Cerebral heterotopia appearing as an extranasal polyp.

Cerebral heterotopia represents a rare tumorlike lesion related in its development to the encephaloceles but differing from these by non-connection to the cranial cavity. We describe an unusual case of cerebral heterotopia appearing as a pedunculated polyp on the upper lip of a newborn. As the distinction between encephaloceles and cerebral heterotopias is based on the correlation of radiologic and clinical findings, the surgical treatment of tumorlike lesions of the nasal and pharyngeal region in children requires a careful preoperative evaluation of their possible connection to the cranial cavity.

Radiosensitization in sarcoma cell lines with a p53 missense mutation correlates with prevention of irradiation G2/M arrest but not with induction of apoptosis.

We analysed the effects of caffeine and taxol on the radiobiological behaviour of two human sarcoma cell lines (RD, SK-LMS-1) each with a p53 missense mutation. Treatment with 2 mM caffeine resulted in an inhibition of the irradiation induced G2/M arrest in both cell lines. This effect was coupled with a radiosensitization in cell line SK-LMS-1 after an irradiation with 6 Gy (enhancement factor of 5.0). However, the effect of radiosensitization was not correlated with an induction of apoptosis. Incubation with 20 nM taxol increased the irradiation induced apoptosis almost 3-fold in cell line SK-LMS-1, but not in cell line RD. However, taxol had no effect on the irradiation induced G2/M arrest or radiosensitivity in either cell line. The results support the hypothesis that the prevention of irradiation induced G2/M arrest but not the induction of apoptosis plays a critical role in determining radiosensitivity in sarcoma cell lines with p53 mutations.

Tumor hypoxia, p53, and prognosis in cervical cancers.

BACKGROUND: The p53 protein is involved in the regulation of initiation of apoptosis. In vitro, p53-deficient cells do not respond to hypoxia with apoptosis as do p53-normal cells, and this may lead to a relative growth advantage of cells without a functioning p53 under hypoxia. On the basis of this hypothesis, a selection of cells with a functionally inactive p53 may occur in hypoxic tumors. The development of uterine cervical carcinomas is closely associated with infections of human papilloma viruses, which may cause a degradation of the tumor suppressor gene p53, resulting in a restriction of apoptosis. Thus, cervical cancers have often a functionally inactive p53. The purpose of our clinical study was therefore to investigate the association between p53, hypoxia, and prognosis in cervical cancers in which the oxygenation status can be determined by clinical methods. MATERIAL AND METHODS: Seventy patients with locally advanced squamous cell cervical cancer Stages IIB (n = 14), IIIB (n = 49), and IVA (n = 7) were investigated in the period from 1996 through 1999. All were treated with definitive radiotherapy with curative intent by a combination of external radiotherapy plus high-dose-rate afterloading. Before therapy, tumor oxygenation was measured with a needle probe polarographically using the Eppendorf histograph. Hypoxic tumors were defined as those with pO(2) measurements below 5 mm Hg (HF5). Pretreatment biopsies were taken and analyzed immunohistologically for p53 protein expression with the DO-7 antibody. The DNA index was measured by flow cytometry. The statistical data analysis was done with SPSS 9.0 for Windows. RESULTS: The 3-year overall survival was 55% for the whole group of patients. Clinical prognostic factors in a multivariate analysis were pretreatment hemoglobin level (3-year survival 62% for patients with a pretreatment hemoglobin > or =11 g/dl vs. 27% for hemoglobin <11 g/dl, p = 0.006) and FIGO stage (Stage IIB: 65%; Stage IIIB: 60%; Stage IVA: 29%, p = 0.01). Sixty of the 70 tumors showed positive immunohistologic staining for p53 protein (transformed p53 = tp53), and 10/70 were negative (wild-type p53 = wtp53); p53 expression had no significant impact on survival (50% for tp53 vs. 79% for wtp53, p = 0.11). FIGO stage and anemia had no impact on p53 expression. Forty-nine of 70 tumors were hypoxic (HF5+), and 21 showed no hypoxia (HF5-). Hypoxic carcinomas were more frequently positive for p53 as compared to nonhypoxic tumors (27% vs. 13%, p = 0.011) and showed a trend toward a lower survival (48% vs. 70%, p = 0.07). In a further multivariate analysis, the impact of a combination of p53 expression and hypoxia on survival was examined. After adjusting for FIGO stage and pretreatment anemia, patients with wtp53 tumors had the best prognosis (3-year survival 79%) followed by tp53-HF5(-) patients (57%), and the most unfavorable prognosis was observed for tp53-HF5(+) patients (47%). The DNA index was higher in tp53 carcinomas compared to wtp53 tumors, 1.97 +/- 0.4 vs. 1.67 +/- 0.1, p = 0.05. The highest DNA index was found in hypoxic tumors with transformed p53 (2.2 +/- 3.1). CONCLUSIONS: Advanced stage and pretreatment hemoglobin level are independent prognostic factors in cervical carcinomas. The immunohistologic detection of (a functionally inactive) p53 and the presence of hypoxia had no prognostic impact, if analyzed as single parameters. However, the combination of both parameters was able to discriminate different prognostic subgroups. Moreover, hypoxic cancers were more often immunohistologically positive for tp53 protein and had a higher DNA index with the highest DNA index in tumors with both hypoxia and tp53 protein expression. These findings in summary support the theory that the tumor's microenvironment may influence the biologic behavior via hypoxia.

