Robert Sedlacek

Pentobarbital anesthesia and the response of tumor and normal tissue in the C3Hf/sed mouse to radiation

Studies of the effect of pentobarbital anesthesia on the radiation response have been performed using early generation isotransplants of three spontaneous tumors of the C3H mouse: a mammary carcinoma (MCaIV), a fibrosarcoma (FSaII), and a squamous cell carcinoma (SCCVII). The enhancement ratio of pentobarbital [ER(PB)] for TCD50 as the end point was greater than or equal to 1 for all conditions tested. The ER(PB) for O2 3 ATA conditions and two equal doses was 1.46, 1.72, and 2.21 for MCaIV, FSaII, and SCCVII, respectively. The ER(PB) using MCaIV was the same for O2 and carbogen at 1 or 3 ATA. Also, tumor size of MCaIV did not significantly affect the ER(PB) for O2 3 ATA conditions. Further, with the two-dose protocol the anesthesia and the hyperbaric oxygen needed to be used at the second dose; condition at the first dose was not critical. For fractionated irradiation of MCaIV (10 and 15 equal doses) the ER(PB) was smaller than for two-dose treatment; also the effect was less for intratumor temperature of 35 degrees C than 26-27 degrees C. There was no effect of the anesthesia on the acute response of normal skin of the leg. Lung damage by hyperbaric oxygen was not an important factor in these results. Additionally, ERs were computed for O2 at 3 ATA. This ER(O2 3 ATA) was larger for anesthesized than conscious mice. The ER(O2 3 ATA) for MCaIV was high (greater than 1.5) even for radiation given in 10 or 15 equal doses.

Radiation response of xenografts of a human squamous cell carcinoma and a glioblastoma multiforme : a progress report

Human cell lines derived from squamous cell carcinomas of the pharynx (FaDu and HSCC6) and glioblastoma multiforme (U87, A2, A7, MMC-1, MMC-2) have been studied in vitro as monolayers in exponential (all 7 cell lines) or plateau phase (FaDu and U87), and as 1 mm diameter spheroids in vitro (FaDu and U87) and as 6 mm diameter xenografts growing in the legs of athymic NCr(nu/nu) nude mice (FaDu, HSCC6, U87, A7 cells). For SF2s and D values, there was broad overlap of values between SCC and glioma cell lines. In contrast, the D0 values were higher for U87, A2, A7, and MCC-1 than the two SCC cell lines, while the extrapolation numbers were greater for the two SCC lines than any of the glial tumor lines (these differences were not regularly significant). Complete dose response assays for local control of FaDu, HSCC6, U87, and A7 xenografts have been performed under conditions of normal blood flow and clamp hypoxia for tumors growing in mice which had received 6 Gy WBI at 24 hr before transplantation. Under the latter circumstances, irradiations have been performed on FaDu and U87 as single doses or as 2, 4, or 8 equal doses; for the fractionated irradiation, treatments were given on a BID basis with 4 hr between the treatments on any 1 day. For irradiation of 1 mm diameter spheroids, radiation was administered as single doses under conditions of equilibration with AIR. The TCD50 for the FaDu was significantly higher and the dose response curve steeper for tumors growing in immune suppressed (6 Gy WBI 24 hr prior to transplantation) than in control nude mice. Tumors, exponential or plateau phase cells, and spheroids derived from U87 were significantly and substantially more resistant under all conditions and fractionation schedules than for FaDu. Thus, the in vitro results do not indicate a clearly greater resistance by the glioma cell lines, while the more limited TCD50 data (single dose and 8 fractions irradiation) show more resistance in vivo by the glial tumors. We noted that the TCD50 values for U87 and A7 glial tumors overlap those for spontaneous tumors of the C3H mouse but are higher than the human squamous cell carcinoma xenografts in the nude mice. Substantial additional data from xenografts are needed to determine if the higher TCD50 values for GBMs, especially for fractionated irradiation, is a regular finding and is of sufficient magnitude to be pursued by studies to explain the observed differences.

