Camille Mallouh

GLYOXALASE I ACTIVITY IN HUMAN PROSTATE CANCER: A POTENTIAL MARKER AND IMPORTANCE IN CHEMOTHERAPY

PURPOSE To provide information on the activity of Gly-I in prostate cancer. MATERIALS AND METHODS We performed qualitative Gly-I assay on prostate tissues. RESULTS Gly-I activity between prostate cancer and noncancerous specimens differed substantially and significantly, although such activity also varied somewhat among cancer specimens. CONCLUSIONS Gly-I activity is indeed higher in cancerous than in noncancerous specimens, suggesting that it may play a role in prostate cancer homeostasis and survival.

TNF‐mediated cytotoxicity and resistance in human prostate cancer cell lines

The contribution of TNF receptor (TNF‐R) expression was investigated with respect to TNF sensitivity or insensitivity for androgen‐dependent and androgen‐independent human prostate cancer (PCA) cell lines, respectively.

Effect of tumor necrosis factor-α and interferon-γ on the growth of human prostate cancer cell lines

Human recombinant tumor necrosis factor-α (rTNF-α, 10-12–10-8 M) inhibited the proliferation of androgen-dependent LNCaP cells by 32–56%. In contrast, proliferation of androgen-independent PC-3 and JCA-1 cells was only slightly inhibited, or not inhibited at all, respectively. Human recombinant interferon-γ (rIFN-γ, 500 U/ml) decreased proliferation of PC-3 and JCA-1 cells by 35% and 53%, respectively, but had no effect on LNCaP cells. Interestingly, the combination of rIFN-γ and TNF-α had greater antiproliferative effects on JCA-1 cells than treatment with either cytokine alone. However, the antiproliferative effects of this combination were similar to those observed for PC-3 or LNCaP cells treated with rIFN-γ or TNF-α alone, respectively. These data suggest that some forms of androgen-independent prostate cancer may benefit from a combination therapy of IFN-γ and TNF-α, while the use of IFN-γ alone may be more efficacious in others.

The effects of brefeldin A (BFA) on cell cycle progression involving the modulation of the retinoblastoma protein (pRB) in PC-3 prostate cancer cells.

PURPOSE To investigate the effects of brefelding A (BFA) on the growth of the androgen-independent human prostate cancer PC-3 cells, focusing on cell cycle regulation. MATERIALS AND METHODS BFA is a fungal macrocyclic lactone with an antiviral activity. PC-3 cells were cultured with various concentrations of BFA for indicated times and cell growth was monitored at each time point. Cell cycle analysis was performed to explore the mechanism of BFA-induced growth inhibition. To further investigate the cell cycle regulation, cell cycle-controlling factors, such as the retinoblastoma gene product (pRB) and its regulatory components cdk2, cdk4, and cyclin D1, were analyzed by Western immunoblots. RESULTS BFA was a potent growth inhibitor at a concentration of 30 ng./ml., resulting in a > 70% reduction in cell number at 3 days. Cell cycle analysis revealed a cell arrest in the G1 to S phase transition. Western blots further showed that BFA induced dephosphorylation of pRB accompanied by down regulation of cdk2, cdk4, and cyclin D1 expression. The extended pRB dephosphorylation in control cell lysates was also observed by the addition of BFA-treated lysates, but was prevented by the inclusion of phosphatase inhibitors in assay mixtures. CONCLUSION These results suggest that BFA may be a potent cell cycle modulator, which post-translationally regulates pRB phosphorylation possibly by down-regulating cdk2, cdk4, and cyclin D1 and/or by up-regulating a phosphatase(s) capable of dephosphorylating pRB. Thus, BFA-induced growth inhibition in PC-3 cells appears to be at least partially due to the modulation of a pRB-mediated growth pathway.

ANALYSIS OF CATHEPSIN D FORMS AND THEIR CLINICAL IMPLICATIONS IN HUMAN PROSTATE CANCER

PURPOSE To assess cathepsin D (Cat.D) status in the prostate, we analyzed the different Cat.D forms in human prostate tissues using Western immunoblots. MATERIALS AND METHODS Cell extracts were prepared from prostate tissues (n = 42) obtained from radical prostatectomy, adopting the tissue homogenization method. Expression of the different Cat.D forms was analyzed using Western blots. The catalytic activity of Cat.D was assayed by acid treatment, in which cell extracts were incubated in acidic buffer (pH 3 to 4) at 37C for 1 hour. RESULTS Pathologically confirmed normal (NML), benign prostatic hyperplasia (BPH) and cancer (CAP) specimens all expressed Cat.D, but as two distinct forms. Both NML and BPH predominantly expressed an inactive procathepsin D (Pro.Cat.D), while CAP notably exhibited an active mature Cat.D. The assessment of Cat.D activity, using PSA (prostate specific antigen) as a physiological substrate, showed that such activity was consistently higher in CAP than in NML/BPH specimens. Further studies revealed that the mode of Cat.D activation in CAP specimens appeared to be primarily due to acid-induced autoproteolysis (self-degradation) of mature Cat.D. CONCLUSION This study demonstrates that expression and activity of Cat.D varies among prostate specimens. A greater expression of mature Cat.D with a higher catalytic activity in CAP specimens is the most notable difference from NML/BPH. Therefore, the differential expression/activity of Cat.D forms may be a useful indicator for assessing prostate cancer status.

