Cell and Gene Therapy

Plasmid DNA and Viral Vector Applications and Discovery Services

Cell and gene therapies are transformative for the future of medicine and patient success, particularly for treating diseases that currently have limited or no current treatment options. They hold the potential for personalized (read more about plasmid DNA and viral vectors in precision medicine), durable, and even curative outcomes.

What exactly are cell and gene therapies? We are glad you asked!

Cell and gene therapies are incredibly advanced medical treatments designed to treat or potentially cure diseases by addressing the underlying genetic causes or by enhancing cellular functions by introducing or inducing optimized cellular behavior. This medical advancement has many applications and different types of patient outcomes because of how limitless the effects of gene editing can have on patients.

Cell Therapy

Regenerative Medicine: Cell therapy is used to replace or repair damaged or diseased cells and tissues from that may occur from inherited diseases, serious injury, or infectious diseases. For example, stem cell therapies are applied in regenerative medicine to treat conditions like heart disease, spinal cord injuries, and certain types of blood disorders. Read more about plasmid DNA and viral vectors in regenerative medicine and stem cell therapies.
Immunotherapy: Cells such as T-cells are modified or engineered to target and destroy cancer cells. CAR-T cell therapy is a key example, where a patient’s own T-cells are engineered to recognize and attack cancer cells. Read more about CAR-T and lentivirus applications further down.
Tissue Engineering: Cells can be used to create tissues or even organs that can be transplanted into patients. This approach could address organ shortages met through bioprinting or treat conditions like severe burns or degenerative diseases.

Gene Therapy

Correcting Genetic Disorders: Gene therapy involves introducing, removing, adding, silencing or altering in other ways, genetic material within a patient’s cells to treat diseases caused by genetic mutations. This is particularly applicable for monogenic disorders like cystic fibrosis, hemophilia, or sickle cell anemia.
Oncology: Gene therapy can be used to introduce new genes, through viral and non-viral vectors, into cancer cells to either kill them or make them more susceptible to other treatments like chemotherapy or radiation. Learn more about plasmid DNA and viral vectors in oncology.
Vaccination and Preventive Therapies: Genetic material can be introduced into the body to stimulate an immune response, similar to vaccines. Learn more about plasmid DNA and viral vectors in vaccine development.

The real question is how cell and gene therapies can help patients recover from disease, improve the quality of their life, and ultimately have better outcomes. The ultimate patient outcome depends on what specific illness, ailment, or preventative measure, the cell or gene therapy is targeting.

Curative Potential: Unlike traditional treatments, cell and gene therapy often aim for a one-time or limited-duration treatment that could result in a long-term cure. For example, patients with inherited genetic diseases may have their faulty genes corrected, potentially curing the condition.
Target Cells: Used to transduce host T cells. Once modified, these T cells can recognize and attack cancer cells.
Application: Involves extracting T cells from a patient, modifying them with the CAR-T lentivirus, and then reintroducing them into the patient to fight cancer.

Plasmid DNA in Cell and Gene Therapy Molecular Cloning and Mutagensis

These molecular biology techniques, molecular cloning and mutagenesis, are integral to creating and modifying the genetic material in plasmid DNA used in these therapies for the following reasons:

Gene Delivery: In gene therapy, plasmid DNA is used to deliver therapeutic genes into target cells. These genes can correct a defective gene, produce a therapeutic protein, or regulate gene expression in the patient’s cells.
Viral Vector Production: Plasmids are used to produce viral vectors (such as AAV or lentivirus) that are widely used to deliver therapeutic genes in gene therapy. Plasmid DNA is transfected into producer cells, which are then used to package viral vectors containing the therapeutic gene.

