Summary:

Many chemotherapeutic anticancer agents target vital components of cell division and inhibit survival and growth of cancer cells. Anti-tubulin agents for example, target the microtubule complex formed during mitosis and result in cell apoptosis. While, chemotherapy can be effective initially, relapse due to drug resistance is a frequent occurrence due to the multidrug resistance (MDR) phenomenon. MDR is associated with decreased drug uptake, increased drug efflux and other cellular mechanisms that render drugs ineffective. One of the well-known factors involved with the aforementioned mechanisms is P-glycoprotein (Pgp) which is usually overexpressed in MDR and functions and an efflux pump that prevents drug molecules from diffusing through the cell membrane. It is speculated that by encapsulating potent drugs such as anti-tubulins in nanoscale drug carriers, nanoparticles (NPs), drug-molecule interaction with the cell membrane and thus the Pgp mechanism can be bypassed. Additionally, the NPs can improve the toxicity profile of drugs and help deliver drugs that are too hydrophobic in their native form. Polymeric NPs are a class of nanomedicine that can be engineered for safety and biodegradability. They are also great drug carriers for extremely hydrophobic molecules and can potentially modulate the cellular uptake pathways to bypass Pgp and MDR. In this article, the Li lab from the University of British Columbia describe a polymeric NP system that is highly effective in MDR tumors in vivo and significantly increases survival rates in the animal model.

The Li lab first assessed the cytotoxic activity of a number of known anti-tubulin agents against MDR tumor cells and found that Podophyllitoxin (PPT) was highly potent against all 3 different MDR tumor cell lines tested. PPT is however highly toxic and is not water soluble. Therefore, despite its high potency, PPT is not commonly used for cancer treatment. Chemical modification approaches to improve PPT safety and efficacy profiles have previously failed. The Li group therefore, developed an NP delivery system to improve safety and solubility issues of PPT. They conjugated PPT and varying amounts of polyethylene glycol (PEG) to a carboxymethyl cellulose (CMC) backbone. The polymer-drug conjugates were then formulated into NPs using the NanoAssemblr^TM Benchtop Instrument to achieve high uniformity and consistency in NP size. The authors found that the PPT/PEG ratio determined the size and drug release rate. Higher PPT/ PEG led to larger particles and slower drug release rates which may be due to the formation of a stable complex hydrophobic core in high PPT NPs.

The authors then in vivo-tested 3 candidate PPT-NPs with 3 different PPT/ PEG ratios of 20, 5 and 2 with sizes of 120, 30 and 20 nm respectively. The 3 formulations were fluorescently labeled and injected intravenously into MDR tumor bearing mice. Imaging of the harvested tissues indicated that the 20 nm NPs’ tumor accumulation was significantly higher than the 120 nm NPs which predominantly accumulated in the liver. The 20 nm NPs led to tumor accumulation of 8-fold higher than any other tissues while the 30 nm particles achieved a 3-fold increase and the 120 nm NPs tumor accumulation was 7 times lower than the liver. Additionally, tumor tissue analysis indicated different intra-tumoral distribution patterns for the 3 sizes of NPs. The 20 nm NPs were able to penetrate the core of tumors whereas the 30 and 120 nm stayed in tumor periphery. Tumor core penetration is thought to be of critical importance for treatment of cancer and inhibiting recurrence of tumors. The 20 nm NPs developed by the Li lab had a 20-fold uptake increase in the core compared to the other 2 sizes.

The PPT-NPs were then further assessed through maximum tolerated dose and antitumor efficacy studies. All three NP candidates were tolerated when dosed 3 times at 180 mg PPT/kg while the native PPT was very toxic at one dose of 20 mg/kg. The NPs were then compared side by side with native PPT and two well-known cancer drugs, docetaxel and Cabazitaxel. Following the treatments, the tumors rebounded in all treatments except the 20 mg/kg PPT-NP treated mice. Additionally, the mice in the 20 nm NP group had a median survival of >60 while in all the other groups the animals reached the end point before 38 days post treatment. Mice treated with 20 nm NPs showed no sign of toxicity and no palpable tumor was found in 7 out of 10 animals.

In conclusion, nanoparticles (NPs) can be efficient drug carriers to remodel drug safety and efficacy profiles while not requiring major chemical modifications on the drug molecule. NPs can assist with delivery of drugs that are too toxic or hydrophobic and can also help bypass cellular drug resistance mechanisms. The polymeric NPs developed in this paper efficaciously eradicated drug resistance tumors and did not impose any toxicity issues on the treated animals. Also, the NP backbone polymer modification provided a means of controlling the properties of NPs such as size, release rate and tumor penetration, indicating the great potential of customizing these systems as needed based on tumor characteristics. Additionally, these findings highlight the importance of particles size in the efficacy of the drug. Differences as small as 10 nm in the average diameter of the NPs produced appreciable differences in the survival of model animals. This highlights the importance of having NPs with well-defined size, which can be achieved with NanoAssemblr technology. More broadly, the findings in this paper demonstrate the great potential of nanomedicine for development of systemic drug delivery vehicles for treatment of cancer.

Abstract:

Podophyllotoxin (PPT) exhibited significant activity against P-glycoprotein mediated multidrug resistant (MDR) tumor cell lines; however, due to its poor solubility and high toxicity, PPT cannot be dosed systemically, preventing its clinical use for MDR cancer. We developed a nanoparticle dosage form of PPT by covalently conjugating PPT and polyethylene glycol (PEG) with acetylated carboxymethyl cellulose (CMC-Ac) using one-pot esterification chemistry. The polymer conjugates self-assembled into nanoparticles (NPs) of variable sizes (20–120 nm) depending on the PPT-to-PEG molar ratio (2–20). The conjugate with a low PPT/PEG molar ratio of 2 yielded NPs with a mean diameter of 20 nm and released PPT at ∼5%/day in serum, while conjugates with increased PPT/PEG ratios (5 and 20) produced bigger particles (30 nm and 120 nm respectively) that displayed slower drug release (∼2.5%/day and ∼1%/day respectively). The 20 nm particles exhibited 2- to 5-fold enhanced cell killing potency and 5- to 20-fold increased tumor delivery compared to the larger NPs. The biodistribution of the 20 nm PPT-NPs was highly selective to the tumor with 8-fold higher accumulation than all other examined tissues, while the larger PPT-NPs (30 and 120 nm) exhibited increased liver uptake. Within the tumor, >90% of the 20 nm PPT-NPs penetrated to the hypovascular core, while the larger particles were largely restricted in the hypervascular periphery. The 20 nm PPT-NPs displayed significantly improved efficacy against MDR tumors in mice compared to the larger PPT-NPs, native PPT and the standard taxane chemotherapies, with minimal toxicity.