Virus clearance validation across continuous capture chromatography

Multicolumn capture chromatography is gaining increased attention lately due to the significant economic and process advantages it offers compared with traditional batch mode chromatography. However, for wide adoption of this technology in clinical and commercial space, it requires scalable models for executing viral validation studies. In this study, viral validation studies were conducted under cGLP guidelines to assess retro- (X-MuLV) and parvo-virus (MVM) clearance across twin-column continuous capture chromatography (CaptureSMB). A surrogate model was also developed using standard batch mode chromatography based on flow path modifications to mimic the loading strategy used in CaptureSMB. The results show that a steady state was achieved by the second cycle for both antibody binding and virus clearance and that the surrogate model using batch mode chromatography equipment provided impurity clearance that was comparable to that obtained during cyclical operation of CaptureSMB. Further, the log reduction values (LRVs) achieved during CaptureSMB were also comparable to the LRVs obtained using standard batch capture chromatography. This was expected since the mode of virus separation during protein A chromatography is primarily based on removal during the flow through and wash steps. Finally, this study also presents assessments on the resin cleaning strategy during continuous chromatography and how the duration of clean-in-place solution exposure impacts virus carryover.     

Mechanistic insights into viral clearance during the chromatography steps in antibody processes by using virus surrogates

Viral safety is required for biological products to treat human diseases, and the burden of inactivation and or virus removal lies on the downstream purification process. Minute virus of mice (MVM) is a nonenveloped parvovirus commonly used as the worst-case model virus in validation studies because of its small size and high chemical stability. In this study, we investigated the use of MVM-mock virus particle (MVP) and bacteriophage ΦX174 as surrogates for MVM to mimic viral clearance studies, with a focus on chromatography operations. Based on structural models and comparison of log reduction value among MVM, MVP, and ΦX174, it was demonstrated that MVP can be used as a noninfectious surrogate to assess viral clearance during process development in multiple chromatography systems in a biosafety level one (BSL-1) laboratory. Protein A (ProA) chromatography was investigated to strategically assess the impact of the resin, impurities, and the monoclonal antibody product on virus removal.     

Renal function at steady state in a toad (Bufo viridis) acclimated in hyperosmotic NaCl and urea solutions

Kidney function of the euryhaline toad Bufo viridis was studied in animals acclimated to tap water and solutions of NaCl (230 and 500 mosmol · kg^-1 H₂O) and urea (500 mmol · l^-1) in steady-state conditions. An ureter was eatheterized for continuous urine collection and blood was sampled from an iliac artery. A single injection of ³H-inulin served for estimation of glomerular filtration rate: this was in the range of 15–27 ml · kg^-1 · h^-1 and did not differ significantly in any of the acclimation conditions. Urine flow, on the other hand, varied considerably and was highest in tap water (18.2±3.2 ml · kg^-1 · h^-1; urine/plasma inulin ratio=0.88), lower in 230 mosmol · kg^-1 H₂O NaCl solution (13.5±3.9 ml · kg^-1 · h^-1; u/p inulin ratio=1.73) and lowest in 500 mosmol · kg^-1 H₂O NaCl or urea acclimation solutions (5–7 ml · kg^-1 · h^-1; u/p inulin=3.7–4.2). Clearance of free water was high in the tap water group, lower in 230 mosmol · kg^-1 H₂O NaCl solution, and much lower in the hyperosmotic acclimation conditions. Clearances of both Na⁺ and Cl^- were similar under our experimental conditions, but changed independently in accordance to the composition of the acclimation solution. Potassium clearance was similar in all acclimation conditions, and a constant plasma K⁺ concentration was maintained. Urea clearance was high in tap water and 500 mmol · l^-1 urea acclimation groups and low in the NaCl acclimations. The experiments show that the glomerular filtration rate remains more or less unchanged in all acclimation conditions, and suggest that the different rates of urine flow at steady state must be due mostly to tubular processes. The final composition of the urine is the result of specific and highly selective tubular processes.Abbreviations %T fractional reabsorbance - AVT argine vasotocin - C _{free water} free water clearance - C _osmol osmolyte clearance - GFR glomerular filtration rate - MS-222 methanetricaine sulphonate - U/P urine to plasma inulin ratio - V volume