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N Kyprianou

Publications and source records attributed to N Kyprianou.

At least 37 records · Page 2Linked to original sources
PubMed1999

Loss of cell cycle regulators p27(Kip1) and cyclin E in transitional cell carcinoma of the bladder correlates with tumor grade and patient survival.

The cyclin-dependent kinase inhibitor p27(Kip1) is a powerful molecular determinant of cell cycle progression. Loss of expression of p27(Kip1) has been shown to be predictive of disease progression in several human malignancies. In this study we investigated the expression of two key cell cycle regulators, p27(Kip1) and cyclin E, in the progression of transitional cell carcinoma of the bladder. An immunohistochemical analysis was conducted in a series of 50 bladder tumor specimens, including 3 metastatic lymph nodes, and 7 normal bladder specimens, using specific antibodies against the two regulators of the cell cycle, p27(Kip1) and cyclin E. The degree of immunoreactivity was correlated with the pathological tumor grade, stage, and patient survival. A uniformly intense immunoreactivity for p27(Kip1) and cyclin E was observed in epithelial cells of normal bladder tissue. Malignant bladder tissue demonstrated a heterogeneous pattern of significantly reduced p27(Kip1) and cyclin E immunoreactivity, compared with normal urothelium (P < 0.01). In addition, there was progressive loss of expression of both cell cycle proteins with increasing tumor grade and pathological stage. Expression of p27(Kip1) was significantly lower in the poorly differentiated tumors (grades III) compared to well and moderately differentiated (grades I and II) tumors (P = 0.004). Moreover, the expression of cyclin E was lower in grade III tumors compared to grade I and II lesions, although this difference failed to reach statistical significance. Most significantly, Kaplan-Meier plots of patient survival show increased mortality risk associated with low levels of p27(Kip1) (P = 0.001) and cyclin E (P = 0.002) expression. This is the first evidence that loss of expression of p27(Kip1) and cyclin E in human bladder transitional cell carcinoma cells correlates with advancing histological aggressiveness and poor patient survival. These results have clinical importance, because they support a role for p27(Kip1) and cyclin E as novel predictive markers of the biological potential of bladder tumors that will enable identification of those tumors most likely to progress to muscle invasive disease and of patient survival.

Adult↗
PubMed1999

Alpha 1-adrenoceptor antagonists terazosin and doxazosin induce prostate apoptosis without affecting cell proliferation in patients with benign prostatic hyperplasia.

PURPOSE: Recent evidence indicated that an alpha 1 blocker, doxazosin, induces prostate apoptosis in patients with benign prostatic hyperplasia (BPH). In this study, to determine whether this apoptotic response was mediated by alpha 1 adrenoceptor-dependent mechanism or was specific to doxazosin, we examined the effect of another alpha 1 blocker, terazosin, in addition to doxazosin, on the dynamics of prostate cell growth. MATERIALS AND METHODS: Cell proliferation and apoptosis were evaluated in BPH patients, an untreated (control) group (n = 31), and men treated with terazosin (n = 42) and doxazosin (n = 61) for the relief of the obstructive symptoms. Terazosin (1 to 10 mg./day) and doxazosin (2 to 8 mg./day) treatment varied from 1 week to 3 years. Ki-67 immunostaining and the TUNEL assay were used to evaluate the proliferative and apoptotic indices, respectively, in both the epithelial and stromal components of prostate (biopsy and prostatectomy) specimens. The smooth muscle cell content of the prostatic stroma was identified on the basis of smooth muscle alpha-actin immunoreactivity. RESULTS: A significant induction of apoptosis was observed in both the prostatic epithelial and stromal cells within the first month of terazosin and doxazosin therapy, as compared with untreated controls (p < 0.05). Furthermore, the marked induction of prostatic stroma apoptosis in response to both alpha 1 adrenoceptor antagonists was paralleled by a significant decrease in the smooth muscle alpha-actin expression. This loss of prostatic smooth muscle cells correlated with morphological stromal regression (as detected by trichrome staining) and BPH symptom improvement. Neither terazosin nor doxazosin therapy resulted in significant changes in prostate cell proliferation. CONCLUSIONS: These findings demonstrate that alpha-blockers as a class may regulate prostate growth by inducing apoptosis in both the epithelial and stromal cells, with little effect on cell proliferation. Apoptosis-mediated prostate stromal regression appears as a molecular mechanism underlying the therapeutic response to alpha 1 blockade in the treatment of BPH.

