bluuebunny/crossref_metadata_2025 huggingface dataset:
Dosimetric verification of intensity‐modulated fields
Authors: Wang Xiaohong , Spirou Spiridon , LoSasso Thomas , Stein Jörg , Chui Chen‐Shou , Mohan Radhe
Published: March 2003

The optimization of intensity distributions and the delivery of intensity‐modulated treatments with dynamic multi‐leaf collimators (MLC) offer important improvements to three‐dimensional conformal radiotherapy. In this study, a nine‐beam intensity‐modulated prostate plan was generated using the inverse radiotherapy technique. The resulting fluence profiles were converted into dynamic MLC leaf motions as functions of monitor units. The leaf motion pattern data were then transferred to the MLC control computer and were used to guide the motions of the leaves during irradiation. To verify that the dose distribution predicted by the optimization and planning systems was actually delivered, a homogeneous polystyrene phantom was irradiated with each of the nine intensity‐modulated beams incident normally on the phantom. For each exposure, a radiographic film was placed normal to the beam in the phantom to record the deposited dose. The films were calibrated and scanned to generate 2‐D isodose distributions. The dose was also calculated by convolving the incident fluence pattern with pencil beams. The measured and calculated dose distributions were compared and found to have discrepancies in excess of 5% of the central axis dose. The source of discrepancies was suspected to be the rounded edges of the leaves and the scattered radiation from the various components of the collimation system. After approximate corrections were made for these effects, the agreement between the two dose distributions was within 2%. We also studied the impact of the “tongue‐and‐groove” effect on dynamic MLC treatments and showed that it is possible to render this effect inconsequential by appropriately synchronizing leaf motions. This study also demonstrated that accurate and rapid delivery of realistic intensity‐modulated plans is feasible using a dynamic multi‐leaf collimator.
Generation of arbitrary intensity profiles by combining the scanning beam with dynamic multileaf collimation
Authors: Spirou Spiridon V. , Chui Chen‐Shou
Published: March 2003

An algorithm, which combines the scanning beam with dynamic collimation to generate any arbitrary intensity profile, is presented. The desired intensity profile is assumed to be piecewise linear. The dynamic collimation method used is the “sliding window.” The algorithm can be used either for a given scanning beam profile or to simultaneously determine the scanning beam profile and the leaf motions required to generate the desired intensity profile, which minimize the total treatment time. The limitations imposed by the physics of an elementary beam are taken into account. The algorithm is an iterative one, with typical calculation times being of the order of a few milliseconds.
X‐ray imaging with amorphous selenium: Optimal spectra for digital mammography
Authors: Fahrig Rebecca , Rowlands J. A. , Yaffe Martin J.
Published: March 2003

The optimum x‐ray spectra for acquisition of digital mammographic images using an amorphous selenium (a‐Se) photoconductor are investigated. The recorded images consist of latent charge distributions on the surface of an a‐Se plate, which are then read out using two methods, laser discharge, or flat panel recharge. The investigation is based on a model of the breast previously developed for a phosphor‐based digital readout system, and has been extended to include the effects specific to the use of photoconductors. The effects of plate thickness, x‐ray scatter, readout noise, dose, and the kind of breast tissue on the nature of the optimum spectrum are explored for the two readout methods. The results indicate that use of a kilovoltage setting in the current mammographic range, and a molybdenum target spectrum is appropriate for digital readout of a‐Se detectors. This conclusion contrasts with the appreciably higher kilovoltages traditionally used with the xerographic (toner) readout of latent charge images on a‐Se.
Quantification of multiple sclerosis lesion volumes in 1.5 and 0.5 T anisotropically filtered and unfiltered MR exams
Authors: Mitchell J. Ross , Karlik Stephen J. , Lee Donald H. , Eliasziw Michael , Rice George P. , Fenster Aaron
Published: March 2003

Recently, guidelines for the use of MRI in the monitoring of MS have recommended the use of imaging systems with mid‐field (0.5–1.0 T) or high‐field (greater than 1.0 T) strengths. Higher field strengths provide many advantages, including increased signal‐to‐noise ratios (SNR). SNR also may be increased by post‐processing algorithms that reduce noise. In this paper we evaluate the impact on operator variability of (a) lesion quantification in high‐field (1.5 T) versus mid‐field (0.5 T) exams; and (b) an anisotropic diffusion filter algorithm that reduces image noise without blurring or moving object boundaries. Inter‐ and intra‐operator reliability and variability were studied using repeated quantification of lesions in 1.5 and 0.5 T filtered and unfiltered MR exams of a MS patient. Results indicate that inter‐operator variability in 1.5 T unfiltered exams was 0.34 cm3 and was significantly larger than that in 1.5 T filtered (0.27 cm3), 0.5 T unfiltered (0.26 cm3), and 0.5 T filtered (0.24 cm3) exams. Similarly, intra‐operator variability in 1.5 T unfiltered exams was 0.23 cm3 and was significantly larger than that in 1.5 T filtered (0.19 cm3), 0.5 T unfiltered (0.19 cm3), and 0.5 T filtered (0.18 cm3) exams. In addition, the minimum significant change between two successive measurements of lesion volume by the same operator, was 0.64 cm3 in 1.5 T unfiltered exams, but 0.53 cm3 or less in other exams. For two different operators making successive measurements, the minimum significant change was 0.94 cm3 in 1.5 T unfiltered exams, but only 0.75 cm3 or less in other exams. Finally, the number of lesions to be monitored for an average change in volume at a given power and significance level was greater by 30%–60% for quantification in 1.5 T unfiltered exams. These results suggest that inter‐ and intra‐operator variability are reduced by anisotropic filtering, and by quantification in 0.5 T exams. Reduced operator variabilities may result from higher detail signal‐to‐noise ratios (dSNRs) in 0.5 T and filtered exams.
Tissue–phantom ratios from percentage depth doses
Authors: Bjärngard Bengt E. , Zhu Timothy C. , Ceberg Crister
Published: March 2003

