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C Paraskeva

Publications and source records attributed to C Paraskeva.

At least 91 records · Page 5Linked to original sources
PubMed1993

Molecular events including p53 and k-ras alterations in the in vitro progression of a human colorectal adenoma cell line to an adenocarcinoma.

The aim of the current study was to identify genetic abnormalities in human colorectal adenoma and carcinoma derived cell lines, and to determine whether the genetic changes which occur in vitro are relevant to the in vivo situation. Loss of 1p(33-35) region was shown to be the most common chromosome 1 abnormality and loss of heterozygosity (LOH) of the DCC gene and/or adjacent sequences was detected in all adenoma derived cells as well as the carcinoma cell lines. The level of p53 protein was also investigated as increased cellular p53 protein had previously been associated with mutation of the p53 gene. A further aim was to investigate genetic changes in our in vitro model of tumour progression, where the adenoma derived PC/AA cell line has previously been converted in vitro to two distinct tumorigenic phenotypes, producing either an adenocarcinoma or a mucinous carcinoma in athymic nude mice. Progression to the adenocarcinoma phenotype was shown to involve a specific chromosome 1 rearrangement, loss of both normal copies of chromosome 18 (although DCC gene sequences were retained), loss of the remaining wild type allele of k-ras resulting in homozygosity for the k-ras codon 12 mutation and increased cellular p53 protein as detected by SDS-PAGE Western blotting. The increase in p53 protein was shown not to be due to the acquisition of a mutation in the p53 gene. Interestingly, progression of the adenoma derived PC/AA cell line to the mucinous malignant phenotype did not involve any of these molecular rearrangements, suggesting that different genetically distinct pathways are involved in colorectal carcinogenesis. These studies show that the genetic changes in our in vitro model of human colorectal tumour progression are similar to those observed in in vivo studies.

Adenocarcinoma↗
PubMed1992

Specific cytogenetic abnormalities and k-ras mutation in two new human colorectal-adenoma-derived cell lines.

Two new human epithelial cell lines from sporadic colorectal adenomas designated S/RR and S/BR are reported. Both cell lines have extended growth capacities in vitro, reaching passages 38 and 40 respectively and show no sign of senescence. S/RR and S/BR cell lines have retained the ability to differentiate in vitro, as shown by mucin production from goblet-like cells. S/BR was derived from a large colonic tubular villous adenoma (3 to 4 cm), whereas S/RR was derived from a small rectal adenoma (< 1 cm), and may represent a relatively early-stage adenoma. The parent S/RR cell line has given rise to a clonogenic variant, designated S/RR/Cl, which also has shown no sign of senescence and has currently reached passage 43. Both the S/BR and the S/RR cell lines had mutations in codon 12 of the K-ras gene, while retaining one normal allele. The presence of this mutation, particularly in the cell line S/RR derived from a small adenoma, is consistent with ras mutation being a relatively early event in colorectal carcinogenesis and is perhaps involved in the ability of the adenoma cells to progress and to give rise to an immortal cell line in vitro. The clonal derivatives of the S/RR cells have an isochromosome 1q and abnormalities of chromosome 13 which include an isochromosome 13q. The S/BR cells have a deletion on the short arm of chromosome 1 and trisomy 7. The common abnormality for S/RR and S/BR cells involves chromosome 1. The involvement of different chromosomes in the 2 cell lines also suggests different pathways for malignant progression of the premalignant adenoma cells.

Adenoma↗
PubMed1992

A single human colonic adenoma cell line can be converted in vitro to both a colorectal adenocarcinoma and a mucinous carcinoma.

In a previous study, using a chemical carcinogen, we converted in vitro a non-tumorigenic cell line derived from a human colorectal diploid adenoma, designated PC/AA, into a tumorigenic cell line which, when inoculated into athymic nude mice, produced progressively growing adenocarcinomas. We now report that continuous in vitro passage of the PC/AA adenoma cell line resulted in its spontaneous transformation to a mucinous carcinoma with a modal karyotype of 51, XY, +i(Iq), +8, +9, +13, +i(13q), -21, +mar. These studies show that a single adenoma can be converted along 2 independent pathways, giving rise to either a mucinous carcinoma or an adenocarcinoma, and provide further experimental evidence for the adenoma-carcinoma sequence. Cytogenetic changes which occur along both pathways to tumorigenicity include abnormalities of chromosome I and multiple copies of chromosome 13. These abnormalities may be important in tumour development and progression in colorectal carcinogenesis.

