Technology

Explanation of the technology and its advantages

Regulon has developed a unique liposome encapsulation technology applicable to drugs, small molecules, peptides, proteins and viruses aiming at improving human lives and reducing the side effects of chemotherapy known to exacerbate the quality of life (QOL) of cancer patients. The firm has successfully applied this technology to encapsulate two members of the platinum family of anticancer drugs, cisplatin and oxaliplatin. Platins are the cornerstone of modern chemotherapy, as they have been commonly applied in the effective treatment of the majority of tumor indications for the last 30 years. Cisplatin was approved in 1978 and increased dramatically the prognosis of epithelial malignancies; as a result of its introduction into the arsenals of chemotherapy the cure rate of testicular cancer rose from 10% to 90%. And in spite of over 30 years of “triumphal” discoveries in cancer drugs oncologists are turning back to cisplatin.

Despite their well-documented efficacy, however, platinum treatment, almost invariably, results in pronounced toxicity against the patient's kidneys, nerves, bone marrow, gastrointestinal tract, hearing system and most other vital organs. Thus, generating platins of lower toxicity and / or higher efficacy (higher therapeutic index) is an unmet medical need in cancer chemotherapy. The company’s technology has successfully addressed this need and has already shown clinical proof of concept in humans in advanced Phase III studies.

A major problem in treating cancer and other serious illnesses is that highly toxic drugs are often administered at suboptimal doses because of their side effects on healthy tissues. By targeting drugs specifically to tumor cells or other diseased cells and avoiding normal healthy cells, it may be possible to treat patients with safer and more effective doses. Regulon’s world-wide patented nanotechnology increases the efficacy of current block-buster anticancer drugs, while significantly reducing their serious side-effects on patients. Regulon’s unique liposome encapsulation technology can use most of the 1000 FDA-approved drugs and wrap these into liposomes thus generating branded names of off-patent drugs.

 

The technology is explained in the following Figures:

The mechanism of toxicity reduction is the same responsible for the increased levels of our drug in tumors and its higher efficacy. The 110 nm in diameter nanoparticles have the ability to find the tumors and metastasis in the body and to concentrate inside them; this process known as extravasation, takes advantage of the compromised endothelium of the vasculature of the tumors generated during neoangiogenesis. Lipoplatin has shown an amazing concentration in tumors and metastases at levels up to 200-fold higher compared to the adjacent normal tissue in surgical specimens from patients (Boulikas et al, 2005). This is why all side effects of cisplatin have been reduced by Lipoplatin.

Boulikas T, Stathopoulos GP, Volakakis N and Vougiouka M. (2005) Systemic Lipoplatin Infusion Results In Preferential Tumor Uptake In Human Studies. Anticancer Res 25, 3031-3040.

 

Depiction of a Lipoplatin Nanoparticle (right). Cisplatin molecules are depicted as yellow spheres surrounded by the lipid bilayer with the PEGylated lipid sticking out like hair from the outer surface. Thus, this toxic substance, cisplatin, is camouflaged by its lipid shell as a nutrient. This nanoparticle can pass undetected by macrophages after intravenous injection to human cancer patients because of its PEG coating thus escaping immune surveillance. Its interaction with serum components (a process known as opsonization) is also minimized and Lipoplatin can remain in systemic circulation for long periods with a half-life of 120 hours as shown from Phase I studies. These properties are prerequisites for the nanoparticle to find its target, the compromised endothelium of tumor vasculature, and leak out. For comparison, the non-PEGylated nanoparticle (left) is cleared rapidly by macrophages with a half-life of 20 min.

 

The extravasation of Lipoplatin nanoparticles to tumors (right). The scheme shows a blood vessel in normal tissue (left) and in tumor tissue (right). Lipoplatin nanoparticles are depicted as red spheres. In normal tissue blood vessels are impenetrable by small nanoparticles of 100nm. On the contrary, tumor blood vessels have imperfections (tiny holes) in their walls (called endothelium); tumor blood vessels are established during the process of neo-angiogenesis (meaning sprouting of new blood vessels by a tumor cell mass during its growth phase). Lipoplatin nanoparticles take advantage of these tiny holes to pass through and extravasate inside the tumor. Human studies have shown that the levels of platinum drug in the tumor tissue is up to 200-fold higher compared to its levels in the adjacent normal tissue from surgical specimens.

© CNRS Photothèque/SAGASCIENCE / CAILLAUD François

 

Penetration of Lipoplatin nanoparticles through the cell membrane of tumor cells. Lipoplatin nanoparticles once inside the tumor cell mass can fuse with the cell membrane because of the presence of the fusogenic lipid DPPG in their lipid bilayer; an alternative mechanism proposed is that Lipoplatin is taken by endocytosis by tumor cells as shown from Lipoplatin containing fluorescent lipids and imaging of the tumor cells in culture thus treated with fluorescent microscopy. These processes occurring at the cell membrane level are promoted by the lipid shell of the nanoparticles (disguised as nutrients). The technology allows Lipoplatin to empty its toxic payload (cisplatin) inside the cytoplasm to kill the tumor cell. The cell membrane is considered a significant barrier to trasportation of the toxic molecules of cisplatin across and inside the tumor cell. Thus, delivery of cisplatin molecules by Lipoplatin is a tremendous improvement not only in the efficient targeting but also for the uptake of cisplatin.

 

Lipoplatin or DPPG-liposomes with fluorescent lipids enter rapidly MCF-7 breast cancer cells in culture. Time-course processing of FITC-labeled DPPG-containing liposomes (left) and Lipoplatin (right) using confocal microscopy. At 5 min the majority of the signal is localized in the membrane. Lipids are rapidly internalized and at 4-24 hours, a strong signal is observed in the cytoplasm and at the perinuclear area. These results demonstrate that our nanoparticles (unlike those of Doxil) are able to cross the cell membrane barrier. This property is suggested to lower the side effects of the drug (hand-and-foot syndrome), to bypass resistance in platinum-treated patients and to enhance the efficacy of our drug. Platinum drugs can kill cancer cells only after crossing the cell membrane barrier to damage cellular components.

 

Furthermore, the technology lends itself to gene therapy applications, e.g. for cancer immunotherapy based on a liposomally encapsulated virus carrying the human IL-12 gene. This is a breakthrough in molecular medicine because it allows a number of potential gene therapy treatments to be delivered to patients.

In addition to cancer treatment, the technology can be used against viral infections (such as AIDS and hepatitis) as well as fungal, protozoan and bacterial infections to deliver liposomal antibiotics to the inflammatory area; in addition, the technology has applications in cardiovascular disease, arthritis and autoimmune diseases where alterations of the inflammatory tissue vasculature also enable targeting with Regulon’s nanoparticles as carriers of the appropriate drug. See more in: http://www.regulon.org/: Keynote Presentation on our Technologies.

 

2b. A nanotechnology formulation

Regulon’s nanoparticles represent an advanced nanotechnology formulation. Their composition is natural (lipid shell therefore no problem with cumulative toxicity from the carrier compared to synthetic nanomaterials). Furthermore, Lipoplatin nanoparticles supposedly are uptaken more by tumor compared to normal cells because of their avidity for nutrients; thus Lipoplatin nanoparticles are mistaken as nutrients by tumor cells. The fusogenic lipid on their surface also promotes fusion with the cell membrane.

 

2c. The antiangiogenesis potential of Lipoplatin

Lipoplatin has antiangiogenesis properties as was suggested from animal studies. Regulon plans to demonstrate this on tumor specimens from patients treated with Lipoplatin (Appendix 7, Figure 9).