Effect of sodium butyrate and other differentiation inducers on poorly differentiated human ovarian adenocarcinoma cell lines.

We have studied the effects of sodium butyrate, retinoic acid, and dimethyl sulfoxide on two human ovarian carcinoma cell lines PE04 and PE01. PE04 cells, after treatment with sodium butyrate at cytostatic doses (2-3 mM for 4 days), exhibited phenotypic changes including induction of alkaline phosphatase and determinants recognized by the monoclonal antibodies 123C3 and 123A8. These effects are not simply the result of cytostasis as they were not produced by dimethyl sulfoxide or retinoic acid. Other markers are also modified by sodium butyrate including lipid, acid mucin, and glycogen. Retinoic acid modulated expression of lipid and CA125, while dimethyl sulfoxide reduced expression of CA125. Other short chain fatty acids such as propionic acid and valeric acid (in addition to butyric acid) also induced alkaline phosphatase and the determinants recognized by 123C3 and 123A8 in PE04 cells. Other differentiation inducers and cytotoxic agents studied did not induce these markers at cytostatic concentrations. The effects of sodium butyrate (and related short chain fatty acids) thus appear to be relatively specific for this cell line.

Characterization and properties of nine human ovarian adenocarcinoma cell lines.

Four series of cell lines have been derived from patients with ovarian adenocarcinoma. Nine cell lines have been established at one from a solid metastasis. Six lines were derived from the ascites or pleural effusion of patients with poorly differentiated adenocarcinoma: PEO1, PEO4, and PEO6 from one patient, PEA1 and PEA2 from a second, and PEO16 from a third. Three lines (PEO14 and PEO23 from ascites and TO14 from a solid metastasis) were derived from a patient with a well-differentiated serous adenocarcinoma. Each set of cell lines was morphologically distinct. The five cell lines PEO1, PEO4, PEO6, PEA1, and PEA2 had cloning efficiencies on plastic of 1-2% and only a few cells in these lines expressed alkaline phosphatase or vimentin. Only a low percentage of these cells reacted with the monoclonal antibodies 123C3 and 123A8 but most reacted with OC125. Conversely the cell lines PEO14, TO14, PEO23, and PEO16 were characterized by low cloning efficiency values (less than 0.05%), marked expression of alkaline phosphatase and vimentin, and good reaction with 123C3 and 123A8 but not OC125. These four cell lines also exhibited dome formation. Four of the cell lines, PEO1, PEO4, PEO6, and PEO16, have been xenografted into immune-deprived mice and found to be tumorigenic.

Establishment of ovarian cancer cell lines.

Human tumor cell lines have provided valuable model systems to study a wide variety of tumor characteristics including the cell biology, genetics, and chemosensitivity profiles of disease. A large number of ovarian cancer cell lines have now been established and are in widespread use Table 1) (1-15). Many of these have been selected to reflect specific situations, e.g., pre- and postchemotherapy models or different histo- logical subtypes. Table 1 Properties of Established Ovarian Carcinoma Cell Lines Prior Cell Line Histology Source Treatment Ref. PE01 P.D. Serous adenoca Ascites P/FU/CHL 1 PE04 P.D. Serous adenoca Ascites P/FU/CHL 1 PE06 P.D. Serous adenoca Ascites P/FU/CHL 2 PEA1 P.D. Adenoca Pleural None 2 PEA2 P.D. Adenoca Ascites P/Pred 2 PE016 P.D. Serous adenoca Ascites Radioth 2 PE014 W.D.Serous adenoca Ascites None 2 T014 W.D.Serous adenoca Sol. Met None 2 PE023 W.D.Serous adenoca Ascites P/CHL 2 SKOV-3 Adenoca Ascites T 3 SW626 Adenoca - - 3 OVCAR-2 Adenoca Ascites P/Cy 4 OVCAR-3 P.D. papillary adenoca Ascites P/Cy/Adr 5 OVCAR-4 Adenoca Ascites P/Cy/Adr 6 OVCAR-5 Adenoca Ascites None 7 OAW 28 Adenoca Ascites P / Mel 8 OAW 42 Serous adenoca Ascites P 8 41M Adenoca Ascites None 9 59M Endometr adenoca Ascites None 8 CH1 Papillary adenoca Ascites P/ JM8 8 138D Serous adenoca Ascites Carb 9 180D Adenoca Ascites P 9 200D Serous adenoca Solid None 9 253D Serous adenoca Ascites Cy/MPA 9 HOC-1 W.D. Serous adenoca Ascites None 10 HOC-7 W.D. Serous adenoca Ascites None 10 CAOV-3 Adenoca Tumour Cy/Adr/FU 10 COLO 110 Serous adenoca Sol. Met None 11 COLO 316 Serous adenoca Pleural None 11 COLO 319 Serous adenoca Ascites None 11 COLO 330 Serous adenoca Ascites Mel/Radiother 11 IGROV1 Adenoca Primary None 12 HTOA W.D. serous adenoca Primary None 13 OV-1063 Papillary adenoca Ascites Cy/Adr/P/HMM 14 DO-s W.D. mucinous adenoca Ascites - 15 P.D. = Poorly differentiated; W.D. = Well differentiated; adenoca = adenocarcinoma; pleural = pleural effusion; Sol.met. = solid metastasis; P = cisplatin; FU = 5-fluorouracil; CHL = chlorambucil; Pred = prednimustine; Radioth = radiotherapy; T = thiotepa; Cy = cyclophosphamide; Adr = adriamycin; Mel = melphalan; Carb = carboplatin; MPA = medroxyprogesterone actetate; HMM = examethylmelamine.