When converting fractional (percentage) depth doses to tissue–phantom ratios, one must use a factor that accounts for the different source‐to‐point distances. Two minor correction factors are also involved. One is the ratio of total to primary dose at the two different distances from the source, for the same depth and field size. This factor is usually ignored. It was determined experimentally that this can introduce up to 1.5% error at 6 MV. The second correction factor reflects differences related to scattered photons and electrons at the depth of normalization in the two geometries. This correction is accounted for in published conversion procedures. It was found to be less than 1% provided the normalization depth is sufficient for electron equilibrium, which occurs first well beyond the depth of maximum dose. One may avoid electron‐equilibrium problems by using an interim normalization depth that provides electron equilibrium with some margin, renormalizing to a shallower depth if desired. With this precaution, the accuracy when measured fractional depth doses were converted to tissue–phantom ratios was comparable to that of directly measured tissue–phantom ratios even when the correction factors were ignored.
Three‐dimensional reconstruction of vascular trees. Theory and methodology
Authors: Henri C. J. , Peters T. M.
Published: March 2003

In this paper we examine the few‐view reconstruction problem as it applies to imaging vascular trees. A fully automated reconstruction algorithm is described that circumvents the traditional “correspondence problem,” using only notions of consistency and connectivity. It is assumed that the vascular tree is a connected structure and that its centerlines have been identified in three or more images. The first of three steps in the procedure involves generating a connected structure that represents the multiplicity of solutions that are consistent with any two (different) projections. The second step assigns to each branch in this structure a measure of agreement based on its relationship with one or more additional views of the vasculature. The problem then becomes one of propagating this information, via connectivity relationships and consistency checks, throughout the above structure to distinguish between the branches comprising the imaged structure and the accompanying artifacts. In this paper we present the theory and methodology of the technique, while in a companion paper we address the issue of validation via simulations and experiments. Together, these papers shed some light on why ambiguities arise and often lead to errors in the few‐view reconstruction problem. Strategies to handle these errors are described and results are presented that demonstrate the ability to obtain adequate reconstructions with as few as three distinct views.
Europium‐152 as a potential substitute for cobalt‐60 in radiation therapy
Authors: Adelman S. L.
Published: March 2003

Construction has begun of prototype europium‐152 sources, with the principal goal of providing cost‐effective teletherapy for emergent nations where prices for cobalt‐60 are excessive, owing to replacement needs, and the use of linacs is economically prohibitive and technologically unfeasible. 152Eu sources (T1/2=13.4y vs T1/2=5.26y for 60Co) are designed for absorbed dose rates ≳1.5 Gy/min at 1 m, with specific activities ≳5 TBq/gm, allowing substitution into existing irradiators. Costs of 152Eu compare favorably with 60Co, taking account of extended useful lifetime.
Density resolution in quantitative computed tomography of foam and lung
Authors: Kemerink Gerrit J. , Kruize Han H. , Lamers Rob J. S. , van Engelshoven Jos M. A.
Published: March 2003

This study was performed to assess density resolution in quantitative computed tomography (CT) of foam and lung. Density resolution, a measure for the ability to discriminate materials of different density in a CT number histogram, is normally determined by quantum noise. In a cellular solid, variations in mass in the volumes sampled by CT cause an additional degradation of density resolution by the linear partial volume effect. The sample volume, which is directly related to spatial resolution, can be varied by choosing different section thicknesses and reconstruction filters. Several polyethene (PE) foams, as simple models of lung tissue, and five patients were investigated using various sample volumes. For the uniform PE foams, density resolution could be directly determined as the full width at half maximum of CT number histograms. Density resolution for foams with cell sizes of 0.8–1.5 mm was dominated by effects caused by the limited sample size, not by quantum noise. The relative magnitudes of density resolution could roughly be explained with a model for a hypothetic random cellular solid. Since lungs are not of uniform density, analysis of patient data was more complicated. A combined convolution least‐squares fit procedure, together with information obtained in the studies of foam, were used to determine density resolution in lung studies. Density resolution, both for foams and lung, was strongly dependent on sample volume, and was quite poor for thin sections and sharp filters. Consequently, histogram‐shape related parameters are sensitive to the spatial resolution chosen on CT. Thin section densitometry, using a 1‐mm section with a standard or high resolution filter, is not recommended except in determining average density. When using thicker sections, an in‐plane spatial resolution similar to section thickness is advised.
A totally integrated simulation technique for three‐field breast treatment using a CT simulator
Authors: Butker Elizabeth K. , Helton Daisy J. , Keller James W. , Hughes Lorie L. , Crenshaw Tim , Davis Lawrence W.
Published: March 2003

