BL mice

/III. 1. Raduuion Ondo~ Bd Phys.. Vol. in the U.S.A. All ri&hts trsmvc4. Rimed

IO. pp. 1479-1482 CopYnght

036@3016/84 503.00 + .oO 8 1984 Perpmon Fns Ltd.

??Session III

RADIATION AND CONCURRENT CHEMOTHERAPY FOR THE TREATMENT OF LEWIS LUNG TUMOR AND B16 MELANOMA TUMOR IN C57/BL MICE J. E. PEDERSEN, M.D.‘** AND G. BARRON, B.!k2 ‘Department

of Radiation Oncology, Cross Cancer Institute; and ‘Department University of Alberta, Edmonton, Alberta, Canada

of Radiology,

C57/BLmice bearing eitber Lewis lung tumor or B16 meknome tumor were beated with radiitioa and mt

chemotherapy. Tbe bwtwnt results were determined in rive by tumor regrowth delay assay. Wbea coetinuous I (MU) oc Adriamycia (ADRIA) or Mitomycin-C infusion of either Cycbpbaepbamide (CYCL0) or S-Fl(MITO-C) wmusedincombixmtion withcaatinuoas mdhth at 1 cCy/min, no bcrease ia tumor regrowth delay was observed over tbat of radiatiea alone. When mahipk drug cbemotberapy, FAM (S-FU, ADRIA, MITO-C) was administered ia combinatioa witb radktioo at 80 &y/m& no ioaease ia tumor regrowth deky was observed over tbat of radiatioa alone. In these two marbe tumor medek, when clinkally rekvant coecentratioos of commonly used chemotherapy agents were rombiaed with radiation, no therapeutic advantage was observed. Radiation, chemotherapy, Low dose rate.

INTRODUCI-ION

muscles of 8 to 10 week old C57/BL mice as previously described by Steel and Adams.’ When the leg diameter reached 8 mm (approximately 0.25 gm of tumotb) the animals were prepared for treatment with either continuous tumor irradiation (LDR) and concurrent, continuous drug administration or HDR radiation combined with multiple drug chemotherapy administration. Following the treatment, the leg diameter was measured 3 times a week; when the tumor regrowth reached four times the initial volume, the animals were k&d6 Animals that died before tumor regrowth was complete were not included in the assessment.

The combination of radiation and chemotherapy is theoretically attractive because an improved therapeutic ratio would exist if tumor control is enhanced with little or no increase in host toxicity. Clinically, the combination of chemotherapy and conventional radiation have been tested,’ but the therapeutic advantages have not been obvious. ’Likewise, continuous infusion of chemotherapy and continuous low dose rate (LDR) radiotherapy have been combined in an attempt to improve local tumor control.2 One form of LDR radiotherapy used clinically is brachytherapy, which allows for more localized treatment and continuous radiation.4 Currently the use of interstitial radiation (brachytherapy) is being revived because of improved techniques and newer isotopes.’ Therefore, to assess the value of chemotherapy agents improving the therapeutic effectiveness of radiation, we have combined a number of commonly used chemotherapy agents with either high dose rate (HDR) radiation or LDR continuous radiation. Only clinically relevant drug doses were administered to the tumor bearing mice.’

Radiation technique Forty unanesthetized mice were placed, separately, in a 2.5 cm diameter perspex tube with a slot along the side of the tube to allow the left hind leg to be extended away from the mouse body in order that the mouse body could be shielded with 10 half value thickness (HVT) of lead, yet allowing the tumor-bearing leg to be fully irradiated. The tubes were stacked one on top of the other, 10 high. A cesium source treatment machine* was used for the LDR radiation. Lead attenuation and source-to-tumor distance were used to obtain the desired dose rate of 1 cGy/min. The HDR irradiation (80 cGy/min) was administered with the Cobalt 60 treatment machine.? The

METHODS AND MATERIALS

Tumor model Either Lewis lung tumor or B 16 melanoma tumor was injected as a tumor mush into the left gastrocnemius

11560 University Avenue, Edmonton, Alberta, Canada, T6G 122. Accepted for publication 22 March 1984. ?? Picker TM137. t AECL, Theratron.

