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D A Scudiero

Publications and source records attributed to D A Scudiero.

At least 37 records · Page 2Linked to original sources
PubMed1988

Feasibility of drug screening with panels of human tumor cell lines using a microculture tetrazolium assay.

For the past 30 years strategies for the preclinical discovery and development of potential anticancer agents have been based largely upon the testing of agents in mice bearing transplantable leukemias and solid tumors derived from a limited number of murine as well as human sources. The feasibility of implementing an alternate approach, namely combined in vitro/in vivo screening for selective cytotoxicity among panels of human tumor cell lines derived from a broad spectrum of human solid tumors is under investigation. A group of 30 cell lines acquired from a variety of sources and representing 8 lung cancer pathologies as well as 76 cell lines representing 10 other categories of human cancer (carcinomas of colon, breast, kidney, prostate, ovary, head and neck; glioma; leukemia; melanoma; and sarcoma) have exhibited acceptable growth characteristics and suitable colorimetric profiles in a single, standard culture medium. Measurements of in vitro growth in microculture wells by cell-mediated reduction of tetrazolium showed excellent correlation (0.89 less than r2 less than 0.98) with measurements of cellular protein in adherent cell line cultures as well as viable cell count in suspension cell line cultures (0.94 less than r2 less than 0.99). Since the microculture tetrazolium assay provides sensitive and reproducible indices of growth as well as drug sensitivity in individual cell lines over the course of multiple passages and several months' cultivation, it appears suitable for initial-stage in vitro drug screening.

Antineoplastic Agents↗
PubMed1987

The role of O6-methylguanine in human cell killing, sister chromatid exchange induction and mutagenesis: a review.

O6-methylguanine (O6mG) produced in DNA by such SN1 methylating agents as N-methyl-N-nitrosourea and N-methyl-N'-nitro-N-nitrosoguanidine (MNNG) has been suggested by some to be the lesion that leads to certain biological endpoints in mammalian cells: cell killing, sister chromatid exchange (SCE) production, mutagenesis and cellular transformation. Other evidence is interpreted as inconsistent with this point of view. The finding of Karran & Williams (1985) that O6mG delivered to cells in culture resulted in the depletion of the activity of the protein responsible for repair of O6mG in DNA (O6mG-DNA methyltransferase, O6MT) provided a tool for the assessment of the role of O6mG in producing biological endpoints. In this paper we review much of the literature on human cells pertinent to this question. In addition we present our survival data obtained using the depletion technique of Karran & Williams as well as data supporting a model invoking a mismatch and excision response to O6mG proposed by Sklar & Strauss (1980). Although data linking O6mG to causation are inconclusive, it is premature to conclude that O6mG is not a lesion lethal to certain cultured cells.

Adenoviridae↗
PubMed1985

Human tumor cell strains both unable to repair O6-methylguanine and hypersensitive to killing by human alpha and beta interferons.

Human brain tumor cell strains were previously found by others to be sensitive to growth inhibition by human interferon-beta (HuIFN-beta). We noticed that the sensitive strains were some that we had found deficient in the repair of O6-methylguanine (O6MeG), a characteristic of 20% of the human tumor cell strains we have studied. We confirmed this sensitivity to HuIFN-beta, and have further shown that human brain tumor cells which repair O6MeG are resistant to the growth inhibitory effects of HuIFN-beta. In addition, treatment with HuIFN-alpha or HuIFN-beta resulted in more killing (reproductive inactivation) of six human tumor cell strains deficient in repairing O6-methylguanine in DNA than did such treatment of six strains of cells proficient in such repair. Further, we found two human lines, altered to become O6MeG repair deficient after establishment of the primary tumor cell culture, that were resistant to interferon. IFN treatment produced no DNA damage detectable by either chemical or biological assays. It is suggested that the genes responsible for resistance to IFN treatment and to agents that produce O6MeG are often coordinately shut down.

Brain Neoplasms↗
PubMed1984

Hypersensitivity to DNA-damaging agents in cultured cells from patients with Usher's syndrome and Duchenne muscular dystrophy.

Lymphoblastoid lines from nine Usher's syndrome (recessively inherited retinitis pigmentosa and congenital sensorineural deafness) patients (representing eight kindreds) and from ten Duchenne muscular dystrophy patients (representing seven kindreds) showed a small but statistically significant hypersensitivity to the lethal effects of X-rays, as measured by the cellular ability to exclude the vital dye trypan blue, when compared with lines from 26 normal control subjects. Fibroblast lines from the Usher's syndrome patients, treated with X-rays or the radiomimetic, DNA-damaging chemical N-methyl-N'-nitro-N-nitrosoguanidine, also showed a statistically significant hypersensitivity when compared to normal fibroblast lines. These findings are consistent with the possibility that defective DNA repair mechanisms may be involved in the pathogenesis of these degenerative diseases.

