Reflex PCA3 messenger ribonucleic acid testing: validation of postbiopsy urine samples and correlation with prostate biopsy findings in ∼2000 patients.

OBJECTIVE: To validate post-transrectal ultrasonography (TRUS) prostate biopsy (bx) urine samples for PCA3 messenger ribonucleic acid testing, including correlation of PCA3 score with concurrent bx findings. METHODS: From July 2008 to July 2010, 2015 patients had urine collected immediately after a TRUS-guided prostate bx. Excluded were men with history of prostate carcinoma (CaP), <6 or ≥24 bx cores, and/or prostate-specific antigen (PSA) level ≥50 ng/mL, resulting in 1909 included men. PCA3 and PSA messenger ribonucleic acid were quantitated using transcription-mediated amplification. A PCA3 score of ≥35 was considered positive. RESULTS: Mean and median ages were 66 years. Mean and median PSA levels were 6.7 and 5.1 ng/mL, respectively. Bxs were benign in 970 (50.8%), CaP in 726 (38%), high-grade prostatic intraepithelial neoplasia (HGPIN) in 124 (6.5%), and atypical in 89 (4.7%). PCA3 test was informative in 1887 (98.8%) patients. Means ± standard deviations (median) of PCA3 scores for benign, HGPIN, atypical, and CaP were 22.3 ± 27.9 (12.8), 37.6 ± 43.2 (24.1), 35.7 ± 36.2 (25.7), and 46.9 ± 48.1 (31.6; P <.05 benign vs CaP, benign vs HGPIN and atypical, HGPIN and atypical vs CaP). Sensitivity and specificity of PCA3 for CaP were 46.3% and 78.7%, respectively. CaP risk increased with progressively higher PCA3 score ranges from 14.8% for PCA3 <5 to 66.7% for PCA3 >100. Area under the curve (AUC) for the PCA3 receiver operating characteristics was not significantly different in men without prior bx (AUC = 0.716) compared with men with at least 1 prior nonpositive bx (AUC = 0.702). CONCLUSION: Post-TRUS bx urine is a valid sample for PCA3 testing. Patients with a negative bx and a positive PCA3 test may have a higher likelihood of unsampled CaP.

Comparative analysis of whole mount processing and systematic sampling of radical prostatectomy specimens: pathological outcomes and risk of biochemical recurrence.

PURPOSE: Whole mount processing is more resource intensive than routine systematic sampling of radical retropubic prostatectomy specimens. We compared whole mount and systematic sampling for detecting pathological outcomes, and compared the prognostic value of pathological findings across pathological methods. MATERIALS AND METHODS: We included men (608 whole mount and 525 systematic sampling samples) with no prior treatment who underwent radical retropubic prostatectomy at Vanderbilt University Medical Center between January 2000 and June 2008. We used univariate and multivariate analysis to compare the pathological outcome detection rate between pathological methods. Kaplan-Meier curves and the log rank test were used to compare the prognostic value of pathological findings across pathological methods. RESULTS: There were no significant differences between the whole mount and the systematic sampling groups in detecting extraprostatic extension (25% vs 30%), positive surgical margins (31% vs 31%), pathological Gleason score less than 7 (49% vs 43%), 7 (39% vs 43%) or greater than 7 (12% vs 13%), seminal vesicle invasion (8% vs 10%) or lymph node involvement (3% vs 5%). Tumor volume was higher in the systematic sampling group and whole mount detected more multiple surgical margins (each p <0.01). There were no significant differences in the likelihood of biochemical recurrence between the pathological methods when patients were stratified by pathological outcome. CONCLUSIONS: Except for estimated tumor volume and multiple margins whole mount and systematic sampling yield similar pathological information. Each method stratifies patients into comparable risk groups for biochemical recurrence. Thus, while whole mount is more resource intensive, it does not appear to result in improved detection of clinically important pathological outcomes or prognostication.