[Association between elevated serum VEGF and polarographically measured tumor hypoxia in head and neck carcinomas]. / Assoziation zwischen erhöhten Serum-VEGF-Werten und polarographisch gemessener Tumorhypoxie bei Patienten mit Kopf-Hals-Karzinomen.

PURPOSE: Clinical investigation of a potential relationship between VEGF concentration in serum (sVEGF) and polarographically measured tumor oxygenation in patients with squamous cell carcinoma of the head and neck (SCCHN). PATIENTS AND METHODS: In 56 patients with SCCHN we estimated the classical tumor parameters, the sVEGF concentration (immunoassay) and the tumor oxygenation (Eppendorf pO2 histograph). The platelet count and the tumor volume were evaluated simultaneously. RESULTS: In a unifactorial analysis the total volume (132 cm3 vs. 38 cm3), the hypoxic subvolume (HSV = total volume multiplied with the relative frequency of values < or = 5 mm Hg/63 cm3 vs. 10 cm3) and the platelet count (380 10(9)/l vs. 271 10(9)/l) were significantly higher in the patient group with a sVEGF level > 707 pg/ml compared to the group with a sVEGF below this threshold. The multifactorial analysis confirmed significant effects for the hypoxic subvolume and the platelet count. Regarding hypoxic subvolume and sVEGF as continuous parameters a significantly positive correlation was found. This correlation remained somewhat weaker but significant after inclusion of the platelet count as covariate. CONCLUSION: On base of our data a clinical association between elevated sVEGF and polarographically measured tumor hypoxia could be confirmed. This was possible considering not only the relative grade of hypoxia but also the absolute amount of hypoxic regions. The VEGF released from platelets during blood clotting influences the sVEGF level essentially, however, the hypoxia effect was not completely deleted. Due to the platelet effect an estimation of sVEGF is not able to substitute polarographical measurement of tumor pO2. Therefore in an ongoing study we investigate whether VEGF values estimated in plasma are better correlated with the polarographically measured tumor pO2 than serum VEGF levels.

Loss of G2/M arrest correlates with radiosensitization in two human sarcoma cell lines with mutant p53.

We have examined the modulation of radiosensitivity by using caffeine in two human sarcoma cell lines both with a p53 mutation (US8-93 and LMS6-93). In both cell lines a strong irradiation-induced G2/M arrest was coupled with a low rate of apoptosis. Incubation with caffeine resulted in a low percentage of S and G2/M cells, associated with an accumulation in G1. With a higher caffeine concentration, we detected a lower clonogenic survival with IC(50) at 2 mM. In both cell lines incubation with caffeine completely prevents the irradiation-induced G2/M arrest. This was connected to radiosensitization, but without direct correlation to an induction of apoptosis. The effect of radiosensitization rose with higher irradiation doses. However, in comparison with LMS6-93, it was stronger in cell line US8-93. A higher radiosensitization in US8-93 correlated with the prevention of strong irradiation-induced G2/M response and higher initial DNA damage. Results of Western hybridization reveal a p53-independent mechanism of radiosensitization caused by caffeine. Our findings suggest that modulation in G2/M regulation may affect a common checkpoint for tumor cells with defective p53 function. Furthermore, our results show that the enhancer effect of caffeine is dependent on a strong reduction in the number of G2/M arrested cells and on an inhibition of DNA damage repair after irradiation.

