Increased expression of type 2 3alpha-hydroxysteroid dehydrogenase/type 5 17beta-hydroxysteroid dehydrogenase (AKR1C3) and its relationship with androgen receptor in prostate carcinoma.

Type 2 3alpha-hydroxysteroid dehydrogenase (3alpha-HSD) is a multi-functional enzyme that possesses 3alpha-, 17beta- and 20alpha-HSD, as well as prostaglandin (PG) F synthase activities and catalyzes androgen, estrogen, progestin and PG metabolism. Type 2 3alpha-HSD was cloned from human prostate, is a member of the aldo-keto reductase (AKR) superfamily and was named AKR1C3. In androgen target tissues such as the prostate, AKR1C3 catalyzes the conversion of Delta(4)-androstene-3,17-dione to testosterone, 5alpha-dihydrotestosterone to 5alpha-androstane-3alpha,17beta-diol (3alpha-diol), and 3alpha-diol to androsterone. Thus AKR1C3 may regulate the balance of androgens and hence trans-activation of the androgen receptor in these tissues. Tissue distribution studies indicate that AKR1C3 transcripts are highly expressed in human prostate. To measure AKR1C3 protein expression and its distribution in the prostate, we raised a monoclonal antibody specifically recognizing AKR1C3. This antibody allowed us to distinguish AKR1C3 from other AKR1C family members in human tissues. Immunoblot analysis showed that this monoclonal antibody binds to one species of protein in primary cultures of prostate epithelial cells and in LNCaP prostate cancer cells. Immunohistochemistry with this antibody on human prostate detected strong nuclear immunoreactivity in normal stromal and smooth muscle cells, perineurial cells, urothelial (transitional) cells, and endothelial cells. Normal prostate epithelial cells were only faintly immunoreactive or negative. Positive immunoreactivity was demonstrated in primary prostatic adenocarcinoma in 9 of 11 cases. Variable increases in immunoreactivity for AKR1C3 was also demonstrated in non-neoplastic changes in the prostate including chronic inflammation, atrophy and urothelial (transitional) cell metaplasia. We conclude that elevated expression of AKR1C3 is highly associated with prostate carcinoma. Although the biological significance of elevated AKR1C3 in prostatic carcinoma is uncertain, AKR1C3 may be responsible for the trophic effects of androgens and/or PGs on prostatic epithelial cells.

An enema formulation of alicaforsen, an antisense inhibitor of intercellular adhesion molecule-1, in the treatment of chronic, unremitting pouchitis.

BACKGROUND: Pouchitis is the major long-term complication of ileal pouch-anal anastomosis for ulcerative colitis. The incidence of pouchitis is as high as 50% several years after surgery. Two-thirds of pouchitis patients suffer recurrence. Of those who recur, one-quarter suffer from chronic, unremitting pouchitis. Current treatments for this disorder are disappointing. AIM: To determine whether a topically administered enema formulation of ISIS 2302 (alicaforsen), an antisense inhibitor of intercellular adhesion molecule-1, can improve the clinical symptoms, endoscopic mucosal appearance and mucosal histology in patients with chronic, unremitting pouchitis, a disorder in which this molecule is over-expressed. METHODS: In an open-label, uncontrolled study, 12 patients with chronic, unremitting pouchitis were treated with 240 mg alicaforsen antisense enema nightly for 6 weeks. Clinical evaluation and endoscopy were performed at baseline and at weeks 3, 6 and 10. Pouchoscopy with biopsy was carried out at baseline and at weeks 6 and 10. The primary end-point was the reduction from baseline of the Pouchitis Disease Activity Index (PDAI) at week 6. Secondary end-points included the PDAI at week 10. Safety was evaluated by analysing the adverse events, vital signs and laboratory parameters. RESULTS: After 6 weeks of nightly alicaforsen enema, a statistically significant (n = 12, P = 0.001) reduction in the PDAI from baseline (11.42) to week 6 (6.83) was observed. Mean reductions in the endoscopy sub-score from baseline (5.25) to week 3 (3.08) and week 6 (2.58) were statistically significant (P = 0.0039 and P = 0.0005, respectively). The mean reductions in clinical symptom sub-score from baseline (3.75) to week 3 (2.33) and week 6 (2.25) were also statistically significant (P = 0.0156 and P = 0.0117, respectively). Ten of the 12 patients achieved a mucosal appearance score of 0 or 1 at endoscopy. Five of the 12 patients (42%) had a non-statistically significant decrease in the histology component of their PDAI from baseline to week 6. By week 6, seven of the 12 patients (58%) were in remission, as defined by PDAI < 7, with a mean decrease from baseline in PDAI score of six points. The alicaforsen enemas were well tolerated and no serious side-effects were noted. CONCLUSIONS: Antisense enema to intercellular adhesion molecule-1 is safe and well tolerated. In an open-label trial, it appeared to improve the PDAI score, clinical symptoms and endoscopic mucosal appearance. It may also improve the histology. In the light of the responses observed in this trial, a randomized, placebo-controlled trial is warranted.

