Endothelial vascular cell adhesion molecule 1 expression is suppressed by melanoma and carcinoma.

Vascular cell adhesion molecule 1 (VCAM-1) mediates extravasation of circulating leukocytes into inflamed tissues, and presumably, plays a role in the immigration of cytotoxic effector lymphocytes into tumor metastases. Since metastases are rarely cleared by blood-borne cells from the immune system, we asked whether the tumor may escape host defense by interfering with the mechanism of effector cell extravasation. Here we show that in mice and humans, VCAM-1 expression is repressed on tumor-infiltrating vascular endothelial cells in the lungs. On lung blood vessels distant from the tumor, VCAM-1 is constitutively expressed. When melanoma and endothelioma cells were cultured on either side of a Nucleopore membrane, the expression of VCAM-1 on the endothelioma cells was inhibited and VCAM-1 gene transcription was suppressed. We propose that the downregulation of VCAM-1 is a mechanism by which vascularized melanoma and carcinoma avoid invasion by cytotoxic cells of the immune system.

Expression and modulation of CD44 variant isoforms in humans.

CD44 is a ubiquitous surface molecule that exists as a number of isoforms, generated by alternative splicing of 10 "variant" exons. Little is known about the expression and function of the variant isoforms, except that certain isoforms may play a role in cancer metastasis. We produced mAbs against CD44 variant regions encoded by exons 4v, 6v, and 9v, by immunizing mice with a fusion protein spanning variant exons 3v to 10v. A comprehensive analysis of human tissues revealed that CD44 variant isoforms were expressed widely throughout the body, principally by epithelial cells. However there was differential expression of CD44 variant exons by different epithelia. Most epithelia expressed exon 9v, but much fewer expressed 6v or 4v. The regions of epithelia that expressed the highest levels of the variant isoforms were the generative cells, particularly the basal cells of stratified squamous epithelium, and of glandular epithelium. CD44 variant isoforms were also expressed differentially by leukocytes, with CD44-9v expressed at very low levels and CD44-6v and 4v virtually absent. However, CD44-9v and CD44-6v were the main variants that were transiently upregulated on T cells after mitogenic stimulation and on myelomonocytic cell lines by TNF alpha and IFN gamma treatment. Some epithelial cell lines could preferentially upregulate CD44-6v upon IFN gamma incubation. These results show that CD44 variant isoforms are expressed much more widely than first appreciated, and that expression of the variant isoforms on some cell types can be modulated by particular cytokines.

High expression level of alpha 6 integrin in human breast carcinoma is correlated with reduced survival.

We recently reported that alpha 6 integrin mediates experimental metastasis in mice by functioning in the adhesion of tumor cells to the vascular endothelium. In the current study, we investigated the expression of human alpha 6 integrin in invasive breast carcinomas of 119 women. In 50% of the tumors alpha 6 integrin was expressed in the majority of the cells, and this expression was correlated with reduced survival time. By contrast, the 24% of patients with breast tumors devoid of alpha 6 integrin expression all survived. The tumors were also evaluated for clinical risk factors including histological grading and steroid receptor level. The combination of these factors with alpha 6 integrin expression was superior in predicting overall survival than considering the other factors alone. The correlation with decreased survival time was consistent, regardless of whether the tumors expressed the alpha 6 integrin A or B forms, which differ in their cytoplasmic domain. On the basis of this pilot study we consider alpha 6 integrin expression to be a novel prognostic marker for human breast cancer.

Cytogenetic analysis of multifocal bladder cancer supports a monoclonal origin and intraepithelial spread of tumor cells.