Androgen regulated genes in human prostate xenografts in mice: relation to BPH and prostate cancer.

Benign prostatic hyperplasia (BPH) and prostate carcinoma (CaP) are linked to aging and the presence of androgens, suggesting that androgen regulated genes play a major role in these common diseases. Androgen regulation of prostate growth and development depends on the presence of intact epithelial-stromal interactions. Further, the prostatic stroma is implicated in BPH. This suggests that epithelial cell lines are inadequate to identify androgen regulated genes that could contribute to BPH and CaP and which could serve as potential clinical biomarkers. In this study, we used a human prostate xenograft model to define a profile of genes regulated in vivo by androgens, with an emphasis on identifying candidate biomarkers. Benign transition zone (TZ) human prostate tissue from radical prostatectomies was grafted to the sub-renal capsule site of intact or castrated male immunodeficient mice, followed by the removal or addition of androgens, respectively. Microarray analysis of RNA from these tissues was used to identify genes that were; 1) highly expressed in prostate, 2) had significant expression changes in response to androgens, and, 3) encode extracellular proteins. A total of 95 genes meeting these criteria were selected for analysis and validation of expression in patient prostate tissues using quantitative real-time PCR. Expression levels of these genes were measured in pooled RNAs from human prostate tissues with varying severity of BPH pathologic changes and CaP of varying Gleason score. A number of androgen regulated genes were identified. Additionally, a subset of these genes were over-expressed in RNA from clinical BPH tissues, and the levels of many were found to correlate with disease status. Our results demonstrate the feasibility, and some of the problems, of using a mouse xenograft model to characterize the androgen regulated expression profiles of intact human prostate tissues.

PCA3 urine mRNA testing for prostate carcinoma: patterns of use by community urologists and assay performance in reference laboratory setting.

OBJECTIVES: Multiple trials have shown the high specificity of urine prostate cancer gene 3 (PCA3) compared with serum prostate-specific antigen (PSA) for biopsy detection of prostate carcinoma. We characterized the patterns of use of PCA3 by community urologists and determined the performance of PCA3 testing as a laboratory-developed test in a reference laboratory setting. METHODS: The urine PCA3 and PSA mRNA levels after digital rectal examination were determined using transcription-mediated amplification. The cutoff for a positive PCA3 score (PCA3/PSA mRNA x 10(3)) were pre-established at > or = 35. The PCA3 results were correlated with the serum PSA level, previous biopsy history, and the prostate biopsy findings. RESULTS: A total of 278 PCA3 tests were performed from December 2006 to June 2007. Of the PCA3 tested patients, 55.5% had previously undergone > or = 1 prostate biopsy; 92.7% had a PSA level > or = 2.5 ng/mL. The PCA3 test informative rate was 97.5%. For 50 samples that were also analyzed at a separate laboratory, concordance was achieved in 94%. The mean and median PCA3 score was 44.3 and 21.1, respectively. No correlation was found with the serum PSA level. The PCA3 test was negative in 16 of 19 patients with negative concurrent biopsy findings and positive in 8 of 11 with positive concurrent biopsy findings (sensitivity 72.7% and specificity 84.2%). Of 32 patients (70% with previous biopsy) who had undergone biopsy an average of 56 days after positive PCA3 test results, prostate carcinoma was detected in 41%. CONCLUSIONS: Urine PCA3 testing on the transcription-mediated amplification platform performed well as a laboratory-developed test. The high specificity of PCA3 was confirmed. In patients with elevated PSA levels and negative biopsy findings, PCA3 testing might be useful in choosing between repeat biopsy and more conservative follow-up.

Clinical utility of prostate carcinoma molecular diagnostic tests.