A permanent cell line of the crayfish Orconectes limosus as a potential model in comparative oncology.

A cell line, designated OLGA-PH-J/92, was established from neuronal tissue of the crayfish Orconectes limosus. To date the cell line has been subcultured more than 150 times. From the original cell line two 'daughter' cell lines and one cloned cell line were isolated. Best growth was obtained when the cells were incubated in Eagle's Minimum Essential Medium supplemented with 6% fetal bovine serum at a temperature of 27 degrees C. Under these conditions the population doubling time lasted between 23 and 25 hours. The shape of the cells is dendritic, but can change to spherical when conditions are less optimal. The cell lines showed features of transformation, such as anchorage independence, loss of contact inhibition, and low serum requirement. The number of chromosomes found in the cell lines ranged from 11 to 136, while in the donor species numbered between 98 and 106. It will be of interest to study if the O. limosus cell lines grow malignantly in vivo. OLGA-PH-J/92 and the derivative cell lines should also be suitable for studying viral infections in crustaceans.

[No association between p53 overexpression and polarographically measured tumor oxygenation in patients with head and neck carcinomas]. / Keine Assoziation zwischen p53-Uberexpression und polarographisch gemessener Tumoroxygenierung bei Patienten mit Kopf-Hals-Karzinomen.

PURPOSE: Clinical investigation of a potential relationship between the polarographically measured tumor oxygenation and the p53 status in patients with squamous cell carcinoma of the head and neck. PATIENTS AND METHODS: In 99 patients with mostly advanced, histologically proven squamous cell carcinoma of the head and neck we estimated the classical tumor parameters (TNM stage, histological grading) the immunohistochemical p53-overexpression (DO-7) and the tumor oxygenation status (Eppendorf pO2 Histograph). The tumor volume and the hemoglobin concentration were evaluated simultaneously. RESULTS: No statistically significant difference could be detected between immunohistological p53-positive (p53 > or = 10% stained cells) and p53-negative tumors (p53 < 10% stained cells) regarding both the median pO2 and the relative frequency of values < or = 5 mm Hg. Moreover, no statistically relevant differences could be seen between both p53-groups considering the hemoglobin concentration, the TNM stage, the histological grading and the tumor volume. CONCLUSION: Our data imply that there is no association between p53-overexpression and tumor hypoxia in head and neck carcinomas. However, this is not necessarily in contradiction to experimental or clinical data that confirmed a relationship between hypoxia and p53-mediated increased malignancy of tumor cells in other tumor entities. The comparable oxygenation status of p53-positive and p53-negative tumors in our study is associated with an analogous clinical tumor aggressiveness of both groups. That could be caused by a hypoxia related but p53-independent selection of tumor cells with a more malignant phenotype in head and neck carcinomas. However, further research is needed to prove this possible relationship.

Regulation of proliferation and apoptosis in sporadic and hereditary medullary thyroid carcinomas and their putative precursor lesions.

C-cell hyperplasia (CCH) and medullary thyroid carcinoma (MTC) in patients affected by germline mutations of the RET oncogene represent an exceptional opportunity to study the regulation of proliferation and apoptosis during tumour initiation and progression. In 56 specimens [CCH, n=1; MTC with CCH, n=26; MTC, n=20; lymph-node metastasis (LNM), n=9] from 46 patients [multiple endocrine neoplasia type 2a (MEN2a), n=24; MEN2b, n=2; familiar MTC (FMTC), n=4; sporadic MTC, n-16] and 3 cases of non-neoplastic CCH, proliferation activity (MIB1), the rate of apoptosis [dUTP nick end labelling (TUNEL)] and expression of p53, bcl-2, bcl-x and bax were investigated and compared with clinical data. In MEN-associated CCH and small MTC, bcl-2 was strongly expressed, bcl-x was moderately expressed and bax was only weakly expressed. Advanced tumours and LNM did show a more heterogeneous bcl-2 staining accompanied by an increased bax expression and accelerated proliferation. The rate of apoptosis was extremely low in all investigated tumours. P53 was detectable in three patients with rapidly growing and extensively metastasising MTC. No somatic p53 mutations were found. Hereditary MTC with germline RET mutations at codon 918 (MEN2b) and codon 634 revealed a bias towards a higher proliferation activity at a younger age and are more frequently accompanied by LNM. CCH and MTC are characterised with a preponderance of bcl-2 as a factor blocking the programmed cell death. While MTC, in general, is a slowly growing tumour, a minority of tumours do progress rapidly with high proliferation. The factors leading to an accelerated tumour progression do not seem to take their effect via the regulation of apoptosis. Certain alterations of RET are supposed to have a direct or indirect implication on proliferation and, because of this, an effect on the clinical course.