Bladder cancer is often characterized by a multifocal growth pattern. This observation has given rise to the hypothesis of "field cancerization," predicting a polyclonal origin of multiple tumors rising from an area of independently transformed mucosa cells. On the other hand, genetic studies suggested a monoclonal origin. To address these contradictory hypotheses, we performed comparative genomic hybridization (CGH) on 32 tumors originating from six bladder cystectomy specimens. All tumors derived from the same patient showed a set of 7-13 identical chromosomal aberrations and additional individual alterations. Most striking were the findings of 17p losses in all (32 of 32) tumors of the six cystectomy specimens and 20p gains in all tumors of four bladders, as well as an unexpected high number of chromosomal changes (20.4 alterations per tumor on average). To clarify a possible role of the TP53 tumor suppressor gene on 17p13, we applied immunohistochemistry and sequence analysis on the tumors and additional 52 mucosa samples. Identical TP53 mutations and protein overexpression was found in individual tumors only as well as in mucosa samples from continuous areas. Our results not only provide further evidence for a monoclonal origin of multifocal bladder cancer but also point at intraepithelial migration of tumor cells carrying specific chromosomal aberrations.

CD44 isoforms correlate with cellular differentiation but not with prognosis in human breast cancer.

CD44 is a transmembrane glycoprotein occurring in several isoforms with different extracellular regions. The various transcripts are encoded by one gene locus containing 20 exons, of which at least 10 can be alternatively spliced in nascent RNA. Isoforms encoded by the variant exons (termed CD44v) are highly restricted in their distribution in nonmalignant tissue as opposed to the standard form of CD44 (CD44s) abundant in many tissues. Specific variant isoforms containing exon 6v have been shown to render nonmetastatic rat tumor cells metastatic. Based on the prominent role in rat metastasis formation, CD44v isoforms were suggested to be involved in human tumor progression. Correlations between prognosis and expression of CD44v have been reported for gastric and colon carcinoma, for non-Hodgkin's lymphoma, and recently for breast carcinoma. We evaluated the expression of CD44 isoforms in node-positive (n = 119) and node-negative (n = 108) cases of breast carcinoma by immunohistochemistry using CD44v exon-specific mAbs. In a subset of 43 cases of high-risk patients, reverse transcription-PCR was used to determine the exon composition of the transcripts. Protein and RNA expression data were probed statistically for their correlation to survival of the patients and clinical risk factors. In contrast to recently published data (M. Kaufmann et al., Lancet, 345: 615-619, 1995), in our cohort disease-free and overall survival data did not indicate significant correlations with the expression of the analyzed isoforms in univariate and multivariate analyses. Comparison of CD44 protein expression with established clinical risk factors for survival such as tumor size (pT1+pT2) and histological grading revealed correlations with the presence of CD44s (P = 0.02 and P = 0.03, respectively) and CD44-9v (P = 0.05 for histological grading). Carcinoma tissues with elevated estrogen and progesterone receptor levels showed positive correlation with CD44-6v (P = 0.001), while a trend for significant coexpression of CD44s and CD44-9v isoforms was observed in estrogen receptor-positive tissues (P = 0.08 and 0.06, respectively). In breast cancer, CD44s, CD44-9v, and CD44-6v are apparently markers for cellular differentiation but not for tumor progression. Our data suggest that steroid hormone receptors may be associated with the in vivo expression of CD44-6v-containing isoforms in human mammary carcinoma.

Applied Sciences: Absence of CD44-standard in human neuroblastoma correlates with histological dedifferentiation, N-myc amplification and reduced survival probability.

Expression of CD44 was examined by immunohistochemistry in 205 primary neuroblastomas together with histological grading according to the Shimada classification at the time of diagnosis. In addition, Southern blot analysis to determine N-myc gene amplification was carried out in the same tissue. When compared with clinical data such as stage, age and event-free survival probability it was found that CD44 expression characterizes well differentiated tumours and thus correlates with prognosis (event-free survival probability in CD44 positive patients 0.6 (n = 129) vs. 0.0 (n = 21) in CD44 negative patients). In tumours with N-myc = 1, CD44 positivity was found in 91% of patients as compared to 57% of patients with N-myc > 1 in tumours. All tumours of 13 patients with metastatic stage 4s (with good prognosis) showed CD44s expression. Thus, detection of CD44s expression might serve as a prognostic indicator which can be rapidly detected at diagnosis.

[Contraction of the hip and fecal drainage via a fistula tract 30 years after "appendectomy"]. / Stuhlfistel und Hüftgelenkskontraktur 30 Jahre nach "Blinddarmoperation".

