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Biomedical subjects

T J Dougherty

Publications and source records attributed to T J Dougherty.

At least 19 recordsLinked to original sources
PubMed1992

Photodynamic therapy.

Photodynamic therapy (PDT) has been developed over the past decade into a useful treatment for several types of solid cancers in man. This unique therapy requires a photosensitiser accumulated in tumours and local activation by visible light generally delivered from lasers and delivered to the patient through various types of fibers and endoscopes. PDT appears to be most effective in treating certain superficial, difficult to treat cancers such as carcinoma in situ of the urinary bladder (here complete control is the intent), but also is effectively used in bulkier tumours obstructing bronchi or the oesophagus where palliation can be achieved. The primary mechanism of action is the in situ generation of an active form of molecular oxygen (singlet oxygen) which causes the rapid, local onset of vascular stasis and eventual vascular haemorrhage and tumour wall destruction. This process appears to be mediated through various cytokines such as prostaglandin, lymphokines and thromboxanes. The ultimate clinical value of PDT will be seen over the next few years following health agency approval worldwide.

Carcinoma, Non-Small-Cell Lung↗
PubMed1992

Cloning and identification of the Escherichia coli murB DNA sequence, which encodes UDP-N-acetylenolpyruvoylglucosamine reductase.

The murB gene, which complemented the UDP-N-acetylenolpyruvoylglucosamine reductase (EC 1.1.1.158) mutation in Escherichia coli ST5, was cloned from an E. coli chromosomal library. murB was subcloned on a 2.8-kb PvuII fragment into pUC19 and sequenced. A 1,029-bp open reading frame encoded a 342-amino-acid polypeptide of 37,859 Da. A DNA sequence homology search revealed that murB had almost 100% homology with a previously reported unidentified open reading frame, ORFII, at 89.9 min. Physical and genetic mapping results were consistent with this map position, and minicell analyses of murB subclones showed a plasmid-encoded protein of approximately 37,000 Da, which closely matched the calculated size of the murB protein.

Carbohydrate Dehydrogenases↗
PubMed1992

Photodynamic therapy for the treatment of basal cell carcinoma.

BACKGROUND: Photodynamic therapy is an investigational method for the treatment of a variety of solid tumors. The purpose of this study was to determine the optimum factors and illustrate the effectiveness of photodynamic therapy in the treatment of basal cell carcinomas. This was a prospective study in which patients presenting with primary or recurrent basal cell tumors, particularly but not exclusively widespread tumors or large single tumors, were offered the option of photodynamic therapy in their treatment regimen. RESULTS: Patients were administered 1 mg/kg of a photosensitizer (Photofrin II). Light doses (630 nm) ranged from 72 to 288 J/cm2. A total of 37 patients with 151 sites were treated in this study. A complete response rate of 88% was achieved with one application. Morbidity was low; the most significant side effects were moderate pain and edema. CONCLUSIONS: Photodynamic therapy is a modality that offers localized treatment of primary or recurrent nonmelanoma skin cancer. By applying reciprocal doses of photosensitizer and light, the efficacy of photodynamic therapy in the treatment of skin lesions is demonstrated achieving significant light penetration into tissue with a high complete response rate of the lesions and acceptable normal tissue response.

Basal Cell Carcinoma↗
PubMed1991

Substitution of lysine at position 104 or 240 of TEM-1pTZ18R beta-lactamase enhances the effect of serine-164 substitution on hydrolysis or affinity for cephalosporins and the monobactam aztreonam.

