Search

A logical inference from the recent reports indicating that malignant brain tumors are composed of a heterogeneous cell population is that combination chemotherapy will be required for effective brain tumor control. For several years we have been investigating the use of Bleomycin as an agent to be used in conjunction with radiation therapy and a nitrosourea compound. Since systemically administered Bleomycin does not cross the blood-brain-barrier and has significant toxicity when used parenterally in high doses, we have studied the use of smaller doses of Bleomycin injected directly into the brain tumor cavity. Such an intracerebral dose was more effective in prolonging survival of rats burdened with experimental 9L gliosarcomas than an intravenous dose that is 25 times as great. The combination of intracerebrally administered Bleomycin and radiation therapy was more effective than either modality alone. Furthermore, the combination of Bleomycin delivered intracerebrally and BCNU given systemically was more effective than eitheragent used alone. Finally, in a Phase I clinical trial of Bleomycin given via an Ommaya reservoir to eight patients with recurrent malignant brain tumors, we have demonstrated that individual doses of up to 7.5 units and cumulative doses of up to 255 units can be administered without significant toxicity.

The therapeutic efficacy of combined modality treatment with radiation therapy and bleomycin was investigated in rats burdened with the intracerebral 9L gliosarcoma. Both radiation (single or fractionated exposures) and bleomycin (injected intracerebrally directly into the tumor region) are effective in prolonging survival when used as single agents. Bleomcyin (1.0 mg/kg/week) combined with low-dose radiation therapy (15.3 Gy in 6 fractions in 2 weeks) prolonged survival over that of radiation alone, but not to the extent of high-dose radiation therapy (30.6 Gy in the same schedule). Bleomycin was effective whether given simultaneously or following fractionated radiation therapy-the important factor being delivery of radiation therapy early in the disease process. The greatest enhancement in survival caused by combination therapy compared to that by single agent therapy was observed when single exposure radiation therapy (20 Gy) followed single bleomycin administration by 4 hr. These results suggest the possibility of using bleomycin as an adjunct to radiation therapy for the treatment of patients with malignant brain tumors.

The growth curve of monolayer cultures of ARH-77 cells, a human myeloma cell line propagated in vitro, is represented by an everbending curve on a semilogarithmic plot; however, the curve can be fitted by a straight line on a linear—linear plot. This unusual growth pattern suggests that, instead of a fixed proportion of the population, a fixed number of ARH-77 cells divide per unit time. The following are cell cycle transit time parameters calculated from percent labeled mitosis experiments: TG1, 10.0 ± 3.5 hours; TS, 14.3 ± 2.3 hours; TG2, 4.3 hours; TM, 1.4 ± 1.3 hours; and TC, 30.0 ± 6.1 hours. For cells exposed continuously to 3H-thymidine the values are: growth fraction, 67%; TG1, 6.5 hours; TS, 13.0 hours; and TG2 + M, 3.0 hours. The average doubling time is 4.6 days (range, 3.8–4.7 days); after about 10 to 15 days in culture, the growth rate of freshly passaged cells declines markedly, as reflected by a growth curve with a much shallower slope. The changes are accompanied by a marked decline in the labeling index from 41.3% (range, 28.9%–53.7%) during the first 3 days of culture to less than 5% measured on day 21. Flow cytometry for DNA content-dependent cell cycle compartment distribution demonstrates an obvious decline in the proportion of S-phase cells and a marked accumulation of G2 phase cells as the cultures age. When the supernatant medium of ARH-77 cells grown for 10 days is replaced by fresh medium, a new burst of vigorous cellular growth is observed with a curve slope similar to that observed during the first 5 days of culture. If the 10-day-old supernatant medium is used to set up cultures with freshly harvested ARH-77 cells, their growth curve resembles that of 10-day-old cultures. However, this supernatant medium induces no decrease in the growth rate of other human tumor cells, suggesting that inhibition of cellular growth does not result from exhaustion of nutrients, but that ARH-77 cells produce a molecular mediator that specifically inhibits the growth of these cells. ARH-77 cells could be synchronized with a single treatment of 3 or 5 mM thymidine; (dThd) and cloning efficiency was 2% to 4% in a double-layer soft agar assay. Treatment for 1 hour with increasing concentrations of melphalan produced a threshold exponential survival curve (Dq = 0.45 μg/ml and D0 = 0.35 μg/ml, 1 hour). Heterotransplantation into nude mice and rats failed for every route (subcutaneous, intravenous, intraperitoneal, and intrafemoral) except the intracerebral route.

