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Biomedical subjects

R Gambari

Publications and source records attributed to R Gambari.

At least 91 records · Page 5Linked to original sources
PubMed1991

A set of viral DNA decamers enriched in transcription control signals.

We studied the frequency distribution of oligonucleotides 10 bp long in a sample of 620 Kb of viral genomes, containing 102 sequences from GenBank, with the aim of detecting transcription control signals. Two thousand three hundred decamers had a frequency 10 times higher than the mean and were subjected to further statistical analysis. For each of the 2300 decamers (parents), we counted the individual frequencies of the 30 decamers differing from the parent by one base mutation (progeny) and then calculated two variance/mean chi squares for the progeny, with and without the parent. We then studied the distribution of the ratio between the two chi squares. Out of 2300 decamers, 10 times more frequent than average, 479 decamers had a chi square ratio of 1.9 or larger. In this final set, which corresponds to less than 0.05% of all possible decamers, 58 decamers were found to contain viral and eukaryotic transcription control elements, like NF-kB, Sp1 and others. Furthermore, this set contains an excess of signals of length 5, 6, 7, 8, 9 and 10, when compared to 150 random sets, bootstrapped from the same viral genomes.

Algorithms↗
PubMed1991

Transgenic mice mimic the methylation pattern of the human HLA-DR alpha gene.

The methylation pattern of the human HLA-DR alpha gene has been studied in different tissues of transgenic mice. Offspring from two transgenic lines was selected for this analysis, carrying the integrated HLA-DR alpha gene in either single or multiple (8-10) copies per diploid genome. In transgenic animals two distinct methylation patterns of the HLA-DR alpha gene are generated, due to a complete methylation of all the GCGC and CCGG sites the former, and to unmethylation restricted to one or both the GCGC sites located in the 5' portion of the HLA-DR alpha gene, the latter. Unmethylation restricted to the 5' portion of the HLA-DR alpha gene is a highly conserved feature in human tissues and in vitro cultured cell lines; therefore, it is concluded that the methylation pattern of the human HLA-DR alpha transgene may be faithfully reconstituted in transgenic animals. Northern blotting analysis of the RNA isolated from tissues of the transgenic mouse carrying single-copy HLA-DR alpha transgene demonstrates its tissue specific expression, suggesting that transgenic mice may represent an "in vivo" experimental system to study the relationship between methylation state and transcriptional activation.

Animals↗
PubMed1991

Distamycin inhibits the binding of a nuclear factor to the -278/-256 upstream sequence of the human HLA-DR alpha gene.

In this study we analyse the effects of the anti-tumor compound distamycin on the binding of nuclear factor(s) to a synthetic oligonucleotide (GTATA/IFN-gamma) mimicking a putative regulatory region of the human HLA-DR alpha gene. This region contains the sequence (GTATA), that is required for nuclear protein binding and is likely to interact with distamycin. The present results, by showing that distamycin inhibits the interaction between nuclear factors and the GTATA/IFN-gamma oligonucleotide, suggest that distamycin might alter the binding of transacting factors to cis-elements containing AT/TA sequences. Alterations of nuclear protein binding to specific target sequences could be one of the molecular mechanism(s) by which distamycin exerts its antiproliferative activity on living cells.

Antineoplastic Agents↗
PubMed1991

Tissue-specific expression of the HLA-DRA gene in transgenic mice.

Transgenic mice were produced containing a 33 kilobase (kb) DNA fragment encompassing the five exons and all the known regulatory regions of the class II HLA-DRA gene. The transgene displayed regulated expression [constitutive and interferon-gamma (IFN)-gamma induced] of the human products in most mouse tissues. The tissue distribution of the DRA transgene products more closely resembled that of their mouse homologues, the endogenous H-2 Ea products, than the wider distribution of DRA products in humans. This was evident in several tissues (endothelia of small vessels, especially those of glomerular capillaries, Kupffer cells, and epithelial cells lining the gastrointestinal tract), known to differentially express class II molecules in the two species. Thus, the wider human specific pattern of expression requires an exact cis/trans complementation which is incompletely reconstituted in transgenic mice, suggesting that human-specific cis-acting elements may have arisen during evolution to direct the expression of class II genes to those anatomical regions which usually lack them in the mouse. The only example of aberrant expression of the DRA gene in the present series of transgenic mice was in the dendritic and/or epithelial cells of the thymic cortex, which displayed greatly reduced DR alpha levels in spite of a normal expression of the endogenous E alpha molecules.

Animals↗
PubMed1991

N1-substituted benzamidines: synthesis, antiproteinase activity and inhibition of tumor cell growth.