By site-directed mutagenesis, TEM-1 beta-lactamase was altered to contain single amino acid changes of E104K, R164S, and E240K, in addition to double changes of E104K/R164S or R164S/E240K and the triple change of E104K/R164S/E240K. Hydrolysis rates for cephaloridine and benzylpenicillin were lowered at least 1 order of magnitude for all enzymes containing R164S substitutions. All mutant enzymes exhibited increased kcat values for beta-lactam antibiotics containing an aminothiazole oxime side chain. Hydrolysis of ceftazidime was most affected, with kcat values increased 3-4 orders of magnitude in all enzymes with the substituted R164S moiety. Km values decreased for all substrates except ceftazidime in the enzymes with multiple mutations. Aztreonam was most affected, with Km values lowered 23-56-fold in the enzymes bearing multiple mutations. When the crystal structures of aztreonam and related monobactams were studied and projected into an active-site model of the PC1 beta-lactamase, it became apparent that the two lysine residues might serve equivalent roles by interacting with the carboxylate of the aminothiazole oxime side chain. Hydrogen-bonding interactions involving the oxime and N7 of the lysine, particularly Lys-104, may also be important in some antibiotics. Ser-164 apparently serves an indirect role, since it is somewhat distant from the active-site cleft.

Aztreonam↗
PubMed1991

Preferential binding and aggregation of rabbit C-reactive protein with arginine-rich proteins.

Human and rabbit C-reactive proteins (CRP) are similar in mol. wt, structure and amino acid sequence. In addition to the characteristic phosphoryline (PC)-binding specificity, both CRP molecules bind arginine- and lysine-rich proteins. The human CRP-cationic protein reactivity has been reported to be inhibited by calcium and promoted by PC in the presence of calcium. The present study compares binding and precipitation reactions of rabbit CRP (raCRP) with arginine- and lysine-rich proteins, and demonstrates the differential modulation of these interactions by calcium and acidic pH. Rabbit CRP shows preferential binding and precipitation reactivities with arginine-rich cationic molecules. Binding of raCRP to poly-L-arginine (PLA) and arginine-rich histone (ARH) occurs at pH 6.0, in the presence of calcium and is inhibitable by phosphorylcholine (PC) suggesting an interaction at or near the calcium-modulated PC binding site. The in vitro precipitation of raCRP and arginine-rich cationic molecules is significantly inhibited at pH 6.0, by the non-precipitating lysine-rich ligand PLL, and by physiological levels of calcium, and may reflect the participation of distinct "self-aggregation" sites on CRP in the precipitation response. The significance of the preferential arginine reactivity of raCRP to in vivo functions as a scavenger of chromatin during cell death and/or as a modulator of lipoprotein metabolism during the acute phase response is discussed.

Animals↗
PubMed1991

Bacteriochlorophyll-a as photosensitizer for photodynamic treatment of transplantable murine tumors.

Bacteriochlorophyll-a (bChla), which absorbs light of 780 nm wavelength, was tested for in vivo photodynamic activity in the SMT-F and RIF transplantable mouse tumor systems. High performance liquid chromatography (HPLC) analysis of tissue extracts showed that bChla was rapidly degraded in vivo to bacteriopheophytin-a (bPheoa) and other breakdown products. These were also photodynamically active, and tumor response could be achieved over a wavelength range of 660 to 780 nm, while tumor cure was restricted to wavelengths of 755 (bPheoa) to 780 nm. A photosensitizing product absorbing at 660 nm was also present in isolated tumor cells. Photodynamic cell kill of tumor cells isolated from tumors after bChla accumulation in vivo, using 775 or 780 nm light in vitro, was exponential up to 20-40 J cm-2. Above this light dose little or no further damage could be achieved, which is an indication of the rapid photobleaching of these sensitizers. In vivo, vascular occlusion occurred readily if light treatment was delivered shortly after sensitizer administration, but was delayed if light treatment was carried out 24 h after injection. Although up to 70% of tumor cells were lethally damaged after completion of in vivo light treatment, concurrent severe vascular destruction seemed necessary for tumor cure. Normal tissue photosensitivity totally subsided within 5 days after sensitizer administration.

Animals↗
PubMed1991

Activity and physicochemical properties of Photofrin.

Upon certain conditions of storage, Photofrin samples changed their photodynamic therapy activity in mice. These samples showed similar absorption and high performance liquid chromatography (HPLC) profiles. The Photofrin samples that were active, however, had higher relative fluorescence yields than those which failed. Active and inactive samples also had different characteristics when analyzed by gel filtration chromatography using Sephacryl S-300; the failed samples had 60% or more high-molecular-weight fraction. The change of the amount of this high-molecular-weight fraction with the time of storage at room temperature could be clearly demonstrated. Results from fluorescence, HPLC and mass spectrometry studies suggest that the high-molecular-weight fraction may form by dehydration from or condensation between various components of Photofrin.