Experimental studies on the incidence of metastases after failure of radiation treatment and the effect of salvage surgery

FSaII, a spontaneous fibrosarcoma, and SCCVII, a spontaneous squamous carcinoma, were studied as early generation isotransplants in the right leg of C3Hf/Sed mice. Animals successfully treated, in respect to local control by surgery alone for 6 mm diameter tumors, had an incidence of distant metastases of 2.6% for the FSaII, and 8% for the SCCVII. For 12 mm tumors the incidence of metastases was 14.3% and 41%, respectively. Animals successfully treated with radiation alone for 6 mm tumors had an incidence of distant metastases of 3.1% for the FSaII, and 6.9% for the SCCVII. Animals that developed local recurrence after radiation therapy were treated with salvage surgery when the recurrent tumor was either 6 mm or 12 mm. For those successfully treated the incidence of metastases was 12.5% for the FSaII, and 43% for the SCCVII when salvage surgery was performed on 6 mm tumors. When surgery was delayed and performed on 12 mm tumors, the incidence was 46.6% and 70.3%, respectively. The results indicate that after failure of radiation treatment there is a high incidence of metastases from the recurrent tumors. The incidence, however, can be reduced considerably by carrying out early salvage surgery.

Studies on fractionated hyperthermia in experimental animal systems I. The foot reaction after equal doses: Heat resistance and repopulation

Abstract The response of the mouse foot to fractionated hyperthermia was studied in vivo . Hyperthermia was given by immersing the mouse foot into a constant temperature water bath kept at 43.5 ± 0.1°C. Foot reaction was scored after treatment according to a numerical score system; at 43.5°C the time to induce loss of one toe or a greater reaction in half the treated animals (RD 50 ) was assayed. The equal dose fractions were used throughout the present experiments. The RD 50 for various time intervals (Ti) between two doses demonstrated rapid development of heat resistance which reached a maximum in 48 hours after first treatment. The resistance subsided thereafter and the complete decay of resistance required almost 2 weeks. If 2 doses were given with a Ti of more than 17 days, further increase of the RD 50 was observed, which might be attributed to repopulation of surviving cells. Isoeffect curves for treatment schedules with various Ti were obtained. These studies indicated that heat resistance develops repeatedly after each treatment and is the most important factor in fractionated hyperthermia. The kinetics of the resistance appeared to depend on the Ti and number of fractions as well as fraction size. In general the resistance developed fully in 48 hours after first hyperthermia, whereas it appeared to develop in 24 hours after the second treatment with a Ti of 2 days or after the third treatment with a Ti of 1 day. The resistance was less extensive for a large number of fractions with Ti of 2 days.

Time distributions of recurrences of immunogenic and nonimmunogenic tumors following local irradiation.

Three hundred and fourteen mice received single-dose irradiation of the right leg and thigh as treatment of an 8-mm mammary carcinoma isotransplant, and were then observed until death, usually by 1000 days. The time distributions of death due to local recurrence, radiation-induced sarcoma, distant metastasis in the absence of local regrowth, second primary, intercurrent disease, and unknown causes have been evaluated. The times for the transplant tumor inoculum to grow to an 8-mm tumor and the times of death due to local regrowth, distant metastasis, or induced tumor were all approximately log-normally distributed. Of the 128 recurrences, the latest-appearing 3 were at 300, 323, and 436 days; no recurrences were noted during the time period from 436 to 1000 days. These findings have been interpreted to mean that in some cases absolute cure of mice of the tumor in the leg was achieved by radiation alone at the dose levels employed. Radiation-induced sarcomas began to appear after 300 days. The time of appe...

Evaluation of tumor energy metabolism and microvascular blood flow after glucose or mannitol administration using 31p nuclear magnetic resonance spectroscopy and laser doppler flowmetry

The effects of intraperitoneally administered glucose or mannitol (5 mg/g body weight, 25% solutions) on tumor energy metabolism and tumor red blood cell flux were studied using 31P-nuclear magnetic resonance spectroscopy and laser Doppler flowmetry. Isotransplants of a spontaneous murine fibrosarcoma growing in the hind foot dorsum were used. 31P-nuclear magnetic resonance and laser Doppler flowmetry studies in glucose treated animals were performed on small (congruent to 100 mm3) and large (congruent to 300 mm3) tumors. In mannitol treated animals, tumors with an average volume of congruent to 200 mm3 were used. Using this tumor model, intraperitoneally administration of the hypertonic sugar solutions caused similar declines in tumor microcirculation (mannitol, 60 +/- 8% flow reduction; glucose, 72 +/- 4% flow reduction; t = 60 min). These changes were not glucose-specific and can primarily be explained by a water shift into the abdominal cavity and an associated hypovolemic hemoconcentration. A stable (small tumors) or transiently increased (large tumors) tumor energy metabolism which occurred after glucose administration was probably caused by a transiently increased glucose availability. The decline in energy metabolism after mannitol, a non-metabolized sugar alcohol, and the earlier decline in tumor pH seen in the glucose treated animals, supports this conclusion. The differences in the high energy phosphate response to glucose seen in small compared with large tumors, suggests that the baseline metabolic state of larger tumors includes a glucose deficiency in addition to tumor hypoxia.

