Limitations of reverse-transcriptase polymerase chain reaction analyses for detection of micrometastatic epithelial cancer cells in bone marrow.

PURPOSE: This study was designed to evaluate the potential of reverse-transcriptase polymerase chain reaction (RT-PCR) analyses for the detection of micrometastatic carcinoma cells in bone marrow (BM). PATIENTS AND METHODS: The specificity of RT-PCR assays with primers specific for various tumor-associated and organ-specific mRNA species was examined by analysis of 53 BM aspirates from control patients with no epithelial malignancy. In addition, BM samples from 63 patients with prostate cancer (n = 53) or breast cancer (n = 10) were analyzed by RT-PCR with primers specific for prostate-specific antigen (PSA) mRNA. As a reference method, all samples were analyzed simultaneously by an established immunocytochemical assay, using monoclonal antibodies (mAbs) against cytokeratins (CK) for tumor-cell detection. RESULTS: Seven of eight marker species could be detected in a considerable number of BM samples from control patients: epithelial glycoprotein-40 (EGP-40; 53 of 53 samples), desmoplakin I (DPI I; five of five), carcinoembryonic antigen (CEA; five of 19), erb-B2 (five of seven), erb-B3 (six of seven), prostate-specific membrane antigen (PSM; four of nine), and CK18 (five of seven). Only PSA mRNA was not detected in any of the 53 control BM samples. In serial dilution experiments, the PSA RT-PCR assay was able to detect five LNCaP prostate carcinoma cells in 4 x 10(6) BM cells. CK-positive cells were found in 20 patients (37.7%) with prostate cancer, while PSA mRNA was found in only 15 (28.3%; P = .04). Moreover, despite the recent observation that PSA is also expressed in mammary carcinomas, none of the 10 CK-positive BM samples were PSA mRNA-positive. CONCLUSION: Limiting factors in the detection of micrometastatic tumor cells by RT-PCR are (1) the illegitimate transcription of tumor-associated or epithelial-specific genes in hematopoietic cells, and (2) the deficient expression of the marker gene in micrometastatic tumor cells.

Immunocytochemical detection of isolated tumour cells in bone marrow of patients with untreated stage C prostatic cancer.

The micrometastatic spread of tumour cells is usually missed by conventional diagnostic techniques, although this spread largely determines the prognosis of patients with primary epithelial cancers. By use of the monoclonal antibody, CK2, to epithelial cytokeratin component number 18 (CK18), individual disseminated carcinoma cells present in bone marrow of cancer patients can now be identified. In the present study, this approach has been applied to patients with virginal stage C adenocarcinoma of the prostate. Double-sided aspirates of iliac bone marrow from 24 of 44 evaluable patients (54.4%) exhibited between one and 38 CK18-positive cells per sample of 2 x 10(6) mononuclear cells. In 13 of these 24 positive patients, CK-positive cells were only detected in one of the two aspirates analysed. There was no statistically significant correlation between this finding and established risk factors, such as the volume and histological grade of the primary tumour or the concentration of prostate specific antigen and prostatic acid phosphatase in serum. The follow-up time is too short to provide meaningful data on the prognostic significance of isolated CK18-positive cells in bone marrow, which, however, has been recently demonstrated in other types of primary epithelial cancers. In conclusion, the presence of prostatic tumour cells in bone marrow might be interpreted as an indicator of the metastatic capacity of an individual primary tumour. The immunocytochemical detection of these cells may, therefore, be useful for increasing the precision of current tumour staging, and to monitor minimal residual cancer in an individual patient.

[The susceptibility to urinary tract infections after antireflux operations (author's transl)]. / Zur Infektanfälligkeit des Harntraktes nach erfolgreicher Antirefluxplastik (I).

The results of a prospective bacteriologic follow-up of 21 consecutive female patients, who had an antireflux operation are given. Only those patients with a proven active susceptibility to urinary tract infection at the time of operation were included in this study. Of these patients, 89% had persistent recurrent urinary tract infection, though the reflux was corrected. Bacteriologic controls and long-term chemotherapy seem warranted in those patients after operation.

