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Background and Purpose: Analyses of permanent brachytherapy seed implants of the prostate have demonstrated that the use of a preplan may lead to a considerable decrease of dosimetric implant quality. The authors aimed to determine whether the same drawbacks of preplanning also apply to high-dose-rate (HDR) brachytherapy., Patients and Methods: 15 patients who underwent two separate HDR brachytherapy implants in addition to external-beam radiation therapy for advanced prostate cancer were analyzed. A pretherapeutic transrectal ultrasound was performed in all patients to generate a preplan for the first brachytherapy implant. For the second brachytherapy, a subset of patients were treated by preplans based on the ultrasound from the first brachytherapy implant. Preplans were compared with the respective postplans assessing the following parameters: coverage index, minimum target dose, homogeneity index, and dose exposure of organs at risk. The prostate geometries (volume, width, height, length) were compared as well., Results: At the first brachytherapy, the matching between the preplan and actual implant geometry was sufficient in 47% of the patients, and the preplan could be applied. The dosimetric implant quality decreased considerably: the mean coverage differed by -0.11, the mean minimum target dose by -0.15, the mean homogeneity index by -0.09. The exposure of organs at risk was not substantially altered. At the second brachytherapy, all patients could be treated by the preplan; the differences between the implant quality parameters were less pronounced. The changes of prostate geometry between preplans and postplans were considerable, the differences in volume ranging from -8.0 to 13.8 cm(3) and in dimensions (width, height, length) from -1.1 to 1.0 cm., Conclusion: Preplanning in HDR brachytherapy of the prostate is associated with a substantial decrease of dosimetric implant quality, when the preplan is based on a pretherapeutic ultrasound. The implant quality is less impaired in subsequent implants of fractionated brachytherapy.

Objective: To evaluate the perioperative gene-specific primed nested reverse transcription-polymerase chain reaction (RT-PCR) for prostate-specific antigen (PSA) and prostate-specific membrane antigen (PSMA) for staging patients undergoing radical prostatectomy and predicting biochemical recurrence., Patients and Methods: In 80 consecutive patients undergoing radical prostatectomy for prostate cancer, blood samples were drawn before, during and 1 and 7 days after removing the prostate. After buffy coat and mRNA extraction, gene-specific primed nested RT-PCR was performed for PSA, PSMA and glyceraldehyde-3-phosphate dehydrogenase mRNA, and Southern blot analysis of the PCR reaction., Results: The sensitivity of gene-specific RT-PCR to detect tumour cells was comparable with random primed RT-PCR. In the 80 patients the stage distribution was pT1 in two (2.5%), pT2 in 30 (37.5%) and > or = pT3 in 48 (60%); the nodal status was pN0 in 57 (71%), pN1 in 11 (14%) and pN2 in 12 (15%). The gene-specific RT-PCR reaction for PSA and PSMA was positive in no patients with pT1, 11 (37%) with pT2 and 23 (48%) with stage > or = pT3 disease. The result for PSA was positive in 12 (52%) and for PSMA in 11 (48%) of those with positive nodal status. Neither gene-specific RT-PCR for PSA or PSMA was able to predict organ-confined disease (P > 0.5). After a median (range) follow-up of 37 (11-67) months a biochemical recurrence was predicted in 65% of patients by preoperative RT-PCR for both PSA and PSMA, with a sensitivity, specificity, positive and negative predictive value of 58%, 80%, 87% and 47%, respectively; the assay after surgery predicted a recurrence in 73%, with respective values of 68%, 84%, 84% and 57%., Conclusions: Gene-specific primed nested RT-PCR for PSA and PSMA is a sensitive and simple assay; it might add substantial information for tumour staging in individual patients. RT-PCR before surgery allows the prediction of recurrence in 65% of cases and after surgery in 73%.

Background: The origin of extragonadal retroperitoneal germ cell tumors remains controversial. Whether they develop primarily in the retroperitoneum or whether they are metastases of a primary testicular tumor has long been debated., Patients and Methods: We retrospectively analyzed 26 patients treated as having primary extragonadal retroperitoneal germ cell tumors based upon the findings of testicular palpation by the referring physician. Testicular evaluation was then extended with ultrasonographical and histological examinations., Results: Biopsy of the extragonadal tumor was performed in 25 patients, confirming diagnosis of extragonadal retroperitoneal germ cell tumor. Prior to treatment patients were clinically evaluated by several physicians and the testes were not considered suspicious for testicular cancer. At urological workup, testes were found to be atrophic and/or indurated in 14 (54%) patients, enlarged in one (4%) and unremarkable in 11 (42%). Ultrasound examination of the testes in 20 patients showed pathological findings in all of them. Histology of the testis was available in 25 of 26 patients and revealed active tumor in three, intratubular germ cell neoplasia in four, scar tissue in 12, sclerosis in three, sclerosis and fibrosis in one, and fibrosis alone in two., Conclusions: So-called primary extragonadal germ cell tumors in the retroperitoneum are very likely a rare or non-existing entity and should be considered as metastases of a viable or burned-out testicular cancer until proven otherwise. All of our patients with histologically examined testes had pathological finding, 76% of which were either viable tumor or scars.