Adrenergic alpha-Antagonists↗
PubMed1998

Tumor-targeted apoptosis by a novel spermine analogue, 1,12-diaziridinyl-4,9-diazadodecane, results in therapeutic efficacy and enhanced radiosensitivity of human prostate cancer.

Interference with polyamine transport and biosynthesis has emerged as an important anticancer strategy involving polyamine analogues and specific inhibitors of key biosynthetic enzymes. Because the prostate gland has a high polyamine content, by using the polyamine transporter for selective uptake into cancer cells, alkylating polyamines are likely to be highly effective against prostatic tumors. We have recently synthesized a novel class of spermine analogues, the lead compound of which has efficacy against human cancer cells (P. S. Callery et al., U. S. patent, 5,612,239, Issued March 17, 1997.). In this study, to investigate the potential therapeutic efficacy of the lead spermine analogue 1,12-diaziridinyl-4, 9-diazadodecane (BIS), against advanced prostate cancer, we examined the in vitro effect and in vivo efficacy of the compound in two androgen-independent human prostate cancer cell lines, PC-3 and DU-145. BIS exhibited a dose-dependent cytotoxic effect against prostate cancer cells via induction of apoptosis. Treatment of cells with BIS (1 microM) for 24 h resulted in a significant induction of apoptosis (24%). Exposure of BIS-treated PC-3 prostate cancer cells to gamma-irradiation resulted in a significant increase in the number of cells undergoing apoptosis and a subsequent decrease in the IC50. Furthermore, BIS treatment led to a significant enhancement of loss of clonogenic survival in irradiated prostate cancer cells (both PC-3 and DU-145). In vivo efficacy trials demonstrated a significant antitumor effect of BIS against both PC-3 and DU-145 tumor xenografts in severe combined immunodeficient mice in a dose-dependent pattern at maximally tolerated doses. Terminal transferase end-labeling analysis indicated that BIS-mediated tumor regression in vivo occurs via induction of apoptosis among prostatic tumor cells. These results suggest that the novel spermine analogue BIS: (a) has a potent antitumor effect against prostatic tumors via induction of apoptosis; and (b) increases the radiosensitivity of human prostate cancer cells by decreasing the apoptotic threshold to radiation. This study may have important clinical implications for the manipulation of this antitumor activity of the polyamine analogue for the optimization of the therapeutic efficacy of radiation in patients with advanced prostate cancer.

Animals↗
PubMed1998

Induction of prostate apoptosis by doxazosin in benign prostatic hyperplasia.

PURPOSE: The molecular mechanisms underlying the therapeutic effect of the alpha1 blocker, doxazosin, on benign prostatic hyperplasia (BPH) are poorly understood. We evaluated the effect of doxazosin on cell proliferation and apoptosis in the prostatic glandular epithelium and stroma of patients with BPH. MATERIALS AND METHODS: We examined proliferative and apoptotic activities in prostate specimens of 22 men a mean of 65 years old with BPH before and after doxazosin treatment within the normal therapeutic range. Proliferative and apoptotic indexes were determined using Ki-67 nuclear antigen staining and the terminal transferase end labeling assay, respectively. The smooth muscle cell content in prostatic specimens was identified by smooth muscle alpha-actin, and desmin immunoreactivity and apoptotic indexes were correlated with prostatic stromal tissue regression and improvement in BPH symptoms. RESULTS: In response to doxazosin treatment there were no significant changes in the kinetics of cell proliferation in the prostatic epithelial or stromal cell population. Mean pretreatment baseline apoptosis was 1.9 and 1.0% for the epithelial and stromal prostate components, respectively. Mean apoptotic indexes significantly increased after 3 months of doxazosin treatment in the glandular epithelial (6%) and smooth muscle cells (15%). By 12 months after treatment epithelial apoptosis had decreased to constitutive levels, while the apoptotic index of prostatic stroma cells remained high. Doxazosin induced smooth muscle cell apoptosis correlated with prostatic stromal degeneration, decreased alpha-smooth muscle actin expression and improved BPH symptoms. CONCLUSIONS: These findings implicate the induction of prostate apoptosis by doxazosin as a potential molecular mechanism underlying the acute and chronic therapeutic responses of BPH to alpha1 blockade.

Adrenergic alpha-Antagonists↗
PubMed1998

Effect of estrogen withdrawal on nitric oxide synthase expression and apoptosis in the rat vagina.

