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C Patrick Reynolds

Publications and source records attributed to C Patrick Reynolds.

26 records · Page 2Linked to original sources
PubMed2003

Lytic bone lesions in human neuroblastoma xenograft involve osteoclast recruitment and are inhibited by bisphosphonate.

Neuroblastoma is the second most common solid tumor in childhood and frequently metastasizes to the bone marrow and the bone matrix. The mechanism involved in bone metastasis and destruction in neuroblastoma is poorly understood. Using a model of bone invasion in immunodeficient mice, we demonstrated that neuroblastoma cells recruited osteoclasts to generate osteolytic lesions and invade the bone matrix. In further support of a contributory role for osteoclasts in neuroblastoma bone invasion, we demonstrated that treatment with the bisphosphonate compound, ibandronate, significantly delayed the progression of osteolytic lesions. The data suggest that bisphosphonates may be clinically effective in the treatment of bone metastases in neuroblastoma.

Adrenal Glands↗
PubMed2003

Antagonism of buthionine sulfoximine cytotoxicity for human neuroblastoma cell lines by hypoxia is reversed by the bioreductive agent tirapazamine.

Relapse of neuroblastoma (NB) commonly occurs in hypoxic tissues. Buthionine sulfoximine (BSO), an inhibitor of glutathione (GSH) synthesis, is cytotoxic for NB cell lines in atmospheric oxygen (20% O(2)). Tirapazamine (TPZ) is a bioreductive agent that forms a toxic-free radical in hypoxia. We determined in four NB cell lines cytotoxicity using the DIMSCAN digital imaging fluorescence assay, glutathione (GSH) levels by the DTNB-GSSG reductase method, apoptosis, reactive oxygen species (ROS), and mitochondrial membrane potential (Delta psi(m)) by flow cytometry. Hypoxia (2% O(2)) antagonized BSO-mediated ROS, apoptosis, and cytotoxicity but not GSH depletion. TPZ synergistically enhanced BSO cytotoxicity in hypoxia for all four NB cell lines, achieving 2-4 logs of cell kill. BSO depleted GSH (8-42% of controls) in 20 and 2% O(2), whereas TPZ only decreased GSH in hypoxia. Maximal GSH depletion was induced by BSO + TPZ. N-acetylcysteine abrogated GSH depletion caused by TPZ but not by BSO. BSO increased ROS, decreased Delta psi(m), and caused apoptosis in 20% O(2) (but not in 2% O(2)). TPZ elevated ROS in 2% O(2) (but not in 20% O(2)), whereas BSO + TPZ increased ROS both in 20 and 2% O(2). In hypoxia, TPZ alone or TPZ + BSO caused an 80% decrease of Delta psi(m) at 24 h, preceding apoptosis in 74-86% of cells at 48 h. Thus, hypoxia significantly antagonizes BSO-mediated cytotoxicity for NB cell lines, but TPZ reversed the inhibition of BSO-mediated cytotoxicity in hypoxia, causing increased ROS, Delta psi(m) decrease, GSH depletion, apoptosis, and synergistic cytotoxicity. These data additionally define the role of ROS in BSO-mediated cytotoxicity and suggest that combining BSO with TPZ could have clinical activity against NB in hypoxic sites.

Antineoplastic Combined Chemotherapy Protocols↗
PubMed2003

Transduction of green fluorescent protein increased oxidative stress and enhanced sensitivity to cytotoxic drugs in neuroblastoma cell lines.

Green fluorescent protein (GFP) is employed as a selection marker for gene transduction and to track tumor cells. Transduction of enhanced GFP (eGFP) into human neuroblastoma cell lines via a lentiviral vector significantly sensitized CHLA-20 (wild-type and functional TP53), and to a lesser extent CHLA-90 cells (multidrug-resistant, mutant, and nonfunctional TP53) to carboplatin, doxorubicin, etoposide, or melphalan, relative to cells transduced using the cell surface antigen CD80 as a selection marker. Total glutathione (GSH) was significantly up-regulated (1.8- to 2.8-fold) after eGFP (but not CD80) transduction in cell lines with, but not in those lacking, functional p53. Cytotoxicity of GSH depletion by buthionine sulfoximine in CHLA-20 (but not in CHLA-20-eGFP) was diminished by hypoxia (2% O(2)). Thus, oxidative stress produced by GFP selects for cells with up-regulated GSH in a p53-dependent manner, and also enhanced the cytotoxicity of anticancer drugs in neuroblastoma cell lines. Our data suggest caution when employing GFP-transduced cells to assess drug sensitivity and that using a cell surface antigen as a selection marker for gene transduction may perturb cells less than GFP.

Antineoplastic Agents↗
PubMed2002

Ceramide signaling in fenretinide-induced endothelial cell apoptosis.