A multifactorial prognostic model for adult soft tissue sarcoma considering clinical, histopathological and molecular data.

Soft tissue sarcomas (STS) are malignant mesenchymal lesions with a high degree of prognostic variability. Different prognostic markers such as grading, staging, tumour type and localisation are known. The establishment of these markers was based on the evaluation at results of extensive cohorts of patients. Therefore, only the established markers provide us with information about probabilities in relation to other qualities. Considering as many different markers as possible in one prognostic statement should increase the value of the resultant information. Therefore, we developed a model involving known prognostic markers to formulate an individual prognostic index. In a retrospective analysis, different prognostic factors of 198 adult STS patients with histological tumour free resection margins were evaluated using a multifactorial analysis. On the basis of a Cox-Regression-Model with proportional hazards, the prognostic factors (tumour type, staging, localisation and type of surgical resection) were selected using previous knowledge and a statistical step backward selection procedure adjusting the immunohistochemical status of p53/Mdm2 expression. On the basis of the baseline survival function of our cohort (S0 (t)), the cumulative probability of survival for two S (2) and five S (5) years was estimated. As a result of our analysis the equations S (2) = (e-00393)P and S (5) = (e-00869)P can be used to estimate the individual two and five-year probability of survival in our cohort. Here p is the result of the amount of the estimated regression-coefficients of the exact variables of the respective individual patient. This model makes it possible to include all the evaluated prognostic factors which, in turn, increases the accuracy of the prognostic information for individual patients underlining the proportional hazards assumption.

Simultaneously occurring liver metastases of pheochromocytoma and medullary thyroid carcinoma--a diagnostic pitfall with clinical implications for patients with multiple endocrine neoplasia type 2a.

Malignant pheochromocytoma is an exceptional complication in patients with Multiple Endocrine Neoplasia Type 2a (MEN2a). In this paper, we report on a 53-year-old male patient with an evident RET gene germline mutation, who simultaneously developed hepatic metastases of medullary thyroid carcinoma (MTC) and pheochromocytoma. Comprehensive immunohistochemical investigations were performed to elaborate markers which could be useful for differentiating between MTC metastases and pheochromocytoma, respectively. Calcitonin and CEA, in particular cytokeratins and trefoil factor family 1 (TFF1), were detectable exclusively in MTC, whereas all the other markers revealed a comparable expression in both MTC and pheochromocytoma. The only clues that could indicate a potential malignant course were size, a lack of sustentacular cells and hyaline globules, and a focal spindle cell pattern in pheochromocytoma. Owing to a wide agreement in cellular differentiation and a lack of specific, routinely applicable markers for pheochromocytomas, a comprehensive and goal-directed immunohistochemistry is required to rule out pheochromocytoma metastasis in patients with MEN2a. A misinterpretation could lead to harmful clinical complications, as shown in the present case.

Glycohistochemical monitoring of chemically induced sarcomas at different stages of tumorigenesis.

Malignant soft tissue tumors still represent a source of uncertainty and controversy concerning histogenetic origin and histological behavior. Considering this, chemically induced sarcomas furnish an attractive model for the elucidation of cellular alterations during tumorigenesis. This approach allows us to closely follow cyto- and histological changes within coherent stages of tumor development. The specimen under scrutiny comprised 35 rat tissue samples from day 10 up to day 200 after benzo[a]pyrene injection. Additionally, for comparison and validation two human malignant fibrous histiocytomas (MFH) were investigated. The essential biological significance of protein-carbohydrate interactions warranted the histochemical application of synthetic tools (neo-glycoproteins-NGP) and lectins in order to reveal phenotypical dynamics in this aspect throughout the process of tumor development. Namely, 6 plant lectins (carbohydrate-binding proteins with defined saccharide specificity), 7 custom-made synthetic NGP (as the corresponding ligands visualizing endogenous lectins) and additionally three antibodies were employed. Characteristic cell populations were histochemically demonstrated in four stages of tumor development: exudation (n = 5), mesenchymal proliferation (n = 7), atypical granulation tissue (n = 7) and sarcoma (n = 16). Changes of glycohistochemical binding patterns were in close phenotypic relation to cellular activity, differentiation, local distribution as well as malignant transformation and tumor progression. At present, the new glycobiological features of the malignant phenotype substantiate the assumption that not only glycosylation but also the receptor display is altered upon carcinogenesis. In conclusion, this chronological longitudinal study takes advantage of the combination of a coherent model of tumorigenesis with innovative histochemical tools whose ligands are supposed to act as mediators of cell-cell- and cell-matrix interactions. It clearly demonstrates the suitability of the glycohistochemical method for comparative approaches. The systematic analysis of glycohistochemical determinants will improve our understanding of the early tumor biological processes with potential implications for therapeutic interventions.

