Adenoviral Producer Cells

Imre Kovesdi,Susan J Hedley

doi:10.3390/v2081681

Abstract

Adenovirus (Ad) vectors, in particular those of the serotype 5, are highly attractive for a wide range of gene therapy, vaccine and virotherapy applications (as discussed in further detail in this issue). Wild type Ad5 virus can replicate in numerous tissue types but to use Ad vectors for therapeutic purposes the viral genome requires modification. In particular, if the viral genome is modified in such a way that the viral life cycle is interfered with, a specific producer cell line is required to provide trans-complementation to overcome the modification and allow viral production. This can occur in two ways; use of a producer cell line that contains specific adenoviral sequences incorporated into the cell genome to trans-complement, or use of a producer cell line that naturally complements for the modified Ad vector genome. This review concentrates on producer cell lines that complement non-replicating adenoviral vectors, starting with the historical HEK293 cell line developed in 1977 for first generation Ad vectors. In addition the problem of replication-competent adenovirus (RCA) contamination in viral preparations from HEK293 cells is addressed leading to the development of alternate cell lines. Furthermore novel cell lines for more complex Ad vectors and alternate serotype Ad vectors are discussed.

Highlights

Producer cells for non-replicating first generation Ad vectorsMany therapeutic Ad5 vectors are non-replicating (or replication deficient) whereby the genome is deleted in the E1 region often in combination with the E3 region to provide space for alternate gene expression cassettes (first generation Ad vectors, see Figure 1)
Adenovirus (Ad) vectors, in particular those of the serotype 5, are highly attractive for a wide range of gene therapy, vaccine and virotherapy applications
When the E1 region is replaced with an expression cassette to produce the gene product that is useful in therapy, such as suicide genes, antigens and antibodies, a producer cell line containing adenovirus E1 sequences is required to complement for this region

Summary

Producer cells for non-replicating first generation Ad vectors

Many therapeutic Ad5 vectors are non-replicating (or replication deficient) whereby the genome is deleted in the E1 region often in combination with the E3 region to provide space for alternate gene expression cassettes (first generation Ad vectors, see Figure 1). Due to the RCA issue and potential licensing issue several other laboratories have developed E1 complementary cells that contain smaller adenoviral E1 DNA segments than HEK293 cells and can propagate vectors to high titers similar to those seen on HEK293 [8,9,10,11] (Table 1) All of these cell lines have reduced or eliminated sequence homology between cellular DNA and viral genome to prevent the opportunity of the double homologous recombination crossover occurring. The future for new cell line development is the incorporation of advantageous genes into the genome to improve productivity, adaptability and robustness as described for the 293-VLI model

Producer cells for non-replicating second generation Ad vectors

Producer cells for high capacity Ad vectors

Producer cell lines for novel Ad vector serotypes

Conclusions