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Keith Miller

Publications and source records attributed to Keith Miller.

At least 19 recordsLinked to original sources
PubMed2006

External quality assurance of HER2 fluorescence in situ hybridisation testing: results of a UK NEQAS pilot scheme.

BACKGROUND AND AIMS: Trastuzumab provides clinical benefit for advanced and early breast cancer patients whose tumours over-express or have gene amplification of the HER2 oncogene. The UK National External Quality Assessment Scheme (NEQAS) for immunohistochemical testing was established to assess and improve the quality of HER2 immunohistochemical testing. However, until recently, no provision was available for HER2 fluorescence in situ hybridisation (FISH) testing. A pilot scheme was set up to review the performance of FISH testing in clinical diagnostic laboratories. METHODS: FISH was performed in 6 reference and 31 participating laboratories using a cell line panel with known HER2 status. RESULTS: Using results from reference laboratories as a criterion for acceptable performance, 60% of all results returned by participants were appropriate and 78% either appropriate or acceptable. However, 22.4% of results returned were deemed inappropriate, including 13 cases (4.2%) where a misdiagnosis would have been made had these been clinical specimens. CONCLUSIONS: The results of three consecutive runs show that both reference laboratories and a proportion of routine clinical diagnostic (about 25%) centres can consistently achieve acceptable quality control of HER2 testing. Data from a significant proportion of participating laboratories show that further steps are required, including those taken via review of performance under schemes such as NEQAS, to improve quality of HER2 testing by FISH in the "real world".

Biomarkers, Tumor↗
PubMed2006

Ectopic prostatic tissue in the uterine cervix and vagina: report of a series with a detailed immunohistochemical analysis.

Prostatic tissue has rarely been described in the lower female genital tract. We describe 6 cases of ectopic prostatic tissue: 5 involving the cervix and 1 the vagina. The latter is the first reported example of benign prostatic tissue in the vagina. The age of the patients ranged from 21 to 65 years; and in all cases, the prostatic tissue was located within the cervical or vaginal stroma without involvement of the surface. In all cases, there were both glandular and squamous elements, which varied in prominence. In some cases, the squamous elements predominated to such an extent that the underlying glandular component was easily overlooked. In the glandular areas, a double cell layer of luminal and basal cells was focally apparent. There was little cytologic atypia or mitotic activity. Immunohistochemically, 3 of 6 cases were positive with prostate specific antigen (PSA) and all 6 cases marked with prostatic acid phosphatase (PSAP). In some of the positive cases, staining was focal. Positive staining with prostatic markers was confined to the glandular elements with no staining of the squamous areas. Immunohistochemical staining with the high molecular weight cytokeratin 34betaE12 highlighted the basal cell layer, which often extended into the center of the cellular islands, reminiscent of basal cell hyperplasia involving the prostate gland. All cases tested were CD10 positive (largely restricted to the basal cell layer), alpha-methylacyl-CoA racemase positive, and p16 negative. Estrogen receptor (ER) and progesterone receptor (PR) were negative in the glandular areas, but ER was positive in the squamous elements in all cases and PR was positive in 1 case. All cases tested were androgen receptor positive and exhibited a low MIB-1 proliferation index with only scattered positive nuclei. The presence of ectopic prostatic tissue in the lower female genital tract may be more common than is appreciated. Once the possibility is considered, the diagnosis is easily confirmed using immunohistochemistry, although staining with prostatic markers may be focal and PSA may be negative. Ectopic prostatic tissue in the lower female genital tract is almost certainly a benign condition, based on the morphology, including the presence of a double cell layer, although follow-up of larger numbers of cases is required. Possible theories of histogenesis include a developmental anomaly, metaplasia of preexisting endocervical glands, and derivation from mesonephric remnants.

Acid Phosphatase↗
PubMed2005

Antistaphylococcal activity of the novel cephalosporin CB-181963 (CAB-175).

