Biomedical subjects

T L Timme

Publications and source records attributed to T L Timme.

At least 19 recordsLinked to original sources

PubMed2002

Gene therapy for prostate cancer: toxicological profile of four HSV-tk transducing adenoviral vectors regulated by different promoters.

Adenoviral vector delivery of the Herpes simplex virus thymidine kinase (HSV-tk) gene in combination with the prodrug ganciclovir (GCV) has been tested in phase I clinical trials for prostate cancer and found to exhibit a satisfactory toxicity profile. We have developed additional adenoviral vectors with differing promoters to optimize the expression profile and in the present study evaluate the potential systemic toxicity of these vectors. Four recombinant adenoviral vectors that express the HSV-tk gene were generated using three different promoters: CMV (leftward orientation); RSV (both rightward and leftward orientation); and the mouse caveolin-1 (cav-1) promoter (leftward orientation). Efficacy was determined in vitro by cytotoxicity assays in a mouse prostate cancer cell line, RM-9, and in vivo by treating orthotopic tumors. Potential toxicity was evaluated from liver histology and apoptotic cell counts and enzyme levels in the serum following intravenous adenoviral vector injection. Although there were differences in HSV-tk expression at the protein level among the four vectors there were no significant differences in in-vitro cytotoxicity studies with GCV or in vivo in tumor growth suppression of an orthotopic mouse prostate cancer model in GCV treated mice. Intravenous delivery of high doses of all adenoviral vectors lead to abnormalities in liver function as measured by specific serum markers and histological evaluation of liver tissue and increased levels of apoptosis in the liver. These abnormalities were most prevalent with the vector containing the CMV promoter and the rightward oriented RSV promoter. They were least prevalent in the vector regulated by the cav-1 promoter. Upregulation of specific chemokines, MIP-2 and MIP-1beta was correlated with apoptotic counts. Our results demonstrate that comprehensive toxicological analysis of adenoviral vectors provides internally consistent information that can differentiate vectors with comparable efficacy based on toxicity. In these studies vectors with the cav-1 promoter-driven and leftward RSV-driven HSV-tk gene demonstrated minimal toxicities with cytotoxic effectiveness comparable to more toxic vectors. Our studies further suggest that promoter selection can influence the toxic effects of an adenoviral gene therapy vector.

Adenocarcinoma↗

PubMed2001

Prostate-specific antigen response and systemic T cell activation after in situ gene therapy in prostate cancer patients failing radiotherapy.

In an extended phase I/II study we evaluated 36 prostate cancer patients with local recurrence after radiotherapy who received single or repeated cycles of replication-deficient adenoviral vector (ADV)-mediated herpes simplex virus-thymidine kinase (HSV-tk) plus ganciclovir (GCV) in situ gene therapy with respect to serum PSA levels, alterations in immune cells, and numbers of apoptotic cells in needle biopsies. An initial cycle of HSV-tk plus GCV gene therapy caused a significant prolongation of the mean serum PSA-doubling time (PSADT) from 15.9 to 42.5 months (p = 0.0271) and in 28 of the injected patients (77.8%) there was a mean PSA reduction (PSAR) of 28%. It took a mean of 8.5 months for the PSA to return to the initial PSA (TR-PSA) value. A repeated cycle of gene therapy failed to significantly extend PSADT but did result in significant increases in PSAR (29.4%) and TR-PSA (10.5 months). Moderately increased serum adenovirus antibody titers were generally observed 2 weeks after initial vector injection. Also at this time there was a statistically significant increase in the mean percent of CD8(+) T cells positive for the HLA-DR marker of activation in peripheral blood (p = 0.0088). Studies using prostate biopsies obtained at the same time point demonstrated that vector DNA was detectable by PCR in most samples yet all patients remained positive for prostate cancer in at least one biopsy core. Further analysis demonstrated a correlation between the level of CD8(+) cells and the number of apoptotic cells in biopsies containing cancer cells (p = 0.042). We conclude that repeated cycles of in situ HSV-tk plus GCV gene therapy can be administered to prostate cancer patients who failed radiotherapy and have a localized recurrence. Biological responses to this experimental therapy including increases in PSADT, PSAR, and TR-PSA, and activated CD8(+) T cells present in the peripheral blood, were demonstrated. Interestingly, the density of CD8(+) cells in posttreatment biopsies correlated with the number of apoptotic cells.

