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D Blakey

Publications and source records attributed to D Blakey.

At least 19 recordsLinked to original sources
PubMed2006

Report of the IWGT working group on strategies and interpretation of regulatory in vivo tests I. Increases in micronucleated bone marrow cells in rodents that do not indicate genotoxic hazards.

In vivo genotoxicity tests play a pivotal role in genotoxicity testing batteries. They are used both to determine if potential genotoxicity observed in vitro is realised in vivo and to detect any genotoxic carcinogens that are poorly detected in vitro. It is recognised that individual in vivo genotoxicity tests have limited sensitivity but good specificity. Thus, a positive result from the established in vivo assays is taken as strong evidence for genotoxic carcinogenicity of the compound tested. However, there is a growing body of evidence that compound-related disturbances in the physiology of the rodents used in these assays can result in increases in micronucleated cells in the bone marrow that are not related to the intrinsic genotoxicity of the compound under test. For rodent bone marrow or peripheral blood micronucleus tests, these disturbances include changes in core body temperature (hypothermia and hyperthermia) and increases in erythropoiesis following prior toxicity to erythroblasts or by direct stimulation of cell division in these cells. This paper reviews relevant data from the literature and also previously unpublished data obtained from a questionnaire devised by the IWGT working group. Regulatory implications of these findings are discussed and flow diagrams have been provided to aid in interpretation and decision-making when such changes in physiology are suspected.

Aniline Compounds↗
PubMed2006

Report of the IWGT working group on strategy/interpretation for regulatory in vivo tests II. Identification of in vivo-only positive compounds in the bone marrow micronucleus test.

A survey conducted as part of an International Workshop on Genotoxicity Testing (IWGT) has identified a number of compounds that appear to be more readily detected in vivo than in vitro. The reasons for this property varies from compound to compound and includes metabolic differences; the influence of gut flora; higher exposures in vivo compared to in vitro; effects on pharmacology, in particular folate depletion or receptor kinase inhibition. It is possible that at least some of these compounds are detectable in vitro if a specific in vitro test is chosen as part of the test battery, but the 'correct' choice of test may not always be obvious when testing a compound of unknown genotoxicity. It is noted that many of the compounds identified in this study interfere with cell cycle kinetics and this can result in either aneugenicity or chromosome breakage. A decision tree is outlined as a guide for the evaluation of compounds that appear to be genotoxic agents in vivo but not in vitro. The regulatory implications of these findings are discussed.

Animals↗
PubMed2005

Changes in practice of breast cancer radiotherapy 1998-2002: an Australasian survey.

In order to assess the impact on radiation oncology practice of the publication of evidence-based guidelines for technical aspects of therapeutic radiation for breast cancer, the Radiation Oncology Expert Advisory Group of the National Breast Cancer Centre conducted two postal surveys of radiation oncologists practising in Australia and New Zealand. Results from a survey conducted in 1998, prior to distribution of the guidelines, have been published previously. This article reports on results from a survey undertaken in 2002 and contains data from 102 respondents who manage women with breast cancer. The results show several important changes in practice since 1998, including increased use of CT scanning in breast cancer treatment planning and increased use of immobilization devices for patient treatment. There is also evidence of increased attention to technical aspects of treatment planning that reduce the potential risk of treatment toxicity. The influence of the guidelines, the wider availability of modern equipment and results from landmark clinical trials on change in radiation therapy practice is discussed.

Australia↗
PubMed2002

A phase I trial of antibody directed enzyme prodrug therapy (ADEPT) in patients with advanced colorectal carcinoma or other CEA producing tumours.

