A useful cell system for studying the regulation of 17HSD/KSR type 2 activity and expression in ovarian epithelial cancer.

17ß-Hydroxysteroid dehydrogenase/17-ketosteroid reductase (17HSD/KSR) activity and 17HSD/KSR types 1, 2, 4, and 5 mRNA levels were characterized in ovarian cancer cell lines derived from patients unexposed to radiation or chemotherapy. Activity was at the limit of detection in TOV-112D and TOV-21G cells. Activity in OV-90 was comparable to that in human placental tissue, was predominantly microsomal and was 17HSD/KSR type 2-like in substrate specificity and inhibition patterns. In monolayers, conversion of testosterone (T) to androstenedione (A) was 12-fold greater than that of A to T. Reduction of fetal bovine serum to 0.3% in the culture medium had no effect on 17ß-HSD activity. Significant levels of type 1 and type 2 mRNAs were observed in OV-90 while only trace amounts were detected in TOV-21G. In contrast, type 4 mRNA levels were comparable for OV-90 and TOV-21G. Type 5 mRNA was detected in both cell lines but its level in OV-90 was twice that of TOV-21G. In OV-90, the type 2-like activity was predominant even though the type 5 mRNA level was 2.5-fold higher than that of the type 2. OV-90 cells may be a useful system for studying the regulation of 17HSD/KSR type 2 activity and expression in ovarian epithelial cancer.

Endometrial carcinoma expresses an increased cathepsin B/D ratio.

OBJECTIVES: Cathepsins B and D belong to a family of proteases involved in tumor invasion and metastasis. As such they may function as biomarkers for the aggressiveness of a given tumor. We examined the enzymatic activity of these proteins as well as the cellular and extracellular distribution of cathepsins B and D. METHODS: 39 snap frozen tissue samples were assayed for activity fluorometrically with cathepsin-specific peptide substrates in combination with specific inhibitors. 4 groups were established: benign tissue, stage I/grade 1, stage i/grade 3, and stage IIIC/any grade. IHC staining for cathepsin B with the percentage of counterstained enzyme calculated from each specimen. RESULTS: A significantly increased level of cathepsin B activity was seen in malignant tissue specimens when compared to benign tissue. The cathepsin B/D ratio confirmed and was required to detect the significance of this distinction for each malignant group when compared to benign samples. There were no differences in cathepsin B or D expression detected between the various malignant groups. IHC staining for cathepsin B was more diffuse in the malignant tissues. CONCLUSIONS: Malignant endometrium displays increased cathepsin B activity when compared benign samples. The cathepsin B/D ratio is increased for each of the malignant groups studied when compared directly to benign endometrium. The cathepsin B/D ratio cannot be utilized to distinguish the stage or grade between any of the malignant groups studied. This ratio may serve to distinguish malignant from benign tumor samples and may be a constitutive change in the malignant transformation.

Cathepsins B and D activity and activity ratios in normal ovaries, benign ovarian neoplasms, and epithelial ovarian cancer.

OBJECTIVE: Cathepsins B (CB) and D (CD) belong to a family of proteases felt to be important in tumor metastasis and invasion. It has been suggested that both enzymes play a role the progression of epithelial ovarian cancer and they have been investigated as potential biomarkers for ovarian cancer. Our objective was to determine if activity ratios of these two isoforms might enhance their usefulness as biomarkers. METHODS: Ovarian cancer cell lines and snap-frozen archived tissue samples were sonicated and cathepsin activities were assayed fluorometrically with cathepsin-specific peptide substrates in combination with specific inhibitors. Tissue specimens were divided into four groups: normal ovary, benign neoplasm, early-stage (I/II) cancer, and late-stage (III/IV) cancer. Median CB and CD activity and the ratio of CB to CD (CB/CD) were compared using the Wilcoxon rank sum test. Nonparametric Spearman correlation was used to determine associations between CA-125 and cathepsin activity. Logistic regression was used to test the association between cathepsin activity and malignancy. RESULTS: In cell lines and tissue, CD activity remained relatively constant, while CB activity varied. CB activity was greatest in cancer tissue. Elevated serum CA-125 was associated with elevations in CB activity and CB/CD but not CD activity. Elevated CB activity and CB/CD as well as increasing CA-125 and age are all associated with malignancy. Multiple logistic regression shows that CB activity and age best predict malignancy status. CONCLUSIONS: CB activity is associated with invasive ovarian neoplasm. Our results do not suggest that the ratio of activity between CB and CD provides any additional information than CB activity alone. Both tissue CB activity and CB/CD activity ratios correlate with serum levels of CA-125; however there is no correlation between CD activity and CA-125.

Inhibition of 3alpha-hydroxysteroid dehydrogenase (3alpha-HSD) activity of human lung microsomes by genistein, daidzein, coumestrol and C(18)-, C(19)- and C(21)-hydroxysteroids and ketosteroids.

Epidemiologic data suggest a relationship between dietary intake of phytochemicals and a lower incidence of some cancers. Modulation of steroid hormone metabolism has been proposed as a basis for this effect. It has been shown that aromatase, 3beta-hydroxysteroid dehydrogenase and 17beta-hydroxysteroid dehydrogenase (17beta-HSD) are inhibited by the isoflavones, genistein and daidzein, and by coumestrol. In general, the extent of inhibition has been expressed in terms of IC50-values, which do not give information as to the pattern of inhibition, i.e., competitive, non-competitive, or mixed. Less is known of the effects of these compounds on 3alpha-HSD. The human lung is known to have a high level of 17beta-HSD and 3alpha-HSD activity. During the course of studies to characterize both activities in normal and inflamed lung and lung tumors we noted that 3alpha-HSD activity with 5alpha-DHT of microsomes from normal, adult lung was particularly susceptible to inhibition by coumestrol. To clarify the pattern of inhibition, the inhibition constants Ki and K'i were evaluated from plots of 1/v versus [I] and [S]/v versus [I]. Genistein, daidzein and coumestrol gave mixed inhibition patterns versus both 5alpha-DHT and NADH. In contrast, 5alpha-androstane-3,17-dione and 5alpha-pregnane-3,20-dione were competitive with 5alpha-DHT. NAD inhibited competitively with NADH. Our findings demonstrate that phytochemicals have the potential to inhibit 5alpha-DHT metabolism and thereby affect the androgen status of the human lung. The observation of a mixed inhibition pattern suggests these compounds bind to more than one form of the enzyme within the catalytic pathway.