Adenoviridae↗

PubMed2001

Caveolin-1 mediates testosterone-stimulated survival/clonal growth and promotes metastatic activities in prostate cancer cells.

Previously, we demonstrated that up-regulation of caveolin-1 (cav-1) was associated with prostate cancer metastasis, biochemical recurrence after radical prostatectomy, and androgen insensitivity. The objective of this study was to characterize the regulation of cav-1 by testosterone (T) and to test the effects of cav-1 on prostate cancer cell survival/clonal growth and metastatic activities. Our results demonstrated that T up-regulated cav-1 protein levels in part through transcriptional regulation and significantly enhanced survival of prostate cancer cell lines ABAC3 and LNCaP after serum starvation (>40% and >60% increased viability, respectively) and in an extended clonogenic assay (approximately 4-fold and 6-fold increase in colonies, respectively). Importantly, antisense cav-1 inhibited the survival effects of T in these assay systems. Modest but not high levels of adenoviral vector-mediated cav-1 expression alone also significantly increased viability (>40%) and clonal growth (10-fold increase in colonies) after serum starvation. Analysis of spontaneous metastasis in stably transfected antisense cav-1 mouse prostate cancer cell clones demonstrated reduction of spontaneous lymph node metastasis incidence (13%), spontaneous lymph node metastasis volume (46%), and experimental lung metastasis incidence (40%) compared with vector control cell clones. Surgical castration further reduced spontaneous lymph node metastasis incidence and volume (18% and 28%, respectively) in antisense cancer cell clones, but not in vector control clones. Our studies demonstrate that cav-1 is a downstream effector of T-mediated prostate cancer cell survival/clonal growth and that modest levels of cav-1 can independently promote prostate cancer cell survival/clonal growth and metastatic activities.

Animals↗

PubMed2001

Secreted caveolin-1 stimulates cell survival/clonal growth and contributes to metastasis in androgen-insensitive prostate cancer.

Caveolin-1 is an integral protein of caveolae, known to play important roles in signal transduction and lipid transport. We demonstrate that caveolin-1 expression is significantly increased in primary and metastatic human prostate cancer after androgen ablation therapy. We also show that caveolin-1 is secreted by androgen-insensitive prostate cancer cells, and that this secretion is regulated by steroid hormones. Significantly, caveolin-1 was detected in the MDL(3) fraction of serum specimens from patients with advanced prostate cancer and to a lesser extent in normal subjects. Conditioned media from high passage caveolin-1 secreting, androgen-insensitive, LNCaP cells stimulated increased viability and clonal growth of low passage, caveolin-1-negative, androgen-sensitive, LNCaP cells in vitro, and this effect was blocked by treating the media with caveolin-1 antibody. i.p. injections of caveolin-1 antibody suppressed the orthotopic growth and spontaneous metastasis of highly metastatic, androgen-insensitive caveolin-1-secreting mouse prostate cancer. Overall, our results establish caveolin-1 as an autocrine/paracrine factor that is associated with androgen-insensitive prostate cancer. We demonstrate the potential for caveolin-1 as a therapeutic target for this important malignancy.

Androgen Antagonists↗

PubMed2001

Combination gene therapy with adenoviral vector-mediated HSV-tk+GCV and IL-12 in an orthotopic mouse model for prostate cancer.

