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Mechanistic Understanding of Protein-Silicone Oil Interactions

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ABSTRACT

Purpose

To investigate interactions between protein and silicone oil so that we can provide some mechanistic understanding of protein aggregation in silicone oil lubricated syringes and its prevention by formulation additives such as Polysorbate 80 and Poloxamer 188.

Methods

Interfacial tension values of silicone oil/water interface of abatacept solutions with and without formulation additives were obtained under equilibrium conditions using Attension Theta optical tensiometer. Their adsorption and desorption profiles were measured using Quartz Crystal Microbalancing with Dissipation monitoring (QCM-D). The degree of aggregation of abatacept was assessed based on size exclusion measurement.

Results

Adsorption of abatacept at the oil/water interface was shown. Polysorbat 80 was more effective than Poloxamer 188 in preventing abatacept adsorption. Moreover, it was noted that some of the adsorbed abatacept molecules were not desorbed readily upon buffer rinse. Finally, no homogeneous aggregation was observed at room temperature and a slight increase of aggregation was only observed for samples measured at 40°C which can be prevented using Polysorbate 80.

Conclusions

Interfacial adsorption of proteins is the key step and maybe responsible for the phenomenon of soluble-protein loss when contacting silicone oil and the irreversible adsorption of protein may be associated with protein denaturation/aggregation.

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AcknowledgmentS & DISCLOSURES

Authors would like to thank the support of Drug Product Science and Technology Management of Bristol-Myers Squibb Company and Dr. M. Hussain for critical review.

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Correspondence to Jinjiang Li.

Appendix

Appendix

Some Theoretical Considerations

In this study, it is assumed that the oil/water (buffer) has a sharp interface (19,20) and its interfacial tension value is γ0 which can be measured, and the adsorption of protein molecules or surfactant molecules or both at the oil/water interface can reduce the interfacial tension to γ. Therefore, the surface pressure is expressed as: Π = γ0-γ, and Π can be modeled as the concentration (Г) in the interfacial layer, which is also a function of protein bulk concentration c, and the interfacial molar area (ω). For a surfactant (component 2) in a protein solution (component 1), the following equation can be derived based on the equation of state assuming the protein is in the state with minimal molar area and the surfactant is in a single adsorption state:

$$ \Pi = - \frac{{RT}}{\omega }\left[ {\ln \left( {1 - {\Gamma_{\sum }}\omega } \right) - {a_{{el}}}\Gamma_1^2\omega_1^2} \right] $$
(1)

while the expressions for the adsorption isotherm of protein and surfactant are

$$ {b_1}{c_1} = \frac{{{\Gamma_1}{\omega_1}}}{{{{\left( {1 - {\Gamma_{\sum }}\omega } \right)}^{{{\omega_2}/\omega }}}}}\;{\text{and}}\;{b_2}{c_2} = \frac{{{\Gamma_2}{\omega_2}}}{{{{\left( {1 - {\Gamma_{\sum }}\omega } \right)}^{{{\omega_2}/\omega }}}}} $$
(2)

with

$$ {\Gamma_{{\sum}}} = {\Gamma_{{1}}} + {\Gamma_{{2}}}\;{\text{and}}\;\frac{{{\Gamma_1}{\omega_1}}}{{{\Gamma_2}{\omega_2}}} = \frac{{{b_1}{c_1}}}{{{b_2}{c_2}}}{(1 - {\Gamma_{\sum }}\omega )^{{\frac{{{\omega_1} - {\omega_2}}}{\omega }}}} $$
(3)

The average molar area of adsorbed component 1 and 2 is

$$ \omega = \frac{{{\Gamma_1}{\omega_1} + {\Gamma_2}{\omega_2}}}{{\Gamma {}_1 + {\Gamma_2}}} $$
(4)

Here c1, c2, ω1, ω2, b1, b2 are the concentrations, molar interfacial area, and bulk/interface distribution coefficients of protein and surfactant, and Γ1 and Γ2 are the concentration of protein and surfactant in the interfacial layer. ael is a parameter related to the electrostatic interaction in the solution depending on the dielectric constant of the protein solution, the total concentration of electrolytes, the number of non-bound unit charges in the protein molecules, etc (20,21). Dynamically, the time-dependent adsorption, Γ(t), depends on the diffusion coefficient of the molecules, bulk concentration and time, and adsorption kinetic model (10) is shown as the following:

$$ \Gamma (t) = 2\sqrt {{\frac{D}{\pi }}} \left( {{c_0}\left){\vphantom{1t}}\right.\!\!\!\!\overline{\,\,\,\vphantom 1{t}} - \int\limits_0^t {c(0,t - \tau )d\sqrt {\tau } } } \right) $$
(5)

where D is the diffusion coefficient and c0 is the bulk concentration, t is the time. In the following text, Eqs. 15 will be used as qualitative guidance for discussion. For a simplied system, the above equation can be reduced to

$$ \Gamma (t) = {c_0}\sqrt {{\frac{{Dt}}{\pi }}} $$
(6)

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Li, J., Pinnamaneni, S., Quan, Y. et al. Mechanistic Understanding of Protein-Silicone Oil Interactions. Pharm Res 29, 1689–1697 (2012). https://doi.org/10.1007/s11095-012-0696-6

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