In this study we investigated the levels of two lysosomal
cysteine protease proteins cathepsin B (CB) and
cathepsin L (CL) and the levels of three
cysteine protease inhibitor proteins stefin A (SFA),
stefin B (SFB) and
cystatin C (CNC) in squamous-cell lung
carcinoma (SQCLC) and matched lung parenchyma specimens and examined the inhibition of CB and
cathepsin C (CC) activities by endogenous inhibitors in extracts from SQCLC,
lung adenocarcinoma (LAC) and lung parenchyma specimens. We found that Stage I SQCLCs contained significantly increased levels of CB
protein, CB activity and SFA
protein as compared to matched lungs. Neither the levels of CL
protein nor the levels of SFB
protein nor the levels of CNC
protein in Stage I SQCLCs and the lungs were significantly different, but the levels of CB and CL
proteins as well as the levels of SFA and SFB
proteins showed significant positive correlation in SQCLCs. In SQCLCs as well as in the lungs the level of SFB
protein was significantly higher than the level of SFA
protein or the level of CNC
protein. In the lungs the levels of SFA
protein and CNC
protein revealed a weak negative correlation trend. In extracts from SQCLCs the level of SFA
protein showed a weak negative correlation with the residual CB activity (i.e. the activity remaining after extract preincubation) whereas in extracts from the lungs the level of CNC
protein displayed a weak negative correlation trend with the residual CB activity and with the residual CC activity. We observed that SQCLCs and LACs contained not only a significantly increased activity of CB but also a significantly higher inhibitory potential against the activity of endogenous CB as compared to matched lungs.
Leupeptin, a small inhibitor of CB, was capable to protect CB in lung
carcinoma and lung parenchyma extracts from preincubation-induced inhibition, revealing an active-site directed and competitive nature of CB inhibition by endogenous
cystatins. Ultrafiltration passaged
protein preparations of nominal Mr < or = 30,000 obtained from extracts of SQCLCs inhibited significantly higher quantities of activity of purified bovine spleen CC than did such
protein preparations from matched lungs. Reaction courses of purified bovine spleen CC that had been preincubated with such
protein preparations resembled those of endogenous CC from SQCLC and lung extracts showing a slow steady-state approach. These observations and the relaxation kinetics of CC from SQCLC and lung extracts suggest that CC in the extracts may be complexed with some
cystatins. In conclusion, our results indicate that quantitatively different combinations of
cystatins are the major constituents of the inhibitory potential against CB and CC in SQCLCs and the lungs.