GKA deel 1 college 5

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GKA deel 1 college 5

  1. 2. Hematopoietic Stem Cell Gene Therapy with a Lentiviral Vector in X-Linked Adrenoleukodystrophy Cartier, Hacein-Bey-Abina, Von Kalle, Fischer, Cavazzana-Calvo and others Science (2009) 326: 818 – 823 <ul><li>Demyelination </li></ul><ul><li>Defect in ALD protein </li></ul><ul><ul><li>ATP-binding cassette transporter </li></ul></ul><ul><ul><li>ABCD1 gene </li></ul></ul><ul><ul><li>Peroxisomal protein </li></ul></ul><ul><ul><li>Lipid degradation in oligodendrocytes and microglia </li></ul></ul><ul><li>Two boys treated </li></ul><ul><ul><li>HSC-Lentivirus </li></ul></ul>Inserted Nov 2009
  2. 3. Hematopoietic Stem Cell Gene Therapy with a Lentiviral Vector in X-Linked Adrenoleukodystrophy Cartier, Hacein-Bey-Abina, Von Kalle, Fischer, Cavazzana-Calvo and others Science (2009) 326: 818 – 823 <ul><li>Demyelination </li></ul><ul><li>Defect in ALD protein </li></ul><ul><ul><li>ATP-binding cassette transporter </li></ul></ul><ul><ul><li>ABCD1 gene </li></ul></ul><ul><ul><li>Peroxisomal protein </li></ul></ul><ul><ul><li>Lipid degradation in oligodendrocytes and microglia </li></ul></ul><ul><li>Two boys treated </li></ul><ul><ul><li>HSC-Lentivirus </li></ul></ul>Inserted Nov 2009
  3. 4. Hematopoietic Stem Cell Gene Therapy with a Lentiviral Vector in X-Linked Adrenoleukodystrophy Cartier, Hacein-Bey-Abina, Von Kalle, Fischer, Cavazzana-Calvo and others Science (2009) 326: 818 – 823 <ul><li>Demyelination </li></ul><ul><li>Defect in ALD protein </li></ul><ul><ul><li>ATP-binding cassette transporter </li></ul></ul><ul><ul><li>ABCD1 gene </li></ul></ul><ul><ul><li>Peroxisomal protein </li></ul></ul><ul><ul><li>Lipid degradation in oligodendrocytes and microglia </li></ul></ul><ul><li>Two boys treated </li></ul><ul><ul><li>HSC-Lentivirus </li></ul></ul>Inserted Nov 2009
  4. 5. Genetische therapiën <ul><li>Anti-sense therapie </li></ul><ul><li>Triplex-strategie </li></ul><ul><li>Gentherapie </li></ul><ul><li>RNAi </li></ul><ul><li>Gen-repair </li></ul><ul><ul><li>Via oligonucleotide </li></ul></ul><ul><ul><li>Via zink-vinger geïnduceerde klieving </li></ul></ul>
  5. 6. RNA interference; RNAi Pomerantz 2002, Nature Med 8, 659-660 <ul><li>RNA interference, RNAi </li></ul><ul><ul><li>siRNA </li></ul></ul><ul><ul><ul><li>Small interfering RNA </li></ul></ul></ul><ul><li>Sequence-specific </li></ul><ul><li>Degradation of mRNA </li></ul><ul><ul><li>Cellular mRNA </li></ul></ul><ul><ul><li>Viral mRNA (HIV?) </li></ul></ul><ul><ul><li>Function for the cell? </li></ul></ul><ul><ul><ul><li>Defense? </li></ul></ul></ul><ul><ul><ul><li>Control of gene expression </li></ul></ul></ul><ul><ul><ul><ul><li>Lin4-Lin14 </li></ul></ul></ul></ul><ul><li>Useful for therapy? </li></ul>
  6. 7. Different small RNAs Großhans and Filipowicz, Nature 451, 2008, 414-416 <ul><li>Inserted 15-2-2008 </li></ul>
  7. 8. Different small RNAs Großhans and Filipowicz, Nature 451, 2008, 414-416 <ul><li>Inserted 15-2-2008 </li></ul>
  8. 9. Different small RNAs Großhans and Filipowicz, Nature 451, 2008, 414-416 <ul><li>Inserted 15-2-2008 </li></ul>
  9. 10. Can RNAi be a therapy? Stevenson, Nature Rev Imm 3, 2003, 851-858 <ul><li>Large dsRNAs (>60 nucleotide pairs)  </li></ul><ul><ul><li>IFN: interferon response </li></ul></ul><ul><ul><ul><li>RNaseL </li></ul></ul></ul><ul><ul><ul><ul><li>Degradation RNA </li></ul></ul></ul></ul><ul><ul><ul><li>Induction PKR </li></ul></ul></ul><ul><ul><ul><ul><li>Protein kinase RNA-regulated </li></ul></ul></ul></ul><ul><ul><ul><li>Shut-off protein synthesis </li></ul></ul></ul><ul><ul><ul><ul><li>Non-specific </li></ul></ul></ul></ul><ul><li>DICER: RNase </li></ul><ul><li>RISC: RNA-induced silencing complex </li></ul>Viral (m)RNA
  10. 11. HIV life cycle Stevenson, Nature Rev Imm 3, 2003, 851-858
  11. 12. siRNA inhibits HIV infection <ul><li>Novina et al. 2002, Nature Med. 8, 681-686 </li></ul><ul><li>HIV binds receptors </li></ul><ul><ul><li>CD4 </li></ul></ul><ul><ul><li>CCR5 or CXCR4 (chemokine receptors) </li></ul></ul><ul><li>Targeting RNAi against </li></ul><ul><ul><li>Cellular mRNAs </li></ul></ul><ul><ul><ul><li>Required for viral replication </li></ul></ul></ul><ul><ul><li>Viral mRNAs </li></ul></ul><ul><ul><ul><li>Incoming RNA </li></ul></ul></ul><ul><ul><ul><li>Produced mRNA </li></ul></ul></ul>
  12. 13. How to make RNA? Stevenson, Nature Rev Imm 3, 2003, 851-858 One promoter
  13. 14. How to make RNA? Stevenson, Nature Rev Imm 3, 2003, 851-858 One promoter Two promoters
  14. 15. How to make RNA? Stevenson, Nature Rev Imm 3, 2003, 851-858 One promoter Two promoters Viral vector: adeno, HIV
  15. 16. Factors influencing effect of RNAi Stevenson, Nature Rev Imm 3, 2003, 851-858 <ul><li>Mutation of HIV-RNA </li></ul><ul><ul><li>Loss of complementarity, required for RNAi </li></ul></ul><ul><li>Virus in vesicles </li></ul><ul><li>Viral RNA partly inaccessible </li></ul>
  16. 17. Factors influencing effect of RNAi Stevenson, Nature Rev Imm 3, 2003, 851-858 <ul><li>Mutation of HIV-RNA </li></ul><ul><ul><li>Loss of complementarity, required for RNAi </li></ul></ul><ul><li>Virus in vesicles </li></ul><ul><li>Viral RNA partly inaccessible </li></ul><ul><li>How to deliver? </li></ul>
  17. 18. Delivery to brain cells Nature 448, 2007, 33-34 <ul><li>Use of the acetylcholine receptor to insert dsRNA duplexes </li></ul>Kumar et al ., Nature 448, 2007, 39-43, Transvascular delivery of small interfering RNA to the central nervous system Inserted July 10, 2007 ========

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