Biomedical subjects

William A Brock

Publications and source records attributed to William A Brock.

7 recordsLinked to original sources

PubMed2004

Accumulation of the common mitochondrial DNA deletion induced by ionizing radiation.

Point mutations and deletions in mitochondrial DNA (mtDNA) accumulate as a result of oxidative stress, including ionizing radiation. As a result, dysfunctional mitochondria suffer from a decline in oxidative phosphorylation and increased release of superoxides and other reactive oxygen species (ROS). Through this mechanism, mitochondria have been implicated in a host of degenerative diseases. Associated with this type of damage, and serving as a marker of total mtDNA mutations and deletions, the accumulation of a specific 4977-bp deletion, known as the common deletion (Delta-mtDNA(4977)), takes place. The Delta-mtDNA(4977) has been reported to increase with age and during the progression of mitochondrial degeneration. The purpose of this study was to investigate whether ionizing radiation induces the formation of the common deletion in a variety of human cell lines and to determine if it is associated with cellular radiosensitivity. Cell lines used included eight normal human skin fibroblast lines, a radiosensitive non-transformed and an SV40 transformed ataxia telangiectasia (AT) homozygous fibroblast line, a Kearns Sayre Syndrome (KSS) line known to contain mitochondrial deletions, and five human tumor lines. The Delta-mtDNA(4977) was assessed by polymerase chain reaction (PCR). Significant levels of Delta-mtDNA(4977) accumulated 72 h after irradiation doses of 2, 5, 10 or 20 Gy in all of the normal lines with lower response in tumor cell lines, but the absolute amounts of the induced deletion were variable. There was no consistent dose-response relationship. SV40 transformed and non-transformed AT cell lines both showed significant induction of the deletion. However, the five tumor cell lines showed only a modest induction of the deletion, including the one line that was deficient in DNA damage repair. No relationship was found between sensitivity to radiation-induced deletions and sensitivity to cell killing by radiation.

Adolescent↗

PubMed2004

Radiosensitization of human and rodent cell lines by INO-1001, a novel inhibitor of poly(ADP-ribose) polymerase.

Inhibition of poly(ADP-ribose) polymerase (PARP) by a novel, potent inhibitor, INO-1001, was examined in two rodent and one human fibroblast cell lines, after single and fractionated radiation treatments. Since PARP plays a role in the early events following DNA damage and influences the effectiveness of DNA repair, its inhibition has been proposed to constitute a drug target for the development of novel radiosensitizers. We found that INO-1001 effectively inhibited PARP activity at non-cytotoxic concentrations. Combination treatment of 10 microM INO-1001 and a single dose of radiation resulted in significant radiosensitization of all three cells lines (enhancement ratios 1.4-1.6). This radioenhancement was even greater when the drug and radiation were given as fractionated treatments (enhancement ratio 8.0). Apoptosis (as evaluated by TUNEL staining) was not enhanced by the treatments, suggesting that inhibiting PARP enzyme activity by INO-1001 enhanced radiation-induced cell killing by interfering with DNA repair mechanisms, resulting in necrotic cell death. INO-1001 therefore, appears to have potential as a potent enhancer of radiation sensitivity, without any intrinsic cytotoxicity from the drug alone.

Animals↗

PubMed2004

Predicting radiosensitivity using DNA end-binding complex analysis.