FMD--the most economically significant animal disease in the world during the last two centuries--has caused the last great panzootic from 1965 to 1967 in Europe. Since then it has become possible to eradicate centres of the epidemic still being present on the continent, mainly by means of the annual mass vaccination of cattle combined with rigid antiepizootic measures which include culling of infected animals. During the years after however there has been sporadic outbreaks again and again. They were mainly caused by virus that escaped from FMD laboratories and by the application of vaccines with residual infectiousity but also to an increasing extent they resulted from virus brought in from endemic regions of the world. The now as before high incidence of FMD in Asia and in wide parts of Africa and South America--after all 71 countries in these regions have been affected by outbreaks of FMD, the classic carrier disease, from 1998 to 2000--resulted in the spread of virus over far distances due to the globalization of world trade and the increasing traveling favoured by modern traffic facilities. Since 1980 in Europe particularly virus strains from the Middle East but also from other parts of Northern Africa and Asia have dominated the epidemiological situation such as the current epizootic in the United Kingdom and the outbreaks resulting from in three other member states of the European Union. In accordance with the EU guidelines the control of occurring outbreaks is exclusively carried out by stamping out. The limits of this procedure have become clearly obvious during the current epizootic in Britain. The use of emergency vaccination in the Netherlands shows a practicable alternative to the excessive mass culling of both infected animals and those being suspected of. The plurality and variability of the causative agent require a permanent observation of the epidemiological situation and of the virus strains involved in order to prevent the disease and to ensure the diagnosis and the topicality of the vaccines being available in the vaccine banks. Long-term success in the global combat against FMD can only be achieved on the basis of close international co-operation intended to restrain the disease significantly in the still endemically infected regions.

Purpose: We evaluate the predictive value of urinary cytokine levels of interleukin (IL) 8 and 18 for response in patients receiving intravesical bacillus Calmette-Guerin (BCG) for prevention of recurrences of superficial bladder cancer and treatment of carcinoma in situ., Materials and Methods: In 28 patients with superficial bladder cancer treated with BCG IL-8 expression in the urine during the first 6 hours after the first BCG instillation was determined. In 17 patients IL-18 levels were also evaluated during the first 12 hours after BCG instillation. IL-8 and 18 levels were determined by solid phase double ligand enzyme-linked immunosorbent assay., Results: In 12 of the 28 patients assessed for IL-8 expression disease recurred after a median followup of 66 months. Median IL-8 expression during the first 6 hours for these patients was 851 ng. (range 232 to 8,497). Median IL-8 expression during the first 6 hours in patients without recurrence was 4,200 ng. (range 432 to 12, 232). Of 8 patients with a followup of greater than 36 months 7 (88%) had no recurrent disease and IL-8 levels greater than 4,000 ng. Patients secreting more than 4,000 ng. IL-8 into the urine after BCG have a significantly higher chance of remaining disease-free (p <0.05), and those with elevated IL-18 expression have a significantly longer disease-free survival (p <0.05). After a median followup of 23 months (range 7 to 93) 6 of the 17 patients assessed for IL-18 expression had treatment failure. Median IL-18 expression in those patients during the first 12 hours was 2,632 pg. (range 860 to 8,298). Median IL-18 expression during the first 12 hours in patients without recurrence was 12,258 pg. (range 1,727 to 151,495)., Conclusions: In this study we confirmed the value of quantitative IL-8 expression in the urine during the first 6 hours after BCG instillation for superficial bladder cancer to predict freedom of disease. Furthermore, to our knowledge we report for the first time the potential value of IL-18 expression in the urine during the first 12 hours after BCG to predict freedom from disease. These findings may help improve the treatment of patients with superficial bladder cancer, especially by identifying those with a high risk of disease recurrence and progression after BCG therapy.

