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Int J Radiat Oncol Biol Phys. 2012 Jul 1;83(3):1023-30. doi: 10.1016/j.ijrobp.2011.08.039. Epub 2011 Dec 2.

Intranuclear delivery of a novel antibody-derived radiosensitizer targeting the DNA-dependent protein kinase catalytic subunit.

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  • 1Institute of Molecular Medicine and Genetics, Georgia Health Sciences University, Augusta, GA 30912, USA.

Abstract

PURPOSE:

To inhibit DNA double-strand break repair in tumor cells by delivery of a single-chain antibody variable region fragment (ScFv 18-2) to the cell nucleus. ScFv 18-2 binds to a regulatory region of the DNA-dependent protein kinase (DNA-PK), an essential enzyme in the nonhomologous end-joining pathway, and inhibits DNA end-joining in a cell-free system and when microinjected into single cells. Development as a radiosensitizer has been limited by the lack of a method for intranuclear delivery to target cells. We investigated a delivery method based on folate receptor-mediated endocytosis.

METHODS AND MATERIALS:

A recombinant ScFv 18-2 derivative was conjugated to folate via a scissile disulfide linker. Folate-ScFv 18-2 was characterized for its ability to be internalized by tumor cells and to influence the behavior of ionizing radiation-induced repair foci. Radiosensitization was measured in a clonogenic survival assay. Survival curves were fitted to a linear-quadratic model, and between-group differences were evaluated by an F test. Sensitization ratios were determined based on mean inhibitory dose.

RESULTS:

Human KB and NCI-H292 lung cancer cells treated with folate-conjugated ScFv 18-2 showed significant radiosensitization (p < 0.001). Sensitization enhancement ratios were 1.92 ± 0.42 for KB cells and 1.63 ± 0.13 for NCI-H292 cells. Studies suggest that treatment inhibits repair of radiation-induced DSBs, as evidenced by the persistence of γ-H2AX-stained foci and by inhibition of staining with anti-DNA-PKcs phosphoserine 2056.

CONCLUSIONS:

Folate-mediated endocytosis is an effective method for intranuclear delivery of an antibody-derived DNA repair inhibitor.

Copyright © 2012 Elsevier Inc. All rights reserved.

PMID:
22138455
[PubMed - indexed for MEDLINE]
PMCID:
PMC3698569
Free PMC Article
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