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Biomedical subjects

M Birnstiel

Publications and source records attributed to M Birnstiel.

16 recordsLinked to original sources
PubMed1999

Immunotherapy of metastatic malignant melanoma by a vaccine consisting of autologous interleukin 2-transfected cancer cells: outcome of a phase I study.

We performed a phase I trial to evaluate the safety and tolerability of repeated skin injections of IL-2-transfected autologous melanoma cells into patients with advanced disease. Cell suspensions, propagated from excised metastases, were IL-2 gene transfected by adenovirus-enhanced transferrinfection and X-irradiated prior to injection. Vaccine production was successful in 54% of the patients. Fifteen patients (37%) received two to eight skin vaccinations of either 3 x 10(6) (intradermal) or 1 x 10(7) (half intradermal, half subcutaneous) transfected melanoma cells per vaccination (secreting 140-17,060 biological response modifier program units of IL-2/10(6) cells/24 hr). Analyses of safety and efficacy were carried out in 15 and 14 patients, respectively. Overall, the vaccine was well tolerated. All patients displayed modest local reactions (erythema, induration, and pruritus) and some experienced flu-like symptoms. Apart from newly appearing (4 of 14) and increasing (5 of 14) anti-adenovirus and newly detectable anti-nuclear antibody titers (1 of 15), recipients developed de novo or exhibited increased melanoma cell-specific delayed-type hypersensitivity (DTH) reactions (8 of 15) and vitiligo (3 of 15) and showed signs of tumor regression (3 of 15). This supports the idea of a vaccine-induced or -amplified anti-cancer immune response. None of the patients exhibited complete or partial regressions, but five of them experienced periods of disease stabilization. Three of these individuals received more than the four planned vaccinations and their mean survival time was 15.7 +/- 3.5 months as compared to 7.8 +/- 4.6 months for the entire patient cohort. These data indicate that IL-2-producing, autologous cancer cells can be safely administered to stage IV melanoma patients and could conceivably be of benefit to patients with less advanced disease.

Adult↗
PubMed1979

Sequence organization of the spacer DNA in a ribosomal gene unit of Xenopus laevis.

A detailed restriction map was constructed for a cloned Xenopus laevis rDNA fragment containing the nontranscribed spacer (NTS) and external transcribed spacer (ETS) together with a portion of both the 18S and 28S rRNA genes. The NTS was found to contain at least three distinct repetitious areas. Region 1 has a repeating unit of approximately 100 bp. The primary structure of this unit has been determined by DNA sequencing. Region 2 is very similar in organization to region 3, and both have an alternating 81/60 bp arrangement as revealed by restriction with Alu I and DNA sequencing. It can be shown that the 81 and 60 bp canons are virtually identical to one another excepting a deletion/insertion of a 21 bp segment. Region 3 differs from region 2 in having sites for Sma I with its 81 bp units. Between these repeated DNA sequences there are two identical, nonrepetitive DNA sequences, each of which is centered around a Bam Hl site. Most of the ETS has been sequenced. It was found to be nonrepetitive and extremely rich in Cs. Close to the 5' end of the 18S coding sequence there is a DNA stretch very rich in purines. About 2.25 kb upstream from the Eco Rl restriction site bisecting the 18S structural gene there is a unique sequence which may be homologous to the 5' end of the 40S precursor RNA. Present evidence suggests that the boundaries between NTS and ETS occur farther downstream than was suggested by electron microscopic data. Sequencing has revealed that the spacer DNA of X. laevis contains different kinds of simple DNA sequences, but no evidence has been found that spacer DNA once arose by saltation of a 15 bp segment. The most surprising finding was that the spacer sequences around the Bam restriction sites (the Bam islands) show high homology with a sequence near the NTS/ETS interface. From the restriction and sequencing analyses it can be deduced that in recent evolutionary times the DNA sequences near the 5' end of the ribosomal transcription unit were reduplicated twice and displaced into spacer by saltation of an intervening short DNA sequence (the 60/81 bp canons). Possible implications of these evolutionary events for spacer functions are consisdered. The sequencing has also provided a molecular basis for a whole range of conclusions arrived at previously by indirect approaches, and these are discussed.

Animals↗
PubMed1977

Novel screening procedure for recombinant plasmids.

Lysed bacterial colonies containing potential recombinant plasmids were mixed with molten agar and sealed into slots of an agarose gel. After electrophoresis overnight, the gel was stained with ethidium bromide, which clearly reveals recombinant plasmids. Xenopus laevis ribosomal DNA and histone DNA of Psammechinus miliaris were ligated into pCRI plasmids and screened by this method.

Animals↗
PubMed1976

Partial denaturation mapping of cloned histone DNA from the sea urchin Psammechinus miliaris.

A cloned 5.6-kb DNA repeat unit of Psammechinus miliaris containing all five histone coding sequences of known order and polarity has been shown to fall clearly into a low melting and a high melting half by thermal denaturation. The topoligies of the DNA sequences thus defined were determined by partial denaturation mapping in the electron microscope to gain further insight into the distribution of spacers and genes within histone DNA.

Animals↗
PubMed1976

Molecular analysis of the histone gene cluster of Psammechinus miliaris: I. Fractionation and identification of five individual histone mRNAs.

