Testing candidate plant barcode regions in the Myristicaceae.

The concept and practice of DNA barcoding have been designed as a system to facilitate species identification and recognition. The primary challenge for barcoding plants has been to identify a suitable region on which to focus the effort. The slow relative nucleotide substitution rates of plant mitochondria and the technical issues with the use of nuclear regions have focused attention on several proposed regions in the plastid genome. One of the challenges for barcoding is to discriminate closely related or recently evolved species. The Myristicaceae, or nutmeg family, is an older group within the angiosperms that contains some recently evolved species providing a challenging test for barcoding plants. The goal of this study is to determine the relative utility of six coding (Universal Plastid Amplicon - UPA, rpoB, rpoc1, accD, rbcL, matK) and one noncoding (trnH-psbA) chloroplast loci for barcoding in the genus Compsoneura using both single region and multiregion approaches. Five of the regions we tested were predominantly invariant across species (UPA, rpoB, rpoC1, accD, rbcL). Two of the regions (matK and trnH-psbA) had significant variation and show promise for barcoding in nutmegs. We demonstrate that a two-gene approach utilizing a moderately variable region (matK) and a more variable region (trnH-psbA) provides resolution among all the Compsonuera species we sampled including the recently evolved C. sprucei and C. mexicana. Our classification analyses based on nonmetric multidimensional scaling ordination, suggest that the use of two regions results in a decreased range of intraspecific variation relative to the distribution of interspecific divergence with 95% of the samples correctly identified in a sequence identification analysis.

On the use of virtual simulation in radiotherapy of the intact breast.

In this paper a method of breast cancer treatment planning using virtual simulation implemented at the Department of Human Oncology at the University of Wisconsin is described. All patients in this procedure are placed in a custom vacuum mold in treatment position with both arms up to avoid collision with the CT scanner aperture. For all patients a CT scan of 5-mm-slice thickness is acquired. The ipsilateral and contralateral breast, the ipsilateral lung and the heart are delineated and a three-dimensional plan is generated that tries to minimize the dose to the ipsilateral lung and heart while ensuring adequate coverage of the affected breast. Digitally reconstructed radiographs are used to verify the patient setup on the treatment machine.

Transgenic mice with pigmented intraocular tumors: tissue of origin and treatment.

PURPOSE: To describe the cell of origin, tumor progression, light and electron microscopic appearance, immunohistochemical properties, and response to frequently used anticancer therapies in two transgenic models of intraocular melanoma. METHODS: Two lines of transgenic mice that develop pigmented intraocular tumors were produced with the SV40 T and t antigens under the control of the mouse tyrosinase gene. Tumors were sequentially studied and characterized by light microscopy, electron microscopy, and immunohistochemistry stains. Tumor response to two cycles of dacarbazine was assessed on the basis of tumor size in one group of animals. Response to external beam irradiation was measured by survival time in other animals. RESULTS: Two lines of transgenic mice developed bilateral intraocular tumors with complete penetrance and without primary cutaneous melanomas. Tumors developed first in the retinal pigment epithelial layer, with subsequent retinal and choroidal invasion, extraocular extension, and metastasis. Tumors stained positive for S-100, HMB-45, and Fas-ligand. Electron microscopy revealed polarization of tumor cells with basement membrane formation, microvilli, immature melanosomes, and abundant endoplasmic reticulum. Dacarbazine significantly reduced tumor size in these mice, and a trend toward dose-dependent decrease in survival was found with external beam irradiation. CONCLUSIONS: Tumors developed from the retinal pigment epithelium. Their histology and growth, however, closely resembled that of human choroidal melanoma. This model may be a useful tool for future studies of endogenous primary pigmented tumors limited to the eye. Response to standard therapies suggests it can serve as a model with which to evaluate therapeutic modalities.

Ferromagnetic hyperthermia: functional and histopathologic effects on normal rabbit ocular tissue.