A method was devised to simulate patients with breast cancer in the actual treatment position utilizing a diagnostic CT spiral scanner, 3‐D Image Workstation for virtual simulation, and a laser coordinate system to transfer planning parameters to the patient's skin. It was desired to produce non‐divergent tangential beams through the lung as well as a matched line for tangential and supraclavicular fields. The patients were immobilized in an Alpha CradleTM cast. Radio‐opaque markers were placed on the superior, inferior, medial, and lateral margins of the field so as to afford appropriate initial field set‐up approximations. The patient was scanned. The data set was then transferred to the workstation where an isocenter was chosen. The patient was marked. Virtual simulation was then performed. This method employed a half beam technique for the posterior edge of the tangential fields. Table rotation and blocking of the superior margin of the tangential fields were used to produce a vertical edge to match a supraclavicular field. Using a beam's eye view the lateral tangent was matched to the medial exit. A digitally reconstructed radiograph was created to define the tangent fields and place the supraclavicular block. Our initial experience with 50 patients verifies that this is a reproducible and accurate technique. Time required for immobilization and tangential field simulation is approximately 30 minutes. Data is available for 3‐D treatment planning or 2‐D treatment planning on a reconstructed transverse slice angled to match the collimator angle through the patient. Using a CT simulator for simulation of breast cancer affords accuracy of at least equal magnitude to conventional simulators as determined by beam films and ease of set‐up. This technique also affords greater ease in changing treatment parameters without having to resimulate the patient.
Chelator effect on ion diffusion in ferrous‐sulfate‐doped gelatin gel dosimeters as analyzed by MRI
Authors: Rae William I. D. , Willemse Casper A. , Lötter Mattheus G. , Engelbrecht Johan S. , Swarts Jannie C.
Published: March 2003

Ferrous‐sulfate‐doped gelatin gel dosimeters are useful tools for the measurement of three‐dimensional absorbed radiation dose distributions. The diffusion of ferric ions through these gels causes degradation with time of the dose distribution image. It would be useful to reduce ferric ion diffusion without decreasing gel sensitivity. The amount of ferric ion diffusion is a function of the time delay after radiation, the gel temperature, and the gel concentration. These effects can be quantified by measuring the ferric ion diffusion coefficient. Determination of the diffusion coefficient by irradiating the lower section of a cylinder of gel, which was then imaged repeatedly over time with a clinical magnetic resonance imager, is described. Analysis of the edge spread function formed at each of several times after irradiation by drawing a profile over the imaged junction between the irradiated and unirradiated halves of the cylinder, gave estimates of the variance of the edge spread function. These variances were used to obtain an estimate of the ferric ion diffusion coefficient for the gel. A method of reducing ferric ion diffusion by adding a chelator and the cross linkage agent formaldehyde is suggested. The chelators investigated were 1,10 phenanthroline, xylenol orange, and bathophenanthroline disulfonic acid. These reduced diffusion to varying extents, and influenced the gel sensitivity. The diffusion coefficient in gels containing xylenol orange was found to be 0.44 mm2h−1. The gel sensitivity was 0.0093 s−1Gy−1. This compared with a diffusion coefficient of 0.82 mm2h−1 for the base line gel that did not contain formaldehyde or chelators. The sensitivity of this base line gel was 0.0129 s−1Gy−1. The addition of xylenol orange produced the most improved gel dosimeter of the gels studied. This gel had a decreased ferric ion diffusion coefficient and a decreased sensitivity. It was still sensitive enough to be useful.
Spatial mapping of the percentage cellularity in human bone marrow using magnetic resonance imaging
Authors: Ballon Douglas , Jakubowski Ann A. , Graham Martin C. , Schneider Erika , Koutcher Jason A.
Published: March 2003

A noninvasive assay for the spatial distribution of the percentage cellularity in human bone marrow is presented. Twelve individuals were studied using two magnetic resonance imaging techniques: (1) fast spin echo imaging with frequency selective presaturation, and (2) three‐point chemical shift imaging. The data were compared to results obtained using a previously validated stimulated echo spectroscopic method. The results of this study demonstrate that a measure of the percentage cellularity in bone marrow is possible using magnetic resonance imaging techniques provided that high‐quality water or lipid suppression is achieved across the region of interest. Since the method is applicable to bone marrow at any anatomic location, it may prove useful in dosimetric calculations during and after a course of internal or external beam radiotherapy.
A model for the physical optimization of external beam radiotherapy
Authors: Holmes Timothy William
Published: March 2003

A model for optimizing conformal radiotherapy dose distributions was developed based on the analogy between rotational radiotherapy and single photon emission computed tomography image reconstruction. The optimization model predicts that a large number of intensity modulated beam profiles are required to maximize conformation of the dose distribution to the dose prescription—a solution not practically feasible using early 1990s treatment delivery technology. A new method of treatment realization similar to spiral CT scanning called tomotherapy is described. The tomotherapy unit would incorporate a diagnostic CT scanner for acquiring tomographic images prior to and during a treatment for optimized treatment planning and treatment verification, respectively. The tomotherapy concept represents the first truly integrated approach to optimized treatment planning, delivery, and verification of external beam radiotherapy and could significantly advance the state‐of‐the‐art in radiotherapy.
The role of magnetic susceptibility in magnetic resonance imaging: MRI magnetic compatibility of the first and second kinds
Authors: Schenck John F.
Published: March 2003