Adenocarcinoma↗
PubMed1992

Cell and molecular biology of gastrointestinal tract cancer.

The gene for familial adenomatous polyposis coli (APC or FAP), which has previously been linked to chromosome 5q21 has been identified. The APC gene has been found to be altered by point mutations in the germ line of both adenomatous polyposis coli and Gardner's syndrome patients and somatically in tumors from sporadic colorectal cancer patients. During the hunt for the APC gene, the closely linked MCC (mutated in colorectal cancer) gene was identified and found to be altered somatically in tumors from sporadic cancer patients. These data suggest that more than one gene on chromosome 5q21 may contribute to colorectal carcinogenesis and that mutations at the APC gene can cause both adenomatous polyposis coli and Gardner's syndrome. The identification of these genes should aid in the counseling of patients with genetic predispositions to colorectal cancer. Progress has also been made in identifying specific genetic changes that occur in other gastrointestinal cancers. A mutational "hotspot" in the p53 gene in human hepatocellular carcinomas has been identified that could reflect exposure to a specific carcinogen, one candidate being aflatoxin B1.

Adenomatous Polyposis Coli↗
PubMed1992

Progression of colorectal cancer is associated with multiple tumor suppressor gene defects but inhibition of tumorigenicity is accomplished by correction of any single defect via chromosome transfer.

Carcinogenesis is a multistage process that has been characterized both by the activation of cellular oncogenes and by the loss of function of tumor suppressor genes. Colorectal cancer has been associated with the activation of ras oncogenes and with the deletion of multiple chromosomal regions including chromosomes 5q, 17p, and 18q. Such chromosome loss is often suggestive of the deletion or loss of function of tumor suppressor genes. The candidate tumor suppressor genes from these regions are, respectively, MCC and/or APC, p53, and DCC. In order to further our understanding of the molecular and genetic mechanisms involved in tumor progression and, thereby, of normal cell growth, it is important to determine whether defects in one or more of these loci contribute functionally in the progression to malignancy in colorectal cancer and whether correction of any of these defects restores normal growth control in vitro and in vivo. To address this question, we have utilized the technique of microcell-mediated chromosome transfer to introduce normal human chromosomes 5, 17, and 18 individually into recipient colorectal cancer cells. Additionally, chromosome 15 was introduced into SW480 cells as an irrelevant control chromosome. While the introduction of chromosome 17 into the tumorigenic colorectal cell line SW480 yielded no viable clones, cell lines were established after the introduction of chromosomes 15, 5, and 18. Hybrids containing chromosome 18 are morphologically similar to the parental line, whereas those containing chromosome 5 are morphologically distinct from the parental cell line, being small, polygonal, and tightly packed. SW480-chromosome 5 hybrids are strongly suppressed for tumorigenicity, while SW480-chromosome 18 hybrids produce slowly growing tumors in some of the animals injected. Hybrids containing the introduced chromosome 18 but was significantly reduced in several of the tumor reconstitute cell lines. Introduction of chromosome 5 had little to no effect on responsiveness, whereas transfer ot chromosome 18 restored responsiveness to some degree. Our findings indicate that while multiple defects in tumor suppressor genes seem to be required for progression to the malignant state in colorectal cancer, correction of only a single defect can have significant effects in vivo and/or in vitro.

Alleles↗
PubMed1991

Differential sensitivity of human colonic adenoma and carcinoma cells to transforming growth factor beta (TGF-beta): conversion of an adenoma cell line to a tumorigenic phenotype is accompanied by a reduced response to the inhibitory effects of TGF-beta.

The growth of three non-tumorigenic human colonic adenoma cell lines, designated AA/C1, RG/C2 and RR/C1, was inhibited by low concentrations of transforming growth factor beta (TGF-beta) (0.05-0.5 ng ml-1). However, the growth of five human colon cancer cell lines under identical conditions was resistant to high concentrations of TGF-beta (2-10 ng ml-1). This is the first report of well-characterized premalignant human colonic cells showing sensitivity to TGF-beta. The TGF-beta-sensitive adenoma cell line AA/C1 was derived from a relatively large adenoma with a K-ras gene mutation and represents a relatively late-stage adenoma, indicating that loss of response to TGF-beta occurs at a relatively late stage in colorectal carcinogenesis and that the presence of a ras gene mutation does not necessarily confer resistance to TGF-beta. Of further interest, the RG/CZ cell line has a p53 mutation showing that p53 mutations do not necessarily lead to TGF-B insensitivity. Furthermore, in this paper we show that the conversion of the AA/C1 adenoma cell line to a tumorigenic phenotype [Williams et al., (1990) Cancer Res., 50, 4724] is accompanied by a reduced response to the growth-inhibitory effects of TGF-beta up to 10 ng ml-1. Reduced responsiveness to the inhibitory effects of TGF-beta may be an important event in the loss of growth control in colorectal carcinogenesis.

