LentiTag: A novel approach to the efficient manufacturing of active lentiviral vectors

PROJECT SUMMARY Cell and gene therapies are garnering attention for their remarkable clinical outcomes and their promise to treat diseases previously considered uncurable. Lentiviral vectors (LVs) are used in over one third of applications in cell and gene therapies.2 As with most viral vector and vaccine manufacturing, LVs are challenging and

expensive to manufacture and there are no methods in commercial use that offer specificity, high yield, and scalability. Current manufacturing relies on anion exchange chromatography, which (1) has poor capacities for large viruses that cannot effectively diffuse into small resin pores, (2) necessitates additional purification steps

because its lack of LV specificity causes contaminant co-purification, and (3) requires harsh elution conditions that result in low infectious LV recoveries of only ~50%.15, 1 In this Phase I SBIR proposal, Isolere Bio will develop a fusion protein reagent comprised of an affinity domain that is specific for LV and a proprietary biopolymer

domain that has robust and precisely tunable liquid-liquid phase separation behavior. The LentiTag™ reagent will capture LVs in solution with high specificity and then efficiently sequester them into phase separated droplets on command with a simple environmental trigger — an incremental adjustment in salt. These droplets, highly

pure and concentrated, are easily separated from all excluded contaminants with tangential flow filtration (TFF), a unit operation that scales up simply and enables high volumetric throughput. Once host cell proteins and other cellular contaminants have been washed away, the LV can be gently recovered from the droplets with an elution

buffer that disrupts the affinity interaction at near-neutral pH. To demonstrate LentiTag™’s technical feasibility, Isolere will first design, produce, and characterize an LV-specific capture protein (LCP). The LCP binds the most common LV envelope glycoprotein and phase separates at ambient temperature with a small increase in salt

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