Biomedical subjects

W S Dalton

Publications and source records attributed to W S Dalton.

At least 19 recordsLinked to original sources

PubMed2003

The farnesyl transferase inhibitor, FTI-277, inhibits growth and induces apoptosis in drug-resistant myeloma tumor cells.

Mutations of the ras gene are among the most commonly identified transforming events in human cancers, including multiple myeloma. Farnesyltransferase inhibitors (FTI) were developed to prevent Ras processing and induce cancer cell death. Several FTIs are in phase II and one is in phase III clinical trials. Preclinically, most of the focus has been on solid tumors, and the effects of FTIs in multiple myeloma have not been investigated. In this study we examined the cytotoxic activity and inhibition of Ras processing in three myeloma cell lines with differing Ras mutation status. H929 cells with activated N-Ras were more sensitive to FTI-277 treatment than 8226 and U266 cells with activated K-Ras or wild-type Ras, respectively. A combination of FTI-277 and a geranylgeranyltransferase I inhibitor (GGTI)-2166 inhibited K-Ras processing and enhanced cell death in 8226 cells. U266 cells and Bcl-x(L) transfectants were equally sensitive to FTI-277 treatment. Similarly, 8226 cells selected for resistance to various chemotherapeutic agents, which resulted in either P-glycoprotein overexpression, altered topoisomerase II activity, or elevated glutathione levels, were equally sensitive to FTI-277. These preclinical studies suggest that prenylation inhibitors may represent new therapeutic agents for the treatment of refractory or drug-resistant multiple myeloma.

Alkyl and Aryl Transferases↗

PubMed2003

Bone marrow stromal-derived soluble factors and direct cell contact contribute to de novo drug resistance of myeloma cells by distinct mechanisms.

The tumor microenvironment plays a critical role in determining the fate of tumor cells. We have previously reported that adhesion of human myeloma and leukemia cell lines to the extracellular matrix protein, fibronectin, confers a multidrug-resistant phenotype. Mechanisms associated with this cell adhesion-mediated drug resistance are drug-type specific. In the present study, we examined the influence of bone marrow stromal cells (BMSCs) on myeloma cell response to the topoisomerase II inhibitor, mitoxantrone. Apoptosis was inhibited by more than 50% when cells were adhered to BMSCs as compared to myeloma cells maintained in suspension. To investigate the mechanisms contributing to the resistance of myeloma cells in contact with BMSCs, we examined the protective effects of BMSCs under four separate conditions: (1) direct cell contact; (2) BMSCs conditioned medium; (3) medium conditioned by coculturing myeloma cells in direct contact with BMSCs; and (4) medium conditioned by coculturing myeloma cells and BMSCs without direct physical contact. Conditioned medium from BMSCs alone was not sufficient to protect myeloma cells from drug-induced apoptosis; however, soluble factors produced during the myeloma-BMSCs interaction decreased the sensitivity of myeloma cells to mitoxantrone, suggesting a dynamic interaction between myeloma cells and BMSCs. We also found that myeloma cells in direct contact with BMSCs underwent growth arrest, whereas soluble factors produced by myeloma cells-BMSCs coincubation stimulated the proliferation of myeloma cells. These data show that both cell-cell adhesion of BMSCs with myeloma cells and soluble factors induced by this cell-cell interaction are involved in the protection of myeloma cells from mitoxantrone-induced apoptosis; however, the mechanisms contributing to the drug resistance are different.

ADP-ribosyl Cyclase↗

PubMed2001

Reduction in drug-induced DNA double-strand breaks associated with beta1 integrin-mediated adhesion correlates with drug resistance in U937 cells.