Background: Clinically, the lethal phenotypes of human prostate cancer are characterized by their progression to androgen-independence and their propensity to form osseous metastases. We reported previously on the establishment of androgen-independent (AI) human prostate cancer cell lines derived from androgen-dependent (AD) LNCaP cells, with androgen independence defined as the capability of prostate cancer cells to grow in castrated hosts. One of the sublines, C4-2, was found to be AI, highly tumorigenic, and metastatic, having a proclivity for metastasis to the bone., Methods: We established the AI and bone metastatic cell sublines B2, B3, B4, and B5 from the parental C4-2 subline, using a previously established coinoculating procedure. We determined the biologic behavior of the parental and derivative LNCaP sublines in vivo and in vitro, as well as their molecular and cytogenetic characteristics., Results: Unlike other human prostate cancer models, the LNCaP progression model shares remarkable similarities with human prostate cancer. We observed a comparable pattern of metastasis from the primary to the lymph node and to the axial skeleton, with a predominant phenotype of osteoblastic reaction; 25-37.5% of the animals developed paraplegia. Cytogenetic and biochemical characterizations of LNCaP sublines also indicate close similarities between human prostate cancer and the LNCaP progression model. Additional chromosomal changes were detected in B2-B5 sublines derived from C4-2 bone metastases. These LNCaP sublines were found to grow faster under anchorage-dependent but not -independent conditions. The in vitro invasion and in vivo metastatic potential of these LNCaP sublines surprisingly correlated with anchorage-dependent and not -independent growth. The derivative LNCaP sublines when cultured in vitro produced a substantially higher (20-30-fold) amount of basal steady-state concentrations of PSA than that of the parental LNCaP cells. PSA production was high initially, but was markedly reduced when the derivative cell lines were inoculated and allowed to grow long-term in vivo for the establishment of tumors and metastasis, suggesting that unknown host factors derived either from the prostate or the bone can effectively downregulate PSA expression by prostate tumor epithelium., Conclusions: The LNCaP model of human prostate cancer progression will help improve our understanding of the mechanisms of androgen-independence and osseous metastasis, and tumor-host determinants of PSA expression., (Copyright 2000 Wiley-Liss, Inc.)

Orthotopic bladder substitution is here to stay. Excellent long term functional results can be obtained. However, careful, restrictive patient selection before surgery, strict attention to some special surgical details during surgery and, probably most important, meticulous postoperative instruction and follow-up of the patient with an orthotopic bladder substitute are mandatory.

Human prostate cancer has the propensity to metastasize to the bone where reciprocal cellular interactions between prostate cancer and bone cells are known to occur. Osteopontin (OPN), a noncollagenous bone extracellular matrix, is a secreted adhesive glycoprotein with a functional RGD cell-binding domain that interacts with the alpha(v)beta3 cell surface integrin heterodimer. OPN has been associated with malignant transformation as well as being ligand to the CD44 receptor. Polyclonal antibodies to human OPN (hOPN) were prepared, and specificity was shown by preabsorption with recombinant hOPN. The stimulatory effect of hOPN protein and the inhibitory effect of hOPN antibody on human prostate cancer cell lines LNCaP and C4-2 were assessed by induction or inhibition of anchorage-independent growth, respectively. Expression of hOPN mRNA in prostate cancer cell lines and human prostate cancer tissue specimens were measured by mRNA blot analysis. Protein expression was assessed by immunohistochemistry in human prostate cancer specimens and by Western blot analysis in prostate cancer cell lines. hOPN stimulated anchorage-independent growth of the human prostate cancer cell lines LNCaP and C4-2 in vitro. Antibodies to hOPN inhibited the growth-stimulatory effect by endogenous OPN, which can be overcome by the addition of exogenous hOPN. hOPN mRNA and protein are expressed in human prostate cancer cell lines in vitro and in clinical human prostate cancer specimens. These findings taken together suggest that OPN may act as a paracrine and autocrine mediator of prostate cancer growth and progression.

