Abstract

The objective of this study was to characterize the in vitro and in vivo properties of the F(ab')(2) fragment of panitumumab and to investigate its potential for imaging and radioimmunotherapy. The panitumumab F(ab')(2) was generated by enzymatic pepsin digestion. After the integrity and immunoreactivity of the F(ab')(2) was evaluated, the fragment was radiolabeled. In vivo studies included direct quantitation of tumor targeting and normal organ distribution of the radiolabeled panitumumab F(ab')(2) as well as planar γ-scintigraphy and PET imaging. The panitumumab F(ab')(2) was successfully produced by peptic digest. The F(ab')(2) was modified with the CHX-A"-DTPA chelate and efficiently radiolabeled with either (111)In or (86)Y. In vivo tumor targeting was achieved with acceptable uptake of radioactivity in the normal organs. The tumor targeting was validated by both imaging modalities with good visualization of the tumor at 24 h. The panitumumab F(ab')(2) fragment is a promising candidate for imaging of HER1 positive cancers.

Highlights

  • The objective of this study was to characterize the in vitro and in vivo properties of the F(ab’)2 fragment of panitumumab and to investigate its potential for imaging and radioimmunotherapy

  • To determine the optimal cleavage time, 2% pepsin was added to 250 μg panitumumab and incubated at 37°C

  • As determined by SDS-PAGE, near complete pepsin digestion of panitumumab to a F(ab’)2 fragment appears to occur after 8 h, evident in Figure 1 by the loss of the higher-molecular-weight band of the intact IgG under non-reducing conditions (Figure 1a) and the transition of the heavy-chain band to a lower molecular weight when subjected to reduction with b-mercaptoethanol (Figure 1b)

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Summary

Introduction

The objective of this study was to characterize the in vitro and in vivo properties of the F(ab’) fragment of panitumumab and to investigate its potential for imaging and radioimmunotherapy. While intact monoclonal antibody molecules are still most commonly used, they may not necessarily be the most efficient or desired molecular form depending on the application Because of their relatively large size (approximately 150 kD), intact mAbs tend to have unfavorable imaging kinetics, relatively poor tumor penetration, and present with the potential for eliciting host antibody responses [2,3,4,5,6,7]. The solution to these myriad obstacles has been to reduce the size of intact antibodies to smaller forms or fragments, achieved either through enzymatic cleavage or by genetic engineering. The average molecular weight of the F(ab’) fragment is approximately 110 kD

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