Nanohydrogels show great potential as efficient drug carriers due to their biocompatibility, low toxicity, and high water absorbability. In this paper, we prepared twoO-carboxymethylated chitosan (OCMC)-based polymers functionalized with β-cyclodextrin (β-CD) and amino acid. The structures of the polymers were characterized by Fourier Transform Infrared (FTIR) Spectroscopy. Morphological study was carried out on a Transmission Electron Microscope (TEM), and the results indicated that the two polymers had irregular spheroidal structure with some pores distributed on their surface. The average particle diameter was below 500 nm, and the zeta potential was above +30 mV. The two polymers were further used for preparing nanohydrogels loaded with anticancer drugs lapatinib and ginsenoside Rg1, and the resulting nanohydrogels showed high drug loading efficiency and pH-sensitive (pH = 4.5) drug release behavior.In vitrocytotoxicity investigation revealed that the nanohydrogels exhibited high cytotoxicity against lung cancer (A549) cells.In vivoanticancer investigation was performed in a transgenicTg(fabp10:rtTA2s-M2; TRE2:EGFP-krasV12) zebrafish model. The results showed that the synthesized nanohydrogels significantly inhibited the expression ofEGFP-krasv12oncogene in zebrafish liver, and theL-arginine modified OCMC-g-Suc-β-CD nanohydrogels loading lapatinib and ginsenoside Rg1 showed the best results.