A 46-year-old female was admitted with increasing fecal drainage via a fistula tract in the right inguinal region. She had a history of surgery for appendicitis 30 years previously, from which there was disturbed wound healing resulting in a blunt fistula, and the patient suffered from contraction of the right hip. Computed tomographic scan and ultrasound demonstrated an inflammatory mass in the right inguinal region. Colonoscopy demonstrated a stenosis of the rectosigmoid junction but did not provide any further specific information. Surgery revealed the presumed diagnosis of complicated Crohn's disease, but an advanced squamous cell carcinoma was also identified. The patient died 23 months later due to generalized tumor. Although malignant transformation of a fistula tract is rare, this case demonstrates that long-standing fistulas should be cured as far as possible without significant morbidity. In the case of incurable fistulas, malignancy must definitely be excluded if the clinical appearance of the fistula changes.

Differentiation and prognosis of neuroblastoma in correlation to the expression of CD44s.

Cell-cell and cell-extracellular matrix interactions mediated by cell adhesion molecules (for example CD44) play an important role in the cascade of metastasis and the progression of human malignant tumours. The most important aim of this review was, on the basis of our results and the literature, to show the correlation between the expression of CD44s and differentiation and prognosis of neuroblastoma. Surprisingly and in contrast to most other malignant tumours, neuroblastomas exhibited an inverse correlation between CD44s expression and tumour progression. It can be stated that CD44s is a prognostic marker in neuroblastoma which correlates significantly with the grade of tumour cell differentiation, but not with clinical stage. Moreover, there exists a statistically significant correlation between MYCN oncogene amplification and the lack of CD44s expression.

Comparison of DNA aneuploidy, chromosome 1 abnormalities, MYCN amplification and CD44 expression as prognostic factors in neuroblastoma.

A comparison of the prognostic impact of five molecular variables in a large series was made, including tests of their nonrandom association and multivariate analysis. Molecular data were available for 377 patients and MYCN amplification, cytogenetic chromosome 1p deletion, loss of chromosome 1p heterozygosity, DNA ploidy and CD44 expression were investigated. Their interdependence and influence on event-free survival was tested uni- and multivariately using Pearson's chi 2-test, Kaplan-Meier estimates, log rank tests and the Cox's regression model. MYCN amplification was present in 18% (58/322) of cases and predicted poorer prognosis in localised (P < 0.001), metastatic (P = 0.002) and even 4S (P = 0.040) disease. CD44 expression was found in 86% (127/148) of cases, and was a marker for favourable outcome in patients with neuroblastoma stages 1-3 (P = 0.003) and 4 (P = 0.017). Chromosome 1p deletion was cytogenetically detected in 51% (28/55), and indicated reduced event-free survival in localised neuroblastoma (P = 0.020). DNA ploidy and loss of heterozygosity on chromosome 1p were of less prognostic value. Most factors of prognostic significance were associated with each other. By multivariate analysis, MYCN was selected as the only relevant factor. Risk estimation of high discriminating power is, therefore, possible for patients with localised and metastatic neuroblastoma using stage and MYCN.

Patterns of chromosomal imbalances in muscle invasive bladder cancer.

Cytogenetic investigations of bladder cancer suggested that development and progression is characterized by specific chromosomal aberrations. In order to identify genetic changes linked to muscle invasive tumors and metastatic growth we analyzed 67 bladder carcinomas (30 pT1 and 37 pT2-4) by means of comparative genomic hybridization (CGH). The most frequent changes were gains of chromosome 1q (54%), 8q (54%), 17q (49%), 2p (30%), 12 (30%), 5p (25%), 3q (24%) and 6p (24%) as well as losses of 11p (43%), 8p (42%), 9p (36%), 11q (34%), 2q, 4q, 5q (30% each), 9q (27%) and 10q (27%). Previously not described amplifications were found at 5p11-p13, 7q21-q31, 9p24 and 17q24-q25. Gains of 3q, 7p, and 18p were markedly more frequent in pT2-4 in comparison to pT1 carcinomas but the difference did not reach statistical significance. Non-metastatic tumors showed more aberrations on average than metastatic carcinomas, although no particular change was found to be predominating in either group. Our data confirm previous findings of strong genetic similarities between minimally and deeply invasive bladder carcinomas but argue for differences between metastatic and non-metastatic disease.