OBJECTIVES: We examined the antistaphylococcal activity of the novel cephalosporin CB-181963 (formerly known as CAB-175), with emphasis on its microbiological activity and penicillin-binding protein specificities. METHODS: Using established procedures, we examined the activity of CB-181963 against methicillin-susceptible (MSSA) and methicillin-resistant (MRSA) strains of Staphylococcus aureus in both planktonic and biofilm culture. We also determined whether CB-181963 exhibited a post-antibiotic effect (PAE). A radioactive competition assay with (3)H-labelled benzylpenicillin was used to determine penicillin-binding protein (PBP) affinities of CB-181963, including binding to PBP2a from MRSA. The potential for emergence of CB-181963-resistant mutants in MSSA and MRSA strains was examined using plating procedures. RESULTS: CB-181963 showed excellent activity against MRSA strains resistant to other cephalosporins in both planktonic and biofilm cultures. However, in common with other cephalosporins it was unable to eradicate biofilms. CB-181963 had a short PAE compared with other beta-lactam antibiotics. CB-181963 retained activity against a strain expressing type A beta-lactamase and demonstrated affinity for PBP2a of MRSA. Mutants resistant to CB-181963 were not recovered in either MSSA or MRSA. CONCLUSIONS: CB-181963 is a potent antistaphylococcal agent with better activity against MRSA than other cephalosporins. The anti-MRSA activity is correlated with elevated binding to PBP2a. CB-181963 may have a role in the treatment of staphylococcal infections, including those caused by MRSA and in the prophylaxis of biofilm-associated MSSA and MRSA infections. However, because of its short PAE, CB-181963 may have to be administered more frequently than other beta-lactam antibiotics, or given via prolonged infusion.

Anti-Bacterial Agents↗
PubMed2005

Ecological Assessment of Substance-abuse Experiences (EASE): findings from a new instrument development pilot study.

A newly developed instrument that assesses a client's orientation to addiction or recovery communities using social context referents was pilot tested with a sample of 103 adults seeking treatment for substance abuse at outpatient and residential treatment facilities on the East Coast. Preliminary findings show promising subscale reliabilities, and suggest that drug- and recovery-related social identities are related to drug-use severity and drug-use concern; and drug-related attitudinal congruence between the treatment-seeker and family and treatment-seeker and other significant persons are related to intention to make behavioral changes in reducing substance abuse.

Adolescent↗
PubMed2004

Comparison of assays for detection of agents causing membrane damage in Staphylococcus aureus.

OBJECTIVES: To develop a novel beta-galactosidase leakage assay for Staphylococcus aureus and to evaluate this alongside other simple methods for detection of agents that cause membrane damage in staphylococci. METHODS: Using a PCR-based approach, a derivative of S. aureus RN4220 was constructed carrying the Escherichia coli lacZ gene under the control of the strong staphylococcal promoter, cap1A. Leakage of beta-galactosidase (BG) from this strain was examined after exposure for 10 min to various membrane-damaging agents at 4 x MIC, using a fluorescence assay and the substrate 4-methylumbelliferyl-beta-d-galactoside. Other assays for membrane damage involving protoplast lysis (PL), leakage of material absorbing at 260 nm (OD) and ATP release as well as the BacLight (BL) assay were carried out using established methods. RESULTS: All the assays, with the exception of the PL assay, detected membrane damage induced by cetyltrimethylammonium bromide, nisin, clofazimine and protegrin IB-367. However, the ability to detect membrane damage induced by these agents differed between the assay systems. The assays also varied considerably in their signal-to-noise ratio, with the ATP assay providing values for nisin approaching 100-fold that of the control. CONCLUSIONS: The PL assay is unsuitable for detection of membrane-damaging agents in S. aureus. The other assays, including the BG assay, detect membrane damage. The OD assay should be sufficient for most purposes since it is effective, rapid and cheap to perform. Studies requiring maximum sensitivity and discrimination should employ the ATP assay.

