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Biomedical subjects

R Taetle

Publications and source records attributed to R Taetle.

At least 73 records · Page 4Linked to original sources
PubMed1989

Combination iron depletion therapy.

Iron (Fe) depletion with anti-transferrin (Tf) receptor monoclonal antibodies (MAbs), Fe chelators, or gallium (Ga) salts inhibits in vitro and in vivo growth of tumor cells. The present studies examined the cytotoxic effects of an IgA anti-human Tf receptor MAb, 42/6, combined with parabactin, a powerful Fe chelator, or Ga nitrate. Parabactin inhibited in vitro growth of human hematopoietic and solid tumor cells, and the rank order of their sensitivities to the Fe chelator was identical to their relative sensitivity to MAb 42/6 as demonstrated in previous studies. When the most parabactin and MAb 42/6-sensitive (HL60 leukemia) and -resistant (KB carcinoma) cells were incubated with various concentrations of parabactin, cell killing was time and dose dependent over the first 24 hours. Little additional cytotoxicity occurred, however, when cells were exposed to parabactin for 48 hours. HL60 cells were slightly more sensitive than KB cells to parabactin cytotoxicity. Addition of optimally effective concentrations of anti-Tf receptor MAb 42/6 to parabactin increased cytotoxicity to HL60 cells over a narrow parabactin dose range but had little effect on cytotoxicity to KB cells. Cell cycle analysis of cells treated with parabactin for 24 hours showed that doses causing variable cytotoxicity increased the percentage of cells in S phase, but higher parabactin concentrations consistently arrested cells in G1 phase or at the G1/S interface. MAb 42/6 also increased toxicity of parabactin to granulocyte/macrophage colony-stimulating factors and normal marrow granulocyte/macrophage progenitors. When HL60 or KB cells were treated with MAb 42/6 combined with Ga nitrate, MAb 42/6 increased cytotoxicity of Ga for HL60 cells but had little or no effect on Ga cytotoxicity to KB cells. In contrast, MAb 42/6 had minimal effects on cytotoxicity of the ribonucleotide reductase inhibitor, isoquinaldehyde thiosemicarbazone, to either HL60 or KB cells. Both hematopoietic and solid tumors were killed by Fe depletion, but the present studies suggested that hematopoietic cells are more sensitive than solid tumor cells to cytotoxic effects of Fe depletion. Combined Fe depletion therapy by the use of MAb 42/6 with an Fe chelator or Ga salt increased toxicity to MAb 42/6-sensitive cells, such as HL60, but was not more effective against MAb 42/6-resistant solid tumor cells. Combination Fe depletion therapy of hematopoietic cell tumors merits evaluation in experimental in vivo tumor systems.

Antibodies, Monoclonal↗
PubMed1989

Induction of c-fos by an anti-epidermal growth factor receptor monoclonal antibody.

The effect of epidermal growth factor (EGF) or a monoclonal antibody to the human EGF receptor on c-fos and c-myc gene expression was examined in KB epidermoid carcinoma cells. EGF increased c-fos transcripts within 30 min and c-myc transcripts within 6-24 hours. The antibody transiently increased c-fos expression within 30 minutes, but did not alter expression of c-myc. Cycloheximide potentiated the c-fos induction by both EGF and antibody. C-fos induction by antibody was less consistent, and was of briefer duration and lesser intensity than the induction by EGF. Nuclear transcription of c-fos decreased in cells treated with EGF or antibody, indicating that c-fos induction by EGF or antibody resulted from post-transcriptional mechanisms.

Antibodies, Monoclonal↗
PubMed1989

Cytokines in chronic inflammatory arthritis. II. Granulocyte-macrophage colony-stimulating factor in rheumatoid synovial effusions.