OBJECTIVES: The physiology of the female sexual response and its molecular mediators remain poorly understood. Nitric oxide (NO) is synthesized in neurons and is a potent relaxor of vascular and nonvascular smooth muscle. In this study, we hypothesize that vaginal atrophy and declining sexual function during menopause may be NO dependent. Using the rat as an experimental model, we examined the expression and topologic localization of vaginal NO synthase (NOS) and the concomitant induction of apoptosis under normal and estrogen-depleted conditions. METHODS: Thirty rats were categorized into six groups on the basis of phase of the estrous cycle or estrogen status after oophorectomy. The expression and cellular localization of NOS was examined in frozen sections using specific antibodies against neuronal (N-NOS) and endothelial NOS (E-NOS). Apoptotic cells were identified in situ using the terminal transferase technique (TUNEL). Trichome staining was performed in all specimens to determine smooth muscle/collagen ratios. RESULTS: N-NOS immunoreactivity was localized to nerve fibers supplying vaginal smooth muscle, perivascular nerve plexuses, and lamina propria. E-NOS was localized to vascular endothelium and perivascular smooth muscle fibers. Both E-NOS and N-NOS expression in intact cycling animals was highest during proestrous and lowest during metestrous. After oophorectomy, levels of both N-NOS and E-NOS declined substantially compared with those of intact animals, and there was a parallel induction of apoptosis. Estrogen withdrawal also resulted in increased vaginal atrophy, intramural collagen accumulation, and perivascular wall thickening, as identified by trichome staining. Estrogen replacement resulted in a significant increase in E-NOS and N-NOS expression, as well as diminished apoptosis and vaginal atrophy. CONCLUSIONS: This cellular distribution of NOS in the rat vagina suggests that NO may modulate both vaginal blood supply and vaginal smooth musculature. Estrogen appears to play a critical role in concomitantly regulating vaginal NOS expression and apoptosis in nerves, smooth muscle, vascular endothelium, and epithelium of the rat vagina. These findings may have significant clinical implications for the pathophysiology of postmenopausal female sexual dysfunction.

Animals↗
PubMed1998

bcl-2/bax ratio as a predictive marker for therapeutic response to radiotherapy in patients with prostate cancer.

OBJECTIVES: Markers predictive of therapeutic response of prostatic tumors to radiotherapy may have major significance in optimizing effective treatment of prostate cancer. Because inherent cellular radioresistance plays a critical role in the failure of radiotherapy, in this study, we investigated whether there is a correlation between the ratio of two apoptosis regulators, bcl-2 (apoptosis suppressor) and bax (apoptosis inducer) in prostatic tumors and the clinical response to radiotherapy in patients with localized prostate cancer. METHODS: A retrospective review of records of 41 patients who underwent external beam radiotherapy for prostate cancer was conducted. On the basis of post-treatment prostate biopsy and prostate-specific antigen (PSA) criteria, the cancers of 20 patients were classified as radiation nonresponders and 21 as radiation responders. Immunohistochemical analysis was performed on paraffin-embedded prostate sections to determine the level of expression of the two apoptotic proteins, bcl-2 and bax, in tumor cells. RESULTS: bcl-2 immunoreactivity was significantly higher in prostatic tumors not responsive to radiotherapy (38.6+/-4.1), compared with the radiation responders (24.1+/-4.6) (P <0.001). Expression of bax protein was lower in nonresponders, but values were not significantly different from the responders. The resulting significantly higher bcl-2/bax ratio (P <0.01) correlated with poor therapeutic responsiveness of prostate cancer to radiotherapy (1.12+/-0.12 and 0.56+/-0.13, for nonresponders and responders, respectively). This correlation (r=0.67) was independent of age, PSA, and Gleason score. CONCLUSIONS: These findings suggest that patients with an elevated bcl-2/bax ratio are at increased risk of their cancer failing to respond to radiotherapy. This study suggests a predictive value for the bcl-2/bax ratio as a potential molecular marker for predicting radioresistance of prostatic tumors.

Aged↗
PubMed1998

Overexpression of transforming growth factor (TGF) beta1 type II receptor restores TGF-beta1 sensitivity and signaling in human prostate cancer cells.

