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Nanodiamonds(NDs)have been widely used in drug delivery.It is critical to understand the details of intracellular membrane trafficking and crosstalk with other cellular metabolic processes.Here we exploited the endocytosis and membrane trafficking of NDs labeled with fluorophore or fluorescence protein in several types of tumor cells.We found that cellular uptake of NDs involved multiple endocytotic pathways.A major part of the endocytotic vesicles experienced maturation from early endosome to late endosome and fused with lysosome eventually.Importantly,we observed significant colocalization of NDs with the protein marker of autophagosome LC3,suggesting an interference of the autophagic activity of the cells.We further confirmed the interaction between NDs and autophagic pathway with biochemical assays.Tumor cells often use autophagy as a source of energy to promote their survival and growth.A number of clinical trials have revealed the promising role of autophagic regulators as novel antitumor drugs.Indeed,we proved that NDs increased apoptosis of tumor cells under stress conditions.Our results provide new insights into NDs-based tumor therapy.