Purpose: We prospectively evaluated a decrease in outflow obstruction caused by benign prostatic hyperplasia (BPH) with second generation thermotherapy., Materials and Methods: Transurethral microwave therapy was given with local anesthesia to 134 patients with urodynamically and cystoscopically documented obstruction by BPH and preserved detrusor function. Of 134 patients 67 (50%) had a general health score of 3 or greater., Results: Urgency was the main complaint during thermotherapy. After a median followup of 24 months (minimum 12) 100 patients were evaluable at 6 and 12 months. Of the initial 134 patients 17 (13%) who required additional treatment (repeat thermotherapy, transurethral prostatic resection, permanent cystostomy), 7 who died during followup for treatment unrelated reasons and 10 who were lost to followup or refused evaluation were excluded from further analysis. Mean International Prostate Symptom Score decreased from 22.5 before to 3.6 at 6 months after treatment and remained stable at 12 months. Mean Quality of Life Index improved from 4.3 before to 1 at 12 months after treatment. Mean maximum flow increased from 7.3 ml. per second before to 14.5 at 6 months and 13.9 at 12 months after treatment. Mean post-void residual decreased from 199 to 34.8 and 37.2 ml. at 6 and 12 months, respectively. Urodynamic evaluation of 84 patients after 6 months revealed a decrease in mean detrusor opening pressure from 96.8 to 53 cm. water and mean detrusor pressure at maximum flow from 99.8 to 59.7 cm. water. Mean ultrasonographic prostate volume decreased from 57.6 to 42.4 cc and a cavity in the prostate was documented in 65 of the 84 cases (77%). All changes between the pretreatment and posttreatment values at 6 and 12 months, respectively, were statistically significant (paired t test p<0.00001)., Conclusions: Targeted transurethral thermotherapy with second generation microwave equipment is minimally invasive, easy to apply and generally well tolerated with local anesthesia. Infravesical outlet obstruction and voiding pressures as assessed by pressure flow studies significantly decreased 6 months after treatment. Subjective voiding symptoms as well as post-void residual urine were significantly decreased, and urinary flow was improved 6 and 12 months after treatment of documented BPH.

LNCaP lineage-derived human prostate cancer cell lines C4-2 and C4-2B4 acquire androgen independence and osseous metastatic potential in vivo. Using C4-2 and C4-2B4 the goals of the current investigation were 1) to establish an ideal bone xenograft model for prostate cancer cells in intact athymic or SCID/bg mice using an intraosseous route of tumor cell administration and 2) to compare prostate cancer metastasis by administering cells either through intravenous (i.v.) or intracardiac administration in athymic or SCID/bg mice. Subsequent to tumor cell administration, prostate cancer growth in the skeleton was assessed by radiographic bone density, serum prostate-specific antigen (PSA) levels, presence of hematogenous prostate cancer cells and histopathologic evaluation of tumor specimens in the lymph node and skeleton. Our results show that whereas LNCaP cells injected intracardially failed to develop metastasis, C4-2 cells injected similarly had the highest metastatic capability in SCID/bg mice. Retroperitoneal and mediastinal lymph node metastases were noted in 3/7 animals, whereas 2/7 animals developed osteoblastic spine metastases. Intracardiac injection of C4-2 in athymic hosts produced spinal metastases in 1/5 animals at 8-12 weeks post-injection; PC-3 injected intracardially also metastasized to the bone but yielded osteolytic responses. Intravenous injection of either LNCaP or C4-2 failed to establish tumor colonies. Intrailiac injection of C4-2 but not LNCaP nor C4-2B4 cells in athymic mice established rapidly growing tumors in 4/8 animals at 2-7 weeks after inoculation. Intrafemoral injection of C4-2 (9/16) and C4-2B4 (5/18) but not LNCaP (0/13) cells resulted in the development of osteoblastic bone lesions in athymic mice (mean: 6 weeks, range: 3-12 weeks). In SCID/bg mice, intrafemoral injection of LNCaP (6/8), C4-2 (8/8) and C4-2B4 (8/8) cells formed PSA-producing, osteoblastic tumors in the bone marrow space within 3-5 weeks after tumor cell inoculation. A stepwise increase of serum PSA was detected in all animals. Reverse transcription-polymerase chain reaction (RT-PCR) to detect hematogenously disseminated prostate cancer cells could not be correlated to either serum PSA level or histological evidence of tumor cells in the marrow space. We have thus established a PSA-producing and osteoblastic human prostate cancer xenograft model in mice.

Objectives: A low-pressure ileal bladder replacement does not have any coordinated contraction during micturition, which is why we have evaluated various antireflux mechanisms in the context of a randomized prospective study., Material and Methods: 70 patients undergoing low-pressure ileal bladder replacement were randomized into 2 groups. An antireflux mechanism was performed in 35 patients and an afferent tubular segment was performed in the other 35 patients., Results: After a median follow-up of 57 and 45 months respectively, the functional capacity of the reservoir, incidence of urinary tract infections, urinary continence, voiding havits, and serum urea and creatinine were similar in the two groups. 11/67 (16.5%) evaluable ureteropelvic units with an antireflux mechanism and 2/69 (3%) units with an afferent tubular segment developed major dilatation due to stenosis of the antireflux mechanism or the ureteroileal anastomosis (Fisher's exact test, p < 0.009). No radiological reflux could be demonstrated during micturition in either group. A simultaneous increase of intravesical, intraabdominal and intrapelvic pressure was observed during a Valsalva manoeuvre., Conclusion: Protection of the upper urinary tract by an ileal afferent tubular segment has yet to be confirmed in a larger series of patients with a longer follow-up. Our results show that prevention of reflux is less important in the case of orthotopic low-pressure ileal bladder replacement. Consequently, creation of an antireflux mechanism, associated with a high complication rate, is probably not justified.