TGF-beta1 is a potent negative regulator of cell growth that transduces signals through interaction with type I and type II receptors that form a heteroduplex. Abnormal expression and mutational alterations of these receptors have recently been shown in several human malignancies. In previous studies, we have demonstrated reduced expression of both types of transforming growth factor (TGF) beta1 receptors in human prostate tumors. In this study, using the human prostate cancer cell line, LNCaP, which is refractory to TGF-beta1 and lacks type II receptor (R-II), we investigated whether overexpression of the R-II receptor can restore sensitivity to the negative growth effects of TGF-beta1. LNCaP cells were transfected with plasmid containing the full length of human TGF-beta R-II receptor cDNA sequence. Stable transfectant clones were selected for R-II mRNA and protein expression by Northern and Western analyses, respectively. The effect of TGF-beta on LNCaP R-II overexpressing clones was examined on the basis of: (a) growth inhibition (cell number); (b) DNA synthesis using the [3H]thymidine incorporation assay; (c) induction of cyclin-dependent-kinase inhibitors, p21WAF-1/Cip1, p27Kip1, and p15; and (d) colony-forming ability in soft agar. Both the cell number and the rate of DNA synthesis of R-II-overexpressing clones were significantly suppressed by exogenous TGF-beta1 in a dose-dependent manner, compared with control cell lines. Treatment of R-II cloned transfectants with TGF-beta1 induced a G1 arrest, which was accompanied by a transient increase in p21WAF-1/Cip1 and p27Kip1 expression at the mRNA and protein level. Furthermore, the LNCaP R-II transfectants analyzed exhibited a markedly reduced colony-forming ability. Our results indicate that overexpression of TGF-beta1 R-II receptor in LNCaP prostate cancer cells caused tumor growth inhibition by restoring the TGF-beta signaling mechanism and TGF-beta1 sensitivity.

Carrier Proteins↗
PubMed1997

Transient tyrosine phosphorylation of p34cdc2 is an early event in radiation-induced apoptosis of prostate cancer cells.

BACKGROUND: Previous studies have demonstrated that androgen-independent human prostate cancer cells undergo radiation-induced apoptosis. The present study investigated the early events that trigger the apoptotic response of prostate cancer cells after exposure to ionizing irradiation. METHODS: Human prostate cancer cells (PC-3) were exposed to single doses of ionizing irradiation, and the immediate protein phosphorylation events were temporally correlated with induction of apoptosis. Apoptosis among the irradiated cell populations was evaluated using the fluorescein-terminal transferase assay. RESULTS: The kinetics of phosphorylation of a Mr 34,000 substrate followed a transient course: an initial increase was observed after 10 min postirradiation, reaching maximum levels by 60 min, and the protein subsequently underwent rapid dephosphorylation. Subsequent analysis revealed that the substrate for this tyrosine phosphorylation is the serine/ threonine p34cdc2 protein kinase, a cell cycle regulatory protein that controls cell entry into mitosis. This enhanced phosphorylation temporally preceded the radiation-induced apoptotic DNA fragmentation as detected by the terminal transferase technique. Arresting the cells in G0/G1 phase by pretreatment with suramin totally abrogated radiation-induced phosphorylation of p34cdc2 protein at the tyrosine residue, indicating that this posttranslational modification occurs in cell populations that escape G2 arrest and undergo apoptosis in response to radiation. CONCLUSIONS: These results suggest that a rapid and transient phosphorylation of a protein that controls mitotic progression precedes and potentially triggers radiation-induced apoptosis in prostate cancer cells.

Apoptosis↗
PubMed1997

Down-regulation of protein and mRNA expression for transforming growth factor-beta (TGF-beta1) type I and type II receptors in human prostate cancer.

Transforming growth factor-beta (TGF-beta1) is a potent negative regulator of cell growth that transduces signals through interactions with type I and II receptors. Abnormal expression and mutational alterations of these receptors have been observed in several human malignancies. In this study, we investigated the expression of the two types of TGF-beta1 receptors, R-I and R-II, in a normal human prostate, primary prostate adenocarcinoma and lymph nodes with metastatic deposits. Expression of receptor proteins was examined by immunohistochemical analysis in paraffin-embedded prostatic tissue sections, and mRNA expression was determined by Northern blot and RT-PCR analysis. Uniformly strong immunoreactivity for both TGF-beta receptor proteins, R-I and R-II, was exclusively localized to the prostatic glandular epithelium of normal prostates. In contrast, tumor epithelial cells in primary and metastatic prostatic cancer specimens exhibited a weak heterogeneous immunoreactivity for both R-I and R-II receptors; 25% of primary prostatic tumors and 45% of the lymph nodes with metastases were totally negative for R-I and R-II expression, while the rest exhibited a significantly reduced immunoreactivity for both types of receptors compared to the normal prostate (p < 0.05). Moreover, there was a significant decrease in the expression of R-I and R-II mRNA, in all 20 primary prostatic tumors and 4 lymph nodes positive for metastases, indicating that the decreased protein expression was due to down-regulation of gene expression for the two receptors. Our findings imply that decreased expression of TGF-beta1 type I and type II receptors might be involved in prostate tumorigenesis.