Anti-Bacterial Agents↗
PubMed2004

Anti-staphylococcal activity and mode of action of clofazimine.

OBJECTIVES: Infections caused by Staphylococcus aureus might be treated with agents whose primary indications are for other infections. Clofazimine, an established anti-mycobacterial drug, could be such a candidate. However, the anti-staphylococcal properties of clofazimine have not been fully described and its mode of action, possibly involving inhibition of both RNA polymerase and a membrane-located target, has not been explored in detail. We have now conducted experiments to address these issues. METHODS: Using established procedures, we examined the activity of clofazimine against a range of clinical isolates of S. aureus and determined whether it was bactericidal, exhibited a post-antibiotic effect (PAE), or interacted synergically with other agents. The potential for emergence of clofazimine-resistant mutants was also examined. Mode of action studies involved macromolecular synthesis assays, cross-screening against rifampicin-resistant mutants, susceptibility of RNA polymerase to clofazimine in vitro and several methods to detect drug-induced membrane damage. RESULTS: Clofazimine demonstrated good anti-staphylococcal activity encompassing MSSA, MRSA and GISA. It was bactericidal and resistant mutants could not be isolated. Clofazimine did not exhibit a PAE and failed to act synergically with other drugs. No evidence for specific inhibition of RNA polymerase was obtained. Clofazimine caused non-specific inhibition of DNA, RNA and protein synthesis, consistent with membrane-damaging activity that was detected in three independent assays for membrane disrupting agents. CONCLUSIONS: Clofazimine is a potent anti-staphylococcal agent. It appears to be a membrane-disrupting agent and does not inhibit RNA polymerase.

Anti-Infective Agents↗
PubMed2004

Escherichia coli mutators present an enhanced risk for emergence of antibiotic resistance during urinary tract infections.

Mutators may present an enhanced risk for the emergence of antibiotic resistance in bacteria during chemotherapy. Using Escherichia coli mutators as a model, we evaluated their ability to develop resistance to antibiotics routinely used for the treatment of urinary tract infections (UTIs). Under conditions that simulate therapeutic drug concentrations in humans, low-level resistance to trimethoprim, gentamicin, and cefotaxime emerged more frequently in mutators than normal strains. Resistance to trimethoprim in both cell types arose from a single point mutation in folA (Ile94-->Leu) and cefotaxime resistance resulted from loss of outer membrane porin OmpF. The mechanisms of gentamicin resistance could not be defined, but resistance did not result from mutations in ribosomal protein L6 (rplF). Although similar mechanisms of low-level antibiotic resistance probably arise in these strains, mutators are a risk factor because the increased generation of mutants with low-level resistance enhances the opportunity for subsequent emergence of high-level resistance.

Anti-Bacterial Agents↗
PubMed2004

Recent advances in operant conditioning technology: a versatile and affordable computerized touchscreen system.

We report the construction of a new operant chamber that incorporates modern computer, touchscreen, and display technologies. An LCD display was housed in the front wall of a lightweight Plexiglas chamber. An Apple eMac computer was used to present visual stimuli on the monitor and to control other chamber events. Responses to the stimuli were recorded using a transparent resistive-type touchscreen that overlaid the monitor. The resulting system is simple and inexpensive to construct but powerful and flexible enough to explore a broad range of issues in animal learning and behavior.

Animals↗
PubMed2003

Pyrazolo-[1,5-a]-1,3,5-triazine corticotropin-releasing factor (CRF) receptor ligands.

The syntheses and rat CRF receptor binding affinities of 'retro-pyrazolotriazine' corticotropin-releasing factor (CRF) ligands 4 are reported. Some have high affinity for rat CRF receptors (K(i)< or =10 nM). The data provide additional support for the hypothesis that it is possible to interchange isosteric cores with similar electronic properties in the design of high-affinity CRF receptor ligands, provided the peripheral pharmacophore elements are maintained in the same three-dimensional array.