Animals↗
PubMed1991

Photosensitization by synthetic diporphyrins and dichlorins in vivo and in vitro.

A group of polycarboxylic diporphyrins, two dichlorins and a porphyrin-chlorine dimer, with rings linked by methylene groups, were examined to help identify structures which can mediate photodynamic tumor eradication in vivo. Among the features sought were short persistence of normal tissue photosensitization and substantial absorbance at wavelengths longer than 630 nm. Both objectives were achieved, with pertinent structure-activity relationships partly characterized. The relative hydrophobicity of the different sensitizers was an important determinant of their accumulation in cell culture, but not of in vivo effectiveness. These compounds showed affinity for protein and high-density lipoprotein components of serum. Their distribution may be mediated by a different mechanism than that which occurs with more hydrophobic sensitizers like hematoporphyrin derivative which have greater affinity for low-density lipoproteins and less for protein components of serum, as compared with the products examined in this study.

Animals↗
PubMed1991

Chlorin and porphyrin derivatives as potential photosensitizers in photodynamic therapy.

In order to find a photosensitizer with better optical properties and pharmacokinetics than Photofrin II, a series of new photosensitizers related to methyl pheophorbide-a and chlorin-e6 were synthesized. These compounds absorb at substantially longer wavelengths (lambda max 660 nm) than does Photofrin II (630 nm) and show promise for use in photodynamic therapy. Among the porphyrins, we observed that long carbon chain ether derivatives are better photosensitizers than their ester analogs. These sensitizers were tested for in vivo photosensitizing activity vis-a-vis Photofrin II, using the standard screening system of DBA/2 mice bearing transplanted SMT/F tumors. Most of these photosensitizers were found to have better tumoricidal photosensitizing activity than Photofrin II and demonstrated more rapid attenuation of normal tissue photosensitivity with time after administration vis-a-vis Photofrin II.

Molecular Structure↗
PubMed1991

Comparison of cefepime, cefpirome, and cefaclidine binding affinities for penicillin-binding proteins in Escherichia coli K-12 and Pseudomonas aeruginosa SC8329.

The relative binding affinities of the extended-spectrum cephalosporins cefepime, cefpirome, and cefaclidine for the penicillin-binding proteins (PBPs) of Escherichia coli K-12 and Pseudomonas aeruginosa SC8329 were determined. Affinities were calculated from competition experiments between these antibiotics and [3H]benzylpenicillin in isolated membrane preparations. The concentrations which reduced binding to a PBP by 50% (IC50s) were determined. For E. coli, all three antibiotics displayed good PBP 3 binding (IC50s of 0.5 microgram/ml or less), and MICs roughly correlated with these values. Cefepime had a greater than 20-fold-lower IC50 for PBP 2 of E. coli than the other antibiotics. For P. aeruginosa, all of the antibiotics bound poorly (greater than 25 micrograms/ml) to PBP 2 but showed excellent pseudomonal (less than 0.0025 microgram/ml) PBP 3 binding. No correlations were seen between IC50s and MICs for P. aeruginosa. Despite differences in PBP binding, cefepime, cefpirome, and cefaclidine all displayed similar bactericidal activity for E. coli K-12 over the initial 3 h after antibiotic addition. All three caused E. coli to form filaments at values close to the MICs. In addition, cefepime induced "bleb" formation along the filaments at concentrations greater than 10x the MIC.

Bacterial Proteins↗
PubMed1991

Tobramycin uptake in Escherichia coli is driven by either electrical potential or ATP.