We previously showed that adhesion of myeloma cells to fibronectin (FN) by means of beta1 integrins causes resistance to certain cytotoxic drugs. The study described here found that adhesion of U937 human histiocytic lymphoma cells to FN provides a survival advantage with respect to damage induced by the topoisomerase (topo) II inhibitors mitoxantrone, doxorubicin, and etoposide. Apoptosis induced by a topo II inhibitor is thought to be initiated by DNA damage. The neutral comet assay was used to determine whether initial drug-induced DNA damage correlated with cellular-adhesion-mediated drug resistance. Cellular adhesion by means of beta1 integrins resulted in a 40% to 60% reduction in mitoxantrone- and etoposide-induced DNA double-strand breaks. When the mechanisms regulating the initial drug-induced DNA damage were examined, a beta1 integrin-mediated reduction in drug-induced DNA double-strand breaks was found to correlate with reduced topo II activity and decreased salt-extractable nuclear topo IIbeta protein levels. Confocal studies showed changes in the nuclear localization of topo IIbeta; however, alterations in the nuclear-to-cytoplasmic ratio of topo IIbeta in FN-adhered cells were not significantly different. Furthermore, after a high level of salt extraction of nuclear proteins, higher levels of topo IIbeta-associated DNA binding were observed in FN-adhered cells than in cells in suspension. Together, these data suggest that topo IIbeta is more tightly bound to the nucleus of FN-adhered cells. Thus, FN adhesion by means of beta1 integrins appears to protect U937 cells from initial drug-induced DNA damage by reducing topo II activity secondarily to alterations in the nuclear distribution of topo IIbeta.

Antineoplastic Agents↗

PubMed2001

Ascorbic acid enhances arsenic trioxide-induced cytotoxicity in multiple myeloma cells.

Multiple myeloma (MM) is a clonal B-cell malignancy characterized by slow-growing plasma cells in the bone marrow (BM). Patients with MM typically respond to initial chemotherapies; however, essentially all progress to a chemoresistant state. Factors that contribute to the chemorefractory phenotype include modulation of free radical scavenging, increased expression of drug efflux pumps, and changes in gene expression that allow escape from apoptotic signaling. Recent data indicate that arsenic trioxide (As(2)O(3)) induces remission of refractory acute promyelocytic leukemia and apoptosis of cell lines overexpressing Bcl-2 family members; therefore, it was hypothesized that chemorefractory MM cells would be sensitive to As(2)O(3). As(2)O(3) induced apoptosis in 4 human MM cell lines: 8226/S, 8226/Dox40, U266, and U266/Bcl-x(L). The addition of interleukin-6 had no effect on cell death. Glutathione (GSH) has been implicated as an inhibitor of As(2)O(3)-induced cell death either through conjugating As(2)O(3) or by sequestering reactive oxygen induced by As(2)O(3). Consistent with this possibility, increasing GSH levels with N-acetylcysteine attenuated As(2)O(3) cytotoxicity. Decreases in GSH have been associated with ascorbic acid (AA) metabolism. Clinically relevant doses of AA decreased GSH levels and potentiated As(2)O(3)-mediated cell death of all 4 MM cell lines. Similar results were obtained in freshly isolated human MM cells. In contrast, normal BM cells displayed little sensitivity to As(2)O(3) alone or in combination with AA. Together, these data suggest that As(2)O(3) and AA may be effective antineoplastic agents in refractory MM and that AA might be a useful adjuvant in GSH-sensitive therapies. (Blood. 2001;98:805-813)

Acetylcysteine↗

PubMed2001

Overexpression of a dominant-negative signal transducer and activator of transcription 3 variant in tumor cells leads to production of soluble factors that induce apoptosis and cell cycle arrest.

Gene therapy of B16 tumors with a dominant-negative signal transducer and activator of transcription (Stat3) variant, designated Stat3beta, results in inhibition of tumor growth and tumor regression. Although only 10-15% of the tumor cells are transfected in vivo, the Stat3beta-induced antitumor effect is associated with massive apoptosis of B16 tumor cells, indicative of a potent bystander effect. Here, we provide evidence that blocking Stat3 signaling in B16 cells results in release of soluble factors that are capable of inducing apoptosis and cell cycle arrest of nontransfected B16 cells. RNase protection assays using multi-template probes specific for key physiological regulators of apoptosis reveal that overexpression of Stat3beta in B16 tumor cells induces the expression of the apoptotic effector, tumor necrosis factor-related apoptosis-inducing ligand. These in vitro results suggest that the observed in vivo bystander effect leading to tumor cell growth inhibition is mediated, at least in part, by soluble factors produced as a result of overexpression of Stat3beta in tumor cells.