The concept of magnetic susceptibility is central to many current research and development activities in magnetic resonance imaging (MRI); for example, the development of MR‐guided surgery has created a need for surgical instruments and other devices with susceptibility tailored to the MR environment; susceptibility effects can lead to position errors of up to several millimeters in MR‐guided stereotactic surgery; and the variation of magnetic susceptibility on a microscopic scale within tissues contributes to MR contrast and is the basis of functional MRI. The magnetic aspects of MR compatibility are discussed in terms of two levels of acceptability: Materials with the first kind of magnetic field compatibility are such that magnetic forces and torques do not interfere significantly when the materials are used within the magnetic field of the scanner; materials with the second kind of magnetic field compatibility meet the more demanding requirement that they produce only negligible artifacts within the MR image and their effect on the positional accuracy of features within the image is negligible or can readily be corrected. Several materials exhibiting magnetic field compatibility of the second kind have been studied and a group of materials that produce essentially no image distortion, even when located directly within the imaging field of view, is identified. Because of demagnetizing effects, the shape and orientation, as well as the susceptibility, of objects within and adjacent to the imaging region is important in MRI. The quantitative use of susceptibility data is important to MRI, but the use of literature values for the susceptibility of materials is often difficult because of inconsistent traditions in the definitions and units used for magnetic parameters—particularly susceptibility. The uniform use of SI units for magnetic susceptibility and related quantities would help to achieve consistency and avoid confusion in MRI.
Verification of milled two‐dimensional photon compensating filters using an electronic portal imaging device
Authors: Low Daniel A. , Li Zuofeng , Klein Eric E.
Published: March 2003

A computer‐driven milling machine is being installed at the Mallinckrodt Institute of Radiology to fabricate photon compensating filters for conventional compensation and beam‐intensity modulation. Commissioning and quality assurance procedures are being developed for the design, fabrication, and delivery systems prior to using the milled filters in the clinic. The portion of the quality assurance procedures governing the filter fabrication using a computer‐driven milling machine includes, in part, a comparison of designed and fabricated filters. Test filters include geometrically regular filters, such as flat surfaces and steps. The verification of these shapes is accomplished using spatial measurements. However, to test the mill's ability to generate complex curved surfaces, filters with more complicated surfaces are generated. These filters do not lend themselves to precise verification using physical measurement of the thickness distribution. A method has been developed to verify the thickness distributions of these complex filters by irradiating the filters with high‐energy x rays and comparing the scattered and transmitted fluence to the fluence calculated using the intended filter shape. The fluence measurement is made using a calibrated commercial liquid ionization chamber electronic portal imaging device. The calculated fluence is separated into transmitted primary and scattered fluences and is determined using a convolution of a distributed radiation source kernel with an exponential filter attenuation function. The attenuation coefficient is measured for the filter material (Lipowitz metal) and fit to a second‐order polynomial in filter thickness and off‐axis distance. The distributed source kernel is measured using a split‐field technique and fit to a sum of three two‐dimensional Gaussian distributions. The scattered radiation is modeled by the Klein–Nishina cross section. The algorithm is tested by comparing calculated fluences with measured fluences for a series of machined filters: an open field, flat filters of 9.6‐, 15.8‐, and 31.6‐mm thickness, split‐field filters, and a pyramid‐shaped filter. In each case, the algorithm calculates the fluence to within 3% of the measured values over the entire irradiated field size to within 1.5 cm of the collimated field edges.
The elements of tissue–air ratio and systematic error
Authors: Nizin P. S. , Bellezza D. M. , Mooij R. B.
Published: March 2003

The definition of tissue–air ratio (TAR) is based on the concept of primary dose. To determine TAR, both in‐phantom and in‐air ionization measurements are utilized. To convert ionization in the phantom into dose and that in air into primary dose, correction factors must be applied to chamber readings in both geometries. Due to difficulties in selecting proper correction factors, TAR is subject to systematic error. The error comes from two sources of uncertainty: (1) Primary dose cannot be measured. Therefore approximate methods, such as in‐air ionization measurements, are used. (2) Detectors of ionization are of finite dimensions and they are inhomogeneous. In this study, analytical expression for a systematic error is derived. Because in this derivation systematic error is an accumulative error, it is no longer necessary to convert ionization, both in air and in phantom, into a dose when calculating TARs. A method of determining systematic error is described. This method is based on the ability to produce accurate zero‐field data in photon beams by means of a linear extrapolation technique. Using 60Co γ radiation in water as an example, it is shown how to generate TAR data free of systematic error. A possibility of determining TARs for therapeutic x rays is discussed.
Dosimetry of high energy photon and electron beams with CEA films
Authors: Cheng Chee‐Wai , Das Indra J.
Published: March 2003