Radiation-Induced Osteogenic Sarcoma of C3H Mouse: Effects of Corynebacterium parvum and WBI on its Natural History and Response to Irradiation*

Abstract An osteogenic sarcoma, which appeared at 316 days following single dose 5000 rad to the leg of C 3 H mouse, has been studied as early generation F2 and F3 transplants in syngenic hosts with respect to local growth, pattern of spread, and response to local irradiation in normal, C. parvum treated, and whole body irradiated hosts. Mean survival of untreated mice was 126 days after transplantation. Grossly evident metastatic tumor appeared in 85% of these mice; in 47 of 48 animals metastases were seen in the lung. Corynebacterium parvum given intravenously as a single dose of 350 μg at 96 hr after tumor transplant retarded tumor growth: regression was observed in 10 of 19 tumors, 3 of 19 mice were cured of their osteosarcomas and mean survival was prolonged from 126 to 173 days , in one study where i.v. C. parvum was given when tumor was 5 mm, 1 of 13 mice was cured by C. parvum alone. Although 5000 rad resulted in 100% of tumor destruction in normal mice; 49% died of metastatic tumor to the lung. In C. parvum treatment mice only 16% died of metastatic tumor. The radiation doses which achieved control of half of the irradiated 8 mm diameter tumors were 4350 and 3600 rad for normal and C. parvum treated hosts, respectively. While body irradiation 600 rad given 24 hr prior to tumor transplant had an opposite effect to Corynebacterium parvum .

EXAMINATION FOR A CORRELATION BETWEEN PROBABILITIES OF DEVELOPMENT OF DISTANT METASTASIS AND OF LOCAL RECURRENCE.

Correlation between probability of developing local recurrence and distant metastasis following local irradiation was studied using 12 mm diameter transplants of a mouse mammary carcinoma. Results showed that mice dying of metastatic tumor to lungs with gross local control had subclinical foci of viable tumor cells at the primary site with a frequency expected if local recurrence was not affected by metastatic tumor in lung. Contrariwise 12 of 13 mice dying of local recurrence had metastatic tumor at death. Amputation of the thigh when recurrence was first detected sharply reduced the frequency of metastasis to the lung.

The response of two human tumor xenografts to fractionated irradiation. the derivation of α β ratios from growth delay, tumor control, and in vitro cell survival assays

A series of growth delay experiments was performed to derive alpha/beta ratios for two human neoplasms growing as xenografts in the hind limbs of NCr/Sed nude mice. The tumors were irradiated at 6 mm mean diameter under clamp-hypoxic conditions in one, two, four, or eight fractions, 2 fractions per day with a minimum intertreatment interval of 4 hr and a maximum overall treatment time of 3 days. The alpha/beta ratios derived for the high grade glioma U87 and the pharyngeal squamous carcinoma FaDu were 38 and 20 Gy, respectively. Comparably high values were derived from the same two tumors in a reanalysis of fractionated TCD50 data. The alpha/beta ratios were similarly high whether the TCD50 data were analyzed using the Full Effect plot or the Direct Method. For comparison, cell survival assays were performed on U87 and FaDu irradiated in vitro under plateau-phase, aerobic conditions. The alpha/beta ratios obtained were 9.2 and 15.0 Gy, respectively. Such high alpha/beta values suggest a therapeutic gain could result from the use of small doses per fraction.

Comparison of hyperbaric oxygen and misonidazole in fractionated irradiation of murine tumors.

The enhancement ratios for hyperbaric oxygen (O/sub 2/3ATA) and for misonidazole (0.3 mg/g body wt) for fractionated irradiation (5 or 10 equal doses) of two spontaneous tumors and of normal skin of the C3H mouse have been determined. Acute skin reactions were scored for mice irradiated 18 days after plucking hair from the leg. Enhancement ratios for the TCD/sub 50/ values were virtually the same for O/sub 2/3ATA and misonidazole, 1.45 to 1.55. For acute skin reaction the enhancement ratio was higher for O/sub 2/3ATA, i.e., 1.94 vs 1.54 for misonidazole.