Molecular Cloning in Cell and Gene Therapy:

Therapeutic Gene Cloning: Molecular cloning allows scientists to insert a therapeutic gene (or other genetic elements) into a plasmid vector. This cloned plasmid can then be amplified in bacteria and purified for further use in therapy.
Optimization of Gene Expression: Molecular cloning techniques are used to fine tune plasmids to optimize the expression of therapeutic genes, making sure they are efficiently expressed in the target cells. Researchers can clone different promoters, enhancers, or regulatory sequences into plasmids to achieve the desired expression levels.
Customizing Plasmid Vectors: Researchers can clone multiple genetic elements into plasmids, including therapeutic genes, reporter genes (such as GFP), and selection markers, allowing for tailored solutions in gene therapy. These customized plasmids are often used in viral vector production for gene delivery.

Mutagenesis in Cell and Gene Therapy

Engineering Therapeutic Proteins: Mutagenesis is used to alter specific amino acids in therapeutic proteins, improving their stability, efficacy, or reducing immunogenicity. For instance, mutagenesis can optimize antibodies or enzymes used in gene therapy.
Studying Disease Models: By introducing specific mutations into genes (via plasmid DNA), researchers can create models of genetic diseases, allowing them to study disease mechanisms and test potential therapies.
Optimizing Gene Therapy Vectors: Mutagenesis can be applied to optimize the components of gene therapy vectors, such as improving viral capsid proteins for better targeting and delivery, or modifying regulatory elements to optomize gene expression.

Plasmid DNA Based Cell and Gene Therapy Companies and Therapeutic Candidates

Pfizer: Comirnaty: The COVID-19 mRNA vaccine used plasmid DNA during the manufacturing process to generate the mRNA template. Although this isn’t a direct gene therapy, plasmid DNA is crucial in the production process.
Poseida Therapeutics: P-BCMA-101: A CAR-T cell therapy targeting multiple myeloma. Poseida uses a proprietary piggyBac transposon system, where plasmid DNA is used to modify T-cells without viral vectors, making this a non-viral gene therapy approach.
Genprex: Reqorsa: A gene therapy candidate for non-small cell lung cancer (NSCLC). It uses a plasmid DNA-based vector to deliver the TUSC2 gene (a tumor suppressor gene) directly to cancer cells, which helps re-activate the body’s natural tumor suppression pathways.
Genprex: Reqorsa: A gene therapy candidate for non-small cell lung cancer. It uses a plasmid DNA-based vector to deliver the TUSC2 gene (a tumor suppressor gene) directly to cancer cells, which helps re-activate the body’s natural tumor suppression pathways.

Viral Vectors in Cell and Gene Therapy Overview

AAV, adenoviruses, lentiviruses, and retroviruses are commonly used in cell and gene therapy research and production as vectors for delivering therapeutic genes into target cells.

Adeno-associated viral vectors belong to the adenoviral family yet have their own unique characteristics and advantages particularly in cell and gene therapy research and applications in neurological disorders, rare genetic disorders, oncology, ophthalmology, and cardiovascular areas because of their cell tropism, long term expression, and low immunogenicity.

Adenoviral vectors play a crucial role in cell and gene therapy, particularly in oncology and vaccine development. While their strong immunogenicity and transient expression present challenges, advances in vector design and engineering are enhancing their safety, specificity, and therapeutic utility.

Lentiviral vectors and retroviral vectors belong to the family of retroviruses, but lentiviruses are a subset that can infect both dividing and non-dividing cells, making them particularly versatile. Learn about the differences in lentiviral vs retroviral vectors.

AAV in Cell and Gene Therapy

Low Immunogenicity: Compared to adenovirus, AAV triggers minimal immune response and lower risks of inflammatory reactions and antibody responses.
Safety Profiles: AAV is non-pathogenic in humans and does not integrate into the host cell genome like retrovirus or lentivirus.
Board Cell Tropism: AAV serotypes allow for optimized cell transduction and efficient payload delivery into the host cell’s specific tissue unlike retrovirus and lentivirus which are tissue agnostic.