3T3 Cells↗

PubMed2001

Mechanisms associated with cell adhesion mediated drug resistance (CAM-DR) in hematopoietic malignancies.

The tumor microenvironment is often overlooked when considering tumor response to chemotherapeutic agents. This environment consists of soluble factors, components of the extracellular matrix as well as cell-cell interactions. Recently, it has become clear that cell-cell and cell-matrix interactions result in cytoskeletal reorganization and the activation of multiple signal transduction pathways that directly influence cell survival, growth and differentiation. Experimental evidence shows that anti-apoptotic pathways initiated by cell adhesion are operative in tumor cells and, furthermore, cause resistance to mechanistically distinct cytotoxics. For hematopoietic tumors, cell adhesion to a single matrix, fibronectin is sufficient to inhibit apoptosis induced by mechanistically distinct cyctotoxics. Adhesion of hematopoietic tumors to this matrix blocks cell cycle progression, and for the human multiple myeloma 8226 cell line adhesion to fibronectin resulted in increased p27kip1 levels, which correlated with cell cycle arrest and drug resistance. A decrease in initial DNA damage induced by topoisomerase II inhibitors has also been observed in adherent hematopoietic tumor cell lines. Further studies investigating the mechanisms of cell adhesion mediated drug resistance may reveal novel targets directed at the reversal of de novo drug resistance.

Antineoplastic Agents↗

PubMed2001

Cell adhesion-mediated drug resistance (CAM-DR) protects the K562 chronic myelogenous leukemia cell line from apoptosis induced by BCR/ABL inhibition, cytotoxic drugs, and gamma-irradiation.

Integrin-mediated cellular adhesion to extracellular matrix (ECM) components is an important determinant of chemotherapeutic response of human myeloma cells. Here, we demonstrate that when K562 chronic myelogenous leukemia (CML) cells are adhered to fibronectin (FN), they become resistant to apoptosis induced by the BCR/ABL inhibitors AG957 and STI-571, as well as DNA damaging agents and gamma-irradiation. This phenomenon, termed cell adhesion-mediated drug resistance (CAM-DR), was induced by adhesion through the alpha5beta1 (VLA-5) integrin. Phosphotyrosine analysis demonstrates that anti-apoptotic signaling through integrins in K562 cells is independent of the tyrosine kinases activated by BCR/ABL, with the possible exception of an unknown 80 kDa protein. Cytoprotection of FN-adhered CML cells indicates that tumor-ECM interactions may be critical for the emergence of drug-resistant tumor populations and treatment failure in this disease. Antagonists of beta1 integrin-mediated adhesion or corresponding signal transduction elements may sensitize CML cells to chemotherapy and prevent resistance to the novel BCR/ABL kinase inhibitors being used for the treatment of this disease.

Antineoplastic Agents↗

PubMed2001

Multiple myeloma.

Multiple myeloma (MM) is a malignancy of the plasma cell characterized by migration and localization to the bone marrow where cells then disseminate and facilitate the formation of bone lesions. Unfortunately, while treatment of this disease is effective in palliating the disease, and even prolonging survival, this disease is generally regarded as incurable. Understanding the basic biology of myeloma cells will ultimately lead to more effective treatments by developing target based therapy. In Section I, Dr. Bergsagel discusses the molecular pathogenesis of MM and shares insights regarding specific chromosomal translocations and their role in the genesis and progression of MM. New information regarding FGFR3 as an oncogene as well as how activating mutations may contribute to disease evolution and may be an important target for novel therapeutics of MM is presented. In Section II, Dr. Anderson elaborates on novel therapeutic approaches to MM also targeting fundamental genetic abnormalities in MM cells. Both preclinical and clinical studies of novel agents including PS-341 and IMiDs are highlighted. In Section III, Dr. Harousseau discusses the role of autologous stem cell transplant in MM. He highlights clinical trials addressing the question of conditioning regimens and the impact of tandem transplants. He also addresses the role of allogeneic BMT and the use of attenuated dose conditioning regimens (so called mini-allogeneic transplants) in the treatment of MM. In Section IV, Dr. Dalton provides an overview of the current state of myeloma therapy and summarizes the different and exciting approaches being undertaken to cure this disease.