With all the advantages of film dosimetry in the megavoltage energy range, the use of film as a dosimeter is still limited due to the various difficulties associated with films such as energy dependence, film orientation, and sensitometric nonlinearity. Recently, therapy verification and localization films (CEA TVS and TLF films) from a Swedish manufacturer have become available in vacuum‐sealed water‐proof packaging in the US. The packaging renders the CEA films useful in a water phantom and ideal for photon and electron dosimetry. A systematic study has been carried out to investigate the potential of dosimetric application of the new films for high energy photon and electron beams. For the TVS films, the characteristic curve is generally energy independent but appears to be dependent on the source of the radiation, i.e., whether it is gamma rays or bremsstrahlung x rays. Compared to Kodak Readypack XV films, the CEA TVS film is linear in optical density over a much larger range of radiation dose. The inter‐ and intra‐variation of the TVS films is less than 2%. For electrons, the characteristic curve is linear over a similar density range as photons but exhibit a slight energy dependence. TVS film is slightly directional dependent on the incident radiation for both photons and electrons. The perpendicular orientation results in higher optical density than the parallel orientation. The differences are within ±2% except in the buildup region for photons and in the exponential fall‐off region of the electron beams where differences up to 4% are noted. For the CEA TLF film which is about three times faster than the TVS film, the characteristic curve is reasonably linear over the dose range of 0–15 cGy and energy independent within the experimental uncertainty (±5%). Percent depth dose and isodose measurements with the TVS films are in good agreement with ion chamber results.
Using tissue texture surrounding calcification clusters to predict benign vs malignant outcomes
Authors: Thiele David L. , Kimme‐Smith Carolyn , Johnson Timothy D. , McCombs Marie , Bassett Lawrence W.
Published: March 2003

The positive predictive value of mammography is between 20% and 25% for clustered microcalcifications. For very early cancers there is often a lack of concordance between mammographic signs and pathology. This study examines the usefulness of computer texture analysis to improve the accuracy of malignant diagnosis. Texture analysis of the breast tissue surrounding microcalcifications on digitally acquired images during stereotactic biopsy is used in this study to predict malignant vs benign outcomes. 54 biopsy proven cases (36 benign, 18 malignant) are used. The texture analysis calculates statistical features from gray level co‐occurrence matrices and fractal geometry for equal probability and linear quantizations of the image data. Discriminant models are generated using linear discriminant analysis and logistic discriminant analysis. Results do not differ significantly by method of quantization or discriminant analysis. Jackknife results misclassify 2 of 18 malignant cases (sensitivity 89%) and 6 of 36 benign cases (specificity 83%) for logistic discriminant analysis. From this preliminary study, texture analysis appears to show significant discriminatory power between benign and malignant tissue, which may be useful in resolving problems of discordance between pathological and mammographic findings, and may ultimately reduce the number of benign biopsies.
In Vivo Pharmacodynamics of HMR 3270, a Glucan Synthase Inhibitor, in a Murine Candidiasis Model
Authors: Andes D. , Marchillo K. , Lowther J. , Bryskier A. , Stamstad T. , Conklin R.
Published: March 2003

In vivo pharmacokinetic/pharmacodynamic characterization for numerous antibacterial compounds has had a significant impact upon optimal dosing regimen design and the development of in vivo susceptibility breakpoints. More recently, similar characterization has been undertaken for antifungal drug classes. Very little is known of these pharmacodynamic relationships for the new echinocandin class of compounds. We utilized a neutropenic murine model of disseminated candidiasis to describe the time course antifungal activity of HMR 3270, a new glucan synthase inhibitor. Single-dose in vivo time kill studies with four 16-fold escalating doses demonstrated concentration-dependent killing when drug levels in serum were more than four times the MIC. Postantifungal effects were dose dependent, ranging from 8 to 80 h duration. Multiple dosing regimen studies utilized six total doses, four dosing intervals, and a treatment duration of 6 days. Shortening the dosing interval from every 144 h (q144h) to q36h resulted in a fourfold rise in the dose necessary to achieve a net fungistatic effect. The peak/MIC ratio most strongly correlated with treatment outcomes (peak/MIC ratio,
R
2
= 98%; ratio of the area under the concentration-time curve from 0 to 24 h to the MIC,
R
2
= 79%, percentage of time above the MIC,
R
2
= 61%). Studies were also conducted with five additional
Candida albicans
isolates to determine if a similar peak/MIC ratio was associated with efficacy. In vivo concentration-dependent killing was similarly observed in studies with each of the additional isolates. The peak/MIC ratio necessary to produce efficacy was relatively similar among the strains studied (
P
= 0.42). The peak/MIC ratio (mean ± standard deviation) necessary to achieve a fungistatic effect was 3.72 ± 1.84, and the ratio necessary to achieve maximal killing was near 10.
In Vivo Pharmacodynamics of a New Triazole, Ravuconazole, in a Murine Candidiasis Model
Authors: Andes D. , Marchillo K. , Stamstad T. , Conklin R.
Published: March 2003

