High-Grade Purification of Third-Generation HIV-Based Lentiviral Vectors by Anion Exchange Chromatography for Experimental Gene and Stem Cell Therapy Applications

Olgun, Hazal Banu; Tasyurek, Hale M.; Şanlioğlu, Ahter Dilşad; Şanlıoğlu, Salih

doi:10.1007/7651_2018_154

Cited by 18 publications

(21 citation statements)

References 23 publications

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“…The use of Benzonase or Denarase reduces DNA size and decreases bulk viscosity. [ 41,46 ] These enzymes require magnesium ions as a co‐factor and show optimum activity at 37 °C; however, different experimental conditions (e.g., amount of enzyme, temperature, incubation time, salt concentration, pH, and the presence of enzyme inhibitors) are reported regarding nucleic acid cleavage step. Recently, Oxford Biomedica developed a system (SecNuc) that avoids the addition of nucleases that are co‐produced with LV.…”

Section: Scalable Methods For LV Downstream Processingmentioning

confidence: 99%

“…Viral pelleting and resuspension procedure is often used as a concentration method. [39][40][41] The use of density gradient centrifugation or ultracentrifugation using sucrose [42,43] or iodixanol [44] has been reported and can achieve some degree of purification along with reasonable concentration. [41,45] Another technique for small-scale purification schemes uses polyethylene glycol (PEG).…”

Section: Traditional LV Purification and Concentration Methodsmentioning

confidence: 99%

“…[26] Alternatively, multimodal chromatography, like Capto Core 700, can be used for polishing. [26,41,46,91] This kind of flowthrough chromatography combines the properties of SEC and AEX. The chromatographic media is composed of particles with a functionalized core, to which larger particles cannot access, and a non-functionalized outer layer.…”

Section: Polishing Formulation and Sterilizationmentioning

confidence: 99%

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Advances in Lentivirus Purification

Moreira

Cavaco

Faria

et al. 2020

Biotechnology Journal

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Lentiviral vectors (LVs) have been increasingly used as a tool for gene and cell therapies since they can stably integrate the genome in dividing and nondividing cells. LV production and purification processes have evolved substantially over the last decades. However, the increasing demands for higher quantities with more restrictive purity requirements are stimulating the development of novel materials and strategies to supply the market with LV in a cost-effective manner. A detailed review of each downstream process unit operation is performed, limitations, strengths, and potential outcomes being covered. Currently, the majority of large-scale LV manufacturing processes are still based on adherent cell culture, although it is known that the industry is migrating fast to suspension cultures. Regarding the purification strategy, it consists of batch chromatography and membrane technology. Nevertheless, new solutions are being created to improve the current production schemes and expand its clinical use.

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Section: Scalable Methods For LV Downstream Processingmentioning

confidence: 99%

Section: Traditional LV Purification and Concentration Methodsmentioning

confidence: 99%

Section: Polishing Formulation and Sterilizationmentioning

confidence: 99%

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Advances in Lentivirus Purification

Moreira

Cavaco

Faria

et al. 2020

Biotechnology Journal

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“…The initial downstream stages used in clarification are typically microfiltration and/or centrifugation. Cells and bulk particles can be sedimented and removed by low-speed centrifugation [ 181 ]. Alternatively, acoustic filters can be used to entrap suspended cells in soundwaves to return to the bioreactor during harvest [ 162 ].…”

Section: Downstream Processing Of Lentiviral Vectorsmentioning

confidence: 99%

“…Thus, vector flows through the column, whilst negatively charged DNA or other smaller proteins are captured and removed. This can be applied as a polishing step or contaminant reduction step for vectors and viruses [ 181 , 226 , 227 ].…”

Section: Downstream Processing Of Lentiviral Vectorsmentioning

confidence: 99%

Lentiviral Vector Bioprocessing

Perry

Rayat

2021

Viruses

110

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Lentiviral vectors (LVs) are potent tools for the delivery of genes of interest into mammalian cells and are now commonly utilised within the growing field of cell and gene therapy for the treatment of monogenic diseases and adoptive therapies such as chimeric antigen T-cell (CAR-T) therapy. This is a comprehensive review of the individual bioprocess operations employed in LV production. We highlight the role of envelope proteins in vector design as well as their impact on the bioprocessing of lentiviral vectors. An overview of the current state of these operations provides opportunities for bioprocess discovery and improvement with emphasis on the considerations for optimal and scalable processing of LV during development and clinical production. Upstream culture for LV generation is described with comparisons on the different transfection methods and various bioreactors for suspension and adherent producer cell cultivation. The purification of LV is examined, evaluating different sequences of downstream process operations for both small- and large-scale production requirements. For scalable operations, a key focus is the development in chromatographic purification in addition to an in-depth examination of the application of tangential flow filtration. A summary of vector quantification and characterisation assays is also presented. Finally, the assessment of the whole bioprocess for LV production is discussed to benefit from the broader understanding of potential interactions of the different process options. This review is aimed to assist in the achievement of high quality, high concentration lentiviral vectors from robust and scalable processes.

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Clinical grade lentiviral vector purification and quality control requirements

Shi¹,

Jia²,

Liu

2022

J of Separation Science

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Lentiviral vectors have been proven to be a powerful tool in gene therapies that includes the ability to perform long-term gene editing in both dividing and nondividing cells. In order to meet the rising demand for clinical-grade lentiviral vectors for future clinical trials and requirements by regulatory agencies, new methods and technologies were developed, including the rapid optimization of production and purification processes. However, gaps still exist in achieving ideal yields and recovery rates in large-scale manufacturing process steps. The downstream purification process is a critical step required to obtain a sufficient quantity and high-quality lentiviral vectors products, which is challenged by the low stability of the lentiviral vector particles and large production volumes associated with the manufacturing process. This review summarizes the most recent and promising technologies and enhancements used in the large-scale purification process step of lentiviral vector manufacturing and aims to provide a significant contribution towards the achievement of providing sufficient quantity and quality of lentiviral vectors in scalable processes.

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High-Grade Purification of Third-Generation HIV-Based Lentiviral Vectors by Anion Exchange Chromatography for Experimental Gene and Stem Cell Therapy Applications

Cited by 18 publications

References 23 publications

Advances in Lentivirus Purification

Advances in Lentivirus Purification

Lentiviral Vector Bioprocessing

Clinical grade lentiviral vector purification and quality control requirements

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