Humans↗

PubMed2001

Inhibition of JAK kinase activity enhances Fas-mediated apoptosis but reduces cytotoxic activity of topoisomerase II inhibitors in U266 myeloma cells.

Our previous work demonstrated that the Janus kinase (JAK)-Stat3 pathway regulates expression of Bcl-x(L) in the U266 human multiple myeloma cell line and prevents Fas-mediated apoptosis. Inhibition of this pathway by the JAK selective kinase inhibitor AG490 or dominant-negative Stat3 protein results in down-regulation of Bcl-x(L) expression and enhanced sensitivity to Fas-mediated apoptosis. Because Bcl-x(L) has also been implicated in resistance to chemotherapeutic drugs, we investigated whether inhibition of the JAK-Stat3 pathway and subsequent reduction in Bcl-x(L) expression would also enhance cytotoxic drug activity. Contrary to this prediction, pretreatment of U266 myeloma cells with AG490, followed by exposure to topoisomerase II- inhibiting agents, antagonized drug-induced apoptosis. This effect correlated with reduced cyclin D1 expression and cell cycle arrest. The cell cycle arrest following AG490 pretreatment further correlated with reduced mitoxantrone-induced DNA double-strand breaks and reduced cell death, findings consistent with the critical requirement of DNA damage for drug cytotoxicity. These studies demonstrate that inhibition of the JAK-Stat3 pathway can result in paradoxical effects relative to cytotoxic drug response. These paradoxical responses may be explained by the findings that JAK-Stat3 signaling regulates the expression of multiple genes involved in controlling cell proliferation and apoptosis. Thus, understanding the cellular context of inhibiting signal transduction pathways is essential for the design of novel combination therapies for cancer.

Antineoplastic Agents↗

PubMed2001

Cell adhesion is a key determinant in de novo multidrug resistance (MDR): new targets for the prevention of acquired MDR.

Clinical circumvention of multidrug resistance (MDR) is a Sisyphian task faced in the treatment of many cancers. Identification of several mechanisms of acquired MDR has led to the development of chemosensitizing agents that counter specific mechanisms of MDR. Initial successes in therapy using "chemosensitizers" often culminate in relapse due to the multifactorial nature of acquired MDR. Therefore, it may be important to design therapeutic strategies that focus on mechanisms that allow for cell survival after initial treatments, before the acquisition of MDR. It has been proposed that extracellular effectors such as cytokines, matrix components, and adjacent cells may provide sanctuary to cancer cells by preventing stress-induced cell death. This review focuses on research implicating the cancer cell environment as a particularly important determinant in the emergence of drug resistance. More specifically, we will discuss the role of direct contact between cancer cells and the extracellular matrix or with adjacent cells as extrinsic effectors of de novo MDR. Cell adhesion has been demonstrated to prevent cell death through a number of mechanisms. Identification of cell adhesion-mediated drug resistance as an initial or de novo effector of MDR suggests that therapies targeting interactions between cancer cells and their environment may lead to the sensitization of cancer cells to chemotherapy or radiotherapy before the emergence of acquired mechanisms of MDR.

Animals↗

PubMed2000

Adhesion to fibronectin via beta1 integrins regulates p27kip1 levels and contributes to cell adhesion mediated drug resistance (CAM-DR).

The tumor cell environment may influence drug response through interactions with the extracellular matrix (ECM). We recently reported that adhesion of myeloma cells to fibronectin (FN) via beta1 integrins is associated with a cell adhesion mediated drug resistance (CAM-DR). Activation of beta1 integrins is known to influence both apoptosis and cell growth. We hypothesized that the FN mediated cytoprotection may be in part due to perturbations in cell cycle progression. In this report we demonstrate that adhesion of myeloma cells to FN results in a G1 arrest associated with increased p27kip1 protein levels and inhibition of cyclin A and E associated kinase activity. Disruption of cells from FN adhesion resulted in a rapid recruitment of cells into S phase, a decrease in p27kip1 levels, and reversion to a drug sensitive phenotype. Treatment of cells with p27Kip1 antisense oligonucleotides did not affect FN adhesion; however, p27Kip1 protein levels were reduced and cells became sensitive to cytotoxic drugs. These studies demonstrate that beta1 mediated adhesion of myeloma cells to FN regulates p27kip1 levels and that p27kip1 levels are causally related to CAM-DR. Disruption of beta1 integrin mediated FN adhesion may represent a potential target for the potentiation of drug induced apoptosis.

