Differentiation therapy is potentiated by chemotherapy and hyperthermia in human and canine brain tumor cells in vitro.

Human glioblastoma (U-87MG) and canine glioma (canine brain tumor [CBT]) cell lines were tested in vitro for their therapeutic sensitivity to sequential treatment with differentiating agents and chemotherapy or hyperthermia. Both cell lines responded to the inducer combination dibutyryl adenosine-3',5'-cyclic monophosphate/sodium butyrate by the formation of cytoplasmic processes detectable within 7 hours and attained approximately 90% morphological differentiation within 2 days of exposure. The clonogenicity of CBT and U-87MG cells gradually decreased after 1 to 7 days of exposure to the inducer combination, but this treatment alone failed to kill the cells. After the removal of the inducers, both lines dedifferentiated and the rate of clonogenesis increased. 1,3-bis-(2-Chloroethyl)-1-nitrosourea administered to CBT and U-87MG cells before or after 3 days of treatment with inducers potentiated the antiproliferative effects of the differentiating agents. Cisplatin administered to U-87MG cells enhanced the antiproliferative effect of the differentiating agents to a greater extent when added before the inducers rather than after differentiation was stimulated. The sequential treatment of CBT cells with a 44 degrees C heat pulse for 30 minutes followed by differentiating agents produced an additive potentiation of cell killing, whereas the reverse sequence did not. Hyperthermia pretreatment at 44 degrees C for 15 minutes or at 42 degrees C for 30 minutes failed to enhance the antiproliferative effects of inducing agents.(ABSTRACT TRUNCATED AT 250 WORDS)

In vitro response of human glioblastoma and canine glioma cells to hyperthermia, radiation, and chemotherapy.

Little is known about the sensitivity of human glioblastoma cells to hyperthermia alone and in combination with other therapies. We carried out in vitro cell survival studies on the human glioblastoma cell line U-87MG and our model canine glioma canine brain tumor (CBT) cells after multimodality treatment. Ionizing radiation was administered to flasks of cells in logarithmic growth at 500 rads (5 Gy) with consecutive treatment by hyperthermia, 1,3-bis-(2-chloroethyl)-1-nitrosourea (BCNU), or cisplatin. Cells were treated with single doses of BCNU at 5 microM with sequentially added radiation or hyperthermia and at 1 to 2 micrograms/ml of cisplatin with hyperthermia. Hyperthermia was administered in a precision controlled water bath at 44 degrees C for 30 minutes in combination with chemotherapy or radiation. In general, the sensitivity of U-87MG and CBT cells was similar for all test regimens. For example, colony formation efficiency decreased by 64% in CBT cells and by 64.4% in U-87MG cells after hyperthermia alone at 44 degrees C for 60 minutes. All combinations of BCNU, hyperthermia, and radiation administered in vitro produced enhanced cell killing, but the effects of multiple modalities were generally additive in both cell lines.(ABSTRACT TRUNCATED AT 250 WORDS)

Blood-brain barrier alteration after microwave-induced hyperthermia is purely a thermal effect: I. Temperature and power measurements.

The effect of microwave-induced hyperthermia on the blood-brain barrier was studied in 21 Sprague-Dawley rats. Under sodium pentobarbital anesthesia, animals were place in a stereotactic frame, and an interstitial microwave antenna operating at 2450 MHz was inserted in a bony groove drilled parallel to the sagittal suture. Some antennae were equipped with an external cooling jacket. Temperature measurements were made lateral to the antenna by fluoroptical thermometry, and power was calculated from the time-temperature profile. Five minutes prior to termination of microwave irradiation, horseradish peroxidase (1 mg/20 g body weight) was injected intravenously. Extravasation of horseradish peroxidase was observed in brain tissue heated above 44.3 degrees C for 30 minutes and at 42.5 degrees C for 60 minutes. Microwave irradiation failed to open the blood-brain barrier when brain temperatures were sustained below 40.3 degrees C by the cooling system. Extravasation of blood-borne peroxidase occurred at sites of maximal temperature elevation, even when these did not coincide with the site of maximum power density. The data suggest that microwave-induced hyperthermia is an effective means for opening the blood-brain barrier and that the mechanism is not related to the nonthermal effect of microwaves.

