Background: Cathepsin D is a lysosomal aspartyl protease. Our previous studies have suggested that cathcpsin D plays an important role in invasion and metastasis of nasopharyngeal carcinoma (NPC). Methods: To identify proteins that interact with cathepsin 1) and gain an insight into the role of cathepsin D in invasion and metastasis of NPC, co-immunoprecipitation (co-IP) combined with mass spectrometry (MS)-based proteomics approach was used. Cathepsin D associated proteins were identified by MS. Protein-protein interaction network were analyzed by bioinformatics including gene ontology (GO), function clustering. And co-IP and western blotting confirmation were for protein-protein interaction. The expression of cathepsin D, epidermal growth factor receptor (EGFR) and heat-shock protein 90A (HSP90A) in NPC were detected by immunohistochemistry (IHC). And the invasion and metastasis capability were detected by Transwell invasion assay. Results: One hundred and forty-one cathepsin D associated proteins were identified, including EGFR and HSP90A, proteins clearly associated with tumor invasion and metastasis. The interaction of these two proteins with cathepsin D was further validated by co-IP followed by western blotting. The 141 proteins were classified into 12 function-related groups. Protein-protein interaction network analysis indicated that cathepsin D might have a significance on invasion and metastasis of NPC by interacting with EGFR and HSP90A. Indeed, cathepsin D/EGFR/HSP90A could form complexes in NPC cells. In addition, overexpression of cathepsin D in NPC cells not only up-regulated EGFR and HSP90A, but also increased the invasive ability of NPC cells. Conclusions: Cathepsin D could enhance the invasion and metastasis capability of NPC cells may through binding to EGFR and HSP90A and triggering the activation of the signaling pathways.