Nucleic-acid- and cell-based therapeutics - Pharmaceutical Biotechnology: Concepts and Aapplications - page 433

Binding of the monoclonal antibody to the protein A domain would ensue and the immobilized
monoclonal antibody would dictate the cell type targeted.
Initial studies using this system have proved encouraging. The altered virus (without associated mon-
oclonal antibody) failed to infect a wide variety of human cell lines. By initially incubating with mono-
clonal antibody of the appropriate specifi city, however, the viral particles were capable of effi ciently
transducing cells expressing surface receptors such as CD4, CD33 and human leukocyte antigen.
A number of other issues must now be addressed including determining if the IgG-protein A af-
fi nity is suffi ciently high to keep the antibody associated with the virus in vivo . The full potential of
this approach will also require more detailed characterization of surface markers uniquely associated
with different cell types. However, the approach exemplifi es the types of technical innovation now
being introduced that will make second-generation vectors more suited to their role in gene therapy.

14.3.3 Manufacture of viral vectors

Viral vector manufacture for therapeutic purposes involves initial viral propagation in appropri-
ate animal cell lines, viral recovery, concentration, purifi cation and formulation. A generalized
manufacturing scenario for adenoviral-based vectors is outlined in Figure 14.7. The manufacture
of alternative viral vectors likely follows a substantially similar approach.
Master and working banks of both the viral vector and the animal cell line will have been con-
structed during the drug development process (see Chapter 4). Manufacture of a batch of vector,
therefore, will be initiated by the culture of packing cells in suitable animal cell bioreactors. The

Seed vith vector

Packing cell culture
(animal cell bioreactor)

Vector propigation

Cell lysis
(homogenization)

Harvest of infected cells
(Filtration/ centrifugation)

Clarification
(Filtration)

DNA digestion

Inactivation of potential
contaminant viruses
(Solvent/detergent)

Concentration if required
(Ultrafiltration)

Chromatographic purification
(Ion exchange & gel filtration)

Formulation & fill

Figure 14.7 Large-scale manufacture of adenoviral vectors for use for gene-therapy-based clinical proto-
cols. Refer to text for details