Gene Therapy and Drug Discovery Notes

Subjects Covered by the Class

• Part 1: History and Modern Drug Discovery Process

  • Brief history of drug discovery and diseases
  • Modern drug discovery and development process
  • Recombinant DNA technology:
  • Gene cloning
  • Creation of transgenic organisms

• Part 2: Vaccine as Biological Therapeutics

  • History of vaccine development
  • Immunity and mechanisms of self-protection against pathogens
  • Principles of vaccine-mediated immunization
  • Common forms of vaccines and their preparation
  • Available vaccines: case studies

• Part 3: Protein- and Peptide-Based Therapeutics

  • Chemical and biological basis of protein and peptide drugs
  • Identifying a therapeutic protein
  • Preparation, production, analysis, and metabolism of protein drugs
  • Formulation considerations for protein drugs including PEGylation
  • Protein drugs available for treatment: case studies

• Part 4: Antibody-Based Therapeutics

  • Fundamentals of antibody-based drugs and immunotherapy
  • Development of polyclonal and monoclonal antibodies
  • Humanized and recombinant antibodies
  • Antibody drug discovery and conjugates
  • Antibodies currently used in clinic: case studies

• Part 5: Gene Therapeutics

  • Definition of gene, genome, and genetic diseases
  • Gene replacement therapy
  • Gene therapy through genomic editing to correct genetic defects
  • Gene therapy-based treatment case studies

• Part 6: Oligo- and RNA-Based Therapeutics

  • Applications of oligonucleotide-based therapeutics
  • RNA interference and siRNA
  • Vehicles for delivery of oligo and RNA
  • RNA-based drugs for disease treatment: case studies

• Part 7: Regenerative Medicine and Cell-Based Therapeutics

  • Introduction to cell-based therapy
  • Bone marrow transplantation and stem cells
  • Genetically modified cells e.g., CAR-T cells
  • Case studies of cell-based therapeutics for disease treatment

Gene, Genome, and Genetic Diseases

• Definition of a Gene

  • Genetic information unit: DNA sequence specifying polypeptide or RNA
  • Located at specific loci on chromosomes
  • Entire genetic information of an organism: genome (e.g., human genome)

• Composition of Human Genome

  • Approximately 3.2 billion base pairs (bp); includes 16,569 mitochondrial bp
  • Roughly 20,000 protein-coding genes (~1.5% of genome)
  • 23,500 RNA coding genes (tRNAs, rRNAs, etc.)
  • Approximately 15,000 pseudogenes identified
  • 98% of genome comprises noncoding DNA including introns and regulatory sequences
  • Genetic variation frequency: ~1 in 1,000 bp; indicating 99.9% genetic similarity

Genetic Diseases

• Definition

  • Genetic diseases arise from gene mutations or variations
  • Can be inherited and passed through families
  • Majority arise from complex interactions of multiple genes and environmental factors

• Types of Genetic Abnormalities

  • Macro: Chromosomal mutations (structural and number changes)
  • Micro: Point mutations (alterations in gene sequence)

Mechanisms of Mutation

• Mutation Rate

  • E. coli: 1 in 10^8 nt; implications for humans calculated at ~30 nt/cycle
  • Influences: DNA polymerase errors, environmental factors

• Types of Genetic Changes

  • Classifications: Deletion, insertion, substitution (point mutations)
  • Mechanism of chromosome mutations involves transposition and transposons discovered by Dr. Barbara McClintock

• Pyrimidine Dimerization

  • Occurs from UV light exposure; causes mutations and skin conditions
  • Importance of nucleotide excision repair mechanisms for disease prevention

Gene Therapy and Genome Editing

• Gene Therapy Goals

  • Introduce functional genes into cells to treat or prevent diseases
  • Techniques include gene replacement and genomic editing

• Genome Editing Technologies

  • Utilizes nucleases for specific DNA alteration: ZFN, TALEN, CRISPR/Cas9
  • Mechanisms involve homology-directed repair (HDR) and non-homologous end joining (NHEJ)

CRISPR/Cas9 System

• Structure and Function

  • Gapped RNA sequence guides Cas9 to DNA target
  • PAM sequence (e.g., 5'-NGG-3') is critical for target recognition

• Applications

  • Successful editing in various organisms; potential for treating a range of diseases

• Challenges

  • Off-target effects leading to unintended mutations
  • Need for efficient delivery systems for therapeutic application

Conclusion

• Gene therapy and genome editing continue to advance medicine with prospects for treating genetic conditions through targeted genetic interventions.