The Projects
To help meet it's goals, the NanoTumor Center will focus on six projects, tracked within six categories, outlined in the timeline below. Following the timeline are detailed descriptions, functions and goals of each project.
Project 1 - Nanoparticles In vivo: Interactions with Cells and Tissues
Leader: Erkki Ruoslahti
A main goal of the proposed CCNE is to develop nanosystems that specifically seek out tumor tissue for the purposes of diagnosing, monitoring and treating tumors. Each of these applications requires selective delivery of nanoparticles into tumors and/or selective detection/activity in tumors. Particles capable of specifically entering into parenchymal or stromal tumor cells provide further advantages. In addition, particles designed for certain in vivo diagnostic purposes should be able to exit from tumor cells they have entered, allowing for send and return type approaches. This project (Project 1) focuses on the development of homing peptide-nanoparticle constructs that optimize tumor homing while avoiding non-specific accumulation in non-target tissues and minimizing toxicity. A large collection of peptides and other compounds capable of selectively delivering nanoparticles to tumor vasculature is on hand. The biggest single obstacle to be overcome in optimizing the tumor homing of such particles is the ability of the reticuloendothelial system (RES) and macrophages to eliminate particles from the systemic circulation. The ability of the peptides to effect passage of the particles into the extravascular space, and internalization of the particles and their payload into tumor cells and tumor endothelial cells are also major determinants in targeting efficacy of nanoparticles. We propose to develop new technology to deal with the liver uptake and extravasation issues. A team of tumor biologists, peptide chemists, materials scientists, and engineers endeavor to achieve these goals by identifying and characterizing peptides from combinatorial libraries that promote the binding and intracellular entry in tumors and discourage it elsewhere in the body. We expect this work in Project 1 to provide the basis for rational design of nanoparticle systems with greatly improved in vivo properties for effective tumor targeting due to diminished liver uptake and ability to travel through barriers such as cell layers and membranes. The results from this Project will enable the design of in vivo routing systems for multifunctional nanodevices. The proposed use of in vivo discovery methods and early in vivo testing and optimization of the routing systems will accelerate the construction of devices that are likely to be clinically useful.
These peptides, and the existing collection of homing peptides that recognize specific receptors in tumor vessels and/or on tumor cells, will be available to the other Projects in this CCNE. The validity of the peptides as nanoparticle delivery vehicles and applicability of the results to human cells and tissues will be validated in conjunction with Project 2 (Targeted In vivo micro Platform for nano devices) and Project 6 (Tumor Therapy/Annihilation using a Smart nanoplatform) of this CCNE.
Project 2 - Targeted In vivo micro Platform for Nano Devices
Leaders: Mike Sailor and Sadik Esener
A group of investigators with complementary expertise has been assembled to construct micron-sized devices ("mother ships") that are equipped with nano-sized features and that are capable of homing into cancerous tumors in vivo and performing various tasks in the tumor. These tasks include detecting, identifying, and imaging a tumor, performing measurements on it, and delivering therapies. The system will be based on micron-sized, nanostructured particles made from various soft or hard composites. Mother ship delivery of nanoparticles concept was chosen for a number of reasons: (1) Preliminary results suggest that micron-sized particles coated with tumor-specific vascular homing peptides become lodged in tumor vasculature; (2) Micron-sized particles, unlike nanoparticles, are not taken up by the reticuloendothelial system; (3) The mother ship can be functionalized with a wide range of targeting moieties, nanoparticles, drugs, and/or imaging agents in a controlled fashion; (4) The micron-range size makes it possible, for example, to use active electronic materials such as porous silicon, incorporating unique engineering features such as optical and radio frequency reporting properties, high-capacity nano-porous carrier structure, biocompatibility and bioresorbability; (5) This applicant group has extensive experience with nanostructured microdevices, and prototype mother ship particles resembling red blood cells have already been constructed and tested in vivo.
Project 2 relies on project 1 for targeting, on Cores B, C, and D to determine toxic effects, imaging attributes and fabrication, and will develop unique in vivo drug and nanosensor delivery techniques that will be used in projects 5 and 6.
Project 3 - Tumor Activated Amplification System
Leaders: Roger Tsien and Patrick Daugherty
The main goal of this project is to develop generic chemistries by which very small nanoparticles (<8 nm diameter or 10-200 KDa in molecular weight) will selectively accumulate and aggregate within solid tumors in vivo. A corollary goal is to investigate supralinear contrast mechanisms for imaging and therapeutic reduction of such tumors. The basic approach is to devise nanoparticle coatings that are nonsticky in normal tissue but that become mutually adherent when exposed to proteases or growth factors specific to the tumor microenvironment, using three alternative, mutually nonexclusive strategies: 1) Bacterial surface display will be used to optimize mini-proteins that bind multimeric growth factors enriched in tumors only after activation by a tumor specific protease. Nanoparticles displaying such peptides should become homomerically crosslinked when they encounter both tumor specific proteases and growth factors. 2) Nanoparticles will be coated with protease substrate peptides and heteromeric aggregation will be triggered by tumor-specific proteases or bioreduction in hypoxic tissue. The receptor is vancomycin, which binds D-ala-D-ala with a free carboxy terminus, released from the cleaved peptide by a self-eliminating spacer. 3) The nanoparticles will display leucine zipper sequences that have been fused to protease substrate sequences and circularized, thus preventing alpha-helix formation necessary for homo- or heterodimerization. Exposure to tumor-specific proteases will cut the substrate sequence, linearize the peptides, and allow homo- or heterodimerization with partners on adjacent nanoparticles.