Activin Receptors, Type I↗
PubMed1997

bcl-2 over-expression delays radiation-induced apoptosis without affecting the clonogenic survival of human prostate cancer cells.

In this study we evaluated the effect of over-expression of the bcl-2 gene, a potent apoptosis suppressor, on radiation-induced apoptotic cell death in 2 human prostate cancer cell lines, androgen-independent PC-3 cells and androgen-sensitive LNCaP cells. Cells were transfected with the bcl-2 gene and bcl-2 transfectant clones isolated under neomycin selection; bcl-2 gene integration and level of mRNA and protein expression in the cloned transfectants were examined by Southern, Northern and Western blot analyses, respectively. Parental, neo control and bcl-2-expressing cells were exposed to single or fractionated doses of ionizing irradiation, and the cellular response to radiation was determined at 24, 48 and 72 hr post-irradiation, on the basis of: (i) loss of cell viability, (ii) clonogenic survival and (iii) induction of apoptotic DNA fragmentation. At 24 hr post-irradiation all cell lines, i.e., parental and bcl-2 transfectants, failed to form colonies, though the majority of bcl-2-expressing cells did not exhibit apoptotic morphology; bcl-2 over-expression in both cell lines reduced apoptosis 48 hr post-irradiation from 20-25% to 5% at a dose of 2,000 cGy. By 72 hr, bcl-2 over-expression afforded a 3-fold protection from radiation-induced apoptosis. There was no significant difference, however, in the clonogenic survival of the parental and bcl-2-expressing cells. Furthermore, there was a 24 hr delay in induction of the apoptosis marker gene SGP-2/TRPM-2 in the bcl-2-expressing cells, co-incidental with the delay in apoptotic DNA fragmentation.

Apoptosis↗
PubMed1997

Loss of the cyclin-dependent kinase inhibitor p27(Kip1) protein in human prostate cancer correlates with tumor grade.

Loss of expression or mutational deletion of the cyclin-dependent kinase inhibitor p27(Kip1) has recently been implicated in malignant development. In this study, we investigated the relationship between p27(Kip1) protein expression and tumor grade in human prostate cancer by conducting an immunohistochemical analysis in a series of normal prostate, benign prostatic hyperplasia, and malignant prostate cancer specimens. The proliferative activity of prostatic tumors was determined on the basis of the Ki-67 nuclear antigen staining. A uniformly intense immunoreactivity for p27(Kip1) was localized to the nuclei of glandular epithelial cells of normal prostates. The benign glandular epithelia exhibited moderate immunostaining. In the malignant prostate tissue, however, a heterogeneous pattern of substantially reduced p27(Kip1) immunoreactivity was found among the glandular epithelial cells. The majority of primary prostate cancer specimens (68%) were totally negative for p27(Kip1) immunoreactivity, whereas the rest exhibited a significantly decreased p27(Kip1) expression, compared with the normal prostate (P < 0.01). Moreover, there was progressively diminished p27(Kip1) immunostaining with increased tumor grade. This loss of p27(Kip1) was associated with an increase in the proliferative index of prostatic tumors (r = 0.88). There was no significant relationship between p27(Kip) loss and the transforming growth factor beta receptor status of prostatic adenocarcinomas. These results indicate that frequent loss of the cyclin-dependent kinase inhibitor p27(Kip1) in human prostate cancer cells correlates with advancing histological aggressiveness, implicating deregulation of p27(Kip1) in prostate tumor progression.

Adenocarcinoma↗
PubMed1996

Incidence of apoptosis and cell proliferation in prostate cancer: relationship with TGF-beta1 and bcl-2 expression.

