A novel three-component cascade reaction was discovered and developed to synthesize pyridodiindoles with the assistance of microwave irradiation. A collection of pyridodiindoles was prepared by means of the mild reaction and simple operation procedure, which could be applicable to a broad scope of functional aldehydes. Screening demonstrated that compound 5g exhibited a good potency in HCT116 cell lines, and this work validated the feasibility of this novel reaction for generating promising bioactive compounds.