A liquid culture technique was used to study 23 synovial fluids (SF) (21 from inflammatory joint diseases and 2 noninflammatory SF) and supernatants of two cultured rheumatoid arthritis (RA) synovial tissues for colony-stimulating factor (CSF). The proliferative responses of human peripheral blood macrophage-depleted non-T cells treated with synovial fluids, supernatants of synovial tissue explants, and recombinant granulocyte-macrophage (rGM)-CSF were compared. Aggregates of cells that formed in long-term cultures (15 d) were similar for each applied agent and consisted of macrophages, eosinophils, and large blasts. Tritiated thymidine incorporation was proportional to the concentration of rGM-CSF and was accompanied by an increase in number and size of cellular aggregates formed in the cultures. CSF activity was observed in inflammatory SF, with tritiated thymidine uptake of 3,501 +/- 1,140 cpm in the presence of RA samples (n = 15) compared to 1,985 +/- 628 for non-RA inflammatory SF (n = 7) (P less than 0.05) and 583 +/- 525 for medium (n = 6) (P less than 0.01). The proliferative response to RA SF was often more apparent when the samples were diluted, because at higher concentrations the RA SF was inhibitory. Two RA SF were fractionated by Sephadex G100 column chromatography; low levels of CSF activity were detected in fractions corresponding to Mr of 70-100 kD, but the major CSF activity was found in the 20-24-kD fractions. A polyclonal rabbit anti-GM-CSF antibody eliminated the stimulating activity from both rGM-CSF and RA SF. Finally, a specific RIA identified significant levels of GM-CSF (40-140 U/ml) in the culture supernatants of 3 additional RA synovial tissues. These data document the local production of GM-CSF in rheumatoid synovitis and are the first description of this cytokine at a site of disease activity.

Arthritis↗
PubMed1988

Effects of anti-epidermal growth factor (EGF) receptor antibodies and an anti-EGF receptor recombinant-ricin A chain immunoconjugate on growth of human cells.

The effects of antiepidermal growth factor (EGF) receptor monoclonal antibodies (MAbs) 528 and 225 and a 528-ricin A conjugate on the growth of normal and malignant human cells were tested in vitro. Malignant human cell lines with EGF receptor numbers ranging from 0 to 4 X 10(5) receptors/cell, human fetal fibroblasts, and normal marrow granulocyte/macrophage progenitors (CFU-gm) showed no effect when grown with 10(-12) M to 10(-7) M MAb 225 or 528. MAbs 225 and 528 and EGF also had no effect on the ability of marrow stromal cells to maintain CFU-gm viability in long-term marrow cultures. Reversible growth inhibition of A431 epidermoid and MDA-468 breast carcinoma cells with 2 and 3 X 10(6) EGF receptors/cell, respectively, was observed with both antibodies and with 10(-8) M EGF. In contrast, an immunoconjugate prepared with MAb 528 and recombinant ricin A chain (528-rRA) showed dose-dependent killing over a concentration range of 10(-12) M to 10(-8) M against cells with greater than or equal to 1.2 X 10(5) EGF receptors/cell [concentration that causes 50% inhibition of growth (IC50) values, approximately 10(-12) M to 10(-10) M]. Human fetal fibroblasts (5.6 X 10(4) EGF receptors/cell), melanoma cells without detectable EGF receptors, and human CFU-gm showed IC50 values of greater than 10(-8) M. Killing of KB epidermoid carcinoma cells and 547 ovarian carcinoma cells with 4 and 1.2 X 10(5) EGF receptors/cell by 10(-10) or 10(-11) M 528-rRA was time dependent, but cytotoxicity to 547 cells was not complete even with 48 hours of immunotoxin treatment. Cytotoxicity of 528-rRA was not enhanced by chloroquine or verapamil. In vitro, anti-EGF receptor MAbs cause reversible antiproliferative effects only against malignant cell lines with amplified EGF receptor expression. In contrast, 528-rRA shows potent, specific toxicity to cells with greater than 50,000 EGF receptors/cell. However, kinetics of cell killing with 528-rRA are protracted, suggesting that prolonged exposure may be required for in vivo antitumor effects.

Antibodies, Monoclonal↗
PubMed1988

Uridine pharmacokinetics in cancer patients.

The availability of uridine can alter the sensitivity of tumor cells to antimetabolites such as N-phosphonacetyl-L-aspartic acid (PALA) and acivicin by virtue of the cell's ability to salvage preformed metabolites from its environment. We investigated the pharmacokinetics of physiologically relevant amounts of uridine in cancer patients in a pilot study to further our understanding of uridine metabolism in the human body. Four cancer patients, two males and two females, were given an i.v. bolus of a trace amount of radiolabeled uridine. The nucleoside disappeared from the plasma in a triphasic manner, with initial half-lives of 0.57 +/- 0.28 and 1.79 +/- 0.62 min and a terminal half-life of 17.5 +/- 7.3 min. The volume of distribution was 481 +/- 70 ml/kg, and the plasma uridine clearance was calculated to be 1.70 +/- 0.42 l/min. Simultaneous plasma and bone marrow uridine concentrations were measured in a separate group of seven healthy volunteers. The uridine concentration in plasma was 2.32 +/- 0.58 microM, and that in the bone marrow plasma was 10.44 +/- 5.06 microM. These results suggest a very rapid turnover of uridine in the plasma when the nucleoside is present at physiologic concentrations, and that there is a locally high concentration of uridine available for salvage in the bone marrow.