Dihydroxyanthraquinone (DHAQ, NSC 279836, a new cancer chemotherapeutic agent presently in clinical trials) was tested, alone and in combination with radiation, for therapeutic effectiveness in a rat solid tumor model. The Walker 256 fibrosarcoma was implanted subcutaneously in the leg via trochar so as to yield palpable tumors within 7-11 days. When tumor diameters reached 1 cm, the animals were treated as follows: as controls; with DHAQ (3.0 mg/kg, i.p.); with 300 KVP x-rays (15 Gy to the tumor-burdened leg); or with a combination of DHAQ and radiation. Tumor diameters were measured three times a week to monitor the response to therapy. DHAQ alone had no effect on tumor growth rate. Radiation alone produced a delay in tumor growth. The combination of DHAQ and radiation resulted in a consistently better therapeutic response than did radiation only, but at only a marginal level of statistical significance (0.05 less than p less than 0.10). However, there were 3/10 long-term survivors (greater than 80 days post treatment) without evidence of tumor in the combination therapy group; whereas all animals in the radiation group died or were sacrificed because of progressive tumor growth by day 71. These results suggest that even though DHAQ is only minimally effective against this rat solid tumor when used alone, there may still be some added therapeutic benefit derived from a combination of DHAQ and radiation therapy.

Serum levels of angiotensin-I-converting enzyme (ACE) were assayed before, during, and after radiation therapy in 209 patients receiving treatment for neoplastic disease. Daily fluctuations in the measured ACE levels were minimized by comparing all patient values to that of a simultaneously run sample from a standard source of serum obtained by pooling sera from young, healthy volunteers. Most of the patients tested presented with a normal to low ACE level, with the mean value for all patients being 70% of the standard value. More patients with primary lung cancer displayed values that were low (107 of 120) than did patients with disease outside the lung (44 of 78), this difference being statistically significant at p less than 0.001. In addition, more patients with lung cancer had values less than 70% of the standard than did patients with disease outside the lungs. These initial results suggest that monitoring of serum ACE levels may be useful in the management of patients with malignant disease in the lung.

The cytotoxic activity of various chemotherapy agents was investigated in asynchronous populations of cultured 9L rat brain tumor cells, and as a function of their position in the cell cycle. Representative drugs from the classes of DNA-active agents, alkylating agents, spindle poisons, and antimetabolites were tested. The ability to induce cell lethality in asynchronous populations as a function of drug concentration varied for 1 hr pulse exposures. In order of decreasing cytotoxic activity, DHAQ was the most effective, followed by VCR, VDS, VBL, ADR, BCNU, cis-DDP, BLM, DBD, RZ, and HU. The effect of chemotherapy agents on synchronous 9L cells obtained by mitotic selection also varied with respect to the individual agent and was cell cycle-dependent. Survival age-responses ranged from being minimal to demonstrating significant fluctuations as a function of cell cycle position. For all agents except ADR and HU, the sensitivity of G1 phase was greater than S phase. RZ exhibited essentially a flat age-response. Comparison of the cell cycle age-responses of chemotherapeutic agents to those exhibited by the cytotoxic modalities of radiation and hyperthermia demonstrate several unique differences.

The present report describes the fabrication technique and the dosimetry aspects of a new compensator system that uses the low melting point allow Lipowitz metal. Compensating ratios (CR, mm of tissue compensated per mm of cerrobend) were determined for various field sizes and depths for 60 Co, and X ray energies of 4, 6, 18, and 25 MV. Typical CR for 10 cm × 10 cm field and 10 cm depth were: 60 Co, 4 MV, and 6 MV,1:15;18 and 25 MV, 1:20. Verification of these CR were performed at 6 MV using polystyrene phantoms. Ionization measurements were made for various field sizes and depths and normalized to central axis full-phantom readings for both compensated and non-compensated fields. Without compensation, percent differences ranged as high as 40% for a tissue deficit of 10 cm. With the compensating filters (CF) in place, this difference was reduced to 2–4% Overall, the CF system was capable of producing dose uniformity to within ±10% for a wide range of depths and field sizes.

