Azaya Therapeutics PSN Technology

Introduction:

Azaya Therapeutics, Inc. is an emerging specialty pharmaceutical company with a novel drug delivery system. Its proprietary Protein Stabilized Nanoparticle (PSNTM) technology platform addresses the significant problems associated with delivery of water insoluble drugs. The Company is initially applying its PSN technology to produce a targeted, safer and more efficacious formulation of a marketed chemotherapy drug for the treatment of cancer.

Scientific Background:

Liposomes are vesicular sacs resembling tiny cells—their thin but durable lipid bilayer membranes separate an internal aqueous volume from the external medium. Water-soluble drugs can be encapsulated in the internal compartments, while water-insoluble drugs can be incorporated in the hydrophobic region of the membrane. Depending on the production process used, each vesicle may contain a single membrane (unilamellar) or several layers of membrane (multilamellar). Typical liposome sizes range from 0.05 to several μm in diameter.

Since their discovery more than three decades ago, liposomes have been applied to the delivery of a wide range of drugs by virtually every route of administration. The prospect of targeting liposome encapsulated drugs to sites of disease (such as tumors) generated considerable attention in medical research the late 60’s and early 70’s. Although early liposomal formulations were shown to reduce toxicity in animal models, they were no more active than the standalone active ingredient, partly as a result of the stability of the liposome, typically emptying 50% of its contents as a consequence of rapid binding of plasma proteins. Furthermore, liposomes that survived destabilization in blood were rapidly sequestered by fixed macrophages residing in the liver and spleen (the mononuclear phagocyte system [MPS]), destroying the liposomes.

  1. Advances:
    During the late 70’s and early 80’s liposomes were reengineered to circulate longer in the blood and remain intact while doing so. This was done first by grafting the liposome with glycolipids, followed by a second generation and more robust grafting of the liposome with methoxypolyethylene glycol (mPEG), otherwise referred to as stealth liposomes. The PEG grafted liposomes increased the in vivo liposome stability, reduced MPS uptake and provided longer plasma residence times. In short, the liposome was engineered for use in the pharmaceutical industry, but still remand difficult to produce on large scale for production.
  2. Traditional Manufacturing Process:
    The traditional method used for the production of liposome products (e.g. Doxil) includes a multi step process; pre-formation of the liposome (pre-emulsion), pre-sizing via filter extrusion (0.05–0.45 µm), dialyzing the pre-liposome suspension with a 5% to 15% aqueous sucrose solution, encapsulating the active ingredient by mixing an active ingredient (e.g. doxorubicin) and the liposome suspension, by an aseptic filtration, and a final product dispensed into commercial vials. This lengthy process requires several complex steps that need meticulous attention, resulting in a costly process for manufacturing.

    In addition to the high cost associated with traditional liposomal production, the final product has a limited shelf life. Typically, the final liposome encapsulated drug product is packaged and delivered as a saline suspension that is stored at reduced temperatures. As a result these products have a finite shelf life of approximately 12 months.

  3. Azaya Therapeutics New Technology:
    As a result, Azaya Therapeutics has developed its Protein Stabilized Nanoparticle (PSN™) technology that utilizes a one step manufacturing process. The liposome product is prepared in a single step that encapsulates the active drug Docetaxel (ATI-1123) in the lipid layer of the liposome while forming active nanoparticles in situ (60-80 nm). The active liposomal product is dispensed into vials and lyophilized to yield a dried powder that has an extended shelf life compared to conventional liposomes. Drug stability and release rates are predicted to be <5% in saline after 24 hours, while the reconstituted material that has been stored at 4° C for several months shows complete retention of the encapsulated drug with <5% of the active ingredient released from the liposome.

    While Azaya has successfully demonstrated the encapsulation of Docetaxel (ATI-1123), this process is geared toward the formulation of hydrophobic molecules that would otherwise have limited success as developmental drugs using traditional formulation methodologies. Furthermore, a variety of marketed cytotoxic agents use the more traditional formulations that use toxic carrier i.e. Cremophore EL and Tween 80, while the Azaya liposome formulation utilizes natural and inactive excipients (lipids, proteins and cholesterol), eliminating the need for toxic carriers.

    Although many of the known cyctotoxic agents are poorly soluble, Azaya’s PSN technology offers a potential solution that not only addresses formulation issues of poorly soluble compounds, but it can also enhance the activity (efficacy) of known drug products through a passive targeting mechanism. As it would imply, passive targeting is a result of small particles being preferentially taken up into the cancer tumor, resulting in higher concentration of active drug at the diseased site. This is achieved by the enhanced permeation and retention effect (EPR); liposomes (small particles <200 nm in size) move into the tumor from the bloodstream through holes or gaps in the vessels supplying the tumor (vasculature leakage), accumulating a larger quantity of cyctotoxic agent or nanoparticle within the diseased tissue.

