APH-0804

A combination of Tdp1 inhibitors such as tetracycline and Top1 inhibitors such as topotecan or irinotecan, co-encapsulated in phospholipid nanosomes for camptothecin-resistant cancers

Combination Therapy for Drug Resistant Cancers

Camptothecin is a potent anticancer agent that works by inhibiting topoisomerase 1 (Top1), an enzyme that cleaves DNA and promotes cell division and cancerous growth. Another enzyme Tdp1 (Tyrosyl DNA phosphodiesterase) can repair Top1-DNA covalent complexes by hydrolyzing the tyrosyl-DNA bond. Inhibiting Tdp1 has the potential to enhance the anticancer activity of Top1 inhibitors such as camptothecins (camptothecin, irinotecan, topotecan) and to act as anti-proliferative agents. The development of Tdp1 inhibitors as anticancer agents can be envisioned as combinations of Tdp1 and Top1 inhibitors.

Top1 inhibitors such as camptothecins are extremely hydrophobic, while Tdp1 inhibitors such as the antibiotic tetracyclines are quite hydrophilic. In order for these drugs to work in unison, they should be optimally co-encapsulated and delivered to the cancerous cells as a single package, such as nanoparticles.

Current Co-Encapsulation Challenges

At present, there are no available technologies that can readily co-encapsulate hydrophobic and hydrophilic drugs in phospholipid nanosomes in a single-step, scalable process. Conventional processes for the encapsulation of hydrophobic drugs utilize many processing steps and require large quantities of organic solvents.

These processes are very time consuming, costly and inefficient. Generally, such phospholipid liposomes have a wide dispersion of particle size, and they tend to have a median size greater than 100 microns in diameter. In addition, the exposure of therapeutic agent to the organic solvent may adversely affect the integrity of the final product.

Other conventional processes for the encapsulation of hydrophilic drugs into phospholipid liposomes utilize high pressure homogenization that requires a significant amount of recycling, generates heat with every pass through the homogenizer and could be contaminated with heavy metal particles. These conventional processing methods may also compromise stability.

Tdp1 and Top1 Inhibitor Co-Encapsulation

CFN was used to co-encapsulate Top1 and Tdp1 inhibitors in phospholipid nanosomes (small, uniform liposomes). Camptothecins are hydrophobic and will be packaged in the lipid bilayer. Tdp1 inhibitors such as tetracycline and neomycin are water-soluble and will be packaged in the aqueous core of phospholipid nanosomes.

CFN technology was used to form stable and uniform phospholipid nanosomes containing camptothecin (Top1) and neomycin (Tdp1) as well as irinotecan (Top1) and tetracycline (Tdp1). Nanosomes ranged from 127 to 178 nm and they were isolated by ultracentrifugation, size exclusion chromatography (SEC) or dialysis. By SEC, encapsulation efficiencies were ~46% and ~74%, respectively, for irinotecan and tetracycline.

Benefits and Advantages of Co-Encapsulation for Combination Cancer

• Combination therapy will solve the unmet medical problem of camptothecin-resistant cancers.
• Nanosomal formulations will result in reduced systemic toxicity, due to masking of cytotoxic effects of camptothecins and Tdp1 inhibitors.
• Stability of the lactone ring in nanosomes will be improved as a result of protection from the neutral pH of the blood stream.
• By increasing residence time in the circulatory system, nanosomes will increase therapeutic efficacy of the combination drugs.
• Pegylated phospholipids can be utilized to provide steric hindrance that will increase residence time and therapeutic efficacy as is done with Doxil^®, liposome encapsulated doxorubicin.
• Phospholipids linked with cancer-specific ligands can be utilized to target co-encapsulated camptothecin and Tdp1 inhibitors to cancers of the colon, lung or ovary.
• This smart targeting will further reduce toxicities associated with Top1 and Tdp1 inhibitors while increasing efficacy and therapeutic index.