Serum cholinesterase in the mothers of neural tube defect progeny.

Human Serum Cholinesterase activity and polymorphism at its two gene loci, CHE1 and CHE2, were compared in maternal serum from neural tube defect pregnancies, normal pregnancies and a non-pregnant control group. Variants at the CHE1 locus were identified by dibucaine, fluoride and R02 0683 inhibition. The CHE2 phenotype was demonstrated by DISC polyacrylamide gel electrophoresis. Total HSChE activity in the pregnant groups was slightly less than in the control group but there was no difference in activity between the affected and the normal pregnancies. Three variants were identified. All were found in the non-affected pregnant group. One variant at CHE1 was identified, an I phenotype, and two C5+ phenotypes, the CHE2 variant. No obvious relationships were found between HSChE activity or a particular genetic variant and NTD progeny.

The identification of human chromosomes by quinacrine fluorescence after hybridisation in situ.

A method is described for producing fluorescent bands on human chromosomes by staining with quinacrine after hybridisation in situ. The advantages of the method include the eslimination of artefacts arising from staining before hybridisation, the fact that there is no reduction in sample number between staining and autoradiography, the ease with which autoradiographic grains can be identified and counted. and the reduction in exposure time.

Satellite DNA and cytological staining patterns in heterochromatic inversions of human chromosome 9.

A series of partial inversions of the heterochromatic C-band of chromosome 9 have been stained with distamycin A plus 4',6-diamidino-2-phenyl-indol-2HCl (DA/DAPI) and found to consist of three classes: (a) those in which only the C-band in the long arm fluoresces with DA/DAPI (these are the most frequent), (b) those in which only the C-band in the short arm fluoresces with DA/DAPI, and (c) those in which the C-bands in both arms fluoresce with DA/DAPI. There are also differences in the satellite DNA content of each type of inversion as measured by hybridisation in situ. Types (a) and (b) have satellite DNA contents similar to those of their normal homologues, which type (c) has a satellite DNA content almost double that of the normal homologue. It appears that DA/DAPI specifically stains heterochromatin that contains satellite DNA. The ability to distinguish these three types of inversion may help to resolve the question of the clinical significance of such inversions.

A cloned repeated DNA sequence in human chromosome heteromorphisms.

A sequence derived by ECoRI restriction of human satellite DNA III has been cloned in lambda gt WES. The cloned DNA was used as a template for in vitro synthesis of cRNA, which was hybridized in situ to preparations of human metaphase chromosomes with a range of heterochromatic polymorphisms. Most of the hybridization was found on chromosome 1, and the amount of hybridization was related to the size of the C-band on this chromosome. Hybridization to other chromosomes was not related to the C-band size, although hybridization of total satellite DNA is proportional to C-band size. Total satellite DNAs contain a mixture of sequences, some of which are predominantly located on only one pair of chromosomes. Hybridization in situ is able to discriminate between such chromosome-specific sequences and the bulk of satellite DNA. Further analysis of satellite DNAs may identify sequences specific for every chromosome pair.

Satellite DNA sequences in the human acrocentric chromosomes: information from translocations and heteromorphisms.

