A new kind of magnetic gelatin-encapsulated nanoassembly was fabricated as a multifunctional drug delivery system (DDS) with pH, redox, and enzyme stimuli-responsive drug release for cancer theranostics. Cystamine-modified gelatin was used to encapsulate doxorubicin (Dox) as an anticancer drug and Fe₃O₄ as the magnetic resonance imaging (MRI) contrast agent to assembly into a nanoassembly via the formation of an oil-in-water microemulsion. Such a DDS system can provide long-term Dox release and more effective drug availability, since the acidic environment, high glutathione (GSH) and matrix metalloproteinase (MMP) level of cancer tissues. And it would beneficial for high accumulation around tumor through the enhanced penetration and retention (EPR) effect, and deeper penetration into tumor because of its enzyme-degradable and size-changeable ability. This drug delivery vehicle had good biocompatibility. It can effectively reduce the efflux of drugs when these nanoparticles were uptaken by cells. Besides, the Dox-Fe₃O₄@gelatin nanoparticles can be a contrast agent for T₂-weighted MRI.