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Differentiation between temporary and real non-clearability of biotinylated IgG antibody by avidin in mice.

Dou S, Virostko J, Rusckowski M, Greiner DL, Powers AC, Liu G - Front Pharmacol (2014)

Bottom Line: By comparing the effects of natural clearance at a longer post-injection time and avidin clearance, we demonstrated that avidin clearance is much more effective.By directly attaching avidin to a biotinylated antibody prior to injection, we found that the biotinylated antibody in blood, once bound to the clearing agent, can be removed from the circulation immediately and completely, while the real non-clearable antibody without biotin stays.In conclusion, the use of antibody pretargeting as a tool in this study has improved understanding of the incomplete clearance by avidin and can aid in overcoming this obstacle.

View Article: PubMed Central - PubMed

Affiliation: Department of Radiology, University of Massachusetts Medical School Worcester, MA, USA.

ABSTRACT
Although an increasing number of antibody conjugates are being used in the clinic, there remain many unmet needs in antibody targeting. Normal tissue background is one of the key issues that limits the therapeutic efficacy and the detection sensitivity. Background reduction coupled with dose increase may provide the required target accumulation of the label or toxin at an acceptable normal tissue background. However, the knowledge about the in vivo interaction between antibody and a clearing agent is currently inadequate for designing a rational clearance regimen or system. The current investigation focuses on the clearability of antibody for background reduction, an important topic to antibody targeting in general. The investigation employs pretargeting as a research tool and avidin as a model clearing agent. By comparing the effects of natural clearance at a longer post-injection time and avidin clearance, we demonstrated that avidin clearance is much more effective. By directly attaching avidin to a biotinylated antibody prior to injection, we found that the biotinylated antibody in blood, once bound to the clearing agent, can be removed from the circulation immediately and completely, while the real non-clearable antibody without biotin stays. The study of multiple avidin injections confirmed that the presence of clearable biotinylated antibodies after an avidin injection is due to their temporary inaccessibility and subsequent return from tissue compartments. The collective clearance efficiency of 91% by three avidin injections indicates a continuous IV infusion would be recommended to remove all of the biotinylated IgG molecules. In conclusion, the use of antibody pretargeting as a tool in this study has improved understanding of the incomplete clearance by avidin and can aid in overcoming this obstacle.

No MeSH data available.


Clearance efficiency at different pretargeting intervals in a pretargeting procedure using biotin-CC49-MORF, avidin, and 99 mTc-cMORF. The cMORF level is proportional to and used as a measure of the antibody level. The clearance efficiency is defined as the ratio of blood level with/without avidin.
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Figure 1: Clearance efficiency at different pretargeting intervals in a pretargeting procedure using biotin-CC49-MORF, avidin, and 99 mTc-cMORF. The cMORF level is proportional to and used as a measure of the antibody level. The clearance efficiency is defined as the ratio of blood level with/without avidin.

Mentions: Both prolonging the pretargeting interval and the use of avidin reduced the antibody levels in blood as shown in Figure 1 and normal tissues (not presented), but the use of avidin is more effective. Two-Way ANOVA indicates that the cMORF blood level is significantly decreased both by pretargeting interval prolongation (p = 0.007) and avidin administration (p < 0.00005). With regard to avidin administration, the result of ANOVA is in agreement with the visual judgment based on the large difference relative to the SD at each interval. The clearance efficiency using avidin as a clearing agent is calculated to be 55–58%, with >40% antibody remaining in the circulation. As will be shown, this incomplete clearance is neither due to ineffective binding of avidin nor to the dissociation of the avidin/biotin complex but mainly results from the rapid clearance of avidin from blood to liver.


Differentiation between temporary and real non-clearability of biotinylated IgG antibody by avidin in mice.

Dou S, Virostko J, Rusckowski M, Greiner DL, Powers AC, Liu G - Front Pharmacol (2014)

Clearance efficiency at different pretargeting intervals in a pretargeting procedure using biotin-CC49-MORF, avidin, and 99 mTc-cMORF. The cMORF level is proportional to and used as a measure of the antibody level. The clearance efficiency is defined as the ratio of blood level with/without avidin.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4109569&req=5

Figure 1: Clearance efficiency at different pretargeting intervals in a pretargeting procedure using biotin-CC49-MORF, avidin, and 99 mTc-cMORF. The cMORF level is proportional to and used as a measure of the antibody level. The clearance efficiency is defined as the ratio of blood level with/without avidin.
Mentions: Both prolonging the pretargeting interval and the use of avidin reduced the antibody levels in blood as shown in Figure 1 and normal tissues (not presented), but the use of avidin is more effective. Two-Way ANOVA indicates that the cMORF blood level is significantly decreased both by pretargeting interval prolongation (p = 0.007) and avidin administration (p < 0.00005). With regard to avidin administration, the result of ANOVA is in agreement with the visual judgment based on the large difference relative to the SD at each interval. The clearance efficiency using avidin as a clearing agent is calculated to be 55–58%, with >40% antibody remaining in the circulation. As will be shown, this incomplete clearance is neither due to ineffective binding of avidin nor to the dissociation of the avidin/biotin complex but mainly results from the rapid clearance of avidin from blood to liver.

Bottom Line: By comparing the effects of natural clearance at a longer post-injection time and avidin clearance, we demonstrated that avidin clearance is much more effective.By directly attaching avidin to a biotinylated antibody prior to injection, we found that the biotinylated antibody in blood, once bound to the clearing agent, can be removed from the circulation immediately and completely, while the real non-clearable antibody without biotin stays.In conclusion, the use of antibody pretargeting as a tool in this study has improved understanding of the incomplete clearance by avidin and can aid in overcoming this obstacle.

View Article: PubMed Central - PubMed

Affiliation: Department of Radiology, University of Massachusetts Medical School Worcester, MA, USA.

ABSTRACT
Although an increasing number of antibody conjugates are being used in the clinic, there remain many unmet needs in antibody targeting. Normal tissue background is one of the key issues that limits the therapeutic efficacy and the detection sensitivity. Background reduction coupled with dose increase may provide the required target accumulation of the label or toxin at an acceptable normal tissue background. However, the knowledge about the in vivo interaction between antibody and a clearing agent is currently inadequate for designing a rational clearance regimen or system. The current investigation focuses on the clearability of antibody for background reduction, an important topic to antibody targeting in general. The investigation employs pretargeting as a research tool and avidin as a model clearing agent. By comparing the effects of natural clearance at a longer post-injection time and avidin clearance, we demonstrated that avidin clearance is much more effective. By directly attaching avidin to a biotinylated antibody prior to injection, we found that the biotinylated antibody in blood, once bound to the clearing agent, can be removed from the circulation immediately and completely, while the real non-clearable antibody without biotin stays. The study of multiple avidin injections confirmed that the presence of clearable biotinylated antibodies after an avidin injection is due to their temporary inaccessibility and subsequent return from tissue compartments. The collective clearance efficiency of 91% by three avidin injections indicates a continuous IV infusion would be recommended to remove all of the biotinylated IgG molecules. In conclusion, the use of antibody pretargeting as a tool in this study has improved understanding of the incomplete clearance by avidin and can aid in overcoming this obstacle.

No MeSH data available.