Once aggregation occurs, ideally the imaging or therapeutic contrast should scale more than linearly with the number of assembled nanoparticles. We hope to demonstrate such supralinear contrast for optical imaging and heating of gold nanoparticles, magnetic resonance imaging T2 relaxivity and hysteresis-induced hyperthermia due to iron oxide nanoparticles, or ultrasound scattering of perfluorocarbon-containing nanodroplets.
We therefore propose a new approach by which nanoparticles will be engineered to congregate preferentially within solid tumors rather than in normal tissue. The hope is to detect and kill tumors, especially metastases, at smaller and earlier stages than currently possible.
Project 4 - Ex vivo Sensors and Phenotypes for Cancer cells
Leader: Andrew Kummel
There are three phases of cancer at which there is a need for accurate and sensitive detection: early detection of pre-cancerous cell expansions or malignancy, assessment of variants within a tumor population, and the monitoring of tumor cells during and post therapy. We will develop ex vivo platforms to assess the presence of altered cells at each of these three phases. Two tumor systems will be addressed: Chronic Lymphocytic Leukemia (CLL) and breast cancer. For each there is extensive local clinical expertise, and the UCSD Cancer Center maintains large longitudinal patient sample collections and clinical information databases. The human patient resources are complemented by two mouse models actively under study. The Eu-TCL1 mouse is a model for CLL that recapitulates most of the essential features of this solid/liquid phase tumor. The second model is the HER-2/neu transgenic mouse that develops spontaneous mammary tumors similar to human breast cancer.
This project has two major aims, each with its own platform, and the two platforms are used in an integrated manner. (1) A pulsed dielectrophoretic microarray, which can concentrate cancer cells and DMA fragments from blood without tags will be developed. A unique electric field induced FRET technique is used to detect cancer serum DNA using nanoparticles. (2) A phenotype array detector with quantum dot imaging spectroscopy will be fabricated so that the concentrations of multiple receptors can be determined simultaneously on 60,000 cells. The same technique will be used to test cancer cell response to chemotherapy agent when the cancer cells are surrounded by stromal or other supporting cells. These platforms will be used in conjunction with projects 5 and 6.
Project 5 - Longitudinal Tumor Monitoring with Nanodevices
Leaders: Dennis Carson and Tom Kipps
The neoplastic process that results in incurable cancer is a series of deviations from the normal homeostatic system. The goal of this project is to integrate traditionally biologic measures of these deviations with nanotechnology through computational approaches. The central hypothesis and working model on which this project is based is that time series data contains patterns that, when identified by computational means, can be used for early detection, prognosis, and/or evaluation of the response to anti-cancer therapy. Two types of cancer will be used to address this hypothesis: Chronic Lymphocytic Leukemia (CLL) and breast cancer. For each there is extensive basic and clinical research expertise at the UCSD Cancer Center, which is home for large longitudinal sample collections and clinical information databases. Two mouse models actively under study complement the human patient resources. The first aim of this project is to validate a role for longitudinal measures accomplished with nanodevices in the prediction, prognosis, and/or treatment of tumors in these mouse models. Transgenic mice predisposed to develop breast cancer or leukemia will be assessed for a broad spectrum of tumor and host response indicators during the time of tumor development. The data from the mouse studies will feed into the second aim, which is to create computational models to integrate biologic data with data from nano-sensors. Finally, this project will leverage the information gained in the first two aims to validate ex vivo platforms for the monitoring of CLL and breast cancer, using our large clinical sample collections. This research will benefit public in two ways. Most directly, it will allow for better clinical evaluation of patients with breast cancer and CLL using new nanotechnologies. More generally, it will establish a new paradigm for detecting and monitoring cancer that allow for earlier, more focused treatments that are less debilitating and ultimately more effective than current anti-cancer therapies.
Project 6 - Tumor Therapy/Annihilation using a Smart Nanoplatform
Leader: David Cheresh
The advent of nanotechnology holds the promise of new modalities for the treatment of disease in man. In particular, the multifunctional nature of nanoplatforms is well-suited for complex diseases that involve several different cellular compartments, such as cancer. The overall objective of project 6 is to develop and test programmable, or "smart" nanoplatforms (SNaPs) that are based on common nanoplatform cores. Nanoplatforms have been designed which will undergo spontaneous self-assembly, based on hostguest chemical interactions. The assembly is dependent upon integration of polyethyleneglycol polymer (PEG)-conjugated molecular guests, where the distal terminus of the PEG polymer can be conjugated to "programmable" elements. This host-guest based nanoplatform provides several advantages over our existing platforms: the poor pharmacological properties associated with many potent drugs are actually exploited in this approach, with fewer expected side effects, the design is highly flexible, accommodating multiple therapeutic, imaging, targeting or other effector functions within each nanoplatform, and the nanoplatforms are easily and rapidly programmable by-simple coincubation of the host and guest moieties.
The capacity to incorporate multiple targeting elements into the SNaPs will be used to evaluate whether low affinity/high avidity modes of targeting represent an improvement in target cognition over current high affinity/low avidity approaches. We hypothesize that moderate-to-low affinity, high-avidity dependent interactions offer increased opportunities for true "recognition" of the platform target, particularly if heterogenous recognition of several different "sensor-ligands" is required. Finally, we will test the capacity of the programmable nanoplatform to target distinct cell populations that contribute to tumor growth and malignancy, including both vascular and tumor elements, using syngeneic and transgenic models of disease. A focus of these investigations will be the efficacy of the nanoplatform, when used to eradicate residual and metastatic disease, after ablation of the primary tumor. The capacity for SNaPs to hunt and kill residual disease is a potential strength of the nanoplatform, as recurring and metastatic disease accounts for the majority of disease morbidity. These studies will lay the groundwork for a new generation of easily programmed, multifunctional nanoplatforms, amenable to the treatment of malignancy in human patients.