[Curative radiotherapy in stage C prostatic carcinoma. Experiences with radiotherapy-endocrine treatment]. / Kurative Radiotherapie beim Prostatakarzinom im Stadium C Erfahrungen mit einer radiotherapeutisch-endokrinen Behandlung.

In a prospective study 209 patients with prostatic cancer stage C who were seen between 1968-1974 received endocrine treatment and radiotherapy (6000 r) of the prostate without inclusion of regional lymph nodes. The patients were seen every 6 months up to 8 years. 247 transrectal biopsies and 7 transurethral resections of the prostate were performed. The analysis of the biopsies revealed that the combined endocrine and radiotherapy is followed by tumor neutralisation of about 70% in highly differentiated prostatic cancers and less effective in lower differentiated tumors. 80% of biopsies from tumors responding to therapy still show tumor cells. The 5-year-survival corresponded with tumor regression and tumor size. However a prognosis on the basis of these findings is not possible for the individual case.

[The value of direct localization studies for the surgical treatment of urinary tract infections (author's transl)]. / Der Wert direkter bakteriologischer Lokalisationsmethoden für die operative Behandlung von Harnwegsinfekten.

Techniques to localize an upper urinary tract infection--the bladder wash-out test and the ureteral catheterization method--are described. Case reports are given to demonstrate the usefulness of these methods when surgical treatment of urinary tract infections seems to be indicated.

[Occult reflux (author's transl)]. / Der okkulte Reflux.

Level diagnosis repeatedly performed in patients without roentgenologically demonstrable reflux demonstrated bladder bacteriuria in 80% of the cases. The remaining 20% had supravesical bacteriuria. We called this occult reflux, if reinfection was demonstrated. Contamination of the upper tract by occult reflux can, but must not induce pyelonephritis. Bilateral antireflux surgery frequently eliminates occult reflux of bacteria, so this seems a debatable method of treatment.

[Treatment of prostatic carcinoma stage C with combined perineal and transurethral cryosurgery (author's transl)]. / Kombinierte perineale und transurethrale Vereisung beim Prostatacarcinom im Stadium C.

31 patients with prostatic carcinoma stage C were treated by combined perineal and transurethral cryosurgery. Additionally in all patients a subcapsular orchiectomy was performed. A distinct tumour reduction was found in all patients. Six months after the freezing a palpable tumour could be detected only in 4 cases out of 29. At different time intervalls after the operation 57 biopsies of the prostate were obtained with the tru-cut-needle. In 14 patients no tumour tissue could be detected whereas tumour cells were found in the biopsies of the remaining patients. Combined perineal and transurethral cryosurgery therefore is a palliative form of treatment in prostatic carcinoma.

[The periurethral flora in female patients with recurrent urinary tract infection (UTI) (author's transl)]. / Untersuchungen über die Besiedelung der periurethralen Schleimhäute mit potentiell uropathogenen Keimen (Darmbakterien) bei Patientinnen mit rekurrierenden Harnwegsinfekten.

In 52 girls and 38 women with recurrent nonobstructive UTI 529 bacterial samples were obtained from the periurethral area under the following conditions: no bacteriuria and no chemotherapy for at least 3 weeks before the examination. In 225 samples we found E. coli and enterococci as the most frequent potentially uropathogenic bacteria. In the periurethral region these were found approximately twice as frequently as in urologically normal persons. 36 (40%) of the patients had no further recurrences during the follow-up, though 107 (44%) of the swabs showed an abnormal bacterial colonization. 54 (60%) of the patients, many of them with very frequent recurrences, in only 118 (41%) of the obtained samples showed an abnormal periurethral flora. The findings suggest that the periurethral flora is not the most important factor in the pathogenesis of UTI.