Instead of relying on serum prostate-specific antigen (PSA) to identify patients for prostate biopsy, new laboratory tests are needed that have improved specificity for prostate carcinoma (CaP), allow accurate classification of clinically insignificant CaPs, allow for detection of clinically significant CaP in patients without elevated serum PSA, and allow for identification of aggressive forms of CaP, which may warrant adjunctive or even molecularly targeted therapy in the future. Over the last several years, high-throughput gene expression profiling and proteinomics have led to the identification of genes and proteins that are specifically overexpressed in CaP. Molecular diagnostic techniques readily translated to the clinical laboratory have been incorporated into the development of new tests based on these novel molecular alterations in CaP. Some of these tests already have well-documented clinical utility, such as in facilitating prostate biopsy decisions, and are routinely available. The current review focuses on the biological, clinical, and laboratory aspects of the most promising of these current and near-future molecular CaP tests.

Incidence and location of prostate and urothelial carcinoma in prostates from cystoprostatectomies: implications for possible apical sparing surgery.

PURPOSE: Prostatic carcinoma (Pca) at cystoprostatectomy is usually an incidental finding with the majority thought to be clinically insignificant. Most studies have not specifically addressed the location of Pca or the incidence and location of in situ or invasive urothelial carcinoma (Uca) in prostates of cystoprostatectomy specimens. The frequency of involvement of the apex with these processes has clinical implications. Specifically urinary continence following orthotopic diversion may be enhanced by prostate apical sparing. In this study the pathological features of Pca and Uca, and the frequency of apical involvement were investigated in prostates from cystoprostatectomy specimens. MATERIALS AND METHODS: Whole mounted prostates from 121 consecutive cystoprostatectomy specimens were analyzed. Pca location, tumor volume, grade, stage, surgical margin and pelvic lymph node status of Pcas were assessed. Clinically insignificant Pcas had a volume of less than 0.5 cc without Gleason pattern 4, extracapsular extension, seminal vesicle invasion, lymph node involvement or positive surgical margins. Prostate involvement by Uca or urothelial carcinoma in situ (CIS)/severe dysplasia and its location were assessed. RESULTS: Of 121 prostates 50 (41%) had unsuspected Pca, of which 24 (48%) were clinically significant. Of Pcas 30 of 50 (60%) involved the apex, including 19 of 24 (79%) that were significant and 11 of 26 (42%) that were insignificant. Of 121 prostates 58 (48%) had Uca involving the prostatic stroma, noninvasive Uca or urothelial CIS/severe dysplasia in the prostatic urethra or periurethral ducts, of which 19 (33%) had apical involvement. Overall only 32 of 121 patients (26%) had no Pca or prostate Uca/CIS and only 45 (37%) had no clinically significant Pca or Uca/CIS in the prostate. However, 74 of the 121 patients (61%) had no prostatic apical involvement by Pca or Uca/CIS and 85 (70%) had no apical involvement by clinically significant Pca or Uca/CIS. Patients with prostatic apical involvement by invasive or in situ Uca uniformly had involvement of more proximal (toward the base) portions of the prostate. CONCLUSIONS: The majority of prostates from cystoprostatectomies had no involvement of the prostatic apex by Uca or clinically significant Pca. Hence, most patients may be candidates for prostate apical sparing. However, involvement of the apex by Uca in any patient raises concern about procedures that leave portions of the prostate urethra after cystectomy in an effort to improve continence. In candidates for orthotopic neobladder reconstruction removing all of the prostatic urethra and sparing the remainder of the prostatic apex may allow improved preservation of urinary continence with an acceptable low risk of clinical Pca progression. Whether future strategies for preoperative exclusion of apical Pca and intraoperative assessment of more proximal prostate to help exclude apical urothelial disease may identify patients suitable for prostatic apical sparing remains to be determined. The impact on functional outcomes and cancer control also require additional study.

Bioluminescence imaging of vascular endothelial growth factor promoter activity in murine mammary tumorigenesis.