CDC2-CDC28 Kinases↗

PubMed2000

Monitoring the expression profiles of doxorubicin-induced and doxorubicin-resistant cancer cells by cDNA microarray.

Drug resistance in cancer is a major obstacle to successful chemotherapy. Cancer cells exposed to antitumor drugs may be directly induced to express a subset of genes that could confer resistance, thus allowing some cells to escape killing and form the relapsed resistant tumor. Alternatively, some cancer cells may be expressing an array of genes that could confer intrinsic resistance, and exposure to cytotoxic drugs select for the survival of these cells that form the relapsed tumor. We have used cDNA microarray to monitor the expression profiles of MCF-7 cells that are either transiently treated with doxorubicin or selected for resistance to doxorubicin. Our results showed that transient treatment with doxorubicin altered the expression of a diverse group of genes in a time-dependent manner. A subset of the induced genes was also found to be constitutively overexpressed in cells selected for resistance to doxorubicin. This distinct set of overlapping genes may represent the signature profile of doxorubicin-induced gene expression and resistance in cancer cells. Our studies demonstrate the feasibility of obtaining potential molecular profile or fingerprint of anticancer drugs in cancer cells by cDNA microarray, which might yield further insights into the mechanisms of drug resistance and suggest alternative methods of treatment.

Antineoplastic Agents↗

PubMed2000

Breast cancer resistance protein is localized at the plasma membrane in mitoxantrone- and topotecan-resistant cell lines.

Tumor cells may display a multidrug resistant phenotype by overexpression of ATP-binding cassette transporters such as multidrug resistance (MDRI) P-glycoprotein, multidrug resistance protein 1 (MRP1), and breast cancer resistance protein (BCRP). The presence of BCRP has thus far been reported solely using mRNA data. In this study, we describe a BCRP-specific monoclonal antibody, BXP-34, obtained from mice, immunized with mitoxantrone-resistant, BCRP mRNA-positive MCF-7 MR human breast cancer cells. BCRP was detected in BCRP-transfected cells and in several mitoxantrone- and topotecan-selected tumor cell sublines. Pronounced staining of the cell membranes showed that the transporter is mainly present at the plasma membrane. In a panel of human tumors, including primary tumors as well as drug-treated breast cancer and acute myeloid leukemia samples, BCRP was low or undetectable. Extended studies will be required to analyze the possible contribution of BCRP to clinical multidrug resistance.

ATP Binding Cassette Transporter, Subfamily G, Mem↗

PubMed2000

Clonal variability in CD95 expression is the major determinant in Fas-medicated, but not chemotherapy-medicated apoptosis in the RPMI 8226 multiple myeloma cell line.

CD95 (Fas/APO-1) is a member of the TNFR superfamily that induces apoptosis following cross-linking with its cognate ligand, CD95L (FasL/APO-1L) or agonist antibody. The human myeloma cell line, RPMI 8226, has limited sensitivity to CD95-mediated apoptosis, with a maximum of 65% of the population responding. To determine the source of the limited sensitivity to CD95-mediated apoptosis, we isolated multiple clones from the RPMI-8226 cell line by limiting dilution. Analysis of these clones demonstrated that sensitivity to CD95-mediated cell death directly correlated with CD95 expression. Clones with high levels of CD95 expression had greater than 90% cell death, whereas cells with low levels of expression had less than 10% cell death. In contrast, no correlative differences were identified for other members of the DISC complex, or for members of the anti-apoptotic Bcl-2 family. We further examined the sensitivity of the 8226 clones to various cytotoxic agents. Although modest clonal variability was demonstrated in response to the chemotherapeutic drugs, doxorubicin, etoposide (VP-16), and vincristine, there was no correlation between CD95 function and sensitivity to chemotherapeutic drugs. These results indicate that in this cell line, receptor expression is rate limiting in CD95-mediated apoptosis, whereas CD95 expression was not a determinant in drug-induced programmed cell death.

