| Research backgroundColorectal cancer(CRC),an aggressive primary intestinal malignancy,is the third most common cancer and the second cause of cancer-related death worldwide.Many patients,Such as 50%of KRAS/NRAS/BRAF wild-type,5%-10%of BRAF V600E mutation and 5%of microsatellite instability and mismatch repair deficiency,have already benefited from EGFR-targeted,BRAF-targeted therapies or immunotherapy.However,effective targeted therapies are not yet available for 35%to 40%of patients with KRAS or NRAS mutation.Thus,targeting KRAS is an attractive approach in KRAS-mutated mCRC.The success of KRASG12C inhibitors(such as AMG510)has opened a new era of targeting KRAS protein.The development of KRASG12D inhibitors(such as MRTX1133)is also making new progress.Although MRTX1133 has shown its inhibitory effect on proliferation in KRASG12D mutant pancreatic cancer and colorectal cancer,we found that the Application of MRTX1133 in KRASG12D mutant colorectal cancer could lead to adaptive changes and drug tolerance.Therefore,the molecular mechanism of these adaptive changes and their blocking strategy are worthy of further discussion.Research methods1.Cell clone formation experiment was used to observe the changes of cell growth after drug intervention and elution,and the changes of molecular signals were analyzed by Western Blot and transcriptome data.2.CRISPR library and drug library screening were used to find strategies to block adaptive changes.3.Western blotting,immunohistochemistry,RNA interference experiments,and transcriptome data were used to analyze the molecular mechanism of blocking adaptive changes.4.Verify the strategy and its molecular mechanism in the cell line,organoid,and cell line-derived xenograft models.Research results1.Cell clone formation assay showed that the level of cell quantity recovered after MRTX1133 elution,and phosphorylated extracellular signal-regulated kinase(p-ERK)rebounded was found by transcriptome analysis.2.CRISPR kinase library screening showed that EGFR knockout had a synergistic lethal effect,while drug screening found that EGFR blocking and MRTX1133 aslo had a synergistic effect.3.Western blotting and immunohistochemistry showed that EGFR was activated,and ERBB receptor feedback inhibitor 1(ERRFI1)was down-regulated by analyzing the transcriptome data.4.There is a synergistic effect between EGFR inhibitor and MRTX1133 in cell line,organoid,and cell line-derived xenograft models.Research conclusionThe intervention of MRTX1133 resulted in reversible growth arrest of KRASG12D mutated CRC cells,accompanied by partial reactivation of RAS effect signals.Mechanistically,MRTX1133 treatment down-regulates the expression of ERBB receptor feedback inhibitor 1(ERRFI1),leading to EGFR feedback activation.Wild-type RAS further mediates the activation of signal transduction downstream of EGFR,which leads to the rebound of RAS efffectors and reduces the efficacy of MRTX1133.Therefore,the combination of EGFR inhibitors and MRTX1133 can more effectively inhibit KRASG12D mutant CRC organoids and cell line-derived xenografts proliferation.In summary,our study revealed that the feedback activation of EGER is an important molecular event that limits the efficacy of KRASG12D inhibitors,and established a potential combination therapy of KRASG12D and EGFR inhibitors for CRC patients with KRASG12D mutant. |
