Virus-like particles (VLPs) hold tremendous potential as vaccine candidates. These innovative biopharmaceuticals present the remarkable advantages of closely mimicking the three-dimensional nature of an actual virus while lacking the virus genome packaged inside its capsid. As a result, an equally efficient but safer prophylaxis is anticipated as compared to inactivated or live attenuated viral vaccines. With the advent of successful cases of approved VLP-based vaccines, pharmaceutical companies are indeed redirecting their resources to the development of such products. This paper reviews the current choices and trends of large-scale production and purification of VLP-based vaccines generated through the baculovirus expression vector system using insect cells.

Recombinant baculoviruses (rBVs) are widely used as vectors for the production of recombinant proteins in insect cells. More recently, these viral vectors have been gaining increasing attention due to their emerging potential as gene therapy vehicles to mammalian cells. Their production in stirred bioreactors using insect cells is an established technology; however, the downstream processing (DSP) of baculoviruses envisaged for clinical applications is still poorly developed. In the present work, the recovery and purification of rBVs aiming at injectable-grade virus batches for gene therapy trials was studied. A complete downstream process comprising three steps-depth filtration, ultra/ diafiltration and membrane sorption-was successfully developed. Optimal operational conditions for each individual step were achieved yielding a scalable DSP for rBVs as vectors for gene therapy. The processing route designed hereby presents global recovery yields reaching 40% (at purities over 98%) and, most importantly, relies on technologies easy to transfer to process scales under cGMP guidelines.

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Lentivirus can be engineered to be a highly potent vector for gene therapy applications. However, generation of clinical grade vectors in enough quantities for therapeutic use is still troublesome and limits the preclinical and clinical experiments. As a first step to solve this unmet need we recently introduced a baculovirus-based production system for lentiviral vector (LV) production using adherent cells. Herein, we have adapted and optimized the production of these vectors to a suspension cell culture system using recombinant baculoviruses delivering all elements required for a safe latest generation LV preparation. High-titer LV stocks were achieved in 293T cells grown in suspension. Produced viruses were accurately characterized and the functionality was also tested in vivo. Produced viruses were compared with viruses produced by calcium phosphate transfection method in adherent cells and polyethylenimine transfection method in suspension cells. Furthermore, a scalable and cost-effective capture purification step was developed based on a diethylaminoethyl monolithic column capable of removing most of the baculoviruses from the LV pool with 65% recovery.

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