Over the past 2 years, an afterloading technique has been developed and refined to implant radioactive Ir-192 sources into brain tumors. The implantation procedure integrates a stereotaxic system with computerized tomography (CT), which provides tumor position, volume, and guides the placement of catheters. A radiolucent ring-frame immobilizes the head as holes are made at 1 cm intervals with the aid of a template. Catheters containing dummy sources 1 cm apart are then inserted to the desired depth, and their position verified in three dimensions to insure complete coverage of visible tumor volume as defined by contrast enhancement. Once catheters are secured, the anesthetized patient is moved to the intensive care unit where the dummy sources are replaced by ribbons of Ir-192 seeds (specific activity 0.6–1.0 mg Ra eq). CT scans with the dummy sources in place are used to designate spatial coordinates of the active sources. A computer program converts position data and source strength into isodose contours in any plane. The implant duration (70–100 hours) for the desired dose to the tumor periphery (60–120 Gy) is then calculated. Dose rate contours are superimposed on preimplant CT scans. Maximum and minimum doses are determined in each of the various planes. Verification dosimetry has been carried out with thermoluminescent dosimeters placed in a catheter located in a plane along the tumor periphery. In vivo isodose values compared to idealized plans agree within ±5%–10%.

At the University of California Lawrence Berkeley Laboratory and the Northern California Oncology Group, a preliminary study of heavy charged particle radiotherapy in soft tissue and bone sarcoma has been carried out. Fifty-two patients with bone or soft tissue tumors were treated wholly or in part with heavy charged particles from 1978 to 1985. Eleven patients, considered inevaluable for purposes of this analysis, received less than 50 Gray-equivalents (GyE) because of the following: progressive disease (three patients); palliative treatment due to recurrent disease after previous radiation therapy (three patients); or since they were part of preliminary studies of relative biological effectiveness (RBE) (five patients). Forty-one patients received from 50 to 78.5 GyE, with a mean of 65 GyE. They had an average of 23 months follow-up, ranging from 4 to 78 months. In patients with paraspinal chondrosarcoma 9 of 11 had local control, with a mean follow-up time of 32 months. In the remaining patients with other histologies, 19 of 30 were controlled within the irradiated area, with a mean follow-up time of 20 months. Serious complications were encountered in the CNS (four patients), in the bowel (one patient), and in bone (one patient). Heavy charged particle radiotherapy appears to be of value in treating bone or soft tissue sarcoma; further trials are planned.

In order to measure respiration rates in rats irradiated to the whole thorax, a plethysmograph involving a microphone with extended low‐frequency response and an airtight, whole‐body chamber has been constructed. The microphone response to the respiration pressure waves created by the rats placed in the chamber is markedly greater in amplitude than the response to laboratory sounds and animal noises; thereby filtering complexities are avoided. The output from the microphone went to a frequency counter for measurement of the number of pressure waves per unit time. Control, unirradiated rats, between the ages of 3 and 18 months, demonstrated a respiration rate in the range of 130–160 breaths per minute. Rats that received thoracic radiation doses greater than 12 Gy displayed increased respiration rates and occasional abnormal respiration patterns. These results demonstrate that this whole‐body plethysmograph incorporating a sensitive microphone and simple electronics can be used for the measurement of respi...

The effects of radiation therapy on normal rats and on rats burdened with 9L brain tumors have been studied. The heads of normal rats were X-irradiated with single exposures ranging from 1000 R to 2700 R. Followiag acute exposures greater than 2100 R, all animals died in 8 to 12 days. Approximately 30% of the animals survived beyond 12 days over the range of 1950 to 1950 R; following exposures less than 1850 R, all animals survived the acute radiation effects, and median survival times (MST) increased with decreasing exposure. Three fractionated radiation schedules were also studied: 2100 R or 3000 R in 10 equal fractions, and 3000 R in 6 equal fractions, each schedule being administered over a 2 week period. The first schedule produced a MST of greater than 1 1 2 years; the other schedules produced MSTs that were lower (192 and 145 days, respectively). It was determined that by applying a factor of 1.9, similar survival responses of normal rats were obtained with single as with fractionated radiation exposures. Animals burdened with 9L gliosarcoma brain tumors normally died of the disease process within 18–28 days after tumor inoculation. Both single and fractionated radiation therapy resulted in a prolongation of survival of tumor-burdened rats. This prolongation was found to be linearly dependent upon the dose; but only minimally dependent upon the time after inoculation (7–12 days) at which therapy was initiated, or upon the fractionation schedule that was used. As with normal animals, similar responses were obtained with single as with fractionated exposures when a factor (1.9) was applied. All imam-hearing animals died prior to the time that death was observed in normal, irradiated rats. Thus, the 9L gliosarcoma rat brain tumor model can be used for the pre-clinical experimental investigation of new therapeutic schedules involving radiation therapy and adjuvant therapies.