Antibody-directed enzyme prodrug therapy is a targeted therapy in which a prodrug is activated selectively at the tumour site by an enzyme, which has been targeted to the tumour by an antibody (antibody-enzyme conjugate). Previous clinical trials have shown evidence of tumour response, however, the activated drug had a long half-life, which resulted in dose-limiting myelosuppression. Also, the targeting system, although giving high tumour to blood ratios of antibody-enzyme conjugate (10 000 : 1) required administration of a clearing antibody in addition to the antibody-enzyme conjugate. The purpose of this current study therefore was to attempt tumour targeting of the antibody-enzyme conjugate without the clearing antibody, and to investigate a new prodrug (bis-iodo phenol mustard, ZD2767P) whose activated form is highly potent and has a short half-life. Twenty-seven patients were treated with antibody-directed enzyme prodrug therapy using A5CP antibody-enzyme conjugate and ZD2767P prodrug, in a dose-escalating phase I trial. The maximum tolerated dose of ZD2767P was reached at 15.5 mg m(-2)x three administrations with a serum carboxypeptidase G2 level of 0.05 U ml(-1). Myelosuppression limited dose escalation. Other toxicities were mild. Patients' quality of life was not adversely affected during the trial as assessed by the measures used. There were no clinical or radiological responses seen in the study, but three patients had stable disease at day 56. Human anti-mouse antibody and human anti-carboxypeptidase G2 antibody were produced in response to the antibody enzyme conjugate (A5CP). The antibody-enzyme conjugate localisation data (carboxypeptidase G2 enzyme levels by HPLC on tumour and normal tissue samples, and gamma camera analysis of I-131 radiolabelled conjugate) are consistent with inadequate tumour localisation (median tumour: normal tissue ratios of antibody-enzyme conjugate of less than 1). A clearance system is therefore desirable with this antibody-enzyme conjugate or a more efficient targeting system is required. ZD2767P was shown to clear rapidly from the circulation and activated drug was not measurable in the blood. ZD2767P has potential for use in future antibody-directed enzyme prodrug therapy systems.

Adult↗
PubMed2002

Isolex 300i CD34-selected cells to support multiple cycles of high-dose therapy.

BACKGROUND: We have previously reported that repeated cycles of high-dose therapy (HDT), can be supported by unmanipulated autologous PBPC. Here we investigate whether purified CD34+ cells, obtained by immunomagnetic separation using the Isolex 300i device, can support such therapy. METHODS: Twenty-nine consecutive patients with metastatic breast cancer had PBPC mobilized and harvested following chemotherapy and G-CSF (10 microg/kg per day). Patients with > 4.0 x 10(6)/kg CD34+ cells in the apheresis product underwent CD34-selection using the Isolex 300i (v2.0) device. All cells collected were equally divided into three aliquots and cryopreserved. Patients who did not achieve this threshold had unmanipulated cells collected and stored. Patients subsequently received three cycles of HDT with paclitaxel (175 mg/m2), thiotepa (300 mg/m2) and either ifosfamide (10 g/m2) or cyclophosphamide (4 g/m2). It was intended for patients to receive CD34-selected cells to support each of the three cycles of HDT (i.e 1/3 for each cycle) and to compare hemopoietic recovery between patients receiving CD34-selected cells or unmanipulated cells. RESULTS: Thirteen of the 29 patients (45%) did not mobilize sufficient CD34+ cells to undergo CD34-selection. The remaining 16 patients underwent CD34-selection with a median purity of 84.3% (range: 16.3-96.1%) and yield of 34% (range: 1-60%). Fifteen of these patients proceeded to HDT and 42 of the planned 45 cycles were administered. Nine patients had all three HDT cycles supported by CD34-selected cells. The median number of CD34-selected cells (x 10(6)/kg) infused per cycle was 1.5 (range: 0.04-3.01). Three of the 15 patients required infusion of 'back-up' unmanipulated cells because of delayed neutrophil recovery. Of the 13 patients whose PBPCs did not undergo CD34+ cell selection, 11 proceeded to HDT with a median of 3.2 x 10(6)/kg (range: 2.0-4.4) unselected cells infused per cycle and 31 of 33 planned cycles were delivered. When hemopoietic recovery was compared between cycles of HDT supported by CD34-selected (n = 34) and unmanipulated cells (n = 31), there was a modest slowing in the patients receiving CD34-selected cells; time to ANC > 1.0 x 10(9)/L = 11 days versus 10 days (P = 0.0122) and platelets > 20 x 10(9)/L = 14 days versus 13 days (P = 0.0009). No difference in recovery to 50 x 10(9)/L was observed (P = 0.54). CONCLUSION: We have demonstrated that Isolex 300i CD34-selected cells are capable of supporting multiple cycles of HDT. However, we were unable to acquire sufficient CD34+ cells to perform this processing in 45% (13/29) of patients and further improvements in yield are required to overcome the modest delay in neutrophil and platelet recovery.