Vascular endothelial growth factor (VEGF) is a major inducer of angiogenesis. We generated a transgenic reporter mouse, VEGF-GL, in which an enhanced green fluorescent protein-luciferase fusion protein is expressed under the control of a human VEGF-A promoter. The VEGF-GL mouse exhibited intense bioluminescence throughout the body at 1 week of age. The signals rapidly declined to a relatively low level as the mice grew. The adult VEGF-GL mouse showed restricted bioluminescence to the areas undergoing wound healing. In contrast, the VEGF-GL mice, which were crossed with mouse mammary tumor virus-polyoma virus middle T antigen transgenic mammary tumor mice, exhibited prominent bioluminescence in the tumors, correlating with VEGF transcription. Tumor bioluminescence was observed in the bigenic mice as early as 8 weeks, before tumors were palpable, and the signals increased with tumor growth. In conclusion, the VEGF-GL mouse permits longitudinal and quantitative assessment of VEGF promoter activity in vivo. The model should facilitate understanding of the molecular controls and pathways that regulate VEGF transcription in vivo.

Conditional expression of human 15-lipoxygenase-1 in mouse prostate induces prostatic intraepithelial neoplasia: the FLiMP mouse model.

The incidence and mortality of prostate cancer (PCa) vary greatly in different geographic regions, for which lifestyle factors, such as dietary fat intake, have been implicated. Human 15-lipoxygenase-1 (h15-LO-1), which metabolizes polyunsaturated fatty acids, is a highly regulated, tissue-specific, lipid-peroxidating enzyme that functions in physiological membrane remodeling and in the pathogenesis of atherosclerosis, inflammation, and carcinogenesis. We have shown that aberrant overexpression of 15-LO-1 occurs in human PCa, particularly high-grade PCa, and in high-grade prostatic intraepithelial neoplasia (HGPIN), and that the murine orthologue is increased in SV40-based genetically engineered mouse (GEM) models of PCa, such as LADY and TRansgenic Adenocarcinoma of Mouse Prostate. To further define the role of 15-LO-1 in prostate carcinogenesis, we established a novel GEM model with targeted overexpression of h15-LO-1 in the prostate [human fifteen lipoxygenase-1 in mouse prostate (FLiMP)]. We used a Cre- mediated and a loxP-mediated recombination strategy to target h15-LO-1 specifically to the prostate of C57BL/6 mice. Wild-type (wt), FLiMP+/-, and FLiMP+/+ mice aged 7 to 21, 24 to 28, and 35 weeks were characterized by histopathology, immunohistochemistry (IHC), and DNA/RNA and enzyme analyses. Compared to wt mice, h15-LO-1 enzyme activity was increased similarly in both homozygous FLiMP+/+ and hemizygous FLiMP+/- prostates. Dorsolateral and ventral prostates of FLiMP mice showed focal and progressive epithelial hyperplasia with nuclear atypia, indicative of the definition of mouse prostatic intraepithelial neoplasia (mPIN) according to the National Cancer Institute. These foci showed increased proliferation by Ki-67 IHC. No progression to invasive PCa was noted up to 35 weeks. By IHC, h15-LO-1 expression was limited to luminal epithelial cells, with increased expression in mPIN foci (similar to human HGPIN). In summary, targeted overexpression of h15-LO-1 (a gene overexpressed in human PCa and HGPIN) to mouse prostate is sufficient to promote epithelial proliferation and mPIN development. These results support 15-LO-1 as having a role in prostate tumor initiation and as an early target for dietary or other prevention strategies. The FLiMP mouse model should also be useful in crosses with other GEM models to further define the combinations of molecular alterations necessary for PCa progression.

Tumour volume is an independent predictor of prostate-specific antigen recurrence in patients undergoing radical prostatectomy for clinically localized prostate cancer.

