Pyrrolidine dithiocarbamate exerts anti-proliferative and pro-apoptotic effects in renal cell carcinoma cell lines.
BACKGROUND
The activation of nuclear factor-kappaB (NF-kappaB) has been implicated in the development, progression and metastasis of renal cell carcinoma (RCC). This study investigates the effect of pyrrolidine dithiocarbamate (PDTC), a NF-kappaB inhibitor, on two metastatic human RCC cell lines, ACHN and SN12K1.


METHODS
RCC cell lines and normal cells were exposed to 25 or 50 microM of PDTC. Apoptosis was measured by flow cytometry and TdT-mediated nick end labelling methods. Cell viability and proliferation were measured by MTT and BrdU assays, respectively. Expression of NF-kappaB subunits, IkappaBs, IkappaB Kinase (IKK) complex and apoptotic regulatory proteins were analysed by western blotting and/or immunofluorescence. DNA-binding activity of NF-kappaB subunits were measured by ELISA.


RESULTS
RCC cell lines had a higher basal level expression of all the five subunits of NF-kappaB than normal primary cultures of human proximal tubular epithelial cells or HK-2 cells. PDTC decreased the viability and proliferation of RCC, but not normal cells. Of the two RCC cell lines, ACHN had a higher basal level expression of all the five NF-kappaB subunits than SN12K1 and was more resistant to PDTC. While PDTC induced an overall decrease in expression of all the five NF-kappaB subunits in both RCC cell lines, unexpectedly, it increased the nuclear expression of NF-kappaB in ACHN, but not in SN12K1. PDTC reduced the DNA-binding activity of all the NF-kappaB subunits and the expression of the IKK complex (IKK-alpha, IKK-beta and IKK-gamma) and the inhibitory units IkappaB-alpha and IkappaB-beta. PDTC induced a significant increase in apoptosis in both RCC cell lines. This was associated with a decrease in expression of the anti-apoptotic proteins, Bcl-2 and Bcl-(XL), without marked changes in the pro-apoptotic protein Bax.


CONCLUSIONS
These data suggest that PDTC has the potential to be an anticancer agent in some forms of RCC.
Morais
Christudas
C
Conjoint Renal Laboratory, Bancroft Centre, Queensland Health Pathology Service, Royal Brisbane and Women's Hospital, 300 Herston Road, Brisbane, Queensland 4006, Australia.
Pat
Betty
B
Gobe
Glenda
G
Johnson
David W
DW
Healy
Helen
H
D017209:Apoptosis / Q000187:drug effects*; D002292:Carcinoma, Renal Cell / Q000188:drug therapy* / Q000473:pathology*; D045744:Cell Line, Tumor; D049109:Cell Proliferation / Q000187:drug effects*; D006801:Humans; D007680:Kidney Neoplasms / Q000188:drug therapy* / Q000473:pathology*; D016328:NF-kappa B / Q000037:antagonists & inhibitors*; D011759:Pyrrolidines / Q000627:therapeutic use*; D013859:Thiocarbamates / Q000627:therapeutic use*
D016328:NF-kappa B; D011759:Pyrrolidines; D013859:Thiocarbamates; C020972:pyrrolidine dithiocarbamic acid