Engineering tumor-colonizing E. coli Nissle 1917 for detection and treatment of colorectal neoplasia.

Bioengineered probiotics enable new opportunities to improve colorectal cancer (CRC) screening, prevention and treatment. Here, first, we demonstrate selective colonization of colorectal adenomas after oral delivery of probiotic E. coli Nissle 1917 (EcN) to a genetically-engineered murine model of CRC predisposition and orthotopic models of CRC. We next undertake an interventional, double-blind, dual-centre, prospective clinical trial, in which CRC patients take either placebo or EcN for two weeks prior to resection of neoplastic and adjacent normal colorectal tissue (ACTRN12619000210178). We detect enrichment of EcN in tumor samples over normal tissue from probiotic-treated patients (primary outcome of the trial). Next, we develop early CRC intervention strategies. To detect lesions, we engineer EcN to produce a small molecule, salicylate. Oral delivery of this strain results in increased levels of salicylate in the urine of adenoma-bearing mice, in comparison to healthy controls. To assess therapeutic potential, we engineer EcN to locally release a cytokine, GM-CSF, and blocking nanobodies against PD-L1 and CTLA-4 at the neoplastic site, and demonstrate that oral delivery of this strain reduces adenoma burden by ~50%. Together, these results support the use of EcN as an orally-deliverable platform to detect disease and treat CRC through the production of screening and therapeutic molecules.

Colorectal cancer detection and treatment with engineered probiotics.

Bioengineered probiotics enable new opportunities to improve colorectal cancer (CRC) screening, prevention and treatment strategies. Here, we demonstrate the phenomenon of selective, long-term colonization of colorectal adenomas after oral delivery of probiotic E. coli Nissle 1917 (EcN) to a genetically-engineered murine model of CRC predisposition. We show that, after oral administration, adenomas can be monitored over time by recovering EcN from stool. We also demonstrate specific colonization of EcN to solitary neoplastic lesions in an orthotopic murine model of CRC. We then exploit this neoplasia-homing property of EcN to develop early CRC intervention strategies. To detect lesions, we engineer EcN to produce a small molecule, salicylate, and demonstrate that oral delivery of this strain results in significantly increased levels of salicylate in the urine of adenoma-bearing mice, in comparison to healthy controls. We also assess EcN engineered to locally release immunotherapeutics at the neoplastic site. Oral delivery to mice bearing adenomas, reduced adenoma burden by ∻50%, with notable differences in the spatial distribution of T cell populations within diseased and healthy intestinal tissue, suggesting local induction of robust anti-tumor immunity. Together, these results support the use of EcN as an orally-delivered platform to detect disease and treat CRC through its production of screening and therapeutic molecules.

Understanding and predicting distorted T- versus Y-geometries for neutral chromous complexes supported by a sterically encumbering beta-diketiminate ligand.

A series of three-coordinate Cr(II) complexes sharing the common molecular fragment "(nacnac)Cr" (nacnac(-) = [ArNC((t)Bu)](2)CH, Ar = 2,6-(i)Pr(2)C(6)H(3)) were prepared via salt metathesis with the dimer [(nacnac)Cr(mu-Cl)](2). Single-crystal X-ray diffraction studies revealed that the complexes (nacnac)Cr(L) (L = CH(2)(t)Bu, CH(3), CH(2)CH(3), SiH{2,4,6-Me(3)C(6)H(2)}(2), O{2,6-(i)Pr(2)C(6)H(3)}, N{CH(3)}(2)) represent a rare class of mononuclear, neutral chromium complexes with a three-coordinate high-spin chromous metal center. Depending on the nature of the third ligand, L(-), these complexes can adopt either distorted T-shaped or Y-shaped coordination geometries. Density functional theory calculations and molecular orbital analyses in combination with a detailed molecular fragment energy decomposition were used to establish an intuitive concept of the key electronic structure patterns that determine the coordination geometry of preference. The frontier orbitals of the (nacnac)Cr(II) fragment direct pi-donating ligands to adopt Y-shaped geometry, whereas ligands that are primarily sigma-donors prefer T-shaped coordination. The relationship between electronics at the metal center and coordination geometry was extended to include the putative neutral three-coordinate high-spin complexes of Sc(II) and Mn(II), which are predicted to both adopt Y-shaped geometry.