To avoid the harmful side effects of cetuximab and improve its therapeutic efficacy, egg serum albumin (ESA) was used as a targeting drug carrier moiety for cancer therapy against Caco-2 colon cancer cells. The simple improved desolvation method was used to synthesize ESA nanoparticles (ESA-NPs) and cetuximab-loaded albumin nanoparticles (CET-ANPs) with glutaraldehyde as a crosslinking agent. The ESA-NPs and CET-ANPs were spherically shaped, and their sizes and surface potentials were 100 and − 24 nm and 170 and − 20 nm, respectively, as determined using transmission electron microscopy (TEM) and a Zeta potential analyzer. The specific functional groups of the prepared nanoparticles were revealed by FTIR analysis. In the MTT (3-(4, 5-dimethylthiazolyl-2)-2, 5-diphenyltetrazolium bromide) assay, CET-ANPs exerted the highest antitumor activity after 24 h followed by CET, ESA-NPs, and pure ESA. Combination of CET + ESA-NPs at different IC50 concentrations at ratios of 1:1, 1:2, 2:1, 1:4, 4:1, 1:9, or 9:1 showed significant synergistic effects with a combination index (CI) > 1. Furthermore, the CET either loaded with ESA-NPs or administered in combination (CET + ESA NPs) caused significant apoptotic damage, as well as an S-phase or G2/M cell cycle arrest to the cancer cells, respectively. These were directly linked with a significant upregulation of mRNA expression of Caspase3 and Bax genes and an extreme downregulation of the mRNA expression of Bcl2, particularly in the combination treatment group, as compared to the untreated cells. Finally, ESA-NPs improved the effectiveness of cetuximab, strongly caused apoptotic and antiproliferative action with lower systemic toxicity, and could be suggested for the targeted administration of anticancer medications in various nanosystems.