Generation of angiostatin-like fragments from plasminogen by prostate-specific antigen

Angiostatin, a potent inhibitor of angiogenesis, tumour growth and metastasis, is a biologically active fragment of plasminogen, containing the kringle domains 1-4. It is generated from plasminogen by limited proteolysis. We show that prostate-specific antigen (PSA), a serine proteinase secreted by human prostate and human prostate cancer cells, is able to convert Lys-plasminogen to biologically active angiostatin-like fragments, containing kringles 1-4, by limited proteolysis of peptide bond Glu439-Ala440 in vitro. In an in vitro morphogenesis assay, the purified angiostatin-like fragments inhibited proliferation and tubular formation of human umbilical vein endothelial cells with the same efficacy as angiostatin. This finding might help to understand growth characteristics of prostate cancer, which usually has low microvessel density and slow proliferation.     

Immunohistochemical analysis of midkine expression in human prostate carcinoma.

Midkine (MK) is a growth/differentiation factor frequently expressed at high levels in some types of human malignancies. To investigate whether MK is a useful marker in prostate carcinogenesis, immunohistochemical analysis was performed on samples of both latent and clinical prostate cancers of various stages, as well as on specimens of normal gland and prostatic intraepithelial neoplasia (PIN). Of the 80 clinical cancers examined, 69 specimens (86.3%) were immunoreactive for MK, with metastatic lesions generally showing higher expression than the corresponding primaries; normal prostate tissues were negative or showed only weak staining. Midkine was also detected in 12 of 15 latent cancers (80%) and in 12 of 16 cases of PIN (75%). In sections of whole prostate, MK showed variable expression through tumorous sections, probably in reflection of heterogeneous cell populations. The results demonstrate the possible value of MK as a marker for early and latent disease, as well as for more advanced clinical stages of prostate cancer.     

Matrilysin expression in human prostate carcinoma

The metalloproteinases, a multigene family of metal-requiring enzymes, have been suggested to play a role in tumor invasion and metastasis. Previously, we demonstrated that human primary prostate tumors express higher levels of matrilysin and gelatinase A mRNA than normal prostate does. In the study presented here, we used in situ hybridization and immunohistochemical staining of serial sections of paraffin-embedded primary prostate tumors to compare the sites of matrilysin and gelatinase A expression and protein localization. These results confirmed the epithelial nature of matrilysin expression and protein localization. In contrast, gelatinase A mRNA was localized to the interstitial stroma, whereas the protein was associated with the epithelial tumor cells. In situ hybridization was also used to demonstrate that gelatinase B expression was restricted to macrophages infiltrating the tumors. Proteins isolated from an additional set of frozen tumor specimens were analyzed by western blotting to determine the relative amounts of matrilysin in the active and proenzyme forms. The western analyses demonstrated that in all cases in which matrilysin was detected, at least some of the enzyme was in the active form. These results are discussed with respect to the possible role these enzymes may play in prostate tumor progression. © 1996 Wiley-Liss, Inc.     

Combined c-Myc and caveolin-1 expression in human prostate carcinoma predicts prostate carcinoma progression

BACKGROUND: Over-expression of the oncogene c-Myc has been implicated in the development and progression of human prostate carcinoma. However, previous assessments of c-Myc expression have not revealed its potential for predicting prostate carcinoma progression. Caveolin-1 is associated with prostate carcinoma progression and is a downstream target gene of c-Myc. The observation that caveolin-1 can suppress c-Myc-induced apoptosis suggested the potential for cooperation between c-Myc and caveolin-1 in malignant progression. In this study, the authors evaluated the prognostic potential of combined c-Myc and caveolin-1 expression in human prostate carcinoma progression. METHODS: Immunostaining with c-Myc and caveolin-1-specific antibodies was performed on paraffin sections from 104 radical prostatectomy specimens from men with lymph node negative prostate carcinoma. Combined c-Myc and caveolin-1 immunostaining scores were related with the clinical and pathologic features and the probability of prostate-specific antigen recurrence after surgery. RESULTS: The combination of c-Myc and caveolin-1 immunopositivity correlated positively with Gleason score (rho = 0.219; P = 0.0253) and positive surgical margin (rho = 0.333; P = 0.0006). The combination of positive c-Myc and caveolin-1 in patients with clinically confined prostate carcinoma was a significant prognostic marker for the time to disease progression after surgery in both univariate analysis (P = 0.0039; hazard ratio, 3.035) and multivariate analysis (P = 0.0114; hazard ratio, 2.916). CONCLUSIONS: The coexpression of c-Myc and caveolin-1 showed potential as a useful prognostic marker for human prostate carcinoma. The current results suggest interactions between c-Myc and caveolin-1 in the progression of human prostate carcinoma.     

