Phase I topotecan preparative regimen for high-risk neuroblastoma, high-grade glioma, and refractory/recurrent pediatric solid tumors

We evaluated the toxicity and maximum tolerated dose of topotecan in a novel myeloablative regimen as treatment for high-risk pediatric tumors. Patients received an assigned topotecan dosage in combination with fixed doses of carboplatin and thiotepa, followed by autologous hematopoietic stem cells infusion. Topotecan dose was escalated in cohorts of four patients until the maximum tolerated dose of topotecan was defined or until accrual of 30 patients. Pharmacokinetics of topotecan were examined, and event-free survival was estimated. We describe preliminary results following treatment of 25 pediatric patients with high-risk solid tumors.     

Detection of rare mutant alleles by restriction endonuclease-mediated selective-PCR: assay design and optimization

BACKGROUND: Restriction endonuclease-mediated selective (REMS)-PCR, allows detection of point mutations, deletions, and insertions. Reactions require concurrent activity of a restriction endonuclease (RE) and a DNA polymerase, both of which must be sufficiently thermostable to retain activity during thermocycling. The inclusion of the RE in REMS-PCR inhibits amplification of sequences containing the RE recognition site, thus producing selective amplification of sequences that lack the RE site. METHODS: Assays were used that allowed the selection of conditions that produce concurrent RE/DNA polymerase activity. The RE thermostability assay involved thermocycling a RE under various conditions and assessing residual cleavage activity at various time points. Conditions found to preserve RE activity during thermocyling were then tested for their compatibility with DNA polymerase-mediated PCR. RESULTS: A range of conditions that preserve activity of the RE BstNI over 30 cycles of PCR was identified. A subset of these conditions was subsequently found to mediate specific amplification using Taq DNA polymerase. These conditions were used to develop a REMS-PCR protocol for the detection of mutations at codon 12 of the K-ras gene. This protocol allowed the detection of 1 mutant allele in a background of 1000 wild-type alleles. The presence of primer sets for RE and PCR control amplicons provided unambiguous assessment of mutant status. CONCLUSION: Implementation of the assays described may facilitate development of REMS-PCR assays targeted to other loci associated with disease.     

Encounters between adolescents and general practice in Australia

Aim: To describe the nature of the encounters between adolescents and general practice in Australia. Methods: Data collected by the Bettering the Evaluation and Care of Health programme from 1998–2004 were analysed. Data for 10–14‐year‐old and 15–19‐year‐old males and females were compared with data for 25–29‐year‐olds. The outcome measures included: number of encounters compared with other age groups, reasons for encounter, problems managed, treatments prescribed and referrals made for key problems and types of consultations. Results: Adolescents have the lowest rate of encounter with general practice, compared with all other age groups. Respiratory, skin, musculoskeletal and unspecified (fever, injury, weakness) problems accounted for the great majority of reasons for encounter and problems managed. Management of mental health problems, preventive health care and health education were very infrequently managed problems. Standard surgery consultations were more common among adolescents than among young adults. Conclusions: Adolescents have a relatively low rate of encounter with general practice and the problems managed are primarily physical ailments. There is great scope to improve delivery of preventive health care and to increase management of mental health problems.     

Cochlear microvasculature in normal and damaged ears

The minute vessels of the cochlea were examined post mortem in normal guinea pigs and in others exposed to intense noise or treated with the ototoxic drugs gentamicin and quinine. The benzidine stain was used to display the capillary networks, and osmic acid for surface preparations, “thick” sections, and ultrathin sections for electron microscopic examination. The capillaries of the spiral ligament, i.e., the so-called arteriovenous anastomoses, do not show the characteristics of the muscular shunts described by Chambers and Zweifach. They are divided into two groups, the adstrial capillaries closely applied to the basal surface of the stria vascularis, and the poststrial capillaries which supply the tissue of the spiral ligament. Continuing beneath the floor of the external sulcus they form an alternating pattern with the root cells, and are surrounded by a specialized pericapillary tissue which resembles that found around the capillaries of the spiral prominence. These tissues are readily destroyed by gentamicin and other ototoxic aminoglycosidic antibiotics. Vasoconstriction of the outer and inner spiral vessels in response to noise and quinine appears to involve both contraction of pericytes and swelling of the endothelial cells. After prolonged exposure to noise or treatment with gentamicin, degeneration of capillaries with formation of intervascular strands and avascular channels is found in the suprastrial network of the spiral ligament. Similar capillary changes are seen in presbycusis, and in the deaf Dalmatian dog.     

Intravenous Cereport (RMP-7) Enhances Delivery of Hydrophilic Chemotherapeutics and Increases Survival in Rats with Metastatic Tumors in the Brain

Purpose. The following experiments determined whether intravenous infusions of Cereport enhance delivery of chemotherapeutics and prolong survival in rats with metastatic tumors in the brain. Methods. Autoradiography and scintillation were used to examine uptake of the lipophilic (paclitaxel and carmustine) and the hydrophilic (carboplatin) chemotherapeutic agents, as well as the large hydrophilic marker, 70 kDa dextran. Cereport was also tested in combination with the chemotherapeutic drugs carboplatin, vinorelbine, gemcitabine and carmustine to determine if Cereport could enhance the survival benefit beyond that provided by chemotherapy alone. Results. Cereport enhanced the uptake of carboplatin and dextran, but not paclitaxel or carmustine. The pattern of Cereport's uptake effect with carboplatin revealed that Cereport selectively increased the proportion of highly permeable regions. Survival was significantly enhanced when Cereport was combined with either carboplatin, vinorelbine, or gemcitabine, but not carmustine, compared to each chemotherapeutic agent alone. Conclusions. These data provide the first evidence that Cereport, or any receptor-mediated approach intended to enhance the permeability of the blood-brain tumor barrier, can increase the delivery hydrophilic drugs to metastatic tumors in the brain, increasing survival in tumor-bearing rats.     

How best to express oestrogen receptor activity

Oestrogen receptor (ER) activity, detected and expressed in a variety of ways, is important in breast cancer. Experience in Edinburgh (1973-1996) showed that [1] no single mode of expression was entirely satisfactory, [2] the probability of a good 'outcome' (prognosis or response to endocrine therapy) increased with increasing activity (either fmol ER sites/mg protein or per cent cells staining for ER). Thus the use of a single 'cut-off' should be avoided and activity quantified, or stratified into categories.     

Increased cell survival by inhibition of BRCA1 using an antisense approach in an estrogen responsive ovarian carcinoma cell line

We tested the hypothesis that BRCA1 may play a role in the regulation of ovarian tumor cell death as well as the inhibition of ovarian cell proliferation. Introduction of BRCA1 antisense retroviral constructs into BG-1 estrogen-dependent ovarian adenocarcinoma cells resulted in reduced BRCA1 expression. BRCA1 antisense pooled populations and derived subclones were able to proliferate in monolayer culture without estrogen, whereas control cells began to die after 10 days of estrogen deprivation. In addition, both populations and subclones of BRCA1 antisense infected cells demonstrated a growth advantage in monolayer culture in the presence of estrogen and were able to proliferate in monolayer culture without estrogen, while control cells did not. Furthermore, clonal studies demonstrated that reduced levels of BRCA1 protein correlated with growth in soft agar and greater tumor formation in nude mice in the absence of estrogen. These data suggest that reduction of BRCA1 protein in BG-1 ovarian adenocarcinoma cells may have an effect on cell survival during estrogen deprivation both in vitro and in vivo.