Mycobacterium Bovis (Strain BCG) Effects on the Growth and Metastasis of a Transplantable Hamster Lung Adenocarcinoma

The effect of Bacillus-Calmette-Guerin (BCG) on the growth of an adenocarcinoma of the lung induced in a Syrian hamster was evaluated. It inhibited intradermal growth of the tumor when the tumor cells were mixed with high doses of BCG prior to intradermal injection. The development of lung metastasis after excision of the primary intradermal tumor was inhibited by BCG at doses 100 times lower than those used to inhibit the growth of intradermal tumors. The results suggest that effective immunotherapy may depend on the number of viable BCG cells, the route of administration, and the anatomic location of the tumor. OHIO J. SCI. 79(3): 126, 1979 Syrian golden hamsters when treated intratracheally with benzo(a)pyrene and iron oxide develop respiratory tract malignancies which for the most part resemble the majority of human lung cancers both in location and morphologic characteristics (Saffiotti et al 1968). We have been assessing the effects of BCG preparations on the development of primary lung tumors (Zwilling et al 1977) and have sought in this study to assess the effect of BCG on the growth and metastasis of a transplantable hamster lung adenocarcinoma induced by the intratracheal application of the carcinogenic agent benzo(a)pyrene. MATERIALS AND METHODS Animals. Inbred male Syrian golden hamsters, strain LSH/LAK, were obtained from Charles River Lakeview (Lakeview, N.J.). Animals were housed 5 per cage under standard laboratory conditions and were used when 7-12 weeks of age. Tumor cells. The tumor cell line used in this study was a bronchogenic adenocarcinoma, designated LG 1002. It was induced in an outbred Syrian golden hamster by the intratracheal instillation of benzo(a)pyrene adsorbed to ferric oxide particles and suspended in saline (Saffiotti et al 1968). The tumor was maintained by serial passage in the hamster Manuscript received 23 T 1978 and in revised form 16 November 1978 (#78-37). Supported by USPHS grant CA-16342 from the National Cancer Institute. cheek pouch. Tumor cell inocula were prepared from the 2nd to 10th transplant generations by digesting small tumor pieces with trypsin and suspending the cells at the desired concentration in Hank's balanced salt solution (HBSS). Mycobacterial antigen. The Phipps strain of Mycobacterium bovis, strain Bacillus-CalmetteGuerin (BCG), TMC#1029, Lot #NC734A was obtained from the Trudeau Institute (Saranac Lake, N.Y.). Vials containing 1.2 x 10 colony forming units (cfu) were stored frozen at—70 °C. Dilutions of the BCG for injection were made in Hank's BSS. Concentration of the BCG suspension was accomplished by centrifugation at 200Xg for 20 minutes. Immunizations. Tumor cell inoculations were given intradermally in the intrascapular region of the back or into the hind footpad of animals anesthetized with Brevitol Sodium. Locally administered BCG was admixed with the tumor cell inoculum prior to injection. Intratracheal injections of BCG were performed by the method described by Saffiotti et al (1968) for carcinogen instillation. Monitoring of tumor growth. Growth of the tumor cell inocula given intradermally in the back was monitored by measuring the greatest and the least diameters of the tumor nodule with calipers. Tumor size was expressed as the product of the two diameters. Tumor growth in animals receiving footpad injections was monitored by measuring the footpad thickness with calipers. Footpad tumor growth was expressed as a tumor index which was calculated as the ratio of the thickness of the injected footpad to the thickness of the uninjected footpad. Intradermal tumors were excised along with sufficient surrounding skin to insure the complete removal of the primary tumor mass. The