纤维性肿瘤间质的存在创造了药物传递障碍，并促进了侵袭性的生物学和 胰腺癌细胞的耐药性。我们翻译研究项目的目标是开发 新一代受体靶向和蛋白水解酶活性靶向配体与基质破坏 用于晚期胰腺癌有效治疗的纳米粒给药平台。我们广泛的 研究开发了一种使用重组uPAR靶向配体的新的肿瘤靶向方法 含有uPA的氨基末端片段(ATF)与基质金属蛋白酶的催化结构域融合- 14(ATFmmp14)。ATFMMP14配体的新颖设计不仅确保了增强的肿瘤内递送 治疗药物，但也能打破肿瘤间质细胞和细胞外基质障碍。我们也 开发了一种携带伊立替康活性代谢物SN38的可生物降解的透明质酸纳米粒 (HANP/SN38)。结果表明，ATFMMP14偶联的HANP/SN38对药物有显著的增强作用 KRAS驱动的转基因小鼠和人胰腺PDX肿瘤模型中的肿瘤递送。重要的是， 纳米药物从肿瘤间质中迁移出来并穿透基底膜衬里导管 癌细胞进入肿瘤细胞。结果，ATFMMP14-HANP/SN38的系统递送显著受到抑制 肿瘤生长。与目前的联合疗法(FOLFRINOX或吉西他滨-NAB-紫杉醇)相比， ATFMMP14-HANP/SN38治疗显着延长药物源性PDX肿瘤模型的生存时间 耐药的胰腺癌患者。为了将这一新方法转化为癌症患者的靶向治疗，在这篇文章中 第一阶段研究，我们将开发新一代无标记人类ATFmmp14配体，这些配体针对 具有较高的结合亲和力、靶标特异性和MMP14酶活性。建议数 AIM 1的研究将设计出稳定表达CHO细胞的ATFMMP14(R2)和可分泌的枯草杆菌 大规模生产无标签ATFMMP14(R2)的蛋白表达系统。然后我们将评估和 比较它们的生产程序、蛋白质产量、靶标特异性和体内外MMP14活性 选择最适合药物开发的靶向配体(目标2)。最后，还观察了其疗效。 ATFMMP14(R2)将在胰腺PDX肿瘤模型中确定(AIM 3)。这一阶段的里程碑是 研究项目如下：1)开发ATFMMP14-HANP(R2)作为一种靶向基质破碎性纳米药物 为未来商业化提供平台；以及2)开发靶向纳米药物ATFMMP14(R2)- HANP/SN38，用于旨在实现临床前研究的研究新药(IND)的第二阶段研究 ATFMMP14-HANP/SN38靶向治疗晚期胰腺癌的研究进展

Project Summary Resistance to therapy is the major clinical challenge in pancreatic cancer therapy. Low efficiency in drug delivery and aggressive tumor biology are the major causes of a poor therapeutic response. It is well known that the presence of a dense fibrous tumor stroma creates a drug delivery barrier and promotes aggressive biology and drug resistance in pancreatic cancer cells. The objective of our translational research project is to develop a new generation of receptor-targeted and protease active targeting ligand and stroma-breaking nanoparticle drug delivery platform for effective treatment of advanced pancreatic cancer. Our extensive research led to the development of a new tumor targeting approach using a recombinant uPAR targeting ligand containing the amino terminal fragment (ATF) of uPA fused with the catalytic domain of matrix metalloproteinase- 14 (ATFmmp14). Novel designs of ATFmmp14 ligand not only ensure enhanced intratumoral delivery of therapeutic agents, but also enable breaking tumor stroma cellular and extracellular matrix barriers. We also developed a biodegradable hyaluronic acid nanoparticle carrying an active metabolite of irinotecan, SN38 (HANP/SN38). Our results demonstrated that ATFmmp14 conjugated HANP/SN38 significantly enhanced drug delivery to tumors in KRAS-driven transgenic mouse and human pancreatic PDX tumor models. Importantly, the nanoparticle-drug migrated out of tumor stroma and penetrated through the basement membrane lining ductal cancer cells to enter tumor cells. As a result, systemic delivery of ATFmmp14-HANP/SN38 significantly inhibited tumor growth. In comparison with current combination therapies (FOLFRINOX or gemcitabine-Nab-Paclitaxel), ATFmmp14-HANP/SN38 treatment significantly prolonged survival in the PDX tumor models derived from drug resistant pancreatic cancer patients. To translate this novel approach for target therapy in cancer patients, in this phase 1 study, we will develop a new generation of tagless human ATFmmp14 ligands that are optimized for the GMP production and have high binding affinity, target specificity and MMP14 enzymatic activity. The proposed studies in Aim 1 will engineer a stable ATFmmp14(R2) expressing CHO cell line and a B. subtilis secretable protein expression system for large scale production of tagless ATFmmp14(R2). We will then evaluate and compare their production procedure, protein yield, target specificity, and MMP14 activity in vitro and in vivo to select the most suitable targeting ligand for drug development (Aim 2). Finally, therapeutic effect of ATFmmp14(R2) will be determined in a pancreatic PDX tumor model (Aim 3). The milestones of this phase I research project are: 1) to develop ATFmmp14-HANP(R2) as a stroma-breaking and targeted nanoparticle drug delivery platform for future commercialization; and 2) to develop a targeted nanoparticle drug, ATFmmp14(R2)- HANP/SN38, for the Phase II study aimed at the Investigational New Drug (IND)-enabling preclinical studies for the translational development of ATFmmp14-HANP/SN38 for targeted therapy of advanced pancreatic cancer.