The dedifferentiation of prostate carcinoma

Fifty-four patients with prostate carcinoma, each having 2 TURP (transurethral resection of the prostate) procedures separated by 3 to 11 years, were studied to determine whether the histologic appearance of prostate carcinoma remains the same for the life of the host or whether the histological appearance changes with time. Using the M. D. Anderson (MDAH) method of grading prostate carcinoma, 19 of 26 (73%) Grade 1 lesions, 9 of 12 (75%) Grade 2 lesions, and 7 of 8 (88%) Grade 3 lesions dedifferentiated into another grade at the time of the 2nd TURP. Eight cases that were Grade 4 at the time of the 1st TURP, remained Grade 4 lesions at the time of the 2nd TURP. Although 10 Grade 1, Grade 2, and Grade 3 lesions did not change grades, 8 of these 10 cases were less differentiated at the time of the second TURP than they were at the time of the first TURP. Furthermore, no Grade 1 lesions demonstrated evidence of metastases, but 19% of Grade 2 lesions, 55% of Grade 3 lesions, and 80% of Grade 4 lesions demonstrated evidence of metastases. This study suggests that the usual course of prostate carcinoma is dedifferentiation and that with dedifferentiation, the likelihood of metastases increases.     

人喉癌新的相关基因cDNA片段的克隆与鉴定

目的:寻找喉癌新的相关基因,阐明喉癌发生的分子机制,为喉癌的基因诊治提供研究的靶基因。方法:应用mRNA差异显示方法对喉癌患者的癌旁、癌及颈部转移淋巴结组织差异cDNA进行了筛选、克隆和测序,并对其进行了鉴定和同源性分析。结果:筛选出7个差异cDNA片段,克隆并鉴定了其中两个与喉癌发生相关的cDNA片段,它们与多种肿瘤相关基因的cDNA序列同源。结论:克隆的两个cDNA片段可能是喉癌新的相关基因cDNA片段,全长cDNA克隆及深入研究其功能有助于发现喉癌新的相关基因。     

The dedifferentiation of metastatic prostate carcinoma

Two hundred consecutive staging lymphadenectomies with metastatic prostate adenocarcinoma and 100 consecutive autopsies with widely disseminated metastatic prostate adenocarcinoma were identified. The metastases from 41% of the staging lymphadenectomies were entirely differentiated (gland forming) and an additional 43% were predominantly (50% or more) differentiated. In contrast, the metastases from 70% of the autopsies were entirely undifferentiated (non-gland forming) and an additional 18% were predominantly undifferentiated. Further, five patients with completely or predominantly differentiated metastases in staging lymphadenectomies were found to have widespread completely or predominantly undifferentiated metastases at autopsy 4-7 years later. These findings suggest that dedifferentiation occurs within metastases and that dedifferentiation within metastases may be important in understanding the widespread dissemination of metastatic prostate carcinoma.     