Prognostic importance of the expression of CD44 splice variants in oral squamous cell carcinomas.

Considering squamous cell carcinomas (SCCs) of the oral cavity and oropharynx the molecular mechanisms underlying the infiltration and destruction of adjacent tissue as well as the metastatic spread are largely unknown. In this context, the detection of defective expression of cellular adhesion molecules in the tumour cells, e.g. CD44, might be important and correlated with prognosis. Paraffin-embedded tumour-tissue from 99 patients with primary oral and oropharyngeal SCC, additionally including corresponding lymph-node metastases in nine cases, was analysed for expression of the CD44 splice variants v4, v5, v6, v7, and v9 by means of immunohistochemistry. A diminution of at least one of the examined CD44 isoforms compared to the normal oral epithelium was observed in 39.4% of the squamous cell carcinomas. No correlations could be found between CD44 expression and pT- or pN-stage. However, decreased expression of v9 was correlated with higher histological grade (p < 0.001). Moreover, reduced CD44 expression was a statistically significant independent predictor for shorter survival time (p = 0.002) as well as shorter recurrence-free interval (p = 0.004) in addition to pT- and pN-stage. The separate analysis showed that particularly the decreased v7 (p = 0.04) and v9 (p < 0.02) expression in the tumour cells was associated negatively with survival.

CD44s, E-cadherin and PCNA as markers for progression in renal cell carcinoma.

CD44s, E-cadherin and PCNA play an important role in the tumorigenesis of renal cell carcinoma (RCC), although their significance in the clinical progression of RCC is still not clear. In n = 137 cases of operatively resected RCC the expression of CD44s, E-cadherin and PCNA was semiquantitatively analyzed by the APAAP immunohistochemical method. The mean observation period of the n = 74 (54%) of patients with no evidence of disease was 52.6 months and for the n = 63 (46%) patients with progressive disease was 18.7 months. The mean progression free period occurring with absent or low levels of CD44s and PCNA expression was 79 and 118+ months, and with strong expression was 12 and 18 months (p < 0.001), respectively. With strong E-cadherin expression the progression free period was 110+ months, and with low expression 61 months (p = 0.01). A high CD44s/E-cadherin ratio and an above average PCNA expression were identified as independent prognostic parameters for the progression tendency of RCC.

Cytology and image cytometry after colonic lavage: a complementary diagnostic tool in patients with ulcerative colitis.

BACKGROUND: Patients with extensive, long-standing ulcerative colitis have increased risk of colorectal cancer. AIMS: To improve the detection of high-risk patients, using a combination of colonic cytology, histology, and DNA image cytometry after segmental colonic lavage. PATIENTS: A series of 16 patients (8 high-risk patients) with ulcerative colitis were investigated. METHODS: After segmental lavage step, biopsies were obtained. Gradient centrifugation of the colonic fluid was performed for isolation and purification of epithelial cells. The smears and biopsy specimens obtained were stained for routine interpretation and for DNA image cytometry. RESULTS: Segmental lavage could be performed in all patients. Specimens from two high-risk patients showed low grade dysplasia and atypia by means of histology and cytology, respectively. In one patient, without increased colorectal cancer risk, atypia was detected. Three patients in the high-risk group, two of those diagnosed as positive for dysplasia and atypia, showed aneuploidy histologically and cytologically. DNA aneuploidy, in cytological material, was found exclusively in three low-risk patients, one of those had atypia cytologically. CONCLUSIONS: Isolation and purification of epithelial cells after segmental colonic lavage using density gradient centrifugation can be performed as part of routine endoscopy. It provides information about atypical cells and DNA aneuploidy as additional markers of malignant transformation. The combination of cytologic examination and DNA image cytometry might improve the detection of high-risk ulcerative colitis patients.

Metastatic retroperitoneal paraganglioma in a 16-year-old girl. Case report, molecular pathological and cytogenetic findings.

