Research in Focus: Nanomedicine & Drug Delivery

ISSUE SIX: NOVEMBER 2014

RESEARCH IN

FOCUS Nanomedicine & Drug Delivery Dr. Shyh-Dar Li

THE LABORATORY OF NANOMEDICINE AND DRUG DELIVERY Lab Members Shyh-Dar Li, Principal Investigator Weihsu Claire Chen, Research Associate Jonathan May, Research Associate Anirrudha (Ani) Roy, Postdoctoral Fellow Wei-Lun Tang, Graduate Student Yucheng (Rita) Zhao, Graduate Student INTRODUCTION Compounds need to possess balanced hydrophilicity and hydrophobicity to be efficiently absorbed from oral dosing or safely administered by i.v. injection. Unfortunately, the majority of the potent anticancer compounds in the development pipeline suffer from limited water solubility, and using conventional pharmaceutical formulations to improve the solubility is challenging, resulting in termination of many potent compounds in early development. Moreover, these highly cytotoxic agents often lack selectivity in drug biodistribution and cell killing, inducing severe side effects.

In the drug delivery field, nanomedicine is often referred to as assembling a drug into a pharmaceutical vehicle that is in the nanometer size range (10-200 nm) for improved physiochemical properties (i.e. increased solubility, stability and dissolution), pharmacokinetics, or tissue selectivity (i.e. inflamed tissue and cancer). These nanoparticles encompass liposomes, micelles, polymeric nanoparticles, dendrimers, and macromolecules. Nanomedicine is particularly attractive for tumor diagnosis and therapy owing to its unique feature termed as enhanced permeability and retention (EPR) effect: nanoparticles selectively extravasate into a tumor through its leaky vasculature, while sparing the normal tissues with a rigid capillary structure. Small molecule drugs, on the other hand, permeate into both normal and cancerous tissues with no selectivity, inducing significant side effects. Additionally, tumor targeted drug delivery can be enhanced by conjugating a targeting ligand (i.e. antibody) onto nanoparticles that recognizes a surface receptor on tumor cells, resulting in improved efficacy. The tremendous promise of nanomedicine in non-invasive tumor imaging, early detection and 2 路 RESEARCH IN FOCUS

drug delivery is evidenced with many products in clinical use and trials (Clinical Pharmacology & Therapeutics 2014).

RESEARCH SUMMARY Although nanoparticles can be employed to improve water solubility and tissue selectivity of anticancer compounds, there are several biological barriers to their delivery (Fig 1). Nanoparticles injected intravenously must evade reticuloendothelial (RES) and renal clearance, and remain stable in plasma during systemic circulation, such that a sufficient dose of the nanoparticle and drug can interact with tumor physiology.

Once particles successfully extravasate into the tumor compartment, the particles must travel through the stroma against high interstitial fluid pressure (IFP) gradients, and ultimately interact with the target cells or release the drug payload for pharmacological effect. Therefore, nanoparticle formulations must be carefully designed to overcome these barriers to achieve significant therapeutic activity.

Fig 1: The three phases of drug delivery by nanoparticles.

The Li lab focuses on developing innovative lipid- and polymer-based drug delivery technologies to enhance cancer therapy. The team is particularly interested in how to optimize the nanoparticle delivery systems to better tackle these three major delivery obstacles. The RESEARCH IN FOCUS 路 3

knowledge gain from the research is expected to contribute to enhanced chemotherapy delivery and improved clinical care of cancer patients.

SELECTED PROJECTS The Laboratory of Nanomedicine and Drug Delivery has developed two proprietary drug delivery technologies: a polymeric conjugate technology (NanoCMC) for tumor-targeted delivery of water insoluble drugs, and a thermosensitive liposome technology (HaT) for image-guided local delivery of cytotoxic drugs.