Adult↗
PubMed1988

Gamma-interferon modulates human monocyte/macrophage transferrin receptor expression.

Although circulating human monocytes do not express transferrin (Tf) receptors, cultured adherent blood cells display high-affinity Tf binding sites. In the present studies, effects of various cytokines and biologically active proteins on human monocyte/macrophage Tf receptors were investigated. After culture, Tf receptor expression by adherent blood cells was time dependent and plateaued by 7 days. The addition of interleukin-1 (IL-1), alpha-interferon (alpha-IFN), granulocyte/macrophage-colony stimulating factor (GM-CSF), or human IgG to macrophages cultured for 4 days did not alter Tf receptor expression. Fe-saturated, human Tf caused a significant, dose-dependent decrease in receptor expression. At a dose of 100 U/mL, gamma-interferon (gamma-IFN) significantly increased Tf receptor expression by macrophages cultured for 4 (230% +/- 51% of control) or 7 days (150% +/- 22%). Scatchard analyses showed increased binding sites but no change in receptor affinity. Northern and slot blot analysis of cellular mRNA from macrophages cultured for 4 to 7 days and exposed to gamma-IFN showed a two- to fivefold increase in Tf receptor mRNA, but less than or equal to 30% increase in beta-actin mRNA. Ferritin content of gamma-IFN-treated macrophages was 47% to 63% of control cells. Net uptake of 59Fe from Tf by gamma-IFN-treated cells was 10% to 17% of control, but uptake of radiolabeled Tf was comparable. When macrophages were labeled with 59Fe and then exposed to gamma-IFN, cell-associated Fe was reduced by 43%, indicating that gamma-IFN caused macrophage Fe release. gamma-IFN specifically modulates Tf receptor display by inducing Fe release and reducing cellular Fe content. Regulation of Tf receptor expression in macrophages is controlled by cellular Fe content and is thus similar to regulatory mechanisms in dividing cells.

Colony-Stimulating Factors↗
PubMed1988

Ricin binding and protein synthesis inhibition in human hematopoietic cell lines.

Previous studies showed that human blood cells exhibited varying sensitivities to ricin. To investigate the basis for these differences, ricin binding to human hematopoietic cell lines was assessed and correlated with in vitro ricin sensitivities. Resistant mutants were also isolated and characterized. Ricin binding to CEM cells was rapid, time-dependent, and blocked by unlabeled ricin, but not albumin; ricin binding approached saturation at 3 mumol/L. Scatchard analyses showed multiple classes of binding sites, with maximum and minimum Kd values estimated at 1.5 x 10(-8) mol/L and 2.5 x 10(-7) mol/L. At 4 degrees C, membrane-bound ricin dissociated slowly from the cell surface in the presence of unlabeled ricin, but greater than 95% of the surface-bound ricin was removed with 0.1 mol/L lactose. At 37 degrees C, ricin dissociated from the cell surface with biphasic kinetics. Ricin uptake at 37 degrees C increased linearly for 15 to 30 minutes and plateaued at levels representing 12% to 29% of the amount of ricin bound at 4 degrees C, depending on the cell line. Ricin binding at 4 degrees C varied two- to fivefold among hematopoietic cell lines and was reduced approximately tenfold by incubation with lactose. When compared with parent CEM cells, ricin-resistant CEM variants showed a greater than 95% reduction in ricin binding and showed no detectable binding with lactose added. However, these cells were as sensitive as parent CEM cells to an anti-T-cell ricin immunoconjugate. For all cells examined, there was a close correlation (r = +.9) between ricin bound per cell and in vitro ricin sensitivity. Human hematopoietic cells show widely varying ricin binding, indicating major differences in the carbohydrate content or structure of surface glycoproteins and glycolipids. These variations are probably the major determinant of nonspecific toxicity of ricin immunoconjugates.

Cell Line↗
PubMed1988

Effects of insulin and insulin-like growth factor I on growth of human leukemia cells in serum-free and protein-free medium.

