Practical aspects of in situ 16O (gamma,n) 15O activation using a conventional medical accelerator for the purpose of perfusion imaging.

We report investigations into the feasibility of generating radioactive oxygen (15O, a positron emitter, with half-life 2.05 min) using a tuned Elekta SL25 accelerator, for the end purpose of imaging tumor perfusion. 15O is produced by the "gamma, neutron," (gamma,n) reaction between high-energy photons and normal oxygen (16O) in the body. As most in vivo 16O is bound in water molecules the 15O radio-marker is produced in proportion to water content in tissue. Imaging the washout of the 15O distribution using sensitive positron-emission-tomography (PET) technology can yield spatial information about blood perfusion in the tissue. The aim of this article was to determine the amount of 15O activity that could be produced by the tuned medical accelerator. A further aim was to model the activation process using Monte Carlo and to investigate ways to optimize the amount of 15O that could be generated. Increased activation was achieved by (i) tuning the beam to give higher-energy electrons incident on the target of the accelerator, (ii) increasing dose rate by removing the conventional filtration in the beam and reducing the source to object distance, and (iii) reducing low-energy photons by means of a carbon block absorber. The activity per-unit-dose produced by the tuned beam was measured by irradiating spheres of water to known doses and placing the spheres in a calibrated coincidence-counting apparatus. Peak energy of the tuned bremsstrahlung beam was estimated at 29 MeV, and generated activity up to 0.24/microCi/cc/3Gy in water. The measured amount of 15O agreed to within 10% of the prediction from the Monte-Carlo-computed spectrum, indicating reasonable ability to model the activation process. The optimal thickness of the carbon absorber was found to be about 25 cm. The insertion of a carbon absorber improved spectral quality for activation purposes but at the cost of reduced dose rate. In conclusion, the viability of generating 15O with an Elekta SL25 has been demonstrated. In conjunction with recent advances in high-sensitivity portable PET imaging devices, real potential exists for imaging in situ activated 15O washout as a surrogate measurement of macroscopic tumor perfusion.

A proposed standard data file format for hyperthermia treatments.

One of the most critical areas of research essential to improve the clinical use of hyperthermia in cancer therapy is the determination and evaluation of temperature during therapy. In order to improve the dissemination of newly developed thermometry measurement and analysis programs and facilitate the transport of hyperthermia data files between computers for both intra- and inter- institutional use, we propose a standard file format, the Hyperthermia Data Standard (HDS), which can be used for all hyperthermia data. This data file consists of segments of which nine have been defined. The first is an initial /VERSION segment which identifies the file standard. This is followed by eight major segments: 2) a /IDENT segment which contains file and patient identification information; 3) a /TEXT segment containing additional miscellaneous documentation; 4) a /TUMOR segment which provides tumor and treatment site information; 5) a /HSETUP segment which contains treatment setup information; 6) a /FORMAT segment which defines the data segment structure; 7) a /DATA segment containing the actual thermometry data; 8) a /FOLLOWUP segment for documenting patient response; and 9) an /ANALYSIS segment containing summary information. All segments begin and end with a specified delimiter and, except for the first two (/VERSION and /IDENT), may be repeated more than once. For ease in debugging and verifying adherence to the standard, all information in the file is encoded in printable ASCII characters. All segments consist solely of records containing KEYWORDS and KEYWORD VALUES which identify specified information. Certain KEYWORDS are standard, i.e., having specified names and information formats as defined in this document. The /DATA segment consists of ASCII encoded chronological treatment thermometry and other data whose format and structure is identified in the /FORMAT segment. The /ANALYSIS segment provides the flexibility to store the results of any analysis program along with the actual data file for ease of data management. This thermometry data file standard is designed to provide both flexibility and extendability for all possible forms of hyperthermia treatment data. Furthermore, it is designed to provide a simple format which may be easily read by higher level languages. This document is intended for commercial manufacturers and others who are writing programs to document clinical hyperthermia treatments with the intention that they either use this format for their data storage or provide a means to convert their data into this format.(ABSTRACT TRUNCATED AT 400 WORDS)

Thermal isoeffect dose: addressing the problem of thermotolerance.