The incidence of programmed cell death (apoptosis) and cell proliferation was investigated in the normal and malignant human prostate to define the significance of their potential deregulation in human prostate cancer. The incidence of "spontaneous" apoptosis was analyzed using an in situ end-labeling procedure for detection of nucleosomal DNA fragmentation, as well as the pattern and topological localization of expression of the 2 proteins regulating apoptosis, TGF-beta1, and bcl-2, in 40 primary prostatic adenocarcinomas with varying tumor grades, 17 lymph nodes positive for metastatic prostate cancer and 9 normal prostate specimens. The basal level of cell proliferation of the different prostatic cell populations in the same specimens was determined, utilizing the Ki-67 nuclear antigen. Localized prostate cancer cells exhibited a relatively low rate of apoptosis, which was significantly lower than the apoptotic index of normal prostate glandular epithelial cells. Metastatic prostate tumor cells, however, exhibited a significantly higher apoptotic index compared with localized prostate cancer cells. A significant increase in the proliferative index was detected in prostatic tumors compared with the normal gland (5-fold), and there was an even more marked elevation in the proliferative index of the metastatic prostate tumor cells compared to the normal prostate epithelial cells (approximately 24-fold). Immunohistochemical analysis of normal and malignant prostate specimens revealed a predominant TGF-beta immunoreactivity in the glandular epithelial cells, while the stromal component was totally negative. There was a significant increase in the levels of TGF-beta in primary prostatic tumors compared to the normal prostate. Bcl-2 expression was detected among certain populations of tumor epithelial cells in a mutually exclusive topological distribution pattern for apoptosis. In marked contrast, neither TGF-beta1 nor bcl-2 immunoreactivity was detected in metastatic prostate tumor cells, despite their high proliferative and apoptotic rates. Balancing the prostatic growth equation for the prostatic tumor epithelial cell populations revealed a substantial net increase in cell number in both primary and metastatic prostate cancers. This loss of apoptotic control in favor of cell proliferation may be responsible for prostate cancer initiation and progression.

Adenocarcinoma↗
PubMed1996

Apoptotic versus proliferative activities in human benign prostatic hyperplasia.

Cell growth in the normal prostate is regulated by a delicate balance between cell death and cell proliferation (ie, apoptotic v proliferative activity). Disruption of the molecular mechanisms that regulate these two processes may underline the abnormal growth of the gland leading to benign prostatic hyperplasia (BPH). In this study, the incidence of programmed cell death (apoptosis) and cell proliferation was comparatively analyzed among the various cell subpopulations in the normal and benign hyperplastic human prostate. The authors also examined the relative expression of two proteins involved in the regulation of prostate apoptosis: (1) transforming growth factor (TGF)-beta1, a negative growth factor able to induce prostate apoptosis under physiological conditions; and (2) bcl-2, a potent apoptosis suppressor. Analysis of the incidence of "spontaneous" apoptosis in situ, using the end-labeling terminal transferase staining technique for the detection of nucleosomal DNA fragmentation, revealed infrequent apoptotic staining in isolated basal and secretory prostate epithelial cells. The basal level of cell proliferation was determined on the basis of the Ki-67 nuclear antigen staining, a nuclear protein that appears primarily during the proliferative phases of the cell cycle. The Ki-67-positive nuclei were equally distributed among the basal and secretory epithelial cells of the hyperplastic prostatic acini. The apoptotic index of the secretory and basal cells of the prostate epithelium was higher in the normal prostate compared with BPH tissue, whereas there was a significant increase in the proliferative index of the respective cell populations in the hyperplastic prostate. Balancing the apoptotic versus the proliferative activities revealed a substantial net decrease (fourfold) in the total number of cells dying via apoptosis in both the glandular and basal epithelial cell compartments of the hypertrophic prostate (BPH) when compared with the normal gland. TGF-beta staining was exclusively identified in the secretory epithelial cells, lining the prostatic lumen with minimal involvement of the basal cells and total lack of immunoreactivity among the stroma elements. Statistical analysis revealed a significant elevation in TGF-beta expression in the epithelial cells of BPH tissue compared with the normal prostate (P < .001). Expression of bcl-2 was topologically restricted to the glandular epithelium of the prostate. In the normal prostate, bcl-2 immunoreactivity was predominantly identified in the basal cell layer. An increase in both the intensity of immunoreactivity for bcl-2 and the number of positive epithelial cells (basal and secretory) was detected in BPH specimens relative to the normal prostate (P < .02). These results suggest a potential involvement of enhanced expression of this antiapoptosis protein in deregulation of the normal apoptotic cell death mechanisms in the human prostate, thus resulting in a growth imbalance in favor of cell proliferation that might ultimately promote prostatic hyperplasia.