We have synthesized N1-substituted benzamidines and poly-benzamidines with the aim to produce antitumor drugs retaining differential biological properties with respect to unsubstituted compounds. Antiproliferative activity on in vitro cultured human leukemic cells was exhibited by N1-substituted poly-benzamidines, while N1-substituted benzamidines were found to retain very low antitumor effects. Furthermore, our results suggest that N1-substituted benzamidines and some of poly-benzamidines exhibit low activity on trypsin and kallikrein. Taken together these data indicate that some N1-substituted poly-benzamidines could be of interest for experimental antitumor therapy, since are likely to retain low side effects due to alteration of proteinase activity.

Animals↗
PubMed1991

Effects of aromatic polyamidines on the electrophoretic mobility of HIV-1 genomic regions amplified by polymerase-chain reaction.

In this paper we describe the use of polymerase chain reaction (PCR) to amplify DNA sequences suitable for studies on the activity of DNA-binding drugs of possible interest in anti-tumor as well as anti-viral therapy. To this aim (a) we amplified by PCR two regions of the HIV-1 genome (one localized within the LTR, the other within the env gene), known to bind nuclear factors and (b) we determined whether different aromatic polyamidines are able to differentially affect the electrophoretic mobility of these HIV-1 PCR fragments. We found that aromatic polyamidines differentially affect the electrophoretic migration of PCR-amplified HIV-1 genomic regions. This differential effect, related to a differential DNA-binding activity, could lead to a differential inhibition of protein-DNA interactions.

Base Sequence↗
PubMed1990

Human HLA-DR alpha gene: a rare oligonucleotide (GTATA) identifies an upstream sequence required for nuclear protein binding.

Synthetic oligonucleotides containing putative regulatory sequences are currently employed to identify and isolate genes coding for nuclear binding factors. Upstream DNA sequences of eukaryotic genes required for transcriptional activity and tissue specificity can be identified by means of biochemical techniques as well as computer analysis using homology searching. An alternative approach has been recently proposed by our research group. Scanning DNA sequences 1.8 megabases in length from a Genetic Sequence Data Bank, we have identified rare oligonucleotides 5 base pairs (bp) long, which are localized within or close to regulatory segments in mammalian promoters. In this paper we demonstrate that the rare GTATA sequence identifies an upstream region of the HLA-DR alpha gene which operates in conjunction with the sequence AGAAGTCAG, homologous to a box found in many interferon-inducible genes, in binding nuclear proteins.

Base Sequence↗
PubMed1990

Tetra-amidines exhibiting anti-proteinase activity: effects on oriented migration and in vitro invasiveness of a Chinese hamster cell line transfected with the activated human T24-Ha-ras-1 oncogene.

In the present paper we have investigated the effects of aromatic tetra-amidines on attachment, oriented migration and in vitro invasiveness of the Chinese Hamster FH06T1-1 fibroblast lung cell line, transformed with the activated human T24-Ha-ras-1 oncogene. The FH06T1-1 cell line is tumorigenic in nude mice and displays growth properties and biological features clearly distinct from those of the FH06N1-1 cell line, obtained after transfection of the same fibroblast cells with the normal Ha-ras-1 proto-oncogene. Attachment, oriented migration and invasiveness were analysed by culturing the cells on a reconstituted extracellular matrix, composed of collagen IV, laminin, entactin and heparan sulphate proteoglycans. The results obtained demonstrate that oriented migration is performed only by FH06T1-1 cells and that tetra-benzamidines are effective inhibitors of oriented migration and "in vitro" invasiveness of these tumorigenic cells. These findings should encourage further studies on the possible antimetastatic effects of these antiproteinase tetra-benzamidines on experimental animals.

Amidines↗
PubMed1990

Liposome-associated retinoic acid. Increased in vitro antiproliferative effects on neoplastic cells.

The activity of liposome-associated retinoic acid was analyzed on in vitro cultured tumor cell lines and compared to the antiproliferative effects of free retinoic acid. It was found that liposome-associated retinoic acid is about 300 times more active than free retinoic acid in inhibiting in vitro cell growth of leukemic and melanoma cell lines. An increased activity of retinoic acid (10-20 times) was also obtained after premixing of this compound with empty liposomes, demonstrating that the retinoic acid efficiently interacts with liposomes which may facilitate solubility and cell uptake of retinoids.

Animals↗
PubMed1990

Tumor cell growth inhibition by liposome-encapsulated aromatic polyamidines.

Apart from its antiproteinase activity, the aromatic polyamidine TAPP-Br [the bromo derivative of 1,3-di-(p-amidinophenoxy)-2,2-bis-(p-amidinophenoxymethyl)propane (TAPP-H)] is able to inhibit the in vitro growth of a variety of tumor cell lines, including human melanoma, and breast and kidney carcinoma. We have now shown that TAPP-Br can efficiently be encapsulated into egg phosphatidylcholine vesicles. When incorporated into these liposomes, the inhibitory effect of TAPP-Br is significantly enhanced compared with that of the free drug. Based on these promising results, a proposal is made for the delivery of this antiproliferative agent to tumor cells by using liposomes as the vehicle.