Ferromagnetic (FM) hyperthermia has previously been evaluated in a rabbit tumour model of ocular melanoma. To study the effect of focal heating in normal rabbit eyes, FM seeds were implanted into a 14-mm episcleral plaque an heated to operating temperatures of 48 or 58 degrees C. Thermal induction was performed by placing rabbits in a uniform, oscillating (11 kHz) magnetic field operating at 1200 W and as H-field strength of 265 A/m. Eyes were heated for 60 min with continuous scleral temperature monitoring. Hyperthermic effects were monitored by direct opthalmic examination, fundus photography, serial electroretinography and histopathology. Intraocular temperatures were mapped with direct fiberoptic thermometry. All treatment effects were confined to the area covered by the episcleral plaque. Direct ophthalmoscopic examination revealed early retinal whitening during heat induction followed by localized exudative retinal detachments, limited to the area of the retinal surface overlying the plaque, that resolved spontaneously. Serial electroretinography was virtually indistinguishable between the 48 and 54 degrees C temperature groups. We noted a minimal alteration in a- and b-wave amplitudes with no changes in implicit times. Histopathology at 3 weeks post-treatment documented chorioretinal scarring overlying the thermal plaque treatment zone. No evidence of heamorrhage infection, cataract or scleral thinning was noted. This study documents the apparent focal containment of thermal effects with FM heating utilizing operating temperatures ad high as 54 degrees C for 60 min, and discloses no evidence of diffuse ocular toxicity.

Temperature distributions during thermoradiotherapy: a sensitivity study with a transient numerical model of the rabbit eye.

An approach to the treatment of medium-sized choroidal melanomas combines radiation with ferromagnetic hyperthermia. The study herein discusses results with a numerical thermal model of a choroidal melanoma in the rabbit eye as treated with episcleral, thermoradiotherapy plaques. The sensitivity of a temperature-dependent blood perfusion model is investigated.

Radiation therapy for squamous cell carcinoma in dystrophic epidermolysis bullosa: case reports and literature review.

Dystrophic epidermolysis bullosa (DEB) is a debilitating systemic disease frequently associated with biologically aggressive secondary squamous cell carcinomas arising from affected skin or mucosal surfaces. Treatment of these carcinomas with surgery, chemotherapy, or radiation is complicated by inherently poor wound healing. We report on two DEB patients treated with radiation therapy for locally advanced squamous cell carcinoma, and retrospectively analyze 10 DEB patients treated with radiation, reported in the literature. Of the 11 fully available and described case results from radiation therapy, six (54%) patients demonstrated a partial tumor response. All patients receiving > 4,500 cGy developed moist skin desquamation and delayed skin healing. Radiation therapy may be of benefit in palliating DEB patients who have locally advanced carcinoma, but has been associated with enhanced normal tissue toxicity, suggesting a narrow or absent therapeutic index between irradiated carcinoma and skin.

Radiation therapy and ferromagnetic hyperthermia in the treatment of murine transgenic retinoblastoma.

BACKGROUND: Combined modality therapy for childhood retinoblastoma holds the potential of decreasing treatment-related morbidity while maintaining excellent tumor control rates. OBJECTIVE: To evaluate the efficacy of external beam radiation therapy (EBRT), ferromagnetic hyperthermia (FMH), and the combination of both modalities in the control of ocular tumors in a transgenic murine model of retinoblastoma. METHODS: One hundred sixty-six mouse eyes from 4-week-old animals transgenically positive for simian virus 40 large T antigen were treated with a total dose of 10, 15, 20, 30, 40, 45, or 50 Gy of EBRT in 5-Gy fractions twice daily, with 48 degrees C or 54 degrees C FMH for 20 minutes, or with combined EBRT at 10 or 30 Gy and 48 degrees C or 54 degrees C FMH for 20 minutes. Serial histologic sections, obtained 8 weeks after treatment, were examined for the presence of tumor. RESULTS: The tumor control dose for 50% of eyes (TCD50) treated with EBRT occurred at 27.6 Gy. Ferromagnetic hyperthermia at 48 degrees C cured 30% (6/20) of eyes, while 54 degrees C FMH resulted in a 100% (20/20) cure rate. Combined treatment with 48 degrees C FMH and EBRT exhibited a TCD50 at 3.3 Gy. The thermal enhancement ratio was 8.4. Ferromagnetic hyperthermia at 54 degrees C exhibited tumor cure in all animals, but 25% of eyes were lost owing to secondary treatment complications. CONCLUSIONS: This represents the first documentation of tumor control via EBRT, ocular FMH, and a combination of these treatment modalities in this murine transgenic retinoblastoma model. The extent of treatment synergy in this model suggests that combined treatment application may allow a reduction in total ocular and periocular radiation dose while maintaining excellent local tumor control.

