Targeting MMR-deficient colorectal cancer with a potent small molecule UNI110

Abstract

Mismatch repair (MMR) deficiency is a hallmark of microsatellite instability (MSI) in hereditary non-polyposis colorectal cancer, Lynch syndrome, contributing to resistance against conventional chemotherapy and posing a significant therapeutic challenge. In this study, we introduce UNI110, a novel small molecule derived from Baicalein, engineered for enhanced selectivity against MMR-deficient cancer cells. UNI110 exhibits a remarkable sevenfold increase in potency over Baicalein, demonstrating significantly lower IC50 values and heightened cytotoxic effects in MMR-deficient cell lines. Mechanistically, UNI110 selectively induces DNA damage in MMR-deficient cancer cells, ultimately resulting in cell death. Furthermore, UNI110 disrupts homologous recombination (HR) repair by inhibiting the MSH2-MSH3 complex, specifically blocking the interaction between MSH2 and EXO1, thereby impairing long-range end resection during double-strand break (DSB) repair. These findings establish UNI110 as a promising lead compound for the targeted treatment of MMR-deficient colorectal cancers, offering a potential breakthrough in overcoming chemotherapy resistance and improving patient outcomes.

Keywords: Baicalein; UNI110; end resection; homologous recombination; mismatch repair.

Figure 1.

Synthesis of UNI110. Scheme of UNI110 synthesis. Baicalein (1), 4-oxo-2-phenyl-4H-chromene-5,6,7-triyl triacetate (2), benzyl bromide derivative (3), and UNI110 (4).

Figure 2.

UNI110 selectively kills MutSa deficient cells. A, B. HEC59 and HEC59-2 cells were treated with the indicated doses of UNI110 for 24 hours. Cell viability was measured by Cell Titer-Glo reagent. Data are represented as mean ± SD (n = 3). The IC50 was determined using non-linear regression analysis. C. HEC59 and HEC59-2 cells were treated with 8 µM UNI110 for 24 hours. Isolated proteins were subjected to immunoblotting using the indicated antibodies. D. HEK293T cells were transfected with control and MSH2 siRNAs. The following day, cells were treated with 8 µM UNI110 for 24 hours. Isolated proteins were then analyzed using immunoblotting with the indicated antibodies.

Figure 3.

UNI110 decreases the efficiency of HR. A. HR reporter cell lines were treated with the indicated concentrations of UNI110 for 48 hours. GFP positive cells were analyzed by FACS. RAD51 inhibitor B02 was used as positive control. The data are represented as mean ± SD (n =3), and statistical significance was assessed using an unpaired t-test. B. HEK293T cells were treated with 8 µM UNI110 for 24 hours. The cell cycle profiles were analyzed by FACS. The data are represented as mean ± SD (n =3), and statistical significance was assessed using an unpaired t-test.

Figure 4.

UNI110 decreases the efficiency of HR. A. HEK293T cells were treated with 4 µM UNI110 for 24 hours. Extracts were immunoprecipitated using an MSH2 antibody, and the interacting proteins were subsequently analyzed by immunoblotting with the specified antibodies. B. HEK293T cells were treated with 8 µM UNI110 for 24 hours, followed by treatment with 1 µM CPT and 4 µM Aphidicolin. Subsequently, the cells were permeabilized and incubated with an anti-RPA32 antibody. After staining with an Alexa Fluor 488-conjugated secondary antibody, the cells were analyzed by FACS. The data are presented as mean ± SD (n =3), and statistical significance was assessed using two-way ANOVA. C. U2OS cells were treated with 8 µM UNI110 for 24 hours, followed by a 1-hour treatment with 1 µM CPT. The cells were permeabilized and fixed, then incubated with a primary antibody followed by an Alexa Fluor 488-conjugated secondary antibody. Nuclei were stained with DAPI for visualization. Scale bar: 20 µm. D. Schematic representation of the mechanism.