DNA repair protein MGMT protects against N-methyl-N-nitrosourea-induced conversion of benign into malignant tumors.

Tumor formation is a multi-step process that can be divided into the stages of tumor initiation, promotion and progression. Previously, we showed that overexpression in skin of mice of the DNA repair protein O(6)-methylguanine-DNA methyltransferase (MGMT) protects against N-methyl-N-nitrosourea (MNU)-induced tumor initiation without affecting tumor promotion. This indicated that O(6)-methylguanine, which is specifically repaired by MGMT, is a major tumor-initiating lesion.

Preclinical study on gene therapy of cervical carcinoma using adeno-associated virus vectors.

Approximately 90% of cervical carcinomas are causally linked to infections with high-risk human papillomaviruses (HPVs), whose oncogenicity has been assigned to the continued expression of two early genes, E6 and E7. Reversal of the transformed phenotype by inhibiting E6/E7 gene expression therefore provides a suitable goal for future tumor therapy. Using recombinant adeno-associated virus type 2 (AAV-2) vectors, two types of therapeutic genes were expressed in cervical carcinoma cells with the aim of suppressing the E6/E7 oncogenes: (a) antisense E6/E7 and ribozyme genes and (b) the monocyte chemoattractant protein-1 (MCP-1) gene encoding MCP-1.

Caffeine-derived N-nitroso compounds. V. Carcinogenicity of mononitrosocaffeidine and dinitrosocaffeidine in bd-ix rats.

Mononitrosocaffeidine (MNC) and dinitrosocaffeidine (DNC) are new N-nitroso compounds obtained from in vitro nitrosation of caffeidine, a hydrolysis product of caffeine present in a typically made and widely consumed tea from Kashmir (India), a high incidence area of esophageal and stomach cancer. The chemical synthesis, in vitro metabolic studies and mutagenicity of the compounds has been previously reported. DNC, a nitrosamide is highly mutagenic both with and without metabolic activation whereas MNC, like several other aromatic asymmetric nitrosamines, does not exhibit genotoxic or mutagenic properties.

Using gene carrier probability to select high risk families for identifying germline mutations in breast cancer susceptibility genes.

Germline mutations in highly penetrant autosomal dominant genes explain about 5% of all breast cancer, and heritable mutations in the BRCA1 breast and ovarian cancer susceptibility gene account for 2-3% of breast cancer in the general population. Nevertheless, the presence of such mutations is highly predictive of disease development. Since screening for mutations is still technically laborious, we investigated whether the prior probability of being a carrier of a dominant breast cancer susceptibility gene in the youngest affected family member could be used to identify families in which the probability of finding a mutation is sufficiently high.

The expression of the imprinted genes H19 and IGF-2 in choriocarcinoma cell lines. Is H19 a tumor suppressor gene?

H19 is a paternally imprinted gene with unknown function. It is located in close proximity to the maternally imprinted IGF-2 gene on chromosome 11p15.5.

The product of the imprinted H19 gene is an oncofetal RNA.

Aims/background: The H19 gene is an imprinted, maternally expressed gene in humans. It is tightly linked and coregulated with the imprinted, paternally expressed gene of insulin-like growth factor 2. The H19 gene product is not translated into protein and functions as an RNA molecule.

Epitope mapping and functional characterization of monoclonal antibodies specific for herpes simplex virus type I DNA polymerase.

Three monoclonal antibodies (MAbs 1051a, 1051b and 1051c) were raised against a surface region (residues 597-686) of the herpes simplex virus type 1 (HSV-1) DNA polymerase (HSV pol), and their epitopes were mapped. The MAbs reacted serotype specifically with the native and denatured HSV pol, as shown by Western blot analysis, immunoprecipitation and immunofluorescence microscopy, indicating their usefulness for biochemical studies and clinical diagnosis of HSV-1 infections, MAb 1051c, displaying the least cross-reactivity with cellular proteins in the Western blot analysis, was successfully utilized not only for coimmunoprecipitation, but also for the analysis and three-dimensional modeling of the cellular sites of HSV pol interaction by confocal laser immunofluorescence microscopy.

Latent chromosomal instability in cancer patients.

There is increasing evidence that, similar to what is found with other genetic disorders, genomic instability is one of the most general features of cancer. Different forms of manifestation including latent instability have been suggested. To recognize latent chromosomal instability we treated lymphocyte cultures of cancer patients and healthy persons with caffeine, two different doses of bleomycin, and a combination of bleomycin and caffeine.