NanoCMC Drug Delivery Platform Many nanoparticle systems formulate hydrophobic drugs via passive encapsulation. These nanoparticles often release the drugs rapidly due to efficient partition of the drugs to circulating plasma proteins, resulting in impaired pharmacokinetics and drug delivery. The NanoCMC polymer is composed of a hydrophobic drug and PEG conjugated via ester linkages to acetylated carboxymethylcellulose (Fig 2). The drug is carried by the polymer via a covalent bond, and therefore, the drug release is controlled, leading to improved pharmacokinetics and drug targeting. The docetaxel (DTX)-containing lead nanoparticle, Cellax, has demonstrated superior safety and efficacy compared to approved taxanes, including Taxotere® (native DTX) and Abraxane® (albumin-bound paclitaxel, the only FDAapproved taxane nanoparticle).

Fig 2: Schematic of NanoCMC. When the drug-to-PEG ratio is balanced, the polymer condenses into a nanoparticle in aqueous systems. A TEM image of a typical NanoCMC particle is depicted (~120 nm in diameter).

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Recently, the team discovered that Cellax targets cancer-associated fibroblasts and modulate the tumor microenvironment by improving the perfusion and reducing the IFP, leading to decreased metastasis in multiple breast and pancreatic tumor models (Fig 3).

Fig 3: Anti-metastatic effect of native docetaxel (DTX), nabpaclitaxel (nab-PTX, Abraxane), and Cellax against the 4T1 breast tumor.

This platform technology can be applied to deliver other water insoluble drugs. The most recent example is podophyllotoxin, which is a topically used anti-tubulin agent. The drug can be formulated into ultra-small nanoparticles (20 nm) by the NanoCMC technology, and then given intravenously to regress multi-drug resistant tumors with no significant side effects.

HaT (Heat-activated cytoToxic ) Liposomal Drug Delivery Platform Long-circulating liposomes extravasate to tumor tissues via the leaky vasculature, but often do not penetrate tumor well. The drug is stably incorporated inside the liposome with a low release rate (<2% per day).

These factors lead to heterogeneous drug exposure within the tumor and sometimes limited bioavailability. Thermosensitive liposomes function to concentrate the drug in the blood circulation by minimizing renal clearance and drug uptake and metabolism in other compartments, and rapidly release the drug in the vasculature of a heated (40â&#x20AC;&#x201C;42 °C) tumor, where the high drug concentration gradient forces released drug into the tumor. In this way, the bioavailable drug taken up by the tumor is significantly enhanced compared to a long-circulating liposome approach RESEARCH IN FOCUS ¡ 5

(Fig 4). The thermosensitive formulation is administered in combination with the hyperthermia treatment, causing immediate release of the encapsulated drug within the heated tumor. Therefore, this approach is not dependent on the EPR effect for tumor targeting, a process which can be less efficient and tumor-dependent.

Fig 4: Comparison of the drug delivery approaches of the long circulating nanoparticles (A) and the thermosensitive nanoparticles (B).

The research team has created a new thermosensitive liposomal formulation (HaT) and demonstrated that HaT-liposomes release a concentrated burst of drug when heated to 40-42°C, but is stable at 37-38°C. HaT-liposomes contain only 2 components, dipalmitoyl phosphatidyl choline (DPPC) and Brij78, and they display increased drug release rate constants for many drugs at 40-41°C compared to the lyso-lipid temperature-sensitive liposome (LTSL, same lipid formulation as ThermoDOX: DPPC/MSPC/DSPE-PEG2000 in clinical trials). HaTliposomes improve the delivery of gemcitabine (GEM) into heated tumors (43°C) by 25-fold and 7-fold compared to free GEM and LTSL-GEM, respectively, thus inducing complete tumor remission after a single treatment (Fig 5).

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Fig 5: Antitumor efficacy of GEM in different formulations in combination with localized hyperthermia.

This thermosensitive formulation for GEM is currently being tested for treating locally advanced pancreatic tumors in animal models. Localized pancreatic tumors will be identified by MR imaging, followed by MR-guided focused ultrasound to locally heat the tumor at 42°C to trigger localized release of GEM from HaT-liposomes.

