基因治疗 PPT课件
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Poor selectivity and inefficiency of enrichment in the target cell or tissue.
■ Background
Different delivery tools
Virus Vector: Retrovirus, Lentivirus, Adenovirus, Adeno-associated virus
■ Background
Different treatments to the diseases
Chemical drugs Surgery Physical Therapy Transplantation therapy Immunotherapy Regenerative medicine
Gene therapy
■ Background
Technical problem: Barriers to the target
DNA/RNA is unstable in the bloodstream, can be immunogenic and does not readily cross membranes to enter cells. Nuclease, renal filtration
Non-virus method
Can deliver both siRNA and plasmid DNA
■ Background
Liposome
Protect entrapped oligonucleotides from nuclease degradation and renal clearance Promote cellular uptake and endosomal escape They include the use of cationic or ionizable lipids, shielding lipids, cholesterol and targeting ligands
Retrovirus:
Virus based transduction
Widely studied RNA virus Wide host range Infect dividing cell and just infect once Integrate into host genome Low Immunogenic Long expression period Insertion mutation by random integration and oncogenicity Can’t infect non-dividing cell and low virus titer
Triantennary GalNAc–siRNA
Non-virus method
ASGPR, on hepatocytes
Both subcutaneous and intravenous administration of this conjugate revealed great accumulation of siRNA in the liver and improved knockdown of the target gene.
※ Using DNA to express functional protein ※ Using siRNA or shRNA to attenuate abnormal gene
expression in mRNA level ※ Genome editing to correct mutant gene sequence: ZFN,
Non-virus method
Can deliver both siRNA and plasmid DNA
■ Background
Conjugate delivery
Non-virus method
Dynamic PolyConjugates (DPC)
First reported in 2007 PEG: shielding effect GalNAc ligand was essential for both uptake by hepatocytes and in vivo silencing activity. Other targeting ligands has been explored, including peptides, antibodies, small molecules, glycans, lectins and nucleic acids.
■ Background
Herpes Simplex Virus-1
Virus based transduction
DNA virus High infection efficiency Infect dividing and non-dividing cell Neurotropic virus Large transgene capacity ( up to ~150kb ) Long expression period High immunogenic and inflammatory response and necrosis
■ Background
Lentivirus:
Virus based transduction
RNA virus derived from HIV Can infect non-dividing cell Low Immunogenic Long expression period
Insertion mutation by random integration and oncogenicity Low virus titer
TALEN, CRISPR-Cas9…
■ Background
Gene therapy
Applications: Gene therapy can be applied in many diseases,
such as cancer, viral disease, genetic disease, et al. And mostly appliable to the single-gene disorders.
※ The first approved gene therapy case in the United States took place on 14 September 1990.
※ There are more than 2000 clinical trials have being launched in the past seven years.
Chemical modification can make the RNA be resistant to the nuclease cleavage.
Non-virus method
■ Background
Cyclodextrin polymer nanoparticles
First applied in 1999 Targeted: ligand Low toxicity Steric stabilization No measured innate immune responses when administered intravenously
……
■ Background
Gene therapy Definition: Gene therapy is the use of DNA as a drug to treat
disease by delivering therapeutic DNA into a patient's cells.
■ Background
Hybrid virus vector
HSV/AAV Ad/EBV HSV/EBV Ad/AAV Ad/retrovirus
Virus based transduction
■ Background
siRNA-based gene therapy
■ Background
Chemical modification
(AAV), Herpes Simplex Virus (HSV-1), et al.
Non-virus method: Cyclodextrin Polymers, Lipids, Peptide, Antibodies, Aptamers,
and small mwk.baidu.comlecules
■ Background
Adeno-associated virus (AAV)
DNA virus without pathogenicity Specific host range Infect dividing and non-dividing cell Low integration level, exist as episome in host cell No insertion mutation by random integration and oncogenicity (site-specific integrate into 19 chromosome) Long expression period No immunogenic and inflammatory response Small transgene capacity ( ~3kb ) Low virus titer (1012 VP/ml) Complex procedure and manipulation Host range limitation
99% knockdown of liver genes after a single 0.2 mg per kg dose in non-human primates, with the effect lasting nearly 7 weeks
■ Background
Conjugate delivery
Gene Therapy
■ Background
Human beings fight against all kind of diseases
Cancer CCVD: Cardio-Cerebrovascular Diseases
Viral Disease Genetic Disease: more than 2000 diseases CNS: Central Nervous System Disease Immune Diseases Diabetes ……
■ Background
( Jesse Gelsinger, 18 years old, died from severe immune response caused by adenovirus based gene therapy in 1999)
■ Background
Virus based transduction
■ Background
Adenovirus:
Virus based transduction
DNA virus High virus titer (up to 1014VP/ml) and high infection efficiency Wide host range and large transgene capacity ( ~37kb ) Infect dividing and non-dividing cell Low integration level, exist as episome in host cell No insertion mutation by random integration and oncogenicity Short expression period (5-20 days) Complex procedure and manipulation Potential immunogenic and inflammatory response
■ Background
Different delivery tools
Virus Vector: Retrovirus, Lentivirus, Adenovirus, Adeno-associated virus
■ Background
Different treatments to the diseases
Chemical drugs Surgery Physical Therapy Transplantation therapy Immunotherapy Regenerative medicine
Gene therapy
■ Background
Technical problem: Barriers to the target
DNA/RNA is unstable in the bloodstream, can be immunogenic and does not readily cross membranes to enter cells. Nuclease, renal filtration
Non-virus method
Can deliver both siRNA and plasmid DNA
■ Background
Liposome
Protect entrapped oligonucleotides from nuclease degradation and renal clearance Promote cellular uptake and endosomal escape They include the use of cationic or ionizable lipids, shielding lipids, cholesterol and targeting ligands
Retrovirus:
Virus based transduction
Widely studied RNA virus Wide host range Infect dividing cell and just infect once Integrate into host genome Low Immunogenic Long expression period Insertion mutation by random integration and oncogenicity Can’t infect non-dividing cell and low virus titer
Triantennary GalNAc–siRNA
Non-virus method
ASGPR, on hepatocytes
Both subcutaneous and intravenous administration of this conjugate revealed great accumulation of siRNA in the liver and improved knockdown of the target gene.