Human myeloid leukemia cells (HL60) and malignant lymphocytes (Namalwa) were grown in protein-free, Fe-supplemented media and used to study growth responses to insulin and insulin-like growth factor 1 (IGF-I). HL60 cells previously grown in serum-free medium containing microgram quantities of insulin showed an 18-fold reduction in cumulative cell production when grown without insulin. However, the same cells showed reduced or absent growth stimulation with 1 to 100 ng/mL insulin or IGF-I for at least four days following insulin deprivation, indicating that culture conditions modified insulin and IGF-I responses. When the same cells were grown in Fe-supplemented, protein-free medium (RPMI-Fe), insulin and IGF-I caused dose-dependent stimulation of HL60 cell growth with half-maximal stimulation at nanogram concentrations. Namalwa cells grown in protein-free medium showed no response to either hormone. Radioligand binding showed the presence of insulin and IGF-I receptors on both HL60 and Namalwa cells grown in RPMI-Fe. HL60 cells grown in fetal bovine serum had higher, and cells grown with microgram quantities of insulin dramatically reduced, insulin binding. Competitive binding studies and cultures with anti-IGF-I receptor antibody showed insulin and IGF-I stimulated growth through their respective specific receptors. Both insulin and IGF-I stimulate growth of some cultured human leukemia cells, but the presence of insulin or IGF-I receptors alone does not predict growth responses. Culture conditions affect both cellular responses and ligand binding by these hormones and must be closely controlled to study growth responses.

Antibodies, Monoclonal↗
PubMed1987

Use of nude mouse xenografts as preclinical screens. Characterization of xenograft-derived melanoma cell lines.

Cell lines derived from human melanoma xenografts were characterized for surface markers, karyotype abnormalities, and in vitro drug sensitivity. Xenografts were established using metastatic explants from untreated patients and passaged in nude mice. Cell lines were readily established from melanoma xenografts, and formed colonies when plated in semisolid media. The lines expressed human melanoma-associated and other surface antigens, human lactate dehydrogenase (LDH) isoenzymes, and contained only human chromosomes. They failed to express murine histocompatibility determinants and were negative for murine viruses by mouse antibody production assay. Karyotypes showed abnormalities of chromosomes 3, 6, and 7 similar to other melanomas. In vitro chemosensitivity profiles were compared using cell line and xenograft colony-forming assays. Values were similar for the original xenografts and their cell lines. Xenograft-derived human melanoma lines resemble other melanoma cell lines and primary melanomas with respect to surface antigens and karyotype abnormalities, and are appropriate models for studying in vitro drug sensitivity. When used as a model for transition from solid tumor to cell line, these studies suggest cell lines closely mirror in vitro chemosensitivities of parent tumor cells. However, occasional, unpredictable changes in sensitivity to some drugs occurs during this transition.

Animals↗
PubMed1987

Preclinical evaluation of illudins as anticancer agents.

Illudins are low molecular weight natural products which were previously evaluated as anticancer drugs using rodent tumor models. In the present studies, we used in vitro cultures of human cancer cells to reevaluate their potential as anticancer agents. Using continuous exposure, Illudins S and M were cytotoxic to human leukemia cells at concentrations of 6-100 nM, but dihydroilludin M was 3 orders of magnitude less toxic, thus identifying a ketone site as a structural feature critical for cytotoxicity. Cytokinetic studies showed that illudin S caused a complete block at the G1-S phase interface of the cell cycle. Kinetics of inhibition of radiolabeled thymidine, uridine, and leucine incorporation suggested a primary effect on DNA synthesis. In colony and liquid culture assays, cell killing was time dependent but near maximal with a 2-h exposure. Myeloid and T-lymphocyte leukemia cells were most sensitive (50% inhibitory concentration, 6-11 nM), but B-cell leukemia/lymphoma, melanoma, and ovarian carcinoma cells were at least 10 times more resistant. Bone marrow granulocyte/macrophage progenitors showed intermediate sensitivity. Illudin S was equally effective against CEM T-lymphocyte leukemia cells expressing the multidrug resistance phenotype associated with Mr 180,000 glycoprotein and the parental cell line. CEM cells resistant to doxorubicin, epipodophyllotoxins, and 1-beta-D-arabinofuranosylcytosine showed only a 2-fold increased resistance to illudin S. Illudins are novel and potent cytotoxins which may be preferentially active against human myeloid and T-cell leukemias, including cells resistant to more conventional chemotherapeutic agents. The present studies illustrate the breadth of information which can be obtained on a new agent using present in vitro screening procedures and human cells.