Forty-five consecutive patients with chordoma or chondrosarcoma at the base of skull or cervical spine were treated at the University of California Lawrence Berkeley Laboratory (UCLBL) and University of California School of Medicine, San Francisco (UCSF) between November 1977 and October 1986. All patients had undergone a subtotal surgical resection. Twenty-three patients were treated definitively with charged particles, 13 patients were treated with photons and particles, and 9 patients were treated for recurrent disease. Total doses ranged from 36 to 80 Gray equivalent (GyE). Thirty-three patients are alive with a minimum followup of 1 year. The actuarial survival and local control for all patients at 5 years is 62% and 59%, respectively. Patients treated for primary disease had a 78% actuarial local control rate at 2 years, whereas the rate for patients with recurrent disease was 33%. Patients with smaller visible tumor volumes (less than 20 cc) had a significantly better local control rate than patients with larger tumor volumes (80% vs 33% actuarial rate at 5 years). Patients with chondrosarcoma had the highest local control rate, as did patients treated with particles alone. Complications included 3 patients with unilateral visual loss, two patients who became blind, and 4 patients with radiation injury to the brainstem.

The response of cultured 9L rat brain tumor cells to hyperthermia (45/sup 0/C water bath) and ionizing radiation was investigated as a function of their position in the cell cycle. Similar to other cell lines, 9L cells demonstrated a sensitivity to cell killing by hyperthermia during S phase, and in particular late S phase, in contrast to high survival in early G/sub 1/ phase. Mitotic cells treated prior to selection under proper pH and temperature conditions displayed a survival that was comparable to that of S-phase cells. Using the method of retroactive synchrony, the thermal sensitivity during mitosis was also shown to fluctuate, being greater for cells closer to division than those near the G/sub 2//M boundary. Irradiated 9L cells exhibit only minimal changes in survival across the majority of the cell cycle (except for a radiosensitive mitosis). Combining the two agents resulted in survival levels that were less than expected from the product of the survivals after individual treatments. The degree of enhancement was greatest for those cells in mid-G/sub 1/ phase and S phase. When the two treatments were separated in time, there was an increase in survival that was greater when X rays preceeded heat than formore » the reverse sequence. The degree of recovery was greater for cells initially treated at the S/G/sub 2/ boundary, than at the G/sub 1//S boundary, than at mid-G/sub 1/. Recovery to an independent level was reached within 1 hr for all positions except mid-G/sub 1/ when radiation preceeded hyperthermia; for the reverse sequence, recovery to an independent level was not observed over a 4-hr period.« less

Several responses of cultured 9L rat brain tumor cells to ionizing radiation were investigated as a function of their position in the cell cycle. The mitotic selection procedure for cell cycle analysis was utilized to study the blockade of progression of cells through G/sub 2/ and the resultant division delay. The transition point for this blockade (i.e., the last point in the cell cycle at which cells are blocked, and after which they are refractory to delay) was located approximately at the G/sub 2//M boundary 35 min prior to selection. The duration of division delay was a linear function of the x-ray dose, 33 min of delay/Gy. The survival of cells following exposure to a constant dose of x rays at varous times after incubation of mitotically selected cells demonstrated a definite resistance during the G/sub 1/ phase and slight resistance during the S phase, relative to the level of survival at the G/sub 1//S boundary. As shown with other cell lines there was maximum radiosensitivity during G/sub 2/ and mitosis. Thus the response of 9L rat brain tumor cells in culture to ionizing radiation is similar to that of other cultured mammalian cell lines in regard to induction andmore » duration of division delay, and the sensitivity of M and G/sub 2/ cells to radiation-induced cell lethality. However, the radioresistance exhibited over the remainder of the cell cycle results in a relatively flat age response for radiation-induced lethality. This may in part explain the radiation resistance observed for the 9L gliosarcoma in situ.« less