Animals↗
PubMed2003

The role of mutators in the emergence of antibiotic-resistant bacteria.

Bacteria contain a number of error prevention and error correction systems that maintain genome stability. However, strains exhibiting elevated mutation frequencies have recently been reported amongst natural populations of pathogenic Escherichia coli, Salmonella enterica, Pseudomonas aeruginosa, Neisseria meningitidis, Helicobacter pylori and Streptococcus pneumoniae. The majority of naturally occurring, strong mutators contain defects in the methyl-directed mismatch repair (MMR) system, with mutations in mutS predominating. MMR-deficient strains possess superior genetic backgrounds for the selection of some antibiotic-resistance mutations since mutation frequencies up to 1000-fold higher than normal strains have been reported, and resistance levels achieved in mutators can be greater than those arising in non-mutator hosts. MMR is a major constraint to interspecies recombination events. Removal of this barrier, as in the case of MMR defective mutators, also enhances the frequency of horizontal gene transfer, which is an important mechanism of acquired drug resistance in bacteria. Permanent global mutator status is associated with loss of fitness as mutators accumulate deleterious mutations more frequently than non-mutators. Fitness limitations of mutators may be overcome simply by the high bacterial cell densities that can be achieved during acute infection or by the adoption of transient mutator status. Mutators are a risk factor during the treatment of bacterial infections as they appear to enhance the selection of mutants expressing high- and low-level antibiotic resistance and have the capacity to refine existing plasmid-located resistance determinants.

Animals↗
PubMed2003

Biological properties of novel antistaphylococcal quinoline-indole agents.

The antibacterial properties of novel quinoline-indole (QI) agents were examined. QI agents demonstrated potent bactericidal activities against Staphylococcus aureus, killing by lytic and nonlytic mechanisms. S. aureus mutants resistant to a lytic QI agent (SEP 155342) and a nonlytic QI agent (SEP 118843) arose at frequencies of 1.4 x 10(-9) and 1.2 x 10(-8), respectively, by selection at four times the MICs. Mutants resistant to QI agent SEP 155342 were unstable, but mutants resistant to QI agent SEP 118843 displayed stable resistance. Mutants resistant to QI agent SEP 118843 were not cross resistant to other inhibitors, including QI agent SEP 155342. Addition of QI agents SEP 118843 and SEP 155342 at four times the MIC caused nonspecific inhibition of several macromolecular biosynthetic pathways in S. aureus. Within 10 min, QI agents SEP 118843 and SEP 155342 both interfered with bacterial membrane integrity, as measured by uptake of propidium iodide. Agents from the two classes of the QI agents probably kill staphylococci by separate mechanisms which, nevertheless, both involve interference with cytoplasmic membrane function. Precise structure-activity relationships for the division of QI agents into two classes could not be determined. However, lytic activity was often associated with substitution of a basic amine at position 4 of the quinoline nucleus, whereas compounds with nonlytic activity usually contained an aromatic ring with or without a methoxy substituent at position 4. Nonlytic QI agents such as SEP 118843 may possess selective activity against the prokaryotic membrane since this compound failed to lyse mouse erythrocytes when it was added at a concentration equivalent to four times the MIC for S. aureus.

Animals↗
PubMed2002

Arylamidrazones as novel corticotropin releasing factor receptor antagonists.

The arylamidrazones have been found to be potent corticotropin releasing factor (CRF) receptor antagonists structurally distinct from previously reported CRF1 antagonists. Attempts to modify the arylamidrazone core suggested an important role for the anilino NH moiety. The right-hand-side 2-nitro feature in lead 1 could be replaced with substituents methyl, chloro, cyano, or trifluoromethyl with a 4- to 10-fold reduction in receptor binding. With appropriate left-hand-side modifications, this potency loss could be recovered.

Aniline Compounds↗