Antineoplastic Agents↗
PubMed1987

Effects of monoclonal anti-transferrin receptor antibodies on in vitro growth of human solid tumor cells.

To determine whether anti-transferrin (Tf) receptor monoclonal antibodies might be useful in treatment of human solid tumors, in vitro effects of immunoglobulin A (42/6) and immunoglobulin G (B3/25) anti-Tf receptor antibodies on human solid tumor growth were examined. In colony and liquid cultures containing 10% serum, B3/25 did not inhibit growth of melanoma or ovarian carcinoma cell lines. 42/6 caused modest dose-dependent inhibition in colony cultures (maximum inhibition approximately 50%), and slowed growth of melanoma, ovarian carcinoma and epidermoid carcinoma cells in liquid culture. Inhibition was more pronounced in low (1%) serum, and was abrogated by 200 micrograms/ml iron-saturated Tf or 50 microM ferric nitriloacetate. All cells displayed high affinity Tf receptors (4-20 X 10(4)/cell). Cells grown in 1% serum and epidermoid carcinoma cells displayed more receptors, and susceptibility to 42/6 inhibition appeared related to higher receptor number. After culture with anti-Tf receptor antibodies, solid tumor cells showed a 57-93% reduction in surface Tf-binding sites. Tf uptake by cells grown for 24 h in B3/25 was approximately 50% of control, but was reduced to less than 10% of control with 42/6. Immunofluorescence staining of melanoma and HL60 promyelocytic leukemia cells suggested greater heterogeneity of Tf receptor display on melanoma than on leukemia cells. Previous studies showed 42/6 completely blocked blood cell Tf internalization and is a potent inhibitor of hemopoietic cell growth. In contrast, in solid tumor cells, inhibition of Tf uptake and growth inhibition are subtotal. Solid tumor resistance to 42/6 may be due in part to greater heterogeneity of Tf receptor display by proliferating cells. However, responses to iron-saturated Tf and ferric nitriloacetate in the presence of 42/6 also differ in hemopoietic and solid tumor cells, suggesting possible differences in Tf processing or iron growth requirements.

Antibodies, Monoclonal↗
PubMed1987

Effects of epidermal growth factor on transferrin receptor phosphorylation and surface expression in malignant epithelial cells.

The transferrin (Tf) receptor is a major transmembrane protein which provides iron for normal and malignant cell growth. Epidermal growth factor (EGF) has been reported to rapidly and transiently alter the number of surface Tf receptors in normal and transformed epithelial cells. To investigate mechanisms of EGF-induced changes in surface Tf display, EGF effects on surface Tf receptors were compared in two cell lines which differ in their number of EGF receptors and growth responses to EGF. In cloned A431 cells with high receptor numbers which are growth-inhibited by EGF, EGF caused a 50% decrease in Tf receptor expression after 30 min. In contrast, EGF induced a rapid, transitory increase (within 5 min) in the number of surface Tf receptors on KB carcinoma cells which returned to basal levels by 15 min. The observed changes in Tf receptor display were due to altered receptor distribution and not changes in ligand affinity or total cellular transferrin receptor pools. Anti-EGF receptor monoclonal antibody blocked effects of EGF on transferrin receptor expression. Since the antibody is internalized and causes EGF receptor down-regulation, effects on transferrin receptor expression were independent of these events. EGF-induced alterations in Tf receptor display occurred even when cells were pretreated with colchicine, suggesting that changes in surface Tf binding were not mediated by cytoskeletal components. Na orthovanadate, which mimics some early cellular effects of EGF, duplicated EGF's effects on A431 Tf receptors, but had no effect on KB cells, suggesting these responses occur by differing mechanisms. To determine whether EGF caused changes in Tf receptor phosphorylation, 32P-labelled Tf receptors were immunoprecipitated after EGF treatment. After exposure to EGF, A431 cells showed no change in Tf phosphorylation, but KB cells showed a transient, 6-fold increase in transferrin receptor phosphorylation on serine residues. In both A431 and KB cells, phorbol ester (PMA) also increased phosphorylation on transferrin receptors, but had little effect on surface Tf receptor expression. In malignant cell lines, EGE induces rapid, variable changes in transferrin receptor expression and phosphorylation which differ from the effects of PMA. These early responses to EGF appear to differ with the cell type and correlate poorly with alterations in Tf receptor phosphorylation. These results suggest Tf receptor phosphorylation does not regulate Tf receptor display in all cells.