Adult↗
PubMed2002

CliniMACS CD34-selected cells to support multiple cycles of high-dose therapy.

BACKGROUND: Traditionally, following high-dose therapy (HDT), unmanipulated autologous PBPC are infused. Alternatively, purified CD34+ cells can now be obtained by immunomagnetic separation using the CliniMACS device. Limited data currently exist examining hemopoietic recovery with such cells. METHODS: Ten patients with advanced breast cancer had PBPC mobilized with docetaxel (100 mg/m2) and G-CSF (10 microg/kg per day), harvested and processed using the CliniMACS CD34-selection device and equally divided into three aliquots for cryopreservation. Unmanipulated 'back-up' cells were also collected on a separate day of the same mobilization, divided into three and cryopreserved. Patients subsequently received three cycles of HDT with cyclophosphamide (4 g/m2), thiotepa (300 mg/m2) and paclitaxel (175 mg/m2). The intent was for patients to receive CD34-selected cells to support each of the three cycles of HDT (i.e., 1/3 for each cycle). If, however, hemopoietic recovery was delayed after Cycle 1, 1/3 of the unmanipulated cells were infused following Cycle 2 and the remaining CD34-selected cells (2/3) were used to support Cycle 3. RESULTS: PBPC from 10 patients underwent CD34-selection with a resulting median purity of 93% (range: 76-98%) and yield of 62% (range: 16-93%). Of the 10 patients, only two were able to be supported with CD34-selected cells for all three cycles of HDT. The remaining eight patients required unmanipulated 'back-up' cells to support Cycle 2. Three patients also required infusion of 'back-up' unmanipulated cells because of persistent neutropenia (n = 1) or thrombocytopenia (n = 2) following cycles initially supported by CD34-selected cells. The median number of CD34-selected cells (x 10(6)/kg) infused per cycle was 1.5 (0.7-2.6) (n = 20) and unselected cells was 1.7 (1.4-2.8) (n = 10). Comparing hemopoietic recovery between cycles of HDT supported by CD34-selected (n = 20) and unmanipulated cells (n = 10) there was a significant slowing with the CD34-selected cells; time to ANC > 1.0 = 13 days versus 10 days, platelets > 20 = 17 days versus 13 days, > 50 = 25 versus 17 days (all P values < 0.001). There was no correlation between the dose of CD34-selected cells infused and neutrophil/platelet recovery. DISCUSSION: We have demonstrated that, although unmanipulated PBPC achieve rapid hemopoietic recovery (at modest CD34 doses of < or = 2.8 x 10(6)/kg), CliniMACS-selected CD34+ cells (in the doses utilized in this study of < or = 2.6 x 10(6)/kg) result in significantly prolonged recovery.

Adult↗
PubMed2001

Breast radiotherapy: an Australasian survey of current treatment techniques.

Prior to the dissemination of evidence-based quality assurance guidelines, the Australian National Breast Cancer Centre Radiation Oncology Group conducted a process survey of breast radiotherapy treatment delivery throughout Australia. A process survey was conducted in August/September 1998. This survey comprised questions enquiring about treatment positioning, immobilization devices used, planning strategies, simulation and dose computation methods, treatment prescribing and quality assurance. The survey was sent to 123 Australian fellows of the Royal Australian and New Zealand College of Radiologists (RANZCR) and to the six directors of New Zealand radiation oncology departments. Fifty-eight questionnaires were returned of which 38 were received from individuals and 20 represented a reply from a department with a routine breast radiotherapy protocol (representing an average of 4.5 radiation oncologists per reply). The study identified great consistency between departments with respect to dose and fractionation for breast tangents. The study also identified some areas of treatment planning and delivery that varied between individuals or departments. These mainly reflected a lack of evidence in some areas of radiotherapy treatment delivery. The circulation of quality assurance guidelines will perhaps improve consistency of radiotherapy techniques in which studies have identified that technique changes improve outcome. This study identified that these areas include the taking of simulation and port films and the use of off-axis dosimetry. Further studies are required for areas of radiotherapy treatment delivery that have little evidence for or against their implementation.

