Herein, we report a cerium oxide nanocubes (ncCeO2)-reduced graphene oxide (RGO)-based nanocomposite for the detection of oral cancer biomarker, cytokeratin fragment-21-1 (Cyfra-21-1), using the electrochemical technique. Nanocomposite of ncCeO2-RGO was prepared by the in situ reduction of graphene oxide (GO), in the presence of ncCeO2, using hydrazine hydrate. Raman spectra confirmed the presence of ncCeO2 in the matrix of RGO. The chemical composition of the ncCeO2-RGO nanocomposite was determined by X-ray photoelectron spectroscopy (XPS). X-ray diffraction (XRD) studies have indicated the presence of crystalline ncCeO2 and the amorphous nature of RGO. Thin films of ncCeO2-RGO composites were spin coated onto the indium tin oxide (ITO) coated glass surface and used for the co-immobilization of specific antibody of Cyfra-21-1 by N-ethyl-N-(3-dimethyl aminopropyl)carbodiimide hydrochloride and N-hydroxysuccinimide (EDC-NHS) coupling chemistry. Electrochemical response studies were monitored by using the differential pulse voltammetry (DPV) technique in the range of 0.625 pg mL-1 to 15 ng mL-1. The best linear response was observed in the range of 0.625 pg mL-1 to 0.01 ng mL-1, with a low detection limit of 0.625 pg mL-1. The sensitivity was found to be 14.5 μA ng-1 mL cm-2 with R2 0.98, which was an improvement compared to the results from previously reported work. This BSA/anti-Cyfra-21-1/ncCeO2-RGO/ITO immunosensor shows selectivity towards Cyfra-21-1 in the presence of glucose, sodium chloride (NaCl), mucin 16 (MUC-16) and interleukin 8 (IL-8).