Cathepsin B (CB) is involved in degradation of
extracellular matrix proteins during
tumor progression in human solid organ
tumors (such as colorectal, bladder, and breast
cancers), including human
prostate cancer. Its activities are regulated by endogenous inhibitors (such as
stefins or
cystatins). Increased expression of
cathepsin B message,
protein, and membrane association have been linked to
malignancy, but there are very few studies of their
mRNA expression in
prostate cancer using in situ hybridization techniques. Our objective was to determine the relationship of CB and
stefin A (
cystatin A)
mRNA localization to the Gleason grading system for histologic scores in the hope of distinguishing aggressive and less aggressive variants of
prostate cancer. We used a 25-base biotinylated
oligonucleotide CB
cDNA antisense probe to localize CB message and a 27-base biotinylated
oligonucleotide stefin A cDNA antisense probe to localize
stefin A message. Prostate samples from 41
prostatectomy patients were collected along with their pre-surgery serum PSA levels and clinical stage of the disease. Sections prepared from frozen prostate tissue samples were hybridized with the CB and
stefin A and control pBR 322 probes using techniques reported by Sinha et al. [1] and their distribution quantitated by an image analysis system. Prostate sections treated with
RNAse before hybridization or incubated with the pBR 322 control probe showed little or no reaction products, confirming that localization of CB and
stefin A probes was specific. In
prostate cancer, the reaction products were found in neoplastic and invasive cells and occasionally in stromal cells. The ratios of CB to
stefin A were similar in normal prostate and
benign prostatic hyperplasia (BPH) whereas they varied consistently within and between Gleason histologic scores for
prostate cancer. These variations showed three localization patterns; namely,
prostate cancers with higher levels of CB than
stefin A, lower levels of CB than
stefin A, and similar levels of CB and
stefin A. All three patterns and ratios for CB and
stefin A were found in prostate samples (22/41) represented by the Gleason histologic score 6
tumors. In these
tumors, serum PSA levels ranged from 1 to 78 ng/ml and
prostate cancers showed B, C, and D clinical stages. There was no correlation of CB/
stefin A ratio and serum PSA values or clinical stage in a limited number of
prostate cancer cases. Our data showed that there were
prostate cancer cases within Gleason histologic scores which expressed high, similar, and low levels of CB when compared to
stefin A. We postulate that
prostate cancer cases showing higher levels of CB compared to
stefin A probably represent an aggressive variant of this
cancer within any one Gleason histologic score. If this is the case, aggressive variants of
prostate cancer would occur within Gleason scores 3 to 10 even though higher scores are usually considered more aggressive forms of
prostate cancers. Since our study is based upon a very limited number of frozen prostate samples, we emphasize that a larger series of archival
prostate cancer samples along with their survival data should be analyzed to establish any relationship of CB/
stefin A ratio and aggressive variants of this
cancer. Therefore, our conclusion is tentative. Our study provides a partial explanation for differences in the
clinical course of
prostate cancer in patients. This is the first study to show that determination of CB and
stefin A mRNA ratios may lead to identification of aggressive and less aggressive variants of
prostate cancer within a Gleason histologic score.