Purpose We previously reported that autophagy in tumor cells plays a critical role in cross-presentation of tumor antigens and that autophagosomes are efficient antigen carriers for cross-priming of tumor-reactive CD8+ T cells. Here we sought to characterize further the autophagosome-enriched vaccine named DRibble (DRiPs-containing blebs), derived from tumor cells after inhibition of protein degradation and provide insights into the mechanisms responsible for their efficacy as a novel cancer immunotherapy. Experimental Design DRibbles were characterized by western blot and light or transmission electron microscopy. The efficiency of cross-presentation mediated by DRibbles was first compared with that of whole tumor cells and pure proteins. The mechanisms of antigen cross-presentation by DRibbles were analyzed and the anti-tumor efficacy of the DRibble vaccine was tested in 3LL Lewis lung tumors and B16F10 melanoma. Results The DRibbles sequester both long-lived and short-lived proteins, including defective ribosomal products (DRiPs), as well as damage-associated molecular pattern (DAMP) molecules exemplified by HSP90, HSP94, calreticulin, and HMGB1. DRibbles express ligands for CLEC9A, a newly described C-type lectin receptor expressed by a subset of conventional DCs (cDCs) and plasmacytoid DCs (pDCs) and cross-presentation was partially CLEC9A-dependent. Furthermore, this autophagy assisted antigen cross presentation pathway involved both caveolae- and clathrin-mediated endocytosis and ERAD machinery. It depends on proteasome and TAP1, but lysosome functions of APCs. Importantly, DC loaded with autophagosome-enriched DRibbles can eradicate 3LL Lewis lung tumors and significantly delay the growth of B16F10 melanoma. Conclusion These data documented the unique characteristics and potent anti-tumor efficacy of the autophagosome-based DRibble vaccine. The efficacy of DRibble cancer vaccine will be further tested in clinical trials.