Alumina ceramic as a biomaterial for use in afterloading radiation catheters for hyperthermia.

A major technical challenge to the use of interstitial hyperthermia in malignant brain tumors is the production of a well-defined, uniform hyperthermal field. In theory, A 915-MHz microwave antenna should allow fewer antennas to be used and cause less mechanical brain damage; however, standard radiation afterloading catheters require antennas to be 12 cm long; this is clearly impractical for intracranial use. Since alumina ceramic (Al2O3) catheters permit short microwave antennas (3-5 cm in length) to function properly in neural tissue, it is important to test the biocompatibility of alumina for use in combined interstitial microwave hyperthermia and brachytherapy. A 5-mm length of alumina catheter was implanted into the brains of 15 white rats. The animals were killed at 3, 7, 14, 28, and 56 days. Histological examination revealed only minor mechanical damage and no encapsulation until 1 month; even then, the glial wall was only a few cell layers thick. Five animals received implants and were killed at similar intervals for x-ray microanalysis with the scanning electron microscope. No migration of aluminum into the brain was detected when compared with two control animals that did not receive implants and an alumina blank. Although we measured 50% attenuation of the radiation from iridium-192 sources in alumina catheters as compared with conventional ones, alumina catheters can still be used for interstitial radiation by increasing either the activity of the seeds or the duration of treatment.

Angioarchitecture of the CNS, pituitary gland, and intracerebral grafts revealed with peroxidase cytochemistry.

Blood vessels of the fetal, neonatal, and adult subprimate and primate CNS, including circumventricular organs (e.g., median eminence, pituitary gland, etc.), and of solid CNS and nonneural (anterior pituitary gland) allografts placed within brains of adult mammalian hosts were visualized with peroxidase cytochemistry applied in three ways: to tissues from animals injected systemically with native horseradish peroxidase (HRP) or peroxidase conjugated to the lectin wheat germ agglutinin (WGA) prior to perfusion fixation; to tissues from animals infused with native HRP into the aorta subsequent to perfusion fixation; and to tissues from animals fixed by immersion and incubated for endogenous peroxidase activity in red cells retained within blood vessels. In neonatal and adult animals receiving native HRP intravascularly, non-fenestrated vessels contributing to a blood-brain barrier were outlined with HRP reaction product when tetramethylbenzidine (TMB) as opposed to diaminobenzidine (DAB) was used as the chromogen; fenestrated vessels of circumventricular organs were not discernible due to the density of extravascular reaction product. Fenestrated and non-fenestrated cerebral and extracerebral blood vessels exposed to bloodborne WGA-HRP were visible when incubated in TMB and DAB solutions. Native HRP infused into the aorta of fixed animals likewise labeled non- fenestrated vessels throughout the brain upon exposure to TMB or DAB but obscured fenestrated vessels of the circumventricular organs. Endogenous peroxidase activity of red cells, seen equally well with TMB and DAB, outlined blood vessels throughout the cerebral gray and white matter and all circumventricular organs in fetal, neonatal, and adult animals. Application of the three peroxidase cytochemical approaches to study the development or absence of a blood-brain barrier in intracerebral allografts demonstrated that the vascularization of day 16-19 fetal/1 day neonatal CNS allografts is not well defined prior to 7 days following intracerebral placement of the grafts. CNS allografts secured from donor sites expected to possess a blood-brain barrier exhibited blood vessels that were not leaky to HRP injected intravenously in the host. Fenestrated blood vessels associated with anterior pituitary allografts were evident prior to 3 days posttransplantation within the host brain and permitted blood-borne HRP in the host to enter the graft and surrounding host brain parenchyma.

The morbidity and mortality of brain tumors. A perspective on recent advances in therapy.

