Kurt Yun Mou’s research group in the Institute of Biomedical Sciences at Academia Sinica developed a novel cancer therapy and published their results in “Molecular Therapy” in Apri this year. Taking the advantage of TNF-α as a pro-tumor signal, Mou’s group successfully designed an immunotoxin, which can piggyback on TNF-α and co-internalize into cancer cells for cell killing. The study further developed E. coli as a “live drug”, which can colonize and replicate in tumors, to constantly deliver the immunotoxin and stimulate the TNF-α expression. As a result, the treatment greatly suppressed the tumor growth in a syngeneic murine melanoma model.
TNF-α is a pleiotropic cytokine that is tightly regulated at a very low level in body but greatly increased at the inflammation site. TNF-α transduces its signal through the ligation to the ubiquitously expressed receptor TNFR1. When TNF-α binds TNFR1, the complex is internalized into cells through endocytosis. TNF-α involves in many autoimmune diseases, such as rheumatoid arthritis and inflammatory bowel disease. Many TNF-α blockades are approved by FDA for treating autoimmune diseases. In oncology, TNF-α was also found to play an important pro-tumor role. For example, TNF-α is involved in tumor initiation as TNF-α-/- mice were resistant to skin tumors induced by chemical carcinogens. Cancer cells overexpressing TNF-α resulted in increased vascularity and enlarged tumor sizes in mouse melanoma, lung cancer, and mammary cancer models. However, clinical trials showed that the administration of TNF-α blockades could not effectively suppress the tumor growth.
Bacterial therapy has emerged as a revitalizing modality for cancer treatment since the pioneering works by William Coley in the late 19th century. Modern studies have demonstrated several advantages of bacterial therapy against cancer, including (1) selective colonization in tumors27, (2) pathogen-associated molecular patterns to stimulate the immune system28, and (3) great engineerability. For example, E. coli overexpressing tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) caused a dramatic reduction in tumor size in mouse models. Immune checkpoint blockades can be locally delivered in tumors by E. coli and greatly inhibited the tumor growth. A phase 1 clinical trial for treating advanced solid tumors and lymphoma was recently launched using a E. coli probiotic Nissle 1917 engineered for STING agonist delivery. The treatment was well-tolerated and achieved target engagement and durable disease stabilization.
Here, we propose an unconventional strategy for TNF-α targeting. Instead of using TNF-α blockades widely utilized in clinics, we designed non-neutralizing TNF-α antibodies, which can piggyback on TNF-α and co-internalize into cancer cells for intracellular drug delivery. We employed phage display and yeast surface display to select high-affinity, high-specificity TNF-α antibodies. We further used biolayer interferometry to characterize the non-neutralizing antibodies. In the cell experiments, we showed that the non-neutralizing antibodies can co-internalize into cells with TNF-α. We further formulated the antibody as antibody-drug conjugate or immunotoxin and revealed that they exerted TNF-α-dependent cytotoxicity to cancer cells. We deliberately implemented this immunotoxin into a E. coli vehicle for a synergistic effect, i.e. the bacteria stimulated the TNF-α expression that facilitates the immunotoxin internalization. The immunotoxin-secreting bacteria effectively suppressed the tumor growth in a syngeneic mouse model of melanoma. The tumor-infiltrating leukocyte analysis revealed several beneficial immunomodulations by the treatment, including increased M1 macrophages, decreased M2 macrophages, and increased activated CD4+ and CD8+ lymphocytes.
The Trojan Horse strategy in this study is fundamentally different from the traditional blockade strategy. We anticipate that this new concept can broaden the angles in the antibody drug development and provide alternative mechanisms in the management of various human diseases. In addition, both US and European FDA have drafted the guidance for live biotherapeutic product (LBP). We expect that engineered bacteria can be the next-generation “live drug” for cancer therapy after CAR-T.
Che-Wei Hu, the first author of this study, was the research assistant in the Institute of Biomedical Sciences. The co-authors in order are You-Chiun Chang, Cheng-Hao Liu, Yao-An Yu, and Kurt Yun Mou (corresponding author).