Australia↗
PubMed2000

Docetaxel effectively mobilizes peripheral blood CD34+ cells.

We prospectively evaluated docetaxel (100 mg/m2) with G-CSF (10 microg/kg S.C., daily) for mobilization efficiency in 26 patients with breast cancer. The minimum target yield was >4.5 x 10(6) CD34+ cells/kg (optimum = 9 x 10(6)/kg), sufficient to support the subsequent three cycles of high-dose therapy (HDT). The peak days for peripheral blood (PB) CD34+ cells were day 8 and day 9. Seven collections began on day 7, 16 on day 8 and three on day 9. The median peripheral blood progenitor cell (PBPC) CD34+ cell content ranged from 1.2 to 5.9 x 10(6)/kg per day during days 7 to 11 with a median CD34+ content of the total 72 PBPC collections of 3.4 x 10(6)/kg (0.07-15.6). Fifteen patients obtained a PBPC collection exceeding 5 x 10(6)/kg on a single day of collection. Following a median 3 days collection for each patient (range 2-4), the median total CD34+ for all individual sets of collections was 9.7 x 10(6)/kg (range 1.0-28.4). We were able to achieve the minimum CD34+ cell target yield in 22 of 26 patients with one cycle of mobilisation chemotherapy and in two of these patients a second collection yielded sufficient cells. Twenty-two patients have subsequently received repetitive HDT and PBPC transplantation with 57 cycles of HDT having been delivered. For all 57 cycles, the median time to absolute neutrophil count (ANC) >0.5 x 10(9)/l and 1.0 x 10(9)/l was 10 days (range 8-22) and 11 days (range 8-23), respectively. The median time to platelets greater than 20 x 10(9)/l, 50 x 10(9)/l and 100 x 10(9)/l was 13 days (range 11-23), 17 days (range 12-53) and 23 days (range 18-70), respectively. We conclude that docetaxel with G-CSF effectively mobilises PBPCs with apheresis needing to be commenced approximately 8 days after docetaxel administration.

Adult↗
PubMed2000

Repetitive high-dose therapy with cyclophosphamide, thiotepa and docetaxel with peripheral blood progenitor cell and filgrastim support for metastatic and locally advanced breast cancer: results of a phase I study.

This phase I study was designed to determine the optimal dosages of a novel repetitive high-dose therapy regimen for patients with metastatic breast cancer (MBC). The planned treatment was three cycles of high-dose cyclophosphamide, thiotepa and docetaxel delivered every 35 days with progressive dose-escalation in successive cohorts. Each cycle was supported by peripheral blood progenitor cells (PBPC) and filgrastim. Eighteen patients were entered into this trial. Of the planned 54 treatment cycles, 44 were delivered and 11 patients completed all three cycles. The dose-limiting toxicities were interstitial pneumonitis and mucositis with moderately severe diarrhea (n = 3) and rash (n = 3). There were no treatment-related deaths. Of the 17 patients with evaluable disease, 16 patients responded with six patients achieving a complete remission and an additional four patients achieving no detectable disease (negative restaging including PET scan) but a persistently abnormal bone scan. At a median follow-up of 12 months, median progression-free survival was 11 months with the median overall survival not reached. The recommended doses for phase II/III studies are cyclophosphamide (4 g/m2), thiotepa (300 mg/m2) and docetaxel (100 mg/m2).

Adult↗
PubMed2000

A versatile system for receptor-mediated gene delivery permits increased entry of DNA into target cells, enhanced delivery to the nucleus and elevated rates of transgene expression.