In vivo studies have characterized the pharmacodynamic characteristics of the triazole fluconazole. These investigations demonstrated that the ratio of the area under the concentration-time curve from 0 to 24 h to the MIC (24-h AUC/MIC ratio) is the critical pharmacokinetic/pharmacodynamic (PK/PD) parameter associated with treatment efficacy. Further analysis demonstrated that a fluconazole 24-h AUC/MIC ratio of 20 to 25 was predictive of treatment success in both experimental models and clinical trials. We used a neutropenic murine model of disseminated
Candida albicans
infection to similarly characterize the time course activity of the new triazole ravuconazole. The PK/PD parameters (percent time above the MIC, AUC/MIC ratio, and peak level in serum/MIC ratio) were correlated with in vivo efficacy, as measured by organism number in kidney cultures after 24 and 72 h of therapy. Ravuconazole kinetics and protein binding were performed in neutropenic infected mice. Peak/dose and AUC/dose values ranged from 0.03 to 0.04 and 0.30 to 0.34, respectively. Serum elimination half-life ranged from 3.9 to 4.8 h. Protein binding was 95.8%. Single-dose postantifungal effect studies demonstrated prolonged suppression of organism regrowth after serum ravuconazole levels had fallen below the MIC. Treatment efficacies with the five dosing intervals studied were similar, supporting the argument for the AUC/MIC ratio as the PK/PD parameter predictive of efficacy. Nonlinear regression analysis also suggested that the AUC/MIC ratio was strongly predictive of treatment outcomes (AUC/MIC ratio,
R
2
= 91%; peak/MIC ratio,
R
2
= 85%; percent time above the MIC,
R
2
= 47 to 65%). Similar studies were conducted with seven additional
C. albicans
isolates with various ravuconazole susceptibilities (MIC, 0.016 to 0.12 μg/ml) to determine if a similar 24-h AUC/MIC ratio was associated with efficacy. The ravuconazole free-drug AUC/MIC ratios were similar for all of the organisms studied (10 to 36; mean ± SD = 20.3 ± 8.2;
P
= 0.43). These free-drug AUC/MIC ratios are similar to those observed for fluconazole in this model.
Sublethal Injury and Resuscitation of
Candida albicans
after Amphotericin B Treatment
Authors: Liao Robert S. , Rennie Robert P. , Talbot James A.
Published: March 2003

Amphotericin B treatment was previously shown to inhibit
Candida albicans
reproduction and reduce the fluorescence of vitality-specific dyes without causing a corresponding increase in the fluorescence of the mortality-specific dyes bis-(1,3-dibutylbarbituric acid)trimethine oxonol and SYBR Green Ι. In the present study, we have confirmed these results and have shown that the numbers of CFU are reduced by 99.9% by treatment with 0.5 μg of amphotericin B per ml for 10 h at 35°C. This reduction was not due to fungal cell death. First, the level of reduction of the tetrazolium salt 2,3-bis(2-methoxy-4-nitro-5-sulfophenyl)-5-[(phenylamino)carbonyl]-2
H
-tetrazolium hydroxide increased in the presence of concentrations of amphotericin B that caused greater than 90% reductions in the numbers of CFU. Second, fungal cells treated with amphotericin B at a concentration of 0.5 μg/ml were resuscitated by further incubation at 22°C for 15 h in the continued presence of amphotericin B. Third, recovery of the ability to replicate was prevented by sequential treatment with 20 μg of miconazole per ml, which also increased the fluorescence of mortality-specific dyes to near the maximal levels achieved with 0.9 μg of amphotericin B per ml. Sequential treatment with fluconazole and flucytosine did not increase the levels of staining with the mortality-specific dyes. Itraconazole was less effective than ketoconazole, which was less effective than miconazole. The practice of equating the loss of the capacity of
C. albicans
to form colonies with fungal cell death may give incorrect results in assays with amphotericin B, and the results of assays with caution with other antifungal agents that are lipophilic or that possess significant postantifungal effects may need to be interpreted.
Drug Resistance Is Not Directly Affected by Mating Type Locus Zygosity in
Candida albicans
Authors: Pujol Claude , Messer Shawn A. , Pfaller Michael , Soll David R.
Published: March 2003

Recently, evidence was presented that in a collection of fluconazole-resistant strains of
Candida albicans
there was a much higher proportion of homozygotes for the mating type locus (
MTL
) than in a collection of fluconazole-sensitive isolates, suggesting the possibility that when cells become
MTL
homozygous they acquire intrinsic drug resistance. To investigate this possibility, an opposite strategy was employed. First, drug susceptibility was measured in a collection of isolates selected for
MTL
homozygosity. The majority of these isolates had not been exposed to antifungal drugs. Second, the level of drug susceptibility was compared between spontaneously generated
MTL
-homozygous progeny and their
MTL
-heterozygous parent strains which had not been exposed to antifungal drugs. The results demonstrate that naturally occurring
MTL
-homozygous strains are not intrinsically more drug resistant, supporting the hypotheses that either the higher incidence of
MTL
homozygosity previously demonstrated among fluconazole-resistant isolates involved associated homozygosity of a drug resistance gene linked to the
MTL
locus, or that
MTL
-homozygous strains may be better at developing drug resistance upon exposure to the drug than
MTL
-heterozygous strains. Furthermore, the results demonstrate that a switch by an
MTL
-homozygous strain from the white to opaque phenotype, the latter functioning as the facilitator of mating, does not notably alter drug susceptibility.
Genetic Basis for Differential Activities of Fluconazole and Voriconazole against
Candida krusei
Authors: Fukuoka Takashi , Johnston Douglas A. , Winslow Carol A. , de Groot Marcel J. , Burt Catherine , Hitchcock Christopher A. , Filler Scott G.
Published: March 2003