Adult↗
PubMed1996

Incidence of apoptosis, cell proliferation and bcl-2 expression in transitional cell carcinoma of the bladder: association with tumor progression.

PURPOSE: Apoptosis is the distinctive form of programmed cell death that complements cell proliferation in maintaining normal tissue homeostasis. The significance of constitutive apoptosis in the development and progression of transitional cell carcinoma of the bladder has yet to be investigated. In the present study, the incidence of baseline apoptosis and the expression of 2 genes regulating this molecular process, bcl-2 and TGF-beta 1, as well as the level of cell proliferation, were examined by an intensive immunohistochemical analysis in normal bladder and bladder cancer specimens. MATERIALS AND METHODS: Apoptosis was detected by in situ end-labeling of fragmented DNA using the terminal deoxynucleotidyl transferase reaction in 45 paraffin-embedded primary transitional cell carcinoma specimens, 9 metastatic lymph nodes and 5 normal bladder specimens. The proliferation status of the tumor cells among the same bladder cancer specimens was evaluated by using a monoclonal antibody that recognizes the proliferation-associated nuclear antigen, Ki-67. RESULTS: The apoptotic index of normal transitional epithelium (0.06%) was significantly lower than that of all grades of transitional bladder carcinoma (p = 0.006). Although the apoptotic index of transitional carcinomas increased with increasing grade, this difference failed to achieve statistical significance, ranging from 0.54 +/- .23% in grade I to 1.24 +/- .77% in grade III. The proliferative index, as determined by Ki-67 positivity, also increased with increasing grades of tumor (12.8 +/- 8.4% in grade I to 22.6 +/- 15.2% in grade III) and was significantly greater than in normal urothelium (0.64 +/- 0.52%, p = 0.003). Bcl-2 expression was significantly lower in the normal transitional epithelium and in the well and moderately differentiated tumors (grades I-II) when compared with poorly differentiated (grade III) tumors (p = .004). The incidence of bcl-2 expression in all bladder specimens analyzed was uniformly low (< 5.3%). Transforming growth factor-beta 1 expression was not detected in any of the normal bladder specimens, primary tumors, or metastatic lymph nodes analyzed. CONCLUSIONS: The present findings revealed that no statistically significant correlation exists between the frequency of apoptosis and the pathological stage of bladder tumors, while they clearly demonstrate a strong direct correlation between an increased rate of cell proliferation and bladder cancer progression.

Apoptosis↗
PubMed1995

Enhanced expression of ICAM-1 in a murine fibrosarcoma reduces tumor growth rate.

Intercellular adhesion molecule-1 (ICAM-1) plays an essential role in lymphocyte adhesion to endothelium and migration across endothelial cell barriers. We undertook this study to determine the growth of a murine fibrosarcoma transfected with the ICAM-1 gene. MCA-105 tumor cells were cotransfected with ICAM-1 and the plasmid for neomycin resistance (NeoR). Selected G418-resistant clones were expanded and cell surface ICAM-1 expression was verified using a fluorescence-activated cell sorter. Integration of the ICAM-1 gene and ICAM-1 mRNA expression were verified by Southern and Northern blot hybridization analysis, respectively. C57BL/6 mice were divided into five groups (six animals/group): Control, NeoR only, ICAM-1 (low expressing, Clone 25), ICAM-1 (high expressing, Clone 81), and a 1:1 mixture of NeoR:Clone 81; animals received 1 x 10(6) cells on Day 0 and tumor measurements began on Day 7 and were measured in mm2. At 19 days, tumors from cell lines expressing ICAM-1 were significantly (P < .05) smaller than both the parental cell line and tumor-containing NeoR only (364 mm2 vs 466 mm2 and 527 mm2, respectively). This decrease in tumor growth may be a result of increased lymphocyte migration or increased anti-tumor cytotoxicity by infiltrating lymphocytes. The results from the mixed tumor experiment suggest a possible paracrine effect by cells expressing ICAM-1. Studies are currently under way to investigate the effect of immunotherapy on tumors derived from ICAM-1-cloned transfectants.

Animals↗
PubMed1995

Transfection of a murine fibrosarcoma with intercellular adhesion molecule-1 enhances the response to adoptive immunotherapy.