Morphologic characteristics of the interaction between normal cytotrophoblasts and their malignant counterpart in the development of trophoblastic neoplasia.

Objectives: To define the biology of the tumor-host cell interaction with regard to cellular kinetics and morphologic changes during cell-cell interaction in an in vitro model of trophoblastic neoplasia.

Methods: Using a coculture in vitro system of cytotrophoblasts and choriocarcinoma cells, we investigated the cellular kinetics and the morphologic changes in these interacting cells. A fully automatic time-lapse image system was used to record phase contrast images of the cocultured cells in a tissue culture chamber.

Developmentally imprinted genes as markers for bladder tumor progression.

Purpose: Developmentally imprinted genes, such as H19 and insulin-like growth factor-II (IGF-II), play an important role during human embryogenesis and also have been implicated in the pathogenesis of embryonal tumors of childhood. Since H19 is expressed in human fetal bladder, we evaluated 35 bladder carcinomas for H19 expression by in situ hybridization analysis and correlated expression with tumor grade. As a prelude to gene transfer studies to determine if H19 is a bladder tumor oncogene, we also evaluated bladder cell lines for expression of H19, IGF-II, IGF-I and the type I IGF receptor.

Allelic imbalance on chromosome 13q: evidence for the involvement of BRCA2 and RB1 in sporadic breast cancer.

Recently, the breast cancer susceptibility gene BRCA2 has been identified in chromosome 13q, a region that also contains the retinoblastoma gene RB1. To elucidate a possible role of BRCA2 and RB1 in sporadic breast tumorigenesis, allelic imbalance (AI) at 13q loci was examined in 78 primary sporadic breast tumors. AI was found in 52-63% of tumors.

The effect of the JE (MCP-1) gene, which encodes monocyte chemoattractant protein-1, on the growth of HeLa cells and derived somatic-cell hybrids in nude mice.

To investigate the effect of tumor-associated macrophages on the in vivo growth properties of cervical carcinoma cells, tumorigenic human papilloma virus (HPV) 18-positive HeLa cells were transfected with an expression vector harboring the cDNA for the macrophage chemoattractant protein-1 JE (MCP-1). Although the endogenous gene is present and not structurally rearranged, its expression seems to be negatively affected by a still unknown mechanism. Inoculation of JE (MCP-1)-negative HeLa cells into nude mice led to rapidly growing tumors, where macrophage infiltration into the inner tumor mass was not detectable immunohistochemically.

Trisomy 8 and urogenital malformation in the ACI-rat.

ACI-rats are considered as a model for studying urogenital abnormalities. In order to recognise cytogenetic changes related to these abnormalities 50 male ACI/Seg rats were examined by means of gross macroscopic, histological, and karyotypical investigations. In six of the examined animals (12%) unilateral agenesis of the kidney and ipsilateral hypoplasia of the testes and seminal vesicles were observed.

Evaluation of nuclear morphology in adriamycin sensitive and resistant cells.

The adriamycin sensitive and resistant human breast cancer cells (MCF-7) were investigated by computerized image analysis for establishment of characteristic cell nuclei-morphological differences. Using specially developed algorithms and a large set of subvisual parameters, morphological features characteristic for the resistant cell line were described. The most useful parameters for distinguishing the resistant from sensitive cells are related to chromatin structure and include integral optical density per micron 2, the average area of chromatin region, and the integral density of the chromatin region per micron 2.

Genetic alterations in sporadic renal-cell carcinoma: molecular analyses of tumor suppressor gene harboring chromosomal regions 3p, 5q, and 17p.

On a genetic level, renal-cell carcinoma has been characterized by an abnormality on the short arm of chromosome 3 (3p), which suggests the inactivation of a tumor suppressor gene. One tumor suppressor gene at 3p, the von Hippel-Lindau disease gene, is implicated in tumor development of a whole spectrum of hereditary neoplasms, including renal-cell carcinoma. It is not clear whether the same tumor suppressor gene accounts for all, i.

Subclinical human immunodeficiency virus-related changes in oral mucosa shown by image analysis of tongue smears.