Animals↗
PubMed1987

Monoclonal antibodies to purified ricin A-chain: production and properties.

Ricin A-chain was purified from native ricin using lactosyl-Sepharose. It was non-toxic to whole cells at a concentration of 1 microM yet nearly as effective as an equimolar concentration of ricin in blocking in vitro protein synthesis. Hybridomas secreting monoclonal antibodies to ricin A-chain were produced using the murine myeloma cell line NS-1. These anti-ricin A-chain antibodies cross-reacted with whole ricin but exhibited little cross-reactivity with purified ricin B-chain. Antibodies 2F2 and 2F5 both immunoprecipitated ricin A-chain. Both antibodies also precipitated ricin B-chain, as did the irrelevant control antibodies MOPC-21 and MPC-11. Pre-incubation of B-chain with 0.1 M galactose eliminated greater than 90% of precipitation by 2F2, MOPC-21 and MPC-11 but effected minimally precipitation by 2F5. Enzyme-linked immunosorbent assays using antibodies 2F2 and 2F5 to detect ricin A-chain in murine or human serum were linear between 40 and 800 ng ricin A-chain per ml. Anti-ricin A-chain antibodies 2F2 and 2F5 produced some inhibition of in vitro A-chain catalytic activity. Specific monoclonal antibodies to A-chain hemitoxin will be useful for characterization of functional hemitoxin domains, in in vitro assays for the stability of A-chain immunotoxins, and in characterizing the cellular internalization and processing of conjugates containing ricin or ricin A-chain.

Animals↗
PubMed1987

Regulation of transferrin receptor expression in myeloid leukemia cells.

Surface transferrin (Tf) receptors are displayed by cultured human hematopoietic cells and provide Fe required for cell growth. Cell cycle status, cell density in culture, exposure to Fe, and differentiation alter Tf receptor display by myeloid leukemia cells. To investigate mechanisms controlling Tf receptor expression, rates of receptor synthesis and steady-state mRNA levels were measured in HL60 promyelocytic cells grown in serum and serum-free media or after differentiation in response to dimethylsulfoxide (DMSO). Although surface binding sites were unchanged during the first three days in culture with serum or in serum-free media containing Tf, by the third day, rates of receptor biosynthesis and steady-state mRNA levels declined, consistent with cell density-dependent, receptor regulation. Cells grown with soluble Fe instead of Tf showed reduced Tf binding sites, rates of receptor synthesis, and Tf receptor mRNA. When cells grown with Fe were subcultured, Tf receptor mRNA levels increased within 15 minutes and peaked by 24 hours. This was followed by a decline in receptor and gene expression so that by three days cells grown in the presence of Fe expressed approximately four times fewer receptors and had half the rates of Tf receptor synthesis and mRNA levels of cells grown in serum or Tf. Cells treated with DMSO showed a rapid decline in surface receptors, receptor synthesis, and steady-state mRNA levels. Modulation of Tf receptor expression was not due to redistribution between the cell surface and an internal receptor pool. In each instance, concurrent assessment of N-ras transcripts showed that changes in Tf receptor mRNA levels were not due to generalized alterations in protein synthesis. Exposure of cells grown in Fe or treated with DMSO to cycloheximide did not alter Tf receptor mRNA levels, thereby suggesting that receptor expression was not regulated by posttranscriptional processes dependent on protein synthesis. Actinomycin D inhibition of Tf receptor mRNA was compatible with a transcript half-life of approximately 2.2 hours. Nuclear transcription studies showed reduced rates of Tf receptor transcription after culture with Fe or exposure to DMSO. The present studies show complex patterns of Tf receptor gene regulation in cultured myeloid leukemia cells and demonstrate that transcriptional regulation is a major mechanism controlling Tf receptor gene expression in response to Fe and differentiation.

Animals↗
PubMed1987

Use of nude mouse xenografts as preclinical drug screens: in vivo activity of established chemotherapeutic agents against melanoma and ovarian carcinoma xenografts.