The electrophoretic separation of labeled "9S" histone mRNAs obtained from cleaving sea urchin polysomes was found at first to be highly unreproducible. It became evident that the secondary structure of the individual mRNAs had a greater effect on their relative electrophoretic mobilities than did their molecular weight differentials. We determined the parameters affecting electrophoretic mobility by the novel method of running the labeled polysomal RNA in slab gels across polyacrylamide and urea gradients. The initially complex and species-specific electrophoretic pattern could then, by a judicious choice of denaturing conditions, be simplified to yield five well defined classes of labeled mRNAs. Using optimal conditions for the separation of the RNA components, five messengers were isolated from Psammechinus embryos by preparative disc electrophoresis, four of which, after two electrophoretic separations, exhibited a unimodal distribution. Each of the mRNAs was translated in vitro, four of the five fractions promoting the synthesis of one major protein. The in vitro products were characterized by comparison of their electrophoretic mobilities with those of known sea urchin histones. It was thus possible to correlate individual mRNAs with specific histones. We propose that the five mRNAs designated a-e in order of decreasing electrophoretic mobility code for the histones H4, H2A, H2B, H3, and H1.

Animals↗
PubMed1976

Molecular analysis of the histone gene cluster of psammechinus miliaris: II. The arrangement of the five histone-coding and spacer sequences.

Histone DNA of Psammechinus miliaris was obtained in an enriched form by buoyant density gradient centrifugation and was cleaved into 6 kb repeat units (Birnstiel et al., 1975a) by the action of the specific endonucleases EcoRI and HindIII. Since it was suspected that the 6 kb unit harbored all five histone-coding sequences, the histone DNA unit was subdivided into five segments with the aim of providing five fragments carrying just one coding sequence each. This was achieved by the combined use of EcoRI Hindll, Hindlll, and Hpa I. A physical map was constructed from the overlaps arising in these restriction experiments. Each of the five segments was shown to hybridize uniquely with just one of the five highly purified histone mRNAs (Gross et al., 1976a). By this procedure, the order of the mRNA sequences on the histone DNA was found to be a, c, d, b, e (Gross et al., 1976a), and hence of the protein coding sequences H4, H2B, H3, H2A, and H1. Further evidence is presented that the 6 kb repeat unit, amplified by means of a Murray lambda vector phage, contains AT-rich DNA sequences which would be expected not to code for histone proteins.

Adenine↗
PubMed1976

Molecular analysis of the histone gene cluster of Psammechinus miliaris: III. Polarity and asymmetry of the histone-coding sequences.

Fragments of histone DNA produced by restriction endonucleases contain 5' and 3' termini with defined topologies relative to the histone-coding sequences. After limited resection with lambda-exonuclease, the 6 kb Hindlll histone DNA fragment (see Schaffner et al., 1976) hybridizes to H4 histone mRNA whether or not the DNA has been denatured. This shown that the coding sequence for the histone H4 is proximal to the 3' terminus of the Hindlll restriction fragment. By contrast, lambda-exonuclease digestion of the 6 kb EcoRI histone DNA fragment drastically reduces hybridization of the H4 mRNA. Hence in this molecule, the H4 DNA sequence is near a 5' terminus of the EcoRI restriction fragment. From these results and those described in the preceding paper (Schaffner et al., 1976), the polarity of the H4 gene is therefore 5' H2B leads to H4 leads to H1 3'. Cloned Psammechinus histone DNA (S. G. Clarkson, H. Smith, W. Schaffner, K. Gross, and M. Birnstiel, manuscript submitted for publication) may be strand-separated by electrophoresis. Highly purified histone mRNAs (Gross et al., 1976) all hybridize almost exclusively to the strand of lesser electrophoretic mobility. It follows that all coding sequences are arranged in tandem within the same DNA strand, and that hence they share the same polarity as the H4 DNA sequences. Transcription therefore proceeds in the gene cluster in the direction H4 leads to H2B leads to H3 leads to H2A leads to H1.

Animals↗
PubMed1974

Isolation and some properties of the genes coding for histone proteins.

The DNA coding for histone proteins, together with the intervening spacer DNA, has been isolated from the sea urchin, Psammechinus miliaris, by repeated centrifugation on CsCl-asctinomycin gradients. The biophysical data show that a major portion of the spacer is 37% G+C and very divergent in sequence, while the DNA segments coding for histone proteins are high in G+C and have diverged to a lesser extent within the species.

Animals↗
PubMed1969

Kinetic complexity of chloroplastal deoxyribonucleic acid and mitochondrial deoxyribonucleic acid from higher plants.

1. Chloroplasts and mitochondria were isolated by aqueous and non-aqueous cell-fractionation techniques. In a variety of higher plants the mitochondrial DNA bands in a caesium chloride gradient at 1.706g.cm.(-3), whereas chloroplastal DNA has a buoyant density of 1.697g.cm.(-3). 2. In total cellular DNA of moderate molecular weight, the chloroplastal DNA is found within the Gaussian distribution of the nuclear DNA and is not resolved as a satellite. 3. Both chloroplastal DNA and mitochondrial DNA from lettuce renature rapidly. 4. The kinetic complexity of mitochondrial DNA is > 10(8) daltons. 5. Chloroplastal DNA is made up from fast and slow renaturing sequences with kinetic complexities of 3x10(6) and 1.2x10(8) daltons respectively. 6. From the discrepancy between analytical and kinetic complexity it is concluded that chloroplastal DNA is extensively reiterated.

Cell Nucleus↗