OTHER PROJECTS The team is interested in developing an improved active loading method to stably incorporate water insoluble drugs into lipid nanoparticles and has obtained encouraging results. They expect that this platform technology can be employed to deliver many natural products and enable them to be used as new therapeutics for many diseases. The group is also working on a new prodrug technology to enhance drug delivery to the brain, and will be collaborating with the UBC neurological team to develop new therapeutics for neurological diseases, such as Alzheimer’s disease, Parkinson’s disease and Huntington’s disease.

FUNDING SUPPORTS CIHR, NIH, Prostate Cancer Foundation, MaRS Innovation, Ontario Centres of Excellence and Ontario Institute for Cancer Research.

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RELEVANT PUBLICATIONS Docetaxel-carboxymethylcellulose nanoparticles display enhanced anti-tumor activity in murine models of castration-resistant prostate cancer.Hoang B, Ernsting MJ, Murakami M, Undzys E, Li SD. Int J Pharm. 2014 Aug 25;471(12):224-33.

Carboxymethylcellulose-based and docetaxel-loaded nanoparticles circumvent P-glycoprotein-mediated multidrug resistance. Roy A, Murakami M, Ernsting MJ, Hoang B, Undzys E, Li SD. Mol Pharm. 2014 Aug 4;11(8):25929.

Thermosensitive liposomes for the delivery of gemcitabine and oxaliplatin to tumors. May JP, Ernsting MJ, Undzys E, Li SD. Mol Pharm. 2013 Dec 2;10(12):4499-508.

Docetaxel conjugate nanoparticles that target a-smooth muscle actinexpressing stromal cells suppress breast cancer metastasis. Murakami M, Ernsting MJ, Undzys E, Holwell N, Foltz WD, Li SD. Cancer Res. 2013 Aug 1;73(15):4862-71.

A docetaxel-carboxymethylcellulose nanoparticle outperforms the approved taxane nanoformulation, Abraxane, in mouse tumor models with significant control of metastases. Ernsting MJ, Murakami M, Undzys E, Aman A, Press B, Li SD. J Control Release. 2012 Sep 28;162(3):575-81.

A thermosensitive liposome prepared with a Cu虏+ gradient demonstrates improved pharmacokinetics, drug delivery and antitumor efficacy. Tagami T, May JP, Ernsting MJ, Li SD. J Control Release. 2012 Jul 10;161(1):142-9.

Tumor-targeted drug delivery using MR-contrasted docetaxel carboxymethylcellulose nanoparticles. Ernsting MJ, Foltz WD, Undzys E, Tagami T, Li SD. Biomaterials. 2012 May;33(15):3931-41.

Preclinical pharmacokinetic, biodistribution, and anti-cancer efficacy studies of a docetaxel-carboxymethylcellulose nanoparticle in mouse models. Ernsting MJ, Tang WL, MacCallum NW, Li SD. Biomaterials. 2012 Feb;33(5):1445-54.

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Synthetic modification of carboxymethylcellulose and use thereof to prepare a nanoparticle forming conjugate of docetaxel for enhanced cytotoxicity against cancer cells. Ernsting MJ, Tang WL, MacCallum N, Li SD. Bioconjug Chem. 2011 Dec 21;22(12):2474-86.

MRI monitoring of intratumoral drug delivery and prediction of the therapeutic effect with a multifunctional thermosensitive liposome. Tagami T, Foltz WD, Ernsting MJ, Lee CM, Tannock IF, May JP, Li SD. Biomaterials.

2011 Sep;32(27):6570-8.

Optimization of a novel and improved thermosensitive liposome formulated with DPPC and a Brij surfactant using a robust in vitro system.Tagami T, Ernsting MJ, Li SD. J Control Release. 2011 Sep 25;154(3):290-7.

Efficient tumor regression by a single and low dose treatment with a novel and enhanced formulation of thermosensitive liposomal doxorubicin. Tagami T, Ernsting MJ, Li SD. J Control Release. 2011 Jun 10;152(2):303-9.

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