To evaluate the utility of nude mouse xenografts as preclinical drug screens, the activity of ten established chemotherapeutic agents was evaluated against seven melanoma and three ovarian carcinoma xenografts. Xenografts were established using primary explants from patients who had not received chemotherapy and serially passaged as sc tumors in nude mice. In vivo drug activities for dactinomycin, carmustine, vinblastine, melphalan, amsacrine, cisplatin, bleomycin, mitomycin, doxorubicin, and etoposide were evaluated by 4 weekly ip injections of 10% less than LD10 doses. Plots of relative tumor growth versus time were nearly log-linear. Analysis of in vivo activity was performed using percent control growth (treated/control tumor volume) and by calculation of a novel growth delay index obtained by fitting growth curves to a quadratic regression model. Both modes of data analysis identified alkylating agents (melphalan, carmustine, and mitomycin) as the most active drugs against human melanomas. Melphalan, mitomycin, and cisplatin showed the greatest activity against ovarian xenografts. However, complete tumor regressions were noted only with melphalan, mitomycin, and cisplatin against a single ovarian tumor xenograft. Correlation analysis suggested xenograft tumor growth rate was an important determinant of drug response. These results suggest that preclinical, new-drug screening with melanoma xenografts would identify drugs such as alkylating agents as active, and may not provide an advantage over murine leukemia screens. However, screening with ovarian xenografts may more closely reflect clinical drug activity. Criteria for detecting active drugs in such systems are discussed.

Animals↗
PubMed1986

Use of nude mouse xenografts as preclinical drug screens. Further studies on in vitro growth of xenograft tumor colony-forming cells.

Previous studies suggested tumor colony-forming cells (CFC) grown from xenografts might be useful as a preclinical, in vitro drug screen. To further evaluate this possibility, eight melanoma and six ovarian carcinoma xenografts were established from untreated patients and tested for in vitro CFC growth. For each tumor, linear relationships between cells plated and colony (30 cells or greater than 75 micron diameter) and cluster (10-30 cells or 50-75 micron) growth were observed. All eight melanomas grew sufficient colonies (greater than or equal to 30) for in vitro drug assessment, although four required hypoxic (pO2 = 40) incubation to reliably attain this level of growth. Only one in six of the ovary xenografts consistently grew enough colonies, and growth was not significantly improved by hypoxic incubation, or addition of luteinizing hormone, follicle-stimulating hormone, or steroid hormones. Cloning efficiencies (colonies + clusters/cells plated) for tumors demonstrating adequate growth ranged from 0.01% to 0.3%. For most tumors, no direct relationship was observed between characteristics of xenograft tumors (size) or their resulting cell suspensions (viabilities, cell yield) and CFC growth. Cell suspensions were incubated with a 3 log concentration of nine established chemotherapeutic agents. Resulting dose-effect curves were linear and showed no plateaus of drug effect. Analyzing 447 in vitro drug trials on six melanomas and one ovarian carcinoma, interexperiment variability was high. Cell lines were established from three xenografts using a low concentration of fetal bovine serum (1%), and also examined for in vitro drug sensitivity. Using both liquid culture isotope incorporation and a colony-forming assay, drug sensitivity profiles for the cell lines were nearly identical to those for parent xenograft CFC. However, assays performed using the cell lines were more reproducible than those using xenograft tissue. The authors conclude that tumor CFC can be reliably grown from melanoma xenografts, but in vitro drug assays using these xenografts are poorly reproducible. The xenografts are a resource for establishing cell lines, and drug assays performed using these lines are highly reproducible. Similarities in drug sensitivity profiles for parent xenograft CFC and derived cell lines suggest that, despite poor reproducibility, repetitive assays using melanoma CFC accurately reflect some properties of cells which sustain tumor cell growth.

Animals↗
PubMed1986

[Interrelation between transferrin receptor expression and intracellular ferritin concentration in leukemia cells and normal marrow cells].

Serum-free cultures of HL60 promyelocytic leukemia cells and cultured fresh leukemia and normal marrow cells were used to investigate relationships between proliferation, transferrin receptor (TfR) display and intracellular ferritin (Fer). HL60 cells in serum- und Tf-free medium displayed 3 times less TfR than cells in serum or Tf containing medium. But Fer in Tf-independent cells was 50 times higher than Fer in serum- or Tf-supplemented cells. HL60 cells induced to differentiate by DMSO or vitamin D3 decreased TfR but increased Fer. Expression of TfR with fresh leukemia and normal marrow cells was less clear than in HL60 cells; DMSO or vitamin D3 induced differentiation was associated with a 10-fold Fer increase in leukemia cells and greater than 100-fold increase in marrow cells. TfR-expression and ferritin synthesis may be important events in cell differentiation and growth.

Bone Marrow Cells↗