We previously demonstrated significant therapeutic activities associated with adenoviral vector-mediated Herpes Simplex Virus/thymidine kinase (AdHSV-tk) with ganciclovir (GCV) in situ gene therapy in the RM-1 orthotopic mouse prostate cancer model and interleukin-12 (AdmIL-12) in situ gene therapy in the RM-9 orthotopic mouse prostate model for prostate cancer. In both protocols, local cytotoxicity and activities against pre-established lung metastases were demonstrated. To test whether combined AdHSV-tk+GCV+IL-12 gene therapy would lead to enhanced therapeutic effects when compared to either treatment alone, we used RM-9 mouse prostate cancer cells in both orthotopic and pre-established lung metastases models of prostate cancer. Combined treatment with a single injection of optimal doses of AdHSV-tk+GCV or AdmIL-12 led to significantly increased suppression of orthotopic tumor growth. IL-12 gene therapy alone was more effective than AdHSV-tk+GCV in suppressing spontaneous lymph node metastases and pre-established lung metastases but combination gene therapy did not result in additional anti-metastatic activities. Combination gene therapy also did not achieve significantly better animal survival compared to AdHSV-tk+GCV or AdmIL-12 alone. Analysis of localized antitumor activities demonstrated that AdHSV-tk+GCV therapy induced higher levels of necrosis compared to AdmIL-12 or combination therapy. However, both treatments alone and combination therapy produced similar increases in apoptotic index. To address the possible mechanisms of locally co-operative cytotoxic activities, we analyzed the systemic natural killer (NK) response and the numbers of tumor-infiltrating immune cells using quantitative immunohistochemical analysis. AdHSV-tk+GCV therapy alone led to detectable increases in iNOS-positive cells, CD4+and CD8+T-cells and moderately increased numbers of F4/80 (macrophage selective)-positive cells within treated tumors. In contrast, AdmIL-12 elicited a highly robust pattern of tumor infiltration for all four of these immune cells that was in general mimicked by combination therapy. Further analysis of the accumulation of transforming growth factor-beta1 (TGF-beta1) immunohistochemical staining demonstrated that AdHSV-tk+GCV treatment, but not AdmIL-12 treatment, produced cancer cell-associated increases in this cytokine relative to control Ad-beta-gal injections. Interestingly, local injection with AdHSV-tk+GCV induced significant splenocyte-derived NK cell cytolytic activities with maximal response 7 days following treatment, whereas AdmIL-12 injection produced significantly higher NK activity with maximal response 2 days following injection. The combined treatment produced a higher systemic NK response over the 14-day treatment period. Depletion of NK cells in vivo demonstrated that this immunocyte subpopulation was responsible for early locally cytotoxic activities induced by AdHSV-tk+GCV but not AdmIL-12 and that NK activities were largely responsible for activities against pre-established metastases generated by both gene therapy protocols. Prostate Cancer and Prostatic Diseases (2001) 4, 44-55

Journal Article↗

PubMed2001

In situ gene therapy for prostate cancer: immunomodulatory approaches.

The development of effective treatments for prostate cancer is thwarted by the natural history of the disease. The biological and clinical potential of most individual cancers is uncertain. In many cases the disease will not progress to clinical significance but experimental and clinical studies indicate that prostate cancer can and may metastasis early in the course of the disease from relatively small foci (i.e., not necessarily the largest or index cancer). Localised prostate cancer is potentially curable with localised therapies (radical prostatectomy or irradiation therapy). However, there are no curative therapies for metastatic prostate cancer. Gene therapy, especially those approaches with an immunomodulatory component, may provide additional therapeutic options with the potential to affect both localised and systemic disease. We have pioneered the development and application of in situ gene therapy protocols using adenoviral vectors to transduce specific genes that generate cytotoxic activity and/or a systemic antitumour immune response. In addition we have completed initial studies that demonstrate the therapeutic potential of adenoviral vector-mediated gene modified cell-based vaccines. Our review discusses preclinical studies focused on the development of immunostimulatory in situ gene therapy approaches that hopefully will provide novel and effective treatments for localised and metastatic prostate cancer.