[Localizationstudies in female patients with recurrent urinary tract infection. II. Patients with ureterovesical reflux (author's transl)]. / Lokalisationsdiagnostische Untersuchungen bei Patientinnen mit rekurrierenden Harnwegsinfekten. II. Patientinnen mit vesiko-ureteralem Reflux.

In 17 girls and 5 women with urinary tract infection and low-pressure reflux we performed 23 bladder-washout-tests. 12 were interpreted as supravesical, 11 as vesical bacteriurias. All patients with vesical bacteriurias had a normal IVP. The majority of patients with supravesical bacteriurias had clubbing and scarring. The possibility of interpreting the supravesical bacteriurias as a sign for pyelitis rather than pyelonephritis is discussed.

[Localizationstudies in female patients with recurrent urinary tract infection. III. Patients after successfull antireflux-operation (author's transl)]. / Lokalisationsdiagnostische Untersuchungen bei Patientinnen mit rekurrierenden Harnwegsinfekten. III. Patientinnen nach erfolgreicher Antirefluxplastik.

In 14 girls and 5 women with recurrent urinary tract infections after successfull antireflux-operation we performed 72 bladder-wash-out-tests. Whereas preoperatively 50% of the patients had a supravesical bacteriuria, only 25% had it postoperatively. Most of the postoperativ vesical and supravesical bacteriurias were interpreted as reinfections. The demonstrated postoperativ ascension of bacteria in the upper urinary tract in spite of successfull surgical treatment cannot be taken as an argument against operation. The postoperative supravesical bacteriurias are less frequent and have less morbidity than the prepoerative supravesical infections.

[Familial vesico-ureteral reflux and familial urinary tract infection (author's transl)]. / Familiärer vesico-ureteraler Reflux und familiäre Infektanfälligkeit

In 6 families several members had vesico-ureteral reflux and/or recurrent urinary tract infections. From the analysis of the case reports the following tentative conclusions could be drawn: Familial reflux may be caused. 1. by primary insufficiency of the vesico-ureteral junction. 2. possibly by secondary changes consequent to a familial tendency to recurrent urinary tract infections.

[Potentially uropathogenic enterobacteria on the periurethral mucosa of urologically normal individuals (author's transl)]. / Untersuchungen über die Besiedelung der periurethralen Schleimhäute mit potentiell uropathogenen Keimen (Darmbakterien) bei urologisch gesunden Probanden

Swabs from the periurethra from 160 urologically normal individuals, male and female, of various age groups were examined for potentially uropathogenic enterobacteria. Children from 0--3 years constantly showed (100%) high numbers of different enterobacteria. The frequency of identification and the number of these bacteria diminished abruptly with advancing age. In adult women we found a prevalence of 15%. The periurethral flora with regard to enterobacteria is probably the result of a dynamic equilibrium between contamination and elimination. The important role of contamination is documented by the massive seeding in the 0--3 years age group and the abrupt diminuition after successful bowel training. The mechanism of elimination is unknown. Personal hygiene, the pH of the periurethral mucosa, a special antibacterial substance and the ecological equilibrium of the periurethral flora are discussed as possible factors.

[Kidney damage in sterile reflux]. / Nierenschäden bei sterilem Reflux.

On the basis of the data mentioned and taking into consideration the fact that the whole complex of problems is not yet clarified unequivocally we come to the following conclusions: 1. Up to now there are no proofs for the fact that the most frequent type of reflux, i.e. the simple reflux of the urine from the urinary bladder into the upper urinary passage causes a renal damage. 2. The complicated VUR may certainly cause a urinary stasis kidney. In simultaneous calyco-tubular reflux probably focal shrinkings of the parenchyma may develop.

[Localizationstudies in female patients with recurrent urinary tract infection. I. Patients without ureterovesical reflux (author's transl)]. / Lokalisationsdiagnostische Untersuchungen bei Patientinnen mit rekurrierenden Harnwegsinfekten. I. Patientinnen ohne vesiko-ureteralen Reflux.