UNLABELLED: Authors from the USA sought to establish the relationship between tumour volume, pathological stage and outcomes after radical prostatectomy. In a large series of patients they found that tumour volume was correlated directly with pathological stage, and that it was independently correlated with PSA recurrence. The authors suggested that tumour volume had a potential use for prognostication in patients undergoing radical prostatectomy. Two papers, one from the USA and one from Germany, advise a re-staging TUR in patients with superficial bladder cancer who are at high risk of early tumour progression. In a large series of patients they found that residual tumour after initial resection was commoner than might be expected, and that the second resection indicated the way to earlier radical treatment and a better prognosis. OBJECTIVE: To establish the relationship between tumour volume (TV), pathological stage and outcome after radical prostatectomy (RP), as TV is theoretically an important variable in prostate cancer pathology, but to date it has not been routinely reported and its independent prognostic significance is not well defined. PATIENTS AND METHODS: The study included 431 consecutive patients undergoing RP for clinically localized cancer, from January 2000 to January 2002, who had a pathological examination of totally submitted whole-mount processed RP specimens. In addition to Gleason grade, tumour stage and margin assessment by standard techniques, TV was determined by digital planimetry. The total TV or index TV, for cases with obvious discrete separate tumours, were correlated with pathological stage and prostate-specific antigen (PSA) recurrence. RESULTS: The mean (range) follow-up was 25.4 (6-51) months, and the mean TV for all patients was 3.28 (0.4-38.8) mL. There was a direct correlation between TV and pathological stage (P < 0.001). The TV for organ-confined and extraprostatic disease was 2.09 and 6.02 mL, respectively (P < 0.001). In a multivariate analysis, TV was an independent predictor of PSA recurrence (P = 0.04). The mean TV for patients with PSA recurrence vs no recurrence was 6.8 and 2.6 mL, respectively (P < 0.001). CONCLUSION: TV correlates directly with pathological stage in RP specimens; furthermore, it is independently correlated with PSA recurrence. TV has potential use for prognostication in patients undergoing RP, and may be combined with other well established clinical variables to aid in predicting outcomes.

Significance of atypical and suspicious small acinar proliferations, and high grade prostatic intraepithelial neoplasia on prostate biopsy: implications for cancer detection and biopsy strategy.

PURPOSE: We reviewed our results from a urological pathology reference laboratory with respect to the incidence of HGPIN, and atypical and suspicious lesions in the spectrum of ASAP. Subsequent CaP findings on repeat biopsy with relevant clinical implications were assessed. MATERIALS AND METHODS: A review of 42,667 prostate biopsies was performed. We defined atypical and suspicious as variants of ASAP with suspicious being more worrisome for CaP. Findings were correlated with the location of CaP on repeat prostate biopsy. RESULTS: The rate of subsequent CaP detection was significantly higher for an initial diagnosis of suspicious findings (51% or 54 of 107 cases) than for atypical findings (34% or 39 of 116) or HGPIN (22% or 79 of 358, p < 0.001). CaP was found on the same side of the prostate in 61 of 78 (78%), 30 of 39 (77%) and 41 of 54 patients (76%) with initial HGPIN, atypical and suspicious biopsies, respectively. There was no significant difference among the 3 groups in the likelihood of future CaP at the same site or the same side of the prostate. CONCLUSIONS: Patients with a suspicious biopsy were significantly more likely to have CaP on future biopsy than those with atypical findings or HGPIN, suggesting that there may be divisions with prognostic significance in the larger category of ASAP. To our knowledge the reproducibility of recognizing such divisions remains to be established. Neither atypical nor suspicious lesions were more likely than HGPIN to predict CaP at the same site or side of the prostate as the original diagnosis. Repeat biopsy may be indicated in any patient with HGPIN, or atypical or suspicious lesions and this biopsy should not be limited to the site or side of the original pathological findings.

Expression and role of Foxa proteins in prostate cancer.