Purpose: We assessed whether interleukin-8 (IL-8), one of the first cytokines expressed in the urine after bacillus Calmette-Guerin (BCG) therapy for superficial urothelial tumors, may serve as a prognostic factor for treatment response., Materials and Methods: Of 20 patients with superficial urothelial cancer of the urinary tract treated with BCG 13 had superficial bladder cancer and 7 received BCG perfusion of the upper urinary tract due to inoperability, solitary kidney, renal insufficiency or bilateral disease. After intravesical instillation of 120 mg. BCG or after 2-hour perfusion of the upper urinary tract with 360 mg. BCG urine was collected at 6-hour intervals. IL-8 was determined by solid phase double ligand enzyme-linked immunosorbent assay., Results: IL-8 was stable in the urine for more than 48 hours. At a median followup of 36.5 months treatment failure occurred in 10 of the 20 patients, including 3 with recurrence, 6 with progressive disease and 1 with extensive carcinoma in situ. IL-8 excretion was more than 4,000 ng. in the first 6 hours after BCG therapy in all 10 patients who remained disease-free. In 9 of the 10 patients with recurrent disease IL-8 excretion was less than 4,000 ng. in the first 6 hours after BCG therapy. Patients secreting less than 4,000 ng. IL-8 into the urine during the first 6 hours after BCG therapy had a significantly higher risk of tumor recurrence and progression (p <0.0002)., Conclusions: Due to its stability and kinetics IL-8 determined in urine collected during the first 6 hours after BCG therapy may prove to be a prognostic factor for tumor recurrence and progression after BCG therapy.

Background: We assessed the frequency and molecular basis of p53 mutations in clinically localized prostatic adenocarcinoma., Methods: Prostate specimens were examined from 100 patients with clinically localized prostatic adenocarcinoma and 13 patients with benign prostatic hyperplasia (BPH). Mutations producing nuclear accumulation of p53 were detected immunohistochemically. Exon-specific mutations were analyzed by polymerase chain reaction amplification and single strand conformation polymorphism (PCR-SSCP) and sequenced., Results: p53 accumulation was detected in 5 tumors using antibody DO-1, and in 4 of these using antibody PAb 1801, but not in BPH. PCR-SSCP detected mutations in all 5 tumors, with alterations in exon 5 for 1 tumor, exon 6 for 3 tumors, and exon 7 for 1 tumor. An exon 6 mutation was also found in a tumor with no anti-p53 staining., Conclusions: p53 mutations are uncommon in clinically localized prostatic adenocarcinoma and absent from BPH. 5 of the 6 mutations were derived from locally invasive, prostate carcinomas, supporting the hypothesis that mutation of p53 is a late event in prostate carcinoma progression.

Overexpression of p53, as determined by immunohistochemical staining with the murine monoclonal antibody DO7, was determined in specimens of 46 primary superficial transitional cell bladder tumours (14 TaG2, 10 T1G2, 22 T1G3). A colon cancer specimen served as a positive control and normal mesenchymal cells in the specimens served as an internal negative control. An exceptionally high proportion 36/46 (78%) of the specimens were found to stain positively for p53 in over 20% of the cell nuclei. After a median follow-up of 7 years, ten patients developed progressive disease. Of these ten patients nine demonstrated p53 positivity, resulting in a sensitivity of 90%. However, 27 of the overall 36 patients (75%) with p53-positive tumours did not progress to a higher stage or metastatic disease. These findings suggest that p53 overexpression is not of predictive prognostic value in superficial transitional cell carcinoma. With 7 of 14 specimens (50%) of Ta tumours overexpressing p53, the results were suggestive of p53 mutation being an early event in carcinogenesis. When the threshold was set at 50% of the cell nuclei overexpressing p53, 16/46 (35%) classified as p53 positive. Of the 16 tumours staining positively for p53, 7 (46%) progressed and 9 (56%) did not. None of the Ta and 16 (50%) of the T1 tumours classified as positive. This more stringent definition of positivity still does not identify p53 positivity as a single prognostic factor. With 50% of T1 tumours classifying as positive, we still find that p53 mutation may be an early event in carcinogenesis of bladder cancer.

