Treatment > Gene Therapy

Meeting Abstract

University of Michigan, Ann Arbor, MI; Medical College of Ohio, Toledo, OH

Transfer of the herpes simplex virus type 1 thymidine kinase (HSV-TK) gene into tumor cells allows phosphorylation of the antiviral drug ganciclovir (GCV). 
Incubation with GCV results in cytotoxicity of not only the HSV-TK-expressing cells, but also neighboring bystander cells that do not express HSV-TK. 
Bystander cytotoxicity has been attributed to the transfer of phosphorylated GCV from HSV-TK-expressing cells to neighboring bystander cells through gap junction intercellular communication (GJIC). 
We previously reported that SW620 colon carcinoma cells exhibited good bystander cytotoxicity despite low (<3%) GJIC. 
We have now extended these results to determine if bystander killing can occur with GCV in cells that are completely lacking GJIC. 
For these studies, we used both HeLa cells, which have been reported to be deficient in GJIC connexin proteins and exhibit 0% communication, and U251 human glioblastoma cells transfected with a dominant-negative connexin43 (DN14 cells). 
GJIC of parental U251 cells (80%) was eliminated in DN14 cells as determined by Lucifer Yellow (LY) microinjection. 
GCV-mediated bystander cytotoxicity was decreased in DN14 cells compared to that of the U251 parent cell line. 
In co-cultures of HSV-TK-expressing and non-expressing HeLa cells incubated with GCV, no significant bystander killing was observed. 
However, in these co-cultures there was increased survival of HSV-TK-expressing cells (IC90 = 100 μM) compared to survival in cultures of 100% HSV-TK-expressing cells (IC90 = 2.5 μM) suggesting that the presence of bystander cells decreased the level of phosphorylated GCV in the HSV-TK-expressing cells. 
Further analysis using a double dye flow cytometry technique demonstrated greater GJIC compared to LY microinjection. 
Over a 24 hour period, GJIC was ~ 40% in the DN14 cell line and ~ 25% in the HeLa cell line. 
These data suggest that during prolonged drug incubation, low levels of GJIC are sufficient to transfer phosphorylated GCV from the HSV-TK-expressing cells to neighboring bystander cells resulting in bystander cytotoxicity and/or decreased cytotoxicity to the HSV-TK-expressing cells.

Copyright © 2003 American Association for Cancer Research. All rights reserved.