Animals↗

PubMed2001

In situ prostate cancer gene therapy using a novel adenoviral vector regulated by the caveolin-1 promoter.

Caveolin-1, a structural component of caveolae, is overexpressed in metastatic and androgen-resistant prostate cancer and highly expressed in tumor-associated endothelial cells. The mouse cav-1 promoter was cloned and placed upstream of the HSV-tk gene in an adenoviral vector (Adcav-1tk) and compared with a cytomegalovirus (CMV) or Rous sarcoma virus (RSV) promoter-driven HSV-tk, AdCMVtk and AdRSVtk vectors, respectively. Mouse and human prostate cancer cells and mouse endothelial cells were infected with Adcav-1tk, AdCMVtk or control vectors without the HSV-tk gene (Adcav-1 and AdCMV) and subsequently treated with ganciclovir (GCV). GCV-mediated in vitro cytotoxicity induced by the Adcav-1tk vector was comparable to that for AdCMVtk in multiple mouse and human prostate cancer cell lines. To evaluate the activity of Adcav-1tk in vivo, orthotopic mouse prostate cancer tumors were generated with RM-9 cells and injected in situ with Adcav-1tk, AdCMVtk, AdRSVtk, or AdCMVbetagal (control) and treated with GCV. All three HSV-tk transducing vectors produced statistically significant reductions in wet weight and increased apoptotic indices compared with the control vector. However, only Adcav-1tk produced significant necrosis, and only Adcav-1tk and AdRSVtk caused significant decreases in microvessel density. In conclusion, Adcav-1tk demonstrated efficacy in vitro and in vivo in preclinical models of prostate cancer. Our results suggest that the cav-1 promoter may have unique benefits in targeting gene therapy to prostate cancer and its associated vasculature.

Adenoviridae↗

PubMed2000

Caveolin-1 is regulated by c-myc and suppresses c-myc-induced apoptosis.

Recent data indicating that overexpression of caveolin-1 as well as c-myc are relatively common features of advanced prostate cancer prompted us to test for potential cooperative interactions between caveolin-1 and c-myc that would be consistent with malignant progression. We used the well-characterized Rat1AmycERT cells to show that the caveolin-1 gene is down-regulated at the level of transcription by c-myc. By maintaining relatively high levels of caveolin-1 with an adenoviral vector or in stably transfected clones we show that caveolin-1 can suppress c-myc-induced apoptosis. Further we established human prostate cancer cell lines with the mycER construct and show that clones with increased caveolin-1 are more resistant to myc-induced apoptosis and have increased capacity for growth in soft agar when c-myc is activated.

Animals↗

PubMed2000

Mouse prostate reconstitution model system: A series of in vivo and in vitro models for benign and malignant prostatic disease.

BACKGROUND: An elucidation of the complex, morphological and molecular changes that underlie benign and malignant prostatic disease will likely lead to improved methods of diagnosis and therapy for those disorders. To identify and understand the interrelation of the phenotypic and genetic changes inherent in these important diseases requires the development and use of in vivo and in vitro models that closely mimic specific aspects of the disease process. Once the suspected molecular underpinnings of prostatic disease are uncovered, in vivo and in vitro models will be required for further testing of the functional significance of specific genetic alterations as they are identified. In addition models of prostatic disease are necessary to evaluate novel therapeutic approaches. METHODS: The mouse prostate reconstitution (MPR) model system was developed more than a decade ago with these specific needs in mind. Over the years, specific modifications of the MPR model have demonstrated its versatility and applicability for the study of benign and malignant prostatic disease, including metastatic progression. RESULTS: We discuss various modifications of the MPR model system made for its application to specific aspects of prostatic disease; the clinically relevant information that has been gleaned thus far from the use of this model system; and advances on the horizon for the expansion of its role in prostate research. CONCLUSIONS: The MPR model system has contributed substantially to the understanding and treatment of benign and malignant prostatic diseases. Additional modifications in this series of in vivo and in vitro models will likely lead to further advances.