Purpose: Intestinal low pressure orthotopic bladder substitutes have no major coordinated contractions during micturition. Therefore, the importance and type of reflux prevention were assessed in a prospective randomized study., Materials and Methods: A total of 70 patients with an ileal low pressure bladder substitute was randomized to receive a nipple valve or an isoperistaltic afferent ileal tubular segment for reflux prevention., Results: After median observation times of 57 and 45 months, respectively, the results regarding functional reservoir capacity, incidence of infected urine, urinary continence, voiding habits and serum electrolytes, urea and creatinine were similar in both groups. Severe upper tract dilatation due to ureteroileal or nipple stenosis occurred in 9 of 67 evaluable reno-ureteral units (13.5%) in patients with antireflux nipples and in 2 of 69 (3%) in patients with an afferent tubular segment. This difference in favor of the latter cases is significant (Fisher's exact test p < 0.03). Video urodynamics did not show reflux of contrast medium during voiding in either group. A simultaneous intravesical, intra-abdominal and intrapelvic pressure increase was noted during the Valsalva maneuver., Conclusions: While long-term upper tract preservation by an afferent tubular ileal segment must be confirmed in larger patient series with longer followup, our results indicate that reflux prevention in patients with orthotopic low pressure bladder substitutes is not a major concern and does not justify the use of antireflux mechanisms with a high complication rate.

The lethal phenotypes of advanced prostate cancer are androgen independent (AI) and metastatic to the axial skeleton. Our laboratory has developed an AI mouse model of metastatic human prostate cancer. In this communication, we report the development of tumor suppressor gene therapy in this AI and metastatic (C4-2) cancer model. By using recombinant adenovirus as a delivery vehicle, we introduced a wild-type p53 tumor suppressor gene into prostate cancer cell lines. Despite a silent mutation at codon 152 of the p53 gene, C4-2 cells express functional, but low, levels of p53 protein. However, the other prostatic cell lines, PC-3 and DU145, have a deletion mutation and two point mutations of the p53 gene, respectively. In vitro studies showed that cell growth, as measured by the thymidine incorporation assay, was inhibited in the C4-2, PC-3, and DU145 cells infected with wild-type p53 adenovirus in comparison to control viruses. Recombinant wild-type p53 adenovirus inhibited prostate tumor growth and its production of prostate-specific antigen (PSA) when injected into C4-2 tumors in nude mice. All p53-treated mice were tumor free as long as 12 weeks after cessation of the 8-week treatment regimen. Two of 8 p53-treated mice developed small tumors growing at distant sites after a prolonged period of follow-up observation. Moreover, other AI prostate cancer cells, PC-3 and DU145, treated with Ad5-CMV-p53 failed to develop into tumors in vivo. This gene therapy strategy may be used against AI prostatic cancer regardless of p53 gene mutation status.

Background: Suramin, a polysulfonated naphthylurea and a recognized antitrypanosomal agent, has shown some promise in phase II clinical trials in the management of hormone-refractory human prostate cancer. Reduction of serum prostate-specific antigen (PSA) levels has been proposed as an end point for evaluating the antitumor efficacy of treatments for hormone-refractory prostate cancer., Purpose: We examined the antitumor effect of suramin in an in vivo mouse model of hormone-refractory human prostate cancer to determine whether a decrease in PSA levels reflects a reduction in tumor growth (volume). The tumors were induced in castrated, athymic nude mice by use of the androgen-independent, tumorigenic human prostate cancer cell line C4-2, which is a subline of the androgen-dependent, parental nontumorigenic cell line LNCaP. We also evaluated the effects of suramin in vitro on cell growth and the expression of PSA messenger RNA (mRNA) in both LNCaP and C4-2 cells., Methods: For the in vivo studies, 24 mice were given a subcutaneous injection of 5 x 10(6) C4-2 cells at each of four sites. Animals (n = 20) with tumor volumes greater than 1 mm3 or less than 5 mm3 were divided equally into two groups. Drug treatment was initiated in one group by administration of 1 mg suramin intraperitoneally, followed by 0.1 mg suramin at 10-day intervals to maintain constant serum levels. Tumor growth and PSA expression levels were monitored. For the in vitro studies, both LNCaP and C4-2 cells were exposed to 100-400 microgram/mL suramin, and cell growth was monitored by a quantitative crystal violet assay. PSA mRNA expression was assessed by northern blot analysis in cells treated with either 250 microgram/mL suramin, 400 ng/mL dihydrotestosterone (DHT) (positive control), or 0.5-75 microgram/mL hydrocortisone (to mimic the clinical use of hydrocortisone during suramin treatment to compensate for the loss of adrenocortical function). In some studies, the combined effect of DHT and suramin on PSA mRNA expression was also evaluated. A two-way analysis of variance was performed to evaluate the treatment differences, and P values were obtained from two-sided tests for statistical significance., Results: In vivo, suramin did not significantly affect the growth of androgen-independent C4-2 tumors (relative to the growth of tumors in 5% glucose-treated control animals; P = .76). However, suramin significantly decreased the ratio of PSA level to tumor volume (ng/mL PSA per mm(3) of tumor) (P<.001). Mice developed bone metastases in both treatment arms. Suramin affected the in vitro growth of LNCaP cells but not of C4-2 cells. Suramin diminished PSA mRNA expression in both LNCaP and C4-2 cells grown in vitro. Hydrocortisone had no effect on PSA mRNA levels., Conclusions: Although suramin inhibited the growth of androgen-dependent LNCaP cells, it did not inhibit the growth of androgen-independent C4-2 cells either in vitro or in vivo. Suramin significantly decreased PSA mRNA expression in both cell lines in vitro and depressed serum PSA levels in mice bearing androgen-independent C4-2 tumors., Implications: PSA level should be used with caution as an end point in clinical trials using suramin therapy for hormone-refractory prostate cancer.