Thermoradiotherapy of intraocular tumors in an animal model: concurrent vs. sequential brachytherapy and ferromagnetic hyperthermia.

PURPOSE: To compare concurrent vs. sequential ferromagnetic thermoradiotherapy in vivo. METHODS AND MATERIALS: Greene melanomas were implanted subretinally in rabbits and observed until they were 3-5 mm in diameter. Episcleral plaques were assembled with 125I seeds for radiation therapy, or with ferromagnetic (FM) thermoseeds and nonradioactive I seeds for hyperthermia. Rabbits were implanted by centering a plaque over the intraocular melanoma. After a given dose of radiation had been delivered, the plaque was removed and a nonradioactive plaque containing FM thermoseeds was inserted into the same extrascleral space. One hour later, hyperthermia (46-47 degrees C at the plaque-scleral interface) was initiated and continued for a period of 1 h by placing the rabbits in a magnetic induction coil powered to 1200 W. Tumor size was determined at 1- to 2-week intervals by indirect ophthalmoscopy and by ultrasound. RESULTS: Dose-response analysis of 27 treated eye melanomas showed 50% local tumor control at 43 Gy for 125I alone and 29.4 Gy for 125I followed by FM hyperthermia. The thermal enhancement ratio was 1.4. CONCLUSION: Comparison with a previously published thermal enhancement ratio of 4.4 (for concurrent 125I and FM hyperthermia) leads us to conclude that thermal enhancement of 125I brachytherapy is more efficient in this tumor model system when hyperthermia is delivered during, rather than after, the irradiation process.

Retroperitoneal paragangliomas: natural history and review of the literature.

Retroperitoneal paragangliomas are uncommon neoplasms that arise from neural crest cells. Typical retroperitoneal sites of origin include adrenal medulla, organs of Zuckerkandl, and along sympathetic and parasympathetic nerves. We present a case of a woman diagnosed with retroperitoneal paraganglioma treated with debulking, intraoperative radiotherapy and external beam radiation therapy. She experienced a 7-year disease-free interval prior to symptomatic metastatic recurrence. Such remissions are consistent with the natural history for paragangliomas. The purpose of this report and literature review is to summarize the pertinent biologic and therapeutic options for these uncommon tumors.

The use of generalized cell-survival data in a physiologically based objective function for hyperthermia treatment planning: a sensitivity study with a simple tissue model implanted with an array of ferromagnetic thermoseeds.