Antineoplastic Agents↗

PubMed2000

The tumor microenvironment as a determinant of cancer cell survival: a possible mechanism for de novo drug resistance.

The influence of the microenvironment in the pathogenesis and progression of human cancer has traditionally been considered in the context of solid tumors. More recently, evidence has been accumulating to support the role of the bone marrow microenvironment in hematologic malignancies as well, particularly in multiple myeloma. This review focuses on myeloma as a model to demonstrate that the bone marrow microenvironment provides a sanctuary against programmed cell death and promotes tumor cell survival and progression. Additionally, the protective effects of the bone marrow milieu may confer a protection from cytotoxic drugs, allowing the emergence of drug-resistant tumors. These advances may assist in the design of novel therapeutic approaches to enhance the efficacy of standard chemotherapeutic drugs.

Bone Marrow↗

PubMed2000

Alternative pathways of cell death to circumvent pleiotropic resistance in myeloma cells: role of cytotoxic T-lymphocytes.

Pleiotropic resistance to treatment remains one of the major reasons for therapeutic failures in patients with multiple myeloma. Myeloma cells are frequently resistant to physiological inducers of cell death prior to chemotherapy. Moreover, in the course of treatment cells acquire a multidrug resistant (MDR) phenotype, making eradication of the tumor even more difficult. A necessary prerequisite for circumventing complex pleiotropic resistance is therefore defining the signaling pathways that execute death in myeloma cells. This review discusses evidence that cytokine-expressing autologous tumor cell vaccine may be an efficient tool for elimination of both intrinsically resistant myeloma cells as well as cells with acquired MDR in murine models. The vaccine was similarly potent against wild type cells that were resistant to several death receptor ligands, and their isogenic sublines selected for P-glycoprotein-mediated MDR. The anti-myeloma effect of the vaccine was mediated by granzyme B/perforin-secreting cytotoxic T-lymphocytes. This is an example of therapeutic strategy directed at utilizing death pathways that are preserved in pleiotropically resistant tumor cells.

Cell Death↗

PubMed2000

Integrin-mediated drug resistance in multiple myeloma.

Drug resistance remains a major obstacle to the treatment of many hematopoietic malignancies such as multiple myeloma. Although much research has been focused on acquired resistance phenotypes, we believe that de novo drug resistance mechanisms may be an important component in protecting cells from initial drug exposure. It is now realized that many of the biological processes associated with this disease, including cell survival, may come as a result of the direct interactions of malignant plasma cells with the bone marrow microenvironment. This review examines the role of cell adhesion to one bone marrow component, fibronectin (FN), and the impact it may have on response to cytotoxic drugs. We discuss the influence of the integrin VLA-4 (alpha4beta1) on cell adhesion mediated drug resistance (CAM-DR) as well as the effects of chronic drug exposure on integrin function. Data presented here demonstrates that drug selection can make a non-adherent cell line adherent to FN through inside-out integrin activation and consequently cause a decrease in sensitivity to drug. We also speculate on the possible mediators of this intrinsic mechanism of drug resistance which may, along with the integrins themselves, become promising therapeutic targets in cancer treatment.

Cell Adhesion↗

PubMed2000

Considerations in the management of myeloma.

Multiple myeloma remains an incurable cancer. In recent years, progress in different drug classes has improved outcomes, but management has become more complicated. Areas such as prognostic classification, the increased use of high-dose chemotherapy with autologous stem-cell rescue, and a wider array of ancillary drugs must be integrated into recommendations for a consolidated treatment plan. Estimating prognosis is dependent on both clinical features and a growing list of laboratory tests. Autologous transplantation has been applied to an increasing proportion of patients, at different points in the disease process. Besides the age cut-off issue, there are still significant treatment choices to be made within the transplant technique. Newer drugs, most recently, thalidomide (Thalomid), may offer benefits independent of conventional cytotoxic drugs or steroids. Use of ancillary drugs, such as bisphosphonates, interferon, P-glycoprotein blockers, antibiotics, and growth factors, are also discussed. For the future, immunotherapy in the posttransplant setting appears promising. Ultimately, basic research must identify intracellular targets for the development of specific new-generation drugs.

Clinical Trials as Topic↗