Our laboratory has previously reported on the derivation of LNCaP cell sublines from LNCaP tumors maintained in castrated and intact athymic male mice. These LNCaP sublines differ from the parental line in tumorigenicity and androgen dependence. This paper demonstrates that one of these sublines acquired metastatic potential. When inoculated either s.c. or orthotopically, the C4-2 subline metastasized to the lymph node and bone with an incidence of 11-50%. Interestingly, the incidence of osseous metastasis was higher in castrated than in intact male hosts. We evaluated the chromosomal, immunohistochemical, and biochemical characteristics of the LNCaP sublines derived from C4-2 tumors that metastasized to the lymph node and bone. Cytogenetic analysis showed that all sublines were human and shared common marker chromosomes with the parental LNCaP cells. This experimental human prostate cancer model may permit, for the first time, the study of the molecular mechanisms underlying human prostate cancer metastasis.

The influence of the peptide diacetylsplenopentin (SP5) on an early protection of guinea pigs against foot-and-mouth disease (FMD) was investigated. 80% protection was achieved if SP5 was applied in a dose of 2 mg one day before challenge with FMD virus (FMDV) type O1 Lausanne strain. In comparison with this a conventional commercial adsorbate vaccine protected guinea pigs about 7 days after vaccination. An earlier protection can be obtained in general by vaccination with a higher content of the immunogen. A tenfold increase of the 146 S particle dose in a conventional oil adjuvanted FMDV vaccine protected pigs about 2 days earlier as observed after inoculating a "normal" vaccine (about 11 days).

RGD-containing peptides from the immunodominant region of VP1 between amino acids 135-160 from foot-and-mouth disease virus (FMDV) type O1 Kaufbeuren (O1K) prevented virus adsorption to piglet kidney (PK) cells. The highly conserved amino acid RGD sequence (Arg.-Gly.-Asp.) was a prerequisite of this effect. To prevent infection with 100-200 TCID50 in 10(6) PK cells, 20-250 micrograms of each peptide should have been added.

Swine plays a very particular role in FMD epizootiology. It is, therefore, absolutely necessary to have highly effective vaccines available for this species at all times. They have to ensure early buildup of long-lasting strong immunity even after one single application. Since the effectiveness of conventional adsorbate vaccines had proved to be insufficient, monovalent and trivalent oil emulsion vaccines were specifically developed of swine, using a GDR-made oil adjuvant. Stable immunity is very soon induced by them to endangered pig stock even against the immunologically problematic sub-types O1 and A5 after one single subcutaneous (s.c.) application of 2 ml (monovalent) or 5 ml (trivalent). Application establishes in s.c. connective tissue an oil emulsion depot that leads to formation of a vaccination granuloma. The immunocompetent cells identified in the latter are morphologically correlated to adjuvant action.

Coupled synthetic peptides, representing the sequences of amino acids 130-160, 141-160 and 145-160 of foot-and-mouth disease virus O1K protein VP1, induced virus-binding and virus-neutralizing antibody response in guinea pigs, rabbits, and pigs. We also detected antibody response in guinea pigs after immunization with uncoupled peptides and in cattle with 21 aa-peptide-Keyhole-limpet hemocyanin (-KLH). The best results were obtained from 21 aa-peptide-KLH and 31 aa-peptide with or without KLH or thyroglobulin as carrier. Our preliminary results show the induction of virus-neutralizing antibodies to be obviously influenced by length of the peptide as well as by the kind of carrier and coupling.