Adenovirus in Cell and Gene Therapy

Targeting and Specificity: Unlike retrovirus and lentivirus, adenoviral vectors have engineering fiber proteins or other capsid components to improve cell targeting and reduce off-target effects.
Improved Safety Profiles: Development of helper-dependent (HDAd) or gutted adenoviruses, which lack all viral genes, reducing immunogenicity and prolonging transgene expression. Learn more about helper-dependent adenovirus in adenovirus packaging capacity.
Novel Serotypes: Use of rare or non-human adenovirus serotypes to overcome pre-existing immunity and expand therapeutic potential.

Lentivirus in Cell and Gene Therapy

Gene Delivery to Non-Dividing Cells: Unlike most retroviruses, lentiviruses can infect non-dividing cells such as neurons, muscle cells, and liver cells. This makes them highly useful for delivering therapeutic genes to a wide range of tissues in vivo.
Long-Term Gene Expression: Lentiviruses integrate their genetic material into the host cell’s genome, allowing for stable, long-term expression of therapeutic genes. This is essential for treatments that require sustained gene expression, such as in chronic diseases or for producing therapeutic proteins continuously.
Ex-Vivo Gene Therapy: Lentiviral vectors are often used in ex-vivo gene therapy. In this approach, patient cells (e.g., hematopoietic stem cells or T-cells) are removed, genetically modified using lentiviruses to introduce therapeutic genes, and then returned to the patient. This is the method used in CAR-T cell therapy, where T-cells are modified to express chimeric antigen receptors to attack cancer cells.

Retrovirus in Cell and Gene Therapy

Gene Delivery to Dividing Cells: Retroviruses can integrate their genetic material into the DNA of dividing cells, making them useful in cases where the target cells are rapidly proliferating, such as in some cancers or certain types of stem cell therapies.
Ex-Vivo Gene Therapy for Hematopoietic Stem Cells: Retroviruses are commonly used in ex-vivo gene therapy approaches where hematopoietic stem cells are removed from the patient, modified with a therapeutic gene, and then transplanted back into the patient. These stem cells can then give rise to genetically corrected blood cells, offering potential treatments for blood disorders.
Oncolytic Retrovirus Therapy: Retroviruses are also being engineered to target and kill cancer cells directly. This approach, called oncolytic viral therapy, involves using retroviruses that preferentially infect and destroy tumor cells, while sparing normal cells.

Viral Vector Cell and Gene Therapy Applications

AAV Cell and Gene Therapy Applications

Neurological Disorders: AAV has excellent success in neurological research in areas for genetic brain disorders, Parkinson’s, Alzheimer’s, spinal muscular atrophy, and CNS disorders.
Rare Genetic Disorders: While rare disease was a significant challenge for other modalities for many years, AAV vectors have proved to be effective in treating hemophilia, lysosomal storage disorders, muscular dystrophy, and retinal disease.
Cardiovascular Treatments: AAV’s cardiovascular tropism has allowed it to be applied in gene therapies for inherited heart conditions, cardiac genetic mutations, and potential treatments for heart failure.

Adenovirus Cell and Gene Therapy Applications

Cancer Therapy: Oncolytic adenoviruses are engineered to selectively replicate in and kill cancer cells. Their gene delivery allows the expression of therapeutic genes such as tumor suppressors or pro-drug activating enzymes.
Vaccine Development: Used as vectors for vaccine antigens, including vaccines for SARS-CoV-2.
Gene Therapy: Adenoviruses deliver functional genes to patients in order to treat genetic disorders such as cystic fibrosis.

Lentivirus Cell and Gene Therapy Applications

CAR-T Cell Therapy: Lentiviral vectors are used to modify T-cells in patients with certain types of cancers (e.g., leukemia, lymphoma) to target and kill tumor cells.
Treatment of Genetic Diseases: Lentivirus-mediated gene therapy is used to treat genetic disorders like β-thalassemia and sickle cell anemia by delivering functional copies of defective genes into hematopoietic stem cells.
Neurodegenerative Disease Research: Lentiviruses are also used in research on neurodegenerative diseases such as Parkinson’s and Huntington’s disease. They can deliver therapeutic genes into neurons, offering potential treatments for these conditions.