Antineoplastic Agents↗
PubMed1989

Differential effects of liposome-entrapped desferrioxamine on proliferation and erythroid differentiation of murine erythroleukemic Friend cells.

It is known that iron chelators (such as desferrioxamine) are potent inhibitors of both cell proliferation and erythroid differentiation. We have shown with in vitro studies that in the case of tumor cells desferrioxamine is even more efficient in inhibiting cell proliferation when entrapped in liposomes consisting of egg yolk phosphatidylcholine. At the same time liposome-entrapped desferrioxamine retains only a slight effect on hexamethylenebisacetamide induction of erythroid differentiation and hemoglobin accumulation of murine erythroleukemic Friend cells. Based on these findings, we propose liposome-entrapped desferrioxamine as potential antineoplastic agent as well as a specific chemical for the study of both iron metabolism and distribution in normal and neoplastic cells. In addition, unlike free desferrioxamine, the liposome-entrapped drug could also be used in combination with inducers of differentiation. With respect to this issue, it is possible that liposome-entrapped desferrioxamine, might permit erythroid differentiation of both neoplastic cells as well as normal stem cells.

Animals↗
PubMed1989

Methylation state of cellular genes and oncogenes as a marker of malignancy in human carcinomas.

The methylation pattern of the human HLA-DR alpha gene was analyzed in primary tumors and lymph node metastases isolated from patients with a variety of tumors, including thyroid, pancreas, breast and gastric carcinomas and melanomas. In normal tissues (including breast, muscle, brain, sperm, T-and B-lymphocytes) the HLA-DR alpha gene is hypermethylated at CCGG and GCGC sites. In all tissues studied, the only constantly unmethylated region was located in the 5' portion of the gene. Our results indicate that the HLA-DR alpha gene is hypomethylated in metastatic lymph nodes, as well as in the carcinomas and melanomas studied. These findings lend support to the hypothesis that DNA hypomethylation of the human HLA-DR alpha gene may represent a molecular marker of malignant tumors.

DNA↗
PubMed1989

Induction of murine erythroleukemia cell differentiation by proteinases is inhibited by aromatic poly-amidines.

The effects of the tetra benzamidine serine-proteinase inhibitor 1,3-di-(p-amidinophenoxy) -2,2- bis- (p-amidinophenoxymethyl)propane (TAPP-H) and related compounds, including halo-derivatives, were determined on the erythroid differentiation of murine erythroleukemic cells induced by trypsin and kallikrein. These aromatic poly-amidines and their halo derivatives were found to be strong inhibitors of both trypsin and kallikrein mediated induction of commitment of MEL cells to erythroid differentiation, hemoglobin synthesis and accumulation, globin mRNA production. No inhibitory effects were detected by treating proteinase-induced MEL cells with benzamidine. Only slight inhibitory activity was found after treatment of trypsin-induced MEL cells with other antiproteinase compounds widely used in the control of proteinase-dependent functions, including leupeptin, antipain and Bowman-Birk proteinase inhibitor. MEL cells induced to erythroid differentiation by proteinases could be proposed as an experimental system to test the biological activity of proteinase inhibitors.

Amidines↗
PubMed1989

The frequency of oligonucleotides in mammalian genic regions.

The large body of nucleic acid sequence data now available offers a unique opportunity for the characterization of individual oligonucleotides which may be specific to sequence functional domains. We have prepared algorithms for the study of the frequency distribution of all oligonucleotides of length 2-6 in DNA sequences. We have implemented them in the study of 634 mammalian DNA sequences spanning 1.782 Mb, and have obtained the distribution of the ratio between the observed frequency of oligonucleotides and their expected frequency based on independent nucleotide probabilities. We then studied the distribution of oligonucleotides (or k-tuples) of each length in a subset of 129 complete mammalian genes spanning 0.607 Mb. Eight distinct genomic regions, namely 5'-non-transcribed, first exon, first intron, intermediate exons, intermediate introns, last intron, last exon and 3'-non-transcribed, were considered. We observed that some oligonucleotides show a statistical behaviour and a regional distribution similar to that of known signal sequences. Moreover the frequency distribution of oligonucleotides of length 5 and 6 tends to become bimodal, indicating the existence of a population of very frequent oligonucleotides.

Algorithms↗
PubMed1989

Isolation and characterization of a K562 cell line resistant to 1-beta-D-arabinofuranosylcytosine-mediated erythroid induction.

The isolation of a K562 cell line, K562(S)R, resistant to 1-beta-D-arabinofuranosylcytosine (ara-C)-mediated erythroid induction, is described. Ara-C (10-50 microM) inhibits cell growth of K562(S)R cells but is not able to activate the program of erythroid induction. This failure is associated with the lack in the increase of accumulation of epsilon-globin and gamma-globin mRNA sequences in ara-C-treated K562(S)R cells. This cell line could be of interest for studies focused on molecular mechanisms of activation of globin genes in K562 cells.

Azacitidine↗