Animals↗

PubMed2000

Resistance to lysis by cytotoxic T cells: a dominant effect in metastatic mouse prostate cancer cells.

Better understanding of the immunology of prostate cancer is needed for the development of new therapeutic approaches that can be used in conjunction with current treatment methods. The present study was designed to compare the immunological properties of a genetically matched pair of primary tumor- and metastasis-derived prostate cancer cell lines generated from the mouse prostate reconstitution (MPR) model. Only the primary prostate cancer cells were immunogenic in that prior immunization with irradiated primary but not the metastatic prostate cancer cells delayed the growth of subsequently injected live cancer cells. The lack of immunogenicity of the metastatic cells was not attributable to their inability to induce antitumor cytotoxic T cells. Both primary and metastatic cells induced antitumor CTLs in syngeneic hosts, but unlike the primary cells, the metastatic cells were resistant to CTL lysis. Differential resistance to cytolysis in metastatic versus primary prostate cancer cells was not attributable to the differential expression of molecules such as transporter associated with antigen processing (TAP)-1, TAP-2, low molecular weight protein of the proteasome complex (LMP)-2, and LMP-7 that contribute to antigen presentation by class I MHC. IFN-gamma induced surface class I MHC expression, as well as gene expression of TAP-1, TAP-2, LMP-2, and LMP-7 in the metastatic cells, yet the cells remained resistant to cell lysis induced by CTLs. Interestingly, although in comparison to the primary cells the metastatic cells were resistant to cytolysis, both cell types were susceptible to DNA fragmentation induced by CTLs. Cell fusion between primary and metastatic cancer cells resulted in hybrids that also resisted the cytolytic activity of CTLs. Therefore, there is a dominant factor(s) in the metastatic prostate cancer cells that confers specific protection against CTL cytolysis in this model system.

Animals↗

PubMed2000

Dietary 4-HPR suppresses the development of bone metastasis in vivo in a mouse model of prostate cancer progression.

The effects of the synthetic retinoid N-(4-hydroxyphenyl) retinamide (4-HPR) on prostate cancer metastasis in vivo were evaluated in the mouse prostate reconstitution (MPR) model. MPRs were produced by infection of either heterozygous (+/-) or nullizygous (-/-) p53-mutant fetal prostatic epithelial cells with the recombinant retrovirus Zipras/myc 9. Previous studies have documented that loss of p53 function potentiates metastasis in this model system. MPRs were grafted into homozygous (+/+) p53 male mice, fed a 4-HPR containing diet or a control diet and maintained until the status of tumor progression dictated sacrifice. Under these experimental conditions, treatment with 4-HPR did not have a significant effect on primary tumor wet weight for either p53 +/- or p53 -/- MPRs. For, p53 +/- MPRs the animals fed the 4-HPR diet had a slight improvement in survival and a significant reduction in the number of mesenteric metastases (P = 0.0477, t-test). Notably, in p53 +/- MPRs the incidence of metastasis to lumbar spine and sternum was 92% in the control animals compared to 54% in the 4-HPR treated animals (P = 0.035, chi2-test). In p53 -/- MPRs there was a trend toward a reduction in the number of soft tissue metastases to lung and liver in the 4-HPR group relative to the control diet group and a statistically significant reduction in the incidence of metastasis to bone was demonstrated in that 50% of control animals versus 30% of 4-HPR treated p53 -/- animals harbored bone metastases (P = 0 < 0.05, chi2-test). Cell lines were established from portions of the primary tumor and from selected metastatic deposits in each experimental group. Clonal analysis, by retroviral integration pattern, indicated increased clonal diversity in both the primary tumors and metastasis-derived cell lines from 4-HPR treated animals relative to the control animals. In vitro treatment with 4-HPR did not reveal discriminating differences between cell lines derived from primary tumors and bone metastases or control and treatment groups in regard to growth arrest or apoptotic responses. Overall these studies indicate limited anti-tumor and anti-metastatic activity in this highly aggressive in vivo mouse model of prostate cancer, yet 4-HPR treatment significantly suppressed the development of bone metastases in p53 +/- and p53 -/- MPRs revealing a novel and potentially clinically useful activity of this retinoid.