[Malignant epithelioid hemangioendothelioma of the liver - a very rare tumor in children]. / Das maligne epitheloide Hämangioendotheliom der Leber. Ein sehr seltener Tumor im Kindesalter.

We report on a 12-year old boy suffering from malignant epithelioid hemangioendothelioma of the liver, which is a very rare tumor in childhood. The tumor was detected by ultrasound examination at the age of 10 and appeared at that time as a solitary intrahepatic nodular lesion. During the following 2 years multiple nodular lesions developed in both hepatic lobes. There were neither any suspect anamnestic findings nor abnormal clinical or laboratory data. The tumor showed the typical histomorphological, immunohistochemical, and ultrastructural features of this entity, which is usually seen in older patients. We investigated proliferative activity, apoptotic regulation, and expression of VEGF and VEGF-receptor flk-1 by means of immunohistochemical techniques. According to the known slow growth activity of these tumors we found only a few Ki-67 positive tumor cells. We did not detect any apoptotic cells using TUNEL technique. The positive immunoreaction of the tumor cells with antibodies against VEGF and VEGF-receptor flk-1 may indicate the regulation of tumor growth by angiogenetic factors. We present our findings together with a summary of the most important publications of recent years concerning these tumors.

Significance of proliferative activity and DNA ploidy in pancreatic cancer and chronic pancreatitis.

BACKGROUND: Precise preoperative assessment of diagnosis and prognosis in patients with pancreatic tumors would facilitate improvement of treatment strategies. In this context, we evaluated the significance of the proliferative index and of static DNA cytophotometry in the diagnosis and prognosis of pancreatic tumors. METHODS: Consecutive surgical specimens from 26 patients with ductal pancreatic cancers and eight patients with chronic pancreatitis were investigated by: 1. Staging; 2. Conventional histological and cytological grading; 3. MIB-1 (Ki-67 labeling) proliferating index; and 4. Static DNA cytophotometry. RESULTS: All patients with chronic pancreatitis had a normal MIB-1 labeling index and a euploid DNA content. In contrast, patients with pancreatic cancers rarely had a normal labeling index (1 of 26 patients) or a euploid DNA content (6 of 26 patients). Staging significantly correlated with survival time. However, it did not correlate with cytological criteria. Cytological criteria, such as conventional grading, MIB-1 proliferating index, and DNA ploidy, were not significantly correlated with survival time. Conventional grading was significantly correlated (p < 0.02) with proliferating index, but not with DNA ploidy. CONCLUSION: Proliferating index and DNA ploidy are relevant cytological markers that can help to discriminate between chronic pancreatitis and pancreatic cancer. The prognostic significance of these markers in pancreatic cancer patients, however, seems to be less relevant than tumor stage and of limited relevance for the individual cancer patient.

Presarcomatous lesions of experimentally induced sarcomas in rats: morphologic, histochemical, and immunohistochemical features.