A method of calculating a thermal isoeffect dose by converting thermal exposure into equivalent-minutes at 43 degrees C (EQ43) has been described previously by this investigator and others. Some investigators have suggested variations in the constants of this approach based on evaluations of the available in vivo data. The selection of these constants affect thermal dose calculations most at temperatures below 43 degrees C. Since treatment response appears to be most closely related to the dose in the coolest part of the tumour, the selection of appropriate constants may be quite important. The data suggest that these variations in constants are a consequence of thermotolerance. A more appropriate approach to address this problem is presented. The phenomena of thermotolerance complicates the practical application of this thermal isoeffect dose model. The dose modification caused by chronic thermotolerance which occurs during exposure at mild hyperthermic temperatures can be estimated by calculating the thermotolerance dose ratio (TTDR) between the equivalent-minute dose calculated with and without a transition temperature at 43 degrees C. The data suggest that the TTDR as a function of time can be mathematically described and used to correct for the dose modifying effect of thermotolerance. Using these results in a modification of the thermal isoeffect dose model causes a significant improvement in the fit for in vitro data below 43 degrees C.

Induction of prostaglandin production by hyperthermia in murine peritoneal exudate macrophages.

In response to inflammatory stimuli, macrophages synthesize and secrete prostaglandins (PGE) along with other potent inflammatory mediators. We have studied the effect of hyperthermia on the production of PGE by murine mononuclear phagocytes. Exposure to high temperature induced PGE production by cultured C3H/HeJ exudate macrophages in a time- and temperature-dependent manner. Increase in PGE production was detected when macrophages were treated at 41 degrees C and above for 1 h with a much greater increase at 42 and 43 degrees C. The secretion of PGE into culture supernatants by heat-treated macrophages reached a maximum approximately 24 h after heat treatment. The production of PGE by macrophages after hyperthermia was inhibited either by the addition of 5 X 10(-7) M indomethacin or by the subsequent incubation at 4 degrees C, suggesting that the elevated PGE production by macrophages is mediated through the activation of cyclooxygenase. Heat treatment under the same conditions failed to stimulate the production of PGE by either a human monocyte-like tumor cell line (U-937) or a mouse fibroblast cell line (L-929).

Fluorescence studies of macrophage recognition and endocytosis of native and acetylated low-density lipoprotein.

Macrophage recognition and endocytosis of 1,1'-dioctadecyl-3,3,3',3'-tetramethylindocarbocyanine perchlorate (diI)-labeled low-density lipoprotein (LDL) and acetyl LDL (Ac-LDL) was studied using fluorescence flow cytometry and fluorescence video intensification microscopy. RAW264 macrophages and U937 monocytes were grown in the tissue culture media in the presence and absence of LDL and Ac-LDL. Several lines of evidence indicate that receptor-mediated endocytosis of diI-labeled LDL or Ac-LDL was taking place. Binding can be distinguished from binding plus endocytosis by incubation at 4 and 37 degrees C, respectively. Binding was saturable at 4 degrees C and uptake at 37 degrees C was time- and ligand dose-dependent. Also, unlabeled LDL and Ac-LDL compete for their receptors. Macrophages grown in the presence or absence of LDL demonstrated distinct labeling patterns. LDL receptors were significantly increased by culture in defined medium without serum lipoproteins, while Ac-LDL receptors remained unaffected. Flow cytometry can provide an important tool to examine receptor levels, modulation of these levels and receptor-mediated endocytosis. Video intensification microscopy of similarly labeled cells has been performed. Receptors appear as punctate fluorescence, usually distributed randomly across the cell surface.

Correlations between 31P NMR spectroscopy and 15O perfusion measurements in the RIF-1 murine tumor in vivo.

The tumor physiological environment is one of the least understood and most important factors in determining the response of solid tumors to cancer therapy. To examine several important characteristics of the tumor physiological environment we have used in situ photon activation-15O decay measurements (perfusion characteristics) and 31P surface coil-NMR spectroscopy (metabolic characteristics) to observe in vivo subcutaneous RIF-1 tumors grown in female C3H/Anf mice. The following correlations between the 15O perfusion characteristics and the 31P NMR metabolic characteristics in individual tumors were observed: a negative correlation between pH, as measured by NMR (pHNMR), and the inorganic phosphate to nucleosides triphosphate peak height ratio (Pi:NTP); for the well-perfused fraction of the tumor there is a positive correlation with both pHNMR and the phosphocreatine to nucleosides triphosphate peak height ratio (PCr:NTP), and a negative correlation with Pi:NTP. These correlations are interpreted as evidence for a direct relationship between the distribution of cellular physiological environments and the tumor metabolic state. Because these physiological characteristics affect tumor response to various therapeutic modalities and both measurements can be made on humans, it is suggested that these techniques may be of prognostic value in the clinical management of human cancer.