Androgen receptor gene expression in human prostate carcinoma cell lines

Responses to androgen vary widely among prostate cancers and prostatic carcinoma cell lines. We have explored the basis for this heterogeneity by examining the levels of androgen receptor expression in a prostate carcinoma cell line (LNCaP) that expresses the androgen receptor and two prostate carcinoma cell lines that do not contain detectable androgen receptor. We find that while the LNCaP cell line contains high levels of both the androgen receptor protein and mRNA, the receptor-negative cell lines DU-145 and PC-3 do not express androgen receptor protein as detected by immunoblotting or mRNA as detected by Northern analysis or S1 nuclease protection. These results indicate that the absence of androgen receptor expression in the androgen receptor-negative cell lines is caused by diminished androgen receptor mRNA levels. Genomic Southern analysis indicates that the differences in androgen receptor expression in each of these cell lines is not associated with detectable alterations in the structure of the androgen receptor gene.     

Laminin-5-mediated gene expression in human prostate carcinoma cells

Interactions between extracellular matrix (ECM) proteins and prostate carcinoma cells provide a dynamic model of prostate tumor progression. Previous work in our laboratory showed that laminin-5, an important member of a family of ECM glycoproteins expressed in the basal lamina, is lost in prostate carcinoma. Moreover, we showed that the receptor for laminin-5, the alpha6beta4 integrin, is altered in prostate tumors. However, the genes that laminin-5 potentially regulates and the significance of its loss of expression in prostate cancer are not known. We selected cDNA microarray as a comprehensive and systematic method for surveying and examining gene expression induced by laminin-5. To establish a definitive role for laminin-5 in prostate tumor progression and understand the significance of its loss of expression, we used a cDNA microarray containing 5289 human genes to detect perturbations of gene expression when DU145 prostate carcinoma cells interacted with purified laminin-5 after 0.5, 6, and 24 h. Triplicate experiments showed modulations of four, 61, and 14 genes at 0.5, 6, and 24 h, respectively. Genes associated with signal transduction, cell adhesion, the cell cycle, and cell structure were identified and validated by northern blot analysis. Protein expression was further assessed by immunohistochemistry. Mol. Carcinog. 30:119-129, 2001.     

The radial distance of extraprostatic extension of prostate carcinoma: implications for prostate brachytherapy.

BACKGROUND: Extraprostatic extension (EPE) is an unfavorable prognostic factor in patients with prostate carcinoma. Prior studies have reported the linear extent of EPE measured circumferentially along the edge of the prostate. In this study, the authors defined and evaluated a novel measure of EPE in a large series of radical prostatectomy specimens. These results have important clinical implications in the management of localized prostate carcinoma by brachytherapy and other modalities. METHODS: The authors reviewed the preoperative records and biopsy findings from 376 patients who underwent radical retropubic prostatectomy between September 1991 and June 1993. Whole mount radical prostatectomy specimens were examined, and the location of EPE for each specimen was recorded. The radial EPE distance was measured perpendicular to the edge of the prostate. For specimens with multiple EPE sites, the maximum radial EPE distance was recorded. Established eligibility criteria for prostate brachytherapy were evaluated using these results, with emphasis placed on achieving adequate radiation dose coverage 3-5 mm beyond the capsule or the edge of the prostate. RESULTS: EPE was identified in 105 of 376 specimens (28%) at 248 sites. The radial EPE distance in these specimens had a mean of 0.8 mm (range, 0.04-4.4 mm) and a median of 0.5 mm. Of these 105 patients, the median and mean preoperative prostate specific antigen (PSA) concentrations were 11.8 ng/mL and 17.9 ng/mL, respectively. The mean and range of the Gleason score and prostate volume for all specimens were 6.3 (range, 3-9) and 39 cc (range, 8-294 cc), respectively. In 107 patients who met the selection criteria for prostate brachytherapy eligibility of a PSA level < 10 ng/mL, Gleason score < 7, and gland volume < 60 cc, the maximum and mean radial EPE distances were 0.6 mm and 0.03 mm, respectively. CONCLUSIONS: The radial distance of EPE is an important measure that influences treatment strategies for patients with localized prostate carcinoma. Currently described criteria for the treatment of early stage prostate carcinoma by brachytherapy alone appear satisfactory to ensure effective radiation dose coverage of EPE of prostate tumors. Treating the prostate with a 3-5 mm margin by brachytherapy would encompass all known tumor in approximately 99% of the specimens examined in this study.     