Retrovirus Cell and Gene Therapy Applications

X-linked Severe Combined Immunodeficiency (SCID-X1): One of the early successes of retrovirus mediated gene therapy was in treating SCID-X1, a rare immune disorder. By using retroviruses to deliver a functional copy of the gene responsible for SCID, researchers successfully restored immune function in patients.
Gene Therapy for Cancer: Retroviruses have been used to introduce therapeutic genes into cancer cells, including suicide genes that make cancer cells more susceptible to chemotherapy or other treatments.

AAV Based Cell and Gene Therapy Companies and Therapeutic Candidates

AveXis/Novartis:
- Zolgensma: AAV9 treatment for the treatment of spinal muscular atrophy in children.
Spark Therapeutics:
- Luxturna: AAV2 vectors used to treated inherited retinal dystrophy.
Wet AMD (Rare Genetic Disease):
- RGX-314: ABBV-RGX-314 is being developed as a novel, one-time subretinal treatment that includes the NAV® AAV8 vector containing a gene encoding for a monoclonal antibody fragment.

Adenovirus Based Cell and Gene Therapy Companies and Therapeutic Candidates

Amgen:
- Imlygic: Oncolytic adenovirus engineered to selectively replicate in and lyse tumor cells, with an added GM-CSF gene to boost anti-tumor immunity.
Cend Therapeutics:
- CendR-01: Modified adenovirus targeting tumor microenvironments to enhance drug delivery.
CG Oncology:
- CG0070: Oncolytic adenovirus that selectively replicates in tumor cells with defective retinoblastoma (Rb) pathways and expresses GM-CSF for immune stimulation.

Lentivirus Based Cell and Gene Therapy Companies and Therapeutic Candidates

bluebird bio:
- Zynteglo: An approved gene therapy for beta-thalassemia, a rare genetic blood disorder. Lentiviral vectors are used to deliver a functional copy of the β-globin gene into hematopoietic stem cells.
- Skysona: A lentiviral gene therapy for cerebral adrenoleukodystrophy (CALD), a neurodegenerative disorder. It uses lentivirus to deliver a functional gene into the patient’s HSCs.
Kite Pharma:
- Yescarta: A CAR-T cell therapy for treating large B-cell lymphoma. Lentiviral vectors are used to modify the patient’s T-cells to express chimeric antigen receptors targeting cancer cells.
Orchard Therapeutics:
- Libmeldy: Approved gene therapy for metachromatic leukodystrophy (MLD). It uses lentiviral vectors to deliver a functional copy of the ARSA gene into HSCs.
Novartis:
- Kymriah: A CAR-T cell therapy for treating B-cell acute lymphoblastic leukemia (ALL) and large B-cell lymphoma. It uses lentiviral vectors to engineer the patient’s T-cells.

Retrovirus Based Cell and Gene Therapy Companies and Therapeutic Candidates

GSK:
- Strimvelis: A retroviral gene therapy for adenosine deaminase severe combined immunodeficiency (ADA-SCID). Retroviral vectors are used to insert a functioning ADA gene into the patient’s hematopoietic stem cells.
Rocket Pharmaceuticals:
- RP-L201: A retroviral gene therapy for leukocyte adhesion deficiency-I (LAD-I), using a gamma-retroviral vector to insert a functioning gene into the patient’s stem cells.
Mustang Bio:
- MB-107: Retrovirus-based gene therapy for X-linked severe combined immunodeficiency (SCID-X1), also known as “bubble boy syndrome.” It uses a retroviral vector to insert a corrected version of the IL2RG gene into hematopoietic stem cells.

Molecular Cloning, Mutagenesis, AAV Packaging, Adenovirus Packaging, Lentivirus Packaging, Retrovirus Packaging, and CRISPR-Cas9 Services for Cell and Gene Therapy R&D

Our laboratory specializes in plasmid DNA and viral vector services for cell and gene therapy discovery phase research including:

If you have a question about your plasmid DNA and viral vector based cell and gene therapy research, contact our team.