Biomarker risk assessment and bladder cancer detection in a cohort exposed to benzidine.

BACKGROUND: Cancer screening with highly sensitive, specific biomarkers that reflect molecular phenotypic alterations is an attractive strategy for cancer control. We examined whether biomarker profiles could be used for risk assessment and cancer detection in a cohort of Chinese workers occupationally exposed to benzidine and at risk for bladder cancer. METHODS: The cohort consisted of 1788 exposed and 373 nonexposed workers, followed from 1991 through 1997. We assayed urothelial cells from voided urine samples for DNA ploidy (expressed as the 5C-exceeding rate [DNA 5CER]), the bladder tumor-associated antigen p300, and a cytoskeletal protein (G-actin). Workers were stratified into different risk groups (high, moderate, and low risk) at each examination based on a predefined biomarker profile. For workers who developed bladder cancer, tumor risk assessment was analyzed from samples collected 6-12 months before the cancer diagnosis. The associations between risk group and subsequent development of bladder cancer were analyzed by Cox proportional hazards regression analysis and logistic analysis, after adjustment. All statistical tests were two-sided. RESULTS: Twenty-eight bladder cancers were diagnosed in exposed workers and two in nonexposed workers. For risk assessment, DNA 5CER had 87.5% sensitivity, 86.5% specificity, an odds ratio (OR) of 46.2 (95% confidence interval [CI] = 8.1 to 867.0), and a risk ratio (RR) of 16.2 (95% CI = 7.1 to 37.0); p300 had 50.0% sensitivity, 97.9% specificity, an OR of 40.0 (95% CI = 9.0 to 177.8), and an RR of 37.9 (95% CI = 16.8 to 85.3). The risk of developing bladder cancer was 19.6 (95% CI = 8.0 to 47.9) times higher in workers positive for either the DNA 5CER or p300 biomarkers than in workers negative for both biomarkers and 81.4 (95% CI = 33.3 to 199.3) times higher in workers positive for both biomarkers. G-actin was a poor marker of individual risk. CONCLUSIONS: Occupationally exposed workers at risk for bladder cancer can be individually stratified, screened, monitored, and diagnosed based on predefined molecular biomarker profiles.

Abnormal G-actin content in single prostate cells as a biomarker of prostate cancer.

The distribution of altered G-actin was investigated in prostatic cells obtained by fine needle aspiration (FNA) from 27 excised prostate glands obtained during radical prostatectomy. FNA, which was used to obtain single cells for image analysis, sampled in the region of any nodules and in grossly normal areas of the contralateral lobes. Quantitative fluorescence-image analysis was used to assay the amount of G-actin in individual cells. Abnormal G-actin, a precursor cytoskeletal protein representing cytoskeletal rearrangements accompanying cellular transformation, was associated with the presence of adenocarcinoma in 22 of 27 specimens from the dominant nodule, but only 3 of 20 in the grossly normal specimens (P<.0001). The mean G-actin content of all samples from the dominant nodule was 113.2+/-6.87 and 69.57+/-4.47 from the grossly normal area, the difference being significant at P<.0001. Altered G-actin was not associated with Gleason score (P = .95), grade (P = .26), stage (P = .058), or tumor volume (P = .32), thereby indicating it is a general marker for prostate adenocarcinoma.

Loss of tissue transglutaminase as a biomarker for prostate adenocarcinoma.

BACKGROUND: Additional molecular tissue biomarkers for prostate carcinoma are needed to stratify patients with clinically suspicious findings, such as an elevated prostate specific antigen (PSA) with a negative biopsy, according to risk. METHODS: Prostate tissues from 43 cancer cases and 47 controls with no evidence of cancer were labeled for transglutaminase by immunohistochemistry. Immunoreactivity was quantified using the Autocyte Pathology Workstation. In addition, quantitative fluorescence image analysis was used to compare transglutaminase concentrations in cells obtained by fine-needle aspiration from excised prostates. Loss of gene expression was evaluated by reverse transcriptase-polymerase chain reaction and growth with 5-azacytidine. RESULTS: Visually, benign glands from controls generally expressed tissue transglutaminase, whereas regions with adenocarcinoma generally were negative. With quantitative immunohistochemistry, 41 of 43 adenocarcinoma of the prostate (CaP) cases expressed lower mean percentage areas positive for transglutaminase than did 30 of 30 benign prostatic hyperplasia (BPH) and 17 of 17 prostatitis cases (P < 0.0001; odds ratio [OR], 1577; 95% confidence interval (CI), 74-33, 820; relative risk [RR], 25; 95% CI, 6-95). Quantitative immunofluorescence of 3277 cells collected by FNA from 19 CaP cases and 645 cells from 5 cases of BPH showed that the mean content of transglutaminase was 93 femtograms (fg) for the CaP-derived cells and 138 fg for the BPH cells (P < 0.0001). Receiver operating curve analysis of the immunohistochemistry data showed an optimized threshold produced 95% sensitivity with 100% specificity. Growth of LNCaP cells with 5-azacytidine failed to stimulate transglutaminase expression, suggesting that loss of expression was likely not attributable to promoter methylation. CONCLUSIONS: Measurements of transglutaminase on tissue sections provides additional diagnostic information that is potentially useful for risk assessment of patients with suspicious clinical findings, such as nodules or positive PSA and negative biopsies, without overdetecting disease.