Adolescent↗
PubMed1984

Relationship of DNA repair phenotypes of human fibroblast and tumor strains to killing by N-methyl-N'-nitro-N-nitrosoguanidine.

Two DNA repair assays were used to group human cells. (a) The first assay, survival of N-methyl-N'-nitro-N-nitrosoguanidine (MNNG)-treated adenovirus infecting cellular monolayers, was previously used to define the Mer phenotype of the strain. Strains that supported the growth of MNNG-treated viruses as well as did human fibroblasts were "Mer+"; those that gave rise to clearly less virus survival were "Mer-." (b) The second assay, data from which are presented in this paper, was that of post-MNNG colony-forming ability, and defined the Rem phenotype of the strain. Strains having post-MNNG colony-forming ability like that of human fibroblasts were "Rem+"; more sensitive strains were "Rem-". In all, 22 human cell strains were analyzed for their post-MNNG colony-forming ability. The most resistant strains (eight Mer+ Rem+ strains) had an average inactivation slope of 0.32 "lethal hit"/microM and were those fully able to repair O6-methylguanine (O6mGua) produced in their DNA by a 5 microM dose of MNNG. The most sensitive strains (9 Mer- Rem- strains) had an average inactivation slope of 7.0 "lethal hits"/microM, and were strains that failed to repair O6mGua. Five strains of intermediate sensitivity (Mer+ Rem-) had an average inactivation slope of 0.93 "lethal hit"/microM and were able to repair some labeled O6mGua produced by a 5 microM dose of labeled MNNG, but they repaired significantly less labeled O6mGua if pretreated with unlabeled MNNG. Representative strains from each group were treated with MNNG and assayed for ability: (a) to perform DNA repair synthesis (and DNA repair replication); (b) to support the growth of MNNG-treated adenoviruses; and (c) to restore control levels of tertiary structure to their DNA as assayed by nucleoid sedimentation. The results support the hypothesis that a lesion (both produced by agents that produce O6mGua and repaired by cell strains that repair O6mGua, but not by those that do not) is a lesion lethal to Mer- Rem- strains. This lesion may also initiate induction of excess DNA repair synthesis, the relaxed conformation of nucleoids, the reduced ability to repair MNNG-treated adenovirus, and sister chromatid exchanges as well.

Adenoviridae↗
PubMed1984

Sensitivity of human cell strains having different abilities to repair O6-methylguanine in DNA to inactivation by alkylating agents including chloroethylnitrosoureas.

In order to investigate the mechanisms of cellular damage by alkylating agents, human fibroblasts and tumor cell strains having different sensitivities to killing by N-methyl-N'-nitro-N-nitrosoguanidine [and different abilities to repair O6-methylguanine ( O6mGua ) in their DNA] were treated with other alkylating agents. Methyl methanesulfonate, 1,3-bis(2-chloroethyl)-1-nitrosourea (BCNU), 1-(2-hydroxyethyl)-3-(2-chloroethyl)-3-nitrosourea, and N-ethyl-N'-nitro-N-nitrosoguanidine gave rise to sensitivity differences, but the differences were less than those observed with N-methyl-N'-nitro-N-nitrosoguanidine. After treatment with UV light, the strains showed similar survival. The data show that the DNA repair mechanism(s) responsible for the differential survival of the strains after N-methyl-N'-nitro-N-nitrosoguanidine treatment probably play(s) a role in repairing DNA damage produced by methyl methanesulfonate, N-ethyl-N'-nitro-N-nitrosoguanidine, BCNU, and 1-(2-hydroxyethyl)-3-(2-chloroethyl)-3-nitrosourea but not that produced by UV. Furthermore, the results support the idea that a breakdown product of BCNU, that does not cause damage repairable by O6mGua repair mechanisms, contributes to the lethal effects due to BCNU-produced DNA-damage that is repairable by O6mGua repair mechanisms. The survival data, along with nucleoid sedimentation and adenovirus host-cell reactivation data, are consistent with the hypothesis that the lesion(s) lethal to tumor cells defective in O6mGua DNA repair are lesions in which DNA oxygen atoms are alkylated.

Alkylating Agents↗
PubMed1983

Statistical methods for in vitro cell survival assays.

Statistical methods are presented for estimating and comparing survival curves obtained from experiments in which cells are exposed in vitro to increasing doses of a DNA-damaging agent. These methods, which are applicable in a variety of cell survival assays, are illustrated in the evaluation of two sets of experiments in which the colony-forming ability of fibroblast cell lines from 9 muscular dystrophy patients and 17 normal individuals were studied after exposure to N-methyl-N'-nitro-N-nitrosoguanidine (MNNG).

