Difference Between Lentiviral Vectors, AAV Vectors and Plasmid Vectors

I. Introduction to Lentiviral Vectors

Lentiviral vectors are viral gene therapy vectors derived from human immunodeficiency virus-1 (HIV-1). Unlike typical retroviral vectors, they can infect both dividing and non-dividing cells. Lentiviral vectors are used for direct infection of host cells, where the target gene is reverse-transcribed and integrated into the host genome upon entry, enabling high-level expression of the effector molecule.

II. Introduction to Plasmid Vectors

A plasmid is a small, extrachromosomal DNA molecule that can replicate independently from the cell’s chromosomal or nucleoid DNA. Generally, the presence or absence of a plasmid does not determine the survival of the host cell. It is the most common vector in genetic engineering.

So, what are the differences between lentiviral vectors and plasmid vectors?

• Different Entry Methods into Cells: Plasmids enter target cells via transfection, while viruses enter via infection, which is more specific and has higher efficiency.
• Different Forms of Carried Sequences and Their Functional Mechanisms: Plasmid DNA carries DNA sequences that typically need to enter the cell nucleus to exert their function. Viral vectors can carry either DNA or RNA sequences; RNA sequences usually do not require nuclear entry to function, and for host genome homologous recombination, viral vectors have higher efficiency.
• Different Packaging and Purification Methods: Plasmids are typically amplified in prokaryotic cells and then purified. Viral vectors are usually packaged in eukaryotic cells, expanded through stepwise culture, and then obtained by lysing cells/collecting supernatant followed by purification.
• Different Storage Methods: Plasmids are generally very stable and not sensitive to temperature. Viruses are different; they usually require low-temperature frozen storage, are highly sensitive to temperature, and repeated freeze-thaw cycles greatly affect their viability.
• Different Applications in Vaccines: Plasmid DNA as a carrier for DNA vaccines typically has low immunogenicity and requires liposomes or electroporation guns to enhance DNA entry into cells. Viral vector vaccines have good immunogenicity, often easily inducing strong antibody and T-cell responses, but are often limited by pre-existing immunity.

III. Introduction to AAV Vectors and Comparisons

To provide a more comprehensive overview of common gene therapy vectors, it’s useful to include adeno-associated virus (AAV) vectors, which are widely used alongside lentiviral and plasmid vectors. AAV vectors are non-pathogenic, single-stranded DNA viruses derived from parvoviruses, commonly employed in gene therapy due to their low immunogenicity, broad tissue tropism (via different serotypes), and ability to achieve long-term transgene expression. Unlike integrating vectors, AAV genomes typically remain episomal (non-integrating) in non-dividing cells, with low-frequency random integration possible. They have a packaging limit of ~4.7-5 kb, making them suitable for smaller genes but requiring strategies like dual-vector systems for larger ones. AAVs primarily infect non-dividing cells efficiently, similar to lentiviruses, but do not integrate as readily, reducing insertional mutagenesis risks.

Key Differences Among Lentiviral, Plasmid, and AAV Vectors:

• Integration and Persistence: Lentiviral vectors integrate into the host genome for stable, long-term expression (heritable in dividing cells) but carry mutagenesis risks. Plasmids are episomal, non-integrating, and often transient unless engineered otherwise. AAV vectors are mostly episomal, providing long-term expression in non-dividing cells without integration, though low-level integration can occur.
• Cell Tropism and Infection Efficiency: Lentiviruses transduce both dividing and non-dividing cells (e.g., neurons, stem cells) with high efficiency. Plasmids rely on transfection (less efficient, especially in primary cells). AAVs efficiently transduce non-dividing cells with serotype-specific tropism (e.g., AAV9 for CNS, AAV8 for liver).
• Packaging Capacity: Lentiviruses (~8-10 kb), plasmids (variable, up to 20 kb or more but transfection-limited), AAVs (~4.7 kb single-stranded, ~2.4 kb for self-complementary).
• Immune Response: Lentiviruses may trigger stronger responses due to HIV-1 origins (mitigated by pseudotyping). Plasmids have low immunogenicity but poor delivery. AAVs have mild responses but pre-existing immunity in ~30-50% of humans can neutralize them.
• Applications: Lentiviruses for ex vivo CAR-T or stable integration; plasmids for transient expression or vaccines; AAVs for in vivo gene therapy (e.g., Luxturna for retinal dystrophy, Zolgensma for SMA).
• Safety and Manufacturing: All require GMP-grade production; AAVs often face capsid impurity issues (e.g., empty/full ratios), while lentiviruses need biosafety level 2 handling. Plasmids are simplest to produce but least efficient in vivo.

Summary Comparison Table