Aminoglycoside antibiotics such as streptomycin and tobramycin must traverse the bacterial cytoplasmic membrane prior to initiating lethal effects. Previous data on Escherichia coli, Staphylococcus aureus, and Bacillus subtilis have demonstrated that transport of aminoglycosides is regulated by delta psi, the electrical component of the proton motive force. However, several laboratories have observed that growth of bacterial cells can occur in the apparent absence of delta psi, and we wished to confirm these studies with E. coli and further investigate whether transport of aminoglycosides could occur in the absence of a membrane potential. Treatment of acrA strain CL2 with the protonophore carbonyl cyanide m-chlorophenylhydrazone (CCCP) dissipated delta psi, decreased intracellular ATP levels, and resulted in cessation of growth; after a variable period of time (3 to 7 h), growth resumed, ultimately achieving growth rates comparable to those of untreated cells. Absence of delta psi in these cells was confirmed by absence of [3H]tetraphenyl phosphonium+ uptake as measured by membrane filtration, lack of flagellar motion, and inability of these cells to transport proline (but not methionine). Regrowth was associated with restoration of normal intracellular ATP as measured by luciferin-luciferase bioluminescence assay. Unlike unacclimatized CL2 cells treated with CCCP, these cells transported [3H]tobramycin similarly to untreated cells; aminoglycoside-induced killing was seen in association with transport. These studies suggest that under certain circumstances aminoglycoside transport can be driven by ATP (or other high-energy activated phosphate donors) alone, in the absence of a measurable delta psi. delta uncBC mutants of CL2 incapable of interconverting delta psi and ATP were treated with CCCP, resulting in dissipation of delta psi but no alteration in ATP content. Despite maintenance of normal ATP, there was no transport of [3H] bramycin, confirming that under normal growth conditions ATP has no role in the transport of aminoglycosides.

Adenosine Triphosphate↗
PubMed1991

Unusual [Ca2+] dependence of vascular smooth muscle cell shortening velocity.

The relationship between unloaded shortening velocity and Ca2+ concentration was determined for hog carotid arterial smooth muscle cells, freshly isolated by digestion with papain. Cells were exposed to various [Ca2+] for 60 s at 37 degrees C and then stimulated with 10 microM histamine. Cell length was measured by a video analysis system. Shortening velocity was expressed as an exponential rate constant by fitting the cell lengths to the following equation: length = Lmin + (Lmax-Lmin)exp[-v (time-latency)], where Lmax is length before contraction, Lmin is shortest length reached, time is time elapsed after addition of agonist, latency is time from addition of agonist until contraction starts, and v is the exponential rate constant (s-1). Cells shortened substantially, usually reaching one-fourth to one-third of their initial length within 1 min. Shortening velocities of the cells were much faster than published values of maximum shortening velocity in muscle strips from this same tissue. At 1.6 mM Ca2+, v was 0.173 +/- 0.015 s-1. When Ca2+ was increased to 5 or 10 mM, v was not significantly different. However, when Ca2+ was decreased to 0.5 and 0.16 mM, v increased to 0.288 and 0.258 s-1, respectively. The difference between 0.5 and 1.6 mM was significant. The unexpected increase in shortening velocity at low Ca2+ was also seen when 10 mM caffeine was used as a stimulus: v at 1.6 mM Ca2+ was 0.156 s-1, whereas v at 0.16 mM Ca2+ was 0.272 s-1. The high shortening velocities we measured suggest that measurements made on multicellular tissues seriously underestimate the potential shortening velocity of isolated individual cells.

Animals↗
PubMed1990

Fast atom bombardment mass spectral analyses of Photofrin II and its synthetic analogs.

Fast atom bombardment (FAB) mass spectroscopy was used to characterize the components of Photofrin II. The data indicate that the oligomeric components of Photofrin II contain up to nine porphyrin units. In order to elucidate the nature of covalent linkages between porphyrin units, Photofrin II methyl ester and a porphyrin derivative-2 (2A) were synthesized and characterized by FAB mass spectroscopy. Model studies were also performed by analyzing possible components of Photofrin II, namely, porphyrin dimers 3, 4, 5, 6 and timer 7 with ether or ester linkages. The higher porphyrin oligomers joined by only ether linkages were also synthesized and characterized by FAB mass spectroscopy. The model studies indicate that the porphyrin units in Photofrin II are linked with ether or ether/ester linkages.

Antineoplastic Agents↗