Primary intracerebellar osteosarcoma arising within an epidermoid cyst.

The authors report a case of a primary extraskeletal osteosarcoma arising within an epidermoid cyst in the parenchyma of the cerebellum in a 64-year-old woman. On initial presentation, the tumor involved the midline cerebellum without attachment to the surrounding dura mater or calvarium. Complete medical and radiologic evaluation failed to reveal a primary skeletal or other extraskeletal osteosarcoma. To our knowledge, this is the first reported case of a primary extraskeletal osteosarcoma within the cerebellum. Osteosarcoma as a primary brain tumor is exceedingly rare, and only three cases (all occurring within the cerebral hemispheres) have been reported previously. The histogenesis of primary sarcomas of the brain is not evident. The associated finding of an epidermoid cyst suggests the tumor originated from a teratoma.

Characterization of the DD23 tumor-associated antigen for bladder cancer detection and recurrence monitoring. Marker Network for Bladder Cancer.

Bladder cancer detection, monitoring, and prevention represent major problems that could be addressed with sensitive and specific biomarkers. The antigen recognized by the DD23 antibody, previously developed against a tumor-related antigen, was partially biochemically characterized, and its sensitivity and specificity in cancer detection and recurrence monitoring was evaluated. Quantitative fluorescence image analysis was used to quantify antigen content in exfoliated urothelial cells in a cross-section of patients with bladder cancers of all grades and stages and control populations. The antigen was found in tumor cells as well as normal-appearing urothelial cells, suggesting it represents a marker induced by the altered growth factor environment of a cancer-containing bladder. When used as a quantitative marker, the sensitivity for bladder cancer detection was 85%, and the specificity was 95%. No significant difference was seen between symptomatic and asymptomatic control populations, including patients with previous bladder cancers in the absence of a recurrence. In bladder cancer recurrence monitoring, results were consistently negative until just before detection of a recurrence. The biomarker reflects a "field effect" that occurs very late in tumorigenesis and seems to represent events common to most cancers involving the genitourinary tract. Western blotting showed the antibody recognized a dimeric protein. DD23 quantification in single cells may be particularly useful in targeting cystoscopic intervention for recurrence monitoring and, because of its high specificity, could be a tool for bladder cancer screening in high-risk groups.

Voice and healing after vocal fold epithelium removal by CO2 laser vs. microlaryngeal stripping.

Controversy exists regarding voice recovery after the use of laser vs. microforceps techniques in the removal of benign vocal fold lesions. The purpose of this study is to compare recovery of voice and healing between groups of cats undergoing vocal fold epithelium removal by CO2 laser and those having vocal fold stripping. Fourteen adult female cats underwent standardized unilateral vocal fold injuries by CO2 laser ablation or stripping. After a 6-week recovery period, phonations were evoked by electrical stimulation of the midbrain periaqueductal gray area. Phonations were recorded for acoustic analysis. The larynges were harvested, fixed, and sectioned for histologic correlation. Acoustic analysis showed the mean signal-to-noise ratios in the laser group (19.72) to be significantly higher than those in the stripped group (13.51) (p = 0.04). The stripped group showed significantly greater amplitude perturbation (8.68% vs. 2.43%, p = 0.02). No between-group difference was found for period perturbation. Histologically, the laser group showed minimal Reinke's space scarring and near-normal epithelial regeneration, and the stripped group showed marked subepithelial scarring, often involving the vocalis muscle. These results demonstrate superior recovery of voice and healing in animals undergoing vocal fold epithelium removal with the CO2 laser. Inferior outcomes seen in the stripped group may be related to difficulty in preserving Reinke's space during epithelium removal.

G-actin as a risk factor and modulatable endpoint for cancer chemoprevention trials.

