The Evolution of Genetic Engineering in Medical Genetics

Human Genome project (1990 - 2003) and today contrast

Def: Intl scientific effort that mapped all 3 billion base pairs

Cost: $2.7 billion ($9 per base pair)

Relevance: Establish foundation for modern genetic medicine and revealed approx 20,000-25,000 human genes

Today

Today, whole genome seq costs under $1,000 and can be completed <24 hours.

Reading our genetic code → write/edit it opening medical interventions

Tool for genetic modification: CRISPR-Cas 9 (Discovery, how it works, medical applications)

CRISPR discovery :

• Identified in bacteria as adaptive immune system agains viral infection → adapted into gene-editing tool

• Emmanuelle Charpentier and Jennifer Doudna received 2020 Nobel prize for this

How CRISPR works

• Functions as molecular scissors guided by RNA to target specific DNA sequences with precision

• Cas9 enzyme cuts DNA at targeted location allowing for -, +, or alteration of genetic material

Medical application:

• Clinical trials for genetic blood disorders (sickle cell disease, β- thalassemia) , certain cancers, and inherited blindness.

• Potential to treat thousands of single-gene disorders

Gene therapy timeline (incl cost) :

Which treatment was the 1st approved gene therapy?

Which treatment was 1st FDA approved gene therapy for inherited disease?

Which treatment for a leading cause of infant mortality?

Which treatment genetically modified immune cell to fight 80% blood cancer?

First approved gene therapy in western world (2012)

• Glybera for lipoprotein lipase deficiency approved.

• Cost $1 million per treatment.

First FDA approved gene therapy for inherited disease (2017)

• Luxturna

• Treatment restored functional vision in patients with previously incurable form of blindness (RPE65 mutation-associated retinal dystrophy).

Treatment for a leading genetic cause of infant mortality - spinal muscular atrophy (2019)

• Zolgensma

• Single-dose treatment for spinal muscular atrophy, as 1 time infusion replaces the function of missing or nonworking SMN1 gene

• Cost $2.1 million.

Genetically modified immune cells to fight 80% blood cancer (2017-Present)

• CAR-T Cell Therapies

• Kymriah and Yescarta used genetically modified immune cells to fight cancer. Drugs achieve remission rates of 80% in some previously untreatable blood cancers.

How has genetic testing revolutionized medicine, and what is the impact of precision medicine? Provide examples.

Genetic testing revolution

1. Newborn screening identifies conditions needing immediate intervention

2. Carrier testing determines recessive disease risk before conception

3. Predictive testing assess future disease susceptibility

4. Pharmacogenomics guides medication selection and dosing

Precision Medicine Impact

• FDA approved over 75 drugs with pharmacogenomic biomarkers→ allows treatments tailored to patients' genetic profiles

• Oncology transformed by targeted therapies attack cancer cells based on specific genetic mutations not just tissue type.

• From "one- size-fits-all" to "the right drug for the right patient at the right time."

Ethical consideration in human genetic engineering

1. Somatic vs germline editing

• Somatic affect only patient while germline changes can be inherited by future generation

• Most scientist advocate moratorium (pause) on clinical germline editing until safety & ethical frameworks established (WHO

The World Health Organization has established a global registry to track all human genome editing research and has called for regulatory authorities to approve no clinical applications of human germline genome editing until its implications have been properly considered.

2. Designer baby debate

• 2018 birth of first genetically edited babies in China (modified to resist HIV) sparked outrage so scientific community need to be careful on therapeutic intervention vs enhancement

3. Access and equity

• Genetic therapies often cost millions per treatment so not genetic divide between who can and cant afford

Genetic engineering for rare disease treatment:

How many ppl does rare disease affect and how much is genetic?

How many known rare diseases and how many have no FDA approved treatments?

Average cost of treatments?

Rare disease affects: 400 million ppl worldwide with 80% have genetic origins

Known rare diseases: 7000+ with approx 95% have no FDA approved treatments

Average gene therapy cost: $1-2 million

Stem cells technology and genetic engineering possibilities and example IRL

• Gene-corrected patient-specific stem cells for autologous transplantation

• Engineered organdies for disease modelling and drug screening

• 3D bioprinting of genetically optimized tissues and organs

• Reprogramming immune cells to fight cancer and autoimmune diseases

In 2023, the first patient received a kidney transplant developed from genetically modified pig cells, potentially solving the organ shortage crisis.