    Azaya’s single step manufacturing process incorporates key characteristics that enhance existing liposomal technology for drug delivery. To date, there have been only moderate advances and Azaya’s PSN technology is a large step towards a more efficient and economical drug delivery system. Additionally, standardized manufacturing produces uniform particles that are necessary for passive targeting of cancer tumors via the EPR effect. With this superior technology, Azaya Therapeutics has conducted pre-clinical studies that have demonstrated improved therapeutic potential; enhanced pharmacokinetics, enhanced inhibition of tumor xenograpft models and evidence of increased tumor uptake of active compound.

  4. Other Nanotechnologies:
    1. Emulsions:
      Abraxane is a protein stabilized emulsion that enhances the solubility of water insoluble or hydrophobic molecules, in this case paclitaxel. Although this type of formulation does not enhance the pharmacokinetic profile of the active ingredient, it does reduce toxicities associated with the excipients i.e. Cremophor EL, used to formulate taxol. As a result, the solubility of taxol is improved and the non drug related toxicities is eliminated as a result. This technology enhances the solubility of a limited class of drug product, and has limited affect on the pharmacokinetics. As a result, this technology possesses a major limitation to improve cancer therapies, while the objective is to develop better drug products to treat the disease.
    2. Dendrimers:
      A dendrimer is a polymer that is highly branched monomer leading to a monodisperse, tree-like or generational structure. They have well-defined chemical structures and exhibit mono-dispersity and have potential applications in targeting cancer cells and drug delivery. Currently several products are in development for various indications. A major limitation is the uncertainty about potential product toxicity and process manufacturing (scale-up).
    3. Fullerenes:
      A form of carbon (C-60), fullerenes are composed of carbon atoms arranged in a soccer ball-like shape, hence their other name—buckyballs. Easily manufactured in quantity, they seem to be ideal drug delivery vehicles due to their size and shape. These particles have a great deal of potential, but a major concern that arises is the potential for toxicity. Currently several products are in pre-clinical development.
    4. RNA nanotechnology:
      RNA fragments associated with active compounds that self recognize to form a nanoparticle used for drug delivery. This technology is in early discovery, entering into development.
    5. Nanosomes (SuperFluids CFN technology):
      Lipid like membranes used to encapsulate active drugs. This technology is in early discovery, entering into development.

    All of these technologies have scale-up challenges that have yet to be overcome. As technology advances, many of these tools will potentially become viable tools for drug formulations.

Supporting Data for Azaya’s ATI-1123 PSN Formulation of Docetaxel:

  1. Proof of principle established for Azaya’s lead drug candidate ATI-1123 (a reformulation of Taxotere® (docetaxel), a marketed drug with worldwide annual sales of $2+ billion)
  2. Eliminated toxic carrier used in Taxotere and encapsulated active compound in protein stabilized liposome as a nanoparticle (< 200 nanometers) that allows for tumor targeting through Enhanced Permeation and Retention effect (EPR)
    • Pharmacokinetics (PK) study established a significantly enhanced profile (higher drug concentrations and lower clearance) for ATI-1123 versus docetaxel—better anti-tumor activity.
    • Prostate and lung tumor xenograft studies showed better efficacy for ATI-1123 versus docetaxel.
    • Technology transfer in process with Brookwood Pharmaceuticals, Inc. with expected production of GLP material for IND toxicology study in Q4 2006.

    The Company has successfully completed several key preclinical studies, including a MTD study, a pharmacokinetics study, a plasma concentration and tumor uptake study, biodistribution, prostate tumor xenograft model, a lung tumor xenograft model, pancreases xenograft model and mesothelioma xenograft model. The results of these studies demonstrate that ATI-1123 has (1) a pharmacokinetic profile that is very distinct from docetaxel with a 6-fold increase in area under the curve (AUC) and (2) significantly greater anti-tumor activity with potentially reduced toxicity.

  3. Pharmacokinetics Study:
    An extensive pharmacokinetics (PK) study of ATI-1123 has been completed in rats to compare the distribution and clearance of ATI-1123 vs. docetaxel at three different dose levels. The results demonstrated that the Azaya PSL technology:
    • Significantly improved the amount of docetaxel and its duration in the bloodstream
    • Clearly showed a difference in key parameters such as:
      • Exposure—dose dependant increase in exposure for both drugs with ATI-1123 showing a significant amplification in concentration over an extended period.
      • Clearance—lower clearance observed for ATI-1123.
      • Area Under the Curve (AUC)—showed a linear dose correlation for both drugs with ATI-1123 showing a significant increase in AUC.