PURPOSE: A physiologically based objective function for identifying a combination of ferromagnetic seed temperatures and locations that maximizes the fraction of tumor cells killed in pretreatment planning of local hyperthermia. METHODS AND MATERIALS: An objective-function is developed and coupled to finite element software that solves the bioheat transfer equation. The sensitivity of the objective function is studied in the optimization of a ferromagnetic hyperthermia treatment. The objective function has several salient features including (a) a physiological basis that considers increasing the fraction of cells killed with increasing temperatures above a minimum therapeutic temperature (Tmin,thera), (b) a term to penalize for heating of normal tissues above Tmin,thera, and (c) a scalar weighting factor (gamma) that has treatment implications. Reasonable estimates for gamma are provided and their influence on the objective function is demonstrated. The cell-kill algorithm formulated in the objective function is based empirically upon the behavior of published hyperthermic cell-survival data. The objective function is shown to be independent of normal tissue size and shape when subjected to a known outer-surface, thermal boundary condition. Therefore, fractions of cells killed in tumors of different shapes and sizes can be compared to determine the relative performance of thermoseed arrays to heat different tumors. RESULTS: In simulations with an idealized tissue model perfused by blood at various rates, maxima of the objective function are unique and identify seed spacings and Curie-point temperatures that maximize the fraction of tumor cells killed. In ferromagnetic hyperthermia treatment planning, seed spacing can be based on maximizing the minimum tumor temperature and minimizing the maximum normal tissue temperature. It is shown that this treatment plan is less effective than a plan based on seed spacings that maximize the objective function. CONCLUSIONS: It is shown that under the assumptions of the model and based on a desired therapeutic goal, the objective function identifies a combination of thermoseed temperatures and locations that maximizes the fraction of tumor cells killed.

Effect of interseed spacing, tissue perfusion, thermoseed temperatures and catheters in ferromagnetic hyperthermia: results from simulations using finite element models of thermoseeds and catheters.

Finite element heat-transfer models of ferromagnetic thermoseeds and catheters are developed for simulating ferromagnetic hyperthermia. These models are implemented into a general purpose, finite element computer program to solve the bioheat transfer equation. The seed and catheter models are unique in that they have fewer modeling constraints than other previously developed thermal models. Simulations are conducted with a 4 x 4 array of seeds in a multicompartment tissue model. The heat transfer model predicts that fractions of tumor greater than 43 degrees C are between 8 and 40% lower when seed temperatures depend on power versus models which assume a constant seed temperature. Fractions of tumor greater than 42 degrees C, in simulations using seed and catheter models, are between 3.3 and 25% lower than in simulations with bare seeds. It is demonstrated that an array of seeds with Curie points of 62.6 degrees C heats the tumor very well over nearly all blood perfusion cases studied. In summary, results herein suggest that thermal models simulating ferromagnetic hyperthermia should consider the power-temperature dependence of seeds and include explicit models of catheters.

Temperature-dependent versus constant-rate blood perfusion modelling in ferromagnetic thermoseed hyperthermia: results with a model of the human prostate.

Finite-element solutions to the Pennes bioheat equation are obtained with a model of a tumour-containing, human prostate and surrounding normal tissues. Simulations of ferromagnetic hyperthermia treatments are conducted on the tissue model in which the prostate is implanted with an irregularly spaced array of thermoseeds. Several combinations of thermoseed temperatures with different Curie points are investigated. Non-uniform, constant-rate blood perfusion models are studied and compared with temperature-dependent descriptions of blood perfusion. Blood perfusions in the temperature-dependent models initially increase with tissue temperature and then decrease at higher temperatures. Simulations with temperature-dependent versus constant-rate blood perfusion models reveal significant differences in temperature distributions in and surrounding the tumour-containing prostate. Results from the simulations include differences (between temperature-dependent and constant-rate models) in (1) the percentage of normal tissue volume and tumour volume at temperatures > 42 degrees C, and (2) temperature descriptors in the tumour (subscript t) and normal (subscript n) tissues including Tmax.t, Tmin.t and Tmax.n. Isotherms and grey-scale contours in the tumour and surrounding normal tissues are presented for four simulations that model a combination of high-temperature thermoseeds. Several simulations show that Tmin.t is between 1.7 and 2.6 degrees C higher and Tmax.n is between 2.1 and 3.3 degrees C higher with a temperature-dependent versus a comparable constant-rate blood perfusion model. The same simulations reveal that the percentages of tumour volume at temperatures > 42 degrees C are between 0 and 68% higher with the temperature-dependent versus the constant-rate perfusion model over all seed combinations studied. In summary, a numerical method is presented which makes it possible to investigate temperature-dependent, continuous functions of blood perfusion in simulations of hyperthermia treatments. Simulations with this numerical method reveal that the use of constant-rate instead of temperature-dependent blood perfusion models can be a conservative approach in treatment planning of ferromagnetic hyperthermia.