Adenoma↗
PubMed1990

Characterization of a sialic-acid-rich mucus glycoprotein secreted by a premalignant human colorectal adenoma cell line.

The human colonic cell line PC/AA, derived from an adenoma, retains in vitro colonic cell differentiation, notably the production of mucus glycoproteins. The PC/AA adenoma cells produce an extracellular gel layer in culture. The PC/AA gel could be isolated by extraction of the cell cultures with guanidine hydrochloride. The extracted material was purified by gel filtration and caesium chloride density-gradient centrifugation and showed properties typical of mucus glycoproteins, namely, a carbohydrate content above 60% of dry weight rich in N-acetylgalactosamine and sialic acid and low in mannose; an amino acid composition with high serine threonine and proline content; a molecular weight above 1,000 kDa on Sepharose CL 4B chromatography and on SDS-polyacrylamide gel electrophoresis under reducing conditions (greater than 200 kDa); a buoyant density of approximately 1.48 g/ml and the release of oligosaccharides by the alkaline beta-elimination reaction. Comparison of the gel mucus glycoprotein purified from premalignant PC/AA cells with normal human colon mucin showed that it has a higher sialic acid content. This suggests that higher sialic acid levels may precede the development of malignancy.

Adenoma↗
PubMed1990

p53 gene mutations occur in combination with 17p allelic deletions as late events in colorectal tumorigenesis.

Coordinate loss of one copy of the p53 gene and mutation of the remaining copy occur in colorectal carcinomas and in many other human malignancies. However, the prevalence of p53 gene mutations in carcinomas which maintain both parental copies of p53 has not previously been evaluated. Moreover, it is not known whether p53 gene mutations are limited to malignant tumors or whether they can also occur in benign neoplasms. To answer these questions, a total of 58 colorectal tumors have been examined; in each tumor, allelic losses were assessed using restriction fragment length polymorphisms and p53 gene mutations were assessed by sequencing cloned polymerase chain reaction products. The following conclusions emerged: (a) p53 gene mutations occurred but were relatively rare in adenomas, regardless of size and whether the adenomas were derived from patients with familial adenomatous polyposis; (b) In carcinomas as well as in adenomas, p53 gene mutations were infrequently observed in tumors which contain both copies of chromosome 17p (17% of 30 tumors), while tumors which lost one copy of chromosome 17p usually had a mutation in the remaining p53 allele (86% of 28 tumors); (c) p53 gene mutations were found at similar frequencies in primary tumor samples and in cell lines derived from tumors. These and other data suggest that the rate limiting step in p53 inactivation is point mutation and that once a mutation occurs, loss of the remaining wild-type allele rapidly follows. Both mutations and allelic losses generally occur near the transition from benign to malignant growth, and the p53 gene may play a causal role in this progression.

Adenoma↗
PubMed1990

Neoplastic transformation of a human colonic epithelial cell line: in vitro evidence for the adenoma to carcinoma sequence.

The purpose of this study was to establish an in vitro model for tumor progression in colorectal carcinogenesis, by transforming the premalignant human colonic PC/AA adenoma cell line to the malignant phenotype. A rare clonogenic variant AA/C1 [colony-forming efficiency (CFE) on plastic of 1.05%] was isolated from the diploid PC/AA adenoma cell line (C. Paraskeva, S. Finerty, and S. Powell, Int. J. Cancer, 41: 908-912, 1988). AA/C1 was aneuploid and when treated with 1 mM sodium butyrate for 14 days gave rise to the AA/C1/SB cell line which had an increased CFE on plastic (6.13%) although the cells remained anchorage dependent and nontumorigenic. After exposure of these AA/C1/SB cells to the carcinogen N-methyl-N'-nitro-N'-nitrosoguanidine an anchorage-independent cell line was isolated (AA/C1/SB10). On continuous in vitro passage, the CFE in agarose of AA/C1/SB10 has increased to 17.3% and the cells have become tumorigenic producing adenocarcinomas in athymic nude mice. AA/C1, AA/C1/SB, and AA/C1/SB10 cell lines have common chromosomal abnormalities including a pericentric inversion of chromosome 1 with deletion of part of the short arm and monosomy for chromosome 18. This in vitro progression provides the first reported experimental evidence for the adenoma to carcinoma sequence in the human colon, and the cytogenetic evidence suggests that it is relevant to in vivo carcinogenesis.