Presented at the Conference on Chemical Modifiers of Cancer Treatment. Banff,Alberta, Canada. November 27-December 1, 1983. Supported by the Albena Cancer Board Endowment Research Fund. Reprint requests to: J. E. Pedemn, Cross Cancer Institute, 1479

RadiationOncology 0 Biology 0 Physics

August 1984, Volume IO, Number 8

CONTROt(13)

.-

X-

30 Gy(36) - 5FU+30Gy(16) AADRIA + 30 Gy(ll) ?? - CYCLO +30 Gy(13) @I- MIT0 -C +30 Gy(8)

??

lb

;

DAYS AFTER

1cGy min

;o

1;

TREATMENT

Fig. 1. Combined continuous chemotherapy and LDR radiation for Lewis lung tumor in C57/BL mice. The results are means of all tumors, and the number in brackets indicates the number of tumors per group.

radiation dosimetry assessments were made by members of the Physics Department at the Cross Cancer Institute.

Drug administration techniques Four commonly used clinical drugs CYCLO,* 5-FIJ,t MITO-C$ and ADRIA§ were tested with continuous irradiation. CYCLO and MIT04 were dissolved in sterile

water; ADRIA was dissolved in 0.9 sterile saline, and 5-FU (Roche) was used in the standard parenteral formulation. All drugs were diluted to the appropriate concentration with physiological saline immediately before use, and then were drawn into 3 ml syringes. A Harvard multiple continuous infusion pump was used to inject 20 mice simultaneously at a rate of 0.11 ml per hour for

?? ? CONTROL ? ? ? CYCLO ?

O;-

(10) 40 8 SOmg/Kg 5FU 25mg/Kg (18) ADRIA 4 mg/Kg (11) MITO-C lmg/Kg (10)

30 Gy (23) $1SFU + 30 Gy A-

ADRIA

(13) + 30 Gy (11)

m- CYCLO +30 Gy(5) o- MIT0-C+30 Gy(5) I

1

I

L

I

I

I

I

15 5 10 DAYS AFTER TREATMENT

(14)

1cGy

min

1

20

Fig. 2. Combined continuous chemotherapy and LDR radiation for B16 melanoma tumor in C57/BL mice. The results are means of all tumors and the number in brackets indicates the number of tumors per group.

* Cyclophosphamide, Bristol, Vancouver, B.C., Canada. t Mitomycin C, Bristol, Vancouver, B.C., Canada.

$5 Fiuorouracil, Hoffman LaRoche, Montreal, P.Q., Canada. 9 Adriamycin, Mississauga, Ontario, Canada.

Radiation and chemotherapy 0 J. E. PEDERSEN AND

G.

BARRON

1481

?? - CONTROL O-

FAM

(5) - 2mg/Kg - 1 mg/Kg - 15mg/Kg

20 Gy FAM 24 HRS ?? - FAM SIMULT. A- FAM 24 HRS oO-

10 5 DAYS AFTER TREATMENT

ADRIA (8) MIT0 -C 5 FU BEFORE WITH AFTER

20 Gy(8) 20 Gy (7) 20 Gy (8)

I

80cGy min

15

Fig. 3. Combined combination chemotherapy and HDR radiation for Lewis lung tumor in C57/BL mice. Results are the means of all tumors. The number in brackets indicates the number of tumors per group.

mice. These were clinically relevant and were compatible with survival in all but a few mice.

50 hours. The chemotherapy was infused into the peritoneal cavity of the mouse via intramedic polyethylene tubing (PE-50) which was introduced intraperitoneally while the mice were momentarily anesthetized with methoxyflurane inhalation anesthetic. Figures 1 and 2 indicate the doses of drugs injected into the mice. The control mice received equal volumes of normal saline. Mouse temperatures were maintained at 35 to 37’-‘C (rectal temperature) by heating the ambient air around the perspex mouse holders. Kernel corn provided some nutrition during the 50 hours of treatment. Those mice receiving the multiple drug regime (FAM) in combination with HDR irradiation received the drugs as an i.p. bolus. Control mice received normal saline. Figures 3 and 4 indicate the drug doses injected into the

