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Experimental Therapeutics, Molecular Targets, and Chemical Biology

Targeted Therapy with a Salmonella Typhimurium Leucine-Arginine Auxotroph Cures Orthotopic Human Breast Tumors in Nude Mice

Ming Zhao, Meng Yang, Huaiyu Ma, Xiaoming Li, Xiuying Tan, Shukuan Li, Zhijian Yang and Robert M. Hoffman
Ming Zhao
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Meng Yang
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Huaiyu Ma
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Xiaoming Li
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Xiuying Tan
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Shukuan Li
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Zhijian Yang
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Robert M. Hoffman
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DOI: 10.1158/0008-5472.CAN-06-0716 Published August 2006
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    Figure 1.

    Tumor targeting of S. typhimurium A1 and A1-R in vitro. A, RFP-labeled HT-29 human colon cancer cells were grown in 24-well tissue culture plates. S. typhimurium-GFP, grown to late-log phase, were diluted in cell culture medium (1 × 106), added to the tumor cells, and placed in an incubator at 37°C. After 60 minutes, the cells were rinsed. Adherent bacteria were released and then quantitated by plating for CFU on LB agar medium. The number of A1-R bacteria attached to tumor cells was ∼6 times higher than the parental A1 bacteria. B and C, to measure invasion of bacteria, the cancer cells were first rinsed five times with PBS, and then the cells were cultured in medium containing gentamicin sulfate (20 μg/mL) to kill external but not internal bacteria. After incubation with gentamicin for 12 hours, the cancer cells were washed once with PBS, and the viable GFP-expressing intracellular bacteria were observed by fluorescence microscopy. A1-R (C) bacteria invaded and grew in tumor cells to a greater extent than the parental A1 (B). Bar, 58 μm.

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    Figure 2.

    Growth of S. typhimurium A1-R in MARY-X human breast cancer tissue after tail vein injection. NCR nude mice, 6 to 8 weeks, were s.c. implanted in the midright side with human breast cancer tissue. Bacteria were injected directly into the tail vein (107 CFU/100 μL PBS). The tumors were excised, and samples were observed under fluorescence microscopy. For histology, the tumor tissue was fixed with 10% buffered formalin and processed for paraffin section and HE staining by standard methods. A and B, bacteria growing in tumor tissue on day 2 after infection. Bar, 150 μm in (A and B). C, surface image of bacterial distribution in intact tumor tissue on day 5 after infection. Bar, 180 μm. D, bacterial distribution in transverse section of tumor showing inner surface on day 5 after infection. Bar, 180 μm. E, bacteria growth in the viable MARY-X tumor tissue. Bar, 25 μm. F, bacteria destroyed tumor tissue (day 4 after infection). Bar, 50 μm. G, bacteria growth within the vascular regions of tumors (day 4 after infection). Blue dots, arrows, S. typhimurium A1-R. Bar, 100 μm.

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    Figure 3.

    Efficacy of A1-R on MARY-X human breast cancer growing s.c. in nude mice. A and B, efficacy of A1-R on tumor growth. Six-week-old female nu/nu mice were s.c. implanted on the flank of the nude mice with 2 mm3 MARY-X human breast cancer tissue. The bacteria were grown overnight on LB medium and then diluted 1:10 in LB medium. Bacteria were harvested in late-log phase, washed with PBS, and then diluted in PBS (107 CFU/100 μL PBS). C, efficacy of A1-R on survival of nude mice with MARY-X tumor. Tumor growth was monitored by whole-body imaging. Mice were sacrificed when the tumor mass reached 2 cm3. Survival time was compared with the untreated mice. The survival of treated animals was prolonged, and the 40% cured mice survived as long as non–tumor-bearing mice. n = 10 mice for each group.

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    Figure 4.

    Efficacy of S. typhimurium A1-R on MARY-X human breast cancer growing orthotopically in nude mice. A and B, efficacy of A1-R on orthotopic MARY-X breast cancer. Nude mice, 6 to 8 weeks, were orthotopically implanted in the right second mammary gland fat pad with 2 mm3 MARY-X human breast cancer tissue. A1-R bacteria were injected directly into the tail vein (107 CFU/100 μL PBS). Tumor size was measured at indicated time points after infection. n = 10 animals.

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Cancer Research: 66 (15)
August 2006
Volume 66, Issue 15
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Targeted Therapy with a Salmonella Typhimurium Leucine-Arginine Auxotroph Cures Orthotopic Human Breast Tumors in Nude Mice
Ming Zhao, Meng Yang, Huaiyu Ma, Xiaoming Li, Xiuying Tan, Shukuan Li, Zhijian Yang and Robert M. Hoffman
Cancer Res August 1 2006 (66) (15) 7647-7652; DOI: 10.1158/0008-5472.CAN-06-0716

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Targeted Therapy with a Salmonella Typhimurium Leucine-Arginine Auxotroph Cures Orthotopic Human Breast Tumors in Nude Mice
Ming Zhao, Meng Yang, Huaiyu Ma, Xiaoming Li, Xiuying Tan, Shukuan Li, Zhijian Yang and Robert M. Hoffman
Cancer Res August 1 2006 (66) (15) 7647-7652; DOI: 10.1158/0008-5472.CAN-06-0716
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