We have developed a method for stabilisation of polyelectrolyte gene delivery vectors by crosslinking their surfaces with biodegradable multivalent copolymers based on N-(2-hydroxypropyl)methacrylamide (HPMA). The resulting nanoparticulate vectors resist attack by serum proteins and can be modified for cell-specific delivery by incorporation of targeting ligands onto the polymer coating. Here we show that vascular endothelial growth factor (VEGF), transferrin and basic fibroblast growth factor (bFGF) can each be linked to polyHPMA-coated poly(L-lysine)/DNA complexes. All ligand-targeted complexes demonstrated increased uptake into receptor-positive cells (measured using plasmids containing 32P-dCTP), that could be antagonised with excess free ligand. Targeted complexes also showed increased transfection, resistant to inhibition by serum, suggesting the possibility of effective application in vivo. Analysis using fluorescence microscopy confirmed enhanced uptake of ligand-targeted complexes (using Texas Red-labelled plasmid DNA), although VEGF- and transferrin-targeted complexes were restricted to cytoplasmic or perinuclear distributions. In contrast, bFGF-targeted complexes showed efficient delivery into the nucleus, with accumulation of more than 100000 plasmids per cell within distinct intranuclear compartments. This method permits versatile targeting of genes to selected cells and may also permit manipulation of intracellular trafficking. It should find several important applications in gene delivery systems both in vitro and in vivo.

Cell Nucleus↗
PubMed1999

Repetitive high-dose therapy with ifosfamide, thiotepa and paclitaxel with peripheral blood progenitor cell and filgrastim support for metastatic and locally advanced breast cancer: results of a phase I study.

BACKGROUND: This phase I study was designed to determine the optimal dosages of a novel repetitive high-dose therapy regimen for patients with metastatic breast cancer (MBC). PATIENTS AND METHODS: The planned treatment was three cycles of high-dose ifosfamide, thiotepa and conventional-dose paclitaxel delivered every 28 days with progressive dose-escalation in successive cohorts. Each cycle was supported by peripheral blood progenitor cells (PBPC) and filgrastim. RESULTS: Twenty-three patients were entered into this trial. Of the planned 69 treatment cycles, 59 were delivered and fifteen patients completed all three cycles. The dose-limiting toxicities were renal tubular acidosis, encephalopathy, mucositis and enterocolitis. There was one treatment-related hemorrhagic death. CONCLUSIONS: The recommended doses for phase II or III studies are ifosfamide (10 g/m2), thiotepa (350 mg/m2) and paclitaxel (175 mg/m2).

Adult↗
PubMed1998

Engineered human carboxypeptidase B enzymes that hydrolyse hippuryl-L-glutamic acid: reversed-polarity mutants.

Variants of human pancreatic carboxypeptidase B (HCPB), with specificity for hydrolysis of C-terminal glutamic acid and aspartic acid, were prepared by site-directed mutagenesis of the human gene and expressed in the periplasm of Escherichia coli. By changing residues in the lining of the S1' pocket of the enzyme, it was possible to reverse the substrate specificity to give variants able to hydrolyse prior to C-terminal acidic amino acid residues instead of the normal C-terminal basic residues. This was achieved by mutating Asp253 at the base of the S1' specificity pocket, which normally interacts with the basic side-chain of the substrate, to either Lys or Arg. The resulting enzymes had the desired reversed polarity and enzyme activity was improved significantly with further mutations at residue 251. The [G251T,D253K]HCPB double mutant was 100 times more active against hippuryl-L-glutamic acid (hipp-Glu) as substrate than was the single mutant, [D253K]HCPB. Triple mutants, containing additional changes at Ala248, had improved activity against hipp-Glu substrate when position 251 was Asn. These reversed-polarity mutants of a human enzyme have the potential to be used in antibody-directed enzyme prodrug therapy of cancer.

Alanine↗
PubMed1997

Radiotherapy treatment for isolated loco-regional recurrence of rectosigmoid cancer following definitive surgery: Peter MacCallum Cancer Institute experience, 1981-1990.

PURPOSE: To assess the success of external beam radiation treatment in the management of loco-regional recurrence of rectosigmoid cancer. METHODS AND MATERIALS: A retrospective analysis of 135 patients with locally recurrent rectosigmoid cancer presenting to Peter MacCallum Cancer Institute between January 1981 and December 1990 was undertaken. Patients were treated with three different dose ranges of radiotherapy: 50-60 Gy ("Radical" group), 45 Gy ("High-dose palliative" group), and <45 Gy ("Low-dose palliative" group). Symptomatic response rates and overall survival for each group were determined. RESULTS: Symptomatic response rates of 85, 81, and 56% were achieved in the radical, high-dose palliative, and low-dose palliative groups, respectively. Estimated median survival times were 17.9, 14.8, and 9.1 months for the radical, high-dose palliative, and low-dose palliative groups, respectively.