The molecular mechanism(s) for prostate cancer progression to androgen independence are poorly understood. We have recently shown that Foxa1 and Foxa2 proteins are differentially expressed in epithelial cells during murine prostate development, growth, and adult function. Currently, the role of Foxa proteins in prostate cancer development and progression is unknown. Foxa protein expression was investigated in the LPB-Tag LADY mouse prostate cancer models, in human prostate cancer specimens, and various prostate cancer cell lines using Western blot and immunostaining analysis. In vitro transient transfection, studies were performed to investigate Foxa/prostate-specific gene regulation. Foxa1 was strongly expressed in areas of prostatic intraepithelial neoplasia (PIN) in both the androgen dependent 12T-7f and in the metastatic, androgen independent 12T-10 LADY models. Prominent Foxa1 and Foxa2 expression was observed in 12T-10 invasive undifferentiated neuroendocrine carcinomas, in the hormone independent and metastasizing 12T-10 derived, NE-10 allograft tumors, and in all metastatic lesions isolated from 12T-10 mice. Foxa1 protein expression was always observed in human prostate carcinomas, regardless of Gleason grade score, while Foxa2 was only detected in neuroendocrine small cell carcinomas and in some high Gleason score adenocarcinomas. Foxa proteins were also differentially expressed in three prostate cancer cell lines. Importantly, in vitro functional assays demonstrated that Foxa2 could activate androgen-dependent prostate-specific genes in an androgen receptor and ligand-independent manner. These results suggest that Foxa proteins are important in prostate carcinogenesis. In particular, Foxa2 may be involved in progression of prostate cancer to androgen independence. As such, Foxa proteins may represent novel targets for therapeutic intervention.

Forkhead box A1 regulates prostate ductal morphogenesis and promotes epithelial cell maturation.

We have previously shown that a forkhead transcription factor Foxa1 interacts with androgen signaling and controls prostate differentiated response. Here, we show the mouse Foxa1 expression marks the entire embryonic urogenital sinus epithelium (UGE), contrasting with Shh and Foxa2, which are restricted to the basally located cells during prostate budding. The Foxa1-deficient mouse prostate shows a severely altered ductal pattern that resembles primitive epithelial cords surrounded by thick stromal layers. Characterization of these mutant cells indicates a population of basal-like cells similar to those found in the embryonic UGE, whereas no differentiated or mature luminal epithelial cells are found in Foxa1-deficient epithelium. These phenotypic changes are accompanied with molecular aberrations, including focal epithelial activation of Shh and elevated Foxa2 and Notch1 in the null epithelium. Perturbed epithelial-stromal interactions induced by Foxa1-deficient epithelium is evident, as demonstrated by the expansion of surrounding smooth muscle and elevated levels of stromal factors (Bmp4, Fgf7, Fgf10 and Gli). The prostatic homeobox protein Nkx3.1, a known proliferation inhibitor, was downregulated in Foxa1-deficient epithelial cells, while several prostate-specific androgen-regulated markers, including a novel Foxa1 target, are absent in the null prostate. These data indicate that Foxa1 plays a pivotal role in controlling prostate morphogenesis and cell differentiation.

Use of tissue recombination to predict phenotypes of transgenic mouse models of prostate carcinoma.

Transgenic mouse models of cancer represent a powerful approach for exploring disease processes and testing potential therapeutic interventions. Currently, it is difficult to predict if a specific genetic manipulation will result in a desirable phenotype. The present study tests the idea that tissue recombinants recapitulate the pathologic features of the neoplastic prostate seen in transgenic mice, and would thus be suitable predictive models for new mouse design. The large probasin-large T-antigen (LPB-Tag) transgenic lines 12T-7f and 12T-10 were used as a basis for this study. Tissue recombinants of bladder epithelium (BlE) and urogenital sinus mesenchyme (UGM) were implanted under the renal capsule of athymic mice. Recombinants composed of BlE from 12T-10 LPB-Tag and wild-type (wt) UGM faithfully recapitulated the histopathologic and temporal features of intact transgenic mice of this line. Tissue recombinants using BlE from 12T-7f mice and wt UGM developed epithelial proliferation with atypia that lacked the associated hypercellular stroma seen in the intact 12T-7f line. Recombinants using 12T-7f UGM demonstrated that the hypercellular stroma results from stromal cell expression of the SV40 large T antigen. Corresponding to the recombinant phenotypes, stromal Tag immunostaining was observed in prostate tissues from intact 12T-7f but not 12T-10 mice. Similar stromal expression of Tag was also noted in the hypercellular TRAMP prostatic stroma. Further analysis revealed a previously unreported pattern of SV40T expression in the LADY and TRAMP models including ductus deferens and seminal vesicle stroma as well as region and cell type-specific patterns in the epididymis. The present study demonstrates the utility of using tissue recombination to explore organ-specific phenotypes. Recombination strategies should enable quick and cost-effective screening for likely phenotypes in transgenic animals. This comparison of tissue recombination to existing models shows that this approach can elicit new information on well-characterized models.