A practical alternative to conventional five-field irradiation postmastectomy for locally advanced breast cancer.

A combination of electron and photon beams has been used as an alternative for the conventional five-field method to irradiate patients postmastectomy for locally advanced breast cancer. Anterior and posterior opposed photon beams treat in continuity the lateral chest wall, axilla, and supraclavicular lymph nodes. An adjacent anterior electron beam is used at an energy matched to the depth of the internal mammary nodes. It includes the anterior chest wall, but bolus is used in the lateral aspect to spare underlying lung. This electron beam eliminates the diverging junction between the internal mammary and medial tangential fields used in the conventional five-field technique. Overlaps along the junction between the photon and electron beams are minimized by placing the center of the photon field along its medial border. Measurements with an Alderson-Rando phantom show dose-distribution advantages for this technique over the conventional five-field approach. There is less chance of underdosing tumor cells or of overdosing normal tissue along beam junctions. Clinical studies on 29 patients treated by this technique between July 1985 and December 1989 show increased rates of acute skin reactions, but otherwise similar side effects compared with 57 breast cancer patients treated with the five-field technique over the same time period. Local recurrence rates and patient survival rates were similar for the two groups. Given the dose-distribution advantages of this technique and its simple adaptation to accommodate unusual surgical scars or cancer recurrences, its use should be considered for postmastectomy patients with locally advanced breast cancer in well-equipped cancer treatment centers.

Concurrent ferromagnetic hyperthermia and 125I brachytherapy in a rabbit choroidal melanoma model.

Ferromagnetic (FM) thermoseeds and radioactive (125I) seeds were combined in an episcleral plaque to give concurrent hyperthermia and irradiation for enhanced tumour destruction. A Greene melanoma cell line was utilized to study the interaction between these treatment modalities. We attached five FM thermoseeds (with an operating temperature of 48 degrees C) in parallel with alternating rows of 125I seeds onto the inner surface of each 14 mm Silastic plaque. Plaques were centred over a 3-6 mm (diameter) intraocular melanoma in each rabbit. Some rabbits were then placed within a heating coil, and their eye tumours were warmed rapidly to therapeutic temperatures (43.6 degrees C across the tumour base) while the temperature of normal conjunctiva across the globe did not exceed 38.5 degrees C. Analysis of 49 treated eye melanomas showed 50% local tumour control at 41.7 Gy for 125I alone, whereas only 9.5 Gy were needed to give the same local control rate after 125I with concurrent FM hyperthermia. Thus, a thermal enhancement ratio of 4.4 was obtained. Hyperthermia alone gave a 20% tumour response rate, but responses were only temporary. We conclude that FM thermoseeds can be used to deliver biologically effective hyperthermia concurrently with radiation, thereby reducing the dose of radiation needed for tumour control.

Effect of implant variables on temperatures achieved during ferromagnetic hyperthermia.

Effects of ferromagnetic implant variables on steady-state temperature were studied in both in vitro (phantom) and in vivo (rabbit hind limb musculature) models. Thermoseed implant variables included: (1) the presence and number of thermoseed sleeves; (2) variations in thermoseed alignment within the oscillating electromagnetic field; (3) generator power levels of 300 W, 600 W, and 1200 W; and (4) separation of thermoseed tracks by 0.8 cm versus 1 cm. When the thermoseeds were aligned parallel to the electromagnetic field, temperature distributions in the in vivo model using bare thermoseeds and thermoseeds encased in a single sleeve (0.1 mm wall thickness) of polyethylene tubing were statistically higher than in tests performed with thermoseeds encased in a double sleeve (0.25 mm over 0.1 mm wall thickness) of tubing (p = 0.006). Nonetheless, average steady-state temperatures above a therapeutic minimum (greater than or equal to 42 degrees C) were achieved at all generator power levels using thermoseeds encased in a double sleeve of tubing and aligned parallel to the electromagnetic field. Gross misalignment of thermoseeds with the electromagnetic field was partly compensated for by utilizing higher generator power levels. Thermoseed tracks separated by 0.8 cm and aligned parallel to the electromagnetic field yielded average steady-state temperatures that were 0.4-2.2 degrees C higher than those obtained with a thermoseed track separation of 1 cm.