Morphological studies have been performed mainly on manifest sarcomas, leading to divergent views of its histogenesis. However, histogenesis requires understanding of the tumor precursor cells and cannot be resolved by static morphologic studies. Defining presarcomatous lesions is made more difficult because they do not possess a basement membrane that serves as biologic and nosologic boundary like in epithelial cancers. The present study, therefore, investigated the early phases of experimentally induced rat sarcoma, which closely resembles human malignant fibrous histiocytoma (MFH). Histological, enzyme- and immunohistochemical methods were used to define the sequential events involved in tumorigenesis. A benzo[a]pyrene-oil mixture was injected intramuscularly into the thighs of rats, producing MFH 120 days later. Groups of animals were sacrificed every 10 days. The injections produced soft tissue lesions characterized by three distinct phases that overlapped or existed simultaneously in one animal: an initial acute inflammatory reaction characterized by an infiltration of lymphocytes, monocytes and macrophages, a second mesenchymal fibromatous phase characterized by the predominance of spindle-shaped, focally atypical fibroblast-like cells and collagen, and a third premalignant neovascularization phase characterized by dominant capillary proliferations. Overt MFH developed 120 days after injection and consisted of spindle-shaped fibroblast- and histiocyte-like cells containing atypical mitoses and arranged in a storiform pattern. Control animals injected with olive oil revealed acute inflammatory reactions after 30 days and no signs of chronic inflammation or malignancy after 60 and 120 days. We concluded that these experimentally-induced rat sarcomas develop in a triphasic pattern that resembles non-healing granulation tissue, with neovascularization preceding the occurrence of overt MFH.

G2/M checkpoint is p53-dependent and independent after irradiation in five human sarcoma cell lines.

To determine the role of p53 in G2/M arrest, G2/M transition and apoptosis, we investigated five human sarcoma cell lines with different p53 gene status in their response to X-rays. The p53 status of the cell lines was mutant (US 8-93, LMS 6-93 and RD), null (SAOS-2) and wildtype (A-204). Clonogenic survival of the cell lines varied as the survival fraction at 2 Gy (SF2) ranged from 0.28 to 0.79. Compared with the mutated p53 cell lines (SF2 with a range from 0.46 to 0.79) the clonogenic survival of the wildtype p53 (wt-p53) cell line A-204 (SF2 = 0.34) was lower. The p53 null cell line (SAOS-2) was also sensitive to X-rays (SF2 = 0.28). We detected, in all cell lines a similar behavior in their response to irradiation with G2/M arrest and apoptosis. However, the maximal rate of apoptosis with a range from 7.0 to 18.0% was rather small. The decrease of G2/M cells was coupled with an increased percentage of apoptotic cells. However, a different delay in G2/M did not result in a change of radiation sensitivity. Western analyses showed an increased P53 level only for the cell line A-204 (wt-p53) after irradiation. Our results point out that there is not always a simple relationship between p53 gene status and radiation sensitivity. We suggest, that wt-p53 plays an active role in G2/M arrest and in decreasing the number of G2/M cells as a response to apoptosis. Therefore, p53-dependent regulation in G2/M may be as important as p53-independent mechanisms.

Clinical relevance of pRb and p53 co-overexpression in soft tissue sarcomas.

The goal of this study was to examine the relationship between immunohistochemical pRb detectability and p53 overexpression in 198 soft tissue sarcomas (STS) with regard to its clinical relevance. Distinct pRb detectability multivariately shows a correlation to survival rate (relative risk (RR)=1.59, P=0.037). p53 positivity was also multivariately correlated to poor prognosis (RR=2.17, P=0.0014). Stratification of pRb staining to p53 results shows a prognostical graduation. Patients with negativity for both proteins have the most favorable prognosis (projected 5-year survival rate (psr)=54.5%). In contrast to this, positivity for both antibodies has the highest risk (RR=2.48, P=0.02) and the poorest prognosis (psr=17.4%). To conclude, these results explain that the clinical relevance of immunohistochemical pRb positivity in STS is connected with p53 in the form of having an increasing effect on the known prognostic relevance of p53 overexpression.

Assessment of genomic imbalances in malignant fibrous histiocytomas by comparative genomic hybridization.

In order to investigate genomic imbalances, comparative genomic hybridization was applied to 20 malignant fibrous histiocytomas. Deletions were rare and found mainly in chromosomes 2q33-35, 4q32-qter, 8p, 9p21-pter, 12p and 19p, whereas, over-representations frequently affected chromosomes 3, 4q31, 5p, 6, 7, 14q22-ter, 18p, as well as, five distinct amplifications within the regions 12q12-15 and 15q24-qter. The total number of genetic imbalances per tumor was slightly increased in primary tumors when compared to relapses. No relationship was found between the patterns of gain and loss when compared to the histological subtype, tumor grading, the clinical outcome and the p53 mutation status.

Primary hereditary medullary thyroid carcinoma--C-cell morphology and correlation with preoperative calcitonin levels.

