There are several limitations to the application of nanoparticles in the treatment of cancer, including their low drug loading, poor colloidal stability, insufficient tumor penetration, and uncontrolled release of the drug. Herein, gelatin/laponite (LP)/doxorubicin (GLD) nanoparticles are developed by crosslinking LP with gelatin for doxorubicin delivery. GLD shows high doxorubicin encapsulation efficacy (99%) and strong colloidal stability, as seen from the unchanged size over the past 21 days and reduced protein absorption by 48-fold compared with unmodified laponite/doxorubicin nanoparticles. When gelatin from 115 nm GLD reaches the tumor site, matrix metallopeptidase-2 (MMP-2) from the tumor environment breaks it down to release smaller 40 nm LP nanoparticles for effective tumor cell endocytosis. As demonstrated by superior penetration in both in vitro three-dimensional (3D) tumor spheroids (138-fold increase compared to the free drug) and in vivo tumor models. The intracellular low pH and MMP-2 further cause doxorubicin release after endocytosis by tumor cells, leading to a higher inhibitory potential against cancer cells. The improved anticancer effectiveness and strong in vivo biocompatibility of GLD have been confirmed using a mouse tumor-bearing model. MMP-2/pH sequentially triggered anticancer drug delivery is made possible by the logical design of tumor-penetrating GLD, offering a useful method for anticancer therapy.