Detection and subcellular localization of two 15S-lipoxygenases in human cornea.

PURPOSE: There are two human 15-lipoxygenases (LOX), 15-LOX-1 and -2, which convert arachidonic acid to 15S-hydroxyeicosatetraenoic acid (15S-HETE). The presence of both 15-LOXs in the human cornea prompted this study to delineate their roles in the human corneal epithelium. METHODS: Human corneal epithelia from donor corneas and a human corneal epithelial (HCE) cell line were used in [1-(14)C]arachidonic acid incubations, Western blot analysis, and quantitative real-time RT-PCR. Cell cultures of HCE were treated with 15S-HETE to measure its effect on cell growth. HCE cells were transfected with plasmids to express green fluorescent (GFP) fusion proteins of 15-LOX-1 and -2, and in vivo laser confocal microscopy was performed to determine the subcellular localization of the 15-LOX fusion proteins. RESULTS: [1-(14)C]Arachidonic acid incubations yielded 15S-HETE as the only LOX product. Treatment with 15S-HETE (5-10 microM) reduced growth rate and induced apoptosis in cultured HCE cells in a dose-dependent manner. 15-LOX-2 but not 15-LOX-1 was detected by Western blot analysis, although we were able to detect similar levels of both 15-LOX mRNAs by real-time quantitative RT-PCR. 15-LOX-1 and -2 proteins showed different subcellular expression patterns. 15-LOX-2 GFP was expressed in the cytoplasm and nucleus (actively taken up into the nucleus). 15-LOX-1 GFP fusion protein expression was restricted to the cytoplasm. CONCLUSIONS: These findings indicate that 15-LOX-2 is the predominant 15-LOX protein in human cornea, and its product, 15S-HETE, plays a role in cellular proliferation. Because the two 15-LOXs have different subcellular compartmentalization, the authors hypothesize that their products are also compartmentalized and therefore exert different molecular effects in the human corneal epithelium.

Development and characterization of efficient xenograft models for benign and malignant human prostate tissue.

BACKGROUND: Various research groups have attempted to grow fresh, histologically intact human prostate cancer tissues in immunodeficient mice. Unfortunately, grafting of such tissues to the sub-cutaneous compartment was found to be associated with low engraftment rates. Furthermore, xenografts could only be established using high-grade, advanced stage, but not low- or moderate-grade prostate cancer tissues. METHODS: This paper describes methods for xenografting both benign and malignant human prostate tissue to severe combined immunodeficient (SCID) mice. We examine the efficiency and histopathologic consequences of grafting to the sub-cutaneous, sub-renal capsule, and prostatic orthotopic sites. RESULTS: Sub-renal capsule grafting was most efficient in terms of take rate (>90%) for both benign and malignant tissue. Orthotopic grafts consistently exhibited the best histopathologic differentiation, although good differentiation with continued expression of androgen receptors (AR) and PSA was also seen in the sub-renal capsule site. Sub-cutaneous grafting resulted in low take rates and the lowest level of histodifferentiation in surviving grafts. Grafted benign tissues in all sites appropriately expressed AR, PSA, cytokeratins 8, 18, and 14 as well as p63; carcinoma tissues did not express the basal cell markers. Grafting of tissues to castrated hosts did not affect the graft take rates (but was not practical in the case of the orthotopic site). Grafting followed by host castration resulted in epithelial regression with loss PSA and reduced AR expression at all three sites. CONCLUSIONS: These data suggest that sub-renal capsule and orthotopic grafting of human prostate tissue can be used for many basic scientific and translational studies.

