Synergistic Effect of Cavitation and Agitation on Protein Aggregation

Torisu, Tetsuo; Maruno, Takahiro; Hamaji, Yoshinori; Ohkubo, Tadayasu; Uchiyama, Susumu

doi:10.1016/j.xphs.2016.10.015

Cited by 70 publications

(67 citation statements)

References 26 publications

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“…Due to the lack of stable intermediate structures, protein aggregation generated by cavitation seems to occur spontaneously on the vapor/liquid interface. Similar to our results, instant aggregation behavior onto cavitation was recently suggested in a study were the combined effect of cavitation and shaking was analyzed . No hydroxyl radial associated changes in the antibody structure could be measured with mass spectrometry.…”

Section: Resultssupporting

confidence: 90%

“…Our findings indicate that cavitation in a certain bioprocess at high protein concentrations is most likely overseen. According to Torisu, Maruno, Hamaji, Ohkubo, and Uchiyama aggregates occurring from cavitation serve as a seeds for larger protein aggregates when further stress, like air/liquid interactions, occur. Therefore, addressing cavitation is crucial when designing bioprocesses.…”

Section: Resultsmentioning

confidence: 99%

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Impact of Cavitation, High Shear Stress and Air/Liquid Interfaces on Protein Aggregation

Duerkop

Berger

Dürauer

et al. 2018

Biotechnology Journal

102

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The reported impact of shear stress on protein aggregation has been contradictory. At high shear rates, the occurrence of cavitation or entrapment of air is reasonable and their effects possibly misattributed to shear stress. Nine different proteins (α-lactalbumin, two antibodies, fibroblast growth factor 2, granulocyte colony stimulating factor [GCSF], green fluorescence protein [GFP], hemoglobin, human serum albumin, and lysozyme) are tested for their aggregation behavior on vapor/liquid interfaces generated by cavitation and compared it to the isolated effects of high shear stress and air/liquid interfaces generated by foaming. Cavitation induced the aggregation of GCSF by þ68.9%, hemoglobin þ4%, and human serum albumin þ2.9%, compared to a control, whereas the other proteins do not aggregate. The protein aggregation behaviors of the different proteins at air/liquid interfaces are similar to cavitation, but the effect is more pronounced. Air-liquid interface induced the aggregation of GCSF by þ94.5%, hemoglobin þ35.5%, and human serum albumin (HSA) þ31.1%.The results indicate that the sensitivity of a certain protein toward cavitation is very similar to air/liquid-induced aggregation. Hence, hydroxyl radicals cannot be seen as the driving force for protein aggregation when cavitation occurs. Further, high shear rates of up to 10 8 s À1 do not affect any of the tested proteins. Therefore, also within this study generated extremely high isolated shear rates cannot be considered to harm structural integrity when processing proteins.

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Section: Resultssupporting

confidence: 90%

Section: Resultsmentioning

confidence: 99%

Impact of Cavitation, High Shear Stress and Air/Liquid Interfaces on Protein Aggregation

Duerkop

Berger

Dürauer

et al. 2018

Biotechnology Journal

102

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“…Studies have shown that friability testing is an effective approach for assessing the combined stresses of dropping and shaking. 13,19 Therefore, to assess the effect of dropping in combination with other shipping-relevant stress conditions on SO droplet release, we subjected PFSs to friability testing. The friability testing conditions, which yielded a concentration of micron-size SO droplets of 3 Â 10 5 p/mL or 1.5 Â 10 6 p/mL, as determined by flow imaging (FI) microscopy, were used to prepare SO-containing WFI.…”

Section: Introductionmentioning

confidence: 99%

An Assessment of the Ability of Submicron- and Micron-Size Silicone Oil Droplets in Dropped Prefillable Syringes to Invoke Early- and Late-Stage Immune Responses

Krayukhina¹,

Yokoyama²,

Hayashihara³

et al. 2019

Journal of Pharmaceutical Sciences

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a b s t r a c tA number of biopharmaceuticals are available as lyophilized formulations along with a prefilled syringe (PFS) containing water for injection (WFI). Submicron-and micron-size droplets of lubricating silicone oil (SO) applied to the inner surface of the PFS barrel might migrate into the WFI, to which protein pharmaceuticals can adsorb, potentially inducing an immune response. In the present study, we subjected siliconized cyclo-olefin polymer PFSs filled with WFI to dropping stress to simulate actual shipping conditions as well as evaluated the risk associated with the released SO droplets. The results confirmed the undesirable effects of SO on therapeutic proteins, including adsorption to SO droplets and increased secretion of several innate cytokines from human peripheral blood mononuclear cells of a small donor panel. Assessment of immunogenicity in vivo using BALB/c mice revealed a slight increase in the plasma concentrations of antidrug antibodies over 21 days in response to SO-containing antibody samples compared to the absence of SO. These results indicate that SO droplets form complexes with pharmaceutical proteins that can potentially invoke early-and late-stage immune responses. Therefore, the use of SO-free cyclo-olefin polymer PFSs as primary containers for WFI could contribute to the enhanced safety of reconstituted biopharmaceuticals.

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“…No chemical modifications was found for the antibody while the data for rhGH was inconsistent. Recent research on cavitation did also not find radial associated oxidation of an antibody by cavitation events [45]. However, in both studies protein aggregation could be noticed, which indicates that the driving force behind cavitation induced aggregation is more reasonable the generated surface area and not hydroxyl radicals.…”

Section: Protein Stressmentioning

confidence: 69%

Influence of cavitation and high shear stress on HSA aggregation behavior

Duerkop

Berger

Dürauer

et al. 2017

Engineering in Life Sciences

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Neither the influence of high shear rates nor the impact of cavitation on protein aggregation is fully understood. The effect of cavitation bubble collapse‐derived hydroxyl radicals on the aggregation behavior of human serum albumin (HSA) was investigated. Radicals were generated by pumping through a micro‐orifice, ultra‐sonication, or chemically by Fenton's reaction. The amount of radicals produced by the two mechanical methods (0.12 and 11.25 nmol/(L min)) was not enough to change the protein integrity. In contrast, Fenton's reaction resulted in 382 nmol/(L min) of radicals, inducing protein aggregation. However, the micro‐orifice promoted the formation of soluble dimeric HSA aggregates. A validated computational fluid dynamic model of the orifice revealed a maximum and average shear rate on the order of 108 s−1 and 1.2 × 106 s−1, respectively. Although these values are among the highest ever reported in the literature, dimer formation did not occur when we used the same flow rate but suppressed cavitation. Therefore, aggregation is most likely caused by the increased surface area due to cavitation‐mediated bubble growth, not by hydroxyl radical release or shear stress as often reported.

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Synergistic Effect of Cavitation and Agitation on Protein Aggregation

Cited by 70 publications

References 26 publications

Impact of Cavitation, High Shear Stress and Air/Liquid Interfaces on Protein Aggregation

Impact of Cavitation, High Shear Stress and Air/Liquid Interfaces on Protein Aggregation

An Assessment of the Ability of Submicron- and Micron-Size Silicone Oil Droplets in Dropped Prefillable Syringes to Invoke Early- and Late-Stage Immune Responses

Influence of cavitation and high shear stress on HSA aggregation behavior

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