Tumor-draining lymph nodes (LN) are the first site of metastasis in most types of cancer. The extent of metastasis in the LN is often used in staging cancer progression. Notably, in recent work the investigators described novel nanoscale TRAIL-coated liposomes that when conjugated to human natural killer (NK) cells enhance their endogenous therapeutic potential in killing cancer cells both in vitro and in vivo. In this project, these liposomes will e targeted to the LN by conjugating them to NK cells, and their ability to prevent the lymphatic spread of colon cancer tumors investigated in mice. It will be shown that targeting NK cells with TRAIL liposomes can enhance liposome retention time within the tumor draining lymph nodes to induce apoptosis in cancer cells. The proposed work is organized into three Specific Aims.
Specific Aim 1 : To optimize the TRAIL/anti-NK1.1 liposome formulation and investigate their ability to target lymph node metastases. Liposomes will be produced with varying diameters and PEG chain lengths. The ability of super NK cells to induce apoptosis in colon cancer cells will be tested in an in vitro mimic of the lymph node, measuring cellular apoptosis as a function of effector cell to target cell ratio. The 9 different liposome formulations (3 liposome diameters X 3 PEG chain lengths) will be injected subcutaneously in the left and right abdominal flanks of C57BL/6 mice. Inguinal lymph node cells will be harvested and analyzed via flow cytometry to assess human TRAIL presentation on the surface of NK cells. Subcutaneous colon carcinoma tumors will be grown in B6Rag1 mice, and treated with repeated subcutaneous injections of liposome solution, to identify the most effective formulations for metastasis elimination.
Specific Aim 2 : To characterize the biodistribution, pharmacokinetics and toxicity of the TRAIL/anti-NK1.1 liposome formulations introduced intraperitoneally. Intraperitoneal route of liposome injection will be examined, to enable efficacy studies in the orthotopic colon cancer model of Aim 3. The relationship between liposome properties, biodistribution and pharmacokinetics following intraperitoneal injection is less well understood than either IV or subcutaneous injection, and will be addressed for different liposome formulations. An examination of liver, lymph node, and systemic toxicity in response to repeated liposome treatment will be conducted.
Specific Aim 3 : To evaluate TRAIL/anti-NK1.1 liposome efficacy in an orthotopic model of colon cancer metastasis to the mesenteric lymph nodes. Human colorectal cancer cells will be implanted into the cecal wall of the mouse colon, with orthotopic tumor and LN metastasis monitored noninvasively using bioluminescence imaging. Mice will be treated with repeated intraperitoneal injections of TRAIL/anti-NK1.1 liposomes to determine whether this targeted therapy can reduce or eliminate the formation of mesenteric LN metastases. The work proposed here testing this platform in in vitro culture, a 3D engineered platform and animal models is critical to the translation of this novel technology as a treatment for metastasis.
The majority of cancers metastasize through the lymphatic system and LN metastasis is a negative prognostic factor in many cancers. In this study, it is proposed to demonstrate the ability of TRAIL liposomes conjugated to natural killer cells which deliver them to the tumor-draining lymph nodes to prevent the metastasis of a primary tumor using human xenograft models. By targeting TRAIL liposomes to NK cells, the therapeutic potential of the protein is enhanced by presenting TRAIL in its natural form, capitalizing on high uptake in the tumor-draining lymph nodes, the ability to present the therapeutic substance to cancer cells in its natural form, and low toxicity to local lymph node tissue.
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