Early thyroidectomy offers an opportunity of preventing the development of medullary thyroid carcinoma (MTC) in patients at risk for hereditary MTC. We investigated the thyroid glands of 32 patients with hereditary MTC to identify the changes in C-cell morphology and to correlate these with plasma calcitonin (CT) levels and with clinical data. The entire thyroid gland was processed for histological examination including immunostaining for CT. All glands revealed C-cell hyperplasia (CCH), and MTC was found in 21 patients (66% of 32, youngest patient 6 years, youngest with lymph node metastases [LNM] 17 years). The transition from CCH to MTC was characterized by destruction of the follicular basement membrane and by diminished intensity of CT immunostaining. Normal plasma CT levels after provocation with pentagastrin were found only in patients with CCH. Basally elevated plasma CT levels were restricted to MTC. LNM were only found in multifocal tumours at least 4 mm in diameter. It is not yet clear whether or not CCH in patients at risk for hereditary MTC is a neoplastic change, but in these patients the term 'C-cell hyperplasia' is of doubtful value. All MEN gene carriers reveal CCH, and almost all of them will develop multifocal MTC, so that CCH is probably a precursor lesion of an indubitably malignant tumour. Prophylactic thyroidectomy is justified at the age of 6 to anticipate development of a MTC. Lymphadenectomy is necessary in children if they are older than 10 years or have elevated plasma CT levels.

p53 status in radiation-induced soft-tissue sarcomas.

BACKGROUND: Following therapeutic irradiation after a latency period of many years radiation-induced tumors, often sarcomas, can arise. Results of radiation-induced DNA damage can be 1. p53 over-expression, inducing growth arrest or apoptosis, and 2. occurrence of mutations, frequently including the p53 gene, as one molecular promotor for carcinogenesis. We were interested whether radiation-induced sarcomas are associated with alterations of the p53 status. MATERIAL AND METHODS: Samples from 11 radiation-induced soft-tissue sarcomas (STS) were studied by a non-radioactive PCR-SSCP sequencing analysis and by immunohistochemistry with five antibodies for their p53 status. RESULTS: A tumor of one patient possessed a G->A transition in codon 280 (exon 8). Of 11 tumors, 9 showed nuclear p53 positivity, detected by monoclonal antibody DO-1. Of these 9 patients, 7 died during the observation period, whereas the 2 patients with DO-1 negative tumor samples are still alive. CONCLUSIONS: p53 over-expression and p53 mutation occur in radiation-induced STS. p53 status is expected to have prognostic impact for radiation-induced STS.

Molecular characterization and liposomal transfection of a p53-mutated cell line established from a poorly differentiated leiomyosarcoma.

A human cell line LMS6-93 has been established from a leiomyosarcoma (LMS). Characteristics for ultrastructure, growth characteristics, cell cycle distribution, karyotype, protein expression detected by immunohistochemistry (IHC), p53 mutational status and liposomal transfection behaviour were studied and determined. The primary tumor was clearly positive for á-smooth muscle type actin and desmin in moderately differentiated areas and indicated a loss of myogenic differentiation in other regions and therefore was classified as a poorly differentiated LMS. The cell line LMS6-93 contains mainly polymorphic spindle shaped or polygonal tumor cells which possess the characteristics of primitive mesenchymal cells, based on their morphology and positive reaction with an antibody to vimentin. IHC staining for S100, synaptophysin A, NSE, neurofilament proteins and cytokeratins were negative. Cytogenetic analysis revealed in the cell line diploid karyotypes comparatively close to several structural and numerical aberrations for chromosomes 2, 5, 6, 9, 10, 12, 14, 17, 18, 20, 22, and Y. IHC positivity was found for the tumor suppressor protein Rb and the oncogene product MDM2. In a p53 mutational analysis a 1 bp insertional mutation in exon 6 (G insertion in codon 215) was detected and confirmed in the original primary tumor. The other p53 allele appears to be wild-type as indicated in Western hybridization. Using different cationic lipid formulations complexed with a reporter expression vector (GFP) successful transfection into LMS6-93 cells was observed. The highest transfection rates (20-30% GFP expression in the viable cell population) were obtained with lipofectin. These results suggest that LMS6-93 functions as a good in vitro model for transfection studies on an LMS cell line carrying a heterozygous p53-frameshift mutation.