Satellite III DNA has been located by in situ hybridization in chromosomes 1, 3--5, 7, 9, 10, 13--18, 20--22, and Y and ribosomal DNA (rDNA) in the acrocentric chromosomes 13--15, 21, and 22. In the acrocentric chromosomes, the satellite DNA is located in the short arm. Here we report comparisons by in situ hybridization of the amount of satellite DNA in Robertsonian translocation and "normal variant" chromosomes with that in their homologs. In almost all dicentric Robertsonian translocations, the amount of satellite DNA is less than that in the normal homologs, but it is rarely completely absent, indicating that satellite DNA is located between the centromere and the nucleolus organizer region (NOR) and that the breakpoints are within the satellite DNA. The amount of satellite DNA shows a range of variation in "normal" chromosomes, and this is still more extreme in "normal variant" chromosomes, those with large short arm (p+ or ph+) generally having more satellite DNA than those with small short arms (p- or ph-). The cytological satellites are heterogeneous in DNA content; some contain satellite DNA, others apparently do not, and the satellite DNA content is not related to the size or intensity of fluorescence of the satellites. The significance of these variations for the putative functions of satellite DNA is discussed.

Satellite DNA loss and nucleolar organiser activity in an individual with a de novo chromosome 13,14 translocation.

The distribution of satellite DNA and nucleolar organiser activity have been studied in a female with a new dicentric translocation chromosome derived from the maternal chromosomes 13 and 14. More than half the satellite DNA (60.5%) was lost in the translocation, together with both the nucleolar organiser regions (NOR'S). However, at least one NOR (chromosome 21) which was inactive in the mother (by the AgI reaction) is active in the subject, and this may be an example of functional compensation. The somatic cells of the mother of the subject, which do not have the translocation, show a high frequency of acrocentric associations, but these do not include any obvious excess of associations involving chromosomes 13 and 14, indicating that the high frequency of association in somatic cells is not in itself a predisposition to Robertsonian translocation in germ line cells. The father's chromosomes 9 both have more satellite DNA in the secondary constriction than normal, but this is not reflected in any obviously larger size of the C-band in this region.

The location of DNA homologous to human satellite III DNA in the chromosomes of chimpanzee (Pan troglodytes), gorilla (Gorilla gorilla) and orang utan (Pongo pygmaeus).

Radioactive RNA with sequences complementary to human DNA satellite III was hybridised in situ to metaphase chromosomes of the chimpanzee (Pan troglodytes), the gorilla (Gorilla gorilla) and the orangutan (Pongo pygmaeus). A quantitative analysis of the radioactivity, and hence of the chromosomal distribution of human DNA satellite III equivalent sequences in the great apes, was undertaken, and the results compared with interspecies chromosome homologies based upon Giemsa banding patterns. In some instances DNA with sequence homology to human satellite III is present on the equivalent ("homologous") chromosomes in identical positions in two or more species although quantitative differences are observed. In other cases there appears to be no correspondence between satellite DNA location and chromosome homology determined by banding patterns. These results differ from those found for most transcribed DNA sequences where the same sequence is located on homologous chromosomes in each species.

Oestrogen receptor expression and the effects of oestrogen and tamoxifen on the growth of human ovarian carcinoma cell lines.

To assess the role of oestrogen regulation in the growth of ovarian cancer, we examined the effects of an oestrogen, 17 beta-oestradiol, and an anti-oestrogen, tamoxifen, on oestrogen receptor (ER) -positive and -negative human ovarian carcinoma cell lines. As measured by a dextran-coated charcoal adsorption assay, cell lines PEO1, PEO4 and PEO6 possessed moderate concentrations of ER (96-132 fmol mg-1 protein), PEA1 and PEA2 had low values (12-23 fmol mg-1 protein) and PEO14, TO14, PEO23 and PEO16 were ER-negative. Addition of 17 beta-oestradiol (10 nM or 0.1 nM) to the ER +ve cell line, PEO4, increased the growth rate. This oestrogen stimulation could be blocked by 1 microM tamoxifen. In contrast, the growth rate of the ER -ve cell line PEO14 was unaffected by the addition of 17 beta-oestradiol or tamoxifen. Concentrations of tamoxifen in excess of 8 microM were required to produce complete cytostasis in all lines. This concentration of tamoxifen over 72 hours also inhibited 50% colony formation when cells were plated on plastic. These data indicate that some ovarian carcinoma cell lines contain ER and their growth can be sensitive to oestrogen and anti-oestrogen modulation.

Cellular heterogeneity and drug resistance in two ovarian adenocarcinoma cell lines derived from a single patient.

Two ovarian cell lines were derived from the ascites of a patient before and after the onset of resistance to chemotherapy involving cis-platinum, chlorambucil and 5-fluorouracil. Characterization of these lines shows them to have various features in common and some significant differences. Cytologically the lines cannot be distinguished and they both contain high concentrations of oestrogen receptor. However, they do differ with respect to their growth characteristics, karyotype, glutathione content and sensitivity to cis-platinum. The karyotypes of the 2 lines show several marker chromosomes in common but the resistant line contained a chromosome 8 and a 17 which were absent from the earlier sensitive line. This suggests a clonal origin with subsequent divergence to a heterogeneous population.