Previous reports have suggested that measuring radiosensitivity of normal and tumor cells would have significant clinical relevance for the practice of radiation oncology. We hypothesized that radiosensitivity might be predicted by analyzing DNA end-binding complexes (DNA-EBCs), which form at DNA double-strand breaks, the most important cytotoxic lesion caused by radiation. To test this hypothesis, the DNA-EBC pattern of 21 primary human fibroblast cultures and 15 tumor cell lines were studied. DNA-EBC patterns were determined using a modified electrophoretic mobility shift assay and were correlated with radiosensitivity, as measured by SF2. DNA-EBC analysis identified a rapidly migrating ATM-containing band (identified as "band-A") of which the density correlated with SF2 (0.02 </= SF2 </= 0.41) in primary fibroblasts (r(2) = 0.77). The DNA-EBC pattern of peripheral blood lymphocytes was identical to that of fibroblasts. In addition, band-A density correlated with SF2 (0.35 </= SF2 </= 0.80) in 15 human tumor cell lines (r(2) = 0.91). Densitometry of other bands, or total DNA-EBC binding, correlated more poorly with SF2 (r(2) < 0.45). These data indicate that DNA-EBC analysis may be a practical, clinically relevant predictor of tumor and primary cell radiosensitivity.

Cell Line↗

PubMed2004

Radiosensitivity is predicted by DNA end-binding complex density, but not by nuclear levels of band components.

BACKGROUND AND PURPOSE: We previously determined that the density of a rapidly migrating DNA end-binding complex (termed 'band-A') predicts radiosensitivity of human normal and tumor cells. The goal of this study was first to identify the protein components of band-A and to determine if the protein levels of band-A components would correlate with band-A density and radiosensitivity. PATIENTS AND METHODS: DNA end-binding protein complex (DNA-EBC) protein components were identified by adding antibodies specific for a variety of DNA repair-associated proteins to the DNA-EBC reaction and then noting which antibodies super-shifted various DNA-EBC bands. Band-A levels were correlated with SF2 for a panel of primary human fibroblasts heterozygous for sequence-proven mutations in BRCA1 or BRCA2. The nuclear protein levels of band-A components were determined in each BRCA1 heterozygote by western hybridization. RESULTS: DNA-EBC analysis of human nuclear proteins revealed 10 identifiable bands. The density of the most rapidly migrating DNA-EBC band correlated closely with both BRCA-mutation status and radiosensitivity (r(2)=0.85). This band was absent in cells with homozygous mutations in their ataxia-telangiectasia-mutated protein (ATM) genes. This band was also completely supershifted by the addition of antibodies to ATM, Ku70, DNA ligase III, Rpa32, Rpa14, DNA ligase IV, XRCC4, WRN, BLM, RAD51 and p53. However, the intranuclear concentrations of these proteins did not correlate with either the SF2 or DNA-EBC density. Neither BRCA1 or BRCA2 could be detected in band-A. CONCLUSIONS: DNA-EBC analysis of human nuclear extracts resulted in 10 bands, at least six of which contained ATM. The density of one of the DNA-EBCs predicted the radiosensitization caused by BRCA haploinsufficiency, and this band contains Ku70, ATM, DNA ligase III, Rpa32, Rpa14, DNA ligase IV, XRCC4, WRN, BLM, RAD51 and p53 but not BRCA 1 or 2. The density of band-A was independent of the nuclear concentration of any of its known component.

BRCA1 Protein↗

PubMed2003

Chromosomal fragility syndrome and family history of radiosensitivity as indicators for radiotherapy dose modification.

Beside a few known radiosensitive syndromes, a patient's reaction to radiotherapy is difficult to predict. In this report we describe the management of a pediatric cancer patient presented with a family history of radiosensitivity and cancer proneness. Laboratory investigations revealed a chromosomal fragility syndrome and an increased cellular radiosensitivity in vitro. AT gene sequencing revealed no mutations. The patient was treated with reduced radiation doses to avoid the presumed increased risks of toxicity to normal tissues. The patient tolerated well the treatment with no significant acute or late radiation sequelae. Five years later, the patient remains both disease and complications free. While an accurate laboratory test for radiosensitivity is still lacking, assessments of chromosomal fragility, cell survival and clinical medicine will continue to be useful for a small number of patients.

Cerebellar Neoplasms↗

PubMed2002

Evidence of haplotype insufficiency in human cells containing a germline mutation in BRCA1 or BRCA2.