Invasive infections caused by
Candida krusei
are a significant concern because this organism is intrinsically resistant to fluconazole. Voriconazole is more active than fluconazole against
C. krusei
in vitro. One mechanism of fluconazole resistance in
C. krusei
is diminished sensitivity of the target enzyme, cytochrome P450 sterol 14α-demethylase (CYP51), to inhibition by this drug. We investigated the interactions of fluconazole and voriconazole with the CYP51s of
C. krusei
(ckCYP51) and fluconazole-susceptible
Candida albicans
(caCYP51). We found that voriconazole was a more potent inhibitor of both ckCYP51 and caCYP51 in cell extracts than was fluconazole. Also, the ckCYP51 was less sensitive to inhibition by both drugs than was caCYP51. These results were confirmed by expressing the
CYP51
genes from
C. krusei
and
C. albicans
in
Saccharomyces cerevisiae
and determining the susceptibility of the transformants to voriconazole and fluconazole. We constructed homology models of the CYP51s of
C. albicans
and
C. krusei
based on the crystal structure of CYP51 from
Mycobacterium tuberculosis
. These models predicted that voriconazole is a more potent inhibitor of both caCYP51 and ckCYP51 than is fluconazole, because the extra methyl group of voriconazole results in a stronger hydrophobic interaction with the aromatic amino acids in the substrate binding site and more extensive filling of this site. Although there are multiple differences in the predicted amino acid sequence of caCYP51 and ckCYP51, the models of the two enzymes were quite similar and the mechanism for the relative resistance of ckCYP51 to the azoles was not apparent.
Genome-Wide Expression Profile Analysis Reveals Coordinately Regulated Genes Associated with Stepwise Acquisition of Azole Resistance in
Candida albicans
Clinical Isolates
Authors: Rogers P. David , Barker Katherine S.
Published: March 2003

Candida albicans
is an opportunistic human fungal pathogen and a causative agent of oropharyngeal candidiasis (OPC), the most frequent opportunistic infection among patients with AIDS. Fluconazole and other azole antifungal agents have proven effective in the management of OPC; however, with increased use of these agents treatment failures have occurred. Such failures have been associated with the emergence of azole-resistant strains of
C. albicans
. In the present study we examined changes in the genome-wide gene expression profile of a series of
C. albicans
clinical isolates representing the stepwise acquisition of azole resistance. In addition to genes previously associated with azole resistance, we identified many genes whose differential expression was for the first time associated with this phenotype. Furthermore, the expression of these genes was correlated with that of the known resistance genes
CDR1
,
CDR2
, and
CaMDR1
. Genes coordinately regulated with the up-regulation of
CDR1
and
CDR2
included the up-regulation of
GPX1
and
RTA3
and the down-regulation of
EBP1
. Genes coordinately regulated with the up-regulation of
CaMDR1
included the up-regulation of
IFD1
,
IFD4
,
IFD5
,
IFD7
,
GRP2
,
DPP1
,
CRD2
, and
INO1
and the down-regulation of
FET34
,
OPI3
, and
IPF1222
. Several of these appeared to be coordinately regulated with both the CDR genes and
CaMDR1
. Many of these genes are involved in the oxidative stress response, suggesting that reduced susceptibility to oxidative damage may contribute to azole resistance. Further evaluation of the role these genes and their respective gene products play in azole antifungal resistance is warranted.
Isolation of Three
Mycobacterium ulcerans
Strains Resistant to Rifampin after Experimental Chemotherapy of Mice
Authors: Marsollier Laurent , Honoré Nadine , Legras Pierre , Manceau Anne Lise , Kouakou Henri , Carbonnelle Bernard , Cole Stewart T.
Published: March 2003

By use of a murine model for Buruli ulcer,
Mycobacterium ulcerans
was found to be susceptible to rifampin, with the MIC being 0.5 to 1 μg/ml. Three mutants were isolated after rifampin monotherapy. Two were resistant to rifampin at 8 μg/ml, and one was resistant to rifampin at 32 μg/ml. The mutants harbored Ser416Phe mutations and His420Tyr mutations in the
rpoB
gene, and these mutations have also been found to be responsible for rifampin resistance in the leprosy and tubercle bacilli. The results indicate that while rifampin may be active against
M. ulcerans
, it should never be used as monotherapy in humans.
In Vivo Antiretroviral Activity of Stampidine in Chronically Feline Immunodeficiency Virus-Infected Cats
Authors: Uckun Fatih M. , Chen Chun-Lin , Samuel Peter , Pendergrass Sharon , Venkatachalam T. K. , Waurzyniak Barbara , Qazi Sanjive
Published: March 2003