Adult↗
PubMed1996

Toward antibody-directed "abzyme" prodrug therapy, ADAPT: carbamate prodrug activation by a catalytic antibody and its in vitro application to human tumor cell killing.

Antibody-directed enzyme prodrug therapy, ADEPT, is a recent approach to targeted cancer chemotherapy intended to diminish the nonspecific toxicity associated with many commonly used chemotherapeutic agents. Most ADEPT systems incorporate a bacterial enzyme, and thus their potential is reduced because of the immunogenicity of that component of the conjugate. This limitation can be circumvented by the use of a catalytic antibody, which can be "humanized," in place of the bacterial enzyme catalyst. We have explored the scope of such antibody-directed "abzyme" prodrug therapy, ADAPT, to evaluate the potential for a repeatable targeted cancer chemotherapy. We report the production of a catalytic antibody that can hydrolyze the carbamate prodrug 4-[N,N-bis(2-chloroethyl)]aminophenyl-N-[(1S)-(1,3- dicarboxy)propyl]carbamate (1) to generate the corresponding cytotoxic nitrogen mustard (Km = 201 microM, kcat = 1.88 min-1). In vitro studies with this abzyme, EA11-D7, and prodrug 1 lead to a marked reduction in viability of cultured human colonic carcinoma (LoVo) cells relative to appropriate controls. In addition, we have found a good correlation between antibody catalysis as determined by this cytotoxicity assay in vitro and competitive binding studies of candidate abzymes to the truncated transition-state analogue ethyl 4-nitrophenylmethylphosphonate. This cell-kill assay heralds a general approach to direct and rapid screening of antibody libraries for catalysts.

Aniline Mustard↗
PubMed1996

Postoperative radiotherapy for Dukes' B and C rectal cancer: Peter MacCallum Cancer Institute experience.

This retrospective study reviews the outcome of patients with Dukes' B and C rectal cancer treated with adjuvant post-operative pelvic radiotherapy at the Peter MacCallum Cancer Institute from 1981 to 1990. Sixty-one patients (22 Dukes' B, 36 Dukes' C and 3 unknown stage) received a median dose of 50 Gy of pelvic irradiation. Locoregional relapse occurred in 33% of patients. Estimated median progression-free survival was 1.7 years with 46% surviving without progression at 2 years and 30% at 5 years. There was no difference according to Dukes' stage. The estimated median survival was 2.6 years, with no difference according to disease stage. These results with postoperative radiotherapy alone are inferior to results achievable by combination chemotherapy and radiotherapy as adjuvant therapy which should now be considered standard therapy following surgical resection for Dukes' B and C rectal cancer.

Adenocarcinoma↗
PubMed1995

Low-dose caudal morphine for postoperative analgesia in infants and children: a report of 500 cases.

STUDY OBJECTIVE: To determine the effectiveness of morphine 0.03 mg/kg or 0.04 mg/kg administered caudally to children for postoperative pain relief. DESIGN: Retrospective chart review. SETTING: University-affiliated hospital. PATIENTS: The charts of 500 children who had undergone various surgical procedures and who were given caudal morphine 0.03 mg/kg or 0.04 mg/kg either prior to the surgical procedure or immediately at the conclusion of the surgical procedure. MEASUREMENTS AND MAIN RESULTS: Parameters of respiratory rate, oxygen saturation, nausea and vomiting, voiding problems, and pruritus were recorded for each patient. There was no respiratory depression noted in the review of the 500 patients; 23% had nausea and vomiting, 3% had voiding problems needing bladder catheterization, and 7% reported pruritus, which was treated with either diphenhydramine or naloxone. CONCLUSION: Statistically there were no differences between morphine 0.03 mg/kg and morphine 0.04 mg/kg in all the study parameters. There was no respiratory depression in the 500 cases reviewed. The postoperative pain relief ranged from 6 hours to over 24 hours. This method of immediate postoperative pain management proved to be effective and safe.

Adolescent↗