Three peptides of main epitope of FMD virus O1-Kaufbeuren, VP1 (16, 21, 31), were found to induce in the 130-160 sequence range, in free and/or carrier-bonded form, virus-neutralising antibodies in guinea pig, rabbit, mouse, swine, and cattle. Five carrier proteins were tested, with thyroglobulin, next to keyhole limpet hemocyanin (KLH), being most effective for 16-peptides (145-160) and 21-peptides (141-160 Tyr161). To protect guinea pig from FMD, minimum dosage of 21-peptide was found to be 2 x 8 micrograms. The immunogenic spectrum of peptides and conjugates proved to be broader than that of monovalent vaccines of inactivated virus. Free peptides were found to be also capable in vitro of inhibiting virus infection.

Riems FMD two-component oil emulsion vaccine was subcutaneously applied (5 ml) under field conditions to 855 store pigs of different age groups (trivalent--O1, A5, C). It produced early onset of lasting strong immunity against the three above FMD virus types. General condition of the animals and their body weight development were not adversely affected. Pea-size to walnut-size vaccination granulomas were recorded on slaughter as locally delimited reactions in 15 to 20 percent of vaccinated animals and were found to be morphologically correlated to adjuvant action. They were easily removed from the carcasses by excision of the vaccination point, with only minor loss of slaughter substance.

The GDR in 1977 had been the first country to introduce a laboratory method for potency testing of FMD vaccines on the basis of secured correlations between titres of virus-neutralising serum antibodies of immunised cattle, on the one hand, and their probit-transformed protection against FMD, on the other. This is now the only state-registered method for potency testing. The method has ever since worked well all over the place. An evaluation of results obtained between 1982 and 1988 revealed seasonal variations of anti-FMD immunogenesis in cattle. FMD immunisations from February to July proved more effective than those performed in the rest of the year, in that higher antibody titres were built up within 14 days from vaccination. These results were confirmed by rates of protection of cattle recorded from direct potency testing of immunised animals, between 1969 and 1977.

An analysis is made of the latest developments and importance of immunoprophylaxis against FMD, with reference to the epizootiological situation of the disease and its bearings, today, on industrialized livestock production. Comprehensive immunoprophylactic action (vaccination of all cattle every year) has been taken first by the GDR and later on by more European countries, as well. As a result of such action the disastrous impact of FMD and its associated high loss rate have been considerably mitigated. A decisive role, in that context, has been played by the development of effective inactivated vaccines for swine. An FMD live vaccine has been devised in GDR for after-care of swine. It is not only harmless to the animal, but, as well known, it provides numerous advantages over adsorbate vaccines. FMD immunoprophylaxis for full success depends primarily on the proper selection of virus strains suitable for vaccine produktion, vaccine quality, avoidance of post-vaccination damage or disease, and the availability of an efficient concept for vaccine application. The above problems are expounded in this paper, and derived from them are substantive conclusions for further studies, with the view to improving FMD immunoprophylaxis.

At all 3 studied FMD-viruses typs O2, A5 and C we could show the 73S unit in the analytical ultracentrifuge and in the electron microscope. 73S unit is found in the normal cycle of purification of virus and by density gradient centrifugation separated and purified. In CsCl pH 7.6 its density is 1.308 +/- 0,005 g/ml. Its sedimentation coefficient has a value of 72.7 +/- 1,5S. In electron microscope it show itself as a empty virus capsid. Its diameter is in partial purified preparations with 25 +/- 1 nm the same as of the virion. Its wall diameter is 2 to 3 nm. Further purification induced defiguration of particles and increase of its diameter. 73S unit dissociates in 19S and 12S units and shows a typical protein-UV-absorption spectrum with a maximum at 276 to 278 nm and a minimum at 250 nm. Emax/Emin is 2.3. Extinction coefficient E276nm is 1,4 mg/cm2. By sucrose density gradient centrifugation and titration of fractions in the complement fixation test it was detected, that croude virus solution contained already the 73S unit.

29 pigs (meat type of German Landrace) were kept in groups in a confined space (0.48 m2 a head; group E), in a normal area (1.2 m2 a head; group N) or in a normal area plus daily exercise (group L). After having been kept under these conditions for specified periods, the pigs were made to run on a band moving at 1.1 metres a second for 2 km. During exercise the heart rate increased to 275 beats a minute in group E and only 180 in group L. In group L the recovery pulse rate was greater and the absolute and relative heart weight smaller than in group L; rectal temperature after exercise was also higher than in group L. Values for group N lay between those of groups E and L. The relationship of respiration rate to rectal temperature after exercise was greater in group L than in groups E and N. Physical fitness of groups E and N was inferior to that of group L.