Because tumorigenesis is an ongoing process, biomarkers can be used to identify individuals at risk for bladder cancer, and treatment of those at risk to prevent or slow further progression could be an effective means of cancer control given accurate individual risk assessment. Tumorigenesis proceeds through a series of defined phenotypic changes, including those in genetically altered cells destined to become cancer as well as in surrounding normal cells responding to the altered cytokine environment. A panel of biomarkers for the changes can provide a useful system for individual risk assessment in cancer patients and in individuals exposed to carcinogens. The use of such markers can increase the specificity of chemoprevention trials by targeting therapy to patients likely to respond, and thereby markedly reduce the costs of the trials. Previous studies in our laboratories showed the cytoskeletal proteins G- and F-actin reflect differentiation-related changes in cells undergoing tumorigenesis and in adjacent "field" cells, and a pattern of low F-actin and high G-actin is indicative of increased risk. Actin changes may be a common feature in genetic and epigenetic carcinogenic mechanisms. In a group of over 1600 workers exposed to benzidine, G-actin correlated with exposure, establishing it as an early marker of effect. In another study, a profile of biomarkers was monitored in patients who underwent transurethral resection of bladder tumor (TURBT) and received Bacillus Calmette Guerin (BCG) and/or DMSO. The primary objective was to determine how the defined biomarkers expressed in the tumor and the field correlate with clinical response and recurrence. DMSO, known to modulate G-actin in vitro, was used as an agent. Results strongly support the hypothesis that cytosolic G-actin levels measured by quantitative fluorescence image analysis (QFIA) can be an important intermediate endpoint marker for chemoprevention and that the p300 (M344) and DNA ploidy markers identify a high-risk group that requires more aggressive therapy and recurrence monitoring. Further research with other markers has shown that DD23 and nuclear actin, both of which identify late, specific changes, may increase the battery of useful markers. Taken together these studies show how biomarkers are employed to study individuals at risk, aid in the selection of chemopreventive compounds and assist in the understanding of the pathogenesis of malignancy.

Congenital mesoblastic nephroma: an immunohistochemical and lectin study.

Eight cases of congenital mesoblastic nephroma (CMN) were examined. Three CMNs were of the classical (typical) variant, two were cellular (atypical), and three showed a mixed pattern. A panel of nephron segment-specific tubular epithelial markers (the lectins Tetragonolobus purpureas, Phaseolus vulgaris erythroagglutinin, and Arachis hypogaea and antibodies to epithelial membrane antigen, cytokeratin, and Tamm-Horsfall protein) were used to differentiate epithelial structures within the tumor. Antibodies against vimentin, desmin, and muscle-specific actin were used as mesenchymal markers. A monoclonal antibody to the long (embryonic) form of polysialic acid (PSA) on the neural cell adhesion molecule was used as a putative renal oncodevelopmental marker. An antibody to proliferating cell nuclear antigen also was applied, which revealed increased proliferative rate in cellular CMNs. In addition to clearly entrapped native renal tubules, CMNs contain tubular structures with immature, dysplastic epithelium and occasional epithelial cell clusters embedded deep within the tumor. These immature tubules and clusters express distal nephron, including collecting duct markers and, occasionally, vimentin and PSA. We propose that these primitive tubules and epithelial structures may originate from the ureteric bud. An epithelial differentiation of the tumor cells also is possible. In one pure cellular CMN and two mixed CMNs the cellular component showed diffuse staining for PSA. The PSA (neural cell adhesion molecule) expression of the cellular component suggests that CMN may originate from the uninduced nephrogenic mesenchyme.

Extraskeletal osteosarcoma. A clinicopathologic review of 26 cases.

The clinical records and histopathologic features in 26 cases of extraskeletal osteosarcoma (ESOS) diagnosed at M.D. Anderson Cancer Center (Houston) between 1950 and 1987 were reviewed. Presentation was usually that of an enlarging soft tissue mass. The thigh (11 cases), upper extremity/shoulder girdle (three cases), and retroperitoneum (three cases) were the most common anatomic sites. Tumor size ranged from 2.5 to 30 cm. The predominant histologic pattern was osteoblastic in four cases, chondroblastic in two, fibroblastic or pleomorphic malignant fibrous histiocytoma (MFH)-like in four, giant cell type MFH-like in one, and small cell in one. Various mixtures of these patterns were seen in the remaining 14 tumors. The telangiectatic pattern was not seen as the predominant component in any primary tumor but was observed as a minor component. Thirteen tumors recurred locally and 16 metastasized; five patients had distant metastases at presentation. The lungs, bone, and soft tissue were the most frequent metastatic sites. Sixteen patients died of disease at 2 to 54 months, one patient died of unrelated causes at 61 months, seven patients were alive with no evidence of disease (NED) at 30 to 122 months, and two patients were alive with disease at 28 and 54 months, respectively. Tumor size (less than 5 cm versus greater than or equal to 5 cm) was the main prognostic factor; all patients alive with NED for whom accurate tumor measurements were available (six of seven) had neoplasms measuring less than 5 cm that were amenable to complete surgical excision. Histologic pattern and other clinicopathologic features did not significantly affect outcome.