Effects of sequencing of the total course of combined hyperthermia and radiation on the RIF-1 murine tumor.

The optimal sequence for clinical utilization of combined radiotherapy and hyperthermia is not known. The clinical trials have resulted in similar responses whether hyperthermia is given before or after radiation. Moreover, studies addressing the best sequence for an entire course of multifractionated hyperthermia and radiation are lacking. In these experiments, the importance of sequencing of heat and irradiation in a multifractionated treatment regimen in RIF-1 murine tumors was studied. It was observed that a close sequence of heat and irradiation is more beneficial than separate cytotoxic action. When heat and irradiation were given simultaneously, (within 1 hour) 67% to 75% of the tumors were cured. Heat and irradiation given sequentially (the entire course of one following the entire course of the other, each separated by 72 hours) cured 20% of the tumors. No tumors were cured when treated with heat or irradiation alone. The tumor regrowth time (mean tumor doubling time) is much longer in simultaneous treatment than in sequential treatment. It appears that heating first decreases the effectiveness of subsequent irradiation, causing a shorter growth delay than the opposite sequence. Heat alone does not alter the tumor bed permanently, but irradiation seems to do so, resulting in a slower rate of growth upon recurrence.

In vivo metabolic effects of hyperglycemia in murine radiation-induced fibrosarcoma: a 31P NMR investigation.

The hyperglycemia-induced in vivo metabolic changes produced in subcutaneous murine RIF-1 tumors, grown on female C3H/Anf mice, were examined with 31P surface-coil NMR. Serum glucose levels were elevated 4-fold by bolus intraperitoneal injection of 0.3 ml of an aqueous 50% glucose solution. Tumor pH was calculated from the chemical shift of Pi and relative phosphocreatine and ATP concentrations were determined by Simpson's rule integration of the peak areas. Tumor pH decreased by ca. 0.45 unit over 2 hr while phosphocreatine concentrations decreased by ca. 50% over the same time period (n = 9). Initial tumor pH correlated inversely with the initial peak intensity ratio of Pi:ATP (r = -0.77). In a significant number of tumors (n = 4), two pH populations were observed. In these tumors, one population was unaffected by hyperglycemia and the other showed a decrease in pH. In the other tumors (n = 5), the pH distribution broadened as the pH decreased. In these tumors, the observed decreased in phosphocreatine concentration correlated with that calculated from the effect of measured tumor pH on the intracellular creatine kinase equilibrium (n = 18, r = 0.91). This correlation and consideration of the Pi distribution in the tumor suggest that the pH measured by 31P NMR is weighted heavily by intracellular pH for the RIF-1 tumor. The presence of two distinct tumor pH populations or a broadened pH distribution likely reflects variations in tumor microcellular environment. Control experiments showed negligible changes in tumor pH and high energy phosphate concentrations after bolus intraperitoneal injection of 0.3 ml of isotonic saline. In addition, negligible changes in leg muscle pH and high energy phosphate concentrations were observed after glucose injection into mice with or without tumors. These results indicate that hyperglycemia induced by intraperitoneal glucose injection is effective in lowering the tumor pH of the murine RIF-1 tumor.

Thermal dose expression in clinical hyperthermia and correlation with tumor response/control.

Thermal dose has been identified as one of the most important factors which influence the efficacy of hyperthermia. Adequate temperature must be delivered for an appropriate period of time to the entire tumor volume in order to achieve optimal therapeutic results. Present clinical thermometry systems provide coarse temperature readings, since only selected tumor or normal tissue temperatures are monitored. Experimental in vitro and in vivo data suggest that the minimal temperature observed in the tumor determines therapeutic effectiveness. Unfortunately, at the present time, clinical data documenting these observations are scarce. The inhomogeneity of temperature distribution throughout the tumor volume makes difficult accurate correlations with tumor response and subsequent tumor control. Several mathematical models have been offered to express the time-temperature equivalency in relation to a reference temperature (43 degrees equivalent). Factors such as step-down heating, fractionated hyperthermia, thermal adaptation, and combination with irradiation, in addition to physiological parameters such as blood flow, play a major role in the expression of thermal dose. In order to meaningfully express thermal dose in clinical hyperthermia, several procedures are recommended, such as static phantom studies of specific absorption rate distributions for heat delivery equipment, detailed thermal mapping in hyperthermia sessions, development of reliable predictive biomathematical models to express temperature-time equivalency, and the fostering of research in 3-dimensional noninvasive clinical thermometry.