Animals↗

PubMed2000

Prostate cancer gene therapy: comparison of adenovirus-mediated expression of interleukin 12 with interleukin 12 plus B7-1 for in situ gene therapy and gene-modified, cell-based vaccines.

We have documented previously that adenovirus-mediated interleukin 12 (IL-12) gene therapy is effective for orthotopic tumor control and suppression of pre-established metastases in a preclinical prostate cancer model (Y. Nasu et al., Gene Ther., 6: 338-349, 1999). In this report, we directly compare the effectiveness of an adenovirus that expresses both IL-12 and the costimulatory molecule B7-1 (AdmIL12/B7) with one that expresses IL-12 alone (AdmIL-12) using the poorly immunogenic RM-9 orthotopic murine model of prostate cancer. We document AdmIL-12/B7-mediated secretion of IL-12 and increased surface expression of B7-1 in infected RM-9 tumor cells. A significant reduction in orthotopic tumor size and increased survival was demonstrated in mice treated with a single orthotopic injection of AdmIL-12/B7 compared with AdmIL-12 or controls. Six of 19 animals treated with AdmIL-12/B7 survived long term with apparent eradication of the primary tumor in contrast to one of 38 animals in the AdmIL-12-treated group. Orthotopic treatment of tumors with both vectors led to an infiltration of both CD4+ and CD8+ immunoreactive cells, with AdmIL-12/B7 treatment having a more prolonged infiltration of CD8+ cells. AdmIL-12/B7 was also more effective than AdmIL-12 or controls at suppression of pre-established metastases. We further developed a vaccine model based on s.c. injection of infected, irradiated RM-9 cells and found that both AdmIL-12 and AdmIL-12/B7 are effective at suppressing the development and growth of challenge orthotopic tumors using this protocol.

Adenoviridae↗

PubMed1999

In situ gene therapy for adenocarcinoma of the prostate: a phase I clinical trial.

For patients with local recurrence of prostate cancer after definitive irradiation therapy there is no treatment widely considered safe and effective. After extensive preclinical testing of prodrug gene therapy in vitro and in vivo, we conducted a phase I dose escalation clinical trial of intraprostatic injection of a replication-deficient adenovirus (ADV) containing the herpes simplex virus thymidine kinase gene (HSV-tk) injected directly into the prostate, followed by intravenous administration of the prodrug ganciclovir (GCV). Our goal was to determine safe dose levels of the vector for future trials of efficacy. Patients with a rising serum prostate-specific antigen (PSA) level and biopsy confirmation of local recurrence of prostate cancer without evidence of metastases one or more years after definitive irradiation therapy were eligible for the trial. After giving informed consent, patients received injections of increasing concentrations of ADV/HSA-tk in 1 ml into the prostate under ultrasound guidance. Ganciclovir was then given intravenously for 14 days (5 mg/kg every 12 hr). Patients were monitored closely for evidence of toxicity and for response to therapy. Eighteen patients were treated at 4 escalating doses: group 1 (n = 4) received 1 x 10(8) infectious units (IU); group 2 (n = 5) received 1 x 10(9) IU; group 3 (n = 4) received 1 x 10(10) IU; group 4 (n = 5) received 1 x 10(11) IU. Vector was detected by PCR of urine samples after treatment, increasing in frequency and duration (up to 32 days) as the dose increased. All cultures of blood and urine specimens were negative for growth of adenovirus. Minimal toxicity (grade 1-2) was encountered in four patients. One patient at the highest dose level developed spontaneously reversible grade 4 thrombocytopenia and grade 3 hepatotoxicity. Three patients achieved an objective response, one each at the three highest dose levels, documented by a fall in serum PSA levels by 50% or more, sustained for 6 weeks to 1 year. This study is the first to demonstrate the safety of ADV/HSV-tk plus GCV gene therapy in human prostate cancer and the first to demonstrate anticancer activity of gene therapy in patients with prostate cancer. Further trials are underway to identify the optimal distribution of vector within the prostate and to explore the safety of repeat courses of gene therapy.