In 16 girls and 12 women with recurrent urinary tract infection without ureterovesical reflux we performed 75 bladder washout-tests. 52 (72%) tests indicated a vesical, 19 (27%) of supravesical bacteriuria. The 16 girls had less often (18%) supravesical bacteriurias than the 12 women (58%). If follows that in the absence of reflux, ascension of bacteria from the bladder to the upper tract is a relatively frequent phenomenon, the pathogenic importance of which is not yet clear. Clinical manifestations and urographic aspect correlated rather poorly with the results of the bladder-washout-test.

["Removing of infection" through nephrectomy of the pyelonephritic atrophic kidney]. / "Infektsanierung" durch Nephrektomie pyelonephritischer Schrumpfnieren?

On the basis of own examinations is shown: 1. In female patients with non-obstructive infections of the urinary tract and unilateral pyelonephritic cirrhosis of the kidney the recidivations of the infections are in general not to be traced back to the flaring up of old germinal areas in them. 2. For this reason the removal of the diseased kidney also does not in general effect a sanation of the infection. In most cases it is not necessary, occasionally even harmful. 3. The nephrectomy with the aim of the sanation of the infection is indicated only then, when was proved that kidney is focus of the germ. This is possible only with the help of an exact localisation diagnostics and identification of the germs.

[Antibody-coaded bacteria in urine sediment of children with urinary tract infections (author's transl)]. / Nachweis antikörperbeladener Bakterien im Urinsediment bei kindlichen Harnwegsinfektionen.

In 40 girls and 9 women with recurrent, non-obstructive urinary tract infections, and in 5 patients with nephrolithiasis the site of infection was determined by 108 bladder washout tests. The corresponding sediments were examined for antibody-coaded bacteria using an immunofluorescence test (IFT). The IFT was positive in 18 (72%) of 25 supravesical bacteriurias. Out of 75 vesical bacteriurias 39 infantile and 7 adult (together 61%) specimens contained antibody-coaded bacteria. We think the IFT is of no diagnostic value in localizing recurrent urinary tract infections in girls. For adults such an evaluation is not yet possible due to our small number of cases.

[Possibilities of cytostatic therapy in adenocarcinoma of the kidney. I. Influence of vinblastine on the growth of transplanted tumors]. / Möglichkeiten der zytostatischen therapie beim adenokarzinom der niere. I. Einfluss von vinblastin auf das wachstum transplantierter tumoren.

The effect of vinblastine sulfate on the acceptance and growth of a very malignant, human adenocarcinoma of the kidney (RCCI) on nude mice was investigated. Without any treatment, this tumor has an acceptance rate of 100%. Subcutaneously transplanted pieces of tumor, measuring 3 X 3 X 1 mm, commence to grow rapidly 1 week after transplantation and reach a diameter of approximately 2 cm, 6 weeks thereafter. Tumors with a diameter of approximately 2 cm, treated during 6 weeks with vinblastine (0.6 mg/kg/week), continued to grow during treatment and thereafter. However, when these growing, pretreated tumors were transplanted on new experimental animals, they either were not accepted by them or grew very slowly. When animals with tumor transplants were treated for 6 weeks with vinblastine (same schedule as above), beginning on the 1st day after transplantation, tumor growth was markedly retarded in every case, but only very seldom the tumor was not accepted.

[Pathogenetic significance of the passage of microorganisms into the upper urinary tract in female patients with recurring, nonobstructive urinary tract infections. II. Pyelitis]. / Zur pathogenetischen Bedeutung von Keimaufstiegen in die oberen Harnwege bei Patientinnen mit rezidivierenden, nicht obstrucktiven Harninfekten. Teil II: Pyelitis.

With the help of three arguments which come from the data analysis of a special consulting hour is demonstrated the ascents of germs into the prevesical urinary tracts in female patients with relapsing, non-obstructive urinary infections not in the least in every case lead to a recognizable lesion of the kidney. The thesis is erected that the generally tabooed pyelitis by all means corresponds to a clinical reality. This thesis is discussed in the light of findings from animal experiments and clinical findings from literature.