Here we report the antiretroviral activity of the experimental nucleoside reverse transcriptase inhibitor (NRTI) compound stampidine in cats chronically infected with feline immunodeficiency virus (FIV). Notably, a single oral bolus dose of 50 or 100 mg of stampidine per kg resulted in a transient ≥1-log decrease in the FIV load of circulating peripheral blood mononuclear cells in five of six FIV-infected cats and no side effects. A 4-week stampidine treatment course with twice-daily administration of hard gelatin capsules containing 25 to 100 mg of stampidine per kg was also very well tolerated by cats at cumulative dose levels as high as 8.4 g/kg and exhibited a dose-dependent antiretroviral effect. One of three cats treated at the 25-mg/kg dose level, three of three cats treated at the 50-mg/kg dose level, and three of three cats treated at the 100-mg/kg dose level (but none of three control cats treated with placebo pills) showed a therapeutic response, as evidenced by a ≥1-log reduction in the FIV load in peripheral blood mononuclear cells within 2 weeks. The previously documented in vitro and in vivo antiretroviral activity of stampidine against primary clinical human immunodeficiency virus type 1 isolates with genotypic and/or phenotypic NRTI resistance, together with its favorable animal toxicity profile, pharmacokinetics, and in vivo antiretroviral activity in FIV-infected cats, warrants further development of this promising new NRTI compound.
Single Nucleotide Polymorphisms in Genes Associated with Isoniazid Resistance in
Mycobacterium tuberculosis
Authors: Ramaswamy Srinivas V. , Reich Robert , Dou Shu-Jun , Jasperse Linda , Pan Xi , Wanger Audrey , Quitugua Teresa , Graviss Edward A.
Published: March 2003

Isoniazid (INH) is a central component of drug regimens used worldwide to treat tuberculosis. Previous studies have identified resistance-associated mutations in
katG
,
inhA
,
kasA
,
ndh
, and the
oxyR-ahpC
intergenic region. DNA microarray-based experiments have shown that INH induces several genes in
Mycobacterium tuberculosis
that encode proteins physiologically relevant to the drug's mode of action. To gain further insight into the molecular genetic basis of INH resistance, 20 genes implicated in INH resistance were sequenced for INH resistance-associated mutations. Thirty-eight INH-monoresistant clinical isolates and 86 INH-susceptible isolates of
M. tuberculosis
were obtained from the Texas Department of Health and the Houston Tuberculosis Initiative. Epidemiologic independence was established for all isolates by IS
6110
restriction fragment length polymorphism analysis. Susceptible isolates were matched with resistant isolates by molecular genetic group and IS
6110
profiles. Spoligotyping was done with isolates with five or fewer IS
6110
copies. A major genetic group was established on the basis of the polymorphisms in
katG
codon 463 and
gyrA
codon 95. MICs were determined by the E-test. Semiquantitative catalase assays were performed with isolates with mutations in the
katG
gene. When the 20 genes were sequenced, it was found that 17 (44.7%) INH-resistant isolates had a single-locus, resistance-associated mutation in the
katG
,
mabA
, or
Rv1772
gene. Seventeen (44.7%) INH-resistant isolates had resistance-associated mutations in two or more genes, and 76% of all INH-resistant isolates had a mutation in the
katG
gene. Mutations were also identified in the
fadE24
,
Rv1592c
,
Rv1772
,
Rv0340
, and
iniBAC
genes, recently shown by DNA-based microarray experiments to be upregulated in response to INH. In general, the MICs were higher for isolates with mutations in
katG
and the isolates had reduced catalase activities. The results show that a variety of single nucleotide polymorphisms in multiple genes are found exclusively in INH-resistant clinical isolates. These genes either are involved in mycolic acid biosynthesis or are overexpressed as a response to the buildup or cellular toxicity of INH.
Role of Nutrient Limitation and Stationary-Phase Existence in
Klebsiella pneumoniae
Biofilm Resistance to Ampicillin and Ciprofloxacin
Authors: Anderl Jeff N. , Zahller Jeff , Roe Frank , Stewart Philip S.
Published: March 2003

Biofilms formed by
Klebsiella pneumoniae
resisted killing during prolonged exposure to ampicillin or ciprofloxacin even though these agents have been shown to penetrate bacterial aggregates. Bacteria dispersed from biofilms into medium quickly regained most of their susceptibility. Experiments with free-floating bacteria showed that stationary-phase bacteria were protected from killing by either antibiotic, especially when the test was performed in medium lacking carbon and nitrogen sources. These results suggested that the antibiotic tolerance of biofilm bacteria could be explained by nutrient limitation in the biofilm leading to stationary-phase existence of at least some of the cells in the biofilm. This mechanism was supported by experimental characterization of nutrient availability and growth status in biofilms. The average specific growth rate of bacteria in biofilms was only 0.032 h
−1
compared to the specific growth rate of planktonic bacteria of 0.59 h
−1
measured in the same medium. Glucose did not penetrate all the way through the biofilm, and oxygen was shown to penetrate only into the upper 100 μm. The specific catalase activity was elevated in biofilm bacteria to a level similar to that of stationary-phase planktonic cells. Transmission electron microscopy revealed that bacteria were affected by ampicillin near the periphery of the biofilm but were not affected in the interior. Taken together, these results indicate that
K. pneumoniae
in this system experience nutrient limitation locally within the biofilm, leading to zones in which the bacteria enter stationary phase and are growing slowly or not at all. In these inactive regions, bacteria are less susceptible to killing by antibiotics.