Thermal dose determination in cancer therapy.

With the rapid development of clinical hyperthermia for the treatment of cancer either alone or in conjunction with other modalities, a means of measuring a thermal dose in terms which are clinically relevant to the biological effect is needed. A comparison of published data empirically suggests a basic relationship that may be used to calculate a "thermal dose." From a knowledge of the temperature during treatment as a function of time combined with a mathematical description of the time-temperature relationship, an estimate of the actual treatment calculated as an exposure time at some reference temperature can be determined. This could be of great benefit in providing a real-time accumulated dose during actual patient treatment. For the purpose of this study, a reference temperature of 43 degrees C has been arbitrarily chosen to convert all thermal exposures to "equivalent-minutes" at this temperature. This dose calculation can be compared to an integrated calculation of the "degree-minutes" to determine its prognostic ability. The time-temperature relationship upon which this equivalent dose calculation is based does not predict, nor does it require, that different tissues have the same sensitivity to heat. A computer program written in FORTRAN is included for performing calculations of both equivalent-minutes (t43) and degree-minutes (tdm43). Means are provided to alter the reference temperature, the Arrhenius "break" temperature and the time-temperature relationship both above and below the "break" temperature. In addition, the effect of factors such as step-down heating, thermotolerance, and physiological conditions on thermal dose calculations are discussed. The equations and methods described in this report are not intended to represent the only approach for thermal dose estimation; instead, they are intended to provide a simple but effective means for such calculations for clinical use and to stimulate efforts to evaluate data in terms of therapeutically useful thermal units.

Differential effect of hyperthermia on murine bone marrow normal colony-forming units and AKR and L1210 leukemia stem cells.

Thermal dose-survival curves for normal hematopoietic and leukemia cells were assessed by spleen colony assays after in vitro heat exposure ranging from 41 degrees to 45 degrees. No effect of 43 degrees heat treatment on the fraction of cells lodging in the spleen was observed. Marked differences in heat sensitivity were observed between normal, L1210, and AKR leukemia cells, the first being les sensitive than were the malignant cells. Furthermore, a greater relative difference between normal stem cells and leukemia cells was observed at lower temperatures. Normal bone marrow cells forced into regenerative activity prior to heat treatment were more heat sensitive than was their undisturbed counterpart, suggesting that noncycling hematopoietic cells are less heat sensitive than are proliferating cells.

Use of the Clinac-35 for tissue activation in noninvasive measurement of capillary blood flow.

Through adjustment of operating parameters responsible for the acceleration, steering, and focusing of electrons en route to the target, we have extracted a 30-MV x-ray beam from the Clinac-35 linear accelerator that is suitable for use in photon activation -15O decay studies of tissue perfusion. This beam is significantly more efficient than the "standard" 25-MV beam in producing 15O in situ through activation of tissue oxygen, thereby substantially reducing the dose to tissue required to yield a desired initial 15O activity. Production and characteristics of the higher energy beam are discussed, and data obtained from its application to measurement of capillary blood flow in animal tumors are presented and analyzed.

The effect of chronic and acute heat conditioning on the development of thermal tolerance.

Survival studies with Chinese hamster ovary cells showed that thermal tolerance, which developed during chronic heating (treatment times greater than or equal to 1 hr) or after acute heating (treatment times less than 1 hr) involves similar mechanisms. For example, cells that expressed thermal tolerance during a 6-14 hr chronic heat treatment at 41.5 degrees C or 42 degrees C also expressed thermal tolerance to a subsequent acute treatment at 45.5 degrees C. Also, cells heated acutely for 10 min at 45.5 degrees C and incubated at 37 degrees C for 12 hr showed tolerance to both 45.5 degrees C acute and 42 degrees C chronic hyperthermia. Finally, thermal tolerance developed between fractionated acute heat treatment at 45.5 degrees C and fractionated chronic heat treatments at 42.5 degrees C. These data indicate that when cells are tolerant to chronic hyperthermia they are also tolerant to acute hyperthermia and that the reverse is also true.