Challenges and limitations in genetic engineering applications

Technical challenges

1. Off-target effects and mosaicism in gene editing

2. Unintended Immune reactions to delivery vectors and edited cells

3. Difficulty targeting specific tissues and organs

4. Complex genetics of multigenic disorders

Economic barriers

1. Multimillion-dollar treatment costs

2. Limited incentives for ultra-rare conditions

3. Healthcare system unpreparedness for one-time curative therapies

4. Global inequities in access to genetic technologies

Regulatory hurdles

1. Varying intl reg frameworks

2. Long-term safety monitoring challenge

3. Ethical oversight of rapidly evolving technologies

4. Balancing innovation with precaution

The Future of Medical Genetics: Possibilities and Responsibilities

Possibilities

1. More precise genetic modifications by base editing and prime editing

2. In vivo gene editing directly to affected tissues

3. Epigenetic editing to modify gene expression without changing DNA seq

4. Artificial chromosomes as genetic medicine delivery platforms

Responsibilites

1. Inclusive governance with diverse stakeholder input

2. Equitable access to genetic tech globally

3. Transparent research and regulatory process

4. Continuous ethical reassessment as capabilities evolve

What is gene cloning (Def, Uses, What could this mean for birthing/ cancer)

Def: Gene cloning is process of creating multiple copies of specific DNA segment.

Uses:

1. Molecular scissors: Restriction enzymes to cut DNA at precise sites

2. Bacterial vectors: DNA fragments inserted into plasmids (small circular DNA vectors)

3. Replication: Recombinant DNA molecules replicate inside host cells producing many gene copies

Side effects

1. Birthing: DNA sequence changes so can’t reproduce

2. Cancer: Wrong DNA sequence → Wrong protein → May result in cancer

DNA cloning steps + importance of steps

1. Cutting: Restriction enzymes precisely cut target gene and plasmid vector

2. Ligation: DNA ligase ‘glues’ gene into plasmid forming recombinant DNA

3. Transformation: Recombinant plasmid introduced into bacteria

4. Selection: Antibiotic selection isolates bacteria with cloned gene

5. Amplification: Bacteria replicate producing lots of cloned DNA

Importance of steps : Ensures only bacteria with desired gene survive and multiply, yielding pure samples of the cloned DNA.

Polymerase Chain Reaction /PCR (Definition, Complement cloning, Essential tool for)

Def: PCR is a revolutionary technique that amplifies specific DNA segments rapidly in vitro → creates millions of copy from even tiny DNA sample within hrs

Cloning: PCR complements traditional cloning by ↑speed gene isolation and allowing quick analysis

Essential tool for: DNA sequencing, targeted mutagenesis, and genotyping, (understanding genetic variations and disease mechs in medical research)

Medical Breakthroughs Powered by Gene Cloning (Examples)

1. Human insulin

• Recombinant human insulin purer and safer than animal derived ones for diabetes

2. Clotting factos

• Cloned genes produce clotting factors → help haemophilia patients

3. Growth hormone (GH)

• Human GH & various cytokines made via cloned genes for regulating immune response and addressing deficiencies

These recombinant proteins significantly reduce the risks of allergic reactions and disease transmission more than before

Gene cloning in disease Diagnosis and forensics : Uses in disease research, forensic science and personalised medicine

• Disease Research: Enables detailed gene function studies and mutation analysis to uncover disease mechanisms.

• Forensic Science: Used extensively for suspect identification, paternity testing, and establishing familial relationships in criminal investigations.

• Personalized Medicine: Genotyping techniques reveal individual genetic variations, guiding tailored treatments and drug dosages.

advances in molecular cloning techniques ( synthetic gene synthesis vs multi-fragment assembly, swappable gene cassettes)

1. Synthetic gene synthesis: Direct chem gene synthesis error-free sequences, bypassing traditional cloning steps for custom DNA constructs.

2. Multifragment assembly: Seamless stitching of multiple DNA fragments allowing rapid construction of complex gene circuits

3. Swappable Gene Cassettes: Modular gene cassettes allow increased flexibility for gene function studies and protein expression engineering

These advancements ensure higher fidelity, faster turnaround, and greater versatility in genetic engineering applications.

Molecular Cloning Transforming Medical Microbiology

1. Therapeutic proteins allow production of recombinant cytokines and antimicrobial peptides to bolster immune response

2. Allows rapid new vaccine development and gene therapy vectors for new pathogens

3. Epidemiological tracking of polymicrobial infections to understand spread and evolution

4. Pathogen Identification overcoming limitation of traditional culture based methods

Ethical and Technical challenges of gene cloning

Technical Hurdles

• Cloning complex human genes as size and intricacy of human genome

• Quality of starting material (RNA/DNA) critical for cloning success so need meticulous sample prep

• Continuous improvement needed to reduce errors and ↑efficiency of cloning especially for large-scale

Ethical Dilemmas

• Human germline modification can impact future generation

• Societal debates around responsible use of genetic tech and unintended consequences

• Equitable access to gene based therapies avoid health disparities

Future of gene cloning and DNA analysis in medicine

Gene cloning and DNA analysis wil allow…

1. Accelerated development of drugs and vaccine bringing new treatments to patients sooner

2. Personalised therapies and effective therapeutic intervention by using CRISPR and advanced gene editing

3. Expanded roles in diagnosis , synthetic bio, regenerative medicine