Hyperthermia and irradiation for locally recurrent previously irradiated breast cancer.

From March 1981 to June 1989, 44 patients with locally recurrent, previously irradiated adenocarcinoma of the breast were treated with hyperthermia and irradiation. Treatment sites were chest wall (35) or nodal (nine), with an average tumor area of 29.44 cm2 (range 0.16 to 252 cm2) prior to treatment. Concurrent radiation doses varied from 16 to 56 Gy (mean = 29.4 Gy). Externally applied microwave hyperthermia was given twice weekly, aiming at 43 degrees C for 60 min. Evaluation at one month post treatment revealed 41% complete response (CR), 23% partial response (PR), and 36% no response (NR), and 67% of patients with a CR sustained that response for greater than 12 months. Tumors heated to a mean thermal dose (equivalent-minutes at 42.5 degrees C) greater than 50 had a 53% CR rate, significantly better than the 14% CR rate observed in patients whose tumors received a mean thermal dose less than 50. Among patients with tumors less than or equal to 6 cm2 in area, 65% achieved CR, significantly better than the 26% CR rate noted for patients with tumors greater than 6 cm2 in area. Only four patients (7.4%) experienced complications: one developed a catheter-related infection, two had ulcerated infections, and one had a severe blister. In summary, higher thermal doses delivered and smaller tumor areas were associated with more favorable tumor responses.

Adjunctive therapy (whole body hyperthermia versus lonidamine) to total body irradiation for the treatment of favorable B-cell neoplasms: a report of two pilot clinical trials and laboratory investigations.

Based on earlier clinical and preclinical investigations, we designed two different pilot trials for patients with nodular lymphoma or chronic lymphocytic leukemia. These studies evaluated the use of either 41.8 degrees C whole body hyperthermia (WBH), or the nonmyelosuppressive chemotherapeutic drug, lonidamine (LON), as an adjunct to total body irradiation (TBI) (12.5 cGy twice a week, every other week for a planned total dose of 150 cGy). Whole body hyperthermia was initiated approximately 10 min after total body irradiation; lonidamine was administered orally (420 mg/m2) on a daily basis. Although entry to the studies was nonrandomized, the two patient populations were accrued during the same time frame and were comparable in terms of histology, stage of disease, performance status, and prior therapy. Of 8 patients entered on the TBI/WBH study, we observed 3 complete responses (CR), 4 partial responses (PR), and 1 improvement (i.e., a 48% decrease in tumor burden). Of 10 patients entered in the TBI/LON study, there was 1 CR and 4 PR. For the TBI/WBH study, myelosuppression was not treatment-limiting; there were no instances of infection or bleeding and platelet support was never required. The median survival time for the TBI/WBH study is 52.5 months based on Kaplan Meir estimates. Two patients remain in a CR. The median time to treatment failure (MTTF) is 9.4 months (90% confidence interval = 7-15.4 months). In the TBI/LON study, 50% of patients receiving TBI required treatment modification due to platelet-count depression during therapy, but there were no instances of infection or bleeding. Frequently observed LON-related toxicities included myalgias, testicular pain, photophobia and ototoxicity. For the TBI/LON study, median survival is 7.6 months; MTTF was 2.4 months. In analyzing the results of these pilot studies, our subjective clinical impressions lead to the hypothesis that WBH protected against TBI-induced thrombocytopenia during therapy, whereas LON had no effect on TBI-induced myelosuppression. This speculation was tested and confirmed in a series of in vitro and in vivo experiments.