Stress stimuli can mediate apoptosis by generation of the lipid second messenger, ceramide. Herein we investigate the molecular mechanism of ceramide signaling in endothelial apoptosis induced by fenretinide (N-(4-hydroxyphenyl)retinamide (4-HPR)). 4-HPR, a synthetic derivative of retinoic acid that induces ceramide in tumor cell lines, has been shown to have antiangiogenic effects, but the molecular mechanism of these is largely unknown. We report that 4-HPR was cytotoxic to endothelial cells (50% cytotoxicity at 2.4 microm, 90% at 5.36 microm) and induced a caspase-dependent endothelial apoptosis. 4-HPR (5 microm) increased ceramide levels in endothelial cells 5.3-fold, and the increase in ceramide was required to achieve the apoptotic effect of 4-HPR. The 4-HPR-induced increase in ceramide was suppressed by inhibitors of ceramide synthesis, fumonisin B(1), myriocin, and l-cycloserine, and 4-HPR transiently activated serine palmitoyltransferase, demonstrating that 4-HPR induced de novo ceramide synthesis. Sphingomyelin levels were not altered by 4-HPR, and desipramine had no effect on ceramide level, suggesting that sphingomyelinase did not contribute to the 4-HPR-induced ceramide increase. Finally, the pancaspase inhibitor, t-butyloxycarbonyl-aspartyl[O-methyl]-fluoromethyl ketone, suppressed 4-HPR-mediated apoptosis but not ceramide accumulation, suggesting that ceramide is upstream of caspases. Our results provide the first evidence that increased ceramide biosynthesis is required for 4-HPR-induced endothelial apoptosis and present a molecular mechanism for its antiangiogenic effects.

Acyltransferases↗
PubMed2002

Antidisialoganglioside/granulocyte macrophage-colony-stimulating factor fusion protein facilitates neutrophil antibody-dependent cellular cytotoxicity and depends on FcgammaRII (CD32) and Mac-1 (CD11b/CD18) for enhanced effector cell adhesion and azurophil granule exocytosis.

Polymorphonuclear leukocytes (PMNs) mediate antibody-dependent cellular cytotoxicity (ADCC), which is increased by the addition of granulocyte-macrophage colony-stimulating factor (GM-CSF). We sought to determine whether PMN ADCC also would be increased by the addition of an antibody/GM-CSF fusion protein and whether this would be associated with the up-regulation and activation of Mac-1 (CD11b/CD18) and with azurophil granule exocytosis. ADCC against LA-N-1 human neuroblastoma cells was evaluated with 4-hour calcein acetoxymethyl ester (calcein-AM) microcytotoxicity assay, electron microscopy, and multi-parameter flow cytometry. With the calcein-AM assay, LA-N-1 cell survival was 10%, 55%, and 75% when PMN ADCC was mediated by the antidisialoganglioside (anti-GD2) immunocytokine hu14.18/GM-CSF, by monoclonal antibody (mAb) hu14.18 mixed with GM-CSF, and by hu14.18 alone. Function-blocking mAbs demonstrated that FcgammaRII and FcgammaRIII were required for ADCC with hu14.18 alone or mixed with GM-CSF, but that only FcgammaRII was required for ADCC with hu14.18/GM-CSF. ADCC mediated by hu14.18 and hu14.18/GM-CSF was Mac-1 dependent. Electron microscopy demonstrated the greatest PMN adhesion, spreading, and lysis of targets with hu14.18/GM-CSF. Monoclonal antibodies blocking Mac-1 function allowed the tethering of PMN to targets with hu14.18/GM-CSF but prevented adhesion, spreading, and cytolysis. Flow cytometry showed that hu14.18 with or without GM-CSF and hu14.18/GM-CSF all mediated Mac-1-dependent PMN-target cell conjugate formation but that GM-CSF was required for the highest expression and activation of Mac-1, as evidenced by the mAb24-defined beta(2)-integrin activation epitope. Hu14.18/GM-CSF induced the highest sustained azurophil granule exocytosis, almost exclusively in PMNs with activated Mac-1. Thus, hu14.18/GM-CSF facilitates PMN ADCC against neuroblastoma cells associated with FcgammaRII and Mac-1-dependent enhanced adhesion and degranulation.

Antibodies↗
PubMed2002

Targeted delivery in primary and metastatic brain tumors: summary report of the seventh annual meeting of the Blood-Brain Barrier Disruption Consortium.

The November 2000 NIH report of the Brain Tumor Progress Review Group identified delivering and targeting therapeutic agents as a priority in the treatment of malignant brain tumors. For this reason, the seventh annual Blood-Brain Barrier Disruption Consortium meeting, partially funded by an NIH R13 Grant, focused on recent advances in targeted delivery to the central nervous system, clinical trials for primary and metastatic brain tumors using enhanced chemotherapy delivery, and strategies to lessen the toxicities associated with dose intensive treatments, using thiols.

Antineoplastic Agents↗
PubMed2002

Testing of new agents in childhood cancer preclinical models: meeting summary.

A workshop on pediatric preclinical testing, sponsored by the National Cancer Institute and the Children's Oncology Group Phase 1 Consortium, was held on June 26-27, 2001 in Bethesda, Maryland. Drs. Peter Adamson, Peter Houghton, and Malcolm Smith organized and hosted the meeting. There were 20 participants from 12 institutions. The primary objectives of the workshop included: (a) development of a working inventory of available preclinical models (including human tumor xenografts in immunodeficient mice, transgenic and syngeneic tumors, and selected in vitro models), with a basic understanding of the strengths and weaknesses of each as possible components of a preclinical testing program; (b) identification of the key scientific issues related to establishment of a program for preclinical testing of new agents for their applicability to childhood cancers; and (c) identification of the key infrastructure requirements for a program for preclinical testing of new agents for their applicability to childhood cancers. This report is a synthesis of the workshop's presentations and discussions.

Animals↗