Non-viral ocular delivery


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PEG-PLA microparticles for encapsulation and delivery of Tat-EGFP to retinal cells.

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Non-viral ocular delivery

  1. 1. CANADIAN RESEARCH FOCUS Interview with Dr. Mehrdad Rafat “ PEG-PLA microparticles for encapsulation and delivery of Tat-EGFP to retinal cells”, Biomaterials (2010). 31: 3414-3421. doi:10.1016/j.biomaterials.2010.01.031 May 7 th , 2010 conducted by Patricia Comeau
  2. 2. <ul><li>Presentation Contents </li></ul><ul><li>Brief background on article Slides 3 - 5 </li></ul><ul><li>Interview with Dr.Rafat Slides 6 - 23 </li></ul><ul><li>Dr. Rafat’s Biography Slides 24 - 27 </li></ul>
  3. 3. <ul><li>PEG-PLA microparticles for encapsulation and delivery of Tat-EGFP to retinal cells </li></ul><ul><li>Polyethylene glycol-polylactic acid (PEG–PLA) microparticles were used for encapsulation and delivery of a Transactivator of transcription-enhanced green fluorescent protein fusion (Tat-EGFP) to retinal cells. </li></ul><ul><li>Main objective was to develop a system that delivered Tat-EGFP with an initial rapid release (within 24 h) followed by a sustained release </li></ul>
  4. 4. Figure 1: Tat-EGFP encapsulated nano/microparticles subretinally injected into the outer nuclear layer of the retina Image courtesy of Rafat, M., University of Ottawa; and Kolb, E., University of Utah, 2010
  5. 5. <ul><li>Concerns for delivery of an effective therapy to the retinal cells include restricted permeability of the corneal and conjunctival epithelia, and the presence of the blood-retina barrier. </li></ul><ul><li>The size of any delivery vehicle must be small enough not to negatively impact the sensitive retinal cells. Microparticles and nanoparticles in particular offer the advantage of a controlled and sustained subcellular drug release. </li></ul>
  6. 6. Interview with Dr. Rafat Vision Program, Ottawa Hospital Research Institute and Department of Cellular and Molecular Medicine, University of Ottawa
  7. 7. What is the need for controlled release technologies in the eye?  <ul><li>Drugs and therapeutic agents have been traditionally administered to the eye as topical liquid drops. One of the main problems in ocular therapeutics is the delivery of an optimal concentration of a therapeutic agent at the target site for a prolonged period of time. It is believed that less than 5% of a therapeutic agent administered topically is ocularly absorbed. </li></ul>…continued on next slide ->
  8. 8. <ul><li>This low ocular absorption is due to the loss in the tear film, and the corneal layers as well as the restricted permeability of the corneal and conjunctival epithelia. </li></ul><ul><li>These limitations are more critical for the retina, as most of the retinal diseases involve cells in the back of the eye. In addition, due to the presence of the blood-retina barrier, drug delivery to the retina by conventional methods poses a challenge. </li></ul>
  9. 9. What are the key target diseases? <ul><li>Key target diseases include: </li></ul><ul><ul><li>age-related macular degeneration, </li></ul></ul><ul><ul><li>diabetic retinopathy, </li></ul></ul><ul><ul><li>glaucoma, </li></ul></ul><ul><ul><li>retinal ischemia, </li></ul></ul><ul><ul><li>retinal detachment, </li></ul></ul><ul><ul><li>cataract, and </li></ul></ul><ul><ul><li>ocular herpes </li></ul></ul>
  10. 10. How and where are the microparticles injected into the retina? <ul><li>The microparticles can be delivered into the eye by subretinal injection. It is performed by creating a sclerotomy (surgical incision of the sclera) about 2 mm posterior to the limbus. A coverslip coated with 0.3% hypromellose is placed on top of the eye to provide magnification and visualization of the back of the eye. </li></ul>…continued on next slide ->
  11. 11. <ul><li>Tat-EGFP encapsulated microparticles is dispersed in Dulbecco's Phosphate Buffered Saline and transferred to a 10-mL syringe with a 33-gauge blunt needle attached. The needle is then inserted through the scleral puncture, guided lateral to the lens, and inserted through the retina and microparticles are injected to the subretinal space of the eye. </li></ul>
  12. 12. What other characteristics of the microparticles, aside from size and morphology, influence subretinal delivery? <ul><li>Polarity and hydrophilicity of the microparticles influence the delivery and release mechanisms. For example, more hydrophilic polymers tend to absorb more water resulting in faster degradation of microparticles and faster release of therapeutic agents. </li></ul>
  13. 13. Clinically, what would you replace Tat-EGFP with to treat a retinal disease? <ul><li>Our plan is to replace Tat-EGFP with x-linked inhibitor of apoptosis protein (XIAP) for clinical applications. We have previously reported that XIAP confers structural neuroprotection of photoreceptors for at least 2 months after retinal detachment, which is also associated with AMD. </li></ul>…continued on next slide ->
  14. 14. <ul><li>XIAP is a key member of the inhibitors of apoptosis gene family and is a promising therapeutic agent as it suppresses caspases 3, 7, and 9, whose activation has been shown to cause apoptotic cell death in retinal detachment animal models. </li></ul>