Cell Survival↗
PubMed1982

Lymphoblastoid lines and skin fibroblasts from patients with tuberous sclerosis are abnormally sensitive to ionizing radiation and to a radiomimetic chemical.

Lymphoblastoid lines, derived by transforming peripheral blood lymphocytes with Epstein-Barr virus, and skin fibroblast lines were established from two patients with tuberous sclerosis. The number of viable lymphoblastoid cells was determined by their ability to exclude the vital dye trypan blue after their irradiation with x-rays or 254 nm ultraviolet light. The growth of fibroblasts was determined by their ability to form colonies after treatment with the radiomimetic, DNA-damaging chemical N-methyl-N'-nitro-N-nitrosoguanidine. The tuberous sclerosis lymphoblastoid lines were hypersensitive to x-rays but had normal sensitivity to the ultraviolet radiation. The tuberous sclerosis fibroblast lines were hypersensitive to the N-methyl-N'-nitro-N-nitrosoguanidine. The hypersensitivity of tuberous sclerosis cells to x-rays and to N-methyl-N'-nitro-N-nitrosoguanidine is believed to reflect defective repair of DNA damaged by these agents and may provide the basis for in vitro, including prenatal, diagnostic tests for tuberous sclerosis.

Adolescent↗
PubMed1981

Dependence of mammalian DNA synthesis on DNA supercoiling. III. Characterization of the inhibition of replicative and repair-type DNA synthesis by novobiocin and nalidixic acid.

Novobiocin and nalidixic acid, inhibitors of the bacterial enzyme DNA gyrase, inhibit DNA, RNA and protein synthesis in several human and rodent cell lines. The sensitivity of DNA synthesis (both replicative and repair) to inhibition by novobiocin and nalidixic acid is greater than that of protein synthesis. Novobiocin inhibits RNA synthesis about half as effectively as it does DNA synthesis, whereas nalidixic acid inhibits both equally well. Replicative DNA synthesis, as measured by incorporation of [3H]thymidine, is blocked by novobiocin in a number of cell strains; the inhibition is reversible with respect to both DNA synthesis and cell killing, and continues for as long as 20--30 h if the cells are kept in novobiocin-containing growth medium. Both novobiocin and nalidixic acid inhibit repair DNA synthesis (measured by BND-cellulose chromatography) induced by ultraviolet light or N-methyl-N'-nitro-N-nitrosoguanidine (but not that induced by methyl methanesulfonate) at lower concentration (as low as 5 micrograms/ml) than those required to inhibit replicative DNA synthesis (50 micrograms/ml or greater). Neither novobiocin nor nalidixic acid alone induces DNA repair synthesis. Incubation of ultraviolet-irradiated cells with 10--100 micrograms/ml novobiocin results in little, if any, further reduction of colony-forming ability (beyond that caused by the ultraviolet irradiation). Novobiocin at sufficiently low concentrations (200 micrograms/ml) apparently generates a quiescent state (in terms of cellular DNA metabolism) from which recovery is possible. Under more drastic conditions of time in contact with cells and concentration, however, novobiocin itself induces mammalian cell killing.

Cell Line↗
PubMed1981

Hypersensitivity to N-methyl-N'-nitro-N-nitrosoguanidine in fibroblasts from patients with Huntington disease, familial dysautonomia, and other primary neuronal degenerations.

Cells from patients with ataxia telangiectasia, a rare autosomal recessive disease characterized by primary neuronal degeneration, are abnormally sensitive to the DNA-damaging chemical N-methyl-N'-nitro-N-nitrosoguanidine. We have conducted experiments to determine whether more common primary neuronal degenerations also have a hypersensitivity to this radiomimetic chemical. Fibroblast strains from 13 control donors and from 13 patients with inherited primary neuronal degenerations were treated in vitro with the chemical, and the strains' sensitivity to the chemical was then determined by measuring their ability to divide and form colonies. Twelve of the 13 patient strains, including the 6 Huntington disease and the 4 familial dysautonomia strains, were abnormally sensitive. This hypersensitivity, which is believed to reflect defective repair of the chemically-induced DNA damage, might provide the basis for presymptomatic and prenatal diagnostic tests for these disorders and for elucidating their pathogenesis.

Adolescent↗
PubMed1980

Defective repair of alkylated DNA by human tumour and SV40-transformed human cell strains.