Adenoma↗
PubMed1989

Possible involvement of chromosome 1 in in vitro immortalization: evidence from progression of a human adenoma-derived cell line in vitro.

We have previously reported that continuous in vitro passage in the presence of 3T3 feeders of a non-tumorigenic adenoma-derived epithelial cell line, designated PC/AA, resulted in its becoming immortal. At early passage PC/AA was normal diploid, whereas every cell of PC/AA late passage had an isochromosome 1(q) which led us to suggest that abnormalities of chromosome 1 may be involved in tumour progression. We now report the isolation of a 3T3-feeder-independent variant of early-passage PC/AA, designated PC/AA/FI, which was immortal in vitro and remained non-tumorigenic. Each cell of PC/AA/FI again has an isochromosome 1(q), like the late-passage PC/AA. However, with PC/AA/FI it is the other chromosome 1 of the homologous pair which is involved in the formation of the isochromosome 1(q). This is possible to determine because of the polymorphic centromeric heterochromatin on chromosome 1 of the early-passage PC/AA. With the late-passage PC/AA (grown with 3T3 feeders) the homologue with the large C-band has given rise to an isochromosome 1(q) whereas with PC/AA/FI it is the other homologue with the smaller C-band which has given rise to this isochromosome. Both the immortal PC/AA/FI and the immortal PC/AA late passage, therefore, have independent abnormalities involving chromosome 1. These results indicate that chromosome 1 may be involved in in vitro immortalization.

Adenoma↗
PubMed1989

Immunodetection of cathepsins B and L present in and secreted from human pre-malignant and malignant colorectal tumour cell lines.

Pre-malignant and malignant human colorectal tumour epithelial cell lines both secreted precursor forms of the 2 cysteine proteinases, cathepsins B and L. The amount of proteinases secreted by these cell lines varied according to the cell density. Comparison at similar cell densities showed that the pre-malignant, adenoma-derived cell line (PC/AA) secreted as much, or more, of both cathepsin B and L precursors as did the malignant, carcinoma-derived cell line (PC/JW/FI). However, mature forms of cathepsins B and L were detected in the culture media of only the carcinoma-derived cell line, thus indicating that the invasive potential of a tumour may be related to its ability to process extracellularly the secreted precursor enzyme to a mature and consequently active enzyme, rather than to the amount of proteinase synthesized and/or secreted. Similar results were obtained using 2 other epithelium-derived tumour cell lines, HT/29 (carcinoma) and SP/AN (adenoma). Immunolocation studies showed that cathepsin B was lysosomal while cathepsin L appeared to have a distribution more consistent with a plasma membrane association. Purified human cathepsins B and L (mature form) were capable of solubilizing an isolated basement membrane matrix (bovine anterior lens capsule) in vitro, thus indicating that the secreted mature enzymes and the membrane-associated cathepsin L could potentially degrade basal laminae or sub-endothelial basement membranes in vivo.

Adenoma↗
PubMed1989

Specific cytogenetic abnormalities in two new human colorectal adenoma-derived epithelial cell lines.

Two new epithelial cell lines from sporadic human colorectal adenomas designated S/AN and S/RG are reported. S/AN was from a villous adenoma and S/RG from a tubular adenoma. Both cell lines have extended growth capacities in vitro reaching passages 18 and 15, respectively, so far and show no signs of senescence. S/AN and S/RG have retained in vitro the ability to form mucin-producing goblet-like cells. Every cell of S/AN has a deletion on the short arm of chromosome 1 and one normal copy of chromosome 1. S/AN is also monosomic for chromosome 18. The majority of cells of S/RG only have one normal copy of chromosomes 6, 7, 14, 17, 18, and 22. S/RG also has several marker chromosomes. Although aneuploid S/AN and S/RG are nontumorigenic in athymic nude mice, these cytogenetic abnormalities are insufficient for the fully tumorigenic phenotype. The common abnormality for S/AN and S/RG is monosomy for chromosome 18, indicating that this is a central and important step in colorectal carcinogenesis. Our cytogenetic analysis of the adenoma cell lines suggests at least two possible routes by which premalignant colonic cells can develop and progress to malignancy. S/RG, unlike most other adenoma cell lines, is clonogenic. Aneuploidy, clonogenicity, and extended in vitro growth capacity may therefore be useful in vitro markers for adenoma cell lines with a relatively high malignant potential.

Adenoma↗