There is evidence that certain lesions of the oral mucosa, such as hairy leukoplakia (HL), in patients seropositive for human immunodeficiency virus correlate with the subsequent development of acquired immune deficiency syndrome. The authors suggest that HL is a final manifestation of alterations that gradually develop after HIV infection. To recognize inapparent early subclinical changes in oral mucosa, the authors applied methods of digital image analysis to investigate tongue smears from healthy control subjects and immunosuppressed patients after chemotherapy and HIV infection.

Chromatin organization and breast cancer prognosis. Two-dimensional and three-dimensional image analysis.

Background: The assessment of breast cancer prognosis still relies on clinical staging and histologic grading. Among the different prognostic parameters, attention has been focused on features of chromatin structure. Although data from two-dimensional (2-D) analysis of chromatin structure show significant correlation with the survival, their biologic interpretation remains difficult.

Steroidal interactions in the ageing endocrine system: absence of suppression and pathology in reproductive systems of old males from a mixed-sex socially stressful rat colony.

A paradigm using chronic social stress and multiple measures of the reproductive system were used to assess changes with ageing in the dynamics of endogenous steroid interactions. The 22- to 24-month-old male rats lived for 8 weeks in one of four types of colony, in groups of the same sex or groups of mixed sex including familiar or unfamiliar old males. Measures of endocrinology (circulating steroid levels), behaviour (exploration and sociosexual responses), physiology (body and organ weights and epididymal sperm count) and histology (adrenal and ventral prostate glands) served as markers of activation of the hypothalamic-pituitary-adrenal (HPA) or hypothalamic-pituitary-testicular (HPT) axes.

Prevention of tumor metastasis formation by anti-variant CD44.

A splice variant of CD44 (CD44v) originally discovered on metastases of a rat pancreatic adenocarcinoma (BSp73ASML) has been shown by transfection to confer metastatic behavior to nonmetastatic tumor cells (Günthert U., M. Hofmann, W.

Growth of Hodgkin cell lines in severely combined immunodeficient mice.

No animal model exists for the in vivo growth of Hodgkin's-lymphoma-derived cells. Neither unmanipulated Hodgkin's-disease(HD)-derived cell lines nor primary biopsy tissue could be grown in nude mice. Since the severe combined immunodeficient (SCID) mouse has been reported to be a better recipient for transplanted human lymphatic tissue than the nude mouse, we tested whether SCID mice provide suitable conditions for the in vivo growth of HD cell lines.

A tilting device for three-dimensional microscopy: application to in situ imaging of interphase cell nuclei.

The resolution of an optical microscope is considerably less in the direction of the optical axis (z) than in the focal plane (x-y plane). This is true of conventional as well as confocal microscopes. For quantitative microscopy, for instance studies of the three-dimensional (3-D) organization of chromosomes in human interphase cell nuclei, the 3-D image must be reconstructed by a point spread function or an optical transfer function with careful consideration of the properties of the imaging system.

Three-dimensional analysis of cell nucleus structures visualized by confocal scanning laser microscopy.

Studies of the three-dimensional (3-D) organization of cell nuclei are becoming increasingly important for the understanding of basic cellular events such as growth and differentiation. Modern methods of molecular biology, including in situ hybridization and immunofluorescence, allow the visualization of specific nuclear structures and the study of spatial arrangements of chromosome domains in interphase nuclei. Specific methods for labelling nuclear structures are used to develop computerized techniques for the automated analysis of the 3-D organization of cell nuclei.

Regressing nude mouse grafts of Burkitt's lymphoma x lymphoblastoid cell hybrids show deregulation of the c-myc gene and expression of the EBV latent membrane protein.

Somatic cell hybrids between the malignant Burkitt lymphoma cell line BL 60 and the non-malignant Epstein-Barr virus (EBV) immortalized lymphoblastoid cell line (LCL) IARC 277 demonstrate the deregulated c-myc transcription pattern of the parental BL cell line during exponential growth in tissue culture. Subcutaneous nude mouse grafts of these hybrids, however, completely regress after an initial growth phase. To investigate whether regression of these grafts is mediated by a down-regulation of the BL-60-derived deregulated c-myc gene in vivo, c-myc transcription was analyzed in growing versus regressing hybrid grafts.

Quantitative features of chromatin structure in the prognosis of breast cancer.

In prognosis of breast cancer different parameters are in current use. Along with clinical staging the most important parameter appears to be histologic grading. Features of the grading such as nuclear pleomorphism proved to correlate closely with the proliferative activity and aggressiveness of the tumors.