We have identified a group of 8 (among 39) human tumour cell strains deficient in the ability to support the growth of adenovirus 5 preparations treated with N-methyl-N'-nitro-N-nitrosoguanidine (MNNG), but able to support the growth of non-treated adenovirus normally. This deficient behaviour defines the Mer- phenotype. Strains having the Mer- phenotype were found to arise from tumours originating in four different organs. Relative to Mer+ strains, Mer- tumour strains showed greater sensitivity to MNNG-produced killing, greater MNNG-stimulated "DNA repair synthesis and a more rapid MNNG-produced decrease in semi-conservative DNA synthesis. Here we report that (1) Mer- strains are deficient in removing O6-methylguanine (O6-MeG) from their DNA after [Me-14C]MMNG treatment (Table 1); (2) Mer- tumour strains originate from tumours arising in patients having Mer+ normal fibroblasts (Fig. 1a, b); (3) SV40 transformation of (Mer+) human fibroblasts often converts them to Mer- strains (Fig. 1c, d); (4) MNNG produces more sister chromatid exchanges (SCEs) in Mer- than in Mer+ cell strains (Fig. 2).

Adenoviruses, Human↗
PubMed1980

Decreased DNA repair synthesis and defective colony-forming ability of ataxia telangiectasia fibroblast cell strains treated with N-methyl-N'-nitro-N-nitrosoguanidine.

DNA repair synthesis and posttreatment colony-forming ability were measured in six human fibroblast cell strains derived from patients with the autosomal recessive disorder, ataxia telangiectasia (AT), and five cell strains from normal individuals. All 11 cell strains showed approximately equal levels of DNA repair synthesis after methyl methanesulfonate and ultraviolet light treatments assayed by the benzoylated naphthoylated diethylaminoethyl-cellulose method. Four AT cell strains exhibited 27 to 41% of the repair shown by normal cell strains after treatment with N-methyl-N'-nitro-N-nitrosoguanidine (MNNG). Two AT cell strains had normal levels of repair synthesis after MNNG treatment. All the AT strains tested had normal survival after treatment with methyl methanesulfonate and ultraviolet light as assayed by posttreatment colony-forming ability. The six AT cell strains exhibited diminished survival after MNNG treatment. The inhibition of [3H]thymidine uptake as a function of MNNG concentration was similar for all the normal and AT cell strains tested.

Alkylating Agents↗
PubMed1973

Physiological modifications in the production and repair of methyl methane sulfonate-induced breaks in the deoxyribonucleic acid of Escherichia coli K-12.

The medium in which Rec(+) strains of Escherichia coli K-12 are grown affected their sensitivity to treatment with methyl methane sulfonate (MMS). Rec(+) cells grown to the stationary phase in glucose-enriched nutrient broth (GNB) were more resistant to MMS than cells grown in nutrient broth (NB). The repair of MMS-induced breaks (or alkali-labile bonds) in the deoxyribonucleic acid (DNA) from E. coli K-12 strains AB1157, AB1886 uvrA6, and SR111 recA13 recB21 grown in GNB and NB media was examined by means of alkaline sucrose gradient centrifugation. It appeared that essentially all of the repair of breaks that occurred, as evidenced by an increase in "molecular weight," took place within 10 min after treatment with MMS under our conditions. Cell survival was highest in cells for which the size of the DNA after the post-treatment incubation was the largest. The largest DNA after post-treatment incubation was found in Rec(+) cells grown in GNB medium. The results suggest that these cells may have an enhanced capacity for repairing breaks in DNA.

Alkanesulfonates↗
PubMedDate not supplied

A protein expression database for the molecular pharmacology of cancer.

In the last six years, the Developmental Therapeutics Program (DTP) of the US National Cancer Institute (NCI) has screened over 60,000 chemical compounds and a larger number of natural product extracts for their ability to inhibit growth of 60 different cancer cell lines representing different organs of origin. Whereas inhibition of the growth of one cancer cell type gives no information on drug specificity, the relative growth inhibitory activities against 60 different cells constitute patterns that encode detailed information on mechanisms of action and resistance (as reviewed in Boyd and Paull, Drug Devel. Res. 1995, 34, 19-109 and Weinstein et al., Science 1997, 275, 343-349). In order to correlate the patterns of activity with properties of the cells, we and other laboratories are characterizing the cells with respect to a large number of factors at the DNA, mRNA, and protein levels. As part of that effort, we have developed a two-dimensional gel electrophoresis (2-DE) protein expression database covering all 60 cell types (Buolamwini et al., submitted). Here we present analyses of the correlations among protein spots (i) in terms of their patterns of expression and (ii) in terms of their apparent relationships to the pharmacology of a set of 3989 screened compounds. The correlations tend to be stronger for the latter than for the former, suggesting that the spots have more robust signatures in terms of the pharmacology than in terms of expression levels. Links to pertinent databases and tools of analysis will be updated progressively at http:@www.nci.nih.gov/intra/lmp/jnwbio.htm and http:@epnwsl.ncifcrf.gov:2345/dis3d/dtp.++ +html.

Antineoplastic Agents↗