Gene therapy

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Gene therapy

  1. 1. Article: "Noninvasive and Targeted Gene Delivery into the Brain Using Microbubble-Facilitated Focused Ultrasound" Malova Anna 06.04.2013 Malova Anna Article: "Noninvasive and Targeted Gene Delivery into the Brain Using Microbubble-Facilit06.04.2013 1 / 16
  2. 2. Introduction rAAV Gene therapy is a potentially powerful means of treatment of various diseases with genomic causes. Recombinant adeno-associated viral (rAAV) vectors No pathogenicity Typical persistence of the transgene Low immunogenicity Complete removal of all viral genes Long-term gene expression Malova Anna Article: "Noninvasive and Targeted Gene Delivery into the Brain Using Microbubble-Facilit06.04.2013 2 / 16
  3. 3. Introduction Using AAV serotype 2 (AAV2) vectors uses for the treatment of various CNS diseases as Canavan’s disease Batten’s disease Parkinson’s disease Alzheimer’s disease Malova Anna Article: "Noninvasive and Targeted Gene Delivery into the Brain Using Microbubble-Facilit06.04.2013 3 / 16
  4. 4. Introduction Blood-Brain Barrier Separation of circulating blood from the brain extracellular fluid in the central nervous system Protects the brain tissue from circulating microorganisms and toxins Actively transport metabolic products such as glucose across the barrier with specific proteins Malova Anna Article: "Noninvasive and Targeted Gene Delivery into the Brain Using Microbubble-Facilit06.04.2013 4 / 16
  5. 5. Introduction Problems Existence of the blood-brain barrier (BBB) Intravenous injection is ineffective Need local, direct injection into the brain Region of recombinant gene-expression is severely limited Transfected cells cannot be widely spread Additional risks Burr holes Specific targeted gene expression cannot be controlled Using IV injection other organs would be significantly infected Malova Anna Article: "Noninvasive and Targeted Gene Delivery into the Brain Using Microbubble-Facilit06.04.2013 5 / 16
  6. 6. Introduction Microbubble-enhanced focused ultrasound (FUS) Locally and temporally disrupt the BBB Noninvasive method Contrast-enhanced magnetic resonance imaging can be used to observe, monitor and guide Low viral titer of 1 × 109 viral genomes Malova Anna Article: "Noninvasive and Targeted Gene Delivery into the Brain Using Microbubble-Facilit06.04.2013 6 / 16
  7. 7. Goal Experimental scheme Malova Anna Article: "Noninvasive and Targeted Gene Delivery into the Brain Using Microbubble-Facilit06.04.2013 7 / 16
  8. 8. Goal Preparing Outbred mice were separated into two groups Group 1 were used to optimize the time course of AAV infection from 1 to 6 weeks Group 2 used to evaluate the efficiency of approach at the optimized time for infection of 2–3 weeks. Malova Anna Article: "Noninvasive and Targeted Gene Delivery into the Brain Using Microbubble-Facilit06.04.2013 8 / 16
  9. 9. Goal Experiment Catheter was inserted into the tail vein for injections Viral vectors with the titer of 1 × 109 viral genomes per gramm were bolus injected through catheter Immediately followed a microbubbles bolus injection Ultrasonic energy was delivered to the brain transcranially using a spherically focused transducer Animals were sacrificed after FUS sonication. The brains of these mice were quickly removed and frozen Malova Anna Article: "Noninvasive and Targeted Gene Delivery into the Brain Using Microbubble-Facilit06.04.2013 9 / 16
  10. 10. Results GFP Expression Malova Anna Article: "Noninvasive and Targeted Gene Delivery into the Brain Using Microbubble-Facilit06.04.2013 10 / 16
  11. 11. Results GFP Expression. Explanation Determine the kinetics of infection in the BBB-opened brain region (top) In the absence of BBB-opening, GFP expression did not detect GFP expression could be observed within the first week Reached a maximum level at week 3, after which it decayed. AAV was only capable of transducing cells located in the vicinity of the BBB-breakdown injection track region but could not infect the parenchyma containing an intact BBB structure Malova Anna Article: "Noninvasive and Targeted Gene Delivery into the Brain Using Microbubble-Facilit06.04.2013 11 / 16
  12. 12. Results Correlation between MRI and GFP expressions Malova Anna Article: "Noninvasive and Targeted Gene Delivery into the Brain Using Microbubble-Facilit06.04.2013 12 / 16
  13. 13. Results Correlation between MRI and GFP expressions. Explanation GFP expression sites colocalized well with the contrast-enhanced regions observed in MRT images from the same animals High correlation between GFP expression and MRI signal increase Higher degree of BBB-opening induced a greater level of AAV transfection and expression Malova Anna Article: "Noninvasive and Targeted Gene Delivery into the Brain Using Microbubble-Facilit06.04.2013 13 / 16
  14. 14. Discussion Discussion Unlike current invasive procedures involving direct local injection of viral vector, FUS procedure concentrated ultrasound transcranially to induce local BBB opening for expressing genes at specific target regions in a noninvasive manner. GFP expression correlated well with MRI signal enhancement, suggesting the possibility of using MRI-monitored BBB-opening Since viral transduction of specific cells relies on the promoters that drive viral vectors, it is important to select promoters that will result in efficient gene expression in the selected target cells Malova Anna Article: "Noninvasive and Targeted Gene Delivery into the Brain Using Microbubble-Facilit06.04.2013 14 / 16
  15. 15. Discussion Future Directon The given viral vector in this study should be of no safety concern, and there should be highly possible to further improve the gene expression rate and transduction distribution when higher titer of viral vector is employed Combined use of viral-vector intravenous administration with FUS-BBB opening is a potential technique to achieve targeted gene delivery for CNS disease treatment noninvasively. Malova Anna Article: "Noninvasive and Targeted Gene Delivery into the Brain Using Microbubble-Facilit06.04.2013 15 / 16
  16. 16. Questions Questions Thank you for your attentions! Questions? Malova Anna Article: "Noninvasive and Targeted Gene Delivery into the Brain Using Microbubble-Facilit06.04.2013 16 / 16

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