BACKGROUND: Increasing the ability of antitumor effector cells to leave the vasculature and gain access to tumor cells may improve therapeutic efficacy. We undertook this study to determine whether increased expression of intercellular adhesion of molecular-1 (ICAM-1) by gene transfection would result in an improved response to adoptive immunotherapy in vivo. METHODS: C57BL/6 mice received 1 x 10(6) tumor cells on day 0. Tumor cells examined were MCA-105 (parental), NeoR (MCA-105 transfected with the neomycin resistance gene), or Clones 81 or 149 (MCA-105 cotransfected with NeoR and the gene for ICAM-1 and highly express ICAM-1). Animals were treated by use of no treatment, interleukin-2 alone (days 10 through 14), hyperthermia alone (days 10 and 13), or interleukin-2 + hyperthermia, and tumor growth was reported as a ratio to size on day 10. In vitro cytotoxicity was assayed by using murine lymphokine-activated killer cells. RESULTS: Tumors transfected with ICAM-1 and treated with hyperthermia + immunotherapy grew significantly (p < 0.05) slower (mean, 0.78 +/- 0.16 on day 19) than parental tumor (size, 1.35 +/- 0.22) or tumor cells transfected with NeoR alone (1.21 +/- 0.19). Tumors containing both MCA-105 and Clone 81 treated with hyperthermia + immunotherapy grew significantly slower (1.58 +/- 0.49 on day 19, p < 0.05) than untreated Clone 81 (2.38 +/- 0.46) or treated MCA-105 (2.49 +- 0.29) but more rapidly than treated Clone 81 (1.18 +/- 0.08), suggesting a paracrine efect for ICAM-1. CONCLUSIONS: These findings show that increased expression of ICAM-1 by tumor cells results in a significant increase in antitumor efficacy of combined interleukin-2 and hyperthermia in a murine model. Although the mechanism has yet to be elucidated, modulation of cellular adhesion may play a role in the therapeutic efficacy of cellular immunotherapy.

Animals↗
PubMed1995

Reversion of human prostate tumorigenic growth by azatyrosine.

OBJECTIVES: Azatyrosine, an antibiotic isolated from a Streptomyces species, has been previously shown to have antitumor activity against ras- and neu-transformed fibroblasts and human epithelial cells. In this study, we investigated the effect of azatyrosine on human prostate cancer cell growth and the reversion potential of this antibiotic on prostate tumorigenic cell lines. METHODS: Three androgen-independent human prostate cancer cell lines (TSU-Prl, DU-145, and PC-3) were cultured in the presence of azatyrosine and their growth rates were determined over a 7-day period. Following exhaustive treatment with azatyrosine for 5 weeks, three azatyrosine-resistant colonies were cloned from the PC-3 cell line and were subsequently established as stable cell lines. The growth characteristics of these azatyrosine-resistant clones were examined both in vitro and in vivo to establish their "potentially revertant" profiles. RESULTS: Incubation with azatyrosine (for 7 days) resulted in greater than 95% in vitro growth inhibition of the three parental prostate cancer cell lines. Analysis of the biologic properties of these azatyrosine-resistant cell lines revealed: (1) a significant reduction in in vitro growth rates; (2) a decreased rate of DNA synthesis as measured by thymidine uptake; and (3) a decreased ability for colony formation in soft agar. Moreover all three azatyrosine-resistant clones exhibited suppressed tumorigenicity in severe combined immunodeficient (SCID) mice when compared with the parental cell line. An important observation was that one revertant clone demonstrated complete loss of tumorigenicity. On the basis of this biologic behavior, these cell lines were characterized as revertants. Cytogenetic analysis revealed gross chromosomal differences between the revertant clones and the parental cell line. Northern hybridization analysis demonstrated elevated expression of the K-rev-1 and bcl-2 but not the rrg mRNA transcripts in the revertant cell lines. CONCLUSIONS: These results suggest that azatyrosine inhibits prostate tumorigenic growth; it has a high reversion efficiency on human prostate cancer cells; and the K-rev-1 suppressor gene and the bcl-2 proto-oncogene could be potentially involved in the reversion mechanism mediated by azatyrosine. This reversion of prostate cancer cells to an apparently nontumorigenic phenotype points to a potentially significant therapeutic role for azatyrosine in the treatment of advanced prostate cancer.

Adenocarcinoma↗