NE-10 neuroendocrine cancer promotes the LNCaP xenograft growth in castrated mice.

Increases in neuroendocrine (NE) cells and their secretory products are closely correlated with tumor progression and androgen-independent prostate cancer. However, the mechanisms by which NE cells influence prostate cancer growth and progression, especially after androgen ablation therapy, are poorly understood. To investigate the role of NE cells on prostate cancer growth, LNCaP xenograft tumors were implanted into nude mice. After the LNCaP tumors were established, the NE mouse prostate allograft (NE-10) was implanted on the opposite flank of these nude mice to test whether NE tumor-derived systemic factors can influence LNCaP growth. Mice bearing LNCaP tumors with or without NE allografts were castrated 2 weeks after NE tumor inoculation, and changes in LNCaP tumor growth rate and gene expression were investigated. After castration, LNCaP tumor growth decreased in mice bearing LNCaP tumors alone, and this was accompanied by a loss of nuclear androgen receptor (AR) localization. In contrast, in castrated mice bearing both LNCaP and NE-10 tumors, LNCaP tumors continued to grow, had increased levels of nuclear AR, and secreted prostate-specific antigen. Therefore, in the absence of testicular androgens, NE secretions were sufficient to maintain LNCaP cell growth and androgen-regulated gene expression in vivo. Furthermore, in vitro experiments showed that NE secretions combined with low levels of androgens activated the AR, an effect that was blocked by the antiandrogen bicalutamide. Because an increase in AR level has been reported to be sufficient to account for hormone refractory prostate cancers, the NE cell population ability to increase AR level/activity can be another mechanism that allows prostate cancer to escape androgen ablation therapy.

15-Lipoxygenase-2 expression in benign and neoplastic lung: an immunohistochemical study and correlation with tumor grade and proliferation.

15-Lipoxygenase-2 (15-LOX-2) is an arachidonic acid-metabolizing enzyme expressed in prostate, lung, skin, esophagus, and cornea. In the benign prostate, it is expressed in differentiated secretory epithelial cells, where its enzymatic product 15-HETE may regulate transcription by activating the nuclear receptor peroxisome proliferator-activated receptor gamma (PPARgamma). 15-LOX-2 and 15-HETE formation are reduced in prostate carcinoma. The distribution of 15-LOX-2 in the normal lung and its expression in lung carcinomas has not been reported and was investigated in the current study by using immunohistochemistry and tissue microarrays (TMAs). In benign lung, 15-LOX-2 immunostaining was noted exclusively in type II pneumocytes, which are known to express PPARgamma. Of 160 lung carcinomas, 15-LOX-2 was expressed in non-small cell carcinomas (NSCLC), including 33 of 69 (48%) adenocarcinomas, with 10 of 16 (63%) bronchioloalveolar carcinomas immunopositive. Fourteen of 55 (25%) squamous cell carcinomas and 2 of 14 (14%) large cell carcinomas showed weak immunostaining. All 19 neuroendocrine tumors were negative. Better differentiated NSCLCs showed greater 15-LOX-2 expression, with a significant inverse correlation between 15-LOX-2 immunostaining and tumor grade (P < 0.03). A significant inverse correlation was also noted between 15-LOX-2 immunostaining and tumor cell proliferation (Ki-67 immunostaining; P < 0.0001). These findings suggest a possible role of 15-LOX-2 in regulating secretory differentiation and proliferation in benign lung and NSCLCs, particularly adenocarcinomas.