BLT Reagent Supports CRC Immunoresistance Research: Nanoprodrug Targeting Tumor-Associated Intracellular Bacteria Enhances Colorectal Cancer Immunotherapy

New progress has been made in overcoming immunotherapy resistance in colorectal cancer.

Professor Yunjian Yu’s team from Tiangong University, in collaboration with Tianjin Medical University General Hospital and other institutions, published their findings in the international authoritative journal “Journal of Controlled Release” (IF = 11.5, Q1 journal).

This study provides a novel therapeutic strategy for microbiota-driven tumors by developing infection-specific ciprofloxacin prodrug nanoparticles (MTCP-NPs) to overcome immunotherapy resistance in colorectal cancer, particularly in the pMMR/MSS subtype, through targeted elimination of intracellular Fusobacterium nucleatum and remodeling of the immunosuppressive tumor microenvironment.

The study demonstrated that MTCP-NPs nanoprodrugs, through the dual mechanisms of IL4 receptor (IL4R)-mediated targeted delivery and CTSB-responsive drug release, could efficiently target M2-type tumor-associated macrophages with an uptake rate of 80%. The nanoparticles precisely released ciprofloxacin at sites infected with Fusobacterium nucleatum, eliminating 90% of intracellular pathogenic bacteria within 48 hours, which was significantly superior to conventional antibiotics and non-targeted nanocarriers. Meanwhile, MTCP-NPs markedly downregulated IDO secretion, reduced the infiltration of myeloid-derived suppressor cells and the proportion of regulatory T cells, and promoted dendritic cell maturation as well as CD8⁺ T-cell infiltration, successfully converting “cold tumors” into “hot tumors” and effectively remodeling the immunosuppressive tumor microenvironment.

In vivo experiments further showed that the nanoprodrug reduced the survival rate of intratumoral Fusobacterium nucleatum by 95% in a subcutaneous colorectal cancer model and significantly prolonged the median survival time of mice to 21 days. In the orthotopic model, combination therapy with anti-PD-L1 antibody exhibited synergistic enhancement, extending the median survival time to 44 days. In addition, the treatment enriched splenic central memory T cells to maintain long-term antitumor immune responses. MTCP-NPs also demonstrated excellent biocompatibility, with a hemolysis rate below 5% and minimal toxicity toward normal cells. Moreover, the nanoparticles specifically eliminated pathogenic bacterial populations while enriching beneficial bacteria such as Lactobacillus, thereby maintaining gut microbiota balance and avoiding the damage caused by broad-spectrum antibiotics to commensal flora. These findings suggest that MTCP-NPs represent a promising therapeutic strategy with both efficacy and safety for immunotherapy-resistant tumors such as pMMR/MSS colorectal cancer.

Experiments using BLT Reagent

In the study, the research team used D-luciferin potassium salt from Biolight Biotechnology to monitor the progression of CT26-Luc cells in orthotopic CRC models while evaluating the therapeutic efficacy of MTCP-NPs alone and in combination with anti-PD-L1 immune checkpoint inhibitors. Experimental results showed that, compared with PBS treatment, anti-PD-L1 monotherapy exhibited minimal therapeutic effects, whereas MTCP-NPs alone significantly suppressed tumor bioluminescence. Notably, the MTCP-NPs/anti-PD-L1 combination therapy demonstrated synergistic enhancement, and endpoint analysis confirmed that both tumor weight and metastatic nodules were significantly reduced compared with monotherapy groups.

Doi: 10.1016/j.jconrel.2025.114512