Pseudomonas aeruginosa NfsB and nitro-CBI-DEI--a promising enzyme/prodrug combination for gene directed enzyme prodrug therapy.

Abstract

BACKGROUND:

The nitro-chloromethylbenzindoline prodrug nitro-CBI-DEI appears a promising candidate for the anti-cancer strategy gene-directed enzyme prodrug therapy, based on its ability to be converted to a highly cytotoxic cell-permeable derivative by the nitroreductase NfsB from Escherichia coli. However, relative to some other nitroaromatic prodrugs, nitro-CBI-DEI is a poor substrate for E. coli NfsB. To address this limitation we evaluated other nitroreductase candidates from E. coli and Pseudomonas aeruginosa.

FINDINGS:

Initial screens of candidate genes in the E. coli reporter strain SOS-R2 identified two additional nitroreductases, E. coli NfsA and P. aeruginosa NfsB, as being more effective activators of nitro-CBI-DEI than E. coli NfsB. In monolayer cytotoxicity assays, human colon carcinoma (HCT-116) cells transfected with P. aeruginosa NfsB were >4.5-fold more sensitive to nitro-CBI-DEI than cells expressing either E. coli enzyme, and 23.5-fold more sensitive than untransfected HCT-116. In three dimensional mixed cell cultures, not only were the P. aeruginosa NfsB expressing cells 540-fold more sensitive to nitro-CBI-DEI than pure cultures of untransfected HCT-116, the activated drug that they generated also displayed an unprecedented local bystander effect.

CONCLUSION:

We posit that the discrepancy in the fold-sensitivity to nitro-CBI-DEI between the two and three dimensional cytotoxicity assays stems from loss of activated drug into the media in the monolayer cultures. This emphasises the importance of evaluating high-bystander GDEPT prodrugs in three dimensional models. The high cytotoxicity and bystander effect exhibited by the NfsB_Pa/nitro-CBI-DEI combination suggest that further preclinical development of this GDEPT pairing is warranted.

• Authors: Laura K Green, Sophie P Syddall, Kendall M Carlin, Glenn D Bell, Christopher P Guise, Alexandra M Mowday, Michael P Hay, Jeffrey B Smaill, Adam V Patterson, David F Ackerley
• Journal: Molecular cancer (Mol Cancer) Vol. 12 Pg. 58 (Jun 10 2013) ISSN: 1476-4598 [Electronic] England
• PMID: 23758947 (Publication Type: Journal Article, Research Support, Non-U.S. Gov't)

Chemical References

• Prodrugs
• Nitroreductases

Topics

• Bystander Effect
• Cell Line, Tumor
• Cell Proliferation (drug effects)
• Enzyme Therapy
• Gene Expression
• Genetic Therapy
• HCT116 Cells
• Humans
• Inhibitory Concentration 50
• Nitroreductases (genetics, metabolism)
• Prodrugs (metabolism, pharmacology, toxicity)
• Pseudomonas aeruginosa (enzymology, genetics)
• Tumor Stem Cell Assay