Activated ras alleles in human carcinoma of the prostate are rare

Although oncogenically activated ras alleles are common in some types of human cancers, the frequency in human carcinoma of the prostate (CaP) has not previously been addressed. In this paper, we report a comprehensive screening of 19 CaPs and 3 CaP cell lines for activating point mutations in the sequences of the 12th and 61st codons of c-Ha-ras-1 and the c-Ki-ras-2 genes, as well as the 12th, 13th, and 61st codons of the c-N-ras gene. The 19 CaPs were chosen to represent a wide range of stages (B through D), Gleason scores (3 through 10), and DNA ploidy (diploid with low proliferation to nontetraploid-aneuploid). Fifteen of the tumors had been untreated prior to resection and 4 were obtained postradiation therapy. The polymerase chain reaction was used to amplify genomic DNA sequences and transcribed mRNA sequences of the ras genes prior to allele-specific oligonucleotide probe hybridization. We have detected a mutant allele with a point mutation in the second nucleotide of the 61st codon of the c-Ha-ras-1 gene in one specimen. Thus, we conclude that the activation of ras alleles is not significant in the development or progression of most CaPs.     

前列腺癌患者治疗前后PSA水平

目的　探讨PSA在前列腺癌临床诊疗中的应用价值。方法　采用ELISA法测定 3 3例健康者、3 6例良性前列腺增生及 3 3例前列腺癌治疗前后T PSA和F PSA并动态观察 2 0例放疗后T PSA水平变化。结果　前列腺癌T PSA和F PSA、F/T均显著高于良性前列腺增生 (P <0 .0 1) ;前列腺癌治疗后T PSA和F PSA显著下降 (P <0 .0 1) ,而F/T无明显改变 (P >0 .0 5 ) ;前列腺癌放疗后病情稳定T PSA下降 ,病情进展则T PSA升高。结论　动态测定前列腺癌患者治疗前后PSA水平变化 ,可判断近期疗效 ,监测病情变化。     

Construction and selection of human anti-idiotypic antibody single chain variable fragments or CDR3 fragments of nasopharyngeal carcinoma

Peripheral blood mononuclear cells (PBMCs) of patients with NPC were immunized in vitro by anti-NPC monoclonal antibody FC2 and transformed by Epstein-Barr virus (EBV). Detection showed that of 10 NPC patients, 8 patients' B cells immunized by FC2 and transformed by EBV produced anti-idiotypic antibodies to NPC. Five types of VH genes and 7 types of VL genes were obtained by RT-PCR amplification and then connected with (Gly4Ser)3 linker to form 14 types of scFv genes. ScFv genes digested with Sfi I were cloned into vector fUSE5 and transformed into E. coli MC 1061. Phage anti-idiotypic antibody library with 1.5 x 10(8) clones was obtained. After four rounds of panning, 270 phage clones were selected randomly and 91 FC2-positive clones were obtained by Sandwich ELISA, the positive ratio was 33.7%. 5 clones (D83, E92, G22, I50, I54), which might display beta type Ab2 scFv, were selected by binding inhibition test. These 5 phage anti-idiotypic antibodies were further analyzed by DNA sequencing. The VDJ regions of G22, I50, I54 belonged to VH4-39-D4-11-JH3-linker-V1-19-JL2, VH4-4-D4-11-JH6 and VH4-31-D4-11-JH6, respectively. E92 had the same VDJ regions with G22; D83 had the same VDJ regions with 150. So, a strategy for preparing and selecting beta type Ab2 scFv or CDR by means of immunization in vitro, EBV transformation and phage display technique is feasible, which paves a way for preparing cancer vaccine using beta type Ab2 scFv.