RESULTS The effects on Lewis lung tumor and B 16 melanoma tumor when continuous chemotherapy was administered concurrently with LDR radiation are shown in Figures 1 and 2. CYCLO, 5-FU, ADRIA, or MITO-C, when administered without radiation, produced no observable effect on tumor growth. Thirty Gy at 1 cGy/min. produced significant tumor regrowth delay; however, when continuous infusion of either CYCLO, 5-FU, ADRIA or MITG-C was combined with the radiation, no extra re-

?? - CONTROL a-

0aaA1

1

1

I

I

I

I

5 DAYS AFTER

10

15

(5) FAM - 2 mg/Kg - lmg/Kg - lSmg/Kg

ADRIA (5) MITO-C 5FU

20 Gy (5) FAM 24 HRS BEFORE 20 Gy (10) FAM SIMULT. WITH 20 Gy (IO) FAM 24 HRS AFTER 20 Gy (9)

BOcGy min

i

20

TREATMENT

Fig. 4. Combined combination chemotherapy and HDR radiation for B16 melanoma in C57/BL mice. Results are the means of the tumors and the number in brackets indicates the number of tumors per group.

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0 Biology 0 Physics

growth delay was observed. This was observed for both the Lewis lung tumor and the B16 melanoma tumor. Figures 3 and 4 show the effects on Lewis lung tumor and B 16 melanoma tumor when combination chemotherapy (FAM) is administered to mice at clinically relevant concentrations. Twenty Gy at 80 cGy/min. produced significant tumor regrowth delay in both Lewis lung tumor and B 16 melanoma tumor, however, when FAM chemotherapy was administered either simultaneously with radiation, 24 hours before, or 24 hours after radiation, no increase in tumor regrowth delay was observed over radiation alone.

DISCUSSION When Lewis lung tumor and B16 melanoma tumor were treated with radiation therapy doses in the range of 20 to 30 Gy, significant tumor regrowth delay could readily be demonstrated. However, when radiation was

August 1984, Volume 10, Number 8

combined with a clinically relevant dose of chemotherapy no increase in tumor cell killing was observed as indicated by the tumor regrowth delay assay. One possible explanation for this observation is that chemotherapy alone had no appreciable effect on tumor regrowth in these experiments. The chemotherapy doses chosen for these experiments represent clinical situations in which chemotherapy often has no observable tumor effect. A number of authors have reported an increase in tumor cell killing in some animal tumor models when chemotherapy and radiation treatments are combined.’ For example, Steel et al.’ reported that 300 mg/kg CYCLO reduced the radiation dose required for cure of Lewis lung tumor. However, 300 mg/kg of Cyclophosphamide is a concentration considerably above a clinically safe dose3 and therefore it is not possible to make clinical inferences from not entirely clinically relevant experiments. The findings from the experiments discussed in this paper are in agreement with many clinical observations.

REFERENCES Reny, R.J.: Interaction of drugs and radiation-promise or pitfall. Clin. Rudiol. 33: 12 l-129, 1982. Fu, K.K., Rayner, P.A., Lam, K.: Modification of the effects of continuous low dose rate irradiation by chemotherapy (Abstract). Proceedings of the Seventh International Congress of Radiation Research. Amsterdam, July 3-8, 1983. Freireich, E.J., Gehan, E.A., Rall, D.F., Schmidt, L.H., Skipper, H.E.: Quantitative comparison of toxicity of anticancer agents in mouse rat, hamster, dog, monkey and man. Cancer Chemo. Rep. SO(4): 2 19-244, 1966. Hilaris, B.S., Nori, S.: Brachytherapy Oncology, 1982. Hal-

aris, B.S., Novi, S. @is.). New York, N.Y., Memorial SloanKettering Cancer Center, 1982. 5. Pierquin, B., Chassagne, D.J., Chahbaxian, C.M., Wilson. J.F.: Brachyrherupy. Pierquin, B., Chassagne, D.J., Chagbaxian, C.M., Wilson, J.F. @is.). St. Louis, Missouri, Warren H. Green, Inc., 1978. 6. Steel, G.G., Adams, K.: Stem cell survival and tumor control in the Lewis lung carcinoma. Cancer Res. 35: 1530-1535, 1975. 7. Steel, G.G., Hill, R.P., Peckham, M.J.: Combined radiotherapy-chemotherapy of Lewis lung carcinoma. Inr. J. Radiat. Oncol. Biol. Phys. 4: 49-52, 1978.