  15. 15. Why is there a difference between the cellular uptake at 48 and 96hours? <ul><li>This phenomenon may be caused by the biodegradation of PEG-PLA nano/microparticles resulting in the break down of larger particles into smaller ones. Also, please note that there might be some image to image variations as the images for various time points were not taken at exactly the same spot in the culture dish. </li></ul>
  16. 16. How do electroretinograms determine the biocompatibility of micro-particles? <ul><li>Electroretinography (ERG) measures the electrical responses of various retinal cell types, including the photoreceptors, inner retinal cells (bipolar cells), and the ganglion cells. E.g. an A-wave is generally caused by extracellular ionic currents generated by photoreceptors and B-wave is generated by bipolar cell activity. </li></ul>…continued on next slide ->
  17. 17. <ul><li>If microparticles had caused any cell toxicity or death, the responses that we would get on ERG would have been different than those of normal healthy cells. Because no significant differences between PEG-PLA-treated eyes and control healthy eyes were observed in our ERG study, it suggested that the particles were biocompatible toward the retinal cells. </li></ul>
  18. 18. Does the presence of the microparticles in the retina pose any potential risks?  <ul><li>According to our findings so far, the presence of PEG-PLA microparticles in the outer nuclear layer of the retina did not cause toxicity or adverse side effects. However, one potential risk factor for these particles is their non-transparent nature. This phenomenon may temporarily cause blurred vision until the particles are fully degraded in the eye, which may take up to few months. </li></ul>
  19. 19. When do you plan for the particles to release the drug? <ul><li>One of the goals is to have the particles release their protein once they become embedded in the retina. The microparticles and their release profile, however, need to be customized for each ocular disease. For example, chronic, progressive disorders need a continuous moderate release of the therapeutic agents over time while acute insults require immediate intervention. </li></ul>
  20. 20. How are the polymer degradation products removed from the eye? <ul><li>As the particles are directly injected into the retina, we know that there is blood circulation in the retina, e.g., it is continuously supplied with oxygenated blood via the retinal artery and drained of deoxygenated blood via the central retinal vein. Therefore, it is very likely that biodegradation products leave the eye via the retinal vein and capillaries. </li></ul>
  21. 21. What major hurdles remain to be overcome before patients can benefit from its application? <ul><li>Despite the promising nature of this technology, we need to conduct 3-5 more years of research to refine and engineer various formulations using different proteins (e.g. XIAP) and polymers and tailoring them for various ocular diseases. We also need to test these formulations in bigger animal models for the proof of concept prior to moving into human trials. </li></ul>…continued on next slide ->
  22. 22. <ul><li>To achieve these goals we will need more public and private funding. </li></ul><ul><li>Other obstacles include the regulatory matters involved with clinical trials, the high cost of clinical trials, and development of manufacturing facilities and protocols that are in compliance with Good Manufacturing Practice requirements. </li></ul>
  23. 23. CC-CRS Question #8 Thank you for the interview!
  24. 24. Dr. Mehrdad Rafat Biography of
  25. 25. <ul><li>Dr. Rafat received his Masters and Ph.D. degrees in Chemical Engineering from the University of Ottawa with specialization in Biomaterials and Tissue Engineering. After completing his Ph.D., Dr. Rafat joined Dr. Tsilfidis’s group as a post-doctoral fellow (PDF) at the Ottawa Hospital Research Institute (OHRI) and worked on the development of nanoparticles for controlled gene delivery to retinal cells for prevention of retinal blindness. Dr. Rafat is currently a PDF at OHRI/UOttawa with Dr. Tsilfidis and Dr. Isabelle Catelas working on the development of nanoparticles systems for controlled release and delivery of proteins/drugs for retina, and bone regeneration applications, respectively. </li></ul>…continued on next slide ->
  26. 26. <ul><li>As a result of his work towards the invention of the first clinically-tested bioengineered cornea he was awarded NSERC’s 2008 Innovation Challenge Award and the Ontario Centers of Excellence Industrial Fellowship Award (OCE/CMM ) in 2006. </li></ul><ul><li>In addition to his academic endeavors, Dr. Rafat has also worked with the Medical Devices Bureau at Health Canada for evaluation and regulation of medical devices, as well as been a senior scientific consultant to biotech industries including the Hawaii-based firm, Cellular Bioengineering Inc, and the start-up company, Bioconstrux Inc. of Ottawa. </li></ul>…continued on next slide ->
  27. 27. <ul><li>Dr. Rafat’s research interests are mainly focused on the development of bioengineered materials as implantable scaffolds and nano and microparticles systems for controlled delivery of cells, drugs, and proteins for biomedical applications. More specifically, he is interested in the application of hybrid nanomaterials in regenerative medicine – particularly that involving ocular and cardiovascular therapies. Beyond the development of bioengineered materials he is also interested in the evaluation, regulation, and commercialization of medical device and therapeutic technologies for various medical applications. </li></ul>