Adenocarcinoma↗

PubMed1999

Adenovirus-mediated interleukin-12 gene therapy for prostate cancer: suppression of orthotopic tumor growth and pre-established lung metastases in an orthotopic model.

Interleukin-12 (IL-12) can elicit potent antitumoral effects that involve the recruitment of specific immune effector cells. We investigated the efficacy of a single injection of a recombinant adenovirus expressing murine IL-12 (AdmIL-12) directly into orthotopic mouse prostate carcinomas generated from a poorly immunogenic cell line (RM-9) derived from the mouse prostate reconstitution system. Significant growth suppression (> 50% reduction of tumor weight) and increased mean survival time (23.4 to 28.9 days) were observed compared with controls. Suppression of pre-established lung metastases was also observed following the injection of AdmIL-12 into the orthotopic tumor. Cytolytic natural killer cell activity was markedly enhanced 1-2 days after virus injection. Immunohistochemical analysis showed significantly elevated intratumoral infiltration of CD4+ and CD8+ T cells 7 days after virus injection. However, splenocyte-derived cytotoxic T lymphocytes were not detected during the 14 days following treatment. Increased numbers of nitric oxide synthase-positive macrophages were seen in the AdmIL-12 treated group 7 days following injection. Systemic inhibition of natural killer cells with antiasialo-GM1 serum led to increased numbers of lung metastases in AdmIL-12-treated orthotopic tumors but did not affect local tumor growth. In this model system the antitumor effects of a single injection of adenovirus-mediated IL-12 appears to be based to a large extent on the activation of nitric oxide synthase in macrophages and possibly T cell activities, whereas the relatively early cytolytic activity of natural killer cells are largely but not exclusively responsible for the antimetastatic effects.

Adenoviridae↗

PubMed1999

Elevated levels of circulating interleukin-6 and transforming growth factor-beta1 in patients with metastatic prostatic carcinoma.

PURPOSE: Specific cytokines have been found to be secreted by and influence the growth of prostate cancers in cell culture. Interleukin-6 (IL-6), tumor necrosis factor alpha (TNFalpha), granulocyte macrophage-colony stimulating factor (GM-CSF) and transforming growth factor-beta1 (TGF-beta1) have all been closely associated with prostate cancer. We analyzed the levels of these cytokines in the systemic circulation of patients with varying stages of prostate cancer compared to controls. MATERIALS AND METHODS: Serum IL-6, TNFalpha and GM-CSF were measured using commercially available enzyme linked immunosorbent assays in 5 groups of patients, including controls-19 men presenting to prostate cancer screening with normal digital rectal examination and serum prostate specific antigen (PSA) no greater than 2.0 ng./ml., stage pT2-19 with cancer confined to the prostate in the radical prostatectomy specimen, stage pT3-10 with extraprostatic extension and/or seminal vesicle involvement, stage N1-12 with lymph node metastases at pelvic lymph node dissection, and stage M1-9 with bone metastases. Platelet poor plasma TGF-beta1 was measured using a commercially available enzyme linked immunosorbent assay in controls and patients with stage M1 disease only because it was not available for patients with stages pT2, pT3 and N1 disease. No patient had a history of any other malignancy. All blood specimens were collected before surgery and/or androgen ablation. Statistical analysis was done with the Kruskal-Wallis analysis of variance. RESULTS: Serum IL-6 and platelet poor plasma TGF-beta1 were significantly elevated in patients with clinically evident metastases (p = 0.0008 and 0.0412, respectively) while serum GM-CSF and TNFalpha were not. IL-6 and TGF-beta1 correlated with increasing serum PSA (p = 0.0335 and 0.0386, respectively). GM-CSF did not correlate with PSA or age. In multivariate analysis TNFalpha correlated with age but not PSA. CONCLUSIONS: IL-6 and TGF-beta1 correlate with tumor burden as assessed by serum PSA or clinically evident metastases. Further research is needed to determine the response to androgen ablation as well as the source(s) and actions of these cytokines.