Retroperitoneal paraganglioma is a rare tumor, especially occurring in childhood and adolescence, with a marked tendency to become biologically malignant. It has not been possible to predict the clinical outcome of paraganglioma patients by conventional histology, hence malignancy can only be demonstrated by the occurrence of metastatic lesions. Currently, only limited information on the genetics of this tumor is available. We report on a 16-year-old girl with a large retroperitoneal paraganglioma and an osseous metastasis to the first lumbar vertebra. In addition to morphological and immunohistochemical examinations, a molecular cytogenetic analysis was performed. Comparative genomic hybridization (CGH) revealed imbalanced chromosomal aberrations with a loss of chromosome 1p and a gain of 1q, indicating isochromosome 1q. A loss of chromosome 3 as well as low-level gains of chromosomes 4, 5, 6q, 11q and 13q were detected. A PCR-based microsatellite analysis of 1p confirmed the loss of heterozygosity, including NB1 and NB2 putative tumor-suppressor gene regions. Telomerase activity, which is found in the majority of malignant tumors, could not be detected. The case presented here is the first more comprehensive molecular genetic analysis of a sporadic malignant paraganglioma.

The immunohistochemical reactivity of an anti-epithelial monoclonal antibody (b-12) in meningiomas.

Studies on immunohistochemical localization of the epitope b-12 were performed in 15 meningiomas using an indirect immunoperoxidase method. 2 of 15 meningiomas stained for b-12. The b-12 antigen represents a further epithelial marker detectable in meningiomas.

[Amelanotic melanoma of the rectum]. / Das amelanotische Melanom des Rectums.

We report the very rare case of an amelanotic anorectal melanoma. Until now there are only three other well documented cases of that tumor-type reported in the literature. The course of disease is described from initial symptoms till 10 months after discharge from hospital and suggestions for staging, therapy and follow-up are made.

Adenocarcinoma of the ileoanal pouch for ulcerative colitis--a complication of severe chronic atrophic pouchitis?

BACKGROUND: The appearance of a carcinoma in the ileal pouch after restorative proctocolectomy with ileal pouch-anal anastomosis (IPAA) for ulcerative proctocolitis is rare. Most of these adenocarcinomas previously described in literature develop from residual viable rectal mucosa. We report a case of an adenocarcinoma arising in all probability from the ileal pouch after malignant transformation of the ileal pouch mucosa based on a chronic atrophic pouchitis. PATIENT AND METHODS: A 34-year-old man developed an adenocarcinoma after a double-stapled ileorectal J-pouch for ulcerative colitis (UC) proceeded from malignant ileal transformation. Before surgery, he had a 20-year history of UC refractory to medical therapy, but no occurrence of backwash ileitis, dysplasia or colitis-associated illness. He experienced severe pouchitis after IPAA since the ileostomy closure. Carcinoma was ensured by endoscopy, and the patient underwent an abdominoperineal pouch extirpation combined with excision of perirectal tissues and anal canal. Histology after surgery showed a pT4,pN2(4/16)pM0,G3 adenocarcinoma with global severe chronic atrophic pouchitis (CAP), villous atrophy and malignant ileal transformation. No metaplasia of the rectal mucosa was found, not even malignant epithelial transformation of the anal canal. CONCLUSION: This case suggests that a malignant transformation of the ileal pouch mucosa may occur as a pure complication of severe CAP, even in the absence of backwash ileitis or a previous history of cancer. The absence of metaplasia of the rectal mucosa revealed the passage from CAP to dysplastic epithelium and to cancer. A multifactorial development of carcinogenesis is supposed, but we emphasize the importance of severe CAP, and that careful surveillance is needed in patients after IPAA. We must submit that this is just a case report and cannot stand for general cancer development in ulcerative colitis, but it may point out the risk factor of chronic inflammation and leads the surgeon to skillful working when building the pouch.

[Significance of adhesion molecules in oncology]. / Bedeutung von Adhäsionsmolekülen in der Onkologie.

Cell-Cell-and cell-extracellular matrix interactions are important in the process of tumor cell invasion and metastasis. These interactions are mediated by adhesion molecules e.g. CD44, integrins, E-cadherin and N-CAM. The role of adhesion molecules along metastatic cascade as well as their importance in tumor differentiation and progression are discussed herein.