Methylglyoxal–Induced Apoptosis in Human Prostate Carcinoma: Potential Modality for Prostate Cancer Treatment

Objective: To examine the cellular effects of methylglyoxal (MG), a toxic physiological metabolite, on human prostatic cancer PC–3 cells.Methods: The effects of MG on cell growth and viability were evaluated first, and then its effects on the cell cycle and the glycolytic process were analyzed by Western blots and specific assays. Possible MG–induced apoptosis was also assessed by DNA analysis using agarose gel electrophoresis.Results: MG ≥3 mM caused severe growth inhibition, resulting in nearly 100% cell death by 24h. The time course study revealed that expression of cyclin D1, cdk2, and cdk4 was significantly (>50%) downregulated in 3 h of MG (3 mM) exposure, followed by the dephosphorylation of retinoblastoma protein by 6 h. Both the glyceraldehyde 3phosphate dehydrogenase activity and the cellular lactate level were also reduced by ∼50 and 80%, respectively, following 6–hour MG exposure. Induction of apoptosis by MG was indicated by partial degradation of poly(ADP–ribose) polymerase and further confirmed by discrete DNA fragmentation detected on an agarose gel. Conclusion: MG is capable of inducing apoptosis in prostatic cancer PC–3 cells, due primarily to a blocking of the cell cycle progression (G1 arrest) and glycolytic pathway. Therefore, MG could be a potent apoptosis inducer, which may have a potential for prostate cancer treatment.

Glyoxalase I phenotype as a potential risk factor for prostate carcinoma

OBJECTIVES To elicit a possible link between glyoxalase I (Gly-I), a detoxifying enzyme, and the incidence of prostate cancer (PCa), we investigated Gly-I phenotypic expression in the prostatic tissue and red blood cells (RBCs) from patients with PCa. METHODS Eighty-seven clinical specimens, including 42 PCa tissue samples, 20 RBC samples, and 25 matched pair (prostate and RBC) samples from patients at prostatectomy were examined. The Gly-I phenotypes in these specimens were assessed by nondenaturing starch-polyacrylamide gel electrophoresis. RESULTS Of the 87 patients, 63 (72.4%) were white, 15 (17.2%) were black, and 9 (10.4%) were another ethnicity (eg, Hispanic, Asian, Indian). Three Gly-I phenotypes were detected in these specimens as fast, intermediate, and slow-moving bands on the gel. The fast phenotype was the most common form found in the white (34 [54%] of 63) and black (8 [53.3%] of 15) patients, but the third ethnic group was too small for proper analysis. To validate this finding, the data from the white patients were compared with the Gly-I phenotypic frequencies in U.S. populations. The data analysis confirmed that a higher incidence (54%) of the fast type in our white patients was statistically significant (P <0.0001) compared with its phenotypic frequency of 30.6% in the general U.S. white population. CONCLUSIONS The significantly high frequency (P <0.0001) of the fast Gly-I phenotype was detected among patients with PCa, suggesting it is a potential risk factor for PCa. Whether its increased incidence in whites reflects the lack of sample numbers for other ethnic groups needs additional investigation.

Unusual primary prostatic malignancies

Atypical prostatic cancer is defined as any malignancy of prostatic origin, excluding the common acinar prostatic adenocarcinoma. These atypical prostate cancers account for less than 5 percent of all prostatic malignancies. Because atypical prostatic malignancies occur rarely, little is known about their biologic behavior. To date, no large series of patients have been reported, and specific treatment protocols have yet to be defined. Herein we describe 5 new cases of atypical prostatic cancer. By reporting the occurrence of new cases, we believe a better understanding of the natural history of these lesions will emerge.

Role of cathepsin D in prostatic cancer cell growth and its regulation by brefeldin A

Abstract We investigated a possible relationship between brefeldin A (BFA), an antibiotic, and cathepsin D (Cat.D), a lysosomal protease, in prostate cancer proliferation. Effects of BFA (30 ng/ml) were examined on the growth of three human prostatic cancer cell lines, PC-3, DU-145, and LNCaP cells. Its effect on Cat.D in these cancer cells was assessed by Western blots and compared with Cat.D expressed in clinical prostate specimens (n=55). BFA profoundly (>70%) inhibited the growth of all three cancer cell lines. Western blots revealed that expression of procathepsin D (Pro.Cat.D) was markedly increased with BFA, whereas actively proliferating (control) cells greatly exhibited mature Cat.D. Analysis of prostate specimens then showed predominant Pro.Cat.D expression in non-cancerous tissues while also showing enhanced expression of mature Cat.D in all cancer specimens. Therefore, BFA-induced growth inhibition in prostatic cancer cells is associated with a blocking of Cat.D maturation (activation), suggesting a possible role of Cat.D in prostate cancer proliferation/development.

Depressed myocardial function after transurethral resection of prostate.

Cardiovascular physiologic monitoring was undertaken in 12 patients undergoing transurethral resection of the prostate with the aid of flow-directed Swan-Ganz catheter and the Automated Physiologic Profile. Cardiac and pulmonary pressures and physiologic parameters were derived pre- and postoperatively. Resecting time, body temperature, intravenous fluid administered, serum hemoglobin, and sodium also were recorded. Of the 12 patients studied, 66 per cent experienced a drop in their cardiac index as well as their left ventricular function after surgery. Myocardial function curves revealed that 7 patients (58 per cent) had decreased cardiac function, 2 had no change, and 3 had increased function. Four patients with preoperative pulmonary wedge pressures (PAW) over 9 mm. Hg experienced depressed cardiac function. Three patients were resected for over sixty minutes, and all experienced depressed cardiac function. Vital signs, serum hemoglobin, or serum sodium did not reflect this change. We believe that relative hypervolemia, undetected elevation of pulmonary wedge pressure. We believe that relative hypervolemia, undetected elevation of pulmonary wedge pressure, and prolonged resection are factors that depress cardiac function and increase the risk of cardiovascular complication in transurethral surgery.