※ Using DNA to express functional protein ※ Using siRNA or shRNA to attenuate abnormal gene
expression in mRNA level ※ Genome editing to correct mutant gene sequence: ZFN,
Non-virus method
Can deliver both siRNA and plasmid DNA
■ Background
Conjugate delivery
Non-virus method
Dynamic PolyConjugates (DPC)
First reported in 2007 PEG: shielding effect GalNAc ligand was essential for both uptake by hepatocytes and in vivo silencing activity. Other targeting ligands has been explored, including peptides, antibodies, small molecules, glycans, lectins and nucleic acids.
■ Background
Herpes Simplex Virus-1
Virus based transduction
DNA virus High infection efficiency Infect dividing and non-dividing cell Neurotropic virus Large transgene capacity ( up to ~150kb ) Long expression period High immunogenic and inflammatory response and necrosis
■ Background
Lentivirus:
Virus based transduction
RNA virus derived from HIV Can infect non-dividing cell Low Immunogenic Long expression period
Insertion mutation by random integration and oncogenicity Low virus titer
TALEN, CRISPR-Cas9…
■ Background
Gene therapy
Applications: Gene therapy can be applied in many diseases,
such as cancer, viral disease, genetic disease, et al. And mostly appliable to the single-gene disorders.
※ The first approved gene therapy case in the United States took place on 14 September 1990.
※ There are more than 2000 clinical trials have being launched in the past seven years.
Chemical modification can make the RNA be resistant to the nuclease cleavage.
Non-virus method
■ Background
Cyclodextrin polymer nanoparticles
First applied in 1999 Targeted: ligand Low toxicity Steric stabilization No measured innate immune responses when administered intravenously
……
■ Background
Gene therapy Definition: Gene therapy is the use of DNA as a drug to treat
disease by delivering therapeutic DNA into a patient's cells.
■ Background
Hybrid virus vector
HSV/AAV Ad/EBV HSV/EBV Ad/AAV Ad/retrovirus
Virus based transduction
■ Background
siRNA-based gene therapy
■ Background
Chemical modification
(AAV), Herpes Simplex Virus (HSV-1), et al.
Non-virus method: Cyclodextrin Polymers, Lipids, Peptide, Antibodies, Aptamers,
and small mwk.baidu.comlecules
■ Background
Adeno-associated virus (AAV)
DNA virus without pathogenicity Specific host range Infect dividing and non-dividing cell Low integration level, exist as episome in host cell No insertion mutation by random integration and oncogenicity (site-specific integrate into 19 chromosome) Long expression period No immunogenic and inflammatory response Small transgene capacity ( ~3kb ) Low virus titer (1012 VP/ml) Complex procedure and manipulation Host range limitation
99% knockdown of liver genes after a single 0.2 mg per kg dose in non-human primates, with the effect lasting nearly 7 weeks
■ Background
Conjugate delivery
Gene Therapy
■ Background
Human beings fight against all kind of diseases
Cancer CCVD: Cardio-Cerebrovascular Diseases
Viral Disease Genetic Disease: more than 2000 diseases CNS: Central Nervous System Disease Immune Diseases Diabetes ……
■ Background
( Jesse Gelsinger, 18 years old, died from severe immune response caused by adenovirus based gene therapy in 1999)
■ Background
Virus based transduction
■ Background
Adenovirus:
Virus based transduction
DNA virus High virus titer (up to 1014VP/ml) and high infection efficiency Wide host range and large transgene capacity ( ~37kb ) Infect dividing and non-dividing cell Low integration level, exist as episome in host cell No insertion mutation by random integration and oncogenicity Short expression period (5-20 days) Complex procedure and manipulation Potential immunogenic and inflammatory response