Adult↗

PubMed1998

Reduced lysyl oxidase messenger RNA levels in experimental and human prostate cancer.

To identify genes associated with prostate cancer progression, we developed a strategy involving the use of differential display PCR and a panel of genetically matched primary tumor- and metastasis-derived mouse prostate cancer cell lines. We analyzed sequences that were differentially stimulated by transforming growth factor-beta1 in primary tumor-versus metastasis-derived cell lines, based on our previous studies indicating that acquisition of differential responses to this growth factor could result in phenotypic traits that facilitate tumor metastasis from specific cell clones within the primary tumor. Using this system, we isolated and sequenced a cDNA fragment that encoded mouse lysyl oxidase (LO) and was induced by transforming growth factor-beta1 in primary tumor but not in metastasis-derived cells. Northern blotting analysis revealed increased LO expression in a panel of primary tumor cell lines but significantly reduced or nondetectable expression in their matched metastatic counterparts. Further in situ hybridization analysis revealed LO expression in normal mouse prostate epithelium but, in most cases, progressive loss of expression in primary prostate cancer and associated metastatic lesions. Importantly, in situ hybridization studies of normal human prostate and prostate malignancies revealed a similar loss of expression during progression to metastasis. The progressive loss of LO expression during prostate cancer progression provides information that may increase our understanding of the mechanisms that underlie this disease. In addition, LO may provide a useful molecular marker and/or establish a novel therapeutic target for prostate cancer.

Animals↗

PubMed1998

Suppression of caveolin expression induces androgen sensitivity in metastatic androgen-insensitive mouse prostate cancer cells.

Although prostate cancer cells are often initially sensitive to androgen ablation, they eventually lose this response and continue to survive, grow and spread in the absence of androgenic steroids. The mechanism(s) that underlie resistance to androgen ablation therapy remain mostly unknown. We have demonstrated that elevated caveolin protein levels are associated with human prostate cancer progression in pathological specimens. Here we show that suppression of caveolin expression by a stably transfected antisense caveolin-1 cDNA vector converted androgen-insensitive metastatic mouse prostate cancer cells to an androgen-sensitive phenotype. Orthotopically grown tumors and low-density cell cultures derived from antisense caveolin clones had increased apoptosis in the absence of androgenic steroids, whereas similarly grown tumors and cells from vector (control) clones and parental cells were not sensitive to androgens. Studies using a representative antisense caveolin clone showed that selection for androgen resistance in vivo correlated with increased caveolin levels, and that adenovirus-mediated caveolin expression blocked androgen sensitivity. Our results identify a new candidate gene for hormone-resistant prostate cancer in man and indicate that androgen insensitivity can be an inherent property of metastatic prostate cancer.

Androgens↗

PubMed1998

Gene expression of prothrombin in the human kidney and its potential relevance to kidney stone disease.

OBJECTIVE: To determine whether urinary prothrombin fragment 1 (UPTF1), which shows considerable promise as a critical determinant of calcium oxalate (CaOx) stone formation, is manufactured by the human kidney. MATERIALS AND METHODS: Ribonucleic acid was isolated from eight kidneys, two spleens and one liver. Using reverse transcriptase-polymerase chain reaction, mRNA corresponding to the UPTF1 portion of prothrombin was analysed by agarose-gel electrophoresis and Southern blotting. RESULTS: Six kidney specimens showed clear evidence of prothrombin gene expression; expression in the kidney was less than that in the liver. CONCLUSION: This is the first demonstration of prothrombin gene expression within the human kidney, a finding that not only has implications for CaOx stone disease but also potentially for blood coagulation.

Biomarkers↗