A Novel Cell-Penetrating Antibody Fragment Inhibits the DNA Repair Protein RAD51

Scientific Reports
Landon PastushokC Ronald Geyer

Abstract

DNA damaging chemotherapies are successful in cancer therapy, however, the damage can be reversed by DNA repair mechanisms that may be up-regulated in cancer cells. We hypothesized that inhibiting RAD51, a protein involved in homologous recombination DNA repair, would block DNA repair and restore the effectiveness of DNA damaging chemotherapy. We used phage-display to generate a novel synthetic antibody fragment that bound human RAD51 with high affinity (KD = 8.1 nM) and inhibited RAD51 ssDNA binding in vitro. As RAD51 is an intracellular target, we created a corresponding intrabody fragment that caused a strong growth inhibitory phenotype on human cells in culture. We then used a novel cell-penetrating peptide "iPTD" fusion to generate a therapeutically relevant antibody fragment that effectively entered living cells and enhanced the cell-killing effect of a DNA alkylating agent. The iPTD may be similarly useful as a cell-penetrating peptide for other antibody fragments and open the door to numerous intracellular targets previously off-limits in living cells.

References

Apr 1, 1991·Trends in Biotechnology·R E Bird, B W Walker
Aug 1, 1988·Proceedings of the National Academy of Sciences of the United States of America·J S HustonR Crea
Sep 1, 1966·Clinica Chimica Acta; International Journal of Clinical Chemistry·K Itaya, M Ui
Mar 14, 1995·Proceedings of the National Academy of Sciences of the United States of America·T HaafD C Ward
Jun 14, 1996·Biochemical and Biophysical Research Communications·T TakiT Morita
Jun 25, 1996·Proceedings of the National Academy of Sciences of the United States of America·T Tsuzuki MoritaT
Dec 31, 1997·Molecular and Cellular Biology·S J XiaR J Shmookler Reis
Apr 16, 1998·The Journal of Biological Chemistry·E A Namsaraev, P Berg
May 8, 1998·Biochemical and Biophysical Research Communications·T OhnishiT Morita
Aug 11, 1998·Trends in Biochemical Sciences·P Baumann, S C West
Nov 28, 2000·International Journal of Cancer. Journal International Du Cancer·H MaackeH W Stürzbecher
Jun 19, 2001·Journal of Immunological Methods·Y ZhaoH Kohler
Jul 19, 2001·Proceedings of the National Academy of Sciences of the United States of America·X YuE H Egelman
Feb 13, 2002·The Journal of Biological Chemistry·Gregory TomblineRichard Fishel
Aug 13, 2003·The Journal of Biological Chemistry·Patrick SungMichael G Sehorn
Oct 7, 2004·The Journal of Biological Chemistry·Markus RaschlePatrick Sung
Mar 10, 2005·The Journal of Gene Medicine·Makoto ItoYasufumi Kaneda
Jun 21, 2006·Proceedings of the National Academy of Sciences of the United States of America·Konstantin KiianitsaWolf-Dietrich Heyer
Oct 13, 2006·Molecular and Cellular Biology·Cindy W FungLorraine S Symington
Jun 15, 2007·Nucleic Acids Research·Xuan LiWolf-Dietrich Heyer
Sep 11, 2007·Journal of Molecular Biology·Frederic A FellouseSachdev S Sidhu
Feb 8, 2008·Nature Reviews. Cancer·Thomas HelledayRicky A Sharma
Feb 9, 2008·Pharmacology·Howard R Mellor, Richard Callaghan
Jul 12, 2008·Molecular Carcinogenesis·Erika T Brown, Jeffrey T Holt
Apr 2, 2009·Nucleic Acids Research·Takako IshidaHitoshi Kurumizaka
May 19, 2009·Journal of Molecular Biology·Tomohiko Sugiyama, Noriko Kantake
Oct 27, 2009·Nature Structural & Molecular Biology·Karina B FalboXuetong Shen

❮ Previous
Next ❯

Citations

Jul 15, 2020·Annual Review of Genetics·Braulio BonillaKara A Bernstein
Apr 21, 2021·Expert Opinion on Drug Discovery·Ly PoroskÜlo Langel
Oct 6, 2020·NAR Cancer·McKenzie K GrundyKara A Bernstein
Jul 21, 2021·International Journal of Radiation Biology·Jac A NickoloffRobert Hromas

❮ Previous
Next ❯

Methods Mentioned

BETA
phage display
biosensors
biolayer interferometry
biosensor
Biolayer
light microscopy
Flow cytometry
PCR
Protein Assay
phage-display

Software Mentioned

SPSS
ForteBio Data Analysis

Related Concepts

Trending Feeds

COVID-19

Coronaviruses encompass a large family of viruses that cause the common cold as well as more serious diseases, such as the ongoing outbreak of coronavirus disease 2019 (COVID-19; formally known as 2019-nCoV). Coronaviruses can spread from animals to humans; symptoms include fever, cough, shortness of breath, and breathing difficulties; in more severe cases, infection can lead to death. This feed covers recent research on COVID-19.

Blastomycosis

Blastomycosis fungal infections spread through inhaling Blastomyces dermatitidis spores. Discover the latest research on blastomycosis fungal infections here.

Nuclear Pore Complex in ALS/FTD

Alterations in nucleocytoplasmic transport, controlled by the nuclear pore complex, may be involved in the pathomechanism underlying multiple neurodegenerative diseases including Amyotrophic Lateral Sclerosis and Frontotemporal Dementia. Here is the latest research on the nuclear pore complex in ALS and FTD.

Applications of Molecular Barcoding

The concept of molecular barcoding is that each original DNA or RNA molecule is attached to a unique sequence barcode. Sequence reads having different barcodes represent different original molecules, while sequence reads having the same barcode are results of PCR duplication from one original molecule. Discover the latest research on molecular barcoding here.

Chronic Fatigue Syndrome

Chronic fatigue syndrome is a disease characterized by unexplained disabling fatigue; the pathology of which is incompletely understood. Discover the latest research on chronic fatigue syndrome here.

Evolution of Pluripotency

Pluripotency refers to the ability of a cell to develop into three primary germ cell layers of the embryo. This feed focuses on the mechanisms that underlie the evolution of pluripotency. Here is the latest research.

Position Effect Variegation

Position Effect Variagation occurs when a gene is inactivated due to its positioning near heterochromatic regions within a chromosome. Discover the latest research on Position Effect Variagation here.

STING Receptor Agonists

Stimulator of IFN genes (STING) are a group of transmembrane proteins that are involved in the induction of type I interferon that is important in the innate immune response. The stimulation of STING has been an active area of research in the treatment of cancer and infectious diseases. Here is the latest research on STING receptor agonists.

Microbicide

Microbicides are products that can be applied to vaginal or rectal mucosal surfaces with the goal of preventing, or at least significantly reducing, the transmission of sexually transmitted infections. Here is the latest research on microbicides.

Related Papers

Biochemical and Biophysical Research Communications
B V BronkJ D Regan
Radiotherapy and Oncology : Journal of the European Society for Therapeutic Radiology and Oncology
Helen ZhaoRobert G Bristow
Nihon yakurigaku zasshi. Folia pharmacologica Japonica
Naoki Kajiwara, Futoshi Shibasaki
© 2021 Meta ULC. All rights reserved