The BRCA1 and BRCA2 gene products are thought to play important roles in the processing of DNA damage. To assess whether heterozygous mutations in these genes are associated with cellular radiosensitivity, we performed an in vitro radiation clonogenic survival assay on dermal fibroblasts obtained from 8 sequence-proven BRCA heterozygotes (6 BRCA1, 2 BRCA2). These data were compared to results obtained from a previous set of 17 prospectively studied cancer patients who had a negligible risk for a BRCA mutation. In addition, results from radiation-induced chromatid break assay performed on lymphocytes obtained from 9 BRCA heterozygotes (8 BRCA1, 1 BRCA2) were compared to results from a control group of 18 women with no cancer history. Results from both assays suggested that cells containing a heterozygous mutation in BRCA1 or BRCA2 were more radiosensitive than controls. For the fibroblast studies, the mean surviving fraction at 2 Gy (SF2) for carriers was 0.279 vs. 0.348 for the control set (p = 0.007). For the lymphocyte studies, the mean number of chromatid breaks after 125 cGy of radiation was 0.79 breaks per cell for the carriers vs. 0.45 for the controls (p = 0.0005). There was no apparent difference in the radiosensitivity between cells with BRCA1 vs. BRCA2 mutations (p = 0.769), although the small sample size minimizes the certainty of this observation. These preliminary results are consistent with a relationship between a germline mutation in BRCA1 or BRCA2 and a hypersensitivity to radiation. This phenotype could possibly predispose to an increased risk of radiation-induced mutagenesis and carcinogenesis.

BRCA1 Protein↗

PubMed2002

Biofeedback therapy expedites the resolution of reflux in older children.

PURPOSE: In older children the spontaneous resolution rate of low grade vesicoureteral reflux is low and currently its management is controversial in regard to surgery versus prophylaxis versus observation alone. Bladder dysfunction in children with neurogenic bladders and to a less declarative degree in neurologically intact children has a role in the etiology or persistence of reflux. We determine the impact of biofeedback therapy on neurologically intact children with vesicoureteral reflux and detrusor-sphincter dyssynergia. MATERIALS AND METHODS: Vesicoureteral reflux was detected by voiding cystourethrography in children evaluated for urinary tract infections. Children with breakthrough infections or dysfunctional voiding based on history underwent uroflowmetry with concomitant patch electromyography of the external sphincter. Dyssynergia was defined as increased or steady electromyography activity during micturition. Biofeedback was initially performed weekly and the interval increased as indicated. The goals were to eliminate dyssynergia and reduce or eliminate post-void residual urine. Voiding cystourethrography was performed 1 year later to determine the status of the reflux. Ureteral reimplantation was performed during the period of biofeedback when indicated. RESULTS: From February 1997 to March 2001, 25 children 6 to 10 years old (mean age 9) with vesicoureteral reflux and detrusor-sphincter dyssynergia were treated with biofeedback therapy. There were 31 units (5 bilateral) with reflux, which was grade I in 10, II in 15, III in 5 and IV in 1. Children underwent an average of 7 sessions of biofeedback (range 2 to 20). On followup voiding cystourethrography, vesicoureteral reflux resolved in 17 units (55%), grade improved in 5 (16%) and reflux remained unchanged in 9 (29%). All cured vesicoureteral reflux was grade I (8 cases) or II (9). Four children (5 renal units) underwent reimplantation. In cured children there were no breakthrough infections during or since therapy and post-void residual urine decreased from an average of 40% before to 10% after therapy. Symptoms of urgency, daytime wetting and hoarding of urine improved or were eliminated in all children with resolved vesicoureteral reflux. CONCLUSIONS: Treating external detrusor-sphincter dyssynergia in older children with low grade vesicoureteral reflux, with biofeedback results in 1-year resolution rates that are considerably greater than historical resolution rates. External detrusor-sphincter dyssynergia should be screened for in children when surgery or discontinuation of chemoprophylaxis is considered so that biofeedback can be started.

Biofeedback, Psychology↗