    These critical findings show significant differentiation of ATI-1123 from docetaxel with ATI-1123 having potentially improved anti-tumor benefits.

    Concluding remarks from IDD (the Institute for Drug Development (IDD), a division of The Cancer Therapy and Research Center):
    The purpose of this nonclinical study was to identify potential differences in docetaxel pharmacokinetics when administered using the Azaya liposomal drug delivery system (ATI-1123) compared with standard docetaxel formulated in Tween 80 and ethanol. The study findings show clear and marked differences in docetaxel plasma kinetics following administration of ATI-1123 compared with standard vehicle docetaxel. Specifically, plasma concentrations of docetaxel were consistently higher for ATI-1123 over the entire exposure period. This results in a 4- to 5-fold elevation of total plasma docetaxel AUC associated with ATI-1123 administration compared with an equivalent dose of docetaxel using a standard vehicle. This difference corresponds to a decrease in systemic docetaxel clearance for ATI-1123. Higher plasma concentrations are also consistent with a smaller volume of distribution for the ATI-1123 liposome preparation.

    These marked differences in plasma docetaxel kinetics are in the range and direction expected from a successful liposomal drug delivery system. This pharmacological alteration traps docetaxel in the intravascular compartment, decreases the volume of drug distribution, and prolongs the duration and magnitude of systemic drug exposure. If the kinetic profile of docetaxel arising from ATI-1123 was identical or overlapped that seen with standard vehicle docetaxel, the integrity of the liposomal delivery system would have to be questioned. These finding show clear and marked differences in plasma kinetics for ATI-1123 and as such, are highly promising. These data support further studies of this agent to analyze the alterations in drug distribution in tissues and most importantly, difference in toxicity and overall antitumor efficacy.

  4. Preclinical Xenograft Studies:
    To further differentiate and benchmark ATI-1123 to the standard docetaxel formulation, a number of xenograft studies were completed to show biological activity using non-GMP research drug material.

    The most exciting study results to date have been seen in a human prostate (PC-3) xenograft model.  Single agent ATI-1123 administered every other day for 3 doses (3 total injections) produced a mean estimated tumor growth inhibition of greater than 100% on Day 36 of the study. In this regard, at Day 36 of the study, all animals treated with ATI-1123 at 25 mg/kg had achieved either a partial (8/9) or complete (1/9) response or regression. In comparison, docetaxel at the maximum tolerated dose produced a mean tumor growth inhibition of 85% with no partial or complete responses observed in any animal. By Day 36 of this study, tumors had begun to regrow in the group treated with docetaxel, while tumors in the group treated with ATI-1123 remained stable.

    This data suggest that ATI-1123 is highly effective against the PC-3 human prostate tumor model. In comparison to docetaxel, these results imply that a higher dose of ATI-1123 may potentially be administered, which produces enhanced antitumor activity in this model.

  5. Bio-Distribution Study:
    Early evidence in preclinical proof of principle studies indicates that the concentration of docetaxel in tumor tissue is higher in ATI-1123 vs. docetaxel (2–3 fold), again increasing the potential for higher efficacy.

In Summary:

Azaya Therapeutics has developed a unique, one step process to manufacture nanosized liposomes encapsulated with active compound; docetaxel (ATI-1123). This formulation technology has revealed a striking improvement in activity and pharmacokinetic profile of the parent drug, taxotere. These results warrant further development to examine product safety for a new therapy for cancer treatment.

Supporting Literature:

  1. M. N. Khalid, P. Simard, D. Hoarau, A. Dragomir, J. Leroux, Long Circulating Poly(Ethylene Glycol)Decorated Lipid Nanocapsules Deliver Docetaxel to Solid Tumors, Pharmaceutical Research, 23(4), 2006.
  2. Yoshio Nakano, Phospholipids, Drug Delivery Technologies, 6(6), 2006, 60-63.
  3. Jason McKinnie, Nanobiotechnology, Drug Delivery Technologies, 6(6), 2006, 40-44.
  4. Cindy H. Dubin, Formulation Strategies for Poorly Soluble Drugs, Drug Delivery Technologies, 6(6), 2006, 34-38.
  5. Literature: Long Circulating Poly(Ethylene Glycol)-Decorated Lipid Nanocapsules Deliver Docetaxel to Solid Tumors
  6. Literature: Drug Delivery Technology, June 2006 Vol 6 No 6