This article reviews current morbidity and mortality statistics for the major classes of primary brain tumors including malignant astrocytoma, glioblastoma, low-grade astrocytoma, oligodendroglioma, meningioma, and other benign tumors and metastatic tumors. Innovations in therapy are discussed for surgery, radiation, chemotherapy, and such newer areas as hyperthermia, immunotherapy, and phototherapy.

Interstitial microwave hyperthermia for brain tumors. Results of a phase-1 clinical trial.

The technical feasibility and clinical safety of interstitial microwave hyperthermia was evaluated in six patients with glioblastoma and malignant astrocytoma. Prior to entry into the study, each patient had received surgery, radiation and nitrosourea chemotherapy. All patients were implanted at open craniotomy with a flexible microwave radiator/sensor (o.d. 1.5 mm) and transcutaneously connected to a 245- MHz microwave generator. Intraoperative thermal field plots and cooling curves were obtained with the aid of non-perturbing probes (o.d. 1.2 mm) perpendicularly driven into the tumor at fixed radial distances from the central antenna. In comparison to similar measurements carried out in normal feline brains, human gliomas were unable to efficiently dissipate heat as demonstrated by doubling of the effective diameter of the thermal field to 4 cm and by prolongation of the decay time in all cooling curves. Patients were also implanted with subarachnoid ICP monitors over the contralateral hemisphere. Two postoperative treatments were given at 45 degrees C for 60 min on the night of surgery and 48 hr later. No patient was aware of power on/power off, there were no permanent neurologic sequelae and there were no significant changes in the ICP. Power was manually controlled with visual feedback in the first three patients and automatically controlled by a computer-based system in the final three patients. Four of the six patients have lived 18 months after implantation and two of these have negative CT scans at 18 and 27 months since recurrence. It appears that interstitial microwave hyperthermia is both feasible and safe within the intracranial cavity and that combined interstitial irradiation and hyperthermia deserves clinical study.

Hyperthermia for brain tumors: biophysical rationale.

Hyperthermia has great potential as an antineoplastic agent because: (a) it is effective against relatively radioresistant hypoxic cells and cells in S phase; (b) unlike most chemotherapeutic agents, it is effective against poorly vascularized and metabolically quiescent tissues; (c) as a physical agent, its biological effect is related to the duration and intensity of its application; (d) it seems to have no cumulative toxicity; and (e) it potentiates the effects of both chemotherapy and ionizing radiation at the cellular level. The use of hyperthermia for malignant brain tumors is constrained by a relatively narrow therapeutic index and the considerable thermal sensitivity of normal neural tissue. Glioblastoma multiforme, by virtue of its low growth fraction and heterogeneous cell populations, seems to be an ideal candidate for hyperthermia administered as part of a combined modality treatment program. Focal hyperthermia can be produced by a number of energy sources, including those utilizing ultrasound, microwave, and radiofrequency generators. The clinical safety and feasibility of a miniature microwave radiator/sensor system for direct implantation have been demonstrated. In comparison to normal feline brain, malignant brain tumors in humans are unable to dissipate heat efficiently.

Clinical hyperthermia trials: design principles and practice.

The initial development of a significant risk device is predicated upon demonstrable medical need and an adequate biophysical rationale. Although microwave and radiofrequency thermotherapy systems strongly meet these requirements, their clinical evaluation has often suffered from inadequate or inappropriate preclinical laboratory experimentation. It is suggested that devices be tested in animals rather than phantoms and that thermal profiles, cooling curves, power vs. temperature studies et al be carried out in the organ or organs of interest. Clinical trials can then be designed in which an attempt is made to replicate laboratory measurements in humans in order to develop physical dose-response relationships and toxicity data (i.e. a phase-I or feasibility study). The experimental paradigm for clinical drug testing can also be applied, with some modifications, to the further evaluation of devices for the determination of therapeutic response rates (phase-II) and for controlled evaluation against available treatments in a homogeneous patient population (phase-III). It is extremely important that early clinical trials not be contaminated by the possible effects of other concurrent therapies and that sophisticated statistical design be employed to protect human subjects from unnecessary exposure to experimental treatments. The ethical issues involved are best dealt with by good scientific design.